Clarissa Silva Martins*, Daniel Elias, Leandro Machado Colli, Carlos Eduardo Couri, Manoel Carlos L A Souza, Ayrton C. Moreira, Lucila Elias, Milton Cesar Foss and Margaret De Castro
Ribeirao Preto Medical School - University of Sao Paulo, Ribeirao Preto, Brazil

 

Introduction: A hypothetical role of glucocorticoid (GC) in the pathogenesis of human Metabolic Syndrome (MetS) has been suggested since hypercortisolism shares several similarities with MetS. It has been postulated that excessive activity of CG within the normal physiological range, either by increased GC receptor (GR) sensitivity to the hormone or by altered cortisol metabolism, may contribute to the pathogenesis of insulin resistance and the development of cardiovascular risk factors.

Objectives: To evaluate the hypothalamic-pituitary-adrenal (HPA) axis sensitivity to dexamethasone (DEX), NR3C1 polymorphisms (SNPs), gene expression of GRα and GRβ isoforms, and a panel of cytokines (IL1B, IL2, IL4, IL6, IL8, IL10, TNF and IFNG) in peripheral immune cells of MetS patients and controls.

Materials and Methods: Prospective study with 40 MetS patients and 40 controls was conducted at the School of Medicine of Ribeirao Preto University Hospital. Plasma (PF) and salivary (SF) cortisol were measured by RIA in basal conditions and after 0.25, 0.5 and 1mg of DEX given at 2300h. In addition, p.N363S (rs6195), p.ER22/23EK (rs6189-6190), and BclI (rs41423247) SNPs were evaluated by qPCR allelic discrimination. Exons 3 to 9 and exon/intron boundaries of NR3C1 were sequenced. GR isoforms and cytokines expression were assessed by qPCR.

Results: PF (nmol/l) and SF (nmol/l) after 0,5mg DEX were higher in MetS patients compared to controls (PF: 119.0±24 vs. 57.5±2.9; p=0.07 and SF: 7.6±2.0 vs. 4.5±1.3; p=0.05) and also after 1mg DEX (PF: 70.2±17.3 vs. 37.9±2.6; p=0.02 and SF: 4.9±1.7 vs. 2.2±0.3; p<0.0001). After all DEX doses, a lower number of MetS patients suppressed PF and SF compared to controls. The BclI genotypic frequencies (%) differed between patients and controls (CC:56/CG:44 and CC:50/CG:32.5/GG:17.5; respectively, p=0.03). TheGRβ was overexpressed (fold=100.0; p=0.002) and IL4 (fold=-265.0; p<0.0001) was underexpressed in MetS patients.

Conclusions: MetS patients exhibited decreased hypothalamus-pituitary sensitivity to GC feedback, confirming the HPA axis activation in this condition. The lower frequency of the BclI polymorphism may contribute to this HPA axis dysregulation. Moreover, the MetS low grade chronic inflammation appears to promote GRβ overexpression, resulting in tissue-specific GC resistance, one of the mechanisms through which GCs may consume peripheral subcutaneous adipose tissue and promote the preferential expansion of visceral adipose tissue. Our data support the emerging concept that HPA axis disruption might be involved in the incompletely understood pathogenesis of MetS.

 

Nothing to Disclose: CSM, DE, LMC, CEC, MCLAS, ACM, LE, MCF, MD

PP06-1 21184 1.0000 THR-400 A HPA Axis Dysregulation, NR3C1 Polymorphisms and Glucocorticoid Receptor Isoforms Imbalance in Metabolic Syndrome 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 11:30:00 AM PP06 5906 11:15:00 AM Adrenal Tumors and Glucocorticoid Action Poster Preview


Gabriela R V Sousa*1, Tamaya C Ribeiro1, Andre M Faria1, Beatriz M P Mariani2, Antonio M Lerario1, Maria Claudia N Zerbini2, Ibere C Soares3, Alda Wakamatsu2, Venancio A F Alves2, Berenice B Mendonca4, Maria Candida B V Fragoso5, Ana Claudia Latronico1 and Madson Q. Almeida5
1Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil, 2Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil, 3Hospital do Câncer de Barretos, Porto Velho, Brasil, 4School of Medicine, Sao Paulo University, São Paulo, Brazil, 5Hospital das Clínicas & Instituto do Câncer do Estado de São Paulo (ICESP), Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil

 

Introduction: Deregulation of microRNA (miRNA) expression in adrenocortical carcinomas (ACCs) has important prognostic and therapeutic implications. DICER1, an endoribonuclease in the RNase III family, is a key component of the miRNA processing machinery. In cooperation with its cofactor, transactivation response (TAR) RNA-binding protein (TRBP2), DICER1 cuts the pre-miRNA to meet the specification. Low DICER1 expression was associated with poor clinical outcome in ovarian, breast and lung cancer. Recently, DICER1 mutations in the RNase IIIb domain were found in steroidogenic Sertoli–Leydig cell and embryonic tumors. Aim: In this study, we assessed the mRNA and/or protein expression of DICER1 and TARBP2 in adult adrenocortical tumors (ACTs). Additionally, we investigated DICER1 and TARBP2 hot spot mutations. Expression of miRNAs (miR-103, miR-107 and miR-497) involved in DICER1 and TARBP2 regulation was also determined. Patients and methods: DICER1 protein expression was analyzed in 151 adult ACTs (77 adenomas and 74 carcinomas). Messenger DICER1 and TARBP2 expression was assessed in 55 ACTs (30 adenomas and 25 carcinomas) by quantitative real-time PCR. Expression of miR-103, miR-107 and miR-497 was determined in the same cohort. In addition, we sequenced four metal-binding sites (codons 1709 and 1705 in exon 24; codons 1810 and 1813 in exon 25) within the RNase IIIb catalytic center of DICER1 gene, and the (C)5 coding microsatellite repeat of exon 5 of TARBP2 gene in 61 ACTs. Results: Low DICER1 protein expression was significantly associated with reduced overall (p= 0.01) and disease-free survival (p= 0.01) in ACC patients. Among ACC patients with stage 3 or 4, 22 out of 32 (69%) displayed low DICER1 protein expression (X2= 10.1, p= 0.01). DICER1 protein expression was not significantly different between adenomas and carcinomas. At mRNA level, DICER1 expression had an important superposition between adenomas and carcinomas, but it was higher in carcinomas than in adenomas [median (range); 1.96 (-1.4 to 4.54) vs. 0.81 (-1.5 to 99) respectively, p= 0.03]. Among ACC patients, DICER1 mRNA levels did not predict outcome. TARBP2 expression was not associated with histological and clinical parameters. In addition, expression of miR-103, miR-107 and miR-497 did not correlate with DICER1 and TARBP2 expression in ACTs. No variant was identified in the hot spot mutation region of DICER1 and TARBP2 genes. Conclusion: Low DICER1 expression was significantly associated with poor outcome in adult ACC patients. However, DICER1 deregulation in ACCs was not caused by mutations within the RNase IIIb domain and not associated with expression of its regulatory miRNAs. Support: FAPESP (2012/21272-6; 2013/09621-8)

 

Nothing to Disclose: GRVS, TCR, AMF, BMPM, AML, MCNZ, ICS, AW, VAFA, BBM, MCBVF, ACL, MQA

PP06-2 18893 2.0000 THR-384 A Low DICER1 Expression Is Associated with Poor Clinical Outcome Adrenocortical Carcinoma 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 11:30:00 AM PP06 5906 11:15:00 AM Adrenal Tumors and Glucocorticoid Action Poster Preview


Petra Bullova*1, Jan Schovanek1, Yasin Tayem1, Alessio Giubellino1, Robert Wesley2, Nikoletta Lendvai1, Svenja Nolting1, Zdenek Frysak3, Shivaani Kummar1 and Karel Pacak4
1National Institutes of Health, 2National Institutes of Health, Bethesda, MD, 3Department of Internal Medicine III – Nephrology, Rheumatology and Endocrinology, University Hospital of Olomouc, Czech Republic, 4National Institutes of Health, Eunice Kennedy Shriver National Institute of Child Health and Human Development, Bethesda, MD

 

Pheochromocytomas and paragangliomas (PHEO/PGL) are neuroendocrine tumors derived from chromaffin cells. Up-to-date, the only curative option for primary tumors is surgery. In case of metastatic disease, CVD treatment and 131I-MIBG radiation therapy remain the most effective ways of stabilization of the disease and decreasing the tumor burden. Unfortunately, these options are effective only in a small portion of the patients as 131I-MIBG therapy is limited to the tumors showing its uptake and tumor cells often become resistant to CVD with time. Therefore, there is an extensive research focused on finding a permanent and effective cure for metastatic PHEO/PGL.

In the present study, we studied a novel topoisomerase I inhibitor, LMP-400, as a potential treatment for this disease. First, we evaluated presence of topoisomerase I in human metastatic tumors. We found a significantly higher expression of topoisomerase I in these tumors compared to normal adrenal medulla and primary tumors, which provided us with a basis for further studies. LMP-400 inhibited cell growth of established mouse and rat PHEO cell lines and also of human primary tumor tissue cells in vitro. In a study performed in athymic female mice, LMP-400 showed a significant inhibitory effect on tumor growth and metastases formation in two evaluated drug administration regimens. Furthermore, we studied effects of the compound on hypoxia pathway and we found that low doses of LMP-400 decreased protein levels of hypoxia-inducible factor 1 alpha (HIF-1α), one of the transcription factors studied as potential metastatic drivers in these tumors. HIF-1α decrease resulted in changes in the mRNA levels of HIF-1α transcriptional targets. In addition, we performed in vitro cell growth inhibition studies combining LMP-400 with other chemotherapeutic drugs currently used in clinics and it showed an increased growth inhibitory effect in the combination than when treated individually.

In conclusion, we believe that based on our results showing the inhibitory effect of LMP-400 on tumor growth in vitro and in vivo and affecting hypoxia pathway, which plays a major role in these tumors, the compound is a promising treatment option for patients with metastatic PHEO/PGL. In addition, its effect at low concentrations in combination with other drugs makes it a very potential candidate for its use in future clinical trials.

 

Nothing to Disclose: PB, JS, YT, AG, RW, NL, SN, ZF, SK, KP

PP06-3 21003 3.0000 THR-385 A Inhibitory Effect of Topoisomerase I Inhibitor LMP-400 on in Vitro and in Vivo Animal and Human Pheochromocytoma Cells and Models 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 11:30:00 AM PP06 5906 11:15:00 AM Adrenal Tumors and Glucocorticoid Action Poster Preview


Annabel Sophie Berthon*1, Fabio R Faucz2, Mihail Zilbermint1, Guillaume Assie3, Maya Beth Lodish1, Eva Szarek1, Giampaolo Trivellin4, Ninet Sinaii2, Rossella Libe3, Stéphanie Espiard1, Ludivine Drougat3, Bruno Ragazzon3, Benjamin Feldman2, Mihaela Serpe5, Jerome Yves Bertherat6 and Constantine A Stratakis1
1National Institutes of Health (NIH), Bethesda, MD, 2National Institutes of Health, Bethesda, MD, 3INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, France, 4National Institute of Health, Bethesda, MD, 5National Institut of Health, 6INSERM U 1016, Cochin Institute, Paris Descartes University, Paris, France

 

Recently, Armadillo Repeat Containing 5 (ARMC5) gene has been identified as a potential tumor suppressor gene for cortisol-producing primary macronodular adrenal hyperplasia (PMAH). These results were confirmed in our cohort of PMAH patients in which germline ARMC5 mutations were found in 15 out 34 patients. According to the expected tumor suppressor role of ARMC5, in silico analysis identified 7 mutations (20.6%) that are predicted to affect ARMC5 expression. Moreover, all patients carrying ARMC5 mutations had clinically significant Cushing’s syndrome. To elucidate the function of ARMC5, which remains unknown, we are using two different strategies. We first developed two transient Danio rerio models: a loss-of-function model created using antisense morpholino-oligonucleotide (MO) to knock down the zebrafish armc5 ortholog, and a gain-of-function created by injection of in vitro synthetized zebrafish armc5 mRNA. To further investigate which signaling pathways are modified in these two models, we performed RNA-seq on both armc5 MO and mRNA-injected embryos at 30 hours post fertilization. The comparative analysis of these two conditions suggests an important role of Armc5 in apoptotic processes, which corroborates in vitro human findings. We also used the CRISP/Cas9 methodology to mutate the Drosophila melanogaster armc5 ortholog. The armc5 null mutant flies appear to be developmental lethal. The results obtained from these two animal models lead to the identification of the ARMC5 pathway that is essential for tumor development in humans. We are also in the process of generating a KO mouse in the laboratory to better understand the role of this gene in adrenocortical pathology.

 

Disclosure: JYB: Investigator, Ipsen, Investigator, Novartis Pharmaceuticals, Advisory Group Member, atterocor. Nothing to Disclose: ASB, FRF, MZ, GA, MBL, ES, GT, NS, RL, SE, LD, BR, BF, MS, CAS

PP06-4 21417 4.0000 THR-386 A Functional Investigation of a New Aimah Suppressor Gene, ARMC5, Using Animal Models 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 11:30:00 AM PP06 5906 11:15:00 AM Adrenal Tumors and Glucocorticoid Action Poster Preview


Cadence True*1, Diana Takahashi1, M Susan Smith2 and Kevin L. Grove1
1Oregon National Primate Research Center, Beaverton, OR, 2OR National Primate Rsrch Ctr, Beaverton, OR

 

The hypothalamic neurocircuitry of metabolically-gated reproductive function has been well-studied in the rodent.  However, recent neuroanatomical data has demonstrated significant differences in the colocalization patterns for some of these key neuropeptides in the human brain.  To determine whether the non-human primate is a better anatomical model of the human we investigated the immunohistochemical localization of several key neuropeptides regulating reproduction and feeding in the infundibular nucleus, the primate equivalent of the arcuate nucleus (ARH), in ovariectomized adult female rhesus macaques.  Kisspeptin, neurokinin B and dynorphin are three important reproductive neuropeptides that are all coexpressed (KNDy neurons) in the ARH of rodents. While colocalization of kisspeptin-immunoreactivity (-ir) and NKB-ir was observed in the non-human primate as previously reported, very few fibers containing both kisspeptin-ir and dynorphin-ir were detected, consistent with recent data from humans.  To investigate the neurocircuitry implicated in food intake we examined the anorexigenic neuropeptides proopiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART), which are coexpressed in the ARH of the rodent.  Additionally, we investigated the orexigenic neuropeptides, neuropeptide Y (NPY) and agouti-related peptide (AgRP), which are also coexpressed in the rodent.  Colocalized NPY-ir and AgRP-ir fibers were observed indicating preservation of this coexpression pattern in rodents, non-human primates and humans.  As previously reported no coexpression of CART-ir and alpha-melanin-stimulating hormone-ir (a POMC gene product) was observed in the non-human primate. Instead there was abundant colocalization of CART-ir and NPY-ir fibers in the infundibular nucleus similar to observations in humans. CART-ir was also observed in kisspeptin-ir fibers, once again similar to humans, indicating that these cells may be critical for linking metabolic and reproduction function in the primate.  Overall these findings highlight major neuroanatomical differences between rodents and primates including lack of evidence for canonical kisspeptin/neurokinin B/dynorphin and POMC/CART coexpression patterns in the primate.  We propose that compared to the rodent, the non-human primate is a better-suited model organism to study the hypothalamic mechanisms by which negative and positive metabolic states lead to inhibition of reproductive function in humans.

 

Disclosure: KLG: Advisory Group Member, Novo Nordisk, Principal Investigator, Novo Nordisk, Consultant, Ember, Principal Investigator, ERX, Consultant, Sanofi, Employee, Novo Nordisk. Nothing to Disclose: CT, DT, MSS

PP05-2 19082 2.0000 THR-471 A Coexpression of Key Neuropeptides Regulating Reproduction and Feeding in the Non-Human Primate: Differences from the Rodent 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 11:30:00 AM PP05 5912 11:15:00 AM GnRH & Neuroendocrinology Poster Preview


Darwin Omar Larco*1, Madelaine J Cho-Clark1, Shailaja K Mani2 and T John Wu1
1Uniformed Services University, Bethesda, MD, 2Baylor College of Medicine, Houston, TX

 

Gonadotropin-Releasing Hormone (GnRH) neurons originate outside the central nervous system (CNS) in the nasal placode where their migration to the basal forebrain is dependent on the integration of multiple signaling cues during development. The proper migration and establishment of the GnRH neuronal population within CNS is critical for normal pubertal onset and reproductive function. The endopeptidase EP24.15 is expressed along the migratory path of GnRH neurons and cleaves the full-length GnRH to generate the metabolite GnRH-(1-5). Using the GN11 cell model, which is considered an immature GnRH neuronal cell line, we recently demonstrated that GnRH-(1-5) inhibits cellular migration in a wound closure assay by binding the orphan G protein-coupled receptor 173 (GPR173). GnRH-(1-5) activating GPR173 initiates the formation of a complex with b-arrestin 2 and PTEN to subsequently inhibit the STAT3 pathway to regulate migration. In this study, we examined changes in genes, matrix metalloproteinase (MMP) activity, and secreted chemokines, which are implicated in the regulation of cell migration, in GN11 cells treated with GnRH-(1-5). First, we used a PCR array (Qiagen) to measure changes in 84 genes in cells treated with GnRH-(1-5) for 30 min. Acute GnRH-(1-5) treatment significantly decreased members of the cytokine signaling pathway such as IL-1alpha (0.62 relative to 1.0) and IL-4Ralpha (0.73 relative to 1.0), which correlates with our previous finding that GnRH-(1-5) inhibits the normally cytokine-activated STAT3 pathway. We also measured the activity of MMP-2 and MMP-9 by zymography in GN11 cells exposed to GnRH-(1-5) and found no significant changes. Furthermore, we measured the secretion of chemokines that are important regulators of migration in cells treated for 1h and 24h with GnRH-(1-5). Similarly, GnRH-(1-5) did not alter the levels of CXCL1, CXCL2, CXCL9, CXCL10, ccl2, and ccl5. In addition, the TGF-beta family was measured and we found that GN11 cells secrete TGF-beta1 significantly more that TGF-beta2 and -3. However, GnRH-(1-5) did not significantly change the levels of the TGF-beta family members. These results suggest that GnRH-(1-5) may modulate the response of migrating GnRH neurons to external cues present in the extracellular milieu (ECM). Therefore, we examined the effect of GnRH-(1-5) on the migration of GN11 cells in the presence of an in vitro ECM using a matrigel invasion assay. Interestingly, GnRH-(1-5) (100 nM) significantly (p < 0.05) inhibited the ability of GN11 cells to migrate through matrigel environment relative to control cells and is dependent on GPR173 as determined by siRNA experiments. Collectively, our studies demonstrate that the downstream mechanism of GnRH-(1-5) binding GPR173 to decrease cell migration and invasion is ECM-dependent and likely involves changes in the response and sensitivity of GnRH neurons to external cues.

 

Nothing to Disclose: DOL, MJC, SKM, TJW

PP05-3 19928 3.0000 THR-459 A The Extracellular Environment Regulates the Effect of GnRH-(1-5) on the Migration of GN11 Cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 11:30:00 AM PP05 5912 11:15:00 AM GnRH & Neuroendocrinology Poster Preview


Ali Abbara*1, Channa N Jayasena2, Shakunthala Narayanaswamy3, Chioma Izzi-Engbeaya3, Alexander N Comninos1, Ailish Coblentz4, Zainab Malik3, Subair Sarang4, Mathini Sridharan4, Deborah Peters4, Mohammad A Ghatei3, Stephen R Bloom2 and Waljit S Dhillo1
1Imperial College NHS Healthcare Trust, London, United Kingdom, 2Imperial College London, United Kingdom, 3Imperial College London, London, United Kingdom, 4Imperial College London

 

Introduction: Ageing in the male is commonly associated with a fall in serum testosterone termed Late-Onset Hypogonadism (LOH). LOH is thought to occur partly due to decreased testicular function, however the compensatory luteinizing hormonal response is also often blunted. This suggests that ageing may be associated with disordered gonadotropin releasing hormone (GnRH) release from the hypothalamus, or reduced gonadotropin release from the pituitary gland. It has not previously been possible to make a distinction between examining hypothalamic and pituitary function in the male reproductive axis; however the recent identification of kisspeptin, which potently stimulates GnRH release, makes this possible. We therefore used kisspeptin-54 (KP54) and GnRH administration to investigate whether hypothalamic or pituitary reproductive function was altered in older males when compared with a young control group.

Methods: Following ethical approval, we carried out a single-blinded placebo-controlled study. A group of older men (mean age 61.0yrs) and a control group of younger men (mean age 28.9yrs) each underwent 3 study visits at least 1 week apart, during which they received vehicle, kisspeptin or GnRH (n=5 per group). Ten minutely blood sampling for luteinizing hormone (LH), follicle stimulating hormone (FSH) and testosterone was performed throughout each 4h study visit. After 1h of baseline blood sampling, a 3h intravenous infusion of vehicle, 0.1 nmol/kg/h of KP54 or GnRH, was commenced and reproductive hormonal response assessed. Mean AUC of change in each reproductive hormone from baseline was compared by unpaired t test with P<0.05 regarded as significant.

Results: Mean BMI and baseline testosterone were 23.6 kg/m2, 20.6 nmol/l in the young men (YM) and 25.4 kg/m2, 16.9 nmol/l in the older male (OM) groups, respectively. No significant differences were observed in the LH responses between young and older men following either KP54 or GnRH infusion (mean±SEM AUC LH change in iU.h/L: KP54 YM 12.1±1.6, KP54 OM 12.1±2.0, P=ns vs. KP54 YM; GnRH YM 19.6±3.2, GnRH OM 18.6±3.4, P=ns vs. GnRH YM). FSH response was not significantly different in the older age group to both KP54 and GnRH (mean±SEM AUC change in FSH in iU.h/L: KP54 YM 1.6±0.3, OM 2.5±0.8, P=ns vs. KP54 YM; GnRH YM 2.3±0.4, OM 4.1±1.8; P=ns vs GnRH YM).

Conclusion: This pilot study suggests that older men without known LOH have similar reproductive hormonal response to KP54 when compared with younger men. Thus KP54 offers a potential novel tool for examining hypothalamic function in older men.

 

Nothing to Disclose: AA, CNJ, SN, CI, ANC, AC, ZM, SS, MS, DP, MAG, SRB, WSD

PP05-4 21182 4.0000 THR-469 A Kisspeptin- a Novel Test of Hypothalamic Function in Older Men 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 11:30:00 AM PP05 5912 11:15:00 AM GnRH & Neuroendocrinology Poster Preview


Susan Jung*, Mandy Richter-Kraus, Florian Brackmann, Helmuth G. Doerr and Regina Trollmann
University Hospital Erlangen, Erlangen, Germany

 

Background: Perinatal cerebral hypoxia is a main risk factor for acquired brain injury and neurological disabilities. Stabilization of hypoxia-inducible transcription factors (HIF) modulates mitochondrial respiration, glycolysis, angiogenesis, and vasculogenesis mediating adaptation to reduced oxygen concentration. However, HIF stabilization is associated with decreased growth hormone (GH) gene expression in pituitary cells suggesting suppression of GH-induced secretion of neurotrophic factors.

Objective: We hypothesized that exogenous GH modulates hypoxia-induced disturbances of the GH/IGF-1 axis mediating protective effects under global hypoxia during early development.

Methods: A total of 179 mice were randomized at postnatal day 7 (P7) according the complete block design into treatment groups of at least 8 animals and exposed either to normoxia or hypoxia (8% O2) for 6h using an INVIVO400 hypoxia workstation (Baker Ruskinn). Within 30 min after reoxygenation and once daily for three consecutive days, recombinant hGH (Genotropin®, Pfizer) at doses of 0, 10, 50, 250, 1000, and 4000 µg/kg was injected intraperitoneal (i. p.) in the pups. Body weight and length (nose-to-tail length) were monitored and mGH and IGF-1 plasma protein concentrations were quantified at P14 and P21.

Results: Whereas no physiological differences between the treatment groups were observed at P7, treatment with low hGH doses adversely affected perinatal growth and weight gain under normoxia. Mice dosed with 10–250 µg/kg hGH gained 72% less weight (P<0.0001) until P10 and were significantly shorter than non- or sham-treated mice. Thereby, growth retardation persisted until P21 in mice treated with 10 and 50 µg/kg hGH and were correlated to significantly decreased plasma concentration of mGH and IGF-1. Hypoxia-exposed non- and sham-treated animals revealed significantly lower weight gain (P<0.0001) between P7 and P10 than normoxic animals. Simultaneously, exposure to hypoxia significantly decreased plasma mGH (P<0.05) and IGF-1 levels (P<0.05) for at least one week. Notably, treatment with 1000 and 4000 µg/kg hGH restored hypoxia-induced disturbances of the GH/IGF-1 axis compensating growth deficiency in neonatal mice.

Conclusion: Our data indicate that i) perinatal, hypoxia-induced disturbances of the GH-/IGF-1 axis are transient and that ii) hGH treatment stabilizes the somatotropic axis after perinatal hypoxia. Further studies are focused on modification of cerebral neurotropic factors by stabilization of the GH/IGF-1 axis and thus preventing neuronal loss in the developing hypoxic rodent brain.

 

Nothing to Disclose: SJ, MR, FB, HGD, RT

PP03-1 18400 1.0000 THR-146 A Restoration of Hypoxia-Induced Changes of Plasma mGH and IGF-1 Concentrations in Neonatal Mice in Response to hGH Treatment 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 11:30:00 AM PP03 5919 11:15:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Poster Preview


María Gabriela Ballerini*, Paula A. Scaglia, Ignacio Bergadá, Alicia Martinez, Mercedes Altube, Ana C. Keselman, Débora Braslavsky, María Gabriela Ropelato, Hector G. Jasper and Horacio M Domene
Hospital de Niños Dr. Ricardo Gutiérrez, Buenos Aires, Argentina

 

Background: More than 90 GHR gene mutations have been described in complete GH insensitivity (GHI), nevertheless, only few heterozygous GHR genetic variants have been associated to GHI. Little information regarding the impact of common polymorphisms within coding (Ex) and intronic (In) GHRregions on height and components of the GH/IGF-I/IGFBPs system is available.

Aim: To test if GHR gene polymorphisms are associated to height and components of the GH/IGF-I/IGFBPs system in ISS children.

Methods: GHR gene (Ex/In flanking regions) was PCR-amplified and sequenced in 75 unrelated ISS children. The GHR pseudoexon (6Psi) mutation was investigated by RFLP. GHR gene variants were classified as rare (minor allele frequency (MAF) ≤ 1%), less frequent (1-10%) and common (≥ 10%). To test the genotype influence of SNPs with MAF > 10% (HapMap-Project) on height and GH/IGF-I/IGFBPs system, ISS children were grouped as homozygous for the major allele and carriers of one or two copies of the minor allele. The proportion of variants (less than 3 versus 4 or more) was evaluated according to the median value of: height (-2.86 SDS), GHBP (-0.9 SDS; in house functional assay), maximal stimulated-GH (14.9 ng/L), IGF-I (-1.4 SDS), IGFBP-3 (-1.0 SDS, ICMA) and ALS (-1.3 SDS; RIA). Hardy-Weinberg equilibrium, Fisher´s exact test and Mann-Whitney analysis were used.

Results: Six common and 5 less frequent variants were identified: Ex3 deletion (MAF: 26%), rs6179 (Ex6, 23%), rs6180 (Ex10, 42%), rs12521020 (In1, 32%), rs10941579 (In2, 32%), rs33972388 (In7, 37%), rs34223737 (In7, 1%), rs6880730 (In8, 5%), rs6182, rs6184 and rs115376349 (Ex10, 2%). In ISS, MAF distribution was not different from 41 control children or the HapMap database. We identified 4 heterozygous rare variants in 4/75 children (5.3%): p.R229H (Ex7) and p.R386C (Ex10) with normal IGF-I, IGFBP-3 and ALS levels, 1 of them with low GHBP (<-1.9 SDS); rs143475648 (In3) with low GH-dependent biomarkers and c.618+700_705dupCAGCCA (In6) with low IGF-I levels. These variants were not found in control children. Neither individual GHR gene genotypes, nor the sum of GHR polymorphic variants were associated to height, GHBP, IGF-I, IGFBP-3 or ALS (p>0.06).

Conclusions: The sum of polymorphic variants has no effect on GH sensitivity, measured by levels of serum GHBP, IGF-I, IGFBP-3 and ALS. While p.R229H variant may result in a slight reduction in GHR expression as reflected by low GHBP levels, p.R386C, rs143475648 and c.618+700_705dupCAGCCA variants remain to be characterized by in vitro functional studies.

 

Nothing to Disclose: MGB, PAS, IB, AM, MA, ACK, DB, MGR, HGJ, HMD

PP03-2 21111 2.0000 THR-149 A Influence of GH Receptor Gene Variants within Coding and Intronic Regions in Children with Idiopathic Short Stature (ISS) on Height and Components of the GH/IGF-I/IGFBP System 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 11:30:00 AM PP03 5919 11:15:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Poster Preview


Chiara De Leonibus*1, Pierre Chatelain2, Chris Knight3, Peter Clayton4 and Adam Stevens1
1University of Manchester and Royal Manchester Children’s Hospital, Manchester, United Kingdom, 2Université Claude Bernard, Lyon, France, 3University of Manchester, 4University of Manchester & Manchester Academic Health Sciences Centre, Manchester, United Kingdom

 

Background

The rate of normal growth tends to increase in mid-spring/summer and is greater in children living at higher latitudes(1). This study examined i) the impact of living at different latitudes on height velocity (HV) and ii) the interactions between environmental [latitude, summer daylight exposure (SDE)] and genomic factors [single nucleotide polymorphisms (SNPs), gene expression (GE) profiles] that effect HV in response to treatment with recombinant-human growth hormone (r-hGH) in children with growth hormone deficiency (GHD).

Methods

Pre-pubertal naïve GHD patients (n=118) from the PREDICT prospective long-term follow-up study (NCT00699855) were analyzed. The effect of treatment over one year with r-hGH was measured by HV (cm/yr) and assessed in relation to SDE, carriage of seven SNPs previously associated with high growth response(2) and basal GE determined prior to treatment. Data were submitted from 28 centers in 14 countries SDE and HV were correlated with basal GE using rank regression and network models were constructed from the GE overlap to investigate causal interactions.

Results

GHD patients from latitudes with higher SDE had a better HV in response to r-hGH compared to those with intermediate and lower SDE [median (Q1, Q3): 9.8 (8.5, 11.4) vs 8.1 (7.0, 9.8) and 8.2 (7.0, 10.3) cm/yr, respectively; p=0.019], with HV over all groups correlating with SDE (r=0.256, p=0.006). For SNPs within GRB10, IGFBP3, TGF-α, CYP19A1 and TP53 genes, there was a significant interaction between the SNP and SDE (p<0.05) in relation to growth response. The effect of carriage of SNPs in IGFBP3, TGF-α and TP53 on HV was greater in patients from locations with higher SDE; while the converse was found for GRB10 and CYP19A1 Network models derived from SDE and HV associated basal GE identified i) the developmental transcription factor NANOG as a key regulator, and ii) up-regulation of growth-related and “circadian clock” pathways (p=3.0x10-3).

Conclusion

This study showed complex gene-environment interactions between genes modulating growth and SDE in response to r-hGH, with the biological regulation of the circadian clock as a possible linking mechanism. The transcriptional regulator NANOG was found to be a master regulator of these interactions implicated in the development of circadian oscillator action and bone growth.

 

Disclosure: PC: Investigator, Merck Serono, Speaker, Merck Serono, Advisory Group Member, Merck Serono. PC: Investigator, Merck Serono, Speaker, Merck Serono. AS: Investigator, Merck Serono. Nothing to Disclose: CD, CK

PP03-3 21710 3.0000 THR-145 A Effect of Summer Daylight Exposure and Genetic Background on Growth in Growth Hormone Deficient (GHD) Children 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 11:30:00 AM PP03 5919 11:15:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Poster Preview


Eric G. Vajda*1, Diane J. Plotkin2, Douglas K. Logan3, Yong-Xi Li3, David G Orloff3, J. D. Pipkin4, Lin Zhi4 and Keith B. Marschke4
1Ligand Pharmaceuticals Inc, La Jolla, CA, 2Diane Plotkin Clinical Development Consultation Services, Poway, CA, 3Medpace, Inc., Cincinnati, OH, 4Ligand Pharmaceuticals Incorporated, La Jolla, CA

 

In T2DM, elevated levels of glucagon exacerbate hyperglycemia and its associated complications. Antagonism of glucagon action reduces blood glucose levels and hemoglobin A1c in T2DM patients. We have discovered a novel, selective, orally bioavailable glucagon receptor antagonist, LGD-6972, and investigated its pharmacokinetics (PK), safety, and pharmacodynamics (PD) in a first-in-human, ascending, single-dose, double blind, placebo‑controlled study in normal healthy (NH) and T2DM subjects (NCT01919684). A total of 56 participants were enrolled, 48 NH subjects and 8 T2DM subjects. The dose range was 2 to 480 mg for NH subjects. The T2DM subjects received a dose of 40 mg. LGD-6972 was well tolerated and considered safe up to the highest dose tested in NH and T2DM subjects. LGD-6972 was well-absorbed after oral administration; peak plasma concentrations were reached approximately 5 to 8 hours post dose with a long elimination half-life of about 50 hours, supporting once-daily dosing. The effects of LGD-6972 on fasting and post-prandial plasma glucose, glucagon, insulin, and glucagon-like peptide-1 (GLP-1) were evaluated in NH and T2DM subjects, and a meal tolerance test (MTT) and continuous glucose monitoring (CGM) were performed in T2DM subjects. In NH subjects, treatment with LGD-6972 resulted in small decreases in glucose levels approximating dose-dependency, and also lowered glucose levels in T2DM subjects. The decrease in glucose was most obvious in the fasting plasma glucose (FPG) levels 24 and 48 hours post-dose. The magnitude of decrease in FPG was greater in subjects with T2DM than in NH subjects. No glucose <60 mg/dL was observed in any group during the study. No consistent trend was revealed by CGM in the change in daytime glucose compared to baseline. However, a within-group trend towards lower nighttime glucose was observed, as well as a reduction in 24-hour glucose AUC. The mean glucagon and total GLP-1 levels were increased in NH subjects in a dose-dependent manner up to 240 mg, especially apparent between 24 to 96 hours post-dose. The glucagon and total GLP-1 levels after 480 mg were slightly below the expected levels, indicating that response to LGD-6972 may have reached a maximum. The mean glucagon and total GLP-1 levels were increased compared to placebo treatment in subjects with T2DM; most obvious 24 to 72 hours post-dose and greater than that in NH subjects. LGD-6972 treatment did not generate a consistent trend in the changes in the active GLP-1 or insulin levels across all dose groups after a single dose. The mean changes in glucose, glucagon, insulin, and GLP-1 levels from pre-meal did not generate marked consistent trends at 2 hours post-dose for most of the treatment groups, including subjects with T2DM. LGD-6972 is a promising agent for the treatment of T2DM, demonstrating PD effects after a single dose, and is currently in a multiple-ascending dose clinical trial (NCT02250222).

 

Disclosure: EGV: Employee, Ligand Pharmaceuticals. JDP: Employee, Ligand Pharmaceuticals Incorporated. LZ: Employee, Ligand Pharmaceuticals Incorporated. KBM: Employee, Ligand Pharmaceuticals Incorporated. Nothing to Disclose: DJP, DKL, YXL, DGO

PP01-1 21114 1.0000 THR-669 A Pharmacodynamic Effects of Single Doses of the Glucagon Receptor Antagonist LGD-6972 in Healthy Subjects and Subjects with Type 2 Diabetes Mellitus (T2DM) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 11:30:00 AM PP01 5938 11:15:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Poster Preview


Cecilia C. Low Wang*1, Rebecca Sanagorski2, R Matthew Hawkins1, Joanna Gibbs3, Kenneth Tompkins4, Mark Bridenstine5, Stacey Seggelke6 and Boris Draznin7
1University of Colorado Anschutz Medical Campus School of Medicine, Aurora, CO, 2Denver VA Medical Center, Denver, CO, 3Univ of Colorado AMC School of Medicine, Aurora, CO, 4Univ of Colorado School of Medicine, Aurora, CO, 5Univ Colorado AMC School of Medicine, Aurora, CO, 6Univ Colorado AMC School of Medicine, 7University of Colorado School of Medicine, Aurora, CO

 

Consistently effective strategies to transition hospitalized patients from intravenous (IV) insulin infusion to a subcutaneous (SC) insulin regimen are lacking. Problems that can occur include rebound hyperglycemia, recurrent diabetic ketoacidosis, and hypoglycemia. We conducted a randomized controlled trial of SC detemir insulin added to continuous IV insulin infusion vs continuous IV insulin infusion without added detemir to determine whether initiation of and concurrent therapy with a basal insulin would be an effective and safe strategy for transitioning patients with diabetes from IV to SC insulin. Methods: After approval of the study protocol by the Colorado Multiple Institutional Review Board, patients with diabetes on IV insulin infusion admitted to the University of Colorado Hospital or the Denver Veterans Affairs (VA) Medical Center who provided written, informed consent were enrolled and randomized to detemir intervention (DET) vs control (CON). Patients randomized to DET were started on detemir insulin 0.25 units/kg/day within 24 hours (hr) of starting the IV insulin infusion, and continued on this until discontinuation of the IV insulin infusion. Patients randomized to CON continued to receive usual care without any changes. For both groups, all decisions regarding titration of insulin infusion rates, timing of discontinuation of IV insulin, and transitioning to a SC insulin regimen were made by the primary team without intervention by the study team. Point-of-care blood glucose (BG) levels were checked as part of usual care: every 1-2 hours (hr) during IV insulin infusion, and every 4-6 hr during SC insulin therapy. Rebound hyperglycemia was defined as any BG over 180 mg/dL in the first 12 hr after discontinuation of IV insulin. Results: 30 patients were randomized (23 males, 7 females), 14 to CON and 16 to DET. Average age was 52.3±11.2 years and 54.1 ±16.2 years, and average weight was 91.0 ±22.1 kg and 91.4 ±20.6 kg in CON and DET, respectively. The incidence of rebound hyperglycemia was 100% in the CON group and 18.8% in the DET group. The overall average BG in the first 12 hr after discontinuation of IV insulin was 219.6 ±57.8 mg/dL in the CON group and 137.0 ±39.9 mg/dL in the DET group (p<0.0001 between groups). Average glycemia was significantly lower in the DET group than the CON group in each of the three 4-hour time periods after discontinuation of insulin infusion. Three patients manifested BG between 50-70 mg/dL, one at 0.5 hr and two at 6-7 hr after stopping IV infusion. Conclusions: The use of SC detemir insulin added to continuous IV insulin infusion was effective for reducing rebound hyperglycemia after discontinuation of IV insulin and during the transition to a SC insulin regimen. Larger studies are needed to confirm these findings and to determine optimal doses to avoid hypoglycemia, for patients with a variety of inpatient diagnoses requiring continuous IV insulin infusion.

 

Nothing to Disclose: CCL, RS, RMH, JG, KT, MB, SS, BD

PP01-2 21323 2.0000 THR-662 A Impact of Subcutaneous Detemir Insulin on Rebound Hyperglycemia in Hospitalized Patients with Diabetes Transitioning from a Continuous Intravenous Insulin Infusion 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 11:30:00 AM PP01 5938 11:15:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Poster Preview


Robert C Hood*1, Richard F Arakaki2, Carol H Wysham3, Ying Grace Li4, Julie A Settles5 and Jeffrey A Jackson5
1Endocrine Clinic of Southeast Texas, Beaumont, TX, 2University of Hawaii at Manoa, Honolulu, HI, 3Rockwood Center for Diabetes and Endocrinology, Spokane, WA, 4Eli Lilly and Company, Indianapolis, IN, 5Lilly USA, LLC, Indianapolis, IN

 

Use of recombinant human regular U-500 (U-500R) insulin has increased dramatically in recent years despite gaps in clinical knowledge regarding its initial dosing and optimal dose titration as well as efficacy and other clinical outcomes in severely insulin-resistant patients with inadequately controlled T2D. The unique prandial/basal actions of U-500R allow its use as insulin monotherapy. The current study is the first to compare efficacy and safety of 2 specific dosing regimens of U-500R replacing high-dose U-100 insulins in this target population.

The study was a 24-week, open-label, parallel-arm trial in 325 patients taking 201 to 600 units/day U-100 insulin therapy with or without oral antihyperglycemic agents. Patient demographics (means ± SD) included age, 55.4 ± 9.8 years; duration of diabetes, 15.2 ± 7.4 years; BMI, 41.9 ± 7.5 kg/m2; HbA1c, 8.7 ± 1.0%; U-100 insulin dose, 287.5 ± 80.5 units administered in 5 injections/day (median; range, 2 to 10). Patients were randomized to either thrice-daily (TID, n = 162) or twice-daily (BID, n = 163) U-500R after a 4-week lead-in period. The transition formula from U-100 insulins (predominantly basal–bolus analogue therapy) to U-500R reduced the total daily dose (TDD) by 20% for those with baseline HbA1c ≤8.0% or mean self-monitored plasma glucose (SMPG) <183 mg/dL. Otherwise, a 1:1 transition was used. Initial U-500R dosing proportions were 40:30:30 (breakfast:lunch:dinner) for TID and 60:40 (breakfast:dinner) for BID. Both algorithms adjusted TDD at each visit up to +30% (‑20% for hypoglycemia) to achieve pre-meal SMPG 71 to 130 mg/dL.

After 24 weeks, both treatments demonstrated significant and comparable reductions in HbA1c from baseline (TID, ‑1.12% [endpoint HbA1c, 7.53% ± 1.1%]; BID, ‑1.22% [endpoint HbA1c, 7.41% ± 1.0%]; P < 0.001 for both) and clinical equivalence (difference [BID vs. TID], ‑0.10% [P = 0.37]; 95% CI, ‑0.33% to 0.12%) at the non-inferiority margin 0.4%. Proportions of patients reaching HbA1c target values were similar between the 2 regimens (<8.0%: TID 70%, BID 69%; <7.5%: TID 55%, BID 47%; and <7.0%: TID 29%, BID 31%). Comparable increases in U-500R TDD (TID, 242.7 to 343.1 units; BID, 249.0 to 335.0 units) were observed. Severe hypoglycemia was uncommon and occurred with similar incidence (TID: 3 patients [1.9%]; BID: 6 patients [3.7%]). Incidence and rate of documented symptomatic hypoglycemia (≤70 mg/dL) were lower for TID compared with BID (= 0.003 and P = 0.02, respectively). Weight gain was similar between regimens (TID, 5.4 ± 0.4 kg; BID, 4.9 ± 0.4 kg).

In summary, initiation and titration of U-500R using either algorithm (TID or BID) improves glycemic control effectively and safely with fewer injections in patients with T2D on high-dose/high-volume U-100 insulin. These results provide clinicians with a practical framework for using U-500R in severely insulin-resistant patients with suboptimally controlled T2D.

 

Disclosure: RCH: Consultant, Eli Lilly & Company, Consultant, Novo Nordisk, Speaker Bureau Member, Eli Lilly & Company, Speaker Bureau Member, Astra Zeneca, Speaker Bureau Member, Jansen Pharmaceuticals, Speaker Bureau Member, Novo Nordisk, Speaker Bureau Member, Takeda, Speaker Bureau Member, Boehringer-Ingelheim, Speaker Bureau Member, GlaxoSmithKline, Medical Advisory Board Member, Takeda. RFA: Research Funding, Eli Lilly & Company, Research Funding, AbbVie, Research Funding, Merck & Co., Speaker Bureau Member, Astra Zeneca. CHW: Speaker, Jansen Pharmaceuticals, Consultant, Jansen Pharmaceuticals, Speaker, Lilly USA, LLC, Speaker, Novo Nordisk, Consultant, Sanofi, Speaker, Sanofi, Speaker, Astra Zeneca, Consultant, Astra Zeneca, Speaker, Boehringer Ingelheim Pharmaceuticals, Consultant, Boehringer Ingelheim Pharmaceuticals. YGL: Employee, Eli Lilly & Company. JAS: Employee, Lilly USA, LLC. JAJ: Employee, Lilly USA, LLC.

PP01-3 18681 3.0000 THR-658 A A Randomized Clinical Trial Comparing Efficacy and Safety of 2 Titration Algorithms for Human Regular U-500 Insulin in Severely Insulin-Resistant Patients with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 11:30:00 AM PP01 5938 11:15:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Poster Preview


Michael Grimes*1, Peter L Perreiah1, Laura Potoski1, Denise Plis2 and R Harsha Rao3
1VA Pittsburgh Healthcare System, Pittsburgh, PA, 2VA Healthcare Network, Wilkes-Barre, PA, 3Division of Endocrinology, VA Pittsburgh Healthcare System, Pittsburgh, PA

 

Background:  The parallel epidemics of obesity and diabetes are responsible for the rising prevalence of markedly insulin resistant diabetes (MIRD, total daily dose of insulin [TDD] ≥200 units), a potentially high-risk condition that poses a significant therapeutic challenge, with limited options for glycemic management.

Objective:  To determine the impact of U500 insulin on outcomes in MIRD.

Methods:  Retrospective chart reviews of 324 patients on TDD ≥200 units, identified from pharmacy records, who were either transitioned to U500 insulin therapy (U500 cohort, n=180) or maintained on U100 insulin therapy (U100 cohort, n=144) by patient/provider choice.

Outcome Measures: Mortality, microvascular and macrovascular complications between MIRD onset (verified from prescription signatures/progress notes) and date of last contact/death.

Results:  The number of patients with MIRD rose 10-fold between 2001 and 2013, while U500 insulin use rose 25-fold.  MIRD was associated with a high prevalence of diabetes-related vascular complications at onset, both macrovascular (259/324 [80%], principally CAD: 167/324 [51.5%]) and microvascular (199/324 [61.4%], principally nephropathy: 156/324 [48.2%]).  Patients transitioned to U500 insulin had a significantly lower mortality compared to those who remained on U100 insulin (27/180[15%] vs 39/144[27.1%], p=0.011), reflecting a 41% survival benefit on KM analysis (Hazard Ratio=0.59, p=0.0336) over similar follow-up duration (U500:1431±61days [Mean±SE], U100:1501±84), despite virtually identical clinical characteristics at MIRD onset.  Microvascular  complications were significantly reduced in the U500 cohort (105/180[58.3%] vs U100: 107/144[74.3%], p=0.003), specifically diabetic nephropathy (U500: 53/180[29.4%) vs U100: 79/144[54.9%], p<0.0001), both new onset (U500: 25/100[25%] vs U100: 33/68[48.5%], p=0.0016) and progressive (U500: 28/80[30%] vs U100: 33/68[48.5%], p=0.0014).  Macrovascular disease was impacted only marginally (U500: 53/180[29.4%] vs U100: 57/144[39.6%], p=0.056), principally from preventing peripheral arterial disease (PAD; U500:4/180[2.2%] vs U100:16/144[11.1%], p=0. 005), but not CAD (U500: 46/180[25.6%] vs U100: 41/144[28.5%],p=NS) or CVA (U500:6/180[3.3%] vs U100: 8/144[5.6%], p=NS).

Hemoglobin A1c in the U500 cohort decreased 0.91±0.1% (p<0.01), associated with an increase in TDD of 108±10units (p<0.01) and weight gain of 6.9±0.9kg (p<0.01), but A1c, weight and TDD did not change significantly in the U100 group. 

Conclusions:  MIRD is associated with high morality and frequent vascular complications, but substantial reductions in mortality and onset/progression of nephropathy and PAD can result from higher TDD and lower A1c associated with U500 therapy.  By contrast, worse outcomes may be attributable to “dose inertia” associated with U100 therapy in MIRD.

 

Nothing to Disclose: MG, PLP, LP, DP, RHR

PP01-4 21786 4.0000 THR-657 A The Burgeoning Problem of Markedly Insulin Resistant Diabetes Associated with Morbid Obesity: Impact of U500 Insulin Therapy on Outcomes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 11:30:00 AM PP01 5938 11:15:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Poster Preview


La Shondra Ellis*1, Robert Romano2, Jessica Costa-Guda2, Justin Bellizzi3 and Andrew Arnold2
1University of Connecticut Health Center, Farmington, CT, 2University of Connecticut School of Medicine, Farmington, CT, 3University of Connecticut School of Medicine

 

The molecular abnormalities leading to sporadic parathyroid adenomas are heterogeneous and incompletely understood. Previous investigations identified recurrent, clonally selected driver mutations in the cyclin D1 (PRAD1, CCND1) proto-oncogene and MEN1 tumor suppressor gene, and a combination of rare germline variants and somatic alterations in several cyclin-dependent kinase inhibitor genes. Subsequent identification of novel, recurrently altered genes through unbiased sequencing approaches has yielded only a few good candidates, all with low frequencies of observed mutations such as EZH2 (mutated in 1% of adenomas). Recently, compelling genetic evidence was reported implicating the ZFX gene, part of the Krueppel C2H2 type zinc finger protein family, as a likely parathyroid adenoma oncogene involved in almost 5% of cases (Oncoscience 2014; 1:360-366). Recurrent somatic mutations in ZFX were strikingly specific, focused on a hotspot of two consecutive highly conserved arginine residues in the most C-terminal zinc finger domain of the protein. ZFX is situated on the X-chromosome but escapes X-inactivation and is thus transcribed from both alleles in females; in males the highly homologous gene ZFY, on the Y-chromosome, is expressed and provides dosage compensation. Since analogous mutations in ZFY could have similar tumorigenic effects as those in ZFX, and with one such ZFY mutation already noted in a colon carcinoma (COSMIC), we analyzed ZFY in a series of typical sporadic parathyroid adenomas from male patients. Specifically, the 13th zinc finger domain, containing codons R781/R782 (corresponding to mutated ZFX codons R786/R787), was PCR-amplified and Sanger sequenced in 34 adenomas. No mutations were identified. Furthermore, the entire coding region of ZFY was fully sequenced in 20 of these tumors, again with no mutations identified. Therefore, it does not appear that R781/R782 missense mutations in ZFY commonly contribute to the molecular pathogenesis of parathyroid adenomas. However, analyses of more tumors will be necessary to exclude the possibility that ZFY may be involved in a similar percentage of adenomas as is the case for ZFX. The biology of ZFX, and perhaps ZFY, oncogenic mutations in the context of parathyroid tissue will certainly further illuminate the pathogenesis of these neoplasms.

 

Nothing to Disclose: LSE, RR, JC, JB, AA

PP08-1 19904 1.0000 THR-225 A Mutational Analysis of ZFY in Sporadic Parathyroid Adenomas 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 11:30:00 AM PP08 5965 11:15:00 AM Parathyroid and Bone Metabolism Poster Preview


Richard C. Lindsey*1, Charles M. Abreu1, Andy Cheng2, Sheila Pourteymoor1, Catrina Alarcon1 and Subburaman Mohan3
1Jerry L. Pettis VA Memorial Medical Center, Loma Linda, CA, 2Jerry L. Pettis VA Memorial Medical Center, Redlands, CA, 3VA Loma Linda Healthcare System, Loma Linda, CA

 

Recent studies involving mutant mouse models that are deficient in thyroid hormone (TH) action have provided unequivocal evidence that there is an important window in time prior to puberty when TH plays a critical role in the regulation of skeletal growth and development. TH regulated total body fat negatively during the prepubertal growth period, an observation which is consistent with the well-established effects of TH to accelerate resting energy expenditure. Because adipocytes and osteoblasts are both derived from bone marrow mesenchymal stromal (BMMS) cells, and because a relationship between increased bone marrow adiposity and decreased bone formation has been suggested, we examined whether TH deficiency during the postnatal growth period affects bone marrow adiposity in mice. We found bone marrow adiposity, measured by DEXA, was increased by >20% (P < 0.01) in TH deficient (Tshrhyt/hyt) mice at 3 weeks of age at multiple skeletal sites, and 10 day treatment with T3/T4 at a time when serum levels of T3/T4 normally increase (day 5-14) rescued this phenotype. To examine the mechanism for TH regulation of bone marrow adiposity, we tested the effects of TH treatment on mRNA expression levels of markers of adipocyte differentiation using primary cultures of mouse BMMS cells and the ST2 mouse stromal cell line in vitro. We found that treatment with 10 ng/mL T3 significantly increased expression levels of PGC-1α and UCP1, genes that have been established as markers of brown/beige fat. Real time RT-PCR measurements revealed 7.8-, 7.5-, and 2.5-fold increases in PGC-1α mRNA levels (all P < 0.05) at days 1, 4, and 6 after treatment with T3 in BMMS cells. UCP1 mRNA expression was increased by 3.5-fold (P = 0.01) after 6 days of treatment with T3.  By contrast, expression levels of the white adipocyte markers HOXC8 and HOXC9 were unaffected by 6 days of treatment with T3 in BMMS cells. Similarly, a 24 hour treatment with 10 ng/mL T3 increased expression of PGC-1α (6.0-fold, P < 0.01) but not HOXC8 or HOXC9 in ST2 cells. Based on our data, we conclude that 1) TH is an important regulator of bone marrow adiposity, and 2) the TH effect on energy metabolism may in part be mediated via increased production of thermogenic brown/beige fat.

 

Nothing to Disclose: RCL, CMA, AC, SP, CA, SM

PP08-2 20012 2.0000 THR-200 A Thyroid Hormone Regulation of Bone Marrow Adiposity in Mice 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 11:30:00 AM PP08 5965 11:15:00 AM Parathyroid and Bone Metabolism Poster Preview


Presley Garrison*1, Jeffrey Hanson2, Youn Hee Jee3, Ola Nilsson3, Jeffrey Baron4 and Julian Lui3
1NICHD, 2NCI, 3NICHD, Bethesda, MD, 4NIH, Bethesda, MD

 

Articular and growth plate cartilage each comprise three histologically distinct zones of chondrocytes. Articular cartilage is divided into superficial (SZ), mid (MZ), and deep (DZ) zones while growth plate cartilage is divided into resting (RZ), proliferative (PZ), and hypertrophic (HZ) zones. The molecular mechanisms that underlie the spatial regulation of chondrocyte differentiation are not well understood. To explore the role of BMP signaling, we used laser capture microdissection (LCM) and solution hybridization with barcoded probes (NanoString) (1) to quantify mRNA expression of 33 BMP-related genes in all zones of articular and growth plate cartilage of 7-day old mice (n=6). Previously identified zonal markers validated the accuracy of microdissection: Col10a1 mRNA was higher in HZ than in adjacent PZ (180-fold, P<0.01) and Prg4 was higher in SZ than MZ (24-fold, P<0.01). In articular cartilage, specific BMP agonists were primarily expressed in SZ: Bmp2 (3.6-fold greater than MZ) and Bmp6 (3.9-fold) (both P<0.01) whereas BMP functional antagonists were primarily expressed in MZ and DZ: gremlin (6.4-fold MZ vs. SZ), noggin (2.8-fold MZ vs. SZ), and Bmp3 (2.0-fold MZ vs. SZ, 2.7-fold DZ vs. MZ) (all P<0.01), suggesting an overall gradient in BMP signaling. However, antagonist Gdf10 was primarily expressed in SZ (3.8-fold greater than MZ, P<0.01). In growth plate cartilage, several BMP agonists were primarily expressed in HZ: Bmp1 (3.0-fold greater than PZ), Bmp2 (14-fold), and Bmp6 (55-fold) (all P<0.01). However, agonists Bmp5 showed greatest expression in RZ (2.1-fold greater than PZ) and Bmp7 showed greater expression in RZ and PZ (4.2-fold PZ vs. HZ) (all P<0.01). Some BMP antagonists were primarily expressed in RZ:  gremlin (3.7-fold greater than PZ) and Bmp3 (11-fold) (both P<0.01). However, functional antagonists Noggin and Smad7 showed greatest expression in HZ (2.4-, 3.3-fold, respectively, both P<0.01), and Gdf10 showed greatest expression in PZ (9.4-fold PZ vs. RZ, 6.5-fold PZ vs. HZ, both P<0.01). BMP receptors (Bmpr1a, Bmpr1b, and Bmpr2) and downstream effectors of the BMP system (Smad1, Smad4, and Smad5) did not show major expression differences between zones in articular or growth plate cartilage. In summary, we found that expression of multiple BMP agonists and functional antagonists showed striking spatial regulation in postnatal articular and growth plate cartilage. The observed expression patterns suggest that a BMP signaling gradient is present across postnatal articular cartilage such that greater BMP activity occurs in SZ. In the growth plate, some individual genes showed expression patterns analogous to that of articular cartilage, but the overall pattern of BMP agonist and antagonist expression showed greater complexity.

 

Nothing to Disclose: PG, JH, YHJ, ON, JB, JL

PP08-3 21008 3.0000 THR-217 A Spatial Regulation of Bone Morphogenetic Proteins (BMPs) in Postnatal Articular and Growth Plate Cartilage 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 11:30:00 AM PP08 5965 11:15:00 AM Parathyroid and Bone Metabolism Poster Preview


Blandine Poulet1, Andrew A Pitsillides2 and Abir Mukherjee*2
1University College London, United Kingdom, 2Royal Veterinary College, London, United Kingdom

 

Transforming growth factor β (TGFβ) ligands are important in regulating bone mass. Activin, a TGFβ ligand, plays crucial roles in bone development, turnover and mass and deletion of activin in mice leads to skeletal changes including delayed bone development. Control of the activin signalling pathway may, therefore, represent a novel target for limiting excessive bone loss during ageing and osteoporosis. Follistatin-like 3 (FSTL3) is an endogenous inhibitor of activin and related TGFβ ligands. FSTL3 gene deleted mice (FSTL3 KO) are viable and present multiple age-onset phenotypes that appear to be at least partially linked to activin action. We hypothesise that FSTL3 deletion in mice will generate an age-dependent modulation of bone architecture and mass. Cortical and metaphyseal bones from the tibiae of young (8wks), mature (30wks) and old (70wks) FSTL3 KO and WT control mice were analysed by micro-computerised tomography (microCT). Dissected tibiae were fixed in neutral buffered formalin and stored in 70% ethanol. Bones were scanned by microCT with an isotropic 5µm voxel size (µCT, Skyscan 1176). Each dataset was analysed using 3D algorithms in CTAn software (Skyscan), for cortical (0.5mm region at 50% tibial length) and trabecular (between 10 and 15% of total bone length from the proximal end) compartments. Cortical bone area / tissue area, bone perimeter, cortical medullary area and cross sectional thickness and also trabecular bone volume / total volume, trabecular thickness, separation and number, were measured. Our results show that tibiae were significantly longer in FSTL3-deficient mice compared to age-matched WT mice at 8wks but significantly shorter at 30 and 70 wks. Cortical bone parameters at 8wks showed that the tibiae of FSTL3 KO mice had increased bone area/tissue area, which was still evident at 30wks but not at 70wks. Cross sectional thickness was increased at 30wks and medullary area decreased at 30 and 70wks compared to WT controls. The trabecular compartment showed an increase in trabecular thickness at 8wks of age only and no other changes in other trabecular parameters were seen. Taken together our findings suggest that FSTL3 can control bone growth and architecture, particularly in relation to cortical bone. The mechanism by which lack of FSTL3 increases bone mass in young animals is being investigated.

 

Nothing to Disclose: BP, AAP, AM

PP08-4 22028 4.0000 THR-221 A Follistatin-like 3 Regulates Bone Structure in Mice 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 11:30:00 AM PP08 5965 11:15:00 AM Parathyroid and Bone Metabolism Poster Preview


W. Patricia Bandettini1, Alexander Karageorgiadis2, Ninet Sinaii3, Margaret Farmar Keil4, Charalampos Lyssikatos5, Marie Helene Schernthaner-Reiter6, Douglas R Rosing7, Vandana Sachdev7, J Aidan Carney8, Andrew E Arai9, Maya Beth Lodish*3 and Constantine A Stratakis6
1National Heart, Lung and Blood Institute, NIH, Bethesda, MD, 2NICHD, NIH, 3National Institutes of Health, Bethesda, MD, 4Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH)., Bethesda, MD, 5Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH), Bethesda, MD, 6National Institutes of Health (NIH), Bethesda, MD, 7National Heart, Lung and Blood Institute, NIH, 8Mayo Clinic, Rochester, MN, 9NHLBI

 

Carney Complex (CNC) is a well-described multiple neoplasia syndrome with a constellation of endocrine manifestations, pigmented skin lesions, and tumors of multiple organ systems, including endocrine, cardiac, cutaneous, and neural, associated with mutations in PRKAR1A (1).  The cardiac myxomas are of particular concern due to the potential for systemic embolization and risk of hemodynamically significant cardiac mechanical obstruction. Up to 15% of patients with CNC present with growth hormone (GH) excess due to GH-producing pituitary macroadenomas and antecedent pituitary hyperplasia is even more prevalent. Substantial evidence supports the role of the GH-IGF-1 axis in cancer incidence, and acromegaly is associated with an increased risk of colon cancer. The current study was performed to evaluate whether a relationship existed between GH excess and the development of cardiac myxomas in patients with CNC that we have followed for the last 20 years (1994-2014) at the National Institutes of Health (NIH) (n=99). These patients underwent serial cardiac imaging, including dynamic magnetic resonance imaging to screen for cardiac myxomas. We used cardiac screening data, and also data of prior myxoma resection, to identify those with a positive history of myxoma. The patients also underwent laboratory testing for GH excess. We found that 60% of the patients with GH excess (n=27/45) had a cardiac myxoma, compared to 37% of those who did not have GH excess (n=20/54, p=0.03). Additionally, patients with a history of GH excess were more likely to have a cardiac mass versus those without GH excess (OR=2.55, 95% CI:1.13–5.75, p=0.02). This is the first study describing the association between GH excess and an increased risk of cardiac myxomas in patients with CNC. The findings have important clinical implications because cardiac myxomas pose the most serious mortality risk in CNC. This association raises consideration of early surveillance for and treatment of GH excess in CNC patients.

 

Nothing to Disclose: WPB, AK, NS, MFK, CL, MHS, DRR, VS, JAC, AEA, MBL, CAS

PP02-1 19378 1.0000 THR-339 A Growth Hormone Excess Is Associated with an Increased Risk of Cardiac Myxomas in Carney Complex: A 20 Year Study (1994-2014) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 11:30:00 AM PP02 5977 11:15:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Poster Preview


Carolina R.C. Pieterman*1, Joanne M de Laat1, Jos W.R. Twisk2, Menno R Vriens1, Ad RMM Hermus3, Olaf M Dekkers4, Wouter W. de Herder5, Anouk N A Van der Horst-Schrivers6, Madeleine L. Drent2, Peter H Bisschop7, Bastiaan Havekes8 and Gerlof D Valk1
1University Medical Center Utrecht, Netherlands, 2VU University Medical Center, Netherlands, 3Radboud University Nijmegen Medical Centre, Netherlands, 4Leiden University Medical Center, Netherlands, 5Erasmus Medical Center, Rotterdam, Netherlands, 6University of Groningen, University Medical Center Groningen, Groningen, Netherlands, 7Academic Medical Center, Netherlands, 8Maastricht University Medical Centre, Netherlands

 

Pancreatic neuroendocrine tumors (pNET) are seen in approximately half of the patients with MEN1. PNETs are an important cause of MEN1-related morbidity and mortality. Up to thirty percent of the MEN1 patients with pNETs develop distant metastases, which is the most important prognostic factor for overall survival (1). Size of MEN1-related pNETs and tumor growth are reasons for surgical intervention to prevent metastasis (2). However, at present the natural course of pNETs in MEN1 is incompletely understood, hampering risk stratification and personalized care. In this present study we assess growth rate and determinants of growth in MEN1-related pNETs.

Patients with a pNET were selected from the MEN1 database of the DMSG including >90% of the Dutch MEN1 population >16 years with an average of 21 years of follow-up (n=323) (3). Patients were considered to have a pNET if found at pathology and/or diagnosed on the base of CT,MRI or EUS and confirmed at least once on subsequent imaging. For growth analysis CT/MRI was used and the size (mm) of the largest tumor in the pancreatic head and/or body-tail was followed over time. Size was analyzed from the first moment a tumor was identified until the end of follow-up, surgical removal or start of systemic anti-tumor therapy. Growth analysis was performed using Linear Mixed Models analysis to account for clustering of observations within patients. Gender, age, genotype, use of somatostatin analogues (SSA) and serum calcium were investigated as possible effect modifiers. To correlate tumor growth with disease related outcome, adverse outcome (pNET-related liver metastases or death) was analyzed for possible effect modification.

PNET was present in 159 (49%) patients. Ninety-two tumors from 82 patients were available for growth analysis. Thirthy-six patients (44%) were male. Median age at diagnosis of the tumor was 40 years. Baseline tumor size was 12 mm (range 3-82 mm). Twenty (22%) tumors were eventually surgically removed, baseline tumor size for these tumors was 17 mm (range 5-82). No clinically relevant growth was observed during a median follow-up of 4 (range 0.5-16.5) years:  the average growth rate of pNETs was <1 mm/year. Tumors that were surgically removed had a statistically significant higher growth rate, however this was still <1 mm/year. Growth rate did not differ by gender, age, genotype, use of SSA or serum calcium. Tumors from patients in whom during follow-up an adverse outcome was documented had a statistically significant somewhat higher growth rate, but still <1 mm/year.

In this large patient group during long term follow-up, the growth rate of pNETs in MEN1 appears to be slower than previously thought and was not influenced by gender, age, genotype, use of SSA and serum calcium. Present results do not support a clinically relevant relation between growth rate and adverse outcome indicating that tumor growth rate cannot be used as indicator for surgery.

 

Nothing to Disclose: CRCP, JMD, JWRT, MRV, ARH, OMD, WWD, ANAV, MLD, PHB, BH, GDV

PP02-3 20880 2.0000 THR-325 A Growth Rate of MEN1-Related Pancreatic Neuroendocrine Tumors: New Insights Based on the Results from the DutchMEN1 Study Group (DMSG) National MEN1 Database 1990-2010 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 11:30:00 AM PP02 5977 11:15:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Poster Preview


Paul J Newey*1 and Rajesh V Thakker2
1Ninewells Hospital & Medical School, University of Dundee, Dundee, Scotland, 2University of Oxford, Oxford, United Kingdom

 

Recent large-scale DNA sequencing projects have provided unprecedented insight into the frequency and spectrum of rare coding variants present within populations. Indeed, the majority of protein-coding variation is rare, evolutionarily recent, and enriched for deleterious alleles1. The contribution of such rare variation to disease susceptibility represents one of the major challenges of contemporary medicine. However, the high frequency of rare coding variation within the population has important implications for the rational interpretation of clinical genetic studies; a failure to consider the inherent frequency of rare variation within control populations may result in a reporting bias when evaluating germline variants in disease-associated genes. To address these potential biases, analysis of sequence data from large control populations may allow the estimation of rare variant frequencies. In this study, 27 genes associated with endocrine disease (e.g. MEN1, AIP, RET, VHL) were evaluated for the frequency of rare (minor allele frequency (MAF): <0.5%), very rare (MAF: <0.01%) and singleton (i.e. occurring only once) germline non-synonymous variants in a population of ~4300 European-Americans using publicly available data from the NHLBI GO Exome Sequencing Project (evs.gs.washington.edu/). Analysis of these genes demonstrated significant variability in the frequency of rare coding variants. As expected, a positive correlation was observed between the number of rare variants in a given gene and protein length. For example, high rare variant frequencies were observed in NF1 (1 in 44 individuals) and RET (1 in 56 individuals), consistent with their large size (i.e. >1000 amino acids). However, even when corrected for amino-acid length, a ~10-fold difference was observed between genes with the lowest (i.e. CDC73) and highest (i.e. CDKN1A) singleton variant frequency rate, whilst genes with no rare coding region variants were also identified (e.g. AP2S1). On average, a singleton missense variant was observed once every ~180,000 amino acids (5.4x105 nucleotides sequenced). Thus, for a protein of 500 amino acids, a singleton variant would be anticipated ~1:360 individuals. To model the situation in which individuals undergo multiple gene testing, collective rare variant frequencies were calculated. For example, using a panel of 11 genes for phaeochromocytoma, a rare variant would be anticipated once in every 8 individuals sequenced, whilst a singleton variant once in every 30 individuals. This analysis confirms the unexpectedly high rate of rare coding-region variation in control populations, and highlights the need to consider this background frequency when undertaking genetic studies. In the absence of robust methods for predicting variant pathogenicity, these studies emphasize the clinical challenges faced as next generation sequencing move into the clinical arena.

 

Nothing to Disclose: PJN, RVT

PP02-4 19775 3.0000 THR-329 A Frequency and Spectrum of Rare Germline Coding Variants in Hereditary Endocrine Disease Genes: Implications for Clinical Genetic Studies 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 11:30:00 AM PP02 5977 11:15:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Poster Preview


Naoki Hattori*1, Takashi Ishihara2 and Akira Shimatsu3
1Ritsumeikan University, Kusatsu-City, Shiga, Japan, 2Kobe City General Hospital, Ashiya-Shi, Japan, 3National Hospital Organization Kyoto Medical Center, Kyoto, Japan

 

Objective TSH is a sensitive indicator of thyroid function. In subclinical hypothyroidism, however, serum TSH concentrations are elevated despite normal thyroid hormone levels. Macro TSH is a large molecular sized TSH on gel filtration chromatography. We hypothesized that the bioactivity of macro TSH might be low like macro PRL and that macro TSH may be accumulated in the circulation. Thus, patients with macro TSH may show elevated TSH levels despite normal thyroid hormone levels, similar to subclinical hypothyroidism. This study aimed to characterize the nature of macro TSH.

Patients We screened macro TSH in 1083 patients with subclinical hypothyroidism (634 women and 449 men, aged 64.6 ± 17.7 years) followed at our hospital, and in 54 serum samples that were sent to our laboratory from doctors in other hospitals, suspecting the presence of macro TSH.

Measurements Macro TSH was screened by polyethylene glycol (PEG) method, in which macro TSH was precipitated with 12.5% PEG. The PEG-precipitable TSH (%), which may represent the amount of macro TSH, was calculated as (total TSH - free TSH) / total TSH × 100. Serum samples that exhibited PEG-precipitable TSH ratios greater than 75% (mean + 1.5 SD in controls) were subjected to gel filtration chromatography to confirm macro TSH. The biological activity of macro TSH was examined in the FRTL-5 rat thyroid cell line. Protein G column and HAMA (human anti-mouse antibodies) blockers were used to characterize macro TSH. Macro PRL was examined in patients with macro TSH using the same method as macro TSH.

Results Among 1083 patients with subclinical hypothyroidism, 14 patients had macro TSH (1.3%). Among 54 serum samples that were requested to examine the presence of macro TSH, 5 serum samples were found to containe macro TSH. The nature of macro TSH in these 19 serum samples included 14 anti-TSH autoantibodies of IgG class, 2 non-IgG-associated, and 3 HAMA. Macro TSH showed low bioactivity compared to the immunoreactivity. Three patients with macro TSH also had macro PRL.

Conclusions Macro TSH was heterogeneous, but it is mostly comprised of TSH and anti-TSH autoantibodies. Because 3 of 19 patients with macro TSH also had macro PRL, common mechanisms may be involved in the generation of macro TSH and macro PRL in some patients. When PEG-precipitable TSH exceeds 90% in serum samples with TSH above 10 mU/l, clinicians should strongly suspect the presence of macro TSH and confirm it by gel chromatography. Because macro TSH exhibited low bioactivity, thyroid hormone replacement therapy may not be required in patients with subclinical hypothyroidism due to macro TSH except for those with high serum free TSH levels.

 

Nothing to Disclose: NH, TI, AS

PP11-1 19723 1.0000 THR-012 A Etiology and Bioactivity of Macro TSH 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 11:30:00 AM PP11 6034 11:15:00 AM Thyroid Development, Clinical and Autoimmunity Poster Preview


Thomas Daniel Milner*1, Alexander Lloyd Berry2, Suchitra Raj,3, Ralf Clauss3, John Wright3 and Sue Davidson3
1Royal Surrey County Hospital, Hull, United Kingdom, 2Hull York Medical School, Hull, United Kingdom, 3Royal Surrey County Hospital, Guildford, United Kingdom

 

Background: Establishing an appropriate radioiodine dose in the treatment of hyperthyroidism caused by Grave’s disease, multinodular goitre (MNG) or solitary toxic nodule (STN) requires a balance between effective cure and risk of complications. Several clinical and patient factors are also thought to affect radioiodine outcomes, in particular the use of antithyroid drugs (ATDs) at the time of treatment. This study aims to identify factors associated with optimal radioiodine outcomes

 

Methodology: The introduction of a higher radioiodine dose into the Royal Surrey County Hospital in 2008 generated two treatment cohorts for hyperthyroid patients: a low dose cohort (n=62) receiving 350MBq, and a high dose cohort (n=102) where Grave’s disease patients received 415MBq and MNG or STN patients received 475MBq. A retrospective cohort study was conducted analysing the effect of this dose increase, as well as various other factors, on proportion hyperthyroid and proportion hypothyroid at one year, and time to absence of hyperthyroidism.

 

Results: At one year post-radioiodine, 32 (19.5%) patients were hyperthyroid, 37 (22.6%) patients were euthyroid, and 95 (57.9%) were hypothyroid. The higher radioiodine dose had no impact on all treatment outcomes. The use of ATDs post-radioiodine (OR=2.346; 95%CI=1.071-5.138; p=0.033), and a high free thyroxine (fT4) in the year prior to treatment (OR=8.499; 95%CI=1.231-58.707; p=0.03) were associated with increased risk of hyperthyroidism at one year. ATDs post-radioiodine (adj. HR=0.445; 95%CI=0.308-0.643; p<0.0005) and high fT4 (HR=0.627; 95%CI=0.420-0.935; p=0.022) also significantly prolonging time to absence of hyperthyroidism, with the effect of ATDs being maintained on multivariate analysis.

 

Conclusions: As higher dose radioiodine therapy did not alter treatment outcomes, contrary to current guidelines, the use of lower dose (350MBq) therapy in hyperthyroidism would be most appropriate, to mitigate any risks of long-term complications. In addition, provided a patient’s clinical status permits, the use of carbimazole or PTU post-radioiodine should be avoided due to their effects on radioiodine uptake.

 

Nothing to Disclose: TDM, ALB, SR, RC, JW, SD

PP11-2 21327 2.0000 THR-023 A Optimal Outcomes with the Use of Low Dose Radioiodine Therapy for Hyperthyroidism and the Avoidance of Antithyroid Medications Post-Treatment 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 11:30:00 AM PP11 6034 11:15:00 AM Thyroid Development, Clinical and Autoimmunity Poster Preview


David Martin Carty*, Paul Welsh, Elaine Butler and Christian Delles
University of Glasgow, Glasgow, United Kingdom

 

Treatment of overt hypothyroidism (OH) in pregnancy leads to improved foetal and maternal outcomes. Recent guidelines from US and European thyroid associations also indicate that women with subclinical hypothyroidism (SCH, TSH above the trimester-specific reference range, or ≥2.5 mU/L) should be treated with thyroxine. In these women, however, there is limited evidence of improvement in outcomes. In this study we measured thyroid function in early pregnancy to examine the frequency of OH and SCH in a general obstetric population, and to explore its relationship with obstetric outcomes.

4300 women were recruited from obstetric clinics in Glasgow, UK between 2007-2010 as part of the “Proteomics in Pre-eclampsia” study. Plasma samples and clinical data were obtained at the initial antenatal hospital visit at gestational week 12-14. Data on pregnancy outcomes was obtained from hospital databases. Birthweight centiles, corrected for maternal age, ethnicity, baby sex and gestational age were used. Thyroid stimulating hormone (TSH) and free thyroxine (fT4) levels were analysed in 2014 using clinically validated platforms (Roche, UK). In women with abnormal thyroid tests in pregnancy, hospital databases were used to identify more recent thyroid problems.

Delivery information was available on 4179 (97.2%) women: 3889 (90.4%) were Caucasian, 36 miscarried, and 87 (2%) developed pre-eclampsia. 73 women were known to have pre-existing thyroid disease and were not included in the analysis; of them 28 (38%) had a TSH above 2.5mU/L.

58 women were identified as having OH (TSH>5 mU/l), of whom 2 were later diagnosed during pregnancy and 7 have been diagnosed since pregnancy. 8 women were found to have isolated hypothyroxinaemia (fT4 <9 pmol/l) . 406 women (10.6%) were identified as having SCH (TSH >2.5 mU/L but <5mU/L, and fT4 within the non-pregnant reference range). Women with OH delivered babies of lower birthweight than those with normal thyroid function (3196g ± 805 vs 3403g ± 402, p=0.02, corrected centile 37±12 vs 44±14, p=0.05), there was no difference in birthweight between women with SCH and those with normal thyroid function. There was no relationship between early pregnancy thyroid tests and rates of pre-eclampsia, gestational diabetes, miscarriage, preterm delivery or Caesarean section.

Retrospective screening of thyroid function in this unselected obstetric cohort identified 1.4% of women having OH, and these women delivered infants of lower birthweight. In women known to have thyroid disease, more than a third had a TSH above the pregnancy reference range. 10% were identified as having SCH, which did not impact upon obstetric outcomes. Recent guidelines suggest that if identified, women with SCH should be treated with thyroxine. This treatment, and the subsequent monitoring required would have major implications for design and costing of obstetric services.

 

Nothing to Disclose: DMC, PW, EB, CD

PP11-3 20383 3.0000 THR-006 A Frequency of Overt and Subclinical Hypothyroidism in Pregnancy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 11:30:00 AM PP11 6034 11:15:00 AM Thyroid Development, Clinical and Autoimmunity Poster Preview


Stephanie Clare Cox*1, John V Conaglen2 and Marianne Susan Elston3
1Waikato District Health Board, Hamilton, New Zealand, 2Waikato Clinical School, University of Auckland, Hamilton, New Zealand, 3Waikato Hospital, Hamilton, New Zealand

 

Context

There have been advances in our understanding of Graves’ disease over the last two decades, particularly with regard to the risks of anti-thyroid drugs and increased concern over the long term effects of radiation exposure, as well as advances in surgical techniques. Research into the effects of the disease and it’s treatments during pregnancy have also led to shifts in practice over this time. Multiple surveys were undertaken in the late 1980’s and early 1990’s looking at clinical practice patterns amongst physicians in the US, Europe and Australasia and recently several of these surveys have been repeated to assess the shifts in practice over time.

Objective

The aim of this study was to assess the current management of Graves’ disease by New Zealand (NZ) endocrinology/internal medicine specialists and to compare findings with those of contemporary international studies and a previous 1991 New Zealand survey1.

Methods

We conducted an online case-based survey of all NZ physicians practising in the area of internal medicine, diabetes and/or endocrinology. Physicians were identified from the NZ Medical Council register. The case studies and questions from the previous 1991 NZ study were reformatted into an online survey and the link was emailed to the list of physicians.  A total of 117 physicians were invited to participate. Of these six were unable to be located, two had retired, two were on extended leave throughout the survey time period and one had permanently left NZ. Respondents who treated <2 patients/year with Graves’ disease were asked only to complete the sections on demographics and access to services. 

Results

In comparison to the 1991 NZ survey1 the first-line use of Radioiodine was less, at only 3%, compared with 41%, using the same clinical scenario. This corresponded with an increase in the use of antithyroid drugs, while the rates of surgery as a first line treatment remained fairly static over time. This is more in line with the current practice of our European colleagues than with Endocrinologists in North America who continue to report higher rates of RAI use2. In the index case, all respondents opted for carbimazole as their drug of choice (methimazole is not available in NZ). When the case was altered to a woman desiring pregnancy 28% would instead use PTU pre-pregnancy, while 56% would switch to PTU once the patient became pregnant. Both of these figures suggest NZ physicians use PTU less in pregnancy compared with international practice. Access to services, particularly nuclear medicine imaging, was an issue for 23% of respondents.

Conclusions

Practice in NZ has changed over the last twenty years, with a move away from radioactive iodine treatment. This parallels changes in practice in Europe and to a lesser extent America. NZ physicians appear to use PTU less commonly pre-pregnancy, although the majority would switch to PTU in early pregnancy.

 

Nothing to Disclose: SCC, JVC, MSE

PP11-4 21266 4.0000 THR-007 A The Management of Graves’ Disease in New Zealand 2014 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 11:30:00 AM PP11 6034 11:15:00 AM Thyroid Development, Clinical and Autoimmunity Poster Preview


Ruchi Mathur*1, Manpreet Mundi2, Kathleen Shari Chua1, Paul Lorentz2, Eugenia Lin1, Gillian M Barlow1, Miguel Burch1, Scott Cunneen1, Adrienne Youdim1 and Mark Pimentel1
1Cedars-Sinai Medical Center, Los Angeles, CA, 2Mayo Clinic, Rochester, MN

 

Background

Gut microbes have been shown to play significant roles in metabolic dysfunction and the development of obesity. Of these, the methanogenic archaea have been specifically linked to altered metabolism and weight gain. We have previously shown that the presence of both methane and hydrogen on breath test (a surrogate for gastrointestinal colonization with methanogens) is associated with greater body mass index (BMI), percent body fat, as well as glucose intolerance. Since alterations in gut microbial populations are also associated with weight loss following gastric bypass surgery, we investigated whether methane and hydrogen on breath test was associated with differential weight loss after Roux- en-Y gastric bypass.

Methods

Obese subjects aged 18 to 70 years with a pre-surgery BMI ≥ 35 were recruited at ≥ 4 months after bariatric surgery (Roux-en-Y or gastric sleeve). Pre- and post-surgery weights and BMIs were recorded, and single sample breath tests were performed to determine post-surgery methane and hydrogen levels. BMI and body weight changes were normalized to 6 months post surgery.

Results

A total of 156 subjects were recruited (112 females, 44 males). The average age was 47.9 ± 0.9 years and the average pre-surgery BMI was 42.6 ± 0.3. For all subjects, % change in weight and % change in BMI were normalized for pre-surgery BMI. Using cutoffs of methane ≥3 ppm and hydrogen ≥20 ppm to define methane and hydrogen positivity (M+/H+), M+/H+ subjects (n=13) had a lower % change in BMI than all other subjects (n=144) following surgery (20.6±1.9 vs. 23.5±0.6 respectively, p=0.13), and also had a significantly lower % change in weight (20.05±1.8 vs. 23.9±0.5 respectively, p=0.036). When cutoffs of methane ≥3 ppm and hydrogen ≥10 ppm were used to define methane and hydrogen positivity, M+/H+ subjects (n=18) still exhibited a lower prorated % change in BMI following surgery than all other subjects (n=139) (20.4±1.6 vs. 23.6±0.6 respectively, p=0.057), as well as a significantly lower prorated % change in weight than (20.1±1.5 vs. 24.0±0.5 respectively, p=0.014).

Conclusions

Subjects with gastrointestinal methanogen colonization (as determined by methane and hydrogen levels on breath test) exhibited less % weight loss and % BMI reduction than subjects without detectable methanogen colonization following Roux-en-Y gastric bypass surgery. This was shown regardless of whether a breath hydrogen level of ≥20 ppm or the more stringent level of ≥10 ppm was used to define hydrogen positivity.  Altering methanogenic colonization via pharmacotherapy and/or dietary interventions may provide additional benefit in those achieving less than optimal weight loss post gastric bypass surgery.

 

Nothing to Disclose: RM, MM, KSC, PL, EL, GMB, MB, SC, AY, MP

PP07-1 20644 1.0000 THR-550 A Intestinal Methane Production Is Associated with Decreased Weight Loss Following Bariatric Surgery 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 11:30:00 AM PP07 6045 11:15:00 AM Obesity: Clinical Studies Poster Preview


Jean-Marc Schwarz*1, Susan M Noworolski2, Michael J Wen3, Grace Marie Jones1, Ewan Sinclair1, Artem Dyachenco1, Viva Tai3, Moises Velasco Alin1, Ayca Erkin-Cakmak3, Alejandro Gugliucci1, Kathleen Mulligan3 and Robert H Lustig4
1Touro University California, Vallejo, 2University of California San Francisco, San Francisco, 3University of California San Francisco, 4University of California - San Francisco, San Francisco, CA

 

Background: Previous studies have shown that high sugar (specifically fructose) consumption is associated with liver fat accumulation and/or hypertriglyceridemia, which may increase risk for Type 2 diabetes and cardiovascular disease. The conversion of sugar to fat in the liver (hepatic de novolipogenesis [DNL]) may be an important pathogenic mechanism.

Objective: To determine the effect of 10 days of a fructose- but not calorie- restricted diet on DNL in obese Latino and African American children with high habitual dietary sugar intake.

Design/Methods: Latino (8F, 9M) and African American (4F, 1M) children (ages 9-18; BMI z-score 2.4 ± 0.1), who were high dietary sugar consumers at baseline (average fructose intake >50 g/day), had all meals provided for 10 days with the same caloric and macronutrient composition as their standard diet, but with other carbohydrate substituted for sugar. Subjects were weighed daily and diets adjusted to maintain baseline weight. Fractional DNL and DNL area under the curve (DNL-AUC) were measured over 8 hours of test meal feeding on Day 0 (high fructose) and Day 10 (low fructose). Test meals contained 1-13C sodium acetate tracer.  Post-prandial blood samples were analyzed by gas chromatography/mass spectrometry, and DNL calculated by mass isotopomer distribution analysis. Liver fat percentage was determined by magnetic resonance spectroscopy.

Results:DNL during feeding was significantly reduced with fructose restriction, beginning 50 minutes after initiation of tracer/feeding (2.5±0.3 vs.1.6±0.1% on days 0 and 10 respectively, P<0.003 by paired t-test) and continuing throughout the tracer/feeding procedure (13.3±1.2 vs. 5.8±0.6% at 8 hours on days 0 and 10, P<0.001).  Integrated DNL-AUC decreased by 58.7% from 53.5±6.4 on day 0 to 22.1±3.3 on day 10 (P<0.001). The decrease in DNL-AUC over only 10 days of fructose restriction was accompanied by a 29.5% reduction in liver fat from 11.9±2.3% on day 0 to 9.5±2.2% on day 10 (P <0.001; n=20). These effects remained statistically significant after adjusting by ANCOVA for minor weight loss over the 10 days (1.0±0.3 kg, P<0.001).

Conclusions: Isocaloric dietary fructose restriction for 10 days decreased hepatic DNL and liver fat in Latino and African American children irrespective of weight loss. These results suggest that hepatic DNL is an important mechanism leading to liver fat accumulation in children, which can be reversed by short-term fructose restriction. These data support public health efforts to reduce sugar consumption.

 

Nothing to Disclose: JMS, SMN, MJW, GMJ, ES, AD, VT, MV, AE, AG, KM, RHL

PP07-3 19571 3.0000 THR-549 A Isocaloric Fructose Restriction for 10 Days Reduces Hepatic De Novo Lipogenesis and Liver Fat in Obese Latino and African American Children 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 11:30:00 AM PP07 6045 11:15:00 AM Obesity: Clinical Studies Poster Preview


Ajay Chawla*
University of California San Francisco, San Francisco, CA

 

 

Nothing to Disclose: AC

AL10-1 22249 1.0000 A The Intersection of Inflammation and Metabolism in Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 11:45:00 AM AL/OR10 6078 11:15:00 AM Richard E. Weitzman Outstanding Early Career Investigator Award Lecture


Florian Le Billan*, Junaid Ali Khan, Khadija Lamribet, Say Viengchareun, Jérôme Fagart and Marc Lombes
INSERM U693, Le Kremlin Bicêtre, France

 

Aldosterone exerts numerous pleiotropic functions, notably in the kidney – where it regulates fluid and electrolyte homeostasis – but also in the cardiovascular and central nervous systems. Aldosterone acts mainly by activating the mineralocorticoid receptor (MR), a transcription factor that interacts with multiple transcription coregulators to regulate target gene expression. Dysfunctions in the mineralocorticoid pathway are involved in various pathological disorders such as hypertension, cardiovascular, renal or metabolic diseases.

Aldosterone mechanism of action is extremely complex, involving several intricate aspects (1) whereby kinetic and cross-talk dynamics appeared to play a critical role in the control of vectorial ionic transport. In this context, an overall assessment of MR genomic targets by new genomic tools was highly desirable. Here, we setup the chromatin immunoprecipitation (ChIP) with a specific anti-MR antibody in a highly differentiated human renal cell line expressing GFP-MR treated or not with aldosterone. This approach coupled with the innovative high-throughput sequencing technology (ChiP seq, HiSeq) allowed identification of hundreds of direct genomic MR targets (MACS Software), including the well known SCNN1A gene coding for the alpha subunit of the epithelial sodium channel (αENaC). Computational analysis of these genomic sites defined a specific MR response element (MRE) in which the AGtACAgxatGTtCt sequence was the most prevalent motif. Of interest, genomic MR binding sites were located at various distances (up to 150 kb) and directions from the transcriptional start sites of target genes. Specific aldosterone-induced recruitment of MR on the first most relevant genomic sequences was further validated by ChIP-qPCR and correlated with concomitant and positive aldosterone-activated transcriptional regulation of the corresponding gene as assayed by RT-qPCR. Analyses of the dynamics of MR recruitment and of its transcriptional co-regulators together with the evaluation of the functional role of these genes in the renal mineralocorticoid signaling pathway are underway.

This work provides new insights into aldosterone, MR-mediated signaling and opens germane perspectives for mineralocorticoid-related pathophysiology.

 

Nothing to Disclose: FL, JAK, KL, SV, JF, ML

OR06-1 19789 1.0000 A Identification of Genomic Targets of the Mineralocorticoid Receptor and Transcriptional Regulation of Aldosterone-Regulated Genes in Human Renal Cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 1:00:00 PM OR06 5902 11:30:00 AM HPA Axis and Adrenal: Receptors To Clinical Impact Oral


Tracy Choi Sze Mak*1, Dawn EW Livingstone2, Brian R Walker1 and Ruth Andrew1
1University of Edinburgh, Edinburgh, United Kingdom, 2University of Edinburgh, Edinburgh Scotland, United Kingdom

 

Background

Glucocorticoids induce insulin resistance in multiple tissues. Systemic glucocorticoid receptor (GR) antagonism improves insulin and glucose homeostasis in ob/ob mice but the site of action is uncertain, with effects confounded by compensatory hypercorticosteronemia. A-348441, a GR antagonist targeted to the liver by conjugation to a bile acid, is used here for the first time to evaluate the role of hepatic GR in the metabolic complications of short-term diet-induced obesity in mice.

Experimental Design

C57BL6/J male and female mice (n = 6-12 per group; age 12 weeks) were given control, high fat (HF, 58% fat, 26% sucrose) or HF with A-348441 (~100mg/kg/day, kindly gifted by KaroBio AB) diet for 4 weeks. Glucose tolerance tests (ip GTT; 2mg/g body weight) were performed after 3 weeks. After 4 weeks, mice were culled (8am). Plasma insulin, non-esterified fatty acids (NEFA), and corticosterone were analyzed using ELISA, and plasma glucose and liver triglycerides (TG) spectrophotometrically. Data are control; HF; HF+A348441, mean±SEM *p<0.05, **p<0.01 vs control; #p<0.05, ##p<0.01 vs HF.

Results

Plasma corticosterone concentrations were not changed in either gender by HF diet or A-348441.

In male mice, A-348441 prevented HF-induced bodyweight gain (31.8±0.5; 34.3±0.5*; 31.1±0.8g#), expansion of the liver (1.47±0.05; 1.66±0.05*; 1.40±0.04g##) and total adipose depot weight gain (1.26±0.07; 2.46±0.14**; 1.58±0.14g##). A-348441 also attenuated HF-induced elevations in fasting plasma insulin (0.83±0.12; 2.06±0.20**; 0.99±0.17ng/mL##), fasting glucose (132.8±7.5; 166.1±8.2**; 125.1±7.0mg/dL##), and insulin response to GTT (91.7±8.2; 261.3±16.3**; 145.2±20.2ng/mL.min##). Neither HF diet nor A-348441 influenced liver triglycerides, fasting plasma NEFA or insulin-mediated NEFA suppression in male mice.

In female mice, HF diet did not significantly increase body or tissue weights, fasting plasma insulin, or post-GTT plasma insulin; A-348441 also had no effect. HF diet did increase fasting and post-GTT plasma glucose, but the only effect of A-348441 was a trend for reduced fasting glucose (113.6±7.5; 164.9±13.5*; 129.6±10.5 mg/dL). HF diet also increased liver TG and decreased NEFA suppression in female mice, but A-348441 did not influence these parameters.

Conclusion

Liver-specific GR antagonism not only improves insulin sensitivity and glucose tolerance in male mice with short-term diet-induced obesity, but also attenuates weight gain. Further dynamic studies are required to assess effects on energy balance and/or liver triglyceride export and its uptake in peripheral adipose. Females were relatively protected from metabolic dysfunction with diet-induced obesity, and GR antagonism had no discernible effect. These results suggest targeting hepatic GR may have a beneficial role in metabolic homeostasis in diet-induced obesity.

 

Nothing to Disclose: TCSM, DEL, BRW, RA

OR06-2 19257 2.0000 A Liver-Selective Glucocorticoid Receptor Antagonism Improves Insulin Sensitivity and Prevents Adipose Expansion in Diet-Induced Obesity 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 1:00:00 PM OR06 5902 11:30:00 AM HPA Axis and Adrenal: Receptors To Clinical Impact Oral


Kevin C.J. Yuen*1, Frank Buttgereit2, Dorothy McCabe3, Sheela Kolluri4, Brinda Tammara4, Riccardo Rojo4 and Juliana Hendrika Hey-Hadavi5
1Swedish Neuroscience Institute, Seattle, WA, 2Charite University hospital, Berlin, Germany, 3Pfizer Inc, New York, 4Pfizer Inc, 5Pfizer Inc, New York, NY

 

Background

PF-04171327 is a non-steroidal ‘dissociated” or partial agonist of the glucocorticoid receptor (DAGR) currently under investigation for anti-inflammatory efficacy, while mitigating unwanted side-effects of glucocorticoids(GCs). Administration of GCs and DAGR is expected to suppress the HPA axis, causing feedback inhibition of cortisol and ACTH secretion. In an 8-week Phase 2b dose-finding study to evaluate the efficacy and safety of DAGR in treating patients with rheumatoid arthritis (RA), 4 doses of DAGR were compared to 2 doses of prednisone (Pred) or placebo (PBO).  The effect of DAGR treatment on HPA axis suppression was evaluated with serial morning cortisol measurements and an ACTH stimulation test. 

Methods

323 adults with active RA therapy were randomized to receive DAGR 1, 5, 10 or 15 mg, Pred 5 or 10 mg, or PBO daily for 8 weeks followed by a 4-week taper. Any use of GCs within 6 weeks of screening was prohibited.  Fasting cortisol levels were measured at each 2-weekly study visit for 8 weeks, and a 250 mcg ACTH stimulation test was performed at Week 13.

Results

Across the dose groups, mean age ranged from 50.4 to 57.4 yrs; proportion of females ranged from 73% to 89.1%; and proportion of Caucasians ranged from 82.6% to 95.6%. Based on aggregate efficacy data, DAGR 10 and 15 mg was found to be efficacious (superior to PBO and comparable to Pred 10 mg, with 20% improvement using the American College of Rheumatology core set measures, and 28-point disease assessment scale, at Week 8). Median suppression of plasma cortisol levels were observed as early as Week 2 with  DAGR 1, 5, 10 and 15 mg doses showing suppression from baseline of  101, 122,  96.4,  110.5 ng/ml by  -14.3%, -79.6%,  -93.7%, -95.3%  respectively; suppression for Pred 5 mg and 10 mg from baseline of 113 and 123 ng/ml by -25.1% and  -42.7%  respectively; and that for PBO  from baseline of 106 ng/ml by  -6.4%. This magnitude of suppression was sustained throughout the 8 weeks of active treatment.  Prompt HPA axis recovery was observed during the 4-week DAGR taper, as morning cortisol levels normalized and a normal response to the ACTH stimulation test was evident in >95% of the patients.  All DAGR doses were well-tolerated, with no clinical manifestations suggestive of adrenal suppression reported.

Conclusions

Profound suppression of plasma cortisol was observed with DAGR 5, 10, and 15 mg, while partial suppression was seen with Pred 5 and 10 mg.  No subjects reported clinical symptoms of adrenal insufficiency at any time.  HPA axis recovery in the taper period was demonstrated by normalization of morning cortisol levels and the ACTH stimulation test. The clinical implications of cortisol suppression by DAGR require further exploration.

 

Disclosure: KCJY: Principal Investigator, OPKO, Ad Hoc Consultant, Pfizer, Inc., Ad Hoc Consultant, Corcept. FB: Scientific Board Member, Pfizer, Inc.. DM: Employee, Pfizer, Inc.. SK: Employee, Pfizer, Inc.. BT: Employee, Pfizer, Inc.. RR: Employee, Pfizer, Inc.. JHH: Employee, Pfizer, Inc..

OR06-3 21844 3.0000 A Profound Suppression of Endogenous Cortisol Secretion with Pf-04171327 (a dissociated agonist of glucocorticoid receptor) Compared to Prednisone 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 1:00:00 PM OR06 5902 11:30:00 AM HPA Axis and Adrenal: Receptors To Clinical Impact Oral


Stéphanie Espiard*1, Ludivine Drougat1, Rossella Libe1, Guillaume Assie1, Karine Perlemoine1, Laurence Guignat2, Gaelle Barrande3, Francoise S Brucker-Davis4, Stephanie L Lopez5, Emmanuel D Sonnet6, Florence Torremocha7, Nathalie Chabbert-Buffet8, Marie-Laure Raffin-Sanson9, Lionel Groussin10, Francoise Borson-Chazot11, Xavier Bertagna1, Constantine A Stratakis12, Felix Beuschlein13, Bruno Ragazzon1 and Jerome Yves Bertherat14
1INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, France, 2Department of Endocrinology, Referral Center for Rare Adrenal Diseases, Assistance Publique Hôpitaux de Paris, Hôpital Cochin, Paris, 3Department of Endocrinology, Regional Hospital of Orléans, Orleans, France, 4Department of Endocrinology, Diabetology and Reproductive Medicine, University Hospital of Nice, Nice, France, 5Department of Internal Medicine B, Endocrinology and Metabolic Diseases, University Hospital of Limoges, Limoges Cedex, France, 6Department of Endocrinology, Hôpital de la Cavale Blanche, University Hospital of Brest, Brest Cedex, France, 7Department of Internal Medicine and Endocrinology, University Hospital of Poitiers, Poitiers, 8Unit of Endocrinology, Obstetrics, Gynecology and Reproductive Medicine Department, Assistance Publique Hôpitaux de Paris, Hôpital Tenon, Paris, France, 9Assistance publique hôpitaux de Paris, Hôpital Ambroise Paré, Service d'Endocrinologie, France, 10INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, 11Hospices Civils de Lyon, Groupement Hospitalier est, Bron Cedex, France, 12National Institutes of Health (NIH), Bethesda, MD, 13Klinikum der Universität München, Ludwig-Maximilian University, Munich, Germany, 14INSERM U 1016, Cochin Institute, Paris Descartes University, Paris, France

 

Context: Primary bilateral macronodular adrenal hyperplasia (PBMAH) is a rare cause of Cushing’s syndrome (CS) and bilateral adrenal tumors. Germline mutations of ARMC5 have been recently identified as a frequent cause of sporadic and familial PBMAH (1-5). 

Objective: The aim of this study was to determine the frequency of ARMC5 mutations in a large cohort of PBMAH index cases with subclinical or clinical CS. Genotype-phenotype correlation was established.

Patients and Methods: 98 patients with PBMAH were investigated. Leukocytes and/or tumor DNA was genotyped for ARMC5 by Sanger sequencing. Clinical data were collected (age, sex, CS, hypertension). Basal (midnight cortisol level, ACTH, androgens, mineralocorticoids hormones) and post-test parameters (overnight dexamethasone suppression test, ACTH 1-24 stimulation test, and illegitimate receptor responses) were assessed. Computed tomography and histological reports were analyzed.

Results: ARMC5 inactivating mutations were characterized for 24 patients (26%). In total, 36 different mutations were found by the sum of the germline and somatic DNA analysis in addition to one germline microdeletion and somatic losses of heterozygosity. The pathogenic role of the 10 missense mutants and the p.F700del deletion was demonstrated in H295R and Hela cell lines using immunofluorescence staining and flow cytometry. ARMC5 mutants were unable to induce apoptosis unlike the wild-type gene. In addition, 23 patients (23.5%) harbored missense variants which are predicted “benign” by in silico models and previously reported in 1000 genomes and Exome Variants Server databases. These patients and those without variations were considered as wild-type. Seven patients were not included in the comparison because of variants of uncertain significance. ARMC5 mutated patients exhibited more severe CS and were younger than wild-type patients. Overt clinical CS, higher midnight plasma cortisol, lower plasma ACTH and higher plasma cortisol after dexamethasone suppression was observed in the mutated patients. Adrenals of mutated patients were bigger with a higher number of nodules. No mutated patients, but 9 wild-type patients presented with food dependent CS.

Conclusions: More than one quarter of unrelated PBMAH patients present a pathogenic germline ARMC5 defect and these index cases present more severe CS and larger adrenal. Systematic genotyping of ARMC5 may help for early diagnosis of PBMAH, familial counseling, and patients’ management.  

 

Disclosure: JYB: Advisory Group Member, atterocor, Investigator, Novartis Pharmaceuticals, Investigator, Ipsen. Nothing to Disclose: SE, LD, RL, GA, KP, LG, GB, FSB, SLL, EDS, FT, NC, MLR, LG, FB, XB, CAS, FB, BR

OR06-4 20748 4.0000 A Armadillo Repeat Containing 5 Gene (ARMC5) in a Large Cohort of Primary Bilateral Macronodular Adrenal Hyperplasia (PBMAH): Genotype-Phenotype Correlations 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 1:00:00 PM OR06 5902 11:30:00 AM HPA Axis and Adrenal: Receptors To Clinical Impact Oral


Guillaume Assie*1, Anne Jouinot2, Eric Letouze3, Olivia Barreau4, Martin Fassnacht5, Windy Luscap2, Hanin Omeiri6, Stephanie Rodriguez7, Karine Perlemoine1, Fernande rené-Corail8, Nabila Elarouci9, Silviu Sbiera5, Matthias Kroiss5, Bruno Allolio5, Jens Waldmann10, Marcus Quinkler11, Massimo Mannelli12, Franco Mantero13, Thomas Papathomas14, Ronald R de Krijger15, Antoine Tabarin16, Veronique Kerlan17, Eric Baudin18, Frederique Tissier6, Bertrand Dousset19, Lionel Groussin20, Laurence Amar21, Eric Laurent Clauser22, Simon Faillot20, Xavier Bertagna1, Bruno Ragazzon1, Felix Beuschlein23, Rossella Libe1, Aurélien de Reynies24 and Jerome Yves Bertherat25
1INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, France, 2INSERM U1016, CNRS UMR 8104, Institut Cochin Université Paris Descartes, Paris, 3Ligue Nationale Contre Le Cancer, Paris, France, 4Inserm U1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, 5University Hospital Wuerzburg, Wuerzburg, Germany, 6INSERM U1016, CNRS UMR 8104, Institut Cochin Université Paris Descartes, 7Institut Cochin, INSERM U1016, CNRS 8104, Université Paris Descartes, 8INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, Paris, 9Ligue Nationale Contre Le Cancer, Paris, 10Visceral-, Thoracic and Vascular Surgery, University Hospital Giessen and Marburg, Marburg, Germany, 11Charite Campus Mitte, Berlin, Germany, 12Univ of Florence, Florence, Italy, 13University of Padova, Padova, Italy, 14Department of Pathology, Josephine Nefkens Institute, Erasmus MC-University Medical Center, Rotterdam, Netherlands, 15Erasmus MC, Rotterdam, Netherlands, 16University hospital of Bordeaux, Pessac, France, 17CHUBrest - Hopital De La Cavale Blanche, Brest, France, 18Institut Gustave-Roussy, Villejuif, France, 19Department of Digestive and Endocrine Surgery, Assistance Publique Hôpitaux de Paris, Hôpital Cochin, Paris, France, 20INSERM U 1016, CNRS 8104, Institut Cochin, Paris Descartes University, Paris, 21Hopital Europeen Georges Pompido, Paris, France, 22Assistance Publique Hôpitaux de Paris, Hôpital Cochin, Paris, France, Paris, France, 23Klinikum der Universität München, Ludwig-Maximilian University, Munich, Germany, 24Ligue Contre Le Cancer, Paris, 25INSERM U 1016, Cochin Institute, Paris Descartes University, Paris, France

 

The integrated genomics of adrenocortical carcinomas (ACCs) was recently reported (1), showing the existence of two molecular types, with different outcomes. The tumors with poor outcome include ACCs with global hypermethylation in the CpG islands of the gene promoter regions, and ACCs with mutations in a few recurrent genes, including mainly genes from the Wnt-βcatenin pathway (the newly identified tumor suppressor ZNRF3, and CTNNB1) and from the p53 pathway (TP53, CDKN2A and RB1).

Aim: To build a molecular prognostic predictor, based on tumor DNA alterations, including hypermethylation and mutations of these genes.

Methods:

A cohort of 130 ACCs collected within the COMETE and ENSAT networks were included. Targeted sequencing of 9 genes recurrently altered in ACC (ZNRF3, CTNNB1, TP53, RB1, CDKN2A) was performed by PCR-based capture(LifeTechnologies Ampliseq), followed by next generation sequencing (NGS, LifeTechnologies PGM). Homozygous deletions were identified by SNP arrays (Illumina). CpG island methylation status of each ACC was determined by MS-MLPA (methylation-specific multiplex-ligation-dependent probe amplification), using a commercially available kit (ME002-B1 kit, MRC Holland), and targeting CpGs islands of 4 genes (PAX5, GSTP1, PYCARD, PAX6) previously demonstrated as best reflecting the global methylation status (2); tumors were considered hypermethylated when the mean proportion of methylated alleles was >12% for these 4 genes.

Results:

MS-MLPA showed that 53% of the ACCs were hypermethylated. Hypermethylation was associated with worse survival (Cox p<0.001).

Targeted NGS and SNP arrays showed that mutations and or homozygous deletions of ZNRF3, CTNNB1, TP53, RB1, and CDKN2A were found in 20, 16, 15, 6 and 11 6% of the ACCs respectively. An alteration affecting the Wnt-βcatenin pathway (ZNRF3 or CTNNB1) was observed in 36% of ACCs. An alteration affecting the p53 pathway (TP53, RB1 or CDKN2A) was present in 29%.  Patients with an alteration affecting either the Wnt-βcatenin or p53 pathway presented a worse survival (Cox p=0.019).

Tumors of the poor outcome molecular subtype, defined as the tumors presenting either an hypermethylation, or an alteration of the Wnt-βcatenin or p53 pathways, corresponded to 71% of the cohort. These patients presented a worse survival (Cox p=0.002).

Conclusion:

Tumor DNA can be used to determine the ACC molecular subtype, and therefore the prognosis of ACC patients. This will help to design new prognostic molecular tools.

 

Disclosure: MF: Advisory Group Member, Atterocor. AT: Advisory Group Member, Ipsen, Advisory Group Member, Novartis Pharmaceuticals, Investigator, Novartis Pharmaceuticals, Speaker, Ipsen, Speaker, Novartis Pharmaceuticals. JYB: Advisory Group Member, atterocor, Investigator, Novartis Pharmaceuticals, Investigator, Ipsen. Nothing to Disclose: GA, AJ, EL, OB, WL, HO, SR, KP, FR, NE, SS, MK, BA, JW, MQ, MM, FM, TP, RRD, VK, EB, FT, BD, LG, LA, ELC, SF, XB, BR, FB, RL, AD

OR06-5 22107 5.0000 A Survival of Adrenocortical Carcinoma Determined from the Tumor DNA : An Application of the Post-Genomics Area 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 1:00:00 PM OR06 5902 11:30:00 AM HPA Axis and Adrenal: Receptors To Clinical Impact Oral


Richard J. Auchus*1, Adina F. Turcu2, Joanna L. Spencer-Segal3, Robert H. Farber4, Rosa Luo4, Dimitri E. Grigoriadis4 and Chris F. O'Brien4
1University of Michigan, Ann Arbor, MI, 2The University of Michigan, Ann Arbor, MI, 3University of Michigan, Ann Arbor, NY, 4Neurocrine Biosciences, San Diego, CA

 

Background: Classic CAH is an autosomal recessive genetic disorder that results in little or no cortisol biosynthesis, which eliminates feedback inhibition of pituitary adrenocorticotropic hormone (ACTH) secretion, increases production of precursor steroids, and leads to adrenal-derived androgen excess.  NBI-77860 is a selective corticotropin-releasing factor type 1 (CRF1) receptor antagonist that has demonstrated an ability to reduce ACTH release and is being developed as a novel oral treatment for classic CAH due to 21‑hydroxylase deficiency (21OHD). 

Methods: This study was a single-blind, placebo-controlled, single center, fixed-sequence, single-dose trial in adult female 21OHD patients.  A total of 8 classic 21OHD females, ages 19 to 58, were administered single doses of NBI-77860 300 mg, 600 mg, and placebo at 2200h during three separate treatment periods separated by a 3-week washout period.  Blood samples were collected over 24-hours to evaluate the pharmacokinetics (PK) of NBI-77860 and the hypothalamic-pituitary-adrenal (HPA) axis biomarkers ACTH, 17-hydroxyprogesterone (17OHP), androstenedione, and testosterone.  The primary analysis of biomarker data was an examination of grouped subject data, for each biomarker, expressed as a mean percent change from predose levels for the two active dosing conditions relative to the placebo condition. 

Results: Plasma levels of NBI-77860 were consistent with previous Phase 1, PK experience with this compound.  There were clinically meaningful reductions from predose levels of both 17OHP and ACTH following administration of NBI-77860 relative to placebo.  The 300 mg and 600 mg doses yielded nearly identical effects on mean percent change from predose in 17OHP and ACTH.  Variable reductions in androstenedione and testosterone levels were observed.  Individual subject responses were also evaluated, and treatment “responders” were conservatively defined as subjects with at least a 50% decrease in 17OHP and ACTH 8-12 hours following NBI-77860 dosing relative to placebo during the peak morning period.  This responder rate, following a single dose, was 50% for the study.  The single bedtime doses of NBI-77860 were well-tolerated in these adult female 21OHD patients.  The overall incidence of adverse events was low and similar among the three treatment conditions. 

Conclusions: Meaningful reductions in HPA axis biomarkers, particularly 17OHP and ACTH, were observed following NBI-77860 dosing in these patients.  These data are promising and provide a rational basis for continuing investigations of the CRF1 receptor antagonist, NBI-77860, and its activity in reducing ACTH and steroid biomarkers in the target population.  Studying the pharmacodynamics and PK of repeated doses of NBI-77860 will be helpful in this regard.

 

Disclosure: RJA: Investigator, Quest Diagnostics, Advisory Group Member, Laboratory Corporation of America, Investigator, Neurocrine Biosciences, Investigator, Novartis Pharmaceuticals, Consultant, Corcept. RHF: Employee, Neurocrine Biosciences. RL: Employee, Neurocrine Biosciences . DEG: Employee, Neurocrine Biosciences . CFO: Employee, Neurocrine Biosciences . Nothing to Disclose: AFT, JLS

OR06-6 20127 6.0000 A A Pharmacokinetic and Biomarker Study of the Corticotropin-Releasing Factor Receptor Antagonist NBI-77860 in Adult Females with Classic, 21-Hydroxylase Deficiency, Congenital Adrenal Hyperplasia (CAH) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 1:00:00 PM OR06 5902 11:30:00 AM HPA Axis and Adrenal: Receptors To Clinical Impact Oral


Kristy M. Heppner*1, Arian F. Baquero1, Camdin Bennett1, Melissa A. Kirigiti2, Sarah R. Lindsley1, Martha A. Bosch3, Oline K. Ronnekleiv3, Kevin L. Grove1 and M. Susan Smith1
1Oregon National Primate Research Center, Oregon Health & Science University, Beaverton, OR, 2Oregon National Primate Research Center, Beaverton, OR, 3Oregon Health & Science University, Portland, OR

 

Reproductive function is tightly linked to energy status, and reproductive disorders are often associated with metabolic dysfunction. Kisspeptin (Kiss1) neurons in the hypothalamic arcuate nucleus (ARC) are key components of the reproductive neuroendocrine axis as they regulate the basal pulsatile release of gonadotropin releasing hormone (GnRH). The identity of factors that regulate energy metabolism that might also modulate ARC Kiss1 neurons is largely unknown. One possible factor is glucagon-like peptide-1 (GLP-1), a neuropeptide produced by preproglucagon (PPG) expressing neurons in the brainstem. PPG neurons project to brain regions that regulate energy metabolism, including the ARC where GLP-1 regulates feeding and body weight. Although GLP-1 action on energy metabolism is well known, its action on the reproductive neuroendocrine axis has been largely unexplored. As GLP-1 fiber projections and the GLP-1 receptor (GLP-1R) are abundant in the ARC, we hypothesized that GLP-1 could modulate ARC Kiss1 action. Using immunohistochemistry, we found that GLP-producing neurons came in close apposition with ARC Kiss1 cells of ovariectomized (OVX) mice. Furthermore, single cell real-time PCR demonstrated that ARC Kiss1 neurons contained Glp-1r mRNA. Loose patch recordings from brain slices revealed that liraglutide (a long-acting GLP-1R agonist) increased action potential firing in ARC Kiss1 cells.  Moreover, current clamp recordings showed that liraglutide caused a membrane depolarization of ARC Kiss1 cells in the presence of presynaptic blockers.    

Calorie restriction (CR) potently suppresses ARC Kiss1 expression and downstream GnRH/luteinizing hormone (LH) release. As we identified GLP-1 as a positive regulator of ARC Kiss1 neurons, we hypothesized that a suppression of CNS GLP-1 action is contributing to reduced ARC Kiss1 action during CR.  First, we found that brainstem Ppg mRNA is reduced following a 48h fast in OVX mice.  However, restoring GLP-1 action using subcutaneous injections of liraglutide, which is known to enter the ARC, did not prevent LH inhibition during a 48h fast in OVX mice. Taken together, these studies demonstrate that GLP-1 is a positive regulator of ARC Kiss1 neurons.  Although suppression of brainstem Ppg mRNA may be contributing to the suppression of ARC Kiss1 and downstream GnRH/LH during CR, restoring GLP-1 action using peripheral injections of liraglutide was not sufficient to prevent LH inhibition during a 48h fast. PPG neurons do not express the GLP-1R, and therefore studies using methods that directly stimulate brainstem PPG neurons will give us a more accurate assessment of whether central PPG activation during CR will prevent LH inhibition. As a whole, more studies are necessary to understand the physiological significance of GLP-1 interaction with ARC Kiss1 neurons.

 

Disclosure: KLG: Principal Investigator, ERX, Consultant, Ember, Principal Investigator, Novo Nordisk, Advisory Group Member, Novo Nordisk, Consultant, Sanofi, Employee, Novo Nordisk. Nothing to Disclose: KMH, AFB, CB, MAK, SRL, MAB, OKR, MSS

OR05-1 19512 1.0000 A Glucagon-like Peptide-1 (GLP-1) Action on the Reproductive Neuroendocrine Axis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR05 5908 11:30:00 AM GnRH & Neuroendocrinology Oral


Alexander N Comninos*1, Jelena Anastasovska2, Meliz Sahuri-Arisoylu2, Xiao Feng Li3, Shengyun Li3, Minghan Hu3, Channa N Jayasena2, Mohammad A Ghatei4, Stephen R Bloom2, Paul Matthews2, Kevin O'Byrne3, Jimmy D Bell2 and Waljit S Dhillo1
1Imperial College NHS Healthcare Trust, London, United Kingdom, 2Imperial College London, United Kingdom, 3King's College London, London, United Kingdom, 4Imperial College London, London, United Kingdom

 

Kisspeptin is a crucial activator of reproductive function. The role of kisspeptin has been studied extensively within the hypothalamus but little is known about its significance in other areas of the brain. Exploring the extra-hypothalamic roles of kisspeptin is of key importance for our understanding of kisspeptin signaling and for the development of kisspeptin as a therapeutic.

Kisspeptin and its cognate receptor are expressed within the amygdala, a key limbic brain structure that has important roles in social and reproductive behaviours. In addition, the amygdala exerts an ‘inhibitory brake’ on reproduction via inhibitory projections to hypothalamic centers involved in reproductive hormone release. We therefore hypothesized that kisspeptin affects neuronal activation and reproductive pathways beyond the hypothalamus and particularly within the amygdala.

In a first test of our hypothesis, we mapped brain neuronal activity (using Manganese-Enhanced MRI) in adult rodents during peripheral (ip) administration of kisspeptin. In a second test of our hypothesis we investigated functional relevance in adult rodents by assessing the gonadotrophin response to direct intra-amygdala (MeA) administration of kisspeptin or kisspeptin antagonist.

Peripheral kisspeptin administration resulted in a marked decrease in neuronal activity in the amygdala compared to vehicle (20% decrease left amygdala kisspeptin vs. vehicle p=0.026; 22% decrease right amygdala kisspeptin vs. vehicle p=0.009)(n=7-8/group). This was associated with a simultaneous increase in luteinizing hormone (LH) secretion (vehicle +0.6±0.2ng/ml, kisspeptin +3.7±0.5ng/ml, p<0.0001 vs. vehicle). In addition, direct intra-MeA administration of kisspeptin resulted in a dose-dependent increase in LH secretion (kisspeptin 100pmol vs. vehicle, p=0.007; kisspeptin 1nmol vs. vehicle, p=0.002; kisspeptin 1nmol vs kisspeptin 100pmol, p=0.002)(n=5-7/group). Finally, blocking endogenous kisspeptin signaling specifically within the amygdala by administering intra-MeA kisspeptin antagonist decreased LH secretion (p=0.041 vs. vehicle) as well as LH pulse frequency (p=0.001 vs. vehicle)(n=5-7/group).

Collectively, these data provide the first evidence for a novel pathway in which kisspeptin inhibits neuronal activity in the amygdala, thereby releasing the amygdala’s ‘inhibitory brake’ on reproduction, resulting in stimulation of gonadotrophin secretion and pulsatility. In addition, this is the first report of kisspeptin signaling outside the hypothalamic-pituitary-gonadal axis influencing reproductive hormone secretion. Therefore, these data have important implications for our understanding of reproductive biology, as well as the development of kisspeptin as a therapeutic.

 

Nothing to Disclose: ANC, JA, MS, XFL, SL, MH, CNJ, MAG, SRB, PM, KO, JDB, WSD

OR05-2 18810 2.0000 A Kisspeptin Signaling in the Amygdala Modulates Reproductive Hormone Secretion 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR05 5908 11:30:00 AM GnRH & Neuroendocrinology Oral


Graeme L Fraser*1, Steven Ramael2, Hamid R Hoveyda3 and Jean Combalbert2
1Euroscreen SA, Gosselies, Belgium, 2Euroscreen SA, 3Euroscreen SA, Belgium

 

The role of NK3 receptor (NK3R) signaling in the central neuroendocrine control of the hypothalamic-pituitary-gonadal (‘HPG’) axis is widely recognized based upon both recent genomic and pharmacological research.  We have previously demonstrated in preclinical models that NK3R antagonists selectively inhibit LH, but not FSH, and thereby modulate the HPG axis in a more refined, ostensibly non-castrating, manner than the GnRH ligands currently used for the treatment of sex-hormone disorders. The objective of the current study was to evaluate the safety, pharmacokinetics and key efficacy biomarkers following 21 days of ESN364 administration to healthy premenopausal women.  This was a double-blind, placebo-controlled, multiple-dose study with sequential dose escalation where subjects (N=6/group) were administered placebo, 20, 60 or 180 mg once daily with treatment initiation on Day 3 (±2) after onset of menses.  ESN364 was well-tolerated and rapidly bioavailable after oral administration. The LH surge was delayed in a dose-dependent manner consistent with a significant prolongation of the menstrual cycle, delayed presentation of a dominant follicle and delayed endometrial thickening (p<0.05, in all cases). Dose-dependent decreases in progesterone (up to >70%) and estradiol (up to >50% in both the follicular and luteal phases) were observed, although at no time did estradiol levels decline below 20 pg/mL (levels associated with bone mineral density loss). No inhibition of FSH levels occurred at any time throughout the menstrual cycle. ESN364 effects were rapidly reversed after discontinuation as demonstrated by the immediate restoration of normal menstrual cycle length and associated hormone levels. This is the first demonstration of the clinical effects of an NK3R antagonist to suppress the reproductive endocrine axis in healthy premenopausal women. In total, these data suggest that ESN364 may offer a safe, effective alternative to GnRH ligands in the treatment of reproductive hormone-dependent disease states in premenopausal women.

 

Disclosure: GLF: Chief Scientific Officer, Euroscreen SA. SR: Clinical Researcher, Euroscreen SA. HRH: Management Position, Euroscreen SA. JC: Management Position, Euroscreen SA.

OR05-3 21733 3.0000 A Suppression of Lutenizing Hormone, Estradiol and Progesterone in Premenopausal Women By Oral Administration of the NK3 Receptor Antagonist ESN364 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR05 5908 11:30:00 AM GnRH & Neuroendocrinology Oral


Karolina Skorupskaite*1, Jyothis T George2 and Richard Alexander Anderson3
1The Queen's Medical Research Institute, Edinburgh, United Kingdom, 2University of Oxford, Oxford, United Kingdom, 3University of Edinburgh, Edinburgh, United Kingdom

 

Background

The hypothalamic neuropeptide kisspeptin is a key regulator of GnRH and thus gonadotropin (LH and FSH) secretion. Gonadotropin response to exogenous kisspeptin is increased in the late follicular phase, suggesting a central role of kisspeptin in mediating pre-ovulatory positive estrogen feedback. We hypothesised that exogenous kisspeptin, administered concurrently with appropriately timed estrogen, would further increase gonadotropin secretion, mimicking the physiological mid-cycle surge.

Methods

We used a model of follicular phase administration of transdermal estradiol (200µg/day) to induce positive feedback and increase LH secretion 48 hr later. Eight women with regular menstrual cycles received this regimen on cycle day 9-10 and at 24 hr were randomised to receive kisspeptin-10 (4µg/kg/hour iv) or saline infusion for 7 hours, at the end of which estradiol treatment was discontinued. All women returned in a subsequent cycle to receive the alternate treatment allocation.  Hormone concentrations were compared by ANOVA with Bonferroni multiple comparison post hoc analysis.

Results

Model Validation

In control (saline infusion) cycles, estradiol increased from 302±39 pmol/l pre-administration to 744±98 at 24 hr after patch administration (p<0.0001); and after removal of estradiol patches at 32 hr fell to 351±83 pmol/l at 48 hr. LH increased after 48hr but not 24hr of estradiol treatment (time 0: 4.3±0.6; 24 hr: 3.2±0.3; 48 hr: 8.5±1.2 IU/l, p=0.0007). FSH initially fell during estrogen treatment, then increased significantly after 48hr in the saline group (time 0: 2.6±0.3; 24 hr: 1.8±0.3; 48hr: 3.4±0.6 IU/l, p=0.005).

Kisspeptin Response

Kisspeptin-10 acutely stimulated LH secretion, from 3.8±0.6 to 18.1±4.7 IU/l (p<0.0001) at the end of infusion, while saline had no effect (3.2±0.3 to 2.7±0.3 IU/l, ns). LH remained significantly elevated at 48 and 72 hr after the start of estrogen treatment (16.4±4.4 and 8.5±1.4 IU/l, both p<0.01 vs start of infusion); at those time points LH concentrations were similar to the estrogen-stimulated concentrations in the control group.

Kisspeptin-10 also acutely increased FSH secretion: from 1.7±0.1 IU/l pre-infusion to 3.9±0.6 IU/l post-infusion (p<0.001) while saline had no such effect (1.8±0.3 to 1.3±0.2 IU/l, ns). FSH remained elevated following kisspeptin-10 at 48 and 72 hr (4.0±0.5 and 3.8±0.3 IU/l, both p=0.0001) and was similar to FSH concentrations in controls.

Conclusions

Kisspeptin-10 amplified the positive feedback effect of estrogen with substantial LH secretion in this model of estrogen-induced gonadotropin secretion in women.  Notably, kisspeptin-10 also increased FSH secretion. These data suggest a central role for kisspeptin signalling in mediating the positive estrogen feedback which facilitates the ovulatory midcycle surge in gonadotropin secretion.

Funding: Wellcome Trust-STMTI

 

Disclosure: JTG: Clinical Researcher, Astra Zeneca, Clinician, Eli Lilly & Company, Clinician, Sanofi, Clinical Researcher, Takeda, Advisory Group Member, Novo Nordisk, Advisory Group Member, Bristol-Myers Squibb, Clinician, Astra Zeneca. RAA: Consultant, Astra Zeneca. Nothing to Disclose: KS

OR05-4 19196 4.0000 A Kisspeptin-10 Enhances the Effects of Positive Estrogen Feedback on LH and FSH Secretion in Women 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR05 5908 11:30:00 AM GnRH & Neuroendocrinology Oral


Magali van Leckwyck*, Weilin Kong, Kathryn Jane Burton, Marco Giacomini, Nathalie Vionnet and Francois Pierre Pralong
Lausanne University Hospital, Lausanne, Switzerland

 

The activity of the female neuroendocrine reproductive axis is intimately linked to nutritional status. Undernutrition has clearly been associated with hypogonadotropic hypogonadism. Obesity is also associated with decreased fertility, but the pathophysiological consequences of excess weight on the hypothalamic control of reproduction are far less understood. To gain insight into these mechanisms, we investigated the effects of short-term hypercaloric intake on the hypothalamic GnRH secretion in seven healthy, lean female volunteers. They were characterized by a mean age of 24.0 ± 0.8 years, a mean BMI of 22.0 ± 0.7 kg/m², and regular menstrual cycles lasting 29.4 ± 0.7 days. Hypothalamic GnRH secretion was evaluated in the follicular phase by assessing LH pulsatility on two different days (baseline assessment and hyperinsulinemic, euglycemic clamp), using frequent (q10’) blood sampling protocols. These protocols were performed at the end of two distinct nutritional interventions: the first intervention consisted of one week of controlled isocaloric diet (calculated as 1.5 times the resting metabolic rate). The second intervention (hypercaloric diet) lasted four weeks: ~50% extra calories were added in the form of sucrose (3 g/kg BW/day) and fat (1g/kg BW/day) to the usual diet of the volunteers (first three weeks), and to the controlled isocaloric diet (last week). This hypercaloric diet induced an average weight gain of 2 ± 0.3 kg (p<0.05), corresponding to a BMI increase of 0.7 ± 0.1 kg/m² (p<0.05). A significant decrease of 14.5 ± 4.5 % in insulin sensitivity was also observed (D = -1.8 ± 0.7 mg glc/kg/min; p<0.05). In these conditions, LH peak amplitude was significantly decreased by 17.9 ± 9.3% (D = -0.4 ± 0.13 U/l; p<0.05), while no change in the LH peak frequency was observed. These results demonstrate for the first time a significant effect of short-term hypercaloric intake on parameters of GnRH pulsatile secretion in normal, lean female volunteers, providing insight into the pathophysiological consequences of obesity on fertility.

 

Nothing to Disclose: MV, WK, KJB, MG, NV, FPP

OR05-5 20305 5.0000 A Short-Term Overfeeding Induces Alterations in Pulsatile LH Secretion in Healthy, Lean Young Women 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR05 5908 11:30:00 AM GnRH & Neuroendocrinology Oral


Brent S. Abel*1, Ranganath Muniyappa1, Monica C. Skarulis2, Phillip Gorden3 and Rebecca J. Brown4
1NIDDK, Bethesda, MD, 2DEOB, NIDDK, NIH, 3NIDDK, NIH, 4National Institute of Health, Bethesda, MD

 

Background: Leptin replacement in patients with leptin gene mutations improves hypogonadotropic hypogonadism (1-2). Exogenous leptin therapy has also been shown to increase luteinizing hormone (LH) pulsatility and restore ovulation in women with hypothalamic amenorrhea, a model of relative leptin deficiency (3). Patients with lipodystrophy have leptin deficiency due to lack of adipose tissue, and are a natural model to study effects of leptin deficiency and replacement in humans. The effects of leptin replacement on LH secretion in patients with lipodystrophy are unknown.

Objective: We examined LH secretory dynamics in male and female patients with lipodystrophy, on and off exogenous leptin therapy.

Methods: This was a 2 period, non-randomized study, including previously leptin-treated (n=4) and leptin-naïve (n=8) subjects (5 generalized lipodystrophy; 7 partial lipodystrophy).  In period 1 (5 days) the leptin-treated group continued leptin; leptin was withdrawn for the next 14 days (period 2).  Leptin-naïve subjects were studied without leptin in period 1, and with leptin replacement in period 2. Nocturnal LH secretory dynamics were assessed [23:00-7:00, q10 min sampling, analyzed by multiparameter deconvolution and approximate entropy (ApEn) algorithms] at the end of each period. Early morning follicle stimulating hormone (FSH), estradiol (E2), and total testosterone (T) were measured at the end of each period.

Results: There were no period effects between patients initiated or withdrawn from leptin, so data were analyzed in aggregate. Mean (on: 5.0 ± 3.1 U/L, off: 3.2 ± 1.3, p=0.02) and integrated nocturnal LH concentrations (on: 2403 ± 1495 U·L-1·min-1, off: 1534 ± 642, p=0.02) were higher on leptin therapy. LH burst frequency (on: 0.77 ± 0.26 hr-1, off: 0.67 ± 0.24, p=0.04) and secretory burst mass (on: 9.7 ± 15.4 U/L, off: 7.0 ± 11.2, p=0.04) were higher on leptin. Consequently, leptin therapy increased pulsatile production rate (on: 64 ± 101 U·L-1·8hr-1, off: 57 ± 73, p=0.01) but not basal secretion of LH (on: 20 ± 27 U·L-1·8hr-1, off: 21 ± 30, p=0.48). Orderliness of LH release (ApEn) tended to be higher on leptin therapy (p=0.09). FSH tended to be higher on leptin (on: 6.0 ± 6.2 U/L, off: 4.4 ± 3.9, p=0.05). In males (n=4), T was higher on leptin (on: 507 ± 286 ng/dL, off: 360 ± 174, p=0.04), and in females (n=8), E2 was higher on leptin (on: 74 ± 36 pg/mL, off: 29 ± 24, p=0.01).

Discussion: Leptin increases spontaneous nocturnal LH secretion in patients with lipodystrophy. This is consistent with rodent and in vitro studies showing a direct stimulatory effect of leptin on LH secretion. Increased LH frequency suggests additional hypothalamic effects. These novel findings may explicate some of the salutary effects of leptin therapy on the hypothalamic-pituitary-gonadal axis in patients with lipodystrophy.

 

Nothing to Disclose: BSA, RM, MCS, PG, RJB

OR05-6 19226 6.0000 A Recombinant Human Leptin (Metreleptin) Administration Augments Nocturnal LH Secretion in Lipodystrophy Patients 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR05 5908 11:30:00 AM GnRH & Neuroendocrinology Oral


María Teresa Muñoz-Calvo1, Vicente Barrios1, Jesús Pozo1, Gabriel Ángel Martos-Moreno1, Federico G Hawkins2, Horacio M Domene3, Hector G. Jasper4, Shoshana Yakar5, Cheryl A Conover6, John J Kopchick7, Julie Ann Chowen1, Ron G Rosenfeld8, Luis A Pérez-Jurado9 and Jesús Argente*1
1Hospital Infantil Universitario Niño Jesús. Universidad Autónoma de Madrid. CIBERobn, Instituto de Salud Carlos III, Madrid, Spain, 2Hospital Universitario 12 de Octubre, Madrid, Spain, 3R Gutierrez Children's Hosp, Buenos Aires, Argentina, 4Division of Endocrinology, Ricardo Gutierrez Children´s Hospital, and Center for Endocrinology Research, CEDIE/CONICET, Buenos Aires, Argentina., 5New York University College of Dentistry, New York, NY, 6Mayo Clinic, Rochester, MN, 7Ohio University, Athens, OH, 8Oregon Health and Science University, Portland, OR, 9Universitat Pompeu Fabra.Hospital del Mar Research Institute (IMIM). CIBERER, Instituto de Salud Carlos III., Barcelona, Spain

 

PAPP-A2 is a protease that cleaves IGF-binding protein (IGFBP)5 and to a lesser extent IGFBP3. Pappa2 knock-out (KO) mice have a reduction in body and organ size with skeletal abnormalities. However, in humans, mutations in this protease have not been described. We report here, for the first time, a family in which 2 of 4 siblings show blunted postnatal growth due to a homozygous frameshift mutation in exon 3 of the PAPPA2 gene (c.1927_1928insAT, p.D643fs25X) resulting in a premature stop codon. Both affected children; a 10 year old female with height in the 10th percentile and her brother (6 yrs old) in the 25th percentile, while their target height is in the 75thpercentile, were thin (BMI: -1.5 SDS and -1.8 SDS) and microcephalic (-2.78 SDS and -3.3 SDS), but with apparently normal intellectual abilities. Initial biochemical studies showed [represented as case 1 and 2 (normal range)]: IGF-1 [1162 and 917 (102-256 ng/ml)], IGFBP3 [5912 and 4850 (2206-4200 ng/ml)] and ALS [3745 and 3625 (999-2634 mU/ml)]. Mutation analysis of the IGF receptor gene was normal. Exome analysis revealed the above mentioned mutation in the PAPP-A2 gene. The parents (nonconsanguineous) and one sibling, all heterozygous for this mutation, as well as the homozygous wild type 4th sibling, all had normal growth parameters and no other clinical phenotype. Further biochemical analysis of the affected siblings showed: undetectable PAPP-A2, spontaneous GH secretion tested during 8 h in the normal upper range (mean 4.5 ng/ml), low free IGF-1[0.32 and 0.13 (0.45-0.86 ng/ml)]; reduced PAPP-A1 [0.55 and 0.54 (0.82-3.26 ng/ml)]; high insulin [15.1 and 27.0 (4.0-12.95 μU/ml)]; normal IGFBP1 [1.52 and 6.37 (1.22-22.32 ng/ml)] and decreased-normal IGFBP2 [75.6 and 340.6 (176.7-353.9 ng/ml)]; increased intact IGFBP4 [65.5 and 68.4 (6.8-32.9 ng/ml)] and total IGFBP4 [89.5 and 92.6 (18.4-60.7 ng/ml)]; low-normal IGFBP5 [197 and 280 (211-707 ng/ml)] with increased IGF1 binding activity [(87 and 93 (43-70 % active)], but with reduced overall concentrations bound to IGF1 as detected by cross-linking studies (43 and 58 % of control values). Functional studies also showed increased IGF binding capacity of IGFBP3. No other endocrine abnormalities were found and the hypothalamic-pituitary axis was normal by MRI. X-rays showed very thin long bones and normal bone density (DEXA), which is similar to that reported in the Pappa2 KO mice models. In contrast, organ size was found to be normal by ultrasound in these patients. A therapeutic trial with hrIGF1 with monitoring of growth velocity will be attempted in these patients. In conclusion, we have described a new syndrome due to a PAPP-A2 gene mutation, where despite very high total IGF1 levels, lack of this protease activity results in reduced IGF1 bio-availability that produces decreased longitudinal growth, microcephaly and skeletal abnormalities.

 

Nothing to Disclose: MTM, VB, JP, GÁM, FGH, HMD, HGJ, SY, CAC, JJK, JAC, RGR, LAP, JA

OR03-1 21121 1.0000 A A New Syndrome of Short Stature, Mild Microcephaly, Skeletal Abnormalities and High Circulating IGF1, IGFBP3 and ALS Associated with a Homozygous Mutation in the Gene for Pregnancy-Associated Plasma Protein A2 (PAPP-A2, pappalysin2) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 1:00:00 PM OR03 5917 11:30:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Oral


Isabelle Maystadt1, Shayne F Andrew2, Jean De Schepper3, Nathalie Wauters4, Valerie Benoît1, Pascal Joset5, Beatrice Oneda5, Andrew Dauber2, Ron G Rosenfeld6, Anita Rauch5 and Vivian Hwa*2
1Institut de Pathologie et de Génétique, Gosselies, Belgium, 2Cincinnati Children's Hospital Medical Center, Cincinnati, OH, 3Ghent University Hospital, Ghent, Belgium, 4Hopital Civil de Charleroi, Charleroi, Belgium, 5University of Zurich, Zurich-Schwerzenbach, Switzerland, 6Oregon Health and Science University, Portland, OR

 

Background: Heterozygous mutations in the insulin-like growth factor 1 receptor (IGF1R) are often associated with congenital IGF1 resistance, causing variable degrees of intrauterine growth retardation (IUGR) and postnatal short stature. Only one homozygous IGF1Rmutation has been described, associated with a child who presented with severe growth failure, mild intellectual impairment, microcephaly, dysmorphic features, cardiac malformations and disturbed glucose tolerance (1).

Clinical Case: Two siblings, an 11-year-old girl (P1) and a 7-year-old boy (P2), presented with postnatal growth failure (height -6.0 SDS) and dysmorphic features. They were born with severe IUGR (birth weight and length: -6 SDS) to healthy consanguineous short Yemenite parents (mother’s height, 145cm; father’s height, 160cm). A moderate delayed bone age and epiphyseal abnormalities were noted. Severe microcephaly (head circumference, -7 SDS), mild intellectual disability, deafness, congenital cardiac malformations, oligodontia and enamel hypoplasia, and a partial vermis hypoplasia (brain MRI) were also documented in both patients. Endocrine evaluations revealed normal basal growth hormone, elevated IGF1 and normal IGFBP3 concentrations. Fasting glucose levels were normal, although OGTT (2hr) for P1, was in the diabetic range, with elevated glucose (216, normal:70-110 mg/dl), insulin (66.5, normal:5-25 mcU/ml) and c-peptide (9.8, normal:1.1-3.6 ng/ml) levels, consistent with insulin insensitivity. P2 produced a disturbed glucose tolerance OGTT profile. Molecular evaluation by exome sequence analysis identified a novel homozygous, deletion of 12 nucleotides (exon 10) in the IGF1Rgene, which generated an in-frame p.A711_E714del. Functional analysis by fluorescence-activated cell sorting (FACS) of live primary fibroblasts derived from the patients demonstrated normal expression and cell surface localization of the mutant IGF1R. IGF-I-induced signal transduction, however, was significantly reduced, but not abrogated.

Conclusion: Homozygous IGF1R mutations identified to date do not completely ablate IGF1R expression/functions. Severe pre- and postnatal growth failure, mild intellectual disability, microcephaly and dysmorphic features were pronounced compared to patients with heterozygous IGF1R defects. These features, together with congenital cardiac anomalies and marked insulin insensitivity, support critical roles of IGF1R in human development and maintenance of glucose homeostasis, mechanism(s) to be elucidates.

 

Nothing to Disclose: IM, SFA, JD, NW, VB, PJ, BO, AD, RGR, AR, VH

OR03-2 19830 2.0000 A Novel Homozygous IGF1 Receptor (IGF1R) Mutation, p.A711_E714del, in Two Siblings with Severe Growth Failure, Congenital Malformations, Deafness and Insulin Insensitivity 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 1:00:00 PM OR03 5917 11:30:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Oral


Giampaolo Trivellin*1, Adrian F Daly2, Fabio R Faucz3, Bo Yuan4, Liliya Rostomyan2, Darwin Omar Larco5, Marie Helene Schernthaner-Reiter1, Eva Szarek1, Leticia F. Leal1, Jean-Hubert Caberg6, Emilie Castermans6, Chiara Villa6, Aggeliki Dimopoulos7, Prashant Chittiboina8, Paraskevi Xekouki1, Nalini Samir Shah9, Daniel Metzger10, Philippe A Lysy11, Emanuele Ferrante12, Natalia Strebkova13, Nadia Mazerkina14, Maria Chiara Zatelli15, Maya Beth Lodish1, Anelia Dafinova Horvath1, Rodrigo Bertollo de Alexandre1, Allison D Manning3, Isaac Levy1, Margaret Farmar Keil16, Maria De La Luz Sierra1, Leonor Palmeira2, Wouter Coppieters17, Michel Georges17, Luciana Ansaneli Naves18, Tim D. Cheetham19, Gustavo Barcelos Barra20, Mauricette Jamar6, Vincent Bours6, Tao-Yiao J Wu5, Catherine S. Choong21, Jerome Yves Bertherat22, Philippe Chanson23, Peter Kamenický23, William E. Farrell24, Anne Barlier25, Martha M Quezado26, Ivana Bjelobaba3, Stanko S Stojilkovic7, Jurgen Wess27, Stefano Costanzi28, Pengfei Liu4, James R Lupski4, Albert Beckers2 and Constantine A Stratakis1
1National Institutes of Health (NIH), Bethesda, MD, 2CHU de Liège-University of Liège, Liège, Belgium, 3National Institutes of Health, Bethesda, MD, 4Baylor College of Medicine, Houston, TX, 5Uniformed Services University, Bethesda, MD, 6University of Liège, Liège, Belgium, 7Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH), Bethesda, MD, 8National Institute of Neurological Diseases and Stroke (NINDS), National Institutes of Health, Bethesda, MD, 9KEM Hospital, Mumbai, Maharashtra, India, 10BC Children's Hospital, Vancouver, BC, Canada, 11Université Catholique de Louvain, Brussels, Belgium, 12University of Milan, Milan, Italy, 13Institute of Pediatric Endocrinology, Moscow, Russia, 14Burdenko Neurosurgery Institute, Moscow, Russia, 15University of Ferrara, Ferrara, Italy, 16Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH)., Bethesda, MD, 17GIGA-Genomics, Liège, Belgium, 18University of Brasilia, Brasilia, Brazil, 19Royal Victoria Infirmary, Newcastle upon Tyne, United Kingdom, 20Laboratorio Sabin, Brasilia, Brazil, 21Princess Margaret Hospital for Children, Subiaco WA, Australia, 22INSERM U 1016, Cochin Institute, Paris Descartes University, Paris, France, 23Univ Paris-Sud, UMR-S693, Assistance Publique-Hôpitaux de Paris, Le Kremlin Bicêtre, France, 24Keele University, Stoke on Trent, Staffordshire, United Kingdom, 25Aix-Marseille University, Marseille, France, 26National Cancer Institute, Bethesda, MD, 27NIH-NIDDK, Bethesda, MD, 28American University, Washington, DC

 

Background: High growth hormone (GH) secretion is a rare condition that leads to gigantism in childhood and acromegaly in adults; the causes of gigantism and acromegaly are poorly understood.

Aim of the study: To study the genetic defects underlying infantile gigantism and acromegaly.

Patients and methods: We performed genomic and genetic studies in patients with gigantism (n=46), and then sequenced an implicated gene in an international cohort of acromegaly patients (n=248). In vitro functional studies were performed in rat somatomammotrope GH3 cells to assess the role of an identified genetic variant.

Results: We detected by array-CGH a novel microduplication at chromosome Xq26 in two unrelated kindreds and twelve sporadic cases of various ethnic origins with infantile gigantism (n=16; 11 females). Two smallest regions of overlaps (SRO) were shared by the duplications; breakpoint junctions characterization revealed microhomology, suggesting a replicative mechanism for formation. The common duplicated genomic interval extends for approximately 500 Kb and contains four protein-coding genes. Only one of these genes, encoding a G-protein coupled receptor (GPCR) that strongly activates the cAMP signaling pathway was consistently over-expressed in patients’ pituitary lesions both at the RNA and protein level. We identified a recurrent variant in this gene in 4% of patients with acromegaly, mostly in tumors, whereas no pathogenic mutations were observed in the other three genes located within the SRO. When transfected into GH3 cells the mutation led to increased GH secretion and cell proliferation.

Conclusions: We describe a previously unrecognized genomic disorder caused by Xq26 microduplication (X-LAG for X-linked acro-gigantism) and characterized by early-onset gigantism resulting from GH excess that can be transmitted as a dominant trait. The gene is a GPCR that is dosage-sensitive and over-expressed in patients’ pituitary lesions; it activates the cAMP pathway, whose mitogenic effects in pituitary somatotroph cells are well established; and a recurrent mutation is found in patients with sporadic acromegaly.

G.T. and A.F.D. share the first authorship; A.B. and C.A.S. share the senior authorship of this abstract

 

Disclosure: AFD: Ad Hoc Consultant, NPS, Clinical Researcher, Pfizer, Inc.. PC: Researcher, Ipsen, Researcher, Novartis Pharmaceuticals, Researcher, Novo Nordisk, Researcher, Pfizer, Inc., Advisory Group Member, Ipsen, Advisory Group Member, Novartis Pharmaceuticals, Advisory Group Member, Viropharma, Speaker, Ipsen, Speaker, Novartis Pharmaceuticals, Speaker, Pfizer, Inc.. AB: Investigator, Ipsen, Medical Advisory Board Member, Novartis Pharmaceuticals, Clinical Researcher, Pfizer Global R&D. Nothing to Disclose: GT, FRF, BY, LR, DOL, MHS, ES, LFL, JHC, EC, CV, AD, PC, PX, NSS, DM, PAL, EF, NS, NM, MCZ, MBL, ADH, RBD, ADM, IL, MFK, MDLLS, LP, WC, MG, LAN, TDC, GBB, MJ, VB, TYJW, CSC, JYB, PK, WEF, AB, MMQ, IB, SSS, JW, SC, PL, JRL, CAS

OR03-3 20120 3.0000 A X-Linked Acro-Gigantism (X-LAG) Due to Microduplications of Chromosome Xq26: A New Disorder and Implications for Acromegaly 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 1:00:00 PM OR03 5917 11:30:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Oral


Ana Paula Abreu*1, Victor M. Navarro2, Aysegul Eren3, Joy N. Liang1, John C Gill4, Sekoni D. Noel4, Iain Robert Thompson4, Stephanie A. Roberts5, Rona S. Carroll4, Ana Claudia Latronico6 and Ursula B. Kaiser1
1Brigham and Women's Hospital/Harvard Medical School, Boston, MA, 2Harvard Medical School and Brigham and Women's Hospital, Boston, MA, 3Western Reserve Health Education/ Northside Medical Center, Boston, MA, 4Brigham and Women's Hospital/Harvard Med School, Boston, MA, 5Boston Children's Hospital/Harvard Med School, Boston, MA, 6Hospital das Clinicas da Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil

 

The identification of loss-of-function MKRN3 mutations in 30% of a cohort of families with central precocious puberty linked this gene to the reproductive axis and GnRH regulation. Subsequent studies in patients with and without family histories of central precocious puberty confirmed these findings. Mkrn3 expression in the arcuate nucleus (ARC) of mice is high in the juvenile period and markedly decreases before puberty. Taken together, human and mouse data show an inhibitory effect of MKRN3 on GnRH secretion, but the mechanism of action of MKRN3 is yet to be understood. Furthermore, factors contributing to regulation of Mkrn3 expression are not known. We hypothesized that sex steroids might regulate Mkrn3 expression to contribute to its decline at the onset of puberty in mice. We assessed Mkrn3 expression in the hypothalamic ARC and anteroventral periventricular nucleus (AVPV) of wild type (WT) and hypogonadal (hpg) male and female mice to determine: (1) if the absence of sex steroids would alter the Mkrn3 expression pattern; and (2) if the Mkrn3 expression pattern is different between these two nuclei. hpg mice are GnRH deficient, resulting in lack of pubertal development and almost absent circulating sex steroids. Mkrn3 expression decreased across pubertal maturation in the AVPV of WT mice, similar to the expression pattern observed in the ARC. There was no difference in Mkrn3 expression levels between WT and hpg male and female mice across pubertal maturation in either the ARC or the AVPV. Estradiol (E2) has a positive feedback action on Kiss1 expression in the AVPV of female mice and is implicated in the LH surge. Conversely, E2 has a negative feedback action on Kiss1 expression in the ARC. We hypothesized that E2 treatment might decrease AVPV Mkrn3 expression to contribute to the pubertal increase in AVPV Kiss1 expression. To further investigate the possibility of E2 regulation of Mkrn3 expression, we treated WT female mice at postnatal day 11 (P11) with 2 μg E2 or vehicle subcutaneously and measured AVPV and ARC Mkrn3 and Kiss1 expression 24 hours later. E2 treatment did not alter Mkrn3 expression in the AVPV or in the ARC of female juvenile mice. As expected, Kiss1 expression showed a significant increase in the AVPV and a significant decrease in the ARC of E2-treated female mice compared to controls. In summary, the decrease in Mkrn3 expression in the AVPV across pubertal maturation suggests that Mkrn3 may be playing an inhibitory role in the AVPV, as previously also suggested for the ARC. Furthermore no global regulation of Mkrn3 expression by sex steroids was detected in the ARC or AVPV, suggesting that the decline in Mkrn3 at the onset of puberty may occur independently of the pubertal increase in gonadal sex steroid production.

 

Nothing to Disclose: APA, VMN, AE, JNL, JCG, SDN, IRT, SAR, RSC, ACL, UBK

OR03-4 21239 4.0000 A Regulation of Mkrn3 Expression By Sex Steroids 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 1:00:00 PM OR03 5917 11:30:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Oral


Julian Lui*1, Kevin Barnes1, Presley Garrison2, Abigail Paulson2, Vijay Shimoga2, Ola Nilsson3 and Jeffrey Baron4
1NICHD, Bethesda, MD, 2NICHD, 3NICHD, NIH, Bethesda, MD, 4NIH, Bethesda, MD

 

EZH2 (Enhancer of zeste 2) is a key member of the Polycomb Repressor Complex 2 (PRC2), which is responsible for methylation of histone 3 lysine 27 (H3K27), a histone modification important for chromatin condensation and gene silencing. In humans, heterozygous mutations in EZH2 cause Weaver syndrome, an overgrowth disorder with skeletal abnormalities and advanced bone age. In addition, the EZH2 gene lies in a locus associated with adult human height variation, suggesting that EZH2, and perhaps H3K27 methylation, play important roles in skeletal and overall body growth. We therefore sought to investigate the regulation of skeletal growth by EZH2. Because Ezh1 and 2 have partially redundant functions in the PRC2 histone methylase complex, we generated a mouse model with a complete homozygous knockout of Ezh1 and cartilage-specific homozygous knockout of Ezh2 (Ezh1-/-, Ezh2fl/fl, Col2­a1-cre). Complete knockout of Ezh1 or cartilage-specific Ezh2 knockout alone showed no growth phenotype. However, the combined knockout mice exhibited postnatal growth retardation that gradually becomes more prominent from birth to 2-wks of age (body weight at 2-wks old, KO 3.7 ± 0.6 vs WT 8.2 ± 0.5 g, P<0.01). Immunostaining showed that dimethyl- and trimethyl-H3K27 were absent in the cartilage of Ezh1-/-, Ezh2fl/fl, Col2­a1-cre mice, indicating loss of PRC2 function in cartilage. Longitudinal bone growth of these mice was significantly impaired by 3 days of age, with decreased tibial length (5.44 ± 0.08 vs WT 6.00 ± 0.11 mm, P<0.01) and decreased proliferation in the proliferative columns (5.07 ± 0.26 vs WT 6.19 ± 0.19 BrdU-labeled cells/column, P<0.01). Knockout mice also exhibited decreased proliferative zone height (291 ± 8mm vs WT 395 ± 11mm, P<0.01), decreased number of proliferative chondrocytes per column (56 ± 2 cells vs WT 73 ± 3 cells, P<0.01), decreased terminal hypertrophic cell size (15.4 ± 0.4mm vs WT 20.5 ± 0.5mm, P<0.01), but increased number of hypertrophic cells per column (13.9 ± 0.5 cells vs WT 11.3 ± 0.5 cells, P<0.01). Collectively, these data indicate that loss of histone methyltransferases Ezh1 and 2 in the growth plate impairs longitudinal skeletal growth by inhibiting chondrocyte proliferation and causing premature hypertrophic differentiation and therefore suggest that H3K27 methylation by the PRC2 complex is essential for normal mammalian skeletal growth.

 

Nothing to Disclose: JL, KB, PG, AP, VS, ON, JB

OR03-5 19252 5.0000 A Histone Methyltransferases Ezh1 and 2 Are Required for Normal Skeletal Growth 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 1:00:00 PM OR03 5917 11:30:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Oral


Adam Stevens*1, Chiara De Leonibus1, Philip Murray1, Ekaterina B Koledova2, Pierre Chatelain3 and Peter Clayton4
1University of Manchester and Royal Manchester Children’s Hospital, Manchester, United Kingdom, 2Merck, Darmstadt, Germany, 3Université Claude Bernard, Lyon, France, 4University of Manchester & Manchester Academic Health Sciences Centre, Manchester, United Kingdom

 

Background: The growth promoting effects of growth hormone (GH) are linked with insulin sensitivity and lipid metabolism; the underlying biological networks are unclear.

Objectives: To identify gene clusters and pathways linking growth and metabolic response in children with growth hormone deficiency (GHD) treated with recombinant human growth hormone (r-hGH).

Methods: Pre-pubertal children with GHD (n=125) were enrolled from the PREDICT short-term (NCT00256126) and long-term follow-up prospective study (NCT00699855). Whole blood gene expression (GE) was determined prior to treatment with r-hGH. One year height velocity (HV) and 1 month change (Δ) in IGF-I on r-hGH were used as clinical markers of growth response; Δ of serum fasting glucose, insulin, triglycerides, LDL and HDL levels were used as clinical markers of metabolic function; the relationships between these variables were identified by partial correlation. Associations were made with basal GE using rank regression (p<0.05). The GE overlap (p<0.05) between clinical markers was used to construct network models and identify gene clusters. Clusters were ranked using their network centrality score (Moduland algorithm) to define associated biological functions (hypergeometric test with Benjamini-Hochberg false discovery rate modification, q<0.05). Causal network analysis (CNA), mapping known literature to the network models, was performed to identify master regulators of GE (Ingenuity Pathway Analysis).

Results: Six paired correlations of clinical markers were identified (p<0.05); HV with ΔIGF-I, HV with ΔLDL and ΔIGF-I/HDL/triglycerides/glucose (individually) with Δinsulin. The intersection between the network models defined by these correlations identified five clusters of genes that linked growth and metabolism. These clusters were associated with adipogenesis, cell cycle checkpoint control and AMPK/mTOR signaling pathways (q<0.001) linked to the master regulators RXR, PLK1 and TSC1, respectively (p<0.001).

Three genes associated with the network models were found in the overlap of the GE correlated with the markers HV, ΔIGF, ΔLDL and ΔInsulin: SPTBN1, RRBP1 and UGGT2. These genes are putative functional markers of the metabolic response to r-hGH and have been previously associated with adult height, obesity and insulin sensitivity.

Conclusions: This study has identified network models, biological pathways and genomic markers linking growth and metabolic response to r-hGH therapy in GHD children.

 

Disclosure: AS: Investigator, Merck Serono. EBK: Employee, Merck Serono. PC: Investigator, Merck Serono, Speaker, Merck Serono, Advisory Group Member, Merck Serono. PC: Investigator, Merck Serono, Speaker, Merck Serono. Nothing to Disclose: CD, PM

OR03-6 21692 6.0000 A Adipogenesis, mTOR and AMPK Pathways Link the Growth and Metabolic Actions of Growth Hormone Action in Children with GH Deficiency (GHD) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 1:00:00 PM OR03 5917 11:30:00 AM From the Genome to the Clinic: Growth and Puberty Disorders Oral


Joanna Klubo-Gwiezdzinska*1, Elaine Cochran2, Robert Kenneth Semple3, Rebecca J. Brown1 and Phillip Gorden2
1National Institute of Health, Bethesda, MD, 2NIH, Bethesda, MD, 3University of Cambridge, Cambridge, United Kingdom

 

Background

 Type B insulin resistance due to autoantibodies to the insulin receptor is characterized by diabetes refractory to massive doses of insulin, hypercatabolism resulting in dramatic weight loss, severe hyperandrogenism, and widespread acanthosis nigricans.

The 10 year mortality rate is 54%. In the present study we show the continued efficacy of combination therapy for type B insulin resistance.

 Trial design

 Inclusion criteria– patients with type B insulin resistance confirmed by the presence of insulin receptor antibodies.

 Study design- The patients underwent routine hematological, immunological, and biochemical testing at baseline and in regular intervals during the treatment. The treatment protocol consisted of:

-          Rituximab750 mg/meter body surface area iv in two consecutive doses 2 weeks apart, repeated every 3-4 months if the disease persisted

-          Cyclophosphamide 100 mg po daily continuously until remission was achieved

-          Dexamethasone 40 mg po  daily for 4 days or methylprednisolone 1 g iv for 2 days, repeated  every 3-4  weeks if the disease remained active

-          Once patients achieved remission- maintenance therapy with azathioprine 100 mg daily.

Primary endpoints - remission was defined as the amelioration of the hyperglycemia and discontinuation of insulin therapy and/or normalization of hyperandrogenemia.

Results

Seventeen patients fulfilled inclusion criteria – 14 women and 3 men with mean age of 38.4+/-17.6 years.

The underlying autoimmune background consisted of mixed connective tissue disease in 6/17 patients, lupus in 8/17 patients, Sjogren disease in 1 patient, uveitis in 1 patient and undetermined in 1 patient.

Mean baseline fasting glucose level was 307.5 +/-149 mg/dl, mean HgbA1c 11.3+/- 2.7%, mean baseline insulin level 1428+/-1708 mcU/ml (normal (N) 6.0-27.0), c-peptide 4.8+/-3.9 ng/ml (N 0.9-7.1), average daily insulin requirements ranged between 200 and 19000  units per day, mean total testosterone 463+/-346 ng/dl (N 8-60 for female patients), androstendione 374.4+/-426 ng/dl (N 30-200), total cholesterol 134.6+/-36.4 mg/dl, LDL 58.9+/-36.4 mg/dl, HDL  64+/-20.13 mg/dl, triglycerides 57.13+/-20.13 mg/dl. 

14/17 patients achieved remission after a median 5.5 months of therapy (range 2.5-27.25 months) as documented by normalization of fasting glucose (mean 88.8+/-26.8 mg/dl), HgbA1c (mean 6.1+/-2%), decreased to zero insulin requirements and normalization of testosterone levels to105.3+/-29.25 ng/dl and androstendione to 105.7+/-113 ng/dl. Three patients have yet to achieve remission had been treated most recently for only 1 to 5 months. During the course of the follow up (median 65 months), 2/17 patients developed disease recurrence with hyperandrogenemia, but again responded to combination therapy.

Conclusions

This standardized approach represents a personalized targeted therapy that is effective in this most extreme form of diabetes.

 

Nothing to Disclose: JK, EC, RKS, RJB, PG

OR01-1 21553 1.0000 A Continued Efficacy of Combination Therapy for Type B Insulin Resistance  Due to Autoantibodies to the Insulin Receptor 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR01 5933 11:30:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Oral


Donald C Simonson*1, Su Ann Ding2, Florencia Halperin1, Marlene Wewalka2, Kathleen Foster2, Katherine Kelly2, Jennifer Panosian2, Ann Goebel-Fabbri2, Osama Hamdy2, Kerri Clancy3, David Lautz3, Ashley Vernon1 and Allison B Goldfine2
1Brigham and Women's Hospital, Boston, MA, 2Joslin Diabetes Center, Boston, MA, 3Emerson Hospital, Concord, MA

 

Weight loss improves patient-reported outcomes in obese patients with type 2 diabetes (T2D); however, differences between surgical and medical weight management and the durability of these effects are not well established.  We randomized 40 obese patients with T2D (22M / 18F; weight 109 ± 15 kg; BMI 36.5 ± 3.7 kg/m²; age 51 ± 10 yrs; diabetes duration 9 ± 5 yrs; HbA1c 8.2 ± 1.2%; 40% on insulin) to laparoscopic adjustable gastric band (LAGB; n = 18) or a 12-week intensive medical diabetes and weight management program (IMWM; n = 22) with follow-up for 12 months.  At baseline, both groups had comparable SF-36 physical health (PH) (76 ± 14) and mental health (MH) (75 ± 13) status scores, and moderately elevated Impact of Weight on Quality of Life (IWQOL) (59 ± 17) and Problem Areas in Diabetes (PAID) (46 ± 14) scores.  After 10% weight loss or 3 months (if 10% loss was not achieved), LAGB and IWMW achieved similarly significant weight loss (-9.7 ± 0.8 vs. -9.2 ± 0.8 kg) and improvement in HbA1c (-1.0 ± 0.3 vs. -1.7 ± 0.3%; p = 0.06 between groups), respectively.  SF-36 PH (2 ± 2 vs. 6 ± 1) and MH (2 ± 2 vs. 6 ± 2) improved minimally from baseline in LAGB and IMWM, with no differences between groups.  IWQOL (-11 ± 2 vs. -9 ± 2) and PAID (-7 ± 2 vs. -9 ± 2) improved significantly from baseline (p<0.05 to p<0.01), but improvements were similar in both groups.  At 12 months, weight loss was significantly greater after LAGB (-13.5 ± 1.7 vs. -8.5 ± 1.6 kg; p < 0.05) but there was no difference in lowering of HbA1c (-1.2 ± 0.3 vs. -1.0 ± 0.3%).  SF-36 PH and MH changed minimally compared with the earlier assessment with no differences between groups.  Changes in IWQOL (-14 ± 2 vs. -11 ± 2) and PAID (-13 ± 2 vs. -13 ± 2) further improved significantly from baseline in both LAGB and IMWM, respectively (p<0.01 vs. baseline for both groups), but the effects were similar in the surgical and medical treatment arms.  In both groups combined, the improvement in HbA1c correlated with the improvement in patient’s self-assessment of diabetes-specific emotional distress as assessed by PAID (r = 0.51, p < 0.01).  We conclude that in obese patients with T2D: 1) both LAGB and IMWM result in significant weight loss and reduction in HbA1c, 2) weight loss is greater after LAGB, but the improvement in HbA1c is similar, 3) both treatments are associated with small but comparable changes in self-reported physical and mental health status as measured by the SF-36, and 4) both treatments are associated with significant reductions in the impact of weight on QOL and problem areas in diabetes, but there were no differences between groups.  These results should be considered when planning LAGB vs. intensive medical weight management for patients with type 2 diabetes.

 

Disclosure: OH: Research Funding, Metagenics, Research Funding, Neurometrix, Consultant, Abbott Laboratories, Consultant, Merck & Co.. ABG: Advisory Group Member, Novo Nordisk. Nothing to Disclose: DCS, SAD, FH, MW, KF, KK, JP, AG, KC, DL, AV

OR01-2 21557 2.0000 A Patient-Reported Outcomes One Year after Randomization to Laparoscopic Adjustable Gastric Banding or Intensive Weight and Diabetes Management in Obese Patients with Type 2 Diabetes Mellitus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR01 5933 11:30:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Oral


Michelle L Katz*1, Gabriela Heiden Telo2, Julia Beatrice Cartaya1, Carly E Dougher2, Ming Ding2 and Lori M Laffel2
1Massachusetts General Hospital, Boston, MA, 2Joslin Diabetes Center, Boston, MA

 

Background: Cardiovascular disease (CVD) remains a significant source of premature morbidity and mortality in persons with type 1 diabetes (T1D). However, T1D providers of youth and young adults, who focus on glycemic management, often inadequately treat nonglycemic CVD risk factors.  Large pediatric cohorts in the USA (SEARCH for Diabetes in Youth, Type 1 Diabetes Exchange) have documented widespread under-treatment of CVD risk factors, such as hypertension and hyperlipidemia. 

Methods:  In a T1D cohort of 645 youth and young adults followed for a median of 12.9 years (interquartile range 9.1-16.5), 212 (33%) had hyperlipidemia (LDL ≥130 mg/dl) and underwent detailed chart review. Clinical data, lab results, and provider management recommendations were extracted from 3,222 encounters. Analyses included assessment of patterns of persistent hyperlipidemia, effectiveness of provider recommendations, and acceptance and adherence to lipid-lowering treatments (Rx).  Comparisons of LDL levels before and after provider recommendations were assessed by T test.

Results:   At baseline, youth (37% male) were 16.0±4.8 years old with T1D duration 9.2±5.1 years, A1c 9.9±2.0%, and LDL 149±22 mg/dl. Over a mean of 13.2 years of follow-up, providers appropriately noted LDL elevations in 80.6% of encounters when LDL ≥130 mg/dl. In 51 of youth, ≥50% of subsequent LDLs continued to be ≥130 mg/dl.  To lower LDL, providers commonly recommended improving glycemic control (30% of encounters); dietary changes (24% of encounters); enhancing exercise (16% of encounters); and starting, changing, or reinforcing adherence with Rx (11% of encounters).  Only Rx advice yielded improved LDL levels at subsequent visits (mean improvement of 26.9±51.5 mg/dl vs 10.9±35.5 mg/dl in those without Rx, p=.008).  In our sample, 67 individuals (32%) were eventually started on Rx, and 22 (10%) (n=22) refused Rx. Rx was started after a median of 4 elevated LDLs (interquartile range 2-7) and 3.6 years (interquartile range 0.8-8.8 years), and at a mean age of 20.5±4.4 years. After starting Rx, 61% of youth continued to have 50% or more of subsequent LDLs ≥ 130 mg/dl.

Conclusions:  Youth with T1D frequently have hyperlipidemia. Provider recommendations for lifestyle do not produce major improvements in LDL levels. While Rx is effective, initiation is often delayed, titration inadequate, and patient acceptance limited.  Changes in clinical practice are needed to improve management of hyperlipidemia in youth and young adults with T1D.

 

Nothing to Disclose: MLK, GHT, JBC, CED, MD, LML

OR01-3 21870 3.0000 A Under-Management of Hyperlipidemia in Young Persons with Type 1 Diabetes (T1D) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR01 5933 11:30:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Oral


Mohamed I. Husseiny Elsayed*1, Leah Selby2, Alexander Kaye1, Eba Hathout2, Fouad R Kandeel1 and Kevin George Ferreri3
1Diabetes & Metabolic Research Institute, Beckman Research Institute of City of Hope, Duarte, CA, 2Loma Linda University School of Medicine, 3City of Hope, Duarte, CA

 

Type 1 diabetes (T1D) results from the immune-mediated destruction of the insulin-secreting beta cells of the pancreas. Our previous study demonstrated that the DNA encoding the human insulin gene promoter is uniquely unmethylated in beta cells. We developed a methylation-specific PCR (MSP) assay for unmethylated insulin DNA to identify circulating beta cell DNA as a way to measure beta cell death. This study monitored 10 patients with new onset of T1D age 12y and older starting within the first 3 months of diagnosis, with evaluations throughout their first year post-diagnosis focusing on evidence of beta cell loss as well as glycemic control. All T1D patients were diagnosed based on ADA criteria of elevated blood glucose and, HbA1c, as well as the presence of one or more positive autoantibody titers (insulin, GAD65 and IA-2 antibodies). Blood samples were collected at diagnosis, 1, 2, 4, 6, 9, and 12 months post-diagnosis and analyzed by MSP assays. In addition, 90 min stimulated C-peptide level following a mixed-meal tolerance tests (MMTT) were measured at baseline and quarterly to measure of residual beta cell mass. The longitudinal relationship between these metabolic parameters and the appearance of beta cell DNA in circulation were analyzed using the MSP assay. Our results showed that 70% and 80% of patients were positive for GAD65 and IA-2 autoantibodies, respectively. HbA1C ranged between 9.1% and 18.5% at diagnosis and subsequently decreased after initiation of insulin therapy. Stimulated C-peptide levels at diagnosis were very low but increased with meal stimulation once the patient started insulin therapy, but then in most cases declined again over time. The low levels of C-peptide at diagnosis might have been a reflection of the toxicity of hyperglycemia to the beta cells and the subsequent improvement with initiation of insulin therapy and control of hyperglycemia probably reflected improvement of beta cell function with glucose control. In this trial, most of the patients showed a high signal of C-peptide after stimulation but this was insufficient to control hyperglycemia, indicating that patients still had residual partial beta cells function. Using our MSP assay, we found a significantly increased relative unmethylation ratio (RUR) of insulin DNA that is compatible with the known timeline of beta cell death in early onset T1D. We noted a decrease in the RUR following the development of diabetes suggesting the arrest of further beta cell death in the pancreas. The MSP assay raised the possibility that destruction of beta cells may occur as a result of episodic attacks of autoimmunity in T1D. We conclude that these patients were in a “honeymoon period” during which the ongoing beta cell death was delayed. Therefore MSP assay can provide an effective method to monitor beta cell destruction in early T1D and may be useful in tracking established and innovative measures to ameliorate the disease.

 

Nothing to Disclose: MIHE, LS, AK, EH, FRK, KGF

OR01-4 19100 4.0000 A Methylation-Specific PCR Assay for Detection of Beta Cell Death in Newly Onset Type 1 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR01 5933 11:30:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Oral


Ingrid Libman*1, Kellee M Miller2, Linda A DiMeglio3, Kathleen Bethin4, Michelle L Katz5, Avni Shah6, Jill H Simmons7, Michael James Haller8, Sripriya Raman9, William V Tamborlane10, Julie Coffey11, Ashleigh M Saenz2 and Roy W Beck12
1Children's Hospital of Pittsburgh of UPMC, Pittsburgh, PA, 2Jaeb Center for Health Research, Tampa, FL, 3Indiana Univ Sch of Med, Indianapolis, IN, 4University at Buffalo School of Medicine and Biomedical Sciences, Buffalo, NY, 5Joslin Diabetes Center, Boston, MA, 6Stanford University, 7Vanderbilt University Medical Center, Nashville, TN, 8University of Florida, Gainesville, FL, 9Children's Mercy Hospital, Kansas City, MO, 10Yale Univ Sch of Med, New Haven, CT, 11University of Iowa, 12Jaeb Center for Health Research Tampa FL, Tampa, FL

 

Achieving optimal glycemic control is especially challenging in overweight and obese adolescents with T1D, highlighting a need for additional approaches to therapy.  This study was designed to evaluate the efficacy and safety of metformin as adjunct therapy in these difficult to control teenagers who had HbA1c levels above the target range.  

At 26 T1D Exchange pediatric diabetes treatment  centers, 140 adolescents 12-19 years old (mean±SD 15.3 ± 1.7 years ) with T1D duration >1 year (7.0± 3.3 years), body mass index (BMI)  ≥ 85th percentile for age and gender (94± 4th %), total daily insulin dose ≥0.8 units/kg (1.1 ± 0.2 units/kg) and HbA1c 7.5%-9.9% (8.8%± 0.7%) were randomly assigned to either metformin (maximally tolerated dose 1000-2000 mg/day) or placebo for 6 months, in addition to their basal/bolus insulin therapy.  The primary outcome was change in central lab- measured HbA1c after 26 weeks. 

Baseline HbA1c was 8.8% in each group.  At 13 weeks, change in HbA1c from baseline in the metformin group (-0.2%±0.8%) was significantly better than placebo (+0.1%±0.8%, p=0.02). However, this differential effect was not sustained at 26 weeks where mean change in HbA1c from baseline was +0.2%± 0.8 in the metformin group and +0.2%± 0.9 in the placebo group (p=0.92).   At 26 weeks, a 25% or more reduction from baseline in total daily insulin per kg of body weight was achieved by 23% of the metformin group compared with 1% of the placebo group (p=0.003); 24% of the metformin group and 7% of the placebo group had a reduction in BMI z-score >10% from baseline to 26 weeks (p=0.01).    Differences in body composition measures by DEXA scan also favored the metformin group at 26 weeks. More participants in the metformin group reported gastrointestinal adverse events (70% vs. 35%, p<0.001) and at least one severe hypoglycemic event defined as cognitive impairment requiring assistance to treat (8% vs. 0 events; p=0.03).

Six months of adjunctive metformin therapy does not improve glucose control for overweight adolescents with T1D, but does reduce insulin requirements and BMI.

 

Nothing to Disclose: IL, KMM, LAD, KB, MLK, AS, JHS, MJH, SR, WVT, JC, AMS, RWB

OR01-5 19331 5.0000 A Metformin As an Adjunct Therapy Does Not Improve Glycemic Control Among Overweight Adolescents with Type 1 Diabetes (T1D) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR01 5933 11:30:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Oral


Ashu Rastogi*1 and Anil Bhansali2
1Post Graduate Institute of Medical Education and Research, Chandigarh, India, 2PGIMER, Chandigarh, India

 

Context: Diabetic foot osteomyelitis (DFO) and Charcot’s neuroarthropathy (CN) are limb-threatening complications and require quite different treatments. Since the approach to treatment largely determines the outcome, it is important to distinguish each of these entities, early. The clinical and laboratory markers are often nonspecific in the early stages of DFO. MRI for the diagnosis of DFO is relatively sensitive but has poor specificity in the background of CN. 99m Tc  and 111In  labeled leukocyte  scintigraphy have been used to identify DFO but suffer from poor spatial resolution.18F-Flouride is a highly sensitive bone seeking PET tracer and combining 18F-Fluoride PET/CT  (18F- PET) and 18F-FDG labeled autologous leukocyte PET/CT (FDG-LL PET/CT) may accurately diagnose bone infection in presence of acute and/or chronic CN.

Methods:Twenty nine patients with suspected DFO underwent CEMRI, 18F- PET, FDG-LL PET/CT. Patient's plasma was obtained for autologous leukocytes and labelled with 314.5–555 MBq (8.5– 15 mCi) of 18F-FDG. Bone biopsy was considered gold standard for  the diagnosis of DFO. A lesion localized to a bony structure on FDG-LL PET/CT corresponding to clinically suspected site along with 18F-PET/CT uptake was classified as true positive for osteomyelitis, and as a soft-tissue infection if it did not involve the bone. Increased 18F-Flouride uptakes in periarticular areas or underlying multiple bones with or without that of adjacent soft tissues were labeled as acute on CN.

Results: FDG-LL PET/CT showed increased tracer uptake [SUV (max) 8.4±4.7] at clinically involved site in 10 patients who had bone biopsy-proven DFO (true positive). No abnormal labeled leukocyte localization was observed in three patients with biopsy-positive infection (false negative). 10 subjects with suspected osteomyelitis had no abnormal uptake on FDG-LL PET/CT and bone cultures were sterile (true negative). There was no false positive on FDG-LL PET/CT study. CEMRI identified 9 out of 13 cases of osteomyelitis and was false positive in 6 cases. The sensitivity and specificity of concurrent 18F-Flouride and FDG-LL PET/CT is 83.3% and 100% compared to 81.8% and 45.5% of CEMRI, respectively, for the diagnosis of DFO on the background of CN of foot.

Discussion and conclusions: CEMRI had poor specificity for the diagnosis of DFO on background of CN as it relies on identification of marrow edema for diagnosing bone infection, which could be of non-infective etiology like Charcot’s neuroarthropathy.  [18F]-2-fluoro-2-deoxy-D-glucose (FDG)  as the radiotracer  is a non-specific indicator of increased intracellular metabolism compared to 18F-Flouridewhich has affinity for bone. The present study shows high accuracy of 18F-Flouride PET/CT with FDG-LL PET/CT for the diagnosis of osteomyelitis in diabetic CN of foot. Patients showing  PET/CT localization of labeled leukocytes may be spared of bone biopsy.

 

Nothing to Disclose: AR, AB

OR01-6 18487 6.0000 A 18F-Flouride PET/CT and 18F -FDG Labeled Autologous Leukocyte PET/CT for Diagnosis of Osteomyelitis in Diabetic Charcot's Neuroarthropathy of Foot 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR01 5933 11:30:00 AM Clinical Issues in Type 1 and Type 2 Diabetes Oral


Jose Manuel Garcia*1, Jennifer Temel2, David Currow3, Kenneth Fearon4, Ying Yan5, John Friend5 and Amy Abernethy6
1MEDVAMC/Baylor College of Medicine, Houston, TX, 2Massachusetts General Hospital, Boston, MA, 3Flinders University, Adelaide, Australia, 4Western General Hospital, Edinburgh, United Kingdom, 5Helsinn Therapeutics, Inc., Bridgewater, NJ, 6Duke University, Durham, NC

 

Cancer anorexia-cachexia syndrome (CACS) is a multifactorial, debilitating condition common among non-small cell lung cancer (NSCLC) patients. In CACS, altered metabolism and reduced food intake contribute to decreased body weight, mainly through loss of lean body mass (LBM) but also through loss of fat mass (FM). Ghrelin has been shown to activate key pathways in the regulation of body composition, appetite and metabolism. Anamorelin HCl (ANAM) is a novel investigational ghrelin receptor agonist.

Two global, double-blind, Phase III trials (ROMANA 1, NCT01387269, N=484; ROMANA 2, NCT01387282, N=495) assessed ANAM efficacy/safety in patients with unresectable stage III/IV NSCLC and cachexia (≥5% weight loss within prior 6 months or BMI <20 kg/m2). Patients were randomized (2:1) to receive daily oral ANAM (100 mg) or placebo for 12 weeks. Co-primary endpoints were change from baseline over 12 weeks in LBM (measured by DXA) and in hand grip strength (HGS). Secondary endpoints included change from baseline in the anorexia-cachexia domain of the Functional Assessment of Anorexia/Cachexia Treatment (FAACT). Hunger was evaluated using an exploratory Hunger Assessment Scale. Post-hoc analyses evaluated additional body composition parameters from DXA, including total body mass (TBM) and FM.

Over 12 weeks, ANAM significantly increased LBM vs placebo in both studies (p<0.0001). In ROMANA 1, median change in LBM was 1.10 kg (95% CI 0.76; 1.42) for ANAM vs -0.44 kg (95% CI -0.88; 0.20) for placebo; similarly, changes in ROMANA 2 were ANAM 0.75 kg (95% CI 0.51; 1.00) vs placebo -0.96 kg (95% CI -1.27; -0.46). Post-hoc analyses on additional DXA parameters revealed that ANAM significantly increased TBM vs placebo at week 12 in both studies (median changes from baseline of 2.87 vs 0.07 kg, p<0.0001; and 2.04 vs -0.59 kg, p<0.0001). This change was driven by LBM gains, but also included significant increases in FM vs placebo (median changes from baseline of 1.21 vs -0.13 kg, p<0.0001; and 0.77 vs 0.09 kg, p=0.012). Changes in HGS were not statistically different between treatment arms. In both studies, ANAM significantly improved FAACT anorexia/cachexia domain scores vs placebo (mean changes of 4.12±0.8 vs 1.92±0.8; p=0.0004; and 3.48±0.9 vs 1.34±1.0; p=0.0016). Mean changes at week 12 in the Hunger Assessment scores for ANAM vs placebo were 0.93±1.9 vs 0.47±1.8 in ROMANA 1 and 1.00±2.3 vs 0.60±2.4 in ROMANA 2. The most frequent drug-related adverse event (AE) for the ANAM arm in both ROMANA 1 and 2 was hyperglycemia (5.3% and 4.2%), and both studies had few drug-related grade ≥3 AEs in the ANAM arm (0.9% and 2.7% vs 1.2% and 2.5% in the placebo arm).

ANAM for 12 weeks was well tolerated in two global, large-scale, Phase III studies. In both studies ANAM significantly increased LBM and FM, suggesting a restoration of energy balance. Also, ANAM significantly improved patients’ symptoms and concerns related to anorexia-cachexia.

 

Disclosure: JMG: Investigator, Helsinn Therapeutics Inc., Investigator, Aeterna Zentaris Inc. , Investigator, Department of Veterans Affairs , Consultant, Helsinn Therapeutics Inc., Consultant, Aeterna Zentaris Inc. . YY: Employee, Helsinn Therapeutics, Inc.. JF: Employee, Helsinn Therapeutics, Inc.. AA: Director, athenahealth, Inc. , Vice President, Flatiron Health, Inc. , Consultant, ACORN Research, Consultant, Bristol-Myers Squibb, Investigator, Pfizer, Inc., Investigator, Bristol-Myers Squibb, Investigator, Kanglaite, Investigator, Dendreon, Investigator, Helsinn Therapeutics, Inc., Investigator, Celgene, Investigator, GlaxoSmithKline, Investigator, DARA Biosciences, Owner, Advoset LLC , Owner, Orange Leaf Associates LLC. Nothing to Disclose: JT, DC, KF

OR04-1 19188 1.0000 A Analysis of Body Composition Parameters in Patients from ROMANA 1 and 2: Phase III Studies of Anamorelin in NSCLC Patients with Cachexia 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR04 5947 11:30:00 AM GH, Prolactin and Metabolism Oral


Laura E. Dichtel*1, Mette Bjerre2, Miriam A. Bredella1, Anu V. Gerweck3, Ariana D. Riccio3, Jan Frystyk2 and Karen K. Miller1
1Massachusetts General Hospital/Harvard Medical School, Boston, MA, 2Aarhus University, Aarhus, Denmark, 3Massachusetts General Hospital, Boston, MA

 

Obesity is characterized by a relative reduction in GH secretion and serum IGF-1, but normal IGF-1 bioactivity. This suggests that obese individuals have a higher proportion of bioactive to total IGF-1 than their lean counterparts. We investigated whether the preservation of IGF-1 bioactivity in obesity is linked to low GH levels, abdominal adiposity, or insulin resistance.  We also investigated how GH administration, which modulates these factors in different directions, influences absolute and relative IGF-1 bioactivity. Finally, we hypothesized that increases in bioactive IGF-1, not total IGF-1, would drive changes in body composition with GH administration.

We studied 50 women with abdominal obesity before and after randomization to GH (n=28) or placebo (n=22) administration for 3 mos. Bioactive IGF-1 was measured by a kinase receptor activation assay; relative IGF-1 bioactivity was expressed as “% bioactive IGF-1” [(bioactive IGF-1/total IGF-1) x100]. Body composition was assessed by DXA and insulin resistance by OGTT.

Prior to treatment, visceral adipose tissue (VAT, r= -0.39, p=0.006) and insulin resistance (OGTT glucose area under the curve) (r= -0.46, p=0.001) were negative determinants of bioactive IGF-1; peak GH on glucagon stimulation testing was not a determinant. VAT (r=0.29, p=0.04) was a positive determinant of % bioactive IGF-1, demonstrating a relative preservation of IGF-1 bioactivity with visceral adiposity. GH treatment resulted in an increase in total IGF-1 (144 ± 56 to 238 ± 75 ng/mL, p<0.0001), IGF-1 z score (-1.6 ± 0.5 to -0.4 ± 1.1, p <0.0001), absolute IGF-1 bioactivity (1.29 ± 0.39 to 2.60 ± 1.12 ng/mL, p<0.0001), and a trend toward an increase in % bioactive IGF-1 (1.00 ± 0.45 to 1.14 ± 0.43%, p=0.06).  The increase in absolute IGF-1 bioactivity with GH administration predicted an increase in lean mass (r= 0.31, p=0.03), decrease in VAT (r= -0.33, p=0.03) and increase in fasting glucose (r= 0.41, p=0.005). In contrast, there was no association between increase in total IGF-1 and any variable. The increase in % bioactive IGF-1 with GH treatment was associated with an increase in insulin resistance [OGTT 2H glucose (r= 0.38, p=0.05)] and a decrease in trunk fat (r= -0.54, 0.004) over 3 mos. 

In conclusion, bioactive IGF-1 is inversely associated with abdominal fat and insulin resistance in obese women. Percent bioactive IGF-1 is higher in obese women with more VAT, driven by lower total IGF-1. With GH administration, increases in absolute IGF-1 bioactivity, but not total IGF-1, predict changes in body composition. Moreover, % bioactive IGF-1 increases in parallel with increasing insulin resistance, despite a reduction in abdominal fat. Our data suggest that GH treatment increases absolute bioactive IGF-1 and % IGF-1 bioactivity in parallel with increased insulin resistance. In turn, GH may modulate body composition in obesity through increases in bioactive, not total, IGF-1.

 

Nothing to Disclose: LED, MB, MAB, AVG, ADR, JF, KKM

OR04-2 18811 2.0000 A The Effect of Growth Hormone (GH) on Bioactive IGF-I in Obese Women 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR04 5947 11:30:00 AM GH, Prolactin and Metabolism Oral


Edward Owen List*, Darlene E Berryman, Kevin Ray Funk, Jesse Kowalski, Ross Comisford, Mathew Buckman, Miranda Vandargriff and John J Kopchick
Ohio University, Athens, OH

 

Growth hormone receptor gene disrupted (GHR-/-) mice have proven useful for determining numerous actions of GH; however, determining the effects of GH on specific tissues in vivo have been challenging.  Fortuitously, the development of the Cre/Lox system in mice permits individual tissues to be targeted for gene disruption.  Thus, several lines of tissue-specific GHR gene disrupted mice have recently been generated by our laboratory and others in order to better characterize the effects of GH specifically on individual tissues.  Currently, eleven published studies have utilized seven different tissue-specific GHR knockout lines.  Of these, five distinct tissues/cell types have been targeted (adipose tissue, liver, muscle, macrophages and beta cells). From the eleven published studies, only three have compared results in both males and females and all of these were done in liver-specific GHRKO mice, resulting in a lack of data regarding the influence of sex on tissue-specific GHR gene disruption in mice.  To this end, muscle-specific GHR gene disrupted mice have been produced in two separate laboratories; however, both research groups have limited their research to male mice exclusively. As a consequence, nothing is known about the muscle-specific effects of GHR gene disruption in females.  Thus, in the current study we examine the effects of muscle-specific GHR gene disruption (MuGHRKO) on glucose metabolism in both male and female mice. We also for the first time in any of the tissue-specific GHR mouse lines report longevity and cause of death.  As expected, our results reveal that male MuGHRKO mice have improved fasting blood glucose, insulin, c-peptide, and glucose tolerance.  However, we discovered that female MuGHRKO mice show no improvement in any of these measures of glucose metabolism. Since improved glucose metabolism is thought to be key mechanism by which global GHR-/- mice are extremely long lived, we expected that male MuGHRKO mice would also have enhanced longevity. However, mean and median lifespan as well as cause of death in MuGHRKO mice did not differ from controls for either sex.  In conclusion, the data from the current study highlight the importance of using both sexes when performing studies in vivo and suggest that females are resistant to improved glucose metabolism when GH action is disrupted specifically in muscle. In addition, improved glucose metabolism in skeletal muscle devoid of GH action as seen in male MuGHRKO mice is not sufficient to increase lifespan. These data suggest that extended longevity of global GHR-/- mice does not appear to be mediated by direct action of GH on muscle.

 

Nothing to Disclose: EOL, DEB, KRF, JK, RC, MB, MV, JJK

OR04-3 20427 3.0000 A Improvements to Glucose Metabolism in Muscle-Specific Growth Hormone Receptor Gene Disrupted Mice Are Sex-Specific and Do Not Result in Extended Longevity 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR04 5947 11:30:00 AM GH, Prolactin and Metabolism Oral


Guillaume Osterstock1, Paul R. Le Tissier*2, Violeta Mitutsova1, Alexander Barre1, Manon Granier1, Pierre Fontanaud3, Marine Chazalon1, Danielle Carmignac4, Patrice E Mollard3, Iain Robinson4, Malcolm James Low5, Nikolaus Plesnila6, David Jonathan Hodson7, Chrystel Lafont3, Patrick Samper1 and Pierre-François Méry1
1Institute of Functional Genomics, Montpellier, France, 2University of Edinburgh, Edinburgh, United Kingdom, 3IGF-CNRS, Montpellier, France, 4MRC National Institute for Medical Research, London, United Kingdom, 5University of Michigan Medical School, Ann Arbor, MI, 6Royal College of Surgeons in Ireland, Dublin, Ireland, 7Imperial College, London, United Kingdom

 

Subsets of hypothalamic neurons generate highly-ordered neurohormone pulses to properly regulate basic body functions such as growth and metabolism. However, the electrical origin of the hypothalamic pulse generator has so far remained elusive. In the present study, we show that the inhibitory hypothalamic neuropeptide somatostatin (SST) triggered patterned growth hormone-releasing hormone (GHRH) neuron firing when tonically applied to acute brain slices. These firing patterns were fine-tuned by an initial inhibitory action at the GHRH neuron level due to K+ channel (GIRK) activation, followed by a delayed, sst1/sst2 receptor-dependent unbalancing of glutamatergic and GABAergic inputs. Thus, two intermingled feedforward loops trigger oscillatory firing patterns in hypothalamic (GHRH) neurons. To determine the significance for pituitary GH release, we recreated the patterned GHRH neuron spike firing by direct expression of channelrhodopsin2 (ChR2) specifically in GHRH neurons. This was achieved using a novel GHRH-Cre transgenic mouse model in combination with R26-loxSTOPlox-ChR2-tdTomato and direct in vivo manipulation via a fibre optic to the thinned palate bone.  Application of light pulse trains (5 min ON, 3 min OFF) resulted in long-lasting rises in GH, which contrast with application of a sustained (20-30 min) train of light pulses resulting in a single GH pulse that usually peaks before stimulus termination. Thus, a simple tripartite neuronal circuit allows GHRH neurons to deliver a patterned spike firing code, capable of  stimulating release of a robust and sustained GH pulse into the bloodstream. Such neuronal circuits may represent a basic building block which recurs throughout the hypothalamus to finely adjust neuroendocrine activity in response to physiological demand.

 

Nothing to Disclose: GO, PRL, VM, AB, MG, PF, MC, DC, PEM, IR, MJL, NP, DJH, CL, PS, PFM

OR04-4 20377 4.0000 A Hypothalamic Neuroendocrine Feedforward Loops Generate Patterned GHRH Output Regulating GH Secretion 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR04 5947 11:30:00 AM GH, Prolactin and Metabolism Oral


Ronadip R Banerjee*1, Holly A Cyphert2, Xueying Gu1, Roland W. Stein2 and Seung K Kim3
1Stanford University School of Medicine, Stanford, CA, 2Vanderbilt University Medical Center, Nashville, TN, 3Stanford University School of Medicine, HHMI, Stanford, CA

 

Prolactin and placental lactogen signaling through the prolactin receptor (PRLR) are required for the adaptive maternal response to pregnancy.  Lactogens are critical for the expansion of pancreatic beta cell mass necessary to maintain glucose homeostasis in response to increased maternal-fetal metabolic demands.  However, the molecular mechanisms underlying lactogen-induced changes within islets during pregnancy remain poorly understood (1).  As whole-animal PRLR knockout mice (PRLRnull) are infertile, precluding pregnancy studies (2), we have generated a novel conditional loxP allele of PRLR and deleted PRLR specifically within pancreatic beta-cells using RIP-Cre (BPRLRKO mice).  Surprisingly, BPRLRKO mice exhibit no baseline abnormalities in glucose homeostasis, suggesting that the islet defects described in PRLRnull mice are a consequence of PRLR deletion in other tissues (3).  BPRLRKO mice are fertile, and develop gestational diabetes that resolves postpartum but recurs and worsens in a second pregnancy.   At late gestation, BPRLRKO mice show reduced beta-cell proliferation and a 2.5 fold reduction in beta-cell mass compared to littermate controls.  To identify molecular mediators of defective proliferation and beta cell mass expansion we examined gene expression in islets isolated from pregnant BPRLRKO females and controls.  We found BPRLRKO mice failed to induce cyclins A2 and B1 as well as the transcription factor FoxM1, a key regulator of cell cycle progression and proliferation in islets.  Furthermore, BPRLRKO mice did not induce tryptophan hydroxylase 1 (Tph1) expression and lacked the pregnancy-associated increase in islet serotonin, a critical paracrine-autocrine signal promoting beta-cell proliferation (4). Additionally, the transcription factor MafB is normally expressed in a subset of beta cells during pregnancy; the MafB+Ins+ population of beta cells is significantly reduced in pregnant BPRLRKO mice.  To our knowledge, this is the first evidence linking prolactin signaling and regulation of MafB expression.  Taken together, our results demonstrate the requirement for PRLR signaling in the physiologic expansion of beta cells during pregnancy, and identify key mediators of proliferation induced by prolactin signaling.  We also anticipate that the development of the floxed PRLR mouse will allow conditional gene targeting analysis of PRLR signaling and facilitate our understanding of lactogenic hormone action in specific tissues and disease states.

 

Nothing to Disclose: RRB, HAC, XG, RWS, SKK

OR04-5 20102 5.0000 A Loss of Prolactin Receptor Signaling in Pancreatic Beta Cells Causes Gestational Diabetes Mellitus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR04 5947 11:30:00 AM GH, Prolactin and Metabolism Oral


Ying Liu*1, Yue Zhang1, Jing Jiang1, John Fairbanks Langenheim2, Wen Y Chen2, Kurt R. Zinn3 and Stuart J Frank1
1University of Alabama at Birmingham, Birmingham, AL, 2Clemson University, Greenville, SC, 3Univ of Alabama at Birmingham, Birmingham, AL

 

Growth hormone receptor (GHR) and prolactin receptor (PRLR) are transmembrane glycoproteins cytokine receptors that are structurally homologous and activate JAK2-STAT signaling pathways. We previously detected association between GHR and PRLR in T47D human breast cancer cells by coimmunoprecipitation, suggesting that, in addition to forming homodimers, these related receptors may form hetero-assemblages that may have functional consequences. Adding to the potential richness GH and PRL physiology, human GH can activate either GHR or PRLR, while PRL activates only PRLR. We recently reported (1) that the GHR-GHR association can be detected specifically using the split luciferase complementation assay in which coexpressed chimeras encoding GHR fused to Nluc (residues 1-398 of firefly luciferase) and GHR fused to Cluc (luciferase residues 394-550), by virtue of their physical proximity, reconstitute luciferase activity which is detected by bioluminescence imaging and quantitation. Using this assay, we showed that GH acutely augments this bioluminescence, consistent with the induction of conformational changes that bring GHR cytoplasmic tails closer. In the current study, we examined human GHR-Nluc/PRLR-Cluc complementation in a GHR- and PRLR-deficient human fibrosarcoma cell line. We verified expression and signaling capacity of each chimera individually and then found that GHR-Nluc/PRLR-Cluc coexpression yielded specific complementation that was not observed when either chimera was expressed with an erythropoietin receptor-luc fragment chimera. Notably, this basal complementation was monotonically decreased by ~30% by GH treatment over a 40 min period. This GH-induced decrease of complementation was partially inhibited by a GHR-specific antagonist (B2036) alone or a PRLR-specific antagonist (G129R) alone but completely inhibited by a combined antagonist (B2036+G129R) treatment, suggesting GH engages both GHR and PRLR to exert its effect. In contrast, a unique GHR-PRLR heteromer agonist, B2036-G129R (B-G; B2036 and G129R fused in tandem with single binding sites for GHR and PRLR, respectively (2)), augmented GHR-Nluc/PRLR-Cluc complementation. Studies are underway to more fully characterize GHR-PRLR hetero-assemblages using the split luciferase complementation assay, in concert with biochemical and signaling techniques.

 

Nothing to Disclose: YL, YZ, JJ, JFL, WYC, KRZ, SJF

OR04-6 20106 6.0000 A Investigation of Growth Hormone Receptor / Prolactin Receptor Hetero-Assemblage By Split Luciferase Complementation 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR04 5947 11:30:00 AM GH, Prolactin and Metabolism Oral


Amanda Leigh Herberger*1, Hanson Ho2, Alfred Li1, Christian Yuzon Santa Maria2, Jenna Hwong1, Zhiqiang Cheng1, Chia-Ling Tu1 and Wenhan Chang1
1University of California San Francisco, San Francisco, CA, 2University of California, San Francisco, San Francisco, CA

 

Previous observations demonstrated that heteromeric complex formation of the calcium-sensing receptor (CaSR) and the type B g-aminobutyric acid (GABA) receptor 1 (GABABR1) mutually suppressed the expression and signaling ability of the receptors.  Heterozygous (PTCCaSRwt/flox) and homozygous (PTCCaSRflox/flox) knockout (KO) of the Casr gene targeted to parathyroid cells (PTCs) specifically produced mild and severe hyperparathyroidism (HPT) in mice, respectively. The PTCCaSRwt/flox mice were viable, but PTCCaSRflox/flox mice died before 3-4 weeks of age.  The 3-month-old PTCCaSRwt/flox and 3-week-old PTCCaSRflox/flox mice had significantly elevated serum PTH (sPTH, 219±19 and 959±122 pg/ml, respectively) and calcium (sCa, 10.91±0.15 and 16.44±0.05 mg/dL, respectively) levels when compared to 3-month-old control littermates (sPTH: 172±10 pg/ml; sCa: 9.61±0.06 mg/dL, p<0.01), recapitulating phenotypes of human Familial Hypocalciuric Hypercalcemia (FHH) and neonatal severe HPT (NSHPT), respectively.  To test whether the expression and signaling of GABABR1 critically mediates PTH secretion and alters the Ca2+-responsiveness of PTCs at basal and at HPT states, we compared sCa2+ and sPTH levels in the heterozygous and homozygous CaSR KO mice to PTCGABABR1flox/flox mice, which have both alleles of Gabbr1 genes deleted in PTCs, and to double knockout (PTCCaSRwt/flox//PTCGABABR1flox/flox or PTCCaSRflox/flox//PTCGABABR1flox/flox) mice, and compared the secretory capacity and Ca2+-set-point of parathyroid glands (PTGs) acutely cultured from these mice. Interestingly, the PTCGABABR1flox/flox mice showed reduced sPTH (115±8 pg/ml, p<0.01) and sCa (9.42±0.04 mg/dL, p<0.01) levels. Furthermore, concurrent GABABR1 KO rescued the early lethality of PTCCaSRflox/flox mice and suppressed the sPTH (659±117 pg/ml, p<0.01) and sCa (12.12±0.44 mg/dL, p<0.01) levels in the PTCCaSRflox/flox//PTCGABABR1flox/flox mice when compared to those in PTCCaSRflox/flox mice.  Similarly, GABABR1 KO suppressed the sPTH (130±24 pg/ml, p<0.01) and sCa (10.0±0.24 mg/dL, p<0.01) levels in the PTCCaSRwt/flox//PTCGABABR1flox/flox vs PTCCaSRwt/flox mice. Studies of PTGs cultured from the PTCCaSRwt/flox mice showed a profound increase in the maximal PTH secretion (3-4 fold) and a right-shift in Ca2+-set-point from 1.3 to 1.5 mM vs control PTGs, but these effects were completely abrogated by concurrent ablation of the Gabbr1 gene in the PTGs of the PTCCaSRwt/flox//PTCGABABR1flox/flox mice. Our data supports the concept that the parathyroid GABABR1 critically controls mineral homeostasis at basal and at HPT states by sustaining tonic PTH secretion and Ca2+-responsiveness of PTCs and may be the molecular basis for dysregulated PTC functions in human FHH and NSHPT.

 

Nothing to Disclose: ALH, HH, AL, CYS, JH, ZC, CLT, WC

OR08-1 20528 1.0000 A Parathyroid GABABR1 Is the Molecular Basis for PTH Hypersecretion and a Right-Shift in Ca2+ Set-Point in Conditions of Mild and Severe Hyperparathyroidism 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM OR08 5963 11:30:00 AM Parathyroid and Bone Metabolism Oral


Ruiye Bi*1, Tomoyuki Watanabe2, Yi Fan3, Thomas J Gardella2 and Michael Mannstadt1
1Massachusetts General Hospital and Harvard Medical School, Boston, MA, 2Massachusetts General Hospital, Boston, MA, 3Harvard School of Dental Medicine, Boston, MA

 

Hypoparathyroidism (HP) is characterized by inadequate levels of PTH, hypocalcemia, hyperphosphatemia and low bone turnover, and is most commonly caused by damage or removal of the parathyroid glands during neck surgery. Current treatment is limited to oral calcium and active vitamin D, which can lead to hypercalciuria and renal complications. Novel PTH analogs that can mediate sustained normalization of blood and urine calcium are now being explored as potential replacement therapies.
Evaluation of such therapies in vivo requires an effective animal model of HP.  Surgical parathyroidectomy is a proven animal model of acquired HP, but PTX surgery in mice is challenging due to small gland size and variable gland location.  To facilitate this procedure, we developed a mouse model in which the parathyroid gland cells have been engineered to express green fluorescent protein. In these "PTH-GFP" mice, which were derived by crossing PTH-Cre mice with mTmGFPfl/fl mice, non-parathyroid cells fluoresce red while the parathyroids fluoresce green. The glands are thus easily identified, and PTX surgery, as guided by GFP, is greatly facilitated (surgery times < 20 minutes/mouse and no mortality in 80 PTX mice so far). Blood analyses revealed that three days after surgery, GFP-PTX mice, as compared to sham surgery mice, exhibited a dramatic decrease in serum PTH (31 ± 23 vs. 570 ± 57 pg/mL; n=6 per group; p<0.001), low blood Ca++ (1.05 ± 0.05 vs. 1.31 ± 0.04 mmol/L; n=30; p<0.001) and elevated serum phosphorus (9.5 ± 1.0 vs. 7.6 ± 1.8; n=22-24). This hypoparathyroid phenotype remained stable over a three-month observation period.
We then used the model to evaluate the efficacy of LA-PTH, a new long-acting PTH analog, for normalizing blood Ca in conditions of HP. Single s.c. injection of LA-PTH at 1.5, 3, 5, 10, or 20 nmol/kg into GFP-PTX mice led to a dose-dependent increase in blood Ca++ (1.21 ± 0.02, 1.23 ± 0.04, 1.23 ± 0.06, 1.25 ± 0.06, 1.26 ± 0.05, 1.26 ± 0.06 mmol/L; n=5-6 per group).  LA-PTH was about 5-fold more potent on a molar basis than PTH(1-34) (dose=25, 50 nmol/kg; Ca++=1.21 ± 0.05, 1.27 ± 0.09 mmol/L; n≥5). Importantly, a single injection of LA-PTH was able to normalize serum calcium and phosphate levels for more than 72 hours.
These results thus show that GFP-PTX mice provide a novel model of acquired HP, and they further highlight the potential utility of novel long-acting PTH analogs as therapies for this disease.

 

Nothing to Disclose: RB, TW, YF, TJG, MM

OR08-2 21073 2.0000 A A Novel Mouse Model for Acquired Hypoparathyroidism (HP) and Its Application for Studying Long-Acting Parathyroid Hormone (LA-PTH) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM OR08 5963 11:30:00 AM Parathyroid and Bone Metabolism Oral


Hila Bahar*1, Nancy Doyle2, Aurore Varela2, Luc Chouinard3, Elisabeth Lesage2, Robert Guldberg4, Susan Y Smith2 and Gary Hattersley1
1Radius Health, Inc, Waltham, MA, 2Charles River Laboratories, Senneville, Canada, 3Charles River, 4School of Mechanical Engineering, Georgia Institute of Technology

 

Abaloparatide (ABL) is a PTHrP(1-34) analog being developed for the treatment of osteoporosis. In a completed phase 2 clinical study in postmenopausal women with osteoporosis, a high overall BMD responder rate at the lumbar spine, total hip and femoral neck, was observed after 24 weeks of ABL treatment compared to either teriparatide or placebo treatment. We have investigated the mechanisms by which ABL results in BMD gains in aged, osteopenic, ovariectomized (OVX) rats and monkeys. Three months after surgery, sham and osteopenic OVX rats (n=18 per group) were treated daily with vehicle or ABL (1, 5 or 25 μg/kg sc) for 12 months. Treatment with ABL 25 μg/kg significantly increased serum levels of the bone formation markers osteocalcin (OC) and procollagen type I N-terminal propeptide (PINP) after 3 months, which remained elevated throughout the study. In contrast, the bone resorption markers, C-telopeptide (CTX) and deoxypyridinoline (DPD), were not affected throughout the course of the study by treatment with ABL at any dose level. Consistent with these findings, dynamic histomorphometric analysis showed that ABL treatment resulted in linear, dose-dependent significant increases in bone formation parameters without modifying the bone resorption parameters in trabecular bone at the lumbar spine and proximal tibia, and in femur cortical bone, relative to OVX and sham vehicle controls. Following a 9-month bone depletion period, ≥9 years-old sham and osteopenic OVX cynomolgus monkeys (n=16 per group) were treated daily with vehicle or ABL (0.2, 1 or 5 μg/kg, sc) for 16 months. Serum levels of the bone formation marker PINP increased rapidly and remained elevated throughout the study in ABL treated monkeys compared to OVX controls. In contrast, the bone resorption markers CTX and N-telopeptide (NTX) were comparable to OVX controls in ABL treated monkeys. Consistent with these findings, marked gains in BMD were observed at the femoral neck for all dose levels of ABL as early as 4 months of treatment, and were comparable to or above pre-surgery levels. Dynamic histomorphometric analysis indicated a dose dependent increase in bone formation parameters but absence of meaningful changes in bone resorption parameters. In addition, significant increases in femur diaphysis endocortical BFR (Ec.BFR/BS) were noted with ABL 5 μg/kg. Importantly, μCT analysis of humerus cortical beams showed that cortical bone porosity was not affected by ABL treatment at any dose level, compared to OVX control. Thus, the data suggests that ABL exerts a bone anabolic action at least in part due to significant and early bone formation without a corresponding increases in bone resorption in rats and monkeys.

 

Disclosure: HB: Employee, Nuvios, Inc., Employee, Nuvios, Inc.. GH: Employee, Nuvios, Inc., Employee, Nuvios, Inc.. Nothing to Disclose: ND, AV, LC, EL, RG, SYS

OR08-3 21376 3.0000 A Abaloparatide Treatment Increases Bone Formation without a Corresponding Increases in Bone Resorption Resulting in Marked Bone Gains in Osteopenic Ovariectomized Monkeys and Rats 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM OR08 5963 11:30:00 AM Parathyroid and Bone Metabolism Oral


Joshua Nicholas Farr*, Matthew M Roforth, Koji Fujita, Elizabeth J Atkinson, Louise K McCready, James M Peterson, Matthew T Drake, David G Monroe and Sundeep Khosla
Mayo Clinic, Rochester, MN

 

Precise delineation of the specific pathways and genes altered with aging and estrogen (E) deficiency may lead to the development of novel therapeutics to prevent or reverse age-related bone loss.  Further, such studies may identify new skeletal biomarkers.  At present, knowledge of factors relevant to bone metabolism is largely based on rodent models, although gene expression changes in rodents likely do not completely recapitulate those in humans.  The main limitation of human bone studies to date, however, is that only pre-specified genes and pathways have been examined.  High-throughput RNA sequencing (RNAseq), on the other hand, offers an unbiased approach to unveil the entire genome.

Here we present an RNAseq analysis of human bone samples, obtained from iliac crest needle biopsies, to yield the first in vivo interrogation of all potential genes and pathways that may be altered in bone with aging and E deficiency in humans.  60 healthy women were studied, including 20 young women (mean age ± SD, 30.0 ± 5.4 yrs) as well as 40 old women (n = 20 per group) randomized to receive either no therapy (72.9 ± 6.5 yrs) or 3 wks of short-term estrogen (STE) therapy (70.5 ± 5.2 yrs).  Using fairly stringent criteria (median count ≥ 10, P ≤ 0.05, false discovery rate [q] ≤ 0.10), 678 genes were altered with aging including 446 up- and 232 down-regulated genes, respectively (in old vs young subjects).  In bone tissue, aging significantly altered a total of 60 pathways (all P < 0.05), including a subset known to regulate bone metabolism such as Notch, PTEN, HIF1α, and Wnt/β-catenin signaling.  Interestingly, a large number of the identified pathways and genes were previously unrecognized as changing in bone with aging.  Perhaps unexpectedly, many of these pathways and genes have critical functions in other tissues and disease processes, although their role(s) in the pathogenesis of skeletal fragility requires additional investigation.  Notably, SOST levels did not change with aging, but STE therapy tended to lower SOST levels in bone (P < 0.001; q = 0.46).  Importantly, these changes in gene expression mirrored findings in serum (using the MSD sclerostin assay).  Further, in old women, of the 21 unique genes altered in bone by STE, the expression of INHBB (encoding for activin), which markedly decreases with aging (P < 0.001, q < 0.001), is restored to young adult levels in a robust manner (P < 0.001, q < 0.001) in response to E therapy.

In conclusion, our findings demonstrate that aging alters a substantial portion of the skeletal transcriptome, whereas STE therapy appears to have significant, albeit less wide-ranging effects.  To date, our results provide the most comprehensive assessment of the effects of aging and E therapy on the skeleton’s gene expression profile, and serve as a valuable resource for the identification of novel biomarkers associated with age-related bone loss to better target therapies to specific patients.

 

Nothing to Disclose: JNF, MMR, KF, EJA, LKM, JMP, MTD, DGM, SK

OR08-4 20632 4.0000 A Skeletal Biomarkers Associated with Aging and Estrogen Deficiency Revealed By RNA Sequencing 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM OR08 5963 11:30:00 AM Parathyroid and Bone Metabolism Oral


Serra Ucer1, Srividhya Iyer1, Shoshana M Bartell1, Marta Martin-Millan2, Li Han1, Ha-Neui Kim1, Charles A O'Brien1, Maria S Almeida1 and Stavros C Manolagas*3
1University of Arkansas for Med Science/Central Arkansas Veterans Healthcare System, Little Rock, AR, 2University of Cantabria, Santander, Spain, Santander, Spain, 3University of Arkansas for Medical Sciences/Central Arkansas Veterans Healthcare System, Little Rock, AR

 

In men, androgens are critical for the acquisition of bone mass in both the cortical and cancellous bone compartment during growth and for its maintenance throughout life. Consistent with this, male mice with global deletion of the androgen receptor (AR) exhibit low cortical and cancellous bone mass. Nevertheless, male mice with targeted deletion of AR in osteoblasts or osteocytes have lower cancellous bone mass, but no cortical bone phenotype. We have investigated the possibility that the effects of androgens on the cortical compartment result from AR signaling in osteo-progenitors or cells of the osteoclast lineage; or via ERα signaling in either or both of these two cell types upon local conversion of testosterone to estradiol. To this end, we generated mice with targeted deletion of an AR or ERα allele in the mesenchymal (ARflox;Prx1-Cre or ERαflox;Osx1-Cre) or myeloid cell lineage (ARflox;Lysm-Cre or ERαflox;Lysm-Cre) and their descendants. Male ARflox;Prx1-Cre mice exhibited decreased cancellous bone volume and trabecular number (at both 12 and 26 weeks of age), and increased osteoclast number; cortical thickness, on the other hand, was unaffected. Additionally, ARflox;Prx1-Cre mice did not undergo the loss of cancellous bone volume and trabecular number caused by orchidectomy (ORX) in their littermate controls. Nonetheless, control and ARflox;Prx1-Cre male mice lost similar amount of cortical bone following ORX. ERαflox;Osx1-Cre male mice, in difference to their female littermates, had only a transient low bone mass phenotype in the cortex at 6 weeks of age, but that was normalized by 10 weeks of age; and, following ORX lost cortical and cancellous bone indistinguishably from their littermate controls. Finally, ERαflox;Lysm-Cre males had indistinguishable bone phenotype from their littermate controls, including osteoclast numbers in cancellous or cortical bone. Recapitulation of the effects of ORX by AR deletion in the osteoblast lineage demonstrates that the protective effects of androgen on cancellous bone result from AR-mediated effect of androgen in osteoblast progenitors or their descendants.  The protective effects of androgens on cortical bone mass, on the other hand, do not require AR or ERα signaling in any cell type across the osteoblast or osteoclast differentiation lineage; thus, must be exerted indirectly by actions on some other cell type(s). These results also reveal that as in the case of estrogens (Manolagas et al, Nat Rev Endocrinol., 9:699-712, 2013), the cellular targets of androgen action on cancellous and cortical bone are different; and that the cellular targets of estrogen in bone in the female are different than they are in the male.

 

Nothing to Disclose: SU, SI, SMB, MM, LH, HNK, CAO, MSA, SCM

OR08-5 21192 5.0000 A The Effects of Androgens on Cancellous Bone Require Androgen Receptor Signaling in Osteoblasts, but Not Osteoclasts; And in Cortical Bone Require Neither 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM OR08 5963 11:30:00 AM Parathyroid and Bone Metabolism Oral


Jonathan A. Mitchell*1, Alessandra Chesi2, Okan Elci3, Shana E. McCormack2, Sani M. Roy2, Heidi J. Kalkwarf4, Joan M. Lappe5, Vicente Gilsanz6, Sharon E Oberfield7, John A. Shepherd8, Andrea Kelly9, Babette S. Zemel2 and Struan F.A. Grant3
1The University of Pennsylvania, 2The Children's Hospital of Philadelphia, Philadelphia, PA, 3The Children's Hospital of Philadelphia, 4Cincinnati Children's Hospital Medical Center, Cincinnati, OH, 5Creighton University, Omaha, NE, 6Children's Hospital Los Ange, Los Angeles, CA, 7Columbia University College of Physicians and Surgeons, New York, NY, 8University of California San Francisco, San Francisco, CA, 9Children's Hospital of Philadelphia, Philadelphia, PA

 

Background: Maximizing peak bone mass (PBM) in early life is an important strategy to reduce the risk of osteoporosis later in adulthood. Both genetics and physical activity (PA) are known to contribute to PBM. However, it is not known if all children experience increases in bone mineral density (BMD) in response to PA.

Objective: To determine if known GWAS-implicated adult bone density loci interact with PA to modify the association between PA and BMD and bone mineral content (BMC) in childhood.

Design/Methods: We used a six-year prospective observational study design and all participants were of European ancestry, enrolled in the Bone Mineral Density in Childhood Study (N=689, 52.4% female). The following outcomes were estimated by dual energy X-ray absorptiometry: femoral neck (FN), total hip (TH) and spine areal BMD (aBMD), and total body less head (TBLH) BMC, Z-scores corrected for height and age. DNA was extracted from blood/saliva and genotyped using high-throughput technology; we investigated 77 single nucleotide polymorphisms (SNPs) near adult BMD susceptibility loci identified through GWAS. The participants self-reported their PA levels (hr/d) and trained clinicians measured puberty stage (Tanner I, II-IV and V). Interaction effects of PA with each SNP and with both puberty stage and each SNP were examined by sex using linear mixed models. Age, BMI, and dietary calcium were adjusted in these models.

Results: PA exhibited statistical interactions with 30 loci (P<0.05) and 20 loci exhibited statistical interactions with both PA and puberty stage (three-way interaction, P<0.05) to influence aBMD/BMC. For example, SNP rs4283041 near ERC1/WNT5B interacted with PA to influence TBLH-BMC in males (P=1.1x10-4); each hr/d of PA was associated with higher TBLH-BMC Z-score for CC homozygotes (b=0.04, P=3x10-7); similar findings were observed for spine, TH and FN aBMD.  SNP rs7195617 near AXIN1 interacted with PA to influence total hip aBMD in females (P=3.9x10-4); each hr/d of PA was associated with higher TH aBMD Z-score for females carrying one or more G alleles (GG homozygotes: b=0.06, P=4x10-5); similar findings were observed for FN and spine aBMD. The strongest three-way interactions were observed for spine aBMD in males (rs17691610 near MAPT, P=3.6x10-4) and in females (rs7304170 near KLHDC5/PTHLH, P=4.3x10-4). In females, carrying one or more rs7304170 C alleles was associated with increased spine aBMD during Tanner V (CC homozygotes: b=0.15, P=0.002).

Conclusions: PA was associated with higher aBMD/BMC, but this association was dependent on genetic variation at certain known bone density loci. Our data suggests that not all childhood skeletons respond equally to PA. These findings are particularly important for children born into families with a history of osteoporosis; increasing PA in childhood may not always be an efficacious approach to help prevent later osteoporosis.

 

Nothing to Disclose: JAM, AC, OE, SEM, SMR, HJK, JML, VG, SEO, JAS, AK, BSZ, SFAG

OR08-6 21496 6.0000 A The Skeletal Benefits of Physical Activity in Childhood Are Dependent on Genetic Variation at Known Bone Density Loci 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM OR08 5963 11:30:00 AM Parathyroid and Bone Metabolism Oral


Sudheer Kumar Gara*, Yonghong Wang, Dhaval Patel, Yi Liu-Chittenden, Myriem Boufraqech, Paul S Meltzer and Electron Kebebew
National Cancer Institute, NIH, Bethesda, MD

 

Very little is known about the pathogenesis of adrenocortical carcinoma (ACC) and whether the “multi-hit” hypothesis of cancer is true in ACC (progression from normal-to-adenoma-to-carcinoma). The objectives of this study were to investigate the molecular changes associated with normal-to-adenoma-to-carcinoma, and to determine possible new molecular pathways that may lead to ACC initiation and/or progression. We performed genome-wide gene expression analysis in adrenocortical tissue (normal, cortical adenomas, and ACC) samples. Selected samples from the same cohort were subjected to genome-wide DNA methylation array, microRNA expression array, and comparative genomic hybridization (CGH) analysis. We then performed an integrated analysis by a paired-wise comparison (normal-adenoma, ACC-adenoma and ACC-normal) of gene expression profile and selected genes with 4-fold expression differences and an adjusted p<0.05. We found very small differences in the gene expression profile of normal vs. adrenocortical adenoma while the expression profile was markedly different for ACC vs. adrenocortical adenoma and normal adrenal cortex. The majority of the dysregulated genes in ACC were down-regulated. Integration of CGH data with the gene expression showed that 7% and 11% of the differentially expressed genes had copy number alterations in ACC vs. adrenocortical adenoma and normal, respectively. We found the chromosomal loci spanning CDKN1C and GPM6A, a novel potential tumor suppressor candidate gene was frequently deleted in ACC. Similarly, aberrant methylation of dysregulated genes was present in 7% and 10% of ACC vs. normal and adrenocortical adenoma comparisons, respectively. Several genes including CYP1B1 and TBC1D4 were found to be differentially methylated and downregulated in ACC. We identified 14% and 15% of dysregulated genes were targeted by differentially expressed microRNAs and were inversely correlated in ACC vs. adenoma and ACC vs. normal, respectively. We found that miR-9, miR-25, miR-124 and miR-206 had multiple targets and also genes such as ASPH, FOXO1 and MBNL1 are targeted by more than one miRNAs indicating a tight gene regulation. Collectively, these three mechanisms account 28% and 36% of gene dysregulation in ACC vs adenoma and normal respectively. In addition to the well-known altered pathways in ACC, such as IGF signaling and β-catenin, we discovered Oncostatin, EIF2, CDC42 and RXR signaling pathways to be significantly altered and regulated by at least one of these mechanisms. Our data suggest that adrenocortical tumorigenesis is a multi-step process with cumulative epigenetic and copy number changes altering gene expression from normal-to-adenoma-to-carcinoma. In addition, we have identified novel tumor suppressor genes, molecular pathways and the mechanisms associated with ACC.

 

Nothing to Disclose: SKG, YW, DP, YL, MB, PSM, EK

OR02-1 19384 1.0000 A Integration of Whole Genome Wide Approaches to the Analysis of Gene Regulation and Progression of Adrenocortical Carcinoma 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 1:00:00 PM OR02 5972 11:30:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Oral


Anastasia Melin*1, Achim Starke1, Aycan Akca1, Peter Goretzki1, Rajani Maharjan2, Per Hellman2 and Peyman Björklund3
1Lukaskrankenhaus GmbH Neuss, Neuss, Germany, 2Uppsala University, Sweden, 3Uppsala University, Uppsala, Sweden

 

Introduction:

Endocrine pancreatic tumors (p-NETs) occur with an incidence of 0.4 to 1 in 100,000. Insulinomas confine the largest group of p-NETs. Somatic mutations in MEN1 have been found in up to 20% of sporadic p-NETs. Recently, a hot spot mutation was found in the Ying Yang 1 gene in 30% of insulinomas. The YY1 gene can act either as a tumorsupressor or an oncogene in other tumours such as breast, prostate and colorectal cancer. The mutation (YY1 T372R) was associated with a later onset of insulinomas. 

Material and methods: 

Benign sporadic insulinomas (n=39) were snap frozen after surgery. DNA was extracted, subjected to PCR and subsequent automated Sanger sequencing of YY1 and MEN1

Oral glucose tolerance test (OGTT) was performed before the surgery. Fischer exact test and the Mann-Whitney-U-test were used for statistical analysis.

Results:

We found the YY1 T372R mutation in 28% (11/39) of the tumours, mutually exclusive from mutations in MEN1 found in 13% (5/39). The median age of patients with tumour harboring YY1 mutation was 63.0 ± 13.4 years and 45.5 ± 17.2 years in in those without YY1 mutation (p=0.03). There was no significant difference in sex, or the extent of the tumors. OGTT could not show any significant differences between the two groups 36 ±11.5 (mg/dl) in the YY1 mutated group and 41 ± 15.3 (mg/dl) in those without YY1 mutation, (p=1.00) The maximum suppressed insulin value at symptomatic hypoglycemia was 13.4 ± 14 (mU/L) versus 18,2 ± 21,7 (mU/L) p=0.61.

There was no significant difference concerning the time until hypoglycemia of <45 (mg/dl) was achieved in the OGTT test (5 hours versus 4 hours, p=0.15). 

Conclusion:

In accordance to the previously published data, we found the YY1 T372R mutation in 28.2% of  the analyzed sporadic benign insulinomas. We demonstrated a significantly later onset of insulinomas harboring YY1 mutation. Pathophysiological symptoms in the patients investigated by OGTT were not correlated to mutation status. Further investigations are warranted to determine the biological behavior of insulinomas with the YY1 T372R mutation.

 

Nothing to Disclose: AM, AS, AA, PG, RM, PH, PB

OR02-2 21178 2.0000 A YY1 Mutations Are Frequent in Insulinomas, Mutually Exclusive from MEN1 Mutation and Do Not Correlate to Pathophysiological Symptoms 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 1:00:00 PM OR02 5972 11:30:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Oral


Gerard V Walls*1, Mark Stevenson1, Ben Soukup1, Kate E Lines1, Paul T Christie1, Ashley B. Grossman1, Herbert A Schmid2 and Rajesh V Thakker1
1University of Oxford, Oxford, United Kingdom, 2Novartis Pharmaceuticals, Basel, Switzerland

 

Improved therapies for pancreatic and pituitary neuroendocrine tumors (NETs), which may occur in Multiple Endocrine Neoplasia type 1 (MEN1), are needed. We therefore assessed the effects of pasireotide, a somatostatin analogue that has high affinity for 4 of the 5 somatostatin receptor sub-types, in a mouse model for MEN1. Mice deleted for an Men1 allele (Men1+/-) are representative of MEN1 in man, as Men1+/- mice develop pancreatic and pituitary NETs in association with parathyroid tumors from 12 months of age. To study the effects of pasireotide on NET proliferation and apoptosis, we treated Men1+/- mice aged 12 months with either pasireotide 40 micrograms/g body weight intramuscularly (i.m.) once per month (treatment group, n=71) or phosphate-buffered saline (PBS) i.m. (control group, n=73) for 9 months. All Men1+/- mice (n=144) had magnetic resonance imaging (MRI) at 12 and 21 months of age, to assess tumor development, and oral 1 mg/ml 5-bromo-2-deoxyuridine (BrdU) in drinking water for one month from 20 months of age, to assess tumor proliferation. NETs were collected by necropsy at 21 months of age, and proliferation and apoptosis were assessed by immunohistochemical quantification of nuclei containing BrdU and TUNEL assays, respectively. Pasireotide-treated Men1+/- mice had a significantly increased survival, assessed by Kaplan-Meier analysis, compared to PBS-treated mice (82% versus (vs) 64%, respectively, P<0.05), and this was associated with the development of fewer NETs in the pasireotide-treated group vs the PBS-treated group (86% vs 98% pancreatic NETs, and 52% vs 72% pituitary NETs, P<0.05 in both). Furthermore, MRI scans revealed that pasireotide treatment diminished the increase in pituitary NET volumes (pre-treated vs post-treated = 0.933 ±0.099 mm3 vs 3.525 ±1.349 mm3 in the pasireotide-treated group, compared to 0.975 ±0.161mm3 vs 10.583 ±3.187mm3 in the PBS-treated group, P<0.05), while necropy revealed the pasireotide-treated group to have significantly fewer pancreatic NETs when compared to PBS-treated mice (2.36 ±0.25 vs 3.72 ±0.32, respectively, P<0.001). Pasireotide treatment also decreased NET proliferation when compared to PBS treatment (PBS-treated vs pasireotide-treated = 1.81 ±0.15% vs 0.73 ±0.07% in pituitary NETs, and 0.78 ±0.08% vs 0.35 ±0.03% in pancreatic NETs, respectively, P<0.0001), but increased apoptosis (2.35 ±0.44% vs 14.75 ±1.58% in pituitary NETs, and 0.19 ±0.03% vs 0.42 ±0.05% in pancreatic NETs, respectively, P<0.001 for both). Thus, pasireotide treatment in Men1+/- mice increased survival by ~20%, inhibited NET growth and reduced the number and volume of pancreatic and pituitary NETs, thereby indicating its potential use as an anti-proliferative and pro-apoptotic therapy for pancreatic and pituitary NETs.

 

Disclosure: GVW: Researcher, Novartis Pharmaceuticals. ABG: Ad Hoc Consultant, Lecture fees and Advisory Board, Ad Hoc Consultant, Lecture fees and Advisory Board. HAS: Employee, Novartis Pharmaceuticals. RVT: Principal Investigator, Novartis Pharmaceuticals. Nothing to Disclose: MS, BS, KEL, PTC

OR02-3 18709 3.0000 A Treatment with Pasireotide, a Somatostatin Analogue, Results in Decreased Proliferation and Increased Apoptosis in Pancreatic and Pituitary Neuroendocrine Tumors (NETs), in a Mouse Model for Multiple Endocrine Neoplasia Type 1 (MEN1) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 1:00:00 PM OR02 5972 11:30:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Oral


Andrea Estrada*1, Aradhana Venkatesan2, Christopher L Wolfgang3, Elliot K Fishman4, Ihab Kamel4, Syed Z Ali4, Michael Goggins3, Anirban Maitra5, Lori C. Guthrie6, Beth Brillante7, Rachel I Gafni7, Anne Marie Lennon3 and Michael T. Collins8
1NIDCR/NICHD, NIH, Bethesda, MD, 2National Institutes of Health, Bethesda, MD, 3Johns Hopkins University School of Medicine, Baltimore, MD, 4Johns Hopkins University School of Medicine, 5MD Anderson, 6Section on Skeletal Disorders and Mineral Homeostasis, Bethesda, MD, 7National Institutes of Health, 8NIDCR, NIH, Bethesda, MD

 

Background:  The McCune Albright syndrome (MAS) exists along a clinical spectrum that includes fibrous dysplasia of bone, café-au-lait macules, and hyperfunctioning endocrinopathies, including precocious puberty, hyperthyroidism, GH excess, FGF23-mediated hypophosphatemia and hypercortisolism.  It is due to somatic activating mutations in GNAS.  The same GNAS mutations have been found as driver mutations in 66% of intraductal papillary mucinous neoplasms (IPMNs). IPMNs are cystic neoplasms of the pancreas that commonly present in the 7th decade of life, and are thought to be pre-cancerous.  Pancreatic ductal lesions, including IPMNs, have been reported in MAS, however the incidence and natural history is not well characterized.

Objective: Determine the prevalence and characteristics of pancreatic neoplasms in MAS.

Patients and Methods:  Thirty patients (aged 7 to 67 years, median 18.5) in the National Institutes of Health cohort of subjects with MAS underwent abdominal MRI and magnetic resonance cholangiopancreatography (MRCP) screening for pancreatic neoplasms.  Identified IPMNs were further categorized according to the 2012 International Consensus Guidelines into those with either high risk or worrisome features; including main duct dilatation ≥5 mm, cysts ≥ 3 cm, mural nodule, thickened wall or solid component, jaundice secondary to the cyst or history of acute pancreatitis.

Results: Nine patients (30%) had pancreatic abnormalities.  Eight of these patients had cystic lesions of the pancreas; one with simple cysts and seven with IPMNs.  Five of the seven IPMNs exhibited worrisome or high-risk stigmata.  IPMN subtypes included the following: one main pancreatic duct IPMN, three branch duct IPMN and three mixed- IPMN.  The median age of patients with pancreatic pathology was 47 (range 17 to 67).  Three patients had pancreatic divisum, including 2 of the subjects with IPMNs. None of them had evidence of exocrine dysfunction. Surprisingly, 5 of the 9 subjects with pancreatic pathology (55%) were diagnosed with diabetes.  The mean BMI and ages were 29 and 33. The mean BMI and age of onset of type 2 diabetes in the general population are 37 and 45, respectively.  <5% of the remaining >170 subjects in the NIH MAS cohort have diabetes   

Conclusions:  Gastrointestinal neoplasms, particularly IPMNs, appear to occur more frequently and at a younger age in patients with MAS compared to the general population. This suggests that pancreatic neoplasms are a clinical feature of MAS.  Given the malignant potential of IPMNs in the general population, screening for pancreatic neoplasms in patients with MAS may be warranted.  Further determination of the natural history and spectrum of IPMNs in MAS is needed. Finally, the unexpected high rate of diabetes in subjects with MAS and anatomical pathology warrants further investigation.

 

Nothing to Disclose: AE, AV, CLW, EKF, IK, SZA, MG, AM, LCG, BB, RIG, AML, MTC

OR02-4 21953 4.0000 A Pancreatic Pathology in the Mccune-Albright Syndrome 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 1:00:00 PM OR02 5972 11:30:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Oral


Idit Dotan*1, Elliot Jonathan Mitmaker1, Philip Jonathan Rupert Roche2, Lenore Katherine Beitel3, Michael Tamilia3, Mark Anthony Trifiro4 and Miltiadis Paliouras5
1McGill University Health Center, Montreal, QC, Canada, 2Lady Davis Institute for Medical Research-Jewish General Hospital, Montreal, QC, Canada, 3McGill University, Montreal, QC, Canada, 4The Jewish General Hospital, McGill University, Lady Davis Institute for Medical Research, Montreal, QC, Canada, 5Lady Davis Institute for Medical Research-Jewish General Hospital, McGill University, Montreal, QC, Canada

 

Background. The incidence of papillary thyroid carcinoma (PTC) has risen steadily over the past few decades, while surgical and medical treatments are still associated with significant comorbidities. Importantly, recurrence rates of up to 20% expose patients to additional morbidities and increased death risk. The Achilles heel of chemotherapy and biological therapies is heterogeneity within the tumor itself, generated by accumulation of mutations that create multiple sub-populations in a given tumor. These issues emphasize the need for new technology to treat local recurrence and metastatic disease, with higher success rates and minimal compromise to quality of life. It is proposed that targeted ablative physical therapy could represent a therapeutic modality, with limited side effects. Targeted functionalized multi-walled carbon nanotubes (BioNanofluid) act by efficiently converting light energy to heat locally, representing a promising new cancer therapeutic advancing beyond conventional laser ablation and other nanoparticle approaches.

Methods. The presence of the TSH-Receptor (TSH-R) on various cells was established using Western blot analysis. BioNanofluid was conjugated to TSH-R antibody (Ab), purified TSH (Thyrotropin) and recombinant TSH (Thyrogen) through covalent amide bonds to create 3 different types of conjugates. A laser at 532nm was used to illuminate cells for set exposure times. Cell death was assessed using trypan blue staining.

Results. BCPAP, a PTC cell line was TSH-R positive, while NSC-34, a hybrid neuronal mouse cell line was TSH-R negative. A 2:1 BCPAP cell:conjugate ratio and 30 second exposure time eliminated ~60%, 65% and >70%  of the BCPAP cells when using TSH-R Ab, Thyrotropin and Thyrogen conjugates, respectively, with minimal non-targeted killing. Increasing conjugate ratios and longer exposure (40 seconds) yielded higher targeted cell killing rates; however, more than 30% non-targeted killing in the BCPAP Unconjugated BioNanofluid control group was observed, suggesting bulk heating. NSC-34 (control) cell killing was less than 10% under all conditions. With engineered improvement of nanoparticles, chemistry and ligand presentation, targeted killing rate exceeded 90% with ~10% non-specific cell death.

Conclusion. A BioNanofluid platform offering a potential therapeutic path for PTC has been investigated, offering an alternative for more complicated patients, who currently undergo suffering caused by reoperative neck exploration, repeated administration of radioactive iodine (RAI), external beam radiation or chemotherapy. In addition, BioNanofluid treatment may serve as an alternative to RAI therapy, directed at both PTC cells and normal thyroid tissue. This work represents one demonstration of BioNanofluid therapeutic development, where targeted physical therapy can overcome the challenge of tumor heterogeneity.

 

Nothing to Disclose: ID, EJM, PJRR, LKB, MT, MAT, MP

OR02-5 19875 5.0000 A Engineering Bionanofluids to Target Papillary Thyroid Cancer Cell Lines 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 1:00:00 PM OR02 5972 11:30:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Oral


Aleck Hercbergs*1, Thangirala Sudha2, Murat Yalcin3, Hung-Yun Lin4, Mary Koeppe Luidens5, Shaker A. Mousa6 and Paul J Davis7
1Cleveland Clinic, 2Pharmaceutical Research Institute, Albany College of Pharmacy and Health Sciences, 3Uludag University, Bursa, Turkey, 4Taipei Medical University, Taiwan, 5Albany Medical Center, Albany, NY, 6Albany College of Pharmacy and Health Sciences, Rensselaer, NY, 7Albany College of Pharmacy and Health Sciences

 

Glioblastoma multiforme (GBM) is an aggressive brain tumor with a less than 10% five-year survival rate.  The tumor is relatively chemo- and radioresistant and, despite its vascularity, has shown disappointing clinical response to anti-angiogenic management.  Nanoparticulate tetraiodothyroacetic acid (Nanotetrac) is an anti-cancer/anti-angiogenic agent that acts exclusively at a thyroid hormone-tetrac receptor site on plasma membrane integrin αvβ3 and does not gain entry to cells.  In this formulation, tetrac is covalently bound via a linker to 200 nm poly(lactic-co-glycolic acid)(PLGA) nanoparticles.  From the integrin, the agent disorders a panel of tumor cell defense pathways by downstream actions on expression of differentially-regulated genes and is pro-apoptotic by several mechanisms.  In the C6 rat glioma subcutaneous xenograft in the mouse, standard clinical GBM chemotherapy with temozolomide (TMZ) at 10 mg/kg was wholly ineffective (tumor volumes at 14 d), as was subtherapeutic dosage of Nanotetrac (0.3 mg/kg).  Added to TMZ, Nanotetrac at 0.3 mg/kg reduced tumor volume by 53% at 14 d.  In the subcutaneous  human U87MG (U87-luc) GBM xenograft, Nanotetrac as a single dose at 1 mg/kg injected s.c. together with the implanted cells inhibited tumor growth by 79% at 16 d.  Luminescent scanning revealed no viable cells in the Nanotetrac-exposed tumors, with full viability in the void PLGA-injected lesions.  Systemic (s.c.) Nanotetrac at 1 mg/kg q2 d reduced subcutaneous tumor U87MG xenograft volume by 46% at 10 d.  All Nanotetrac results were significant at p < 0.05.  Studied in vitro, U87MG cells proliferated in response to physiological concentrations of L-thyroxine (T4) and supraphysiological levels of 3,5,3-triiodoo-L-thyronine.  The T4 effect was associated with activation of mitogen-activated protein kinase (pERK1/2), but phosphatidylinositol 3-kinase.  Taken together, these results indicate that 1) GBM is a thyroid hormone-responsive tumor, consistent with clinical observations of the action of induced hypothyroxinemia on GBM (A Hercbergs et al., Anticancer Res 23:617-626, 2003; Oncologist, in press, 2015) and 2) Nanotetrac administered systemically or intratumorally is effective in reducing GBM xenograft volumes/weights and in decreasing tumor cell viability.

 

Nothing to Disclose: AH, TS, MY, HYL, MKL, SAM, PJD

OR02-6 21888 6.0000 A Activity of Nanoparticulate Tetrac (Nanotetrac) on Models of Glioblastoma 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Tumor Biology Thursday, March 5th 1:00:00 PM OR02 5972 11:30:00 AM Endocrine Neoplasia: Tumorigenesis and Therapeutics Oral


Peter James Trainer*1, John Newell-Price2, John Ayuk3, Simon J B Aylwin4, Aled Rees5, William Drake6, Philippe Chanson7, Thierry Brue8, Susan M. Webb9, Carmen Fajardo10, Javier Aller11, Ann I McCormack12, David J Torpy13, George Tachas14, Lynne Atley15 and Martin Bidlingmaier16
1The Christie NHS Foundation Trust, Manchester, United Kingdom, 2Univ of Sheffield, Sheffield, United Kingdom, 3University Hospital Birmingham, Birmingham, United Kingdom, 4King's College Hospital, London, United Kingdom, 5Cardiff University, Cardiff, United Kingdom, 6St Bartholomew's Hospital, London, United Kingdom, 7Univ Paris-Sud, UMR-S693, Assistance Publique-Hôpitaux de Paris, Le Kremlin Bicêtre, France, 8Hopital de la Conception, Marseille, France, 9Hosp de Sant Pau, Barcelona, Spain, 10Hospital Universitario La Ribera, Alzira. Valencia, Spain, 11Hospital Universitario Puerta de Hierro, Madrid, Spain, 12St. Vincent Hospital, Naremburn - NSW, Australia, 13Royal Adelaide Hospital, Adelaide, Australia, 14Antisense Therapeutics, Melbourne, Australia, 15Antisense Therapeutics Limited, Melbourne, Australia, 16Ludwig-Maximilians University, Munich, Germany

 

ATL1103 is a second generation antisense oligomer directed at the GH receptor. It is a 20mer with a phosphorothioate backbone and 2’-O-methoxyethyl modifications of the five nucleotides at either end intended to increase its plasma half-life and affinity for the target RNA to allow post-hybridization RNaseH degradation of the GHr RNA strand.  ATL1103 is relatively rapidly distributed from plasma (Cmax in 2-4 hrs) into peripheral tissues, with a tissue clearance half-life of 2-4 weeks in primates.  We report a phase 2 randomised, open-label, parallel group study of subcutaneously administered ATL1103 in patients with active acromegaly.  Appropriate ethical approval was obtained in every jurisdiction and the study is registered as EudraCT 201200314730.  Patients gave written informed consent.   The protocol entailed appropriate washout from any ongoing medical therapy after which IGF-I had to be at least >1.30 times age-related ULN.  Patients were randomised to receive either ATL1103 200 mg once or twice weekly for 13 weeks (3 doses in the first week).  After completion of drug administration, patients were monitored for a further eight weeks.  The primary objectives as per protocol were to evaluate the safety and to understand the single dose and multiple dose pharmacokinetic profiles of ATL1103 in patients with acromegaly.  35 patients were recruited in 13 centres in Australia, France, Spain and UK. 26 (Mean age 50.4 yr; range 26 to 80, 11 male) were randomised, all of whom completed treatment.  ATL1103 was well tolerated with mild to moderate injection site reactions being the most common drug-related AE.  Four SAEs were reported, of which three occurred in a single patient, but none were felt to be study drug related.   Two patients withdrew from the study at completion of dosing with study drug. There was a significant fall in serum IGF-I of 26% by week 14 (one week past the last dose) with 200 mg twice weekly (577 ± 198 v 411 ± 174 ng/ml (mean ± SD), P <0.0001) although the nadir had not been reached.  Once weekly dosing did not result in a significant fall in IGF-I.   The fall in IGF-I with twice weekly dosing was associated with a mean reduction in ring size of 0.92 (P=0.02) and an increase in GH (AUC during an OGTT, P=0.001).  This study provides proof-of-concept that ATL1103 is able to significantly lower IGF-I in patients with acromegaly and offers a novel therapeutic approach.

 

Disclosure: PJT: Clinical Researcher, Antisense Therapeutics, Clinical Researcher, Antisense Therapeutics, Clinical Researcher, Novartis Pharmaceuticals, Clinical Researcher, Chiasma, Clinical Researcher, Ipsen, Speaker, Novartis Pharmaceuticals. AR: Advisory Group Member, Novartis Pharmaceuticals, Advisory Group Member, Ipsen, Speaker, Ipsen. WD: Clinical Researcher, Antisense. PC: Researcher, Ipsen, Researcher, Novartis Pharmaceuticals, Researcher, Novo Nordisk, Researcher, Pfizer, Inc., Advisory Group Member, Ipsen, Advisory Group Member, Novartis Pharmaceuticals, Advisory Group Member, Viropharma, Speaker, Ipsen, Speaker, Novartis Pharmaceuticals, Speaker, Pfizer, Inc.. TB: Investigator, Pfizer, Inc., Consultant, Pfizer, Inc., Speaker, Pfizer, Inc., Investigator, Novo Nordisk, Speaker, Novo Nordisk, Investigator, Novartis Pharmaceuticals, Speaker, Novartis Pharmaceuticals, Consultant, Novartis Pharmaceuticals, Advisory Group Member, Novartis Pharmaceuticals, Consulting speaker, Merck Serono, Consulting speaker, Lilly USA, LLC, Investigator, Ipsen, Speaker, Ipsen, Advisory Group Member, Ipsen, Consultant, Ipsen, Speaker, Sandoz, Investigator, Sandoz. SMW: Ad Hoc Consultant, Antisense Therapeutics, Clinical Researcher, Antisense Therapeutics. JA: Advisory Group Member, Novartis Pharmaceuticals, Speaker, Novartis Pharmaceuticals, Advisory Group Member, Pfizer, Inc., Speaker, Pfizer, Inc., Speaker, Ipsen, Ad Hoc Consultant, Ipsen, Advisory Group Member, Bayer, Inc.. GT: Director Drug Discovery and Patents, Antisense Therapeutics Ltd. LA: Employee, Antisense Therapeutics Ltd. Nothing to Disclose: JN, JA, SJBA, CF, AIM, DJT, MB

OR09-1 20897 1.0000 A A Phase 2 Study of Antisense Oligonucleotide Therapy Directed at the GH Receptor Demonstrates Lowering of Serum IGF-I in Patients with Acromegaly 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR09 5992 11:30:00 AM Pituitary Tumors-Therapeutics Oral


Lillian Ting*1, Anadya Prakash Tripathi2, Christelle Darstein3, Tracy White1 and Nicholas Sauter1
1Novartis Pharmaceuticals Corporation, East Hanover, NJ, 2Novartis Healthcare Pvt Ltd, Hyderabad, India, 3Novartis Pharma AG, Basel, Switzerland

 

Background: LCI699 (osilodrostat) is a potent oral inhibitor of 11β-hydroxylase currently in Phase III clinical development for the treatment of patients with Cushing’s disease. In vitro assessments of osilodrostat suggested potential drug-drug interactions (DDIs) with cytochrome P450 (CYP) enzyme metabolism. Of the numerous CYP enzymes, only five (CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4) are responsible for metabolizing approximately 90% of medications, with CYP3A4 accounting for nearly half this total. This clinical study evaluated the inhibitory effect of osilodrostat on the pharmacokinetics (PK) of the CYP1A2, CYP2C19, CYP2D6, and CYP3A4 probe substrates, caffeine, omeprazole, dextromethorphan, and midazolam, respectively.

Methods: Single-center, open-label, Phase I, DDI study in healthy adult volunteers. On study day 1 volunteers received a single dose of probe substrates (cocktail approach; caffeine [100 mg], omeprazole [20 mg], dextromethorphan [30 mg], and midazolam [2 mg]), followed by PK assessments over the next 48 h. After a 6-day washout period, subjects then received a single dose of osilodrostat 50 mg (planned therapeutic dose range 2–30 mg bid), followed 0.5 h later with a single dose of probe substrates. PK assessments were conducted over the next 48 h.

Results: 10 males and 10 females with an overall mean age, body weight, and body mass index of 41.8 years, 72.95 kg, and 24.4 kg/m2 were enrolled. One subject discontinued (withdrew consent). AUClast geometric mean ratio (GMR) and 90% confidence intervals (90% CI) of probe substrate exposure with osilodrostat administration were 2.33 (2.10–2.59), 1.91 (1.74–2.11), 1.48 (1.34 –1.63), and 1.50 (1.41–1.60) for caffeine, omeprazole, dextromethorphan and midazolam, respectively. Corresponding values for AUC GMR (90% CI) were 2.54 (2.34–2.75), 1.86 (1.61–2.15), 1.54 (1.40–1.69), and 1.50 (1.41–1.59). Cmax GMR (90% CI) for the change in substrate exposure was 1.07 (0.988–1.15), 1.61 (1.40–1.84), 1.35 (1.21–1.50), and 1.47 (1.32–1.62) for caffeine, omeprazole, dextromethorphan and midazolam, respectively.

Osilodrostat is a moderate inhibitor of CYP1A2 and CYP2C19 (exposures of the respective probe substrates increased ~2.5- and 2-fold, respectively) and a weak inhibitor of CYP2D6 and CYP3A4 (exposures of the respective probe substrates increased ~1.5 fold).

Conclusions: Considering that a 50 mg single dose of osilodrostat is reasonably representative of the upper end of the daily drug exposure expected for clinical use (maximum 30 mg bid), it is encouraging that the most clinically important CYP enzyme, CYP3A4, was only weakly inhibited. As most patients with Cushing’s disease in a Phase II study achieved normal urinary free cortisol levels with ≤10 mg bid osilodrostat, osilodrostat is unlikely to have a clinically relevant impact on the exposures of other medications cleared by CYP3A4.

 

Disclosure: LT: Employee, Novartis Pharmaceuticals. APT: Employee, Novartis Pharmaceuticals. CD: Employee, Novartis Pharmaceuticals. TW: Employee, Novartis Pharmaceuticals. NS: Employee, Novartis Pharmaceuticals.

OR09-2 21061 2.0000 A Evaluation of the Effects of LCI699 (Osilodrostat) on Cytochrome P450 Enzymes in Healthy Volunteers: Low Drug-Drug Interaction Potential 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR09 5992 11:30:00 AM Pituitary Tumors-Therapeutics Oral


Shlomo Melmed*1, Nienke R. Biermasz2, Peter J. Trainer3, Gary Patou4 and Asi Haviv5
1Cedars-Sinai Medical Center, Los Angeles, CA, 2Leiden University Medical Center, Netherlands, 3The Christie NHS Foundation Trust, Manchester, United Kingdom, 4Chiasma Inc,, Newton, MA, 5Chiasma, Jerusalem, Israel

 

Background: Oral octreotide capsules (OOC) contain a transient permeability enhancer (TPE®) which enables effective drug absorption. In a multicenter phase 3 trial, subjects previously controlled on injectable somatostatin receptor ligands (SRLs) were switched to OOC for up to 13 months. OOC was shown to be safe and effective in maintaining IGF-1 and GH control in 62% of subjects enrolled vs. 89% at baseline. The present evaluation analyzes determinants of OOC efficacy.

Methods: At baseline, of 151 evaluable enrolled patients, 93 (62%), were fully controlled (IGF-1 < 1 ULN and integrated GH < 2.5 ng/mL); 41 subjects (27%) were partially controlled with IGF-1 levels between 1 – 1.3XULN and GH < 2.5 ng/mL; and 17 subjects (11% ) were not controlled. 86 subjects (57%)  were previously treated with high doses of parenteral SRLs (Octreotide LAR® ≥30mg/month or Lanreotide Autogel®=120mg/month), and 65 were previously treated with   low to mid doses of injectables (LAR 10 or 20mg or Autogel 60 or 90mg).   In this descriptive analysis we studied the relationship between pre-study SRL dose (post hoc), or baseline disease activity (pre-defined), to OOC efficacy up to 13 months. The correlation between prior SRL dose and oral octreotide dose was also studied.

Results:The degree of baseline control while on injectable SRLs predicted subsequent OOC response. Respective OOC response rates were 76 % (full baseline control), 66% (full or partial baseline control) and 24% (uncontrolled at baseline). Previous injectable SRL dose also predicted OOC response. 72% of subjects previously treated with low to mid SRL doses responded to OOC vs.  56% response for subjects previously treated with high doses. Full biochemical control at baseline and low to mid doses of prior injectable SRLs yielded 85% response on OOC (49 of 58 subjects). The OOC dose distribution in all enrolled subjects was shown to be related to prior injectable SRL dose. While 74% of subjects previously treated with low SRL doses required 40mg OOC, 31% of subjects treated with high SRL dose were controlled by 40 mg OOC. 110 subjects were initially controlled on OOC following dose titration, and entered a fixed dose phase (average 12 weeks from first OOC dose). At the beginning of the fixed-dose phase, 51, 25 and 34 subjects were treated with OOC 40, 60 and 80mg respectively. Initial response to low to mid doses of OOC predicts sustainable response over time as the response up to 13‑months was 88, 84, and 47% for each respective OOC dose.

Conclusions: High dose injectable SRL or partial baseline control portend favorable efficacy to OOC. Full baseline biochemical control, and/or low to mid injectable SRL dose requirements portend  higher OOC response rates. OOC was effective in a subset of patients, uncontrolled at baseline while on injectable SRLs. OOC efficacy can reliably be determined within 12 weeks of initiating therapy and its effect is sustainable for up to 13 months.

 

Disclosure: SM: Planning Group Member, Ipsen, Ad Hoc Consultant, Novartis Pharmaceuticals, Principal Investigator, Pfizer, Inc., Ad Hoc Consultant, ISIS, Advisory Group Member, Chiasma. NRB: Coinvestigator, chiasma. PJT: Clinical Researcher, Chiasma, Clinical Researcher, Antisense Therapeutics, Clinical Researcher, Antisense, Clinical Researcher, Novartis Pharmaceuticals, Speaker, Novartis Pharmaceuticals, Clinical Researcher, Ipsen. GP: Management Position, Chiasma. AH: , Chiasma.

OR09-3 21142 3.0000 A Determinants of Oral Octreotide Capsules Efficacy in Acromegaly 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR09 5992 11:30:00 AM Pituitary Tumors-Therapeutics Oral


Aart J. van der Lely*1, Peter J. Jonsson2, Patrick Wilton3, Jose Francisco Cara4 and Ezio Ghigo5
1Erasmus University Medical Center, Rotterdam, Netherlands, 2Endocrine Care, Pfizer Health AB, Sollentuna, Sweden, 3Stockholm, Sweden, 4Pfizer Inc, New York, NY, 5University of Turin, Turin, Italy

 

Introduction:

Pegvisomant (PEGV) is a competitive antagonist of growth hormone (GH) approved for the treatment of patients with acromegaly. The recommended starting dose of PEGV is 10 mg/day, and doses are titrated upwards until normalization of serum levels of insulin-like growth factor-I (IGF-I) is attained. The purpose of this analysis was to evaluate data from ACROSTUDY, a Pfizer-sponsored non-interventional post marketing study on the long-term outcomes of pegvisomant therapy, on the clinical characteristics of patients who required PEGV at a daily dose above 30 mg/day for IGF-I normalization and to compare these with the characteristics of patients who attained IGF-I control with a PEGV dose of 10 mg/day.

Methods:

All data available as of June 2014 were included in this analysis. Baseline was defined as start of PEGV. IGF-I was measured at various local labs and expressed as IGF-I /upper normal level (ULN).  A normal IGF-I was defined as an IGF-I/ULN below 1.2. All patients treated for at least six weeks with a PEGV at a dose above 30 mg/day who had two consecutive normal IGF-I values were included in the “High” group (H). Patients who had two consecutive normal IGF-I values and who never received treatment with a dose of PEGV above 10 mg/day were included in the “Low” group (L). Descriptive statistics are expressed as mean (±SD).  Significance level was set to 5%.

Patients:

56 patients (39% males) with a mean (SD) age of 43.6 (13.0) yrs at PEGV start in H and 368 patients (46% male) with a mean age of 51.9 (14.6) yrs (p<0.05) yrs in L were included in the analysis.

Results:

At baseline 84% of H and 77% of L was treated before with surgery and 29% of H and 26% L had received radiotherapy. Time since diagnosis of acromegaly was 6.3 (8.9) yrs and 8.6 (9.0) in H and L respectively (p<0.05).  Of 6 clinical symptoms (arrhythmia, diabetes mellitus, hypertension, osteoarthritis, sleep apnea, surgery for carpal tunnel syndrome) collected in ACROSTUDY, hypertension and sleep apnea, were more frequent in the H group. At baseline 27% and 44% (p<0.05) were on long-acting somatostatin analogs and 18% and 11% were on dopamine antagonists in H and L resp. IGF/ULN at baseline was 2.8 (1.3) and 1.4 (0.7) (p<0.05) in patients in whom IGF-I levels were available (82% in H and 71% in L). The mean starting dose of PEGV was 16.6 (12.9) mg/d in H and 8.8 (2.4) mg/day in L (p<0.05). Duration of PEGV treatment was 6.9 (2.5) yrs in H and 4.4 (2.5) yrs in L (p<0.05). Duration of treatment with a dose >30 mg was 3.5 (2.0) years. IGF/ULN at the end of follow up was 1.0 (0.5) in H and 0.8 (0.4) in L (p<0.05).

Conclusion:

Data from ACROSTUDY suggest that the group of subjects that need high daily doses of PEGV to normalize their serum IGF-I levels have a higher incidence of hypertension and sleep apnea and higher baseline values of IGF-I, which might suggest that they have a more severe form of acromegaly.

 

Disclosure: AJV: Speaker, Pfizer, Inc., Consultant, Pfizer, Inc., Advisory Group Member, Pfizer, Inc.. PJJ: Employee, Pfizer, Inc.. PW: Consultant, Pfizer, Inc.. JFC: Employee, Pfizer, Inc.. EG: Advisory Group Member, Pfizer, Inc., Consultant, Pfizer, Inc., Speaker, Pfizer, Inc..

OR09-4 20373 4.0000 A Treatment with High Doses of Pegvisomant in 56 Patients with Acromegaly: Experience from Acrostudy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR09 5992 11:30:00 AM Pituitary Tumors-Therapeutics Oral


Greisa Vila*1, Ann-Charlotte Akerblad2, Anders F. Mattsson3, Michaela Riedl4, Susan M. Webb5, Vaclav Hana6, Eigil H Nielsen7, Beverly M.K. Biller8 and Anton Luger9
1Medical University of Vienna, Vienna, Austria, 2Endocrine Care, Pfizer Health AB, Sollentuna, Sweden, 3Pfizer Health AB, Sollentuna, Sweden, 4Medical University of Vienna, Vienna, Austria, 5Hospital Sant Pau, Centro de Investigación Biomédica en Red de Enfermedades Raras (CIBER-ER, Unidad 747), ISCIII and Universitat Autònoma de Barcelona (UAB), Barcelona, Spain, 6Charles University, Prague, Czech Republic, 7Aalborg University Hospital, Aalborg, Denmark, 8Massachusetts General Hospital, Boston, MA, 9Medical University and General Hospital of Vienna, Vienna, Austria

 

Background: Growth hormone (GH) replacement in growth-hormone deficient (GHD) women is not approved during pregnancy. To date, only a few reports on growth hormone replacement therapy (GHRT) before and during pregnancy are available.

Aim and Methodology: Here we describe pregnancies in a large cohort of patients with hypopituitarism and investigate potential factors determining pregnancy outcomes. All pregnancies reported in KIMS (Pfizer International Metabolic Database), pregnancy outcomes and complications, as well as their relationship to use of GHRT during pregnancy were analyzed with simple and multiple regression methods. Significance level was set to 5%.

Results: A total of 201 pregnancies were reported from centers in fifteen countries, 173 in female patients aged 22-43 years (preliminary data on 115 shown at ENDO12), and 28 in partners of male patients. Ninety percent of female pregnant patients had at least one additional pituitary deficiency and two-thirds of women underwent fertility treatment in order to achieve pregnancy. GHRT was stopped before pregnancy in 7.5% of the female patients, as soon as pregnancy was confirmed in 40.1%, and at the end of the second trimester in 24.7% of the patients, while 27.6% of the female patients continued GHRT throughout pregnancy. GH treatment patterns during pregnancy were not related to the etiology of pituitary disease, extent of pituitary deficiencies or conception method. Birth of a healthy child was reported in 79% of the female pregnancies.  Non-elective abortions occurred mainly in the first trimester, and one fetal malformation (cystic hygroma) was diagnosed in the second trimester. Regression analyses performed to identify parameters related to pregnancy outcomes showed that they were not related to GHRT treatment patterns, IGF-I SDS (last before pregnancy), method of conception or number of additional pituitary deficiencies. Negative pregnancy outcomes were associated with a history of depression and with maternal age at conception below 30 or above 35 years. Pregnancy-associated complications were not related to GHRT during the pregnancy.

Conclusion: These data on pregnancies in a large cohort of women with hypopituitarism receiving GHRT reveal different treatment patterns across countries. Of note is the large number of patients requiring assisted reproductive techniques, extending beyond ovulation induction/ovarian stimulation, indicating that successful reproduction in women with pituitary deficiency often requires more than adequate hormone replacement therapy. In the clinical practice setting, nearly all patients taking GH replacement continue treatment during the time when they seek fertility, and more than half remain on treatment (fully or partially) during the pregnancy. Based on the current data, growth hormone replacement regimens during pregnancy do not appear to affect pregnancy outcomes.

 

Disclosure: GV: Speaker, Novartis Pharmaceuticals, Speaker, Pfizer, Inc.. ACA: Employee, Pfizer, Inc.. AFM: Employee, Pfizer, Inc.. MR: Speaker, Novo Nordisk. SMW: Consultant, Pfizer, Inc., Speaker, Pfizer, Inc.. VH: Ad Hoc Consultant, Pfizer, Inc., Speaker, Ipsen, Speaker, Novartis Pharmaceuticals. BMKB: Principal Investigator, OPKO, Consultant, Novo Nordisk, Consultant, Versartis, Consultant, Pfizer, Inc.. AL: Speaker, Serono, Consultant, Ipsen, Speaker, Ipsen, Consultant, Novo Nordisk, Speaker, Novo Nordisk, Consultant, Pfizer, Inc., Speaker, Pfizer, Inc., Consultant, Serono. Nothing to Disclose: EHN

OR09-5 20337 5.0000 A Pregnancy Outcomes in Women with Growth Hormone Deficiency Are Not Related to Growth Hormone Treatment during Pregnancy - a Long-Term Observational Study 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR09 5992 11:30:00 AM Pituitary Tumors-Therapeutics Oral


Katarzyna Kryzanowska-Mittermayer1, Anton Luger*2, Dominique Maiter3, Ulla Feldt-Rasmussen4, Anders F. Mattsson5, Ann-Charlotte Akerblad6, Nina Camacho-Hubner7 and Roger Abs8
1Rudolfstiftung Hospital, Vienna, Austria, 2Medical University and General Hospital of Vienna, Vienna, Austria, 3UCL St Luc Hospital, Brussels, Belgium, 4Copenhagen University Hospital, Rigshospitalet, Copenhagen, Denmark, 5Pfizer Health AB, Sollentuna, Sweden, 6Endocrine Care, Pfizer Health AB, Sollentuna, Sweden, 7Pfizer Inc, New York, NY, 8Edegem, Belgium

 

Growth hormone deficiency (GHD) may be caused by a malignant lesion of the hypothalamic-pituitary area or the treatment of a cranial tumor or hematological malignancy. It is unclear whether patients who survived such malignancies and who are under GH replacement treatment (GHRT) are at increased risk for a second malignant neoplasia.

We assessed the incidence of new malignant neoplasia in adults on GHRT with GHD caused by a malignancy or its treatment enrolled in KIMS (Pfizer International Metabolic Database). Childhood onset (CO) and adult onset (AO) cancer survivors (CS) were compared to patients with CO idiopathic GHD (IGHD) and with AO non-functioning pituitary adenoma (NFPA), respectively.

We compared 349 CO-CS (176 f, age at disease onset 10.4 yr, age at adult GHRT start 24.5 yr, BMI 26.6 kg/m², IGF-I SDS -2.4) with 619 IGHD (221 f, age at disease onset 9.0 yr, age at adult GHRT start 28.8 yr, BMI 24.9 kg/m², IGF-I SDS -3.4). CO-CS composition: germ cell tumor (112), medulloblastoma (68), astrocytoma (53), glioma (49), leukemia or lymphoma (47), nasopharyngeal tumor (10), chordoma (3), sarcoma (7). Radiotherapy was used in 69% of CO-CS. After a mean 5.2 yr GHRT (3908 patient-years), 3 new malignant neoplasias were observed in IGHD (1 leukemia, 1 breast cancer, 1 melanoma) with a Standardized Incidence Ratio (SIR) of 0.47 (95% CI, 0.09-1.37). After a mean 4.0 yr GHRT (2192 patient-years), 16 CO-CS developed a new neoplasia (7 non-melanoma skin cancer (NMSC), 6 malignant brain neoplasia, 2 cervix uteri cancer, 1 papillary thyroid cancer). The overall SIR for CO-CS was 10.39 (95% CI, 5.93-16.87) and 6.52 (95% CI, 2.97-12.37), excluding NMSC.

We compared 174 AO-CS (74 f, age at disease onset 30.1 yr, age at GHRT start 36.0 yr, BMI 29.4 kg/m², IGF-I SDS -1.4) with 2449 NFPA (933 f, age at disease onset 46.6 yr, age at GHRT start 53.2 yr, BMI 28.9 kg/m², IGF-I SDS -1.0). AO-CS composition: germ cell tumor (59), leukemia or lymphoma (35), astrocytoma (29), glioma (18), nasopharyngeal tumor (13), medulloblastoma (9), chordoma (7), sarcoma (4). Radiotherapy was used in 59% of AO-CS and 34% of NFPA. After a mean 4.1 yr GHRT (1024 patient-years), a new malignant neoplasia occurred in 3 AO-CS (liver cancer, prostate cancer, malignant histiocytosis) SIR 1.11 (95% CI, 0.22-3.24). After a mean 5.3 yr GHRT (15215 patient-years), NFPA patients developed a new malignant neoplasia in 153 cases (34 skin lesions, 35 prostate cancers, 13 lung cancers). The SIR for AO-NFPA was 1.05 (95% CI, 0.89-1.23).

This study shows an increased risk of second malignant neoplasia in CO-CS but not in AO-CS within the time frame of this study. This illustrates the need to follow CO-CS patients for longer time periods, especially when previously irradiated.

 

Disclosure: AL: Speaker, Serono, Consultant, Ipsen, Speaker, Ipsen, Consultant, Novo Nordisk, Speaker, Novo Nordisk, Consultant, Pfizer, Inc., Speaker, Pfizer, Inc., Consultant, Serono. UF: Advisory Group Member, Pfizer, Inc.. AFM: Employee, Pfizer, Inc.. ACA: Employee, Pfizer, Inc.. NC: Employee, Pfizer, Inc.. RA: Advisory Group Member, Pfizer, Inc.. Nothing to Disclose: KK, DM

OR09-6 20890 6.0000 A Incidence of New Neoplasia in Growth Hormone Treated Adult Patients with Pituitary Deficiency Following Childhood or Adulthood Malignancy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 1:00:00 PM OR09 5992 11:30:00 AM Pituitary Tumors-Therapeutics Oral


Cheuk Wun Li*1, Francesca Menconi1, Roman Osman1, Erlinda Concepcion1, Chella David2 and Yaron Tomer3
1Icahn School of Medicine at Mount Sinai, 2Mayo Clinic, 3Icahn School of Medicine at Mount Sinai and James J. Peters VA Medical Center, New York, NY

 

Graves’ disease (GD) is characterized by hyperthyroidism, production of thyroid stimulating hormone receptor (TSHR)-stimulating antibodies (TRAb), as well as infiltration of thyroid by T and B cells reactive to thyroid antigens. It is well established that the extracellular domain (ECD) of the human TSHR is the crucial antigen in GD targeted by both T-cells and TRAb. Our lab has previously shown that an HLA-DR variant that contains arginine at position 74 of the DRβ1 chain (DRb1-Arg74) is the specific MHC variant conferring risk for GD while the presence of glutamine at position 74 is protective (Ban et al., Genes Immun, 2004). While several TSHR peptides have been previously proposed as key epitopes, the major TSHR peptide triggering GD remains to be determined. We hypothesized that the key T-cell epitope in GD will bind specifically to the HLA-DRb1-Arg74 pocket. We screened 39 TSHR peptides spanning the ECD using a novel in vitro binding assay developed in our lab. This is an ELISA utilizing Baculovirus-generated recombinant HLA-DRb1-Arg74 and biotinylated TSHR peptides. We identified TSHR.132 and TSHR.197 as the best binders to DRb1-Arg74, with TSHR.132 binding with higher affinity. We then tested these peptides using a “humanized” mouse model, NOD-DR3, which are NOD mice that are null for murine MHC class II and expressing human HLA-DR3 (DRb1-Arg74 positive, confirmed by sequencing). We immunized NOD-DR3 mice with TSHR.132 and TSHR.197 and assessed T-cell responses to the peptides using CFSE test of proliferation and evaluating their cytokine responses. NOD-DR3 mice injected with TSHR.132 showed T-cell proliferation, accompanied by strong cytokine responses, but mice injected with TSHR.197 did not show T-cell responses. In conclusion, our study identified 2 TSHR epitopes; TSHR.132 and TSHR.197 bound in vitro with high affinity to HLA-DRb1-Arg74 which is the key HLA-DR pocket for development of GD. Our data suggest that TSHR.132, that has both high affinity for DRb1-Arg74 and could stimulate T-cell responses, is a major TSHR T-cell epitope. Our findings set the stage of designing inhibitors of binding of TSHR epitopes to HLA-DRb1-Arg74 as a potential novel therapeutic modality in AITD.

 

Nothing to Disclose: CWL, FM, RO, EC, CD, YT

OR11-1 21098 1.0000 A Discovering Thyroid Stimulating Hormone Receptor (TSHR) T-Cell Epitopes in Autoimmune Thyroiditis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 1:00:00 PM OR11 6031 11:30:00 AM Thyroid Development, Clinical and Autoimmunity Oral


Enrico Papini*, Rinaldo Guglielmi, Irene Misischi, Filomena Maria Graziano, Agnese Persichetti, Roberta Rendina, Silvia Taccogna and Giancarlo Bizzarri
Regina Apostolorum Hospital, Albano Laziale, Italy

 

Aim of the study: to establish, in a prospective controlled series of benign thyroid cysts, long-term efficacy of percutaneous ethanol injection (PEIT), predictive factors of favourable outcome, tolerability and side-effects of the procedure.

Materials and Methods: from February 2000 to March 2004, 174 of 260 consecutive cystic thyroid lesions were included in the study. Enrolment criteria were:  a. baseline volume > 5.0 mL; b. fluid component > 50% ; c. relapse after complete drainage; d. normal thyroid function; e. repeat benign cytology. Eighty-six patients (33%) were not eligible because of indeterminate or suspicious cytology, large multinodular goiter, or thyroid hyperfunction. Cyst volume at baseline was 5 - 10 mL in 51%, 10 - 20 mL in 27%, and > 20 mL in 22% of cases. Fluid content was > 75% in 56% and 50 - 75% in 44% of the cases. The following parameters were evaluated after the baseline drainage: characteristics of fluid content, rapid refilling after aspiration, uni or multilocular structure. Treatment was performed, according to the previously described technique, by injection under US monitoring of a volume of 95% ethanol corresponding to 50% of the drained fluid. At the 1 month control, a second treatment was performed in case of a fluid relapse > 30%. Clinical and US controls were performed at 1 and 6 months, and at 1, 5 and 10 years.

Results: Mean volume decrease was -38,2% at 1 month (p=0.05), -52,1% at 6 months (P=0.04), -57,5% at 1 year (p=0,03), -56,9% at 5 years (p=0,04) and -56,4% at 10 years (p=0,04). After 10 years volume reduction was -60,8% in cysts <10 mL (p=0,04),  -59,5% in cysts 10 - 20 mL (p=0,01) and 49,3% in cysts >20mL (p=0,02). Ten-year volume reduction was significantly greater in cysts with a fluid content > 75% than in those < 75% (81% vs  51%, respectively; p=0.01). The most favourable outcome was observed in unilocular cysts with > 75% colloid content and the worst in multilocular cysts with 50-75% fluid component and rapidly refilling hemorrhagic content. No changes of thyroid function or autoimmunity were observed. The procedure was described as well tolerated in 94% of cases. Two minor complications were registered during 229 procedures (0.8%): one self- resolving neck hemathoma and one temporary dysphonia. No malignancies were observed during the follow-up period.

Conclusions: PEIT induces a rapid and long-lasting reduction of the volume of benign cystic thyroid lesions. Favourable predictive factors are a fluid content > 75%, unilocular structure and no rapid refilling after drainage. PEIT should be considered as the first line treatment for prevalently cystic benign thyroid nodules.

 

Nothing to Disclose: EP, RG, IM, FMG, AP, RR, ST, GB

OR11-2 18895 2.0000 A LONG-TERM Outcome and Predictive Factors of Efficacy of Ultrasound-Guided Ethanol Injection for Benign Cystic Thyroid Lesions 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 1:00:00 PM OR11 6031 11:30:00 AM Thyroid Development, Clinical and Autoimmunity Oral


Sanjay Gawade*1, Katrin Hafen1, Urs Zumsteg2, Werner Krenger1 and Gabor Szinnai2
1Pediatric Immunology, Basel, Switzerland, 2University Children's Hospital Basel, Basel, Switzerland

 

In the differentiating tissues, reversible histone acetylation represents a major epigenetic mechanism regulating gene expression. Acetylation of core histones is regulated by histone deacetylases (HDACs) and histone acetyltransferases (HATs) in a reciprocal way. The aim of the study was to investigate the role of histone deacetylase activity for normal thyroid differentiation.

The effects of pharmacological HDAC inhibition by valproic acid (VPA 0.5-1.5mM) were investigated in an ex vivo murine embryonic thyroid organ culture model. Undifferentiated thyroids were microdissected at embryonic day 13.5 (E13.5), and analyzed at day 3 and 7 (E13.5+3d and +7d) of culture by HDAC activity assay, immunohistochemistry, morphometry, and flow cytometry (FACS).

The effect of HDAC inhibition by VPA 1.0mM induced the following epigenetic changes at E13.5+3d: First, general HDAC activity decreased significantly by 45% vs. control. Second, we observed by FACS a significant epigenetic switch on the histone tail mark H3K9 from trimethylation (H3K9me3) to acetylation (H3K9ac) compared to control in the complete thyroid as well as in the epithelial (EpCAM+), endothelial (CD31+) and mesenchymal (PDGRFalpha+) subpopulations. HDAC inhibition resulted in disturbed thyroid differentiation: By immunohistochemistry VPA treated organs showed 1) significantly lower BrdU+ cells, 2) lower number of blood vessels, and 3) decreased follicle number. These results were quantified by FACS and morphometry. By FACS, we observed at E13.5+7d a significantly decreased number of BrdU+ epithelial and endothelial cells while the number of BrdU+ mesenchymal cells did not change. As a consequence, absolute cell numbers of epithelial and endothelial cells decreased significantly (-57% and -79%, respectively) at day 7 of culture, while the mesenchymal cell mass remained constant. Morphometric analysis revealed a significant and dose dependent decrease of follicle numbers in VPA treated thyroids.

Our results show that normal HDAC activity levels are critical for growth, angiogenesis and follicle formation during thyroid differentiation in the mouse and suggest a putative new molecular mechanism for thyroid dysgenesis in the human.

 

Nothing to Disclose: SG, KH, UZ, WK, GS

OR11-3 21218 3.0000 A Histone Deacetylase Activity Is Critical for Embryonic Thyroid Growth and Differentiation 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 1:00:00 PM OR11 6031 11:30:00 AM Thyroid Development, Clinical and Autoimmunity Oral


Tim IM Korevaar*1, Marco Medici2, Henning Tiemeier1, Edward Visser1, Theo J Visser3 and Robin P. Peeters2
1Erasmus Medical Center, Rotterdam, 2Erasmus Medical Center, Rotterdam, Netherlands, 3Erasmus University Medical Center, Rotterdam

 

Context: A decrease in Childhood Intelligence Quotient (IQ) is strongly and independently associated with adverse health and economic outcomes including overall mortality, life-time achievements and gross domestic product. In vitro and animal studies have shown that thyroid hormone (TH) plays a crucial role in early brain development. In humans, neurogenesis starts at the 5th week of pregnancy whereas the fetal thyroid is not fully functional until week 18-20. As a consequence, the fetus heavily depends on the supply of maternal TH during the critical early stages of intrauterine brain development. 
All published clinical studies have investigated the consequences of low TH levels, and therefore little is known about the effects of TSH and FT4 levels over the entire range, including high TH levels. For these reasons, we investigated the association between maternal TSH and FT4 levels during early pregnancy and child IQ.

Participants and Methods: Maternal serum TSH and FT4 levels were determined between pregnancy week 9 and 18, and child IQ was measured at median age 6 years in 3839 mother-child pairs from the Generation R study, an iodine sufficient population. All analyses were adjusted for hCG, TPO-antibody positivity, gestational age, maternal age, parity, smoking, ethnicity, education level, child gender and birth weight. Additionally, in a subset of mother-child pairs (N=2460) child TSH and FT4 levels at birth and at time of IQ measurement were available.

Results: There was a strong and independent, U-shaped association between maternal FT4 levels and child IQ (P=0.003). A similar association with the risk of borderline intellectual functioning (defined as IQ <85) was present (P=0.003).
We performed sensitivity analyses which investigated the maternal FT4 threshold value for significantly decreased mean child IQ levels. Low FT4 levels at or below the 8th percentile were associated with a significantly decreased mean child IQ (range -2.1 to -3.8 points; P≤0.01). For high FT4 levels, mean child IQ levels were significantly decreased at or above the 89th percentile (range -2.0 to -3.8 points; P≤0.01).
TSH levels were also associated with child IQ levels (P=0.04), but this was driven by their reflection of FT4 levels and thus not independent (P=0.37 after adjustment). Results remained unchanged after correction for hCG levels or child TSH and FT4 levels.

Conclusions: This is the first study to show that maternal FT4 levels during early pregnancy are continuously associated with child IQ levels in a U-shape manner. According to our sensitivity analyses, brain development during early life may be affected by maternal FT4 levels in up to 19% of all children. Because high FT4 levels have similar detrimental effects as low FT4 levels, L-T4 treatment aiming for high-normal thyroid function tests may not be completely without risks. 

 

Nothing to Disclose: TIK, MM, HT, EV, TJV, RPP

OR11-4 18809 4.0000 A Novel Insights into the Effects of Maternal Thyroid Function on Child IQ Reveal Detrimental Effects of High FT4 Levels 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 1:00:00 PM OR11 6031 11:30:00 AM Thyroid Development, Clinical and Autoimmunity Oral


Alon Grossman*1, Avraham Weiss2, Nira Koren Morag3, Ilan Shimon4 and Joseph Meyerovitch5
1Rabin Medical Center Beilinson Campus, Petah tikva, Israel, 2Rabin medcial Center, Beilinson Campus, Petah Tikva, Israel, 3Sackler Faculty of Medicine, Tel Aviv University, Israel, 4Rabin Medical Center and Sackler School of Medicine, Tel-Aviv University, Petah-Tiqva, Israel, 5Schneider Children's Medical Center of Israel and Sackler Faculty of Medicine, Tel Aviv University, Ramat Hasharon, Israel

 

Introduction: The association between subclinical hypothyroidism (scH) and hyperthyroidism (scHy) and mortality in the elderly is poorly defined. In this study this association was evaluated and the thyroid stimulating hormone (TSH) values associated with excess mortality in the elderly were defined.

Methods:  A computerized database of a large health care organization (Clalit Health Medical Organization) was reviewed.  All cases of known thyroid disease or cases in which thyroid medications were dispensed were excluded.  Patients older than 65 years evaluated in the years 2002-2012 (end of follow-up 31.12.12) with documented normal free T4 values were included in the analysis and were divided into 3 groups based on TSH values: normal (normal TSH), scH (TSH > 4.2 mIU/L) and scHy (TSH <0.35 mIU/L). Long term Mortality HR was compared between the three groups and a sub analysis according to TSH values was performed in those with scH and scHy.

Results:  Final analysis was performed on 17,440 individuals with subclinical thyroid disease {538 scHy (3.1%), 1956 scH (11.2%), normal 14,946 (85.7%) , average age 83±6.5 years} who were followed-up for 10 years.  Both scH (HR= 1.75 CI 1.63-1.88) and scHy (HR= 2.33 CI 2.08-2.63) were associated with significantly increased mortality that persisted on multivariate analysis for age, gender, chronic kidney disease, chronic obstructive lung disease, smoking, dementia, Parkinson's disease, cerebrovascular disease, congestive heart failure, diabetes mellitus and hypertension (scH HR=1.68 CI 1.56-1.8, ScHy HR=1.93 CI 1.7- 2.17 ).  TSH values > 6.35 mIU/L were associated with the highest mortality in those with scH, whereas in scHy, no threshold for increased mortality was identified.

Conclusions:  Both scH and scHy are associated with increased mortality in the elderly. A threshold TSH value (>6.35 mIU/L) exists for increased mortality in scH, but not in scHy.

 

Disclosure: IS: Advisory Board, Novartis Pharmaceuticals, Researcher, Novartis Pharmaceuticals, Speaker, Novartis Pharmaceuticals, Consultant, Pfizer, Inc., Researcher, Pfizer, Inc., Speaker, Pfizer, Inc., Consultant, Chiasma. Nothing to Disclose: AG, AW, NK, JM

OR11-5 20227 5.0000 A Subclinical Thyroid Disease Increases Mortality in the Elderly 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 1:00:00 PM OR11 6031 11:30:00 AM Thyroid Development, Clinical and Autoimmunity Oral


Steffy Wilhelmina Jansen1, Abimbola Akintola1, Ferdinand Roelfsema2, Evie van der Spoel1, Christa M Cobbaert2, Bart EPB Ballieux1, Peter Egri3, Zsuzanna Kvarta-Papp3, Balazs Gereben3, Csaba Fekete4, Petronella E. Slagboom1, Jeroen van der Grond5, Barbara Demeneix6, Hanno Pijl2, Rudi G J Westendorp1 and Diana van Heemst*1
1Leiden University Medical Center, Netherlands, 2Leiden University Medical Center, Leiden, Netherlands, 3Hungarian Academy of Sciences, Hungary, 4Institute of Experimental Medicine, Budapest, Hungary, 5Leiden University Medical Center, 6Muséum National d'Histoire Naturelle, Paris CEDEX 05, France

 

Only few studies have included subjects with a propensity to reach old age in good health, aiming to disentangle mechanisms contributing to staying healthier for longer. Greater longevity has been associated with higher TSH levels in humans and lower thyroid hormones (TH) levels in animals(1), but the mechanisms underlying TSH/TH differences and longevity are unknown. In this study, we assessed TSH and thyroid hormone regulation in relation with familial longevity. We included 112 subject of the Leiden Longevity study cohort consisting of 61 middle-aged offspring of nonagenarian siblings (offspring) together with 51 of their partners (controls) not originated from long-lived families. All subjects underwent measurements of energy metabolism consisting of an indirect calorimetry measurement and continuous core body temperature measurements. We measured bone resorption and bone formation markers as indicators of bone tissue turnover. In a subset of 20 offspring and 18 controls, blood was frequently sampled over 24 hours, from which circulating TSH levels were measured every 10 minutes and levels of TH every hour. Using deconvolution analysis specific features of HTP-axis function were assessed, including secretion rates, number of pulses, and regularity of pulses. Total geometric mean (95% confidence interval (CI)) TSH secretion was significantly (P < 0.007) higher in the offspring (55.0 (43.9-68.9) mU/l/24h) compared to the controls (34.4 (27.1-43.7) mU/l/24h). In line, offspring had on average, 0.8 mU/L higher serum levels of TSH at all time points. In contrast, circulating TH levels were similar between groups. TSH bioactivity was determined by measuring cAMP production in cultured Chinese hamster ovary cells stably transfected with the human TSH receptor. Offspring and partners had similar cAMP/TSH ratios indicating that their TSH molecules had similar bioactivity. Offspring and controls had similar resting metabolic rate and core body temperature. However, mean (95% CI) levels of the bone resorption marker beta-crosslaps were significantly (P = 0.02) lower in offspring (0.30 (0.28-0.34) ng/ml) compared to controls (0.36 (0.33-0.40) ng/ml), indicating a decreased bone turnover. Moreover, markers of bone renewal also tended to be lower in offspring from long-lived siblings. In conclusion, offspring of exceptionally long-lived humans are characterized by high total TSH secretion, decreased bone turnover, without changes in energy metabolism or circulating TH levels. We propose that pleiotropic effects of differences in thyroid status favor longevity by slowing rates of tissue turnover, without altering whole body metabolism. Our findings provide new targets for studying mechanisms to improve healthy life expectancy, including experimental studies to determine the effects of TSH on tissue regeneration and repair.

 

Nothing to Disclose: SWJ, AA, FR, EV, CMC, BEB, PE, ZK, BG, CF, PES, JV, BD, HP, RGJW, DV

OR11-6 21703 6.0000 A Human Longevity Is Characterized By Higher Total Thyroid Stimulating Hormone Secretion and Similar Levels of Thyroid Hormone 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Thyroid/HPT Axis Thursday, March 5th 1:00:00 PM OR11 6031 11:30:00 AM Thyroid Development, Clinical and Autoimmunity Oral


Corey J Lager*1, Nazanene H Esfandiari1, Angela R Subauste2, Andrew T Kraftson3, Morton B Brown1, Oliver A Varban1, Catherine K Nay3, Amy L Lockwood1 and Elif Arioglu Oral1
1Univ of Michigan, Ann Arbor, MI, 2University of Mississippi Medical Center, Jackson, MS, 3University of Michigan, Ann Arbor, MI

 

Background:Roux-en-Y Gastric Bypass (GB) and Sleeve Gastrectomy (SG) are the most common bariatric procedures used to treat obesity and its metabolic complications. It has been proposed that these surgeries result in comparable long-term outcomes.

Methods:Using data from the UMHS Post-Bariatric Surgery Program, we performed a retrospective analysis to compare weight loss and change in BP, BMI, A1C, lipids, blood indices as well as common vitamin levels between surgeries. Serum creatinine was evaluated as an indirect marker of lean mass. Data were collected at baseline and 2, 6, 12, and 24 months (m) post-surgery. This analysis does not include the early (30-day) post-operative period and the data on immediate surgical complications. Changes between time points and baseline were compared between groups by the Wilcoxon rank sum test. Results are reported as mean±SE.

Results: Data from 392 patients post-GB (age 44±1 years, 81% females) and 337 patients post-SG (age 45±1 years, 77% females) who had their surgeries at our institution from 1/1 2008 to 11/20/2013 were included. To date, 269 GB and 203 SG patients completed 12m follow up and of these 186 GB and 89 SG had 24m follow up. At baseline, BMI and weight were significantly lower in the GB group compared to the SG group (47.2±0.4 kg/m2 and 133±1 kg vs. 49.8±0.5 kg/m2 and 141±2 kg, p<0.0001). GB experienced greater and more rapid weight loss: GB group lost 68.6±2.6 % of excess body weight by 12m vs. 55.0±1.4 % noted in SG group (p<0.0001). This difference was preserved at 24m (65.6±4.1% vs.52.0 ±2.2 %, respectively, p<0.0001). HbA1C improved in both groups with a decrease of 1.40±0.2% following GB and 0.75±0.15% following SG (p =0.01 at 12m). The GB group displayed a fall in serum creatinine that reached statistical difference at 2m and sustained for long-term while the SG group experienced a less remarkable decrease in serum creatinine (0.10±0.01 mg/dL following GB vs 0.04±0.01 following SG, p<0.0001 at 12 m). Both groups had a fall in Hemoglobin (Hb) at 2m, with GB patients displaying significantly greater fall compared to SG (1.14±0.08 g/dL vs. 0.71±0.07 g/dL, p= 0.0002). The fall in Hb was not protracted and improved back to baseline by 6m. We did not observe a significant difference with respect to circulating B12 or Vitamin D levels at baseline or longitudinally; however, patients completing studies especially at later time points were not sufficient to conclude on true differences between the two groups.

Conclusions: Our data demonstrate greater weight loss and metabolic improvements in patients undergoing GB when compared to SG after 2 years. GB patients experienced a larger decrease in Hb within the first two months, and a sustained decrease in creatinine. A consideration of immediate surgical complications and prospective longitudinal data are required in order to truly assess risks and benefits of SG and GB.

 

Disclosure: EAO: Principal Investigator, GI Dynamics. Nothing to Disclose: CJL, NHE, ARS, ATK, MBB, OAV, CKN, ALL

OR07-1 19028 1.0000 A The Two Obesity Surgeries: Are They Truly Comparable? 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 1:00:00 PM OR07 6038 11:30:00 AM Obesity: Clinical Studies Oral


David C Lau*1, Melanie Davies2, F Xavier Pi-Sunyer3, John Wilding4, Linda Shapiro Manning5, Christine Bjørn Jensen5 and Sten Madsbad6
1University of Calgary, Calgary, AB, Canada, 2University of Leicester, Leicester, United Kingdom, 3St Luke's Roosevelt Hosp, New York, NY, 4University of Liverpool, Liverpool, United Kingdom, 5Novo Nordisk A/S, Søborg, Denmark, 6Hvidovre Hospital, Hvidovre, Denmark

 

Liraglutide is a glucagon‐like peptide‐1 analog approved for type 2 diabetes (T2D) at doses up to 1.8 mg. We present here data on the effects of liraglutide 3.0 mg on glycemic excursions and hypoglycemia in overweight and obese adults without T2D from the 56-week SCALE Obesity and Prediabetes trial (Clinicaltrials.gov ID:NCT01272219).

Individuals (BMI ≥27 kg/m2 with ≥1 comorbidity, or ≥30 kg/m2; 78.5% female, 61.2% with prediabetes, 45.1 yrs, body weight 106.2 kg, BMI 38.3 kg/m2 [all means]) were randomized 2:1 to receive once-daily liraglutide 3.0 mg (n=2487) or placebo (n=1244), both as adjunct to 500 kcal/day deficit diet and increased physical activity (≥150 min/week). Changes in fasting (FPG) and postprandial plasma glucose (PPG) (determined during an oral glucose tolerance test; OGTT), were assessed at baseline and week 56. Hypoglycemic events were reported as adverse events (AEs), by individuals based on symptoms (spontaneous, not biochemically confirmed) between visits, or by investigators based on glucose values ≤70 mg/dL during visits (including OGTT), irrespective of symptoms (paleness, shaking, sweating, increased pulse/heartbeat, hunger, vision disorder, unusual behavior or drowsiness).

From baseline to 56 weeks, greater reductions were seen with liraglutide vs placebo in body weight (-8.0% vs -2.6%), FPG (-7.1 mg/dL vs. 0.1 mg/dL) and PPG (-46.8 mg/dL vs -10.4 mg/dL) [observed means LOCF; p<0.0001, ANCOVA]. Improvements in glycemia were more pronounced in individuals with prediabetes than without.

No hypoglycemia AEs required third-party assistance. Similar proportions of individuals on liraglutide (1.3%) and placebo (1.0%) reported hypoglycemia based on symptoms between visits. More individuals with liraglutide vs. placebo had biochemical hypoglycemia AEs during FPG visits (3.6% vs 0.8%) or OGTT (8.3% vs 1.4%), consistent with the observed effects of liraglutide on FPG/PPG. Of biochemical events reported during FPG and OGTT visits, respectively, 31.3% and 41.5% with liraglutide vs. 30.0% and 33.0% with placebo were asymptomatic. As expected with the glucose dependency of liraglutide, only a small proportion of events reported at visits were associated with PG <56 mg/dL; a more clinically relevant threshold for hypoglycemia in individuals without T2D (FPG visits: 0.1% vs 0%, OGTT visits: 2.3% vs 0%, for liraglutide and placebo, respectively). Overall hypoglycemia AEs were less common in those with prediabetes than without (liraglutide: 9.4% vs 15.9%; placebo 2.6% vs 4.3%, respectively), consistent with a higher mean baseline FPG and PPG.

In conclusion, in overweight and obese individuals without diabetes, liraglutide 3.0 mg, as adjunct to diet and exercise, was associated with greater reductions in fasting and postprandial glucose compared with diet and exercise alone. Clinically significant hypoglycemia with glucose <56 mg/dL was rare with both treatments.

 

Disclosure: DCL: Investigator, Astra Zeneca, Investigator, Boehringer Ingelheim, Investigator, Bristol-Myers Squibb, Investigator, Eli Lilly & Company, Investigator, Novo Nordisk, Advisory Group Member, Amgen, Advisory Group Member, Astra Zeneca, Advisory Group Member, Boehringer Ingelheim, Advisory Group Member, Bristol-Myers Squibb, Advisory Group Member, Janssen, Advisory Group Member, Eli Lilly & Company, Advisory Group Member, Novo Nordisk, Advisory Group Member, Roche Pharmaceuticals, Advisory Group Member, Valeant, Speaker Bureau Member, Amgen, Speaker Bureau Member, Astra Zeneca, Speaker Bureau Member, Boehringer Ingelheim, Speaker Bureau Member, Bristol-Myers Squibb, Speaker Bureau Member, Janssen, Speaker Bureau Member, Eli Lilly & Company, Speaker Bureau Member, Merck & Co., Speaker Bureau Member, Novo Nordisk, Speaker Bureau Member, Valeant. MD: Advisory Group Member, Novo Nordisk, Consultant, Novo Nordisk, Investigator, Novo Nordisk, Speaker Bureau Member, Novo Nordisk, Advisory Group Member, Sanofi, Consultant, Sanofi, Investigator, Sanofi, Speaker Bureau Member, Sanofi, Advisory Group Member, Eli Lilly & Company, Consultant, Eli Lilly & Company, Investigator, Eli Lilly & Company, Speaker Bureau Member, Eli Lilly & Company, Advisory Group Member, Merck Sharp & Dohme, Consultant, Merck Sharp & Dohme, Speaker Bureau Member, Merck Sharp & Dohme, Advisory Group Member, Boehringer Ingelheim, Consultant, Boehringer Ingelheim, Speaker Bureau Member, Boehringer Ingelheim, Advisory Group Member, Astra Zeneca, Consultant, Astra Zeneca, Speaker Bureau Member, Astra Zeneca, Advisory Group Member, Janssen, Consultant, Janssen, Speaker Bureau Member, Janssen, Speaker Bureau Member, Mitsubishi Tanabe Pharma Corp. FXP: Advisory Group Member, Novo Nordisk, Advisory Group Member, Weight Watchers International, Advisory Group Member, Johnson &Johnson, Advisory Group Member, Vivus USA, Advisory Group Member, Eisai, Advisory Group Member, Zafgen. JW: Advisory Group Member, Astra Zeneca, Speaker Bureau Member, Astra Zeneca, Investigator, Astra Zeneca, Speaker Bureau Member, Boehringer Ingelheim, Advisory Group Member, Bristol-Myers Squibb, Investigator, Bristol-Myers Squibb, Speaker Bureau Member, Bristol-Myers Squibb, Advisory Group Member, Janssen, Speaker Bureau Member, Janssen, Advisory Group Member, Novo Nordisk, Speaker Bureau Member, Novo Nordisk, Investigator, Novo Nordisk, Speaker Bureau Member, Merck & Co., Consultant, Pfizer Global R&D. LSM: Employee, Novo Nordisk. CBJ: Employee, Novo Nordisk, Employee, Novo Nordisk. SM: Advisory Group Member, Novo Nordisk, Speaker Bureau Member, Novo Nordisk, Investigator, Novo Nordisk.

OR07-2 19540 2.0000 A Hypoglycemia Reported with Liraglutide 3.0 Mg in Overweight and Obese Adults with and without Prediabetes: Results of the Randomized, Double-Blind, Placebo-Controlled 56-Week Scale Obesity and Prediabetes Trial 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 1:00:00 PM OR07 6038 11:30:00 AM Obesity: Clinical Studies Oral


Jovanna Dahlgren*1, Torsten Olbers2, Eva Gronowitz2, Carl-Erik Flodmark3 and Claude N Marcus4
1Inst of Clinical Sci, Gothenburg, Sweden, 2The University of Gothenburg, Gothenburg, Sweden, 3Skånes University hospital SUS, Malmö, Sweden, 4Karolinska Institute, Stockholm, Sweden

 

Introduction

Few studies have followed prospectively the long-term outcome of gastric bypass surgery in children and adolescents. In morbid obese adults, gastric bypass improves significantly cardiovascular profiles and the weight loss obtained is maintained over several decades. Here we present five-year data from the prospective non-randomized controlled study; Adolescent Morbid Obesity Surgery study (AMOS) using laparoscopic Roux-en-Y gastric bypass (LRYGB).

Methods

Eighty-one young pubertal non-syndromic obese adolescents (13.0 to 17.9 years of age, Tanner stage >3) underwent LRYGB. Inclusion criteria was BMI >40 or >35 with co-morbidities. Follow up visits were scheduled at one, two and five years post-surgery. Body composition was examined with DEXA (Lunar). A group of BMI and age matched adolescents were identified in a national registry and were assessed after five years. A BMI and gender matched adult group (35-45 years of age) undergoing LRYGB was also followed prospectively.

Results

Mean inclusion BMI was 45.5 kg/m2 in young surgically treated (YST), 42.2 kg/m2 in adolescent controls (AC) and 43.5 kg/m2 in adult surgically treated (AST). At five years postop, BMI of YST was 32 kg/m2 (total weight loss 27%, p<0.001) while BMI of AC was 43 kg/m2 (gained 10% in weight) and 31 of AST (weight loss 29%, p<0.001). In YST, waist circumference decreased from 134 to 98 cm (p<0.001) and body composition changes included a reduction in the percentage fat mass (51.8% to 40.9%, p<0.0001) as well as a relative increase in lean mass (47.0% to 56.8%, p<0.0001) over the two post-surgery years. Boys lost more fat mass than girls (-17.3% vs. -9.5%, p<0.0001). Fasting insulin in YST decreased from 32 to 7 mU/L the first post-surgery year and continued unchanged low during two and five years post-surgery, whereas levels in AC were above 30 mU/L (p<0.001). The same pattern was seen for supersensitive CRP, which decreased significantly in YST from 7.2 mg/L to 2.5 the first year post-surgery. 

Conclusion

Laparoscopic gastric bypass appears equally effective in adolescents as in adults regarding weight loss and metabolic improvements up to five years after surgery.  There seems to be substantial gender difference in changes in body composition, with increased loss of fat mass in males. If this has an implication on differences in long-term metabolic outcomes between genders merits further evaluation.

 

Nothing to Disclose: JD, TO, EG, CEF, CNM

OR07-3 21767 3.0000 A A Swedish Nationwide Study on Metabolic Longterm Outcome of Laparoscopic Roux-En-Y Gastric Bypass for Children with Severe Obesity 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 1:00:00 PM OR07 6038 11:30:00 AM Obesity: Clinical Studies Oral


Ovidiu A Galescu*1, Andrew Paul Demidowich1, Annie M Altschul1, Sheila M Brady1, Sara A Armaiz Flores1, Van S Hubbard2 and Jack Adam Yanovski1
1Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH), Bethesda, MD, 2National Institute of Diabetes and Digestive and Kidney Diseases/NIH, Bethesda, MD

 

Background:Pediatric obesity is associated with advancement of skeletal maturation and increase in height during childhood. The accuracy of skeletal age-based adult height predictions among obese children has not been well established. We hypothesized that skeletal age-based height predictions would overestimate final adult height among obese (OB) as compared to non-obese (NO) children.

Methods: A sample of NO (BMI percentile <95th %) and OB (BMI percentile ≥95th %) children ages 5.5- 15y were prospectively enrolled for longitudinal investigation. Age- and sex-specific BMI percentiles were calculated according to CDC 2000 standards. Skeletal age was assigned using the Greulich and Pyle atlas and predicted adult height (PH) was calculated according to the Bayley-Pinneau tables. Participants were followed to near-final adult height (FH) as determined by age / skeletal age >14y for girls; >15.5y for boys).

Statistics:The primary analysis was ANCOVA, with the difference between PH and FH (PH-FH) as the dependent variable, and age, sex, race, obesity status, and the sex * obesity status interaction as independent variables.

Results:A total of 191 patients (mean age 9.3±1.8 y; 52.9% female and 47.1% male; 60.2% white, 37.7% black, 2.1% Asian) were enrolled and followed to FH.  At baseline, the mean bone age was 9.9±1.9y; 50% were prepubertal and 34% were obese.  There were no statistically significant differences between the NO and OB groups in sex, race, or mid-parental target height. At baseline, the OB children were younger than NO (8.7±1.4 vs. 9.5±1.8y, p=0.002) but the bone age relative to chronological age was significantly greater among the OB group (+1.5±1.1 vs. +0.3±0.9y, p<0.0001) and height SD score was significantly greater in the OB group (+1.45±0.97 vs. +0.29±0.99, p<0.0001).  Final adult height was documented at mean age 19.9 ±3.2y. In ANCOVA, PH-FH was significantly predicted by sex, age, and the interaction between sex and obesity status. Among obese girls FH was significantly underestimated by PH (-2.3±1.1, CI -4.5 to -0.08cm, p=0.035); however, among obese boys, FH appeared to be overestimated by PH (+2.4±1.2, CI -0.01 to +4.7cm). Among non-obese children, there were no significant differences found between FH and PH.

Discussion: In a large pediatric cohort enriched for obesity, followed prospectively to adult height, obese children were initially taller and had advanced skeletal age, consistent with adipose tissue aromatase action increasing circulating estrogens. Surprisingly in obese girls, despite their tendency to have earlier onset of puberty, FH was underestimated, while in obese boys, who generally show a delayed entry into gonadarche, FH showed a trend towards being overestimated by the Bayley-Pinneau method. These data suggest the Bayley Pinneau tables for predicting adult height using skeletal age may need to be revised to take into account obesity.

 

Disclosure: JAY: Principal Investigator, Zafgen. Nothing to Disclose: OAG, APD, AMA, SMB, SAA, VSH

OR07-4 19385 4.0000 A Obesity and Predicted Height in Children: A Longitudinal Analysis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 1:00:00 PM OR07 6038 11:30:00 AM Obesity: Clinical Studies Oral


Lena MS Carlsson*1, Kajsa Sjoholm1, Per-Arne Svensson1, Peter Jacobson1, Lars Sjöström1 and Markku Peltonen2
1The Sahlgrenska Academy at University of Gothenburg, Gothenburg, Sweden, 2National Institute for Health and Welfare, Helsinki, Finland

 

Background: Obesity increases the risk for diabetes, and diabetes is associated with microvascular and macrovascular complications. We have shown that bariatric surgery strongly reduces the risk for diabetes in obese patients (1) and prevents microvascular and macrovascular complications in obese patients with diabetes (2). It is unknown if the prevention of diabetes by bariatric surgery also leads to reduced long-term risk for diabetes-associated complications when compared to non-surgical obesity care.

Aim: To determine the effects of bariatric surgery on the long-term risk of microvascular and macrovascular events in obese patients without diabetes at baseline.

Methods: The Swedish Obese Subjects (SOS) study is a prospective matched intervention study conducted at 25 surgical departments and 480 primary healthcare centers in Sweden. Participants were 37 to 60 years of age, and BMI was 34 or more in men and 38 or more in women. The current analyses included 3429 patients in the SOS study that did not have diabetes at baseline. Of them, 1658 patients underwent bariatric surgery (19% banding, 69% vertical banded gastroplasty, 12% gastric bypass) and 1771 obese matched controls given usual care. Microvascular (eyes, kidneys and peripheral nerves) and macrovascular (legs, heart and brain) events requiring hospital or specialist outpatient treatment or that were associated with death during follow-up were traced by searching the Swedish Cause of Death Register and the Swedish National Patient Register. Median follow up time was 18 years.

Results: Bariatric surgery reduced the incidence of microvascular plus macrovascular events, whichever came first, in patients without diabetes at baseline (566 and 411 events in the control and surgery groups, respectively; HR=0.73; 95%CI: 0.64-0.82; P<0.001). When the incidence of microvascular and macrovascular events were analyzed separately, both were reduced in the surgery group compared to controls (HR=0.48; 95%CI:0.38-0.60; P<0.001 and HR=0.80; 95%CI:0.70-0.92; P<0.002 for microvascular and macrovascular events, respectively). The reduction of the incidence of microvascular plus macrovascular events was more pronounced in patients with prediabetes at baseline compared to those with normal glucose status (HR=0.52; 95%CI: 0.40-0.68; P<0.001 and HR=0.79; 95%CI: 0.68-0.91; P<0.001, respectively; interaction P-value=0.005).

Conclusion: Bariatric surgery reduced the long-term risk of diabetes-associated vascular disease in obese patients without diabetes at study start.

 

Disclosure: LMC: Speaker, Johnson &Johnson. LS: Speaker, Astra Zeneca, Speaker, Johnson &Johnson. Nothing to Disclose: KS, PAS, PJ, MP

OR07-5 19794 5.0000 A Bariatric Surgery Reduces the Long-Term Incidence of Diabetes-Associated Vascular Disease in Initially Non-Diabetic Obese Patients 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 1:00:00 PM OR07 6038 11:30:00 AM Obesity: Clinical Studies Oral


Ralph DeFronzo1, Richard Mauritz Bergenstal2, Melanie Davies3, Robert Kushner4, Trine Vang Skjøth5, Birgitte Claudius5 and Ofri Mosenzon*6
1University of Texas Hlth Science Ctr, San Antonio, TX, 2HealthPartners Institute, Minneapolis, MN, 3University of Leicester, Leicester, United Kingdom, 4Northwestern University Feinberg School of Medicine, Chicago, IL, 5Novo Nordisk A/S, Søborg, Denmark, 6Hadassah University Hospital, Jerusalem, Israel

 

SCALE Diabetes (NCT01272232); a 56-week, randomized, double-blind, placebo-controlled trial evaluating the efficacy and safety of liraglutide for weight management in overweight/obese adults with T2D. 846 individuals were randomized 2:1:1 to liraglutide 3.0 mg (n=423), 1.8 mg (n=211) or placebo (n=212) as an adjunct to diet and exercise. 50% male, mean age 54.9 years, BMI 37.1 kg/m2, A1c 7.9%, and T2D duration 7.3 years.

This subgroup analysis compares efficacy and safety in responders (weight loss of ≥5% from baseline at week 56) vs non-responders. Changes from baseline data are least-squares means (efficacy) or observed means (safety) with LOCF. Pulse data are observed means for completers.

Mean weight loss for responders vs non-responders was 10.3% vs 1.6% with liraglutide 3.0 mg, 10.4% vs 1.3% with liraglutide 1.8 mg and 9.7% vs 0.7% with placebo, although more individuals in the liraglutide 3.0 mg (49.9%, n=205) and 1.8 mg (35.6%, n=72) groups than the placebo group (13.8%, n=29) were responders (p<0.0001).

Changes in A1c from baseline with liraglutide 3.0 mg, 1.8 mg and placebo were -1.6 and -1.5 vs -1.1% for responders, and -1.0, -1.0% and -0.2% for non-responders. SBP was reduced by -5.0, -6.2 and -5.3 mmHg in responders with liraglutide 3.0 mg, liraglutide 1.8 mg and placebo respectively and -0.5, -1.9 and 0.5 mmHg in non-responders. Physical function scores using IWQoL-LITE improved by 19.0, 16.4 and 15.5 with liraglutide 3.0 mg, liraglutide 1.8 mg and placebo, respectively, in responders vs 11.1, 10.4 and 7.7 in non-responders.

In responders vs non-responders adverse events (AEs) were reported by 96% vs 92% with liraglutide 3.0 mg, 96% vs 90% with liraglutide 1.8 mg and 94% vs.85% with placebo. For serious AEs no marked differences between responders and non-responders were observed. The most frequent AEs were GI disorders, which were more frequent in responders vs non-responders for liraglutide 3.0 mg (76% vs 55%) but not different for liraglutide 1.8 mg (58% vs 57%) or placebo (41% vs 39%). Rates (events/patient year) of documented symptomatic hypoglycemia (plasma glucose ≤56 mg/dL) were similar in responders vs non-responders across all groups, liraglutide 3.0 mg: 0.9 vs 0.8; liraglutide 1.8 mg: 0.8 vs 1.1; placebo: 0.1 vs 0.3. Mean (SD) change in pulse rate from baseline for liraglutide 3.0 mg, liraglutide 1.8 mg and placebo was 1.3 (9.5), 2.0 (10.7) and -4.4 (10.1) bpm for responders and 3.5 (10.2), 3.1 (10.1) and -1.2 (9.0) bpm for non-responders.

In conclusion, a higher proportion of individuals achieved a clinically meaningful ≥5% weight loss at 56 weeks with liraglutide 3.0 mg and 1.8 mg, compared with placebo. The responder groups had a total weight loss at 56 weeks of approx 10%. Responders in the liraglutide groups had greater improvements in A1c, indicating an additional benefit of treatment above that attributed to weight loss. Similar overall incidence of AEs was observed in subgroups.

 

Disclosure: RD: Advisory Group Member, Bristol-Myers Squibb, Advisory Group Member, Takeda, Advisory Group Member, Amylin Pharmaceuticals, Advisory Group Member, Novo Nordisk, Advisory Group Member, Boehringer Ingelheim, Advisory Group Member, Jansen Pharmaceuticals, Advisory Group Member, Lexicon Pharmaceuticals, Inc., Advisory Group Member, Astra Zeneca, Investigator, Amylin Pharmaceuticals, Investigator, Takeda, Investigator, Bristol-Myers Squibb, Investigator, Boehringer Ingelheim, Speaker Bureau Member, Bristol-Myers Squibb, Speaker Bureau Member, Astra Zeneca, Speaker Bureau Member, Novo Nordisk, Speaker Bureau Member, Jansen Pharmaceuticals. RMB: Advisory Group Member, Abbott Laboratories, Investigator, Abbott Laboratories, Advisory Group Member, Bayer, Inc., Investigator, Bayer, Inc., Advisory Group Member, Valeritas Inc, Investigator, Becton Dickinson, Advisory Group Member, Boehringer Ingelheim, Investigator, Boehringer Ingelheim, Advisory Group Member, Bristol-Myers Squibb, Investigator, Bristol-Myers Squibb, Advisory Group Member, Calibra, Investigator, Calibra, Advisory Group Member, Eli Lilly & Company, Investigator, Eli Lilly & Company, Advisory Group Member, Halozyme, Investigator, Halozyme, Advisory Group Member, Helmsley Trust, Investigator, Helmsley Trust, Advisory Group Member, Hygieia, Investigator, Hygieia, Advisory Group Member, Johnson &Johnson, Investigator, Johnson &Johnson, Advisory Group Member, Medtronic Minimed, Investigator, Medtronic Minimed, Investigator, Merck & Co., Advisory Group Member, Novo Nordisk, Investigator, Novo Nordisk, Investigator, NIH, Investigator, ResMed, Advisory Group Member, Roche Pharmaceuticals, Investigator, Roche Pharmaceuticals, Advisory Group Member, Sanofi, Investigator, Sanofi, Advisory Group Member, Takeda, Investigator, Takeda. MD: Advisory Group Member, Novo Nordisk, Consultant, Novo Nordisk, Investigator, Novo Nordisk, Speaker Bureau Member, Novo Nordisk, Advisory Group Member, Sanofi, Consultant, Sanofi, Investigator, Sanofi, Speaker Bureau Member, Sanofi, Advisory Group Member, Eli Lilly & Company, Consultant, Eli Lilly & Company, Investigator, Eli Lilly & Company, Speaker Bureau Member, Eli Lilly & Company, Advisory Group Member, Merck Sharp & Dohme, Consultant, Merck Sharp & Dohme, Speaker Bureau Member, Merck Sharp & Dohme, Advisory Group Member, Boehringer Ingelheim, Consultant, Boehringer Ingelheim, Speaker Bureau Member, Boehringer Ingelheim, Advisory Group Member, Astra Zeneca, Consultant, Astra Zeneca, Speaker Bureau Member, Astra Zeneca, Advisory Group Member, Janssen, Consultant, Janssen, Speaker Bureau Member, Janssen, Speaker Bureau Member, Mitsubishi Tanabe Pharma Corp. RK: Advisory Group Member, Novo Nordisk, Advisory Group Member, Vivus USA, Advisory Group Member, Zafgen, Advisory Group Member, Retrofit, Consultant, Eisai, Consultant, Takeda, Investigator, Weight Watchers, Investigator, Aspire Bariatrics. TVS: Employee, Novo Nordisk, Employee, Novo Nordisk. BC: Employee, Novo Nordisk, Employee, Novo Nordisk. OM: Advisory Group Member, Eli Lilly & Company, Advisory Group Member, Novo Nordisk, Advisory Group Member, Bristol-Myers Squibb, Advisory Group Member, Astra Zeneca, Speaker Bureau Member, Merck Sharp & Dohme, Speaker Bureau Member, Novartis Pharmaceuticals, Speaker Bureau Member, Sanofi, Speaker Bureau Member, Eli Lilly & Company, Speaker Bureau Member, Novo Nordisk, Speaker Bureau Member, Bristol-Myers Squibb, Speaker Bureau Member, Astra Zeneca, Investigator, Novo Nordisk, Advisory Group Member, Sanofi, Advisory Group Member, Novartis Pharmaceuticals.

OR07-6 19542 6.0000 A Efficacy and Safety of Liraglutide 3.0 Mg and 1.8 Mg in Weight Loss Responders from Overweight/Obese  Adults with Type 2 Diabetes (T2D): A Subgroup Analysis of the Scale Diabetes Trial 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Obesity, Adipocyte Biology and Appetite Thursday, March 5th 1:00:00 PM OR07 6038 11:30:00 AM Obesity: Clinical Studies Oral


Adi Cohen*1, David W Dempster2, Hua Zhou2, Robert R Recker3, Joan M. Lappe3, Serge Cremers4, Ralph Müller5, Alexander Zwahlen5, Mafo Kamanda-Kosseh1, Mariana Bucovsky1, Emily M. Stein1, Tom Nickolas1 and Elizabeth Shane1
1Columbia University, New York, NY, 2Helen Hayes Hospital, West Haverstraw, NY, 3Creighton University, Omaha, NE, 4Columbia University, College of Physicians and Surgeons, New York, NY, 5ETH Zurich, Institute for Biomechanics

 

Excess visceral adipose tissue (VAT) is associated with low BMD and poor bone structure. We recently reported that healthy premenopausal women with high DXA trunk fat, a surrogate for VAT, had significantly lower trabecular (Tb) bone volume fraction (BV/TV), Tb number and thickness, lower bone formation rate (BFR) on bone biopsies and serum bone formation markers. In postmenopausal women, serum LDL is inversely and HDL directly related to BMD. We therefore investigated relationships between lipids, BMD, bone microstructure and remodeling in premenopausal women.

In 40 healthy premenopausal women (37 ± 8 yrs; BMI 20-39 kg/m2) with normal BMD by DXA and no history of fracture, we measured body composition by DXA; Tb BV/TV, microstructure and BFR/BS on tetracycline-labeled bone biopsies; and serum IGF-1, sclerostin, bone turnover markers (BTM), HDL, TG, and calculated LDL on fasting morning serum.

As expected, LDL (±SD: 100 ± 25 mg/dL) and TG (87 ± 53 mg/dL) were directly and HDL (50 ± 15 mg/dL) inversely related to BMI and DXA trunk fat. HDL was directly related to Tb BV/TV (r=0.40; p=0.01) and BFR (0.40; p=0.01). We compared 23 women with HDL <50 mg/dL, the metabolic syndrome cutoff, to 17 with normal HDL (63 ± 11 mg/dL). BMI (27.8 ± 4.9 vs 23.0 ± 2.3 kg/m2) and %DXA trunk fat (36 ± 8 vs 27 ± 8) were higher in the low HDL group (both p<0.001). Age, BMD, HOMAIR, serum glucose, PTH, 25-OHD and reported exercise did not differ. The low HDL group had significantly lower BV/TV (21.0 ± 7.3 vs 27.3 ± 6.8%; p=0.009) and Tb thickness (148 ± 36 vs 179 ± 32 µm; p=0.008), and lower BFR (0.006 ± 0.004 vs 0.012 ± 0.008 mm2/mm/yr; p=0.01) and serum markers of resorption (C-telopeptide, 214 ± 78 vs 386 ± 241 pg/mL; p=0.01) and formation (osteocalcin, 13 ± 5 vs 19 ± 10 ng/mL, p=0.04; and PINP, 42 ± 11 vs 53 ± 23 µg/L, 0.07). The low HDL group also had significantly lower IGF-1 (168 ± 40 vs 206 ± 65 ng/mL; p=0.03) and higher sclerostin (0.50 ± 0.19 vs 0.36 ± 0.10 ng/mL; p=0.01), both potential mediators of the detrimental relationship between fat and bone. Relationships between HDL and Tb BV/TV and BFR were unaffected by controlling for reported exercise but no longer significant after controlling for DXA trunk fat.  Relationships between HDL and Tb BV/TV remained significant after controlling for serum IGF-1 (0.38; p=0.018) and sclerostin (0.33; p=0.046). Relationships between HDL and BFR remained significant after controlling for serum IGF-1 (0.36; p=0.029) but were attenuated (0.28; p=0.093) by controlling for sclerostin.  

In summary, healthy premenopausal women with HDL levels consistent with the definition of metabolic syndrome (< 50 mg/dL) were distinguished by significantly lower Tb bone formation and bone volume on bone biopsies. We conclude that HDL is an easily measured marker of excess VAT that may identify young women at risk for poor bone health for studies of the skeletal effects of interventions that promote weight loss or increase exercise.

 

Nothing to Disclose: AC, DWD, HZ, RRR, JML, SC, RM, AZ, MK, MB, EMS, TN, ES

LB-OR01-1 22931 1.0000 A Healthy Premenopausal Women with Low HDL (less than 50 mg/dL) Have Significantly Lower Trabecular Bone Volume and Bone Formation Rate 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM LB-OR01 6268 11:30:00 AM New Discoveries in Bone Health and Disease Oral


Skorn Ponrartana*1, Patricia C Aggabao1, Naga L Dharmavaram1, Carissa L Fisher1, Philippe Friedlich1, Sherin U Devaskar2 and Vicente Gilsanz1
1Children's Hospital Los Angeles, Keck School of Medicine, University of Southern California, Los Angeles, CA, 2Mattel Children's Hospital, UCLA, Los Angeles, CA

 

Background: The cross-sectional area (CSA) of the vertebra is smaller in females than in males throughout childhood, adolescence, and at the timing of peak bone mass – a major determinant of osteoporosis and future fracture risk. We examined whether these sex-related differences in the development of the axial skeleton are also present at birth.

Methods: Vertebral CSA, vertebral height, and intervertebral disc height were measured from the sixth thoracic to the fifth lumbar vertebrae using magnetic resonance imaging (MRI) in 70 healthy full-term newborns (35 male and 35 female). Additionally, measures of the length and CSA of the humerus, musculature, and adiposity were obtained.

Results: Weight, body length, head and waist circumferences, and MRI measures of musculature and adiposity did not significantly differ between sexes (all P’s ≥ 0.060). Compared to newborn boys, girls had significantly smaller vertebral cross-sectional dimensions (1.465 ± 0.11 vs. 1.309 ± 0.12; P < 0.0001) – a disparity that was independent of gestational age, birth weight, and body length. In contrast, sexes were monomorphic with regard to vertebral height, intervertebral disc height, and spinal length (all P’s ≥ 0.108). There were also no sex differences in the length or cross-sectional dimensions of the humerus (both P’s ≥ 0.151).

Conclusions: Factors related to sex influence fetal programming of the axial skeleton. The smaller vertebral CSA in females is associated with greater flexibility of the spine that could represent the human adaptation to fetal load. Unfortunately, it also imparts a mechanical disadvantage that increases stress within the vertebrae for all physical activities and the susceptibility for fragility fractures later in life.

 

Nothing to Disclose: SP, PCA, NLD, CLF, PF, SUD, VG

LB-OR01-2 22524 2.0000 A Differential Effect of Sex in Newborn Vertebrae and Risk of Osteoporosis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM LB-OR01 6268 11:30:00 AM New Discoveries in Bone Health and Disease Oral


Paul D Miller*1, Benjamin Zev Leder2, Gary Hattersley3, Edith Lau4, Peter Alexandersen5, Tomas Hala6, Sorica Mustatea7, Bettina Storgaard Nedergaard8, Annesofie Krogsaa9, Jan Slesinger10, Cristiano AF Zerbini11, Ivo Valter12, Zydrune Visockiene13, Beata Jendrych14, Carolina Aguiar Kulak15, Farid Marquez16, Alan G Harris17, Gregory C Williams3, Ming-yi Hu18, Bente Juel Riis19, Luis Augusto Russo20 and Claus Christiansen21
1Colorado Center for Bone Research, Lakewood, CO, 2Harvard Medical School, Boston, MA, 3Radius Health, Inc, Waltham, MA, 4CCBR Hong Kong, China, 5CCBR Vejle, Denmark, 6CCBR Pardubice, Czech Republic, 7CCBR Bucharest, Romania, 8CCBR Aalborg, Denmark, 9CCBR Ballerup, Denmark, 10CCBR Brno, Czech Republic, 11Centro Paulista de Investigação Clinica, São Paulo, Brazil, 12Center for Clinical and Basic Research, Tallinn, Estonia, 13CCBR Vilnius, Lithuania, 14CCBR Warsaw, Warsaw, Poland, 15Federal University of Parana, Curitiba, Brazil, 16Palm Springs Research Center, Hialeah, FL, 17Radius Health, Inc., Waltham, MA, 18Radius Health Inc, Waltham, MA, 19Nordic Biosciences, Copenhagen, Denmark, 20CCBR Rio De Janeiro, Rio de Janiero, Brazil, 21Nordic Bioscience, Copenhagen, Denmark

 

Abaloparatide (ABL) is an investigational osteoanabolic PTHrP analog being developed for the treatment of postmenopausal osteoporosis (PMO). In the ACTIVE double-blind, placebo-controlled (PBO) Phase 3 fracture prevention trial, 2463 PMO women were randomized to receive 18-months of either daily ABL 80-mcg SC, PBO SC or open label teriparatide (TPT) 20-mcg SC. All patients (PTS) received calcium and vitamin D supplements. 2,463 PTS were randomized (ITT population) and 1,901 (77.9%) completed. The groups were well matched for baseline demographics. PTS were 68.8 years old and the mean BMI was 25.1. 16.3% of PTS had 1 vertebral fracture, 28.2% had 2 or more vertebral fractures and 46.8% had at least 1 non-vertebral fracture. The mean spine, femoral neck and total hip baseline T-scores were -2.90, -2.14 and -1.90, respectively. Among the 2,118 PTS who had baseline and post-therapy X-rays, the fracture rate (FR) among those receiving ABL (n=690) was 0.58%, representing a reduction of new incident vertebral FR by 86% as compared to PBO PTS [n=711, FR 4.22%; (p<0.0001)]. The TPT group FR of 0.84% (n=717) was reduced by 80% (p<0.0001 versus PBO). ABL reduced non-vertebral fracture (Kaplan-Meier estimated (KM) FR=2.7%, hazard ratio=0.57, p=0.0489) and clinical fractures (KM FR=3.9%, hazard ratio=0.55, p=0.0112) versus PBO (KM non-vertebral FR=4.7%, KM clinical FR=8.3%). TPT was not significantly different from PBO in non-vertebral fracture (KM FR=3.3%, hazard ratio=0.72, p=0.2157) or clinical fracture (KM FR=4.8%, hazard ratio=0.71, p=0.1127). For wrist fractures ABL (n=824, KM FR 0.5%) and TPT (n=818, KM FR 2.0%) were not were significantly reduced compared to PBO (n=821, KM FR 1.5%); wrist FR was significantly less in ABL than the TPT group (p=0.0149). BMD increased more in ABL and TPT-treated PTS compared to PBO at the spine, femoral neck and hip at 6, 12 and 18 months (p<0.0001 for all comparisons). ABL also increased BMD at the hip at months 6, 12 (both p<0.0001) and 18 (p=0.0003), in the femoral neck at 6, 12 (both p<0.0001) and 18 (p=0.0016) and in the spine at 6 (p<0.0001) and 12 (p=0.0087) significantly more than TPT. ABL was well tolerated the most frequently reported adverse events were back pain, arthralgia, upper respiratory tract infection, hypercalciuria and dizziness. The overall incidence of hypercalcemia (HPC) based on albumin corrected serum calcium measured pre and post injection (4 hours) were 0.37%, 3.41%, and 6.36% for the PBO, ABL and TPT groups, respectively. Both ABL and TPT groups had significantly higher rates of HPC than PBO (p<0.0001), but for ABL was significantly less frequent than for TPT (p=0.0055). These results demonstrate that ABL reduces the incidence and risk of vertebral, non-vertebral, and clinical fractures in in women with PMO and suggest that ABL may prove to be an effective and safe treatment option in the management of PMO.

 

Disclosure: PDM: Principal Investigator, Radius. BZL: Principal Investigator, Radius. GH: Employee, Radius, Employee, Radius. EL: Principal Investigator, Radius. PA: Principal Investigator, Radius. TH: Principal Investigator, Radius. SM: Principal Investigator, Radius. BSN: Principal Investigator, Radius. AK: Principal Investigator, Radius. JS: Principal Investigator, Radius. CAZ: Principal Investigator, Radius. IV: Principal Investigator, Radius. ZV: Principal Investigator, Radius. BJ: Principal Investigator, Radius. CAK: Principal Investigator, Radius. FM: Principal Investigator, Radius. AGH: Employee, Radius, Employee, Radius. GCW: Employee, Radius, Employee, Radius. MYH: Employee, Radius, Employee, Radius. BJR: Collaborator, Radius. LAR: Collaborator, Radius. CC: Collaborator, Radius.

LB-OR01-3 22577 3.0000 A Effects of Abaloparatide on Vertebral and Non-Vertebral Fracture Incidence in Postmenopausal Women with Osteoporosis - Results of the Phase 3 Active Trial 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM LB-OR01 6268 11:30:00 AM New Discoveries in Bone Health and Disease Oral


Michael P. Whyte*1, Katherine L Madson2, Craig Frank Munns3, Amy L Reeves2, Kenji Fujita4, Hui Zhang5 and Nick James Bishop6
1Shriners Hosp for Children, Saint Louis, MO, 2Shriners Hospital for Children, St. Louis, MO, 3The Children's Hospital at Westmead, Sydney, Australia, 4Alexion Pharmaceuticals, Inc., Cheshire, CT, 5Alexion Pharmaceuticals, Cheshire, CT, 6Univ of Sheffield, Sheffield, United Kingdom

 

Hypophosphatasia (HPP) is the rare disease caused by loss-of-function mutation(s) in the tissue-nonspecific alkaline phosphatase (ALP) gene. Resultant low ALP activity can cause a wide spectrum of sequelae in children including premature loss of primary teeth, rickets, poor growth, muscle weakness, fractures, and pain (1 ,2). The clinical course of these manifestations is not well characterized.

This is a retrospective, multi-national, non-interventional chart review of patients (pts) with childhood HPP (symptom onset ≥6 months [mo] to <18 years [y]). Data were collected from 5 to 15 years of age or until Tanner stage >2, whichever occurred first. Study inclusion required HPP-related skeletal abnormalities, ≥1 set of paired radiographs 6 mo–5 y apart, and 2 height measurements ≥3 y apart. Children who previously received experimental or other treatments impacting bone or growth were excluded. Co-primary outcome measures were change in bone health (evaluated by the 7-point Radiographic Global Impression of Change (RGI-C) scale (-3 = severe worsening; +3 = complete healing) and change in height Z-score (Centers for Disease Control and Prevention [CDC] growth charts). Data are reported as median (min, max).

32 pts (69% boys, 97% white, 78% North American) were enrolled at 9 clinical sites, including 22 children from a large natural history database maintained at one site (2). Age at HPP symptom onset was 1.3 y (0.6, 3.4); age at diagnosis was 3.3 y (1.0, 13.1). Common HPP-related disease characteristics in this cohort included gait disturbance (59%), arthralgia (53%), bone pain (50%), muscle weakness (47%), muscle pain (38%), and fracture (34%). 28% of pts reported HPP-related hospitalizations, primarily for surgical correction of HPP-related deformity. 94% required non-pharmacological interventions such as physiotherapy (34%), orthotics (31%), and mobility aids (13%). 63% received medications for HPP-related complaints, most frequently for relief of pain and discomfort. No significant change between first and last assessment (LA) (time elapsed: 4.25 y [0.66, 7.95]) in radiographic assessment of bone health was observed (RGI-C was +0.33 [-2.33, +2.33; p=0.08]). Height and weight z-scores at first assessment varied considerably within the cohort (-0.86 [‑4.9, +2.6]; -0.86 [-5.0, +2.1], respectively) and all subsequent assessments, with no significant change to LA (p>0.05); at LA, height was -0.92 (-4.9, +1.8) and weight -0.98 (-5.7, +2.1).

As demonstrated by this retrospective chart review study, these children with childhood HPP had morbidity and no significant change in rickets or height z-score during childhood and early adolescence.

 

Disclosure: MPW: Principal Investigator, Alexion. KLM: Coinvestigator, Alexion. CFM: Advisory Group Member, Alexion. KF: Employee, Alexion. HZ: Employee, Alexion. NJB: Study Investigator, Alexion. Nothing to Disclose: ALR

LB-OR01-4 22822 4.0000 A A Retrospective, Multi-National, Non-Interventional, Natural History Study of the Childhood Form of Hypophosphatasia 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM LB-OR01 6268 11:30:00 AM New Discoveries in Bone Health and Disease Oral


Kevin J Basile*1, Alessandra Chesi2, Jonathan A. Mitchell3, Shana E. McCormack2, Sani M. Roy2, Heidi J. Kalkwarf4, Joan M. Lappe5, Vicente Gilsanz6, Sharon E Oberfield7, John A. Shepherd8, Andrea Kelly2, Babette S. Zemel2 and Struan F.A. Grant9
1Children's Hospital of Philadelphia, Philadelphia, PA, 2The Children's Hospital of Philadelphia, Philadelphia, PA, 3University of Pennsylvania, 4Cincinnati Children's Hospital Medical Center, Cincinnati, OH, 5Creighton University, Omaha, NE, 6Children's Hospital Los Angeles, Keck School of Medicine, University of Southern California, Los Angeles, CA, 7Columbia University College of Physicians and Surgeons, New York, NY, 8University of California San Francisco, San Francisco, CA, 9The Children's Hospital of Philadelphia, Philadelphia

 

Background: Height is a complex trait influenced by environmental and genetic factors operating in a number of biological pathways, including those related to hormone activity and bone accretion. The most recent meta-analysis of genome-wide association studies of adult height identified 697 variants corresponding to 423 distinct genomic loci, which collectively account for up to one-fifth of this trait’s heritability. However, the physiological route by which these genetic variants influence an individual’s overall height is not known.

 

Objective/Methods: Stature reflects skeletal size, which is achieved by bone mineral accretion during childhood growth. Thus, we investigated the effects of genetic variants known to associate with adult height on pediatric bone mineral content and density. We leveraged a prospective observational study of 691 participants of European ancestry enrolled in the Bone Mineral Density in Childhood Study (52% female), then sought further support from an additional cohort of 472 Caucasian subjects (51% female). Bone mineral content (BMC) and areal bone mineral density (aBMD) of the radius, hip, spine and total body (to estimate head and whole body BMC/aBMD) were assessed by dual energy X-ray absorptiometry (DXA) and expressed as age and sex-specific Z-scores. We further tested bone Z-scores adjusted for height-for-age Z-score to account for known effects of size on DXA outcomes. DNA was extracted from blood/saliva and genotyped using high-throughput technology. Genotyping of subjects was performed at the Children’s Hospital of Philadelphia using the Illumina Infinium™ II OMNI Express plus Exome BeadChip technology.

 

Results: Three of the loci previously linked to adult height yielded Bonferroni-corrected significance for association with pediatric bone accretion at specific skeletal sites. These loci were rs3750972 at TPCN2 for head aBMD in males (P = 5.48x10-5; ß = 0.241), rs6952113 at C7orf58 (also known as CPED1) for both BMC and aBMD at the distal radius in females (P = 1.63x10-5; ß = -0.250 and P = 6.30x10-8; ß = -0.317, respectively), and rs2781373 at MAX with aBMD at the distal radius in females (P = 3.22x10-5; ß = 0.243). These loci did not show any degree of significant association with height Z-score, nor did height adjustment of BMC/aBMD outcomes dramatically alter the significance of association of these loci to their respective skeletal phenotypes. Additionally, 19 other adult height loci demonstrated at least a nominally significant, directionally consistent association in both the discovery and replication cohorts for a variety of other skeletal phenotypes.

 

Conclusions: These results indicate that at least some of the genetic variants associated with adult height may be exerting their influence via alterations in pediatric bone accretion. Additionally, our data have implicated TPCN2 and MAX as novel pediatric bone density loci.

 

Nothing to Disclose: KJB, AC, JAM, SEM, SMR, HJK, JML, VG, SEO, JAS, AK, BSZ, SFAG

LB-OR01-5 22633 5.0000 A Multiple GWAS-Implicated Adult Height Loci Operate in the Context of Pediatric Bone Mineral Density and Content Determination 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM LB-OR01 6268 11:30:00 AM New Discoveries in Bone Health and Disease Oral


James Koh*1, Joyce Hogue1, Yuli Wang2, Matthew DiSalvo2, Nancy Lynn Allbritton2, Yuhong Shi3, John A Olson Jr.3 and Julie Ann Sosa4
1Duke University Medical Center, Durham, NC, 2University of North Carolina at Chapel Hill, 3University of Maryland School of Medicine, Baltimore, MD, 4Duke University, Durham, NC

 

We have recently shown that human parathyroid adenomas derived from patients with primary hyperparathyroidism (PHPT) are comprised of distinct cellular subpopulations with variable clonal origin and calcium responsiveness [1].  To gain a clearer understanding of the relationship between cellular identity, tumor composition, and clinical biochemistry in PHPT, we developed a novel single cell platform and computational analysis tool for quantitative evaluation of parathyroid tumor behavior. Dispersed parathyroid adenoma cells prepared from surgically resected tumor tissue are plated onto an indexed array composed of a large number of micron-scale elements (microrafts) doped with paramagnetic nanoparticles. The microrafts serve as releasable, optically transparent, individually addressed culture sites for isolated parathyroid cells.  Quantitative live-cell image analysis of calcium flux visualized by Fluo-4-AM epifluorescence was performed, and the cells were subsequently probed in situ for expression of the calcium sensing receptor (CASR), a canonical component in the extracellular calcium-signaling pathway.  Using this system, we found that the reactivity of individual parathyroid tumor cells to extracellular calcium stimulus is highly variable, with discrete patterns of kinetic response observed both between and among parathyroid tumor isolates. When challenged with 3 mM calcium, the proportion of cells exhibiting a rapid and transient response ranged from 7.1-27.7%, while sustained signaling responses were seen in 1.0–48.1%.  Non-responders at this calcium concentration ranged from 30.8-68.2%. When examined in relationship to each individual cell’s behavior, CASR abundance was not found to be a determinant of calcium responsiveness.  56.2 percent of rapid transient responder cells and 38.4 percent of sustained responder cells did not express detectable CASR protein.  Conversely, 32.3 percent of non-responsive cells were CASR-positive. Finally, we generated dose-response curves to calculate calcium EC50 values from a series of parathyroid adenomas and found that the tumors appeared to group into two distinct categories with respect to calcium responsiveness.  One group manifested a mean EC50 of 2.40 mM (95% CI 2.37–2.41), closely aligned to the published normal range [2].  The second group was clearly less responsive to calcium stimulus, with a mean EC50 of 3.61 mM (95% CI 3.45–3.95). This binary distribution indicates the existence of a previously unappreciated biochemical sub-categorization of PHPT tumors, possibly reflecting distinct etiological mechanisms.  Prospective identification of quantitative differences in calcium sensing could have important implications for the clinical management of PHPT.

 

Disclosure: NLA: Founder, Cell Microsystems. Nothing to Disclose: JK, JH, YW, MD, YS, JAO Jr., JAS

LB-OR01-6 22542 6.0000 A Live Single Cell Imaging of Human Parathyroid Tumor Tissue Reveals Functional Heterogeneity in Calcium Sensing 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Bone, Calciotropic Hormones & Vitamin D Thursday, March 5th 1:00:00 PM LB-OR01 6268 11:30:00 AM New Discoveries in Bone Health and Disease Oral


Dominik Pesta*1, Rachel Perry1, Dongyan Zhang1, Michael J. Jurczak2, Andreas Birkenfeld3, Sanjay Bhanot4, Varman T Samuel5 and Gerald I. Shulman2
1Yale University School of Medicine, 2Yale University, 3Technischen Universität Dresden, 4Isis Pharmaceuticals, Carlsbad, CA, 5Yale University, New Haven, CT

 

INDY as part of the SLC13 protein family is a high-affinity di- and tricarboxylate plasma membrane transporter involved in citrate import. In Drosophila, genetic deletion of INDY alters energy metabolism and extends lifespan. Mice lacking INDY are protected from both diet-induced and age-associated hepatic insulin resistance. Here, we examined the impact of selective hepatic knockdown of mammalian Indy protein (mINDY) expression using 2’-O-methoxyethyl chimeric anti-sense oligonucleotides (ASOs) on hepatic glucose metabolism in 4 week high fat fed rats (n=15 per group) assessed by hyperinsulinemic-euglycemic (HEC) clamp studies. After 4 weeks of 2’-O-methoxyethyl chimeric ASO treatment, mINDY mRNA expression was reduced by 91% (P<0.001) in the treatment group. The mINDY 2’-O-methoxyethyl chimeric ASO treated rats showed a 34% reduction in fasting plasma insulin concentrations compared to the control group (14.5 vs. 9.6 mU/ml, P<0.05) and was associated with ~30% reduction in basal rates of endogenous glucose production [5.9 ± 0.6 vs. 8.4 ± 0.8 mg/(kg-min), Furthermore hepatic insulin responsiveness was increased in the mINDY 2’-O-methoxyethyl chimeric ASO rats as reflected by increased suppression of hepatic glucose production during the HEC [19.7 vs. 61.6%, P<0.05]. Taken together these data suggest that hepatic mINDY may be a novel therapeutic target for the treatment of hepatic insulin resistance and type 2 diabetes.

 

Disclosure: SB: Clinical Researcher, Isis Pharmaceuticals. Nothing to Disclose: DP, RP, DZ, MJJ, AB, VTS, GIS

OR10-1 19199 1.0000 A Prevention of Diet-Induced Hepatic Insulin Resistance By Antisense Oligonucleotides Targeted to Mindy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR10 5926 11:45:00 AM The Intersection of Inflammation and Metabolism in Diabetes Oral


Alexsandra C Malaquias*1, Xenia C C S Couto1, Michelle B Moraes2, Mariana F A Funari3, Alexandre C Pereira4, Sandra M F Villares5, Debora R Bertola6 and Alexander A L Jorge7
1Faculdade de Ciencias Medicas da Santa Casa de Sao Paulo, Sao Paulo, Brazil, 2Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo, Brazil, 3Hospital das Clinicas da Faculdade de Medicina da Universidade de Sao Paulo, Sao Paulo - SP, Brazil, 4Instituto do Coraçao (InCor) da FMUSP, Sao Paulo, Brazil, 5Hospital das Clinicas da Faculdade de Medicina da USP, Sao Paulo, Brazil, 6Instituto da Crianca da Faculdade de Medicina da USP, Sao Paulo, Brazil, 7Faculdade de Medicina da Universidade de Sao Paulo (FMUSP), Sao Paulo

 

Introduction: Noonan syndrome (NS) and Noonan-like syndromes (NLS) are autosomal dominant disorders caused by heterozygous mutations in genes of the RAS/MAPK pathway. Insulin and leptin, two important hormones involved in satiety signals, act also through the RAS/MAPK pathway. It raises the intriguing possibility that mutations associated with NS and NLS could have an influence on metabolism and control of energy storage. Because of that, it will be of great interest to characterize the metabolism of NS and NLS.Objectives: The aim of this study was to describe metabolic profile in children with NS and NLS. Subjects and Methods: We selected 58 children (36M:22F) with previously identified pathogenic mutation in NS/NLS genes (41 PTPN11, 5 SOS1, 5 BRAF, 4 RAF1, 1 SHOC2, 1 KRAS and 1 NRAS) and 96 age-matched controls (41M:55F) to undergo anthropometric measurements, glycemia, insulin and serum lipid levels. Height and BMI were expressed as SDS for age and sex. The differences between controls and genotypes were analyzed by t-test and ANOVA. Results: Patients with NS and control ones had similar sex distribution and age (10.5±3.7 vs 10.6±1.5 yr). Patients with NS were shorter (height-SDS= -2.3 ± 1.2 vs. 0.1 ± 1.0, p<0.001) than control group, whereas BMI-SDS were similar between groups. Both NS and control patients showed normal glycemia and insulin levels. Patients with NS presented total cholesterol (142.4 ± 27.0 vs. 156.4 ± 24.7 mg/dl, p=0.001) and high-density lipoprotein cholesterol levels (HDL-C; 41.4 ± 12.6 vs. 58.0 ± 12.4 mg/dl, p<0.001) lower than controls. Low-density lipoprotein cholesterol (LDL-C) levels were similar in both groups. Triglyceride levels were higher in patients with NS than the control ones (78.3 ± 34.5 vs. 66.2 ± 23.0 mg/dl, p=0,004). Patients with PTPN11 mutation had a slight increase in HOMA-IR (1.4 ± 1.3 vs. 1.0 ± 0.6, p=0.04) than control children. Additionally, RAF1 patients showed the lowest HDL-C levels (36.0 ± 8.7 mg/dl) in comparison to PTPN11 patients (40.8 ± 12.2 mg/dl) and other genotypes (50.8 ± 14.6 mg/dl, p=0.043). Patients with NS were more likely to have a low HDL-C (odds ratio 18.6; 95%CI 7.5-46, p<0.001) and higher triglyceride levels (odds ratio 3.8, 95%CI 1.4-10.7, p=0.011) comparing with control children (corrected by sex, age, BMI). Conclusion: Despite NS and NLS BMI was within normal range, patients with NS presented a low HDL-C and higher triglyceride levels, a lipid profile that resembles features of metabolic syndrome and type 2 diabetes. Since SHP2 and SOS1 seem to have a role in insulin signaling through PI3K/AKT pathway, it is worth noting that other mutated molecules involved in NS could influence serum lipid levels suggesting a role of RAS/MAPK pathway mutations in insulin signaling.

 

Nothing to Disclose: ACM, XCCSC, MBM, MFAF, ACP, SMFV, DRB, AALJ

OR10-2 20679 2.0000 A Metabolic Profile in Patients with Noonan and Noonan-like Syndromes Suggests a Role of RAS/MAPK Pathway Mutations in Insulin Signaling 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR10 5926 11:45:00 AM The Intersection of Inflammation and Metabolism in Diabetes Oral


Philippe Klee*1, Emmanuel Somm2, Audrey Guerardel3 and Valerie M Schwitzgebel1
1University Hospitals of Geneva, Geneva, Switzerland, 2University of Geneva School of Medicine, Geneva, Switzerland, 3University Hospital of Geneva, Geneva, Switzerland

 

Background
Type 1 Diabetes (T1DM) results from the auto-immune destruction of > 90% of the pancreatic β-cells. Identifying methods to prevent this process could open interesting perspectives in the prevention of T1DM.

Hypothesis
This work focuses on strategies to increase the resistance of β-cells against auto-immune destruction. We consider the possibility that activation of nicotinic acetylcholine (nACh) receptors protects β-cells against auto-immunity. NACh receptors have been previously implicated in the control of cell death, by the activation of their α7 subunit (1) and are expressed on β-cells.

Methods
Islets isolated from wild type (WT) mice, from mice in which either the α7 or β2 subunit of nACh receptors had been disrupted, as well as Human islets were cultured in presence or absence of nicotine or choline. After 24h, a cocktail of IL-1β+TNFα+IFNg was added. Cell death was quantified by TUNEL analysis. The expression of genes implicated in the regulation of mitochondrial permeability transition was quantified by qPCR.

Results
The exposure of WT mouse islets to the above-mentioned cytokines increased β-cell death, but the pre-culture with nicotine significantly decreased the effect of cytokines (median: 0.7 vs. 1.0 arbitrary units (a.u.); p < 0.01). The experiment was repeated with islets form mice in which either the α7 or the β2 subunit of nACh receptors had been disrupted. We found that nicotine significantly decreased cell death also in the absence of the β2 subunit (median: 0.67 vs. 1.0 a.u.; p < 0.01), but not in the absence of the α7 subunit, indicating that the effect was dependent on this latter subunit. We then repeated the experiment on Human and WT murine islets, with a pre-culture with choline, a specific agonist of the α7 subunit. We found that choline also attenuated (median: 0.78 vs. 1.0 a.u.; p < 0.05) the effect of the cytokines, confirming the importance of the α7 subunit for the protective effect. We then quantified the expression of genes implicated in the regulation of mitochondrial permeability transition by qPCR and found that the cytokines induced the expression of CHOP, a marker of endoplasmic reticulum (ER) stress (2) as well as of PUMA, a protein of the Bcl-2 family (2). For both genes, the effect of the cytokines on their expression was significantly attenuated (4.6 ± 0.3 vs. 11.3 ± 1.8 a.u.; p < 0.01 for CHOP and 0.7 ± 0.07 vs. 1.5 ± 0.2 a.u.; p < 0.05 for PUMA) by pre-incubation with choline.

Conclusion
Our results show that the activation of nACh-receptors via their α7 subunit significantly reduces cytokine-induced β-cell death in mouse and Human islets. This effect was linked to attenuation of ER-stress and modulation of PUMA a pro-apoptotic protein implicated in the regulation of mitochondrial permeability transition. We believe that these results open the exciting perspective of a pharmacological modulation of beta-cell resistance against auto-immunity.

 

Nothing to Disclose: PK, ES, AG, VMS

OR10-3 21756 3.0000 A Choline Protects Human and Murine Beta-Cells in Vitro Against Cytokines By Decreasing Mitochondrial Permeability Transition 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR10 5926 11:45:00 AM The Intersection of Inflammation and Metabolism in Diabetes Oral


Anu Verma*1, Sabareesh Natarajan2, Ajay Chaudhuri3, Kenneth Snyder2, Nitesh D Kuhadiya2, Antoine Makdissi4 and Paresh Dandona5
1State University of New York -Buffalo, University at Buffalo, Williamsville, NY, 2University at Buffalo, Buffalo, NY, 3Diabetes and Endocrinology Center of Western New York, Buffalo, NY, 4State Univ of New York at Buffal, Buffalo, NY, 5State Univ of NY, Buffalo, NY

 

Background: Hyperglycemia at the time of ischemic stroke presentation has been shown to be associated with poor clinical outcomes (1); however, the  effect of hyperglycemia on cerebral blood flow patterns has not been studied.   At our stroke unit, all patients presenting with symptoms of stroke undergo CT Stroke protocol study (CSPS) that includes a 360 slice brain scan, Perfusion CT and CT angiography.  Perfusion CT  delineates  the  ischemic  tissue by  showing increased time to peak (TTP), decreased cerebral blood flow (CBF) and normal or increased cerebral blood volume (CBV), whereas infarcted tissue manifests with markedly decreased CBF and CBV.

Hypothesis: We   hypothesized   that hyperglycemia   induces abnormal   patterns of   cerebral blood flow including diminished cerebral tissue perfusion as reflected in increased TTP in patients investigated for but not found to have ischemic stroke.

Methods: We  studied  the perfusion images of 75 patients who were not found  to have stroke and correlated with their blood glucose level at admission (BGA). 25 patients with BGA <100mg/dl served as the control group whereas 25 with BGA 100-140mg/dl constituted Group A and 25 patients with BGA >140 formed Group B.  Each group was then studied in detail for the cortical and hypothalamic blood flow parameters (CBV, CBF, TTP) using the Vitrea software.

Results: We found that the cortical TTP increased by 17% in both groups A (p=0.019) and B (p=0.033) when compared with the control and  there was a statistically significant rise in TTP  in each sector of the cortex in both hemispheres. Hypothalamic TTP also increased by 12% in group A (p=0.031) and by 1.8% in group B (p=0.386), when compared with  the control group (BGA <100).  There was no significant change in cortical or hypothalamic CBF or CBV.

Conclusion: We  conclude that  in patients without stroke, a blood glucose >100mg/dl, is associated with significant increases in TTP in the cortex and the hypothalamus. Since TTP  is  inversely related  to cellular and  tissue perfusion, our study shows for the first time a reduction in cerebral tissue perfusion at glucose concentrations >100mg/dl. These observations  have  implications  for the pathogenesis of  adverse  outcomes related to hyperglycemia in patients with ischemic stroke and, therefore, similar studies need to be carried out in patients with stroke.

 

Nothing to Disclose: AV, SN, AC, KS, NDK, AM, PD

OR10-4 18771 4.0000 A Reduction in Brain Tissue Perfusion with High Blood Sugars 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR10 5926 11:45:00 AM The Intersection of Inflammation and Metabolism in Diabetes Oral


Mary A. Venneri, Elisa Giannetta, Daniela Fiore, Giuseppe Panio, Rita De Gaetano, Daniele Gianfrilli, Riccardo Pofi, Silvia Masciarelli, Francesco Fazi, Manuela Pellegrini, Andrea Lenzi, Fabio Naro and Andrea M Isidori*
Sapienza University of Rome, Rome, Italy

 

Context: Diabetes is regarded as a state of chronic inflammation. Vascular complications associated with type-2-diabetes mellitus (T2DM) are the leading cause of morbidity and mortality. Our and other groups recently showed that chronic inhibition of phosphodiesterase-5 (PDE5i) has immunomodulatory effects and offers tissue-protection, however the underlying cellular and molecular mechanisms remains to be elucidated.

Aim:To study a subset of proangiogenic monocytes/macrophages expressing TIE2 receptor (TEMs) and its natural ligand, Angiopoietin-1 (ANG1), before and after chronic PDE5i by sildenafil (SILD) in patients with T2DM and mouse models of type1- and -2 diabetes mellitus (STZ and db/db).

Results: We found specific changes in TEMs recruitment in both human and mouse models of diabetes. In this context PDE5i: 1) normalized ANG-1 levels (2.1±0.6 ng/dl; after SILD 6.8±2.8 P<0.01) and circulating TEMs in men with T2DM (TEM frequency 46.1±18.9; after SILD, 76.0±21.5, P <0.01), and restored the angiogenic function of their serum; 2) lowered circulating pro-inflammatory monocytes in patients (CD14+CD16- cells: 29.0±7.3 T2DM; after SILD 12.4±6.4, P<0.01) and in mice (CD11b+Gr-1+cells in STZ: 53.7±7.9 vs. STZ+SILD: 30.4±8.3, P<0.01; db/db 46.1±10.0 vs. db/db+SILD 26.5±7.5, P<0.01); 3) reversed renal and heart organ damage, restoring pericytes regular coverage of endothelial cells, normalizing vascular architecture and increasing proangiogenic TEMs (TEM frequency,  in STZ kidney 6.9±2.7 vs. 30.9±3.6 in STZ+SILD, P<0.01; in STZ heart 5.8± 0.9 vs. 25.1±14.0 in STZ+SILD, P<0.01; in db/db heart: 16.5±2.1 vs. 53.0±16.9 in db/db+SILD, P<0.05; in db/db isoproterenol treated heart 14.0±2.8 vs. 52.2±17.9 in isoproterenol+SILD, P<0.01) 4) antagonized isoproterenol-induced cardiac hypertrophy (p<0.05); 4) inhibited vascular inflammation modulating ANG1/TIE2 pathway.

Conclusion: In summary, we demonstrate that circulating, renal and cardiac TIE2+ proangiogenic myeloid cells are defective in diabetes, especially in injured tissues, and that sildenafil normalizes their levels, by promoting the shift from classic (M1-like) to alternative (M2-like)/TIE2+ macrophage polarization via ANG1. Restoration of TEMs via PDE5i stimulation of angiopoietin-1 improves microcirculation and could help prevent diabetic complications associated with defective tissue repair.

 

Nothing to Disclose: MAV, EG, DF, GP, RD, DG, RP, SM, FF, MP, AL, FN, AMI

OR10-5 20308 5.0000 A Chronic PDE5 Inhibition Restores Defective Angiopoietin-1 and Proangiogenic TIE2-Expressing Monocytes Recruitment in Human and Murine Models of Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 1:00:00 PM OR10 5926 11:45:00 AM The Intersection of Inflammation and Metabolism in Diabetes Oral


Levent Ozsari*1, Liem Phan1, Enrique Fuentes-Mattei2, Guermarie Velazquez-torres2, Kanishka Sircar1, Camilo Jimenez1, Gilbert J Cote1, Marie-Claude Hofmann3, Mong-Hong Lee2, Sai-Ching Jim Yeung1 and Mouhammed Amir Habra1
1The University of Texas MD Anderson Cancer Center, Houston, TX, 2The University of Texas MD Anderson Cancer Center, 3MD Anderson Cancer Center, Houston, TX

 

Background: Adrenocortical carcinoma (ACC) is a rare malignancy with limited treatment options. Hepatocyte growth factor (HGF) and its receptor, c-Met, have been reported to contribute to cancer invasiveness and chemotherapy resistance in different solid tumors, but their significance in ACC has never been studied. Experimental Design: c-Met mRNA expression data were retrieved from an external ACC databases and analyzed. We studied the expression of HGF, c-Met, and activated phosphorylated c-Met in benign and malignant human adrenocortical tissues and an ACC cell line (H295R). We measured circulating HGF levels in ACC patients. The effect of HGF on H295R cells growth was evaluated. The association between HGF/c-Met expressions and markers of cellular proliferation and apoptosis was assessed. We also conducted in-vivo experiments in mouse xenograft model to assess the effect of targeting c-Met on ACC growth by using anti c-Met shRNA or cabozantinib (a multitarget small molecule tyrosine kinase inhibitor including c-Met).Results: Transcriptomics analysis revealed c-Met overexpression in ACC. We confirmed c-Met mRNA upregulation in ACC by using quantitative real-time polymerase chain reaction. An immunohistochemical analysis revealed that HGF, c-Met, and phosphorylated c-Met proteins were more abundant in ACC than in benign adrenal adenomas. Serum HGF levels were significantly higher in ACC patients than in adrenal adenoma and healthy control groups, and HGF was detected in the culture medium of H295R cells. ACC cell growth was stimulated dose-dependently by HGF. HGF/c-Met expression positively correlated with Ki-67% and negatively with apoptosis marker (cleaved caspase 3). Knockdown of cMET mRNA using shRNA and cabozantinib significantly decreased in vitro cell proliferation, induced cell cycle arrest and reduced both mitochondrial respiration and glycolytic metabolism. In xenograft mouse model, targeting c-Met (both by shRNA or cabozantinib) significantly decreased tumor growth. Conclusions:  HGF/c-Met pathway has growth-promoting mechanism in ACC.  Considering the availability of c-Met targeting drugs, targeting this pathway alone or in combination with other therapy is a reasonable therapeutic target that warrants further evaluation.

 

Nothing to Disclose: LO, LP, EF, GV, KS, CJ, GJC, MCH, MHL, SCJY, MAH

20662 1.0000 THR-380 A Hepatocyte Growth Factor/cMET Pathway Activation Enhances Cancer Hallmarks in Adrenocortical Carcinoma: A Potential Therapeutic Target 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Naoko Tsukamoto-Yamauchi*, Tomohiro Terasaka, Kenichi Inagaki and Fumio Otsuka
Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan

 

Melatonin secreted from pineal gland contributes to the formation of circadian rhythm.  Circulating concentrations of melatonin are low in the daytime and high in the night.  However, in Cushing’s syndrome, the fluctuation of circulating melatonin was shown to be abnormal.  We earlier reported that melatonin MT1 activation augments BMPR-Smad signaling through Akt pathway in cooperation with BMP-4 in corticotrope cells.  In the present study, we investigated the role of melatonin and BMP-4 in the regulation of ACTH and PRL secretion by focusing on the clock gene expression including Bmal, Clock, Per and Cry, which are involved in controlling the circadian rhythm.  First, we examined the correlation of mRNA levels of POMC and clock genes using mouse corticotrope AtT20 cells for 3- to 48-h durations.  As a result, the significant correlations between the expression of POMC and Per mRNA levels were revealed, in particular, at 24-h culture conditions.  The knockdown of Per expression by siRNA resulted in the significant reduction of POMC mRNA in AtT20 cells regardless of the presence or absence of CRH stimulation.  Of note, treatments with BMP-4 and melatonin, both of which suppress POMC expression, reduced Per mRNA as well as protein levels in AtT20 cells.  On the other hand, in rat lactosomatotrope GH3 cells, another correlation was found between the expression of PRL and Clock or Bmal mRNA levels at 6-h culture conditions, although this correlation was attenuated in the presence of forskolin treatment.  The siRNA-mediated knockdown of Clock gene, but not that of Bmal gene, significantly reduced PRL mRNA expression by GH3 cells.  Interestingly, Bmal mRNA and protein expressions were significantly increased by forskolin treatment, whereas Clock mRNA and protein levels were not altered by any of treatment with melatonin, BMP-4 or forskolin.  Collectively, the expressional correlation between POMC and Per and that between PRL and Clock were uncovered in the corticotrope and lactosomatotrope cells, respectively.  The Per expression was inhibited by POMC regulators such as CRH, melatonin and BMP-4, whereas the Clock expression was not affected by any of PRL modulators including forskolin, melatonin and BMP-4.  Thus, the effects of melatonin and BMP-4 on the clock gene expression may imply the difference of stability of circadian fluctuations of ACTH and PRL secreted from the anterior pituitary.

 

Nothing to Disclose: NT, TT, KI, FO

20772 2.0000 THR-381 A Circadian Regulation of Pomc and Prolactin By Melatonin and BMP-4 in Anterior Pituitary Cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Karla Margarita Rodriguez*1, Erica Stevenson2 and Courtney Elizabeth Stewart1
1Kent State University, Kent, OH, 2Kent State University, Kent

 

Fibroblast growth factor (FGF) 8 is a potent morphogen during early embryonic development of neuroendocrine cells that reside in the hypothalamus. Previously, we and other researchers have shown that newborn heterozygous (+/neo) and homozygous (neo/neo) Fgf8 hypomorphic mice harbor approximately a 40% reduction in vasopressinergic cells in the paraventricular nucleus (PVN) compared to wildtype (WT) controls. Interestingly, recent studies show that adult Fgf8+/neo hypomorphic mice are more anxious than WT mice, thus suggesting that Fgf8 deficiency in the PVN may have a negative effect on the function of the hypothalamic-pituitary-adrenal (HPA) stress axis. In our first study, we used restraint stress to elicit and measure corticosteroid levels as a function of HPA-function in adult WT and Fgf8+/neo mice. Secondly, we examined brain tissue using immunocytochemistry to visualize, and analyzed corticotropin-releasing hormone (CRH) levels in the PVN of adult WT and Fgf8+/neo mice. These studies showed that Fgf8+/neo mice mounted a faster HPA response compared to their WT counterparts. Moreover, we found that the PVN in Fgf8+/neo mice had higher levels of CRH-immunostaining than WT mice. In general, our study showed significant evidence for the importance of FGF8 function in the development of neuronal circuitry, which enables an animal to cope and regulate the effects of external stressors. Future studies will investigate the molecular and cellular mechanisms that underlie FGF8-dependent embryonic and early perinatal development of CRH PVN neurons in the mouse hypothalamus.

 

Nothing to Disclose: KMR, ES, CES

21561 3.0000 THR-382 A Fibroblast Growth Factor 8 Regulates Corticotropin-Releasing Hormone Expression in the Mouse Paraventricular Nucleus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Waldemar Kanczkowski*1, Ales Neuwirth1, Sylvia Großklaus1, Kai Zacharowski2, Triantafyllos Chavakis1 and Stefan Richard Bornstein3
1Technische Universität Dresden, Dresden, Germany, 2Universitätsklinikum Frankfurt, Frankfurt am Main, Germany, 3University Hospital Carl Gustav Carus, Department of Medicine III, Technische Universität Dresden, Dresden (Germany), Dresden, Germany

 

Sepsis and septic shock in response to bacterial or viral infections remain the major health problem worldwide.  Activation of the adrenal gland stress response is crucial to cope with and to survive these adverse conditions. However, in many critically ill patients, the production of adrenal glucocorticoids, their peripheral action and metabolism are often impaired. Previously, the key role of immune-neuroendocrine interactions including expression and actions of toll-like receptors (TLR) in the regulation of hypothalamic-pituitary-adrenal (HPA) axis during endotoxemia was demonstrated.

In the present study, we have studied whether an intact action of TLRs in the adrenocortical cells or in the immune system within adrenal microenvironment is crucial for activation of adrenal glucocorticoid production, inflammation, immune cell infiltration, and adrenal cell death during endotoxemia-provoked SIRS and polymicrobial sepsis.

In particular, we engaged mice with inactivation of a key adapter molecule involved in TLR signaling (Myeloid differentiation primary response gene 88, MyD88) in adrenocortical cells (Akr1b7-Cre-MyD88 fl/fl) and in the hematopoietic cells only (Vav-Cre-MyD88 fl/fl). By subjecting these mice to LPS, we demonstrated that an intact TLR-signaling in immune but not adrenocortical cells is the major regulator of adrenal gland inflammation, including enhanced expression of pro-inflammatory mediators and immune cell infiltration. In addition, we identified that an intact TLR signaling in immune cells was necessary for a proper activation of the HPA axis.

Our data suggest that an intact immune rather than adrenocortical TLR signaling is critically involved in the regulation of the immune-adrenal crosstalk regulating the adrenal glucocorticoid production and inflammation.

 

Nothing to Disclose: WK, AN, SG, KZ, TC, SRB

20899 4.0000 THR-383 A The Role of Immune and Adrenocortical Cell Specific TLR Inactivation in Adrenal Gland Function and Dysfunction during Systemic Inflammation 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Lu Jian*
The Second Military Medical University, Shanghai, NU, China

 

The mechanism of pro-adhesion and anti-apoptotic effects of Dex in human ovarian cancer cells

Li-juan Yin, Fan Fan, Yan Wang, Xinlei Sun , and Jian Lu*

Department of Pathophysiology, the Second Military Medical University, 800 Xiangyin Road, Shanghai 200443, People’s Republic of China.

 

Abstract

Glucocorticoid (GC) can affect tumor biology not only for its immunosuppression and antiinflammatory effects, but also play a direct role in regulating proliferation, differentiation, adhesion, invasion and apoptosis of tumor cells. Our previous works demonstrated that dexamethasone (Dex), a synthetic GC significantly increased cell adhesion to extracellular matrix (ECM) by increasing levels of integrins b1, a4 and  a5, thereby enhancing cell resistance to chemotherapeutics in human ovarian cancer cells. However it is unclear whether Dex also affect the expression of components of ECM as ligands of integrin b1, and whether other membrane molecules besides integrin are involved in pro-adhesion effect of Dex in ovarian cancer cells. In order to further explore the mechanism of pro-adhesion and anti-apoptotic effects of Dex, in the present study we investigated the effect of Dex on the expression of components of ECM, cell adhesion glycoprotein CD44 and mucin antigen 1(MUC1), a large transmembrane epithelial mucin glycoprotein by QRT-PCR, Western blot and Elisa /Radioimmunoassay(RIA). The results demonstrated that 100 nM Dex treatment significantly up-regulated the expression of fibronectin and MUC1 in ovarian cancer HO-8910 and SKOV3 cells, but did not change the expression of collagen I, III and IV,and laminin, as well as CD44 and its principal ligand hyaluronan(HA). Since in addition to binding to integrin a5b1, fibronectin also binds to CD44. While we demonstrated that treatment of ovarian cancer cells with CD44 blocking antibody significantly decreased the pro-adhesion effect of Dex, suggesting that up-regulation of fibronectin may be also involved in the pro-adhesion effect of Dex by fibronectin-CD44 interaction. Moreover Dex could activate PI-3K –AKT pathway probably through enhanceing cell adhesion, which contributed to enhancing cell resistance to chemotherapy. Finally we found that  inhibiting the expression of  fibronectin and MUC1 with specific siRNAs significantly attenuated the pro-adhesion and activation of AKT induced by Dex. In conclusion, these results indicated that  up-regulation of fibronectin and MUC1 and activation of PI-3K-AKT pathway  by Dex are involved in pro-adhesion and anti-apoptotic effects of Dex in human ovarian cancer cells.  

1. Yu-Xia Chen,et al. Endocrine-Related Cancer. 2010, 17 (1): 39 -50; 2. Herr I, et al. Cancer Research. 2003, 63: 3112–3120. 3. Sui M, et al. International Journal of Cancer 2006, 119: 712–717.

Source of Research support: the National Natural Science Foundation of China (91029722 and 81472690)).

 

Nothing to Disclose: LJ

20238 8.0000 THR-387 A The Mechanism of Pro-Adhesion and Anti-Apoptotic Effects of Dex in Human Ovarian Cancer Cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Karijn J. Pijnenburg-Kleizen*1, Christiaan F Mooij2, Aliesha Griffin3, Nils P. Krone3, Paul N. Span1, Antonius E Van Herwaarden2, Fred C.G.J. Sweep1 and Hedi Claahsen-van der Grinten1
1Radboud University Medical Centre, Nijmegen, Netherlands, 2Radboud University Medical Center, Nijmegen, Netherlands, 3University of Birmingham, Birmingham, United Kingdom

 

Background Congenital adrenal hyperplasia (CAH) patients may show less clinical signs of glucocorticoid deficiency than would be expected based on their low serum cortisol levels. We hypothesize that adrenal steroid precursors and metabolites that accumulate in untreated or poorly controlled CAH patients have clinically relevant glucocorticoid activity and can partially restore cortisol deficiency. We studied the effects of 17-hydroxyprogesterone (17OHP), progesterone (P), androstenedione (Δ4) and 21-deoxycortisol (21DF) on the human glucocorticoid receptor (hGR) in comparison with the effects of cortisol.

Methods In vitro transactivation of the hGR was studied in COS-7 cells co-transfected with the hGR and the luciferase reporter vectors MMTV and pRL-TK. Cells were treated with either cortisol or one of the test compounds in increasing concentrations. Transactivation of the hGR was measured using a dual luciferase assay. Dose-response curves were calculated and the half maximal effective concentration (EC50) was determined for each steroid. Maximal transactivation by the steroids relative to the maximal transactivation by cortisol was calculated. In the second part of our study, nuclear translocation of the hGR was studied by transfection of COS-7 cells with a GFP-tagged hGR and fluorescence microscopy after treatment of the transfected cells with the steroids.

Results The EC50 of cortisol was 1.7 x 10-8 M. 17OHP, P and 21DF transactivated the hGR with EC50s of 2.2 x 10-6 M,  3.0 x 10-6 M and 1.0 x 10-7 M, respectively. For 17OHP and 21DF maximal transactivation was not significantly different from the maximal transactivation by cortisol (respectively 86%, p=0.06 and 97%, p=0.8), but for P maximal transactivation was significantly lower (73%, p=0.004). Δ4 did not transactivate the hGR. Treatment of the cells transfected with the GFP-tagged hGR with 21DF led to complete nuclear translocation of the hGR, whereas treatment with 17OHP or P resulted in partial nuclear translocation of the hGR.

Conclusions Our data suggest that 17OHP, P and 21DF are able to transactivate the hGR and thus may have clinically relevant glucocorticoid activity. For 17OHP and P, high concentrations are needed to achieve transactivation of the hGR. For 21DF however, the concentration needed to reach the EC50 is only approximately 6-fold the concentration of cortisol. Serum concentrations of 21DF reached in untreated CAH patients (up to approximately 4.5 x 10-7 M), might therefore have a clinically relevant agonistic effect on the hGR and could partially compensate the cortisol deficiency in these patients.

 

Nothing to Disclose: KJP, CFM, AG, NPK, PNS, AEV, FCGJS, HC

20848 9.0000 THR-388 A Glucocorticoid Receptor Transactivation By 21-Deoxycortisol, 17-Hydroxyprogesterone and Progesterone in Congenital Adrenal Hyperplasia 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Ana Tiganescu*, Melanie Hupe, Yoshikazu Uchida, Theodora M Mauro, Peter M Elias and Walter M Holleran
University of California San Francisco, San Francisco, CA

 

Glucocorticoids (GC) are frequently prescribed for their anti-inflammatory properties, however side effects including skin thinning and poor wound healing (WH) preclude their chronic use. We previously reported improved WH in aged C57BL/6 mice with a global deletion of the GC-activating enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) and recently demonstrated increased 11β-HSD1 expression during early WH. Here, we topically treated healthy 10 week and 18 month old C57BL6 mice with the 11β-HSD1 inhibitor carbenoxolone (CBX, 30mM) 1 week prior to wounding by double 5mm punch biopsy (n=4-7).

CBX inhibited 11β-HSD1 activity (pmol active corticosterone from inactive 11-dehydrocorticosterone/h) by 66-89% in unwounded skin and during WH (p<0.001). 11β-HSD1 activity was 50% greater in aged mice 2 days post-wounding (5.4 vs. 3.6pmol/h, p<0.01). Aged mice displayed impairments in epidermal barrier function i.e. increased baseline transepidermal water loss (TEWL, g water/h/m2) 7.3 vs. 4.5 (p<0.001) and integrity i.e. increased TEWL per tape strip 9.9 vs. 5.4 (p<0.001). CBX further impaired integrity in aged mice (12.8, p<0.05) with a similar trend for baseline TEWL (9.4, p=0.09). Wounds of aged vs. young mice were 40% smaller at day 2 (6.5 vs. 10.7mm2, p<0.001) but comparable at days 4 and 8. CBX-treated wounds in aged mice were 30% larger at day 2 (p<0.05) with trends indicating altered mRNA expression of pro-inflammatory cytokines (IL1β, 3.9-fold, p=0.12, TNFα, 5.3-fold, p=0.1), extracellular matrix remodelling (COL1, 0.15-fold, p=0.2, MMP9, 5-fold, p=0.17, TIMP1, 0.29-fold, p=0.18) and mesenchymal-epidermal signalling (FGF7, 0.26-fold, p=0.13, IGF1, 0.3-fold, p=0.09). CBX had no effect on any measured outcomes in young mice.

These findings suggest increased local GC activation modestly improve barrier function and early WH in healthy aged mice partly in response to/by suppressing increased inflammation. However, this could predispose aged individuals to the adverse effects of systemic stress e.g. delayed WH. Associations between 11β-HSD1, stress and ageing merit further exploration.

 

Nothing to Disclose: AT, MH, YU, TMM, PME, WMH

20916 10.0000 THR-389 A Increased Local Glucocorticoid Activation Improves Epidermal Function and Early Wound Healing in Healthy Aged Mice 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Takao Hashimoto*1, Li Du1 and Anthony P Heaney2
1University of California, Los Angeles, Los Angeles, CA, 2University of California (UCLA), Los Angeles, CA

 

Cushing’s disease is a life-threatening neuroendocrine disorder due to excess pituitary tumor-derived adrenocorticotrophic hormone (ACTH). No safe and effective medical therapy exists currently to target the corticotroph tumors. Glucorticoid resistance is a common yet largely unexplained phenomenon in many medical conditions and significantly limits therapeutic efficacy of administered glucocorticoids. We previously demonstrated that the orphan nuclear receptor TR4 (nuclear receptor subfamily 2, group C, member 2) binds the proopiomelanocortin (POMC) promoter to potently induce POMC expression and ACTH secretion form corticotroph tumor cells. Our further studies demonstrate that TR4 blocks both glucocorticoid and GR-mediated suppression of POMC transcription. Co-immunoprepicitation studies indicate that TR4 binds the GR in corticotroph tumor cells. In addition to abrogation of GR-directed POMC suppression, our quantitative RT-PCR studies show that TR4 modifies downstream expression of several GR target genes. Our results demonstrate that TR4 binds GR to play an important role in modulating GR actions. Further characterization of this pathway may offer important insights into glucocorticoid resistance.

 

Nothing to Disclose: TH, LD, APH

21983 11.0000 THR-390 A Testicular Receptor-4 (TR4) Modulates Proopiomelanocortin Gene Transcription By Binding the Glucocorticoid Receptor (GR) in Corticotroph Tumor Cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Liting Zhu*1, Jing Zhu2, Gen cheng Wu2 and Zhan zhuang Tian2
1Fudan University, 2Fudan University, shanghai, China

 

Background: Electroacupuncture (EA) has been shown to have anti-stress effects in breast surgery, colorectal surgery, prostatectomy and craniotomy in clinical studies. During the surgical trauma induced stress response, hypothalamic corticotropin-releasing factor (CRF) directly activates the hypothalamus-pituitary-adrenal (HPA) axis. This study investigated the hypothesis that gamma-aminobutyric acid (GABA) receptors mediate the effects of EA treatment on hypothalamic CRF in rats experiencing surgical trauma. 

Methods: The surgical trauma model was induced by partial hepatectomy. Ninety-six male rats were randomly divided into 4 groups: the intact group (INT; n=24); the model group, which received partial hepatectomies (M, n=24); the sham group, which included model animals that received acupuncture without an electric circuit (Sham, n=24); and the EA group, which comprised model animals that received EA (EA, n=24). EA treatment was given twice: 24 h before the surgery and immediately after the partial hepatectomy was completed. The expression of hypothalamic CRF and CRFR, the glucocorticoid receptor (GR), the GABA receptor, plasma adrenocorticotropic hormone (ACTH) and plasma corticosterone (Cort) were observed at 2 h, 4 h, 8 h and 24 h after surgery.

Results: Strikingly, there were higher levels of plasma ACTH in M at 4 h and 24 h after surgery, while the plasma Cort expression was higher than INTs at 2 h, 4 h, 8 h and 24 h after surgery. And the ACTH and Cort expressions in EA presented lower than those of Ms. The hypothalamic CRF and GR increased in M at 2 h and 4 h after surgery, while those in EA decreased to normal levels. The expression of the hypothalamic GABAA α2 subunit and the GABAB receptor were enhanced in M compared with INTs, while were normalized with EA treatment compared with M. 

Conclusion: Our data suggest that EA attenuated the hyperactivity of the HPA axis by the regulation of hypothalamic GABAA and GABAB receptors on hypothalamic CRF. The hypothalamic GABAA α2 subunit may play an important role in inhibitory regulation of  hypothalamic CRF expression.

Keywords: Electroacupuncture; Surgical Trauma; CRF; GABAA Receptor; GABAB Receptor

 

Nothing to Disclose: LZ, JZ, GCW, ZZT

18701 12.0000 THR-391 A Electroacupuncture Speeds up the Regulation of HPA Axis Dysfunction in Acute Surgical Trauma Rats: Mediated By Hypothalamic GABAA and Gabab Receptors 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Jing Zhu*1, Zhan zhuang Tian1 and Liting Zhu2
1Fudan University, shanghai, China, 2Fudan University

 

The hypothalamic-pituitary-adrenal (HPA) axis is a neuroendocrine system that regulated by corticotrophin-releasing hormone (CRH) and arginine vasopressin (AVP). Electroacupuncture (EA) at zusanli (ST36) and sanyinjiao (SP6) can attenuates HPA axis hyperactivity in hepatectomy rats, but the mechanisms still need to clarify. EA as a traditional Chinese treatment, can maintain homeostasis of neuroendocrine. In this study, the role of AVP in hypothalamus on EA attenuated HPA axis hyperactivity was investigated in hepatectomy rats and the effect of EA was observed in this model. ACTH and CORT level in the serum was detected via RIA at 2 hour and 4 hour after surgery. Besides, AVP and GR level in hypothalamus and hippocampus were detected by RT-PCR, Immunofluorescence and Western Blot. The results showed that a distinct increase of AVP and AVPR1a mRNA and protein level was observed between intact and hepatectomy rats, while AVP and AVPR1a mRNA and protein were significant decreased in EA group compared to hepatectomy rats. Overall, EA application at ST36 and SP6 can ameliorate the hyperactivity of HPA axis via AVP/AVPR1a.

 

Nothing to Disclose: JZ, ZZT, LZ

18896 13.0000 THR-392 A Electroacupuncture Attenuated HPA Axis Disorder By Regulating AVP and AVPR1 in Hepatectomy Rats 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Shingo Murasawa1, Kazunori Kageyama*1, Aya Sugiyama1, Yutaka Oki2 and Makoto Daimon1
1Hirosaki University Graduate School of Medicine, Hirosaki Aomori, Japan, 2Hamamatsu Univ Schl of Med, Hamamatsu, Shizuoka, Japan

 

Cushing’s disease is primarily caused by pituitary corticotroph adenomas, which autonomically secrete adrenocorticotropic hormone (ACTH). ACTH production may be associated with tumor cell proliferation; however, the effects of cell cycle progression on ACTH production are little known in corticotroph tumor cells. A DNA polymerase inhibitor, aphidicolin, arrests cells at the entrance to the S phase and blocks the cell cycle; aphidicolin also induces apoptosis in tumor cells. In the present study, we determined ACTH production and cellular proliferation of AtT-20 corticotroph tumor cells following treatment with aphidicolin. Aphidicolin decreased proopiomelanocortin mRNA levels in AtT-20 cells and the levels of ACTH in the culture medium of these cells. Aphidicolin also decreased cell proliferation and induced apoptosis in AtT-20 cells. Fluorescence-activated cell sorting analyses revealed that this agent increased the percentage of G0/G1 phase cells, and decreased S phase cells. Aphidicolin decreased the phosphorylation of cyclic adenosine monophosphate response element-binding protein and Akt. Aphidicolin increased the levels of tumor protein 27 (p27) and 53 (p53), while it decreased cyclin E levels. Aphidicolin suppresses the synthesis and secretion of ACTH in corticotroph tumor cells. Aphidicolin also increased the levels of Zac1, an upstream regulator of p53, and the mRNA levels of the stress response gene growth arrest and DNA damage-inducible 45β (GADD45β), a putative downstream target of p53. Thus, aphidicolin inhibits ACTH production and cellular proliferation in AtT-20 cells.

 

Nothing to Disclose: SM, KK, AS, YO, MD

18898 14.0000 THR-393 A Aphidicolin Inhibits Adrenocorticotropic Hormone Production and Cellular Proliferation in AtT-20 Corticotroph Tumor Cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Ruth Anna Morgan*, John A Keen, Patrick Hadoke and Brian R Walker
University of Edinburgh, Edinburgh, United Kingdom

 

Metabolic syndrome and Cushing’s disease occur in domesticated horses, but whether cortisol contributes to their pathophysiology is uncertain. Equine Cushing’s disease (Pituitary Pars Intermedia Dysfunction, PPID) is characterised by tumours of the pars intermedia, abnormal fat deposition, hirsutism, insulin resistance and elevated plasma ACTH, but paradoxically normal serum cortisol concentrations. We hypothesised that, as in human obesity, increased peripheral cortisol metabolism is a primary abnormality in equine metabolic syndrome (EMS) and in PPID, causing secondary ACTH secretion and increased cortisol and adrenal androgen production.

Body condition score (BCS, obesity) was recorded in female or castrated male healthy horses (n=7), EMS (n=5) and PPID cases (n=6) destined for euthanasia. Plasma ACTH, serum cortisol and insulin were measured by immunoassay; plasma testosterone by LC-MS/MS. Glucocorticoid and androgen metabolites in post-mortem urine samples were quantified by GC-MS/MS and corrected for creatinine. mRNA transcript levels of 11β-HSD1, 5α-reductase (type 1) and 5β-reductase in adipose (neck crest, linea alba, peri-renal) and liver were quantified by qPCR.

20β-Dihydrocortisol, a negligible metabolite in humans, was the most abundant urinary cortisol metabolite (60% of total metabolite excretion) in horses; testosterone was the major urinary androgen. Compared with healthy horses, EMS cases had higher BCS (P<0.001), hyperinsulinaemia (P=0.002) and increased total urinary androgens (1.6 ± 1.5 v 11.4 ± 7.5µg/mmol, P=0.02) and cortisol metabolites (41±3.4 v 132.4 ± 21.0µg/mmol, P=0.002), largely due to increased 20b-dihydrocortisol. Plasma ACTH, serum cortisol and androgens were normal. 11β-HSD1 transcript levels were higher in all adipose depots (P=0.006) but not in liver. Both 5α and 5β reductase were increased in liver (P=0.004). PPID cases were older (P=0.01) than healthy horses with elevated plasma ACTH (257.8 ± 217.1 v 39.6 ± 19.7pg/ml, P<0.001) but were not obese and had normal plasma cortisol (98±50 v 109±20nmol/L) and androgens; urinary cortisol and androgen metabolites were elevated, with increased testosterone (P=0.01), 20b-dihydrocortisol (P=0.02) and, disproportionately, 5b-reduced cortisol metabolites (P=0.03). Transcripts of 11β-HSD1 were increased in peri-renal adipose only (P=0.001).

In conclusion, EMS and PPID are both characterised by enhanced clearance of cortisol and adrenal androgens. This increased clearance may cause activation of the HPA axis contributing to elevated ACTH in PPID and adrenal stimulation in EMS. Given the normal plasma cortisol and androgen concentrations, the equine Cushing’s phenotype may result from altered local steroid metabolism, including by an unidentified 20β-oxoreductase. Glucocorticoid clearance may be a therapeutic target in these horses.

 

Nothing to Disclose: RAM, JAK, PH, BRW

18994 15.0000 THR-394 A Increased Cortisol Clearance As a Primary Abnormality in Equine Metabolic Syndrome and Equine Cushing's Disease 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


James Hawley*1, Laura Owen2, Annie Armston3, Phillip Monaghan4, Carrie Chadwick5 and Brian G. Keevil6
1University Hospital South Manchester, Manchester, United Kingdom, 2University Hospital of South Manchester, Manchester, United Kingdom, 3University Hospital Southampton, Southampton, United Kingdom, 4Christie Hospital NHS Foundation Trust, 5Aintree University Hospital, 6Univ Hospital of South Manchester, Manchester, United Kingdom

 

Background:The accurate measurment of serum cortisol is essential in the investigation of the HPA axis. It has been well documented that routine immunoassays are liable to both under- and over-recovery leading to inaccurate results and subsequent inappropriate investigations. This inaccuracy is proposed to be attributed to both the non-specificity of the complementary antibody and an inability to completely liberate cortisol from its binding globulin. This study seeks to provide an up-to-date assessment of the accuracy of the major immunoassay platforms and compares results to a liquid chromatography-tandem mass spectrometry (LC-MS/MS) candidate reference method.

Design: Serum remaining from routine analysis was aliquoted and distributed to four different centres for cortisol analysis by their respective immunoassay. In addition an aliquot was analysed by our routine LC-MS/MS assay and candidate LC-MS/MS reference method to provide a traceable result for comparison of assay bias. Cohort groups included: normal subjects, patients taking the oral contraceptive pill (OCP), pregnant patients, patients taking prednisolone and patients taking the the 11β-hydroxylase inhibitor metyrapone. Cortisol binding globulin (CBG) was measured in the normal, OCP and pregnant cohorts.

Results: Considerable bias was observed in the normal cohort between the immunoassays when compared to the LC-MS/MS candidate reference method whereas excellent agreement was observed between the routine LC-MS/MS assay and LC-MS/MS candidate reference method. Over recovery was greatest in patients taking prednisolone with the Roche platform particularly affected. Analysis of CBG demonstrated that elevated concentrations are observed during pregnancy and in patients taking the OCP when compared to the normal cohort. Elevated concentrations of CBG were associated with under-recovery in the Abbott Architect immunoassay. Of the patients taking metyrapone positive bias was observed across all immunoassay platforms when compared to the LC-MS/MS reference method.

Conclusions: The accurate quantification of cortisol by current immunoassays is compromised by both the non-specificity of the antibody for cortisol and matrix effects including elevated concentrations of CBG. Users should be aware of the limitations of their current assay and consider these when interpreting results, this is especially true of patients taking metyrapone where the dosage is titrated against cortisol concentration.

 

Nothing to Disclose: JH, LO, AA, PM, CC, BGK

21333 16.0000 THR-397 A Serum Cortisol: What Is Your Laboratory Measuring? 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Sisitha Udara Jayasinghe*1, Gavin W. Lambert2, Susan Torres1, Steve Fraser1 and Anne Isabella Turner1
1Deakin University, Melbourne, Australia, 2Baker IDI Heart and Diabetes Institute, Melbourne, Australia

 

The magnitude of the stress response may be important in the development of chronic disease (Chrousos 2009). It has been shown in men that increased levels of fitness can attenuate HPA axis and SAM system reactivity to stress (Rimmele, Seiler et al. 2009).  Nevertheless, it is not clear if increased physical activity is associated with lower HPA axis and SAM system reactivity to psychological stress in women.

We tested the hypothesis that women who engaged in higher levels of physical activity will have lower cortisol, adrenaline (Adr), noradrenaline (NA) and dopamine (DA) responses to a TSST compared with their relatively less active counterparts.

High and low fit women (n=22 per group; in the follicular phase of the menstrual cycle) were subjected to a TSST (Kirschbaum, Pirke et al. 1993) at 1500h. Concentrations of cortisol, Adr, NA and DA were measured in samples of blood collected every 7-15min from 1400h-1700h.  Cortisol, Adr, NA and DA were compared within and between groups using repeated measures ANOVA.

Physical activity levels (2.7±0.5 vs 7.1±1.3 hours per week; p=0.004) and VO2 max (27.4±1.0 vs 41.9±1.6 ml/kg*min; p<0.001) were significantly higher in high fit women compared with low fit women Both groups responded to the TSST with a substantial elevation of cortisol (107%; p<0.001), Adr (146%; p<0.002), NA (92%; p<0.001) and DA (44%; p<0.001) but this response did not differ significantly between high and low fit women (time * treatment for cortisol, Adr, NA and DA; p=0.987, p=0.118, p=0.169, p=0.392, respectively). Furthermore, there were no significant differences between high and low fit women in pre-treatment values, peak height, reactivity or area under the curve for cortisol, Adr and NA (p>0.05 for all).  For DA, reactivity was higher in the low fit women (p=0.009) compared with the high fit women.  DA pre-treatment values, peak height, reactivity or area under the curve were similar between the groups.

While both groups had substantial responses to psychological stress, high fit women did not have an attenuated HPA axis and SAM system responses to psychological stress compared to low fit women. These findings suggest that, for low fit women who are aged between 30-50 years, the response of plasma cortisol, Adr, NA and DA to a potent acute psychological stressor is not compromised compared to that in high fit women.  Nevertheless, it is possible that differences between the groups may have been seen with a less potent stressor.  Further research is needed to clarify the role of physical activity in stress responsiveness.

 

Nothing to Disclose: SUJ, GWL, ST, SF, AIT

21379 17.0000 THR-398 A Sympatho-Adrenal Medullary (SAM) System and Hypothalamo-Pituitary Adrenal (HPA) Axis Responses to Trier Social Stress Test (TSST) Is Independent of Physical Fitness in Women Aged 30-50 Years 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Steffy Wilhelmina Jansen*1, Ferdinand Roelfsema2, Abimbola Akintola1, Nicole Oei3, Christa M Cobbaert2, Jeroen van der Grond4, Rudi G J Westendorp1, Hanno Pijl2 and Diana van Heemst1
1Leiden University Medical Center, Netherlands, 2Leiden University Medical Center, Leiden, Netherlands, 3University of Amsterdam, 4Leiden University Medical Center

 

The hypothalamic-pituitary-adrenal (HPA)-axis is the most important allostatic system of our body and is of critical importance for survival. Disability to maintain allostasis is associated with age-related diseases. Previously, we found that subjects enriched for familial longevity, have less frequently cardiovascular disease and diabetes compared to age-matched controls (1). In addition they exhibit a lower area under the curve for morning and evening salivary cortisol levels compared to controls (2). These results indicated that HPA-axis might be a potential candidate mechanism underlying healthy familial longevity. In this study, we aimed to compare features of the HPA-axis between subjects enriched for familial longevity and controls. We included 38 subjects of the Leiden Longevity Study cohort consisting of 20 middle-aged offspring of nonagenarian siblings (offspring) together with 18 of their partners (controls). Circulating ACTH and cortisol levels were measured every 10 min for 24h. Using deconvolution analysis (3) and ACTH-cortisol-dose response curve modeling (4), specific features of HPA-axis function were assessed, including secretion rates, number of pulses, regularity of pulses, and adrenal gland responsivity to ACTH. Total geometric mean (95% confidence interval (CI)) ACTH secretion over 24-hours was comparable in offspring (1333 (1091-1629) ng/l per 24h)) and partners (1235 (1000-1525) ng/l per 24h) (P = 0.60). Also, total geometric mean (95%CI) cortisol secretion over 24-hours was comparable in offspring (5481 (4803-6248) µmol/l per 24h) and partners (5324 (4638-6118) µmol/l per 24h) (P = 0.76). Moreover, no differences in ACTH and cortisol secretory parameters were found, including pulse number, pulse mass, pulse mode and regularity. Furthermore differences were found comprising a significant (P = 0.03) lower geometric mean (95% CI) recovery potency in female offspring -36.1 (-30.6- -42.7) ng/l compared to female controls -27.4 (-22.7- -33.0) ng/l, and a tendency towards a slightly increased sensitivity of the adrenal gland in male offspring expressed by a higher geometric mean (95% CI) of the sensitivity in male offspring 0.81 (0.54-1.2) slope units compared to male controls (0.53 (0.36-0.79) slope units)(P = 0.14). These results suggest that basal HPA-axis activity is not likely playing a key role in familial longevity.

 

Nothing to Disclose: SWJ, FR, AA, NO, CMC, JV, RGJW, HP, DV

21681 18.0000 THR-399 A Familial Longevity Is Not Marked By Differences in Hypothalamic-Pituitary-Adrenal-Axis Function 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Katie Kubek1, Keith Mansfield2, Maria Magnifico2, Amy Erickson2, Salah-Dine Chibout3, Francois Pognan3, Jennifer Marlowe2 and Patrick John Devine*1
1Novartis Institutes for BioMedical Research, Inc, Cambridge, MA, 2Novartis Institutes for BioMedical Research, Inc, 3Novartis

 

The dog is an important species for safety testing, so an in vitro adrenal cell model was established. Dog primary adrenocortical cells were isolated following manual removal of the medulla, characterized and compared to similar primary rat cultures for use in investigative toxicity studies. Cell numbers and viability were similar from adrenals shipped overnight or those processed within 2 hr after resection.  The average viability of the cells was 65%, with about 13 million cells obtained per gland. Collagenase digestion provided more viable cells than when using papain. Centrifugation in a Percoll gradient removed dead cells and permitted adrenocortical cells to be concentrated, but could not separate cells from specific adrenal zones. Cells were plated in 96-well collagen-coated plates and allowed to attach overnight before medium was replaced and experiments begun. Aldosterone production over 24 hr decreased from 198 to 28 to 15 ng/million cells/day on days 1, 2 and 3 of culture, respectively.  Cortisol production also decreased over time, from 517 to 175  to 77 ng/million cells/day on days 1 through 3.  Isolation of primary rat adrenal cells were similar to dog cells.  Cell viability from rat adrenals was 70%, with about 1.5 million cells obtained per gland. Rat adrenocortical cells produced less aldosterone than dog cells, (4 ng/million cells/day on day 2) but similar amounts of corticosteroid (101 ng/million cells/day, corticosterone on day 2).  Both adrenocorticotropic hormone (ACTH) and forskolin increased and prolonged steroidogenesis and expression of certain steroidogenic enzymes. Affymatrix gene expression arrays were used to evaluate gene expression in dog adrenocortical cells over time in culture, normalizing to whole dog adrenals. The isolation protocol enriched for cortical adrenocytes, reducing gene expression of chromaffin cells 20-fold over intact dog adrenals. Culture duration was associated with an progressive loss of gene expression associated with steroidogenesis but an increasing rise in genes involved in cholesterol synthesis, and ACTH only partially abrogated these changes. Cyp11B2 gene expression suggested zona glomerulosa (ZG) enrichment with isolation and increased expression with ACTH stimulation, but loss over time in culture. Over time, a ZG-specific gene expression signature was slightly (2-fold) decreased and zona fasciculata genes remained unchanged; neither was affected by addition of ACTH. These results demonstrate functionality of primary adrenocortical cells for both species, reducing over time. Results suggest these models could be used to make interspecies comparisons of adrenal function, investigate mechanisms of adrenal-specific toxicants, drugs or inhibitors of steroidogenesis. Time-dependent changes in adrenocortical cells in culture should be considered when using these models, as optimal function was seen in the first few days.

 

Nothing to Disclose: KK, KM, MM, AE, SDC, FP, JM, PJD

21921 20.0000 THR-401 A Characterization of an in Vitro Primary Dog Adrenocortical Cell Model and Comparison with Primary Rat Adrenocortical Cells. 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Adrenal HPA Axis Thursday, March 5th 3:00:00 PM THR 380-401 5904 1:00:00 PM Adrenal tumours, glucocorticoid regulation and action Poster


Amir H Hamrahian*1, Kevin C.J. Yuen2, Murray B. Gordon3, Karen J Pulaski-libert4, James Bena1 and Beverly M.K. Biller5
1Cleveland Clinic Foundation, Cleveland, OH, 2Swedish Neuroscience Institute, Seattle, WA, 3Allegheny Neuroendocrinology Center, Allegheny General Hospital, Pittsburgh, PA, 4Masscheusetts General Hospital, Boston, MA, 5Neuroendocrine Unit, Boston, MA

 

Context:

The GST is used to evaluate GH and HPA axes in adults with pituitary disorders. Two prospective studies have reported that a GH cut-off level of 3 ng/mL differentiates those with GHD from healthy adults. The Endocrine Society Guidelines recommend a GH cut-off value of 2.5-3 ng/mL for the diagnosis of adult GHD. Recent studies suggest a lower GH cut-off especially in obese patients. Weight-based GST has been used in children, but data are limited in adults.

Objective:

To define GH cut-off values to diagnose adult GHD by GST, and to compare fixed dose (FD, 1 mg, 1.5 mg in patients >90 kg) with weight based (WB, 0.03 mg/kg, maximum 3 mg) GST.

Methods:

28 patients with adult-onset hypothalamic-pituitary disease and 1-2 (G1, n=14) or ≥3 (G2, n=14) pituitary hormone deficiencies were randomized to ITT, FD- and WB-GST (4 hrs) 5-28 days apart at 4 centers. A 3rd group of 14 control subjects (G3) was matched for age (±10 yr), sex, estrogen status and BMI (±5 kg/m2).  A GH response <3 ng/mL to ITT was used to classify patients as GH deficient.  Three subjects aged >65 years classified as GHD based on ≥3 pituitary deficiencies and low IGF-1 did not undergo the ITT.

Results:

Age and sex ratio were comparable between the 3 groups. Median (range) BMI was similar among the groups: G1 28.2 (18.6-36.3), G2 30.2 (19.8-46.2), and G3 27.3 (23.0-46.6). Mean (± SD) standardized IGF-1 levels were higher in controls compared with G1 and G2 (-0.5 ± 1.3 vs. -1.6 ± 1.1 and -1.9 ± 0.7, respectively). Glucagon dose used during the FD- and WB-GST was 1.2 ± 0.2 and 2.4 ± 0.5 mg, respectively.

The best GH cut-point for GHD diagnosis was 1.0 (92% sensitivity, 100% specificity) and 2.0 ng/mL (96% sensitivity and 100% specificity) for FD- and WB-GST, respectively. No significant change in these cut-off values were found if a GH <5 ng/mL was used to define GHD during ITT. Peak GH and BMI negatively correlated during FD- and WB-GST (r= -0.15 to -0.20, p= 0.21 to 0.33), but did not reach statistical significance. Age negatively correlated with the peak GH during FD-GST (r= -0.32, p= 0.04), but not WB-GST (r= -0.21, p= 0.21). Late peak GH (after 3 hours) was seen in 4.8% and 9.5% of the subjects during FD- and WB-GST, respectively.

During FD- and WB-GSTs, 7.1-21.4% of patients developed hypoglycemia (glucose <50 mg/dL) vs. 0% in controls. Nausea was the most common side effect (FD-GST 38% and WB-GST 50% of patients). One patient had a grand mal seizure during FD-GST without hypoglycemia. The FD-GST was preferred by patients over the other two tests, while no significant difference was seen between ITT and WB-GST.

Conclusion:

Our data do not support the use of GH cut-point of 3 ng/mL during FD- or WB-GST. Such a cut-off value may lead to misclassification of some adults as GH deficient. It is unclear whether a lower cut-point is required specifically for patients with higher BMIs. FD-GST was not inferior to WB-GST for GHD diagnosis and was preferred by most patients over the WB-GST and ITT.

 

Disclosure: AHH: Principal Investigator, Novo Nordisk, Consultant, Versartis. KCJY: Principal Investigator, OPKO, Ad Hoc Consultant, Novo Nordisk, Ad Hoc Consultant, Pfizer, Inc., Ad Hoc Consultant, Versartis. MBG: Principal Investigator, OPKO, Principal Investigator, Novo Nordisk, Principal Investigator, Pfizer, Inc.. BMKB: Principal Investigator, OPKO, Consultant, Novo Nordisk, Consultant, Pfizer, Inc., Consultant, Versartis. Nothing to Disclose: KJP, JB

PP05-1 21617 1.0000 THR-457 A Revised GH Cut-Point Needed for the Glucagon Stimulation Test (GST) in the Evaluation of Adult Growth Hormone Deficiency (GHD): Results from a Prospective Randomized Multicenter Study 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Vivien Bonert*1, John D. Carmichael2, Odelia Cooper3, James Mirocha4, Richard E Reitz5, Anne L Caston-Balderrama6, Michael J. McPhaul7, Michael Phillip Caulfield8 and Shlomo Melmed9
1Cedars Sinai Medical Center, Los Angeles, CA, 2University of Southern California, Los Angeles, CA, 3Cedars sinai Medical Center, Los Angeles, CA, 4Cedars-Sinai Medical Center, Los Angeles, CA, 5Quest Diagnostic Inc, San Juan Capistrano, CA, 6Quest Diagnostics Inc, San Juan Capistrano, CA, 7Medical Director, Endocrinology, San Juan Capistrano, CA, 8Quest Diagnostics Nichols Institute, San Juan Capistrano, CA, 9Cedars-Sinai Med Ctr, Los Angeles, CA

 

Background: Accurate measurement of insulin like growth factor 1 (IGF-1) is critical for diagnosis and management of pituitary growth hormone axis abnormalities. As anomalous results, poor reproducibility , low precision and assay bias are often reported with currently  available IGF-1 radioimmunoassays(RIA), mass spectrometry, was employed as an alternative  assay technology.

 Objective: IGF-1 was measured by both RIA and mass spectrometry (LC-MS/MS ) in serum samples from patients with pituitary disorders .

 Methods: Serum samples  prospectively obtained from 101 patients with both naïve and treated secretory and non-secretory pituitary tumors, cysts, hypophysitis,and craniopharyngioma, were evaluated by IGF-1 mass spectrometry (MS) and by Siemens Immulite 2000 IGF-1 assay (reagent lot #512; solid phase, enzyme-labeled chemiluminscent immunometric assay).  After sample accrual, samples were submitted for each assay as a single batch.  

Results: Although results of the two assays were highly correlated (R2=0.97), a significant positive bias was observed in the RIA  compared to the MS assay, reflecting a systematic proportional difference observed between  Siemens and LCMS assays, with the Siemens typically reporting higher results than the LCMS. This is reflected in the slope of the relationship observed in Deming regression y=1.57x – 54.0 and linear regression analyses y=1.55x – 48.3, . where y is the  Siemens assay  value . For classification into normal or abnormal values from each assay, results  were interpreted according to their respective reference intervals. IGF-1 MS assay results were interpreted using published age-adjusted ranges (1) and for the Siemens assay, according to age-adjusted ranges provided in the directional insert. Twenty six of 99 (27.2%) samples for which reference ranges were available for both assays, were classified differently (in range, low, or elevated) by the Siemens and the IGF-1 LCMS assay.  All 26 disagreements occurred with LCMS in the reference range: 14 were with elevated RIA  and 12 with low RIA.  LCMS was in the reference range significantly more frequently than RIA, 72.7% versus 46.5% of the cases, McNemar P < 0.0001.

 Conclusion: While results of  IGF-1 testing with the RIA  and LC-MS/MS assays  were highly correlated, a strong positive bias with higher results was reported using RIA compared to LC-MS/MS.  Twenty seven percent of IGF-1 samples measured by the two assays  were differently classified indicating a large degree of inter-assay variability. Inter-assay variability for IGF-1 results is well established and caution should be used when interpreting results of differing methodologies.The  accuracy of the IGF-1 LCMS methodology, large reference ranges, and concordance of  published LCMS reference ranges with a large published reference population (2 ) reinforces the clinical utility  of the mass spectrometry based IGF-1 assay

 

Disclosure: JDC: Ad Hoc Consultant, Genentech, Inc., Ad Hoc Consultant, Ipsen, Ad Hoc Consultant, Novartis Pharmaceuticals. RER: Coinvestigator, Quest Diagnostics. ALC: Employee, Quest Diagnostics. MJM: Principal Investigator, Quest Diagnostics. MPC: Employee, Quest Diagnostics. SM: Advisory Group Member, Chiasma, Ad Hoc Consultant, ISIS, Principal Investigator, Pfizer, Inc., Ad Hoc Consultant, Novartis Pharmaceuticals, Planning Group Member, Ipsen. Nothing to Disclose: VB, OC, JM

21909 2.0000 THR-458 A Immunometric IGF-1 Measurements Frequently Misclassify Clinical Samples Compared to a Mass Spectrometry IGF-1 Assay 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Kevin C.J. Yuen*1, Jan Frystyk2, Sharon A Rhoads3 and Martin Bidlingmaier4
1Swedish Neuroscience Institute, Seattle, WA, 2Aarhus University, Aarhus, Denmark, 3Oregon Health and Science University, Portland, OR, 4Klinikum der Universität München, Medizinische Klinik und Poliklinik IV, Munich, Germany

 

Context: Pegvisomant (PegV) is a selective GH receptor antagonist that effectively decreases IGF-I levels in healthy adults and normalizes IGF-I production in more than 90% of patients with acromegaly (1). However, although chronic GH receptor blockade with high dose PegV is known to increase endogenous GH secretion (2) without reducing systemic GH clearance (3), data are limited on its acute effects on GH and IGF kinetics in humans. 

Aims: We assessed the impact of a single bolus dose of PegV on the GH/IGF axis in adults suspected of GH deficiency with low IGF-I levels.

Study Design: In this proof-of-concept pilot study, 10 adults suspected of adult GH deficiency received a single bolus dose of SC PegV (1 mg/kg). Blood GH, GH binding protein (GHBP), total IGF-I, IGF-I bioactivity, IGFBP-2, and IGFBP-3 levels were measured at baseline and 72 hrs after SC PegV administration.

Methods: GH was measured using an automated assay that excludes PegV interference, where a monoclonal antibody that does not cross-react with PegV was combined with a monoclonal antibody specific for 22-kD GH; IGF-I was measured by commercial immunoassays using the automated iSYS platform; GHBP was measured by a modification of the ligand immunofunctional assay with an in-house monoclonal anti-GHBP antibody; in vitro IGF-I bioactivity was measured by an in-house IGF-I receptor kinase receptor activation assay, where the assay detects the ability of serum to phosphorylate the IGF-I receptor in vitro; PegV was measured by a two-site competitive immunofluorometric assay, where serum samples were diluted 100-fold to minimize potential GH interference.

Results: A total of 10 patients [5M/5F; age (mean ± SE) 40.8 ± 2.5 yrs; BMI 29.6 ± 2.5 kg/m2] with hypothalamic-pituitary disease, 1-3 pituitary hormone deficiencies and low IGF-I (IGF-I SDS -2.0 ± 0.3) were recruited. Administration of PegV increased GH (prePegV 0.13 ± 0.04 vs postPegV 0.41 ± 0.15 ng/mL; P < 0.01) and GHBP (626 ± 114 vs 1073 ± 174 pmol/L; P < 0.001), decreased total IGF-I (85.1 ± 8.3 vs 64.8 ± 10.2 ng/mL; P < 0.05) and IGF-I bioactivity (1.09 ± 0.08 vs 0.93 ± 0.08 µg/L; P < 0.05), but did not alter IGFBP-2 and IGFBP-3 levels. Serum PegV levels positively correlated with GH (r = 0.83; P < 0.01), ΔGH (r = 0.85; P < 0.01), ΔGHBP (r = 0.85; P < 0.01), and negatively correlated with IGF-I bioactivity (r = -0.61; P< 0.05). No subject reported any adverse reactions to the PegV injections.

Conclusion: A single bolus high dose PegV not only exerts acute GH receptor blockade and lowers total IGF-I levels but also decreases IGF-I bioactivity acutely, independent of altering IGFBP-2 and IGFBP-3 levels even in states of low IGF-I levels. Further studies are needed to verify if the reduction in IGF-I bioactivity with a single bolus high dose PegV is sustainable over time when high PegV doses are administered daily, as this is clinically relevant especially when treating patients with acromegaly.

 

Disclosure: KCJY: Ad Hoc Consultant, Novo Nordisk, Ad Hoc Consultant, Teva, Principal Investigator, Pfizer, Inc.. Nothing to Disclose: JF, SAR, MB

21428 4.0000 THR-460 A Impact of Acute GH Receptor Blockade with Pegvisomant on GH and IGF Kinetics in Adults Suspected of GH Deficiency with Low IGF-I Levels 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Camelia M. Saffarini*1, Cadence True2, Lacey C Plummer1, Margaret Flynn Lippincott1 and Stephanie Beth Seminara1
1Massachusetts General Hospital, Boston, MA, 2Oregon National Primate Research Center, Beaverton, OR

 

Mutations in POLR3B, encoding the second largest catalytic subunit within RNA polymerase III, have been identified in patients with leukodystrophy syndromes, some of which include pubertal delay.  However, broader populations have not yet been tested.  Therefore, probands with isolated hypogonadotropic hypogonadism (n = 166) and no evidence of ataxia, hypodontia, or chorioretinal dystrophy were screened by whole exome sequencing for mutations in POLR3B to evaluate the prevalence of mutations and define genotype/phenotype relationships.  The remaining 3 probands carried 5 rare coding sequence variants that were considered to be deleterious in 4 prediction programs (SIFT, pMUT, MutationTaster, and PANTHER).  Proband #1 carried two missense changes, one inherited from each parent: p.V523E (c.1568T>A), and p.F400S (c.1199T>C).  The p.V523E (c.1568T>A) change has been previously identified in a patient with Gordon Holmes syndrome (GHS, HH and ataxia) and is present in very low frequency (0.05%) in European controls (Exome Sequencing Project).  Proband #2 carried p. M415T (c.1244T>C) (0.1% European controls) which also been previously identified in a patient with GHS as well as a frameshift (c.2633insT; p.L880SfsX41).  Proband #3 carried the same missense variant (c.1244T>C; p.M415T) as well as a splice-site (c.2818-2 A>T) mutation.  Proband #3 is of particular interest, as he was diagnosed with the “fertile eunuch” form of IHH with initial testicular volumes of 11 cc bilaterally.  At age 27, he underwent therapy with exogenous, pulsatile GnRH (escalating doses) for 29 weeks, but was unable to raise his testosterone into the normal range.  His gonadotropins remained in the low/normal range throughout his therapy.  This data suggests that Proband #3 had resistance to exogenous GnRH, consistent with a primary pituitary abnormality.  Thus, the finding of rare, deleterious nucleotide variants in POLR3B in patients with isolated hypogonadotropism but no neurologic defects extends the phenotype spectrum of this gene.

 

Nothing to Disclose: CMS, CT, LCP, MFL, SBS

21856 5.0000 THR-461 A POLR3B Mutations in Subjects with Isolated Normosmic Hypogonadotropic Hypogonadism 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Ali Abbara*1, Channa N Jayasena2, Georgios Christopoulos3, Shakunthala Narayanaswamy4, Chioma Izzi-Engbeaya4, Gurjinder M Nijher2, Alexander N Comninos1, Deborah Peters5, Adam John Buckley5, Risheka Ratnasabapathy4, Julia K Prague4, Mohammad A Ghatei4, Stephen R Bloom2, Geoffray Howard Trew6 and Waljit S Dhillo1
1Imperial College NHS Healthcare Trust, London, United Kingdom, 2Imperial College London, United Kingdom, 3Hammersmith Hospital, London, United Kingdom, 4Imperial College London, London, United Kingdom, 5Imperial College London, 6IVF Unit Hammersmith Hospital, London, United Kingdom

 

Background:In Vitro Fertilization (IVF) treatment is an effective therapy for couples suffering from infertility, but may result in the potentially life-threatening complication termed ‘Ovarian Hyperstimulation Syndrome’ (OHSS). We have previously reported that the hormone kisspeptin-54 can be used to trigger reproductive hormonal release and induce high rates of oocyte maturation in women undergoing IVF treatment. We hypothesized that kisspeptin-54 could be used to safely and effectively trigger oocyte maturation in women at high risk of OHSS.

Methods:Sixty women with subfertility at high risk of OHSS, as identified by high serum anti-mullërian hormone >40 pmol/l and antral follicle count >23 on ultrasound, were randomized to receive a single subcutaneous dose of kisspeptin-54 to trigger oocyte maturation following a standard recombinant FSH / GnRH antagonist protocol. An adaptive dose allocation protocol was utilized, randomizing the first 15 participants to 3.2, 6.4 or 12.8 nmol/kg kisspeptin-54 (n=5 per group). Following interim analysis, remaining participants were randomized to receive either 6.4, 9.6 or 12.8 nmol/kg of kisspeptin-54 (n=15 per group). Oocytes were retrieved 36h after kisspeptin-54 administration, assessed for maturation (primary outcome), and fertilized by intracytoplasmic sperm injection with subsequent transfer of 1-2 embryos. All women were screened for the development of both early and late OHSS by assessment with symptomatology, pelvic ultrasound and blood analysis.

Results:Oocyte maturation was observed in 95% (57/60) of women following kisspeptin-54 administration. Embryo formation did not occur in 3 women, and a clinical decision was made to electively freeze embryos in a further 3 women; thus 51 of 60 women had fresh embryo transfer. Biochemical and clinical pregnancy rates per transfer at all doses of kisspeptin-54 tested were 63% (32/51) and 53% (27/51), respectively. Highest pregnancy rates were seen following 9.6 nmol/kg of kisspeptin-54, which resulted in a 77% (10/13) clinical pregnancy rate per transfer. Eight from 60 women had mild to moderate OHSS, but no woman had severe or life-threatening OHSS requiring medical admission or intervention.

Conclusion: Use of kisspeptin-54 to trigger oocyte maturation with subsequent fresh embryo transfer resulted in high clinical pregnancy rates and low rates of OHSS. Thus kisspeptin-54 shows great potential for future clinical use to safely and effectively trigger oocyte maturation in IVF therapy, even in women at high risk of OHSS.

 

Nothing to Disclose: AA, CNJ, GC, SN, CI, GMN, ANC, DP, AJB, RR, JKP, MAG, SRB, GHT, WSD

21219 6.0000 THR-462 A Kisspeptin-54 Safely and Effectively Triggers Oocyte Maturation during IVF Treatment in Women at High Risk of Developing Ovarian Hyperstimulation Syndrome (OHSS) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Martin Bidlingmaier*1, Sara Harperscheid2, Elahu S Sustarsic3, Riia Karoliina Sustarsic3, Amon Horngacher4, Herbert A Schmid5 and Maximilian Bielohuby6
1Klinikum der Universität München, Medizinische Klinik und Poliklinik IV, Munich, Germany, 2Endocrine Research Unit, Medizinische Klinik und Poliklinik IV, Klinikum der Universität, Ludwig-Maximilians University, 3Endocrine Research Unit, Medizinische Klinik und Poliklinik IV, Klinikum der Universität, Ludwig-Maximilians University, Munich, 4Klinikum der Universität München, Ludwig-Maximilians-Universität, Munich, Germany, 5Novartis Pharma AG, Basel, Switzerland, 6Medizinische Klinik und Poliklinik IV, Munich, Germany

 

The SSA Pasireotide (PAS) is used to treat acromegaly by controlling GH-excess and thereby normalizing IGF-I secretion. Recently, the long-acting release formulation (“LAR”) of PAS has become available reducing the injection frequency in patients to once a month. Because it is not clear if the degree of drug-mediated suppression of GH is directly correlated to the percentage of IGF-I suppression, we investigated the relation between GH and IGF-I secretion in rats treated with different doses of PAS-LAR.

Methods: Male Lewis rats (n=6/group) were injected with either vehicle (controls) or PAS-LAR (0.1, 0.5, 1, 4, 8, 80mg/kg). Blood samples were collected 24h (all doses) and 14 days (only 4, 8 and 80mg/kg) after a single injection of different doses of PAS-LAR. Serum IGF-I and GH were measured by immunoassay. GH concentrations were determined from serial blood samplings (10 samples during 5h) and analyzed by rank-plots. Genes of interest were quantified in liver tissue by real-time PCR (4, 8 and 80mg/kg groups).

Results: Starting with a dose of 0.5mg/kg, either dose of PAS-LAR strongly suppressed GH secretion after 24h and lowered IGF-I by about 20% (0.5mg/kg), 40% (1mg/kg) or 65% (4, 8 and 80mg/kg) vs. controls (p<0.001). Injection of 0.1mg/kg did not show an effect on GH and IGF-I secretion. 14 days after injection, GH was still suppressed in rats treated with any dose (albeit to a smaller degree compared to GH-suppression after 24h). 80mg/kg was most effective, while 4mg/kg and 8mg/kg did not differ significantly. In contrast, IGF-I was suppressed in a dose-dependent manner 14days after treatment. As expected, hepatic IGF-1 expression was significantly lower after 24h and 14days in all PAS-groups. Interestingly, after 14days, each PAS-LAR dose lowered (approx. 50%) expression of the hepatic GH-receptors (p<0.01).

Conclusion: Suppression of GH and IGF-I was significantly higher 24h after the injection of PAS-LAR compared to 14days after injection pointing towards a tachyphylaxis effect. As expected, PAS-LAR dose-dependently suppressed serum IGF-I after 24h and 14days. In contrast, suppression of GH did not show a dose-dependency. Therefore we may suggest direct effects of PAS-LAR on suppression of serum IGF-I which are independent from pituitary GH inhibition. Furthermore, the observation of significantly lower hepatic GH-receptor expression with PAS-LAR fuels the speculation that PAS may also inhibit IGF-I secretion through extra-pituitary mechanisms.

 

Disclosure: HAS: Employee, Novartis Pharmaceuticals. Nothing to Disclose: MB, SH, ESS, RKS, AH, MB

19840 7.0000 THR-463 A Effects of the Somatostatin Analog Pasireotide on GH- and IGF-I-Concentrations: Correlations and Dissociations 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Irina V Trigolosova1, Alexander V Dreval*1, Anna V. Vinogradova1, Irena A Ilovayskaya1 and Bruce H.R. Wolffenbuttel2
1Moscow Regional Research & Clinical Institute, Moscow, Russia, 2University Medical Center Groningen, Groningen, Netherlands

 

BACKGROUND: Patients with acromegaly frequently develop diabetes mellitus (DM), and treatment with long-acting somatostatin analogues (SSA) may worsen blood glucose control.

AIM: To evaluate the efficacy of metformin in newly-diagnosed acromegaly patients during treatment with SSA.

Material and methods: 19 newly-diagnosed acromegaly patients (4 men, 15 women; median  age 55 [IQR 50 -62] years) without known DM underwent an oral glucose tolerance test (OGTT) with glucose and insulin levels assessment every 30 minutes for 2 hours. OGTT performed at baseline and after 6 months of SSA therapy. Early carbohydrate metabolism disturbance  and DM  were diagnosed according to WHO recommendations. From the fasting plasma glucose (FPG) and  insulin (FPI), HbA1c,  index of insulin resistance (НОМА-R), index of insulin sensitivity (MATSUDA-index), area under insulin curves (AUCins) were calculated. Examined patients were randomized into 2 groups: without metformin treatment (group 1), n=13, and with metformin treatment (group 2), n=6.

Results: With SSA treatment, we observed a significant reduction in growth hormone (from 32.4 to 8.4 ng / ml, p <0.01) and the percentage of IGF-1 upper normal limit (from 171 to 26.9 %, p <0.01). In group 1 we observed a tendency of HOMA-R reduction from 2.4 to 2.2 (p=0.08) and increase in the level of Matsuda index from 2.7 to 4.5 (NS). FPI fell from 71 pmol / l to 57.2 pmol / l (p <0.05) and the AUCins ( in 1.3 times), especially its first phase secretion (in 3.7 times) (p <0.05). HbA1c levels and FPG increased from 5.8% to 6.0% (p <0.05), and from 5.3 mmol / l to 6.1 mmol / l (p <0.05) respectively. In group 2 (metformin group) we observed a tendency of a decrease of  HOMA-R from 7.8 to 1.5 (NS ) and the increase of  the Matsuda index from 1.7 to 8.3 (NS), reduction of FPI from 166.5 pmol / l to 38.0 pmol / l (p <0.05), AUCins ( in 2.5 times) (p <0.05), especially its first phase (in 4 times) (p <0.05). HbA1c levels and FPG decreased from 6.5% to 6.1% and  from 8.0 mmol / l to 7.0 mmol / l respectively, (NS).

Conclusions: SSA therapy reduced insulin resistance on one hand, but also reduced the first phase of insulin secretion. Metformin treatment  reduced FPG and HbA1c, but had not effect on the first phase insulin secretion.

 

Nothing to Disclose: IVT, AVD, AVV, IAI, BHRW

21661 8.0000 THR-464 A The Effectiveness of Metformin Therapy in Newly-Diagnosed Acromegaly Patients Receiving Therapy with Somatostatin Analogues 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Chrysanthi Fergani*1, Nora Newcomb2, Robert L. Goodman3, Lique M Coolen1 and Michael N Lehman1
1University of Mississippi Medical Center, Jackson, MS, 2University of Mississippi Medical Center, 3West Virginia University School of Medicine, Morgantown, WV

 

Kisspeptin neurons located in the arcuate nucleus (ARC) of the hypothalamus are primary targets of the tachykinin, neurokinin B (NKB) and this interaction is considered necessary for gonadotropin release. Specifically, kisspeptin cells co-localize NKB and its high affinity receptor neurokinin-3 (NK3R), and it has been proposed that this feedback loop stimulates kisspeptin, and by extension, GnRH/LH release. However, recent evidence suggests that other members of the tachykinin family, namely substance P (SP), may also be involved in the regulation of GnRH/LH release in rodents, via unknown mechanisms. Indeed, SP administered icv, can stimulate LH secretion in the ewe, but only at high doses (Goodman abstract Endo 2015). In the present study, we examined the co-localization of SP and/or its high affinity receptor neurokinin-1 (NK1R) in kisspeptin neurons in the sheep hypothalamus and compared this expression in adult males and females.  Brain tissue from gonadal-intact male and ovariectomized artificial follicular phase female sheep were perfused (n=7/group) and tissue sections containing ARC were processed for double-label immunofluorescent detection of kisspeptin and SP or NK1R.  Preliminary data has revealed an abundance of SP fibers and cell bodies in the ARC, with greater numbers in females compared to males (42±4.1 vs. 18±5.1, respectively, per hemisection).  No co-localization between kisspeptin and SP was observed, however, a high density of fibers was found in close apposition with kisspeptin cells. NK1R immunoreactivity (ir) was observed only in the rostral portion of the ARC, which contains few kisspeptin–ir cells. In the rostral ARC, there was no difference in the number of NK1R-ir cells between sexes (16.5±2.3 vs. 18.5±3.9, in females and males, respectively) and the % co-localization with kisspeptin was low in both sexes (10.6±1.5% vs. 7.5±2.2%). Thus, these results demonstrate that arcuate kisspeptin neurons receive SP-containing inputs, but NK1R expression is limited. Hence, we hypothesize that in the ewe, SP may stimulate LH secretion via a kisspeptin-independent manner. Alternatively, since there is some cross-reactivity among ligands for tachykinin receptors, SP may act via NK3R in kisspeptin cells to stimulate kisspeptin and hence LH secretion. In agreement, an abundance of SP fibers is found in close apposition with kisspeptin cells, and a high dose of SP (100x that of NKB) is needed to increase LH secretion (Goodman abstract Endo 2015).

 

Nothing to Disclose: CF, NN, RLG, LMC, MNL

22178 9.0000 THR-465 A Anatomical Relationships of Kisspeptin to Substance P and Neurokinin-1 Receptor in the Sheep Arcuate Nucleus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Shannon B.Z. Stephens*1, Nagambika Munaganuru2 and Alexander S Kauffman1
1University of California, San Diego, La Jolla, CA, 2University of California San Diego, La Jolla, CA

 

Kisspeptin, encoded by the Kiss1 gene, regulates reproduction by signaling directly to GnRH neurons through the kisspeptin receptor (Kiss1r). Kiss1/kisspeptin neurons are detected in several brain regions, primarily in the hypothalamic anteroventral periventricular (AVPV) and arcuate (ARC) nuclei, as well as a smaller population outside the hypothalamus in the medial amygdala (MeA). As in the AVPV, in the MeA, Kiss1 expression is increased by sex steroids acting via estrogen receptor pathways. However, the pathways and mechanisms by which estradiol signals to increase Kiss1 expression in the MeA is unknown, as is the functional significance of such up-regulation. In the hypothalamus, Kiss1 neurons are regulated primarily through estrogen estrogen receptor α (ERα), despite also coexpressing ERβ. Both ERα and ERβ are highly expressed in the MeA, suggesting that either or both ERs may be involved in Kiss1 regulation in this region. Here, we used mice lacking ERα to examine whether ERα is necessary for estradiol stimulation of Kiss1 expression in the MeA. Heterozygous ERαKO breeding pairs were used to generate male and female ERαKO and WT control littermates. To equalize estradiol levels between genotypes, all ERαKO and WT controls of both sexes were gonadectomized at 7 weeks of age and one week later, half the mice in each genotype received constant estradiol treatment via subcutaneous Silastic capsules; the remaining gonadectomized mice received no hormonal treatment.  Mice were sacrificed 5 days later and blood and brains were collected. Blood serum was analyzed for LH and estradiol levels, while brains were examined for Kiss1 expression in the MeA using in situ hybridization. Kiss1 expression in the MeA was very low in gonadectomized WT and ERαKO male mice that did not receive estradiol treatment. WT males treated with estradiol showed a large increase in MeA Kiss1 expression, both cell number and mRNA per cell. In contrast, MeA Kiss1 levels were significantly lower in estradiol-treated ERαKO males relative to WTs, despite comparable estradiol levels between the two groups.  Thus, ERα appears to be necessary for the ability of estradiol to induce maximal Kiss1 expression in the MeA of male mice, similar to estradiol regulation of AVPV Kiss1 neurons. Current experiments are determining if the same pattern holds true for MeA Kiss1 in females. Uncovering the necessity of ERα for MeA Kiss1 expression may help identify and understand possible functions of this unique extra-hypothalamic kisspeptin population. 

 

Nothing to Disclose: SBZS, NM, ASK

20656 10.0000 THR-466 A Estradiol Regulation of Kiss1 in the Medial Amygdala Involves Estrogen Receptor Alpha 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Charlotte Höybye*1, Jens Sandahl Christiansen2, Birgitte Tønnes Pedersen3, Effie Pournara4 and Beverly M.K. Biller5
1Karolinska University Hospital, Stockholm, Sweden, 2Aarhus University Hospital, Denmark, 3Novo Nordisk A/S, Søborg, Denmark, 4Novo Nordisk Health Care AG, Zurich, Switzerland, 5Massachusetts General Hospital/Harvard Medical School, Boston, MA

 

Background

It is recognised that women are less sensitive to exogenous GH than men, and therefore need higher GH doses than men to achieve the same levels of insulin-like growth factor-I (IGF-I). However, it is not clear whether this knowledge is applied in the clinical practice of GH replacement for patients with GHD.

Objective

To evaluate the changes in IGF-I, serum lipids, waist circumference, fasting blood glucose (FBG) and metabolic status relative to gender and GH dose, during 4-years of GH therapy in adults with GHD. 

Methods

Patients with adult-onset GHD receiving GH therapy (Norditropin®, somatropin, Novo Nordisk A/S) enrolled in NordiNet® IOS with 4 years of follow-up data were evaluated. Descriptive statistics were applied. A t-test was used to analyse change from baseline in GH dose, IGF-I standard deviation score (SDS), BMI, waist circumference, lipids and FBG. Stratification by baseline FBG enabled analysis of conversion of metabolic status (normal: 3.9–5.5 [70–100]; prediabetes/impaired glucose tolerance: 5.6–7.0 [101–126]; diabetes: ≥7.0 [126] mmol/L [mg/dL]) during GH therapy.

Results

Data were analyzed for 822 adult patients (44.9% female). Mean (SD) GH dose (females 0.31 [0.23]; males, 0.28 [0.22] mg) increased significantly from baseline to 4 years, but to a greater extent in females (mean difference [95% CI]: 0.07 [0.03; 0.10] mg/day; p<0.001). A significant increase (and to similar levels) in mean (SD) IGF-I SDS from baseline values (females, –0.88 [1.32]; males, –0.53 [1.38]) was observed at 4 years (females, 1.51 [1.28; 1.75]; males, 1.48 [1.21; 1.75]; p<0.001 vs. baseline for both) in both genders. During 4 years of follow-up, a significant reduction in LDL-cholesterol from baseline was observed in males (p<0.0316) but not in females (females, –0.07 [–0.32; 0.17]; males, –0.24 [–0.46; –0.02] mmol/L) and FBG significantly increased in females but not in males (females, 0.44 [0.15; 0.73]; p=0.0033; males, 0.13 [–0.16; 0.41] mmol/L; ns). Most (84.1%) patients had normal metabolic status at baseline. The proportion of patients with prediabetes at baseline who had normal metabolic status at 4 years was higher in males than in females (females 55% vs. 25%).  

Conclusions

Despite a higher baseline GH dose and dose increase in female than in male patients, 4-year GH treatment outcomes were generally better in males than in females. Although there seems to be clinical awareness of women needing higher GH doses than men, GH doses used may still be insufficient to overcome gender-related GH-resistance.

 

Disclosure: CH: Coinvestigator, Novo Nordisk. JSC: Advisory Group Member, Novo Nordisk. BTP: Employee, Novo Nordisk. EP: Employee, Novo Nordisk. BMKB: Consultant, Pfizer, Inc., Consultant, Versartis, Consultant, Novo Nordisk, Principal Investigator, Inventiv.

18432 11.0000 THR-467 A Is There Still Insufficient Growth Hormone (GH) Replacement of Women with GH Deficiency (GHD)? Analysis of Data from Nordinet International Observational Study (IOS) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Nicole H Bellefontaine*1, Margaret B Allison2, Martin Grosvenor Myers Jr.3 and Carol F Elias1
1University of Michigan, Ann Arbor, MI, 2University of Michigan, 3Univ of Michigan, Ann Arbor, MI

 

Central leptin action is an essential signal for energy-intensive homeostatic processes, including reproduction. The ventral premammilary nucleus (PMv) of the hypothalamus has been shown to be a key site for the metabolic control of reproductive physiology. The PMv abundantly expresses the leptin receptor (LepRb), contains sex steroid receptors, responds to olfactory stimuli, and is putatively involved in the sexually dimorphic circuitry.  Nevertheless, relatively little has been described about the specific PMv LepRb expressing cell populations, sex differences, and their potential role in reproductive functioning. Here we set out to define innervation sites and sexual differences of two cell populations within the PMv using transgenic mouse models. The PMv of female and male LepRb-cre mice were targeted with an adeno-associated virus (AAV) expressing synaptophysin-mCherry fusion protein, uniquely allowing for the analysis of synaptic terminals. Our findings indicate that males have a greater number of infected LepRb cells within the PMv when compared to females. As previously described, the PMv had dense and widespread projections across several nuclei of the hypothalamus, including the arcuate (ARH), ventromedial (VMH), paraventricular (PVH), and preoptic area (POA).  Further, the PMv innervates several extrahypothalamic sites such as the central amygdala, periaqueductal gray, and lateral septum (LS). Interestingly, we found that LepRb PMv neurons of males have far denser projections and that the dorsal LS contains mCherry labeled fibers only in males.  We also describe here a novel subset of LepRb cells within the PMv that express the dopamine transporter (DAT). To determine the innervation of the PMv DAT expressing neurons, we targeted the PMv of DAT-Cre mice with the viral tracer as described above. Fewer mCherry labeled cells were seen the PMv of DAT-cre mice compared to the LepRb mice. Terminals were observed in the VMH, ARH, and PVH. However, no fibers were noted rostrally in the POA or in extrahypothalamic sites, suggesting that the DAT expressing neurons of the PMv have a more restricted innervation pattern. Together, we have mapped the innervation pattern of LepRb expressing cells within the PMv and described striking sex differences, lending further evidence for the role of the PMv in sexually dimorphic circuitry. Moreover, we have described for the first time a novel cell population within the PMv, those expressing DAT. Further studies are needed to elucidate the role of the LepRb expressing DAT cells of the PMv, as well as their potential role in the control of reproductive physiology by leptin.

 

Nothing to Disclose: NHB, MBA, MGM Jr., CFE

20518 12.0000 THR-468 A Molecular Mapping of the Sexually Dimorphic Leptin Receptor Populations within the Ventral Premammilary Nucleus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Johannes D Veldhuis*1, Thomas P. Olson2, Paul Y. Takahashi2, John M. Miles2, Michael J. Joyner2, Rebecca Y. Yang3 and Jean R. Wigham3
1Mayo Clinic & Graduate School of Medicine, Rochester, MN, 2Mayo Clinic, Rochester, MN, 3Mayo Cinic, Rochester, MN

 

Context.  Exercise evokes pulsatile GH release acutely, which is followed by autonegative feedback, whereas glucose suppresses GH release acutely followed by rebound-like GH release (feedforward escape).

Objective.  To test the hypothesis that age, sex steroids, insulin, body composition and physical power determine dynamic GH responses, viz., GH autofeedback and feedforward.

Design. Prospectively randomized glucose-blinded design.

Setting.  National Center for Advancing Translational Sciences.

Participants.  Healthy men ages 19-77 yr (N = 23).

Interventions.  Fasting/rest/saline, fasting/exercise/saline and fasting/rest/iv glucose infusions were used to drive GH dynamics during 10-min blood sampling for chemiluminescence GH assay.

Outcomes.  Stepwise correlation analysis to relate peak/nadir GH dynamics to age, sex steroids, insulin, CT-estimated abdominal fat and physical power (work per unit time).

Results.  Compared with the fasting/rest/saline (control) day, fasting/exercise/saline infusion evoked peak GH within 1 h, followed by negative feedback 3-5 h later.  The dynamic GH excursion was strongly (coefficient of determination, R2 = 0.634) influenced by (i) insulin negatively (P = 0.011), (ii) power positively (P = 0.0008), and (iii) E2 positively (P = 0.001).  The dynamic range of glucose-modulated GH release was determined by insulin negatively (P = 0.0039) and power positively (P = 0.0034) [overall R2 = 0.454].  Under control conditions, power (P = 0.031) and total abdominal fat (P = 0.012) [R2 = 0.267] were the dominant correlates of GH excursions. 

Conclusion.  In healthy men, dynamic GH perturbations induced by exercise and glucose are strongly related to physical power, insulin, estradiol, and body composition, thus forging a network of regulatory pathways.

 

Nothing to Disclose: JDV, TPO, PYT, JMM, MJJ, RYY, JRW

18369 14.0000 THR-470 A Multipathway Modulation of Exercise and Glucose Stress upon GH Secretion in Healthy Men 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Ravikumar Balasubramanian*1, Sheena Chew2, Sarah E Mackinnon2, Peter B Kang2, Caroline Andrews2, Wai-Man Chan2 and Elizabeth C Engle2
1Massachusetts General Hospital, Boston, MA, 2Boston Children's Hospital, Boston, MA

 

Background:  A heterozygous de novo c.1228G>A mutation (E410K) in the TUBB3 gene encoding the neuronal-specific beta-tubulin isotype 3 (TUBB3) causes the TUBB3 E410K syndrome characterized by congenital fibrosis of the extraocular muscles (CFEOM), facial weakness, intellectual and social disabilities, and Kallmann syndrome (anosmia with hypogonadotropic hypogonadism)1. All TUBB3 E410K subjects reported to-date are sporadic cases.

Objective:To report the clinical, genetic, and molecular features of a familial presentation of the TUBB3 E410K syndrome 

Methods: Case report of a mother and three affected children with clinical features of the TUBB3 E410K syndrome. Genetic analysis of the TUBB3 gene was done by Sanger sequencing. Endocrine and non-endocrine phenotypes of the proband and family members were ascertained by clinical history and physical examination.

Results:  A de novo TUBB3 c.1228G>A mutation arose in a female proband who displayed CFEOM, facial weakness, intellectual and social disabilities, and anosmia. However, she underwent normal sexual development at puberty and had three spontaneous pregnancies with subsequent autosomal dominant inheritance of the mutation by her three boys, thus suggesting incomplete penetrance of hypogonadotropic hypogonadism. All sons displayed non-endocrine features of the TUBB3 E410K syndrome similar to their mother, but in contrast to their mother,  had variable features suggestive of additional neuroendocrine abnormalities including hypogonadotropic hypogonadism, hypoadrenalism, growth hormone deficiency and central hypothyroidism.

Conclusions:  This first report of a familial autosomal dominant inheritance of the TUBB3 c.1228G>A mutation  provides new insights into the spectrum of endocrine phenotypes associated with the TUBB3 E410K syndrome. The presence of anosmia without hypogonadotropic hypogonadism in the proband also demonstrates that TUBB3 mutations may have dissociated effects on the ontogeny of olfactory axons and GnRH neurons. These observations emphasize the need for appropriate clinical evaluation and complicate genetic counseling of patients and families with this syndrome.

 

Nothing to Disclose: RB, SC, SEM, PBK, CA, WMC, ECE

21344 16.0000 THR-472 A Expanding the Phenotypic Spectrum and Variability of Endocrine Abnormalities Associated with TUBB3 E410K Syndrome 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


IIonka Kreitschmann-Andermahr*1, Sonja Siegel2, Bernadette Kleist1, Johannes Kohlmann3, Christa Menzel3, Sven-Martin Schlaffer3, Rolf Buslei3 and Michael Buchfelder3
1University of Duisburg-Essen, Essen, Germany, 2University Hospital Essen, Essen, Germany, 3University Hospital Erlangen, Erlangen, Germany

 

Introduction: We recently conducted a large postal survey on the diagnostic process of acromegaly from the patients’ perspective1. An analysis of the data yielded a mean self-reported time from symptom onset to diagnosis of 4.3 ± 6.1 years. In an earlier study of our group with only 40 participants, the time delay of the diagnostic process was significantly associated with psychosocial impairment, which we hypothesized to be caused by embitterment disorder2. This is defined as a prolonged, adverse psychological reaction on a stressful life event, which is perceived as unjust and humiliating. Against this background, it was the aim of the present study to investigate the possible impact of belated diagnosis of acromegaly on quality of life (QoL), depression and, specifically, embitterment in a large patient group.

Methods:  Patients who had been operated between 2000 and 2012 in a large tertiary neurosurgical referral center and had completed the survey on the diagnostic process also filled in self-rating questionnaires on depression (Beck Depression Inventory II; BDI-II), QoL (Short Form 36; SF-36) and embitterment (Bern Embitterment Inventory; BEI). The data of 165 valid answer sets were analyzed using SPSS.

Results: Patients in the study did not suffer, on average, from increased embitterment or an overall reduced QoL. Male patients had significantly higher performance related embitterment scores than women, who in turn, had a significantly worse psychological Qol than men and were more depressed. Number of doctors consulted in the diagnostic process was significantly associated with emotional embitterment, pessimism and hopelessness, (BEI), physical and mental quality of life (SF-36) and depression (BDI-II). Consistently, time delay from first seeking medical advice to diagnosis was also significantly related to the same parameters of the BEI, SF-36 and BDI-II (p ≤ 0.05 – 0.001).

Conclusion: Delayed diagnosis of acromegaly has severe consequences in terms of psychosocial impairment and is associated with higher scores on many subscales of the embitterment questionnaire in patients with acromegaly. The results underscore the need to maximize efforts to establish better diagnostic algorithms for speedier diagnosis. Furthermore, the results should be taken into account for developing psychological supportive treatments for the affected patients.

 

Disclosure: IK: Clinical Researcher, Novartis Pharmaceuticals, Clinical Researcher, Pfizer, Inc., Clinical Researcher, Ipsen. MB: Investigator, Pfizer, Inc., Speaker, Novartis Pharmaceuticals, Speaker, Ipsen, Speaker, Novo Nordisk. Nothing to Disclose: SS, BK, JK, CM, SMS, RB

19565 17.0000 THR-473 A Depression, Embitterment and Impaired Quality of Life Are Sequelae of Belated Diagnosis of Acromegaly 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Flavia Lucia Conceicao*1, Amanda Araujo Laudier2 and Monique Morgado Loureiro3
1Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil, 2Universidade Federal do Rio de Janeiro, Rio de Janeiro, 3Universidade Federal do Rio de Janeiro

 

Growth and development abnormalities have long been studied in children with sickle cell anemia. Almost all have detectable growth failure that affects weight more than height. Normal height is often achieved by adulthood but weight remains lower than that of controls. The etiology for the growth disturbances in this population seems to be multifactorial. Primary hypopituitarism, hypogonadism, and hypothalamic insufficiency, besides chronic anemia have been described. Growth hormone and insulin-like growth factor I axis abnormalities were found in children with sickle cell disease and growth failure, however there aren’t studies in adult populations. Our objective was to study the somatotropic axis in adolescent and adult patients with sickle cell disease. Forty-eight patients with sickle cell disease were submitted to a clinical interview and examination. Hematological parameters and IGF-1 were assessed and all patients were submitted to the insulin tolerance test (ITT). Growth hormone deficiency (GHD) was defined as a GH peak < 3ng/mL. Patients with fetal hemoglobin > 8.6g/dL were classified as “good prognosis”, and those with fetal hemoglobin < or = 8.6g/dL were classified as “poor prognosis”. The median age was 25.5 years (range 15-50) and 58% were female. Four patients had GHD (8.3%), 16 had low IGF-1 (33.33%). Six patients were below the genetic target for stature. Patients in the  “poor prognosis” group (fHb < = 8,6) had lower values of IGF-1 (215.72 X 136.07 ng/dL; p=0.003), suggesting that the reduced levels of IGF-1 are correlated with the severity of the sickle cell disease. In conclusion,  it was the first time that the prevalence of somatotropic axis abnormalities in adult patients with sickle cell disease was evaluated. Patients with sickle cell disease in the “poor prognosis” group had lower IGF level, suggesting a chronic state disease, and not GHD.

 

Nothing to Disclose: FLC, AAL, MML

21357 18.0000 THR-474 A Somatotropic Axis Abnormalities in Patients with Sickle Cell Disease 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Megan Doczi*1, Alexandra Palmer1 and Carl Vitzthum2
1Norwich University, Northfield, VT, 2Colby College, Waterville, ME

 

Shaker family voltage-gated potassium channels (Kv1) contribute to the maintenance of resting membrane potential, action potential characteristics, and neurotransmitter release in several populations of neurons. Evidence has shown that a specific Shaker channel, Kv1.3, can be modulated by circulating hormonal factors such as insulin, suggesting that this channel may be a key determinant of neuronal pathways that regulate metabolic function. Since hypothalamic neurons mediate the homeostatic balance between food intake and energy expenditure, the expression and function of insulin-sensitive Kv1.3 channels during early development may directly influence hypothalamic patterning. To determine whether Kv1.3 channels are expressed in the hypothalamus during early development, we performed quantitative RT-PCR analysis on intact microdissections of embryonic avian hypothalamic tissue. Kv1.3 channels were detected at various developmental time points including embryonic day 8 (E8) and embryonic day 12 (E12). Although no significant difference was detected in Kv1.3 mRNA expression between E8 and E12 (p = 0.0661; n = 3), these data indicate the presence of Kv1.3 mRNA in the developing hypothalamus. Previous research in olfactory bulb neurons has demonstrated that tyrosine phosphorylation-dependent current suppression of Kv1.3 can occur via insulin receptor kinase activity. Therefore, we also tested the hypothesis that insulin receptors (IR) are expressed in the embryonic avian hypothalamus at the same developmental time points as Kv1.3. RT-PCR analysis indicated the presence of IR mRNA in both E8 and E12 intact microdissections of embryonic hypothalamic tissue (n = 3), suggesting that IR signaling may play a role in the regulation of hypothalamic Kv1.3 current during development. To determine whether primary neuronal cultures could be used to study Kv1.3 function, we performed RT-PCR analysis on dissociated hypothalamic neurons from E12 embryos. Consistent with the data obtained from intact hypothalamic tissue, dissociated E12 hypothalamic neurons also revealed the expression of both Kv1.3 and IR in vitro (n = 3). These data indicate that gene expression in the dissociated hypothalamic culture model parallels the results obtained from intact tissue and suggests that future studies implementing this model can be used to examine the electrophysiological function of Kv1.3 in response to various metabolic factors. This study is the first to demonstrate hypothalamic mRNA expression of both Kv1.3 and IR in developing avian embryos and suggests a role for voltage-gated ion channel regulation in the physiological patterning of embryonic hypothalamic circuits.

 

Nothing to Disclose: MD, AP, CV

22005 19.0000 THR-475 A Shaker Family Voltage-Gated Potassium Channel (Kv1) and Insulin Receptor (IR) Expression in the Embryonic Avian Hypothalamus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Sean Stackhouse*1 and Siddharth Ramakrishnan2
1University of Puget OSund, Tacoma, WA, 2University of Puget Sound, Tacoma, WA

 

While the role of Gonadotropin releasing hormone (GnRH) neurons in sexual behavior is well studied, their regulation of social behavior is poorly understood. In this research, we examined the effect of early embryonic exposure of R-F-amide Related Protein3 (RFRP-3), a GnRH antagonist, on social behavior in the teleost medaka (Oryzias latipes). We used the mirror approaching behavior, previously demonstrated as shoaling, a basic social behavior found in many teleosts. Using a transgenic line of medaka with GnRH3-neurons tagged with the green fluorescent protein (GFP), we examined the effects of embryonic exposure to RFRP-3 on both GnRH3 neuron GFP intensity and effects on larval social behavior. Unlike the preoptic area GnRH1 neurons (associated with reproduction) or the midbrain GnRH2 neurons (involved in metabolism), the role of the GnRH3 system associated with the terminal nerve (TN) is poorly understood. This work aims to further investigate the role of the GnRH3 neuronal system and its putative role in modulating social behavior.

Transgenic medaka embryos were treated with 1µM of RFRP-3 from day 0 until hatch. GnRH3-GFP intensity in the TN and trigeminal (TG) areas were recorded using an epifluorescent microscope on 2-4 days post fertilization. Social behavior of larvae was tested at 3 weeks post hatch in a 10cm x 8cm enclosure with 2cm of normal fish water. A fish larva was placed in the chamber under the following conditions with a/an – i) opaque wall; ii) mirror; iii) mirror on one side and a live fish separated by a clear barrier on the other; iv) opaque wall on one side and a live fish separated by a clear barrier on the other. Both RFRP-3 exposed and vehicle treated control fish were exposed to each condition. For tracking, the tank was divided into three equal segments, the mirror/wall region, center region, and a non-mirror/live-fish region. Fish behavior was recorded for 3 minutes after starting the trial and analyzed using the Noldus EthoVision software.

RFRP-3 treated fish showed no significant difference in GnRH3-GFP fluorescence intensity at 3dpf but showed 56% increased intensity at 4dpf (n=12, p<0.01) in the TN and 36% increase the TG (n=12, p<0.05). While there was no overall difference between motion between control and RFRP-3 treated medaka, treated fish were found to spend 70% more time near the mirror under condition (ii) as compared to control fish (p<0.05). We suggest that RFRP-3 exposure may be affecting the GnRH3 neuron development, which could then lead to altered social behavior.

 

Nothing to Disclose: SS, SR

21667 20.0000 THR-476 A Rfrp-3 Affects Gnrh3 Neuronal Development and Larval Social Behavior in the Teleost Oryzias Latipes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Fabiana Lucio-Oliveira*1, Juliana Lima2, Fernanda Vechiato2, Ricardo Coletti2, Tatiane Franco2, Lucila Elias3 and Jose Antunes-Rodrigues3
1School of Medicine of Ribeirao Preto, University of Sao Paulo, Ribeirao Preto, Brazil, 2Universidade de São Paulo, Ribeirão Preto, Brazil, 3Ribeirao Preto Medical School - University of Sao Paulo, Ribeirao Preto, Brazil

 

The hydroelectrolytic homeostasis is constantly regulated by information concerning the body fluids volume and the concentration of ions. Conformational changes in several regions of CNS cells activate neurons located in the PVN and SON nuclei of the hypothalamus, whose secretion includes oxytocin (OT) and vasopressin (AVP). Several neurotransmitters are involved in the control of AVP and OT release, such as nitric oxide (NO). NO is an important signaling molecule which seems to modulate several physiological processes as neurotransmission, vasodilation and behavioral responses, and has been suggested to exert a prominent regulatory role on hypothalamic-pituitary system. However, there are conflicting data on the role of NO in the release of AVP and OT and studies are scarce and controversial. Thus, the aim of this study was to evaluate the effect of N-Propyl-L- Arginine (NPLA), a selective inhibitor of neuronal NOS, on plasma OT and AVP levels in rats submitted to salt loading. Also, NOS activity was evaluated in medial basal hypothalamus (MBH). Seven days before experiments, every adult male Wistar rat weighing 250 g was unilaterally implanted with a cannula into the right lateral cerebral ventricle (i.c.v). Four days before experiment, the animals were maintained with free access to food and tap water (controls) or 0.3M NaCl solution during 96 hours (salt loading). One hour and thirty minutes before decapitation, the rats were treated with an icv injection of NPLA (250 µg/5 µl) or vehicle (saline NaCl 0,9%). One hour after icv injection, tap water and 0.3M NaCl were reoffered. Trunk blood was collected for determination of osmolality and hormones concentration. The brain was removed and MBH was collected for NOS activity measurement. Our data showed, as expected, an increase in plasma osmolality (p<0.0001; F3,49=20.71) of rats that were submitted to salt loading when compared with control animals. The augment in osmolality was followed by an increase in NOS activity in MBH of salt load rats treat with  icv injection vehicle when compared with control animals under the same treatment (p<0.0001; F2,44=13.10). However, animals submitted to salt loading treated with NPLA had a decrease in NOS activity around 44% when compared with animals treated with vehicle (p<0.0001; F1,44=282.9). In addition, there was a significant increase in plasma AVP (p<0.0001; F3,57=38.30 ) and OT (p<0.0001; F3,51=18.53) concentrations in animals submitted to salt loading when compared with control animals in both treatments, but no difference was observed in treatment with NPLA. These results indicate either that modulation of NO on the secretion of AVP and OT observed in several studies is not likely performed through the neuronal isoform of NOS or that the level of inhibition of NOS (about 44%) was not effective in modulating the response of these hormones during osmotic stimuli such as salt loading.

 

Nothing to Disclose: FL, JL, FV, RC, TF, LE, JA

19838 21.0000 THR-477 A Inhibition of NOS Neuronal Isoform Does Not Alter the Levels of Neurohypophyseal Hormones in Salt Loaded Rats 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Noriyuki Koibuchi*1, Syutaro Toya2, Izuki Amano2, Noriaki Shimokawa3 and Yusuke Takatsuru2
1Gunma Univ Grad Sch of Med, Gunma, Japan, 2Gunma University Graduate School of Medicine, Maebashi, Japan, 3Gunma Univ Grad Sch of Med, Maebashi, Japan

 

Exposure to various stresses during neonatal period may induce several neuropsychological disorders in adulthood.  Such disorders may be induced by functional alteration of glutamatergic system.  However, underling mechanisms have not yet been fully clarified.  Furthermore, the involvement of glucocorticoids that are representative stress hormone has not yet been fully studied.  In the present study, we used maternal deprived (MD) male mice as an early life stress model, and studied the change in glutamatergic system and glucocorticoid homeostasis in adulthood.  Neuronal activity of the somatosensory cortex (SSC) was measured by electrophysiology for cortical field potential, and mechanical nociceptive threshold level by von Frey hair somatosensory test.  Glutamate and corticosterone concentrations were measured by in vivo microdialysis.  The neuronal activity, and glutamate and corticosterone concentration in the SSC increased in MD mice.  Nociceptive threshold level decreased in MD mice.  On the other hand an increase in coticosterone concentration by stressful physical stimulation (SPS) was not observed in MD mice, whereas the concentration of glutamate increased extremely during SPS.  Expression levels of AMPA/NMDA receptor were also changed from those of control mice after stimulation.  These findings indicate that early life stress induces disruption of homeostasis of glutametergic synapses in the SSC.  Such alteration may alter the somatosensory response.  Decreased sensitivity of glucocorticoid to acute stress by maternal deprivation may be involved in this disruption.

 

Nothing to Disclose: NK, ST, IA, NS, YT

18635 22.0000 THR-478 A Disruption of Glucocorticoid Homeostasis and Glutamatergic Neuronal Circuit in Adulthood of Male Mice By Maternal Deprivation 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Sarah Orton*1, Marisa Censani2, Alexis Jamie Feuer3 and Maria George Vogiatzi1
1Weill Cornell Medicine New York Presbyterian, New York, NY, 2Weill Cornell New York Presbyterian, New York, NY, 3New York Presbyterian Hospital Weill Cornell Medical Center, New York, NY

 

The use of gonadotropin releasing hormone analogs (GnRHa) is increasing in pediatrics. Previous studies have shown that pubertal suppression with leuprolide acetate may result in an increase in weight and body mass index (BMI). However, the data is limited, conflicting and focused primarily on female subjects with a diagnosis of central precocious puberty (CPP). Sparse data is available on the effect of histrelin on BMI. We hypothesized there would be an increase in BMI during GnRHa treatment. This study aims to establish effects of GnRHa therapy on BMI in both genders and examines patients treated with both leuprolide and histrelin for multiple indications. 

This is a retrospective study of pediatric patients who received GnRHa therapy for 12 months. Age, gender, diagnosis and anthropometric data, including BMI standard deviation score (SDS) were obtained at baseline and at three month intervals.

29 patients (23 females, 6 males; age at start of therapy 10.89 years + 2.3, BMI SDS 0.61 + 0.87 kg/m2) received leuprolide acetate monthly (leuprolide group). 9 patients (31%) had CPP, 9 (31%) had early puberty (puberty between the ages of 8-9 for females and 9-10 for males) and 11 (38%) had short stature treated with leuprolide and growth hormone (GH) (one patient had CPP and GHD). 9 subjects were treated with histrelin (7 females, 2 males; age at start of therapy 11.8 years + 2.3, BMI 0.7 + 0.9 SDS kg/m2, all with the diagnosis of CPP) (histrelin group).

No significant change in BMI SDS was detected in the children with CPP treated with either leuprolide or histrelin. Mean BMI SDS increased significantly from baseline after 12 months of leuprolide therapy in subjects without CPP by 0.165 + 0.073 (p=0.035). When stratified for gender, males had a mean BMI SDS increase of 0.22 + 0.099 (p=0.018). A transient increase in BMI SDS was observed at 6 months only in females treated with both leuprolide and growth hormone (BMI SDS increase of 0.191 + 0.692, p=0.022), whereas no change in BMI SDS was noted after 12 months of therapy.

Our data support that GnRHa therapy does not adversely affect BMI in children with CPP. An increase in BMI SDS, which involved primarily males, was detected in subjects without a diagnosis of CPP treated with leuprolide acetate. These findings need to be confirmed with larger studies and can have significant clinical implications for children who receive such therapy.

 

Nothing to Disclose: SO, MC, AJF, MGV

21867 23.0000 THR-479 A GnRH Agonists May Transiently Decrease BMI in Male and Female Pediatric Patients 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Guangfu Hu, Mu-Lan He, Wendy K.W. Ko and Anderson On-Lam Wong*
University of Hong Kong, Hong Kong, China

 

Neurokinin B (NKB) acting via Type III neurokinin receptor (NK3R) is a key component regulating pulsatile release of luteinizing hormone in mammals.  Recently, the carp TAC3 gene products, NKB and NKB-related peptide (NKBRP), were shown to up-regulate somatolactin α(SLα) expression via NK3R activation in carp pituitary cells.  Although NK3R plays a crucial role in mediating the effect of TAC3 gene products, whether NK3R expression can serve as a regulatory target at the pituitary level to modulate the pituitary actions of NKB/NKBRP is still unclear.  In this study, carp NK3R was cloned and found to be expressed at high level in the pituitary.  Functional expression of carp NK3R also revealed that the receptor exhibited ligand binding selectivity and coupling with post-receptor signaling pathways similar to that of mammalian NK3R.  In carp pituitary cells, NK3R mRNA level could be elevated by IGF-I/-II induction via IGF-I receptor (IGF-IR) coupled to MAPK and PI3K/Akt pathways.  Co-treatment with IGF-I/-II, interestingly, was found to markedly enhance NKB/NKBRP-induced SLα mRNA expression at the pituitary level.  This potentiating effect was dependent on NK3R expression and activation of cAMP/PKA, PLC/PKC and Ca2+/CaMK-II cascades. Co-treatment with IGF, however, did not trigger noticeable enhancement in post-receptor signaling via cAMP/PKA, Ca2+/PKC, MAPK or PI3K/Akt pathways.  These results, as a whole, suggested that the synergistic effect on SLα gene expression induced by IGF and TAC3 gene products is mediated by NK3R up-regulation at the pituitary level but not through modulations in functional crosstalk at the level of post-receptor signaling.

 

Nothing to Disclose: GH, MLH, WKWK, AOLW

19743 24.0000 THR-480 A Novel Synergistic Action of Insulin-like Growth Factor and TAC3 Gene Products on Somatolactin á Gene Expression Via up-Regulation of Type III Neurokinin Receptor at the Pituitary Level 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Thiti Snabboon*1, Natnicha Houngngam2, Paisith Piriyawat3, Sarat Sunthornyothin4 and Sompongse Suwanwalaikorn5
1Faculty of Medicine, Chulalongkorn University, Bangkok, Thailand, 2Excellence Center for Diabetes, Hormone, and Metabolism, King Chulalongkorn Memorial Hospital, Bangkok, Thailand, Bangkok, Thailand, 3Texas Tech University Health Sciences Center El Paso, El Paso, TX, 4Hormonal and Metabolic Disorders Research Unit, Excellence Center for Diabetes, Hormone, and Metabolism, and Division of Endocrinology and Metabolism, Chulalongkorn University, and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, Thailand, 5Chulalongkorn University, Bangkok, Thailand

 

Introduction: Isolated hypogonadotropic hypogonadism (IHH) is a clinically and genetically heterogeneous disorder. The underlying cause may be due to developmental defects or impaired functional activities of GnRH neurons, disturbed interaction between the GnRH ligand and its receptor, or the release of intact gonadotropins. Advances in genetic study indicate nearly 40% of patients carrying a mutation in one of the susceptible genes.

Materials and Methods: We examined the clinical and genetic features of 13 IHH patients: 3 cases with anosmia (Kallmann syndrome, KS) and 10 cases normosmic IHH (nIHH). Additional clinical parameters were recorded: testicular volume, cryptorchidism, cleft lip and/or palate, neurological, dental, renal and craniofacial anomalies. All patients were screened for mutations by using direct sequencing of the entire coding region of the KAL1, FGFR1, FGF8, PROK2, PROKR2, NELF, GnRH1, GnRHR, TAC3, TACR3, KISS1, KISS1R, and CHD7 genes.

Results: Of all 13 patients, 11 were sporadic cases and 2 were from one consanguineous family. All patients, aged 20-45 year, were men and were found to have micropenis and delayed puberty. Five patients had cryptorchidism. Among patients with KS, 2 had renal agenesis and 1 had septo-optic dysplasia (SOD) feature. Mutations were identified in all KS patients and the one family of nIHH. The KS group harbored mutations in KAL1 (c.653_654delTG; p.Val218fsX11) and FGFR1 (c.303C>G; p.Cys101Trp). Digenic mutations, KAL1 (c.1540G>A; p.Glu514Lys) and FGFR1 (c.203G>A; p.Arg68Gln), were identified in the patient with SOD feature. One of the nIHH patients was found to carry a novel homozygous mutation in GnRHR (c.721_742+1delGTCCTTCATCAGGACCCCCCACG).

Conclusions: Our study demonstrated the clinical and genetic features of the Thai patients with IHH. Furthermore, we reiterated the expanding phenotypic spectrum of IHH, in particular its impact on the development of the anterior midline forebrain

 

Nothing to Disclose: TS, NH, PP, SS, SS

19534 25.0000 THR-481 A Clinical and Genetic Characteristics of Thai Patients with Isolated Hypogonadotropic Hypogonadism 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Louise Cheryl Gregory*1, Mark J McCabe1, Rodrigo Bancalari2, Vaitsa Tziaferi1, Helen Spoudeas3 and Mehul Tulsidas Dattani4
1UCL Institute of Child Health, London, United Kingdom, 2Pontificia Universidad Catolica de Chile, Santiago, Chile, 3Great Ormond Street Children's Hospital, London, United Kingdom, 4UCL GOS Institute of Child Health, London, United Kingdom

 

Background Hypogonadotrophic hypogonadism (HH) is a complex developmental disorder characterized by a reduction in gonadotrophins (LH, FSH) that are released from the anterior pituitary. The phenotype is characterized by a delay in onset or complete absence of puberty, often accompanied by genital abnormalities. Kallmann syndrome (KS) is characterized by the association of HH with anosmia/hyposmia, and variably cleft palate, sensorineural hearing loss, and renal abnormalities.

Methods/Cohort We screened 63 HH/KS (M:F 50:13) patients in our cohort, 41 with isolated HH and 22 KS, with 40% of the total having associated abnormalities, for variants in the five most commonly mutated genes, KAL1, PROKR2, FGFR1, FGF8 and GnRHR, using PCR and direct sequencing analysis.

Results We identified 4 variants, two of which were novel. A novel hemizygous missense substitution, KAL1 c.257G>A, p.C86Y, was identified in a male KS patient. A novel FGFR1 heterozygous deletion (c.915delG, p.[Leu305LeufsX6]), causing a frameshift, was identified in two siblings who presented with anosmia: a male with cleft lip/palate and short stature, and a female with a microform cleft lip. The mutation was inherited from the unaffected father. Two previously described variants, that have not been functionally characterized, were also identified in our cohort: a hemizygous early stop codon, KAL1 c.1267C>T, pR423X, in a KS patient and a heterozygous missense substitution, FGFR1 c.2059 G>A, p.G687R, in a male patient with isolated HH who is currently showing signs of reversal after stopping treatment. His mother had primary amenorrhoea and also carries the variant, his father was unaffected and his DNA was unavailable. All four variants are located at highly conserved residues across multiple species and were not present on any control databases.

Conclusion We report two novel and two previously described variants in two different candidate genes in four unrelated HH/KS patients. The potential genetic cause remains unknown in 94% of our HH cohort, signifying the need for further investigation into genes involved in GnRH neuronal migration and secretion, which might be potentially responsible for the pathogenicity.

 

Nothing to Disclose: LCG, MJM, RB, VT, HS, MTD

19198 26.0000 THR-482 A Genetic Screening of Children and Adolescents with Kallmann Syndrome/Hypogonadotrophic Hypogonadism 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Neuroendocrinology and Pituitary Thursday, March 5th 3:00:00 PM THR 457-482 5910 1:00:00 PM Neuroendocrinology Poster


Delanie B. Macedo*1, Danielle de Souza Bessa2, Ana Paula Abreu3, Vinicius N. Brito4, Ursula B. Kaiser5 and Ana Claudia Latronico6
1Unidade de Endocrinologia do Desenvolvimento e Laboratório de Hormônios e Genética Molecular/LIM42, Disciplina de Endocrinologia, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Sao Paulo, Brazil, 2Hospital das Clínicas, Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil, 3Brigham and Women's Hospital and Harvard Medical School, Boston, MA, 4Laboratório de Hormônios e Genética Molecular/LIM42, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, Sao Paulo SP, Brazil, 5Brigham and Women's Hospital/Harvard Medical School, Boston, MA, 6Unidade de Endocrinologia do Desenvolvimento, Laboratório de Hormônios e Genética Molecular/LIM42, Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brasil.

 

Background: Loss-of-function mutations in the coding region of the imprinted gene MKRN3 have recently been recognized as an important cause of familial central precocious puberty (CPP). The promoter region of MKRN3 is notable for potential transcription factor motifs.

Objective: To investigate potential pathogenic variants in the promoter region of MKRN3 in patients with GnRH-dependent pubertal disorders. Variants in the MKRN3 promoter region could result in changes in gene expression and lead to CPP or constitutional delay of growth and puberty (CDGP)

Methods: We studied fifty patients with idiopathic CPP and 25 patients with CDGP. A family history of precocious or delayed sexual development was present in 28% and 32% patients, respectively. Inactivating mutations in the coding region of MKRN3 were excluded in all patients with CPP. Genomic DNA was extracted from peripheral blood leucocytes and a 1000 bp region (-750 to +350 relative to the transcriptional site) of the MKRN3 gene, including potential transcription factor recognition site motifs (PEA3, SRE, SRF, C/EBP, AP2, testis-R), was amplified and automatically sequenced.

Results: We identified a variant, g.23565509T>A (rs182933790), in the promoter region of MKRN3 in a female patient with CPP, who had the onset of pubertal development at age 6.6 years. This variant was also identified in one male patient with CDGP and in one out of 20 control individual suggesting a lack of genotype-phenotype correlation. Furthermore the variant was identified in 1% of the African population from phase 1 of the 1000 Genome Project and the minor allele frequency of this variant was < 0.01, indicating that it is a rare nucleotide change.

Conclusion: No pathogenic variants were detected in the MKRN3 promoter region of patients with idiopathic GnRH-dependent pubertal disorders.

 

Nothing to Disclose: DBM, DDSB, APA, VNB, UBK, ACL

PP03-4 21493 1.0000 THR-144 A Analysis of the MKRN3 Promoter Region in Patients with GnRH- Dependent Pubertal Disorders 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Pierre Chatelain*1, Oleg Malievsky2, Klaudziya Radziuk3, Heba Hassan Elsedfy4, Evgenia Mikhailova5 and Michael Beckert6
1University Claude Bernard, Lyon, France, 2Bashkir State Medical University, Ufa, Russia, 32nd Children City Clinic, Minsk, Belarus, 4Ain Shams University Hospital, Cairo, Egypt, 5Samara State Medical University, Samara, Russia, 6Ascendis Pharma A/S, on behalf of the TransCon hGH study group

 

Background: TransCon hGH is a long-acting prodrug of recombinant human Growth Hormone (hGH) that releases into the blood compartment fully active unmodified hGH.  Previously presented three-month data demonstrated that TransCon hGH has comparable safety and efficacy (height velocity) to daily hGH following repeated dosing in GHD children.

In addition, TransCon hGH was shown in two Phase 1 studies in Healthy Volunteers and a Phase 2 study in adults with hGH Deficiency (AGHD) to: 1) be safe and well tolerated, 2) possess dose dependent, predictable levels of growth hormone, 3) be suitable for a once-weekly dosing regimen, 4) induce an IGF-I pharmacodynamic (PD) response within the normal range throughout the dosing period.

Objectives: The objective of this study is to investigate 1) safety and tolerability, 2) pharmacokinetics (PK) and pharmacodynamics, and 3) efficacy of TransCon hGH in children with Growth Hormone Deficiency.

Design and methods: Pre-pubertal, treatment naïve GHD children received s.c. injections of one of three once-weekly TransCon hGH doses (0.14, 0.21 and 0.30 mg hGH/kg/week) or daily hGH (Genotropin®; 0.03 mg hGH/kg/day = 0.21 mg/kg/week) over a six-month treatment period, in a  randomized, comparator-controlled dose response Phase 2 study. The patients’ GHD diagnosis were established in accordance with international consensus guidelines, based on auxology (height & height velocity), GH stimulation tests & IGF-I. Children Small for Gestational Age (SGA) were excluded.

Results: Safety and efficacy (annualized height velocity), as well as PK and PD data, of up to 25 patients treated over a six-month period with TransCon hGH or daily hGH will be presented. In an interim analysis conducted following three months of treatment all TransCon hGH doses demonstrated an excellent safety / local tolerability profile and an excellent growth within expected ranges (above 10 cm annualized HV) that was comparable to daily hGH.

Conclusions: To date, TransCon hGH has demonstrated efficacy and safety comparable to that observed with daily hGH. Injection site reactions have generally been mild and similar to what is expected with daily hGH injections, with no nodule formation or lipoatrophy noted. This TransCon hGH Phase 2 study conducted in a pediatric population confirms the excellent safety and efficacy profile observed in previous clinical trials.

 

Disclosure: PC: Investigator, Ascendis Pharma. OM: Investigator, Ascendis Pharma. KR: Investigator, Ascendis Pharma A/S. MB: Employee, Ascendis Pharma. Nothing to Disclose: HHE, EM

20252 4.0000 THR-147 A Six-Month Interim Safety and Efficacy of Different Dose Levels of Transcon HGH Administered Once Weekly Versus Standard Daily Human Growth Hormone Replacement Therapy in Pre-Pubertal Children with Growth Hormone Deficiency (GHD) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Zvi Zadik,1, Ron G Rosenfeld*2, Klaudziya Radziuk3, Nataliya Zelinska4, Oleg Malievsky5, Violeta Iotova6, Julia Skorodok7, Ronit Koren8, Leanne Amitzi9, Oren Hershkovitz10, Gili Hart9 and Eyal Fima10
1Kaplan Medical Center, 2Stat 5 LLC, Los Altos, CA, 32nd Children City Clinic, Minsk, Belarus, 4Children Specialized Clinical Hospital, Kiev, Ukraine, 5Bashkir State Medical University, Ufa, Russia, 6UMHAT, Varna, Bulgaria, 7St. Petersburg State Pediatric Medical Academy, St. Petersburg, Russia, 8Prolor, Nes Ziona, Israel, 9OPKO Biologics, Nes Ziona, Israel, 10Prolor Biotech, Nes Ziona, Israel

 

Objective: Growth Hormone (GH) replacement therapy currently requires daily injections, which may cause poor compliance, inconvenience and distress for patients. CTP-modified hGH (MOD-4023) has been developed for once-weekly administration in Growth Hormone Deficient (GHD) adults and children. Pharmacokinetics (PK), pharmacodynamics, (PD) efficacy and safety analysis of weekly treatment with MOD-4023 in GHD naïve children was performed and compared to daily hGH.

 

Design and methods: A randomized, controlled Phase 2 study was conducted in up to 56 pre-pubertal, naïve GHD children receiving subcutaneous injections of one of MOD-4023 doses as a once-weekly regimen (range: 0.25 - 0.66 mg/Kg per week) or daily hGH (34 µg/Kg/day) as a control arm. MOD-4023 and IGF-1 levels were monitored throughout the study. Annual Height Velocity (HV) assessment was evaluated at 12 months.

Results: The baseline characteristics were comparable between all groups. PK/PD profile following administration of MOD-4023 demonstrated a significantly extended half-life as with a dose dependent PK/PD (IGF-1) response was observed between MOD-4023 dose cohorts, reaching steady state with no accumulation or excessive levels by 12 months.  All cohorts demonstrated expected “catch-up” growth, in line with reported age- and GHD severity-matched data. All cohorts demonstrated 12m annual HV above 11 cm/year, correlated with the PK/PD profile in those patients.  Sub analysis of HV response which was based on baseline characteristics confirmed an excellent response is all sub populations.  The 12 months safety analysis suggests a clean and promising profile with no unexpected adverse events.

Conclusions: This is the first report describing PK, PD, efficacy and safety results of extended, 12 months, treatment period with MOD-4023 in pediatric patients with GHD. MOD-4023 administration to GHD children further confirmed its long acting properties. This study further affirmed that a single weekly injection of MOD-4023 has the potential to replace seven consecutive daily injections of currently marketed hGH in pediatric GHD patients and provided data for dose selection for the company’s upcoming Phase 3 trial.

 

Disclosure: KR: Investigator, Ascendis Pharma A/S. OM: Investigator, Ascendis Pharma. Nothing to Disclose: ZZ, RGR, NZ, VI, JS, RK, LA, OH, GH, EF

20955 5.0000 THR-148 A Top Line Results of 12 Months of Once-Weekly Administration of CTP-Modified Human Growth Hormone (MOD-4023): Phase 2 Dose Finding Study in Children with Growth Hormone Deficiency (GHD) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Peter A. Lee*1, Judith L. Ross2, Ricardo Maamari3, Thomas Henschel3 and John Germak3
1Penn State College of Medicine, The Milton S. Hershey Medical Center, Hershey, PA, 2Thomas Jefferson University/duPont Hospital for Children, Philadelphia, PA, 3Novo Nordisk Inc., Plainsboro, NJ

 

Treatment with growth hormone (GH) has been approved by the United States Food and Drug Administration (FDA) for a number of pediatric growth disorders, including Noonan syndrome (NS), and Turner syndrome (TS). Although the pathogenesis of NS and TS is different, they share certain phenotypic features and are generally considered non-GH deficient conditions. The present analysis was undertaken to describe the mean GH dose administered over time, and compare the changes in height standard deviation score (HSDS) and corrected HSDS (HSDS minus target HSDS) in NS versus TS patients enrolled in the American Norditropin Studies: Web-enabled Research (ANSWER) Program registry, from treatment start through 5 years of treatment with Norditropin® (somatropin [rDNA origin] injection, Novo Nordisk A/S, Denmark). As of July 2014, 148 patients with NS (112 males, 36 females) and 550 females with TS, naïve to GH therapy, were enrolled in the ANSWER Program registry. Cross-sectional data were analysed and descriptive statistics applied. Data are presented as mean value ± SD. The mean age at treatment start was 9.2±3.8 years versus 8.5±4.0 years for NS and TS patients, respectively. Mean GH dose (mg/kg/day) at initiation of GH therapy and year (y) 5, respectively, were 0.046±0.011 and 0.050±0.012 for NS and 0.051±0.010 and 0.053±0.013 for TS patients. Over 5 years of GH treatment, <20% of NS, and <15% of TS patients, received GH doses up to the maximum currently approved by the FDA. At treatment start the mean HSDS were –2.61±0.74 and –2.53±0.90, and at y5 were –1.21±0.98 and –1.55±0.92 for NS and TS respectively. Mean change in HSDS from baseline (ΔHSDS) increased over 5 years in NS patients (y1: 0.42±0.39, n=97; y2: 0.77±0.57, n=67; y3: 1.05±0.53, n=50; y4: 1.21±0.63, n=36; y5: 1.30±0.71, n=24). After an initial increase to y3 in TS patients, mean ΔHSDS did not increase further (ΔHSDS: y1: 0.46±0.43, n=373; y2: 0.75±0.55, n=293; y3: 0.93±0.66, n=227; y4: 0.93±0.72, n=164; y5: 0.91±0.76, n=113). Mean corrected HSDS improved over 5 years in NS patients (y1: −1.87±1.14, n=82; y2: −1.50±1.28, n=57; y3: −1.40±1.21, n=45; y4, −0.94±1.21, n=30; y5:–0.76±1.14, n= 21). In contrast TS patients had a plateau in corrected HSDS after y3 (y1: −2.12±1.05, n=312; y2: −1.84±1.04, n=255; y3: −1.76±1.01, n=196; y4: −1.76±1.05, n=140; y5:–1.79±1.06, n=94). Negative correlations between ΔHSDS and baseline age were observed at y1, y2 and y5 of GH treatment in NS patients, but only at y1 in patients with TS. NS patients showed increases in mean HSDS and corrected HSDS over 5 years of GH treatment, in contrast to TS patients, for whom further incremental increases appeared to plateau after y3. Differences in GH responsiveness in NS and TS patients may potentially involve gonadal dysgenesis, skeletal dysplasia related to SHOX deficiency in TS and sub-optimal individual GH dose titration.

 

Disclosure: PAL: Advisory Group Member, Novo Nordisk. JLR: Researcher, Novo Nordisk, Researcher, Versartis, Researcher, Eli Lilly & Company, Advisory Group Member, Novo Nordisk. RM: Employee, Novo Nordisk. TH: Employee, Novo Nordisk. JG: Employee, Novo Nordisk.

18914 7.0000 THR-150 A Effect of Growth Hormone Therapy in Patients with Noonan Syndrome or Turner Syndrome: Analysis of 5-Year GH Dosing and Treatment Outcomes Data from the Answer Program 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Gili Hart*1, Zvi Zadik,2, Klaudziya Radziuk3, Nataliya Zelinska,4, Oleg Malievsky5, Violeta Iotova6, Julia Skorodok7, Ronit Koren8, Leanne Amitzi1, Oren Hershkovitz9 and Eyal Fima9
1OPKO Biologics, Nes Ziona, Israel, 2Kaplan Medical Center, 32nd Children City Clinic, Minsk, Belarus, 4Ukrainian Children Specialized Clinical Hospital, Kyev, Ukraine, 5Bashkir State Medical University, Ufa, Russia, 6UMHAT, Varna, Bulgaria, 7St. Petersburg State Pediatric Medical Academy, St. Petersburg, Russia, 8Prolor, Nes Ziona, Israel, 9Prolor Biotech, Nes Ziona, Israel

 

Objective: Growth Hormone (GH) replacement therapy currently requires daily injections, which may cause poor compliance, inconvenience and distress for patients. CTP-modified hGH (MOD-4023) is being developed for once-weekly administration in Growth Hormone Deficient (GHD) adults and children. In the present study the pharmacokinetics (PK) and pharmacodynamics (PD) profile of MOD-4023 in GHD naïve children was assessed for 12 months of treatment.

Design and methods: A randomized, comparator-controlled Phase 2 study was conducted in up to 56 pre-pubertal, naïve GHD children receiving one of three MOD-4023 doses as once-weekly regimen (0.25, 0.48, 0.66mg/Kg per week) or daily hGH (34µg/Kg/day) as comparator arm in subcutaneous injections. In order to introduce naïve patients to the allocated MOD-4023 dose in a gradual manner, a stepwise dose increase approach was implemented. All patients randomized to receive one of the three MOD-4023 doses started treatment for 2 weeks with the low MOD-4023 dose and based on the patient's dose allocation, followed by a dose increase to the next dose level every two weeks until the final allocated dose was reached. Subsequently to the second dose administration of the targeted dose MOD-4023, GH, IGF-1 and IGF-BP3 concentrations were measured on monthly basis and PK-PD analysis was conducted utilizing a population based approach up to 12 months of treatment.

 

Results: MOD-4023 administration confirmed its long acting properties and superior properties compared to daily hGH. The weekly profile suggests extended half-life, increased exposure and reduced clearance for all patients. Pre-dose and trough levels indicated no accumulation of circulating MOD-4023. IGF-1 and IGF-1 SDS following MOD-4023 administration also increased at a dose proportional manner.

The MOD-4023 profile of throughout the 12 months study reached steady state during the first few weeks of treatment with no indication for excessive levels or accumulation as monitored at 6 month by evaluating trough levels. IGF-1 seem to be normalized and stabilized within the normal range during study and prior to completion the 52 weeks of treatment reaching an optimal value of 0 SDS for the two higher MOD-4023 cohorts which was comparable to the daily hGH arm. As anticipated, IGF-BP3 levels increased following MOD-4023 administration reaching steady state levels by week 15 post study initation.

Conclusions: MOD-4023 once-weekly treatment at three different doses for 52 weeks demonstrated an excellent PK and PD profile supporting once weekly injection in pediatric GHD population and therefore can potentially promote proper growth. In addition, the changes observed in IGF-1 and IGF-BP3 demonstrate adequate stimulation of the GH-IGF-1 axes which were shown to be comparable to that observed with daily hGH treatment

 

Disclosure: KR: Investigator, Ascendis Pharma A/S. OM: Investigator, Ascendis Pharma. Nothing to Disclose: GH, ZZ, NZ, VI, JS, RK, LA, OH, EF

20947 8.0000 THR-151 A Twelve Months Pharmacokinetics and Pharmacodynamics Profile of Once-Weekly, CTP-Modified Human Growth Hormone (MOD-4023): Phase 2 Study in Children with Ghd Deficiency 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Annabel Sophie Berthon*1, Angela Delaney2 and Constantine A Stratakis1
1National Institutes of Health (NIH), Bethesda, MD, 2National Institutes of Health, Bethesda, MD

 

Knockout mice for the kisspeptin receptor, kiss1r, (KRKO) and its ligand kisspeptin, kiss1, (KissKO) replicated the phenotype of isolated hypogonadotropic hypogonadism (IHH) that was first described in association with these genes in humans. Although what leads to adrenarche in humans is unknown, the process precedes and is inextricably linked to pubarche. In this study we analyzed adrenocortical development and function in KRKO and KissKO. At about 5 months of age, in both mouse models, we observe the gradual accumulation of eosinophilic cells at the adrenomedullary boundary. This cell population expressed 20aHSD by immunohistochemistry; Cyp17 is also overexpressed suggesting that these cells are derived from the mouse X-zone which is the transient fetal zone normally disappearing at puberty in male and during the first pregnancy in female mice. Moreover, the KissKO females (but not the males) show corticosterone and aldosterone hypersecretion, which could be due to the documented overexpression of Star. Ongoing studies are focusing at further characterizing the adrenocortical phenotype of these mice. We conclude that KRKO and KissKO mice have a previously unrecognized adrenocortical phenotype that is potentially very interesting and may provide links to the understanding of the long suspected but never proven common regulatory mechanisms of adrenarche and pubarche in humans.

 

Nothing to Disclose: ASB, AD, CAS

21006 9.0000 THR-152 A Fetal Zone Persistence in Adrenal Cortex of Kisspeptin and Kisspeptin Receptor Knockout Mice: Implications for Human Puberty and Adrenal Development 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Kennett Sprogoe*1, Michael Beckert2, David Gilfoyle3, Thomas Kurpiers4, Thomas Wegge4 and Harald Rau4
1Ascendis Pharma, Inc., 2Ascendis Pharma A/S, on behalf of the TransCon hGH study group, 3Ascendis Pharma A/S, Hellerup, Denmark, 4Ascendis Pharma GmbH

 

Background

TransCon hGH is a once-weekly prodrug of recombinant human growth hormone (hGH) intended to improve compliance and treatment outcome of patients requiring growth hormone therapy. TransCon hGH is designed to maintain the same mode of action and distribution within the body as daily administered hGH. TransCon hGH liberates unmodified hGH into the bloodstream via a non-enzymatic self-cleaving process relying only on physiological pH and temperature conditions, which ensures predictable release of unmodified hGH. The hGH released from TransCon hGH has been demonstrated to be identical to hGH for daily administration, for which safety and efficacy has been established through decades of therapy. In clinical studies, TransCon hGH has demonstrated an efficacy, safety, tolerability and immunogenic profile that is comparable to that of daily hGH.

Objectives

To compare the structure and in vitrobiopotency of growth hormone released from TransCon hGH with hGH for daily administration, and compare the exposure of hGH released from TransCon hGH with daily administered hGH, together with the resulting IGF-I levels in GHD adults.

Design and methods

hGH released from TransCon hGH was characterized by high resolution mass spectrometry, circular dichroism, N-terminal amino acid sequencing, peptide mapping and cell based biopotency assays. Pharmacokinetic and pharmacodynamic data of TransCon hGH and daily hGH was obtained in a Phase 2 study in GHD adults.

Results

Characterization of hGH released from TransCon hGH confirmed that it is unmodified and structurally identical to hGH for daily administration.  Cell based biopotency assays confirm that full activity of the growth hormone is restored when it is released from the prodrug. PK/PD data obtained from GHD adults show that maximum levels of both hGH and IGF-I are comparable between weekly administered TransCon hGH and daily administered hGH, with no supraphysiological exposure to either hGH or IGF-I.

Conclusion

Whether released from the pituitary gland or administered as daily injections, growth hormone distributes throughout the body to exert important effects in growth hormone responsive tissue, including bone, muscle, liver and fat tissue. These include anabolic and metabolic effects. Growth hormone released from TransCon hGH has been shown to be identical to hGH in daily administered products. Clinical studies in adults with GHD demonstrate that TransCon hGH has comparable efficacy to daily hGH when administered at the same cumulative weekly dose. Importantly, TransCon hGH does not cause supraphysiological exposure to hGH or IGF-I. 

Designed to maintain the same mode of action and distribution to growth hormone responsive tissues as daily hGH, TransCon hGH has the potential to offer patients requiring growth hormone therapy a long-acting alternative to daily injections

 

Disclosure: KS: Employee, Ascendis Pharma. MB: Employee, Ascendis Pharma. DG: Employee, Ascendis Pharma. TK: Employee, Ascendis Pharma. TW: Employee, Ascendis Pharma. HR: Employee, Ascendis Pharma.

20549 10.0000 THR-153 A Transcon Human Growth Hormone – a Once-Weekly Prodrug of Recombinant Human Growth Hormone: Design, Characterization and PK/PD in Growth Hormone Deficient (GHD) Adults 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Youn Hee Jee*1, Kun Song Lee2, Alan T Remaley3, Young Pyo Chang4, Anton Wellstein5, Jack Adam Yanovski6 and Jeffrey Baron7
1NICHD, Bethesda, MD, 2Dankook University, Korea, Republic of (South), 3NHLBI, NIH, Bethesda, MD, 4Dankook University Hospital, Korea, Republic of (South), 5Georgetown University, Washington, DC, 6Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health (NIH), Bethesda, MD, 7NIH, Bethesda, MD

 

Heparin-Binding Growth Factor Concentrations in Children with Idiopathic Short Stature

Youn Hee Jee, Kun Song Lee, Alan T. Remaley, Young Pyo Chang, Anton Wellstein, Jack A. Yanovski, Jeffrey Baron

Background: Midkine (MDK) and pleiotrophin (PTN) are two closely-related heparin-binding growth factors that in rodents are highly expressed in early life in multiple organs and decrease to undetectable levels by adulthood. The roles of MDK and PTN in human growth and development are unknown.

Objective: To examine associations of plasma MDK and urine PTN with age, sex, adiposity, and growth in children.

Methods: Plasma MDK was measured by ELISA in150 healthy children (mean age, 11.7 y; range 0 to 24 y; BMI SDS range, -2.3 to +3.2) and random urine PTN was measured by ELISA in 87 healthy children (mean age, 15.0 y; range 3 to 24.7 y; BMI SDS range, -2.3 to +2.7). Plasma MDK and urine PTN were also measured in 41 children referred for short stature (mean height SDS, -2.4), including idiopathic short stature (ISS, n=25), constitutional delay (n=12), growth hormone (GH) deficiency (n=1), and small for gestational age (n=3).

Results: In healthy children, plasma MDK concentrations declined with age (R2=0.41, p< 0.001) paralleling height velocity, with values highest in infants. Plasma MDK did not differ significantly between males and females and did not correlate with BMI SDS or percent total body fat. In most of the children with short stature, MDK concentrations did not differ significantly from normal subjects, regardless of diagnosis or GH treatment status, and did not correlate with height SDS or IGF-1 SDS. One 15-year old girl with ISS had 20 fold greater plasma MDK than the mean for age-matched controls (2.9 vs 0.14 ± 0.1 ng/mL, mean ± SD, n=29), and one newborn with intrauterine growth retardation had a low MDK concentrations compared to healthy newborns (0.17 vs 0.9 ± 0.5 ng/mL, mean ± SD, n=9). In healthy children, urine PTN normalized to creatinine (PTN/Cr) also declined with age (R2=0.3, p<0.001). Urine PTN did not differ significantly between males and females and did not correlate with BMI SDS or percent total body fat. However, 11 children with constitutional delay showed significantly higher PTN than age-matched controls or children with other causes of short stature (p<0.001).

Conclusions: In healthy children, plasma MDK concentrations and urine PTN/Cr ratio declined with age, paralleling the physiological decline in growth velocity and were not significantly associated with sex or adiposity. Plasma MDK concentration was markedly increased in one child with ISS and decreased in one SGA newborn. Urine PTN/Cr was elevated in constitutional delay. To establish whether MDK and PTN regulate human growth and whether abnormalities involving MDK or PTN could cause human growth disorders will require further investigation.

 

Disclosure: JAY: Principal Investigator, Zafgen. Nothing to Disclose: YHJ, KSL, ATR, YPC, AW, JB

19343 11.0000 THR-154 A Heparin-Binding Growth Factor Concentrations in Children with Idiopathic Short Stature 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


In Ah Jung1, Yeon Jin Jeon*1, Won-Kyoung Cho1, Jae Wook Lee1, Shin-Hee Kim2, Nak-Gyun Chung1, Kyoung Soon Cho1, Bin Cho1, So Hyun Park1, Min-Ho Jung1, Hyo-Jin Kim1 and Byung-Kyu Suh1
1College of Medicine, The Catholic University of Korea, Seoul, Korea, Republic of (South), 2The Catholic University of Korea, Seoul, Korea, Republic of (South)

 

Background: Long-term survivors of childhood acute leukemia can suffer from growth impairment. The purpose of this study was to evaluate longitudinal growth and final height of pediatric patients who were treated with acute leukemia and factors that can cause growth impairment.

Methods: Of 234 patients (133 males and 101 females) who were diagnosed as acute lymphoblastic leukemia (ALL; n=162) or acute myeloblastic leukemia (AML; n=72) before age 18 between June 1996 and June 2009, 27 patients who were diagnosed as hypogonadism or growth hormone deficiency were excluded and 207 patients (123 males and 84 females) were included. The mean age at enrollment was 14.97 ± 4.36 years. Among them, 155 patients treated with chemotherapy only and 52 patients received hematopoietic stem cell transplantation (HSCT). We reviewed height standard deviation scores (SDS) from diagnosis to 5 years after the diagnosis. Final height SDS of 89 patients who reached final height at enrollment and risk factors such as sex, age at diagnosis, diagnosis, pubertal status at diagnosis, radiation and chronic graft-versus-host-disease (cGVHD) that can affect growth were evaluated.

Results: The mean height SDS at diagnosis of 207 patients was 0.47 ± 1.01 and at 5 years after the diagnosis was -0.06 ± 1.06. The mean height SDS from the diagnosis to 5 years after the diagnosis were significantly decreased (Dheight SDS = -0.53 ± 0.82, p<0.001). And the mean final height SDS of 89 patients was -0.64 ± 1.06 that was significantly lower than the height SDS at diagnosis (Dheight SDS = -1.01 ± 1.10, p<0.001). The height loss was more severe in the patients who received HSCT than the patients who received chemotherapy only (-1.42 ± 1.37 vs. -0.81 ± 0.87, p=0.031). In chemotherapy only patients (n=59), the changes in height SDS from the diagnosis to final height were positively correlated with age at diagnosis (r=0.450, p<0.001) and negatively correlated with cranial irradiation (r=-0.297, p=0.022). In multiple regression analysis, younger age at diagnosis was the only significant risk factor of loss in final height in chemotherapy group. In HSCT patients (n=30), the differences of height SDS between at diagnosis and final height were showed positive correlation with age at diagnosis (r=0.664, p<0.001) and negative correlation with pubertal status at diagnosis (r=-0.510, p=0.004) and total body irradiation (TBI) for conditioning (r=-0.422, p=0.020). In multiple regression analysis, younger age at diagnosis and TBI were   significant risk factors cause loss of final height in HSCT patients (p=0.006 and p=0.010, respectively).

Conclusions: Growth impairment in survivors of childhood acute leukemia was significantly related with younger age at diagnosis.

 

Nothing to Disclose: IAJ, YJJ, WKC, JWL, SHK, NGC, KSC, BC, SHP, MHJ, HJK, BKS

20160 12.0000 THR-155 A Growth Impairment in Children with Acute Leukemia 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Jung Min Ahn*1, Ah Reum Kwon2, Duk Hee Kim3 and Ho-Seong Kim4
1Yonsei University College of Medicine, Seoul, Korea, Republic of (South), 2Severance Hospital, Seoul, Korea, Republic of (South), 3Sowha Children's Hospital, Seoul, Korea, Republic of (South), 4Severance Children's Hospita, Seoul, Korea, Republic of (South)

 

Purpose: The purpose of this study was to screen the gene mutations in Korean children with isolated growth hormone deficiency (GHD) and analyze their characteristics.

Methods: One hundred four patients with isolated GHD were recruited from Severance Hospital. The diagnosis GHD was based on the following criteria: short stature as defined by height less than -2 SD of the age- and sex-matched population; reduced growth velocity; and blunted GH response (<10 ng/ml) to two different pharmacological stimulation test. DNA was extracted from peripheral lymphocytes and amplified using primers designed to include coding and splicing regions of individual exons for GH1, GH releasing hormone receptor (GHRHR) and GH secretagogue receptor (GHSR).

Results: Mutations in GH1, GHRHR and GHSR were identified in 12.5% (13/104). These included previously described mutations in GH1 (c.69 A>G, p.T3A), GHRHR (c.214C>T, p.P16L, c.721G>A, p.V225I) and GHSR (c.144T>C, p.L34P). All ten patients with GH1 mutation led to GHD type IB. There were no differences in the age (8.3± 2.9 vs. 8.5 ± 3.6) (P = 0.44), degree of growth failure ( -3.7 ± 0.9 SDS vs. -3.6 ± 0.5 SDS) (P = 0.12), or endocrinologic characteristics (IGF-1, -0.27 ± 0.17 SDS vs. -0.26 ± 0.21 SDS) (P = 0.29) between patients with and without GH1 mutation. MRI results were available from all patients, there was no difference in the incidence of pituitary abnormalities between patients with and without GH1 mutation (15.3% vs. 14.2%).    

Conclusion: We screened the mutations in GH1, GHRHR and GHSR genes in a large cohort of Korean patients with GHD. In our study, the prevalence of mutations was 12.5%, however we did not find any difference in clinical manifestations between patients with and without mutations.

 

Nothing to Disclose: JMA, ARK, DHK, HSK

20184 13.0000 THR-156 A Genetic Analysis in Korean Children with Isolated Growth Hormone Deficiency 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Mengjie Wang*1, Youjie Zhang1, Dan Lan2 and Jennifer Wootton Hill3
1University of Toledo, Toledo, OH, 2Guangxi Medical University, Nanning, China, 3University of Toledo School of Medicine, Toledo, OH

 

Objective: Our aim was to systematically review and evaluate the efficacy and safety of the addition of recombinant human growth hormone (rhGH) to gonadotropin-releasing hormone analogs (GnRHa) treatment in Chinese children with central precocious puberty (CPP).
Method: We performed a thorough search of the databases (Medline, Embase, the Cochrane Library, Chinese Biomedical Literature Service System, Wanfang Data, Chinese Scientific Journals Fulltext Database, and China National Knowledge Infrastructure). Relevant studies were included, divided and analyzed based on their research arms. Outcome measurements were height, predicted adult height (PAH), the height standard deviation score for bone age (HtSDS-BA) and growth velocity (GV). Adverse effects, including body mass index (BMI) and insulin like growth factor-1 (IGF-1) were evaluated. All analyses were conducted using the software of Stata 11.0.
Results: A total of 13 case-control studies and/or observational studies containing 404 patients were included and analyzed. Compared with placebo groups, administration of GnRHa plus rhGH led to a significant increase in height, PAH, HtSDS-BA and GV, corresponding to a weighted mean difference (WMD) (95% confidence interval (95%CI)) of 8.12 cm (5.70, 10.54), 6.26cm (4.02, 8.49), 0.76 (0.41, 1.11), 3.39 cm/year (2.57, 4.22), respectively. On meta-regression, a negative association was seen between the change in PAH and subjects’ age (β coefficient=-1.489, 95%CI -2.77 to -0.20, P=0.027). Even though meta-regression did not show a statistical significance between the outcomes (height and HtSDS-BA) and subjects’ age, negative trends were seen between the outcomes and subjects’ age. No statistically significant association was found between the changes in the following outcomes (height, PAH, HtSDS-BA and GV) and the therapy duration. In the subgroup analysis, we found that the combined therapy had increased efficacy in subjects younger than 10 years old. However, there was no significant difference between treatment lasting less than 12 months and over 12 months. To further identify the effect of the addition of rhGH to GnRHa, we compared the combined treatment with use of GnRHa alone. The combined treatment was superior with regard to height [3.76 cm (2.46, 5.05)], PAH [3.68 cm (2.82, 4.54)] and HtSDS-BA [0.56 (0.40, 0.73)]. No obvious adverse effects were observed during the GnRHa and rhGH adjunctive therapy.
Conclusion: The results of this meta-analysis may allay concern about the efficacy and safety of rhGH plus GnRHa treatment. The efficacy of the combined therapy could be somewhat increased with use in subjects younger than 10 years old. However, the efficacy did not depend on the therapy duration. Multi-center, long-term and well-designed studies are greatly needed to learn more about the mechanism, the efficacy, and the safety of the addition of rhGH to GnRHa during CPP therapy.

 

Nothing to Disclose: MW, YZ, DL, JWH

19098 14.0000 THR-157 A The Efficacy of Gonadotropin-Releasing Hormone Analogs Alone or in Combination with Recombinant Human Growth Hormone for the Treatment of Chinese Children with Central Precocious Puberty: A Systemic Review and Meta-Analysis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Filippina Filia Dimitriadi*, Anita Azam, Francesco De Luca and Rita Ann Kubicky
St. Christopher's Hospital for Children, Drexel University College of Medicine, Philadelphia, PA

 

Background:  X-ALD is described as a disorder with many different phenotypes: Addison disease, cerebral ALD, adrenomyeloneuropathy (AMN) and carriers.

Clinical Case: J.I. is an AA male who presented at the age of 10 5/12 years with one episode of generalized tonic clonic seizure complicated by a prolonged postictal state and followed by rapid deterioration of mental status requiring endotracheal intubation and ICU admission at St. Christopher’s Hospital for Children. Blood glucose, calcium, and electrolytes were within normal limits. Family reported deterioration in school performance, memory loss and unsteady gait in the past year.

Review of systems was negative for headaches, abdominal pain, and fatigue.

Family history was positive for febrile seizures in the 2 year old sister and epilepsy in the maternal grandfather.

During his hospital stay, he received a 3-day course of solumedrol due to concern of acute disseminated encephalomyelitis. On hospital day 2, he was extubated and his mental status returned to baseline. EEG was normal. MRI of his brain revealed white matter signal abnormality consistent with demyelinating process.

When seen in the Endocrinology Clinic at SCHC 1 month after his discharge, he had a normal neurological exam; and he lacked skin or buccal hyperpigmentation.

Standard dose ACTH stimulation test revealed a peak serum cortisol of 2.6 mcg/dL. PRA was normal (0.15 ng/mL/h) and baseline ACTH elevated at 277.5 pg/mL (normal range, 7.2-63.3); these findings were consistent with the diagnosis of primary adrenal insufficiency with no mineralocorticoid deficiency. J.I. was started on hydrocortisone 10 mg/m2/day.

VLCFAs showed elevation in C26:0, ratio of C24/22 and C26/22 consistent with X-ALD or AMN. Genetic testing of the ABCD1 gene found him to be hemizygous for the c1415_1416delAG mutation.

Given J.I.’s diagnosis, his older brother (although asymptomatic at the age of 16 1/12 years) underwent a diagnostic evaluation that showed elevated VLCFAs, normal MRI brain and he was also found hemizygous for the same mutation. His peak stimulated cortisol was 8.5 mcg/dL with normal PRA level of 0.89 ng/mL/h. He was started on hydrocortisone.

Conclusion: X-ALD is a disorder still incompletely understood, very challenging to diagnose and with very limited treatment options. It should be considered in the differential diagnosis of myelopathy in both males and females. Our case presentation of variable phenotype in two siblings with X-ALD underlines the importance of genetic testing in family members and appropriate counseling and management when the diagnosis is established.

 

Nothing to Disclose: FFD, AA, FD, RAK

20020 15.0000 THR-158 A Variable Phenotype in Two Siblings with X-Linked Adrenoleukodystrophy (X-ALD) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Filippina Filia Dimitriadi*1, Shufang Wu2, Wei Yang2, Bruce Bernstein3, Francesco De Luca4 and Rita Ann Kubicky2
1St. Christopher's Hospital for Children, Drexel University College of Medicine, Philadelphia, PA, 2Section of Endocrinology and Diabetes, St. Christopher's Hospital for Children, Drexel University College of Medicine, Philadelphia, PA, 3St. Christopher's Hospital for Children, Philadelphia, PA, 4Section of Endocrinology and Diabetes, St. Christopher's Hospital for Children, Department of Pediatrics. Drexel University College of Medicine, Philadelphia, PA

 

Background:Fibroblast growth factor 21 (FGF21) has been shown to mediate metabolic adaptation to fasting in rodents and humans. In previous studies, we have also demonstrated that increased FGF21 causes Growth Hormone (GH) insensitivity and, in turn, growth failure. Based on this evidence, we hypothesized that serum FGF21 in prepubertal children with short stature are positively associated with serum GH levels, and negatively associated with serum IGF-1, height, weight, and growth velocity.

Methods:To test our hypothesis, we prospectively enrolled prepubertal children undergoing evaluation for short stature in the Section of Endocrinology and Diabetes at St. Christopher’s Hospital for Children. Blood samples were collected between 8 am and 5 pm. Serum FGF21 was measured using an FGF21 Human ELISA kit (Millipore). GH was measured by RIA, while IGF-1 was measured by LC/MS.

Exclusion criteria included diagnosis of chronic disorders, prolonged (> 6 weeks in the preceding 6 months), administration of medications known to be associated with underweight or poor growth velocity/short stature, or pubertal stage of ≥ Tanner stage 2.

Anthropometric data such as height, weight, BMI/Weight-for-length, gender and age (in months) were collected. Using these data, we calculated Height and BMI Z-scores. Growth velocity Z-score was calculated using the height measurements upon enrollment and within the 6 months preceding enrollment.

Results:22 patients (18 males and 4 females) were recruited. Their mean age was 5.6 ± 2.9 years.   Height Z-score -2.6 ± 0.85; BMI Z-score 0.01 ± 1; Growth velocity Z-score  -0.5 ± 1.9. 

Mean FGF21 level was 132.5 ± 162.6 pg/mL (range 0- 591pg/mL, with undetectable level set to 0 pg/mL). No significant difference was identified in the median FGF21 levels between genders (Mann Whitney U test, p = .99). Mean GH level was 3 ± 3.8 ng/mL and mean IGF-1 level was 89.2 ±  41.5 ng/mL.

No significant correlations with height Z-score, growth velocity Z-score, weight Z-score, serum IGF-1 or serum GH levels were found (Spearman’s rank order statistics).

Discussion:To our knowledge this is the first study reporting circulating FGF21 levels in prepubertal children with short stature.

Our results suggest that there is no association between serum FGF21, serum GH and IGF-1 levels in prepubertal children. In addition, we did not find any association between FGF21 levels and anthropometric measures. In contrast to our results, these associations have been demonstrated in two studies on premature infants.  It is possible that differences in age and nutritional status between our patient population and others may explain these discordant findings.

Longitudinal studies on a larger cohort of prepubertal short children with a wider BMI range may further elucidate the role of FGF21 in human growth.

 

Nothing to Disclose: FFD, SW, WY, BB, FD, RAK

19818 16.0000 THR-159 A Serum FGF21 in Prepubertal Children with Short Stature: Relationship to IGF-1 and GH Levels and Growth Parameters 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Peter Davies*1, Andrea Luczay2, Evangelia Charamndari3, Jeremy Kirk4, Ho-Seong Kim5, Ludmila Kostalova6, Marc Nicolino7, Martin Borkenstein8, Maria Dolores Rodriguez Arnao9, Raúl Calzada-León10, Svante Norgren11, John VanderMeulen12, Sandro Loche13, Alicia Belgorosky14, Minlian Du15, George Stoyanov16, Ekaterina B Koledova17, Jeff Zhao18 and Jan Lebl19
1University of Queensland, Herston, Australia, 2Semmelweis University, Budapest, Hungary, 3University of Athens Medical School, Athens, Greece, 4Birmingham Children's Hospital, Birmingham, United Kingdom, 5Yonsei University, Seoul, Korea, Republic of (South), 6Comenius University Medical School, Bratislava, Slovakia, 7Hôpital Femme-Mère-Enfant, Lyon, France, 8Medical University of Graz, Graz, Austria, 9Hospital Universitario Gregorio Marañón, Madrid, Spain, 10Instituto Nacional de Pediatría, México City, Mexico, 11Karolinska University Hospital, Stockholm, Sweden, 12McMaster Children’s Hospital and McMaster University, Hamilton, ON, Canada, 13Ospedale Microcitemico - ASL Cagliari, Cagliari, Italy, 14Hospital de Pediatría Garrahan, Buenos Aires, Argentina, 15The First Affiliated Hospital of SUN Yat-Sen University, Guangzhou 510080, China, 16EMD Serono, Mississauga, ON, Canada, 17Merck, Darmstadt, Germany, 18EMD Serono, Billerica, MA, 19Charles University in Prague, Prague, Czech Republic

 

Background: The easypod™ auto-injector device enables accurate records of patients’ adherence to recombinant human growth hormone (r-hGH) to be collected, providing real-world data for evaluation. ECOS is an observational study, started in 2010 and now active in 23 countries, with 2,403 patients enrolled. ECOS will evaluate adherence and the factors that may influence it in paediatric patients prescribed r-hGH via easypod™.

Objectives: The primary objective of the ECOS study is to evaluate the level of adherence of paediatric patients receiving r-hGH via easypod™. Secondary objectives include assessment of the impact of adherence on clinical outcomes and the concentrations of insulin-like growth factor 1 (IGF-1) and identification of factors that may influence adherence to this form of treatment.

Methods: ECOS will follow children with Growth Hormone deficiency (GHD), Small for Gestational Age (SGA) and Turner Syndrome (TS) receiving r-hGH therapy for up to 5 years, with interim analyses each year. Demographic, auxological and diagnostic data are obtained from medical notes, with accurate adherence data obtained directly from the patients' easypod™ auto-injectors. Adherence during the study period is defined as the number of days with injections received, divided by the number of days with injections planned, expressed as a percentage. An interim analysis was completed in May 2014.

Results: At the time of analysis, 1,972 patients had been enrolled. The majority were male (57.7%), Caucasian (73.1%), with a diagnosis of GHD (65.7%), SGA (15.0%) or TS (7.7%). Baseline mean [Standard Deviation] age was 9.8 [3.7], 8.9 [3.3], 9.5 [3.8] years and height 118.7 [20.9], 113.4 [18.1], 109.7 [19.7] cm for GHD, SGA and TS, respectively. Investigator-assessed mean growth velocity at start of GH treatment was 4.5 [2.6], 4.7 [1.8], 6.3 [12.1] cm/yr for GHD, SGA and TS.

Individual levels of adherence prospectively measured with easypod™ (median [Q1, Q3], 93.0% [82.8%, 97.5%]) were higher than those previously reported in retrospective studies based on questionnaires and were maintained over time. After 1 year, height velocity (HV) was 7.9 [2.1], 8.0 (1.9) and 6.9 [2.4] cm/yr and HV SDS was 2.13 [2.76], 1.81 [2.17] and 1.51 [2.92] for GHD, SGA and TS. Spearman's product-moment correlation for adherence rate and change in height (0.085, 0.101 0.471 for GHD, SGA and TS) and height velocity (0.132, 0.149, 0.488 for GHD, SGA and TS) was only weakly positive. Age at start of treatment was the most important predictor of growth and growth rates improved less in patients who were not naïve to GH treatment or the easypod™ device.

Conclusions: Adherence rates with the easypod™ device are high and maintained over time. The younger the patient, the better was the first-year response to GH treatment. Inclusion of older children with previous GH therapy may have reduced the correlation between adherence rates and growth outcomes.

 

Disclosure: PD: Committee Member, Merck Serono, Investigator, Merck Serono. AL: Committee Member, Merck Serono, Investigator, Merck Serono. EC: Committee Member, Merck Serono, Investigator, Merck Serono. JK: Committee Member, Merck Serono, Investigator, Merck Serono. HSK: Committee Member, Merck Serono, Investigator, Merck Serono. LK: Committee Member, Merck Serono, Investigator, Merck Serono. MN: Committee Member, Merck Serono, Investigator, Merck Serono. MB: Committee Member, Merck Serono, Investigator, Merck Serono. MDR: Committee Member, Merck Serono, Investigator, Merck Serono. SN: Committee Member, Merck Serono, Investigator, Merck Serono. JV: Committee Member, Merck Serono, Investigator, Merck Serono. SL: Committee Member, Merck Serono, Investigator, Merck Serono. AB: Committee Member, Merck Serono, Investigator, Merck Serono. MD: Committee Member, Merck Serono, Investigator, Merck Serono. GS: Employee, Merck Serono. EBK: Employee, Merck Serono. JZ: Employee, Merck Serono. JL: Committee Member, Merck Serono, Investigator, Merck Serono. Nothing to Disclose: RC

21735 17.0000 THR-160 A The Easypod<sup>TM</sup> Connect Observational Study (ECOS): Results from the 2014 Interim Analysis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Charikleia Stefanaki*1, George Paltoglou2, Flora Bacopoulou1, Dario Boschiero3 and George P Chrousos4
1University of Athens Medical School, Athens, Greece, 2Athens University Medical School, Athens, Greece, 3Biotekna, Venice, Italy, 4Athens University Medical School

 

Growth hormone is fundamental in skeletal growth during puberty, however detailed studies of body composition analyses in adolescents on growth hormone replacement therapy are scarce. In this case-control study, we aimed to investigate differences in body composition, based on the measurement of bio-impedance, between adolescents with idiopathic growth hormone deficiency (GHD) who were treated with recombinant human growth hormone (rhGH) for at least 6 months (treatment group) and adolescents with GHD who had not yet received rhGH (control group). Over a period of 1 year, 10 adolescents (8 males, 2 females; mean age±SD: 12.8±2.6 years) on rhGH treatment and 10 controls (5 males, 5 females; mean age±SD: 13.3 ±2.2 years) matched for age, height, body mass index (BMI) and bone age, were enrolled. Participants underwent dual frequency bio-impedance assessment with the use of the BIA-ACC, BIOTEKNA© device. Statistical analysis revealed significant differences in tissue hydration, as total (p=0.017), intracellular (p=0.023) and extracellular water volume (p=0.018) were significantly increased in the treatment vs. the control group. GH anabolic effects were demonstrated in increased body cell mass (p=0.022), extracellular fluid protein mass (p=0.02), skeletal muscle mass (p=0.02), fat-free mass (p=0.02), total body protein (p= 0.023), BMI (p=0.018) and glycogen mass (p=0.022). The most interesting findings were the increased bone (p=0.015), bone mineral (p=0.018) and soft tissue mineral (p=0.016) content, along with total body magnesium (p=0.02) and calcium concentrations (p=0.017). Metabolism, in terms of kcal per day, was also increased in the treatment group (p=0.02), mainly due to the increased metabolism of the bones (p=0.01) and skeletal muscles (p=0.02). Interestingly, no statistically significant differences in fat mass were found between the two groups. In the treatment group, extracellular potassium concentrations (p=0.019) and mass (p=0.02) were increased in relation to the control group but below the maximum critical value that was given by the manufacturer. These findings probably reflect increased metabolism resulting in decreased exchangeable potassium in the body (1). In conclusion this study reveals that bio-impedance analysis may be a potent, non-invasive tool for assaying body composition, while also confirming the expected beneficial changes in muscle and bone mass and bone mineralization following rhGH treatment. Further prospective studies with large numbers of patients are needed to confirm our results.

 

Disclosure: DB: Chief Scientific Officer, BioTekna srl. Nothing to Disclose: CS, GP, FB, GPC

21690 18.0000 THR-161 A Bio-Impedance Body Composition Analysis in Adolescents with Idiopathic GH Deficiency Confirmed Significant Increase in Hydration, Skeletal Muscle Mass, Bone Mineralization, and Metabolism but No Change in Fat Mass 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Gabriela Sobrero*1, Maria Cecilia Aguirre2, Liliana Silvano3, Cintia Soledad Tarifa2, Alejandra Paez Nuñez2, Mariana Ochetti2, Maria Julia Alvarez2, Graciela María Testa1, Silvia Edith Martin4, Mirta Beatriz Miras1 and Liliana Muñoz3
1Hospital de Niños de la Santisima Trinidad, Cordoba, Argentina, 2Hospital de Niños de la Santísima Trinidad. Córdoba, 3Hospital de Niños de la Santísima Trinidad. Córdoba, Córdoba, Argentina, 4Hospital de Niños de la Santísima Trinidad. Córdoba, Argentina

 

Background: The diagnosis of growth hormone deficiency in children is based on clinical, auxological, radiographic and biochemical criteria which include response to pharmacological stimulation tests (PhT). It is well known that GH concentrations vary according to the assay method used. Currently, the proposed cut-off value of serum GH PhT is 4,70 ng/mL measured by ICMA using the IRP 98/574, however there are no data about cut-off using ECLIA methodology with the same IRP in our population.

Objetive: The aims of this study were to define GH cut-off level by ECLIA in our population and to compare the concentrations measured by ECLIA and ICMA.

Method: We analyzed 245 PhT (72 F, 173 M) from 192 children with short stature aged 0,8 to 16 years, arginine PhT n: 175 and clonidine PhT n: 70. GH concentrations was determined by ICMA (Immulite 2000) and ECLIA (Cobas e 601) using the IRP 98/574.

Results: Linear regression analysis was performed in 1120 samples measured by ICMA vs. ECLIA showing highly significant linearity (y= 0.28+0.98 x; R2=0.95). The ECLIA cut-off of 4,65 + / - 0,22 ng/mL is the best match with the proposed by ICMA (association coefficient phi = 0,88). We found 96% agreement in the interpretation of PhT using the cut-off for each method.

Conclusion: GH measurements by ICMA and ECLIA using the IRP 98/574 were highly correlated. The use of uniform standardization and establishment of cut-off values for each methodology and population allow a consistent interpretation of the results and assist in the accurate diagnosis of GH deficiency.

 

Nothing to Disclose: GS, MCA, LS, CST, AP, MO, MJA, GMT, SEM, MBM, LM

21922 19.0000 THR-162 A Serum GH Cut-Off Levels By Eclia Performed in Pharmacological Stimulation Tests in Children with Short Stature 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Byung-Ho Kang*1, Mun Suk Park2, You Jung Yang3, Sung Jig Lim3 and Kye Shik Shim2
1Kyung Hee University College of Medicine, Seoul, Korea, Republic of (South), 2Kyung Hee University Hospital, Seoul, Korea, Republic of (South), 3Kyung Hee University Hospital, Seoul, South Korea

 

Purpose

The purpose was to get the basic data of optimum serum concentration of estrogen in pubertal growth spurt, minimizing the decrease of bone mineral density or acceleration of epiphyseal closure of long bones.

Method

1. Fifteen female SD rats (4-week aged) were ovariectomized to inhibit their endogenous estrogen effect and randomly divided into 3 groups. After 1 week, the group 1 were injected subcutaneously with sesame oil, as a control, group 2 were with 100µg/kg/week of estradiol depo as a high-dose, and group 3 were with 200µg/kg/week of it as a super-high-dose for 10 weeks on their posterior neck area.

2. Their crown-lump length, body weight, and bone mineral density of spine and long bones were checked weekly from 1 week before through 10 weeks after injections.

3. Serum levels of GH and estradiol were checked with ELISA before and after injections.

4. After 10 weeks of injections, they were euthanized, pituitary glands were dissected and their pituitary Gh1 mRNA levels were checked with quantitative RT-PCR.

5. Their proximal tibia were dissected and stained with hematoxylin-eosin.

6. The thicknesses of epiphyseal plate including proliferative and hypertrophic zone of the proximal tibias were measured in 20 evenly divided sites with microscope.

7. Statistical analyses were done among the 3 groups before and after injections using ANOVA with multiple comparisons for auxological data, and Kruskall Wallis test for seum levels of GH, estradiol, and pituitary Gh1 mRNA levels with SPSS ver.13.0.

Results:

1. There were no significant differences in body lengths among 3 groups.

2. The body weights were decreased, and the bone mineral densities were increased in group 3 but there were no significant differences.

3. Serum GH levels and pituitary Gh1 expressions were significantly increased in group 3.

4. There was no significant difference in the thickness of total epiphyseal plate, proliferative zone, but that of hypertrophic zone of the epiphyses was significantly increased in group 3.

Conclusion

1. GH secretion and Gh1 gene expression were increased after super-high dose estrogen treatment.

2. There were tendencies that body weight is negative and bone mineral density is positive relation with estrogen dosage, but with no significant differences.

3. The thickness of hypertrophic zone in epiphyseal plate was relatively increased after super-high-dose estrogen treatment, maybe because of increased transdifferentiation of osteoblast to osteocyte.

4. The effects of estrogen on epiphyseal plate in rodents may be different with human.

 

Nothing to Disclose: BHK, MSP, YJY, SJL, KSS

21433 20.0000 THR-163 A Change of Growth Pattern and Bone Mineral Density in Ovariectomized Female Rats According to Estrogen Dosage 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Christopher J Child*1, Charmian A Quigley2, Alan G Zimmermann3, Cheri L Deal4, Judith L. Ross5, Eckhard Schoenau6 and Werner F Blum7
1Eli Lilly and Company, Windlesham, United Kingdom, 2Sydney Children’s Hospital, Sydney, Australia, 3Eli Lilly and Company, Indianapolis, IN, 4Sainte-Justine Hospital, Montreal, QC, Canada, 5Thomas Jefferson University/duPont Hospital for Children, Philadelphia, PA, 6University of Cologne, Cologne, Germany, 7University of Giessen, Giessen, Germany

 

GH treatment in short children born small for gestational age (SGA) has shown a growth-promoting effect in both the short- and long-term, and is an approved indication in both Europe and the USA; recommended GH dosages range from 0.25 to 0.47 mg/kg/wk.

To examine long-term outcomes in patients (pts) receiving GH treatment in routine clinical practice, we assessed final height (FH) and safety outcomes in pts born SGA by using data collected in a multinational observational study (Genetics and Neuroendocrinology of Short Stature International Study [GeNeSIS]). 

At the time of analysis, 1208 GH-treated pts born SGA (114 due to Russell-Silver syndrome; 111, other known causes; 932, unknown SGA cause) were enrolled in GeNeSIS. FH (defined by one or more of the following: closed epiphyses, height velocity <2 cm/year, last bone age >14 years (y) for girls/>16 y for boys) was available for 203 pts (FHPop1). Sub-analyses were performed for a) 62 pts with baseline age ≥4 and <11 y who had ≥5 y GH treatment (FHPop2) and b) a subset of FHPop2 who had initial GH dose ≥0.2 and <0.3 mg/kg/wk (FHPop3, N=26).

For FHPop1, FHPop2 and FHPop3, respectively, key baseline data (mean±SD) were: age (y), 10.9±3.1, 8.3±1.7, 8.9±1.9; height standard deviation score (SDS), -2.6±0.9, -2.8±0.9, -2.6±0.8; and BMI SDS, -0.9±1.6, -1.3±1.8, -1.5±1.8. Mean initial GH dose was 0.28±0.10 mg/kg/wk, increasing at FH by ≤12% for all FH populations. FH SDS, BMI SDS, age, and GH duration at FH for each population were as follows: FHPop1: -1.5±0.9, -0.3±1.5, 16.1±1.5 y, 5.0±2.9 y; FHPop2: -1.3±0.8, -0.4±1.6, 16.0±1.2 y, 7.5±1.8 y; and FHPop3: -1.1±0.8, -0.8±1.9, 16.0±1.2, 6.8±1.5 y. Height gains from baseline were 1.1±1.0, 1.5±0.8, and 1.4±0.9 SDS respectively.  

Of 1111 GH-treated pts born SGA with ≥1 follow-up visit (3.2±2.2 y of follow-up), ≥1 adverse event was reported for 283 (25%) vs. 5552/20060 (28%) for GeNeSIS overall (all short stature diagnoses). Adverse events reported for ≥1.0% were precocious puberty (3%), headache (2%), hypothyroidism (2%), arthralgia (2%), attention deficit hyperactivity disorder (2%), scoliosis (1%), asthma (1%), and acne (1%). Two deaths were reported [1 associated with displaced ventriculoperitoneal shunt; 1 with MELAS syndrome (1)]; one case of malignancy (b-cell lymphoma) and 4 cases of diabetes (2 type 2, 1 type 1, and 1 unknown type) were reported. In pts with ≥1 follow-up visit and available IGF-I data mean IGF-I SDS was -1.5±1.6 at baseline, 0.8±1.8 at 3 y of follow-up, and was >+2SDS for 21% of pts at 1 or more follow-up visits.

The height gains observed for pts born SGA enrolled in GeNeSIS were similar to those in previous studies using GH doses similar to the European approved dose (2,3); younger age at treatment start was associated with greater height SDS gain. No new safety concerns were identified.

 

Disclosure: CJC: Employee, Eli Lilly & Company, Employee, Eli Lilly & Company. CAQ: Other activities, please specify:, Eli Lilly & Company, Retiree, Eli Lilly & Company. AGZ: Employee, Eli Lilly & Company, Employee, Eli Lilly & Company. JLR: Researcher, Novo Nordisk, Researcher, Versartis, Researcher, Eli Lilly & Company, Advisory Group Member, Novo Nordisk. ES: Collaborator, Eli Lilly & Company. WFB: Retiree, Eli Lilly & Company, Retiree, Eli Lilly & Company. Nothing to Disclose: CLD

18545 21.0000 THR-165 A Final Height and Safety Outcomes in Growth Hormone (GH)-Treated Children Born Small for Gestational Age (SGA): Experience from a Large, Multinational, Prospective Observational Study 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Hwal Rim Jeong*1, Jungsub Lim2, Young Seok Shim3, Hae Sang Lee1 and Jin Soon Hwang4
1Ajou University School of Medicine, Suwon, Korea, Republic of (South), 2Korea Cancer Ctr Hosp, Seoul, Korea, Republic of (South), 3Hallym University Medical Center, College of Medicine, Hallym University, Seoul, Korea, Republic of (South), 4Ajou Univ School of Med, Suwon City, Korea, Republic of (South)

 

Background: Idiopathic short stature (ISS) is defined auxologically by a height below 2 SD score (SDS) without findings of disease as evident by a complete evaluation by a pediatric endocrinologist including stimulated GH levels. GH responses in ISS are highly variable and are dose dependent. It has been recognized that higher doses of GH are required in ISS patients to achieve similar responses and IGF-1 levels to those seen in GHD.

Objective: To investigate response to same dose of growth hormone (GH) in growth hormone deficiency and idiopathic short stature patient

Methods: This study was conducted retrospectively. Subjects diagnosed with ISS and GHD at Ajou University Hospital were included. ISS group (n= 35) and GHD group (n = 66) were treated with same dose of growth hormone (0.23mg/kg/week SC injection, 6 times a week). All children were prepubertal, and treated with growth hormone for 24months. We reviewed their auxological data, laboratory findings, bone age and change of height SDS (Δ Ht SDS) and growth velocity. And clinical factor associated with 1st year Δ Ht SDS was analyzed.

Result: There was no difference in pretreatment Ht SDS, IGF-BP3 SDS, mid parental height, BMI and bone age delay. ISS group presented higher IGF-1 SDS and GH stimulated peak GH than GHD (-0.19 ± 0.92 and 0.22 ± 0.81, 7.05 ± 2.07 and 21.93 ± 10.00 (ng/ml), p < 0.05, respectively). The same dose of GH presented no difference in GV in both group, but significantly higher 1st year Δ Ht SDS in ISS group than GHD group (0.67 ± 0.31 and 0.82 ± 0.32, p < 0.05). Age was found to be negatively associated with 1st year Δ Ht SDS in both groups.

Conclusion: ISS children can achieve optimal response with same dose of GH compared with those of GHD. Early intervention with GH treatment is best help to attain optimal response in both ISS and GHD.

 

Nothing to Disclose: HRJ, JL, YSS, HSL, JSH

19726 22.0000 THR-166 A A 2 Year Single Center Experience with Same Dose of Growth Hormone on Idiopathic Growth Hormone Deficiency and Idiopathic Short Stature 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Mitchell E. Geffner*1, Anders Lindberg2, Nina Camacho-Hubner3 and Michael B Ranke4
1Children's Hospital Los Angeles, The Saban Research Institute, Los Angeles, CA, 2Pfizer Health AB, Sollentuna, Sweden, 3Pfizer Inc, New York, NY, 4Univ Klinikum Tübingen, Tübingen, Germany

 

Background: Children born SGA are thought to have early and/or rapid tempo puberty. The pubertal pattern of children with Silver-Russell syndrome (SRS), a specific cause of SGA, is less well-characterized. 

Objective: To assess pubertal timing and tempo in SGA children. 

Methods/Subjects: Using the KIGS (Pfizer International Growth data base) pharmaco-epidemiological survey, we undertook a comparative study of pubertal timing and tempo in GH-treated children born with SRS, non-specific SGA, and idiopathic short stature (ISS). Using ANOVA, we compared, by gender (M/F), median (n) growth data SDS at birth and 2 yr, target heights, BMI at pubertal onset, and ages (yr) at which various pubertal milestones were achieved and time intervals between them.

Results: Birth weight [SRSM -3.0 (348), SRSF -2.8 (248), SGAM -2.5 (3049), SGAF -2.4 (1958), ISSM -0.8 (1579), ISSF -0.8 (700) (p <0.01)] and length [SRSM -3.1 (245), SRSF -2.9 (195), SGAM -2.6 (2562), SGAF -2.5 (1723), ISSM -0.5 (991), ISSF -0.3 (449) (p<0.01)] followed the pattern of SRS<SGA<ISS, while target height [SRSM -0.8 (338), SRSF -0.5 (229), SGAM -1.3 (2830), SGAF -1.3 (1817), ISSM -2.0 (1534), ISSF -2.1 (689) (p<0.01)] was reversed (SRS>SGA>ISS). Between birth-2 yr, all groups lost weight, albeit less so in SRSM and SRSF. Rank order for BMI at start of puberty was: SRS<SGA<ISS [SRSM -1.1 (58), SRSF -1.8 (51), SGAM -0.6 (406), SGAF -0.9 (346), ISSM -0.4 (261), ISSF -0.7 (188) (p <0.01, except SGAM vs ISSM, p=NS)].

Age of pubertal onset in all 3 groups was normal. Significant differences included: breast TS2 [SRS 10.4 (119), SGA 11.2 (798), ISS 11.9 (363)] (SRS<SGA<ISS; p<0.01); female pubic hair (PH) onset [SRS 10.6 (113), SGA 11.2 (764), ISS 12.4 (271)] (SRS<SGA<ISS; p<0.01); and menarche [SRS 13.0 (69), SGA 13.3 (434), ISS 13.9 (210)] (SRS<SGA<ISS; p<0.01); male PH onset [SRS 11.4 (151), SGA 12.4 (1077), ISS 13.5 (566)] (SRS<SGA<ISS; p<0.01) and testis 4 mL [SRS 12.3 (144), SGA 13.3 (914), ISS 13.9 (542)] (SRS=SGA<ISS; p<0.01). Prepubertal GH treatment duration correlated with earlier onset of puberty only in the SGAF (r=-0.18, p=<0.01) and in SGAM (r=0.10, p<0.05). Ré pubertal progression, rates for girls moving from TS2 to TS4 breasts [SRS 1.9 (50), SGA 2.0 (329), ISS 2.0 (144)] were not different, whereas boys with SRS had significantly faster testicular growth from 4 to 15 mL [SRS 2.6 (47), SGA 2.3 (326), ISS 2.2 (188)] compared to other groups (SRS>SGA=ISS; p<0.05).

Conclusions: In subjects born SGA enrolled in KIGS, timing and tempo of pubertal events in both sexes occurs within normal age ranges, albeit for most parameters SRS<SGA<ISS. Although prepubertal GH treatment duration marginally correlated with earlier onset of puberty only in the SGA groups, there did not appear to be any accelerator effect on pubertal tempo thereafter.   

 

Disclosure: MEG: Principal Investigator, Endo Pharmaceuticals, Principal Investigator, Ipsen, Principal Investigator, Novo Nordisk, Principal Investigator, Versartis, Advisory Group Member, Endo Pharmaceuticals, Advisory Group Member, Ipsen, Advisory Group Member, Pfizer, Inc., Advisory Group Member, Sandoz, Principal Investigator, Genentech, Inc., Principal Investigator, Novo Nordisk, Editor, Up To Date, Editor, McGraw Hill, Principal Investigator, Eli Lilly & Company, Consultant, Daiichi Sankyo. AL: Employee, Pfizer, Inc.. NC: Employee, Pfizer, Inc.. MBR: Advisory Group Member, Pfizer, Inc..

20298 23.0000 THR-167 A Timing and Progression of Puberty in Children Born SGA 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Carole Fournier*, Rene Rizzoli and Patrick Ammann
Geneva University Hospitals and Faculty of Medicine, Geneva, Switzerland

 

In developing countries, chronic malnutrition is a major factor of child mortality, frequently characterized by insufficient dietary protein intakes. Similarly to kwashiorkor, a low protein diet precludes optimal whole body growth and leads to fatty liver in growing rats. To combat this scourge, low-cost and local food strategies should be developed. Spirulina alga (Spi) has great potential as supplementation since it contains 60% protein, has all essential amino acids and easily grows in tropical regions. Its specific impact on bone growth and the related secretion of hepatic hormones have not yet been studied. To answer this question, one-month old female rats were fed an isocaloric diet containing 10% casein (Cas10, N=8), 5% casein (Cas5, N=8) or 5% casein + 5% Spi into dried powder form (Cas5+Spi, N=8) during 8 weeks. Tibia geometry and liver mass were evaluated. Trabecular bone microarchitecture in proximal tibia was analyzed by microCT and BMD by DXA. Bone strength was evaluated by proximal tibia compression test. Hepatic gene expressions of IGF-1, FGF21, Follistatin and Inhibin A were assessed by real-time RT-PCR and hepatic steatosis by Oil-Red O staining and image analysis. Serum IGF-I and FGF21 were measured. Low protein diet decreased tibia diameter (-5%, p<0.05) and length (-2%, p<0.01) vs. Cas10 group. It reduced proximal tibia BMD (-10%, p<0.001) and bone trabecular volume (BV/TV; -41% p<0.01) vs. 10Cas group, resulting in a lower bone strength (Maximal load: -18%, p<0.01). Spi completely prevents these effects and even leads to higher energy values for breaking bone vs. 5Cas and 10Cas groups (+60% and +43% respectively, p<0.001). The Cas5 group had higher liver weight ratio (+18%, p<0.0001), serum FGF21 (+500%, p<0.001) and hepatic mRNA FGF21 levels (+400%, p<0.01) while serum and mRNA IGF-1 levels were lower vs. Cas10 group (-13% and -26%, p<0.05). Spi completely prevents these effects and even leads to lower liver weight ratio vs. Cas10 group (-8%, p<0.05). Spi also specifically increased mRNA levels of follistatin (+60% and +66% vs. Cas10 and Cas5 group respectively; p<0.05) and reduced those of Inhibin A (-45% and -41%; p<0.05), a subunit protein of Activin A, while low protein diet did not affect these two parameters. We demonstrate that Spi is an effective nutrient to prevent alterations of bone growth, liver fat content, FGF21 and IGF-1 hepatic production in rats fed a low protein diet. As compared to casein, Spi has specific beneficial impact on hepatic fat content and gene expression of follistatin which, if secreted as hepatokine, may positively act at the bone level. This study supports the use of Spi in undernourished children.

 

Nothing to Disclose: CF, RR, PA

21819 24.0000 THR-168 A Spirulina Alga Prevents Impairment of Peak Bone Mass Acquisition and Fatty Liver Induced By an Isocaloric Low Protein Diet 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Chantacha Sitticharoon*1, Maynart Sukharomana1, Supawadee Likitmaskul2, Malika Churintaraphan1 and Pailin Maikaew1
1Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand, 2Faculty of Medicine Siriraj Hospital, Mahidol University

 

Central precocious puberty is found to be associated with childhood obesity and was diagnosed increasingly in obese girls. There are many hormones that might be involved in early onset of puberty including kisspeptin, leptin, and adiponectin. This study would like to compare serum high-molecular weight (HMW) adiponectin, total adiponectin, kisspeptin, leptin, and neuropeptide Y (NPY) between central precocious puberty (CPP) and age-matched control girls with or without excluding obese girls. 25 CPP girls and 29 aged-match control girls were recruited.  CPP girls had higher BMI and body weight (p<0.05) than control girls even exclusion of obese girls; the higher BMI and body weight might be caused from higher fat-free mass (p=0.059) rather than fat mass (p=0.181). Serum leptin levels were note different between CPP and non-CPP girls and were positively correlated with obesity-related parameters (R≈0.5-0.8, p<0.001 all) indicating that leptin was closely related to obesity but might not be a major factor determining precocious puberty. Serum HMW adiponectin was significantly higher in CPP girls compared to control girls (p<0.05) but serum total adiponectin was lower in CPP girls than in control girls (p<0.05). Furthermore, serum total adiponectin was strongly and positively correlated with HMW adiponectin in CPP girls (R=0.915, p<00.1) while much less correlated in control girls (R=0.371, p=0.081) suggesting that other forms of adiponectin rather than HMW adiponectin might be higher in control girls. Interestingly, total adiponectin was negatively correlated with obesity parameters and Tanner stage, (R≈-0.3, p<0.05 all) but was positively correlated with QUICKI (R=0.339, p<0.05) in control girls, not in CPP girls. Moreover, HMW adiponectin was positively correlated with Tanner stage (R=0.351, p<0.05) and LH levels (R=0.369, p<0.01). These results indicated that while entering puberty, HMW adiponectin might play essential roles in progression of puberty whereas before entering puberty, other forms of adiponectin (as represented by total adiponectin) was associated with increased insulin sensitivity and decreased obesity. Kisspeptin was lower in CPP girls when compared to control girls and was positively correlated with SBP and DBP in CPP girls, not in control girls (R≈0.4, p<0.05 all). Kisspeptin was positively correlated with estrogen in control girls (R=0.574, p<0.01), not in CPP girls but was negatively correlated with FSH (R=-0.372, p=0.067) and LH (R=-0.363, p=0.070) in CPP girls and, not in control girls indicating that kisspeptin might plays roles in increased blood pressure in girls entering puberty and increased estrogen levels in girls before entering puberty. NPY was not different between CPP and control girls. In conclusion, HMW adiponectin might be used as a predictive factor to determine pubertal progression or diagnosis of CPP.

 

Nothing to Disclose: CS, MS, SL, MC, PM

20759 25.0000 THR-169 A Increased High-Molecular Weight Adiponectin and Obesity, but Decreased Total Adiponectin and Kisspeptin in Central Precocious Puberty Girls Compared to Aged-Match Control Girls 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Ah Reum Kwon*1, Duk Hee Kim2, Ho-Seong Kim3, Jung Min Ahn4 and Hyun-wook Chae4
1Severance Hospital, Seoul, Korea, Republic of (South), 2Sowha Children's Hospital, Seoul, Korea, Republic of (South), 3Severance Children's Hospita, Seoul, Korea, Republic of (South), 4Yonsei University College of Medicine, Seoul, Korea, Republic of (South)

 

Introduction: Kisspeptin is well known gatekeeper of puberty onset to date. However, several neuroendocrine factors are also discovered to be associated with puberty onset, especially neurokinin B and dynorphin A participate in the neuronal network integrating reproduction with kisspeptin. Therefore Kisspeptin, neurokinin B and dynorphin A were called ‘KNDys’ and became an object of investigation. However, the interactions between KNDys and the reproductive axis have not yet been fully explored. We report herein the expression profile of KNDy gene in the rat at different developmental stages. Methods: Spraque-Dawley (SD) strain female rats were used. To analysis expression of KNDys  mRNA, samples were obtained from hypothalamus in female rats at 4 day, 8 day, 14 day, 23 day, 27 day, 34 day, 38 day and 40 day. At the same time, blood samples were collected for analysis serum level of kisspeptin and luteinizing hormone (LH). The expression of KNDys mRNAs were assessed by RT-PCR and the serum levels of kisspeptin and LH were analyzed by ELISA. Results: The expression of Neurokinin B mRNAs in hypothalamus was low in neonate and infant stages and steadily increased prepubertal and pubertal stages (between day 27 and day 40). The expression of KiSS-1 mRNA was also increased as according to developmental stages. However, Neurokinin B mRNAs expression was increased more slowly than KiSS-1 mRNA expression. Meanwhile, the expression of dynorphin A mRNA in hypothalamus was also increased until infantile stage and decreased from infantile to prepubertal stages, but were increased again and maintained high level in pubertal and adult stage. Conclusion: KiSS-1, NKB and DYN gene contributed pubertal onset by interaction with each other. Pubertal onset may be induced by increased KiSS-1 mRNA, NKB mRNA expression and decreased DYN mRNA expression. However, KiSS-1, NKB, DYN mRNA were moderately increased in pubertal and adult stages, these may make estrus cycles.

 

Nothing to Disclose: ARK, DHK, HSK, JMA, HWC

19727 26.0000 THR-170 A The Expression of Kndys mRNA in Rat at Different Developmental Stages 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Eirini Meimaridou*1, Michelle Goldsworthy Goldsworthy2, Roger D Cox3 and Louise A Metherell4
1William Harvey research Institute, Queen Mary University of London, London, United Kingdom, 2Medical Research Council (MRC) Harwell, Oxfordshire, 3MRC Harwell Mammalian Genetics Unit, United Kingdom, 4William Harvey Research Institute, Barts and the London School of Medicine and Dentistry, Queen Mary University of London, London, United Kingdom

 

In humans nicotinamide nucleotide transhydrogenase (NNT) dysfunction leads to an adrenal specific disorder, glucocorticoid deficiency. NNT, a ubiquitous protein of the inner mitochondrial membrane, produces high concentrations of NADPH for detoxification of reactive oxygen species by glutathione and thioredoxin pathways.  Certain sub-strains of C57BL/6J mice contain a spontaneous Nnt mutation (an in-frame 5 exon deletion) and display glucocorticoid deficiency along with glucose intolerance and reduced insulin secretion. A BAC transgenic rescue of this mouse strain results in overexpression of Nnt (1.9 fold over WT). To understand the underlying mechanism(s) behind the glucocorticoid deficiency we performed comprehensive RNAseq on adrenals from WT (C57BL6N), mutant (C57BL6J) and overexpressing (BAC transgenic) Nnt mice.

This revealed differential expression (fold change ≥1.5; p value <0.001) of 500 genes between WT and mutant mouse adrenals. For 80 of these genes the expression levels were reversed/rescued in the BAC. A further 200 changed their expression levels in the same direction for the mutant and BAC vs WT and the remaining 220 were unaltered when comparing WT vs BAC.

No alterations in antioxidant genes of the glutathione and thioredoxin pathways (PRDX3, GPX1, SOD2, TXNRD2, GR); in genes involved in the steroidogenic pathway (CYP11A1, STAR, CYP11B1) or in components of the ACTH signalling pathway, with the exception of MRAP, were noted. These findings are in keeping with the findings of our in vitro studies in NNT-knockdown H295R adrenocortical cells.

Pathway analysis identified significant enrichment of several pathways including those involved in 1) metabolic processes; PPARy, HSL, PCK1, adiponectin, IRS2 and Glut4 were upregulated in both mutant and overexpressing mice, indicating that fine tuning of Nnt expression is required for the regulation of this pathway 2) upregulation of haemoglobins (Hba-a1, Hbb-b1) in mutant mice, perhaps representing a compensatory increase to improve the antioxidant capacity of adrenal cells. These levels were restored to WT levels in the BAC and 3) upregulation of heat shock proteins in mutant mice that were rescued by the BAC transgene suggesting that ROS damage to proteins is activating chaperone pathways. Replacement of Nnt by the BAC transgene reverses this process implying this process involves more than simply the clearance of mutant Nnt protein itself.

Our data suggest that loss of Nnt and the resultant reduction in NADPH production affects a number of gene networks within the adrenal. It reveals the up- or down-regulation of important pathways presumably to combat the sustained adversity due to mitochondrial NADPH restriction and the concomitant increase in reactive oxygen species that this causes. Overexpression of Nnt reverses some of these changes but not all suggesting that for some pathways a delicate redox balance is key.

 

Nothing to Disclose: EM, MGG, RDC, LAM

21976 27.0000 THR-171 A High-Throughput RNA Sequencing of Adrenals from Mice with Knockout or Overexpression of Nicotinamide Nucleotide Transhydrogenase Reveals Novel Pathways Perturbed By Redox Imbalance 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Carlos Manuel Cossio1, Romina Sainz1, Cristina Patricia Nemer2, Claudia Cannizzaro2, Marco A. Rivarola1, Alicia Belgorosky*3 and Nora Isabel Saraco4
1Garrahan Pediatric Hospital, Buenos Aires, Argentina, 2Garrahan Hospital and R Sarda Hospital, Buenos Aires, Argentina, 3Hospital de Pediatría Garrahan, Buenos Aires, Argentina, 4Hospital de Pediatria Garrhan, Buenos Aires, Argentina

 

Aromatase (Aro) is the key enzyme for estrogen biosynthesis from androgens and is encoded by CYP19A1 gene. In human placenta (Pl), Aro is expressed exclusively in the syncytiotrophoblast.

Several studies have described that abnormal fetal development were related to altered intrauterine environment and particularly placenta deregulation.  It has been reported that small newborns as well as large newborns tend to develop prevalence for metabolic syndrome in adult life. Moreover, patients with Aro deficiency develop lipid and carbohydrate metabolic alterations in adulthood. Our group has previously described in Pl a splicing variant of Aro mRNA (Intron9) that would be translated into inactive Aro protein (1).

The aim of this study was to analyze the expression of Aro mRNA variants in Pl from preterm (PT) (<35 weeks) and term LGA compared to term AGA  newborns. We proposed that the expression of Aro mRNA variants might be involved in the regulation of Aro activity and hence estrogen-androgen balance in intrauterine environment associated with fetal development.

Total RNA was isolated from Pl of PT (GA: 30-35, n=4), LGA (GA: 39-41, n=5) and AGA (GA: 37-41, n=17). Aro mRNA variant expression was analyzed by Real-time RT-PCR with specific primers for total (TotAro, Ex2-Ex3), intron 9 (IN9, Ex8-In9) and active (ActAro, Ex9-Ex10) Aro. β-actin (bAc) was used as housekeeping gene. Statistics (ANOVA and Bonferroni test) were performed on ΔCt data.

TotAro/bAc  was significantly higher in PT (12.83 ± 5.83, AU, mean ± SEM),  compare to AGA (2.38 ± 0.59) and LGA (3.00 ± 1.38) , p<0.05. However, ActAro/bAc  and IN9/bAc were significantly higher in PT (16.29 ± 8.05 and 1.95 ± 0.98), compare to AGA (1.25 ± 0.61 and 0.21 ± 0.09) but not compare to LGA (3.66 ± 1.61 and 0.51 ± 0.23).

Analysis of each transcript variant related to total Aro showed that ActAro/TotAro  and IN9/TotAro were significantly higher in PT (1.33 ± 0.26 and 0.14 ± 0.01) and in LGA (1.22 ± 0.20 and 0.17 ± 0.02) compared to AGA (0.33 ± 0.07 and 0.07 ± 0.01), p<0.05.

The high Aro mRNA expression, particularly Active Aro, in preterm placentas agrees with previous reports of increments in maternal salivary estriol and plasma estradiol in preterm parturition suggesting that Pl Aro plays a role modulating Pl estrogen production associated to prematurity.  Not only in PT but also in LGA higher Active Aro related to Total Aro was observed compare with AGA. These results suggest that Pl Aro mRNA variants might play a role regulating Pl Aro activity and hence androgen-estrogen balance in intrauterine environment. 

1)  Sainz R et al.ENDO2011. Endocr Rev 2011 32: P2-177

 

Nothing to Disclose: CMC, RS, CPN, CC, MAR, AB, NIS

19221 28.0000 THR-172 A Aromatase Transcript Variants Expression in Human Placental Tissues from Preterm Deliveries and Term Deliveries of Large for Gestational Age (LGA) and Adequate for Gestational Age (AGA) Newborns 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Manoj Prasad*1, Randy M Whittal2 and Himangshu S Bose1
1Mercer University, Savannah, GA, 2University of Alberta, Canada, AB, Canada

 

Translocase of the inner membrane (TIM) machinery mediates translocation of proteins into inner mitochondrial membrane (IMM) and matrix. CYP11A1, a mitochondrial IMM resident protein catalyzes the conversion of cholesterol to pregnenolone, the first step of steroid hormone biosynthesis. We found that CYP11A1 interacts with inner mitochondrial translocase Tim50, a part of TIM23 complex at the IMM. Tim50 is anchored to the IMM and exposes a large hydrophilic domain in the intermembrane space which possibly helps docking of CYP11A1. The docking with Tim50 provides CYP11A1 a longer stay into the IMM before its disposal into mitochondrial matix. Native-PAGE experiments followed by antibody shift experiments showed that Tim50 and CYP11A1 are the part of same native complex. Results were further conformed by chemical crosslinking and co-immunoprecipitation experiments. Absence of Tim50, ablated the complex formation and altered pregnenolone synthesis. Furthermore, deletion of more than 50 AA from the N-terminus of CYP11A1 halted its interaction with Tim50. So, we suggest that Tim50 interacts with N-terminal domain of CYP11A1at the IMM to facilitate its translocation and activity.

 

Nothing to Disclose: MP, RMW, HSB

19363 29.0000 THR-173 A Interaction of CYP11A1 with Tim50 at the Mitochondrial Inner Membrane Is Essential for Pregnenolone Synthesis 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Galit Pinto1, Moshe Phillip2 and Galia Gat-Yablonski*3
1Tel Aviv University, Tel Aviv, Israel, 2Schneider Children's Medical Center of Israel, Petah Tikva, Israel, 3Schneider Children's Medical Center, Rosh Haayin, Israel

 

Our previous data suggested that the histone deacetylase (HDAC) SIRT1 is involved in mediating the effect of nutrition on growth; therefore, the aim of the present research was to study the mechanism by which additional HDACs may be involved in nutrition-induced linear growth. The in vivo studies were performed in young male Sprague Dawley rats that were either fed ad libitum (AL) or subjected to 10 days of 40% food restriction (RES) and then re-fed (CU). For in vitro studies, Huh7 hepatoma cells were used. Food restriction led to significant reduction in liver weight, concomitant with increased autophagy (i.e. a decrease in the levels of P62 and an increase in the expression level of AMBRA1 and ATG16L2 genes in the RES group). At the same time, we found that the level of HDAC10 was significantly increased. Over expression of HDAC10 in Huh7 hepatoma cells led to reduced cell viability and increased autophagy as shown by increased conversion of LC3-I to LC3-II. An increase in the level of HDAC10 was also obtained when mTOR was inhibited by Rapamycin. siRNA directed against HDAC10 abolished the effect of Rapamycin on cell viability. These results suggest that increased levels of HDAC10 may mediate the effect of malnutrition on growth attenuation and autophagy. Deciphering the role of epigenetic regulation in the nutrition–growth connection may pave the way for the development of new forms of treatment for children with growth disorders

 

Disclosure: MP: Medical Advisory Board Member, Astra Zeneca, Medical Advisory Board Member, Bristol-Myers Squibb, Study Investigator, Bristol-Myers Squibb, Medical Advisory Board Member, Eli Lilly & Company, Study Investigator, Eli Lilly & Company, Speaker, Medtronic Minimed, Medical Advisory Board Member, Medtronic Minimed, Study Investigator, Medtronic Minimed, Speaker, Novo Nordisk, Study Investigator, Novo Nordisk, Study Investigator, Merck & Co., Study Investigator, Roche Diagnostics, Study Investigator, Pfizer, Inc., Study Investigator, Sanofi, Medical Advisory Board Member, Sanofi, Speaker, Sanofi, Consultant, Andromeda Biotech, Board Member, CGM3 Ltd, Board Member, DreaMed-Diabetes Ltd, Chairman, NG Solutions Ltd. Nothing to Disclose: GP, GG

20251 30.0000 THR-174 A Food Restriction Increases Histone Deacetylase 10-Induced Autophagy in the Liver 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Pediatric Endocrinology Thursday, March 5th 3:00:00 PM THR 144-174 5918 1:00:00 PM Pediatric Endocrinology & Growth Disorders Poster


Magdalena Szopa*, Agnieszka ludwig-Galezowska, Julita Machlowska, Jan Skupien, Piotr Radkowski, Tomasz Klupa, Beata kiec-Wilk, Iwona Wybranska, Pawel Wolkow and Maciej Malecki
Ujcm

 

Specific molecular diagnosis of monogenic diabetes mellitus is important for individualized patient care - by understanding the pathophysiology of the disease one can optimize hypoglycemic treatment and define prognosis for the entire family. Next Generation Sequencing-based exome-sequencing (NGS) might provide additional diagnostic potential as it enables simultaneous analysis of multiple genes in a single test.
Our aim was to assess feasibility of NGS usage for detection of mutations in a set of earlier described diabetic genes in a group of patients from the Polish Registry of MODY. We designed a custom Agilent Sure Select exon-capture assay with baits for 13 known MODY genes (GCK, HNF1A, HNF4A, HNF1B, NEUROD1, INS, CEL, PDX1, PAX4, BLK, KLF11, KCNJ11, ABCC8), two genes mutations of which cause lipodystrophy (LMNA, PPARG), mitochondrial genome and 20 neonatal diabetes genes.
A total of 96 DNA samples were tested. Samples were fragmented using Bioruptor (Diagenode), indexed for multiplexing and hybridisied. Sequencing was performed with an Illumina using 75 or 150bp paired end reads.
The analysis was performed for chromosomal genes only. High or medium impact mutations were identified in 74% of the studied patient samples. We identified 16 high impact mutations: 12 of them were located in HNF1A gene, 1 in ABCC8, 1 in GCK, 1 in EIF2AK3 and 1 in LMNA. We identified 107 medium impact mutations. All known mutations in the positive control samples were detected.
Our pilot experiment using NGS for monogenic diabetes screening in the MODY cohort confirmed that its use is feasible in routine genetic testing.

 

Nothing to Disclose: MS, AL, JM, JS, PR, TK, BK, IW, PW, MM

21759 1.0000 THR-586 A Monogenic Diabetes and Targeted Next-Generation Sequencing in the Mody Registry Cohort of Poland 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Anat Jaffe*1, Shmuel Giveon2, Liat Wulffhart3, Bernice Oberman4, Laurence Freedman4, Arnona Ziv4 and Ofra Kalter-Leibovici4
1Hille Yaffe Medical Center, Hadera, Israel, 2Clalit Health Services, Even Yehuda, Israel, 3Gertner Institute for Epidemiology & Health Policy Research, Sheba Medical Center, Tel-Hashomer, Israel, 4Gertner Institute for Epidemiology & Health Policy Research

 

OBJECTIVE

The prevalence of diabetes mellitus in Ethiopian Jewish Immigrants [EJI] was <0.4% on arrival in Israel, while it was 8.3% in new immigrants from the former Soviet Union [FSU]. Both groups immigrated to Israel beginning in the 1980's. We aimed to compare diabetes risk among EJI, non-Ethiopian Jews [NEJ] and Jews from the FSU living in the same geographical area in Israel and assess whether the differences in diabetes incidence are explained by differences in baseline levels of the metabolic syndrome [MetS] components. 

 

RESEARCH DESIGN AND METHODS

Computerized database of the Sharon-Shomron District of Clalit Health Services between 2007 and 2011 was retrieved for 7,325 EJI and age and sex matched controls-NEJ. The controls included 10,374 "born in Israel" NEJ and 1,443 FSU subjects. "Born in Israel" NEJ served as the comparison group. Age and sex-adjusted hazard ratios [HRs] for incident diabetes were calculated in selected age-groups, and further adjusted for the MetS components. The difference in BMI between those diagnosed with diabetes and non-diabetics was determined within each age and ethnic group.

RESULTS

The HR for incident diabetes in FSU participants was not different from the "born in Israel", for all age groups. However, EJI had  greater diabetes risk at a young (<50y) age, HR- 1.8 (95% CI: 1.5, 2.2) and moderate risk at middle age (50 to 59y) HR- 1.3 (95% CI: 1.0, 1.6).  Diabetes risk in the elderly (60y and above) was not significantly higher among EJI HR-0.792 (95% CI: 0.6, 1.1). Adjustments for the MetS components except BMI, in the EJI, did not significantly change the HR estimates. But adjusting for BMI in EJI, increased HR to 2.3 (95% CI: 1.8, 2.8) in the young age group and to 1.4 (95% CI: 1.1, 1.8) in the middle age group. We further compared the BMI increment between non-diabetic and diabetic persons in the young age group and found it lower in EJI compared to NEJ. The BMI of young EJI at diabetes incidence was 25.88 + 4.21 kg/m2, similar to the BMI of non-diabetic NEJ and FSU participants. BMI at diabetes incidence of NEJ and FSU was 30.14+ 5.6 kg/mand 29+5.8 kg/mrespectively.

CONCLUSIONS

Younger Ethiopian immigrants are at greater risk for diabetes as compared to age-matched FSU subjects, and develop diabetes at lower BMI.  

 

 

Nothing to Disclose: AJ, SG, LW, BO, LF, AZ, OK

20887 2.0000 THR-587 A Younger Ethiopian Immigrants Are at Greater Risk for Diabetes As Compared to Age-Matched New Immigrants from the Former Soviet Union 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Luis Hernandez-Vazquez*1, Carmen V Rivera2 and Rafael Trinidad2
1San Juan City Hospital, 2San Juan City Hospital, San Juan, PR

 

The total number of patients with Human Immunodeficiency Virus is 33 million, with 2.7 million new infections in 2007(1). Puerto Rico has an increasing prevalence trend of Diabetes Mellitus of 12.8% in 2010(3). As treatment of HIV continues to develop, and access to therapy improves, the incidence of HIV associated diabetes is bound to grow. This incidence has augmented with cumulative exposure to combination antiretroviral therapy, as showed in several research studies(1). Concurrent use of opiates may alter beta cell function, while heroin addiction is also associated with insulin resistance. We investigate the prevalence of Diabetes Mellitus and its associated risk factors in a determinate HIV positive population.

In this retrospective study, we reviewed the medical records of 146 HIV positive patients that were evaluated at a hospital of the metropolitan area of San Juan during the years of 2007 to 2010, for the concurrent diagnosis of DM.  For the purpose of this study, a subject was considered to be diabetic if the provider documented a diagnosis of DM in the chart, or the subject was receiving a hypoglycemic and/or insulin-sensitizing agent to treat DM. The prevalence of DM was statistically measured and a Logistic Regression with Pearson X² Square and Fisher’s exact test was used to assess the association between DM and variables such as gender, intravenous drug abuse (IVDA), age intervals, BMI, and Highly Active Antiretroviral Treatment in this population.  The prevalence of DM in the studied population was 13.7% (n=20). There were 59%(n=86) males, 43%(n=63) of patients treated with HAART, 46%(n=67) IVDA, the mean age was 47; with 29% older than 50 years old, and 68% of the patients had a BMI of less than 25.  Gender, IVDA, HAART, BMI, and age were not associated as risk factors for the prevalence of DM in the studied population.  Our data revealed a slightly higher prevalence of DM (13.7%) in HIV infected patients, when compared to that of the general population in Puerto Rico (12.8%).  Still our observations may underestimate reality due to limitations of retrospective analysis; including the possibility of missed testing opportunities for diagnosis. Despite previously reported data(1,2), we observed no significant association between DM and gender, opioid abuse, HAART, overweight, or obesity this time.  This raises concern for yet unrecognized risk factors contributing to a higher prevalence of the disease in this population.  Results of our study alert physicians on the importance of DM screening in the HIV positive patient population.

 

Nothing to Disclose: LH, CVR, RT

22034 3.0000 THR-588 A Prevalence of Diabetes Mellitus in Human Immunodeficiency Virus Positive Patients in Puerto Rico 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Eun Byul Kwon*1, Hwal Rim Jeong2, Young Suk Shim3, Hae Sang Lee2, Jungsub Lim4 and Jin Soon Hwang5
1Ajou University Hospital, Suwon, Korea, Republic of (South), 2Ajou University School of Medicine, Suwon, Korea, Republic of (South), 3Hallym University College of Medicine, Seoul, Korea, Republic of (South), 4Korea Cancer Ctr Hosp, Seoul, Korea, Republic of (South), 5Ajou Univ School of Med, Suwon City, Korea, Republic of (South)

 

Purposes : It is well-known fact that a increase of type 2 diabetes in children with diabetes mellitus in Korea. But there is no statistical review about the changes of rate in subtype of diabetes in the last 10 years. Therefore, in this study, we looked for changes in subtypes of diabetes in children and evaluated the characteristics of the each group in the past 20 years since 1994. In addition, we also examined the correlation between the glycated hemoglobin values at the time of diagnosis and lipid profile.

Methods : We divided patients into 2 groups in 190 patients under the age of 20 who first dignosed with diabetes in Ajou university hospital. Group I was the patients diagnosed with diabetes from September 1994 to December 2004 and Group II was the patients diagnosed with diabetes from January 2005 to April 2014. Each groups ware classified as type 1 diabetes, type 2 diabetes, and maturity onset diabetes of the young(MODY). We compared the changes of diabetic subtypes in each groups. MODY subtype included the patients who confirmed by genetic test and the patients who had non-immune type diabetes, good responses to treatment of sulfonylurea, and a family history of diabetes onset before the age of 25 over 3 generations. We used multiple regression analysis about the correlation between the glycated hemoglobin values at the time of diagnosis and lipid profile.

Results : 52 patients belonged to Group I (44 with type 1 diabetes(84.6%), 6 with type 2 diabetes(11.5%), and 2 with MODY(3.8%)). And 138 patients belonged to Group II (91 with type 1 diabetes(65.9%), 33 with type 2 diabetes(23.9%), and 14 with MODY(10.1%)). The change of composition in each groups had a statistical significance (p-value=0.040). There was no significant difference in the epidemiological values as followings; the age at the time of diagnosis, gender, height, weight, and body mass index(BMI). The serum cholesterol level, and the serum low-density lipoprotein(LDL) level had a correlation with the serum glycated hemoglobin level in the correction of age, the standard deviation score of weight, and the standard deviation score of BMI. Glycated hemoglobin had a regression coefficient 2.984 with the level of cholesterol (p-value=0.013) and a regression coefficient 4.391 with the level of LDL.

Conclusions : The proportion of type 2 diabetes has been increased in patients diagnosed with diabetes over the past 10 years, and the cases of clinically suspected with MODY also increase. In addition, we will need more research to blood sugar control in the diabetic patients with dyslipidemia because it is that the increase in the level of glycated hemoglobin at the time of diagosis has correlations with total cholesterol level and LDL level.

 

Nothing to Disclose: EBK, HRJ, YSS, HSL, JL, JSH

19304 4.0000 THR-589 A The Change of Proportion in Type 2 Diabetes Mellitus in Korean Children and Adolescents 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Khaled Abdullah Alswat*1, Turki M. Almalki2, Naif R. Almalki2 and Khalid Balbid3
1Taif University, School of Medicine, Taif, Saudi Arabia, 2Taif University, 3King Abdulaziz Specialist Hospital

 

Background:

Recent cross-sectional studies suggested that the prevalence of T2D in Saudi range from 10-30%.Diabetes knowledge has been shown to improve self-management skills and glycemic control in patients with T2D. There is a lack of studies to assess the level of knowledge among diabetics in Saudi and more specifically in Taif city.  

Methods:

We conducted a cross-sectional study at the King Abdulaziz Specialist Hospital, Taif, Saudi Arabia, Division of Endocrinology. T2D age ≥18 years who had a routine visit to the Endocrine clinic Jun-Oct 2014 were asked to participate. Baseline characteristics and measurement were obtained at the time of visit. Laboratory data was collected from patient’s EMR. We excluded patients with T1D. We used Michigan Brief Diabetes Knowledge Test to assess patient’s knowledge. Those answered ≥65% of the questions considered to have good knowledge about diabetes. The primary goal is to assess the Diabetes knowledge and its impact on diabetes control and complications.

Result:

Total of 200 patients participated, 96 (48%) were male and 104 (52%) were female, mean age 52.62 yrs old (SD 13.7 years), 50% has diabetes >10 yrs and 22.6% has it for 5-10 yrs, mean A1c 9.19%, mean BMI 31.1%, 73.5% were married, 64.5% did high school or less and 35.2% did college degree or higher, 43.5% considered to have low income, 35.2% were on oral medications only, 41.2% were on insulin, 23.5% were on insulin+/-oral, and the mean correctly answered knowledge questions was 47.95%.

25% of the participants think that A1c reflect BG control over the past week, 50% don’t know what is A1c. 29.5% think that diet soda can be used to treat low BG, 36.5% don’t know but 34% don’t think so.

45 patients (22.5%) considered to have good knowledge about diabetes. Compare to those with poor-knowledge, diabetics with good knowledge has mean age of 49.9 yrs vs 53.43 yrs(p .12), 62.2% were male vs 43.9%(p .03), 53.3% vs 29.9% has college degree or higher(p <.05).

81.8% of the diabetics with good knowledge report sedentary lifestyle compare to 82.7% in the poor-knowledge (p .89). 22.86% of the good knowledge diabetics were active smoker compare to 11.3%(p.51).

Diabetics with good knowledge has mean A1c of 7.7 vs 9.6 (p<.05), mean BMI 31.3% vs 31.1%, mean SBP was 138.4 vs 148.4 (p.21), mean resting HR 73.1 vs 73.8 (p.75), mean TC 167.2 vs 176.3 (p.16), mean LDL 108.6 vs 114.2 (p.37), mean HDL 42.7 vs 42.1(p.79), and mean TG 166.7 vs 181 (p.27).

All diabetics with good knowledge aware that poorly controlled diabetes cause retinopathy compare to 69.1%, CHD 84.4% vs 43.4%, and nephropathy 86.7% vs 53.95 (all p<.05) in patients with poor knowledge respectively.

Conclusion:

The majority (77.5%) of the screened T2D patients considered to have poor knowledge about diabetes. Poor knowledge associated with higher A1c, non-significant increase in most of the measured cardiovascular markers, and those are less aware about diabetes related complications.

 

Nothing to Disclose: KAA, TMA, NRA, KB

20629 5.0000 THR-590 A Diabetes Knowledge and Its Impact on Diabetes Control and Complications Awareness 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Joseph Sack*1, Uri Reicher2 and Abraham Goldstein3
1Bar Ilan University, Tel Aviv, Israel, 2Sack and Reicher, 3Gonda Brain Research Centre, Bar Ilan University

 

Background:

Since 1978 we started screening all the newborns in Israel for Congenital Hypothyrodism (CH). We became interested in seasonality of CH in the different populations in Israel, and therefore the comparative study of overall birth seasonality.

Human populations show seasonal patterns of birth rates, which are stable in any particular population, but vary widely across geographical locations. These differences have been previously attributed mainly to climatic factors, yet additional factors have been proposed. 

Now, we report how a population changes considerably its birth seasonality in only 30 years, and synchronising with another population in the same location. This corresponds to changes in socio-economic factors in the population.

Methods: 

Monthly birth rates were extracted from the Israel Central Bureau of Statistics, and corrected for month length and de-trended using a moving average of 12 months. These were compared with climate, Religious calendar, and socio-economic changes.

Results: 

Over the last 40 years the monthly birth fluctuations of the Arab-Muslim and Jewish populations in Israel changed from qualitatively different patterns to similar ones. 

The peak birth rate of the Arab population moved from winter to late summer, to resemble the Jewish pattern, whereas the Jewish seasonality remained relatively stable. 

Between 1971-80, in January there was a deviation of +12% from the monthly average. In September the deviation was 0%. 

Between 1981-90, in January +5%, and in September +5%. 

Between 1991-2000, in January +2%, and in September +8%.

2001-2010 are similar to 1991-2000 – showing the stabilisation of Arab birth seasonality and synchronisation to the Jewish population.

These changes could not be attributed to climate, religious holidays, or marriage seasonality.

Over the same period, in the Arab population the percent of farmers dropped from 18.8% to 3.2% (in the Jewish from 5.3% to 1.4%), the average number of children per household changed from 3.8 to 2.5 (Jewish - 1.6 to 1.3), the average years of education increased from 8.2 to 11.2 (Jewish - 11.2 to 14).

Conclusions: 

We suggest that the synchronisation of Arab birth seasonality, changing in 40 years to resemble the Jewish one, is probably due to the urbanisation of Arab population, as can be seen in changes in education, profession, birth rate, etc. This is in spite of the Arabs being a native population in Israel, while the Jews are mixed immigrants. The original Arab-Muslim birth seasonality was similar to that of Arabs in neighbouring countries. It is only in Israel that the changed occurred - probably due to the influence of the majority Jewish population.

Changes in seasonal patterns have implications for health services and for research proposing season of birth as a risk factor in various disorders.


 

Nothing to Disclose: JS, UR, AG

21683 6.0000 THR-591 A Change and Synchronisation of Birth Seasonality in the Arab-Muslim Population of Israel (1971 - 2010), Affected By Socio-Economic Factors 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Steven W. Taylor*1, Richard E Reitz1, Nigel J. Clarke2 and Michael J. McPhaul3
1Quest Diagnostics Nichols Institute, San Juan Capistrano, CA, 2Quest Diagnostics Nichols Inst, San Juan Capistrano, CA, 3Medical Director, Endocrinology, San Juan Capistrano, CA

 

The incidence of type 2 diabetes mellitus has increased markedly in the United States and worldwide in the past 2 decades. This increase is traceable largely to the dramatic increase in the prevalence of obesity and insulin resistance in developed nations.

Elevated fasting insulin levels have been shown to reflect the presence of insulin resistance and to predict future development of diabetes mellitus. However, the clinical measurement of insulin levels has been largely relegated to the realm of specialized testing employed primarily in research and epidemiological studies examining insulin resistance, prediabetes, and early Type 2 diabetes. Although insulin measurements have a direct relation to more sophisticated assessments of insulin resistance, researchers have not yet defined specific insulin levels that are associated with specific degrees of insulin resistance. This is at least in part to the marked variability among available assays.  C-peptide measurements are less variable than insulin assays, but also suffer from a lack of standardization.

To address these aspects, we developed a multiplexed method to measure insulin and C-peptide using an LC tandem mass spectrometry assay. The assay involves enrichment of the peptides from patient sera using 2 different monoclonal antibodies immobilized on magnetic beads and processing on a robotic liquid handler.  Eluted peptides are directly analyzed by LC-MS/MS.  The assay has a clinical reportable range from 2.5 to 320 µIU/mL for insulin and 0.11 to 27.2 ng/mL for C-peptide.   Intra- and inter-day assay variation is less than 11% for both peptides. Of the 5 insulin analogues commonly prescribed for diabetes treatment, only the recombinant human insulin drug Humalog® (insulin lispro) causes significant interference for the determination of endogenous insulin. There were no observed interferences for C-peptide.

In summary, we have developed a multiplexed method to measure insulin and C-peptide using an LC tandem mass spectrometry assay. This approach offers a highly accurate and reproducible tool to measure insulin and C-peptide. Applying this tool to clinical samples will permit the standardization of both measurements using physicochemical properties of the analytes. This in turn will facilitate comparison of results obtained in different studies and to highly characterized standard reference materials.

 

Disclosure: SWT: Researcher, Quest Diagnostics. RER: Coinvestigator, Quest Diagnostics. NJC: Coinvestigator, Quest Diagnostics. MJM: Principal Investigator, Quest Diagnostics.

21568 7.0000 THR-592 A A High-Throughput Mass Spectrometry Multiplexed Assay to Measure Insulin and C-Peptide 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Frederick G Hamel*1, Matthew Purbaugh2, Robert Heineman3, Robert G. Bennett4, Gerri Siford2 and Cyrus V Desouza5
1Omaha VA Med Ctr, Omaha, NE, 2University of Nebraska Medical Center, Omaha, NE, 3University of Nebraska Medical Center, 4VA Nebraska-Western Iowa Health Care System, Omaha, NE, 5Univ of Nebraska Med Center, Omaha, NE

 

Recent reports indicate that IDE is present in the bloodstream, and at least a portion of it is associated with exosomes.  We hypothesized that obesity and/or type 2 diabetes (T2DM) would alter the insulin degrading activity in plasma as a response to changes in insulin concentrations.  We recruited healthy (BMI<30, no significant disease), obese (BMI>30) and obese diabetic (T2DM; ICD-9 code) patients and obtained fasting blood samples.  Plasma was centrifuged (100,000 x g) to prepare a microvesicular fraction and supernatant.  Insulin degrading activity was assayed by trichloroacetic acid precipitation of 125I-iodoinsulin, and differences analyzed by ANOVA with a Bonferroni posttest. The presence of IDE in the vesicular and soluble fractions was confirmed by Western blot.  There was little activity detected in the soluble fraction. For the microvesicular fraction, there were no significant differences among the 3 groups, although patients with the highest BMI tended to have lower degrading activity. We saw no statistically significant correlations of degrading activity with a number of clinical parameters including: fasting glucose; triglycerides, age, GFR and albumin. We then post hoc separated healthy and obese patients who were significantly dysglycemic (HbA1c of 6.0 to 6.4) into an additional group, and separated diabetic patients who were on insulin therapy.  Although there was still no difference in degrading activity between healthy and obese patients, there was a decrease in the dysglycemic patients and diabetic patients not on insulin were significantly (p<0.05) lower than healthy.  Interestingly, insulin treatment reversed the effect of diabetes and returned the microvesicular degrading activity to normal levels (p<0.01; diabetic vs insulin-treated diabetic).  As T2DM is associated with a chronic inflammatory state, we examined the effect of lipopolysaccharide (LPS) treatment on plasma IDE in mice.  Mice were treated with LPS or vehicle and 24 hours later plasma isolated.  Insulin-degrading activity was 50% less in animals treated with LPS.  One could speculate that inflammation and/or insulin resistance result in a decrease of vesicular IDE activity and that insulin treatment reverses this through its anti-inflammatory properties, or by overcoming insulin resistance. The results indicate that: a) IDE is present in the non-vesicular portion of human plasma, but is inhibited or inactive; b) a portion of enzymatically active IDE is associated with a fraction consistent with exosomes and is inhibited in diabetes; and c) insulin treatment reverses this inhibition.  Altered IDE activity in the blood could affect clearance of insulin and contribute to hyperinsulinemia.

 

Nothing to Disclose: FGH, MP, RH, RGB, GS, CVD

19099 8.0000 THR-593 A Insulin-Degrading Enzyme Activity in Human Plasma; Alteration with Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Lynn Ang*1, Nicole Nevarez2, Mamta Jaiswal1 and Rodica Pop Busui1
1University of Michigan, Ann Arbor, MI, 2University of Michigan Medical School, Ann Arbor, MI

 

Introduction: Cardiovascular autonomic neuropathy (CAN) is a serious diabetes complication, and an independent predictor of mortality. Currently available methods to detect CAN, although reliable and sensitive, are either too expensive, time-consuming or require large sample sizes. Diabetic CAN and sudomotor dysfunction result from damage of the small unmyelinated cholinergic sympathetic C-fibers. Sensitive, specific, simple, non-invasive, and affordable tests to detect the earliest stages of autonomic dysfunction most susceptible to therapeutic interventions are needed. We evaluated the correlation between sudomotor function and measures of cardiovascular autonomic dysfunction in patients with T1DM.

 Methods: Cross-sectional study in 38 subjects with T1DM (mean diabetes duration >5 years) and 10 age-and-sex-matched healthy non-obese control subjects. Sudomotor function was assessed by measuring electrochemical skin conductance (ESC) of hands and feet with the SUDOSCAN (Impeto Medical; Paris, France). CAN was assessed with cardiovascular reflex tests, heart rate variability (HRV) studies and by positron emission tomography (PET) scans with the sympathetic analogue [11C] meta-hydroxy ephedrine (HED), that were analyzed as left ventricle retention index (RI). Correlation between mean hands and feet ESC and the measures of HRV and mean RI were estimated using Pearson correlation.

 Results: At baseline mean age was similar in T1D and control subjects, while resting heart rate, blood glucose and HbA1c were higher in T1D. There were no differences in the mean hands and feet ESC between groups (75± 11 mcS vs. 77 ± 11 mcS; P=0.61 and 74 ± 14 mcS vs. 77 ± 10 mcS; P =0.53, in T1D vs. control). In preliminary analyses, we found a significant correlation between the E/I ratio with mean hands ESC (r=0.37, P=0.02), and with standard deviation of normal R-R interval (SDNN). No correlations were found between feet ESC and any measure of CAN.

Conclusion: These preliminary data suggest that assessment of the ESC with the SUDOSCAN may not be the ideal tool for assessing early CAN in T1D. However, these findings may be partly due to a relative limited number of subjects studied. These studies are ongoing and longitudinally followed for at least 36 months.

 

Disclosure: RPB: Consultant, Astra Zeneca. Nothing to Disclose: LA, NN, MJ

20451 9.0000 THR-594 A Measures of Cardiovascular Autonomic Neuropathy and Sudomotor Dysfunction in Patients with Type 1 Diabetes (T1D) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


Lynn Ang*1, Jeffrey W Innis1, Joanna L. Spencer-Segal2, Eleni V Dimaraki1, Shane Quinonez1 and Elif A Oral1
1University of Michigan, Ann Arbor, MI, 2University of Michigan, Ann Arbor, NY

 

Background: MELAS is a multisystem disorder of mitochondrial dysfunction caused by mitochondrial DNA (mtDNA) mutations, and it is transmitted vertically in a non-Mendelian manner by mitochondrial inheritance. The A→G transition at nucleotide position 3243 is the most frequent mutation and accounts for 80% of the detected mutations in patients with MELAS syndrome. The reported age of onset varies between the ages of 5-15 years. Common clinical features include normal early development, short stature, hearing loss, recurrent headache and seizures, muscle weakness, easy fatigability, episodic vomiting, congestive heart failure, cardiac conduction block, optic atrophy, pigmented retinopathy, diabetes and nephropathy. Multiple other endocrinopathies have also been reported. Diagnosis is based on clinical findings and molecular genetic testing. Pathogenic mutations can be detectable from leukocytes, skin, hair follicles or most reliably, skeletal muscle. We report an unusual case with a nearly homoplasmic large mtDNA deletion in nucleotides 12439-14594 affecting MT-ND5 and MT-ND6.

Case: A 25 year old white female with a history of end stage renal disease status post renal transplant, bilateral sensorineural hearing loss, hypoparathyroidism, hypopituitarism, retinitis pigmentosa, short stature, and diabetes was admitted to our institution in April 2014 for recurrent seizures, stroke-like episodes, delusion and increased lactate (5.5 mmol/L).  MRI showed restricted diffusion involving the corpus callosum and cerebellum compatible with acute infarcts. A stroke workup was negative including vessel imaging. Given suspicion for MELAS, reevaluation of a previously obtained muscle biopsy revealed ragged red fibers and type 2 fiber atrophy. Respiratory chain enzyme analysis showed no deficiency in respiratory chain complexes. Mitochondrial genome sequencing identified the presence of a nearly homoplasmic large mtDNA deletion of 2156 base pairs that spanned from m.12439-14594 affecting MT-ND5 and MT-ND6. Given a low plasma concentration of arginine, L-Arginine, levocarnitine and Coenzyme Q10 were started with subsequent improvement in mental status. The family history was largely negative with the mitochondrial deletion not present in maternal peripheral leukocytes, suggesting a de novo mutation in the proband.

Conclusion: Recognition of the various phenotypic presentations and endocrinopathies of MELAS is crucial for the diagnosis of a young patient with a stroke-like episode. No specific consensus criteria for the treatment of MELAS have been established because of the genetic heterogeneity and variable natural history of this syndrome. Sensorineural hearing loss has been treated with cochlear implant; diabetes with insulin or oral agents; seizures respond to conventional anticonvulsant therapy; L-Arginine shows promise in treating stroke-like episodes.

 

Nothing to Disclose: LA, JWI, JLS, EVD, SQ, EAO

19996 10.0000 THR-595 A Rare Case of Nearly Homoplasmic Large mtDNA Affecting NADH Dehydrogenase Subunit 5 (MT-ND5) and Subunit 6 (MT-ND6) in a Patient with Mitochondrial Encephalopathy, Lactic Acidosis, Stroke- like Episodes (MELAS) and Multiple Endocrine Anomalies 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 586-595 5923 1:00:00 PM Diabetes Genetics & Epidemiology Poster


M. Michael Wolfe*1, Patricia A. Glazebrook2, Milos Tatalovic2 and Michael O. Boylan2
1MetroHealth Medical Center and Case Western Reserve University, Cleveland, OH, 2MetroHealth Medical Center, Cleveland, OH

 

Background: Past studies have indicated that GIP functions as an efficiency factor by enhancing both nutrient uptake and storage. Thus, a reduction in GIP signaling should prevent the development of obesity in mice fed a high-fat diet (HFD). AIM: To develop a safe and effective biological agent with the capacity to neutralize circulating GIP. Methods: A hybridoma library was created by fusing splenic B cells isolated from mice immunized with a GIP fragment to mouse myeloma SP2/0 cells. Supernates from hybridomas were screened for GIP-specific monoclonal antibodies (mAbs) using a specific ELISA. To identify GIP-neutralizing mAbs, positive supernates were analyzed with a cell-based reporter assay. After purifying the mAbs using protein A agarose, their effects on intraperitoneal (i.p.) glucose tolerance, oral glucose tolerance (OGT), and diet-induced weight gain were evaluated. For i.p. glucose tolerance tests (IPGTTs), mAbs (30 mg/kg body weight [BW]) were administered i.p. 60 min before i.p. administration of glucose (18 mmol/kg BW) and human GIP (2.5 nmol/kg BW). For OGT tests, mAbs (30 mg/kg BW) were injected i.p. prior to oral glucose (2 mg/g BW) gavage. PBS or GIP mAbs (10 mg/kg BW) were also injected i.p. 5 times/week to mice fed a HFD, and both body weight and food consumption were monitored weekly. At the end of the 17-week study period, magnetic resonance (MR) was used to assess fat deposition, and blood was obtained for measurements of insulin, leptin, glucagon-like peptide-1 (GLP-1), and lipids. Results: 2000 hybridoma clones were generated, with 21 positive for GIP binding, one of which yielded a mAb that effectively neutralized 1 nM of GIP. Although serum glucose levels were unchanged, GIP mAbs given i.p. nearly abolished the insulin response to GIP in the IPGTT and reduced the insulin response to oral glucose by 70%. After 17 weeks on the HFD, control mice gained 21.5±1.0 g, while mice receiving GIP mAbs gained 10.5±0.5 g, a reduction in weight gained of 46.5% (P=0.00000007). When corrected for BW, no difference in the quantity of food consumed was detected between the 2 groups. In addition, MR demonstrated that mice treated with GIP mAbs had significantly less subcutaneous (P=0.0002), omental (P=0.0005), and hepatic fat (P=0.030) than untreated animals. In response to GIP mAbs, serum insulin and leptin levels decreased significantly, but no significant changes in serum GLP-1 levels were detected. Total cholesterol (TC), LDL, and triglycerides were decreased, while the HDL:TC ratio increased, significantly in the treatment group. Conclusion: Immunoneutralization of GIP in mice using a specific mAb effectively attenuates weight gain in mice fed a HFD while decreasing fat deposition and improving the lipid profile. The results of these studies support the hypothesis that a reduction in GIP signaling might provide a useful method for the treatment and prevention of obesity and related disorders.

 

Nothing to Disclose: MMW, PAG, MT, MOB

19568 1.0000 THR-620 A Immunoneutralization of Glucose-Dependent Insulinotropic Polypeptide (GIP) Attenuates the Development of Obesity in Mice 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Gaetano Santulli*1, Gennaro Pagano2, Celestino Sardu3, Steven Ross Reiken4, Michele Cannone5, Salvatore D Ascia6, Nicola Marziliano7, Bruno Trimarco8, Theresa Ann Guise9, Alain Lacampagne10 and Andrew Marks11
1Columbia University, New York, NY, 2Federico II University, Italy, 3Leiden University Medical Center (LUMC), Netherlands, 4Columbia University Medical Center, 5Tatarella Hospital, 6Città Study Hospital, 7Ospedale Niguarda, 8Federico II University, Naples, Italy, 9Indiana University School of Medicine, Indianapolis, IN, 10INSERM - Montpellier University, France, 11Columbia University Medical Center, NY

 

Type two ryanodine receptor (RyR2) is a Ca2+ release channel on the endoplasmic reticulum (ER) of many types of cells including pancreatic β-cells. However, the functional role of RyR2 in insulin secretion remains controversial and elusive. To determine the mechanistic role of RyR2 channels in glucose homeostasis we took advantage of RyR2 mutations that render the channel “leaky” in humans, triggering a form of exercise-induced sudden cardiac death known as catecholaminergic polymorphic ventricular tachycardia (1). We found that patients with mutant leaky RyR2 have previously unrecognized yet profound glucose intolerance. To determine whether leaky RyR2 channels in these patients cause glucose intolerance we generated knock-in mice harboring the leaky RyR2 mutations found in humans. These mice exhibited glucose intolerance, significant mitochondrial dysmorphology (fragmented cristae, swelling, lamellar degeneration and outer membrane disruption) and dysfunction (reduced glucose-stimulated and glutamine/leucine-stimulated ATP production, augmented reactive oxygen species generation, decreased mtDNA/nDNA and aconitase) in pancreatic beta cells, impaired insulin secretion, and glucose intolerance. Islets of Langerhans isolated (2) from knock-in mice displayed intracellular Ca2+ leak via RyR2, Ca2+ depleted ER, upregulation of common markers of ER stress BiP and spliced XBP1, and increased expression, most likely compensatory, of Kir6.1. Ca2+ imaging assays (3) confirmed that caffeine-induced Ca2+ release via RyR2 led to mitochondrial Ca2+ accumulation. Intriguingly, human islets from diabetic patients (cadaveric donors) exhibited oxidation of RyR2 and depletion of calstabin2 (an established stabilizer of RyR2) from the channel complex, similar to what observed in knock-in murine islets. We have developed a 1,4-benzothiazepine derivative (Rycal S107) which inhibits stress-induced or genetic mutation induced dissociation of calstabins from RyR channels. Crucially, S107 treatment (10 μM, 4 hr) prevented the loss of calstabin2 from RyR2 channels. Moreover, chronic pharmacological inhibition of intracellular Ca2+ leak (S107, 50 mg/kg/d in drinking water, 1 month) improved mitochondrial fitness and normalized insulin secretion and glucose homeostasis in knock-in mice. We also tested S107 in an animal model of type 2 diabetes mellitus, the ob/ob mouse: oral S107 treatment markedly improved glucose tolerance and increased insulin secretion in islets from ob/ob mice. Taken together, our in vivo and ex vivo data establish that leaky RyR2 channels cause ER stress, mitochondrial dysfunction, impaired insulin secretion, and glucose intolerance.

 

Disclosure: AM: Board Member, ARMGO. Nothing to Disclose: GS, GP, CS, SRR, MC, SDA, NM, BT, TAG, AL

20750 2.0000 THR-621 A Intracellular Calcium Leak Via RyR2 Induces ER Stress and Impairs Mitochondrial Fitness in Pancreatic Beta Cells Leading to Glucose Intolerance 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Stanley Hsia1, Monica DesNoyers1, Martin L. Lee1, Candice Goldstein1 and Theodore C Friedman*2
1Charles R. Drew University, 2Charles R. Drew Univeristy of Medicine and Science, Los Angeles, CA

 

Cigarette smoking has been correlated with insulin resistance, but smoking may also lead to weight loss while smoking cessation may lead to weight gain; the exact metabolic changes occurring with smoking or smoking cessation have not been well studied. The address this knowledge gap, we prospectively studied these metabolic changes using hyperinsulinemic euglycemic clamps coupled with stable isotope tracers to measure hepatic glucose output (HGO) and indirect calorimetry to measure substrate utilization, within an innovative study design of enrolling healthy, ½-to-2 pack-per-day smokers into an 8-week smoking cessation program of behavioral counseling plus oral bupropion (Phase I), followed by a 16-week maintenance period without counseling or bupropion wherein subjects either remained abstinent or naturally resumed/increased smoking (Phase II). Cigarettes per day, breath carbon monoxide (CO), urine nicotine metabolites, weight, body composition, fat distribution, free fatty acids, rates of HGO, total and non-oxidative glucose uptake, glucose oxidation, and the respiratory quotient (RQ), adjusted for mean daily caloric intake and expenditure, were measured in 22 subjects before and after Phase I; 19 subjects continued through Phase II and were assessed again at study end. Changes in metabolic measures over time were also correlated to changes in smoking severity within each phase and overall (24-weeks), and adjusted for caloric intake and expenditure. The cessation program (Phase I) reduced the median (interquartile range) cigarettes per day from 8.8 (6.5-12.3) to 1.4 (0.1-3.3), along with reduced CO and nicotine metabolite levels; there were no further significant changes through Phase II. The ratio of central-to-peripheral fat trended higher during Phase I (correlating inversely with CO changes) but then fell significantly through Phase II. Unadjusted basal HGO fell over 24 weeks; changes in weight- and fat mass-adjusted HGO correlated directly with CO changes. Body weight did not change significantly over time, but changes directly correlated with changes in nicotine metabolites in Phase II and overall. Over 24 weeks, changes in CO and/or nicotine metabolite levels correlated inversely with changes in rates of insulin- and weight- or lean mass-adjusted glucose uptake, and basal and insulin-stimulated glucose oxidation and RQ. In summary, smoking cessation over 8 weeks was associated with a transient worsening of central fat distribution, followed by a larger, favorable reversal over subsequent months. Over 24 weeks, HGO improved in relation to lifestyle changes, weight change correlated directly with reduced nicotine metabolites, and reduced CO and/or nicotine metabolites correlated with increased uptake and oxidation of carbohydrate substrates. The metabolic effects of smoking cessation appear to be complex but generally favorable, and should be encouraged.

 

Nothing to Disclose: SH, MD, MLL, CG, TCF

21451 3.0000 THR-622 A Metabolic Effects of Smokers Undergoing Smoking Cessation 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Chunxia Lu* and Vasantha Padmanabhan
Univ of Michigan, Ann Arbor, MI

 

The inappropriate programming of the metabolic system by developmental exposure to excess steroids is of concern, especially in the female. Our previous findings indicate that testosterone (T) excess from days 30-90/ 60-90 of gestation a) induces maternal hyperinsulinemia [1] and culminate in insulin resistance in the female offspring [2] and b) changes mRNA levels of members of insulin signaling pathway in a tissue specific manner consistent with the liver and muscle but not abdominal fat being insulin resistant [3]. Whether similar changes are evident at the functional level and the role androgen and insulin plays in disrupting insulin signaling at the tissue level is unclear. The aim of the present study was to determine whether prenatal T treatment disrupts phosphorylation of insulin signaling molecules in a tissue specific manner and postnatal treatment with androgen antagonist, flutamide or insulin sensitizer, Rosiglitazone would reverse these effects. Four groups of female offspring were studied: control (C), prenatal T (intramuscular injections of 100 mg T propionate twice a week from days 60-90 of gestation that produces T levels in the female fetus comparable to that found in normal male fetus), prenatal T plus postnatal flutamide (15 mg/kg/day; T+F) and prenatal T plus postnatal Rosiglitazone (0.11 mg/kg/day; T+R). Postnatal treaments began at weaning. Muscle, liver, abdominal and subcutaneous fat tissues collected at 3 years of age were pre-incubated with saline at 37 °C for 10 min and then incubated with insulin (100 nM/mL) at 37 °C for 2 and 5 min respectively. Prior to insulin stimulation, prenatal T treatment did not alter phosphorylation of AKT in all 3 tissues. Phospho-AKT/AKT ratio increased 150% in control livers after two minutes of insulin stimulation (p=0.03).  This increment was not evident in livers obtained from prenatal T-treated females.  Similarly, phospho-AKT levels were increased to ~243% (p=0.009) and ~221% (p=0.025) after 2 and 5 min after insulin stimulation in muscle from C females.  This increase was also not evident in the muscle of prenatal T-treated females. Postnatal flutamide or Rosiglitazone treatment failed to restore insulin response in muscle and liver. Phospho-AKT/AKT ratio increased to a similar degree in response to insulin stimulation in abdominal and subcutaneous fat from both C and prenatal T-treated animals.  These findings indicate that insulin resistance induced by prenatal T excess is programmed via AKT signaling pathway in muscle and liver and that postnatal androgen antagonist or insulin sensitizer treatment alone are incapable of restoring insulin sensitivity. Altered insulin sensitivity at the level of liver and muscle may account for the peripheral insulin resistance in prenatal T-treated sheep. These findings may be of translational relevance to human PCOS, the characteristics of whom prenatal T- treated sheep recapitulate.

 

Nothing to Disclose: CL, VP

21087 4.0000 THR-623 A Developmental Programming: Prenatal Testosterone Excess Decreases Insulin Sensitivity in Muscle and Liver but Not Fat Via AKT Signaling Pathway in Female Sheep 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Nanyoung Yoon*, Keith Dadson and Gary Sweeney
York University, Toronto, ON, Canada

 

Decreased adiponectin levels or action have been established to contribute to diabetes and its cardiovascular complications, including heart failure. The endothelium acts as a barrier which regulates the movement of endocrine hormones between the vascular compartment and the interstitial space; a critical yet underestimated event in allowing functional effects in target tissues. Little is known about adiponectin biodistribution in diabetes. Using fluorescent molecular tomography and injection of adiponectin labeled with VivoTag-750 to wildtype and streptozotocin-induced diabetic Nu/Nu mice, we conducted temporal analysis of adiponectin biodistribution. Whole mouse images taken every 10 min from 0-90 min indicated, both visually and quantitatively, elevated tissue adiponectin content in the diabetic mice. This was confirmed by post-mortem ex vivo tissue analysis and was most apparent in liver, kidney and heart. We also performed in vitro studies using monolayers of endothelial cells grown on inserts and found that high glucose levels enhanced paracellular and transcellular flux of adiponectin across this monolayer from apical to basolateral side. Our preliminary data suggests that altered adiponectin flux may be mediated via changes in claudin-7 (a barrier-forming tight junction protein) and claudin-10 (a channel-forming protein) expression. In conclusion, in addition to the well accepted lack of adiponectin contributing to disease pathogenesis in obesity and diabetes we believe that the ability of adiponectin to move from circulation to tissue interstitial space is an important determinant of its physiological actions.

 

Nothing to Disclose: NY, KD, GS

21290 5.0000 THR-624 A Transendothelial Flux of Adiponectin Is Altered in Diabetes: Mechanisms and Physiological Significance 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Chan-Hee Jung*1, Kyujin Kim2, Bo yeon Kim3, Chul-Hee Kim3, Sung-koo Kang3 and Jioh Mok1
1Soonchunhyang University School of Medicine, Bucheon hospital, Bucheon, Korea, Republic of (South), 2Soonchunhyang University School of Medicine, Bucheon, 3Soonchunhyang University Bucheon Hospital, Soonchunhyang University College of Medicine, Bucheon city, Gyeonggi-do, Korea, Republic of (South)

 

It has been shown that omentin is associated with insulin resistance, inflammation and endothelial dysfunction. Visceral adiposity index (VAI) is a recently proposed marker of both visceral fat distribution and dysfunction. Until now, there is no published study to investigate the associations of serum omentin with visceral adiposity index (VAI), cardiac autonomic neuropathy in type 2 diabetes mellitus (T2DM). Therefore, we investigated the relationships between serum omentin, VAI, and vascular complications including cardiac autonomic neuropathy (CAN) in T2DM. We recruited 97 patients (men 56, women 41, mean age: 57.6 years) who evaluated diabetic microvascular complications (nephropathy, retinopathy and peripheral neuropathy) including CAN. CAN was assessed by the five standard cardiovascular reflex tests according to the Ewing’s protocol. CAN was defined as the presence of at least two abnormal tests or an autonomic neuropathy points ≥2.  Serum omentin levels were assessed by ELISA and VAI was calculated by the formula :

(Males:[WC/(39.68+(1.88×BMI))×(TG/1.03)×(1.31/HDL)],Females:[WC/(36.58+(1.89×BMI)×(TG/0.81)×(1.52/HDL)]. We estimated intraabdominal fat area using abdominal dual bioelectrical impedence analysis (BIA). Atherosclerotic burden was evaluated by brachial-ankle pulse wave velocity (baPWV) and ankle brachial index (ABI). Prevalence of CAN was increased borderline significantly across omentin tertiles (p=0.05) and CAN points were significantly increased progressively across omentin tertiles (p=0.01). However, prevalence of diabetic microvascular complications (nephropathy, retinopathy and peripheral neuropathy) was not significantly different according to tertiles of omentin levels. The mean levels of baPWV were significantly increased progressively according to the omentin tertiles (p=0.001). Serum omentin levels showed significant positive correlations with baPWV, CAN points, and HDL-cholesterol, and negative correlations with VAI. The mean levels of serum omentin were not significantly different according to the presence of each micovascuclar complications including CAN. This present study showed that levels of serum omentin are significantly associated with CAN, VAI and arterial stiffness assessed by baPWV, while there are no associations with each microangiopathies in patients with T2DM. Future prospective studies with larger numbers of patients are required to establish a direct relationship between serum omentin levels, VAI and vascular complications in patients with T2DM.

 

Nothing to Disclose: CHJ, KK, BYK, CHK, SKK, JM

19050 6.0000 THR-625 A Associations of Serum Omentin Levels with Cardiac Autonomic Neuropathy and Visceral Adiposity Index in Patients with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Anne E Newell-Fugate*1, Monica Jarboe1, Corinne Bromfield1, Andrea Braundmeier-Fleming2, Sherrie G Clark3, Robert L Rosenfield4, Janice M Bahr1 and Romana A Nowak1
1University of Illinois at Urbana-Champaign, Urbana, IL, 2Southern Illinois University, Springfield, IL, 3Virginia-Maryland Regional College of Veterinary Medicine, Blacksburg, VA, 4The University of Chicago, Chicago, IL

 

Fat location is a linked with hyperandrogenemia in polycystic ovary syndrome (PCOS). Short-term administration of testosterone (T) lowers insulin sensitivity in healthy women, and may contribute to insulin resistance in PCOS, but little is known of this relationship in female adipose tissue. We hypothesized that administration of depot-testosterone (cypionate) (DT) to female pigs would reduce insulin signaling. DT-treated females (n=5) were given intramuscular DT (100 mg/m2) every 6 days, with the first of 3 doses administered on the day of estrus (day 0). Six age-matched, cycling females were controls (C). Fasting blood was collected on days 0, 3, 7, 11 and 13 of the estrous cycle and assayed for serum total testosterone (TT), insulin, and glucose. Liver, skeletal muscle, visceral (V) and subcutaneous (SC) adipose tissues were collected post-mortem (D13-15 of the cycle) for RTPCR of INSR and IRS1 and assessment of protein kinase B (Akt) activity by western blot (n=3) in adipose tissue only. Data were assessed for normality and logarithmically transformed as needed. Serum TT, insulin, and glucose were analyzed by repeated measures ANOVA. Gene transcript levels were analyzed relative to housekeeping gene HRPT1 by ANOVA. Akt activity was analyzed by normalization of phosphorylated Akt (pAkt) and Akt to the loading control (GAPDH), followed by calculation of the pAkt/Akt ratio by ANOVA. TT was higher in DT pigs on days 3, 7, 11 and 13 of the estrous cycle, approaching masculine concentrations on D13 (p<0.0001; DT13: 13.42 ± 0.83 ng/ml; C13: 0.09 ± 0.93 ng/ml). There was no significant difference in serum glucose in both groups. Serum insulin was higher in DT pigs on D3, but lower on D13 (p<0.01; DT3: 16.14 ± 3.12 μU/ml; C3: 3.36 ± 3.49 μU/ml; DT13: 2.50 ± 3.12 μU/ml; C13: 9.83 ± 3.49 μU/ml). INSR and IRS1 transcript levels were up-regulated in SC adipose tissue from DT pigs (p<0.01; INSR-DT: 2.52 ± 0.34 fold change; C: 1.00 ± 0.38 fold change; IRS1-DT: 2.75 ± 0.41 fold change; C: 1.00 ± 0.46 fold change). INSR transcript level was up-regulated in V adipose tissue from DT pigs (p<0.001; DT: 2.83 ± 0.34 fold change; C: 1.00 ± 0.53 fold change). There was no significant difference in the fold change of either INSR or IRS1 in skeletal muscle or liver. There was a trend for decreased Akt activity in V adipose tissue from DT pigs (p=0.1; DT: 1.56 ± 1.19 pAkt/Akt ratio; C: 4.07 ± 1.09 pAkt/Akt ratio) but no difference in Akt activity in SC adipose tissue (p=0.5; DT: 1.73 ± 0.53; C: 1.76 ± 0.49 pAkt/Akt ratio). In conclusion, administration of high dosages of DT to female pigs up-regulates INSR and IRS1 transcript levels in SC adipose tissue and may decrease Akt activity in V adipose tissue. These results suggest that virilizing T levels in females may differentially affect insulin signaling in V and SC adipose tissue depots, with V adipose demonstrating insulin resistance and SC adipose demonstrating increased insulin sensitivity.

 

Nothing to Disclose: AEN, MJ, CB, AB, SGC, RLR, JMB, RAN

21271 7.0000 THR-626 A Virilizing Concentrations of Serum Testosterone in Female Pigs May Differentially Affect Insulin Signaling in Visceral and Subcutaneous Adipose Tissue 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Maria Clarissa Yasmin Ong Tio*, Sarah E. Seiler, Dorothy H. Slentz, Timothy R. Koves and Deborah M. Muoio
Duke Molecular Physiology Institute, Durham, NC

 

Skeletal muscle insulin resistance (IR) plays an important role in the progression of diabetes, as the muscle is the primary site of insulin action and glucose uptake in states of feeding and exercise. Animal models and human studies of diabetes have shown signatures of incomplete ß-oxidation to be associated with the disease. Moreover, mounting evidence suggests that the development of IR is linked to mitochondrial-derived oxidative stress. Glutathione (GSH) is the most abundant endogenous antioxidant that mitigates mitochondrial oxidative stress. Diminution of GSH has been reported in aging, diabetes, and other disease states, whereas restoration of GSH status is associated with improved mitochondrial function and metabolic control. We hypothesized that redox imbalance would lead to intracellular acylcarnitine accumulation and IR symptoms in primary human skeletal myocytes (HSkMC). Cultured HSkMC obtained from young subjects with low BMI were utilized in the experiments. Intracellular glutathione depletion was achieved with buthionine sulfoximine (BSO) treatment, and parameters measured included oxidative stress via 2’,7’-DCFDA (2’,7’-dichlorofluorescein diacetate) fluorescence, 2-deoxyglucose uptake, and insulin signaling via Akt phosphorylation. Acylcarnitine profiling experiments were done using flow injection tandem mass spectrophotometry in the Metabolomics Core of the Duke Molecular Physiology Institute. Our results showed that oxidative stress secondary to GSH depletion exacerbated intracellular accumulation of medium chain acylcarnitines induced by fatty acid treatment. Metabolic profiling pointed to impairments in cellular acylcarnitine efflux as a possible cause of metabolite accumulation. Finally, GSH depletion led to diminished insulin-stimulated phosphorylation of Akt. These findings support a role for oxidative stress in tissue accumulation of acylcarntines and concomitant development of insulin resistance in HSkMC.

 

Nothing to Disclose: MCYOT, SES, DHS, TRK, DMM

21052 8.0000 THR-627 A Glutathione Depletion Alters Acylcarnitine Flux and Insulin Signaling in Primary Human Skeletal Myocytes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Akari Utsunomiya*1, Vladislava Paharkova2, Joe Capri3, Whitelegge Julian4 and Kuk-Wha Lee5
1Mattel Children's Hospital,UCLA, Los Angeles, CA, 2Mattel Children's Hospital, UC, Los Angeles, CA, 3The NPI-Semel Institute, David Geffen School of Medicine,UCLA, Los Angeles, CA, 4The NPI-Semel Institute,David Geffen School of Medicine,UCLA, CA, 5Mattel Children’s Hospital UCLA, David Geffen School of Medicine at UCLA, Los Angeles, CA

 

Humanin (HN) is a putative 24 amino acid polypeptide transcribed from an open reading frame within a region of mtDNA. It has been shown to have cytoprotective properties against Alzheimer’s disease-, myocardial infarction-,and diabetes-associated cellular insults. We have previously reported that HN improves survival of βcells and delays onset of diabetes in the NOD mouse. To understand the potential role of HN inβcells, we investigated binding partners of rat Humanin (rHN) in insulin-producing cells.

All experiments were conducted using rat INS-1 cells in vitro. We performed immunoprecipitation utilizing an anti-rHN antibody to identify potential binding partners of rHN.  MS/MS analysis was performed on the product.  Insulin molecules were identified as binding partners of rHN in INS-1 cells. Additionally, confirmatory immunoprecipitation with anti-rHN antibody was done followed by immunoblot for insulin. We detected bands with molecular weights consistent with insulin molecules,βChain, insulin dimer and insulin hexamer. These results suggest that rat HN directly associates with insulin molecules in insulin-producing cells.  We further performed western blotting in order to examine the glucose-dependent effect of HN expression in INS-1 cells. INS-1 cells were incubated in glucose-containing media at 5, 11, 16, and 32mM glucose in a time course experiment, and cell lysates were subjected to western blot analysis for rHN expression. rHN expression increased in a dose- and time-dependent manner.   These are the first evidence that glycemia regulates rHN expression in insulin-producing cells. 

In summary,we show the rat HN associates directly with insulin molecules and glycemia regulates rat HN expression in insulin- producing cells. Futher experiments will delineate the specific role of rHN in insulin-secreting cells.                                                     

 

Nothing to Disclose: AU, VP, JC, WJ, KWL

20036 9.0000 THR-628 A Glycemia regulates Rat Humanin (rHN) and Rhn associates with insulin molecules in insulin-producing cells 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Isaac Asare-Bediako*, Rebecca Paszkiewicz, Miguel Burch, Richard N Bergman and Stella P Kim
Cedars-Sinai Medical Center

 

Hyperinsulinemia is an important mechanism by which glucose tolerance is maintained during insulin resistance.  It has become increasingly evident that hepatic insulin clearance can be a primary factor in determining insulin levels, making it of upmost importance to have an accurate method by which insulin clearance is assessed.  While indirect methods of measuring liver insulin extraction are available, they are difficult to validate in humans since it is not possible to access the hepatic portal vein. To evaluate indirect methods, we compared 2 indirect protocols based on the hyperinsulinemic euglycemic clamp (EGC) and the frequently sampled intravenous glucose tolerance test-based (FSIGT) with the direct method of measuring hepatic insulin clearance, the paired portal/peripheral insulin infusion (PPII) in the dog model. Protocols: Indirect: 1) metabolic clearance rate (MCR) from the EGC, which is the ratio between insulin infusion rate and plasma insulin levels at steady-state and 2) fractional disappearance rate of insulin (FCR) during a FSIGT, determined from the rate of decline of plasma insulin after intravneous insulin injection during the test.  Direct: A paired portal or peripheral insulin infusion protocol (PPII), in which insulin is infused on separate days at 3 different rates into either the portal (one day) or peripheral vein (other day).   Insulin clearance is given as (1-mpo/mpe), where m is the slope of the best-fit line relating insulin infusion rates to steady-state plasma insulin concentrations for the 2 routes of delivery (1).  Indirect FCR measured from the FSIGT had a very strong correlation with the direct PPII method (r = 0.64).  Surprisingly, MCR during an EGC was not significantly correlated with hepatic extraction (r = 0.045).  Thus, insulin clearance from the FSIGT is an excellent measure of insulin clearance; whereas MCR from the EGC may not be an acceptable surrogate indirect measure of hepatic insulin clearance.

 

Nothing to Disclose: IA, RP, MB, RNB, SPK

19958 10.0000 THR-629 A Insulin Clearance Cannot be Accurately Measured from a Hyperinsulinemic Clamp 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Brian D Piening*, Wenyu Zhou, Kimberly R Kukurba, Gucci Gu, Kevin Contrepois, Colleen Craig, Dalia Perleman, Tracey L McLaughlin and Michael P Snyder
Stanford University, CA

 

Obesity is a worldwide health epidemic and a major cause for cardiovascular disease and type 2 diabetes mellitus (T2DM). Despite this, our ability to identify the subset of overweight individuals that are at an elevated risk for developing T2DM is woefully inadequate. Here, we developed a longitudinal, multi-omic personalized medicine pipeline for the comprehensive molecular profiling of blood-based analytes that differ between healthy overweight and prediabetic individuals both at baseline and in response to a controlled overfeeding perturbation. We sequenced the genomes of a cohort of 30 individuals (15 insulin-resistant (IR) and 15 insulin-sensitive (IS)) and generated a personalized risk profile for each individual based on known and novel variants associated with T2DM. Multi-omic profiling (transcriptome, DNA methylome, proteome, metabolome etc.) revealed significant differences in multiple ‘omes between IR and IS individuals at baseline, implicating pathways related to chronic inflammation and insulin regulation as well as novel connections to the disease. Participants were then placed on a short-term high caloric diet, followed by additional multi-omic profiling. The dietary perturbation was associated with a wealth of biomolecular expression changes concomitant with weight gain and spanning multiple ‘omes, and this response differed between IR and IS individuals. In total, these large-scale longitudinal data offer a novel view of the dysfunction in cellular networks associated with the progression to T2DM and may offer new strategies for predicting and preventing the disease.

 

Nothing to Disclose: BDP, WZ, KRK, GG, KC, CC, DP, TLM, MPS

21927 11.0000 THR-630 A Comprehensive Longitudinal Multi-Omic Profiling Reveals Molecular Signatures Associated with Prediabetes at Steady State and during Weight Gain 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Hamza Amine1, Yacir Benomar2, Arieh Gertler3 and Mohammed Taouis*4
1University Paris-Sud, 2University of Paris-Sud, France, 3Hebrew Univ Inst of Biochemist, Rehovot, Israel, 4University of Paris XI, Chilly Mazarin, France

 

Palmitic acid enhances TLR4 expression and promotes resistin/TLR4 signaling.

Hamza Amine 1,2, Yacir Benomar, PhD 1,2, Arieh Gertler, PhD3 and Mohammed Taouis, PhD 1,2

1Neuroendocrinologie Moléculaire de la Prise Alimentaire, University of Paris-Sud, UMR 8195, Orsay, F-91405, France.

2 Neuroendocrinologie Moléculaire de la Prise Alimentaire, CNRS, Centre de Neurosciences Paris-Sud UMR8195, Orsay, F-91405, France.

3 The Institute of Biochemistry, Food Science, and Nutrition, Faculty of Agricultural, Food and Environmental Quality Sciences, The Hebrew University of Jerusalem, PO Box 12, 76100 Rehovot, Israel.

Toll-like receptor 4 (TLR4) has a critical role in innate immunity, and the activation of inflammatory pathways play an important role in the induction of insulin resistance. Indeed, we have recently demonstrated that TLR4 is implicated in resistin-induced inflammation and insulin resistance in the hypothalamus (1). We have also shown that TLR4 is up-regulated in the hypothalamus of mice fed a high-fat diet.

Here, we aim to decipher the molecular mechanisms implicated in the regulation of TLR4 expression. For this purpose, human neuroblastoma cells (SHSY-5Y) were exposed during 4h to either palmitic acid (a saturated fatty acid) or the omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA). Cells were then treated with resistin.

Firstly we analyzed the effect of resistin, palmitic acid and DHA on inflammation markers. We show that only resistin was able to activate NF-κB and to increase the phosphorylation of Akt and p38 MAPK. However, palmitic acid pretreatment increases the expression of inflammatory cytokines (IL-6 and TNF-α), similar to resistin. Interestingly, DHA pretreatment suppresses palmitic acid and resistin induced up-regulation of IL-6 and TNF-α.

Secondly, we studied the possible synergistic interaction between resistin and palmitic acid on TLR-4 expression. We show that palmitic acid pretreatment increases TLR4 expression, at both protein and mRNA levels, while DHA pretreatment had no effect. Importantly, palmitic acid pretreatment potentiates resistin effects.

In conclusion, we show for the first time, to our knowledge, that palmitic acid induces TLR4 expression and this leads to the amplification resistin effects promoting then insulin resistance at the neuronal level.

(1) Benomar et al., Diabetes. 2013, 62:102-14.

 

Nothing to Disclose: HA, YB, AG, MT

18977 12.0000 THR-631 A Palmitic Acid Enhances TLR4 Expression and Promotes Resistin/TLR4 Signaling 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Greg Arsenis*
VA Medical Center, Seminole, FL

 

   Epidydimal fat cells are very sensitive to pH changes; alkaline pH stimulates, while acidic pH inhibits, both the insulin receptor binding affinity (IRBA) and glucose transport (GT) activity (1,2). Fat cells possess an amiloride-sensitive Na+/H+ exchange mechanism which regulates intracellular pH (pHi) by a Na+-specific and pH-dependent mechanism (3). The Na+/H+ exchanger is very responsive to insulin and isoproterenol (Iso), the main regulators of GT (3, 4). We studied the activity of the Na+/H+ exchanger, a major regulator of pHi, in the absence of HCO3- and in the presence of ouabain, an inhibitor of Na+/K+ ATPase.  Alterations of the pHi were monitored with 2’,7’-bis (carboxyethyl)-5 (6)-carboxy-fluorescein (BCECF) and Na+ transport was monitored by measuring 22Na+ uptake. The results have shown that Iso (300 nM), which is known to inhibit both IRBA and GT by 50% (5), decreases the resting pHi from 7.26 ± 0.05 to 7.03 ± 0.09, p<0.05. Also, Iso completely inhibited the recovery of the acidified pHi and the 22Na+ uptake by 81% by blocking the Na+/H+ exchanger (control: 0.365 ± 0.019, Iso 0.069 ± 0.006 nmoles/105 cells/2.5 min, p<0.0001). Both of these effects are induced via b2-adrenergic receptor stimulation, adenylate cyclase activation, and a cAMP-dependent mechanism (4). Insulin (1 nM), which is known to increase the IRBA and GT (6), raised pHi by 0.1-0.2 units and increased 22Na+ uptake by >15% over control cells by stimulating the Na+/H+ exchanger. Moreover, insulin blocked the inhibitory effect of Iso and alkalinized the Iso-acidified pHi from 7.03 ± 0.09 to 7.18 ± 0.04, p<0.05; an effect mediated via the activation of the Na+/H+ antiport. Also, insulin blocked the inhibitory effect of Iso on 22Na+ uptake and increased Na+ influx by >400% (Iso: 0.059 ± 0.007, insulin+Iso: 0.244 ± 0.014 nmoles/105 cells/2.5 min, p<0.009). Thus, pHi is regulated mainly through the antagonistic actions of insulin and Iso on the Na+/H+ exchanger. The effects of insulin and Iso on pHi and GT occur simultaneously, suggesting the regulatory role of pHi on GT: acidic pHi inhibits while alkaline pHi stimulates glucose transport. Whether the two functions are taking place at the same time in parallel without affecting each other or whether pHi changes affect one of the intermediate steps of the insulin or Iso signaling pathways, still needs to be elucidated.

 

Nothing to Disclose: GA

19366 13.0000 THR-632 A Novel Counter Regulatory Effects of Insulin and Isoproterenol on Na+/H+ Exchange in Rat Adipocytes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Kathryn Hinchee-Rodriguez*, Rekha Kar, Martin L Adamo, Linda J Roman and Bettie Sue Masters
Univ of Texas Hlth Sci Ctr, San Antonio, TX

 

Type 2 Diabetes (T2DM) is the seventh leading cause of death in the United States, and results from reduced insulin sensitivity coupled with a relative failure of insulin secretion. Reduced insulin sensitivity itself has been associated with decreased nitric oxide synthase (NOS) activity and impaired glucose uptake in T2DM human skeletal muscle. Upon insulin stimulation, NO synthesis increases in normal adult skeletal muscle, whereas no such increase is observed in T2DM adults. We have previously shown that neuronal NOS (nNOS) is phosphorylated at Ser1446, resulting in its activation, in response to insulin in mouse skeletal muscle in vitro and in vivo. But it is still unclear which kinase is responsible, and how insulin resistance in skeletal muscle affects nNOS Ser1446 phosphorylation. In C2C12 myotubes, nNOS phosphorylation was dependent on one or more AKT isoforms, as AKTi, which prevents AKT1/2 activation, and the AKT2-specific inhibitor, AKT2i, both inhibited nNOS phosphorylation. As AKT's activity is inhibited in insulin resistance, we utilized a 45% high fat diet (HFD) mouse model of obesity-induced insulin resistance. In this HFD model, basal levels of phospho-nNOS increased compared to those in a control diet. However, insulin failed to stimulate nNOS phosphorylation in skeletal muscle of the HFD mice, whereas insulin robustly increased phospho-nNOS in control diet mice. Taken together, these results suggest that while nNOS may depend on AKT2 for Ser1446 phosphorylation, this phosphorylation is attenuated under insulin resistance in mice. However, basal nNOS phosphorylation overall is increased, suggesting that alternative mechanisms for nNOS phosphorylation exist in mice compared to humans.

 

Nothing to Disclose: KH, RK, MLA, LJR, BSM

22129 14.0000 THR-633 A Neuronal Nitric Oxide Synthase Phosphorylation Under Normal and Insulin-Resistant Conditions in Mouse Skeletal Muscle 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Samim ALI Mondal*1, Manoj Kumar2, Deep Dutta3, Parna Choudhury4, Sasanka Chakrabarti5 and Satinath Mukhopadhyay6
1Institute Of Post Graduate Medical Education & Research,SSKM Hospital, Kolkata, India, 2Institute of Post Graduate Medical Education & Research, SSKM Hospital, Kolkata, 3Post Graduate Institute of Medical Education & Research & Dr. Ram Manohar Lohia (RML Hospital, 4Institute of Post Graduate Medical Education & Research and Seth Sukhlal Karnani Memorial Hospital, Kolkata, 5Institute Of Post Graduate Medical Education & Research,SSKM Hospital, Kolkata, 6IPGME&R, Calcutta

 

Serum free vitamin-D levels may be better predictors of insulin resistance in individuals with prediabetes


Background:

High prevalence of Vitamin-D deficiency (VDD) in some populations have recently been attributed to low vitamin-D binding protein (DBP) levels. Although studies from India report one of the highest prevalence rates of VDD seen anywhere in the World, the possibility of underlying DBP deficiency has never been studied in these populations. The relationships between free, bioavailable and total 25-hydroxy-vitamin-D (25OHD) with metabolic syndrome (MetS)were evaluated across the spectrum of glycemia in 102 treatment naïve subjects with glucose intolerance.

Methods:

Individuals with normoglycemia (NGT, n=34), prediabetes (preDM, n=64)and type-2 diabetes (T2DM, n=38) were selected from the ‘Eastern India vitamin-D in prediabetes study’ (CTRI/2011/091/000192). Serum 25OHD levels were measured using125I radioimmunoassay kit (Cat. no. REF68100E, Diasorin, USA). Serum DBP levels were measured using sandwich ELISA (Cat. no. MBS161429; My Biosource; San Diego, CA, USA), having detection range 5-1000 ng/L,Intra and inter assay coefficient of variation was <10% & <12% .Free, bioavailable, and DBP-bound 25OHD were calculated using equations adapted from those described by Vermeulen and colleagues.Serum insulin was estimated using CLIA (Immunite-1000, Gwynedd, UK). IL6, IL1b, sTNFR1 and sTNFR2 were estimated using sELISA. IR was estimated using HOMA2-IR (homeostatic model of insulin resistance).

Results:

NGT (age: 50.29±13.2 yrs), prediabetes (age: 46.9±12.3 yrs)and T2DM (age: 49.73±13.4 yrs) patients had significant difference in mean serum DBP (276.42±32.69,318.18±62.65 and 279.41±49.01mcg/ml, P=0.001), total 25OHD (30.50±18.37, 31.95±23.25, 42.07±20.23ng/ml, P=0.075), free 25OHD (9.04±5.33, 8.44±5.91 and 13±6.78pg/ml, P=0.005) and bioavailable 25OHD (3.52±2.08, 3.29±2.32, 5.08±2.64ng/ml, P=0.005). Free 25OHD had significant inverse correlation with HOMA2-IR in NGT (r=-0.532, P=0.007) and prediabetes (r=-0.36, P=0.010). DBP had positive correlation with HOMA2IR in prediabetes (r=0.349, P=0.013) only. 25OHD had inverse correlation with HOMA2IR only in NGT (r=-0.468,P=0.021). Regression analysis revealed free 25OHD to be the best predictor of HOMA2IR at baseline(β=-2.024, P=0.007) and after adjusting for IL6, IL1b, sTNFR1 and sTNFR2 (β=-2.620, P=0.003).  Free, total 25OHD and DBP were not predictive of HOMA2IR in T2DM

Conclusion:

Free 25OHD, in contrast to  total 25OHD may be a better predictor of insulin resistance prediabetes among Indians. Further studies are warranted to study the impact of changes in serum free 25OHD following vitamin-D supplementation on glycemic outcomes in prediabetes. In addition, serum DBP levels may predict the severity of insulin resistance in prediabetes.

 

Nothing to Disclose: SAM, MK, DD, PC, SC, SM

21633 15.0000 THR-634 A Serum Free Vitamin-D Levels May be Better Predictors of Insulin Resistance in Individuals with Prediabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Shiliu Tian1, Xiwen Liu2, Yong Wu3, Adaku Ume3, Guadalupe Navarro2, Desean L. Lee4, Michael Mangubat2, Theodore C Friedman3 and Yan Jun Liu*3
1Shanghai University of Sport, Shanghai, China, 2Charles R. Drew Univeristy of Medicine and Science, Los Angeles, CA, 3Charles R. Drew University of Medicine and Science, Los Angeles, CA, 4Charles R Drew University of Medicine and Science, Los Angeles, CA

 

The pre-receptor activation of glucocorticoid (GC) metabolism in adipose tissue by NADPH-dependent 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) has emerged as a potential mechanism in the pathogenesis of visceral obesity and metabolic syndrome. Hexose-6-phosphate dehydrogenase (H6PDH) mediates 11ß-HSD1 amplifying intracellular GC production by driving NADPH exposure to 11ß-HSD1 and requires glucose-6-phosphate transporter (G6PT) to maintain its activity. Here, we examined whether specific modulation of G6PT affects tissue GC action by examining the possible effects of G6PT inhibitor ASO (G6PT ASO) on adipose 11ß-HSD1 and H6PDH in diabetic db/db mice. We observed that specific G6PT ASO treatment of db/db mice markedly reduced adipose G6PT mRNA and protein expression and substantially decreased adipose 11ß-HSD1 and H6PDH levels. Reduction of G6PT expression was correlated with the suppression of adipose G6Pase and AMPK and corresponded to the improvement of hyperglycemia and insulin resistance in db/db mice. These findings suggest that G6PT plays an important role in the modulation of pre-receptor activation of glucocorticoid and provides new insights into the role of G6PT in the development of type 2 diabetes.

 

Nothing to Disclose: ST, XL, YW, AU, GN, DLL, MM, TCF, YJL

19504 16.0000 THR-635 A Reduction of Glucose-6-Phosphate Transporter Expression in Adipose Tissue Ameliorates Glucose Homeostasis and Insulin Resistance in Diabetic Mice 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Thais Bento Bernardes*1, Jéssika Geisebel Neto2, Thaiane Gadioli Gaique2, Renata Frauches Medeiros2, Carmen C Pazos Moura3 and Karen Jesus Oliveira4
1Federal University of Rio de Janeiro, Rio de Janeiro, 2Federal Fluminense University, 3Federal University of Rio de janeiro, Brazil, 4Federal Fluminense University, Niteroi

 

Cinnamon is a spice widely used in cooking and in alternative medicine in many cultures. Studies showed that cinnamon can influence key endocrine-metabolic systems. Furthermore, it is well known that the maternal nutrition during pregnancy and lactation is crucial for the health of their offspring. Thus, we hypothesized that the maternal cinnamon intake during lactation may modify endocrine parameters of the mothers and their offspring. In order to investigate this, one day after birth female Wistar rats received water (Control group-Ct) or aqueous cinnamon extract (Cinnamon group-Cin, 400mg cinnamon/kg body weight/day) by gavage during the entire lactation. After weaning to adulthood (180 days of life, P180) the Control and Cinnamon male offspring received standard diet. Maternal cinnamon intake did not affect body mass gain or food intake of dams or of their offspring. However, cinnamon intake decreased lipid content in the dams carcass (Ct: 7.08 ± 0.6 n=8; Cin: 4.26 ± 0.6% n=10; p=0.007). Moreover, Cinnamon dams showed no differences in serum levels of adiponectin, leptin, insulin, estradiol, but showed higher serum progesterone (Ct: 12.69 ± 2.5 n=8; Cin: 43.44 ± 13.2ng/mL n=9; p=0.04). Cinnamon offspring at P180 exhibited higher lipid content in the carcass (Ct: 14.26 ± 1.0 n=10; Cin: 20.73 ± 1.5% n=10; p=0.002) without differences in serum adiponectin but higher serum levels of leptin (Ct: 11.50 ± 1.4 n=10; Cin: 33.05 ± 5.15ng/mL n=15; p=0.002).  Cin P180 group exhibited higher serum insulin (Ct:1.80 ± 0.3 n=10; Cin: 2.70 ± 0.2ng/mL n=15; p=0.02) without differences in fasting blood glucose. In soleus skeletal muscle the cinnamon offspring at P180 showed lower expression of insulin receptor beta subunit (IRβ) (Ct: 1.0 ± 0.1 n=11; Cin: 0.73 ± 0.04 a.u.; n=16; p=0.02) associated with increased pIRβ (phospho T1162/1163 Ct: 1.0 ± 0.03 n=11; Cin: 1.44 ± 0.05 a.u.; n=16; p<0.0001). In addition, the expression of the protein threonine phosphatases 1 B (PTP1B), which has an essential role inactivating the pIRβ signaling, was lower (Ct: 1.00 ± 0.05 n=9; Cin: 0.79 ± 0.05 a.u.; n=12; p=0.01). The Akt/protein kinase B protein expression was similar among the offspring groups, although the Cin P180 group exhibit higher protein levels of pAKT (phospho S473) (Ct: 1.01 ± 0.07 n=10; Cin: 1.41 ± 0.1  a.u.; n=13; p<0.05). Despite the lower IR protein expression, the insulin actions may not be completely compromised due to the higher pIRβ and one of its target, the pAKT. Therefore, the maternal cinnamon intake during lactation promoted mild hormonal and metabolic changes in dams. Although, their male offspring exhibit negative alterations of endocrine parameters in adulthood and changes in insulin signaling pathway expression pattern in skeletal muscle. Despite the visceral obesity and higher serum insulin in the offspring they probably have compensatory mechanisms which leads to an euglycemia.

 

Nothing to Disclose: TBB, JGN, TGG, RFM, CCP, KJO

22140 17.0000 THR-636 A Maternal Cinnamon Intake during Lactation Leads to Higher Serum Insulin and Leptin Levels of Their Adult Male Offspring in Rats 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Bruna Kelly Sousa Hirata*, Renata Mancini Banin, Ana Paula Segantine Dornellas, Luciana Chagas Caperuto, Lila Missae Oyama, Eliane Beraldi Ribeiro and Monica Marques Telles
Universidade Federal de São Paulo

 

Due to the high incidence and severity of obesity and its related disorders, it is highly desirable to develop new treatment strategies. Ginkgo biloba extract (GbE), one of the most worldwide used plant extracts, has been pointed as an alternative therapy for the treatment of hyperglycemia and insulin resistance.  In this context, it was aimed to evaluate the effect of GbE on insulin signaling in retroperitoneal white adipose tissue depot of diet-induced obese rats. Rats were fed with high fat diet for 2 months and thereafter treated for 14 days with 500mg/kg of GbE. Rats were then euthanized and samples from retroperitoneal fat depot were removed before and 90 seconds after insulin i.v. injection for the measurement of IR and Akt phosphorylation levels by Western blotting. It was also evaluated the gene expression of both adiponectin receptors Adipo-R1 and Adipo-R2 in retroperitoneal fat depot. The treatment with GbE promoted a significant reduction on both food/energy intake (6.3%, p=0.031) and body weight gain (62%, p=0.013). In addition, the treatment significantly increased Adipo R1 gene expression (33%, p=0.013) and the phosphorylation levels of both IR (281%, p=0.004) and Akt (67%, p=0.039) in retroperitoneal fat depot. The data suggest that GbE might have potential as a therapy to treat obesity and its comorbidities, especially for obese subjects resistant to adhere to a nutritional education program. Other studies are needed for better understand the beneficial effect of GbE herein described.

 

Nothing to Disclose: BKSH, RMB, APSD, LCC, LMO, EBR, MMT

21172 18.0000 THR-637 A Ginkgo Biloba Extract Improves Insulin Signaling in Retroperitoneal Adipose Tissue Depot of Obese Rats 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Jennifer B Hao*1, Arthur Schuna2 and Juan Carlos Jaume3
1University of Wisconsin-Madison, Madison, WI, 2University of Wisconsin-Madison, 3University of Toledo, College of Medicine and Life Sciences, Toledo, OH

 

Anti-insulin antibodies have been described long ago in patients with insulin-treated diabetes (before the introduction of human insulin and insulin analogues) and in patients with Insulin Autoimmune Syndrome (Hirata’s Disease), where the presence of antibodies manifests itself as postprandial hyperglycemia (insulin bound to antibody) and fasting hypoglycemia (dissociation of bound insulin from antibodies).

We present the case of a 65 year-old man with type 2 diabetes and extreme insulin resistance.  At the time he was first seen, patient’s insulin regime was 930 units/day with a hemoglobin A1C (HbA1C) of 10.  Because of the presence of many signs of insulin resistance (acanthosis nigricans included), patient was assessed for type B diabetes with antibodies to the insulin receptor.  Antibodies to the insulin receptor were undetectable however, insulin antibody titers were present at 8.7 U/ml (normal <0.4).  In order to control patient’s hyperglycemia and decrease his insulin needs, we attempted to treat him with prednisone 10 mg daily for 1 month.  We noticed a paradoxically modest decrease in insulin requirements with the steroid treatment.  However, because of the hyperglycemic effects of steroids, we decided not to increase the prednisone dose.  Instead, we offered patient treatment with anti-CD20 monoclonal antibody (Rituximab®).  Patient agreed and received two Rituximab infusions 1 month apart.  Patient’s insulin requirements significantly decreased to 220 units/day.  Repeat insulin antibody titers after infusion were 7.6 U/ml (1:2 dilution was 6.5 U/ml).  Subsequent insulin autoantibody titers were somewhat lower at 5.0 U/ml (1:2 dilution was 2.5 U/ml).  Patient’s glucose control was steadily improving with average insulin requirements below 200 units daily (HbA1C of 8.7) for two months.  Ten weeks after the second infusion of rituximab, patient again experienced poor glycemic control.  Patient’s HbA1C went up to 13.1%.  Patient was then started on mycophenolate mofetil.  Repeat anti-insulin antibody titers measured 3U/ml two weeks after and 0.4 U/ml a month after the start of therapy.  Patient also had remarkable glycemic control and more than a 50% decrease in daily insulin requirements.  Five months went by and glycemic control started to deteriorate again.  Hence, a third dose of Rituximab was given.  Repeat insulin antibody titers were low at 0.4 U/ml two weeks after the infusion and total daily insulin requirements decreased to 325 units daily.

This is the first case report in the literature of “extreme” insulin resistance because of insulin antibodies in an adult patient successfully treated with combination immunosuppressive therapy.  The unexpected presence of antibodies against the insulin molecule in a patient with severe insulin resistance and the response to immunosuppressive treatment raises the question of screening for these antibodies in similar patients.

 

Nothing to Disclose: JBH, AS, JCJ

18838 19.0000 THR-638 A Extreme Insulin Resistance from Insulin Antibodies (not Insulin Receptor Antibodies) Successfully Treated with Combination Immunosuppresive Therapy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Cemre Robinson*1, Elaine Cochran2, Phillip Gorden2 and Rebecca J. Brown3
1The Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, 2NIH, Bethesda, MD, 3National Institute of Health, Bethesda, MD

 

Background:  Management of diabetic ketoacidosis (DKA) in extreme insulin resistance (IR) is not well-described in the literature.

Clinical case: The patient is an 18-year old young man with extreme IR due to Rabson-Mendenhall Syndrome (mutation of the insulin receptor). Two weeks prior to a routine visit at the National Institutes of Health (NIH), he underwent a root canal for an abscessed tooth, and did not take prescribed antibiotics. At NIH, labs showed A1c 14%, bicarbonate 26-30, and chronic glucosuria and ketonuria. He continued his home insulin regimen of 1500 units/day of U-500.  Antibiotics were initiated; he was discharged in hemodynamically stable condition.  Two days later, he was admitted to another hospital with DKA, pH 7.08, pCO2 27, bicarbonate 8, and worsened jaw pain.  Insulin drip was started at 100 U/hr, increased on day 1 to 1000 U/hr, and 2000 U/hr on day 2 without resolution of acidosis.  CT showed a dental abscess extending to adjacent soft tissue, the likely trigger for DKA.  Due to lack of improvement despite IV antibiotics, bicarbonate was given, and dental extraction was performed, after which the patient developed septic shock requiring pressor support for 24 hrs.  He improved thereafter, and was transitioned back to subcutaneous insulin. 

Conclusion:  This case demonstrates the complexity of managing DKA in a patient with extreme IR.  Routine diabetes care in extreme IR requires high-dose insulin, typically >200 U/day.  Despite high-dose insulin, good glycemic control is often not achieved.  DKA management in extreme IR is a major challenge due to the unusually high doses of insulin required, raising major safety concerns for providers not experienced with extreme IR.  As seen in this case, insulin as high as 2000 U/hr can be safely administered. At very high insulin doses, the dose-response curve is extremely shallow, requiring aggressive dose increase.  Doubling of the insulin dose every 1-2 hours was recommended in the context of worsening acidosis.  The primary safety concern with high-dose insulin is hypoglycemia; however, patients with extreme IR are at very low risk for this during DKA.  If hypoglycemia occurs, it can be managed with IV dextrose. Due to the rarity of DKA in extreme IR, there are no guidelines regarding other toxicity associated with high-dose insulin use. This case highlights the importance of aggressive management of refractory DKA in extreme IR, including high-dose insulin, fluids, bicarbonate, and treatment of underlying infection, regardless of the risks associated with these therapies.  Ultimately, the key to resolving DKA was treating the underlying infection; insulin alone was not sufficient. In patients with extreme IR, aggressive evaluation and early treatment of infections should be considered to reduce the risk of DKA, as once DKA occurs, management is extremely challenging.

 

Nothing to Disclose: CR, EC, PG, RJB

19014 20.0000 THR-639 A Management of Diabetic Ketoacidosis in Extreme Insulin Resistance 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 620-639 5929 1:00:00 PM Insulin Signaling and Resistance Poster


Kanika Shanker*, Kelly Rouster-Stevens, Tanicia Cheri Daley Jean-Pierre and Jee-Young Nina Ham
Emory University, Atlanta, GA

 

Autoimmune Hypoglycemia in a young girl with Autoimmune Overlap Syndrome

 

Kanika Shanker, M.D.1, Kelly Rouster-Stevens, M.D. 1, Tanicia Daley, M.D. 1,J Nina Ham, M.D.1

 1Emory School of Medicine, Emory University, Atlanta, GA 30322

Background: Evaluation of hypoglycemia in a patient with type 1 diabetes mellitus is challenging. Type B insulin resistance syndrome is a rare heterogeneous metabolic disease, described in middle aged women, associated with autoimmune disorders, characterized by abnormalities in glucose homeostasis, with poor response to treatment, high relapse and mortality rate.

Clinical case: A 8 year-old Hispanic female with a known diagnosis of dermatomyositis was diagnosed with antibody positive type 1 diabetes mellitus. Three months later, she presented with progressive severe hypoglycemia, and she continued to have post-prandial hyperglycemia despite withdrawal of insulin therapy. She later developed autoimmune overlap syndrome constituted by dermatomyositis, systemic lupus erythematosus with class 5 lupus nephritis, C4 deficiency and Raynaud’s phenomenon. On physical examination, she had thin body habitus (BMI-15Kg/m2), severe acanthosis nigricans in multiple skin folds, hypertrichosis of upper body and lower extremity edema. Following five hours of diagnostic fasting, her blood sugar dropped from 81mg/dl to 46mg/dl with a positive glycemic response to glucagon challenge (delta of 56mg/dl). Further laboratory evaluation showed paradoxical hyper-adiponectinemia, normal triglyceride levels inappropriately elevated insulin level (937.6uIU/ml, normal: 1.7-55.9) and C-peptide (7.1ng/ml, normal: 0.8-3.5). With signs of insulin resistance, hyperandrogenism, simultaneous hypoglycemia and post prandial hyperglycemia, we hypothesized occurrence of type B insulin resistance due to insulin receptor auto autoantibodies. Laboratory evaluation confirmed the presence of a high titer of insulin receptor auto antibodies by western blot.

For the prevention of severe hypoglycemia, she was managed initially by frequent feeding followed by overnight G-tube feeds. She received cycles of rituximab, IVIG, plasmapheresis, and bortezomib. Episodes of hypoglycemia decreased but continued highlighting poor response to B cell and plasma cell immunomodulation. We present a unique case of autoimmune hypoglycemia, type insulin resistance given the young age at presentation, presence of autoimmune overlap syndrome and treatment with bortezomib, a novel therapy targeting proteasomes causing apoptosis of monoclonal plasma cells.

Conclusions: Presence of hypoglycemia in the setting of multiple rheumatological disorders should prompt consideration of autoimmune-mediated hypoglycemia. More studies are needed to understand mechanisms of simultaneous occurrence of hypoglycemia and hyperglycemia.

Nothing to Disclose: KS, KRS,TCDJP, JNH

 

Nothing to Disclose: KS, KR, TCD, JYNH

18749 1.0000 THR-596 A Autoimmune Hypoglycemia in a Young Girl with Autoimmune Overlap Syndrome 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Undine Schubert*, Susann Lehmann, Henning Morawietz, Stefan Richard Bornstein and Barbara Ludwig
University Hospital Carl Gustav Carus, Dresden, Germany

 

A major limiting factor for long-term survival and function of islet transplants is the inappropriate microenvironment after intraportal transplantation. We aim to evaluate the microenvironment of the adrenal as a potentially beneficial transplantation site that promotes beta cell engraftment, survival, proliferation and long-term function.

For in vitro analysis of reactive oxygen species (ROS) a co-culture system of adrenal cells and islets was established. Basal ROS levels were similar in islets cultured alone compared to islet-adrenal cell culture. However, treatment with rotenone induced significantly higher ROS levels in the co-culture group compared to control. A high response in ROS production following rotenone treatment has been shown to be indicative for mitochondrial integrity.

For in vivo studies, Streptozotocin induced diabetic NuNu-mice were used as islet recipients. For islet transplantation, the adrenal was exteriorized via retroperitoneal incision and 300 islets were injected through the upper pole of the gland or the kidney. Animals showed a fast decrease in blood glucose levels within the first days after transplantation in both groups, at around 10 days the curves between adrenal and kidney site drifted apart in favor of the adrenal site. Regardless of the transplantation site, islets showed a well preserved morphology and intense insulin staining. The intra-adrenally engrafted islets show higher vascularization compared to the kidney capsule control.

The preliminary work underlined the feasibility of islet transplantation into the adrenal with first promising results on the restoration of normoglycemia. The results achieved could prove the beneficial effect of the adrenal microenvironment on islet engraftment and function in vitro and in vivo and elucidate the underlying mechanisms in regards to promoting islet revascularization and protection from oxidative stress.

This novel concept might allow reducing the islet mass that is currently needed to reverse diabetes.

 

Nothing to Disclose: US, SL, HM, SRB, BL

20856 2.0000 THR-597 A Engraftment, Function and Beta Cell Regulation in the Adrenal Transplant Model 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Husam Ghanim1, Manav Batra2, Nitesh D Kuhadiya2, Sanaa Abuaysheh3, Kelly Green4, Antoine Makdissi1, Ajay Chaudhuri5 and Paresh Dandona*5
1State University of New York at Buffalo, Buffalo, NY, 2University at Buffalo, Buffalo, NY, 3Suny at Buffalo, 4SUNY at Buffalo, 5Diabetes and Endocrinology Center of Western New York, Buffalo, NY

 

Glucocorticoids have been used as the major class of anti-inflammatory agents for over 60 years. However, they have well known side effects including weight gain, adiposity, and a series of catabolic effects with loss of musculature osteoporosis and diabetogenicity. We have recently described the induction of expression of specific pro-inflammatory genes including HMG-B1, TLRs, LIGHT, LTBR and MMP-9 by a single large dose of hydrocortisone (HC), In addition, these subjects experienced a significant increase in plasma glucose and FFA concentrations. Since our group discovered the anti-inflammatory actions of insulin in 2001, we have now hypothesized that a combination of insulin with a high dose of a glucocorticoid may neutralize the proinflammatory effects of HC and offer an ideal and safe anti-inflammatory combination. Healthy subjects were randomized into a cross over study to be injected in two separate days (a week apart) with 300 mg of hydrocortisone (=60 mg prednisolone) intravenously with or without an infusion of 2U/h of insulin with 5% dextrose for 8 hours. HC injection alone induced a significant anti-inflammatory effect including suppression of CCR-2 (by 69±6%), IL-4 (by 52±7%) and TNF-α (by 55±8%) expression in mononuclear cells (MNC) and plasma MCP-1 (by 64±9%) and TNF-α (by 32±8%) concentrations. However, the HC injection also induced an increase in the MNC expression of HMG-B1, TLR2, TLR9 and LIGHT (by 102±13%, 67±11%, 84±7% and 126±14% over the baseline, respectively, P<0.05) and in plasma levels of MMP-9, LIGHT and TGF-b1 (by 186±15%, 94±9% and 167±17% over the baseline, respectively, P<0.05). Plasma glucose and FFA concentrations also increased significantly following the HC injection alone. Insulin infusion along with the HC injection increased the magnitude of the overall anti-inflammatory effect and completely inhibited the increase in glucose and FFA. In addition, the infusion of insulin with HC injection inhibited the HC induced increase in TLR-2 and TLR-9 expression and significantly reduced the HC induced increases in the MNC expression of HMG-B1 and LIGHT (to only 21±11% and 34±14% over the baseline, respectively, NS) and in plasma LIGHT and TGF-b1 concentrations (to only 21±12% and 12±10% over the baseline, respectively, NS) but not in plasma MMP-9 levels. Thus, the infusion of insulin at a low dose with a high dose of HC or other corticosteroids may constitute an ideal anti-inflammatory cocktail in the in- patient setting.

 

Nothing to Disclose: HG, MB, NDK, SA, KG, AM, AC, PD

21040 3.0000 THR-598 A Insulin and Hydrocortisone Combination As an Anti-Inflammatory Cocktail 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Munmun Chattopadhyay* and Vikram S Thakur
Texas Tech University Health Sciences Center, EL PASO, TX

 

Peripheral sensory neuropathy is one of the most common complications of diabetes. Accumulating evidence suggests that chronic low-grade inflammation involved in the pathogenesis of the disease. We hypothesize that continuous release of inflammatory mediators in the peripheral nervous system causes the sensory neuropathy in diabetic animals; therefore blocking this increase will prevent or delay the development of neuropathy. High mobility group box 1 (HMGB1), a nuclear protein released by injured and severely stressed cells, promotes cytokine release via its interaction with the Toll-like receptor (TLR). In this study we investigated the changes in cytokine/chemokine profiles in the dorsal root ganglia (DRG) and compared the changes in behavior with treatment with TLR4 inhibitor in Type 1 diabetic (T1D) model of pain. At 6 weeks after hyperglycemia, T1D rats demonstrated significant changes in thermal hyperalgesia manifested by a decrease in withdrawal latency to heat, mechanical hyperalgesia measured by the Randall Sellito method of paw pressure. T1D rats exhibited marked increases in IL1β and TLR4 at 4 weeks after diabetes as determined by the Western blot analysis and increases in the levels of TNFα, pp38, HMGB1, RAGE by 6 weeks after diabetes. To determine whether increased TLR4 level is responsible for the painful neuropathy in diabetic animals, we injected TLR4 antagonist TAK242 for 3 days at a dose of 1.5 mg/kg per day I.P. in STZ-diabetic animals, at 4 weeks after diabetes. We tested the pain behaviors in these animals one day before and one day after the injection. The behavior testing shows that animals treated with TAK242 had significant decrease in mechanical hyperalgesia. This preliminary study suggests that TLR4 plays an important role in the inflammatory aspect of the painful neuropathy in T1D animals. Understanding the mechanisms of diabetic neuropathy may provide a novel treatment approach for this difficult-to-treat complication of diabetes.

 

Nothing to Disclose: MC, VST

22179 4.0000 THR-599 A Alterations of Inflammatory Mediators in DRG of Type 1 Diabetic Animals with Neuropathy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Alice Liu*1, Cindy A Lamendola1, Sun H Kim1, Danit Ariel1, Fahim Abbasi1, James Cardell1, Shiming Xu1, Shailja Patel1, Vanessa Tomasso1, Hafasa Mojaddidi1, Kaylene Grove1, Philip S Tsao2, Clete A Kushida3 and Gerald M Reaven1
1Stanford University School of Medicine, CA, 2VA Palo Alto Health Care System, 3Stanford Sleep Medicine Center

 

Obstructive sleep apnea (OSA) is frequently associated with insulin resistance, type 2 diabetes (T2DM), and cardiovascular disease (CVD), yet their pathophysiological relationships have not been fully elucidated. Traditional investigations have focused on potential mechanisms by which sleep-disordered breathing may promote insulin resistance. However, there is also evidence that fasting hyperinsulinemia predicts the development of observed apneas, suggesting that insulin resistance per se may play a role in the pathogenesis of OSA. The study aim was to evaluate whether enhancing insulin sensitivity in individuals with untreated OSA would improve sleep measures, as well as decrease risk factors for T2DM and CVD.

Insulin-resistant, nondiabetic, overweight/obese adults with untreated OSA were randomized (2:1) to pioglitazone (n=30) or placebo (n=15) for 8 weeks in this single-blind study. Insulin resistance was directly quantified by the insulin suppression test. Insulin secretory function was measured by the graded glucose infusion test. Change in the apnea-hypopnea index (AHI) by overnight polysomnography was the primary outcome measure. Fasting lipid/lipoprotein concentrations and hsCRP were measured. Peri-umbilical subcutaneous adipose tissue (SAT) biopsies were performed.

Whole-body insulin sensitivity improved 31% in the pioglitazone-treated group. However, pioglitazone therapy did not alter quantitative (i.e. AHI, minimum or mean oxygen saturation, oxygen desaturation index) or qualititative (i.e. Functional Outcomes of Sleep Questionnaire, Beck Depression Inventory) measurements related to OSA symptomatology. On the other hand, pioglitazone treatment led to decreases in fasting plasma glucose (P<0.01), reduction in the integrated glucose-stimulated insulin secretion rate (GS-ISR) by 20% (P<0.001), and increase in the metabolic clearance rate of insulin by 22% (P<0.001). Insulin-mediated suppression of isoproterenol-stimulated lipolysis in SAT adipocytes improved by 1.5-fold in response to pioglitazone (P<0.05). Decreases in hsCRP, triglycerides, VLDL, LDL4, ApoB, and increases in HDL cholesterol and its subgroups (HDL2, HDL3) and ApoA1 (P<0.05) were also observed with pioglitazone.

While enhanced insulin sensitivity via pioglitazone therapy in insulin-resistant patients with untreated OSA did not ameliorate OSA severity or symptoms, risk for T2DM and CVD was substantially reduced as manifested by a right shift in the GS-ISR dose response curve, increased adipose tissue sensitivity to insulin, and mitigation of inflammation and an atherogenic lipoprotein profile.

 

Nothing to Disclose: AL, CAL, SHK, DA, FA, JC, SX, SP, VT, HM, KG, PST, CAK, GMR

19228 5.0000 THR-600 A Effect of Pioglitazone on Obstructive Sleep Apnea, Insulin Sensitivity, and Cardiovascular Disease Risk Markers: A Randomized Placebo-Controlled Study 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Peng Chin Kek*1, Hong Chang Tan2, Yst Kee2, Su-Yen Goh1 and Yong Mong Bee3
1Singapore General Hosp, Singapore, Singapore, 2Singapore General Hospital, Singapore, 3Singapore General Hospital, Singapore, Singapore

 

New onset diabetes after transplantation (NODAT) is a serious metabolic complication of kidney transplantation that predisposes patients to graft dysfunction and death. The aim of this study was to determine the effect of degree of glycemia, as measured by HbA1c, at diagnosis of NODAT on long term graft and patient survival after kidney transplantation in our centre.  This was a single centre retrospective review of solitary kidney recipients transplanted in our centre between January 1998 and December 2007. Of a total of 432 patients, 388 were eligible for the study. Reasons for exclusion were pre-existing diabetes mellitus (n=16), death (n=3) and graft loss (n=10) within the first 2 weeks after transplantation, and follow-up duration of less than 6 months (n=15). Data were collected from electronic patient records and transplant charts. NODAT was diagnosed in patients who had at least two abnormal glucose concentrations (i.e. fasting plasma glucose ≥7.0 mmol/l or random plasma glucose ≥11.1 mmol/l) after the first two weeks post-transplant. NODAT were reported in 94 of the 388 recipients (24.2%). HbA1c on diagnosis of NODAT were not available in 33 subjects and they were excluded from analysis. Higher glycemic exposure was defined as HbA1c ≥8.0%. Mean age at transplant was 46.9 ± 7.7 years and 52.5% were female. The median follow up time was 48.2 ± 39.5 months. Mean HbA1c at diagnosis was 9.0 ± 3.1% and mean glucose level at diagnosis was 20.9 ± 15.1 mmol/l.  Median time to onset of NODAT was 7.4 months and 50.9% developed within the first 6 months. Thirty-three (54.1%) had HbA1c ≥8.0%. Subjects with HbA1c ≥8.0% had poorer long term survival (7 deaths) using Kaplan-Meier analysis (p=0.041) compared to those with HbA1c <8.0% (1 death). Overall patient survival was 97.0%, 78.2% and 54.1% at 1-, 5-, 10-year post-transplant for those with HbA1c ≥8.0% and 100.0%, 96.2% and 96.2% for those with HbA1c <8.0% at 1-, 5-, 10-year post-transplant. No difference in graft survival was noted in the 2 groups. Patients with NODAT had poorer outcomes in both graft and patient survival. Our data suggest that the degree of glycemic exposure at the onset of diabetes, as measured by HbA1c level, might be associated with a poorer long term survival. It is thus important to closely monitor glucose levels early post-transplantation so that high glycemic exposure can be avoided to improve the long term outcomes.

 

Nothing to Disclose: PCK, HCT, YK, SYG, YMB

20333 6.0000 THR-601 A Higher Glycemic Exposure at Diagnosis of New Onset Diabetes after Kidney Transplant May be Associated with Reduced Patient Survival 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Tatiana Gandrabura*1, Shant Manoushagian2, Jeydith Gutierrez-Perez2, David Berman2 and Daniel J Rubin1
1Temple University Hospital, Philadelphia, PA, 2Temple University Hospital

 

Background: Diabetic myonecrosis, also known as diabetic muscle infarction, is a rare condition that usually occurs in patients with long-standing and uncontrolled diabetes. The exact pathogenesis is unknown but may involve hypoxia-reperfusion injury, atherosclerotic occlusion, or vasculitis with thrombosis. The disease is self-limited, and most patients recover in several weeks with supportive treatment. Most often it affects the thigh muscles, although other areas are occasionally involved. Diabetic myonecrosis can be difficult to diagnose as a similar presentation may also be seen in soft tissue abscess, necrotizing fasciitis, deep vein thrombosis, polymyositis, and acute compartment syndrome. Although biopsy has been done in many cases for diagnosis, some authors have argued against biopsy because of potential complications and delayed recovery.

Clinical Case: We present the case of a 64-year-old male with type 1 diabetes mellitus who presented with severe left anterior thigh pain and leukocytosis, bandemia, tachypnea and tachycardia, but no fever. On examination, the left thigh was edematous, warm, and tender, without erythema or subcutaenous emphysema. The patient was in rapid atrial fibrillation and was admitted to the cardiac intensive care unit for heart rate control and vasopressor support. Broad-spectrum antibiotics were initiated. Venous and arterial duplex ultrasounds of the legs were negative for a blood clot, and initial blood cultures were negative. Creatine kinase was moderately elevated at 1,187 U/L (NL 30-390 U/L). A CT scan of the left leg demonstrated thickening and loss of the normal pattern of the left vastus lateralis muscle, which suggested a differential diagnosis of diabetic myonecrosis or infectious myositis. MRI showed enlargement and loss of the normal fibrillary architecture of the quadriceps with diffusely increased T2 signal, which favored diabetic myonecrosis. Subcutaneous air was absent on imaging. A muscle biopsy was surgically obtained to confirm the diagnosis. The pathology results were consistent with a combination of necrotizing fasciitis and myonecrosis. The patient’s hospital course was complicated by an iatrogenic wound infection and bacteremia with Methicillin-resistant Staphylococcus aureus, which required multiple surgical debridements.

Conclusion: Most cases of diabetic myonecrosis can be diagnosed by the clinical presentation and imaging. MRI is 90% sensitive for the condition. Although biopsy remains the gold standard for diagnosis, it generally should be avoided as biopsy can lead to iatrogenic complications. Biopsy should be reserved for atypical presentations.

 

Nothing to Disclose: TG, SM, JG, DB, DJR

21429 7.0000 THR-602 A Diabetic Myonecrosis: Is Biopsy Needed for Diagnosis? 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Faraj Kargoli1, Fady Hannah-Shmouni*2 and Tom Nickolas3
1Montefiore Medical Center, Bronx, NY, 2Yale-New Haven Hospital, New Haven, CT, 3Columbia University, New York, NY

 

Increased Fracture Risk in Co-Prevalent Diabetes and Chronic Kidney Disease in the United States

Background: Over the past twenty years, fracture rates have doubled in patients with chronic kidney disease (CKD) (1). Etiologic mechanisms driving increased fracture rates are not clear. Diabetes mellitus (DM), most common cause of CKD in the United States (U.S.), has reached epidemic proportions, and both DM and CKD increase fracture risk compared to the general population (1). Whether co-prevalent CKD and DM (CKD-DM) is associated with higher fracture rates than CKD or DM alone and is partly responsible for increasing fracture rates observed in CKD populations is not known. We used a large nationally representative sample of the U.S. non-institutionalized civilian population to test the hypothesis that CKD-DM is associated with higher fracture prevalence rates than either disease alone.

Methods: We conducted a cross-sectional analysis of the continuous National Health and Nutrition Examination Survey (NHANES) of subjects ≥40 years old enrolled from 2000 to 2010 (n=26,070, weighted sample= 22,088,010). DM status and fracture status were ascertained from responses to NHANES questionnaires; CKD status was based on an estimated glomerular filtration rate (eGFR) <60 ml/min. The cohort was divided three groups: DM, CKD and CKD-DM. Uni-and multivariate logistic regression models were used to determine associations.

Results: In the DM group (n=2048): age 60±14, female 48% [25% black], eGFR 98±32 ml/min, body mass index (BMI) 32±7; CKD group (n= 1239): age 74±11, female 54% [12% black], eGFR 47±12, BMI 28±5); CKD-DM group (n=529): age 71±10, female 54% [22% black], eGFR 44±14, BMI 32±8). Fracture prevalence at any-site was 14%, 16% and 15%, respectively (p<.0001). Multivariate models adjusted for age (>65), gender, race, BMI, serum calcium, phosphorus and creatinine demonstrated that: (1) compared to DM alone, co-prevalent CKD-DM was associated with 60% and 66% increased risk of hip and spine fractures respectively (hip: OR 1.6, CI 1.59-1.62; spine: OR 1.66, CI 1.64-1.67); and (2) compared to CKD alone, co-prevalent CKD-DM was associated with 43% and 14% increased risk of hip and spine fractures respectively (hip: OR 1.43, CI 1.41-1.44, spine: OR 1.14, CI 1.13-1.15).

Conclusion: These data suggest that co-prevalent CKD-DM is associated with higher rates of hip and spine fractures than CKD or DM alone. These data have high public health impact and epidemiologic and mechanistic studies are required to evaluate the effects of CKD-DM on the biochemical and micro architectural basis of fragility.


 

Nothing to Disclose: FK, FH, TN

18465 8.0000 THR-603 A Increased Fracture Risk in Co-Prevalent Diabetes and Chronic Kidney Disease in the United States 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Kathryn M Thrailkill*1, Robert Clay Bunn1, Jeffry S Nyman2, Mallikarjuna Rao Rettiganti1, Gael Cockrell1, Elizabeth Wahl1, Charles K Lumpkin1 and John Leslie Fowlkes1
1University of Arkansas for Medical Sciences, Little Rock, AR, 2Vanderbilt University Medical Center, Nashville, TN

 

Renal glucose resorption is regulated by the sodium glucose co-transporter 2 (SGLT2), a high affinity transporter expressed in the proximal tubule.  SGLT2 inhibitors are FDA-approved for treatment of type 2 diabetes (T2D), due to their anti-hyperglycemic properties resulting from enhanced urinary glucose excretion.  Clinical trials are also examining their efficacy in T1D.  Persons with T1D or T2D have increased risk of fracture, related to deficits in the microarchitecture and strength of diabetic bone; therefore, we examined the effects of an SGLT2 inhibitor on glucose and bone homeostasis in a model of diabetic bone disease.  Male, 10-week old, DBA/2J mice with streptozotocin (STZ)-induced diabetes were fed Teklad 8640 chow containing the SGLT2 inhibitor, Canagliflozin (CANA, 50 ppm, n=19) or control chow without CANA (n=18).  Non-diabetic mice were fed chow containing 62.5 ppm CANA, to offset the polyphagia in diabetic mice (n=20), or control chow (n=20).  CANA intake during the 10 week study was 17.7-20.6 mg/kg/day in the STZ (diabetes) group and 14.2-19.6 mg/kg/day in the non-diabetes group.  At sacrifice, serum glucose, PTH, P1NP and RatLAPs levels were measured; the left femur was harvested to assess fracture resistance by combined µCT analysis and three-point bending tests. In the metaphysis, long-term diabetes (DM) induced significant deficits in trabecular bone microarchitecture, including decreased trabecular bone volume fraction (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th) and trabecular tissue mineral density (Tb.TMD), along with increased trabecular spacing (Tb.Sp; p<0.0001 for all parameters).  In the diaphysis, a decrease in cortical bone area (Ct.AR), cortical thickness (Ct.Th) and minimal moment of inertia (Imin), occurred in DM (p<0.0001 for all), in addition to an increase in cortical porosity (Ct.Po; p<0.0001). These changes in DM were associated with reduced fracture resistance (decreased material strength and toughness as well as decreased structural strength and rigidity; p<0.001 for all).  DM was also associated with a ~4-fold increase in PTH (p<0.0001) and a ~2-fold increase in RatLAPs, a marker of bone resorption (p=0.0002), and RatLAPs was negatively correlated with Ct.Th, Tb.Th, and Tb.N (p<0.05 for all).  CANA treatment improved blood glucose in DM mice by ~40% (CONTROL: 101.2 ± 18.8 mg/dL; STZ: 525.2 ± 53.6; STZ + CANA: 338.9 ± 131.4; p<0.001 for STZ vs. STZ+CANA).  Glucose improvement, however, was not associated with significant improvement in trabecular microarchitectural parameters.  Instead, in CANA-treated diabetic mice, a further increase in RatLAPs was seen in the STZ + CANA group (CONTROL: 15.2 ± 3.4 ng/mL; CONT+CANA: 15.3 ± 4.0; STZ: 28.8 ± 6.3; STZ + CANA: 39.1 ± 17.9; p=0.005 for STZ vs. STZ+CANA).  This finding might suggest a drug-related further increase in bone resorption, perhaps induced by increased renal loss of bone minerals.

 

Nothing to Disclose: KMT, RCB, JSN, MRR, GC, EW, CKL, JLF

18680 9.0000 THR-604 A SGLT2 Inhibitor Treatment Improves Blood Glucose but Does Not Correct Diabetic Bone Disease in a Mouse Model 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Hyun Ae Seo* and Eui Hyun Kim
Daegu Fatima Hospital, Korea, Republic of (South)

 

Aims: Coronary artery disease is the major cause of morbidity and mortality in patients with type 2 diabetes. The coronary artery calcium score infers the presence of coronary atherosclerosis by measuring the total amount of calcium in the coronary arteries and has been known to predict the risk for cardiovascular disease. In this study, we investigated the relationship between coronary artery calcium scores and biochemical markers, especially lipid profiles and apolipoproteins.

Methods: We calculated the coronary calcium scores of 496 subjects with type 2 diabetes (229 males and 267 females) via multi-detector row computed tomography (MDCT). Coronary calcium levels were represented by Agatston score. Height, body weight, blood pressure, biochemical markers including lipid profiles (low-density lipoprotein, high-density lipoprotein, triglycerides and total cholesterol) and apolipoproteins (apolipoprotein A-1 and apolipoprotein B) were assessed concurrently.

Results: The mean age of total subjects was 58.46±12.44. The mean Agatston score was 229.81±675.56. The square root of Agatston score was made to achieve a normal distribution curve. Systolic blood pressure and blood urea nitrogen levels were significantly related to Agatston score. In patients with type 2 diabetes, Agatston score was highly associated with high density lipoprotein (HDL) (r = -0.212, P = <0.001). We performed subgroup analysis according to gender. Agatston score and HDL were significantly negatively correlated in both males and females (r = -0.183, P = 0.005 and r = -0.217, P = <0.001 respectively). The negative correlation between Agatston score and HDL after adjustments for age, BMI, systolic blood pressure, blood urea nitrogen, smoking status,  and use of lipid lowering drugs was retained in both males and females (r = -0.151, P = 0.034 and r = -0.229, P = <0.001 respectively).

Conclusion: These results suggest that HDL is a useful independent indicator of coronary artery calcification in patients with type 2 diabetes.

 

Nothing to Disclose: HAS, EHK

19650 11.0000 THR-607 A The Association Between Coronary Artery Calcium Score and High Density Lipoprotein 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Yassin M Mustafa*, Ana Shahnazi, Armana Saeed, Hassan Brim, Gail Nunlee-Bland, Saha Shrabonti, Seyed-Mehdi Nouraie and Hassan Ashktorab
Howard University Hospital, Washington, DC

 

Background: Type 2 diabetes mellitus (T2DM) patients are at increased risk for colorectal carcinoma (CRC). Obesity, insulin resistance and hyperinsulinemia either endogenous or exogenous are believed to be some of the risk factors. Metformin (MT), an insulin sensitizer, was suggested to decrease the risk of colorectal neoplastic lesions. As a result MT may reduce the mortality and increase survival in patients with established CRC.

Aim: To evaluate the association between MT therapy and colorectal neoplasia among African Americans (AAs) with T2DM

Methods: We retrospectively reviewed the medical records of patients who underwent diagnostic colonoscopy in our institution between 2000-2012. A group of predominantly AAs diabetic patients who had colonic neoplasia and a control group with no colonic neoplasia were identified. Data was analyzed using chi-square tests and multivariate logistic regression.

Results:We identified 214 patients with T2DM who were on anti-diabetic medications for at least 6 months prior to colonoscopy. Of these patients 156 (72.9%) had histopathologically confirmed colorectal neoplastic lesions and 58 (27.1%) were lesions free. Of patients with CRC or colorectal adenoma, 67 (43%) were on MT therapy compared to 35 (60%) patients without evidence of colorectal neoplasia(P=0.02). Multivariate logistic regression adjusted for age and gender demonstrated that patients on MT for more than 5 years had significantly decreased risk OR=0.2(95% CI: 0.1-0.4, P<0.001).

Conclusion: Our findings suggest that MT therapy is associated with a lower risk of colorectal neoplastic lesions in AAs with T2DM. Its protective effect appears to increase with duration of therapy.

 

Nothing to Disclose: YMM, AS, AS, HB, GN, SS, SMN, HA

21394 12.0000 THR-608 A Chronic Use of  Metformin Is Associated with a Reduced Colorectal Neoplasia Risk in Type 2 Diabetes Mellitus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Pinar Karakaya*1, Meral Mert2, Yildiz Okuturlar2, Asuman Gedikbasi2, Filiz Islim2, Didem Acarer2, Nursel Kocamaz2, Ozlem Soyluk2, Teslime Ayaz3, Pinar Alarslan4, Ozlem Harmankaya2 and Abdulbaki Kumbasar2
1Bakirkoy Dr. Sadi Konuk Training and Research Hospital, Istanbul, 2Bakirkoy Dr. Sadi Konuk Training and Research Hospital, Istanbul, Turkey, 3Recep Tayyip Erdogan University Training and Research Hospital, Rize, Turkey, 4Katip Celebi University Ataturk Training and Research Hospital, Izmir, Turkey

 

Human paraoxonase 1/arylesterase (PON1) is a calcium dependent ester hydrolase with paraoxonase, arylesterase and diazoxonase activities along with antioxidant and anti-atherogenic properties (1-3). Low PON-1 activity has been associated with susceptibility to coronary artery disease and increased risk of diabetic complications among patients with diabetes (3-5). Therefore the present was designed to evaluate the relation of serum PON-1 activity with biochemical variables as well as brachial artery diameter (BAd) and intima media thickness (BA-IMT) in diabetic patients with or without diabetes related complications. A total of 201 diabetic patients (mean±SD age: 52.4±13.4 years, 73.6% were females) with (n=50) or without (n=150) microvascular and with (n=91) or without (n=108) macrovascular complications were included. Data on patient characteristics, microvascular (neuropathy, retinopathy and nephropathy) and macrovascular [hypertension, coronary artery disease (CAD), past history of coronary artery bypass (CABG), peripheral artery disease (PAD), stroke and past history of myocardial infarction (MI)] complications, blood biochemistry, HOMA-IR, microalbuminuria and BAd and BA-IMT were evaluated with respect to serum PON-1 activity. Mean paraoxonase (107.5±30.7 vs. 123.9±38.8 U/L, p=0.007) and arylesterase (132.1±30.2 vs. 154.7±41.2 U/L, p=0.001) values were significantly lower in patients with than without microvascular complications. There was a negative correlation of HOMA-IR to paraoxonase (r=-0.383, p=0.001) and arylesterase (r=-0.252, p=0.035) levels in patients with macrovascular complications, whereas no correlation of HOMA-IR to paraoxonase or arylesterase was evident in patients with microvascular complications (r=-0.247, p=0.083 and r=-0.196, p=0.173, respectively). Arylesterase values was positively correlated with right BA-IMT in patients with microvascular complications (r=0.374, p=0.042). In conclusion, our findings in  a cohort of diabetic patients revealed no influence of gender, age and macrovascular complications on serum paraoxonase and arylesterase activities, whereas significantly lower PON-1 activity in patients with microvascular complications and negative correlation of both serum paraoxonase and arylesterase activities to HOMA-IR only in patients with macrovascular complications. Serum paraoxonase activities were not correlated to BAd and BA-IMT, regardless of diabetic complications. Our findings emphasize the possible role of low serum paraoxonase and arylesterase activities in predicting diabetic microvascular complications among diabetic patients and poor glycemic control in patients with diabetic macrovascular complications along with the role of arylesterase but not paraoxonase values in predicting early atherosclerosis in patients with diabetic microvascular complications.

 

Nothing to Disclose: PK, MM, YO, AG, FI, DA, NK, OS, TA, PA, OH, AK

18636 13.0000 THR-609 A Relation of Serum Paraoxonase-1 Activity with Biochemical Variables and Brachial Artery Diameter and Intima Media Thickness in Diabetic Patients with or without Diabetes Related Complications 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Gilberto J. Paz-Filho*, Teresa Neeman, Claudio Alberto Mastronardi and Alexander J. Rodriguez
The Australian National University, Canberra, Australia

 

Background: Diabetes is a significant cause of global morbidity. Patients with type 2 diabetes are at an increased risk of developing microvascular complications, including retinopathy, nephropathy and neuropathy. The adipocytokine leptin has been associated with vascular inflammation involved in these complications, and some cross-sectional studies have shown positive correlations between serum leptin levels and prevalence of diabetic microvascular complications. However, there is no comprehensive data examining blood leptin concentration with the presence of these complications.

Methods: A systematic review of the Scopus, MEDLINE and the Cochrane Library databases was conducted in accordance with the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) statement. Clinical studies were sought that assessed the association between circulating leptin concentrations and diabetic microvascular complications. Study quality was evaluated using a modified version of QUADAS 2, an instrument for the quality assessment of studies included in systematic reviews. Meta-analysis was performed on pooled data using an inverse-variance random-effects model, using RevMan v5 software. The standardized mean difference (SMD) and 95% confidence intervals (CI) were calculated for concentrations of leptin in diabetic patients with and without microvascular complications. Heterogeneity was determined by the Q and I2 statistics.

Results: A total of 262 unique abstracts were identified, of which 13 satisfied our inclusion and exclusion criteria. Study quality ranged from 37.5% to 87.5%, and most studies failed to report medication use. Higher leptin levels were associated with nephropathy [0.51 SMD (95% CI: 0.12, 0.90), p=0.01; n=1,165 participants from nine studies]. A leave-one-out sensitivity analysis showed the result remained significant. Leptin was not associated with retinopathy [0.17 (-0.30, 0.65), p=0.48; n=1,012 from five studies] or neuropathy [0.12 (-0.19, 0.43), p=0.45; n=791 from four studies]. Heterogeneity (I2) for studies reporting nephropathy, retinopathy and neuropathy were 87%,  89% and 73%, respectively.

Discussion: Adipocytokines have been variously associated with diabetic microvascular complications. This meta-analysis suggests that relative to patients without complications, higher leptin is associated with nephropathy. A sensitivity analysis of leptin results demonstrates the strength of this finding. Therefore, adipocytokines are clinically relevant to diabetic microvascular complications and may have future potential as novel therapeutic targets or biomarkers for diagnostics, surveillance or risk prediction. Studies were limited by their cross-sectional design and thus large prospective analyses are required to confirm these findings independent of other risk factors.

 

Nothing to Disclose: GJP, TN, CAM, AJR

20210 14.0000 THR-610 A Associations Between Leptin and Diabetic Microvascular Complications: Systematic Review and Meta-Analysis of Cross-Sectional Data 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Akshata Desai*1, Manisha Garg2, Sartaj Sandhu3, Simmanjeet Mangat3, Garima Thapar3, Rujuta Katkar3, Aditya Mehta2, Naven Kang3, Nitesh D Kuhadiya2, Sandeep S Dhindsa3, Ajay Chaudhuri2 and Paresh Dandona2
1SUNY at Buffalo, NY, 2Diabetes and Endocrinology Center of Western New York, Buffalo, NY, 3Diabetes and Endocrinology Center of Western New York, Williamsville, NY

 

Our recent data show that exenatide treatment in type 2 diabetes induces an increase in the expression of the genes of key anti-oxidant enzymes, NQO-1, GST-P1 and HO-1, which are modulated by the key transcription factor, Nrf-2, and suppresses Keap-1, the inhibitor of Nrf-2 (1).These observations are important since Nrf-2 regulated genes reduce oxidative stress and protect the kidneys from developing nephropathic changes in mice rendered diabetic by streptozotocin. These actions are in addition to the potent and comprehensive anti-inflammatory effects exerted by exenatide previously demonstrated by us (2). Furthermore, our data also show that exenatide suppresses TGFβ-1, the key cytokine involved in the pathogenesis of diabetic nephropathy (1). Since our initial observations on the cellular and molecular actions of liraglutide confirm these data, these actions thus are probably applicable to other GLP-1 receptor agonists. In animal models, streptozotocin induced diabetes related nephropathy has been inhibited by GLP-1 receptor agonists through their anti-inflammatory and ROS suppressive actions.

 On the basis of the above, we retrospectively analyzed data from patients treated at our center and found that albuminuria in diabetic patients taking GLP-1 receptor agonists was significantly better than that in those on other anti-diabetic drugs. Thus, in those who presented with micro-albuminuria and were treated with GLP-1 receptor agonists, 61% became normoalbuminuric and no one developed macro-albuminuria. In the other group, 11% developed macro-albuminuria while 42% became normoalbuminuric (p<0.01)(3). In normoalbuminuric patients, 11.3% developed either microalbuminuria or macroalbuminuria in the group on GLP-1 receptor agonists while in the other group, 20% developed either microalbuminuria or macroalbuminuria. These differences were independent of glycemic control since HbA1c levels were similar in the two groups. BP, lipids, use of ACE inhibitors/ARB and statins were also similar between the two groups. No patient in either group developed ESRD. Thus, clearly, the evolution of albuminuria in patients treated with GLP-1 receptor agonists was significantly better than those in the rest as shown below in Figure 1. Clearly, a prospectively randomized study investigating the effect of a GLP-1 receptor agonist on albuminuria or GFR in patients with diabetes needs to be carried out.

 

Nothing to Disclose: AD, MG, SS, SM, GT, RK, AM, NK, NDK, SSD, AC, PD

19227 15.0000 THR-611 A GLP-1 Receptor Agonists Reduce the Progression of Proteinuria in Type 2 Diabetes Mellitus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Jung Soo Lim*1, Ji Hye Huh1, Young Ju Choi2, Byoung Wook Huh2, Mi Young Lee1, Jang Yel Shin1, Choon Hee Chung1, Eun Jig Lee3 and Kap Bum Huh2
1Yonsei University Wonju College of Medicine, Wonju, Korea, Republic of (South), 2Huh’s Diabetes Center and the 21C Diabetes and Vascular Research Institute, Seoul, Korea, Republic of (South), 3Yonsei University College of Medicine, Seoul, Korea, Republic of (South)

 

Background Calcium homeostasis has been shown to affect insulin resistance and secretion. Moreover, several studies have reported that elevated serum calcium is associated with an increased risk of developing metabolic abnormalities, including type 2 diabetes mellitus (DM). However, it remains unclear whether serum calcium level can affect the presence of nonalcoholic fatty liver disease (NAFLD), which has been considered as the hepatic expression of metabolic syndrome. The aim of this study was to investigate the relationship between serum calcium level and NAFLD in Korean type 2 DM with normocalcemia.

Methods A total of 5,711 diabetic subjects who visited our clinic from Jan 2003 to May 2012 were enrolled. Subjects were divided into four groups according to the baseline quartiles of serum calcium levels: Q1 (8.4-8.8 mg/dL), Q2 (8.9-9.2 mg/dL), Q3 (9.3-9.5 mg/dL), and Q4 (9.6-10.4 mg/dL). Insulin sensitivity was assessed by the rate constant of plasma glucose disappearance (Kitt, %/min) using a short insulin tolerance test. Moreover, fatty changes of liver were examined by ultrasonography.

Results The prevalence of NAFLD was 58.8% in the whole study population. As serum calcium levels increased, NAFLD also significantly increased (53.5% in Q1, 58.8% in Q2, 59.7% in Q3, and 61.5% in Q4, respectively). In addition, blood pressure, total cholesterol, LDL cholesterol and plasma triglyceride levels were significantly higher in subjects in the highest quartile compared with those in the lowest, although there were no differences in age, gender and body mass index among the groups. Moreover, insulin sensitivity was significantly reduced in the highest quartile than in the lowest one. Multiple regression analyses showed that the odds ratio (OR) for NAFLD was about 33% higher in individuals with calcium concentration≥9.6 mg/dL than in those with calcium concentration <8.9 mg/dL (OR 1.33 [95% confidence interval 1.08-1.63]).

Conclusion These findings suggest the possibility that increased serum calcium may in part be involved in developing NAFLD in subjects with type 2 DM, even in normal levels. Further investigations are needed to establish a casual relationship.

 

Nothing to Disclose: JSL, JHH, YJC, BWH, MYL, JYS, CHC, EJL, KBH

21475 16.0000 THR-612 A Association Between Serum Calcium Level and Nonalcoholic Fatty Liver Disease in Korean Type 2 Diabetes Mellitus with Normocalcemia 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Ho Jin Kim*, Jun Ho Lee, Jae Ho Cho, Jun Sung Moon, Ji Sung Yoon, Kyu Chang Won and Hyung-Woo Lee
Yeungnam University College of Medicine, Daegu, Korea, Republic of (South)

 

Introduction : Red blood cell (RBC) deformability is an ability of RBC to change shape under stress. RBC deformability has been demonstrated to be impaired in diabetes mellitus. But, little is known about the association between impaired RBC deformability and type 2 diabetes mellitus (T2DM). The aim of this study was to determine the influence of RBC deformability on T2DM.

Methods :We conducted a cross-sectional study with 198 patients with T2DM who visited in Yeungnam university hospital from Mar. to Jul. 2014. Patients with end stage renal disease and who are taking a pentoxifylline and ginkgo biloba were excluded. RBC deformability was measured by using a Rheoscan-D (Rheo-Meditech, Seoul, Korea), and expressed as elongation index (EI). The EI was measured at 3 Pa. We divided the EI into quartile (Q1, Q2, Q3 and Q4 from lowest to highest EI).

Results : 193 patients (mean age 59.82 ± 12.29 years, M:F = 100:93) were finally included. EI had significantly negative correlation with the levels of glycated hemoglobin, and positive correlation with HOMA-B, respectively (β -23.52 , P=0.01 and β 520.03, P=0.02, respectively). Patients with micro complications had lower EI compared with patients without complications (EI 0.303623 vs. 0.310637, p=0.01). Of them, patients with retinopathy had lower EI compared with patients without retinopathy (EI 0.300449 vs. 0.309653, p=0.00), whereas patients with nephropathy or neuropathy and macro complications had no significant difference in EI. After adjustment for age, sex, hypertension, smoking, and lipids, lower EI remained significantly associated with the prevalence of diabetic retinopathy (Odds ratio for Q1 compared with Q4, 4.16; 95% confidence interval, 1.43-12.13).

Conclusion : In patients with T2DM, there are significant relationship between RBC deformability and glycemic control, beta cell function and diabetic retinopathy. These results suggest that decreased RBC deformability is a useful surrogate marker for predicting diabetic retinopathy.

 

Nothing to Disclose: HJK, JHL, JHC, JSM, JSY, KCW, HWL

20382 17.0000 THR-613 A Association Between Red Blood Cell Deformability and Diabetic Complications in Patients with Type 2 Diabetes Mellitus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Mohamed I. Husseiny Elsayed*1, Weiting Du1, Ding Wang1, Jeffrey Rawson1, Alexander Kaye1, Fouad R Kandeel1 and Kevin George Ferreri2
1Diabetes & Metabolic Research Institute, Beckman Research Institute of City of Hope, Duarte, CA, 2City of Hope, Duarte, CA

 

Type 1 diabetes (T1D) is a chronic autoimmune disease characterized by destruction of pancreatic insulin-producing beta cells due to loss of tolerance to specific self-antigens (autoantigen). The attack on the beta cells is accompanied by the development of antigen-specific antibodies and cytotoxic T lymphocytes (CTLs). One of the most promising approaches to restore balance within the immune system is oral administration of diabetic autoantigens, which can result in deletion of destructive islet-specific responses and induction of regulatory responses. Studies in animal models and in the clinic have shown that administration of autoantigens, especially orally to the gut associated lymphoid tissue (GALT), can slow or even stop the progression of disease. We recently reported the development of an oral vaccine based on live attenuated Salmonella for the prevention of diabetes in non-obese diabetic (NOD) mice. That study used mouse preproinsulin (mPPI) gene as the autoantigen fused to SseF effector protein of type-III secretion system (T3SS) encoded by Salmonella Pathogenicity Island-2 (SPI2) under the control of an intracellular regulated promoter (Pro sseA) required for intracellular survival and replication. The SPI2-T3SS prevents extracellular expression and allows preferential delivery of fused protein into the cytosol of antigen-presenting cells (APCs) for optimal immunogenicity. The Salmonella-based delivery of autoantigen was co-administered with Salmonella-based delivery of DNA for mammalian expression of transforming growth factor beta (TGFbeta) by the host APCs. We showed that oral vaccination with the combined mPPI+TGFbeta prevented diabetes in the majority of mice and restored normal glucose tolerance, while untreated or autoantigen alone mice became diabetic. In this study we extended the method to another diabetic autoantigen, Glutamic Acid Decarboxylase (GAD)-65. Similar to preproinsulin, oral co-vaccination with Salmonella carrying GAD65 and TGFbeta prevents diabetes in the majority of animals but GAD65 alone does not. To better understand the mechanism of vaccine action the populations of CD4, CD8, CD25, and FOXP3 were quantified as well as the levels of cytokines in circulation and cytokines released after autoantigen stimulation. We conclude that a Salmonella-based oral vaccine expressing autoantigens in combination with tolerogenic cytokines is a promising therapy for prevention of T1D.

 

Nothing to Disclose: MIHE, WD, DW, JR, AK, FRK, KGF

19112 18.0000 THR-614 A Oral Immunotherapy for Type 1 Diabetes Based on Live Attenuated Salmonella 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Mingfeng Zhang*1, Qing Lin1, Limin Wu1, Nainong Li1, Jeremy Racine1, Qi Tong2 and Defu Zeng1
1Beckman Research Institute of City of Hope, Duarte, CA, 2City of Hope, Duarte, CA

 

We previously reported that induction of mixed chimerism under a radiation-free anti-CD3/CD8 conditioning regimen cures new-onset type 1 diabetes (T1D), and induction of mixed chimerism in combination with administration of growth factors cures late-stage T1D in autoimmune NOD mice. Due to unavailability of anti-human CD3 and CD8 mAb, we need to develop a new regimen with clinically available reagents. Cyclophosphamide, Pentostatin, and ATG has been used to condition patients in clinical hematopoietic cell transplantation (HCT). In the current studies, we conditioned NOD mice with cyclophosphamide (50mg/Kg, daily, 14 days), pentostatin (1mg/Kg, every 3 days, total four injections), and ATG (50mg/Kg, every 4 days, total three injections). The conditioned NOD mice were transplanted with donor BM (50x106) and CD4+ T-depleted spleen cells (25x106) from MHC-mismatched C57BL/6 donor mice. About 85% (10/12) of overt diabetic NOD recipients developed mixed chimerism with no signs of graft versus host disease (GVHD). The mixed chimerism cured new-onset (3 days after onset) T1D via elimination of insulitis and replication of residual β cells. The mixed chimerism in combination of administration of gastrin and EGF cured more than 50% of late-stage (3 weeks after onset) T1D via augmenting β cell replication and neogenesis. These results indicate that induction of mixed chimerism with clinically applicable conditioning regimen has the potential to cure overt T1D.

 

Nothing to Disclose: MZ, QL, LW, NL, JR, QT, DZ

19685 19.0000 THR-615 A Induction of Mixed Chimerism for Cure of Overt Type 1 Diabetes in NOD Mice, Using Clinically Available Reagents without Radiation 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Mustafa Tosur*1, Pamela J Lupo2, Jianli Dong2, Ketan N Patel2 and Phillip D K Lee2
1Texas Children's Hospital / Baylor College of Medicine, Galveston, TX, 2University of Texas Medical Branch, Galveston, TX

 

Introduction: We report an adolescent male with incidentally discovered non-ketotic diabetes mellitus (NKDM), who was also found to have involuntary muscle cramping and severe hypertriglyceridemia.  

Clinical Case: During evaluation for possible viral meningitis, a 14 year-old lean athletic male was incidentally found to have NKDM with negative anti-glutamic acid decarboxylase and islet cell antibodies, and elevated C-peptide. Treatment with glipizide XL 30 mg daily, metformin XR 500 mg bid and carbohydrate limited diet reduced HbA1c from 9.0 to 7.4%. During the course of the diabetes evaluation, he complained of pre-existing painful muscle cramping, intermittently and unpredictably affecting all muscle groups; he had been reluctant to describe this previously. Electromyography was technically difficult due to severe cramping, and was reported as widespread fasciculations and cramp potential without any evidence of myopathy. Lipemic serum noted during a blood draw resulted in a diagnosis of hypertriglyceridemia (>1500 mg/dl) without significant hypercholesterolemia; this has been partially controlled with omega-3 oil and diet with triglyceride 572 mg/dl after a year of treatment. Other clinical findings include unilateral pelvic kidney, moderate proteinuria and elevated blood pressure; renal biopsy pending. Chromosomal microarray analysis showed loss of heterozygosity on chromosomes 4p15.2, 5q13.2, and 19q13.11, which includes total of 24 disease-causing genes in OMIM, increasing the risk of recessive conditions. A 1.28 Mb loss of chromosome 16p11.2 was also identified without disease-causing genes in OMIM. The clinical significance of the microarray results is unknown. LMNA sequencing and targeted gene testing for Kennedy disease were negative. Whole exome sequencing revealed a heterozygous ATM mutation with unknown clinical significance (c.1595G>A); expanded report pending. The patient’s clinical condition has shown virtually no change over 3 years of treatment except for partial biochemical response to medical treatment and diet.

Conclusion: We report a unique association of NKDM with intrinsic muscle disorder, severe hypertriglyceridemia, proteinuria and possibly unrelated renal anomaly. ATM mutation has been previously shown to be associated with insulin resistance and diabetes; however, the role of patient’s heterozygous mutation on his clinical condition is not clear.

 

Nothing to Disclose: MT, PJL, JD, KNP, PDKL

19008 20.0000 THR-616 A An Unusual Case of Non-Ketotic Diabetes in an Adolescent Male 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Sharad S Singhal, James L Figarola, Jyotsana Singhal*, Samuel Rahbar and David Horne
Beckman Research Institute of the City of Hope, Duarte, CA

 

Growing evidence indicates that oxidative stress is increased in the diabetic condition due to overproduction of reactive oxygen radicals and decreased efficiency of antioxidant defenses.  Glutathione S-transferases (GSTs) are a large, multi-gene family of proteins that may play an important role in protecting tissues from oxidative damage because they function in the transport of cellular components or metabolites (or conjugation of metabolites with glutathione) formed during diabetes.  The present study was designed to evaluate the effect of “COH-SR4” on glutathione (GSH)-dependent xenobiotic metabolism and oxidative stress in mice with diabetes.   Profound changes in the levels of GSH-linked antioxidant enzymes such as GST, glutathione peroxidase, glutathione reductase, glucose 6-phosphate dehydrogenase and gamma-glutamyl cysteine synthetase, and oxidative stress markers such as lipid hydroperoxides (LOOH) and thiobarbituric acid reactive substances (TBARS) were assessed in the liver of normal and diabetic (db/db) mice with and without treatment with “COH-SR4”.  In diabetes mellitus, lipid peroxides formed due to oxidative stress serve as endogenous substrates for GSTs.  Mammalian GSTs are highly efficient in the detoxification of these compounds.  Lower levels of GSH and GST, and higher levels of oxidative stress markers were found in the diabetic mice.  The concentration of malondialdehyde (MDA) measured in the liver of normal mice was 630 ± 103 nmol/g tissue.  The changes in the concentration of MDA measured in the liver of db/db mice (2428 ± 235 nmol /g tissue, p < 0.01) and db/db mice treated with SR4 (894 ± 76 nmol /g tissue, p < 0.01) were statistically different from the db/db mice.  SR4 feeding to db/db mice (5 mg/kg b.w., 4-weeks treatment on alternate days by oral gavage) significantly decreases oxidative stress markers, brought the level of LOOH and MDA approximately to the level found in control mice, which could be explained by its anti-hyperglycemic effect.  GSH concentration was significantly lower of db/db mice compared to control (~ 58%; p < 0.01).  Feeding of COH-SR4 to db/db mice markedly increased GSH concentration.  These results are consistent with prediction of our model that higher sugar levels caused by lower AMPK expression in db/db mice should result in accumulation of their precursor lipid hydroperoxides and their degradation products.  These results suggest that the antioxidant deficiency and excessive peroxide-mediated damage may appear early on in non-insulin-dependent diabetes mellitus, before the development of secondary complications.  In conclusion, SR4 is a safe and inexpensive product, causes a significant reduction of the glycation of proteins in diabetic animals.  In summary, the present study on the anti-diabetic effects of a novel dichlorophenyl urea compound “COH-SR4” holds great future promise for further development in diabetes treatment.

 

Nothing to Disclose: SSS, JLF, JS, SR, DH

20465 21.0000 THR-616 A Role of COH-SR4 in Diabetes: Glutathione-Linked Enzymes and Oxidative Stress 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Vera Fernandes*1, Maria Teresa Pereira2, Susana Garrido2, Andre Carvalho2, Claudia Amaral3, Cláudia Freitas2, Helena Neto2, Joana Martins2, Isabel Gonçalves2, José Muras2, Sara Pinto2 and Rui Carvalho2
1Hospital de Braga, Braga, Portugal, 2Hosp Santo Antonio, Porto, Portugal, 3Hospital de Santo Antonio, Porto, Portugal

 

Introduction: The proportion of diabetic patients undergoing a major amputation, preceded by an unsuccessful minor amputation, is not known. Therefore, it’s not clear if a conservative or aggressive attitude is being adopted at the beginning, and what are the consequences.

Aim: To compare diabetic patients undergoing a major amputation preceded by a recent minor amputation (same limb, in the last 30 days), with patients undergoing a major amputation at the beginning, concerning demographic and clinical characteristics. 

Methods: Observational, analytical and retrospective study of diabetic patients undergoing a major amputation at a central hospital, between 2011 and 2013.  The chi-square, exact Fisher’s test, Mann-Whitney and Kruskal-Wallis tests were used.

Results: Of the 195 major amputations included in this study, 51% occurred in females, 35% were previously autonomous and 67.7% were 65 years or over. In 44 major amputations (22.6%) there had been a previous minor amputation and the time between amputations was mostly less than 14 days. Only 6 amputations were performed in patients considered purely neuropathic and none of these subjects had a recent minor amputation. The patients with a recent minor amputation showed to be younger (p=0.028) and autonomous (p=0.025); they presented higher HbA1c levels (p=0.003) and this group had a longer hospital stay (p<0.001). Regarding diabetic complications, it was found that diabetics with major amputation at the beginning had more often cerebrovascular disease (p=0.047) and they presented also a higher number of macrovascular complications (no statistical significant). The proportion of major amputation preceded by a minor was 20.3% in 2011, 16.9% in 2012 and 29.2% in 2013.

Conclusion: This study shows that an important number of major amputations is preceded by a unsuccessful minor amputation, especially in younger and autonomous patients. The older patients and those with more chronic diabetic complications appear to be more often targeted to have a major amputation from the beginning.

 

Nothing to Disclose: VF, MTP, SG, AC, CA, CF, HN, JM, IG, JM, SP, RC

21221 22.0000 THR-617 A Diabetes & Lower Extremity Amputation - Major Amputation: An Expectable Result?- 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Muge Keskin1, Nese Ersoz Gulcelik*2, Cavit Culha3 and Yalcin Aral4
1Ankara Training & Res Hosp, Ankara, Turkey, 2Ankara Training Hospital, 3Ankara Training and Research Hospital, 4Ankara Training and Research Hospital, Ankara, Turkey

 

Fetuin- A levels predict cardiovascular risk in young diabetic patients

Objective: Young adults with type 2 diabetes (T2DM) were demonstrated to have increased cardiovascular disease risk. Fetuin-A (FETUA) levels are associated with insulin resistance, impaired glucose tolerance, hepatosteatosis, subclinical inflammation and increased cardiovascular risk. Apolipoprotein (apo) B/Apo A1 ratio is a well-defined cardiovascular risk assessment marker. In this study, we aimed to investigate the potential role of FETUA in demonstrating cardiovascular disease risk in young adults with type 2 diabetes.

Material and methods: We performed a prospective study on 18 controls, 18 diabetic and 19 prediabetic patients (age 20-40 years). History, physical examination and anthropometric measurements were done for each subject. Fasting serum samples were obtained from all subjects and glucose, insulin, FETUA, LDL, triglyceride, HDL, VLDL, apoB, apoA1, lipo A, HSCRP levels were measured. Serum FETUA levels were determined by ELISA.

Results: FETUA levels were significantly lower in control patients than diabetic and prediabetic patients (33,46±22,10 vs. 51,68±17,10ng/ml, p<0,05 and 33,46±22,10 vs 47,70±10,47ng/ml, p<0,05, respectively). FETUA levels did not differ between diabetic and prediabetic patients (51, 68±17, 10 vs. 47, 70±10, 47ng/ml, p>0, 05). BMI adjusted correlations revealed a positive correlation between FETUA levels and glucose, Apo B levels and Apo B/Apo A1 ratio.

Conclusions: Studies showed that FETUA levels may play a role in the pathophysiology of cardiovascular diseases. In our study, we found a significant increase in FETUA levels in young type 2 diabetic and prediabetic patients. Thus, FETUA may be a useful marker in determining cardiovascular risk of young patients with diabetes or at risk for diabetes.

 

Nothing to Disclose: MK, NE, CC, YA

19590 23.0000 THR-618 A Fetuin- a Levels Predict Cardiovascular Risk in Young Diabetic Patients 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Mohammed Abdulrahman Alshehri*1, Khaled Abdullah Alswat2, Sameer R. Alharthi3 and Ahmed A. Alsuwat3
1King Abdulaziz Specialist Hospital, Taif, 2Taif University, School of Medicine, Taif, Saudi Arabia, 3Taif University

 

Background:

The IDF has released an estimates 382 million with T2D in 2013.The T2D is associated with an increased risk of depression with subsequently increased risk of diabetes complications. Few studies assessed the impact of media consumption and metformin (MET) on risk of depression.

Methods:

We conducted a cross-sectional study at the King Abdulaziz Specialist Hospital, Taif, Saudi Arabia. T2D patients (pts) > 18 years who had a routine visit to the Endocrine clinic from Jun 2013 to Aug 2014 were asked to participate. Baseline characteristics and measurement was obtained. Laboratory data was collected from the pts EMR. We excluded pts with T1D, existing psychiatric illness, untreated hypothyroidism and history of active cancer. We used PHQ-9 to screen for depression and Berlin questionnaire to screen for obstructive sleep apnea (OSA). We measured media mass by hours that include watching TV, using computer, video game and using smart devices for fun. The primary goal of this study is to assess the impact of media mass and MET on the risk of depression and OSA and its relation to the cardiovascular (CV) markers in T2D.

Result:

283 pts participated, 134 (47.3%) were male vs 149 (52.7%), mean age 56.8yrs, mean HbA1c 8.54%, mean duration of T2D of 11.17 yrs, mean BMI 31.8 kg/m2,88.3% were married, 18% did college degree or higher, 65.7% were low income, 41% were on OHA only, 52.7% were on insulin +/- OHA, and 3.9 % were on diet control only. Mean media consumption were 3.3 hrs and 25.4% were exercising regularly.

41.3% met the criteria for depression and 49% were at high risk for OSA. 53.7% consume media ≤ 2 hrs vs 46.3% consume it > 2hr/d, with mean age of 57.1yrs vs 56.5yrs (p .66), 42.5% were male vs 53.1% (p .07), mean BMI 31.4 vs 32.3 kg/m2(p.23), 40.5% vs 42.3% were depressed (p .76), and 50.3% vs 49.2% were at high risk of OSA (p .85).

In regards to the CV markers, those who consume it ≤ 2 vs > 2 hrs/day, their mean HbA1c 8.4% vs 8.6% (p .54), SBP 141.7 vs 139.1 mmHg (p .35), DBP 80.4 vs 81.1 mmHg (p .68), resting HR 80.9 vs 82.0 bpm (p .5), LDL 99.1 vs 103.4 mg/dl (p .42), HDL 43.8 vs 41.4 mg/dl (p .26), and triglyceride 145.8 vs 162.2 mg/dl (p .15).

Those who consume media ≤ 2 hrs, 27.5% exercise regularly compare to 23.1% (p .4), 5.9% are active smoker compare to 10.8% (p .75), to those who consume it > 2 hrs/d respectively.

188 (66.4%) pts were on MET, 37.2% of pts on MET compare to 49.5% of those who weren’t met the criteria for depression (OR 0.66, CI 0.37-0.99). After adjustment of age, gender, duration of T2D, income, education, media mass, FHx of depression, BMI and HbA1c, this association remains significant (p 0.046). 46.3% in the MET group vs 56.8% in those who weren’t, considered as high risk for OSA.

Conclusion:

Pts who consume media >2 hrs/d were non-statistically significant at higher risk of depression, poor glycemic control, worse most of the measured CV markers and reports more unhealthy habits .Pts on MET were at lower risk of depression.

 

Nothing to Disclose: MAA, KAA, SRA, AAA

20609 24.0000 THR-619 A Impact of Media Consumption and Metformin on Risk of Depression and Sleep Apnea in Diabetics 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 596-619 5931 1:00:00 PM Diabetes Immunology, Diabetes Complications Poster


Grigorios Rombopoulos1, Elena Panitti1, George S Vernikos*1, Katopodis Panagiotis2 and Skoutas Dimitrios3
1Novartis Hellas S.A., Athens, Greece, 2Peristeri Clinic, Athens, Greece, 3Sarafianos Lysimachos General Clinic of Thessaloniki, Thessaloniki, Greece

 

Introduction: Management of type 2 diabetes mellitus (T2DM) poses unique challenges in the rapidly growing elderly patient population due to age-related morbidity and mortality. There is limited real world evidence on DPP-4 inhibitors in elderly T2DM patients in Greece. This study evaluates the effectiveness and tolerability of vildagliptin in addition to metformin in elderly patients with T2DM in Greece.

Methods:This was a multicenter observational study with duration of 12-14 weeks. Data recording was completed in two visits (Day 1, Week 12-14/end of study). Patients aged ≥70 years with a diagnosis of T2DM who have, within the previous 2 weeks, been prescribed vildagliptin as add on to metformin therapy or sulfonylurea on top of metformin monotherapy, or T2DM patients inadequately controlled HbA1c ≥7.0% and ≤9.0% on stable doses of metformin monotherapy for at least 3 months prior to entering the study, were recruited. The primary endpoint was change in HbA1c from baseline after 12 weeks of treatment with vildagliptin in addition to metformin or vildagliptin plus metformin single pill combination. Secondary assessments included proportion of patients reaching the glycemic target of HbA1c ≤7.0% and ≤6.5%, change in fasting plasma glucose (FPG) and body mass index (BMI), safety and tolerability, estimation of the incidence rate of confirmed hypoglycemic events and description of the demographic characteristics of the patient population. To assess the incidence rate of confirmed hypoglycemic events, a descriptive control arm of patients who recently initiated sulfonylurea on metformin monotherapy was also exploited.

Results: Of the total of 336 patients at study entry, 50.3% of patients were men, mean age (± SD) was 75.9 ± 4.8 years, mean BMI 28.5 ± 4.1 kg/m2, mean baseline HbA1c 7.9 ± 0.5% and mean FPG 167.8 ± 33.1 mg/dl. For the 285 patients receiving vildagliptin and metformin (245 receiving a fixed combination and 40 receiving a free combination), the mean change in HbA1c from baseline to 12 weeks was -0.97% (95% CI -1.04, -0.91), mean change in FPG was -42.8 mg/dl (95% CI -46.4, -39.3) and mean change in BMI was -0.69 kg/m2(95% CI -0.81, -0.57); p<0.0001 in all cases. Overall, 55.6% of patients reached HbA1c ≤7.0% and 20.4% reached HbA1c ≤6.5%. Overall, two patients reported three adverse events namely, lung infection, hyperpyrexia and constipation; according to the investigator these were not related to the drug. Hypoglycemia occurred in 3 (1.2%) patients receiving vildagliptin and metformin.

Conclusions: Vildagliptin in combination with metformin showed improved glycemic control in daily clinical practice in elderly Greek patients with no increased incidence of adverse events.

 

Disclosure: GR: Employee, Novartis Pharmaceuticals. EP: Employee, Novartis Pharmaceuticals. GSV: Employee, Novartis Pharmaceuticals. KP: Principal Investigator, Novartis Pharmaceuticals. SD: Principal Investigator, Novartis Pharmaceuticals.

19159 1.0000 THR-640 A Effectiveness and Safety of Vildagliptin in Addition to Metformin in Elderly (>=70 years) Greek Patients - the Esteem Study 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Byung-Wan Lee1, Jae Hyeon Kim2, Se Eun Park3, Chang Hee Jung4, Seung-Hwan Lee5, Sunghwan Suh6, Woo Je Lee4, Younghwan Jang7, Sung-Ho Kim7 and Cheol-Young Park*3
1Department of Internal Medicine, Severance Hospital, Endocrine Research Institute, Yonsei University College of Medicine, Seoul, Korea, Republic of (South), 2Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea, Republic of (South), 3Kangbuk Samsung Hospital, Sungkyunkwan University School of Medicine, Seoul, Korea, Republic of (South), 4Asan Medical Center, University of Ulsan College of Medicine, Seoul, Korea, Republic of (South), 5Seoul St. Mary’s Hospital, The Catholic University of Korea School of Medicine, Seoul, Korea, Republic of (South), 6Dong-A University Medical Center, Seoul, Korea, Republic of (South), 7LG Life Sciences, Seoul, Korea, Republic of (South)

 

Background and aims: Glycemic variability and chronic sustained hyperglycemia are the main components of dysglycemia in diabetes. The purpose of this study was to assess whether there is a difference between- or within drug classes on glycemic variability and glucose control as initial combination therapy with metformin in drug-naive patients with type 2 diabetes.

Materials and methods: A multi-center, randomized, active controlled, open-label, parallel design study was performed in 69 patients with HbA1c greater than 7.5%. Subjects were randomized (1:1:1) to receive gemigliptin 50 mg qd, sitagliptin 100 mg qd, or glimepiride 2 mg qd for 12 weeks. The mean amplitude of glycemic excursions (MAGE) and standard deviation (SD) were used for assessing glucose fluctuations at baseline and after 12 weeks. Glycosylated hemoglobin (HbA1c), glycated albumin (GA), fructosamine, C-reactive protein and other metabolic parameters were also measured. Safety and tolerability based on adverse events (AEs) were assessed. To strengthen the reliability of the study results, data from continuous glucose monitoring system (CGMS) was evaluated independently by a blinded central evaluator.

Results: A total of 69 subjects were randomized to receive gemigliptin 50 mg (n = 24), sitagliptin 100 mg (n = 23) or glimepiride 2 mg (n = 22). Mean baseline characteristics were similar across the groups (age, 50.0 years; HbA1c, 9.4%; GA, 26.4%; fructosamine, 412.8 mol/L). At 12 weeks, MAGE was significantly lower in the DPP-4 inhibitor groups, gemigliptin and sitagliptin, than in the glimepiride group (-43.1, -38.3, and -21.7 mg/dl, respectively). Furthermore, the SD of mean glucose was significantly lower in patients with gemigliptin (vs sitagliptin p=0.01; vs glimepiride p=0.007) when compared with sitagliptin and glimepiride. Mean HbA1c was reduced from baseline by 2.75%, 2.24% and 2.75% for gemigliptin, sitagliptin and glimepiride, respectively. A similar profile was also observed in other glycemic control parameters (FPG, glycated albumin, and fructosamine). A greater decrease in total- and LDL-cholesterol and nitrotyrosine was observed for DPP-4 inhibitor groups versus glimepiride, although there were no significant differences between the groups. In addition, only gemigliptin significantly decreased C-reactive protein levels from baseline. Drug-related AEs including symptomatic hypoglycemia were reported more frequently in glimepiride group than in other groups.

Conclusion: In summary, gemigliptin was more effective than glimepiride and sitagliptin in reducing glucose variability as initial combination therapy with metformin in drug-naïve patients with T2DM.

 

Disclosure: CYP: Investigator, Merck & Co.. Nothing to Disclose: BWL, JHK, SEP, CHJ, SHL, SS, WJL, YJ, SHK

21425 2.0000 THR-641 A Effects of Gemigliptin Versus Sitagliptin or Glimepiride on Glycemic Variability As Initial Combination Therapy with Metformin in Drug-Naïve Patients with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Andrew J Lewin*1, Ralph DeFronzo2, Sanjay Patel3, Dacheng Liu4, Renee Kaste4, Hans-Juergen Woerle5 and Uli C Broedl5
1National Research Institute, Los Angeles, CA, 2Univ of Texas Hlth Science Ctr, San Antonio, TX, 3Boehringer Ingelheim Ltd., Bracknell, United Kingdom, 4Boehringer Ingelheim Pharmaceuticals Inc, Ridgefield, CT, 5Boehringer Ingelheim Pharma GmbH & Co. KG, Ingelheim, Germany

 

A randomized, double-blind, parallel group Phase III study evaluated the efficacy and safety of combinations of empagliflozin/linagliptin (EMPA/LINA) in subjects with type 2 diabetes (T2DM) who were not receiving anti-diabetes therapy for ≥12 weeks. Subjects were randomized to EMPA 25 mg/LINA 5 mg (n=137), EMPA 10 mg/LINA 5 mg (n=136), EMPA 25 mg (n=135), EMPA 10 mg (n=134), or LINA 5 mg (n=135) for 52 weeks. Efficacy was evaluated in 667 subjects (mean [SD] age 54.6 [10.2] years; weight 87.9 [20.1] kg; BMI 31.6 [5.6] kg/m2; HbA1c 8.02% [0.96]). At week 52, adjusted mean (LOCF [SE]) changes from baseline in HbA1c were −1.17% (0.08) with EMPA 25 mg/LINA 5 mg, −1.22% (0.08) with EMPA 10 mg/LINA 5 mg, −1.01% (0.08) with EMPA 25 mg, −0.85% (0.08) with EMPA 10 mg, and −0.51% (0.08) with LINA 5 mg. Compared with LINA 5 mg, HbA1c was significantly reduced with EMPA 25 mg/LINA 5 mg (difference: −0.66% [95% CI: −0.90, −0.43; p<0.0001]) and EMPA 10 mg/LINA 5 mg (difference: −0.71% [95% CI: −0.94, −0.48; p<0.0001]). Compared with EMPA 10 mg, EMPA 10 mg/LINA 5 mg led to significant reductions in HbA1c (difference: −0.37% [95% CI −0.60, −0.14; p=0.0017]). There were no significant differences in HbA1c changes from baseline with EMPA 25 mg/LINA 5 mg vs EMPA 25 mg. In subjects with baseline HbA1c ≥7% the percentages of subjects with HbA1c <7% at week 52 were 50.4% with EMPA 25 mg/LINA 5mg, 50.8% with EMPA 10 mg/LINA 5 mg, 45.8% with EMPA 25 mg, 33.1% with EMPA 10 mg, and 27.6% with LINA 5 mg. Compared with LINA 5 mg, adjusted mean (LOCF) body weight was significantly reduced at week 52 with EMPA 25 mg/LINA 5 mg (difference: −1.7 kg [95% CI: −2.8, −0.7]; p=0.0016) and EMPA 10 mg/LINA 5 mg (difference: −1.3 kg [95% CI: −2.4, −0.2]; p=0.0172). Combinations of EMPA/LINA did not significantly reduce weight vs their respective EMPA monotherapies. Adverse events (AEs) were reported in 75.7%, 72.8%, 68.9%, 81.5% and 71.9% of subjects on EMPA 25 mg/LINA 5 mg, EMPA 10 mg/LINA 5 mg, EMPA 25 mg, EMPA 10 mg and LINA 5 mg, respectively, over 52 weeks. Confirmed hypoglycemic AEs (glucose ≤70 mg/dL and/or requiring assistance) were reported in 1 patient each on EMPA 25 mg and LINA 5 mg and 4 subjects on EMPA 10 mg; none required assistance. AEs consistent with urinary tract infection were reported in 12.5%, 15.4%, 10.4%, 16.3% and 10.4% of subjects on EMPA 25 mg/LINA 5 mg, EMPA 10 mg/LINA 5 mg, EMPA 25 mg, EMPA 10 mg and LINA 5 mg, respectively, and AEs consistent with genital infection were reported in 5.9%, 2.9%, 4.4%, 5.2% and 3.0% of subjects in these groups, respectively. In drug-naïve subjects with T2DM, EMPA 25 mg/LINA 5 mg and EMPA 10 mg/LINA 5 mg for 52 weeks significantly reduced HbA1c vs LINA 5 mg. HbA1c reductions were significant with EMPA 10 mg/LINA 5 mg vs EMPA 10 mg, but not with EMPA 25 mg/LINA 5 mg vs EMPA 25 mg. Combinations of EMPA/LINA were well tolerated, with overall safety profiles similar to those known for the individual components.

 

Disclosure: RD: Advisory Group Member, Boehringer Ingelheim, Advisory Group Member, Jansen Pharmaceuticals, Advisory Group Member, Astra Zeneca, Advisory Group Member, Novo Nordisk, Advisory Group Member, Lexicon Pharmaceuticals, Inc., Advisory Group Member, Bristol-Myers Squibb, Investigator, Jansen Pharmaceuticals, Investigator, Astra Zeneca, Investigator, Boehringer Ingelheim, Investigator, Bristol-Myers Squibb. SP: Employee, Boehringer Ingelheim. DL: Employee, Boehringer Ingelheim. RK: Employee, Boehringer Ingelheim. HJW: Employee, Boehringer Ingelheim. UCB: Employee, Boehringer Ingelheim. Nothing to Disclose: AJL

19873 3.0000 THR-642 A Empagliflozin and Linagliptin As Initial Combination for 52 Weeks in Subjects with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Sanjay Patel*1, Andrew J Lewin2, Ralph DeFronzo3, Dacheng Liu4, Renee Kaste4, Hans-Juergen Woerle5 and Uli C Broedl5
1Boehringer Ingelheim Ltd., Bracknell, United Kingdom, 2National Research Institute, Los Angeles, CA, 3Univ of Texas Hlth Science Ctr, San Antonio, TX, 4Boehringer Ingelheim Pharmaceuticals Inc, Ridgefield, CT, 5Boehringer Ingelheim Pharma GmbH & Co. KG, Ingelheim, Germany

 

A randomized, double-blind, parallel group Phase III study evaluated the efficacy and safety of combinations of empagliflozin/linagliptin (EMPA/LINA) as second-line therapy in subjects with type 2 diabetes (T2DM). Subjects were randomized to EMPA 25 mg/LINA 5 mg (n=137), EMPA 10 mg/LINA 5 mg (n=136), EMPA 25 mg (n=141), EMPA 10 mg (n=140), or LINA 5 mg (n=132) as add-on to stable-dose metformin (MET) for 52 weeks. Efficacy was evaluated in 674 subjects (mean [SD] age 56.2 [10.2] years; weight 86.1 [18.7] kg; BMI 30.9 [5.5] kg/m2; HbA1c 7.97% [0.85]). At week 52, reductions in HbA1c with EMPA/LINA were greater to those with EMPA or LINA alone. At week 52, adjusted mean (LOCF [SE]) changes from baseline in HbA1c were −1.21% (0.07) with EMPA 25 mg/LINA 5 mg, −1.05% (0.07) with EMPA 10 mg/LINA 5 mg, −0.64% (0.07) with EMPA 25 mg, −0.69% (0.07) with EMPA 10 mg and −0.48% (0.07) with LINA 5 mg. Compared with LINA 5 mg at week 52, HbA1c was reduced with EMPA 25 mg/LINA 5 mg (difference: −0.73% [95% CI −0.93, −0.53]; p<0.0001) and EMPA 10 mg/LINA 5 mg (difference: −0.57% [95% CI −0.77, −0.37]; p<0.0001). Compared with EMPA 10 mg, HbA1c was reduced with EMPA 10 mg/LINA 5 mg (difference: −0.36% [95% CI −0.56, −0.17]; p=0.0003). Compared with EMPA 25 mg, HbA1c was reduced with EMPA 25 mg/LINA 5 mg (difference: −0.57% [95% CI −0.77, −0.37]; p<0.0001). In subjects with baseline HbA1c ≥7% the percentages achieving HbA1c <7% at week 52 were 48.0% with EMPA 25 mg/LINA 5 mg, 51.6% with EMPA 10 mg/LINA 5 mg, 32.6% with EMPA 25 mg, 32.0% with EMPA 10 mg, and 28.6% with LINA 5 mg. Compared with LINA 5 mg, adjusted mean (LOCF) body weight was significantly reduced at week 52 with EMPA 25 mg/LINA 5 mg (difference: -2.9 kg [95% CI −3.8, −2.0]; p<0.0001) and EMPA 10 mg/LINA 5 mg (difference: −2.4 kg [95% CI −3.3, −1.5]; p<0.0001). Combinations of EMPA/LINA did not significantly reduce body weight vs their respective EMPA monotherapies. Adverse events (AEs) were reported in 71.5%, 69.1%, 73.0%, 68.6% and 68.9% of subjects on EMPA 25 mg/LINA 5 mg, EMPA 10 mg/LINA 5 mg, EMPA 25 mg, EMPA 10 mg and LINA 5 mg, respectively, over 52 weeks. In these groups, confirmed hypoglycemic AEs (glucose ≤70 mg/dL and/or requiring assistance) were reported in 3.6%, 2.2%, 3.5%, 1.4% and 2.3% of subjects, respectively; none required assistance. AEs consistent with urinary tract infection were reported in 10.2%, 9.6%, 13.5%, 11.4% and 15.2% of subjects on EMPA 25 mg/LINA 5 mg, EMPA 10 mg/LINA 5 mg, EMPA 25 mg, EMPA 10 mg and LINA 5 mg, respectively, and AEs consistent with genital infection were reported in 2.2%, 5.9%, 8.5%, 7.9% and 2.3% of subjects, respectively. As second-line therapy for 52 weeks in subjects with T2DM, EMPA 25 mg/LINA 5 mg and EMPA 10 mg/LINA 5 mg significantly reduced HbA1c compared with their respective monotherapies. Combinations of EMPA/LINA were well tolerated, with overall safety profiles similar to those known for the individual components.

 

Disclosure: SP: Employee, Boehringer Ingelheim. RD: Advisory Group Member, Boehringer Ingelheim, Advisory Group Member, Jansen Pharmaceuticals, Advisory Group Member, Astra Zeneca, Advisory Group Member, Novo Nordisk, Advisory Group Member, Lexicon Pharmaceuticals, Inc., Advisory Group Member, Bristol-Myers Squibb, Investigator, Jansen Pharmaceuticals, Investigator, Astra Zeneca, Investigator, Boehringer Ingelheim, Investigator, Bristol-Myers Squibb. DL: Employee, Boehringer Ingelheim. RK: Employee, Boehringer Ingelheim. HJW: Employee, Boehringer Ingelheim. UCB: Employee, Boehringer Ingelheim. Nothing to Disclose: AJL

21286 4.0000 THR-643 A Combination of Empagliflozin/Linagliptin for 52 Weeks As Add-on to Metformin in Subjects with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Molly Corbett Carr*1, June Ye2 and Jane E Reusch3
1GlaxoSmithKline, King of Prussia, PA, 2GlaxoSmithKline, Research Triangle Park, NC, 3Denver Veterans Administration Medical Center, Denver, CO

 

Background

Type 2 diabetes (T2DM) prevalence increases with age, and ~ 26% of people > 65 have T2DM.[i] Diabetes management in older adults with T2DM can be more difficult due to complex medication regimens needed to treat multiple comorbidities. For this post-hoc analysis, we hypothesized that albiglutide would be efficacious across all age groups.

Methods

In the Phase III program, albiglutide was evaluated in 1391 patients included in the pooled population of placebo (PBO) comparator studies (analyses funded by GSK): 923 patients in the albiglutide group and 468 in the PBO group. Pooled studies included patients treated with albiglutide or PBO monotherapy or in combination with metformin, thiazolidinediones or SUs (Harmony studies 1, 2, 3 and 5).The primary efficacy endpoint of the studies varied between 1 and 2 years, but the pooled efficacy analyses presented here represent data after 6 months of treatment. Demographics and baseline characteristics were representative of patients with T2DM and were, in general, well-balanced between the albiglutide and PBO groups. Treatment effects were assessed across 3 age categories (<65 years, ≥65 to <75 years, and ≥75 years). The number of elderly subjects was well represented (15% were ≥65 to <75 years of age, and 1.4% were aged ≥75), median duration of T2DM was 5.5 years, and approximately 68% had some degree of renal impairment (eGFR <90).

Results

In patients < 65 years old, change from baseline in HbA1c was -0.85% ± 0.85 in the albiglutide group (n = 757) and +0.01% ± 1.0 in the PBO group (n = 383); treatment difference = -0.87 (95%CI -0.98,-0.77). In patients ≥65 to < 75 years, change from baseline in HbA1c was -0.72 ± 0.68% in the albiglutide group (n = 136) and
-0.21 ± 0.79% in the PBO group (n = 68); treatment difference of -0.50% (95%CI -0.75,-0.26).  Although there were small numbers of patients aged ≥75 years (n = 19), HbA1c reduction was consistent with younger subgroups: change from baseline = -0.94 ± 0.68% in the albiglutide group (n = 11) and +0.06 ± 0.4% in the PBO group (n = 8) with a treatment difference of -0.97% (95%CI -1.73,-0.2). The incidence of on-therapy SAEs differed among age categories (<65, ≥65 to <75, ≥75 years) for both the albiglutide (10.5%, 14.4%, and 8.3% for the ascending age groups) and placebo (11.8%, 21.4%, and 37.5%) groups. Incidence of on-therapy AEs differed among the age categories for both the albiglutide (85%, 89%, and 75% for the ascending age groups) and placebo (82%, 83%, and 100%) groups.  

Conclusions

Efficacy and safety variables were assessed with albiglutide therapy across 3 age categories, and there were few subjects in the ≥75 year category, which limited interpretation of the data in this subgroup. The efficacy and safety profile of once weekly albiglutide was generally consistent across all age groups.

 

Disclosure: MCC: Employee, GlaxoSmithKline, Employee, GlaxoSmithKline. JY: Employee, GlaxoSmithKline, Employee, GlaxoSmithKline. JER: Advisory Group Member, GlaxoSmithKline, Researcher, BMS, Merck, Astra‐Zeneca;, Researcher, VA Merit, Denver Research Institute, ADA, CVP T32 (5 T32 HL7171‐35), CVP HL‐14985, CCTSI (UL1RR025780), and the Center for Women's Health Research).

21332 5.0000 THR-644 A Positive Treatment Effect of Once Weekly Albiglutide Across All Age Subgroups 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Susan Johnson*1, Michelle A Anderson2, Robert Enns3, Jason M Mallory4, Margaret Sowell5, Tim Wilson1 and Firas H AL-Kawas6
1GlaxoSmithKline, Research Triangle Park, NC, 2University of Michigan Hospital and Health Systems, MI, 3Pacific Gastroenterology Associates; St Paul's Hospital, Canada, 4GlaxoSmithKline, King of Prussia, PA, 5GlaxoSmithKline, Collegeville, PA, 6Johns Hopkins University; Sibley Memorial Hospital, Washington, DC

 

GLP-1 receptor agonists (GLP-1Ra) and DPP-IV inhibitors have been associated with acute pancreatitis (AP). Although the nature of the association remains unclear, labels for these agents contain precautions regarding pancreatitis.

Albiglutide (albi) is a once-weekly GLP-1Ra approved to improve glycemic control in adults with T2DM. The Phase 3 (P3) development program included 8 clinical studies evaluating the efficacy and safety of albi in a broad spectrum of patients with T2DM. The comparators included placebo (pbo) and/or active comparators (SU, TZDs, DPP-IV inhibitors, insulin or another GLP-1R agonist).  

To prospectively adjudicate suspected AP events, the sponsor (GSK) established an independent, blinded pancreatitis adjudication committee (PAC), comprised of 3 external gastroenterologists. Cases of potential pancreatitis were identified using a broad search strategy that encompassed investigator reported AEs of pancreatitis, other potentially associated AEs, and abnormal amylase and lipase values (³3 × ULN) which were routinely collected in one study.  

Classification by the PAC as definite/probable pancreatitis was consistent with clinical practice guidelines for the diagnosis of acute pancreatitis, requiring at least 2 of the following features: 1) characteristic abdominal pain, 2) serum lipase >3X ULN and/or amylase >5X ULN, and 3) characteristic findings on imaging (e.g., positive CT, MRI, or US). The PAC also assessed the relationship to study treatment considering available relevant information.

The P3 program enrolled 2365 patients on albi (4263.8 person-years [PY] of exposure; mean exposure 86.1 weeks) and 2530 treated with active or pbo comparators (4605.4 PY of exposure; mean exposure 87.0 weeks).

Across the program, the PAC adjudicated events in 11 patients treated with albi and 10 patients treated with pbo or active comparators.  In addition, there were 14 patients on albi and 9 patients on comparators with post‑randomization lipase values ³3 × ULN. Among these 44 total patients, the incidence of AP was 1.4 cases per 1000 PY in albi (6/2365 patients [0.3%]) and 0.4 cases per 1000 PY in comparator treated patients (0/468 patients on pbo and 2/2062 patients on active comparators [0.1%]); Chi-square p=0.15. The 2 active comparator cases were in patients treated with a GLP-1Ra.  For this analysis, cases of AP were those adjudicated as definite or probable pancreatitis at least possibly attributed to study drug.

In conclusion, in the albi P3 program, adjudicated cases of AP were uncommon.  The frequency of pancreatitis was higher among patients treated with albi compared to pbo and active comparators.  Of note, both cases in the active comparator group adjudicated as AP were in patients treated with a GLP-1Ra.  Within the limitations of available data, the incidence rate for AP with albi appears to be in the range described for other GLP-1Ra.

 

Disclosure: SJ: Employee, GlaxoSmithKline, Employee, GlaxoSmithKline. MAA: Consultant, GlaxoSmithKline. RE: Consultant, GlaxoSmithKline, Consultant, Jansen Pharmaceuticals. JMM: Employee, GlaxoSmithKline, Employee, GlaxoSmithKline. MS: Employee, GlaxoSmithKline, Employee, GlaxoSmithKline. TW: Employee, GlaxoSmithKline, Employee, GlaxoSmithKline. FHA: Consultant, GlaxoSmithKline.

20501 6.0000 THR-645 A Incidence of Adjudicated Acute Pancreatitis Across the Albiglutide Phase 3 Clinical Program 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Leticia Ferri* and Elise Hardy
AstraZeneca, Wilmington, DE

 

Multiple clinical studies have investigated the glucagon-like peptide-1 receptor agonist (GLP-1RA) exenatide once weekly (QW), a novel microsphere-based therapy, in patients with T2DM. In this analysis, we summarize the efficacy and safety of exenatide QW across 8 comparator-controlled, phase 3, 24–30-week clinical studies including 6 trials from the DURATION clinical program and 2 trials in solely Asian populations (1-8). Patients received exenatide QW as monotherapy or with oral antidiabetic drugs. Efficacy data were summarized while safety data were pooled across all trials. In total, 1934 patients received exenatide QW 2.0 mg; 2396 patients received comparators (non-GLP-1RAs [metformin, sitagliptin, pioglitazone, insulin glargine; n=1338]), exenatide twice daily (BID), or liraglutide. The mean age of patients was 53–58 y, and 52–68% were male. Mean duration of diabetes was 5.7–9.0 y, except in one  study in treatment-naïve patients (mean: 2.7 y). Exenatide QW reduced A1c in 7 of 8 trials with patients on stable background therapy (mean change from baseline: −1.3% to −1.9%); proportions of patients achieving A1c ≤7.0% ranged from 42–73%. Exenatide QW resulted in fasting glucose reductions ranging from −25 mg/dL to −47 mg/dL and mean body weight reductions  from −2.3 kg to −3.7 kg in 6 non-Asian studies and −1.6 kg and −1.7 kg in 2 Asian studies. Efficacy has been observed in all patient subpopulations (eg, by age, sex, baseline A1c, baseline BMI, renal function, exenatide antibody status) and therapy combinations studied to date. Overall, 71.5% of patients treated with exenatide QW experienced ≥1 treatment-emergent adverse event (AE), 4.4% discontinued due to AEs, and 3.2% had ≥1 serious AE. In patients receiving a non-GLP-1RA , 65.8% experienced ≥1 treatment-emergent AE, 2.4% discontinued due to AEs, and 4.5% had ≥1 serious AE. The most common AEs with exenatide QW were gastrointestinal issues (34.6%), which subsided over time. Nausea, diarrhea, and vomiting occurred in 14.4%, 10.5%, and 6.7% of exenatide QW patients, respectively. Minor hypoglycemia was more frequent when exenatide QW was taken with sulfonylureas (SU) than without SU (12.9% vs 2.0%). Major hypoglycemia was experienced by one patient treated with exenatide QW (without SU), one treated with exenatide BID (with SU), and two treated with insulin glargine (one with SU and one without SU). Exposure-adjusted incidence per 100 patient-years calculated for AEs of renal failure-related AEs, thyroid neoplasms, and pancreatitis, was 0.1, 0.2, and 0.5 with exenatide QW, respectively, and 0.3, 0.5, and 0.5 with non-GLP-1RA comparators, respectively. No cases of pancreatic cancer, thyroid cancer, or medullary c-cell carcinoma were reported. In conclusion, 8 studies show consistent efficacy for exenatide QW with A1c reductions of 1.3%–1.9% in patients with T2DM with no new or unexpected safety findings in the integrated population.

 

Disclosure: LF: Consultant, Astra Zeneca. EH: Employee, Astra Zeneca, Employee, Astra Zeneca.

20066 7.0000 THR-646 A Clinical Trial Results for Exenatide Once Weekly: Summary of Efficacy and Safety Data from Eight Randomized Trials of 4330 Patients with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Carol H Wysham*1, Denise Vieke2, Marion Vetter3, Yaohua He3, Nayyar Iqbal3, Elise Hardy4, Anna Ryden5 and Julio Rosenstock6
1University of Washington, 2Mercury Street Medical, Butte, MT, 3Bristol-Myers Squibb, Princeton, NJ, 4AstraZeneca, Wilmington, DE, 5AstraZeneca, Gothenburg, Sweden, 6Dallas Diabetes and Endocrine Center, TX

 

Once-weekly (QW) exenatide reduces A1C and body weight in patients with T2DM, but the original QW microsphere aqueous solution formulation requires reconstitution. A QW formulation containing exenatide dispersed in microspheres suspended in a triglyceride carrier was developed to eliminate the need for reconstitution and allow delivery via an autoinjector pen to facilitate patient education, administration, and acceptance. In the open-label DURATION-NEO-1 study, patients with inadequately controlled T2DM on diet/exercise or oral medications (metformin, sulfonylurea and/or pioglitazone) were randomized (3:2 ratio) to EQW-SAI 2 mg (n=229) or EBID 10 mcg (n=148). The primary endpoint was change from baseline in A1C at Week 28. Secondary outcomes included change from baseline in patient-reported treatment satisfaction and health-related quality of life (HRQoL) at Week 28 (exploratory analysis; nominal P-values). The Diabetes Treatment Satisfaction Questionnaire status version (DTSQs) evaluates treatment satisfaction and perceived frequencies of hyper- and hypoglycemia. Each of the 8 items is scored from 0-6, with higher scores representing greater satisfaction, except for frequency of hyper- and hypoglycemia (which do not contribute to total score [range 0-36]). The Impact of Weight on Quality of Life-Lite version (IWQOL-Lite; validated in T2DM) assesses 5 domains (physical function, self-esteem, sexual life, public distress, and work). Scores range from 0 (worst) to 100 (best). The EQW-SAI group vs the EBID group had a greater least-squares (LS) mean reduction in A1C (–1.39% vs –1.02%; P=0.007) and similar body weight reduction (–1.49 vs –1.89 kg; P=NS) at Week 28. The EQW-SAI group had significantly greater LS mean improvement than the EBID group on the DTSQs total score (+4.8 vs +2.9; P=0.002; mean score Week 28, 32.2 vs 30.2); notably, improvements in item scores for treatment convenience (+0.7 vs +0.3; P=0.002) and flexibility (+0.8 vs +0.2; P=0.0002) were greater with EQW-SAI. Scores for satisfaction with current treatment, understanding of diabetes, continuing treatment, and perceived frequencies of hyper- and hypoglycemia also improved more with EQW-SAI (all P<0.05 vs EBID). Both groups reported improvements on some IWQOL-Lite domains, with no significant between-group differences. Thus, EQW-SAI and EBID both improved patient-reported treatment satisfaction, but the simplified administration of EQW-SAI was associated with greater improvement in treatment satisfaction (including satisfaction with flexibility and convenience), potentially improving treatment acceptance, adherence and compliance.

 

Disclosure: CHW: Consultant, Boehringer Ingelheim Pharmaceuticals, Speaker, Boehringer Ingelheim Pharmaceuticals, Speaker, Jansen Pharmaceuticals, Consultant, Jansen Pharmaceuticals, Speaker, Lilly USA, LLC, Speaker, Novo Nordisk, Consultant, Sanofi, Speaker, Sanofi, Speaker, Astra Zeneca. MV: Employee, Bristol-Myers Squibb. YH: Employee, Bristol-Myers Squibb. NI: Employee, Bristol-Myers Squibb. EH: Employee, Astra Zeneca, Employee, Astra Zeneca. AR: Employee, Astra Zeneca. JR: Advisory Group Member, Sanofi, Advisory Group Member, Novo Nordisk, Advisory Group Member, Eli Lilly & Company, Advisory Group Member, GlaxoSmithKline, Advisory Group Member, Merck & Co., Advisory Group Member, Daiichi Sankyo, Advisory Group Member, Takeda, Advisory Group Member, Jansen Pharmaceuticals, Advisory Group Member, Novartis Pharmaceuticals, Advisory Group Member, Boehringer Ingelheim, Advisory Group Member, MannKind, Advisory Group Member, Halozyme, Advisory Group Member, Intarcia , Advisory Group Member, Lexicon Pharmaceuticals, Inc., Investigator, Merck & Co., Investigator, Pfizer, Inc., Investigator, Sanofi, Investigator, Novo Nordisk, Investigator, Roche Pharmaceuticals, Investigator, Bristol-Myers Squibb, Investigator, Eli Lilly & Company, Investigator, GlaxoSmithKline, Investigator, Takeda, Investigator, Novartis Pharmaceuticals, Investigator, Astra Zeneca, Investigator, Amylin Pharmaceuticals, Investigator, Jansen Pharmaceuticals, Investigator, Daiichi Sankyo, Investigator, MannKind, Investigator, Boehringer Ingelheim, Investigator, Intarcia . Nothing to Disclose: DV

19324 8.0000 THR-647 A Patient-Reported Treatment Satisfaction with Exenatide Once Weekly Suspension for Autoinjection (EQW-SAI) Vs Exenatide Twice Daily (EBID) in Patients with Inadequately Controlled Type 2 Diabetes Mellitus (T2DM) 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Maria Batool*1, Sarah Bellefeuille2, Melissa Atwood2, Thomas S Rector3 and Nacide G Ercan-Fang1
1University of Minnesota Medical Center, Minneapolis, MN, 2Minneapolis VA Health Care System, 3Minneapolis VA Health Care System, Minneapolis, MN

 

Background:

Exenatide is a glucagon-like peptide (GLP-1) receptor agonist approved for use in type 2 diabetes (T2DM) regardless of insulin use.  The purpose of this study was to evaluate whether concurrent insulin treatment affects weight, glycemic control and insulin dose in patients treated with exenatide (EXE+INS)  compared to patients on exenatide only (EXE).

Hypothesis:

Patients treated with both exenatide and insulin (EXE+INS) have less improvement in their glycemic control and lose less weight compared to patients using exenatide without insulin (EXE).

Methods:

Minneapolis Veterans Affairs Health Care System  electronic medical records reviewed from 2006 identified 154 eligible patients (Hemoglobin A1c (A1C) >9%, body mass index >35), prescribed exenatide either with (n= 83) or without (n=71) insulin. A1c, weight, basal and bolus insulin doses were collected at the beginning and end of exenatide treatment or the most recent follow up. Changes in A1c and weight in each group were compared by linear regression adjusting for influential baseline characteristics. 

Results:

Patients were mostly white (91%) males (97%). Mean age was 62 (±8) years. They were treated with EXE for a median (interquartile range) of 69 (27- 157) weeks. At baseline, the mean A1c was 9.7±1.7% and 9.5±2.3 % in the EXE+INS and EXE groups, respectively. Mean (95% confidence interval) A1C changes at the end of treatment were -0.7% (-0.2 to -1.1) and -1.3% (-0.7 to -1.8) for EXE+INS and EXE groups, respectively. Adjusting for age, baseline A1c  and duration of exenatide therapy, the decrease in mean A1c in the EXE+INS group was significantly less by 0.8% (0.2 to 1.4; p = 0.005) than the decrease in the EXE group. In the EXE+INS group, a 10 unit increase in the total daily insulin dose was associated with a 0.05% (0.01 to 0.1; p=0.01) smaller decrease in A1c.

At baseline, the mean weight was 299±50 lbs (EXE+INS) and 262±52 lbs (EXE). Mean changes in weights were -12 lbs (-7 to -17) and -10 lbs (-6 to -15) for EXE+INS and EXE groups, respectively. Adjusting for baseline weight, duration of exenatide therapy, use of weight altering medications and participation in an intensive VA weight management program , the weight loss in the EXE+INS group was less, 6.0 lbs (-0.9 to 13; p = 0.09) but not statistically significant. In the EXE+INS group, a 10 unit increase in the total daily insulin dose was not associated with less weight loss.

At baseline, the median total daily insulin dose was 114 (60 to 195) units. The median change was 0 (-32 to 40) units during exenatide treatment. The basal and bolus doses of insulin did not change significantly either.

Conclusion:

In patients with T2DM, exenatide was associated with improved glycemic control and weight loss regardless of insulin use. However, the glycemic response was attenuated as the dose of insulin increased.  Surprisingly, increasing doses of insulin did not affect ongoing weight loss.


 

Nothing to Disclose: MB, SB, MA, TSR, NGE

18947 9.0000 THR-648 A Effect of Insulin on Changes in Glycemic Control and Body Weight during Treatment with Exenatide in Patients with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Sophia Y Ali*1, Aditya Mehta2, Nitesh D Kuhadiya2, Husam Ghanim3, Antoine Makdissi3, Manav Batra4, Jeanne Hejna5, Ajay Chaudhuri2 and Paresh Dandona2
1Wayne State University, Dearborn, MI, 2Diabetes and Endocrinology Center of Western New York, Buffalo, NY, 3State University of New York at Buffalo, Buffalo, NY, 4University at Buffalo, Buffalo, NY, 5Suny at Buffalo

 

Title:  Reversal of Effects of Liraglutide As Additional Treatment to Insulin in Patients With Type 1 Diabetes After Cessation of Therapy

Abstract Body:

Objective

In the first prospectively randomized study, we have recently demonstrated that addition of 1.2mg and 1.8mg of liraglutide to insulin in patients with type 1 diabetes, significantly reduced HbA1c, mean blood glucose, insulin dose, body weight, carbohydrate intake, systolic BP and CRP over a period of 12 weeks; it also improved quality of life significantly. We have now conducted this study to demonstrate the reversal of effects of liraglutide after its discontinuation. 

Methods

Out of 72 patients randomized in this 12 weeks study, 30 patients (liraglutide group =20 and placebo=10) were followed up at 12±1 weeks after completing the study and discontinuing liraglutide.  These thirty patients (placebo=10; liraglutide = 20) with T1D for at least one year, on insulin therapy and with no detectable c-peptide in plasma (mean BMI: 30±1; mean body weight: 195±8 lbs; mean HbA1c: 7.66±0.11%; mean age: 50±2 years; mean age of T1D diagnosis: 23±2 years; 18 GAD positive, 13 GAD negative, 16 males, 14 females, 29 Caucasians, 1 African American) were randomized to receiving placebo(n=10), 0.6mg(n=8), 1.2mg(n=7) and 1.8mg(n=5) of liraglutide daily for 12 weeks. All patients in liraglutide group have been combined for the purpose of this analysis.

Results

In liraglutide group HbA1c that fell by 0.42% from 7.58±0.14% to 7.16±0.14% (p=0.004) rose by 0.63% to 7.79±0.22% (p=0.003, p=0.06 vs placebo). The average total insulin dose had fallen by 8.54 units from 62.39±4.21 to 53.85±4.69 (p=0.01), which then increased by 7.16 units to 61.01±4.42 (p=0.003,p=0.03 vs placebo). The body weight which fell by 10 lbs from 194±8 to 184±7.91 (p<0.0001) went up by 7 lbs to 191±8 (p=0.001, p=0.01 vs placebo). The daily carbohydrate intake which was reduced by 33 grams from 167±16g to 134±15g (p=0.006) went up by 21 grams to 155±13g (p=0.08). The systolic blood pressure (SBP) which was reduced by 10 mmHg from 125±2.88 to 115±3.54mmHg (p=0.0003) went up by 9mmHg to 124±2.8mmHg (p=0.07). In 1.8 mg group, SBP which had fallen by 20 mmHg from 123±5 to 103±6 (p=0.016) went up by 21 mmHg to 124±4 (0.004). There was no change in any of these indices in the placebo group.

Conclusion

Our data demonstrate the near total reversal of the effects of liraglutide on glycemia, body weight, carbohydrate intake and systolic blood pressure in patients with type 1 diabetes three months after cessation of liraglutide therapy. Thus, while the addition of liraglutide therapy benefits these patients dramatically, it needs to be continued in order to maintain its benefits. These findings have important implications for future treatments of patients with type 1 diabetes.

 

Nothing to Disclose: SYA, AM, NDK, HG, AM, MB, JH, AC, PD

21975 10.0000 THR-649 A Reversal of Effects of Liraglutide As Additional Treatment to Insulin in Patients with Type 1 Diabetes after Cessation of Therapy 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Alaaeldin Bashier1, Azza Abdulaziz Bin Hussain*2, Elamin Abdelgadir2, Ahmed Altinay2, Puja Murli Thadani3, Fatheya Fardallah Alawadi4 and Salah Abusnana5
1Dubai Hospital, Dubai Health Authority, Dubai, United Arab Emirates, 2Dubai Hospital, Dubai Health Authority, 3Dubai Hospital, Mumbai, 4Dubai Hospital, Dubai Health Authority, Ash Shariqah, United Arab Emirates, 5Rcdr Centre. Ajman, UAE.

 

Background: The pathophysiology of type2 diabetes differs between different ethnic groups, with Asians developing type2 diabetes at younger age, at lower body mass index, and in relatively short time. Not only that, some ethnicities has different responses and dosing regimens to different classes of anti-diabetic agents. Data from Japanese population showed that the optimal doses used are smaller than other population and that weight loss is not as effective as seen in Caucasians. Objective: we aimed to assess liraglutide efficacy in reducing weight and HbA1c in Arab population when used as add on to other anti-diabetic agents. Methodology: we prospectively followed patients who were recruited to treatment with liraglutide for a six months period, at the start of the study we checked patients demographics, weight, BP, fasting blood glucose, HbA1c, lipid panel, LFTs and creatinine. Patients were checked at 3 months and at the end of the study at 6 months. Results: mean weight was 96.01±19.2; after 3 months of starting Liraglutide the mean weight reduction was highly significant (94.8±20 with (P< 0.001)). At 6 months the mean weight was 94.5±19 with (p<0.001). Mean HbA1c at baseline was 8.3±1.7 dropped to 7.7±1.4 (p<0.001), and 7.6+1.6  (p<0.001) at 3 and 6 months respectively. Conclusion: Liraglutide is effective in reducing weight and HbA1c as well as other metabolic parameters in Arab population with type2 diabetes.

 

Nothing to Disclose: AB, AAB, EA, AA, PMT, FFA, SA

19564 11.0000 THR-650 A Liraglutide Effect in Reducing Hba1c and Weight in Arab Population with Type2 Diabetes, a Prospective Observational Trial. Short Title: Lead-Arab Trial 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Kristine Santos de Luna*1, Erick Sta. Rosa Mendoza2 and Leilani Basa Mercado-Asis2
1University of Santo Tomas Hospital, Manila, Philippines, 2University of Santo Tomas, Manila City, Philippines

 

INTRODUCTION
Liraglutide, a glucagon-like peptide 1 agonist, stimulates insulin and suppresses glucagon secretion producing significant reductions in blood glucose and beneficial effects on blood pressure. This drug is used in combination with other anti-diabetic medications or as monotherapy because of its low risk for hypoglycemia and moderate weight loss. In a developing country like the Philippines, this efficacy may be limited by its cost leading to noncompliance and treatment failure. 

METHODOLOGY
This is a observational study of patients with type 2 diabetes mellitus who were on liraglutide for 12 months. Weight, body mass index (BMI), blood pressure (BP) and glycosylated hemoglobin (HbA1C) were the primary parameters determining the efficacy of the drug. The measurements were obtained at baseline and at follow-up after 3, 6 and 12 months. Descriptive statistics and measures of proportions were used in the analysis.

RESULTS
A total of 16 patients on liraglutide were included in the study. The age range was 43 to 72 years (6 males and 6 females). The mean duration of diabetes was 8.4 years. Seven patients were on insulin and all were taking one or more oral anti-diabetic drugs. The baseline weight, BMI, BP and HbA1C were 85.0 kg (74.5 to 95.0), 34.8 kg/m2 (28.3 to 49.3) 128/72 mm Hg (110 to160/66 to 83) and 7.6% (5.2 to13.0), respectively. The reduction from baseline in weight (14% at 3 moths and 57% at 6 months) and BMI (21% at 3 months and 69% at 6 months) were sustained. A sustained reduction in HbA1C from baseline was also observed (19% and 52% at 3 and 6 months, respectively). There was also a modest decrease in blood pressure. However, a rebound increase at 12 months in weight (85.1 kg), BMI (32.1 kg/m2), BP (137/81 mm Hg) and HbA1C (7.15%) comparable to baseline levels was observed. The main reason for the deterioration among these parameters were poor compliance (n=11) or discontinuation (n=5) of the drug because of the cost.

CONCLUSION
Liraglutide is an attractive armamentarium for glucose control among diabetic patients with added weight loss and favorable blood pressure control. However, the sustainability of efficacy is limited by cost especially in a developing country. Compliance may be improved by emphasis on its benefits outweighing the cost of hospitalization for possible future complications.

 

Nothing to Disclose: KSD, ESRM, LBM

21654 12.0000 THR-651 A Sustainability of Weight Loss and Glycemic Control with Liraglutide Is Limited By Cost and Compliance in a Developing Country: A One-Year Observational Study 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Bruno Guerci*1, David A D'Alessio2, Carol H Wysham3, Nan Jia4 and Fady T Botros5
1Hôpital Brabois Adultes, 2Duke University, Durham, NC, 3Rockwood Center for Diabetes and Endocrinology, Spokane, WA, 4Lilly USA, LLC, Indianapolis, IN, 5Eli Lilly and Company

 

Dulaglutide (DU) is a once weekly glucagon-like peptide-1 (GLP-1) receptor agonist that has been recently approved by the US Food and Drug Administration for the treatment of type 2 diabetes. Glycemic efficacy of DU has been demonstrated in six Phase 3 studies. The objective of this analysis is to determine major baseline factors that are associated with the reduction in glycosylated hemoglobin A1c (A1C) in response to treatment with DU and to evaluate the role of baseline glycemic factors.

DU patient (pt) covariates from across the 6 DU Phase 3 clinical trials were analyzed using gradient boosting analysis to assess their relative contribution to the change in A1C at 26 weeks of treatment. Glycemic variables which had the greatest amount of influence on A1C change were further evaluated in univariate and multivariate modeling to assess the magnitude and direction of impact. Analyzed baseline factors included age, race, ethnicity, gender, duration of diabetes, serum glucose measures, fasting serum C-peptide and insulin (FSI), weight, BMI, cardiovascular disease history, estimated glomerular filtration rate (eGFR), and albuminuria.

This analysis included 2806 DU treated pt with mean age 56.3 ± 9.8 years, duration of diabetes 7.8 ± 6.0 years, baseline A1C 8.0 ± 1.0%, BMI 32.4 ± 5.2 kg/m2, eGFR 89.5 ± 16.9 mL/min/1.73m2. Based on gradient boosting, the top 5 baseline factors associated with reduction in A1C (and their relative influence) are baseline A1C (48.8%), age (9.1%), fasting serum glucose (FSG) (8.2%), FSI (6.7%), and eGFR (5.4%). The role of baseline glycemic factors (A1C, FSG, FSI) was evaluated. Univariate regression demonstrated that higher baseline A1C and FSG values and lower FSI values were associated with greater reduction in A1C (coefficient estimates of A1C change: -0.598%, -0.101%, and 0.002%, respectively, p<0.0001 for all). Multivariate regression showed similar results for baseline A1C (-0.594 ± 0.027%) and FSI (-0.197 ± 0.045% for FSI ≤55 pmol/L), but greater baseline FSG was associated with smaller decrease in A1C when adjusted for the other 4 factors (0.047 ± 0.010%, p<0.0001). Subgroup analysis by baseline A1C confirmed that higher FSG is associated with smaller decreases in A1C within a subgroup of pts with comparable baseline A1C. 

In conclusion, these data indicate that baseline factors associated with poorer glycemic status (higher A1C and lower FSI) are associated with greater reduction in A1C in response to treatment with DU.

 

Disclosure: BG: Consultant, Abbott Laboratories, Consultant, Menarini Diagnostic, Consultant, Medtronic Minimed, Consultant, Roche Diagnostics, Consultant, MSD, Consultant, Pfizer, Inc., Consultant, Metacure, Consultant, Intarcia, Consultant, Jansen Pharmaceuticals, Consultant, Boehringer-Ingelheim, Consultant, Eli Lilly & Company, Consultant, Novo Nordisk, Consultant, Novartis Pharmaceuticals, Consultant, GlaxoSmithKline, Consultant, Sanofi, Consultant, Bristol-Myers Squibb, Consultant, Vitalaire, Consultant, Dinno Sante, Consultant, Orkyn. DAD: Consultant, Intarcia, Consultant, Boehringer-Ingelheim , Consultant, Jansen Pharmaceuticals, Consultant, Eli Lilly & Company, Consultant, Merck & Co., Consultant, Novo Nordisk. CHW: Consultant, Astra Zeneca, Consultant, Boehringer-Ingelheim, Consultant, Eli Lilly & Company, Consultant, Jansen Pharmaceuticals, Consultant, Novo Nordisk, Consultant, Sanofi, Investigator, Intarcia , Investigator, Merck & Co.. NJ: Employee, Eli Lilly & Company, Employee, Eli Lilly & Company. FTB: Employee, Eli Lilly & Company, Employee, Eli Lilly & Company.

19892 13.0000 THR-652 A Baseline Glycemic Factors Are Associated with the Glycemic Response to Treatment with Once Weekly Dulaglutide in Patients with Type 2 Diabetes 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Jaime A Davidson*1, Guntram Schernthaner2, Laura Hieronymus3, Holly Jodon4, Ujjwala Vijapurkar5, Gary Meininger5 and William Canovatchel5
1Touchstone Diabetes Center, The University of Texas Southwestern Medical Center, Dallas, TX, 2Rudolfstiftung Hospital-Vienna, Vienna, Austria, 3DiabetesCare and Communications, Lexington, KY, 4Metabolic Disease Associates, Erie, PA, 5Janssen Research & Development, LLC, Raritan, NJ

 

Many patients with type 2 diabetes mellitus (T2DM) are obese or overweight, so treatments that improve glycemic control and reduce body weight may be beneficial.  Canagliflozin (CANA), a sodium glucose co-transporter 2 (SGLT2) inhibitor approved for the treatment of T2DM, inhibits renal glucose reabsorption by lowering the renal threshold for glucose (RTG) and increasing urinary glucose excretion (UGE), resulting in lowered plasma glucose levels and a net caloric loss.  CANA has been shown to provide reductions in A1C and body weight compared with sitagliptin (SITA) in patients with T2DM inadequately controlled on metformin (MET) or MET plus sulfonylurea (SU; CANA 300 mg only) in two 52-week, randomized, double-blind, active-controlled, Phase 3 studies.  This analysis evaluated the proportion of patients who responded to treatment in terms of both A1C and body weight.  In Study 1 (N = 1,284; mean baseline A1C, 7.9%; body weight, 87.2 kg), least squares (LS) mean changes from baseline in A1C at Week 52 were –0.73%, –0.88%, and –0.73% with CANA 100 and 300 mg and SITA 100 mg, respectively; LS mean percent changes in body weight were –3.8%, –4.2%, and –1.3%, respectively.  In Study 2 (N = 755; mean baseline A1C, 8.1%; body weight, 88.3 kg), LS mean changes from baseline in A1C were –1.03% and –0.66% at Week 52, and LS mean percent changes in body weight were –2.5% and 0.3% with CANA 300 mg and SITA 100 mg, respectively.  In Study 1, the proportion of patients with reductions in A1C were 77.8%, 84.7%, and 80.5% with CANA 100 and 300 mg and SITA 100 mg, respectively; 82.7%, 84.7%, and 58.8% of patients had reductions in body weight.  A greater proportion of patients in Study 1 had reductions in both A1C and body weight with CANA 100 and 300 mg versus SITA 100 mg (67.7%, 74.7%, and 50.6%, respectively; differences [95% confidence interval (CI)] of 17.1% [9.7, 24.5] and 24.2% [17.1, 31.3], respectively).  In Study 2, the proportion of patients with reductions in A1C with CANA 300 mg and SITA 100 mg were 86.6% and 74.2%, respectively; 78.6% and 47.1% of patients had reductions in body weight.  Similar to Study 1, a greater proportion of patients in Study 2 had reductions in both A1C and body weight with CANA 300 mg (70.6%) versus SITA 100 mg (37.8%; difference [95% CI] of 32.8% [25.7, 39.8]).  The overall incidence of adverse events (AEs) was similar across treatment groups in both studies; CANA was associated with increased rates of genital mycotic infections and osmotic diuresis–related AEs compared with SITA.  In Study 1, the incidence of documented hypoglycemia (≤70 mg/dL) was 7%, 7%, and 4% with CANA 100 and 300 mg and SITA 100 mg, respectively; in Study 2, the incidence of documented hypoglycemia was 43% and 41% with CANA 300 mg and SITA 100 mg, respectively.  In conclusion, CANA provided greater attainment of reduction in both A1C and body weight compared with SITA at 52 weeks and was generally well tolerated in patients with T2DM as add-on to MET or MET plus SU.

 

Disclosure: JAD: Advisory Group Member, Amgen, AstraZeneca, Eli Lilly, Janssen, Merck, Novo Nordisk, and Aspire Bariatrics, Consultant, Amgen, AstraZeneca, Eli Lilly, Janssen, Merck, Novo Nordisk, and Aspire Bariatrics, Speaker, AstraZeneca, Janssen, and Novo Nordisk. LH: Speaker, Janssen. HJ: Speaker, Janssen. UV: Employee, Janssen Research & Development, LLC. GM: Employee, Janssen Research & Development, LLC. WC: Employee, Janssen Research & Development, LLC. Nothing to Disclose: GS

21070 14.0000 THR-653 A Canagliflozin Is Superior to Sitagliptin in Reducing Both A1C and Body Weight in Patients with Type 2 Diabetes Mellitus 2015-03-08 San Diego 2015-03-02 97th Annual Meeting of the Endocrine Society Diabetes & Glucose Metabolism Thursday, March 5th 3:00:00 PM THR 640-672 5935 1:00:00 PM Type 2 Diabetes Therapies Poster


Farid Saad*1, Gheorghe Doros2, Abdulmaged M Traish3 and Aksam A Yassin4
1Bayer Pharma AG, Berlin, Germany, 2Boston University School of Public Health, Boston, MA, 3Boston University School of Medicine, Boston, MA, 4Segeberger Kliniken, Norderstedt, Germany

 

Introduction and Objectives:

In men with T2DM, achieving and maintaining stable glycaemic control remains challenging. Several short-term studies on the use of testosterone (T) in men with T2DM have shown mixed results, with longer-term studies demonstrating significantly better results than short-term studies. We investigated effects of long-term treatment with TU in 77 hypogonadal men with T2DM.

Methods:

Single-center, cumulative, prospective, registry study of 262 hypogonadal men. 77 men (29.4%) had T2DM. All patients had presented with ED and were found hypogonadal (total T 12 nmol/L (~350 ng/dl). All men received TU for up to 11 years. In almost all men, T2DM had been diagnosed previously and was being treated by their respective family physicians. Patients were enrolled into the registry once they had received 1 year of TU. 77 men were treated for a minimum of 2 years, 76 for 3 years, 75 for 4 years, 73 for 5 years, 56 for 6 years, 23 for 7 years, 18 for 8 years, 17 for 9 years, and 7 for 10 years. Measures were taken at every other visit. Results at the end of 10 years follow-up are reported.

Results:

Mean age was 61.54 ± 7.28 years (minimum: 41; maximum: 74 years). 9 of 77 men (12%) were overweight, and 68 (88%) were obese.

Total T increased from 7.59 ± 1.89 nmol/L to trough levels (measured prior to the following injection) between 17 and 20 nmol/L, free T from 150.31 ± 65.24 to 400-500 pmol/L.

Waist circumference decreased from 115.03 ± 10.61 to 96.43 ± 4.72 cm. Weight decreased from 109.52 ± 13.01 to 86 ± 8.98 kg. Mean BMI decreased from 34.66 ± 3.87 to 27.54 ± 2.61 kg/m2 (p<0.0001 vs baseline, statistical significance vs previous year for the first 6 years for all). Weight reduction was -18.97%.

Fasting glucose decreased from 146.26 ± 41.48 to 76.21 ± 5.34 mg/dl (p<0.0001 vs baseline). HbA1c declined progressively from 7.87 ± 1.05 to 7.17 ± 0.96 after 1 year, 6.76 ± 0.92 after 2 years, 6.54 ± 0.9 after 3 years, 6.35 ± 0.85 after 4 years, 6.17 ± 0.66 after 5 years, 6.13 ± 0.76 after 6 years, to 5.69 ± 0.33% after 10 years (p<0.0001 vs baseline, statistical significance vs previous year for the first 5 years). At baseline, 16 patients (20.8%) were within the HbA1c target of 7.0%, 9 (10.7%) within 6.5%. At the last observation for each individual patient, 70 men (90.9%) were within the HbA1c target of 7.0%, 57 (74.0%) within 6.5%. Only 7 patients (9.1%) did not reach either of these HbA1c targets.

The triglyceride:HDL ratio as a surrogate marker of insulin resistance decreased from 8.05 ± 4.29 to 2.67 ± 0.39 (p<0.0001 vs baseline). The major decrease occurred during the first 3 years with statistical significance each year vs previous year.

No patient dropped out.

Conclusions:

Long-term T therapy improved glycaemic control in hypogonadal men with T2DM. The majority of patients achieved the HbA1c targets previously not achieved on standard therapy. Increased lean mass and sustained weight loss may have contributed to the observed improvements.

 

Disclosure: FS: Employee, Bayer Schering Pharma. GD: statistical analyses, Bayer Schering Pharma. AAY: Investigator, Bayer Schering Pharma. Nothing to Disclose: AMT

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