Discovering Thyroid Stimulating Hormone Receptor (TSHR) T-Cell Epitopes in Autoimmune Thyroiditis

Presentation Number: OR11-1
Date of Presentation: March 5th, 2015

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

Abstract

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