Unbiased discovery of autoreactive type 1 diabetes T-cell receptors that bind specific hybrid insulin peptides.

While recent studies have revealed much about the pathogenesis of type 1 diabetes (T1D), the self-antigens and immune receptors that drive cellular autoimmunity have not been precisely determined. In this study, we identify disease-associated T-cell receptor (TCR) sequences, provide evidence that they are involved in T1D pathogenesis, and connect them to specific antigens implicated in beta-cell autoimmunity. We discovered 264 T1D-associated TCRs by comparing TCR repertoires in blood from T1D cases and controls, without any bias toward predefined cell phenotypes or antigens. Multiple lines of evidence support the relevance of these TCRs to T1D pathogenesis: they form convergent sequence clusters linked to the class II HLA risk alleles HLA-DQ8 and HLA-DQ2.5, are found almost exclusively in T1D cases and largely absent in matched controls, can be detected in blood before autoantibodies, have baseline frequencies that stratify post-treatment clinical outcomes, and are enriched in disease-relevant tissues with effector phenotypes; by contrast, the same TCRs, when rarely detected in healthy controls, more often exhibit regulatory phenotypes. We reverse engineered (deorphanized) these TCRs to find their antigenic targets using nucleic acid-based and peptide-based workflows. These independent approaches converged on a narrow set of antigen targets across TCR clusters: a C-peptide-derived hybrid insulin peptide (HIP) hotspot, with representative receptors preferring HIPs over native peptides and remaining specific in proteome-scale testing. Together, these findings connect multiple threads from the T1D literature: the central importance of effector T cells to disease pathogenesis; the role of insulin-derived hybrid peptides as neoantigens; and the potential to use public clusters of TCRs to serve as biomarkers, guide antigen discovery efforts, and elucidate disease mechanisms.