A microenvironment-driven HLA-II-associated insulin neoantigen elicits persistent memory T cell activation in diabetes

The antigenic landscape of autoimmune diabetes reflects a failure to preserve self-tolerance. Yet, how novel neoantigens emerge in humans remains incompletely understood. Here, we designed an immunopeptidomics-based approach to probe HLA-II-bound, islet-derived neoepitopes in patients with type 1 diabetes (T1D). We uncovered a Cys→Ser transformation, conserved between mice and humans, that reshapes autoreactivity to insulin at the single-residue level. This transformation, which we call “C19S,” arises from oxidative remodeling of insulin in stressed pancreatic islets and also occur in cytokine-activated antigen-presenting cells, contributing to a feed-forward loop of neoepitope formation and presentation. Despite involving just one amino acid, C19S is recognized by HLA-DQ8-restricted, register-specific CD4+ T cells that expand at diabetes onset. These neoepitope-specific CD4+ T cells lack regulatory potential but acquire a poised central memory phenotype that persists across disease progression. These findings reveal a distinct, microenvironment-driven route of neoantigen formation that fuels sustained autoreactivity in diabetes.