Dual CD8 and TCR editing in regulatory T cells mediates HLA-A2-restricted tissue-specific homing

Type 1 diabetes (T1D) is marked by the overexpression of class I major histocompatibility complex (MHC) antigens in pancreatic islets, which are targeted by islet-specific CD8⁺ T cells. Here, we aimed to improve regulatory T cell (Treg) infiltration into pancreatic islets by redirecting their specificity toward class I-restricted islet antigens. We functionally validated two public islet specific HLA-A2 (*02:01) restricted TCRs, one specific for ZnT8₁₈₆₋₁₉₄ (clone D222D), the second for IGRP₂₆₅₋₂₇₃ (clone 32) by dual locus (TRAC/CD4) homology-directed editing. Clone D222D was peptide-specific and CD8αβ dependent while clone 32 exhibited antigen promiscuity and showed CD8α dependency. Engineered CD4to8 TCR Tregs maintained stable phenotypes, in vitro suppressive function compared to their polyclonal counterparts, and showed co-receptor-dependent migration in vivo. This approach demonstrates that TCR specificity, reflected by its functional activity, is crucial for tissue-specific trafficking, paving the way to improve the efficacy of Treg therapies for T1D.