Proinsulin multi‐peptide immunotherapy induces antigen‐specific regulatory T cells and limits autoimmunity in a humanized model

Summary Peptide immunotherapy (PIT) is a targeted therapeutic approach, involving administration of disease‐associated peptides, with the aim of restoring antigen‐specific immunological tolerance without generalized immunosuppression. In type 1 diabetes, proinsulin is a primary antigen targeted by the autoimmune response, and is therefore a strong candidate for exploitation via PIT in this setting. To elucidate the optimal conditions for proinsulin‐based PIT and explore mechanisms of action, we developed a preclinical model of proinsulin autoimmunity in a humanized HLA‐DRB1*0401 transgenic HLA‐DR4 Tg mouse. Once proinsulin‐specific tolerance is broken, HLA‐DR4 Tg mice develop autoinflammatory responses, including proinsulin‐specific T cell proliferation, interferon (IFN)‐γ and autoantibody production. These are preventable and quenchable by pre‐ and post‐induction treatment, respectively, using intradermal proinsulin‐PIT injections. Intradermal proinsulin‐PIT enhances proliferation of regulatory [forkhead box protein 3 (FoxP3 + )CD25 high ] CD4 T cells, including those capable of proinsulin‐specific regulation, suggesting this as its main mode of action. In contrast, peptide delivered intradermally on the surface of vitamin D3‐modulated (tolerogenic) dendritic cells, controls autoimmunity in association with proinsulin‐specific IL‐10 production, but no change in regulatory CD4 T cells. These studies define a humanized, translational model for in vivo optimization of PIT to control autoimmunity in type 1 diabetes and indicate that dominant mechanisms of action differ according to mode of peptide delivery.