Engineering Tregs for autoimmune diabetes therapy - SUMMARY Type 1 diabetes (T1D) is an autoimmune disease caused by the immune mediated destruction of insulin-producing beta cells, significantly impacting human health. Tolerance to beta cells is normally maintained by regulatory T cells (Tregs). In T1D, Tregs stop protecting beta cells and leave them vulnerable to immune attack. Work in T1D models has shown that therapeutic restoration of Tregs can prevent disease progression, and clinical studies have shown the safety of this approach in humans. Despite these successes, infusion of polyclonal Tregs has limited clinical efficacy, representing key knowledge gaps. These challenges could be solved by creating “designer” Tregs that are engineered for islet antigen specificity. If successful, this technology would lead to a more personalized and effective T1D immunotherapy. We recently developed an efficient way to generate monoclonal antibodies that recognize peptides bound to MHC molecules. Using this approach we created a series of antibodies that are specific for islet-reactive peptide-MHCI and peptide-MHCII complexes, called ‘TCR-mimics’, relevant for autoimmunity in NOD mice and humans. We used these reagents as chimeric antigen receptors (CAR) to develop Tregs so they were antigen-specific. We demonstrated these CAR Tregs were specific and elicited suppressive functions. In preliminary studies we have shown in vivo feasibility by preventing autoimmune diabetes in mice. Our central hypothesis is that re-directing the specificity of Tregs can be harnessed to prevent and treat T1D. We hypothesize that engineered p:MHC CAR Tregs will provide superior suppression relative to TCR Tregs in autoimmune diabetes. We will test three specific aims; 1) Test efficacy of MHCI and MHCII CAR Treg-mediated suppression in vitro and in vivo, 2) Determine the mechanisms and differential gene expression mediating CAR Treg suppression, and 3). Evaluate synergy between CAR Tregs and anti-CD3 therapy in autoimmune diabetes. Our innovative approach, using p:MHC mAbs to generate CAR Tregs targeting islet antigens allows us to determine if engineered antigen-specific Treg will be optimal to prevent or reverse T1D pathogenesis. The results from this study will also determine the mechanism(s) for suppression for these engineered Tregs. Finally we will compare MHCI- or MHCII- restricted CAR Tregs and determine if both are required for optimal diabetes therapy either alone or in combination with anti-CD3 therapy. We are applying our knowledge to human CAR Tregs to test in preclinical models and are making additional CAR Tregs monthly. We strongly believe CAR Tregs are the next T1D therapy, and the time to focus on this therapeutic option is now.
