Chimeric antigen receptor-modified human regulatory T cells that constitutively express IL-10 maintain their phenotype and are potently suppressive.

Clinical trials of regulatory T cell therapy in transplantation are currently entering phases IIa and IIb, with the majority of these employing polyclonal Treg populations which harbour a broad specificity. Enhancing Treg specificity is possible with the use of chimeric antigen receptors (CAR), which can be customized to respond to a specific human leukocyte antigen (HLA). In this study we build on our previous work in the development of HLA-A2 CAR-Tregs by further equipping cells with the constitutive expression of interleukin 10 (IL-10) and an imaging reporter as additional payloads. Cells were engineered to express combinations of these domains and assessed for phenotype and function. Cells expressing the full construct maintained a stable phenotype after transduction, were specifically activated by HLA-A2 and suppressed alloresponses potently. The addition of IL-10 provided an additional advantage to suppressive capacity. This study therefore provides an important proof-of-principle for this cell engineering approach for next-generation Treg therapy in transplantation.