Wang, Z.; Sarkar, A.; Ge, X.

Chimeric antigen receptors (CAR) that mimic T cell receptors (TCR) on eliciting peptide-major histocompatibility complex (pMHC) specific T cell responses hold great promise in the development of immunotherapies against solid tumors, infections, and autoimmune diseases. However, broad applications of TCR-mimic (TCRm) CARs are hindered to date largely due to lack of a facile approach for the effective isolation of TCRm CARs. Here, we establish a highly efficient process for de novo discovery of TCRm CARs from human naive antibody repertories by combining recombinase-mediated large-diversity monoclonal library construction with T cell activation-based positive and negative screenings. Panels of highly functional TCRm CARs with peptide-specific recognition, minimal cross-reactivity, and low tonic signaling were rapidly identified towards MHC-restricted intracellular tumor-associated antigens MAGE-A3, NY-ESO-1, and MART-1. Transduced TCRm CAR-T cells exhibited pMHC-specific functional avidity, potent cytokine release, and efficacious and persistent cytotoxicity. The developed approach could be used to generate safe and potent immunotherapies targeting MHC-restricted antigens.