Harnessing CD3 diversity to optimize CAR T cells

Current US Food and Drug Administration-approved chimeric antigen receptor (CAR) T cells harbor the T cell receptor (TCR)-derived zeta chain as an intracellular activation domain in addition to costimulatory domains. The functionality in a CAR format of the other chains of the TCR complex, namely CD3 delta, CD3 epsilon and CD3 gamma, instead of zeta, remains unknown. In the present study, we have systematically engineered new CD3 CARs, each containing only one of the CD3 intracellular domains. We found that CARs containing CD3 delta, CD3 epsilon or CD3 gamma cytoplasmic tails outperformed the conventional zeta CAR T cells in vivo. Transcriptomic and proteomic analysis revealed differences in activation potential, metabolism and stimulation-induced T cell dysfunctionality that mechanistically explain the enhanced anti-tumor performance. Furthermore, dimerization of the CARs improved their overall functionality. Using these CARs as minimalistic and synthetic surrogate TCRs, we have identified the phosphatase SHP-1 as a new interaction partner of CD3 delta that binds the CD3 delta-ITAM on phosphorylation of its C-terminal tyrosine. SHP-1 attenuates and restrains activation signals and might thus prevent exhaustion and dysfunction. These new insights into T cell activation could promote the rational redesign of synthetic antigen receptors to improve cancer immunotherapy. Minguet and colleagues systematically examine how individual CD3 chains of the TCR-CD3 complex can improve chimeric antigen receptor (CAR) T cell performance.