TCR activation impairs CAR-T cell function in heterogeneous target cell contexts

Chimeric antigen receptor (CAR) T cells are powerful therapeutics against cancer and autoimmunity, but whether the endogenous T cell receptor (TCR) is beneficial, detrimental or irrelevant for CAR‑T function remains unclear. TCRs of CAR T cells are used as natural barcodes for clonotype tracking, but may also cause graft-versus-host disease. Reversely, virus-specific TCRs could enhance CAR‑T efficacy. We here traced anti-CD19 CAR‑T clonotypes in patients with B‑cell malignancies pre- and post-infusion using single-cell RNA-, TCR-, and CITE-seq. A cytotoxic phenotype, but not ex vivo [KS1] reactivity to tumor cells, predicted CAR‑T persistence. To test the functional impact of endogenous TCR activity on CAR‑T behavior, we combined CAR transduction with orthotopic TCR replacement. This revealed that TCR signaling adds to activation of CAR T cells, but compromises CAR-mediated cytotoxicity, when TCR and CAR antigens are presented by different target cells. Therefore, spatial antigen separation alters TCR/CAR interplay with implications for therapeutic CAR‑T design. [KS1]Would in vitro be easier to understand than ex vivo? If yes, we should change it consistently.