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TCR activation impairs CAR-T cytotoxicity against separate target cells

Moosmann, C.; Drost, F.; Kristensen, N. P.; Boettcher, M.; Pavlova, A.; Hudecek, M.; Reinecke, V.; Bruns, H.; Voelkl, S.; Mackensen, A.; Busch, D. H.; Schubert, B.; Mougiakakos, D.; Schober, K.

2025-10-07 immunology
10.1101/2025.10.07.680634 bioRxiv
Show abstract

Chimeric antigen receptor T cells (CAR-T) are effective therapeutics against cancer and autoimmunity, but whether the endogenous T-cell receptor (TCR) is beneficial, detrimental or irrelevant for CAR-T function and patient outcome remains unclear. 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-sequencing. A cytotoxic phenotype, but not CAR-mediated in vitro 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 gradually 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.

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