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.

doi:10.1101/2025.10.07.680634

bioRxiv · 10.1101/2025.10.07.680634

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.

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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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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. TCR activation impairs CAR-T cytotoxicity against separate target cells. https://doi.org/10.1101/2025.10.07.680634

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