B cell directed CAR-T cell therapy results in activation of CD8+ cytotoxic CAR-negative bystander T cells in both non-human primates and patients

Kaminski, J.; Fleming, R. A.; Alvarez-Calderon, F.; McGuckin, C.; Ho, E. E.; Eng, F.; Rui, X.; Keskula, P.; Cagnin, L.; Charles, J.; Zavistaski, J.; Margossian, S. P.; Kapadia, M. A.; Rottman, J. B.; Lane, J.; Baumeister, S. H. C.; Tkachev, V.; Shalek, A. K.; Kean, L. S.; Gerdemann, U.

doi:10.1101/2023.09.28.559936

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bioRxiv · 10.1101/2023.09.28.559936

B cell directed CAR-T cell therapy results in activation of CD8+ cytotoxic CAR-negative bystander T cells in both non-human primates and patients

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Abstract

There is growing appreciation for the emergence of CARneg bystander T cells after CAR-T cell infusion. However, their phenotypic and transcriptomic hallmarks and mechanisms of activation remain uncertain. We performed single-cell RNA-Seq (scRNA-Seq) on non-human primate (NHP) and patient-derived T cells to interrogate CARneg T cells following B cell targeted CAR-T cell therapy. In a NHP model, we observed a distinct population of activated CD8+ CARneg T cells emerging during CAR-T cell expansion. These bystander CD8+ CARneg T cells exhibited a unique transcriptional signature with upregulation of NK-cell markers (KIR3DL2, CD160, KLRD1), chemokines and chemokine receptors (CCL5, XCL1, CCR9), and downregulation of naive T cell-associated genes (SELL, CD28). A transcriptionally similar population was identified in patients following Tisangelecleucel infusion. Mechanistic studies revealed that IL-2 and IL-15 exposure induced bystander-like CD8+ T cells. These T cells efficiently killed leukemic cells through a TCR-independent mechanism. Together, these data identify bystander CD8+ T cells as a novel mechanism by which CAR-T cell infusion can induce further anti-leukemic activity, measurable in both NHP and in patients. Statement of SignificanceWe have deeply interrogated CARneg bystander CD8+ T cells during CAR-T cell expansion in non{-}human primates and patients receiving Tisangelecleucel to identify the unique transcriptomic signature defining these cells, and to determine that IL-2-and IL-15-induced cytotoxic bystander T cells are capable of killing in a TCR-independent manner. These data highlight the potential of bystander T cells for leukemia control and provide a critical foundation for their future analysis.

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Kaminski, J., Fleming, R. A., Alvarez-Calderon, F., McGuckin, C., Ho, E. E., Eng, F., Rui, X., Keskula, P., Cagnin, L., Charles, J., Zavistaski, J., Margossian, S. P., Kapadia, M. A., Rottman, J. B., Lane, J., Baumeister, S. H. C., Tkachev, V., Shalek, A. K., Kean, L. S., Gerdemann, U.. 2023-09-29. B cell directed CAR-T cell therapy results in activation of CD8+ cytotoxic CAR-negative bystander T cells in both non-human primates and patients. https://doi.org/10.1101/2023.09.28.559936

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