SRP303844 PRJNA697929 GSE165797 Crosstalk between CAR T cell subsets and the tumor microenvironment is essential for sustained cytotoxic activity Chimeric antigen receptor (CAR) T cell therapy relies on the activity of a large pool of tumor-targeting cytotoxic effectors. Whether CAR T cells act autonomously or require interactions with the tumor microenvironment (TME) is unclear. Here, we report an essential crosstalk between CAR T cell subsets and the TME for tumor control. Using single-cell RNA sequencing, we revealed profound modification of the TME during CAR T cell therapy. IFN-gamma produced by CAR T cells and host-derived IL-12 not only enhanced endogenous T and NK cell activity but were also essential for sustaining CAR T cell cytotoxicity as revealed by intravital imaging. Compared to CD8+ CAR T cells, CD4+ CAR T cells were more efficient at host immune activation but less capable of tumor killing. In sum, CAR T cells are not acting alone in vivo but rely instead on a cytokine-mediated crosstalk with the TME for optimal activity. Invigorating CAR T cell interplay with the host represents an attractive strategy to prevent relapses. Overall design: B cell lymphoma was established by i.v. injection of 0.5x10^6 Eµ-myc-DEVD cells in C57BL/6J mice after sublethal irradiation (4 Gy). Six days later, mice were injected i.v. with 20x10^6 CAR4 or CAR8 T cells or left untreated. Single cell RNA-seq of bone marrow cells were performed on these 3 samples. GSE165797 pubmed 33771887 pubmed 37595589