Gene expression profiles of CD4-derived (CAR4) and CD8-derived (CAR8) chimeric antigen receptor T cells after stimulation through the CAR, TCR or both receptors. Mus musculus

Chimeric antigen receptor (CAR)-expressing T-cells induce durable remissions in patients with relapsed/refractory B-cell malignancies. CARs are artificial constructs introduced into mature T-cells conferring a second, non-MHC restricted specificity in addition to the endogenous T-cell receptor (TCR). The impact of TCR activation on CAR T-cell efficacy in vivo has important implications for clinical optimization of CAR T-cell therapy, but cannot be systematically evaluated in xenograft models. Using an immunocompetent, syngeneic murine model of CD19-targeted CAR T-cell therapy for pre-B cell ALL, we demonstrate loss of CD8 CAR T-cell mediated clearance of leukemia associated with T-cell exhaustion and apoptosis when TCR antigen is present. CD4 CAR T-cells demonstrate equivalent cytotoxicity, as compared to CD8 CAR T-cells, and in contrast, retain in vivo efficacy in the presence of TCR stimulation. Gene expression profiles confirm increased exhaustion and apoptosis of CAR8 upon dual receptor stimulation compared to CAR4, and indicate inherent differences in T-cell pathways. Chimeric antigen receptor (CAR) T cells express two activating receptors, the CAR and the endogenous T cell receptor (TCR). CAR T cells can be derived from either CD8 or CD4 T cells to generate CAR8 and CAR4 cells, respectively. In vivo, CAR8 and CAR4 cells respond differently when simultaneously stimulated through the CAR and TCR. Overall design: Murine CAR8 and CAR4 cells were generated with a CAR that recognizes murine CD19 and TCRs that recognize HY-minor histocompatibility antigens. CAR8 and CAR4 T cells were co-incubated for 10 hours with B cells that were either possitive or negative for the expression of CD19 and/or HY-antigen. Through this CAR8 and CAR4 T cells were stimulated through the CAR, the TCR, CAR+TCR (simultaneiously) or neither receptor (CD19 negative, HY-negative B cells). Gene expression profiles of CD4-derived (CAR4) and CD8-derived (CAR8) chimeric antigen receptor T cells after stimulation through the CAR, TCR or both receptors