T-bet overexpression modulates CAR T cell effector functions

Abstract

Abstract Adoptive transfer of T cells modified with a chimeric antigen receptor (CAR) are highly effective in patients with hematologic malignancies. However, many patients relapse and CAR T cells have shown minimal efficacy against solid tumors. Our lab and others have shown that standard CAR designs fail to promote differentiation into canonical T cell subsets. CAR T cells generate both type-1 and type-2 cytokines simultaneously, challenging the traditional paradigm that Th1 and Th2 differentiation programs are mutually exclusive. We sought to control this by overexpressing the transcription factor T-bet to drive a more canonical type-1 effector response. CD28-costimulated CAR T cells overexpressing T-bet showed a reduction in type-2 cytokine production. In an immune-competent syngeneic mouse model of B cell acute lymphoblastic leukemia, T-bet overexpressing CAR T cells showed increased expansion, increased generation of KLRG1 +CD127 −effector T cells, and decreased expression of exhaustion-associated markers PD1, TIM3, and LAG3 through the initial CAR T cell expansion and contraction phases Despite this, Tbet overexpression drove late CAR T cell hypo-functionality and leukemic relapse. However, in 4-1BB-costimulated CAR T cells, which tend to have a weaker effector response than CD28-costimulated CAR T cells, T-bet overexpression led to increased CAR T cell potency, prolonging survival over mice treated with a 4-1BB-costimulated CAR. Furthermore, T-bet overexpression led to increased CAR T cell polyfunctionality against low-antigen density leukemia when compared to standard CAR T cells. These results demonstrate the potential of T-bet-overexpression as a strategy to modulate the strength of the CAR T cell effector response. Supported by funding from the University of Colorado School of Medicine Human Immunology and Immunotherapy Initiative