IFNγ IS REQUIRED FOR OPTIMAL THERAPEUTIC EFFICACY OF KIR-CAR ENGINEERED T CELLS

Abstract

In solid tumours, T cell infiltration triggers adaptive immune resistance in the tumour microenvironment (TME), which oppresses subsequent anti-tumor T cell function. Interferon gamma (IFNγ) plays an important role in the anti-tumor function of T cells and is also a significant contributor to adaptive resistance. Here, we investigated the contribution of IFNγ to the anti-tumor function of T cells engineered with synthetic antigen receptor based on KIR2DS2 (KIR-CARs). We generated IFNγ knockout (KO) KIR-CAR T cells and evaluated their functionality both in-vivo and in-vitro. Treatment of established xenograft tumors with IFNyKO KIR-CAR T cells was associated with a commensurate reduction in adaptive immune resistance; however, we did not observe enhanced efficacy. Rather, the IFNyKO KIR-CAR T cells were less efficacious than wild type KIR-CAR T cells. Further investigation revealed a significant decrease in cytotoxicity of the IFNyKO KIR-CAR T cells at limiting T cell:tumor ratios. Interestingly, this loss of efficacy was not accompanied by a decrease in proliferative capacity of the IFNyKO KIR-CAR T cells or change in their persistence in-vivo. Supplementation of recombinant human IFNy to IFNyKO KIR-CAR T cell cultures completely rescued cytotoxic efficacy. To determine whether optimal cytotoxicity relied upon auto/paracrine IFNyR signalling to the engineered T cells, we generated IFNyR1KO T cells. Both in-vitro and in-vivo, IFNyR1KO T cells performed equitably to their WT counterparts, indicating that autocrine IFNyR signalling did not influence T cell performance. Neutralizing IFNy in both control and IFNyR1KO cultures resulted in an equivalent loss of cytotoxicity, confirming that cytotoxicity is dependent upon IFNy signaling to the tumor. This work confirms the opposing roles of IFNy in the TME and indicates a need for further understanding and optimization of cytokine profiles in engineered T cell therapies.