Patient-derived ascites reveals functional advantages of γδCAR T-cells within the ovarian cancer-like tumor microenvironment

Abstract

Chimeric antigen receptor (CAR) T-cells engineered from {gamma}{delta} T-cells show limited clinical benefit in solid tumors such as ovarian cancer (OVCA). Although {gamma}{delta} T-cells can detect malignant transformation and exert cytotoxicity, these properties confer potent preclinical efficacy but limited therapeutic activity and the basis for this discrepancy remains unclear. Here, we established an ex-vivo culture consisting of highly inflammatory patient-derived malignant ascites and hypoxia conditions, to simulate key obstacles encountered by CAR T-cells in the immunosuppressive OVCA tumor microenvironment (TME). Using this condition, we first demonstrated the presence of CAR-expressing {gamma}{delta} T-cells within conventional (conv)CAR T-cells, which displayed greater proliferative capacity than {gamma}{delta}neg T-cells. We therefore produced {gamma}{delta}CAR T-cells for parallel comparison with conventional (conv)CAR T-cells, evaluating differences in gene expression and functionality. Under TME-like conditions, {gamma}{delta}CAR T-cells demonstrated lower viability but consistently outperformed convCAR T-cells, demonstrating favorable gene expression profiles, enhanced cytotoxicity, improved retention of cytokine secretion and degranulation and increased proliferative capacity. Importantly, we characterized an ascites protein profile that impacted the viability of both cell types comparably. Together, these findings reveal specific functional advantages of {gamma}{delta}CAR T-cells over convCAR T-cells in the immunosuppressive OVCA TME-like condition while facing different cellular fitness constraints, providing insights into the use and limitations of {gamma}{delta}CAR T-cells.