Abstract 5233: Engineered NK-like NKG2C+ CD8 T cells mediate superior anti-tumor efficacy over conventional CD8 T cells and NK cells

Abstract We have recently characterized a unique CD8 T cell subset that expresses NKG2C, as well as multiple other natural killer cell markers. These NKG2C+ CD8 T cells demonstrate NK-like effector functions against various tumor targets, can be efficiently expanded and engineered using GMP-compatible techniques, and most importantly, display beneficial qualities unique to T and NK cells while minimizing the therapeutic deficits of each. NKG2C+ CD8 cells can be easily expanded using a genetically modified K562 feeder line and can be efficiently engineered via retroviral transduction. We genetically engineered NKG2C+ CD8 T cells to express an anti-CD19-28-z CAR construct to target B cell malignancies and compared effector functions of the CAR-engineered NKG2C+ CD8 T cells against CAR+NKG2C- (conventional) CD8 T cells and CAR+ NK cells in vitro and in vivo. We examined expansion, engineering efficiency, tumor control, and extent of cytokine release syndrome (CRS). We also engineered NKG2C+ CD8 T cells with a transgenic NY-ESO-1 TCR to target melanoma and compared the functional responses and efficacy of TCR-engineered NKG2C+ CD8 T cells against TCR-engineered conventional CD8 T cells. CAR+NKG2C+ CD8 T cells demonstrated superior in vitro effector function when compared to CAR+ conventional CD8 T cells and CAR+NK cells and did not upregulate characteristic T cell exhaustion markers, such as PD-1, TIM-3, and LAG-3. These CAR+NKG2C+ CD8 T cells demonstrated superior control of a CD19+ NALM6 tumor model in NRG mice, which was further enhanced with administration of IL-15. CAR+NKG2C+ CD8 T cells with IL-15 demonstrate dramatic and enhanced persistence, as well as superior cytotoxicity and activation in vivo when compared to CAR+ conventional CD8 T cells and CAR+ NK cells. Using a SCID-beige CRS mouse model, we observed reduced CRS in mice treated with CAR+ NKG2C+ CD8 T cells than in mice treated with CAR+ conventional CD8 T cells, with lower levels of proinflammatory cytokines and minimal expansion of murine myeloid subsets. Similarly, TCR-engineered NKG2C+ CD8 T cells demonstrated enhanced tumor control and in vivo persistence in an A375 melanoma-bearing NRG mouse model compared to TCR-engineered conventional CD8 T cells. We have successfully demonstrated that CAR- or TCR-engineered NKG2C+ CD8 T cells have superior anti-tumor efficacy, in vitro and in vivo, to conventional CD8 T cell and NK cell counterparts due to their innate tumor killing and greater in vivo persistence. These cells mediate less CRS and provide a safer alternative to traditional engineered T cell therapeutics. Ultimately, our NKG2C+ CD8 T cells represent a unique platform for engineered T cell therapies to target both solid tumors and hematologic malignancies, that is easily expanded ex vivo, does not require CD4 T cells for in vivo persistence, and is resistant to exhaustion. Citation Format: Kyle Lupo, M. Kazim Panjwani, Anthony Daniyan, Christopher Klebanoff, Katharine Hsu. Engineered NK-like NKG2C+ CD8 T cells mediate superior anti-tumor efficacy over conventional CD8 T cells and NK cells abstract. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts) ; 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84 (6Suppl): Abstract nr 5233.