Selective refueling of CAR T cells using ADA1 and CD26 boosts antitumor immunity

Abstract

CAR T cell therapy has revolutionized the treatment of hematologic malignancies, but its efficacy in solid tumors remains limited due to the immunosuppressive nature of the tumor microenvironment and the inability of T cells to persist and traffic to the tumor site. While current strategies focus on enhancing CAR T cell activity through costimulatory molecules and cytokines, a critical yet often overlooked factor is the competition for nutrients between tumor cells and T cells in the nutrient-deprived tumor microenvironment. To address this challenge, we employed a selective metabolic refueling (MR) strategy by providing T cells with inosine as an alternative fuel source for growth and functionality. In this study, we engineered CAR T cells to co-express a membrane-bound CD26 and a cytoplasmic adenosine deaminase 1 (ADA1) fused to an anti-CD3 scFv. ADA1 irreversibly converts both intracellular and extracellular adenosine to inosine, overcoming adenosine-mediated immunosuppression and providing T cells with inosine for growth. The inclusion of an anti-CD3 scFv fusion partner and overexpressing CD26 boosts ADA1 capture in a membrane proximal manner, providing inosine for T cells and minimizing feeding the tumor cells. We demonstrate that ADA1 is conditionally secreted only in stress conditions and that it activates CAR T cells through trans-signaling in a tumor-specific manner. In addition, we show that, compared to unmodified CAR T cells, CD26-overexpressing CAR T cells have better migration capacity and are less susceptible to TGF-β1 suppression. Finally, we found that, in mice models of human hepatocellular carcinoma (GPC3-MR-CAR) and human non-small cell lung cancer (HER2-MR-CAR), metabolically refueled CAR T cells (MR-CAR) are more efficient in reducing tumor growth than unmodified CAR T cells. Thus, selective refueling CAR T cells using ADA1 and CD26 holds promise for improving the efficacy of CAR T cell therapy of solid tumors.

Advances in research on factors affecting chimeric antigen receptor T‐cell efficacy

“…However, in the tumor microenvironment, TGF‐β‐1 inhibits the expression of CD26, thereby reducing T cell activity. Consequently, in CAR‐T therapy for malignant hematological tumors, increased levels of TGF‐β may weaken the effector function of CAR‐T cells 69 . Studies have shown that knocking out the endogenous TGF‐β receptor II (TGFBR2) in CAR T cells using CRISPR/Cas9 technology can reduce the induction of Treg transformation and prevent exhaustion of CAR T cells 70 .…”

“…Consequently, in CAR-T therapy for malignant hematological tumors, increased levels of TGF-β may weaken the effector function of CAR-T cells. 69 Studies have shown that knocking out the endogenous TGF-β receptor II (TGFBR2) in CAR T cells using CRISPR/Cas9 technology can reduce the induction of Treg transformation and prevent exhaustion of CAR T cells. 70 Thus, the increase of TGF-β in the tumor microenvironment during CAR-T therapy may diminish the cytotoxic effects of CAR-T cells, limiting their ability to recognize and eliminate tumor cells, thereby affecting treatment outcomes.…”