GPC3-targeted CAR-T cells expressing GLUT1 or AGK exhibit enhanced antitumor activity against hepatocellular carcinoma

Chimeric antigen receptor-expressing T (CAR-T) cells induce robust antitumor responses in patients with hematologic malignancies. However, CAR-T cells exhibit only limited efficacy against solid tumors such as hepatocellular carcinoma (HCC), partially due to their limited expansion and persistence. CD8⁺ T cells, as key components of the adaptive immune response, play a central role in antitumor immunity. Aerobic glycolysis is the main metabolic feature of activated CD8⁺ T cells. In the tumor microenvironment, however, the uptake of large amounts of glucose by tumor cells and other immunosuppressive cells can impair the activation of T cells. Only when tumor-infiltrating lymphocytes (TILs) in the tumor microenvironment have a glycolytic advantage might the effector function of T cells be activated. Glucose transporter type 1 (GLUT1) and acylglycerol kinase (AGK) can boost glycolytic metabolism and activate the effector function of CD8⁺ T cells, respectively. In this study, we generated GPC3-targeted CAR-T cells overexpressing GLUT1 or AGK for the treatment of HCC. GPC3-targeted CAR-T cells overexpressing GLUT1 or AGK specifically and effectively lysed GPC3-positive tumor cells in vitro in an antigen-dependent manner. Furthermore, GLUT1 or AGK overexpression protected CAR-T cells from apoptosis during repeated exposures to tumor cells. Compared with second-generation CAR-T cells, GPC3-targeted CAR-T cells overexpressing GLUT1 or AGK exhibited greater CD8⁺ T-cell persistence in vivo and better antitumor effects in HCC allograft mouse models. Finally, we revealed that GLUT1 or AGK maintained anti-apoptosis ability in CD8⁺ T cells via activation of the PI3K/Akt pathway. This finding might identify a therapeutic strategy for advanced HCC.