EGFRvIII-CAR-T Cells with PD-1 Knockout Have Improved Anti-Glioma Activity

Abstract

Glioblastoma multiforme (GBM) is the most malignant form of the brain tumors. EGFR variant III (EGFRvIII) is expressed in about 30% of GBM specimens, but not expressed in normal brain tissues. Therefore, EGFRvIII protein offers an ideal CAR-T therapeutic target for EGFRvIII-positive GBM patients. PD-L1 is expressed in a variety of cancer cells, including GBM. Tumor-associated PD-L1 can bind to PD-1 on T cells and promote apoptosis of T cells, thus suppressing the anti-cancer immune response. In our current studies, PD-1^WT EGFRvIII-CAR-T cells and PD-1^KD EGFRvIII-CAR-T cells were generated. Cytokine production and lytic activity of these two CAR-T cells against to PD-L1^WT EGFRvIII⁺ U373 cells or PD-L1^KO EGFRvIII⁺ U373 cells were evaluated. The results showed that PD-1^KD EGFRvIII-CAR-T cells and PD-1^WT EGFRvIII-CAR-T cells showed same levels of interferon-γ (IFN-γ) and interleukin-2 (IL-2) production as well as cytolytic activity against PD-L1^KO EGFRvIII⁺ U373 cells; however, PD-1^KD EGFRvIII-CAR-T cells exhibited higher levels of IFN-γ and IL-2 production as well as cytolytic activity against PD-L1⁺ EGFRvIII⁺ U373 cells than that of PD-1^WT EGFRvIII-CAR-T cells. PD-1^KD EGFRvIII-CAR-T cells also exhibited higher anti-glioma activity and longer survival in mice in vivo than that of PD-1^WT EGFRvIII-CAR-T cells. Taken together, our findings indicate that PD-1 knockout enhances lytic activity of EGFRvIII-CAR-T cells against PD-L1⁺ EGFRvIII⁺ GBM cells. These might provide a new insight into strategy of GBM CAR-T cell therapy.

Keywords: CAR-T; EGFRvIII; Glioblastoma multiform; PD-1; PD-L1.