Stereotactic Radiation Enhances Efficacy of CAR-T Cells in a Murine Glioma Model

Abstract

T cell tumor infiltration is an important driver of antitumor immune response and tumor cell killing in immune checkpoint inhibitor (ICI) therapy. Many modalities have been investigated to increase T cell tumor infiltration and thereby enhance antitumor immune response including cytotoxic therapies such as radiation. Radiotherapy promotes local inflammation, increases tumor antigen release, and can enhance T cell priming leading to a more robust T cell immune response. Many studies have investigated the combination of radiotherapy and ICI therapy, but limited work has been done examining the combination of targeted CAR-T cell therapy with the T cell immune enhancing effects of radiotherapy. To explore this question, we investigated the anti-tumor efficacy of intra-cranial IL13Rα2 directed CAR-T in combination with tumor irradiation in an immune-competent murine glioma model. Using an immunocompetent murine glioma model, we first determined the sub-therapeutic dose of radiation for our model by using increasing doses of single fraction irradiation (5 Gy, 10 Gy, and 20 Gy). We performed gene-expression panel analysis on brain tumor tissue at each dose to evaluate for changes in tissue expression of chemo-attractant molecules following irradiation. After identifying an appropriate subtherapeutic radiation dose, we tested the combination efficacy of focal radiation and I.C.T delivered IL13Rα2 CAR-T in the glioma model by injecting I.C.T IL13Rα2 CAR T cells 48 hours after 10 Gy tumor irradiation and monitoring tumor growth and response. We then assessed immunologic memory after therapeutic cure by focal radiation, I.C.T. delivered IL13Rα2 CAR-T cell therapy, and combination therapy by re-challenging the cured mice with intracranially implanted IL13Rα2 (+) tumors. In an immunocompetent murine glioma model single fraction doses of 5 Gy and 10 Gy were found to be subtherapeutic. mRNA tissue analysis following irradiation demonstrated statistically significant upregulation of mRNA encoding chemoattractant molecules such as Cxcl2. Mice treated with focal radiation and I.C.T. delivered IL13Rα2 CAR-T exhibited improved survival when compared to mice treated with mock CAR-T cell injection followed by irradiation or with CAR-T cell therapy alone. Mice previously cured with 10 Gy and IL13Rα2 CAR-T treatment demonstrated immunologic memory and rejected IL13Rα2 (+) tumor re-challenge in 5 of 5 mice as compared to naïve mice. Irradiation upregulates mRNA expression of chemoattractant molecules in an immunocompetent murine glioma model. Combination treatment with focal radiation and I.C.T. IL13Rα2 CAR T cell therapy shows improved survival compared to CAR T cell therapy or radiation alone. Combination therapy also leads to enhanced immune memory and improved tumor rejection with tumor re-challenge.