Optimum fractionation of radiation dose to combine anti-PD-1 mAb in MC38 mice model

Abstract

Background: The irradiated tumor cell death can enhance antitumor immunity by inducing antigen expression on tumor cells and activating lymphocytes. Radiotherapy (RT) combined with immunotherapy has revealed promising outcomes in various animal models. However, the optimum fractionation of radiation for priming immune response is controversial. This study aimed to explore the fractionation of radiation to maximize immunity in combinatorial treatment. Methods: Mice bearing MC38 murine colon cancer were treated with up to 24Gy radiation given in various sized fractions as 24Gy x 1f, 8Gy x 3f, 8Gy x 1f followed by 2Gy x 8f and 2Gy x 12f, and tumor growth followed. The immune response in the tumor, drainage lymph node(dLN) and spleen at 48h after radiation were assessed. 8Gy x 3f was chosen to combine anti-PD-1 immunotherapy. The abscopal effects and immune response were assessed by flow cytometry and immunohistochemistry(IHC). Results: Single dose of 24Gy and 8Gy x 3f brought best tumor control. No abscopal effects was observed after radiotherapy alone. Fractionation of 8Gy x 3f increased the irradiated tumor infiltrating lymphocytes(TILs). However, conventional 2Gy doses decreased CD4+TILs and CD8+TILs and increased myeloid myeloid-derived suppressor cell (MDSC) in spleen significantly. As the optimal fractionation to maximize immunity, 8Gy x 3f was chosen to combine anti-PD-1 mAb. Compared to radiotherapy or anti-PD-1 mAb alone, 8Gy x 3f combining with anti-PD-1 mAb brought obvious abscopal effect. CD8+T cells in the dLNs of the irradiated tumors were increased significantly in the combining group. Also, the combining treatment regimen increased CD4+T cells and CD8+T cells and decreased MDSC in the spleen. No serious toxicity of heart, liver, spleen, lung and kidney in each group was observed by using IHC. Conclusions: Hypofractionation of 8Gy x 3f was the fractionation of radiation dose to maximize immunity, compared to single dose of 24Gy and conventional 2Gy doses. Radiation with 8Gy x 3f combining with anti-PD-1 mAb had synergistic antitumor effect.