Regulatory T cells shape the differential impact of radiation dose-fractionation schedules on host innate and adaptive anti-tumor immune defenses: Immunosuppressive implications of radiotherapy.

Abstract Purpose: We examined how radiation dose per fraction (DPF) and total dose, as represented by biological effective dose (BED), can independently and differentially affect the immunomodulatory capacity of radiation therapy (RT). Materials and Methods: AT3-OVA mammary and MC38 colorectal tumors in C57BL/6 mice were irradiated with rationally selected dose-fractionation schedules, alone or with immune-modulating or depleting agents. Tumor growth was monitored as a readout of therapeutic response. Flow cytometry and RNA sequencing of mouse tumors and analysis of transcriptomic datasets from irradiated human cancers were used to dissect the immunomodulatory effects of the different radiation schedules. Results: In AT3-OVA tumors, radiation DPF rather than BED determined the ability of RT to evoke local anti-tumor CD8+ T cell responses and synergize with anti-PD-1 therapy. Natural killer (NK) cell-mediated control of irradiated tumors was more sensitive to radiation BED. Radiation-induced regulatory T cell (Treg) responses, which were detected in both mouse and human tumors, were a major factor underlying the differential activation of adaptive immunity by radiation DPF and the activity of NK cells during the early phase of response to RT. Targeted inhibition of Treg responses within irradiated tumors rescued and enhanced local tumor control by RT and permitted the generation of abscopal and immunological memory responses, irrespective of radiation schedule. Intriguingly, MC38 tumors did not support the induction of an amplified Treg response to RT and were highly vulnerable to its immunoadjuvant effects. Conclusion: Local radiation-induced Treg responses are influenced by radiation schedule and tumor type and are a critical determinant of the immunoadjuvant potential of RT and its ability to synergize with T cell-targeted immunotherapy.