135 - IL-17-Producing Gd T Cells Recruited By Cgas-Sting-Dependent CCL20 in Macrophages Drive Radioresistance

Purpose/Objective(s): The immunosuppressive tumor microenvironment (TME) is a key driver of radioresistance, and its interaction with radiotherapy remains a major research focus. Unlike conventional aß T cells, ?d T cells recognize targets in a non-MHC-restricted manner and play pivotal roles in immune regulation and antitumor response. However, their role in modulating tumor radiosensitivity remains poorly understood. This study aims to elucidate the regulatory effects and mechanisms of ?d T cells on radiotherapy sensitivity.

Materials/Methods: Subcutaneous LLC lung cancer and B16-F10 melanoma models were established in C57BL/6 mice. After 10 Gy radiotherapy, flow cytometry evaluated ?d T cell infiltration and cytokine profiles in the TME. The role of ?d T cells in radiosensitivity was investigated using TCR d chain knockout mice (TCR d^-/-). Single-cell RNA sequencing was performed to analyze changes in various cell subpopulations post-radiotherapy, aiming to identify key ?d T cell subsets, chemokines, and cytokines. Neutralizing antibodies were administered to assess the functional roles of critical subsets and cytokines in vivo. Mechanisms were explored through in vitro immune cell migration assays, RT-qPCR, Western blot, and RNA sequencing.

Results: Compared to lower-dose (2 Gy) radiotherapy or short-term (24 hours) post-radiotherapy, the proportion of ?d T cell infiltration in the TME was significantly increased at 96 hours following higher-dose (10 Gy) radiotherapy. Depletion of ?d T cells markedly enhanced the radiosensitivity of subcutaneous LLC lung cancer and B16-F10 melanoma tumors. Single-cell RNA sequencing revealed that the ?d T cell population in the TME post-radiotherapy was predominantly characterized by the expression of Zbtb16, Il23r, and Il17a, and these cells were identified as the primary source of IL-17A in the TME. Functionally, ?d T cells promoted radioresistance by facilitating the recruitment of myeloid-derived suppressor cells (MDSCs) and suppressing T cell-mediated antitumor immune activation. Mechanistically, the recruitment of ?d T cells was driven by radiated tumor cell-released microparticles (RT-MPs) containing double-stranded DNA (dsDNA), which upregulated CCL20 expression in macrophages via the cGAS-STING/NF-?B signaling pathway.

Conclusion: Radiotherapy-induced tumor cell-derived microparticles activate the cGAS-STING/NF-?B pathway in macrophages, enhancing the chemokine CCL20 secretion and ?d T cell recruitment. These ?d T cells drive radioresistance by producing IL-17A, which recruits MDSCs and inhibits T cell activation. Depleting ?d T cells or IL-17A neutralization significantly enhances tumor radiosensitivity. These results highlight the pivotal role of ?d T cells and IL-17A in mediating radioresistance and suggest potential therapeutic targets to improve radiotherapy efficacy.