Abstract 6510: Irradiation promotes hepatocellular carcinoma immune cloaking via PD-L1 upregulation induced by cGAS-STING activation

Abstract

Abstract Objective. Radiotherapy (RT) is one of the main treatments for patients with unresectable hepatocellular carcinoma (HCC). Emerging evidence indicates that the cGAS-STING pathway is crucial in RT-induced antitumor immune response. Here, we discovered that activation of the cancer cell-intrinsic cGAS-STING pathway mediated immune cloaking resulting from RT-induced DNA damage. This provided a rationale for combining RT with programmed cell death 1 (PD-1) and its ligand (PD-L1) checkpoint blocking in treating HCC. Design. We analyzed changes in PD-L1 expression using 20 fixed HCC tissues from patients treated with preoperative RT and serum soluble PD-L1 (sPD-L1) expression after RT in another cohort of 118 HCC patients, and obtained survival data. We knocked out or knocked down key proteins in the cGAS-STING pathway in Huh7, HCCLM3, MHCC97H Huh7, H22, and Hepa1-6 HCC cell lines. H22 cells were transplanted into wild-type, cGAS-deficient (Mb21d1−/−), and STING-deficient (Tmem173gt) C57BL/6 mice. We performed RNA sequence, immunoblot, dual-luciferase reporter, and chromatin precipitation analyses of mouse liver and cell lines. Hep1-6 cells were used to generate orthotopic intrahepatic tumor models. Mice were administered combinations of RT and antibodies against PD-L1 (or PD-1) or antibodies to deplete T cells. Tumors were collected and analyzed by immunohistochemistry and flow cytometry. Results. RT upregulated PD-L1 in HCC patients, which correlated with poor survival. RT activated cGAS-STING and increased immune checkpoint PD-L1 expression in human and mouse liver cancer cells. Ionizing radiation activated the STING/TBK1/IRF3 innate immune pathway, leading to PD-L1 upregulation in HCC cells, which inhibits cytotoxic T lymphocyte activity and protects tumor cells from immune-mediated eradication. Knockdown of cGAS,STING, TBK1, and IRF3 reversed the antitumor effect of cytotoxic T lymphocyte-mediated cytotoxicity after ionizing radiation in vitro or in vivo. RT potentiated the antitumor effect of PD-1/PD-L1 axis blockade and augmented cytotoxic T-cell infiltration in HCC tumors in immunocompetent mice. Similar synergetic effects were observed in HCC tumors in Mb21d1−/− and Tmem173gt C57BL/6 mice, suggesting that the effects are independent of the host cGAS-STING pathway. CD8 depletion compromised the synergetic antitumor effect of combined RT and anti-PD-L1 blockade, demonstrating that CTLs are required for antitumor immunity induced by the combination therapy. Conclusion Our results identified an immune-cloaking mechanism of RT-activated innate immune cGAS-STING and show that RT could empower HCC immunotherapy. Citation Format: Shisuo Du, Genwen Chen, Ping Yang, Yixin Chen, Yong Hu, Zhaochong Zeng. Irradiation promotes hepatocellular carcinoma immune cloaking via PD-L1 upregulation induced by cGAS-STING activation [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6510.