RIG-I is an intracellular checkpoint that limits CD8+ T-cell antitumor immunity

Abstract

Abstract Background Although a variety of immune checkpoint monoclonal antibodies have been approved for the treatment of cancers, the low response rate of immune checkpoint inhibitors limits their wide clinical application. Therefore, to explore new and broader immunotherapy targets to improve the response rate of CD8+ T cells will provide theoretical basis for clinical treatment of refractory malignant solid tumors.Methods We used single-cell sequencing data to screen for genes associated with exhaustion of infiltrating CD8+ T cells in the tumor microenvironment and validated them by immunoblotting and multiplex immunohistochemistry. We used flow cytometry to analyze the effects of RIG-I knockout in spleen and lymph nodes on the secretion of anti-tumor cytokines and the differentiation and development of CD8+ T cells in vitro. We analyzed the effect of RIG-I knockdown on the growth of hepatocellular carcinoma, colon cancer, and melanoma in vivo, then analyzed the secretion of anti-tumor cytokines by CD8+ T cells in the tumor microenvironment by flow cytometry. We analyzed the possible factors inducing the up-regulation of RIG-I and the signaling pathways downstream of RIG-I that negatively regulate the anti-tumor function of CD8+ T cells in vitro by flow cytometry and immunoblotting. We validated the effect of RIG-I targeting human CD8+ T cells on human tumors in immunodeficient mice. We evaluated the effect of PD-1 antibody in combination with targeted RIG-I on tumor inhibition in mice.Results Here, we demonstrate that RIG-I is upregulated in tumor-infiltrating CD8+ T cells, where it functions as an intracellular checkpoint to negatively regulate CD8+ T-cell function and limit antitumor immunity. Mechanistically, upregulation of RIG-I in CD8+ T cells is induced by retinoic acid, a metabolite of vitamin A in the tumor microenvironment, and directly inhibits the AKT/glycolysis signaling pathway. In addition, deletion of RIG-I enhances the efficacy of adoptively transferred T cells against solid tumors, and inhibition of RIG-I enhances the response to PD-1 blockade.Conclusions Overall, our study identifies RIG-I as an intracellular checkpoint and a potential target for alleviating inhibitory constraints on T cells in cancer immunotherapy, either alone or in combination with an immune checkpoint inhibitor.