Background:ADAR1, the major enzyme for RNA editing, has emerged as a tumor-intrinsic key determinant for cancer immunotherapy efficacy through modulating interferon-mediated innate immunity. However, the role of ADAR1 in innate immune cells such as macrophages remains unknown.
Methods:We first analyzed publicly accessible patient-derived single-cell RNA-sequencing and perturbed RNA sequencing data to elucidate the ADAR1 expression and function in macrophages. Subsequently, we evaluated the combined effects of ADAR1 conditional knockout in macrophages and interferon (IFN)-γ treatment on tumor growth in three distinct disease mouse models: LLC for lung cancer, B16-F10 for melanoma, and MC38 for colon cancer. To gain the mechanistic insights, we performed human cytokine arrays to identify differentially secreted cytokines in response to ADAR1 perturbations in THP-1 cells. Furthermore, we examined the effects of ADAR1 loss and IFN-γ treatment on vessel formation through immunohistochemical staining of mouse tumor sections and tube-forming experiments using HUVEC and SVEC4-10 cells. We also assessed the effects on CD8+ T cells using immunofluorescent and immunohistochemical staining and flow cytometry. To explore the translational potential, we examined the consequences of injecting ADAR1-deficient macrophages alongside IFN-γ treatment on tumor growth in LLC-tumor-bearing mice.
Results:Our analysis on public data suggests that ADAR1 loss in macrophages promotes antitumor immunity as in cancer cells. Indeed, ADAR1 loss in macrophages combined with IFN-γ treatment results in tumor regression in diverse disease mouse models. Mechanistically, the loss of ADAR1 in macrophages leads to the differential secretion of key cytokines: it inhibits the translation of CCL20, GDF15, IL-18BP, and TIM-3 by activating PKR/EIF2α signaling but increases the secretion of IFN-γ through transcriptional upregulation and interleukin (IL)-18 due to the 5'UTR uORF. Consequently, decreased CCL20 and GDF15 and increased IFN-γ suppress angiogenesis, while decreased IL-18BP and TIM-3 and increased IL-18 induce antitumor immunity by enhancing cytotoxicity of CD8+ T cells. We further demonstrate that combination therapy of injecting ADAR1-deficient macrophages and IFN-γ effectively suppresses tumors in vivo.
Conclusion:This study provides a comprehensive elucidation of how ADAR1 loss within macrophages contributes to the establishment of an antitumor microenvironment, suggesting the therapeutic potential of targeting ADAR1 beyond the scope of cancer cells.
Keywords:cytokines; immunity, innate; immunotherapy; macrophages; tumor microenvironment.