G9a promotes immune suppression by targeting the Fbxw7/Notch pathway in glioma stem cells

• Published in: CNS neuroscience & therapeutics Vol. 29; no. 9; pp. 2508 - 2521
• Main Authors: Cao, Yufei, Liu, Bin, Cai, Lize, Li, Yanyan, Huang, Yulun, Zhou, Youxin, Sun, Xingjian, Yang, Wei, Sun, Ting
• Format: Journal Article
• Language: English
• Published: England John Wiley & Sons, Inc 01.09.2023; John Wiley and Sons Inc
• Subjects: Antibodies; Apoptosis; CD4 antigen; CD8 antigen; Cdc4 protein; Chemotherapy; Cholecystokinin; Chromatin; Cytotoxicity; DNA methylation; Epigenetics; Fbxw7; Filtration; Flow cytometry; G9a; Genes; Glioblastoma; Glioma; Glioma cells; glioma stem cells; Growth factors; Histones; immune suppression; Immunofluorescence; Immunohistochemistry; Immunoprecipitation; Immunotherapy; Ligands; Lymphocytes T; Macrophages; Major histocompatibility complex; notch; Original; PD-1 protein; PD-L1 protein; Proteins; Radiation therapy; Stem cells; γ-Interferon; notch; Fbxw7; immune suppression; glioma stem cells; G9a
• ISSN: 1755-5930; 1755-5949
• DOI: 10.1111/cns.14191

Aim Immunotherapy for glioblastoma multiforme (GBM) is limited because of a strongly immunosuppressive tumor microenvironment (TME). Remodeling the immune TME is an effective strategy to eliminate GBM immunotherapy resistance. Glioma stem cells (GSCs) are inherently resistant to chemotherapy and radiotherapy and involved in immune evasion mechanism. This study aimed to investigate the effects of histone methyltransferases 2 (EHMT2 or G9a) on immunosuppressive TME and whether this effect was related to changes on cell stemness. Methods Tumor‐infiltrating immune cells were analyzed by flow cytometry and immunohistochemistry in orthotopic implanted glioma mice model. The gene expressions were measured by RT‐qPCR, western blot, immunofluorescence, and flow cytometry. Cell viability was detected by CCK‐8, and cell apoptosis and cytotoxicity were detected by flow cytometry. The interaction of G9a and F‐box and WD repeat domain containing 7 (Fbxw7) promotor was verified by dual‐luciferase reporter assay and chromatin immunoprecipitation. Results Downregulation of G9a retarded tumor growth and extended survival in an immunocompetent glioma mouse model, promoted the filtration of IFN‐γ + CD4+ and CD8+ T lymphocytes, and suppressed the filtration of PD‐1+ CD4+ and CD8+ T lymphocytes, myeloid‐derived suppressor cells (MDSCs) and M2‐like macrophages in TME. G9a inhibition decreased PD‐L1 and increased MHC‐I expressions by inactivating Notch pathway companying stemness decrease in GSCs. Mechanistically, G9a bound to Fbxw7, a Notch suppressor, to inhibit gene transcription through H3K9me2 of Fbxw7 promotor. Conclusion G9a promotes stemness characteristics through binding Fbxw7 promotor to inhibit Fbxw7 transcription in GSCs, forming an immunosuppressive TME, which provides novel treatment strategies for targeting GSCs in antitumor immunotherapy. G9a promotes histone H3 lysine 9 methylation on Fbxw7 promotor, resulting in the inhibition of Fbxw7 transcription. Downregulated Fbxw7 activates Notch pathway in glioma stem cells to suppress immune response by regulating the expression of immune‐associated molecules.