Cinobufacini Inhibits Immune Escape of Acute Myeloid Leukemia Cells Through Regulating and Controlling MYH9/USP7/c-MYC Pathway
10.13359/j.cnki.gzxbtcm.2024.05.031
- VernacularTitle:华蟾素通过调控MYH9/USP7/c-MYC通路抑制急性髓系白血病细胞免疫逃逸
- Author:
Rong HUANG
1
;
Kai LIU
;
Jing-Quan HAO
;
Li-Huai WANG
;
Zhuo GAN
Author Information
1. 湖南中医药大学第一附属医院血液肿瘤科,湖南长沙 410007
- Keywords:
cinobufacini;
acute myeloid leukemia;
immune escape;
MYH9/USP7/c-MYC pathway;
nude mice;
HL-60 cells
- From:
Journal of Guangzhou University of Traditional Chinese Medicine
2024;41(5):1298-1306
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the effect of cinobufacini on immune escape of acute myeloid leukemia(AML)by regulating myosin heavy chain 9(MYH9)/ubiquitin-specific protease 7(USP7)/cellular-myelocytomatosis viral oncogene(c-MYC)pathway.Methods(1)In vivo experiment:a nude mouse xenograft tumor model was established to evaluate the effect of cinobufotalin on the growth and immune escape of AML cells in vivo.(2)In vitro experiments:human AML cell line HL-60 was treated with different concentrations of cinobufacini,cell viability was detected by cell counting kit 8(CCK-8),and HL-60 cell invasion was detected by Transwell assay.HL-60 cells were co-cultured with activated CD8+ T cells,the expression of CD25,the surface marker of CD8+ T cells,was detected by flow cytometry,the levels of cytokines[interleukin-2(IL-2)and interferon(IFN-γ)]in the co-culture supernatant were detected by enzyme-linked immunosorbent assay(ELISA).CytoTox96 non-radioactive cytotoxicity assay was used to evaluate the cytotoxicity of CD8+ T cells to HL-60 cells.The protein expressions of MYH9,USP7 and c-MYC in HL-60 cells were detected by Western Blot.The interaction between MYH9,USP7 and ubiquitination was detected by co-immunoprecipitation(Co-IP)assay.The MYH9 overexpression plasmid was tranfected to verify the mechanism of cinobufacini in AML.Results Cinobufacini treatment inhibited xerograft tumor growth in nude mice and enhanced the anti-tumor ability of CD8+ T cells.Cinobufacini treatment inhibited HL-60 cell viability and invasion in a concentration-dependent manner.Cinobufacini treatment up-regulated the expression of CD25,a surface marker of CD8+ T cells,and also up-regulated the levels of IL-2 and IFN-γ.Cinobufotalin enhanced the toxicity of CD8+ T cells to HL-60 cells.Cinobufacini inhibits the protein expressions of MYH9,USP7 and c-MYC in HL-60 cells.MYH9 promotes c-MYC deubiquitination by recruiting USP7,but cinobufacini inhibits MYH9-mediated c-MYC deubiquitination.Conclusion Cinobufacini can reduce the recruitment of c-MYC by deubiquitinating enzyme USP7 by inhibiting the expression of MYH9,and promote the ubiquitination and degradation of c-MYC,thereby inhibiting the immune escape of AML cells.
