Study On The Mechanism Of Class Ⅰ Histone Deacetylase HDAC3 In Inhibiting Apoptosis Of Cholangiocarcinoma Cells

Backgrounds:Class I histone deacetylase(HDAC)plays an important role in the development of many tumors.Histone deacetylase 3(HDAC3),as a class I HDACs,has the effect of inhibiting cancer cell apoptosis and promoting cancer cell proliferation in tumors.MI192 is a novel HDAC3-specific inhibitor that exhibits antitumor activity in many tumor cells.However,the role of HDAC3 and the antitumor activity of its inhibitors are unknown in cholangiocarcinoma(CCA).Objective:The present study aimed to evaluate the expression of Class I HDACs in cholangiocarcinoma and investigate the effects of HDAC3 on the proliferation and apoptosis of cholangiocarcinoma cells,and to explore the possible mechanisms.And then we aim to identify a chemotherapeutic agent with the target of HDAC3 in CCA as well as evaluate its therapeutic efficacy.Methods:First,we collected 60 samples of cholangiocarcinoma samples from the Drum Tower Hospital.Immunohistocal technique was used to detect the expression of histone deacetylase in cancer tissue and adjacent normal bile duct tissues and to analyze the relationship between histone deacetylase and prognosis.In addition,we also used protein blotting to detect the expression of HDAC3 protein in 9 cholangiocarcinoma tissues and 2 normal bile duct tissues.We then transfected two cholangiocarcinoma cells(HuCCT-1 and RBE)with siRNA.HDAC3 and HDAC3 plasmids.Cell proliferation and apoptosis were detected by CCK8,colony formation and flow cytometry,respectively.Then we used HDAC3 inhibitors to treat cholangiocarcinoma cell lines through CCK8 and colony formation method to detect changes in cell viability.Finally,we used nude mice subcutaneous transplanted tumor model to assess the inhibitory activity of MI 192 on HDAC3 in vivo and its antitumor activity.Results:The results of this study show that HDAC1,2 and 8 in class I HDACs have no difference in cancer and adjacent tissues,and HDAC3 is upregulated in CCA tissues compared with adjacent tissues and is associated with decreased survival.The CCK8 and colony formation assay showed that upregulation of HDAC3 could promote proliferation of cholangiocarcinoma cells,whereas down-regulation of HDAC3 inhibited tumor cell proliferation.HDAC3 inhibitor MI 192 can inhibit the growth of cholangiocarcinoma cells,reduce the ability of tumor cell colony formation and induce apoptosis.In addition,MI 192 treatnent of nude mice subcutaneous transplanted tumor model,found that nude mice tumor proliferation was inhibited,while increased apoptosis,confirmed MI 192 has anti-tumor activity.Finally,we demonstrate that HDAC3 selective inhibitors inhibit HDAC3 activity in vitro and in vivo and inhibit deacetylation of their downstream target targets.In conclusion,these results demonstrate for the first time that MI 192 inhibits cholangiocarcinoma cell proliferation and induces apoptosis by targeting HDAC3.Conclusions:These findings indicated that HDAC3,a key regulatory factor for cell proliferation and apoptosis,was associated with poor prognosis in CCA patients.For the first time,our results demonstrated that down-regulation of HDAC3 induces apoptosis in human cholangiocarcinoma cells and HDAC3 inhibitors therefore show promise as a new drug candidate for CCA therapy.