Bmi1 Drives The Genesis And Development Of Intrahepatic Cholangiocarcinoma By Inhibiting HLF

Purpose:Bmi1,as a member of PRC1 complex family,plays an important role in the tissue development and cell differentiation by repressing Ink4A/Arf.Oncogene Bmi1 can drive tumor initiation and promote progression in an Ink4A/Arf dependent or independent manner.However,the role and underlying mechanism of Bmi1 in the pathogenesis of intrahepatic cholangiocarcinoma(ICC)are not clear yet.In this study,using both animal and cell models,we aimed to investigate the role of Bmi1 in the development of.ICC and discover its potential targets in ICC.Methods:We examined the expression of Bmi1 in clinical ICC samples using bioinformatic analysis,Western blotting,qPCR and immunohistochemistry.We established Bmi1/NRas ICC mouse models using hydrodynamic injection via tail vein.The effect of Bmi1 knockdown(KD)on cell proliferation was evaluated in human ICC cells.The effect of Bmi1 knockout(KO)or KD on ICC initiation and development was investigated in Bmi1 KO mice and xenograft tumor models,respectively.We evaluated the therapeutical effect of Bmi1 inhibitor PTC-209 in the AKT/NICD induced primary ICC mice.Finally,Western blotting and q RT-PCR were used to verify the correlation between Bmi1 and Ink4A/Arf.RNA-seq,chromatin immunoprecipitation and luciferase reporter genes were used to examine the expression of the downstream target of Bmi1.Results:We found that Bmi1 mRNA and protein level were both higher than those in the normal tissue.Bmi1 with NRas can drive ICC formation in mice.In vitro,Bmi1 silencing inhibits the cell viability and colony formation of ICC cells and induces cell cycle arrest in G2/M phase.Apoptosis of ICC cells after Bmi1 KD was increased by 20%.In vivo,Bmi1 KO or KD blocked ICC formation and ICC xenograft tumor volume reduced by 1/2.Moreover,Bmi1 inhibitor PTC-209-HBr reduced ICC nodule numbers in AKT/NICD mouse model.We found no correlation between Bmi1 and the expression of p16^Ink4A and p14^ARF(or p19^ARF)in ICC cells and tissues.Negative correlation between Bmi1 and HLF was found.Critically,overexpression of HLF inhibited ICC cell proliferation,reduced ICC xenograft tumor volume and primary ICC nodule numbers.HLF silencing restored the cell proliferation inhibited by Bmi1 knockdown.Conclusion:This study reveals that Bmi1 is highly expressed in human ICC specimen.Bmi1 can cooperate with NRas to drive ICC in mice.Bmi1 KD or KO can significantly inhibit the formation and development of ICC,indicating that Bmi1 is necessary for the occurrence of ICC.Importantly,HLF functions as an oncogene and is the direct target of Bmi1 in ICC.Overexpression of HLF can inhibits the development of ICC.Bmi1 is expected to serve as an ideal target for ICC treatment,thus providing new pathway for translational medicine and ICC drug discovery.