The Effect And Mechanism Of CBX2 On Proliferation And Temozolomide Resistance In Glioma Cells

Background and objective:Glioma is the most common malignant primary brain tumor originating from glial cells.It is characterized by invasive growth,rapid progress,difficulty in complete resection,frequent recurrence and poor prognosis.Although the current clinical comprehensive treatment of glioma includes surgical resection,radiotherapy,chemotherapy,targeted drug therapy and supportive therapy,the prognosis of patients is still very poor,and the median survival time of glioblastoma is only 14.6 months.Temozolomide(TMZ)is an international standard first-line chemotherapeutic drug for glioma,but the resistance of most patients to TMZ is inevitable,which has become a major obstacle to clinical treatment.Chromobox 2(CBX2)is an important regulator of epigenetic modification.As a typical component of polycomb repressive complex 1,it silences tumor suppressor gene expression by epigenetic modification of histone,and then participates in the occurrence,development and drug resistance of many cancers,which is closely related to poor prognosis of patients.Therefore,CBX2 has become a potential target of anti-cancer strategy.However,up to date,there are few studies on the role and mechanism of CBX2 in glioma,and its role and potential mechanism in the occurrence,development,and TMZ resistance of glioma are still largely unknown.The purpose of this study is to explore the effect and mechanism of CBX2 on glioma cell proliferation and TMZ resistance,in order to provide a new target for glioma therapy.Methods:(1)Bioinformatics and immunohistochemistry were used to analyze the expression of CBX2 in glioma and its relationship with tumor grade,TMZ resistance and prognosis;(2)The CBX2 knockdown/overexpression glioma cell line was constructed by plasmid lentivirus transfection,respectively.The effects of CBX2 on glioma cell viability,cell proliferation,TMZ resistance and cell apoptosis were observed by CCK-8 test,colony formation test,Ed U test,TMZ sensitivity test and flow cytometry.In the in vivo study,the mouse glioma model was established,and the effects of CBX2 on glioma growth and TMZ resistance were observed by in vivo fluorescence imaging;(3)QPCR,westernblot,co-immunoprecipitation and chromatin immunoprecipi tation were used to explore the molecular mechanism of the effect of CBX2 on glioma cell proliferation and TMZ resistance.Results:(1)Through bioinformatic analysis and clinical sample immunohistochemical analysis,it was found that the expression of CBX2 was up-regulated in gliomas,which was positively correlated with tumor grade,TMZ resistance and poor prognosis;(2)In terms of cell viability,cell proliferation,TMZ resistance and cell apoptosis,the results of CCK-8 test,colony formation test and Ed U test showed that the viability and proliferation of LN18 glioma cells decreased significantly with the knockdown of CBX2.After TMZ treatment,CCK-8 and flow cytometry were performed.The results showed that knocking down CBX2 enhanced the sensitivity of LN18 cells to TMZ and promoted cell apoptosis.On the contrary,overexpression of CBX2 in LN229 cells promoted cell viability,cell proliferation,enhanced TMZ resistance and inhibited cell apoptosis.The in vivo fluorescence imaging results of the mouse glioma model showed that the tumor growth of mice inoculated with LN229 cells overexpressing CBX2 was significantly faster and TMZ resistance was significantly enhanced;(3)In terms of molecular mechanism,the results of q PCR and westernblot showed that overexpression of CBX2 promoted cell proliferation and TMZ resistance by inhibiting PTEN transcription and expression and activating downstream AKT/m TOR signal pathway.On the contrary,knockdown the expression of CBX2 increased the transcription and expression of PTEN and inhibited the activation of downstream AKT/m TOR signal pathway.Further immunoprecipitation and chromatin immunoprecipitation experiments showed that CBX2 mediates the transcriptional inhibition of PTEN by binding to PTEN promoter and recruiting EZH2 for H3K27me3 modification,and then activates AKT/m TOR signal pathway to promote glioma cell proliferation and TMZ resistance.Conclusion:(1)The expression of CBX2 was up-regulated in glioma,which was positively correlated with tumor grade,TMZ resistance and poor prognosis,and played a carcinogenic role in glioma;(2)Knockdown the expression of CBX2 inhibits the proliferation of glioma cells and enhances the sensitivity of TMZ,while overexpression of CBX2 can promote the proliferation of glioma cells and enhance TMZ resistance;(3)CBX2 mediates the transcriptional inhibition of PTEN by binding to PTEN promoter and recruiting EZH2 for H3K27me3 modification,then activates AKT/m TOR signal pathway,and promotes glioma cell proliferation and TMZ resistance.