Methylation-controlled J Protein Regulate Temozolomide Resistance Through ATM/Chk2-dependent Pathway In Glioblastoma

BackgroundGlioblastoma(GBM)is the most common primary malignant brain tumor in adult humans.Although the maximum safe range of surgical resection combined with postoperative radiotherapy and temozolomide chemotherapy has become the current clinical standard treatment strategy,it has a degree of malignancy.High,invasive,and easily recurring characteristics,the prognosis of patients with glioblastoma is currently not optimistic,the overall median survival is only 14 months.Temozolomide(TMZ)is an alkylating agent,and is currently recognized as the first-line chemotherapy drug for treating glioblastoma.It has high bioavailability and easily penetrates the blood-brain barrier,allowing glioma cells DNA A.Basalization inhibits cell proliferation and promotes apoptosis,thereby exerting its anti-tumor efifects.Studies have shown that temozolomide can effectively prolong the median survival of patients,but a considerable part of patients will show different degrees of acquired resistance to TMZ,and ultimately lead to chemotherapy failure.Therefore,TMZ chemotherapy resistance is a major problem that restricts the therapeutic efficacy of glioma chemotherapy.Methylation-controlled J protein(MCJ/DNAJC15)is a member of the heat shock protein(HSP)40 family located on the mitochondrial inner membrane.Previous studies have shown that low expression or lack of expression of MCJ is closely associated with poor prognosis of various tumors such as breast cancer and gastric cancer.It is particularly important that MCJ participate in chemotherapy resistance of breast cancer,ovarian cancer and other tumors,but in glioma drug resistance no research report.ObjectiveTo clarify the relationship between MCJ protein and glioma TMZ chemotherapy resistance;to explore the role of MCJ in TMZ chemotherapy resistance of glioma;to prove that ATM/Chk2 signaling pathway mediates MCJ regulation of glioma TMZ resistance.MethodsThe expression of MCJ in glioma TMZ-resistant cells and parental cells was detected by qPCR and Western blot.The correlation between MCJ expression and glioma drug resistance was analyzed.The overexpression of shRNA or shRNA interfered with the expression of MCJ in glioma cells.5 CCK-8 cell viability assay,Western blot,flow cytometry,EdU et al.studied the role of MCJ in TMZ resistance of gliomas;Western Blot et al.explored the role of ATM/Chk2 signaling pathways in MCJ-regulated glioma TMZ resistance.Results1)The TMZ-resistant glioma cell line can be successfully constructed by gradually increasing the TMZ dose of GBM cells in vitro and has drug resistance;2)MCJ was significantly down-regulated in glioma TMZ-resistant cells,and its low expression was associated with drug resistance and poor prognosis in gliomas;3)Interfering with the expression of MCJ can promote TMZ resistance in parental glioma cells and inhibit TMZ-induced apoptosis.Overexpression of MCJ can promote chemosensitization of TMZ-resistant cells in gliomas;4)Interference with MCJ expression can reduce TMZ-induced p-H2AX expression,whereas over-expression of MCJ in glioma TMZ-resistant cells can increase p-H2AX expression and reduce p-ATM and p-Chk2 expression.Conclusion1)The glioma TMZ-resistant cell line was successfully constructed with a significant drug resistance phenotype;2)Down-regulation of MCJ expression indicates glioma drug resistance and poor prognosis;3)MCJ can promote TMZ chemosensitization of glioma cells;4)It is initially confirmed that MCJ regulates TMZ resistance of gliomas through ATM/Chk2 signaling pathway,but its deep mechanism still needs further investigation.