Study On Mechanism Underlying Mangiferin-induced Radiosensitization Of Malignant Glioma And In Vivo Validation

Objective:Due to the highly aggressive and recurrent characteristics of glioblastomas(GBM),the prognosis of GBM patients is extremely poor,and the effect of radiotherapy is not significant.DNA damage response(DDR)is the main reason for GBM to develop radiotherapy resistance.DDR includes two pathways: homologous recombination(HR)and non-homologous end joining(NHEJ),among which NHEJ is the main pathway for tumor cell repair after radiotherapy.The previous research of our group found that mangiferin may inhibit GBM cell proliferation and DDR.Based on this,the study aims to explore the molecular mechanism of mangiferin by inhibiting the NHEJ pathway to increase the sensitivity of radiotherapy,and to provide theoretical guidance and scientific basis for the development of anti-GBM targeted drugs with mangiferin as the lead compound.Method:(1)After mangiferin treatment,the key proteins of NHEJ repair pathway were detected by Western Blot,including 53BP1,p-ATM,p-53BP1,γ-H2 AX,in U-87 MG and U-118 MG cells with IR treatment.(2)The DSBs marker protein γ-H2 AX focus in U-87 MG and U-118 MG cells was detected by immunofluorescence.(3)After mangiferin treatment,γ-H2 AX focus was detected by immunefluorescence in rat immortalized Schwann cells with IR treatment,and DNA damage percentages in Schwann cells were obtained by ELISA assay.(4)The tumor-bearing mice were established and treated with DMSO,IR,mangiferin,IR+mangiferin in groups.The 20-day weight and 100-day survival rate of the tumor-bearing mice were obtained.Then the tumor volume and weight were also measured.Result:(1)The phosphorylation levels of ATM,53BP1,and H2 AX were significantly decreased after mangiferin treatment(P<0.01).This showed that mangiferin prevented DNA damage repair by inhibiting the NHEJ pathway.(2)Observations on the average percentages of γ-H2AX-positive cells and the average number of γ-H2AX foci per cell suggested that treatment with mangiferin significantly decreased decreased the number of γ-H2AX foci in GBM cells following IR(P<0.01).Mangiferin can improve the radiosensitivity of GBM by inhibiting DNA damage repair.(3)The number of γ-H2AX-positive Schwann cells was similar to that of the control group after mangiferin treatment(P>0.05),and mangiferin was unable to trigger inhibition of NHEJ repair in normal glial cells.(4)Mangiferin combined with IR can effectively increase the weight and prolong survival time of the tumor-bearing mice(n=10,P<0.05).(5)Mangiferin combined with IR can significantly reduce the volume and weight of subcutaneous tumors,inhibiting the growth of tumor tissues(n=10,P<0.05).Conclusion:Mangiferin has a significant sensitization effect on GBM radiotherapy,which may be mediated by the inhibition of NHEJ repair pathway.Hence,mangiferin can become a potential therapeutic drug for improving the radiation sensitivity of glioblastoma.