| MAGE-A9 expression in epithelial ovarian cancer and its clinical significanceOBJECTIVE: Melanoma-associated antigen A9(MAGE-A9)is a protein abundantly expressed in many human cancer types and has been widely regarded as an important target of cancer immunotherapy.However,the expression of MAGE-A9 in epithelial ovarian cancer(EOC)and its potential role in EOC progression have not been studied so far.The aim of this study was to investigate the expression of MAGE-A9 in epithelial ovarian cancer and evaluate its clinical significance in human epithelial ovarian cancer.METHODS: The expression of MAGE-A9 was detected using quantitative real-time PCR(qRT-PCR)and immunohistochemistry(IHC)in human epithelial ovarian cancer(EOC)tissues and normal ovaries or fallopian tubes,as well as benign ovarian tumors.The follow-up data were used to evaluate the prognosis of EOC by Kaplan-meier survival and Cox regression analyses.RESULTS: The expression of MAGE-A9 was significantly higher in human epithelial ovarian cancer(EOC)than non-tumorous tissues.Immunohistochemical staining showed that MAGE-A9 protein was highly expressed in ovarian cancer tissues,and the immunoreactivity of MAGE-A9 was mainly located in the cytoplasm.Furthermore,we found that high expression of MAGE-A9 protein was significantly associated with FIGO staging,histological grade,CA-125 level and tumor metastasis,but not with other pathological parameters.Cox regression analysis and Kaplan-Meier survival curve showed that overexpression of MAGE-A9 was an independent prognostic factor in patients with ovarian cancer.CONCLUSION: These findings indicate that MAGE-A9 expression may be a valuable biomarker for assessing the prognosis of EOC patients.MAGE-A9 overexpression predicts poor prognosis in patients with ovarian cancer.Effect of MAGE-A9 Gene on the Biological Behavior of Epithelial Ovarian Cancer and the Effect of Chemotherapy ResistanceOBJECTIVE: To study the expression profile of MAGE-A9 in ovarian cancer cell lines,and explore the effect of MAGE-A9 on the biological behavior of ovarian cancer cell lines and the involvement of MAGE-A9 in chemotherapeutic resistance.To elucidate the role of MAGE-A9 in the pathogenesis of ovarian cancer,and provide effective theoretical and experimental basis for ovarian cancer treatment strategy.METHODS: The expression of MAGE-A9 mRNA and protein in ovarian cancer cell lines HO-8910,A2780,SK-OV3 were examined using qRT-PCR and Western blot analyses.The expression of MAGE-A9 were detected using qPCR and Western Blot analyses in HO-8910 cell line transfected with different MAGE-A9-targeting shRNAs(ShRNA-1~4)or control shRNA(shRNA-NC).HO-8910 cells were divided into normal group,shRNA group,shRNA + NC group,DDP group,shRNA + DDP group.MTT,cell invasion and wound scratch assays were carried out to investigate the effect of MAGE-A9 depletion on tumor cell proliferation,apoptosis and,invasion and metastasis,and the effect of cisplatin on cell biological behavior.Western blot analysis was used to detect the expression of apoptosis-related proteins,Cyclins and EMT-related proteins.MAGE-A9 overexpression vector was constructed and subjected to A2780 cell transfection.MTT,cell invasion and wound scratch assays were employed to determine the effect of MAGE-A9 overexpression on tumor cell proliferation,apoptosis,invasion and metastasis.A MAGE-A9 overexpression construct was employed to ectopically express MAGE-A9 in MAGE-A9-low cell line A2780.The proliferation of A2780 cells was assessed using MTT assay.The effect of combined action on the biological behavior of cells was investigated using Western blot analysis.The effect of MAGE-A9 on the expression of proteins involved in apoptosis,autophagy and EMT was investigated by Western blot analysis.RESULTS: 1.The expression of MAGE-A9 mRNA and protein in ovarian cancer cell lines were detected by qRT-PCR and Western Blot analyses.The expression of MAGE-A9 in HO-8910 cells was significantly higher(P <0.05),as compared with other cell lines.The results also showed that MAGE-A9 was lowly expressed in A2780 cells(P <0.05).2.RNA interference experiment revealed that MAGE-A9-targeting shRNA-3 achieved the best interfering efficiency among the four shRNAs in HO-8910 cells(P <0.05).3.MTT assay showed that overexpression of MAGE-A9 contributed to malignant proliferation of EOC cells,and depletion of MAGE-A9 significantly enhanced the cytotoxic effect of cisplatin in EOC cells,and the difference was of statistically significance(P <0.05).The viability of cells in MAGE-A9 depletion plus cisplatin treatment was significantly lower than that in other groups(P <0.05).Interference of MAGE-A9 could inhibit the migration of EOC cells.In wound scratch test,shRNA + DDP group expanded faster than other groups,indicating that depletion of MAGE-A9 could effectively inhibit the migration of tumor cells,and the difference was of statistically significance(P <0.05).Western blot showed that a combination of shRNA-3 transfection and cisplatin treatment could strongly trigger the apoptosis and impair the autophagy of EOC cells.The expression of Ki-67,a proliferative marker,was decreased following apoptosis.4.MTT assay showed that ectopic expression of MAGE-A9 promoted the proliferation of A2780 cells,and the difference was statistically significant(P <0.05).Western blot analysis confirmed that ectopic expression of MAGE-A9 dramatically increased the level of MAGE-A9 in A2780 cells.MAGE-A9 overexpression partially antagonized the cytotoxic effect of cisplatin.However,the viability of cells with MAGE-A9 overexpression plus cisplatin treatment was lower than the control group,and the difference was statistically significant(P <0.05).In would scratch test,the wound healing rate of MAGE-A9 overexpression cells was faster,which indicated that MAGE-A9 could effectively promote the migratory ability of tumor cells,and the difference was statistically significant(P <0.05).Western blot analysis showed that MAGE-A9 overexpression could inhibit the apoptosis of EOC cells and increase the autophagy of tumor cells.The expression of MAGE-A9,Vimentin and Ki67 protein increased,and DDP promoted apoptosis(P <0.05).The expression of MAGE-A9,Vimentin and Ki67 protein was significantly decreased following DDP exposure.Ectopic expression of MAGE-A9 could partially attenuate DDP-induced EOC apoptosis,and the difference was statistically significant(P <0.05).CONCLUSION: 1.Depletion of MAGE-A9 can effectively inhibit the proliferation,invasion and migration of ovarian cancer cells,and the inhibitory effect of DDP is more obvious.2.Interference of MAGE-A9 potentiates the pro-apoptotic effect of DDP in EOC cells.3.Overexpression of MAGE-A9 enhances the malignant behearvior of ovarian cancer cell lines,including accelerated proliferation,reduced level of apoptosis,and enhanced DDP resistance. |
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