Effect Of ERCC1 Gene Silencing On Chemosensitivity Of Non-small Cell Lung Cancer

Objectives This study aims to investigate the effect of ERCC1 silencing on the chemosensitivity of non-small cell lung cancer to cisplatin,paclitaxel,pemetrexed alone,or combination.Methods ERCC1 expression data was extrated from the TCGA database.RT-PCR was used to detect the m RNA level of ERCC1 in H23,H2030 and A549 cell lines.Lentivirus with LV-ERCC1-siRNA1,LV-ERCC1-siRNA2 or LV-NC-siRNA was transfected into A549 cells.Western blot was used to detect the expression of the ERCC1.To determine the effect of ERCC1 silencing on chemosensitivity of non-small cell lung cancer to cisplatin,paclitaxel and pemetrexed.Cells were treated cisplatin,paclitaxel,pemetrexed alone or in combination.CCK method and colony formation assay were used to determine the cell viability.Results 1 We analyzed the expression levels of ERCC1 in 535 lung adenocarcinomas and 59 corresponding paracancerous tissues in the TCGA database.We found that the expression level of ERCC1 gene in lung adenocarcinoma tissues(median 2082)was much higher.The difference was statistically significant(Z=-4.901,P=0.001)in its expression in the paraneoplastic tissues(median 1274).The expression level of ERCC1 gene in squamous cell carcinoma of the lung(502 cases)(median 3004)and its expression in paracancerous tissues(49 cases)(median 2721)were not statistically different(Z=-1.643,P=0.100).2 The relative expression levels of ERCC1 in three lung adenocarcinoma cell lines A549,NCI-H23 and NCI-H2030 were 0.16±0.017,0.037±0.01 and 0.026±0.001,respectively.It can be found that ERCC1 m RNA expression is higher in A549 cells.3 The expression of ERCC1 was significantly decreased in A549 cells transfected with LV-ERCC1-siRNA1 and LV-ERCC1-siRNA2 lentivirus.4 CCK assay detected the proliferation of lung cancer cells treated with different chemotherapy regimens.The results are as follows.The survival rate of the ERCC1 silencing group(transfected with LV-ERCC1-siRNA1 or LV-ERCC1-siRNA2 lentivirus)was low after treatment of lung cancer cells transfected with lentivirus with single agents(cisplatin,paclitaxel,and pemetrexed).In the control group(transfection control LV-NC-siRNA lentivirus vector),the difference was statistically significant(P<0.05).For the combination group,lung cancer cells were treated with a fixed concentration of cisplatin(2 ?M)and a certain concentration gradient of paclitaxel(0,2,4,6,8 and 10 n M)for 72 hours,and cell viability was measured.The lowest cell viability of the LV-A549-siRNA1 and LV-A549-siRNA2 ERCC1 transfected group was significantly lower than that of the LV-NC-siRNA control lentivirus transfection group(57.34±5.81)% at(40.47±5.1)% and(37.86±1.22)%,respectively.Lung cancer cells were treated with a fixed concentration of cisplatin(2?M)and a certain concentration gradient of pemetrexed(0,2,4,6,8 and 10 ?M)for 72 hours,and cell viability was measured.The lowest cell viability of LV-A549-siRNA1 and LV-A549-siRNA1 ERCC1 transfection group was(32.52±1.46)% and(33.53±1.26)%,which was significantly lower than that of LV-NC-siRNA control lentivirus transfection group(40.99±1.79)% Lung cancer cells were treated with a fixed concentration of paclitaxel(2n M)and a certain concentration gradient of cisplatin(0,2,4,6,8 and 10 ?M)for 72 hours and cell viability was measured.The lowest cell viability of LV-A549-siRNA1 and LV-A549-siRNA1 ERCC1 transfected group was(21.01±0.25)% and(21.92±0.27)%,which was significantly lower than that of LV-NC-siRNA control lentivirus transfection group(34.32±1.07)%.Lung cancer cells were treated with a fixed concentration of pemetrexed(2?M)and a certain concentration gradient of cisplatin(0,2,4,6,8 and 10 ?M)for 72 hours,and cell viability was measured.The lowest cell viability of LV-A549-siRNA1 and LV-A549-siRNA1 ERCC1 transfection group was(20.71±0.23)% and(18.96±1.85)%,which was significantly lower than that of LV-NCsiRNA control lentivirus transfection group(36.65±0.32)%.5 The results of colony formation assay showed that cisplatin,paclitaxel,and pemetrexed alone could inhibit the colony formation ability of cells,and the colony formation ability of ERCC1 silencing group was lower than that of the control group,and the differences were statistically significant(P<0.01).Conclusions ERCC1 silencing can increase the chemo-sensitivity of non-small cell lung cancer to cisplatin,paclitaxel,pemetrexed alone,or combination.