PC4 Decreases Lung Adenocarcinoma Sensitivity To Cisplatin Through FOXO1 And FOXO3a

Aim1.To explore whether positive cofactor 4(PC4)participated in mediating the biological functions of lung adenocarcinoma cells.2.To elucidate the regulation of PC4 on forkhead O transcription factor 1(FOXO1)and forkhead O transcription factor 3a(FOXO3a)and how PC4 regulated the cell sensitivity of lung adenocarcinoma cells to cisplatin through FOXO1 and FOXO3 a.3.To investigate the roles and related mechanisms of FOXO1 and FOXO3 a in mediating the cell sensitivity of lung adenocarcinoma cells to cisplatin,and in mediating the synergistic enhancing effect of LY294002,an inhibitor of the PI3K/Akt pathway.MethodsLung adenocarcinoma cells A549,PC-9,H1299 were used for the research.Lentivirus infection was applied to knockdown or overexpress PC4,FOXO1 and FOXO3 a.Flow cytometry was utilized to analyze cell apoptosis and cell cycle.Cell counting kit-8(CCK-8)was used to evaluate cell proliferation.Transwell assay was adopted to evaluated cell invasion.Western Blotting assay was used to analyze the expression of related proteins.Nude mice were subcutaneously injected with tumor cells as the in vitro model.ResultsIn the present study,knockdown of PC4 decreased the PC-9 cell invasion and autophagy,and induced more cell apoptosis and a lower cell viability upon cisplatin treatment.Overexpression of PC4 increased the H1299 cell invasion and autophagy,and led to reduced cell apoptosis and increased the cell viability upon cisplatin treatment.The in vitro study further verified that overexpression of PC4 decreased the cell sensitivity to cisplatin.Besides,knockdown of PC4 increased the expression and nuclear accumulation of FOXO1 and FOXO3 a in lung cancer cells,and overexpression of PC4 inhibited the expression and nuclear accumulation of FOXO1 and FOXO3 a.AS1842856,an inhibitor of the FOXO proteins,abrogated the increased cell sensitivity to cisplatin induced by PC4 knockdown.None of the PI3K/Akt inhibitor LY294002,proteasome inhibitor bortezomib or CBP/p300 inhibitor A-485 reversed the regulation of PC4 on FOXO1 and FOXO3 a.the immunoprecipitation assay revealed that PC4 interacted with FOXO3 a.Furthermore,PC4 stabilized FOXO1 and FOXO3 a upon cisplatin treatment in lung cancer.On the other hand,we noticed that cisplatin initially increased the expression and nuclear accumulation of FOXO1 and FOXO3 a in lung adenocarcinoma cells.Knockdown of FOXO1 and FOXO3 a obviously decreased the cell sensitivity to cisplatin.FOXO1 and FOXO3 a regulated cisplatin-induced cell apoptosis independent of Bim,a pro-apoptotic factor downstream of the FOXOs.Furthermore,LY294002 synergistically increased the cytotoxic effects of cisplatin through increasing the expression and nuclear accumulation of FOXO1 and FOXO3 a.Knockdown of FOXO1 and FOXO3 a obviously attenuated the enhancing effects of LY294002 on cisplatin.Conclusion1.PC4 increases the cell invasion and autophagy,and decreases the cell sensitivity to cisplatin in lung adenocarcinoma.2.PC4 reduces the lung adenocarcinoma cell sensitivity to cisplatin through inhibiting FOXO1 and FOXO3 a,which is independent of the PI3K/Akt pathway,proteasome degradation and CBP/p300 acetylation.3.FOXO1 and FOXO3 a increase the cell sensitivity of lung adenocarcinoma cells to cisplatin and mediate the enhancing effect of LY294002 on cisplatin.