Circular RNA CDR1as Sensitized Bladder Cancer To Cisplatin By Sponging MiR-1270 And Regulating APAF1 Expression

Background:Recently,Circular RNAs(CircRNAs)have emerged as critical regulators in tumorigenesis and progression.Several studies on CDR1 as have demonstrated it as a miRNA sponge in different tumors.However,the role of CircRNAs in cancer chemosensitivity is still unclear.In this study,we aimed to investigate the impact and mechanism of circRNA-CDR1 as on bladder cancer cisplatin chemosensitivity.Methods:We constructed CDR1 as overexpressing T24 and EJ cell lines by adenovirus transfection.The cell apoptosis and cisplatin chemosensitivity of CDR1 as on bladder cancer cells were determined using flow cytometry and CCK-8.CDR1 as might be combined with miR-1270 through Starbase prediction.Fish was performed to identify and locate CDR1 as and miR-1270,while RNA pull down was applied to confirm their sponging effect.qRT-PCR was used to measure the expression levels of miR-1270 in bladder cancer tissues and cell lines.The cell apoptosis,proliferation and cisplatin chemosensitivity of miR-1270 on bladder cancer cells were determined using flow cytometry and CCK-8.Bioinformatic databases,qRT-PCR,Western blot and dual-luciferase reporter assay were performed to validate APAF1 as a target of miR-1270.In addition,flow cytometry and CCK-8 verified whether CDR1 as could induce apoptosis and enhance cisplatin chemosensitivity through indirect regulation of APAF1.Probit regression model was used to calculate IC50 of transfected cells.Finally,the sensitivity of CDR1 as to cisplatin chemotherapy in nude mice xenografts were evaluated by size and volume of tumours.Results:Over-expression of CDR1 as induced apoptosis and enhanced cisplatin chemosensitivity in T24 and EJ cells.miR-1270 was highly expressed in bladder cancer tissues and cell lines,and promoted cell proliferation,inhibited cell apoptosis and reduced cisplatin chemosensitivity in T24 and EJ cells.FISH showed that CDR1 as and miR-1270 were co-located in T24 cells.RNA pull down verified that CDR1 as could bind directly to miR-1270.T24 and EJ cells with co-transfection of CDR1 AS and miR-1270 reduced cisplatin chemosensitivity.The experiments of qRT-PCR,Western blot and double luciferase reporter gene confirmed that APAF1 is the target gene of miR-1270.Silencing APAF1 reduced the sensitivity of bladder cancer cells to cisplatin chemotherapy.Furthermore,CDR1 as could directly sponge miR-1270 and abolish its effect on APAF1 consequently.Finally,the volume and weight of xenograft in cisplatin chemotherapy group were lower than those in normal saline group.Compared with the GFP control group,the volume and quality of xenograft tumors in the CDR1 as overexpressing group were significantly reduced after cisplatin chemotherapy.Conclusions:Our study verified that CDR1 as played a cisplatin chemosensitization effect in bladder cancer through CDR1as/miR-1270/APAF1 axis.Therefore,the newly identified axis is expected to be a potential therapeutic targets for bladder cancer patients.