FOXM1 Evokes 5-fluorouracil Resistance In Colorectal Cancer Depending On ABCC10

Background:Colorectal cancer(CRC)is one of the most prevalent malignancy and the third leading cause of cancer-related death worldwide.Despite therapeutic strategies involving surgical resection,radiotherapy and chemotherapy,the survival rate of CRC patients remains unsatisfactory.5-Fluorouracil(5-FU)is one of the main component of adjuvant and palliative chemotherapy for CRC patients.It elicits cytotoxicity by inhibiting the nucleotide synthetic enzyme thymidylate synthase(TYMS),as well as by incorporating fluoronucleotides into RNA and DNA.It has shown that a fraction of CRC patients are inherently refractory to 5-FU-based chemotherapy,and most of the remaining patients will acquire chemoresistance after treatment,leading the failure of treatment and the tumor recurrence and metatastasis.FOXM1 belongs to alarge Forkhead Box family that share a conserved winged-helix DNA-binding domain,playing important roles in a variety of drug resistance.We hypothesis FOXM1 contribute to the 5-FU resistance of CRC depending on the previous experiment,intend to elucidate the underlying mechanisms and discover potential targets for the treatment of CRC patients with 5-FU resistance.Method:1.The raw gene profile data of colorectal cancer patients were downloaded from GEO data repository(GSE3964).Gene expression profile were explored in 5-FU sensitive group and resistant group.2.Analysis the relationship of FOXM1 expression and IC50 values in CRC cell lines.3.Knocking down FOXM1 expression in CRC cells with high FOXM1 levels by two independent lentiviral-mediated short hairpin RNAs(shRNAs)and overexpressing FOXM1 in CRC cells with low FOXM1 levels.Examining the influence of FOXM1 in 5-FU resistance by CCK8 assay,colony formation assay and apoptosis evaluation.4.To investigate the effect of FOXM1 on the 5-FU sensitivity of CRC in vivo,FOXM1 overexpressing RKO cells were subcutaneously transplanted into nude mice,followed by 5-FU and other drug treatment.Xenograft size was measured every 3 days.5.Concerning that FOXM1 functions as a transcription factor,we screened the promoter regions of genes related to 5-FU metabolism and drug efflux for FOXM1 binding elements,then verified the potential target gene by qRT-PCR and ChIP.Detecting the change of IC50 value and apoptosis rate in CRC cells following FOXM1 alteration as well as rescue experiment.Results:In thisstudy,we found that FOXM1 was upregulated in the CRCtissues of 5-FU nonresponsive patients,the expression of FOXM1 was positively correlated with the IC50 value of 5-FU in CRC cell lines.Silencing of FOXM1 reduced cell viability under 5-FU treatment,inhibited cell proliferation and enhanced cell apoptosis upon 5-FU exposure,whileectopic expressionof FOXM1 exhibited the opposite effect.In vivo data confirmed that FOXM1 improves the 5-FU tolerance of CRC cells.Mechanically,FOXM1 promoted 5-FU resistance by directly enhancing the transcription of ABCC10,the combination of vardenafil and 5-FU achieved a notable growth suppression on FOXM1 overexressing xenografts,which was refractory to 5-FU treatment alone.Discussion:Recent studies have demonstrated that FOXM1 involves in the resistance to cisplatin,paclitaxel,docetaxel and epirubicin,indicating that FOXM1 plays important roles in drug resistance.Our study first determinded the roles of FOXM1 in the 5-FU resistance of CRC.Mechanically,FOXM1 promoted 5-FU resistance by directly enhancingthe transcription of ABCC10 to elevate drug efflux.Therefore,targeting ABCC10 provides a strategy for sensitizing FOXM1-abundant CRC to 5-FU.In conclusion,our results demonstrate that FOXM1/ABCC10 axis contributes to the 5-FU resistance of CRC,and may serve as potential therapeutic targets to overcome 5-FU resistance.