The Effect Of The Promoter Methylation Of RASSF1A、DAPK、MGMT、p16、RARβ On The Cisplatin’s Sensitivity In A549Cell

Objective: Drug resistance was an important reason of failure tochemotherapy in NSCLC. According to the reported papers, methylation ofmultiple genes could be one of important factors which regulate thechemosensitivity of cancers. So we aimed to investigate the methylationsratus of multiple key genes associated with sensitivity to cisplatin in lungadenocarcinoma cell line A549.Methods: A549cell line and its homologous cisplatin-resistant cell line(A549-DDP) were cultured. The methylation status of RASSF1A, MGMT,DAPK, p16, RARβ were determined by methylation specific PCR(MSP), themRNA expression of these5genes were determined by RT-PCR in A549and A549-DDP cell line respectively. The relationship was determinedbetween the changes of related genes treated with demethylationreagent(5-aza-CdR) and sensitivity to cisplatin. To investigate the influenceof demethylation reagent on A549-DDP, cell proliferation and apoptosis wasdetected by MTT and flow cytometry Results: RASSF1A, MGMT, p16were unmethylated and high expressedin A549cell line. On the contrary, these three genes were hypermethylatedand low expressed in A549-DDP cell line, and the mRNA expression ofRASSF1A, MGMT could be upregulated with the treatment of5-aza-CdR, Asfor RARβ, unmethylated status and high level of mRNA expression werefound in A549-DDP, and just opposite to that in A549cell line. But there wasno significant difference in the methylation status and mRNA expression ofDAPK between the two cell lines. The IC50of A549was（3.64±0.71）μmol/l,and the IC50of A549-DDP without or with5-aza-CdR was（25.10±3.23）μmol/l,（6.55±1.02）μmol/l for cisplatin respectively.5-aza-CdR enhancedthe cisplatin’s effect of proliferation Inhibition and apoptosis significantly inA549-DDP cell line.Conclusion: The changes of mRNA expression regulated by themethylation of RASSF1A, MGMT, p16, RARβ might be one of possiblereasons of sensitivity to cisplatin in A549. And the methylation were relatedto proliferation inhibition and apoptosis of DDP in A549. The changes ofmethylation status of multiple genes might be potential laboratory biomarkerto predict the sensitivity to DDP in lung cancer.