PXR基因外显子3甲基化与肠癌细胞对5-氟尿嘧啶的耐药性相关  

Association Between Pregnane X Receptor Gene Exon3 Methylation with 5-Fluorouracil Resistance of the Colon Cancer Cells

作　　者：赵嵘[1] 胡丽玲[1] 孔繁强[2] 左爱军[3] 

机构地区：[1]天津市人民医院,天津300191 [2]天津医科大学附属总医院检验科,天津300052 [3]美国南加州大学,ca91016

出　　处：《中国生物化学与分子生物学报》2013年第1期63-69,共7页Chinese Journal of Biochemistry and Molecular Biology

摘　　要：孕烷X受体(pregnane X receptor,PXR)可通过调节细胞色素P450同工酶3A4—CYP3A4的表达而影响肿瘤细胞对化疗的敏感性,而其表达水平则会受到自身基因甲基化的影响.本文研究了结肠癌组织中pxr基因甲基化的分布情况及其对pxr,cyp3a4表达的影响,并在多种结肠癌细胞系中分析了pxr基因甲基化是否与5-氟尿嘧啶(5-FU)耐药性相关.收集结肠癌病灶区、癌旁区及正常结肠组织样本,分别提取基因组DNA及RNA.PCR-限制性酶切分析检测pxr基因外显子3甲基化;real-time PCR检测pxr及cyp3a4基因的表达.鉴定LOVO、LS180、LS174T、HT29、HCT116等5种结肠癌细胞中pxr外显子3甲基化与pxr,cyp3a4表达的相关性并分别筛选出PXR高/低表达的细胞株进行5-FU耐药性分析.结果显示,结肠癌病灶组织中pxr外显子3甲基化频率显著增加,伴有pxr,cyp3a4表达的增强.在结肠组织及结肠癌细胞系中,pxr与cyp3a4的表达均密切相关,且均与pxr甲基化程度相关.PXR高表达细胞株LS180对5-FU的耐药性显著升高,以siRNA分别下调pxr及cyp3a4的表达,均可增加LS180对5-FU的敏感性.结果提示,pxr基因外显子3区甲基化与PXR及CYP3A4的高表达密切相关,并与结肠癌细胞对5-FU的抗药性相关.Pregnane X receptor （PXR） regulates the expression of cytochrome P450 3A4 （CYP3A4） to influence the sensitivity of cancer cells to chemotherapies. To determine the frequency of pxr exon3 methylation in relation to the expression of pxr and cyp3a4 genes, colon cancer tissues and different human cancer cells were investigated. The results showed that a higher frequency of pxr exon3 methylation was positively correlated to the increased pxr and cyp3a4 expressions in cancerous tissues than in adjacent normal colon tissues. The methylation of pxr exon3 was found in LOVO, LS180 and LS174T, but not in HT29 and HCT116 cells. The expression of pxr and cyp3a4 was positively correlated to the pxr exon3 methylation in LOVO, LS180 and LS174T with higher degrees than in HT29 and HCT116 cells. LS180 and HCT116 cells with high and low pxr and cyp3a4 expression,and correspondingly with and without pxr exon3 methylation were chosen for 5-fluorouracil （5-FU） sensitivity analyses. Compared with HCT116, LS180 cell showed a significantly increased resistance to 5-FU induced apoptosis as well as increased 5-FU IC50 of proliferation inhibition. To demonstrate whether increased expression of PXR and CYP3A4 was involved in the 5-FU resistance in LS180 cell, cyp3a4 and pxr siRNAs were transfected to selectively knock down the target gene expression in LS180. As a result, the resistance of LS180 cell to 5-FU was significantly reduced. We also found that high frequency of pxr exon3 methylation in colon cancer tissues was positively correlated to the expression of pxr and cyp3a4 genes similar to the cell line studies. Elevated PXR and CYP3A4 expression could be an important factor that was able to increase the resistance of colon cancer cells to 5-FU treatment.

关 键 词：孕烷X受体 甲基化 结肠癌 5-氟尿嘧啶 抗药性 

分 类 号：R735[医药卫生—肿瘤]