摘要
目的 探讨10个基因的启动子CpG岛甲基化状态与乳腺癌细胞对5-氟尿嘧啶（5-Fu）敏感性的关系,确定与乳腺癌细胞株5-Fu敏感性相关的启动子CpG岛类基因.方法 采用四甲基偶氮唑蓝（MTT）法检测Bcap-37、T47D和ZR-75-30乳腺癌细胞株对5-Fu的敏感性.应用甲基化特异性PCR（MSP）方法,检测3株对5-Fu敏感性有差异的乳腺癌细胞株中BAG1、C110RF31、CBR1、CBR4、GJA1、FOXL2、IGFBP6、P4HA1、SRI和TYM基因的启动子CpG岛甲基化状态,并用实时定量PCR的方法检测PKSS21、LOX、IGFBP6、ABCC8和CHFR基因的稳定态mRNA表达的水平.结果 IGFBP6和FOXL2基因在对5-Fu敏感和耐药的乳腺癌细胞株中的甲基化状态有差异.除CHFR基因以外,PRSS21、LOX、IGFBP6和ABCC8基因的表达水平与启动子CpG岛甲基化状态有关,处于甲基化状态时表达水平低,而在处于去甲基化状态时表达水平高.结论 在敏感和耐药乳腺癌细胞株中存在甲基化状态差异的基因为IGFBP6和FOXL2基因,这为进一步开展DNA甲基化标志在乳腺癌化疗耐药中的应用研究提供了依据.
Objective To explore the relationship between the methylation status of CpG islands in the promoter region of 10 genes in breast cancer cells and their sensitivity to 5-fluouracil （5-Fu） , and to identify the genes responsible for the 5-Fu resistance in breast cancer. Methods Three cell lines （differently resistant to chemotherapy） were used in this study： Bcap-37 （IC50：289.77μg/ml）, T47D （IC50：134.16μg/ml） and ZR-75-30 （IC50：4.20μg/ml）. The methylation profile of 10 genes （BAG1,C11ORF31, CBR1, CBR4, GJA1, FOXL2, IGFBP6, P4HA1, SRI and TYMS） in the 3 breast cancer cell lines was determined by methylation specific PCR. The steady-state mRNAs of ABCC8, CHFR and IGFBP6 genes were quantified by real-time RT PCR analysis. Results Among the 10 genes, only genes IGFBP6 and FOXL2 displayed differential DNA methylation pattern between the 5-Fu-resistant and 5-Fu-sensitive cell lines. The mRNA expression level of genes PRSS21, LOX, IGFBP6, ABCC8 and CHFR was quantified by real-time RT-PCR analysis. Except for CHFR, the expression level of the other 4 genes was correlated with the methylation status of CpG islands, namely, a lower expression level with methylation status and a higher level with demethylation status. Conclusion The results of the present study have demonstrated that there are 8 genes with differential methylation status in chemosensitive and chemoresistant breast cancer cell lines,i. e. two genes more than the six genes we reported previously. Our findings provide both mechanistic insights for the drug resistance of breast cancer and the basis for further studies on potential application of the DNA methylation in this set of genes for prediction of chemosensitivity of breast cancer.
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
2010年第5期328-333,共6页
Chinese Journal of Oncology
基金
上海市科委资助项目（07DJ14074）
上海市肿瘤医院院外合作项目（YWHZ200601）
