Middle East Technical University, Department of Biological Sciences, 06531, Ankara, Turkey.
Biomed Pharmacother. 2012 Feb;66(1):29-35. doi: 10.1016/j.biopha.2011.09.002. Epub 2011 Dec 28.
Development of resistance against anti-cancer drugs is one of the major obstacles of chemotherapy in the treatment of cancer. Etoposide is a topoisomerase II alpha (TOP2A) inhibitor, which is used in the treatment of breast cancer. Alterations in the expression of drug targets or DNA repair genes are among the important resistance mechanisms against TOP2A inhibitors. In this study, expression changes in TOP2A gene and two important mismatch repair (MMR) genes MSH2 and MLH1 were examined in order to understand the relationship between differential expression of these genes and drug resistance against etoposide.
Resistant cell lines were developed from parental MCF7 cell line by stepwise selection in increasing doses of etoposide. Total RNA was isolated from parental and resistant cell lines by using TriReagent. Expression levels of TOP2A, MSH2 and MLH1 were analysed by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Statistical analyses were performed by one way ANOVA.
Etoposide resistant sublines MCF7/1000E, MCF7/1250E and MCF7/2000E were approximately 2, 3 and 4 fold resistant relative to parental MCF7/S cells, respectively. TOP2A, MSH2 and MLH1 expressions decreased in etoposide resistant sublines relative to MCF7/S cells. Expression levels of TOP2A and MLH1 in resistant sublines differed between 10-95 and 18-58 percent of the expression levels in the parental cells, respectively. MSH2 expression levels were decreased 18-82 percent in resistant cells. A transient 15 percent increase in the expression of this gene was observed in subline MCF7/1250E.
Decrease in the expression levels of TOP2A, MSH2 and MLH1 may play significant roles in the development of chemotherapeutic resistance to etoposide in breast cancer. These genes may be considered for further development of new strategies to overcome resistance against topoisomerase II inhibitors.
癌症化疗的主要障碍之一是抗癌药物耐药性的产生。依托泊苷是拓扑异构酶 II α（TOP2A）抑制剂，用于治疗乳腺癌。药物靶点或 DNA 修复基因表达的改变是对抗 TOP2A 抑制剂的重要耐药机制之一。在这项研究中，检查了 TOP2A 基因和两个重要的错配修复（MMR）基因 MSH2 和 MLH1 的表达变化，以了解这些基因表达差异与依托泊苷耐药性之间的关系。
通过逐步增加依托泊苷剂量，从亲本 MCF7 细胞系中开发出耐药细胞系。使用 TriReagent 从亲本和耐药细胞系中分离总 RNA。通过逆转录定量聚合酶链反应（RT-qPCR）分析 TOP2A、MSH2 和 MLH1 的表达水平。通过单向方差分析进行统计分析。
与亲本 MCF7/S 细胞相比，依托泊苷耐药亚系 MCF7/1000E、MCF7/1250E 和 MCF7/2000E 的耐药性分别约为 2、3 和 4 倍。TOP2A、MSH2 和 MLH1 的表达在依托泊苷耐药亚系中均低于 MCF7/S 细胞。TOP2A 和 MLH1 的表达水平在耐药亚系中分别为亲本细胞表达水平的 10-95%和 18-58%，MSH2 表达水平降低 18-82%。在耐药细胞中观察到该基因的表达短暂增加 15%。
TOP2A、MSH2 和 MLH1 表达水平的降低可能在乳腺癌对依托泊苷化疗耐药性的发展中起重要作用。这些基因可能被考虑用于进一步开发克服拓扑异构酶 II 抑制剂耐药性的新策略。
