Lin Ruo-Kai, Hsu Chun-Hua, Wang Yi-Ching
Department of Life Sciences, National Taiwan Normal University, Taipei, Taiwan, ROC.
Anticancer Drugs. 2007 Nov;18(10):1157-64. doi: 10.1097/CAD.0b013e3282a215e9.
Abnormal CpG island hypermethylation of multiple tumor-suppressor genes (TSGs) can lead to the initiation and progression of human cancer. The cytosine of the CpG island on the promoter region is methylated by 5'-cytosine-methyltransferases (DNMTs). Pharmacologic inhibitors of CpG island methylation provide a rational approach to reactivate the TSGs in tumor cells and to restore the critical cellular pathways in cancer cells. Mithramycin A (MMA) is known to be a GC- and CG-rich DNA-binding agent. We sought to determine whether MMA could inhibit CpG island methylation and DNMT expression in lung cancer cells. We found that MMA reduced the CpG island methylation of antimetastasis TSGs, including SLIT2 and TIMP-3 genes, and was associated with the prevention of metastasis. When highly metastatic CL1-5 lung cancer cells were treated with low doses (10 nmol/l) of MMA for 14 days, they reexpressed mRNA levels for these genes. MMA also inhibited the invasion phenotypes of CL1-5 cells as indicated by its inhibition of cancer cell migration using wound-healing and transwell assays. Molecular docking of MMA onto the DNMT1 catalytic domain revealed that MMA might interact with the catalytic pocket of DNMT1. Western blots showed that DNMT1 protein levels were depleted after MMA. These data support the idea that MMA has demethylation and antimetastasis effects on lung cancer cells. This mechanism might be mediated by the interaction of MMA and DNMT1, leading to the depletion of the DNMT1 protein and the reversal of the metastasis phenotype in lung cancer cells.
多个肿瘤抑制基因（TSG）的异常CpG岛高甲基化可导致人类癌症的发生和发展。启动子区域CpG岛的胞嘧啶由5'-胞嘧啶甲基转移酶（DNMT）甲基化。CpG岛甲基化的药理抑制剂为重新激活肿瘤细胞中的TSG以及恢复癌细胞中的关键细胞通路提供了一种合理的方法。光辉霉素A（MMA）是一种已知的富含GC和CG的DNA结合剂。我们试图确定MMA是否能抑制肺癌细胞中的CpG岛甲基化和DNMT表达。我们发现MMA降低了包括SLIT2和TIMP-3基因在内的抗转移TSG的CpG岛甲基化，并与转移的预防有关。当高转移性CL1-5肺癌细胞用低剂量（10 nmol/l）的MMA处理14天时，它们重新表达了这些基因的mRNA水平。MMA还抑制了CL1-5细胞的侵袭表型，这通过其使用伤口愈合和transwell试验对癌细胞迁移的抑制来表明。MMA与DNMT1催化结构域的分子对接显示，MMA可能与DNMT1的催化口袋相互作用。蛋白质免疫印迹显示，MMA处理后DNMT1蛋白水平降低。这些数据支持了MMA对肺癌细胞具有去甲基化和抗转移作用的观点。这种机制可能是由MMA与DNMT1的相互作用介导的，导致DNMT1蛋白的消耗以及肺癌细胞转移表型的逆转。
