Identification of hypermethylated genes associated with cisplatin resistance in human cancers.
Identification of hypermethylated genes associated with cisplatin resistance in human cancers.
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DOI：
10.1158/0008-5472.can-09-3427
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发表时间：
2010-04-01
期刊：
影响因子：
11.2
通讯作者：
Sidransky D
中科院分区：
文献类型：
--
作者：
Chang X;Monitto CL;Demokan S;Kim MS;Chang SS;Zhong X;Califano JA;Sidransky D
顺铂是实体瘤中最广泛使用的细胞毒性抗癌药物之一，然而，继发性耐药的发展仍然是临床疗效的主要障碍。治疗相关的DNA超甲基化可能通过使细胞毒性所需的基因失活而在产生耐药表型中发挥作用。我们应用了药理学解蔽方法来检测高甲基化基因，其失活有助于顺铂耐药。利用来自两个亲本细胞系（KB-3-1和SCC 25）的三对同基因、顺铂敏感和耐药细胞系，我们鉴定了几百个在每个耐药细胞系中下调并被DNA甲基转移酶抑制剂5-氮杂-2 '-脱氧胞苷（5-Aza-dC）重新激活的基因。其中，30个基因在≥ 2个细胞系中共有，和/或据报道在之前的研究中下调。亚硫酸氢盐测序证实，耐药细胞系中有14个基因高甲基化，但敏感亲本细胞系中没有。14个基因中的6个（SAT、C8 orf 4、LAMB 3、TUBB、G 0 S2、MCAM）在敏感细胞系中是顺铂可诱导的，但在耐药细胞系中不是。siRNA敲除两个基因SAT和S100 P，增加敏感亲本细胞系中顺铂处理的细胞活力。S100 P基因敲低可显著降低亲本敏感细胞系的S期分数（SPF），并减缓细胞增殖，这与顺铂敏感性降低有关。基于这些研究结果，我们得出结论，DNA甲基化是一个频繁的事件，在长期暴露于顺铂的细胞，甲基化诱导的基因沉默可能发挥作用的发展耐细胞毒性化疗药物。
Cisplatin is among the most widely used cytotoxic anti-cancer agents in solid tumors, however, the development of secondary resistance remains a major obstacle to clinical efficacy. Treatment-related DNA hypermethylation may play a role in creating drug resistant phenotypes by inactivating genes that are required for cytotoxicity. We applied a pharmacologic unmasking approach to detect hypermethylated genes whose inactivation contributes to cisplatin resistance. Utilizing three pairs of isogeneic, cisplatin-sensitive and -resistant cell lines derived from two parental cell lines (KB-3-1 and SCC25), we identified several hundred genes that were down-regulated in each resistant cell line and re-activated by the DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (5-Aza-dC). Among them, 30 genes were common to ≥ 2 cell lines, and/or reported to be down-regulated in previous studies. Bisulfite sequencing confirmed that 14 genes were hypermethylated in resistant cell lines, but not in the sensitive parental cell lines. Six of 14 genes (SAT, C8orf4, LAMB3, TUBB, G0S2, MCAM) were cisplatin-inducible in sensitive, but not in resistant cell lines. siRNA knockdown of two genes, SAT and S100P, increased cell viability with cisplatin treatment in sensitive parental cell lines. S100P knockdown significantly decreased the S-phase fraction (SPF) of parental sensitive cell lines and slowed cell proliferation, which was associated with decreased sensitivity to cisplatin. Based on these findings, we conclude that DNA methylation is a frequent event in cells that are chronically exposed to cisplatin, and that methylation-induced gene silencing may play a role in the development of resistance to cytotoxic chemotherapeutic agents.