Integrated analysis of DNA methylation and mRNA expression profiling reveals candidate genes associated with cisplatin resistance in non-small cell lung cancer
Integrated analysis of DNA methylation and mRNA expression profiling reveals candidate genes associated with cisplatin resistance in non-small cell lung cancer
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DOI：
10.4161/epi.28601
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发表时间：
2014-06-01
期刊：
影响因子：
3.7
通讯作者：
Tong, Jian-Dong
中科院分区：
文献类型：
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作者：
Zhang, You-Wei;Zheng, Yun;Tong, Jian-Dong
DNA甲基化在获得性耐药的发展过程中起着至关重要的作用。本研究的目的是确定非小细胞肺癌（NSCLC）顺铂（DDP）耐药的候选DNA甲基化驱动因素。A549/DDP细胞系是通过将A549细胞连续暴露于递增浓度的DDP来建立的。基因表达和甲基化分析通过高通量微阵列确定。在原代肿瘤细胞培养物和癌症基因组图谱（TCGA）样本中验证了甲基化状态与DDP反应的关系。在体外和体内测定细胞增殖、凋亡、细胞周期和对DDP的反应。在A549/DDP细胞中，共有372个基因发生了甲基化和下调，这些基因参与了大多数基本的生物学过程。筛选出10个候选基因（S100 P、GDA、WISP 2、LOXL 1、TIMP 4、ICAM 1、CLMP、HSP 8、GAS 1、BMP 2），它们与顺铂耐药有不同程度的关联。低剂量5-Aza-dC和TSA联合应用可逆转A549/DDP细胞的耐药性，沿着候选基因（GAS 1、TIMP 4、ICAM 1和WISP 2）的去甲基化和恢复表达。通过基因转染法强制表达GAS 1可使A549/DDP细胞对DDP的敏感性增加、增殖抑制、细胞周期阻滞、凋亡增强和体内生长迟缓。总之，我们的研究表明，一组候选基因下调的DNA甲基化诱导顺铂耐药的NSCLC，并表明表观遗传治疗重新敏感的细胞顺铂。
DNA methylation plays a critical role during the development of acquired chemoresistance. The aim of this study was to identify candidate DNA methylation drivers of cisplatin (DDP) resistance in non-small cell lung cancer (NSCLC). The A549/DDP cell line was established by continuous exposure of A549 cells to increasing concentrations of DDP. Gene expression and methylation profiling were determined by high-throughput microarrays. Relationship of methylation status and DDP response was validated in primary tumor cell culture and the Cancer Genome Atlas (TCGA) samples. Cell proliferation, apoptosis, cell cycle, and response to DDP were determined in vitro and in vivo. A total of 372 genes showed hypermethylation and downregulation in A549/DDP cells, and these genes were involved in most fundamental biological processes. Ten candidate genes (S100P, GDA, WISP2, LOXL1, TIMP4, ICAM1, CLMP, HSP8, GAS1, BMP2) were selected, and exhibited varying degrees of association with DDP resistance. Low dose combination of 5-aza-2'-deoxycytidine (5-Aza-dC) and trichostatin A (TSA) reversed drug resistance of A549/DDP cells in vitro and in vivo, along with demethylation and restoration of expression of candidate genes (GAS1, TIMP4, ICAM1 and WISP2). Forced expression of GAS 1 in A549/DDP cells by gene transfection contributed to increased sensitivity to DDP, proliferation inhibition, cell cycle arrest, apoptosis enhancement, and in vivo growth retardation. Together, our study demonstrated that a panel of candidate genes downregulated by DNA methylation induced DDP resistance in NSCLC, and showed that epigenetic therapy resensitized cells to DDP.