ABSTRACT

Testicular germ cell tumours (TGCTs) respond well to cisplatin-based therapy. However, cisplatin resistance and poor outcomes do occur. It has been suggested that a shift towards DNA hypermethylation mediates cisplatin resistance in TGCT cells, although there is little direct evidence to support this claim. Here we utilized a series of isogenic cisplatin-resistant cell models and observed a strong association between cisplatin resistance in TGCT cells and a net increase in global CpG and non-CpG DNA methylation spanning regulatory, intergenic, genic and repeat elements. Hypermethylated loci were significantly enriched for repressive DNA segments, CTCF and RAD21 sites and lamina associated domains, suggesting that global nuclear reorganization of chromatin structure occurred in resistant cells. Hypomethylated CpG loci were significantly enriched for EZH2 and SUZ12 binding and H3K27me3 sites. Integrative transcriptome and methylome analyses showed a strong negative correlation between gene promoter and CpG island methylation and gene expression in resistant cells and a weaker positive correlation between gene body methylation and gene expression. A bidirectional shift between gene promoter and gene body DNA methylation occurred within multiple genes that was associated with upregulation of polycomb targets and downregulation of tumour suppressor genes. These data support the hypothesis that global remodelling of DNA methylation is a key factor in mediating cisplatin hypersensitivity and chemoresistance of TGCTs and furthers the rationale for hypomethylation therapy for refractory TGCT patients.

摘要

睾丸生殖细胞肿瘤 (TGCT) 对基于顺铂的治疗反应良好。然而，顺铂耐药和不良结果确实发生。有人提出，向 DNA 高甲基化的转变介导 TGCT 细胞中的顺铂耐药性，尽管几乎没有直接证据支持这一说法。在这里，我们使用了一系列同基因顺铂耐药细胞模型，并观察到 ​​TGCT 细胞中顺铂耐药与跨越调节、基因间、基因和重复元件的全局 CpG 和非 CpG DNA 甲基化净增加之间的强关联。抑制性 DNA 片段、CTCF 和 RAD21 位点以及纤层相关域的高甲基化位点显着富集，表明染色质结构的全球核重组发生在抗性细胞中。低甲基化 CpG 位点显着富集 EZH2 和 SUZ12 结合位点和 H3K27me3 位点。整合转录组和甲基化组分析显示，基因启动子与 CpG 岛甲基化和抗性细胞中基因表达之间存在很强的负相关，而基因体甲基化与基因表达之间存在较弱的正相关。基因启动子和基因体 DNA 甲基化之间的双向转变发生在多个基因中，这与多梳靶标的上调和肿瘤抑制基因的下调有关。这些数据支持这样的假设，即 DNA 甲基化的整体重塑是介导 TGCT 的顺铂超敏反应和化学抗性的关键因素，并进一步说明了难治性 TGCT 患者的低甲基化治疗的基本原理。整合转录组和甲基化组分析显示，基因启动子与 CpG 岛甲基化和抗性细胞中基因表达之间存在很强的负相关，而基因体甲基化与基因表达之间存在较弱的正相关。基因启动子和基因体 DNA 甲基化之间的双向转变发生在多个基因中，这与多梳靶标的上调和肿瘤抑制基因的下调有关。这些数据支持这样的假设，即 DNA 甲基化的整体重塑是介导 TGCT 的顺铂超敏反应和化学抗性的关键因素，并进一步说明了难治性 TGCT 患者的低甲基化治疗的基本原理。整合转录组和甲基化组分析显示，基因启动子与 CpG 岛甲基化和抗性细胞中基因表达之间存在很强的负相关，而基因体甲基化与基因表达之间存在较弱的正相关。基因启动子和基因体 DNA 甲基化之间的双向转变发生在多个基因中，这与多梳靶标的上调和肿瘤抑制基因的下调有关。这些数据支持这样的假设，即 DNA 甲基化的整体重塑是介导 TGCT 的顺铂超敏反应和化学抗性的关键因素，并进一步说明了难治性 TGCT 患者的低甲基化治疗的基本原理。