UMR 144 CNRS/IC, Institut Curie, 26 rue d'Ulm, 75248 Paris Cedex 05, France.
J Natl Cancer Inst. 2011 Jan 5;103(1):47-60. doi: 10.1093/jnci/djq470. Epub 2010 Dec 20.
Epigenetic silencing can extend to whole chromosomal regions in cancer. There have been few genome-wide studies exploring its involvement in tumorigenesis.
We searched for chromosomal regions affected by epigenetic silencing in cancer by using Affymetrix microarrays and real-time quantitative polymerase chain reaction to analyze RNA from 57 bladder tumors compared with normal urothelium. Epigenetic silencing was verified by gene re-expression following treatment of bladder cell lines with 5-aza-deoxycytidine, a DNA demethylating agent, and trichostatin A, a histone deacetylase inhibitor. DNA methylation was studied by bisulfite sequencing and histone methylation and acetylation by chromatin immunoprecipitation. Clustering was used to distinguish tumors with multiple regional epigenetic silencing (MRES) from those without and to analyze the association of this phenotype with histopathologic and molecular types of bladder cancer. The results were confirmed with a second panel of 40 tumor samples and extended in vitro with seven bladder cancer cell lines. All statistical tests were two-sided.
We identified seven chromosomal regions of contiguous genes that were silenced by an epigenetic mechanism. Epigenetic silencing was not associated with DNA methylation but was associated with histone H3K9 and H3K27 methylation and histone H3K9 hypoacetylation. All seven regions were concordantly silenced in a subgroup of 26 tumors, defining an MRES phenotype. MRES tumors exhibited a carcinoma in situ-associated gene expression signature (25 of 26 MRES tumors vs 0 of 31 non-MRES tumors, P < 10⁻¹⁴), rarely carried FGFR3 mutations (one of 26 vs 22 of 31 non-MRES tumors, P < 10⁻¹⁶), and contained 25 of 33 (76%) of the muscle-invasive tumors. Cell lines derived from aggressive bladder tumors presented epigenetic silencing of the same regions.
We have identified an MRES phenotype characterized by the concomitant epigenetic silencing of several chromosomal regions, which, in bladder cancer, is specifically associated with the carcinoma in situ gene expression signature.
表观遗传沉默可延伸至癌症中的整个染色体区域。很少有全基因组研究探索其在肿瘤发生中的作用。
我们使用 Affymetrix 微阵列和实时定量聚合酶链反应，分析了 57 例膀胱癌与正常尿路上皮相比的 RNA，寻找受表观遗传沉默影响的染色体区域。通过用 DNA 去甲基化剂 5-氮杂-2′-脱氧胞苷和组蛋白去乙酰化酶抑制剂曲古抑菌素 A 处理膀胱细胞系，来验证表观遗传沉默。通过亚硫酸氢盐测序和组蛋白甲基化和乙酰化染色质免疫沉淀研究 DNA 甲基化。聚类用于区分具有多个区域性表观遗传沉默（MRES）的肿瘤与没有 MRES 的肿瘤，并分析这种表型与膀胱癌的组织病理学和分子类型的关联。使用第二组 40 个肿瘤样本进行验证，并在体外通过 7 个膀胱癌细胞系进行扩展。所有统计检验均为双侧检验。
我们鉴定了七个连续基因的染色体区域，这些区域被表观遗传机制沉默。表观遗传沉默与 DNA 甲基化无关，但与组蛋白 H3K9 和 H3K27 甲基化以及组蛋白 H3K9 低乙酰化有关。在一个包括 26 个肿瘤的亚组中，所有七个区域都被一致地沉默，定义了一个 MRES 表型。MRES 肿瘤表现出原位癌相关的基因表达特征（26 个 MRES 肿瘤中有 25 个，而非 MRES 肿瘤中有 0 个，P<10⁻¹⁴），很少携带 FGFR3 突变（26 个中有 1 个，而非 MRES 肿瘤中有 31 个中有 22 个，P<10⁻¹⁶），并且包含 33 个中的 25 个（76%）肌层浸润性肿瘤。来源于侵袭性膀胱癌的细胞系表现出相同区域的表观遗传沉默。
我们鉴定了一种 MRES 表型，其特征是几个染色体区域的同时表观遗传沉默，在膀胱癌中，该表型与原位癌基因表达特征特异性相关。
