Hinshelwood Rebecca A, Huschtscha Lily I, Melki John, Stirzaker Clare, Abdipranoto Andrea, Vissel Bryce, Ravasi Timothy, Wells Christine A, Hume David A, Reddel Roger R, Clark Susan J
Garvan Institute of Medical Research, Darlinghurst, New South Wales, Australia.
Cancer Res. 2007 Dec 15;67(24):11517-27. doi: 10.1158/0008-5472.CAN-07-1284.
Human mammary epithelial cells (HMEC) grown under standard cell culture conditions enter a growth phase referred to as selection, but a subpopulation is able to escape from arrest and continue to proliferate. These cells, called post-selection or variant HMECs, may be derived from progenitor cells found in normal mammary epithelium that subsequently acquire premalignant lesions, including p16(INK4A) promoter hypermethylation. Epigenetic silencing of tumor suppressor genes through DNA methylation and histone modification is an early event in tumorigenesis. A major challenge is to find genes or gene pathways that are commonly silenced to provide early epigenetic diagnostic and therapeutic cancer targets. To identify very early epigenetic events that occur in breast cancer, we used microarrays to screen for gene pathways that were suppressed in post-selection HMECs but reactivated after treatment with the demethylation agent 5-aza-2'-deoxycytidine. We found that several members of the transforming growth factor beta (TGF-beta) signaling pathway were consistently down-regulated in the post-selection HMEC populations, and this was associated with a marked decrease in Smad4 nuclear staining. Gene suppression was not associated with DNA methylation but with chromatin remodeling, involving a decrease in histone H3 lysine 27 trimethylation and an increase in histone H3 lysine 9 dimethylation and deacetylation. These results show for the first time that TGF-beta2, its receptors TGF-beta R1 and TGF-beta R2, and activator thrombospondin-1 are concordantly suppressed early in breast carcinogenesis by histone modifications and indicate that the TGF-beta signaling pathway is a novel target for gene activation by epigenetic therapy.
在标准细胞培养条件下生长的人乳腺上皮细胞（HMEC）进入一个称为“筛选”的生长阶段，但有一个亚群能够摆脱停滞并继续增殖。这些细胞，称为筛选后或变异型HMEC，可能源自正常乳腺上皮中的祖细胞，随后获得包括p16(INK4A)启动子高甲基化在内的癌前病变。通过DNA甲基化和组蛋白修饰实现肿瘤抑制基因的表观遗传沉默是肿瘤发生的早期事件。一个主要挑战是找到通常被沉默的基因或基因途径，以提供早期表观遗传诊断和治疗癌症靶点。为了鉴定乳腺癌中发生的非常早期的表观遗传事件，我们使用微阵列筛选在筛选后HMEC中被抑制但在用去甲基化剂5-氮杂-2'-脱氧胞苷处理后重新激活的基因途径。我们发现转化生长因子β（TGF-β）信号通路的几个成员在筛选后HMEC群体中始终下调，这与Smad4核染色的显著减少相关。基因抑制与DNA甲基化无关，而是与染色质重塑有关，涉及组蛋白H3赖氨酸27三甲基化的减少以及组蛋白H3赖氨酸9二甲基化和去乙酰化的增加。这些结果首次表明，TGF-β2及其受体TGF-β R1和TGF-β R2以及激活剂血小板反应蛋白-1在乳腺癌发生早期通过组蛋白修饰被一致抑制，并表明TGF-β信号通路是表观遗传治疗激活基因的新靶点。
