PADI2对雌激素应答基因转录调控机制及其在乳腺癌发生发展中的作用

Breast cancer is the most common malignant tumor in women, and lifetime exposure to estrogen constitutes a major risk factor for breast cancer initiation and development, though the detailed molecular mechanisms are far more from complete. Accumulating evidence suggest that some cofactors for estrogen receptor α (ERα) can modulate target gene activities in estrogen-ERα signaling pathway by posttranslationally modifying histone tails at gene promoters, therefore, drugs targeting such epigenetic modifiers which are often overexpressed in hormonal breast cancers can be potentially used for therapeutic targeting. In our previous studies, for the first time, we identified that peptidylarginine deiminase (PADI) 2 catalyzes citrullination of histone H3 arginine 26 (H3R26) at decondensed chromatin loci surrounding the estrogen-response elements of target promoters and promotes target gene transcription upon estrogen stimulation; Our preliminary data also showed that PADI enzyme inhibitor, Cl-amidine, inhibits MCF-7 cell proliferation, suggesting that PADI2, which plays an important role in ERα signaling, represents a promising target for hormonal breast cancer treatment. To prove this hypothesis, we propose to further explore the molecular mechanisms underlying the interaction between PADI2 and ERα, and the role PADI2 plays in regulation of estrogen response gene transcription in ERα-positive breast cancer cells. Then, the expression of PADI2 and PADI2-catalyzed H3Cit26 will be assessed in breast caner progression tissue microarray in order to indicate the relevance to the clinical progression of breast cancer malignant behavior. Finally, Cl-amidine will be used to treat ERα-positive breast cancer cells, especially the cells that are resistant to endocrine therapy. Our proposals may provide both novel epigenetic target and treatment regimen(s) in breast cancer to overcome endocrine drug resistance.

长期雌激素(E2)刺激是乳腺癌的一个主要诱发因素,机制尚未彻底阐明。研究表明,E2受体(ERα)辅助因子在ERα通路中发挥重要作用，其抑制剂是乳腺癌治疗新靶点。我们前期研究就发现这样一个因子，即肽基精氨酸脱亚胺酶II(PADI2)，它通过催化组蛋白H3 N-末端第26位精氨酸的鸟氨酸化(H3Cit26)促进E2应答元件区染色质开放，影响靶基因转录；我们预试验也证实PADI抑制剂能抑制乳腺癌细胞增殖,提示PADI2可能成为乳腺癌防治新靶点。为验证此假说,我们将进一步阐明PADI2与ERα相互作用机制,揭示PADI2在ERα阳性乳腺癌细胞中基因转录调控分子机理；在此基础上，研究PADI2及H3Cit26与乳腺癌发生发展关系,观察PADI抑制剂对ERα阳性乳腺癌细胞及对内分泌治疗耐受乳腺癌细胞的影响。本课题将为乳腺癌,尤其对内分泌治疗耐受乳腺癌的防治提供新的思路和药物靶点，弥补内分泌治疗的不足。

雌激素受体(ERα)辅助因子在ERα通路中发挥重要作用，其抑制剂是乳腺癌治疗新靶点。其中肽基精氨酸脱亚胺酶II(PADI2)是我们最近发现的一个ERα辅助因子，它通过催化组蛋白H3 N-末端第26位精氨酸的鸟氨酸化(H3Cit26)促进E2应答元件区染色质开放，影响靶基因转录。为进一步阐明PADI2与ERα相互作用机制,揭示PADI2在ERα阳性乳腺癌细胞中基因转录调控分子机理以及PAD12及其催化的组蛋白修饰在乳腺肿瘤中的作用，我们首先证实了PADI2 的LLXXL蛋白结构域参与雌激素受体结合，并发现一种新的PADI2蛋白复合体组分成员，参与雌激素受体信号通路在雌激素处理下的激活。雌激素作用前，带正电的H3R26由于与雌激素应答元件结合阻碍了雌激素受体与雌激素应答元件的结合；当给予雌激素作用后，由于 H3R26的正电被中和，改变了组蛋白与核小体DNA的结合，暴露出雌激素应答元件，从而使得雌激素受体更容易结合。雌激素作用早期，H3Cit26与转录活化基因的增强子区结合，而且与基因转录调控区的P300蛋白结合位点重叠，共同调控雌激素早期应答基因的转录。由于H3R26cit能中和组蛋白H3尾的正电荷，从而改变染色体的空间结构，以利于雌激素受体募集转录复合体，在雌激素应答基因活化中起到重要的调控作用，我们的研究为雌激素应答基因的转录活化提供了新的机制。鉴于H3Cit26以及雌激素受体在雌激素受体阳性乳腺癌细胞中高度特异的相关性，我们推测其可能在临床乳腺肿瘤标本中可能也存在相关性。通过临床样本以及乳腺癌肿瘤数据库分析，发现H3Cit26以及PAD2在预测Luminal A型乳腺肿瘤中可能会成为一个好的检测标记分子。体外细胞药物处理试验证明，PADI抑制剂联合三苯氧胺对乳腺癌细胞，尤其是对三苯氧胺耐药MCF-7乳腺癌细胞的增殖起到抑制作用，细胞周期阻滞在S期，细胞凋亡明显增加，说明PADI抑制剂联合三苯氧胺可能是三苯氧胺耐药乳腺癌的一种有效治疗手段。