Reactivation of GSTP1 Gene by Green Tea Polyphenols

描述（由申请人提供） 通过DNA甲基化的基因表达修饰和通过组蛋白质的染色质重塑被认为是在前列腺癌的发生和发展中观察到的最重要的表观遗传变化。一种这样的改变是通过5 '调控区CpG岛中脱氧胞苷残基的甲基化沉默谷胱甘肽S-转移酶-pi（GSTP 1）（致癌物防御的关键酶）的表达。GSTP 1功能的丧失似乎是前列腺上皮内瘤病变的特征，始终代表前列腺癌的前体。近年来，绿色茶作为癌症预防剂已经获得了相当大的兴趣，并且我们最近已经证明，以人类可达到的剂量口服消耗绿色茶多酚（GTP）导致小鼠前列腺转基因腺癌（TRAMP）模型中前列腺癌发生的显著抑制，并且重要地导致这些小鼠的无肿瘤和总体存活率增加（Proc.Natl.Acad.Sci. Acad. Sci. USA 98：10350-5，2001）。这项研究提供了理论基础，绿色茶可以开发为补充和替代药物（CAM）的前列腺癌。 在细胞培养系统和有限的动物模型中进行的广泛的实验室研究进一步证明，绿色茶多酚对不同类型的致癌物具有保护作用，并诱导II期酶活性，这可能导致增强的解毒过程。重要的是，如果喝绿色茶可以恢复或补偿前列腺中的GST 1活性，并可以减缓前列腺癌发生的过程，这可能与预防和/或治疗这种疾病有关。由于DNA甲基化受DNA甲基转移酶（DNMT）和DNA组蛋白去乙酰化酶（HDAC）的相对活性的影响，因此本发明中待测试的创新想法是绿色茶多酚具有通过调节人前列腺癌LNCaP和MDA PCa 2b细胞中DNMT和HDAC的酶活性来重新激活GST 1表达的潜力。重要的是，这些酶的基础水平比良性前列腺增生和正常人前列腺上皮细胞高2 - 3倍，并含有完全高甲基化的GSTP 1CpG岛等位基因.具体而言，我们将研究绿色茶多酚在以下方面的潜力：i）GSTP 1基因启动子区域中高甲基化CpG岛等位基因的去甲基化及其再激活，ii）DNMT和HDAC活性和表达的调节，以及iii）LNCaP和MDA PCa 2b细胞中总组蛋白H3和乙酰化组蛋白H4蛋白表达的改变。该提案的成功完成将建立一个平台，以确定绿色茶多酚在调节前列腺癌中的表观遗传事件中的作用，这可能有助于绿色茶作为CAM用于前列腺癌患者的发展。

DESCRIPTION (provided by applicant) Modifications in gene expression through DNA methylation and remodeling of chromatin via histone proteins are believed to be the most important epigenetic changes observed in initiation and development of prostate cancer. One such alteration is silencing of the expression of glutathione S-transferase-pi (GSTP1), a critical enzyme of carcinogen defense, through methylation of deoxycytidine residue in CpG islands in the 5'- regulating region. Loss of GSTP1 function appears to be the characteristic of prostatic intraepithelial neoplasia lesions, throughout to represent prostate cancer precursors. In recent years, green tea has gained considerable interest as a cancer preventive agent and we have recently demonstrated that oral consumption of green tea polyphenols (GTP) at human achievable dose results in significant inhibition of prostate carcinogenesis in transgenic adenocarcinoma of the mouse Prostate (TRAMP) model and importantly lead to increased tumor free and overall survival of these mice (Proc. Natl. Acad. Sci. USA 98: 10350-5, 2001). This study provides rationale that green tea could be developed as complementary and alternative medicine (CAM) for prostate cancer. Extensive laboratory studies in cell culture systems and in limited animal models have further demonstrated that green tea polyphenols afford protective effects from diverse types of carcinogens and induce phase II enzyme activity that could lead to enhanced detoxification process. Importantly, if drinking green tea can restore or compensate the GSTP1 activity in the prostate and can slow down the process of prostate carcinogenesis that may have relevance for prevention and/or treatment of this disease. Since DNA methylation is influenced by the relative activities of DNA methyltransferases (DNMT) and DNA histone deacetylases (HDAC), the innovative idea to be tested in this proposal is that green tea polyphenols has potential to reactivate the expression of GSTP1 by modulating the enzyme activity of DNMT and HDAC in human prostate carcinoma LNCaP and MDA PCa 2b cells. Importantly, the basal levels of these enzymes are two- to three- folds higher than benign prostatic hyperplasia and normal human prostate epithelial cells and contain completely hypermethylated GSTP1 CpG island alleles. Specifically, we will investigate the potential of green tea polyphenols in i) demethylation of the hypermethylated CpG island alleles in the promoter region of GSTP1 gene and its reactivation, ii) modulation of DNMT and HDAC activity and expression, and iii) alteration in the total and acetylated histone H3 and H4 protein expression in LNCaP and MDA PCa 2b cells. Successful completion of this proposal will set a platform to define the role of green tea polyphenols in modulating epigenetic events in prostate cancer that may contribute to the development of green tea as CAM for its use in prostate cancer patients.