Mechanistic role of P4501 enzymes in the prevention of PAH carcinogenesis by omega 3 fatty acids

项目摘要 细胞色素P450（CYP）是参与药物代谢和生物活化的主要酶。公 已知几种脂肪酶将ω-3脂肪酸代谢成它们的环氧代谢物， 血管生成、肿瘤生长和转移。许多多环芳烃（PAH）是人类 致癌物质。PAH-DNA加合物可能导致DNA损伤和关键基因突变，最终导致 到癌症PAH-DNA加合物水平与肿瘤发生率之间存在显著的正线性回归 在我们实验室的动物实验中观察到。我们还发现ω-3脂肪酸 二十碳五烯酸（EPA）、二十二碳六烯酸（DHA）抑制CYP 1B 1、EZH 2、DNMT 3a、miR 17， miR 19 b-1和显著降低肺和肝PAH-DNA加合物以及肿瘤发生率。的 本申请的中心假设是ω 3-脂肪酸和它们的环氧代谢物将减弱 肺癌的多种机制，需要衰减PAH-DNA加合物的形成， 调节CYP以及抑制肿瘤发生，可能通过调节表观遗传基因 (e.g., EZH2、DNMT3a、miR-17、miR-19b-1）。我们提出以下具体目标。目标1：测试 CYP 1A 1和CYP 1B 1在预防小鼠PAH致癌中机制假说 与一氧化碳饮食相比， 这些小鼠对BP的作用机制进行了研究。目的2：验证小鼠可溶性胰岛素缺乏的假设 环氧化物水解酶（sEH）对EPA/DHA介导的PAH预防作用比WT小鼠更强 致癌作用，因为已知sEH快速水解环氧代谢物，如17，18-环氧二十碳四烯酸 19，20-环氧二十二碳五烯酸（EDP）转化为无活性代谢物。在 在一些实验中，我们将用特异性sEH抑制剂t-TUCB或新的t-TUCB样抑制剂治疗WT小鼠 (that可能很快进入人体临床试验），然后用EPA/DHA和BP治疗小鼠。目标3： 为了验证内源性ω-3脂肪酸，特别是其环氧代谢产物，将在体内发挥作用的假设， 在体内多环芳烃预防肺癌发生中起着关键作用，并且存在机制联系 CYP 1和sEH之间。Fat-1-转基因（Fat-1-Tg）小鼠，其将内源性ω-6脂肪酸转化为 脂肪酸（富含CO）转化为omega-3脂肪酸并降低omega-6/omega-3的比例，将用于此 study.我们还将建立Fat-1-Tg/sEH基因敲除小鼠，以探讨CYP 1和sEH在体内表达的机制。 酶有助于ω-3脂肪酸介导的PAH致癌预防。如果我们假设 CYP 1和sEH酶在ω-3脂肪酸中起重要作用，即EPA/DHA介导的预防 PAH诱导的癌症被证明是正确的，那么它将在目前的理解中开辟新的领域， 人类癌症预防。如果成功，拟议的研究将导致饮食中的新机制。 对多环芳烃诱发肺癌的干预措施。

PROJECT SUMMARY The cytochrome P450s (CYPs) are the major enzymes involved in drug metabolism and bioactivation. It is well known that several CYP enzymes metabolize omega-3 fatty acids to their epoxy metabolites that inhibit angiogenesis, tumor growth, and metastasis. Numerous polycyclic aromatic hydrocarbons (PAH) are human carcinogens. PAH-DNA adducts may lead to DNA damage and mutations in critical genes, eventually leading to cancer. A significant positive linear regression between levels of PAH-DNA adducts and tumor incidence was observed in animal experiments in our laboratory. We also discovered that omega-3 fatty acids eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) inhibited CYP1B1, EZH2, DNMT3a, miR 17, miR19b-1 and significantly decreased pulmonary and hepatic PAH-DNA adducts, and tumor incidence. The central hypothesis of this application is that omega 3-fatty acids and their epoxy metabolites will attenuate pulmonary carcinogenesis by multiple mechanisms entailing attenuation of PAH-DNA adduct formation by modulating CYPs, as well as by suppression of tumorigenesis, probably via modulation of epigenetic genes (e.g., EZH2, DNMT3a, miR-17, miR-19b-1). We propose the following Specific Aims. Aim 1: To test the hypothesis that CYP1A1 and CYP1B1 play mechanistic roles in prevention of PAH carcinogenesis in mice maintained on EPA, DHA, or EPA + DHA diets, compared to those on a CO diet, followed by exposure of these mice to BP for the study of the mechanisms. Aim 2: To test the hypothesis that mice deficient in soluble epoxide hydrolase (sEH) will confer more protection than WT mice to EPA/DHA-mediated prevention of PAH carcinogenesis, as sEH is known to rapidly hydrolyze epoxy metabolites such as 17,18-epoxy eicosatetraenoic acid (EEQ) and 19,20-epoxy docosapentaenoic acids (EDP) in serum and tissues to inactive metabolites. In some experiments, we will treat WT mice with the specific sEH inhibitor, t-TUCB, or a new t-TUCB-like inhibitor (that is likely to go to human clinical trials soon), followed by treatment of mice with EPA/DHA and BP. Aim 3: To test the hypothesis that endogenous omega-3 fatty acids, especially their epoxy metabolites, will play a pivotal role in the prevention of pulmonary carcinogenesis by PAHs in vivo, and that there is a mechanistic link between CYP1, and sEH. Fat-1-transgenic (Fat-1-Tg) mice, which will convert endogenous omega-6 fatty acids (rich in CO) into omega-3 fatty acids and decrease the ratios of omega-6/omega-3, will be used in this study. We will also create Fat-1-Tg/sEH-null mice for exploring the mechanisms by which CYP1 and sEH enzymes contribute to omega-3 fatty acid-mediated prevention of PAH carcinogenesis. If our hypothesis that CYP1 and sEH enzymes play important roles in omega-3 fatty acids, i.e. EPA/DHA-mediated prevention of PAH-induced cancers turns out to be correct, then it will break new grounds in the current understanding of human cancer prevention. If successful, the proposed studies should lead to novel mechanisms in dietary interventions against lung cancers induced by PAHs.