Human CYP2A and respiratory tract xenobiotic toxicity

说明(申请人提供)：本修订提案的研究目标与降低烟草产品和烟雾的毒性有关，这是与《家庭吸烟预防和烟草控制法案》相关的四个领域之一。具体地说，我们试图更好地了解香烟烟雾(CS)诱导小鼠模型肺肿瘤发生的机制，以提高小鼠模型在测试烟草产品致癌性方面的实用性。我们建议使用PI实验室开发的两个新的转基因小鼠模型来验证以下假设：1)抑制肝脏细胞色素P450(P450)家族的生物转换酶活性会导致循环和组织水平的增加 2)CS诱导的肺肿瘤形成依赖于P450酶对烟草致癌物的靶组织生物激活。这两个小鼠模型都针对细胞色素P450还原酶(CPR)，细胞色素P450还原酶是所有微粒体P450酶活性所必需的酶，它们是肝CPR缺失(LCN)小鼠和肝外CPR低(或xh-CL)小鼠，在LCN小鼠中，CPR在肝细胞中缺失，而在肝外CPR低(或XH-CL)小鼠中，CPR在所有肝外组织(包括肺)中的表达显著降低。提出了两个具体目标。在目标1中，我们将确定肝脏P450活性在CS诱导的肺肿瘤形成中的作用。我们将比较LCN A/J和野生型(WT)A/J小鼠对CS诱导的肺肿瘤形成的敏感性，以及CS诱导的主要烟草致癌物在体内形成的DNA加合物的水平。在肿瘤生物检测中显示更强的致瘤反应的能力将使该检测方法应用于测试新的“减毒”烟草产品以及测试肺肿瘤化学预防的有效性变得更加实用。在目标2中，我们将确定CS诱导的AJ小鼠肺部肿瘤的发生是否依赖于靶组织中P450介导的生物激活。我们将比较xh-CL A/J和WT A/J小鼠对CS诱导的肺肿瘤形成的敏感性，以及CS诱导的肺和肝脏中DNA加合物的水平。预期的结果将解决这一争议，即在CS暴露的小鼠模型中观察到的肺肿瘤是否是CS通过依赖于P450介导的前致癌物的生物激活的遗传毒性途径直接引发肿瘤，而不是CS增加自发肿瘤的促进。 公共卫生相关性：更好地了解吸烟导致小鼠肺肿瘤发生的潜在机制，将增加人们对动物模型患肺癌的预测价值的信心，肺癌是美国癌症相关死亡的主要原因。小鼠模型对香烟诱导的肺肿瘤发生敏感性的普遍提高，将使小鼠肺肿瘤生物测定更适用于各种“减毒”烟草产品的测试，以及肺肿瘤化学预防的有效性测试。

DESCRIPTION (provided by applicant): The research goal of this revision proposal is related to Reducing Toxicity of Tobacco Products and Smoke, one of the four areas relevant to the Family Smoking Prevention and Tobacco Control Act. Specifically, we seek to gain a better understanding of the mechanisms underlying cigarette smoke (CS)-induced lung tumorigenesis in mouse models, in order to improve the utility of mouse models for testing the carcinogenicity of tobacco products. We propose to use two novel transgenic mouse models developed in the PI's laboratory to test the hypotheses that 1) suppression of hepatic cytochrome P450 (P450) family of biotransformation enzyme activity will lead to increased circulating and tissue levels of CS carcinogens and, consequently, increased lung DNA damage and tumorigenesis in mice exposed chronically to CS, and 2) CS-induced lung tumorigenesis depends on target-tissue bioactivation of tobacco carcinogens by P450 enzymes. The two mouse models, both targeting the cytochrome P450 reductase (CPR), an enzyme required for the activities of all microsomal P450 enzymes, are the liver-Cpr-null (LCN) mouse, in which Cpr expression is absent in hepatocytes, and the extra-hepatic Cpr-low (or xh-CL) mouse, in which Cpr expression, and thus P450 activities, in all extrahepatic tissues (including the lung) are greatly decreased. Two Specific Aims are proposed. In Aim 1, we will determine the impact of hepatic P450 activity on CS-induced lung tumorigenesis. We will compare LCN A/J and wild-type (WT) A/J mice for sensitivity to CS-induced lung tumorigenesis, and for levels of CS-induced lung DNA adducts formed by major tobacco carcinogens in vivo. The ability to show increased tumorigenic responses in tumor bioassays would make it more practical to apply the assays to testing of new "reduced-toxicity" tobacco products, as well as for testing efficacy of lung tumor chemoprevention. In Aim 2, we will determine whether CS-induced lung tumorigenesis in AJ mouse depends on P450-mediated bioactivation in the target tissue. We will compare xh-CL A/J and WT A/J mice for sensitivity to CS-induced lung tumorigenesis, and for levels of CS-induced DNA adducts in the lungs and livers. The anticipated results will resolve the controversy about whether the lung tumors observed in CS-exposed mouse models are derived from direct tumor initiation by CS, through genotoxic pathways that depend on P450- mediated bioactivation of procarcinogens, as opposed to increased promotion by CS of spontaneous tumors. PUBLIC HEALTH RELEVANCE: A better understanding of the mechanisms underlying the cigarette smoke-induced lung tumorigenesis in the mouse would increase confidence in the predictive value of the animal model for the human risks of developing lung cancer, which is the leading cause of cancer-related death in the U.S. General improvements in the sensitivity of the mouse models to cigarette smoke-induced lung tumorigenesis would make mouse lung tumor bioassays more applicable to the testing of various "reduced-toxicity" tobacco products, as well as for testing efficacy of lung tumor chemoprevention.