Role of local P450 enzymes in chemical carcinogenesis in mouse mammary gland

描述（由申请人提供）：长期目标是确定乳房组织中化学癌变的机制。我们目前的重点是利用组织选择性基因敲除小鼠模型研究乳腺细胞色素P450 （P450）酶在化学致癌物诱导的乳腺DNA加合物形成中的作用。有充分证据表明，微粒体细胞色素P450 （P450）酶在化学致癌物原的代谢激活中起关键作用。然而，很难确定在全身暴露于某一致癌物后，乳腺组织中的大部分活性致癌物或其DNA加合物是来自局部代谢还是来自肝脏；在前致癌物的代谢激活中，后一器官通常比肝外靶组织更活跃。考虑到肝脏或乳腺组织不仅对最终致癌物的形成有潜在的贡献，而且对中间代谢物，如dba -反式3,4-二氢二醇（3,4-二醇-DMBA）的形成也有潜在的贡献，如7,12-二甲基苯[a]蒽（DMBA），它们是最终致癌物的前体，因此，对于需要多个代谢激活步骤的致癌物，问题变得更加复杂。我们的假设是靶组织P450介导的代谢激活在dba诱导的乳腺（MG） DNA加合物的形成中起重要作用；然而，这种靶组织贡献的程度将根据MG的发育阶段和生理状态而变化。为了验证这一假设，我们将通过条件删除（或抑制）P450还原酶（Cpr）基因来调节肝脏和肝外组织（包括MG）中的P450活性，P450还原酶是所有微粒体P450酶活性所必需的酶。在这项为期两年的试点项目研究中，我们将确定肝脏和肝外（包括MG） P450酶在DMBA（乳腺前致癌物）及其中间代谢产物3,4-二醇DMBA的代谢激活中的相对贡献，在新开发的P450活性组织特异性抑制的小鼠模型中，无论是在肝脏中（指定为肝脏-无crp小鼠），还是在除肝脏外的所有组织中（指定为肝外-低crp小鼠）（Aim 1）。此外，我们将开发和表征MG选择性Cpr无效模型（指定为MG-Cpr无效小鼠），以直接确定MG P450酶在DMBA和3,4-二醇-DMBA诱导的MG dna加合物形成中的作用（目的2）。从研究中获得的知识将改进流行病学研究的设计，这些研究试图将环境化学暴露和/或P450酶的遗传多态性与人类乳腺癌的发病率联系起来。它也可能有助于设计基于机制的方法来预防乳腺癌。

DESCRIPTION (provided by applicant): The long-term objective is to determine the mechanisms of chemical carcingensis in the breast tissue. Our current focus is to study the role of mammary cytochrome P450 (P450) enzymes in chemical carcinogen- induced DNA adduct formation in the mammary gland (MG) using tissue-selective gene knockout mouse models. It is well-documented that the microsomal cytochrome P450 (P450) enzymes play a critical role in the metabolic activation of chemical procarcinogens. However, it has been difficult to determine whether, following systemic exposure to a given procarcinogen, the bulk of the activated carcinogen, or its DNA adducts, in the breast tissue is derived from local metabolism or from the liver; the latter organ is generally much more active than extrahepatic target tissues in the metabolic activation of procarcinogens. The problem becomes even more complicated for procarcinogens, such as 7,12-dimethylbenz[a]anthracene (DMBA), that require multiple steps of metabolic activation, given the potential contributions of hepatic or breast tissues to the formation of not only the ultimate carcinogen, but also the intermediate metabolites, such as DMBA-trans-3,4-dihydrodiol (3,4-diol-DMBA), that are precursors to the ultimate carcinogen. Our hypothesis is that target-tissue P450- mediated metabolic activation plays an important role in DMBA-induced formation of DNA adducts in the mammary gland (MG); however, the extent of this target-tissue contribution will vary according to the developmental stages and physiological states of the MG. To test this hypothesis, we will modulate P450 activities in the liver and extrahepatic tissues (including MG) through conditional deletion (or suppression) of the gene for P450 reductase (Cpr), an enzyme required for the activity of all microsomal P450 enzymes. In this two-year pilot project study, we will determine the relative contributions of hepatic and extrahepatic (including the MG) P450 enzymes in the metabolic activation of DMBA, a mammary procarcinogen, and its intermediate metabolite 3,4-diol-DMBA, in newly developed mouse models with tissue-specific suppression of P450 activities, either in the liver alone (designated as liver-Cpr-null mouse), or in all tissues except the liver (designated as extrahepatic-Cpr-low mouse) (Aim 1). Furthermore, we will develop and characterize MG selective Cpr null models (designated as MG-Cpr-null mice), for direct determination of the role of MG P450 enzymes in the MG DNA-adduct formation induced by DMBA and 3,4-diol-DMBA (Aim 2). The knowledge gained from studies will improve the design of epidemiological studies that attempt to associate environmental chemical exposure, and/or genetic polymorphisms in P450 enzymes, with the incidence of breast cancer in humans. It may also help with designing mechanism-based approaches to the prevention of breast cancer. PUBLIC HEALTH RELEVANCE: This research will further our understanding of the mechanisms of chemical carcinogenesis in the mammary gland induced by environmental chemicals, and provide novel insights for mechanism-based approaches to the prevention of breast cancer.