DETERMINANTS OF DIETARY RISK OF HETEROCYCLIC AMINES

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 在过去的 15 年里，我们一直在研究饮食对癌症的作用。 富含蛋白质的食物的烹饪、热处理和热解会导致形成一组结构相关的杂环芳香胺，这些胺已在许多分析系统中被发现是有效的诱变剂。 这些相同的化合物在雄性和雌性小鼠和大鼠的多个器官部位产生肿瘤，并且100%的非人类灵长类动物在给予其中一种杂环胺后在很短的潜伏期后发展成肝癌。鉴于这些非常令人信服的数据，确定这些饮食诱变剂/致癌物对人类癌症发病率的影响程度并制定限制其影响的策略非常重要。 该 NIH 计划项目试图通过以下方式实现这些目标： - 识别和量化人类饮食中这些杂环胺的摄入量； - 通过分析啮齿动物长期长期喂养接触后的 DNA 结合、细胞遗传学损伤和突变效应，了解这些化合物的慢性毒理学； - 通过表征重要的代谢途径（啮齿动物、非人灵长类动物、人类）来了解动物研究与人类的机制相关性，另外的目标是了解这些杂环胺诱导的肿瘤形成中组织特异性的本质； - 通过评估低剂量摄入这些强效诱变剂的剂量测定，了解高剂量动物研究的剂量相关性，以进行人类风险评估； - 表征与突变相关的致癌物和DNA加合物的结构特征； - 使用在DNA修复和代谢激活方面具有遗传差异的CHO细胞和啮齿动物品系来预测修复和代谢方面个体差异的重要性； - 识别和验证可能对人类风险或易感性测定有用的生物标志物； - 评估该项目生成的数据和文献中的数据，以进行定量癌症风险评估。 显然，AMS 资源对 NIH 资助的计划项目产生了重大影响，因为这项工作的某些方面只能利用加速器质谱法来完成。 特别是，AMS 使我们能够以低剂量进行新陈代谢和 DNA 损伤的研究，这会影响我们计划项目拨款的每个子项目。 当前研究摘要：细胞色素 P450 介导的羟基化和 UDP-葡萄糖醛酸基转移酶 (UGT) 催化的葡萄糖醛酸化是包括杂环胺 (HA) 在内的许多外源物质生物转化的主要代谢途径。研究表明，在人类中，HA PhIP 的生物活化高度依赖于细胞色素 P4501A2 介导的 N-羟基化为相应的 N-羟基-PhIP。随后的N-葡萄糖醛酸化导致形成反应性较低的N-羟基-PhIP-N2-葡萄糖苷酸和N-羟基-PhIP-N3-葡萄糖苷酸，它们可以通过尿液或胆汁排出，或者可以转运到肝外组织，在那里可以发生进一步的代谢。最近的研究表明，在人类中，N-羟基-PhIP 的葡萄糖醛酸化是 PhIP 生物转化的主要途径，而 UGT1A1 同工酶是 N-羟基-PhIP 葡萄糖醛酸化的主要贡献者。此外，几种UGT的多态性表达导致许多底物的代谢差异。 UGT 表达的这种个体间差异可能会改变某些个体中 PhIP 等前致癌物的生物活性。因此，了解人类中 PhIP 和 N-羟基-PhIP 的 N-葡萄糖醛酸化尤为重要，因为未能通过葡萄糖醛酸化结合 N-羟基-PhIP 可能会导致酯化反应进一步激活。这些反应会产生能够结合 DNA 的高反应性化合物，从而可能导致突变。由于 PhIP 生物活化已被证明与动物模型中多个组织中 DNA 加合物的形成有关，通过确定葡萄糖醛酸化对啮齿动物模型中 PhIP 组织特异性生物活化/解毒的作用，可以更好地了解 PhIP 代谢如何促进整个动物中 DNA 加合物的形成。通过使用加速器质谱，我们可以分析肿瘤靶组织中 DNA 加合物的形成以及 PhIP 的饮食相关剂量。 我们假设葡萄糖醛酸化能力减弱的动物和/或组织比 UGT 活性增加的动物更容易受到肿瘤靶组织中 PhIP 诱导的 DNA 加合物的影响。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Over the last 15 years we have been studying the role of diet on cancer. The cooking, heat processing, and pyrolysis of protein-rich foods result in the formation of a group of structurally related heterocyclic aromatic amines that have been found to be potent mutagens in a number of assay systems. These same compounds produce tumors at multiple organ sites in both male and female mice and rats and 100 percent of non-human primates given one of these heterocyclic amines developed hepatocarcinomas after a very short latency. Given these very compelling data, it is important to determine the extent to which these dietary mutagens/carcinogens contribute to the human cancer incidence and to devise strategies to limit their impact. This NIH program project attempts to achieve these goals by: - Identifying and quantifying the human intake of these heterocyclic amines in the diet; - Understanding the chronic toxicology of these compounds by analysis of DNA binding, cytogenetic damage and mutational effects following chronic long-term feeding exposure of rodents; - Understanding the mechanistic relevance of animal studies for humans by characterizing important metabolic pathways (rodents, non-human primates, humans) with the additional goal of understanding the nature of the tissue specificity in tumor formation induced by these heterocyclic amines; - Understanding the dose-relevance of high dose animals studies for human risk assessment by assessing the dosimetry from ingestion of these potent mutagens at low doses; - Characterizing the structural features of carcinogens and DNA adducts that are correlated with mutation; - Predicting the importance of individual differences in repair and metabolism using CHO cells and rodent strains having genetic differences in DNA repair and metabolic activation; - Identifying and validating biomarkers that may be useful for human risk or susceptibility determinations and; - Evaluating data generated from this project and data from the literature to produce quantitative cancer-risk assessment. It is clear that the AMS resource is having a major impact on this NIH funded Program Project, as aspects of this work can only be accomplished utilizing accelerator mass spectrometry. In particular, AMS allows us to conduct studies of metabolism and DNA damage at low dose that impacts every sub-project of our program project grant. Current study abstract: Cytochrome P450-mediated hydroxylation and UDP-glucuronosyltransferase (UGT)-catalyzed glucuronidation are major metabolic pathways in the biotransformation of many xenobiotics including heterocyclic amines (HAs). Studies have shown that, in humans, the bioactivation of the HA PhIP is highly dependent upon cytochrome P4501A2-mediated N-hydroxylation to the corresponding N-hydroxy-PhIP. Subsequent N-glucuronidation results in the formation of the less reactive N-hydroxy-PhIP-N2-glucuronide and N-hydroxy-PhIP-N3-glucuronide, which can be excreted through urine or bile, or can be transported to extrahepatic tissue where further metabolism can occur. Recent studies have shown that, in humans, glucuronidation of N-hydroxy-PhIP is a major pathway in the biotransformation of PhIP and that the UGT1A1 isozyme is a major contributor to N-hydroxy-PhIP glucuronidation. In addition, polymorphic expression of several UGTs has led to differential metabolism of many substrates. This inter-individual variation in UGT expression could potentially alter the bioactivation of pro-carcinogens such as PhIP in certain individuals. Therefore, understanding the N-glucuronidation of PhIP and N-hydroxy-PhIP in humans is especially important because the failure to conjugate N-hydroxy-PhIP by glucuronidation could result in further activation by esterifying reactions. These reactions would result in highly reactive compounds that can bind DNA, potentially causing mutations. Since PhIP bioactivation has been shown to be responsible for the formation of DNA adducts in multiple tissues in animal models, by determining the role glucuronidation has on tissue-specific bioactivation/detoxification of PhIP in rodent models, a better understanding of how PhIP metabolism contributes to DNA adduct formation in whole animals can be established. By using accelerator mass spectrometry we can analyze DNA adduct formation in tumor target tissues and dietary relevant doses of PhIP. We hypothesize that animals and/or tissues with diminished glucuronidation capacity will be more susceptible to PhIP induced DNA adducts in tumor target tissues than animals with increased UGT activity.