DETERMINANTS OF DIETARY RISK OF HETEROCYCLIC AMINES

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 在过去的15年里，我们一直在研究饮食对癌症的作用。富含蛋白质的食物在烹饪、热处理和热解过程中会形成一组结构上相关的杂环芳胺，这些胺在许多检测系统中都被发现是有效的诱变剂。这些化合物在雄性和雌性小鼠和大鼠的多个器官部位都会产生肿瘤，给予其中一种杂环胺的非人类灵长类动物100%在非常短的潜伏期后患上肝癌。鉴于这些非常令人信服的数据，重要的是确定这些饮食诱变剂/致癌物对人类癌症发病率的贡献程度，并制定战略以限制其影响。 美国国立卫生研究院的这个项目试图通过以下方式实现这些目标： -确定和量化人类饮食中这些杂环胺的摄入量； -通过分析长期接触啮齿动物后的DNA结合、细胞遗传损伤和突变效应，了解这些化合物的慢性毒理作用； -通过确定重要的代谢途径(啮齿动物、非人类灵长类动物、人类)的特征，了解动物研究与人类的机制相关性，以及了解这些杂环胺引起肿瘤形成的组织特异性的性质； -通过评估低剂量摄入这些有效诱变剂的剂量学，了解高剂量动物研究对人类风险评估的剂量相关性； -确定与突变相关的致癌物和DNA加合物的结构特征； -利用CHO细胞和在DNA修复和代谢激活方面具有遗传差异的啮齿动物品系，预测个体差异在修复和代谢方面的重要性； -确定和确认可能有助于确定人类风险或易感性的生物标志物； -评估该项目产生的数据和文献中的数据，以产生定量的癌症风险评估。 很明显，AMS资源对NIH资助的这个项目产生了重大影响，因为这项工作的各个方面只能利用加速器质谱学来完成。特别是，AMS允许我们在低剂量下进行新陈代谢和DNA损伤的研究，这些研究影响到我们计划项目拨款的每个子项目。 目前的研究摘要：细胞色素P450介导的羟化和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(HAS)在内的许多异类化合物生物转化的主要代谢途径。研究表明，在人类中，HA PhIP的生物激活高度依赖于细胞色素P4501A2介导的N-羟基化反应。随后的N-葡萄糖醛酸化会导致较不活跃的N-羟基-PhIP-N-葡萄糖醛酸化物和N-羟基-PhIP-N3-葡萄糖醛酸化物的形成，它们可以通过尿液或胆汁排泄，或者可以运输到肝外组织，在那里可以进行进一步的新陈代谢。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的多态表达导致了许多底物的差异代谢。UGT表达的这种个体间差异可能会潜在地改变某些个体中促癌物质的生物活性，如PhIP。因此，了解PhIP和N-羟基-PhIP在人体内的N-葡萄糖醛酸化反应尤为重要，因为如果N-羟基-PhIP不能通过葡萄糖醛酸化反应偶联，则可能会通过酯化反应进一步激活。这些反应将产生高活性的化合物，可以与DNA结合，可能导致突变。由于PhIP生物激活在动物模型中的多个组织中负责DNA加合物的形成，通过确定在啮齿动物模型中葡萄糖醛酸化在PhIP组织特异性生物激活/解毒中的作用，可以更好地理解PhIP代谢如何在整个动物中促进DNA加合物的形成。通过使用加速器质谱仪，我们可以分析肿瘤靶组织中DNA加合物的形成和膳食中PhIP的相关剂量。我们假设，与UGT活性增强的动物相比，葡萄糖醛酸化能力减弱的动物和/或组织将更容易受到PhIP诱导的肿瘤靶组织中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.