Carcinogenic Cyclic Nitrosamine DNA Adducts

说明(由申请人提供)：本建议书集中于四种人类已确定接触的致癌亚硝胺的DNA加合物：A/-亚硝基吡咯烷(NPYR)、A/-亚硝基哌啶(NPIP)、A/-亚硝基琥珀酸(NSAR)和3-(甲基亚硝基)丙酸(MNPA)。所有这些致癌物质都存在于人类环境中，很可能是在人类体内形成的，并在人类尿液中检测到。DNA加合物在亚硝胺致癌过程中起着至关重要的作用。我们的总体假设是，这些亚硝胺在人体组织中产生DNA加合物，因此与人类癌症的原因有关。在这项计划的前五年中，我们已经广泛地在体外表征了由NPYR代谢激活的模型化合物a-乙酰氧基NPYR形成的DNA加合物，以及相关化合物巴豆醛、乙醛及其羟醛缩合产物的DNA加合物。我们还通过NPYR和NPIP的比较代谢实验，为这两种亚硝胺在大鼠体内具有强烈的致癌活性建立了合理的基础。在目前的更新提案中，我们的重点是表征和分析这些亚硝胺在体内形成的DNA加合物，包括在大鼠和人类中形成的DNA加合物。我们假设每个亚硝胺都会产生一组表型的DNA加合物。这些加合物可用作人类接触这些致癌物和代谢激活的生物标志物。我们的具体目标是：1.鉴定NPYR在大鼠肝脏中形成的DNA加合物；2.建立一种综合的LC-ESI-MS方法来定量NPYR在治疗大鼠组织和人类组织中的DNA加合物，并将这些研究扩展到NPIP；3.鉴定大鼠中NSAR和MNPA的DNA加合物，并建立LC-ESI-MS方法来定量它们在大鼠和人体组织中的含量。这项研究的结果将为我们了解这四种亚硝胺与人类接触的DNA加合物的形成和致癌作用提供关键数据。我们的方法是创新的，因为很少有实验室开发出可靠的方法来定量分析特定的DNA加合物，这些加合物反映了特定致癌物在人类体内的代谢激活。这项拟议的研究意义重大，因为已知人类通过饮食、烟草产品和内源性形成接触到这些致癌物质。

DESCRIPTION (provided by applicant): This proposal focuses on DNA adducts of four carcinogenic nitrosamines to which human exposure has been firmly established: A/-nitrosopyrrolidine (NPYR), A/-nitrosopiperidine (NPIP), A/-nitrososarcosine (NSAR), and 3-(methylnitrosamino)propionic acid (MNPA). All of these carcinogens occur in the human environment, are likely formed endogenously in humans, and have been detected in human urine. DNA adducts are absolutely critical in nitrosamine carcinogenesis. Our overall hypothesis is that these nitrosamines produce DNA adducts in human tissues and are therefore involved as causes of human cancer. In the previous five years of this program, we have extensively characterized, in vitro, DNA adducts formed from a-acetoxyNPYR, a model compound for the metabolic activation of NPYR, as well as DNA adducts of the related compounds crotonaldehyde, acetaldehyde, and their aldol condensation products. We have also established, through comparative metabolism experiments of NPYR and NPIP, a reasonable basis for the strongly contrasting carcinogenic activities of these two nitrosamines in rats. In the current renewal proposal, our focus is on the characterization and analysis of DNA adducts formed in vivo, in rats and humans, from these nitrosamines. We hypothesize that each nitrosamine will produce a phenotypic group of DNA adducts. These adducts can be used as biomarkers of human exposure to and metabolic activation of these carcinogens. Our specific aims are: 1. Identify the DNA adducts that are formed in rat liver upon treatment with NPYR; 2. Develop a comprehensive liquid chromatography-electrospray ionization-mass spectrometric (LC-ESI- MS) method to quantify DNA adducts of NPYR in tissues of treated rats and in human tissues, and extend these studies to NPIP; 3. Identify DNA adducts of NSAR and MNPA in rats and develop LC-ESI-MS methods to quantify them in rat and human tissues. The results of this research will provide critical data pertinent to our understanding of DNA adduct formation and carcinogenesis by these four nitrosamines with well-documented human exposure. Our approach is innovative because few laboratories have developed reliable methods for the quantitative analysis of specific DNA adducts which reflect the metabolic activation of particular carcinogens in humans. The proposed research is significant because of the known human exposure to these carcinogens through the diet, tobacco products, and endogenous formation.