Discovery of Genotoxic Biomarkers in Urine for Cancer

描述(由申请人提供)： 该项目旨在开发一种新的方法学程序，有可能提高癌症流行病学研究的质量。这项新程序涉及使用质谱仪(MS)来分析尿液中固有的或源自尿液的DNA加合物。“固有的DNA加合物”将被分析用于两个目的：(1)作为人类接触致癌物的潜在生物标记物，以及(2)作为癌症存在和组织位置的潜在生物标记物。后一种假设是通过综合其他人的以下观察而提出的：(1)每个组织都有独特的亲脂性、明显的内源性DNA加合物模式(如32P标记后/高效液相检测到的)；(2)一些组织DNA溢出到血液中(在癌症中越来越多)，以及(3)血液中的一些游离DNA最终进入尿液。按照已发表的方法，从吸烟者的尿液中提取具有遗传毒性的化学物质，并在S9代谢激活系统与小牛胸腺DNA的存在下进行反应，将获得“衍生DNA加合物”。之前的研究人员认为吸烟者尿液中的遗传毒性化学物质与存在的诱变剂相同。在这个项目中，对三种类型的DNA加合物(固有的、外源性的、固有的-内源性的和衍生的)的检测将通过我们最近开发的一种方法来完成，在这种方法中，在分离后，用咪唑取代的质量标签标记加合物，然后使用基质辅助激光解吸电离飞行时间质谱仪(MALDI-TOFMS)。我们的新技术非常灵敏，可以同时测量广泛的已知和未知DNA加合物的准确质量，不同于任何以前的DNA加合物分析方法。除了来自对照组和吸烟者的尿样外，还将检测肾癌和膀胱癌患者的尿样。尿液是流行病学研究的一个有吸引力的样本。该项目启动了一名分析化学家和一名分子生物学家之间的合作。

DESCRIPTION (provided by applicant): This project seeks to develop a new methological procedure that has the potential for improving the quality of cancer epidemiological research. The new procedure involves the use of mass spectrometry (MS) to analyze DNA adducts inherent to or derived from urine. The "inherent DNA adducts" will be analyzed for two purposes: (1) as potential biomarkers for human exposure to carcinogens, and (2) as potential biomarkers for the presence and tissue location of cancer. This latter hypothesis is suggested by bringing together the following observations by others: (1) each tissue has a unique pattern of lipophilic, apparently endogenous DNA adducts (as detected by 32P-post-labeling/HPLC); (2) some tissue DNA is spilled into the blood (increasingly so in cancer), and (3) some of the free DNA in the blood ends up in the urine. The "derived DNA adducts" will be obtained by following a published method in which genotoxic chemicals are extracted from smoker's urine and reacted in the presence of an S9 metabolic activation system with calf thymus DNA. Previous investigators have considered the genotoxic chemicals in smoker's urine to be the same as the mutagens, which are present. Detection in this project of the three types of DNA adducts (inherentexogenous, inherent-endogenous and derived) will be accomplished by using a method that we have recently developed in which the adducts, after isolation, are labeled with an imidazole substituted mass tag followed by use of matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS). Our new technique is very sensitive and measures the exact masses of a broad range of known and unknown DNA adducts simultaneously, unlike any prior analytical method for DNA adducts. Along with urine samples from control subjects and smokers, urine from patients with renal and bladder cancer will be tested. Urine is an attractive sample for epidemiological studies. The project initiates a collaboration between an analytical chemist and a molecular biologist.