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。新的程序涉及使用质谱法（MS）来分析尿液中固有或衍生的DNA加合物。将出于两个目的分析“固有DNA加合物”：（1）作为人类暴露于致癌物的潜在生物标志物，以及（2）作为癌症存在和组织位置的潜在生物标志物。后一种假设是通过将其他人的以下观察结果结合在一起而提出的：（1）每个组织都具有独特的亲脂性模式，显然是内源性DNA加合物（如通过32 P-后标记/HPLC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。“衍生DNA加合物”将通过以下已发表的方法获得，其中从吸烟者尿液中提取遗传毒性化学物质，并在存在S9代谢活化系统的情况下与小牛胸腺DNA反应。以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。在这个项目中的三种类型的DNA加合物（inherentexogenous，inherent-endogenous和衍生）的检测将通过使用一种方法，我们最近开发的加合物，分离后，用咪唑取代的质量标签，然后使用基质辅助激光解吸电离飞行时间质谱（MALDI-TOFMS）标记。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加合物的精确质量，这与任何先前的DNA加合物分析方法不同。沿着对照受试者和吸烟者的尿液样本，将检测肾癌和膀胱癌患者的尿液。尿液是流行病学研究的一个有吸引力的样本。该项目启动了分析化学家和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.