DNA Adducts of Aromatic Amines: Analysis by HPLC-MS

描述(由申请人提供)：基因毒素与DNA的共价结合形成DNA加合物被许多人认为是化学诱导致癌的第一步。细胞中DNA加合物的测量为暴露提供了直接证据，也是评估人类健康风险的一种手段。然而，DNA加合物的形成与人类癌症发展之间的关系只对几种致癌物质建立起来。这在一定程度上是因为受影响的组织和器官中加合物的水平较低，可用的DNA数量有限，以及与分析相关的困难。为了解决这些分析问题，我们一直在探索使用毛细管分离方法(液相色谱(LC)和毛细管电泳法(CE))与质谱学(MS)相结合的方法来明确检测和表征DNA加合物。利用动物模型，我们的研究证明了毛细管LC-MS/MS检测和定量DNA加合物的能力，其水平类似于人类接触的水平。我们的工作重点是芳香胺，如2-氨基-3-甲基咪唑[4，5-f]喹啉(IQ)，4-氨基联苯(ABP)，2-氨基-1-甲基-6-苯并咪唑[4，5-b]吡啶(PhIP)和2-氨基-3，8-二甲基咪唑[4，5-f]喹恶啉(MeIQx)，存在于香烟烟雾中，在烤鱼或牛肉时形成。肺、结肠、乳房、胰腺和其他组织含有能够激活这些致癌物的酶，这些酶能够将这些致癌物转化为能够与DNA碱基结合的活性形式。我们建议利用我们成功的动物研究来检查肺和其他人类组织，重点是比较吸烟者和不吸烟者之间的DNA加合物含量。我们将进一步研究致癌物的序列识别模式，方法是将它们与模拟已知显示与肺癌相关的特定突变的p53基因序列的合成寡核苷酸反应。我们希望这个项目中产生的数据将有助于确定吸烟和肺癌之间的关系，以及DNA加合物作为遗传毒性癌症指标的价值。

DESCRIPTION (provided by applicant): The covalent binding of genotoxins to DNA to form DNA adducts is considered by many to be the first step in chemically induced carcinogenesis. Measurement of DNA adducts in cells provides direct evidence for exposure and a means for assessing human health risk. However, the relationship between DNA adduct formation and cancer development in humans has only been established for a few carcinogens. This is, in part, because of the low levels at which adducts occur in affected tissues and organs, the limited amounts of DNA available, and the difficulties associated with the analysis. In order to address these analytical problems, we have been exploring the use of capillary separation methods (liquid chromatography (LC) and capillary electrophoresis (CE)) coupled to mass spectrometry (MS) for the unequivocal detection and characterization of DNA adducts. Using animal models, our research has demonstrated the capability of capillary LC-MS/MS for the detection and quantification of DNA adducts at levels similar to those encountered in human exposure. Our work has focused on aromatic amines such as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 4-aminobiphenyl (ABP), 2-amino-1- methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), found in cigarette smoke and formed during the grilling of fish or beef. Lung, colon, breast, pancreas and other tissues contain enzymes capable of activating these carcinogens into reactive forms that can bind to DNA bases. We propose to capitalize on our successful animal studies to examine lung and other human tissues with emphasis on the comparison of DNA adduct content between smokers and non-smokers. We will further examine the sequence recognition patterns of carcinogens by reacting them with synthetic oligonucleotides that mimic p53 gene sequences known to display specific mutations associated with lung cancer. We expect that the data generated in this program will help define the relationship between cigarette smoke and lung cancer and the value of DNA adducts as indicators for genotoxic cancer.