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. ? ? ?
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