The goals of this program are to elucidate the mechanism of carcinogenicity and organotropism of methylazoxymethanol (MAM), a powerful colon-specific carcinogen in most, though not all, rodent species and strains. MAM is the penultimate metabolite of the widely used model colon carcinogen 1,2-dimethylhydrazine (DMH), and the active moiety of the naturally occurring intestinal and liver carcinogen, cycasin. In the previous phase of this continuing program, it was established that MAM is metabolically activated by purified cytochrome P-450j, an isozyme inducible in the liver by administration of ethanol, acetone, isopropanol and other agents. In addition, it was determined that this isozyme, known to metabolically activate N-nitrosodimethylamine and to hydroxylate aniline, can also metabolize azoxymethane (AOM) to MAM, an important step in the metabolic activation of both DMH as well as of AOM. While this isozyme appears to be present in the rat colon mucosa as well as the liver, indications are that the enzymes metabolizing MAM in rat liver and rat colon probably are not identical. The present program seeks to identify the enzymes responsible for the organ-selective metabolic activation of MAM in the rat colon. Evidence exists for the ability of the rat colon mucosa to metabolize MAM via three separate enzymatic pathways: cytosolic alcohol dehydrogenase (ADH), the microsomal mixed function oxidase (MFO) and prostaglandin synthase (PES). These systems will be examined for their ability to metabolically activate or detoxify (1,2-14C)MAM using a unique """"""""hybrid"""""""" HPLC system. The kinetic parameters of the metabolism of MAM by ADH purified from the rat colon will be compared to those obtained from the PES- and MFO-mediated reactions, and inhibition constants for select inhibitors of these reactions will also be obtained. Various inducers of the colonic MFO, including beta-naphthoflavone and gastrointestinal peptide hormones will be used. To obtain further evidence for the involvement or noninvolvement of the MFO in colonic and liver MAM metabolism, MAM metabolism and MAM-induced DNA methylation will be examined in rats and deermice depleted of cytochrome P-450 by cobalt protoporphyrin IX or CoC12. The results of these studies are expected to establish the mechanism of organotropism of MAM for the rodent colon. Further, they are expected to provide important information which will be of use in the more efficient utilization of the DMH, AOM or MAM animal colon cancer models to reflect the human condition.
