The dietary constituents of interest (e.g., methionine, choline, folate, alcohol, selenium, etc.) will be evaluated for their role in the early phases of carcinogenesis (liver and possibly mammary gland) and effects on global and site-specific DNA hypomethylation as well as post-translational modifications (acetylation, methylation) to histones. Animal studies, biochemical and molecular analyses will be performed at the FDA, National Center for Toxicological Research. The NCI will pay for the part of the laboratory costs associated with these analyses. Studies, done in collaboration with Dr. Igor Pogribny (FDA) and Dr. Sharon Ross (NCI), will consist of evaluating epigenetic disregulation (DNA hypomethylation, individual gene hypomethylation and hypermethylation, aberrant expression of related genes, histone and other chromatin modifications, and altered genomic imprinting) in the livers of methyl-deficient rats, determining whether the temporal epigenetic alteration in preneoplastic liver observed with a methyl deficient model also occurs with administration of other non-genotoxic and genotoxic carcinogens. Also, Dr. Pogribny will analyze gene expression and microRNA gene expression profiles in liver of animals fed methyl deficient diet or carcinogen-containing diet, as well as control fed animals will be determined Additionally, we will analyze the effect of corrections of the alterations in cellular epigenetic landscape by diet supplemented with methyl donors (choline, methionine, folic acid, and vitamin B12) on susceptibility to liver carcinogenesis. This will be undertaken to determine whether corrections of preexisting epigenetic changes in a target organ affect the susceptibility to cancer. We expect to provide the evidence that modification of an individual's epigenetic landscape can influence individual's susceptibility to carcinogenesis, and may be a decisive factor in cancer prevention strategies. Dr. Pogribny will also analyze human lymphocytes and rat tissue for histone modifications. Histone extracts will be isolated from stored tissue (i.e., liver, colon) of rats fed various bioactive food components, including diallyl disulfide (DADS), S-allylcysteine (SAC), S-allylmercaptocysteine (SAMC), selenium, selenium and respective controls. Human lymphocytes from individuals exposed to allyl isothiocyanates from cruciferous vegetables or a control diet will be isolated and shipped to Dr. Pogribny's laboratory for histone extraction and analysis of various histone marks. For both the human cells and animal tissue extracts, histone extracts will be examined for histone acetylation and methylation using antibodies specific for various histone marks in Western blot techniques. Histone methyltransferase expression may also be examined in tissues of animals fed various concentrations (deficient, sufficient and supplemental) of selenium and folate. . Funds will be used for laboratory supplies and to partially offset the salary of a post-doctoral research associate to carry out the analyses.
