Susceptibility of rats to PhIP-induced mammary gland cancer varies with age and is associated with differential gene expression.

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2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a carcinogen found in the human diet, induced a 50% incidence of mammary cancer in Sprague-Dawley rats when administered orally during mammary gland development (43-55 days of age). However, mature virgin rats at 150-days of age were resistant to PhIP-induced mammary carcinogenesis. To explore the possible mechanisms for age-related differences in susceptibility, PhIP-DNA adduct levels, mutations, and gene expression were examined in glands from 43-day and 150-day old PhIP-treated rats given identical dosage regimens. Three hours after PhIP, PhIP-DNA adduct levels in the mammary gland (detected by the postlabeling assay) were not statistically different between 43-day and 150-day old rats. By 6 weeks after dosing, adduct levels had declined in rats of both ages (1.1 + 0.1and 2.7 + 0.8, mean + S.E., relative adduct labeling x10⁷, respectively). Mutations in the mammary gland were evaluated in the lacI gene of BigBlue transgenic rats at 6 weeks. Mutant frequency (mean + S.E. (x10^-6), n=4) in 43- and 150-day old rats were not statistically different [195 + 12 and 221 + 25, respectively]. Therefore initial DNA adduct levels, adduct removal, and mutation burden did not appear to account for age differences in susceptibility. To further examine possible mechanisms, the profile of gene expression was examined in 43- and 150-day old rats 6 weeks after PhIP treatment. Expression was examined on a NCI 10K cDNA microarray using analysis tools available from the NCI microarray database. Mammary gland RNA from PhIP-treated rats was arrayed against age-matched control. In glands from PhIP-treated 43-day old rats, 194 clones were differentially expressed. In contrast, only 29 clones were differentially expressed in 150-day old rats. In 150-day old rats, PhIP treatment was primarily associated with overexpression of differentiation-associated genes such as milk protein genes. In PhIP-treated 43-day old rats, many genes in distinct families were overexpressed such as ribosomal genes, proteosomes/protein degradation-related genes, heat shock protein genes, and genes in the Ras superfamily. The results indicate that the molecular responses to PhIP as evaluated by changes in gene expression were largely distinct between rats at susceptible and resistant ages and likely to be responsible for age differences in susceptibility. Elucidating the molecular pathways contributing to differences in susceptibility is expected to shed light on the molecular mechanisms of mammary gland carcinogenesis.