32P-Postlabeling analysis of DNA adduction in mice by synthetic metabolites of the environmental carcinogen, 7H-dibenzo[c,g]carbazole: chromatographic evidence for 3-hydroxy-7H-dibenzo[c,g]carbazole being a proximate genotoxicant in liver but not skin

The DNA adduction by the environmental carcinogen 7H-dibenzo[c,g]carbazole(DBC) and chemically synthesized 2-OH, 3-OH, and 4-OH metabolitesof DBC was investigated in liver and skin of female CD-1 mice.After topical application to the skin of 37 µmol/kg ofDBC or the phenolic metabolites, DNA adducts were measured bya ³²P-post-labeling assay employing carrier-free [-³²P]ATP andATP-deficient conditions. In liver, DBC produced four majorand several minor chromatographically distinct adducts of asyet undetermined chemical structure. The adduct pattern elicitedby 3-OH-DBC was qualitatively similar to the DBC adduct pattern,while this was not the case for 2-OH-DBC and 4-OH-DBC. On thebasis of co-chromatography experiments under various conditions,the DBC and 3-OH-DBC adducts appeared identical, and the totallevel of adduction elicited by these compounds in liver wassubstantial. Similar results were observed when DBC or 3-OH-DBCwere administered i.p. As a major difference between the twocompounds, one 3-OH-DBC adduct (no. 3) was 4.4- and 7.0-foldlower than the corresponding DBC adduct after i.p. and topicaldosing, respectively. In skin, DBC produced two major adductfractions after topical application, one of which could be chromatographicallyresolved into three subcomponents. Prominent adducts producedin skin DNA by each of the three metabolites were differentfrom those elicited by DBC, and the level of adduction by themetabolites was significantly lower than that by DBC. Comparisonof the skin and liver DBC-DNA adduct patterns after topicalapplication of DBC showed that only one of the four major chromatographicallyresolved skin adducts corresponded to a major liver adduct (no.3), and that total adduction in liver was 13.5-fold higher thanin skin. These results suggested that (i) activation of DBCto DNA-binding compounds in liver occurs through at least twopathways with 3-OH-DBC being a proximate carcinogen involvedin the formation of most of the adducts; (ii) 3-OH-DBC and theother two phenolic metabolites investigated play a minor role,if any, in the formation of DBC-DNA adducts in skin; (iii) metabolicactivation of DBC to DNA-binding compounds in liver and skinappears to follow pathways that are different in terms of boththe chemical nature and the amount of the adducts formed; andiv) DBC and 3-OH-DBC exhibit a strong preference for liver versusskin DNA.