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Eight isomeric dGuo and eight dAdo adducts resulting from cis and trans opening of each of the four optically active diol epoxides (DEs) derived from benzo[ a ]pyrene (B a P) and benzo[ c ]phenanthrene (B c Ph) were placed in each of two 16-mer DNA sequences to give 32 modified oligonucleotides, which were ligated into M13mp7L2 and allowed to replicate in SOS-induced Escherichia coli . The effects of parent hydrocarbon, adduct stereochemistry, and sequence context on mutagenic response are highly interdependent. B a P DE adducts are generally more mutagenic than the corresponding B c Ph adducts. The mutational frequency is generally larger for cis - relative to trans -opened DE adducts of both dGuo and dAdo. In a ∼TA*G∼ context, B c Ph DE dAdo adducts (A*) with R configuration at the site of attachment to the adenine base produced very few substitution mutations when compared with adducts having S configuration. This configurational effect is not observed for B a P DE dAdo adducts, nor for B a P or B c Ph...

The effects of two DNA sequence contexts on the mutagenic response to dG and dA adducts derived from the optically active 7,8-diol 9,10-epoxides of benzo[a]pyrene (BaP DEs) and 3,4-diol 1,2-epoxides of benzo[c]phenanthrene (BcPh DEs) were examined. On replication of the adduct-containing, single-stranded vector M13mp 7L2 in E. coli SMH77, frequencies of substitution mutations ranging from 0.05% to 68% were observed. In general, the highest mutational frequencies were observed for BaP DE-dA adducts at the central position in a ∼TAG∼ sequence. The BaP DE adducts showed no relationship between stereochemistry and mutagenic response. In contrast, mutagenicity of the BcPh DE-dA adducts in the ∼TAG∼ sequence depended strongly on the configuration at C-1, the site of attachment of the hydrocarbon to the purine. In this sequence, BcPh DE-dA adducts with 1R configuration were 16-600 times less mutagenic than those with 1S configuration.

The synthesis, structures, and mutagenic properties of benzo[a]pyrene-deoxyadenosine adducts in a sequence context related to the p53 gene are described.

The various methods that have been used in site-specific mutagenesis studies with DNA adducts from hydrocarbon diol epoxides are briefly reviewed. The single-stranded M13 approach pioneered by Lawrence and his colleagues offers the advantage that DNA repair processes should not reduce mutation frequencies since they should lead to vector destruction. The application of this approach to an evaluation of the mutagenic effects arising from a benzo[a]pyrene-deoxyadenosine adduct is shown. Substantial numbers of A→T and A→G base substitution mutations were obtained along with a few A→C mutations.

Dihydrodiol epoxides are carcinogenic metabolites of polycyclic aromatic hydrocarbons. The effects of two such compounds, benzo[g]chrysene 11,12-dihydrodiol 13,14-epoxide and 5-methylchrysene 1,2-dihydrodiol 3,4-epoxide, on the progress of the human mammary carcinoma cell, MCF-7, through the cell cycle was investigated. Unlike other DNA damaging agents, such as actinomycin D, these carcinogens did not arrest MCF-7 cells in the G1 phase of the cell cycle and the cells began DNA replication despite the damage in the DNA template. The dihydrodiol epoxides, like actinomycin D, induced the p53 protein that normally mediates G1 arrest through transcriptional activation of p21wafl/cipl, in these cells. However, increased cellular p21wafl/cipl levels were not found in the hydrocarbon-treated cells. Thus, these carcinogens did not trigger the major ‘guardian of the genome’ cellular defense mechanism mediated by p53. This stealth characteristic may be an important contributor to the carcinogenic action of ...

Anti benzo[s]picene 9,10-dihydrodiol 11,12-epoxide was reacted with calf thymus DNA and with its constituent deoxyribonucleotides. The origin of DNA adducts was established by comparison of their HPLC retention times and UV spectra with those of the nucleotide-derived adduct-markers. The low binding of the dihydrodiol epoxide to DNA (2%) and to deoxyguanylic and deoxyadenylic acids precluded an NMR study, and assignment of cis or trans epoxide ring opening was made by analogy to other anti dihydrodiol epoxide adducts. The adduct ratio for dGuo/dAdo in DNA was 24/76 and the ratios of dGuo cis/dGuo trans and dAdo cis/dAdo trans were 40/60 and 6/94, respectively.

DNA adducts derived from reaction of the racemic bay region syn 7-methylbenz[a]anthracene 3,4-dihydrodiol 1,2-epoxide with calf thymus DNA in vitro were tentatively identified. Eight markers (four deoxyguanosine and four deoxyadenosine adducts) were obtained from separate reactions of the racemic syn dihydrodiol epoxide with deoxyguanylic and deoxyadenylic acids. The nucleoside of origin of individual DNA products was established by comparing HPLC retention times and UV spectra of DNA adducts with those of the adduct markers. All DNA adducts eluted with retention times and had UV spectra that corresponded to the purine nucleoside adducts. Circular dichroism spectra of the marker adducts allowed assignment of S and R configuration at C1, the site of attachment of the hydrocarbon residue to the nucleoside. The CD spectra were comprised of four pairs of spectra that were mirror images of one another. Each pair consisted of the two cis or two trans products resulting from epoxide ring opening of each...

Reactive metabolites from non-planar polycyclic aromatic hydrocarbon carcinogens react extensively with both deoxyadenosine and deoxyguanosine in DNA whereas those from planar molecules react predominantly only with deoxyguanosine. In vitro studies with single adducts in oligonucleotides showed that both types of adduct blocked primer extension and that the limited amount of nucleotide addition opposite the adduct varied with the polymerase, the sequence context of the adduct and the chemical structure of the adduct. When these same single adduct containing-oligonucleotides were introduced into a single-stranded vector that was allowed to replicate in Escherichia coli, the major events observed were blockage of replication, insertion of the correct nucleotide (i.e. T opposite an A adduct and C opposite a G adduct), and insertion of A opposite the adduct. In mouse skin, benzo[c]phenanthrene 4S,3R-dihydrodiol 2S,1R-epoxide initiated substantially more tumors per DNA adduct formed than did the other...

A brief review of the literature on syn dihydrodiol epoxide DNA adducts is presented. NMR studies indicate that bay region substituents influence the conformation of the partially saturated ring in such adducts. However, only substitutions that lead to distortion from planarity are associated with reactivity towards deoxyadenosine residues in DNA and with greater tumorigenicity. This is similar to the situation that obtains for anti dihydrodiol epoxides.