Evidence for the aldo-keto reductase pathway of polycyclic aromatic trans -dihydrodiol activation in human lung A549 cells

Abstract

8-oxo-dGuo ͉ DNA strand breaks ͉ tobacco carcinogens ͉ reactive oxygen species P olycyclic aromatic hydrocarbons (PAHs) are ubiquitous environmental pollutants, which are produced as a result of fossil-fuel combustion and are found in car exhaust and charbroiled and smoked foods (1, 2). They are also present as mixtures in tobacco smoke and are implicated in the causation of human lung cancer (3). To exert their carcinogenic effects, PAHs must be metabolically activated to DNA-damaging agents that will result in the signature mutations in lung cancer. These mutations are G-to-T transversions that either activate the K-ras protooncogene at the 12th and 61st codon (4) or inactivate the p53 tumor suppressor gene at hot spots in its DNA binding domain (5).Using benzo[a]pyrene (B[a]P) as a representative PAH, three pathways of activation have been proposed that lead to these mutations. The first pathway involves the formation of (ϩ)-anti-7␣,8␤-dihydroxy-9␣,10␤-epoxy-7,8,9,10-tetrahydroB[a]P {(Ϯ)- anti-B[a]PDE}.In this pathway there is sequential monoxygenation catalyzed by cytochrome P450 (P450) 1A1/1B1 and hydration to form 7␣,8␤-dihydroxy-7,8-dihydroxy-B[a]P, which undergoes a secondary monoxygenation to form (ϩ)-anti-B[a]PDE (6). This diol-epoxide forms stable (ϩ)-anti-trans-B[a]PDE-N 2 -2Ј-deoxyguanosine (dGuo) adducts, which via trans-lesional bypass DNA polymerases, yield G-to-T transversions (7).The second pathway involves metabolic activation by P450 peroxidases to yield radical cations (8), which can form depurinating adducts that lead to abasic sites. Apurinic/apyrimdinic (AP) sites, if not repaired, can give rise to G-to-T transversions (9). However, it is unlikely that radical cations are sufficiently long-lived to damage DNA in intact cells.The third pathway of PAH activation is the NAD(P ϩ )-dependent oxidation of PAH-trans-dihydrodiols to PAH oquinones catalyzed by dihydrodiol dehydrogenase members of the aldo-keto reductase (AKR) superfamily (10). AKRs divert PAH trans-dihydrodiols to form ketols that spontaneously rearrange to catechols (Scheme 1). The catechols undergo two one-electron oxidation events to produce the corresponding redox-active and electrophilic o-quinones. PAH o-quinones can form stable and depurinating DNA adducts in vitro (11,12), and these adducts may provide a route to G-to-T transversion mutations.In the presence of NAD(P)H, PAH o-quinones also undergo nonenzymatic reduction back to catechols. This event establishes futile redox cycles, which amplify the generation of reactive oxygen species (ROS) at the expense of NADPH and may lead to a prooxidant cellular state. Because a prooxidant state has been associated with tumor initiation and promotion (13), the AKR pathway of PAH activation is attractive in that it could explain how PAHs act as complete carcinogens. In addition, ROS may cause oxidative DNA damage such as 7,8-dihydro-8-oxo-2Ј-deoxyguanosine (8-oxo-dGuo) lesions, which can lead to G-to-T transversions (14). Amplification of ROS by catechol-oquinone interconversion has...

Detoxication of Structurally Diverse Polycyclic Aromatic Hydrocarbon (PAH) o-Quinones by Human Recombinant Catechol-O-methyltransferase (COMT) via O-Methylation of PAH Catechols

“…This metabolic activation led to the formation of ROS and 8-oxo-dGuo lesions in cellular DNA. Importantly, the amount of ROS and 8-oxo-dGuo formed in these experiments was increased in the presence of a COMT inhibitor (22). This suggested that COMT could intercept the catechol and prevent redox cycling.…”

Aryl Hydrocarbon Receptor Facilitates DNA Strand Breaks and 8-Oxo-2′-deoxyguanosine Formation by the Aldo-Keto Reductase Product Benzo[a]pyrene-7,8-dione

“…PAH o-quinones also cause oxidative DNA damage in vitro and in vivo (22)(23)(24)(25). Nanomolar concentrations of PAH o-quinones under redox cycling conditions (NADPH and Cu(II)) lead to significant 8-oxo-dGuo formation in bulk DNA, and the responsible oxidant was found to be singlet oxygen ( 1 O 2 ) (24,26).…”

“…Recently, using either a hOGG1-coupled comet assay or an immunoaffinity capture-stable isotope dilution liquid chromatography/electrospray ionization/multiple reaction monitoring/mass spectrometry (LC/ESI/MRM/MS) assay, it was shown that both the AKR substrate (B[a]P-7,8-trans-dihydrodiol) and the AKR product (B[a]P-7,8-dione) caused significant DNA strand breaks and 8-oxo-dGuo formation in human lung adenocarcinoma A549 cells (25 PAH o-quinones are ligands for the aryl hydrocarbon receptor (AhR) (30). These quinones can promote translocation of AhR to nucleus to induce P4501A1 expression.…”

Synthesis of 13C2-benzo[a]pyrene and its 7,8-dihydrodiol and 7,8-dione implicated as carcinogenic metabolites

“…An investigation of metabolic activation of BP in human lung A549 cells has provided evidence that the AKR-mediated pathway to generate the BP quinone and ROS is operative in these cells. 16 And human bronchoalveolar H358 cells were recently introduced as a model for study of PAH metabolism in normal human lung cells. 17,18 In this connection, development of a stable isotope dilution liquid chromatography tandem mass spectrometric method for analysis of the metabolites of BP and its nucleoside adducts was also described.…”