title: Evidence for the aldo-keto reductase pathway of polycyclic aromatic trans-dihydrodiol activation in human lung A549 cells.
abstract: Polycyclic aromatic hydrocarbons (PAHs) are tobacco carcinogens implicated in the causation of human lung cancer. Metabolic activation is a key prerequisite for PAHs to cause their deleterious effects. Using human lung adenocarcinoma (A549) cells, we provide evidence for the metabolic activation of (+/-)-trans-7,8dihydroxy-7,8-dihydrobenzo[a]pyrene (B[a]P-7,8-trans-dihydrodiol) by aldo-keto reductases (AKRs) to yield benzo[a]pyrene-7,8-dione (B[a]P-7,8-dione), a redox-active o-quinone. We show that B[a]P-7,8-trans-dihydrodiol (AKR substrate) and B[a]P-7,8-dione (AKR product) lead to the production of intracellular reactive oxygen species (ROS) (measured as an increase in dichlorofluorescin diacetate fluores-cence) and that similar changes were not observed with the regioisomer (+/-)-trans-4,5-dihydroxy-4,5-dihydrobenzo[a]pyrene or the diol-epoxide, (+/-)-anti-7,8-dihydroxy-9alpha,10beta-epoxy-7,8,9,10-tetrahydro-B[a]P. B[a]P-7,8-trans-dihydrodiol and B[a]P-7,8-dione also caused a decrease in glutathione levels and an increase in NADP(+)/NADPH ratios, with a concomitant increase in single-strand breaks (as measured by the comet assay) and 7,8-dihydro-8-oxo-2'-deoxyguanosine (8-oxo-dGuo). The specificity of the comet assay was validated by coupling it to human 8-oxo-guanine glycosylase (hOGG1), which excises 8-oxo-Gua to yield single-strand breaks. The levels of 8-oxo-dGuo observed were confirmed by an immunoaffinity purification stable isotope dilution ([(15)N(5)]-8-oxo-dGuo) liquid chromatography-electrospray ionization/multiple reaction monitoring/mass spectrometry (LC-ESI/MRM/MS) assay. B[a]P-7,8-trans-dihydrodiol produced DNA strand breaks in the hOGG1-coupled comet assay as well as 8-oxo-dGuo (as measured by LC-ESI/MRM/MS) and was enhanced by a catechol O-methyl transferase (COMT) inhibitor, suggesting that COMT protects against o-quinone-mediated redox cycling. We conclude that activation of PAH-trans-dihydrodiols by AKRs in lung cells leads to ROS-mediated genotoxicity and contributes to lung carcinogenesis.
authors.affiliation: Center of Excellence in Environmental Toxicology, University of Pennsylvania School of Medicine, Philadelphia, PA 19104-6084, USA.
authors.forename: Jong-Heum
authors.initials: JH
authors.lastname: Park
authors.forename: Dipti
authors.initials: D
authors.lastname: Mangal
authors.forename: Kirk A
authors.initials: KA
authors.lastname: Tacka
authors.forename: Amy M
authors.initials: AM
authors.lastname: Quinn
authors.forename: Ronald G
authors.initials: RG
authors.lastname: Harvey
authors.forename: Ian A
authors.initials: IA
authors.lastname: Blair
authors.forename: Trevor M
authors.initials: TM
authors.lastname: Penning
mesh_terms: D000080242:8-Hydroxy-2'-Deoxyguanosine; D000429:Alcohol Oxidoreductases / Q000378:metabolism*; D000449:Aldehyde Reductase; D000074408:Aldo-Keto Reductases; D001580:Benzopyrenes / Q000494:pharmacology; D001711:Biotransformation / Q000187:drug effects; D065098:Catechol O-Methyltransferase Inhibitors; D045744:Cell Line, Tumor; D020552:Comet Assay; D053903:DNA Breaks, Double-Stranded / Q000187:drug effects; D045647:DNA Glycosylases / Q000378:metabolism; D003849:Deoxyguanosine / Q000031:analogs & derivatives / Q000378:metabolism; D004101:Dihydroxydihydrobenzopyrenes / Q000378:metabolism* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005452:Fluoresceins / Q000378:metabolism; D005453:Fluorescence; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D008168:Lung / Q000201:enzymology* / Q000473:pathology; D010084:Oxidation-Reduction / Q000187:drug effects; D017382:Reactive Oxygen Species / Q000378:metabolism; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization
chemical_list: D001580:Benzopyrenes; D065098:Catechol O-Methyltransferase Inhibitors; D004101:Dihydroxydihydrobenzopyrenes; D004791:Enzyme Inhibitors; D005452:Fluoresceins; D007527:Isoenzymes; D017382:Reactive Oxygen Species; C017228:benzo(a)pyrene 7,8-dihydrodiol; C029569:diacetyldichlorofluorescein; C076588:benzo(a)pyrene-7,8-dione; D000080242:8-Hydroxy-2'-Deoxyguanosine; D000429:Alcohol Oxidoreductases; D000074408:Aldo-Keto Reductases; D000449:Aldehyde Reductase; D045647:DNA Glycosylases; C479058:oxoguanine glycosylase 1, human; D003849:Deoxyguanosine