ABSTRACT
Utilizing molecular dynamics and free energy perturbation, we examine the relative binding affinity of several covalent polycyclic aromatic hydrocarbon - DNA (PAH-DNA) adducts at the central adenine of NRAS codon-61, a mutational hotspot implicated in cancer risk. Several PAHs classified by the International Agency for Research on Cancer as probable, possible, or unclassifiable as to carcinogenicity are found to have greater binding affinity than the known carcinogen, benzo[a]pyrene (B[a]P). van der Waals interactions between the intercalated PAH and neighboring nucleobases, and minimal disruption of the DNA duplex drive increases in binding affinity. PAH-DNA adducts may be repaired by global genomic nucleotide excision repair (GG-NER), hence we also compute relative free energies of complexation of PAH-DNA adducts with RAD4-RAD23 (the yeast ortholog of human XPC-RAD23) which constitutes the recognition step in GG-NER. PAH-DNA adducts exhibiting the greatest DNA binding affinity also exhibit the least RAD4-RAD23 complexation affinity and are thus predicted to resist the GG-NER machinery, contributing to their genotoxic potential. In particular, the fjord region PAHs dibenzo[a,l]pyrene, benzo[g]chrysene, and benzo[c]phenanthrene are found to have greater binding affinity while having weaker RAD4-RAD23 complexation affinity than their respective bay region analogs B[a]P, chrysene, and phenanthrene. We also find that the bay region PAHs dibenzo[a,j]anthracene, dibenzo[a,c]anthracene, and dibenzo[a,h]anthracene exhibit greater binding affinity and weaker RAD4-RAD23 complexation affinity than B[a]P. Thus, the study of PAH genotoxicity likely needs to be substantially broadened, with implications for public policy and the health sciences. This approach can be broadly applied to assess factors contributing to the genotoxicity of other unclassified compounds.
Subject(s)
DNA Adducts , Polycyclic Aromatic Hydrocarbons , Polycyclic Aromatic Hydrocarbons/toxicity , Polycyclic Aromatic Hydrocarbons/chemistry , Polycyclic Aromatic Hydrocarbons/metabolism , DNA Adducts/chemistry , DNA Adducts/metabolism , DNA Adducts/genetics , Humans , DNA Repair , Mutagens/toxicity , Mutagens/chemistry , Molecular Dynamics Simulation , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/chemistry , Thermodynamics , Benzo(a)pyrene/toxicity , Benzo(a)pyrene/chemistry , Benzo(a)pyrene/metabolism , DNA/chemistry , DNA/metabolism , Benzopyrenes/toxicity , Benzopyrenes/chemistry , Benzopyrenes/metabolism , DNA-Binding Proteins/metabolism , DNA-Binding Proteins/genetics , DNA-Binding Proteins/chemistryABSTRACT
In a series of previous studies we reported that black raspberry (BRB) powder inhibits dibenzo[a,l]pyrene (DBP)-induced DNA damage, mutagenesis, and oral squamous cell carcinoma (OSCC) development in mice. In the present study, using human oral leukoplakia (MSK-Leuk1) and squamous cell carcinoma (SCC1483) cells, we tested the hypothesis that BRB extract (BRBE) will enhance the synthesis of glutathione (GSH) and in turn increase GSH conjugation of the fjord-region DBP diol epoxide (DBPDE) derived from DBP leading to inhibition of DBP-induced DNA damage. The syntheses of DBPDE-GSH conjugate, DBPDE-dA adduct, and the corresponding isotope-labeled internal standards were performed; LC-MS/MS methods were used for their quantification. BRBE significantly (p < 0.05) increased cellular GSH by 31% and 13% at 6 and 24 h, respectively, in OSCC cells; in MSK-LeuK1 cells, the levels of GSH significantly (p < 0.05) increased by 55% and 22%, at 1 and 6 h. Since BRBE significantly enhanced the synthesis of GSH in both cell types, subsequent experiments were performed in MSK-Leuk1 cells. Western blot analysis was performed to determine the types of proteins involved in the synthesis of GSH. BRBE significantly (p < 0.05) increased the protein expression (2.5-fold) of the glutamate-cysteine ligase catalytic subunit (GCLC) but had no effect on the glutamate-cysteine ligase modifier subunit (GCLM) and glutathione synthetase (GSS). LC-MS/MS analysis showed that pretreatment of cells with BRBE followed by DBPDE significantly (p < 0.05) increased the levels of DBPDE-GSH conjugate (2.5-fold) and decreased DNA damage by 74% measured by assessing levels of DBPDE-dA adduct formation. Collectively, the results of this in vitro study clearly support our hypothesis, and the LC-MS/MS methods developed in the present study will be highly useful in testing the same hypothesis initially in our mouse model and ultimately in smokers.
Subject(s)
Carcinoma, Squamous Cell , Mouth Neoplasms , Rubus , Humans , Mice , Animals , Carcinogens , Chrysenes , Benzopyrenes/metabolism , Epoxy Compounds , Nicotiana/metabolism , Glutamate-Cysteine Ligase , DNA Adducts , Chromatography, Liquid , Estuaries , Mouth Neoplasms/chemically induced , Tandem Mass Spectrometry , Glutathione/metabolism , Plant Extracts/pharmacologyABSTRACT
Developmental toxicity testing is an animal-intensive endpoints in toxicity testing and calls for animal-free alternatives. Previous studies showed the applicability of an in vitro-in silico approach for predicting developmental toxicity of a range of compounds, based on data from the mouse embryonic stem cell test (EST) combined with physiologically based kinetic (PBK) modelling facilitated reverse dosimetry. In the current study, the use of this approach for predicting developmental toxicity of polycyclic aromatic hydrocarbons (PAHs) was evaluated, using benzo[a]pyrene (BaP) as a model compound. A rat PBK model of BaP was developed to simulate the kinetics of its main metabolite 3-hydroxybenzo[a]pyrene (3-OHBaP), shown previously to be responsible for the developmental toxicity of BaP. Comparison to in vivo kinetic data showed that the model adequately predicted BaP and 3-OHBaP blood concentrations in the rat. Using this PBK model and reverse dosimetry, a concentration-response curve for 3-OHBaP obtained in the EST was translated into an in vivo dose-response curve for developmental toxicity of BaP in rats upon single or repeated dose exposure. The predicted half maximal effect doses (ED50) amounted to 67 and 45 mg/kg bw being comparable to the ED50 derived from the in vivo dose-response data reported for BaP in the literature, of 29 mg/kg bw. The present study provides a proof of principle of applying this in vitro-in silico approach for evaluating developmental toxicity of BaP and may provide a promising strategy for predicting the developmental toxicity of related PAHs, without the need for extensive animal testing.
Subject(s)
Benzo(a)pyrene/administration & dosage , Benzopyrenes/metabolism , Models, Biological , Animals , Benzo(a)pyrene/pharmacokinetics , Benzo(a)pyrene/toxicity , Computer Simulation , Dose-Response Relationship, Drug , Male , Rats , Rats, Sprague-Dawley , Toxicity Tests/methodsABSTRACT
Polycyclic aromatic hydrocarbons (PAHs) are widely distributed in environments, and some of them are causative agents of human cancer. Previous studies concluded that benzo[a]pyrene-7,8-dione (BPQ), which is one kind of carcinogenic PAH metabolites, forms covalently bonded adducts with DNA, and the major adduct formed is a deoxyguanosine adduct. In this work, we investigate the interactions between BPQ and DNA molecules via first-principles calculations. We identify six possible DNA adducts with BPQ. In addition to the four adducts forming covalent bonds, there are two adducts bound purely by van der Waals (vdW) interactions. Remarkably, the two vdW-bound adducts have comparable, if not larger, binding energies as the covalent adducts. The results may help us gain more understanding of the interactions between PAH metabolites and DNA.
Subject(s)
Benzopyrenes/chemistry , DNA Adducts/chemistry , Density Functional Theory , Molecular Dynamics Simulation , Benzopyrenes/metabolism , DNA Adducts/metabolism , Molecular StructureABSTRACT
Resistance of cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis represents the major hurdle to the clinical use of TRAIL or its derivatives. The discovery and development of lead compounds able to sensitize tumor cells to TRAIL-induced cell death is thus likely to overcome this limitation. We recently reported that marine actinomycetes' crude extracts could restore TRAIL sensitivity of the MDA-MB-231 resistant triple negative breast cancer cell line. We demonstrate in this study, that purified secondary metabolites originating from distinct marine actinomycetes (sharkquinone (1), resistomycin (2), undecylprodigiosin (3), butylcyclopentylprodigiosin (4), elloxizanone A (5) and B (6), carboxyexfoliazone (7), and exfoliazone (8)), alone, and in a concentration-dependent manner, induce killing in both MDA-MB-231 and HCT116 cell lines. Combined with TRAIL, these compounds displayed additive to synergistic apoptotic activity in the Jurkat, HCT116 and MDA-MB-231 cell lines. Mechanistically, these secondary metabolites induced and enhanced procaspase-10, -8, -9 and -3 activation leading to an increase in PARP and lamin A/C cleavage. Apoptosis induced by these compounds was blocked by the pan-caspase inhibitor QvD, but not by a deficiency in caspase-8, FADD or TRAIL agonist receptors. Activation of the intrinsic pathway, on the other hand, is likely to explain both their ability to trigger cell death and to restore sensitivity to TRAIL, as it was evidenced that these compounds could induce the downregulation of XIAP and survivin. Our data further highlight that compounds derived from marine sources may lead to novel anti-cancer drug discovery.
Subject(s)
Actinobacteria/metabolism , Aquatic Organisms/metabolism , Down-Regulation/drug effects , Drug Discovery/methods , Drug Resistance, Neoplasm/drug effects , Secondary Metabolism/physiology , Survivin/metabolism , TNF-Related Apoptosis-Inducing Ligand/pharmacology , X-Linked Inhibitor of Apoptosis Protein/metabolism , Apoptosis/drug effects , Benzopyrenes/metabolism , Benzopyrenes/pharmacology , Caspase 8/genetics , Cell Survival/drug effects , Gene Deletion , HCT116 Cells , Humans , Jurkat Cells , Oxazines/metabolism , Oxazines/pharmacology , Prodigiosin/analogs & derivatives , Prodigiosin/metabolism , Prodigiosin/pharmacology , Quinones/metabolism , Quinones/pharmacologyABSTRACT
To reveal the impacts of smoking on genetic architecture of human body weight, we conducted a genome-wide association study on 5,336 subjects in four ethnic populations from MESA (The Multi-Ethnic Study of Atherosclerosis) data. A full genetic model was applied to association mapping for analyzing genetic effects of additive, dominance, epistasis, and their ethnicity-specific effects. Both the unconditional model (base) and conditional model including smoking as a cofactor were investigated. There were 10 SNPs involved in 96 significant genetic effects detected by the base model, which accounted for a high heritability (61.78%). Gene ontology analysis revealed that a number of genetic factors are related to the metabolic pathway of benzopyrene, a main compound in cigarettes. Smoking may play important roles in genetic effects of dominance, dominance-related epistasis, and gene-ethnicity interactions on human body weight. Gene effect prediction shows that the genetic effects of smoking cessation on body weight vary from different populations.
Subject(s)
Body Weight , Genome-Wide Association Study , Smoking/genetics , Atherosclerosis/pathology , Benzopyrenes/chemistry , Benzopyrenes/metabolism , Epistasis, Genetic , Ethnicity/genetics , Gene Ontology , Genotype , Humans , Phenotype , Polymorphism, Single Nucleotide , Quantitative Trait, HeritableABSTRACT
The presence of polycyclic aromatic hydrocarbons (PAHs) in marine environments as a result of contamination is an environmental concern, especially in regions where oil spills such as the Deepwater Horizon have occurred. While numerous PAHs have been studied for their effects on microbes, the family of dibenzopyrenes has yet to be investigated. In this preliminary study, the impacts of these molecules on the community structure of a bacterial consortium isolated from oil-impacted Gulf of Mexico sediment were examined using high-throughput sequencing, demonstrating intriguing negative impacts on species diversity and abundance. While no measurable degradation of the dibenzopyrenes was observed after 28-day incubation, the abundance of known oil-degrading bacteria from orders such as Oceanospirillales, Caulobacterales, Sphingomonadales, and Nitrosococcales were shown to be enhanced. Of the five isomers of dibenzopyrene studied, dibenzo[a,h]pyrene supported the fewer number of microbial species suggesting the isomer was more toxic compared to the other isomers.
Subject(s)
Bacteria/metabolism , Benzopyrenes/analysis , Geologic Sediments/chemistry , Geologic Sediments/microbiology , Polycyclic Aromatic Hydrocarbons/analysis , Water Pollutants, Chemical/analysis , Benzopyrenes/metabolism , Gulf of Mexico , Petroleum Pollution/adverse effects , Polycyclic Aromatic Hydrocarbons/metabolism , Seawater/chemistry , Seawater/microbiology , Water Pollutants, Chemical/metabolismABSTRACT
The aims of this work were to assess the PAH exposure among roofers and to identify relevant biomarkers for monitoring occupational exposure. Several campaigns were conducted between 2004 and 2017, with 28 individual air samples and 240 urinary samples collected from 73 roofers. Seventeen parent PAHs and 14 urinary biomarkers, metabolites of pyrene (1-OHP), benzo(a)pyrene (3-OHBaP and TetraolBaP), naphthalene (1- and 2-naphtols), fluorene (1- 2- 3- 9-fluorenols) and phenanthrene (1- 2- 3- 4- 9-phenanthrols), were analysed. Three exposure groups were considered: soft-applied roofing using polymer-modified bitumen ("PMB"), hot-applied roofing using oxidized bitumen ("OB") and the tearing off of old roof coatings containing coal tar ("CT"). The PAHs containing 2-3 rings were much more abundant, and the highest airborne levels were observed in the "CT" group. The biomonitoring results were consistent with these results, with a large predominance of 2-3 ring PAH metabolites. 1-OHP, 3-fluorenol and 2-phenanthrol were better correlated with airborne levels and less influenced by smoking than the other metabolites. Conversely, 1-/2-naphtol levels were heavily influenced by smoking and not correlated with airborne naphthalene levels. Moreover, 3-OHBaP and TetraolBaP levels were very low when applying bitumen membranes, and much higher exposures were observed during tear-off activities. In this context, the recommended strategy for roofer biomonitoring should include 1-OHP, fluorenols and phenanthrols, as well as carcinogenic BaP metabolites (3-OHBaP or TetraolBaP) when evaluating the occupational exposure of roofers that are tearing off old roof coatings.
Subject(s)
Biological Monitoring/methods , Occupational Exposure/analysis , Polycyclic Aromatic Hydrocarbons/urine , Adult , Air Pollutants, Occupational/analysis , Benzopyrenes/metabolism , Biomarkers/urine , Female , Humans , Male , Middle Aged , Particulate Matter/analysis , Pyrenes/metabolism , Young AdultABSTRACT
Ovarian cancer ranked second in incidence among gynecologic cancers, but it causes more deaths than any other gynecologic cancer; at present there is no curative treatment beyond surgery. Animal models that employ carcinogens found in the human environment can provide a realistic platform to understand the mechanistic basis for disease development and to design rational chemopreventive/therapeutic strategies. We and others have shown that the administration of the environmental pollutant and tobacco smoke constituent dibenzo[ def,p]chrysene (DBP) to mice by several routes of exposure can induce tumors in multiple sites including the ovary. In the present study we compared, for the first time, the tumorigenicity and DNA damage induced by DBP and its metabolites DBP-dihydrodiol (DBPDHD) and DBP-dihydrodiol epoxide (DBPDE) in the mouse ovary. Compounds were dissolved in dimethyl sulfoxide (DMSO) as the vehicle and administered by topical application into the mouse oral cavity three times per week for 38 weeks. No tumors were observed in mice treated with DMSO. At equal dose (24 nmol/30 µL DMSO), the incidence of ovarian tumors induced by DBPDHD was higher (60.7%), although not significantly, than that induced by DBP (44.8%). Similarly the levels of DNA damage induced by DBPDHD in the ovary were higher than those observed with DBP. We did not observe any histological abnormality in the ovary of mice treated with DBPDE, which is consistent with lack of DNA damage. Our results suggested that both DBP and DBPDHD can be metabolized in the mouse ovary leading to the formation of DBPDE that can damage DNA, which is a prerequisite step in the initiation stage of carcinogenesis.
Subject(s)
Benzopyrenes/toxicity , DNA Damage/drug effects , Ovarian Neoplasms/etiology , Ovary/drug effects , Administration, Topical , Animals , Benzopyrenes/metabolism , Carcinogens/metabolism , Carcinogens/toxicity , Chromatography, High Pressure Liquid , DNA Adducts/analysis , Female , Mice , Ovarian Neoplasms/mortality , Ovarian Neoplasms/veterinary , Ovary/pathology , Survival Rate , Tandem Mass SpectrometryABSTRACT
Previously, we showed that oral application of the environmental pollutant dibenzo[a,l]pyrene (DB[a,l]P) induces oral tumors in mice. Thus, in the present investigation we examined the effect of alcohol on DB[a,l]P-induced DNA damage and immune regulation; we showed that alcohol (6.4% v/v in the diet, 35% of Calories) significantly enhanced the levels of (-)-anti-trans-DB[a,l]P-dA while decreased the levels of GSH in the mouse oral tissues. Analysis of RNA expression revealed that DB[a,l]P alone upregulates inflammatory genes while alcohol suppresses several markers of immune surveillance. Collectively, these results suggest that alcohol may enhance oral carcinogenesis induced by DB[a,l]P.
Subject(s)
Alcohol Drinking/adverse effects , Benzopyrenes/metabolism , DNA Damage , Environmental Pollutants/metabolism , Mouth/metabolism , Alcohol Drinking/immunology , Alcoholism , Animals , Carcinogenesis , Mice , Mouth/immunology , Mouth NeoplasmsABSTRACT
Benzo[a]pyrene, a potent human carcinogen, is metabolized in vivo to a diol epoxide that reacts with the N2-position of guanine to produce N2-BP-dG adducts. These adducts are mutagenic causing G to T transversions. These adducts block replicative polymerases but can be bypassed by the Y-family translesion synthesis polymerases. The mechanisms by which mutagenic bypass occurs is not well-known. We have evaluated base pairing structures using atomic substitution of the dNTP with two stereoisomers, 2'-deoxy-N-[(7R,8S,9R,10S)-7,8,9,10-tetrahydro-7,8,9-trihydroxybenzo[a]pyren-10-yl]guanosine and 2'-deoxy-N-[(7S,8R,9S,10R)-7,8,9,10-tetrahydro-7,8,9-trihydroxybenzo[a]pyren-10-yl]guanosine. We have examined the kinetics of incorporation of 1-deaza-dATP, 7-deaza-dATP, 2'-deoxyinosine triphosphate, and 7-deaza-dGTP, analogues of dATP and dGTP in which single atoms are changed. Changes in rate will occur if that atom provided a critical interaction in the transition state of the reaction. We examined two polymerases, Escherichia coli DNA polymerase I (Kf) and Sulfolobus solfataricus DNA polymerase IV (Dpo4), as models of a high fidelity and TLS polymerase, respectively. We found that with Kf, substitution of the nitrogens on the Watson-Crick face of the dNTPs resulted in decreased rate of reactions. This result is consistent with a Hoogsteen base pair in which the template N2-BP-dG flipped from the anti to syn conformation. With Dpo4, while the substitution did not affect the rate of reaction, the amplitude of the reaction decreased with all substitutions. This result suggests that Dpo4 bypasses N2-BP-dG via Hoogsteen base pairs but that the flipped nucleotide can be either the dNTP or the template.
Subject(s)
Benzopyrenes/metabolism , DNA Adducts , DNA Polymerase I/metabolism , DNA Polymerase beta/metabolism , DNA Replication , Deoxyguanosine/analogs & derivatives , Escherichia coli/enzymology , Sulfolobus solfataricus/enzymology , Base Pairing , Catalysis , Deoxyguanosine/metabolismABSTRACT
The aryl hydrocarbon receptor (AhR) pathway plays a key role in receptor activator of NF-κB ligand (RANKL)-mediated osteoclastogenesis. However, the mechanism underlying the regulation of AhR expression in osteoclasts and the signaling pathway through which AhR controls osteoclastogenesis remain unclear. We found that the expression of AhR in bone marrow-derived osteoclasts was upregulated by RANKL at an earlier stage than was the expression of signature osteoclast genes such as those encoding cathepsin K and NFAT, cytoplasmic, calcineurin-dependent 1. In response to RANKL, bone marrow macrophages isolated from AhR-/- mice exhibited impaired phosphorylation of Akt and MAPK as well as NF-κB, whereas their response to M-CSF remained unchanged. Osteoclast differentiation mediated by the AhR signaling pathway was also regulated in an RANKL/c-Fos-dependent manner. Furthermore, ligand activation of AhR by the smoke toxin benzo[a]pyrene accelerated osteoclast differentiation in a receptor-dependent manner, and AhR-dependent regulation of mitochondrial biogenesis in osteoclasts was observed. Moreover, AhR-/- mice exhibited impaired bone healing with delayed endochondral ossification. Taken together, the present results suggest that the RANKL/AhR/c-Fos signaling axis plays a critical role in osteoclastogenesis, thereby identifying the potential of AhR in treating pathological, inflammatory, or metabolic disorders of the bone.
Subject(s)
Mitochondria/metabolism , Osteoclasts/physiology , Osteogenesis , Receptors, Aryl Hydrocarbon/metabolism , Animals , Benzopyrenes/metabolism , Bone Marrow Cells/physiology , Cells, Cultured , Mice , Mice, Inbred C57BL , Mice, Knockout , NF-kappa B/metabolism , Osteogenesis/genetics , Proto-Oncogene Proteins c-fos/metabolism , RANK Ligand/metabolism , Receptors, Aryl Hydrocarbon/genetics , Signal TransductionABSTRACT
Metabolism is a key health risk factor following exposures to pro-carcinogenic polycyclic aromatic hydrocarbons (PAHs) such as dibenzo[def,p]chrysene (DBC), an IARC classified 2A probable human carcinogen. Human exposure to PAHs occurs primarily from the diet in nonsmokers. However, little data is available on the metabolism and pharmacokinetics in humans of high molecular weight PAHs (≥4 aromatic rings), including DBC. We previously determined the pharmacokinetics of DBC in human volunteers orally administered a microdose (29 ng; 5 nCi) of [14C]-DBC by accelerator mass spectrometry (AMS) analysis of total [14C] in plasma and urine. In the current study, we utilized a novel "moving wire" interface between ultraperformance liquid chromatography (UPLC) and AMS to detect and quantify parent DBC and its major metabolites. The major [14C] product identified in plasma was unmetabolized [14C]-DBC itself (Cmax = 18.5 ±15.9 fg/mL, Tmax= 2.1 ± 1.0 h), whereas the major metabolite was identified as [14C]-(+/-)-DBC-11,12-diol (Cmax= 2.5 ±1.3 fg/mL, Tmax= 1.8 h). Several minor species of [14C]-DBC metabolites were also detected for which no reference standards were available. Free and conjugated metabolites were detected in urine with [14C]-(+/-)-DBC-11,12,13,14-tetraol isomers identified as the major metabolites, 56.3% of which were conjugated (Cmax= 35.8 ± 23.0 pg/pool, Tmax = 6-12 h pool). [14C]-DBC-11,12-diol, of which 97.5% was conjugated, was also identified in urine (Cmax = 29.4 ± 11.6 pg/pool, Tmax = 6-12 h pool). Parent [14C]-DBC was not detected in urine. This is the first data set to assess metabolite profiles and associated pharmacokinetics of a carcinogenic PAH in human volunteers at an environmentally relevant dose, providing the data necessary for translation of high dose animal models to humans for translation of environmental health risk assessment.
Subject(s)
Benzopyrenes/metabolism , Benzopyrenes/pharmacokinetics , Adult , Aged , Benzopyrenes/analysis , Chromatography, High Pressure Liquid , Dose-Response Relationship, Drug , Female , Healthy Volunteers , Humans , Male , Mass Spectrometry , Middle Aged , Molecular Structure , Young AdultABSTRACT
Benzo[a]pyrene (BaP) is a by-product of incomplete combustion of fossil fuels and plant/wood products, including tobacco. A physiologically based pharmacokinetic (PBPK) model for BaP for the rat was extended to simulate inhalation exposures to BaP in rats and humans including particle deposition and dissolution of absorbed BaP and renal elimination of 3-hydroxy benzo[a]pyrene (3-OH BaP) in humans. The clearance of particle-associated BaP from lung based on existing data in rats and dogs suggest that the process is bi-phasic. An initial rapid clearance was represented by BaP released from particles followed by a slower first-order clearance that follows particle kinetics. Parameter values for BaP-particle dissociation were estimated using inhalation data from isolated/ventilated/perfused rat lungs and optimized in the extended inhalation model using available rat data. Simulations of acute inhalation exposures in rats identified specific data needs including systemic elimination of BaP metabolites, diffusion-limited transfer rates of BaP from lung tissue to blood and the quantitative role of macrophage-mediated and ciliated clearance mechanisms. The updated BaP model provides very good prediction of the urinary 3-OH BaP concentrations and the relative difference between measured 3-OH BaP in nonsmokers versus smokers. This PBPK model for inhaled BaP is a preliminary tool for quantifying lung BaP dosimetry in rat and humans and was used to prioritize data needs that would provide significant model refinement and robust internal dosimetry capabilities.
Subject(s)
Benzo(a)pyrene/pharmacokinetics , Carcinogens/pharmacokinetics , Lung/metabolism , Models, Biological , Particulate Matter/pharmacokinetics , Administration, Inhalation , Administration, Oral , Animals , Benzo(a)pyrene/administration & dosage , Benzopyrenes/metabolism , Carcinogens/administration & dosage , Humans , Inhalation Exposure , Particulate Matter/administration & dosage , RatsABSTRACT
Benzo[a]pyrene (BP) is a well-known and frequently encountered carcinogen which generates a bulky DNA adduct (+)-trans-10S-BP-N(2)-dG (BP-dG) in cells. DNA polymerase kappa (polκ) is the only known Y-family polymerase that bypasses BP-dG accurately and thus protects cells from genotoxic BP. Here, we report the structures of human polκ in complex with DNA containing either a normal guanine (G) base or a BP-dG adduct at the active site and a correct deoxycytidine. The structures and supporting biochemical data reveal a unique mechanism for accurate replication by translesion synthesis past the major bulky adduct. The active site of polκ opens at the minor groove side of the DNA substrate to accommodate the bulky BP-dG that is attached there. More importantly, polκ stabilizes the lesion DNA substrate in the same active conformation as for regular B-form DNA substrates and the bulky BPDE ring in a 5' end pointing conformation. The BP-dG adducted DNA substrate maintains a Watson-Crick (BP-dG:dC) base pair within the active site, governing correct nucleotide insertion opposite the bulky adduct. In addition, polκ's unique N-clasp domain supports the open conformation of the enzyme and the extended conformation of the single-stranded template to allow bypass of the bulky lesion. This work illustrates the first molecular mechanism for how a bulky major adduct is replicated accurately without strand misalignment and mis-insertion.
Subject(s)
Benzopyrenes/chemistry , DNA Replication , DNA-Directed DNA Polymerase/chemistry , Deoxyguanosine/analogs & derivatives , Benzopyrenes/metabolism , Binding Sites , Catalytic Domain , DNA/chemistry , DNA/metabolism , DNA-Directed DNA Polymerase/metabolism , Deoxyguanosine/chemistry , Deoxyguanosine/metabolism , Humans , Nucleic Acid Conformation , Protein Binding , Protein DomainsABSTRACT
Polycyclic aromatic hydrocarbons (PAHs) are ubiquitous carcinogenic pollutants emitted in complex mixtures in the ambient air and contribute to the incidence of human cancers. Taking into account all absorption routes, biomonitoring is more relevant than atmospheric measurements to health risk assessment, but knowledge about how to use biomarkers is essential. In this work, urinary elimination kinetic of 1-hydroxypyrene (1-OHP) and 3-hydroxybenzo(a)pyrene (3-OHBaP) were studied in six electrometallurgy workers after PAHs exposure. Spot samples were collected on pre- and post-shift of the last workday then the whole urinations were separately sampled during the weekend. Non-linear mixed effects models were built to study inter- and intra-individual variability of both urinary metabolites toxicokinetic and investigate diuresis correction ways. Comparison of models confirmed the diuresis correction requirement to perform urinary biomonitoring of pyrene and BaP exposure. Urinary creatinine was found as a better way than specific gravity to normalize urinary concentrations of 1-OHP and as a good compromise for 3-OHBaP. Maximum observed levels were 1.0 µmol/mol creatinine and 0.8nmol/mol creatinine for 1-OHP and 3-OHBaP, respectively. Urinary 1-OHP concentrations on post-shift were higher than pre-shift for each subject, while 3-OHBaP levels were steady or decreased, and maximum urinary excretion rates of 3-OHBaP was delayed compared to 1-OHP. These results were consistent with the sampling time previously proposed for 3-OHBaP analysis, the next morning after exposure. Apparent urinary half-life of 1-OHP and 3-OHBaP ranged from 12.0h to 18.2h and from 4.8h to 49.5h, respectively. Finally, inter-individual variability of 1-OHP half-life seemed linked with the cutaneous absorption extent during exposure, while calculation of 3-OHBaP half-life required the awareness of individual urinary background level. The toxicokinetic modeling described here is an efficient tool which could be used to describe elimination kinetic and determine diuresis correction way for any other urinary biomarkers of chemicals or metals exposure.
Subject(s)
Benzopyrenes/pharmacokinetics , Environmental Monitoring/methods , Occupational Exposure/analysis , Pyrenes/pharmacokinetics , Adult , Benzopyrenes/metabolism , Biomarkers/urine , Diuresis , Healthy Volunteers , Humans , Male , Metallurgy , Middle Aged , Pyrenes/urineABSTRACT
Benzo(a)pyrene (BaP) is a ubiquitous carcinogen resulting from incomplete combustion of organic compounds and also present at high levels in cigarette smoke. A wide range of biological effects has been attributed to BaP and its genotoxic metabolite BPDE, but the contribution to BaP toxicity of intermediary metabolites generated along the detoxification path remains unknown. Here, we report for the first time how 3-OH-BaP, 9,10-diol and BPDE, three major BaP metabolites, temporally relate to BaP-induced transcriptomic alterations in HepG2 cells. Since BaP is also known to induce AhR activation, we additionally evaluated TCDD to source the expression of non-genotoxic AhR-mediated patterns. 9,10-Diol was shown to activate several transcription factor networks related to BaP metabolism (AhR), oxidative stress (Nrf2) and cell proliferation (HIF-1α, AP-1) in particular at early time points, while BPDE influenced expression of genes involved in cell energetics, DNA repair and apoptotic pathways. Also, in order to grasp the role of BaP and its metabolites in chemical hepatocarcinogenesis, we compared expression patterns from BaP(-metabolites) and TCDD to a signature set of approximately nine thousand gene expressions derived from hepatocellular carcinoma (HCC) patients. While transcriptome modulation by TCDD appeared not significantly related to HCC, BaP and BPDE were shown to deregulate metastatic markers via non-genotoxic and genotoxic mechanisms and activate inflammatory pathways (NF-κß signaling, cytokine-cytokine receptor interaction). BaP also showed strong repression of genes involved in cholesterol and fatty acid biosynthesis. Altogether, this study provides new insights into BaP-induced toxicity and sheds new light onto its mechanism of action as a hepatocarcinogen.
Subject(s)
Benzo(a)pyrene/toxicity , Carcinogens, Environmental/toxicity , DNA Adducts/genetics , DNA Damage , Liver Neoplasms/genetics , Transcriptome/drug effects , 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide/metabolism , 7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide/toxicity , Benzo(a)pyrene/metabolism , Benzopyrenes/metabolism , Benzopyrenes/toxicity , Carcinogens, Environmental/metabolism , DNA Adducts/metabolism , Dihydroxydihydrobenzopyrenes/metabolism , Dihydroxydihydrobenzopyrenes/toxicity , Hep G2 Cells , Humans , Liver Neoplasms/chemically inducedABSTRACT
Dibenzo[def,p]chrysene (DBC) is the most carcinogenic polycyclic aromatic hydrocarbon (PAH) examined to date. We investigated the immunotoxicity of DBC, manifested as spleen atrophy, following acute exposure of adult MutaMouse males by oral gavage. Mice were exposed to 0, 2.0, 6.2, or 20.0 mg DBC /kg-bw per day, for 3 days. Genotoxic endpoints (DBC-DNA adducts and lacZ mutant frequency in spleen and bone marrow, and red blood cell micronucleus frequency) and global gene expression changes were measured. All of the genotoxicity measures increased in a dose-dependent manner in spleen and bone marrow. Gene expression analysis showed that DBC activates p53 signaling pathways related to cellular growth and proliferation, which was evident even at the low dose. Strikingly, the expression profiles of DBC exposed mouse spleens were highly inversely correlated with the expression profiles of the only published toxicogenomics dataset of enlarged mouse spleen. This analysis suggested a central role for Bnip3l, a pro-apoptotic protein involved in negative regulation of erythroid maturation. RT-PCR confirmed expression changes in several genes related to apoptosis, iron metabolism, and aryl hydrocarbon receptor signaling that are regulated in the opposite direction during spleen atrophy versus benzo[a]pyrene-mediated splenomegaly. In addition, benchmark dose modeling of toxicogenomics data yielded toxicity estimates that are very close to traditional toxicity endpoints. This work illustrates the power of toxicogenomics to reveal rich mechanistic information for immunotoxic compounds and its ability to provide information that is quantitatively similar to that derived from standard toxicity methods in health risk assessment.
Subject(s)
Benzopyrenes/toxicity , Carcinogens/toxicity , Gene Expression Profiling , Spleen/drug effects , Animals , Atrophy/chemically induced , Benchmarking , Benzopyrenes/metabolism , Bone Marrow/drug effects , Bone Marrow/metabolism , DNA Adducts/analysis , Dose-Response Relationship, Drug , Male , Membrane Proteins/analysis , Mice , Mice, Transgenic , Mitochondrial Proteins/analysis , Organ Specificity , Reticulocytes/drug effects , Reticulocytes/ultrastructure , Sequence Analysis, RNA , Spleen/metabolism , Spleen/pathology , ToxicogeneticsABSTRACT
Dibenzo[def,p]chrysene (DBP), a representative example of the class of polycyclic aromatic hydrocarbon (PAH), is known to induce tumors in multiple organ sites including the ovary, lung, mammary glands, and oral cavity in rodents. The goal of this study was to test the hypothesis that the levels of DBP and its metabolites that reach and retain the levels for an extended time in the target organs as well as the capacity of these organs to metabolize this carcinogen to active metabolites that can damage DNA may account for its tissue selective tumorigenicity. Therefore, we used the radiolabeled [(3)H] DBP to accurately assess the tissue distribution, excretion, and pharmacokinetics of this carcinogen. We also compared the levels of DBPDE-DNA adducts in a select target organ (ovary) and nontarget organs (kidney and liver) in mice treated orally with DBP. Our results showed that after 1 week, 91.40 ± 7.23% of the radioactivity was recovered in the feces; the corresponding value excreted in the urine was less than 2% after 1 week. After 24 h, the stomach had the highest radioactivity followed by the intestine and the liver; however, after 1 week, levels of the radioactivity in these organs were the lowest among tissues examined including the ovary and liver; the pharmacokinetic analysis of DBP was conducted using a one compartment open model. The level of (-)-anti-trans-DBPDE-dA in the ovaries (8.91 ± 0.08 adducts/10(7) dA) was significantly higher (p < 0.01) than the levels of adducts in kidneys (0.69 ± 0.09 adducts/10(7) dA) and livers (0.63 ± 0.11 adducts/10(7) dA). Collectively, the results of the tissue distribution and pharmacokinetic analysis may not fully support our hypothesis, but the capacity of the target organs vs nontarget organs to metabolize DBP to active intermediates that can damage DNA may account for its tissue selective tumorigenicity.
Subject(s)
Benzopyrenes/metabolism , Environmental Pollutants/metabolism , Animals , Benzopyrenes/chemistry , Benzopyrenes/toxicity , Chromatography, High Pressure Liquid , DNA/chemistry , DNA/metabolism , DNA Adducts/analysis , DNA Damage/drug effects , Environmental Pollutants/chemistry , Environmental Pollutants/toxicity , Feces/chemistry , Female , Half-Life , Mice , Tandem Mass Spectrometry , Tissue Distribution , Tritium/chemistryABSTRACT
BACKGROUND: Protein oxidation is considered to be one of the main causes of cell death, and methionine is one of the primary targets of reactive oxygen species (ROS). However, the mechanisms by which nickel nanoparticles (NiNPs) cause oxidative damage to proteins remain unclear. OBJECTIVES: The objective of this study is to investigate the effects of NiNPs on the methionine sulfoxide reductases (MSR) protein repairing system. METHODS: Two physically similar nickel-based nanoparticles, NiNPs and carbon-coated NiNP (C-NiNPs; control particles), were exposed to human epithelial A549 cells. Cell viability, benzo(a)pyrene diolepoxide (BPDE) protein adducts, methionine oxidation, MSRA and B3, microtubule-associated protein 1A/1B-light chain 3 (LC3) and extracellular signal-regulated kinase (ERK) phosphorylation were investigated. RESULTS: Exposure to NiNPs led to a dose-dependent reduction in cell viability and increased BPDE protein adduct production and methionine oxidation. The methionine repairing enzymatic MSRA and MSRB3 production were suppressed in response to NiNP exposure, suggesting the oxidation of methionine to MetO by NiNP was not reversed back to methionine. Additionally, LC3, an autophagy marker, was down-regulated by NiNPs. Both NiNP and C-NiNP caused ERK phosphorylation. LC3 was positively correlated with MSRA (r = 0.929, p < 0.05) and MSRB3 (r = 0.893, p < 0.05). CONCLUSIONS: MSR was made aberrant by NiNP, which could lead to the dysfunction of autophagy and ERK phosphorylation. The toxicological consequences may be dependent on the chemical characteristics of the nanoparticles.