212 results on '"Eaton, David L."'
Search Results
202. Estradiol metabolites as isoform-specific inhibitors of human glutathione S-transferases.
- Author
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Abel EL, Lyon RP, Bammler TK, Verlinde CL, Lau SS, Monks TJ, and Eaton DL
- Subjects
- Enzyme Inhibitors metabolism, Estradiol metabolism, Glutathione Transferase isolation & purification, Humans, Isoenzymes isolation & purification, Spectrometry, Mass, Electrospray Ionization, Enzyme Inhibitors pharmacology, Estradiol pharmacology, Glutathione Transferase antagonists & inhibitors, Isoenzymes antagonists & inhibitors
- Abstract
Numerous studies have suggested that the lifetime dose of unopposed estrogen is a significant risk factor for breast and uterine cancer. Estradiol (E2) plays a putative role as a tumor promoter through interaction with estrogen receptors but can also be metabolized to redox active and/or mutagenic semiquinones and quinones. Similarly, equine estrogens (components of certain hormone replacement therapy preparations) are converted to quinone metabolites. The use of hormone replacement therapy has also been associated with increased breast and endometrial cancer risk. Recently, metabolites of certain equine estrogens have been shown to inhibit human glutathione S-transferases (hGSTs). Since E2 and equine estrogens share similarities in other biological interactions, we have investigated the inhibitory capacity of endogenously formed E2 metabolites toward various hGSTs. The quinone metabolite of 2-hydroxy-17-beta-estradiol (2-OH-E2) was synthesized, and inhibition of hGST-mediated biotransformation of model substrates was assessed. Inhibition of purified recombinant hGSTM1-1 and hGSTA1-1 occurred in a concentration-dependent manner with IC50-values of approximately 250 and 350 nM, respectively. hGSTs M2-2, P1-1 and T1-1 were significantly less sensitive to inhibition. Specific glutathione-conjugates of the estrogen quinone also potently inhibited hGSTM1-1 and hGSTA1-1. Mass spectrometry data indicate that the inhibition was not mediated via covalent adduction. Although we have demonstrated hGST inhibition via E2 metabolites, our findings indicate that the isoform specificity and potency of GST inhibition by endogenous E2 metabolites is different than that of equine estrogen metabolites.
- Published
- 2004
- Full Text
- View/download PDF
203. Characterization of atrazine biotransformation by human and murine glutathione S-transferases.
- Author
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Abel EL, Opp SM, Verlinde CL, Bammler TK, and Eaton DL
- Subjects
- Adolescent, Adult, Animals, Child, Cytosol enzymology, Female, Glutathione S-Transferase pi biosynthesis, Glutathione Transferase biosynthesis, Humans, In Vitro Techniques, Indicators and Reagents, Isoenzymes metabolism, Liver enzymology, Male, Mice, Models, Molecular, Recombinant Proteins metabolism, Species Specificity, Atrazine pharmacokinetics, Glutathione S-Transferase pi metabolism, Glutathione Transferase metabolism, Herbicides pharmacokinetics
- Abstract
Atrazine is one of the most widely used herbicides in the United States and has been detected, occasionally, at low levels in drinking water sources. The biotransformation of atrazine in humans has not been fully characterized. Rodent studies suggest Phase I-dominated biotransformation with minor Phase II-mediated biotransformation by glutathione S-transferase(s) (GST). In human urine, mercapturates of atrazine are significant metabolites, yet the specific GST form(s) responsible for glutathione (GSH) conjugation have not been identified. Using recombinant alpha, mu, pi and theta class human GSTs, we demonstrated that only hGSTP1-1 displays significant activity toward atrazine (7.1 nmol/min/mg protein). We also confirmed that mouse GST Pi (pi) protein is responsible for the GSH-dependent biotransformation of atrazine in mouse liver; recombinant mGSTP1-1 had a specific activity of 7.3-nmol/min/mg protein. Furthermore, cytosolic fractions from mouse and human liver conjugated atrazine with glutathione at rates of 282.3 and 3.0 pmol/min/mg, respectively. Docking studies of the atrazine-GST conjugate in the hGSTP1-1 substrate-binding site were used to elucidate a basis for the dramatic difference in activity between mouse GSTP1-1 and GSTP2-2 (7.14 versus 0.02 nmol/min/mg protein, respectively). The inactivity of mGSTP2-2 appears to be attributable to an indirect structural disruption of the G-site by Pro12. Possible effects of the hGSTP1 polymorphisms were investigated. No significant differences in catalytic-specific activity were noted among purified proteins corresponding to the four hGSTP1 variants: hGSTP1(*)A (most common form), hGSTP1(*)B (Ile105Val), hGSTP1(*)C (Ile105Val, Ala114Val), and hGSTP1(*)D (Ala114Val). Overall, this work supports a physiological role for GSTs in atrazine biotransformation and indicates a novel diagnostic substrate for human and mouse GSTP1-1 proteins.
- Published
- 2004
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204. Biotransformation of methyl parathion by glutathione S-transferases.
- Author
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Abel EL, Bammler TK, and Eaton DL
- Subjects
- Adolescent, Adult, Animals, Biotransformation, Child, Cytosol enzymology, Cytosol metabolism, Dealkylation, Dinitrochlorobenzene metabolism, Female, Gene Expression Regulation, Enzymologic, Genotype, Glutathione Transferase genetics, Humans, Isoenzymes genetics, Isoenzymes metabolism, Liver metabolism, Male, Mice, Middle Aged, Rats, Rats, Sprague-Dawley, Recombinant Proteins metabolism, Glutathione Transferase metabolism, Insecticides pharmacokinetics, Liver enzymology, Methyl Parathion pharmacokinetics
- Abstract
The organo(thio)phosphate esters are one of the most widely used classes of insecticides. Worldwide, organophosphate insecticides (OPs) result in numerous poisonings each year. In insects, glutathione S-transferases (GSTs) play an important role in OP resistance; limited data suggest that GST-mediated O-dealkylation occurs in humans as well. To characterize the capacity of mammalian GSTs to detoxify OPs, we investigated mammalian GST biotransformation of the widely used OP, methyl parathion (MeP). Cytosolic fractions isolated from rat, mouse, and ten individual adult human livers biotransformed 300 microM MeP at rates of 2.36, 1.76, and 0.70 (mean rate) nmol desmethyl parathion/min/mg, respectively. Our study focused on human GSTs; in particular, we investigated hGSTs M1-1 and T1-1, since deletion polymorphisms occur commonly in these genes. However, we found no correlation between hGSTM1/T1 genotypes and MeP O-dealkylation activities of the ten human liver cytosolic samples. We also measured MeP O-dealkylation activities of several purified recombinant GSTs belonging to the alpha (human GSTs A1-1 and A2-2, mouse GSTA3-3, rat GSTA5-5), mu (human GSTs M1a-1a, M2-2, M3-3, M4-4), pi (human GSTP1-1, mouse GSTs P1-1, P2-2), and theta (human GSTT1-1) classes. At 1 mM glutathione and 300 microM MeP concentrations, hGSTT1-1 and hGSTA1-1 exhibited the highest O-dealkylation activities: 545.8 and 65.0 nmol/min/mg, respectively. When expression level and enzymatic activity are considered, we estimate that hGSTA1-1 is responsible for the majority of MeP O-dealkylation in human hepatic cytosol. In target organs such as brain and skeletal muscle, where hGSTT1-1 is expressed, hGSTT1-1-mediated biotransformation of MeP may be important.
- Published
- 2004
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205. Tellurium-mediated cycloaromatization of acyclic enediynes under mild conditions.
- Author
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Landis CA, Payne MM, Eaton DL, and Anthony JE
- Abstract
The cycloaromatization of acyclic enediynes typically requires very high temperatures (>160 degrees C) and dilute conditions to proceed in a synthetically useful yield. These conditions hinder reaction throughput, inhibiting the use of this reaction for the large-scale production of materials. The reaction of sodium telluride with acyclic arenediynes yields the corresponding tellurepine, which under gentle heating extrudes Te degrees to yield the cycloaromatization product. We have developed conditions that form sodium telluride from inexpensive tellurium metal in situ, and that also perform the desilylation of silylated arenediynes in the same process. Under our conditions, we are able to perform desilylation and cycloaromatization at temperatures as low as 40 degrees C and on a scale as large as 5 g in standard laboratory glassware.
- Published
- 2004
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206. Glutathione S-transferase M1, T1, and P1 polymorphisms and survival among lung cancer patients.
- Author
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Sweeney C, Nazar-Stewart V, Stapleton PL, Eaton DL, and Vaughan TL
- Subjects
- Adolescent, Adult, Aged, Antineoplastic Combined Chemotherapy Protocols therapeutic use, Biomarkers, Tumor genetics, Carcinoma, Large Cell drug therapy, Carcinoma, Small Cell drug therapy, Carcinoma, Squamous Cell drug therapy, Case-Control Studies, Female, Follow-Up Studies, Genotype, Glutathione S-Transferase pi, Glutathione Transferase drug effects, Humans, Isoenzymes drug effects, Lung Neoplasms drug therapy, Male, Middle Aged, Odds Ratio, Polymorphism, Genetic drug effects, Proportional Hazards Models, Smoking drug therapy, Smoking genetics, Smoking mortality, Statistics as Topic, Survival Analysis, Treatment Outcome, Washington epidemiology, Carcinoma, Large Cell genetics, Carcinoma, Large Cell mortality, Carcinoma, Small Cell genetics, Carcinoma, Small Cell mortality, Carcinoma, Squamous Cell genetics, Carcinoma, Squamous Cell mortality, Glutathione Transferase genetics, Isoenzymes genetics, Lung Neoplasms genetics, Lung Neoplasms mortality, Polymorphism, Genetic genetics
- Abstract
Glutathione S-transferase (GST) enzymes detoxify therapeutic drugs and reactive oxidants, so GST polymorphisms may influence survival after diagnosis of cancer. We evaluated survival according to GST polymorphisms in a population-based series of lung cancer patients. The study subjects (n = 274) were men diagnosed with lung cancer from 1993 through 1996 who participated in a case control study and provided a blood sample for genotyping. The presence of the GSTM1 and GSTT1 genes were assayed by multiplex PCR. Genotype at the GSTP1 Ile(105)Val substitution was determined by PCR and oligonucleotide ligation assay. The study subjects were followed for vital status through 2000, and overall survival was evaluated in Kaplan-Meier survival functions and Cox proportional hazards models. Subjects with the GSTM1 null genotype had shorter survival; the proportion of GSTM1 null subjects surviving at 5 years was 0.20 [95% confidence interval (CI) 0.14-0.27], compared with 0.29 (95% CI 0.22-0.37) for GSTM1 present subjects. The relative risk of death associated with GSTM1 null genotype, adjusted for stage at diagnosis and histology, was 1.36, 95% CI 1.04-1.80. There was no association between GSTT1 or GSTP1 genotype and survival in the overall study population, nor in a subgroup of patients treated with chemotherapy (n = 130). For GSTM1, our results are consistent with a previous study, which also observed that the GSTM1-null genotype, which confers susceptibility to lung cancer, was associated with shorter survival. Future studies of lung cancer survival should take into account GSTM1 genotype as well as investigate underlying mechanisms.
- Published
- 2003
207. A population-based study of glutathione S-transferase M1, T1 and P1 genotypes and risk for lung cancer.
- Author
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Nazar-Stewart V, Vaughan TL, Stapleton P, Van Loo J, Nicol-Blades B, and Eaton DL
- Subjects
- Adolescent, Adult, Aged, Case-Control Studies, DNA, Neoplasm, Genotype, Glutathione S-Transferase pi, Humans, Male, Middle Aged, Odds Ratio, Polymerase Chain Reaction, Risk Factors, Washington epidemiology, Genetic Predisposition to Disease, Glutathione Transferase genetics, Isoenzymes genetics, Lung Neoplasms etiology, Lung Neoplasms genetics, Polymorphism, Genetic, Registries statistics & numerical data
- Abstract
A deletion polymorphism for glutathione S-transferase M1 (GSTM1) has been related to risk for lung cancer among smokers in some studies but not in others. We examined GSTM1, a GSTT1 deletion polymorphism and a common GSTP1 gene variant (iso-->val), as risk factors for lung cancer in a population-based case-control study of men. Cases (N=274) were males identified from 1993 to 1996 through the Fred Hutchinson Cancer Research Center Cancer Surveillance System registry for western Washington State. Male age-matched controls (N=501) were selected by random-digit dialing. Subjects participated in a telephone interview and blood draw. GSTM1 and GSTT1 were genotyped with a multiplex PCR assay using beta-globin as a positive control, and GSTP1 single nucleotide variant determined with PCR-based oligonucleotide ligation assays. GSTM1 absence was associated with a modest elevation in risk among all cases (odds ratio=1.27, 95% CI 0.91-1.77) and among non-small cell cancers (adenocarcinoma OR=1.58, 95% CI 0.99-2.52; squamous cell OR=1.40, 95% CI 0.83-2.34). Risk associated with GSTM1 null was increased two to sixfold among heavy smokers. GSTT1 was not associated with lung cancer risk and GSTP1 val was non-significantly associated with a modest reduction in risk, particularly among heavy smokers. No specific combination of GST genotypes was particularly associated with risk. These results support previous reports that the GSTM1 null genotype is associated with a modest increase in risk for lung cancer, particularly among heavy smokers, suggest no role for GSTT1 and the need for further study of GSTP1.
- Published
- 2003
- Full Text
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208. The role of genetic polymorphisms in environmental health.
- Author
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Kelada SN, Eaton DL, Wang SS, Rothman NR, and Khoury MJ
- Subjects
- Humans, Research trends, Research Design, Environmental Exposure, Environmental Health, Genetic Predisposition to Disease, Polymorphism, Genetic
- Abstract
Interest is increasing in the role of variations in the human genome (polymorphisms) in modifying the effect of exposures to environmental health hazards (often referred to as gene-environment interaction), which render some individuals or groups in the population more or less likely to develop disease after exposure. This review is intended for an audience of environmental health practitioners and students and is designed to raise awareness about this rapidly growing field of research by presenting established and novel examples of gene-environment interaction that illustrate the major theme of effect modification. Current data gaps are identified and discussed to illustrate limitations of past research and the need for the application of more robust methods in future research projects. Two primary benefits of incorporating genetics into the existing environmental health research framework are illustrated: a) the ability to detect different levels of risk within the population, and b) greater understanding of etiologic mechanisms. Both offer opportunities for developing new methods of disease prevention. Finally, we describe a basic framework for researchers interested in pursuing health effects research that incorporates genetic polymorphisms.
- Published
- 2003
- Full Text
- View/download PDF
209. Complementary DNA cloning, protein expression, and characterization of alpha-class GSTs from Macaca fascicularis liver.
- Author
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Wang C, Bammler TK, and Eaton DL
- Subjects
- Amino Acid Sequence, Animals, Cloning, Molecular, Gene Expression Regulation, Enzymologic, Gene Library, Glutathione Transferase metabolism, Humans, In Vitro Techniques, Isoenzymes metabolism, Macaca fascicularis, Male, Molecular Sequence Data, RNA, Messenger analysis, Sequence Alignment, Sequence Homology, Amino Acid, Species Specificity, DNA, Complementary genetics, Glutathione Transferase genetics, Isoenzymes genetics, Liver enzymology, Recombinant Proteins biosynthesis
- Abstract
Large species differences exist in sensitivity to aflatoxin B(1) (AFB(1))-induced liver cancer. Mice are resistant to AFB(1)-induced liver cancer because they express an alpha-class GST (mGSTA3-3) that has high activity toward the reactive intermediate aflatoxin B(1)-8,9-epoxide (AFBO). Rats constitutively express only small amounts of a GST with high AFBO activity (rGSTA5-5) and thus are sensitive to AFB(1)-induced hepatocarcinogenesis, although induction of rGSTA5-5 can confer resistance in rats. In contrast to rodents, constitutively expressed human hepatic alpha-class GSTs have little or no AFBO detoxifying activity. Recently, we found that the nonhuman primate, Macaca fascicularis (Mf), has significant constitutive hepatic GST activity toward AFBO and most of this activity belongs to mu-class GSTs. To determine if any alpha-class GSTs in Mf liver have AFBO activity, a cDNA library from a male Mf liver was constructed and screened using the human alpha-class GstA1 cDNA as a probe. Three different cDNA clones with full-length open reading frames were identified from the Mf hepatic cDNA library. Analyses of the cDNA deduced protein sequences indicated that these three alpha-class cDNA clones were 97-98% homologous with each other, and shared 93, 95, and 95% identity with human GSTA1, and were named mfaGSTA1, mfaGSTA2, and mfaGSTA3, respectively. Bacterially expressed mfaGSTA1-1 recombinant protein had similar activities toward classic GST substrates such as DCNB, CHP, and ECA, but slightly lower CDNB conjugating activity relative to human GSTA1-1. However, similar to hGSTA1-1, mfaGSTA1-1 had no AFBO conjugating activity. In addition, similar to human GSTA1 gene, cDNA-derived amino acid sequence analyses demonstrated that all of these Mf alpha-class GSTs genes (mfaGSTA1, mfaGSTA2, and mfaGSTA3) had none of the six critical residues that were identified previously to confer high AFBO activity in mouse alpha-class GSTA3-3. Thus, in contrast to rodents but similar to humans, alpha-class GSTs from the nonhuman primate, Mf, have little conjugating activity toward AFBO.
- Published
- 2002
- Full Text
- View/download PDF
210. Fundamentals are still relevant in toxicology.
- Author
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Eaton DL and Greenlee WF
- Subjects
- Hazardous Substances toxicity, Humans, Politics, Public Health, Risk Assessment methods, Risk Assessment standards, Toxicology methods, Toxicology standards
- Published
- 2002
- Full Text
- View/download PDF
211. A road map to stable, soluble, easily crystallized pentacene derivatives.
- Author
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Anthony JE, Eaton DL, and Parkin SR
- Abstract
[structure: see text] A series of 6,13-disubstituted pentacenes, in which the substituents are functionalized ethyne units, were synthesized and analyzed by X-ray crystallography. The resulting pentacene derivatives were highly soluble and oxidatively stable and exhibited a significant amount of pi-stacking in the crystal.
- Published
- 2002
- Full Text
- View/download PDF
212. Expression of human microsomal epoxide hydrolase in Saccharomyces cerevisiae reveals a functional role in aflatoxin B1 detoxification.
- Author
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Kelly EJ, Erickson KE, Sengstag C, and Eaton DL
- Subjects
- Aflatoxin B1 metabolism, Cytochrome P-450 CYP1A1 drug effects, Cytochrome P-450 CYP1A1 genetics, Cytochrome P-450 CYP1A1 metabolism, Cytochrome P-450 CYP1A2 drug effects, Cytochrome P-450 CYP1A2 genetics, Cytochrome P-450 CYP1A2 metabolism, DNA Adducts drug effects, Epoxide Hydrolases administration & dosage, Epoxide Hydrolases genetics, Gene Expression Regulation, Enzymologic, Humans, Microsomes enzymology, Mutagenicity Tests, Organisms, Genetically Modified, Plasmids, Saccharomyces cerevisiae enzymology, Aflatoxin B1 toxicity, Epoxide Hydrolases metabolism, Microsomes drug effects, Saccharomyces cerevisiae drug effects
- Abstract
The metabolism and genotoxicity of the carcinogenic mycotoxin, aflatoxin B1 (AFB), was studied in the lower eukaryotic yeast Saccharomyces cerevisiae. Recombinant strains of yeast were engineered to express human cDNAs for CYP1A1, CYP1A2, and microsomal epoxide hydrolase (mEH). Coexpression of mEH with CYP1A1 or CYP1A2 resulted in significant decreases in measurements of AFB genotoxicity. In cells expressing CYP1A2 and mEH, the level of AFB-DNA adducts was decreased by 50% relative to cells expressing CYP1A2 alone. Mitotic recombination, as assayed by gene conversion at the trp5 locus, was diminished by 50% or greater in cells coexpressing mEH and CYP1A2 compared to CYP1A2 alone. The mutagenicity of AFB in the Ames assay was also decreased by approximately 50% when AFB was incubated with microsomes containing CYP1A1 or CYP1A2 and mEH versus CYP1A1 or CYP1A2 alone. The biotransformation of AFB by CYPs is known to involve the generation of a reactive epoxide intermediate, AFB-8,9-epoxide, but previous direct biochemical and kinetic studies have failed to demonstrate any functional role for mEH in AFB detoxification. By reconstructing a metabolic pathway in intact yeast, we have shown, for the first time, that mEH may play a role in mitigating the carcinogenic effects of AFB.
- Published
- 2002
- Full Text
- View/download PDF
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