University of Washington

Center for Ecogenetics and Environmental Health

David L. Eaton, Ph.D.
deaton@u.washington.edu
http://depts.washington.edu/ceeh/

Project Description

The theme of this Center is "Biochemical and Molecular Mechanisms Underlying Human Variability in Response to Environmental Exposures". The interactions between genetics and environment are complex, and defy explanation through traditional disciplinary pathways of investigation. The purpose of this NIEHS Center is to provide an administrative infrastructure and technical support to foster the multidisciplinary collaborations necessary to extend basic mechanistic studies on environmental health problems to direct application in human populations. This center consists of 6 research cores: 1) Gastrointestinal and Renal Toxicology, 2) Carcinogenesis, 3) Reproductive and Developmental Toxicology, 4) Neurotoxicology, 5) Cardiovascular Respiratory Toxicology, and 6) Bioinformatics and Biostatistical Methodologies. Each of these Cores consists of 7 - 10 senior investigators and 2 - 5 associate investigators representing several different departments and programs throughout the University. The funded research of these core faculty is enhanced by 5 Facility cores which provide Center investigators access to: 1) Functional Genomics, 2) Functional Proteomics, 3) Analytical Cytology, techniques such as flow cytometry and fluorescence activated quantitative cytometry, 4) Transgenic Animal Services, support for development and maintenance of transgenic animals of value in toxicological research, and 5) Bioinformatics and Biostatistics, provides guidance for research statistical data. The Pilot Projects support five exploratory research projects into innovative new ideas related to the theme of the Center for one year. A Community Outreach and Education Core provides a mechanism to disseminate important research findings of Center investigators to the general community, as well as a coordinating function to extend and enhance existing community education programs to include more emphasis on issues related to environmental health sciences. The Ethical, Legal and Social Issues (ELSI) Core provides the researchers with the latest information dealing with these issues.

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Program Highlights

The Phytochemical, Sulforaphane, is an Effective Antagonist to the Human Steroid and Xenobiotic Receptor (SXR, aka, PXR, NR1I2)

In collaboration with the laboratory of Dr. Ken Thummel, CEEH Director David Eaton's laboratory discovered that the dietary phytochemical, sulforaphane (SFN) acts as a specific antagonist to an important steroid hormone nuclear receptor, the Steroid and Xenobiotic Receptor (SXR, also called the ‘pregnane X receptor, or PXR). SXR is an important regulator of expression of cytochrome P450 3A4 and several other xenobiotic biotransformation enzymes and transporters. Numerous drugs and non-drug chemicals can act as ligands to SXR, thereby stimulating the transcription of CYP3A4. Induction of CYP3A4 protein and activity that follows can result in dramatic changes in the way the liver and intestine metabolize drugs and environmental chemicals that are substrates for CYP3A4. Eaton and colleagues found that 50 uM SFN was capable of completely eliminating rifampicin-mediated induction of CYP3A4 in human hepatocytes. The IC50 for inhibition of induction was approximately 12 uM. These in vitro studies, including a microarray study, were done with the guidance of the Functional Genomics Core of the CEEH. This concentration is attainable in vivo following administration of pharmacological, and perhaps dietary, doses of SFN. In collaboration with the Molecular Modeling service in the Functional Proteomics Core provided by CEEH faculty member Christophe Verlinde, they completed molecular modeling studies that suggest possible mechanisms for the interaction between SFN and certain amino acids in the ligand binding pocket of human SXR. Additional collaborations with CEEH member Johanna Lampe (FHCRC) are underway to explore whether dietary administration of relatively large amounts of sulforaphane (200 umoles per day, in a broccoli sprout extract) will alter the in vivo clearance of the CYP3A4 probe drug, midazolam. Detailed pharmacokinetic analyses of this interaction are being done in collaboration with CEEH member Danny Shen. Finally, the assistance of the Transgenic Animal Core has been instrumental in us obtaining a 'humanized PXR' transgenic mouse from Dr. Frank Gonzalez at the NCI. These humanized PXR mice provide an ideal mouse model to further study the dose-response, time course, and structure-activity relationship between isothiocyanate analogs of SFN and PXR ligand binding.

Zhou C, Poulton EJ, Grun F, Bammler TK, Blumberg B, Thummel KE, Eaton DL. The dietary isothiocyanate sulforaphane is an antagonist of the human steroid and xenobiotic nuclear receptor. Mol Pharmacol. 2007 Jan;71(1):220-9. PubMed (http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=17028159&query_hl=2&itool=pubmed_docsum)

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Do Genetic Polymorphisms in Glutathione Biosynthesis Genes Alter Susceptibility to Beryllium Disease?

Beryllium sensitization (BeS) can occur in as many as 20% of people occupationally exposed to beryllium (Be). Once a person is sensitized to Be, even low, environmentally relevant exposure to Be may contribute to disease. BeS progresses to chronic beryllium disease (CBD) at a rate of approximately 6% to 8% per year. CBD occurs in as many as 3% of individuals exposed occupationally to beryllium (Be) salts or metals. BeS is characterized by a specific immune response to Be antigen, resulting in BeS individuals being generally identified by a positive peripheral blood Be lymphocyte proliferation test (BeLPT) and by definition do not have the pulmonary pathology or physiological abnormalities seen in CBD. CBD is characterized by a chronic inflammatory response to Be and the formation of noncaseating granulomas. The disease results from a type IV delayed hypersensitivity and cell-mediated immune response to Be that appears to begin as a sensitization to Be and then progresses to CBD, even after the removal from occupational exposure. Because glutathione (GSH) has been reported to be increased in chronic beryllium disease (CBD) and is associated with immune modulation, associations between CBD and gene polymorphisms of the rate-limiting enzyme in GSH synthesis, glutamate cysteine ligase (GCL), were investigated by CEEH faculty Terry Kavanagh and Tim Takaro. Glutamate cysteine ligase consists of a catalytic subunit (GCLC) and modifier subunit (GCLM). With assistance from the CEEH Functional Genomics Laboratory (Farin and Viernes), patients with CBD, beryllium-sensitized subjects (BeS), and beryllium-exposed subjects without CBD were genotyped for the GCLC GAG trinucleotide repeat polymorphism (GCLC TNR), the GCLC-129 single nucleotide polymorphism (SNP), and the GCLM-588 SNP. Results indicate that GCLC TNR genotype 7/7 is negatively associated with CBD (odds ratio [OR] = 0.28, 95% confidence interval [CI] = 0.08–0.95) and the GCLM-588 C/C SNP genotype is associated with CBD susceptibility (OR = 3.07, 95% CI = 1.00–9.37). No differences were noted in the BeS group. This study suggests that GSH modulation may play a role in CBD pathogenesis, but not in sensitization to beryllium.

Bekris LM, Viernes HM, Farin FM, Maier LA, Kavanagh TJ, Takaro TK. Chronic beryllium disease and glutathione biosynthesis genes. J Occup Environ Med. 2006 Jun;48(6):599-606. PubMed (http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&dopt=AbstractPlus&list_uids=16766924&query_hl=5&itool=pubmed_docsum)

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