EMS 2001: A Science Odyssey
March 16 - 21, 2001
San Diego, CA
Meeting Program
David M. DeMarini, Program Chair
Friday, March 16, 2001
8:00am -
10:00am Strategic Planning Committee Meeting
1:00pm -
5:00pm EMS Council Meeting
2:00pm -
6:30pm Short Course:
Overview of Molecular Epidemiology
6:00pm -
9:30pm Student Poster Session, Oral Presentations and Reception
{Sponsored by the Genetic Toxicology Association and AstraZeneca}
6:00pm -
8:30pm (Program to be announced)
8:30pm Presentation of the EMS Education Committee Award and Best Student / New Investigator Poster Award
Saturday, March 17, 2001 Theme: Cancer
7:00am -
8:30am EMS Committee Breakfast Meetings
Students Group (S.J. Gould will be present)
Anchorage Program Committee (First Meeting)
Nominating Committee
IAEMS (7:00am - 10:00am)
7:30am -
12:30pm Short Courses:
Overview of Molecular Epidemiology (continued)
Advanced Methods of 32P-Postlabeling for DNA Adducts
New Developments/Applications of the Comet Assay
Quality Assurance Practices in Government, Industrial, and Pharmaceutical Labs
9:00am -
11:00am New Technologies
9:00am Vitotox: A Bioluminescence Assay for Genetoxocity and Cytotoxicity
Studies
Jorma Lampinen, Thermo Labsystems, Franklin, MA
9:30am DNA Sequencing by the Li-Cor System
Barry W. Glickman, University of Victoria, Victoria, BC, Canada
10:00am Toxicity Profiling Using A High-Throughput Gene Expression Microarray
Platform
Mary Jane Cunningham, Genometrix, Inc., The Woodlands, TX
12:45pm -
2:00pm Plenary I: Keynote Lecture
Stephen J. Gould, Harvard University, Cambridge, MA
2:00pm -
5:00pm Molecular Genetics of Cancer
Chair, James C. Fuscoe, US Environmental Protection Agency, Research Triangle
Park, NC
This symposium will on focus on the genetic events that transform a normal
cell to a highly malignant tumor cell. Work over the past couple of decades
has defined a myriad of cellular changes in the progression of various normal
cell types to cancer cells, resulting in an almost overwhelmingly complicated
molecular picture of this disease. Dr. Douglas Hanahan will provide an overview
on the relatively small number of common molecular, biochemical, and cellular
changes shared by most cancers, and how the complexity of the disease may
eventually be understandable in terms of a few underlying principles. Dr.
Webster Cavanee will then discuss the molecular mechanisms resulting in inactivation
of tumor suppressor genes and the biological impact of these genetic alterations.
Nearly every human tumor has re-activated telomerase gene expression, and
Dr. Jerry Shay will present the latest findings on the role that telomeres
play in the cancer process. Finally, Dr. Peter Vogt will discuss the role
of mutations in growth regulatory genes on the oncogenic cellular phenotype.
2:00pm The Hallmarks of Cancer
Douglas Hanahan, University of California, San Francisco
2:50pm Tumor-Suppressor Genes: Mechanisms and Consequences of Gene Inactivation
Webster K. Cavenee, The Ludwig Institute for Cancer Research, La Jolla, CA
3:30pm Coffee Break
4:00pm Telomeres and Cancer
Jerry W. Shay, Univeristy of Texas Southwest Medical Center, Dallas, TX
4:30pm Oncogenes and Cancer
Peter K. Vogt, The Scripps Research Institute, La Jolla, CA
5:00pm -
7:30pm Poster Session I and Visits with Exhibitors
8:00pm -
10:00pm EMS Awards and Reception
Dinner provided!
{Sponsored by BioReliance, Inc. and Covance Laboratories}
Sunday, March 18, 2001 Theme: Genomics
7:00am -
8:30am EMS Committee Breakfast Meetings
Public Relations & Communications Committee
Finance Committee
Education and Student Members Committee
Future Meetings Committee
8:30am -
11:30am Genomics, Proteomics, Bioinformatics
{Co-sponsored by: Pharmacia and the National Institute of Environmental Health
Sciences}
Chairs, Peter J. Stambrook, University of Cincinnati, OH and William Suk, National
Institute of Environmental Health Sciences, Research Triangle Park, NC
The capacity to analyze gene expression at the transcriptional and protein
levels has accelerated exponentially with the advent of new technologies. Changes
in transcription following environmental or other challenge or as a consequence
of a disease state can be assessed rapidly by hybridization to DNA microarrays
on filters, chips, or slides. The expression of genes that behave in a coordinate
fashion can be identified and clustered for further analysis. Similarly, cellular
responses, many of which may not manifest at the level of transcription, can
be monitored at the level of proteins. Post-translational modifications, altered
protein stability, or changes in sub-cellular localization can be monitored
by protein fractionation methods, e.g., 2D gels, and affected proteins can
be identified and characterized by mass spectrometry. Because data acquisition
can outpace our ability to interpret the data, the field of bioinformatics
has become increasingly important. This symposium will address each of these
issues beginning with applications of transcriptional analysis of the behavior
of gene cohorts, followed by approaches to protein fractionation for proteomics,
identification and analysis of proteins by mass spectrometry, and lastly a
session on the application of bioinformatics towards understanding gene function.
8:30am Microarray, Bioinformatics, and Human Health
David Botstein, Stanford University, CA
9:15am From Protein Fractionation to Proteomics
Peter Mose Larsen, Odense University, Denmark
10:00am Mass Spectrometry: From Genomics to Proteomics
John R. Yates III, The Scripps Research Institute, La Jolla, CA
10:45am Bioinformatics: A Pathway to Gene Function
Edward M. Marcotte, University of California, Los Angeles, CA
11:30am -
1:00pm EMS Annual Business Meeting
Lunch provided!
1:00pm -
4:00pm Toxicogenomics
{Sponsored by Eli Lilly and Company}
Chairs, Marilyn J. Aardema, Procter & Gamble; Richard S. Paules, National
Institute of Environmental Health Sciences; James T. MacGregor, US Food and
Drug Administration
Genomic and proteomic technologies are rapidly evolving analytical tools
that have the potential to revolutionize toxicology. These methods hold the
promise of allowing us to answer questions in toxicology that we have not
been able to answer previously, or answer as definitively as desired. The "promise" of
these methods derives from the fact that almost without exception, gene expression
is altered during toxicity -- as a direct or indirect result of exposure
to a toxicant. Further, it is thought that toxicity is preceded by changes
in gene or protein expression. Thus, a measure of changes in gene or protein
expression might be a sensitive, characteristic endpoint that can be assessed
prior to the ultimate toxicity. Importantly, it is hoped that the patterns
of gene/protein expression, i.e., "fingerprints," that are characteristic
for specific mechanisms of toxicity can be identified and then used to address
questions in the risk-assessment process, such as interspecies extrapolation,
low-dose to high-dose extrapolation, and adaptive versus toxic effects. Ultimately,
toxicogenomic "fingerprints" may allow us to predict the toxicity
of an unknown chemical or drug or predict the response of humans to a chemical/drug
exposure. These progress and the promise of this exciting merging of genomics
with toxicology will be covered by the speakers in this symposium.
1:00pm Gene Expression Profiling in Molecular Toxicology
Mark Egerton, Incyte Genomics, Palo Alto, CA
1:30pm Proteomics in Molecular Toxicology
N. Leigh Anderson, Large-Scale Biology Corporation, Rockville, MD
2:00pm Use of Gene-Expression Profiles to Predict Individual Patient Responses
to the Adverse Effects of Drugs
Spencer Farr, Phase I Molecular Toxicology, Santa Fe, NM
2:30pm Toxicogenomics at NIEHS
Richard S. Paules, National Institute of Environmental Health Sciences, Research
Triangle Park, NC
3:00pm International Life Sciences Institute (ILSI) Genomics Project
Marilyn J. Aardema, Procter & Gamble, Cincinnati, OH
3:30pm Toxicogenomics and Proteomics: Potential Impact on Pharmaceutical
Development and Regulation
James T. MacGregor, US Food and Drug Administration, Rockville, MD
4:00pm -
6:30pm Poster Session II and Visits with Exhibitors
6:30pm -
8:30pm Public Symposium: Agricultural Genomics and the Promise of Improved
Nutrition and Healthcare: Safety, Environmental, and Legal Issues
Chairs, Bhaskar Gollapudi and Timothy D. Landry, Dow Chemical Company, Midland,
MI
The application of genomics and recombinant DNA technology may lead to great
benefits in food and healthcare products with direct and environmental benefits.
Yet there is concern about the threat of biotechnology that is related to
its power, rapid development, and lack of public understanding. This symposium
will review the technology and its potential benefits to provide a common
starting point, and then the approaches to biotechnology safety assessment
will be considered. What are the real or perceived threats? Do we have the
knowledge and tools to make suitable safety assessments? How can science
help us to avert safety and environmental problems? As genomics is integrated
into systems biology, what research is needed to help assure human and environmental
safety? Major issues exist relating to property rights and biodiversity.
Applications will require consideration of bioethics as science moves from
vertical to horizontal gene transfer. The developed world can provide aspects
of this technology to underdeveloped and economically depressed areas. How
should this be done? Regulatory bodies such as FDA have been addressing these
issues since the 1980s with overall success. Given the powerful effects and
rapid development of biotechnology, attention will be given to applying scientific
principles to regulatory decisions that must encompass changing technologies.
6:30pm Symposium Overview
Timothy D. Landry, Dow Chemical Company, Midland, MI
6:40pm Agricultural Biotechnology Explained
Steven P. Briggs, Novartis Agricultural Discovery Institute, San Diego, CA
7:05pm Designer Genes in Your Food: Boon or Bane?
Henry I. Miller, Hoover Institute, Stanford Univ., CA
7:30pm The Ethics of Genetically Modified Food: A View from the Developing
World
Angela A. Wasunna, The Hastings Institute, Garrison, NY
7:55pm Genetically Engineered Organisms: A Regulatory Perspective
John C. Matheson, US Food and Drug Administration, Rockville, MD
8:20pm Summary and Questions
Bhaskar Gollapudi, Dow Chemical Company, Midland, MI
Monday, March 19, 2001 Theme: Animal Models/Ethics
7:00am -
8:30am EMS Interest Group Breakfast Meetings
{Sponsored by Taconic}
Risk Assessment Group
DNA Repair Group
Molecular Epidemiology Group
Exhibitor's Breakfast
8:30am -
11:00am Functional Analysis of the Mammalian Genome: Humans and Mice
Chairs, Susan E. Lewis, Cary, NC; Michael D. Shelby, National Institute of
Environmental Health Sciences, Research Triangle Park, NC
As the structure of the human genome is coming close to being elucidated, the
relevant functions of genes are, of course, of primary interest. The similarity
of structure and function of the mouse genome to that of the human permits
the use of mouse models as tools to understand the function of analogous human
genes. One of the approaches to determining the myriad functions of the mammalian
genome is to analyze gene expression and control in mouse mutants. Two of the
speakers, Drs. Eugene Rinchik and Lisa Stubbs, explore the alterations of function
in mice containing experimentally altered genes to demonstrate aspects of genetic
control. Dr. Muriel Davisson will discuss the uses of mouse models of human
disease as well as the importance of genetic background. Dr. Andrew Wyrobek
will discuss the nature of genetic abnormalities arising in male germ cells
and their transmission to subsequent generations.
8:30am Introduction
Susan E. Lewis, Cary, NC
8:40am Risk Factors and Mechanisms of Paternally Transmitted Abnormalities
Andrew J. Wyrobek, Lawrence Livermore National Laboratory, Livermore, CA
9:10am Discovering New Links Between Genes, Diseases, and Susceptibilities
Through Analysis of Heritable Translocations in Mice
Lisa Stubbs, Lawrence Livermore National Laboratory, Livermore, CA
9:50am Functional Annotation of Mammalian DNA Sequence by Large-Scale, Phenotype-Driven
Recovery of Mouse Mutations
Eugene M. Rinchik, Oak Ridge National Laboratory, Oak Ridge, TN
10:30am Mouse Models and Strain-Background Effects
Muriel Davisson, Jackson Laboratory, Bar Harbor, ME
8:30am -
10:30am Ethical Issues Associated with Gene Therapy
Chair, Errol Zeiger, National Institute of Environmental Health Sciences, Research
Triangle Park, NC
In addition to creating new or modified life forms, the ability to transfer
specific parts of genomes or defined genetic material within and between species
creates a number of new social, ethical, and legal issues. This symposium will
address ethical issues associated with genetic manipulation, in general, and
gene therapy, in particular. Because justification, design, and performance
of the scientific procedures should not be separated from consideration of
the ethical issues engendered by the science, and the ethical issues cannot
be addressed properly without a basic understanding of the scientific procedures
and processes involved, the two speakers will address the scientific and ethical
aspects of gene therapy. The symposium is designed to provide scientists with
an overview of the ethical issues, and nonscientists with insight into the
scientific and medical considerations associated with gene therapy.
8:30am Overview of Intersection of Ethics and Gene Therapy
Errol Zeiger, National Institute of Environmental Health Sciences, Research
Triangle Park, NC
8:40am Gene Therapy: Science, Medicine, Art, and Ethics
Robert Erickson, University of Arizona, Phoenix
9:30am Gene Therapy: Ethical and Legal Issues
Vicki Michel, Los Angeles, CA
10:20am Discussion
10:30am -
10:50am Social and Political Origins of the EMS
Scott Frickel, University of Pennsylvania, Philadelphia
10:50am -
11:15am Coffee Break
11:15am -
12:15pm Plenary II: Genomic Biology
Roger Brent, The Molecular Sciences Institute, Berkeley, CA
12:15pm -
12:45pm Robert H. Haynes Memorial Lecture and Toast DNA Repair: A Global
Perspective
Philip C. Hanawalt, Stanford University, Stanford, CA
12:45pm -
6:30pm Free Afternoon
San Diego Old & New Tour
California Coastline & Tidepool Walk Tour
Coronado Mini-Trek Tour
Old Town – Where San Diego Began Tour
Walking Tour of the Gaslamp District Tour
Other San Diego Attractions
6:30pm -
10:00pm Social Event: Evening at the Fleet Science Center
Dinner provided!
Tuesday, March 20, 2001 Theme: Mechanisms of Mutagenesis and Carcinogenesis
7:00am -
8:30am EMS Committee & Interest Group Breakfast Meetings
Membership and Professional Development Committee
Organization Committee
Germ-Cell Effects/Human Genetics/Aneuploidy Group
New Technologies Group
Awards and Honors Committee
8:30am -
12:00pm Chromosomal Aberrations, Somatic Mutations, and Cancer Risk: Past,
Present, and Future
Chairs, Stefano Bonassi, Istituto Nazionale per la Ricerca sul Cancro, Genoa,
Italy; Nathaniel Rothman, National Cancer Institute, Bethesda, MD
The number of papers addressing topics related to the validation of intermediate
biomarkers of disease has greatly increased in the last few years. The reason
for this renewed interest stems mostly from the presumed value of such biomarkers
to predict a disease before its clinical appearance. The possible use of an
early event as a surrogate endpoint of disease would remarkably enhance the
application of preventive measures in groups and even individuals, not to mention
increase insight into the biological process of disease. Among candidate biomarkers
of early effects that have been considered as intermediate steps of the carcinogenic
process, the amount of evidence collected for somatic cell mutations and chromosomal
aberrations is the most sizeable. Mutations are central to human carcinogenesis,
and the study of these events, naturally occurring or environmentally induced,
is a hot topic not only in cancer research. The use of new molecular techniques
to detect mutations in reporter genes and in cancer genes is likely to improve
insight into the association between mutation frequency and cancer risk. A
great interest has recently converged on the chromosomal aberration test. Findings
from epidemiological studies performed in Europe and in Taiwan have provided
direct support to the hypothesis of a causal role of chromosome damage in the
carcinogenic process. These new data, although limited to a few cohorts, have
provided thorough evidence that groups of subjects with increased frequencies
of chromosomal aberrations should be considered at risk of cancer, and, therefore,
may be likely candidates for preventive measures. The causal role of chromosomal
aberrations in cancer risk has been supported by the availability of new technologies
that have greatly enhanced the sensitivity and applicability of chromosomal
aberration tests. The aim of this symposium is to provide an overview of the
biomarkers on early biological effects that have been linked most plausibly
to cancer risk, emphasizing the role of genetic susceptibility and the impact
of new technologies. The possible connection of these findings to public health
issues will be discussed.
8:30am A Molecular Epidemiologic Perspective: Two Years Later
Nathaniel Rothman, National Cancer Institute, Bethesda, MD
8:45am Past, Present, and Future of Biomarkers Relevant to Cancer
Martyn T. Smith, University of California, Berkeley
9:15am Past, Present, and Especially the Future of Chromosomal Aberration
Assays for Predicting Cancer Risk
James D. Tucker, Lawrence Livermore National Laboratory, Livermore, CA
9:45am Genetic Susceptibility to Induced Chromosomal Aberrations and Cancer
Risk
Qingyi Wei, MD Anderson, Houston, TX
10:15am Coffee
10:30am Are Chromosomal Aberrations an Intermediate End-Point of Cancer?
Stefano Bonassi, Istituto Nazionale per la Ricerca sul Cancro, Genoa, Italy
11:00am Somatic Mutation in Humans
Richard J. Albertini, University of Vermont, Burlington
11:30am Chromosomal Aberrations, Somatic Mutations, and Cancer Risk Assessment:
A Public Health Perspective
Dale Hattis, Clark University, Worcester, MA
8:30am -
12:00pm Cell-Cycle Checkpoints and Signal Transduction
Chairs, Toby Rossman, New York University, Tuxedo, NY; Peter J. Stambrook,
University Cincinnati, OH
The principle task of the cell cycle is to replicate DNA without errors and
to segregate the duplicated DNA equally to the two daughter cells. Maintenance
of genomic integrity in eukaryotic cells is ensured by DNA repair systems coupled
with cell-cycle checkpoints. Cell-cycle checkpoints are highly conserved mechanisms
that allow cells to sense DNA damage and to respond by arresting cell-cycle
progression to allow DNA repair. Probably the most familiar is the P53-dependent
G1/S checkpoint that arrests cells for DNA repair prior to DNA replication
to reduce mutagenic events as a consequence of DNA damage. However, other cell-cycle
checkpoints are equally important for maintenance of genomic integrity. This
symposium will provide an overview of checkpoint mechanisms as well as examples
of checkpoints in S phase, G2/mitosis, and a mitotic spindle checkpoint.
8:30am Cell Cycle Overview: Historical Perspectives
Steven Reed, The Scripps Research Institute, La Jolla, CA
9:20am RB-Mediated G1/S Checkpoint Regulation
Erik Knudsen, University of Cincinnati, OH
10:00am Coffee
10:30am S-Phase Checkpoint
Anthony M. Carr, University Sussex, England
11:00am G2/M Checkpoint
Clare McGowan, The Scripps Research Institute, La Jolla, CA
11:30am Mitotic Checkpoints in Animal Cells
Peter Sorger, Massachusetts Institute of Technology, Cambridge, MA
12:00pm -
1:00pm Lunch provided!
{Sponsored by Covance Laboratories}
1:00pm -
3:30pm Poster Session III and Visits with Exhibitors
3:30pm -
6:30pm The Annual Elsevier Mutation Research Symposium: Translesion Synthesis
and SOS Response: How Lesions are Misread
{Sponsored by Elsevier Publishers and the National Institute of Environmental
Health Sciences}
Chair, Gerald P. Holmquist, City of Hope, Duarte, CA
Induced mutations are usually generated by DNA replication past DNA lesions.
Historically, this step in mutagenesis has been the least understood step.
Half of the known DNA polymerases have been discovered in just the last few
years, and most of these are specialized to carry out translesion DNA synthesis.
Without them, replication forks become blocked at cytotoxic lesions. In fact,
the SOS response in E. coli and concomitant upregulation of a bypass polymerase
is initiated by a replication fork stalled at a cytotoxic lesion. In yeast,
deletion of bypass polymerases act as anti-mutators. Presumably, the cells
die in an arrested S-phase rather than survive using replication bypass that
entails misreading of cytotoxic lesions. These translesion DNA polymerases
have a low fidelity for reading normal bases. They also have low processivity;
they fall off quickly so that the high-fidelity replicative polymerases can
take over again. Exactly what bases (A-rule, etc.) are substituted during
bypass of a particular lesion may be determined by the translesion polymerase
responsible for the bypass.
3:30pm Introduction
Gerald P. Holmquist, City of Hope, Duarte, CA
3:40pm DNA Polymerase kappa: A Novel DNA Polymerase in Search of a Function
Errol C. Friedberg, University of Texas Southwest Medical Center, Dallas,
TX
4:20pm Translesion DNA Synthesis: A Conserved Mechanism from Bacteria to
Humans
Roger Woodgate, National Institutes of Health, Bethesda, MD
5:00pm Tea Break
{Sponsored by The Stash Tea Company}
5:30pm The Biochemical Basis of SOS Lesion-Targeted Mutagenesis Involving
Errant DNA Polymerases
Myron F. Goodman, University of Southern California, Los Angeles, CA
6:00pm Translesion Synthesis by RNA Polymerases: Transcriptional Mutagenesis
in Bacterial and Mammalian Cells
Paul W. Doetsch, Emory University School of Medicine, Atlanta, GA
3:30pm -
6:30pm Antimutagenesis/Chemoprevention
{Sponsored by The Stash Tea Company}
Chairs, Rod Dashwood, Oregon State University, Corvallis; Lynnette Ferguson,
University of Auckland, New Zealand
Certain groups of the population have a high risk of cancer, through either
genetic or environmental factors. The possibility that individuals in these
groups could be protected against cancer is enticing. Theoretically at least,
the study of antimutagenesis provides the mechanistic understanding of how
to interfere at various steps in the process. Certain population groups in
China are at high risk of hepatocarcinogenesis through high exposure to aflatoxin.
An enhancement in the activity of detoxifying enzymes such as glutathione S-transferase
(GST) might be predicted to enhance aflatoxin excretion and decrease the risk
to the populations concerned. Oltipraz is a potential chemopreventive agent
designed exactly for this activity. Tom Kensler will provide insights from
recent clinical trials with oltipraz in high-risk Chinese populations, whereas
Fred Kadlubar will speculate on the relative roles of nature vs. nurture in
GST activity. Despite high risks of some cancer types, Chinese, and particularly
Japanese, populations appear to be protected against some forms of cancer that
are prevalent in Western populations. Although there may be a genetic component,
a high intake of green tea has been suggested as a protective factor. Rod Dashwood
will consider mechanistic studies on this effect. Finally, the Polynesian population
in New Zealand appears to provide an interesting contradiction in the epidemiology
of diet and cancer. Eating higher fat, higher calories, and drinking larger
amounts of alcohol in any session are all generally recognised risk factors
for colon cancer. Despite following these high risk diets, Polynesian groups
have approximately one-third the incidence of colorectal cancer as compared
with European populations. Lynn Ferguson will review animal and in vitro studies
that may help interpret whether genotype or diet is the causal factor.
3:30pm Strategies for Protection Against Aflatoxin-Induced Hepatocarcinogenesis:
Insights from Clinical Trials in China
Thomas W. Kensler, Johns Hopkins University, Baltimore, MD
4:15pm Glutathione S-Transferase: Nature versus Nurture
Fred F. Kadlubar, National Center for Toxicological Research, Jefferson,
AR
5:00pm Tea Break
{Sponsored by The Stash Tea Company}
5:30pm Antimutagenesis/Chemoprevention Mechanisms of Tea
Roderick H. Dashwood, Oregon State University, Corvallis
6:00pm Contribution of Animal Studies to Understanding Gene-Diet Interactions
Lynnette R. Ferguson, University of Auckland, New Zealand
6:30pm -
7:30pm Plenary III: Mutability of Repeated DNA Sequences in Humans and Mice
Sir Alec Jeffreys, University of Leicester, Leicester, England
Wednesday, March 21, 2001 Theme: DNA Repair
7:00am -
8:30am EMS Committee & Interest Group Breakfast Meetings
Transgenics Group
Anchorage Program Committee (Second Meeting)
Hollaender Committee
EMS Executive Board Meeting
8:30am -
1:00pm Genotoxic Risks of Perinatal Exposures to Antiretroviral Reverse Transcriptase
Inhibitors (NRTIs)
Chairs, Miriam Poirier, National Cancer Institute, Bethesda, MD; Vernon Walker,
New York State Department of Health, Albany
With the success of the Highly Active Antiretroviral Therapy (HAART) protocols,
each year in the US alone ~750,000 HIV-1-infected individuals, including ~7,000
HIV-1-infected pregnant women, are treated with antiretroviral drugs. These
combinations typically include "dideoxy- type" nucleoside analog
reverse-transcriptase inhibitors (NRTIs), such as zidovudine, lamivudine, didanosine,
and stavudine. Drugs of this class have antiretroviral activity via drug phosphorylation,
incorporation into viral DNA, DNA chain termination, and inhibition of viral
reverse-transcription enzymes. However, the toxic effects of these drugs are
known to occur through both nuclear and mitochondrial mechanisms, and drug-DNA
incorporation appears to be fundamental to both nuclear and mitochondrial manifestations
of toxicity. Nuclear genotoxicity includes drug-DNA incorporation, telomere
shortening, clastogenicity, and mutagenesis. The weak tumorigenicity observed
in adult rodents and the stronger mouse transplacental tumorigenicity that
occurs after zidovudine exposure are likely the result of genotoxicity. However,
in humans there is no epidemiological evidence for zidovudine carcinogenicity.
Mitochondrial toxicity presents clinically as cardiac and/or skeletal muscle
myopathy with occasional lactic acidosis and hepatic steatosis, and it is seen
frequently in adult patients given long-term NRTI treatment. The clinical,
biochemical, and molecular manifestations are similar to those observed in
classic mitochondrial diseases caused by mitochondrial mutations and deletions.
Persistent mitochondrial dysfunction after human transplacental NRTI exposure
has been reported recently in France in a small number of HIV-1-uninfected
children. Two of these children died at ~1 year of age after exposure in utero
and for about a month after birth to zidovudine and lamivudine. Similar effects
have not been observed in American children. Nevertheless, as combination HIV-1
therapies become more complex and effective in preventing maternal-fetal transmission,
the potential risks for long-term health effects from in utero NRTI exposures
need to be fully explored.
8:30am Overview of Antiretroviral Nucleoside Analogs: Clinical Use and Toxicities
Vernon Walker, New York State Department of Health, Albany
8:40am Transplacental Carcinogenicity of AZT in Rodents
Dale Walker, New York State Department of Health, Albany
9:00am Observational Cohort Study of the Genotoxic Risks Arising from Perinatal
NRTI Therapies
Vernon Walker, New York State Department of Health, Albany
9:20am Plasma and Cellular Markers of AZT Metabolism as Indicators of DNA
Incorporation of AZT in Infants Exposed in utero to Combination Antiretroviral
Therapies
Quanxin Meng, New York State Department of Health, Albany
9:40am Pharmacogenomics of AZT
Ainsley Weston, National Institute of Occupational Safety and Health, Morgantown,
WV
10:00am Coffee
10:20am Glycophorin A (GPA) Locus Somatic Mutations in Umbilical Cord Blood
Erythroid Cells of Infants Exposed in utero to NRTIs
William Bigbee, University of Pittsburgh, PA
10:40am HPRT Mutant Frequency and Mutation Spectrum in Newborns with Perinatal
Exposure to Antiretroviral Drugs
J. Patrick O'Neil, University of Vermont, Burlington
11:00am Discussion
11:20am DNA Adduct Formation and Mutation Induction in Neonatal Mice Treated
with Antiretroviral Nucleoside Analog Drugs
Frederick A. Beland, National Center for Toxicological Research, Jefferson,
AR
11:40am Incorporation of NRTIs into Fetal DNA Following Transplacental Drug
Exposures in Humans and Experimental Animal Models
Ofelia Olivero, National Cancer Institute, Bethesda, MD
12:00pm Animal Models of NRTI Mitochondrial Toxicity
Miriam Poirier, National Cancer Institute, Bethesda, MD
12:20pm Mitochondrial Dysfunction in Infants Exposed in utero to NRTIs
Stephane Blanche, Hospital Necker, Paris, France
12:40pm Discussion
8:30am -
10:30am DNA Repair and Genomic Instability
{Sponsored by the National Institute of Environmental Health Sciences}
Chair, Larry H. Thompson, Lawrence Livermore National Laboratory, Livermore,
CA
The goal of this symposium is to provide a broad sampling of the molecular
processes that maintain genomic integrity and, thereby, suppress mutagenesis
and carcinogenesis. Rapid progress in determining the molecular details of
multiple DNA repair and checkpoint pathways has come from biochemical and structural
studies with purified proteins, cell culture systems, and mouse models, which
are presented in this symposium. DNA repair pathways are regulated in the context
of the cell cycle by checkpoint functions that monitor the structural integrity
of chromosomal DNA molecules. Repair pathways consist of nucleotide excision
repair (bulky, helix-distorting base adducts), base excision repair (oxidative
and simple alkylation damage), mismatch repair (mismatched and looped-out bases),
Ku/DNA-PKcs-dependent nonhomologous end joining of double-strand breaks (DSBs),
and homologous recombinational repair of DSBs and interstrand crosslinks. Insights
into how the Ku/DNA-PKcs complex interacts with the ends of DNA molecules and
is activated will be presented. Checkpoint functions acting in response to
damage are initiated by poorly understood mechanisms and mediated by phosphorylation
reactions involving complex kinase cascades that lead to inhibition of cyclin-dependent
kinases, thereby delaying cell-cycle progression and allowing extra time for
repair. Specifically, the ATM kinase, which is defective in radiation-sensitive
ataxia telangiectasia, acts at an early step in the signaling of damage by
phosphorylating multiple substrates, i.e., Tp53, BRCA1, and the checkpoint
kinases Chk1 and Chk2/Cds1. The construction and analysis of numerous mouse
strains carrying single or multiple gene knockouts has revealed interactions
of genes within and between pathways, overlapping pathways for certain types
of lesions, and a central role of Tp53 in promoting genomic stability.
8:30am P53, Oncogene Activation, Genetic Instability, and Tumor Progression
Geoff Wahl, The Salk Institute, La Jolla, CA
9:00am Cancer Predisposition in Mutant Mice Defective in DNA Repair Genes
Lisiane Meira, University of Texas Southwest Medical Center, Dallas, TX
9:30am Coffee
10:00am ATM and Other Kinases in DNA-Damage Signaling Pathways
Eva Lee, University of Texas, San Antonio, TX
10:30am Significance Analysis of Microarrays Applied to the Ionizing Radiation
Response
Gilbert Chu, Stanford University, Stanford, CA
11:00am -
1:00pm Complexity in Genome Stability: Structure and Function of DNA Repair
Complexes
{Sponsored by the National Institute of Environmental Health Sciences}
Chair, John A. Tainer, The Scripps Research Institute, La Jolla, CA
DNA-based information is subject to damage and consequent mutation resulting
from the environment both internal and external to the organism. The balance
reached between the maintenance of genomic integrity by DNA repair and the
evolutionary value of uncorrected mutations for adaptation underlies the biological
organization and efficiency of DNA repair systems. The existence of this dynamic
between optimal DNA repair levels and DNA mutations reveals the importance
of environmental agents and underscores how changes in environmental mutagens
may impact profoundly the maintenance or loss of genetic integrity depending
upon both the types of damage and the nature of DNA repair systems. Fortunately,
the integration of current genetic, biochemical , and structural research on
such DNA repair systems provides the basis for interpreting the complex network
of biologically important interactions among DNA repair proteins required for
their function in maintaining genomic integrity. These results emphasize that
structurally encoded pathways, not individual enzymes, are optimized by evolutionary
selection and determine the current biology of cells and organisms. Thus, a
complete understanding of the biological and medical implication of genetic
polymorphisms will require structural information not only about individual
proteins but also about their interfaces and conformational changes, which
can influence biologically critical connections between repair steps and pathways.
These DNA repair connections are manifested both in transient interactions
that handoff damaged DNA between enzymes and in multi-protein complexes, which
provide high specificity plus tight coupling. This combination of interaction
specificity (without release of DNA repair intermediates) and variability (with
multiple possible DNA repair protein partners) evidently generates the structurally
encoded coordination and regulation of biologically critical connections between
individual DNA repair steps and pathways. The talks in this session will highlight
not only new results on interactions emerging for the function of four major
DNA repair pathways, but, furthermore, provide a useful and testable conceptual
framework for considering common and variable features of DNA repair processes
and for approaching a unified understanding of the structural biology of DNA
repair.
11:00am Complexes Acting in DNA Double-Strand Break Repair
John A. Tainer, The Scripps Research Institute, La Jolla, CA
11:30am Mismatch Repair Protein Interactions in Yeast
Richard Kolodner, University of California, San Diego
12:00pm Recombinational Repair of DNA Breaks
Steve Kowalczykowski, University of California, Davis, CA
12:30pm Pathway Interactions in Transcription-Coupled Repair Directed by
XPG Complexes
Priscilla Cooper, Lawrence Berkeley Laboratory, Berkeley, CA
1:00pm -
5:00pm EMS Council Meeting
EMS 2001 Short Courses
2:00pm -
6:30pm Friday, March 16, 2001
Overview of Molecular Epidemiology
The rapidly increasing number of scientific papers that integrate laboratory
analyses and epidemiologic design is a clear sign of the growing interest
in the young discipline of molecular epidemiology. The simultaneous involvement
of researchers from the different fields has accelerated this momentum and
represents a paradigm shift in medical research, revealing that the multidisciplinary
approach is going to be the standard model for the research in the new century.
Molecular epidemiology integrates the efforts of epidemiologists, molecular
biologists, geneticists, and researchers from many other fields with the
purpose of understanding of the causes and underlying biological mechanisms
of chronic diseases. The ability to disentangle the role of host and genetic
facts that may account for individual susceptibility is a key issue, as well
as the most challenging goal of molecular epidemiology, which is the identification
of individual risk of disease in health people. As a direct consequence of
the interaction between different research areas, the need for a common language
is a fundamental goal that must be achieved to improve the collaboration
between epidemiologists and biologists. Epidemiologists began learning biology,
and laboratory scientists started worrying about bias and confounding. This
reciprocal interest has generated a great demand for education and training
to permit researchers entering this new field to interact quickly with researchers
from other areas. The aim of this Overview of Molecular Epidemiology is to
bring together researchers with different skills, teach them the main concepts
and the basic methods of molecular epidemiology, and invite them to talk
to each other using the same language. This will contribute to the creation
of a new generation of researchers who will be able to apply the power of
epidemiologic and statistical methods to the sensitivity of data coming from
the laboratory. The course is structured in two, 1/2-day sessions, for a
total of 14 lectures, most of them 30-min long. Participants will be introduced
to a molecular epidemiologic view of human studies. Lessons on the epidemiological
design and the statistical analysis of data will be alternated with the description
of biomarkers used most commonly in human biomonitoring, relevant laboratory
methods, and their biological meaning. Leading scientists in the field have
been recruited with the purpose of providing a comprehensive state-of-the-art
overview in the limited time available.
2:00pm Introduction
James D. Tucker, Ph.D., Lawrence Livermore National Laboratory, Livermore,
CA
2:10pm Course Overview
Stefano Bonassi, Ph.D., Istituto Nazionale per la Ricerca sul Cancro, Genoa,
Italy
Methodological Issues
2:30pm Study Design in Molecular Epidemiology
Monsterrat Garcia-Closas, M.D., National Insitutes of Health / National Cancer
Institute, Bethesda, MD
3:15pm Ethical Considerations in Human Studies
Errol Zeiger, Ph.D., Esq., National Institute of Environmental Health Sciences,
Research Triangle Park, NC
3:45pm Biological Sample Collection and Processing
Nina T. Holland, Ph.D., University of California, Berkeley, CA
4:15pm Coffee Break
Biomarkers of Exposure and Early Effect
4:45am Using Biomarkers in Exposure Assessment
Stephen Rappaport, Ph.D., University of North Carolina, Chapel Hill
5:15pm From DNA Damage to Mutation
David M. DeMarini, Ph.D., US Environmental Protection Agency, Research Triangle
Park, NC
5:45pm Somatic Cell Mutations in Cancer Research
Richard J. Albertini, M.D., University of Vermont, Burlington
6:15pm Discussion
7:30am -
12:30pm Saturday, March 17, 2001
Overview of Molecular Epidemiology (continued)
Genetic Susceptibility
7:30am Breakfast
8:00am Cytogenetic Biomarkers
James D. Tucker, Ph.D., Lawrence Livermore National Laboratory, Livermore,
CA
8:30am New Genomic Technologies
Russell Higuchi, Ph.D., Roche Molecular Systems, Alameda, CA
9:00am Genotype vs. Phenotype in Molecular Epidemiology Studies
Frederick F. Kadlubar, Ph.D., National Center for Toxicological Research,
Jefferson, AR
Analysis of Data
9:30am Bias, Confounding, and Effect Modification in Molecular Epidemiology
Studies
Montserrat Garcia-Closas, M.D., National Institutes of Health / National
Cancer Institute, Bethesda, MD
10:00am Coffee Break
10:30am Statistical Analysis of Molecular Epidemiology Studies
Stefano Bonassi, Ph.D., Istituto Nazionale per la Ricerca sul Cancro, Genoa,
Italy
Case Studies
11:00am Studies of Workers Exposed to Benzene and Butadiene
Martyn T. Smith, Ph.D., University of California, Berkeley
11:30am Molecular Epidemiology of Lung Disease
Stephanie J. London, M.D., Dr.P.H., National Institute of Environmental Health
Sciences, Research Triangle Park, NC
12:00pm Discussion
7:30am -
12:30pm Saturday, March 17, 2001
Advanced Methods of 32P-Postlabeling for DNA Adducts
Instructors, Jeffrey Ross and Leon King, US Environmental Protection Agency,
Research Triangle Park, NC
This course will present practical advice for performing, optimizing, and interpreting
results from 32P-postlabeling studies on DNA adducts. Topics to be discussed
include methods for DNA isolation and characterization for postlabeling, optimum
conditions for DNA hydrolysis and postlabeling, alternative methods for adduct
enrichment and resolution of adducts, use of storage phosphor screens for adduct
quantitation, and approaches for troubleshooting the postlabeling process.
This course is designed primarily for those who possess a basic understanding
of the mechanisms of DNA adduct formation. Specific examples from both the
literature and the instructors’ labs will be used to illustrate the influence
of changes in postlabeling parameters on results obtained and provide a framework
for developing new chromatographic procedures to optimize resolution of adducts
that are not resolved readily by standard approaches.
7:30am Breakfast
8:00am Introduction to 32P-Postlabeling: Brief Historical Background and Safety Considerations
8:30am DNA Isolation and Hydrolysis: Extraction Techniques, Criteria for Purity, and Hydrolysis Conditions
9:00am Adduct Enrichment: Butanol Extraction, P1 Disgestion, and Immunoaffinity
9:30am Labeling Conditions: Effect of Phosphatase Activity; Effect of Labeling-Reaction Conditions
10:00am Coffee Break
10:30am HPLC Methods: HPLC and Detector Selection, HPLC Separations, Starting Points, and Optimization
11:00am TLC Methods: Equipment, TLC Separations, Starting Points, Optimization
11:30am Quantitation: X-ray Film vs. Direct-Radioactivity Measurement; Storage-Phosphor Technology; Background Correction; Specific Activity vs. Normal Nucleotides; Data Management
12:00pm Adduct Identification: Co-Chromatography Issues; Preparation of Standards
7:30am -
12:30pm Saturday, March 17, 2001
New Developments and Applications of the Comet Assay
{Sponsored by Integrated Laboratory Systems}
Techniques that permit the sensitive detection of DNA damage and its repair
are critically important in fields of toxicology ranging from aging and clinical
investigations to genetic toxicology and molecular epidemiology. Because the
DNA damage induced by toxic agents is often tissue- and cell-type specific,
an optimal technique would be one that can detect DNA damage and repair in
individual cells obtained under a variety of experimental conditions. In 1988,
Singh and co-workers (Exp. Cell Res. 175:184-191) introduced a microgel technique
involving electrophoresis under alkaline conditions (pH >13) that was capable
of detecting single-strand breaks (SSB) and alkali-labile lesions in the DNA
of individual cells. Since the introduction of the alkaline single-cell gel
(SCG) or Comet assay, a number of advancements have taken place that greatly
increase the flexibility and utility of this technique for detecting various
manifestations of DNA damage (e.g., single- and double-strand breaks, oxidative-induced
base damage, and DNA-DNA/DNA-protein crosslinking) and DNA repair in virtually
any eukaryotic cell. During this course, the range of topics to be covered
include applications in genetic toxicology (both in vitro and in vivo, as well
as standardized guidelines), environmental and human biomonitoring, and cancer
research.
7:30am Breakfast
8:00am Introduction and General Technical Aspects
Raymond R. Tice, Integrated Laboratory Systems, Research Triangle Park, NC
8:30am Genetic Toxicology: General Guidelines
Eva Agurell, AB Astra, Södertälje, Sweden
9:00am Genetic Toxicology: in vitro Applications and High-Throughput Screening
Andreas Hartmann, Novartis Pharma AG, Basel, Switzerland
9:30am Genetic Toxicology: in vivo Studies in Experimental Animals
Yü F. Sasaki, Hachinohe National College of Technology, Hachinohe, Japan
10:00am Coffee Break
10:30am Environmental Biomonitoring: Aquatic Systems
Scott Steinert, Computer Sciences Corporation, San Diego, CA
11:00am Human Biomonitoring: Environmental Pollutants
Emilio Rojas del Castillo, Institute Investigaciones Biomedicas Universidad
Nacional Autonoma de Mexico, Mexico
11:30am Use of the Comet Assay in Cancer Researcu
Peggy Olive, British Columbia Cancer Research Centre, Vancouver, BC, Canada
12:30pm Discussion
7:30am -
12:30pm Saturday, March 17, 2001
Quality Assurance (QA) Practices in Government, Industrial, and Pharmaceutical
Laboratories
{Sponsored by Dow Corning}
The EMS quality assurance (QA) course is aimed at lab associates and new principal
investigators. The course will review and provide examples of quality systems
in government (US EPA), industrial (CIIT, Dow Corning), and pharmaceutical
(GlaxoWellcome) laboratories. Tom Hughes will provide a comprehensive overview
will be provided of the QA practices at the US EPA in RTP, NC, including a
description of QC, peer-review, and QA activities. Descriptions will also be
given of the Agency’s QA Project Plans (QAPPs), Intramural Research Protocols
(IRPs), Technical Systems Reviews (TSRs), and the QA classification of NHEERL/EPA
studies (Category 1-4). Brenda Culpepper will then discuss methods that are
used at the US EPA to classify and archive different types of records (hard
copy, data files, correspondence, notebooks). She will provide a detailed discussion
of master files, storage of records, time periods for holding different types
of files, and how good laboratory record-keeping will insure that records are
reproducible, defensible, and analyzable. Jim Allen will explain implementation
of current regulations, guidelines, and QA standards as they apply to the use
of animals in toxicology research and testing. Emphasis is given to rodents
and will include special considerations related to transgenic and knockout
animal models. Institutional Animal Care and Use Committee (IACUC) functions
and requirements in research protocol review will also be discussed, and educational
and training resources for humane care and use of laboratory animals will be
identified. The three remaining talks will describe industrial and pharmaceutical
QA practices. Patricia Pomerleau will provide an overview of Good Laboratory
Practice (GLP) Regulations. She will explain the many facets and responsibilities
of the study director and staff under a GLP study. A description will be provided
of methods that are unique to a GLP study, as well as details of how to correctly
conduct and manage GLP notebooks and records. Thomas Barfknecht will then discuss
the management of GLP studies at Contract Research Organizations (CROs). Methods
for the placement of studies at CROs will be discussed, which will include
the identification, selection, qualification/re-qualification, and the reasons
for placement of a particular study at a CRO. In the final presentation, John
Haw will present the QA practices for computer technology in the pharmaceutical
industry. He will discuss the techniques to validate computer software and
programs, which will include records management techniques for computer files.
7:30am Breakfast
8:00am Relationships Among QC, Peer Review, and AQ Procedures at the US
EPA
Thomas Hughes, US Environmental Protection Agency, Research Triangle Park,
NC
8:40am QA and Data Management Techniques
Brenda T. Culpepper, US Environmental Protection Agency, Research Triangle
Park, NC
9:20am Regulations and Guidelines for Using Animals in Toxicology Research
James W. Allen, US Environmental Protection Agency, Research Triangle Park,
NC
10:00am Coffee Break
10:20am Overview of GLP Regulations
Patricia Pomerleau, Chemical Industry Institute of Technology, Research Triangle
Park, NC
11:00am Management of GLP Studies at Contract Research Organizations (CROs)
Thomas R. Barfknecht, Dow Corning, Midland, MI
11:40am QA of Computer Technology in the Pharmaceutical Industry
John Haw, GlaxoWellcome, Research Triangle Park, NC
12:20pm Questions
