Reproductive Toxicity Testing: Data Interpretation and Risk Assessment
Rochelle W. Tyl, RTI International, Research Triangle Park, NC

AM04 - Predictive Power of Novel Technologies (Cells to 'Omics'): Promises, Pitfalls and Potential Applications

BASIC
Chairpersons: Srikanth S. Nadadur, USEPA, Research Triangle Park, NC and Mary Jane Cunningham, Houston Advanced Research Center, The Woodlands, TX

Endorsements
*Inhalation Specialty Section
Molecular Biology Specialty Section
Toxicologic & Exploratory Pathology Specialty Section

Abstract Body: Advances in the disciplines of cell and molecular biology have led to the development of novel biotechnologies capable of generating “global molecular profiles” for in situ toxicological assessment. These technologies should accelerate our understanding of molecular basis for potential toxicity and susceptibility. This course will provide both theoretical and practical information on these emerging high throughput technologies, their applicability, interpretation and integration to gain a more comprehensive understanding of cellular responses to chemical/toxicant stress. The first presentation will cover the potential utility of Laser Capture Micro-Dissection in isolating specific cell populations for toxicological assessment at the level of RNA and proteins. The second presentation will provide the ongoing evolution of gene expression profiling approaches from “toxicogenomics” to systems biology. The third presentation will provide an overview of proteomic technologies in analyzing the protein interactions and downstream signaling mediators involved in toxic response pathways. The final presentation will examine the capabilities of high throughput technologies in identifying the single nucleotide polymorphisms (SNPs) and their predictive value for characterizing underlying genetic susceptibilities to toxicants. Overall the course is aimed at educating toxicologists on the array of new technologies available to research toxic outcomes and their underlying mechanisms. The goal is to move towards a better and reliable prediction and extrapolation of toxic responses. This course will also serve as a refresher course for scientists, technical and regulatory staff involved in various stages of compound development or regulation.

Integrating Transgenic Animal Models and Laser Capture Microdissection Applications with 'Micro-omic'-Based Analyses For In Vivo Toxicological Assessments
Kevin L. Dreher, NHEERL, USEPA, Research Triangle Park, NC

Gene Expression Profiling for Toxicity Assessment using Systems Biology
Mary Jane Cunningham, Houston Advanced Research Center, The Woodlands, TX

Clinical Proteomics: Mapping Molecular Networks in Clinical Specimens using Reverse Phase Protein Microarrays
Emanuel F. Petricoin, George Mason University, Manasas, VA

Common DNA Variation in Human Disease and Drug Response
Steven Hamilton, University of California, San Francisco, San Francisco, CA

AM05 - Comparative Endocrine Toxicology

BASIC
Chairpersons: Stephen B. Hooser, Purdue University, West Lafayette, IN and Charles C. Capen, The Ohio State University, Columbus, OH

Endorsements
*Comparative and Veterinary Specialty Section
Reproductive and Developmental Toxicology Specialty Section
Toxicologic & Exploratory Pathology Specialty Section

Hormones secreted by cells of the endocrine system have diverse effects throughout the body. Exposure to xenobiotic compounds can result in profound changes to the endocrine organs and/or their target cells. Significant species differences exist in the structure and function of endocrine and reproductive organs making interpretation of test results and extrapolation from animal models to humans more challenging. In addition to the numerous anatomical and physiological differences, there are also species variations in metabolism and response to toxicants. It is the goal of these presentations to give an overview of the structure, function, regulation, and toxic responses of selected endocrine and reproductive organs. In addition, the speakers will discuss the hormonal assays and other mechanistic approaches necessary to make species comparisons, and to extrapolate the findings from animals to humans. Each presentation will briefly describe important species differences with regard to anatomy, endocrine physiology, and response to different classes of xenobiotic chemicals by selected endocrine and reproductive organs such as the thyroid (follicular cells), ovary, and testis (Leydig cells). One presentation will focus on the principles and pitfalls of hormonal measurements in laboratory animals considering advantages/disadvantages of different methods, species specificity of certain assays, most appropriate sampling times, and other useful items to consider in future protocol development. Following completion of this workshop, attendees should have a more complete understanding of the comparative endocrinology and toxicology of selected endocrine and reproductive organs in laboratory animals.

Comparative Endocrine Toxicology of the Thyroid Gland
Charles C. Capen, The Ohio State University, Columbus, OH

Comparative Ovarian Toxicology
Jodi Flaws, University of Maryland, Baltimore, MD

Comparative Endocrinology and Toxicology of Testicular Leydig Cells
Jon C. Cook, Pfizer, Inc., Groton, CT

Principles and Pitfalls of Hormone Measurements in Toxicology Studies
Terry M. Nett, Colorado State University, Fort Collins, CO

AM06 - Essentials of Metal Toxicology

BASIC
Chairpersons: Wei Zheng, Purdue University, West Lafayette, IN and Michael P. Waalkes, NIEHS, Research Triangle Park, NC

Endorsements
Mechanisms Specialty Section
*Metals Specialty Section
Neurotoxicology Specialty Section
Risk Assessment Specialty Section

Metals are ubiquitously present in environment, essential to human health, and yet toxic upon overexposure. They are neither synthesized by living matter nor destroyed by human endeavour. Metals play a significant role in many human diseases. Unique physical, chemical and biological properties allow metals to persist in the body, cause long-lasting effects, and yet leave few choices for therapeutic intervention. A fundamental question remains, namely what makes metals toxic in promoting carcinogenesis, neurotoxicity, reproductive, liver and kidney injuries, or generalized metabolic disease? This basic course is intended to address the essentials in metal toxicology by providing cutting-edge knowledge on the concepts, theories, clinical outcome, and research methodologies in metal toxicology. The first lecture will highlight the clinical symptoms of metal-induced human diseases, common routes of exposure, and therapeutic intervention. Two subsequent lectures will address metal pharmacokinetics and the general mechanisms of metal-induced cell death, specifically apoptosis. The final lecture will discuss animal models, based on either symptomatic outcomes or biochemical consequences, for studying metal toxicities. Speakers will address these essentials by providing details specific to “hot” metals, such as arsenic, mercury, manganese, lead, and uranium. The course will serve as an introduction to those who desire an expanded knowledge on essentials of metals in human health and diseases, essentials of toxicological actions of metals, and essentials of research approaches in metal toxicological investigation. The course will also be of interest to others engaged in wider aspects of carcinogenesis, neurotoxicology, risk assessment, and occupational health.

Metals in Human Diseases: Symptoms and Clinical Intervention
Howard Hu, Harvard University, Boston, MA

Distribution of Metals: Role of Metal Transporters and Selectivity of Disposition
Michael Aschner, Vanderbilt University, Nashville, TN

Molecular Mechanism of Metal Toxicity: Signal Transduction and Oxidative Stress
Anumantha Kanthasamy, Iowa State University, Ames, IA

Animal Models Used in Metal Toxicological Research
Tomas Guilarte, Johns Hopkins University, Baltimore, MD

AM07 - Practical Strategies for Evaluation of Immunosuppression in Pharmaceutical Development

BASIC
Chairperson: Brian G. Short, Allergan, Inc, Irvine, CA

Endorsements
Immunotoxicology
Regulatory and Safety Evaluation
*Toxicologic & Exploratory Pathology

Increased focus has been placed on evaluating immunotoxicity, particularly immunosuppression, in pharmaceutical development of small molecules and biologics following recent issuance of regulatory guidelines from FDA and CPMP and current ICH discussions on harmonization. Toxicologists, pathologists, immunologists and clinicians in the drug industry are faced with the challenge of addressing the intent of the current guidance documents in a scientifically valid and responsible manner. An integrated approach is necessary to provide a weight of evidence for signal detection in routine toxicity studies as well as conducting immunotoxicity studies or adding endpoints in routine toxicity studies to assess immunophenotyping or functional immune alterations. The toxicologist is also central in communicating the risk to clinicians and regulators following interpretation of nonclinical immunotoxicology data and participating in the search for clinical immune biomarkers to ensure clinical safety of trial participants. The goal of this course is to discuss regulatory guidance and use case examples to illustrate the integration of toxicology, pathology and immunology in addressing both recent scientific advances and practical approaches in the pharmaceutical company setting and how this knowledge impacted clinical trials and labeling, with a focus on drug-induced immunosuppression. Similarities and differences in FDA and CPMP immunotoxicity guidance documents and recent progress in ICH harmonization will be discussed. Signal detection, including data from standard toxicity studies and the controversies among toxicologists and pathologists in following the guidelines will be addressed, including the recently drafted Best Practice Guideline for the Routine Pathology of the Immune System by the STP Immunotoxicology Working Group. Immunophenotyping conducted by flow cytometry or lymphoid immunohistochemistry will cover recent advances with case studies. Functional assays, such as T-cell dependent antibody response, natural killer cell activity, host resistance macrophage/neutrophil function, and cell-mediated immunity will be discussed with regard to design and timing in drug development from an immunotoxicologists perspective. Finally, interpretation of nonclinical immunotoxicology data for risk assessment in clinical trials and use of clinical immune biomarkers will be featured.

Immunotoxicology: What’s New on the ICH Harmonization Front
Kenneth L. Hastings, CDER Office of New Drugs, USFDA, Rockville, MD

Immunosuppression Signal Detection in Standard Toxicity Studies and Immunophenotyping: A Pathologist’s Approach
Patrick J. Haley, AstraZeneca, Wilmington, DE

Assay to Assay: Functional Evaluation of Imunosuppression
Danuta J. Herzyk, GlaxoSmithKline, King of Prussia, PA

Clinical Viewpoint: Interpretation of Nonclinical Immunotoxicity Data, Human Risk Assessment and Clinical Immune Biomarkers
Ian Gourley, Lilly Research Laboratories, Indianapolis, IN

PM10 - Functional Analysis of Gene and Protein Expression: From Experimental Design to Data Analysis

ADVANCED
Chairperson: Richard S. Pollenz, University of South Florida, Tampa, FL

Endorsements
Mechanisms Specialty Section

The mechanistic analysis of cellular responses to xenobiotics requires the functional characterization of changes in both gene and protein expression. The ability to study changes to both genes and proteins in vitro and in vivo has become accessible to more laboratories with the development of molecular tools such as microarrays, siRNA, recombinant protein expression and viral gene delivery. However, the ability to utilize these techniques and generate reproducible results requires a detailed understanding of each procedure. Thus, the goal of this course is to provide the investigator with an overview of experimental design and the use of proper controls for four cutting edge techniques. The first talk will focus on experimental design and analysis of gene expression studies utilizing microarrays. The second presentation will discuss recent advances in the chomatin-immunoprecipitation (ChIP) assay for the analysis of protein interactions at the level of DNA. The third presentation will discuss gene delivery into hepatocytes in vivo utilizing the adenovirus system. Finally, the last presentation will discuss the advantages and disadvantages of recombinant protein expression in cultured cells with emphasis on the level and functionality of the expressed protein and the use of siRNA. The course will be of broad interest to those laboratories considering contemporary mechanistic approaches to gene and protein expression as well as those currently investigating these endpoints.

Design and Analysis of Microarray Experiments
Thomas Sutter, Feinston Center for Genomic Research, University of Memphis, Memphis, TN, USA

Analysis of Protein_DNA Interactions Using Chromatin_Immunoprecipitation (ChIP)
Eli Hestermann, Department of Biology, Furman University, Greenville, SC, USA

Adenovirus_Mediated Gene Delivery to Alter Protein Expression In Vivo
Cornelis Elferink, Department of Pharmacology, UTMB, Galveston, TX, USA

Basics of Recombinant Protein Expression and RNA Interference in Cultured Cells
Richard S. Pollenz, Department of Biology, University of South Florida, Tampa, FL, USA

PM 11 - Xenobiotic Transporters

BASIC
Chairperson: Douglas A. Keller, Sanofi-Aventis

Endorsement
*Drug Discovery Specialty Section

Toxicologists have traditionally considered that chemicals are absorbed and distributed throughout the body largely due to their lipid solubility and diffusivity through cell membranes. We also consider drug metabolism as a process that makes chemicals more water soluble so they will less readily pass cell membranes and be more rapidly excreted. However, it was not understood how the water-soluble metabolite could exit the cell to be excreted. During the last decade a number of transporters have been identified that not only are responsible for the excretion of chemicals, but also for the absorption and distribution of xenobiotics. This course, highlighting the progress made in this research field as both timely and of significant interest to a large number of SOT members who are in academic, government and industrial sectors. This course will give an overview of the families of transporters involved in absorption, distribution and excretion of drugs and chemicals, as well as presentations specific to the major tissues where transporters are known to influence toxicity. The roles of transporters in disposition and toxicity in the liver, kidney, intestine and brain will be discussed.

Overview of the Families of Transporters Responsible for the Absorption, Distribution, and Excretion of Drugs
Curtis D. Klaassen, University of Kansas Medical Center, Kansas City, KS

Pharmacological and Toxicological Significance of Hepatic Transporters
Bruno Hagenbuch, University of Kansas Medical Center, Kansas City, KS

Importance of Renal Transporters in Chemical Disposition and Toxicity
John B. Pritchard, NIEHS, Research Triangle Park, NC

Clinical and Pharmacological Implications of Transporters in the Intestine and Blood-Brain Barrier
Jashvant D. Unadkat, University of Washington, Seattle, WA

PM12 - Neuropathology for the Toxicologist

BASIC
Chairperson: David C. Dorman, CIIT Centers for Health Research, Research Triangle Park, NC

Endorsements
Comparative and Veterinary Specialty Section
Neurotoxicology Specialty Section
*Toxicologic & Exploratory Pathology Specialty Section

This course is designed to provide a basic overview of rodent neuropathology. The course will start off with a review of the normal anatomy and histology of the adult nervous system. This overview will also discuss the ways in which neuropathology and functional assays of motor activity and other behaviors relate to one another. The second lecture relates to tissue handling techniques and basic approaches in neuropathology. This topic is critically important since the ability to detect chemical-induced neuropathology requires proper tissue fixation and processing. Although the second lecture will largely focus on rodent tissues, the approaches and methods to be discussed can be easily adapted to other species. The course will then transition into two presentations focused on common lesions induced by model neurotoxicants. One presentation will focus on central nervous system effects while the latter lecture will address peripheral neuropathies. Our final presentation will discuss morphometric approaches in neuropathology including a discussion of the use of magnetic resonance imaging methods in neurotoxicologic pathology. Participants in this course will gain a greatly improved appreciation of basic neuropathology and applications to toxicology.

An Introduction to the Nervous System
David C. Dorman, CIIT Centers for Health Research, Research Triangle Park, NC

Practical Methods in Rodent Neuropathology
Mark T. Butt, Pathology Associates, Frederick, MD

Oh, My Aching Head: Toxicant-Induced Neuropathology of the Central Nervous System
Brad Bolon, GEMpath Inc., Cedar City, UT

Morphologic Assessment of the Peripheral Nervous System
William M. Valentine, Vanderbilt University, Nashville, TN

Quantitative Morphology in Rodent Neuropathology
Karl Jensen, USEPA, Research Triangle Park, NC

PM13 - Assessing Airway Injury and Remodeling Induced by Inhaled Pollutants Using Magnetic Resonance Imaging, Microscopy and Modeling

BASIC
Chairpersons: Jack R. Harkema, Michigan State University, East Lansing, MI and Richard Corley, Pacific Northwest National Laboratories, Richland, WA

Endorsements
Inhalation Specialty Section

The goal of this course is to present state-of-the-art methods of assessing injury and remodeling of the conducting airways caused by acute or chronic exposures to airborne toxicants including environmental pollutants (e.g., particulate matter, ozone, cigarette smoke, fungal and bacterial toxins). The nature, severity and distribution of airway lesions caused by these inhaled toxicants are due to an integration of several factors including the physicochemical characteristics of the inhaled toxicant (e.g., size of the particles, reactivity of the gas), the exposure regimen (e.g., short-versus long-term, continuous versus episodic), local intra-airway dosimetry (e.g., hot spots of deposition at airway bifurcations, slow clearance from sites of overload) and cellular susceptibility (e.g., sensitive versus resistant epithelial cell types). Toxicant-induced alterations to the airway structure range from oncotic or apoptotic cell death of epithelial cells lining the luminal surface with concurrent acute inflammation to marked remodeling of the airway wall due to intramural fibrosis, epithelial hyperplasia/metaplasia, and chronic active inflammation. The strengths and limitations of a variety of state-of-the-art imaging techniques for visualizing macroscopic changes to the respiratory tract, including magnetic resonance imaging, and computed tomography, will be presented along with those of more routine methods of light, electron and confocal microscopy for assessing tissue and cellular pathology caused by inhaled pollutants. The fundamental principles of airway stereology will also be presented and how these mathematically proven morphometric techniques may be used to quantify, without bias, changes to these tubular structures in order to estimate severity of complex lesions. Finally, presenters will describe the development and implementation of computational models of geometrically correct upper and lower airways used to define, site-specific, dose-response relationships along the respiratory tract of laboratory animals and to estimate the risk of air pollutant exposures to human health.

An Overview of Airway Injury and Remodeling Caused by Inhaled Toxicants: From the Nose to the Lung and from Gases to Particles
Jack R. Harkema, Michigan State University, East Lansing, MI

State-of-the-Art Techniques for Imaging the Upper and Lower Respiratory Tract: Identifying macroscopic to microscopic airway alterations caused by airborne toxicants
Charles Plopper, University of California-Davis, Davis, CA

Sampling, Stereology, and Statistics: Quantifying changes along tubular structures and without bias
Dallas Hyde, University of California-Davis, Davis, CA

Using Computer-Assisted Airway Reconstruction to Define Dose-Response Relationships
Julia Kimbell, CIIT Centers for Health Research, Research Triangle Park, NC

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