Environmental Sciences Institute Research Articles

Characterization and Interlaboratory Comparison of a Gene Expression Signature for Differentiating Genotoxic Mechanisms

The genotoxicity testing battery is highly sensitive for detection of chemical carcinogens. However, it features a low specificity and provides only limited mechanistic information required for risk assessment of positive findings. This is especially important in case of positive findings in the in vitro chromosome damage assays, because chromosome damage may be also induced secondarily to cell death. An increasing body of evidence indicates that toxicogenomic analysis of cellular stress responses provides an insight into mechanisms of action of genotoxicants. To evaluate the utility of such a toxicogenomic analysis we evaluated gene expression profiles of TK6 cells treated with four model genotoxic agents using a targeted high density real-time PCR approach in a multilaboratory project coordinated by the Health and Environmental Sciences Institute Committee on the Application of Genomics in Mechanism-based Risk Assessment. We show that this gene profiling technology produced reproducible data across laboratories allowing us to conclude that expression analysis of a relevant gene set is capable of distinguishing compounds that cause DNA adducts or double strand breaks from those that interfere with mitotic spindle function or that cause chromosome damage as a consequence of cytotoxicity. Furthermore, our data suggest that the gene expression profiles at early time points are most likely to provide information relevant to mechanisms of genotoxic damage and that larger gene expression arrays will likely provide richer information for differentiating molecular mechanisms of action of genotoxicants. Although more compounds need to be tested to identify a robust molecular signature, this study confirms the potential of toxicogenomic analysis for investigation of genotoxic mechanisms.

A HESI consortium approach to assess the human predictive value of non-clinical repolarization assays

Drug-induced ventricular arrhythmia and Torsades de Pointes remain a serious public health issues in bringing safe new pharmaceuticals to the market place. Under the auspices of the International Life Science Institute (ILSI)–Health and Environmental Sciences Institute (HESI), a consortium involving representatives from pharmaceutical companies, regulatory agencies and opinion leaders from the scientific and medical research communities has been initiated. The objectives are (1) to assess the concordance between signals in non-clinical repolarization assays and clinical QT interval prolongation; (2) to investigate the mechanisms for any discrepancy identified between non-clinical and clinical results and to determine viable and successful alternative approaches to identify these compounds; and (3) to assess the proarrhythmic potential of such compounds. At present, the consortium is conducting a retrospective analysis of non-clinical and clinical data from both FDA and contributing companies' databases and supplementing with a literature review. The overall objectives of these initial efforts are to establish a quantitative integrated risk assessment for each compound; to define criteria for concordance and apply them to the database in order to identify non-concordant compounds.

Current developments in reproductive toxicity testing of pesticides

A protocol to evaluate the potential developmental and reproductive effects of test chemicals has been developed by the Life Stages Task Force of the International Life Sciences Institute (ILSI)/Health and Environmental Sciences Institute (HESI) Agricultural Chemical Safety Assessment (ACSA) Technical Committee. Since the original publication, several international groups have provided public comment on conducting the test. The extended one-generation reproductive toxicity test is now under consideration as a potential test guideline. The protocol uses a flexible approach that is markedly different from the current multigenerational guidelines. It encourages the use of toxicokinetics when setting the doses, evaluates more than one rat per sex per litter in the F1 offspring and does not necessarily require mating of the F1 to produce an F2 (F1 mating may be triggered by the presence of effects in the P0 and developing F1 rats). A number of additional reproductive endpoints, and the neurotoxicity and immunotoxicity cohorts are included. The ACSA protocol was developed with the goal of assuring that the methods are scientifically appropriate and the toxicological endpoints and exposure durations are relevant for risk assessment. Compared to existing testing strategies, the proposed approach uses substantially fewer animals, provides additional information on the neonate, juvenile and pubertal animal, and includes an estimation of human exposure potential for making decisions about the extent of testing required. In this paper, the evolution of the protocol since the 2006 publication is discussed. These changes reflect the collective input of a U.S. expert panel of government and industrial scientist convened in 2007 and discussions of an OECD expert group held in Paris, France (October, 2008).

CHEMICAL TESTING: Developmental Immunotoxicity in Review

Much of the current knowledge regarding the adverse effects of environmental pollutants is based on laboratory evaluations performed using adult animals. But new clues now suggest that many diseases—including allergies, asthma, type 1 diabetes, atherosclerosis, leukemia, and autism—may originate in immune system damage caused by exposure to toxicants in early life, possibly even before birth. The routine conduct of safety screens on adult animals misses potential windows of unparalleled immune vulnerability in early life, writes immunotoxicology professor Rodney Dietert of the Cornell University College of Veterinary Medicine, in an invited review of the developmental immunotoxicity (DIT) literature published in the January 2009 issue of Chemical Research in Toxicology. A central tenet of DIT is that the prenatal and perinatal immune systems are uniquely sensitive to toxic insult. For instance, prenatal exposure to dioxin causes immunotoxicity at a dose approximately 100‐fold lower than the exposure that causes damage in adults, as Dietert and colleagues reported in the January 2006 Journal of Toxicology and Environmental Health, Part B Critical Reviews. Moreover, different immune processes are at work in different life stages. Disease‐fighting T cells, for example, develop in the thymus during early life, but by adulthood are produced largely in other tissues. Also, some chemicals produce only transient immune system changes with adult exposure but produce lasting changes with fetal or early neonatal exposure. Some safety screens are conducted across the life span, but DIT testing is usually initiated only after adult data suggest immunotoxicity. “Dietert’s review makes the important point that DIT testing is a complementary approach to toxicology testing in adult immune systems and should be considered when developing immune systems are likely to be exposed to xenobiotics,” says Fred Miller, chief of the NIEHS Environmental Autoimmunity Group. “However, DIT testing raises important questions regarding the appropriate doses, periods of exposure, and routes of exposure that should be tested at different phases of immune system development, all of which would depend on the expected human exposure.” Such questions regarding how to implement DIT testing have been debated worldwide for nearly a decade. But further work is needed to refine the details of screening protocols for DIT testing, says Susan Makris, a toxicologist with the Environmental Protection Agency. Screening procedures may need to be tailored to individual chemicals. “For example, consideration must be given to the pharmacokinetics of the chemical [how the chemical is metabolized] in terms of how it’s being administered,” she says. Otherwise, testing may not closely simulate real‐world exposures. Michael Holsapple, executive director of the Health and Environmental Sciences Institute at the International Life Sciences Institute in Washington, DC, says that what is really needed is development and implementation of DIT testing protocols, including studies to determine “triggers” that would prompt the implementation of such protocols. One such trigger might be the ability of a test compound to cross the placenta or be secreted in milk. “If the fetal or neonatal exposure seems so low as to be basically inconsequential, this could mean that DIT studies would be of little use in assessing risk,” he explains. “These and many other considerations would seem to challenge the premise that DIT testing should become a global policy.” Once protocols are complete, regulatory change must occur across many agencies. “But even where there is agreement on what kind of testing needs to be done, implementing those studies, chemical by chemical, will be very much driven by the regulations that are enforced for each of those groups,” says Makris. Perhaps the biggest obstacle to normalizing DIT testing is that of finding reliable ways to generate clear evidence for an impact of early‐life exposures on human health outcomes in adult life. “Discovering and characterizing the connections between chemical exposure during early‐life stages and disease onset and progression during later‐life stages will only come from developing and implementing DIT testing protocols,” says Holsapple. “And the complexity of these protocols, if they are to cover the full gamut suggested by Dr. Dietert, will be considerable indeed.”

Summary of a workshop on nonclinical and clinical immunotoxicity assessment of immunomodulatory drugs

The number of anti-inflammatory and immunomodulatory drugs being developed in the pharmaceutical industry has increased considerably in the past decade. This increase in research and development has been paralleled by questions from both regulatory agencies and industry on how best to assess decreased host resistance to infections or adverse immunostimulation caused by immunomodulatory agents such as anti-cytokine antibodies (e.g., the tumor necrosis factor-α inhibitors), anti-adhesion molecule antibodies (e.g., anti-α-4 integrin inhibitors) and immunostimulatory molecules (e.g., anti-CD28 antibodies). Although several methods have been developed for nonclinical assessment of immunotoxicity, highly publicized adverse events have brought to light significant gaps in the application of nonclinical immunotoxicity testing in assessing potential risk in humans. Confounding this problem is inconsistent application of immunotoxicology methods for risk assessment within the scientific community, limited understanding of appropriate immunotoxicity testing strategy for immunomodulators and inconsistent testing requests by regulatory agencies. To address these concerns, The Immunotoxicology Technical Committee (ITC) of the International Life Science Institute (ILSI) Health and Environmental Sciences Institute (HESI) organized a workshop on Immunomodulators and Clinical Immunotoxicology in May 2007. The Workshop was convened to identify key gaps in nonclinical and clinical immunotoxicity testing of anti-inflammatory and immunomodulatory agents and to begin to develop consistent approaches for immunotoxicity testing and risk assessment. This paper summarizes the outcome of the HESI ITC Immunomodulators and Clinical Immunotoxicology Workshop. Topics not discussed at the Workshop were outside the scope of this report. Although more work is needed to develop consistent approaches for immunotoxicity assessment of immunomodulators, this Workshop provided the foundation for future discussion.

Research Highlights

Research Highlights

International Life Sciences Institute (Health and Environmental Sciences Institute, HESI) initiative on moving towards better predictors of drug-induced torsades de pointes.

Knowledge of the cardiac safety of emerging new drugs is an important aspect of assuring the expeditious advancement of the best candidates targeted at unmet medical needs while also assuring the safety of clinical trial subjects or patients. Present methodologies for assessing drug-induced torsades de pointes (TdP) are woefully inadequate in terms of their specificity to select pharmaceutical agents, which are human arrhythmia toxicants. Thus, the critical challenge in the pharmaceutical industry today is to identify experimental models, composite strategies, or biomarkers of cardiac risk that can distinguish a drug, which prolongs cardiac ventricular repolarization, but is not proarrhythmic, from one that prolongs the QT interval and leads to TdP. To that end, the HESI Proarrhythmia Models Project Committee recognized that there was little practical understanding of the relationship between drug effects on cardiac ventricular repolarization and the rare clinical event of TdP. It was on that basis that a workshop was convened in Virginia, USA at which four topics were introduced by invited subject matter experts in the following fields: Molecular and Cellular Biology Underlying TdP, Dynamics of Periodicity, Models of TdP Proarrhythmia, and Key Considerations for Demonstrating Utility of Pre-Clinical Models. Contained in this special issue of the British Journal of Pharmacology are reports from each of the presenters that set out the background and key areas of discussion in each of these topic areas. Based on this information, the scientific community is encouraged to consider the ideas advanced in this workshop and to contribute to these important areas of investigations over the next several years.

Histopathology of the Urinary Bladders of Cynomolgus Monkeys Treated with PPAR Agonists

Peroxisome proliferator-activated receptors (PPAR) are involved in the pathogenesis of insulin resistance, diabetes, hyperlipidemias, and related complications. Consequently, a mechanistic understanding of PPAR subtypes and their activation provides promising therapeutic targets for the management of type 2 diabetes mellitus and the metabolic syndrome. Available data from rodent carcinogenicity studies, however, demonstrate that PPAR agonists can be tumorigenic in one or more species of rodents at multiple sites. Sufficient data are not yet available to explain the mode(s) of action for most of these tumor types. There has been information presented by FDA that indicates there are urothelial changes in the monkey (and possibly the dog) in addition to the rat. Outstanding questions exist regarding potency, species differences, safety margins, and other issues. In 2005, the International Life Sciences Institute (ILSI) Health and Environmental Sciences Institute (HESI) PPAR Agonist Project Committee was established to advance research on the modes of action and potential human relevance of emerging rodent tumor data. Additionally, the HESI PPAR Agonist Project Committee authorized a Pathology Working Group (PWG) to examine the urinary bladder from cynomolgus monkeys. The focus of this PWG was to establish consistent diagnostic criteria for urothelial changes and to assess the potential relationship of these changes to treatment. Specific diagnostic criteria and nomenclature were recommended for the diagnosis of urothelial granules, vacuolation, hypertrophy, and hyperplasia in studies conducted with PPARgamma and dual alpha/gamma agonists in cynomolgus monkeys, which will assist investigators performing toxicity studies to provide data in a consistent manner between studies and laboratories. In this review of selected tissues, treatment with PPAR agonists was not associated with urothelial hypertrophy or hyperplasia, but there was an increased incidence in the size and frequency of vacuoles within the superficial urothelial and adjacent intermediate cell layers.

Current and future methods for evaluating the allergenic potential of proteins: International workshop report 23–25 October 2007

The International Life Science Institute’s Health and Environmental Sciences Institute’s Protein Allergenicity Technical Committee hosted an international workshop October 23–25, 2007, in Nice, France, to review and discuss existing and emerging methods and techniques for improving the current weight-of-evidence approach for evaluating the potential allergenicity of novel proteins. The workshop included over 40 international experts from government, industry, and academia. Their expertise represented a range of disciplines including immunology, chemistry, molecular biology, bioinformatics, and toxicology. Among participants, there was consensus that (1) current bioinformatic approaches are highly conservative; (2) advances in bioinformatics using structural comparisons of proteins may be helpful as the availability of structural data increases; (3) proteomics may prove useful for monitoring the natural variability in a plant’s proteome and assessing the impact of biotechnology transformations on endogenous levels of allergens, but only when analytical techniques have been standardized and additional data are available on the natural variation of protein expression in non-transgenic bred plants; (4) basophil response assays are promising techniques, but need additional evaluation around specificity, sensitivity, and reproducibility; (5) additional research is required to develop and validate an animal model for the purpose of predicting protein allergenicity.

Time for a Change: From Exposure Assessment to Exposure Science

During the late 1970s the emerging field of risk assessment spawned a new area of data analysis called “exposure assessment.” It was developed to characterize a process for identifying an exposed population against a measured or estimated outcome—health risk. After publication of the 1983 “Red Book,” Risk Assessment in the Federal Government: Managing the Process [National Research Council (NRC) 1983], it became clear that confidence in our ability to complete complex exposure assessments was low, and “specific components in exposure assessment is complicated by the fact that current methods and approaches … appear to be medium or route specific … and very few components … could be applicable to all media” (NRC 1983). However, the Total Exposure Assessment Methodology (TEAM) study had begun (Wallace et al. 1986). In 1987, the NRC formed the first Committee on Exposure Assessment. It was apparent during the initial meeting that committee members, whose expertise ranged from basic sciences to environmental science and to medicine, were unclear about what exposure meant. A series of discussions and a workshop eventually defined many basic scientific principles, and eventually the committee’s report, Human Exposure Assessment for Air Pollutants [the “White Book” (NRC 1991)], laid the foundation for further development of the field of exposure. An immediate consequence of this effort was the formation of the International Society of Exposure Analysis (ISEA), which provided a home for human exposure research and the mechanistic foundations of the field. Concurrently, a laboratory in the U.S. Environmental Protection Agency (EPA) was renamed the National Exposure Research Laboratory, the Environmental and Occupational Health Sciences Institute started the first graduate program in exposure, and faculty members were hired at some major universities to focus on human exposure research (Lioy 1999). Further, research on single-route and multiroute exposure was supported by the World Health Organization and other international organizations such as the German Environmental Survey (Hoffmann et al. 2000). Recently, it has become apparent that many of the goals of the NRC White Book have been achieved and that analysis of exposure is becoming a priority for national and international strategies to reduce or prevent exposures. Further, the field has passed the point of solely making measurements in support of risk assessment. It has developed into a mature discipline of science, through which a theoretical framework was constructed for developing sophisticated mathematical models linking environmental science to toxicology and public health. An experimental (including observational field studies) foundation has been established to systematically examine how individuals and populations contact contaminants because of their personal activities and behaviors, the microenvironments contacted each day, and the general environment (Lioy 1990, 1999; Ott 1993). An example was the U.S. EPA’s successful National Human Exposure Assessment Survey (Sexton et al. 1995), which was used to focus exposure issues in the National Children’s Study (Needham et al. 2005). In the United States, the efforts to systematically examine problems were significantly enhanced by the U.S. EPA’s shift toward examining cumulative exposures for multiple pollutants and aggregate exposures from multiple media for a single pollutant, and the Centers for Disease Control and Prevention’s implementation of an extensive personal exposure biomonitoring program. Each methodology has reduced uncertainties outlined in the Red Book. In parallel we have seen source-to-dose modeling develop from fundamental scientific principles, such as the Modeling ENvironment for TOtal Risk [MENTOR] (Georgopoulos and Lioy 2006). In 2006, Dana Barr, editor-in-chief of the Journal of Exposure Science and Environmental Epidemiology, published that journal’s working definition for the field of exposure science: “the study of human contact with chemical, physical, or biological agents occurring in their environments, and [advancement of] knowledge of the mechanisms and dynamics of events either causing or preventing adverse health outcomes” (Barr 2006). Finally, in 2008, ISEA was renamed the International Society of Exposure Science (Weisel CW, unpublished information). In light of all the moves forward in the field and the parallel evolution of the Exposure Biology Program at the National Institute of Environmental Health Sciences (Weis et al. 2005), Environmental Health Perspectives will be renaming its classification for publication of research on human contact with contaminants from “exposure assessment” to “exposure science.” The change should continue to encourage and enhance the development of the new discipline and provide new avenues for publication of innovative research that directly impact the environmental health sciences, risk assessment, and risk management. The field has passed the point of solely making measurements in support of risk assessment. It has developed into a mature discipline of science, through which a theoretical framework was constructed for developing sophisticated mathematical models linking environmental science to toxicology and public health.

Sources of variation in baseline gene expression levels from toxicogenomics study control animals across multiple laboratories

BackgroundThe use of gene expression profiling in both clinical and laboratory settings would be enhanced by better characterization of variance due to individual, environmental, and technical factors. Meta-analysis of microarray data from untreated or vehicle-treated animals within the control arm of toxicogenomics studies could yield useful information on baseline fluctuations in gene expression, although control animal data has not been available on a scale and in a form best served for data-mining.ResultsA dataset of control animal microarray expression data was assembled by a working group of the Health and Environmental Sciences Institute's Technical Committee on the Application of Genomics in Mechanism Based Risk Assessment in order to provide a public resource for assessments of variability in baseline gene expression. Data from over 500 Affymetrix microarrays from control rat liver and kidney were collected from 16 different institutions. Thirty-five biological and technical factors were obtained for each animal, describing a wide range of study characteristics, and a subset were evaluated in detail for their contribution to total variability using multivariate statistical and graphical techniques.ConclusionThe study factors that emerged as key sources of variability included gender, organ section, strain, and fasting state. These and other study factors were identified as key descriptors that should be included in the minimal information about a toxicogenomics study needed for interpretation of results by an independent source. Genes that are the most and least variable, gender-selective, or altered by fasting were also identified and functionally categorized. Better characterization of gene expression variability in control animals will aid in the design of toxicogenomics studies and in the interpretation of their results.

Metals: Fresh Track on Indoor Dust

Dust is seldom innocuous. At the very least, it can earn homeowners a reputation for lax housekeeping. More important from a health perspective, it can also exacerbate asthma and allergies. Growing evidence suggests this humble material may pose other health threats caused in part by metals and other potentially toxic substances that hitchhike on core particles. In traditional risk assessment, one common assumption has been that contaminants in indoor dust are just a dilute version of what’s found in the soil outside the building. New evidence from a team of Canadian researchers reveals just the opposite for many metals: indoor dust can be burdened by much higher quantities than in soils outside, and the indoor version can also be more orally bioaccessible (made available to the body through the digestive process). Based on her team’s findings, Pat Rasmussen, a research scientist with Health Canada, says the old assumption “is wrong, and can result in a serious underestimate of indoor exposures in residential risk assessments.” This is one of the early results from the Canadian House Dust Study, scheduled to run from 2007 through 2010, for which Rasmussen is principal investigator. Besides metals, bacteria have been studied in homes in three cities; pesticides, alkylphenols, artificial musks, brominated flame retardants, organotins, parabens, perfluorinated compounds, phthalates, and triclosan also may be studied. Based on samples from about 1,000 homes in 13 representative cities, the study is expected to set the first scientific baseline for various contaminants in urban Canadian homes. Once all study results are available, Health Canada and the Public Health Agency of Canada, two of the study sponsors, are expected to make recommendations on how best to deal with household dust and its associated contaminants. For the pilot phase of the metals study, which was published in volume 14, issue 2 (2008) of Human and Ecological Risk Assessment, Rasmussen and her colleagues determined the copper, nickel, and zinc contents of archived dust and soil samples from 22 homes in one city. They found not only that the concentration of each metal was far higher in indoor dust than in outdoor soil (9, 3, and 7 times, respectively), but that each metal was much more orally bioaccessible in dust than in soil (13, 8, and 23 times, respectively). In addition, they discovered large differences in the organic and inorganic carbon contents, which were 5 and 10 times higher, respectively, in indoor dust than in soil. Through the use of synchrotron X-ray absorption spectroscopy, the team found that copper was more often bound with organic portions of indoor dust (e.g., sulfides such as cysteine, and possibly acetate and oxalate), while zinc was more often bound with the mineral portions. “The X-ray analysis is rather novel, and the results informative,” says Andrew Turner, an associate professor of environmental sciences at the University of Plymouth, United Kingdom. He is not involved with any segment of the study and says he’s unaware of any other study of similar scope. The team hopes its future studies using the synchrotron and novel applications of technology such as Elementar devices, specialized analytical instruments for examining nonmetallic elements, will help it pin down the specific sources of the indoor metals, whose concentrations tend to vary substantially from house to house in the same neighborhood and even from room to room in the same house for reasons unknown. Results from earlier studies indicate that many other metals—including lead, cadmium, arsenic, mercury, aluminum, chromium, iron, manganese, and tin—are usually present in indoor dust. Rasmussen is studying the presence in dust of these other elements and is finalizing results for publication. The mere presence of these metals does not necessarily pose a clear health risk, says Paul Lioy, associate director of the Environmental and Occupational Health Sciences Institute at the University of Medicine & Dentistry of New Jersey. But he says this study, which he did not participate in, adds several nuggets of new information, and he notes there are many indications that this general line of inquiry could prove very important: “Dust is an incredible marker of past exposures to persistent chemicals.” Until more is known about the health effects of house dust, Rasmussen says the best approach for homeowners may be to play it safe and keep dust levels down by vacuuming (a HEPA filter is a good idea but not required, says Rasmussen), damp-wiping surfaces, and removing shoes when entering a house. For situations where the dust is known to be contaminated by a proven toxic source, such as lead paint in older buildings or mercury from a broken compact fluorescent lamp, more specialized cleanup methods are needed.

Guidance for evaluating in vivo fish bioaccumulation data

Currently, the laboratory-derived fish bioconcentration factor (BCF) serves as one of the primary data sources used to assess the potential for a chemical to bioaccumulate. Consequently, fish BCF values serve a central role in decision making and provide the basis for development of quantitative structure-property relationships (QSPRs) used to predict the bioaccumulation potential of untested compounds. However, practical guidance for critically reviewing experimental BCF studies is limited. This lack of transparent guidance hinders improvement in predictive models and can lead to uninformed chemical management decisions. To address this concern, a multiple-stakeholder workshop of experts from government, industry, and academia was convened by the International Life Sciences Institute Health and Environmental Sciences Institute to examine the data availability and quality issues associated with in vivo fish bioconcentration and bioaccumulation data. This paper provides guidance for evaluating key aspects of study design and conduct that must be considered when judging the reliability and adequacy of reported laboratory bioaccumulation data. Key criteria identified for judging study reliability include 1) clear specification of test substance and fish species investigated, 2) analysis of test substance in both fish tissue and exposure medium, 3) no significant adverse effects on exposed test fish, and 4) a reported test BCF that reflects steady-state conditions with unambiguous units. This guidance is then applied to 2 data-rich chemicals (anthracene and 2,3,7,8-tetrachlorodibenzo-p-dioxin) to illustrate the critical need for applying a systematic data quality assessment process. Use of these guidelines will foster development of more accurate QSPR models, improve the performance and reporting of future laboratory studies, and strengthen the technical basis for bioaccumulation assessment in chemicals management.

Predicting Future Human and Environmental Health Challenges: The Health and Environmental Sciences Institute's Scientific Mapping Exercise

To predict important strategic issues in product safety during the next 10 years, the Health and Environmental Sciences Institute (HESI) of the International Life Sciences Institute initiated a mapping exercise to evaluate which issues are likely to be of societal, scientific, and regulatory importance to regulatory authorities, the HESI membership, and the scientific community at large. Scientists representing government, academia, and industry participated in the exercise. Societal issues identified include sensitive populations, alternative therapies, public education on the precautionary principle, obesity, and aging world populations. Scientific issues identified include cancer testing, children's health, mixtures and co-exposures, sensitive populations, idiosyncratic reactions, “omics” or bioinformatics, and environmental toxicology. Regulatory issues identified include national and regional legislation on chemical safety, exposure inputs, new technologies, transitioning new science into regulations and guidelines, conservative default factors, data quality, and sensitive populations. Because some issues were identified as important in all three areas (e.g. sensitive populations), a comprehensive approach to assessment and management is needed to ensure consideration of societal, scientific, and regulatory implications. The resulting HESI Combined Challenges Map is not intended to offer a universal description of challenges in safety assessment, nor is it intended to address, advocate, or manage the prioritized issues. Rather, the map focuses on and predicts issues likely to be central to the strategic agendas of individual companies and regulatory authorities in the developed world. Many of these issues will become increasingly important in the future in rapidly developing economies, such as India and China. The scientific mapping exercise has particular value to the toxicology community because it represents the contributions of key scientists from around the world from government, academia, and industry.

Histopathology Of Hemangiosarcomas in Mice and Hamsters and Liposarcomas/Fibrosarcomas in Rats Associated with PPAR Agonists

Peroxisome proliferator-activated receptors (PPAR) are involved in the pathogenesis of insulin resistance, diabetes, and related complications. Consequently, the identification of PPAR subtypes and the potential for their activation provides promising therapeutic targets for the management of type 2 diabetes mellitus. Available data from rodent carcinogenicity studies, however, demonstrate that PPAR agonists can be tumorigenic in one or more species of rodents at multiple sites. In 2005, the Health and Environmental Sciences Institute (HESI) PPAR Agonist Project Committee was established by a group of pharmaceutical companies to advance research on and to understand the modes of action and human relevance of this emerging rodent tumor data for PPAR agonists. Since the most commonly observed tumor types reported in rodents are hemangiosarcomas, fibrosarcomas and liposarcomas, the PPAR Agonist Project Committee approved a Pathology Working Group (PWG) to develop consensus of morphologic criteria for tumor diagnoses and consistency of diagnoses across multiple studies for hemangiosarcomas in mice and hamsters and liposarcomas/fibrosarcomas in rats. Therefore, the focus of the PWG review was to establish consistent tumor diagnostic criteria, to assess evidence of potentially preneoplastic changes and to identify distinguishing morphologic differences which may exist between spontaneous changes present in control animals with similar changes from treated animals. Specific diagnostic criteria and nomenclature are recommended for the classification of proliferative vascular lesions which may be present in mice or hamsters and for proliferative mesenchymal changes in rats in studies that are conducted with PPAR agonists.

Points to consider emerging from a mini-workshop on cardiac safety: Assessing torsades de pointes liability

A mini-workshop on cardiac safety focusing on assessing drug-induced Torsades de Pointes (TdP) liability was convened as part of the 6th Annual Meeting of the Safety Pharmacology Society. The purpose of this brief publication is to disseminate the salient points emanating from this workshop as a means of engaging the scientific community in the appropriate discussions needed to advance this important field of human safety. The recommendations in this publication extend those of the workshop on “Moving Towards Better Predictors of Drug-Induced Torsades de Pointes” held in November 2005 under the auspice of the International Life Sciences Institute, Health and Environmental Sciences Institute; they fall into four key areas: molecular and cellular biology underlying TdP, dynamics of periodicity, models of TdP proarrhythmia and key considerations for demonstrating utility of non-clinical models. The reader is encouraged to consider the recommendations emanating from the two workshops and align these with ongoing studies in their laboratories. The authors intend to convene a workshop in 2009/2010 to judge advancements in the field of study of drug-induced TdP and make recommendations for a focused validation of those methods holding the greatest promise of improving the predictivity of this unwanted human cardiac risk.

Relevance and follow-up of positive results in in vitro genetic toxicity assays: An ILSI-HESI initiative

In vitro genotoxicity assays are often used to screen and predict whether chemicals might represent mutagenic and carcinogenic risks for humans. Recent discussions have focused on the high rate of positive results in in vitro tests, especially in those assays performed in mammalian cells that are not confirmed in vivo. Currently, there is no general consensus in the scientific community on the interpretation of the significance of positive results from the in vitro genotoxicity assays. To address this issue, the Health and Environmental Sciences Institute (HESI), held an international workshop in June 2006 to discuss the relevance and follow-up of positive results in in vitro genetic toxicity assays. The goals of the meeting were to examine ways to advance the scientific basis for the interpretation of positive findings in in vitro assays, to facilitate the development of follow-up testing strategies and to define criteria for determining the relevance to human health. The workshop identified specific needs in two general categories, i.e., improved testing and improved data interpretation and risk assessment. Recommendations to improve testing included: (1) re-examine the maximum level of cytotoxicity currently required for in vitro tests; (2) re-examine the upper limit concentration for in vitro mammalian studies; (3) develop improved testing strategies using current in vitro assays; (4) define criteria to guide selection of the appropriate follow-up in vivo studies; (5) develop new and more predictive in vitro and in vivo tests. Recommendations for improving interpretation and assessment included: (1) examine the suitability of applying the threshold of toxicological concern concepts to genotoxicity data; (2) develop a structured weight of evidence approach for assessing genotoxic/carcinogenic hazard; and (3) re-examine in vitro and in vivo correlations qualitatively and quantitatively. Conclusions from the workshop highlighted a willingness of scientists from various sectors to change and improve the current paradigm and move from a hazard identification approach to a “realistic” risk-based approach that incorporates information on mechanism of action, kinetics, and human exposure..

ILSI–HESI cardiovascular safety subcommittee dataset: An analysis of the statistical properties of QT interval and rate-corrected QT interval (QTc)

Introduction The Health and Environmental Sciences Institute of the International Life Sciences Institute (ILSI/HESI) Cardiovascular Safety Subcommittee outlined a set of in vivo telemetry studies to determine how well this preclinical model identified compounds known to cause torsades de pointes (TdP) and prolong QT interval in humans. In the original analysis of these data, QT, QTcB (Bazett model), QTcF (Fridericia model), and QTcQ (animal-specific model) were evaluated. We further evaluate the statistical properties of these measurements, using a method that can properly account for the sources of variability in the dataset. Methods The ILSI/HESI telemetry studies were conducted as a double Latin square design where eight dogs each received a vehicle control and three dose levels of a compound on four separate dosing days. We statistically analyzed the QT/QTc intervals using a repeated measures analysis of covariance and evaluate the powers for QT, QTcF and QTcQ based on simulations. Results The analyses for QTcF and QTcB intervals show that all six compounds which were known to cause TdP in humans were identified as positive and all six compounds known to be free of TdP events in their clinical use had no statistically significant treatment-related effects, while the analyses for QTcQ identified all positive compounds except pimozide. The power analysis shows that the method can detect a 7% increment of QT, a 5% increment of QTcF, and a 4% increment of QTcQ, with greater than 80% of power when n = 8. Discussion We describe a repeated measures procedure to perform statistical analysis of covariance on Latin square designs and show that it can be used to detect meaningful changes in the analysis of QT/QTc intervals.

Evaluating emerging issues in epidemiology

Industry and government institutions need a credible approach for evaluating and responding to emerging public health issues. Representatives of industry, government, and academia met under the auspices of the International Life Sciences Institute’s Health and Environmental Sciences Institute (HESI) to develop successful strategies for dealing with emerging issues based on historical case studies. The case studies chosen for evaluation were (1) tampon use and toxic shock syndrome; (2) hazardous waste and childhood cancer risk in Toms River, New Jersey; (3) fenfluramine and phentermine use and valvular heart disease; (4) silicone breast implants and cancer and auto-immune disease; and (5) progestational drugs and birth defects. We identified eight lessons from these case studies. Foremost, we recommend that public and private institutions not defer action until an issue is scientifically resolved and stress that cooperation among issue stakeholders is critical for effective issue resolution. We suggest establishing a research program as an effective way to assure that good science is included in resolution of the issue. We further recommend frequent and timely communication with all stakeholders, and the development of research approaches to fill gaps when the scientific data on an issue are limited.

Integration of human biomonitoring exposure data into risk assessment: HESI initiatives and perspectives

The International Life Sciences Institute (ILSI) Health and Environmental Sciences Institute (HESI) established a Biomonitoring Technical Committee in 2002 to define and characterize appropriate scientific use(s) of biomonitoring tools and/or biomonitoring data needed to characterize exposure to chemicals, and to explore mechanisms for integrating biomonitoring exposure data and toxicology data into a robust risk assessment process. This paper summarizes the Technical Committee's initiatives and perspectives on improving the quality, value, application, and interpretation of human biomonitoring data for exposure and risk assessment purposes. Scientific outreach, collaborative partners, and new projects are described.