National Research Council Vision Research Articles

Acute Oral, Subacute, and Developmental Toxicity Profiling of Naphthalene 2-Yl, 2-Chloro, 5-Nitrobenzoate: Assessment Based on Stress Response, Toxicity, and Adverse Outcome Pathways.

The U.S. National Research Council (NRC) introduced new approaches to report toxicity studies. The NRC vision is to explore the toxicity pathways leading to the adverse effects in intact organisms by the exposure of the chemicals. This study examines the toxicity profiling of the naphthalene-2-yl 2-chloro-5-dinitrobenzoate (SF5) by adopting the vision of NRC that moves from traditional animal studies to the cellular pathways. Acute, subacute, and developmental toxicity studies were assayed according to the Organization for Economic Cooperation and Development (OECD) guidelines. The stress response pathway, toxicity pathway, and adverse effects outcome parameters were analyzed by using their standard protocols. The results showed that the acute toxicity study increases the liver enzyme levels. In a subacute toxicity study, alkaline phosphatase (ALP) levels were raised in both male and female animals. SF5 significantly increases the normal sperm count in the male animals corresponding to a decrease in the abnormality count. Developmental toxicity showed the normal skeletal and morphological parameters, except little hydrocephalus was observed in developmental toxicity. Doses of 20 mg/kg in males and 4 mg/kg in females showed decreased glutathione (GSH) levels in the kidney and liver. MDA levels were also increased in the kidney and liver. However, histopathological studies did not show any cellular change in these organs. No statistical difference was observed in histamine levels, testosterone, nuclear factor erythroid two-related factor-2 (Nrf2), and nuclear factor-kappa B (NF-κB), which showed no initiation of the stress response, toxicity, and adverse effect pathways. Immunomodulation was observed at low doses in subacute toxicity studies. It was concluded that SF5 did not produce abrupt and high-toxicity levels in organs and biochemical parameters. So, it is safe for further studies.

High-throughput screening and environmental risk assessment: State of the science and emerging applications.

In 2007 the United States National Research Council (NRC) published a vision for toxicity testing in the 21st century that emphasized the use of in vitro high-throughput screening (HTS) methods and predictive models as an alternative to in vivo animal testing. In the present study we examine the state of the science of HTS and the progress that has been made in implementing and expanding on the NRC vision, as well as challenges to implementation that remain. Overall, significant progress has been made with regard to the availability of HTS data, aggregation of chemical property and toxicity information into online databases, and the development of various models and frameworks to support extrapolation of HTS data. However, HTS data and associated predictive models have not yet been widely applied in risk assessment. Major barriers include the disconnect between the endpoints measured in HTS assays and the assessment endpoints considered in risk assessments as well as the rapid pace at which new tools and models are evolving in contrast with the slow pace at which regulatory structures change. Nonetheless, there are opportunities for environmental scientists and policymakers alike to take an impactful role in the ongoing development and implementation of the NRC vision. Six specific areas for scientific coordination and/or policy engagement are identified. Environ Toxicol Chem 2019;38:12-26. Published 2018 Wiley Periodicals Inc. on behalf of SETAC. This article is a US government work and, as such, is in the public domain in the United States of America.

Comparison of Points of Departure for Health Risk Assessment Based on High-Throughput Screening Data.

Background:The National Research Council’s vision for toxicity testing in the 21st century anticipates that points of departure (PODs) for establishing human exposure guidelines in future risk assessments will increasingly be based on in vitro high-throughput screening (HTS) data.Objectives:The aim of this study was to compare different PODs for HTS data. Specifically, benchmark doses (BMDs) were compared to the signal-to-noise crossover dose (SNCD), which has been suggested as the lowest dose applicable as a POD.Methods:Hill models were fit to > 10,000 in vitro concentration–response curves, obtained for > 1,400 chemicals tested as part of the U.S. Tox21 Phase I effort. BMDs and lower confidence limits on the BMDs (BMDLs) corresponding to extra effects (i.e., changes in response relative to the maximum response) of 5%, 10%, 20%, 30%, and 40% were estimated for > 8,000 curves, along with BMDs and BMDLs corresponding to additional effects (i.e., absolute changes in response) of 5%, 10%, 15%, 20%, and 25%. The SNCD, defined as the dose where the ratio between the additional effect and the difference between the upper and lower bounds of the two-sided 90% confidence interval on absolute effect was 1, 0.67, and 0.5, respectively, was also calculated and compared with the BMDLs.Results:The BMDL40, BMDL25, and BMDL18, defined in terms of extra effect, corresponded to the SNCD1.0, SNCD0.67, and SNCD0.5, respectively, at the median. Similarly, the BMDL25, BMDL17, and BMDL13, defined in terms of additional effect, corresponded to the SNCD1.0, SNCD0.67, and SNCD0.5, respectively, at the median.Conclusions:The SNCD may serve as a reference level that guides the determination of standardized BMDs for risk assessment based on HTS concentration–response data. The SNCD may also have application as a POD for low-dose extrapolation.Citation:Sand S, Parham F, Portier CJ, Tice RR, Krewski D. 2017. Comparison of points of departure for health risk assessment based on high-throughput screening data. Environ Health Perspect 125:623–633; http://dx.doi.org/10.1289/EHP408

Impacts of 17α-ethynylestradiol exposure on metabolite profiles of zebrafish (Danio rerio) liver cells

Endocrine disrupting chemicals (EDCs) that are frequently detected in bodies of water downstream from sewage treatment facilities can have adverse impacts on fish and other aquatic organisms. To properly assess risk(s) from EDCs, tools are needed that can establish linkages from chemical exposures to adverse outcomes. Traditional methods of testing chemical exposure and toxicity using experimental animals are excessively resource- and time-consuming. In line with EPA's goal of reducing animal use in testing, these traditional screening methods may not be sustainable in the long term, given the ever increasing number of chemicals that must be tested for safety. One of the most promising ways to reduce costs and increase throughput is to use cell cultures instead of experimental animals. In accordance with National Research Council's vision on 21st century toxicity testing, we have developed a cell culture-based metabolomics approach for this application. Using a zebrafish (Danio rerio) liver cell line (ZFL), we have applied NMR-based metabolomics to investigate responses of ZFL cells exposed to 17α-ethynylestradiol (EE2). This analysis showed that metabolite changes induced by EE2 exposure agree well with known impacts of estrogens on live fish. The results of this study demonstrate the potential of cell-based metabolomics to assess chemical exposure and toxicity for regulatory application.

Accelerating the Development of 21st-Century Toxicology: Outcome of a Human Toxicology Project Consortium Workshop

The U.S. National Research Council (NRC) report on “Toxicity Testing in the 21st century” calls for a fundamental shift in the way that chemicals are tested for human health effects and evaluated in risk assessments. The new approach would move toward in vitro methods, typically using human cells in a high-throughput context. The in vitro methods would be designed to detect significant perturbations to “toxicity pathways,” i.e., key biological pathways that, when sufficiently perturbed, lead to adverse health outcomes. To explore progress on the report’s implementation, the Human Toxicology Project Consortium hosted a workshop on 9–10 November 2010 in Washington, DC. The Consortium is a coalition of several corporations, a research institute, and a non-governmental organization dedicated to accelerating the implementation of 21st-century Toxicology as aligned with the NRC vision. The goal of the workshop was to identify practical and scientific ways to accelerate implementation of the NRC vision. The workshop format consisted of plenary presentations, breakout group discussions, and concluding commentaries. The program faculty was drawn from industry, academia, government, and public interest organizations. Most presentations summarized ongoing efforts to modernize toxicology testing and approaches, each with some overlap with the NRC vision. In light of these efforts, the workshop identified recommendations for accelerating implementation of the NRC vision, including greater strategic coordination and planning across projects (facilitated by a steering group), the development of projects that test the proof of concept for implementation of the NRC vision, and greater outreach and communication across stakeholder communities.

Toxicity testing in the 21 century: defining new risk assessment approaches based on perturbation of intracellular toxicity pathways.

The approaches to quantitatively assessing the health risks of chemical exposure have not changed appreciably in the past 50 to 80 years, the focus remaining on high-dose studies that measure adverse outcomes in homogeneous animal populations. This expensive, low-throughput approach relies on conservative extrapolations to relate animal studies to much lower-dose human exposures and is of questionable relevance to predicting risks to humans at their typical low exposures. It makes little use of a mechanistic understanding of the mode of action by which chemicals perturb biological processes in human cells and tissues. An alternative vision, proposed by the U.S. National Research Council (NRC) report Toxicity Testing in the 21st Century: A Vision and a Strategy, called for moving away from traditional high-dose animal studies to an approach based on perturbation of cellular responses using well-designed in vitro assays. Central to this vision are (a) “toxicity pathways” (the innate cellular pathways that may be perturbed by chemicals) and (b) the determination of chemical concentration ranges where those perturbations are likely to be excessive, thereby leading to adverse health effects if present for a prolonged duration in an intact organism. In this paper we briefly review the original NRC report and responses to that report over the past 3 years, and discuss how the change in testing might be achieved in the U.S. and in the European Union (EU). EU initiatives in developing alternatives to animal testing of cosmetic ingredients have run very much in parallel with the NRC report. Moving from current practice to the NRC vision would require using prototype toxicity pathways to develop case studies showing the new vision in action. In this vein, we also discuss how the proposed strategy for toxicity testing might be applied to the toxicity pathways associated with DNA damage and repair.

New Directions in Toxicity Testing

In 2007, the U.S. National Research Council (NRC) published a groundbreaking report entitled Toxicity Testing in the 21st Century: A Vision and a Strategy. The purpose of this report was to develop a long-range strategic plan to update and advance the way environmental agents are tested for toxicity. The vision focused on the identification of critical perturbations of toxicity pathways that lead to adverse human health outcomes using modern scientific tools and technologies. This review describes how emerging scientific methods will move the NRC vision forward and improve the manner in which the potential health risks associated with exposure to environmental agents are assessed. The new paradigm for toxicity testing is compatible with the widely used four-stage risk assessment framework originally proposed by the NRC in 1983 in the so-called Red Book. The Nrf2 antioxidant pathway provides a detailed example of how relevant pathway perturbations will be identified within the context of the new NRC vision for the future of toxicity testing. The implications of the NRC vision for toxicity testing for regulatory risk assessment are also discussed.

The Future of Toxicity Testing

In 2007, the U.S. National Research Council (NRC) released a report, “Toxicity Testing in the 21st Century: A Vision and a Strategy,” that proposes a paradigm shift for toxicity testing of environmental agents. The vision is based on the notion that exposure to environmental agents leads to adverse health outcomes through the perturbation of toxicity pathways that are operative in humans. Implementation of the NRC vision will involve a fundamental change in the assessment of toxicity of environmental agents, moving away from adverse health outcomes observed in experimental animals to the identification of critical perturbations of toxicity pathways. Pathway perturbations will be identified using in vitro assays and quantified for dose response using methods in computational toxicology and other recent scientific advances in basic biology. Implementation of the NRC vision will require a major research effort, not unlike that required to successfully map the human genome, extending over 10 to 20 years, involving the broad scientific community to map important toxicity pathways operative in humans. This article provides an overview of the scientific tools and technologies that will form the core of the NRC vision for toxicity testing. Of particular importance will be the development of rapidly performed in vitro screening assays using human cells and cell lines or human tissue surrogates to efficiently identify environmental agents producing critical pathway perturbations. In addition to the overview of the NRC vision, this study documents the reaction by a number of stakeholder groups since 2007, including the scientific, risk assessment, regulatory, and animal welfare communities.

Implementing the National Academy's Vision and Strategy for Toxicity Testing: Opportunities and Challenges Under the U.S. Toxic Substances Control Act

In 2007, the U.S. National Academy of Sciences, National Research Council (NRC), issued the report Toxicity Testing in the 21st Century: A Vision and a Strategy. This report, which was commissioned by the U.S. Environmental Protection Agency (EPA), called for the U.S. EPA to develop a new approach for how this agency evaluates the toxicity of compounds. The report recommended that the agency move from its current testing system, which is based largely on traditional toxicology, to a testing system that is based primarily on human cell lines and in vitro systems. Successful implementation of the report's vision and strategy will require that scientists, lawyers, and policymakers work together to bridge the gap among disciplines. An important step in building this bridge requires an analysis of the U.S. legal system that frames toxicity testing. If the U.S. laws, regulations, and policies erect barriers that would prevent or impede the U.S. EPA adoption of the NRC vision and strategy, it is important to identify these challenges. At the same time, if existing laws, regulations, and policies are fertile ground for these recommendations, opportunities should be documented. This article discusses and evaluates the challenges and opportunities that arise under key provisions of one major U.S. toxics law, the Toxics Substances Control Act (TSCA).

Behavioral toxicology in the 21st century: Challenges and opportunities for behavioral scientists: Summary of a symposium presented at the annual meeting of the Neurobehavioral Teratology Society, June, 2009

The National Research Council (NRC) of the National Academies of Science recently published a report of its vision of toxicity testing in the 21st century. The report proposes that the current toxicity testing paradigm that depends upon whole-animal tests be replaced with a strategy based upon in vitro tests, in silico models and evaluations of toxicity at the human population level. These goals are intended to set in motion changes that will transform risk assessment into a process in which adverse effects on public health are predicted by quantitative structure–activity relationship (QSAR) models and data from suites of high-throughput in vitro tests. The potential roles for whole-animal testing in this futuristic vision are both various and undefined. A symposium was convened at the annual meeting of the Neurobehavioral Teratology Society in Rio Grande, Puerto Rico in June, 2009 to discuss the potential challenges and opportunities for behavioral scientists in developing and/or altering this strategy toward the ultimate goal of protecting public health from hazardous chemicals. R. Kavlock described the NRC vision, introduced the concept of the ‘toxicity pathway’ (a central guiding principle of the NRC vision), and described the current status of an initial implementation this approach with the EPA's ToxCast® program. K. Crofton described a pathway based upon disruption of thyroid hormone metabolism during development, including agents, targets, and outcomes linked by this mode of action. P. Bushnell proposed a pathway linking the neural targets and cellular to behavioral effects of acute exposure to organic solvents, whose predictive power is limited by our incomplete understanding of the complex CNS circuitry that mediates the behavioral responses to solvents. B. Weiss cautioned the audience regarding a pathway approach to toxicity testing, using the example of the developmental toxicity of phthalates, whose effects on mammalian sexual differentiation would be difficult to identify based on screening tests in vitro. Finally, D. Rice raised concerns regarding the use of data derived from toxicity screening tests to human health risk assessments. Discussion centered around opportunities and challenges for behavioral toxicologists regarding this impending paradigm shift. Opportunities include: identifying and characterizing toxicity pathways; informing the conditions and limits of extrapolation; addressing issues of susceptibility and variability; providing reality-checks on selected positives and negatives from screens; and performing targeted testing and dose-response assessments of chemicals flagged during screening. Challenges include: predicting behavior using models of complex neurobiological pathways; standardizing study designs and dependent variables to facilitate creation of databases; and managing the cost and efficiency of behavioral assessments. Thus, while progress is being made in approaching the vision of 21st century toxicology, we remain a long way from replacing whole-animal tests; indeed, some animal testing will be essential for the foreseeable future at least. Initial advances will likely provide better prioritization tools so that animal resources are used more efficiently and effectively.

Bringing toxicology into the 21st century: A global call to action

Conventional toxicological testing methods are often decades old, costly and low-throughput, with questionable relevance to the human condition. Several of these factors have contributed to a backlog of chemicals that have been inadequately assessed for toxicity. Some authorities have responded to this challenge by implementing large-scale testing programmes. Others have concluded that a paradigm shift in toxicology is warranted. One such call came in 2007 from the United States National Research Council (NRC), which articulated a vision of “21st century toxicology” based predominantly on non-animal techniques. Potential advantages of such an approach include the capacity to examine a far greater number of chemicals and biological outcomes at more relevant exposure levels; a substantial reduction in testing costs, time and animal use; and the grounding of regulatory decisions on human rather than rodent biology. In order for the NRC’s and similar proposals to make a significant impact on regulatory toxicology in the foreseeable future, they must be translated into sustained multidisciplinary research programmes that are well co-ordinated and funded on a multinational level. The Humane Society is calling for a “big biology” project to meet this challenge. We are in the process of forging an international, multi-stakeholder consortium dedicated to implementing the NRC vision.

Exposure as Part of a Systems Approach for Assessing Risk

BackgroundThe U.S. Environmental Protection Agency is facing large challenges in managing environmental chemicals with increasingly complex requirements for assessing risk that push the limits of our current approaches. To address some of these challenges, the National Research Council (NRC) developed a new vision for toxicity testing. Although the report focused only on toxicity testing, it recognized that exposure science will play a crucial role in a new risk-based framework.ObjectiveIn this commentary we expand on the important role of exposure science in a fully integrated system for risk assessment. We also elaborate on the exposure research needed to achieve this vision.DiscussionExposure science, when applied in an integrated systems approach for risk assessment, can be used to inform and prioritize toxicity testing, describe risks, and verify the outcomes of testing. Exposure research in several areas will be needed to achieve the NRC vision. For example, models are needed to screen chemicals based on exposure. Exposure, dose–response, and biological pathway models must be developed and linked. Advanced computational approaches are required for dose reconstruction. Monitoring methods are needed that easily measure exposure, internal dose, susceptibility, and biological outcome. Finally, population monitoring studies are needed to interpret toxicity test results in terms of real-world risk.ConclusionThis commentary is a call for the exposure community to step up to the challenge by developing a predictive science with the knowledge and tools for moving into the 21st century.

Food for thought … on education in alternative methods in toxicology

If you cannot change things, educate those who will. Or, as George Bernard Shaw put it more cynically: “He who can, does. He who cannot, teaches.” the three authors of this Food for thought article have one thing in common: due to the generous support of the Doerenkamp-Zbinden Foundation (http:// www.doerenkamp.ch/en/), we each hold an endowed chair for alternative methods in toxicology, one at each of three prominent universities. Such chairs, established within the faculties of public health, veterinary medicine, and biology, respectively, are the basis for the development of new curricula (Wendel, 2002; Spielmann, 2002; leist, 2006). In the meantime, two more chairs have been established by the DZ Foundation in lausanne, Switzerland, and tiruchirappalli, India, but these do not focus on the field of toxicology. Many subjects considered in this article might be translated to areas other than toxicology, but this would require a case-by-case assessment. The authors have also joined forces with Alan Goldberg of Johns Hopkins University, who established the first academic center for alternative methods 28 years ago (Goldberg, 2009). together these four “ambassadors” form the transatlantic think tank for toxicology (t4) (Fig. 1) aimed at developing concepts for the paradigm shift in toxicology and creating a network for the implementation of the US National Research Council vision (NRC, 2007) of a toxicology for the 21st century (tox21c) (Hartung, 2009a). Here, aspects of teaching alternative approaches and possible synergies shall be explored. We have elaborated, in this series of articles and related publications, on the need for a paradigm shift in toxicology (Hartung and leist, 2008) and on the means to create a new approach, i.e. in vitro (Hartung, 2007a), in vivo (Hartung, 2008a) and in silico (Hartung and Hoffmann, 2009) approaches and their validation (Hartung, 2007b). An assessment of the value of 3Rs approaches has been presented (leist et al., 2008a), as well as opportunities for an evidence-based toxicology (Hartung, 2009b). the case for change has also been made on the basis of economic developments (Bottini et al., 2007; Bottini and Hartung, 2009). Some specifics of the areas of cosmetics (Hartung, 2008b) and food (Hartung and Koeter, 2008) have been addressed. A series of articles outside this Food for thought series complements the picture, e.g., mapping the implementation of tox-21c (Hartung, 2009c), discussing regulatory usefulness of new approaches (Hartung and Daston, 2009), opportunities of stem cells (leist et al., 2008b), the revolutionary character of the ongoing change (Hartung, 2008c), and the needs of ReACH (Rovida and Hartung, 2009; Hartung and Rovida, 2009). But this will not happen and cannot be implemented without changes in mindset and the efforts of people with the right skill sets to implement it. education is the means to achieve these goals. the Swiss cognitive psychologist Jean Piaget said: “the principal goal of education is to create men who are capable of doing new things, not simply of repeating what other generations have done.” A couple of questions come up immediately: Do we need education on alternatives or on a new toxicology? Do we need to create new courses and curricula or modify the current ones? Whom do we need to target, the next generation (students), current practitioners (post-graduate level), senior management (decision makers), or all of them? Which areas of study and work environments should be targeted: biology, chemistry, medicine, veterinary medicine, other life sciences, risk assessors, lawyers, policy makers, regulators, etc.? How can we achieve coverage of large numbers of target audiences? How to network and synergize in a situation of diaspora, i.e. where the different offers and competence centers are widely dispersed? Is “alternative methods” the right catch phrase or should we label it “new toxicology?” Are we teaching alternatives or are we teaching with