John R. Fowle

Perspectives on validation of high-throughput assays supporting 21st century toxicity testing

In vitro high-throughput screening (HTS) assays are seeing increasing use in toxicity testing. HTS assays can simultaneously test many chemicals but have seen limited use in the regulatory arena, in part because of the need to undergo rigorous, time-consuming formal validation. Here we discuss streamlining the validation process, specifically for prioritization applications. By prioritization, we mean a process in which less complex, less expensive, and faster assays are used to prioritize which chemicals are subjected first to more complex, expensive, and slower guideline assays. Data from the HTS prioritization assays is intended to provide a priori evidence that certain chemicals have the potential to lead to the types of adverse effects that the guideline tests are assessing. The need for such prioritization approaches is driven by the fact that there are tens of thousands of chemicals to which people are exposed, but the yearly throughput of most guideline assays is small in comparison. The streamlined validation process would continue to ensure the reliability and relevance of assays for this application. We discuss the following practical guidelines: (1) follow current validation practice to the extent possible and practical; (2) make increased use of reference compounds to better demonstrate assay reliability and relevance; (3) de-emphasize the need for cross-laboratory testing; and (4) implement a web-based, transparent, and expedited peer review process.

Lessons from Toxicology: Developing a 21st-Century Paradigm for Medical Research

Summary Biomedical developments in the 21st century provide an unprecedented opportunity to gain a dynamic systems-level and human-specific understanding of the causes and pathophysiologies of disease. This understanding is a vital need, in view of continuing failures in health research, drug discovery, and clinical translation. The full potential of advanced approaches may not be achieved within a 20th-century conceptual framework dominated by animal models. Novel technologies are being integrated into environmental health research and are also applicable to disease research, but these advances need a new medical research and drug discovery paradigm to gain maximal benefits. We suggest a new conceptual framework that repurposes the 21st-century transition underway in toxicology. Human disease should be conceived as resulting from integrated extrinsic and intrinsic causes, with research focused on modern human-specific models to understand disease pathways at multiple biological levels that are analogous to adverse outcome pathways in toxicology. Systems biology tools should be used to integrate and interpret data about disease causation and pathophysiology. Such an approach promises progress in overcoming the current roadblocks to understanding human disease and successful drug discovery and translation. A discourse should begin now to identify and consider the many challenges and questions that need to be solved.

A primer on systematic reviews in toxicology

Systematic reviews, pioneered in the clinical field, provide a transparent, methodologically rigorous and reproducible means of summarizing the available evidence on a precisely framed research question. Having matured to a well-established approach in many research fields, systematic reviews are receiving increasing attention as a potential tool for answering toxicological questions. In the larger framework of evidence-based toxicology, the advantages and obstacles of, as well as the approaches for, adapting and adopting systematic reviews to toxicology are still being explored. To provide the toxicology community with a starting point for conducting or understanding systematic reviews, we herein summarized available guidance documents from various fields of application. We have elaborated on the systematic review process by breaking it down into ten steps, starting with planning the project, framing the question, and writing and publishing the protocol, and concluding with interpretation and reporting. In addition, we have identified the specific methodological challenges of toxicological questions and have summarized how these can be addressed. Ultimately, this primer is intended to stimulate scientific discussions of the identified issues to fuel the development of toxicology-specific methodology and to encourage the application of systematic review methodology to toxicological issues.

Water disinfection: microbes versus molecules—an introduction of issues

“Water is, apart from the air one breathes, the only nutrient which is, as a matter of necessity, consumed by every human being from the first day to the last day of his existence, and it is consumed in considerably larger quantities than any other nutritional substance.”(1)

FutureTox III: Bridges for Translation

Future Tox III, a Society of Toxicology Contemporary Concepts in Toxicology workshop, was held in November 2015. Building upon Future Tox I and II, Future Tox III was focused on developing the high throughput risk assessment paradigm and taking the science of in vitro data and in silico models forward to explore the question-what progress is being made to address challenges in implementing the emerging big-data toolbox for risk assessment and regulatory decision-making. This article reports on the outcome of the workshop including 2 examples of where advancements in predictive toxicology approaches are being applied within Federal agencies, where opportunities remain within the exposome and AOP domains, and how collectively the toxicology community across multiple sectors can continue to bridge the translation from historical approaches to Tox21 implementation relative to risk assessment and regulatory decision-making.

Instruments for assessing risk of bias and other methodological criteria of animal studies : omission of well-established methods

In response to the systematic review by Krauth et al. (2013) of instruments for assessing animal toxicology studies for risk of bias and other aspects of quality, we propose the need for a broader perspective when appraising—and hopefully improving—such studies. Krauth et al. (2013) reviewed 30 instruments, 4 of which were designed for environmental toxicology studies used to evaluate human and ecological health hazards. The authors noted that these instruments were derived from preclinical pharmaceutical research in animal models. Many of these instruments focus on efficacy and not toxicity, and—as acknowledged by the authors—they may have limited potential application in environmental health research because they often have criteria that are not relevant to hazard and risk assessments. Based on these 30 instruments, Krauth et al. concluded that a limited number of risk of bias assessment criteria have been empirically tested for animal research, including randomization, concealment of allocation, blinding, and accounting for all animals. However, the authors did not discuss which elements of risk of bias criteria have been empirically tested, nor did they discuss how they were tested, leaving the reader with no information on their reliability or usefulness. We would like to bring the readers’ attention to several other important publications in environmental chemical health hazard assessment that are pertinent to this topic (Agerstrand et al. 2011; Hulzebos et al. 2010; Schneider et al. 2009), along with a U.S. Environmental Protection Agency (EPA) approach developed under the High Production Volume Challenge (U.S. EPA 1999b) as well as relevant and potentially eligible guidance developed by the U.S. EPA (1999a) and the Food and Drug Administration (FDA 2003). In addition, the majority of the procedures specified in Good Laboratory Practices and regulatory in vivo toxicity test guidelines (e.g., U.S. EPA 2013; Organisation for Economic Co-operation and Development 1999) were specifically developed to minimize systematic errors, assure high quality data and produce scientifically reliable studies. These additional publications describe design, conduct, and reporting criteria that form the basis of the methodologies employed globally to assure quality and reliability of in vivo toxicological investigations for regulatory assessment of human and ecological health hazards. Because the application of systematic review and related evidence-based approaches in toxicology is still in its infancy, it is especially important at this time to recognize the contributions of these publications. The omission of these publications by Krauth et al. could have major science policy implications. The National Toxicology Program (NTP) (whose parent organization, the National Institute of Environmental Health Sciences, funded the research of Krauth et al.) has begun relying on Krauth et al. (2013) to identify elements of risk of bias in evaluating animal studies of environmental agents as part of its systematic reviews for assessing health effects (NTP 2013a, 2013b). The reliance on criteria that have not been transparently empirically tested instead of well-established methodological criteria developed by authoritative national and international organizations could result in biased systematic reviews that ultimately lead to regulations or classifications not supported by the science. We suggest that further work is warranted in pulling together published perspectives on how to evaluate study quality in animal toxicology studies. Issues in appraising such studies for evaluating environmental hazards to humans and wildlife go well beyond those of human clinical trials, and would benefit from collaboration of experts in animal toxicology with experts in human clinical trials of medical interventions and human epidemiology.

Some Research Needs to Support Mutagenicity Risk Assessments from Whole Mammal Studies

The primary goal of regulatory agencies concerned with human health is to ensure that exposure to chemical substances does not present unreasonable risks. This involves assessing the potential of chemical substances to cause toxic effects and weighing these effects against cost and benefit considerations. If the toxicity and exposure data allow, attempts are made to estimate health damage to the human population. For most environmental chemicals, human data are unavailable. On those occasions when chemical exposure can be associated with a toxicological response in humans, many people have already been exposed and irreparable damage may have occurred. Examples of such events include the kepone incident, neurological disorders; the dibromochloropropane incident, sterility; and the vinyl chloride incident, liver angiosarcomas. Therefore, it is imperative that animal toxicity studies be conducted to assess the likely human health risk before such effects are observed.

CHAPTER 2:Regulatory Testing To Inform Decisions: National and International Requirements

The chapter attempts to convey the complexity and multi-faceted nature of regulatory safety testing to help those interested in improving regulatory testing approaches as they plan and conduct their work to make it most acceptable and useful to the regulatory authorities. National and international government agencies and key non-governmental organizations that use and/or require toxicity testing are described in the context of legislative, legal and historical factors that shaped the development and use of testing to inform decisions. Major testing requirements are described in the context of efforts to move from conventional animal testing approaches to non-animal approaches, and some of the key advances made to date are highlighted. Some of the biggest challenges that must be overcome to advance the refinement, reduction and replacement of animal tests with appropriate non-animal tests in terms of regulatory testing requirements are discussed.

Summary and Perspectives: Panel Discussion on Toxicology and Exposure Assessment: State of the Art

XPOSURE ASSESSMENT IS THE PROCESS of measuring or estimating the magnitude, frequency, E duration, and route of contact with a chemical or physical agent in the environment. Exposure assessment is part of an overall risk analysis process used by governmental and industrial organizations to decide whether various activities associated with potentially hazardous materials pose a threat to human health or the environment. (Figure 1). Although exposure assessment relies on hazard identification, it must be coupled with additional information to characterize the hazard or risk in order to gain a perspective above the significance of the exposure. Uncertainties associated with current methodologies prevent the precise identification of specific environmental and individual exposures, as well as actual intake of substances and subsequent adverse health effects. Given the present state of knowledge about interspecies extrapolation of toxic effects, it appears that major reductions in the uncertainties associated with risk assessment that are likely to be achieved in the near future will be due to improvements in our ability to estimate exposure to chemicals. This symposium reviewed the strength of techniques now available to estimate exposure and discussed what can be done in the future to improve assessment. The presentations in this symposium addressed three generic questions (Table 1). What are we exposed to? (LaGrone and Wallace); How do we estimate exposure to body tissues? (Wogan, Weston et al., Dunn); and What is the link between exposure and disease? (Chapman et al. and Anton-Culver and Burg). The state of the art of exposure measurements and recommendations to improve the science and utility of exposure assessment were discussed. Several themes were identified, including the need to improve methodology, conduct basic research, collect health surveillance data, understand the effects of multiple exposures to pollutants, collaborate among scientific disciplines, and strengthen community education and community relations efforts concerning the public’s exposure to chemicals.