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Alternative Approaches for Identifying Acute Systemic Toxicity: Moving from Research to Regulatory Testing

Jon Hamm, Kristie Sullivan, Amy J. Clippinger, Judy Strickland, Shannon Bell, Barun Bhhatarai, Bas Blaauboer, Warren Casey, David Dorman, Anna Forsby, Natàlia Garcia-Reyero, Sean Gehen, Rabea Graepel, Jon Hotchkiss, Anna Lowit, Joanna Matheson, Elissa Reaves, Louis Scarano, Catherine Sprankle, Jay Tunkel, Dan Wilson, Menghang Xia, Hao Zhu, David Allen.
Toxicology in Vitro (2017) DOI: PMID: 28069485



Acute systemic toxicity testing provides the basis for hazard labeling and risk management of chemicals. A number of international efforts have been directed at identifying non-animal alternatives for in vivo acute systemic toxicity tests. A September 2015 workshop, Alternative Approaches for Identifying Acute Systemic Toxicity: Moving from Research to Regulatory Testing, reviewed the state-of-the-science of non-animal alternatives for this testing and explored ways to facilitate implementation of alternatives. Workshop attendees included representatives from international regulatory agencies, academia, nongovernmental organizations, and industry. Resources identified as necessary for meaningful progress in implementing alternatives included compiling and making available high-quality reference data, training on use and interpretation of in vitro and in silico approaches, and global harmonization of testing requirements. Attendees particularly noted the need to characterize variability in reference data to evaluate new approaches. They also noted the importance of understanding the mechanisms of acute toxicity, which could be facilitated by the development of adverse outcome pathways. Workshop breakout groups explored different approaches to reducing or replacing animal use for acute toxicity testing, with each group crafting a roadmap and strategy to accomplish near-term progress. The workshop steering committee has organized efforts to implement the recommendations of the workshop participants.


Figure 1. Proposed prioritization strategy for testing of chemicals for acute toxicity.

Adapted from (National Research Council, 2015).

Figure 2. Schematic representing the use of alternatives to move away from animal tests.

Putting the pieces together involves multiple approaches and requires increased harmonization and cooperation among regulatory agencies and industry.

Figure 3. A: Sandalore; B: 3-cyclopentene-1-butanol, beta,2,2,3-tetramethyl-.

Figure 4. Overview of the AOP framework.

Adapted from Oki et al., 2016

Figure 5. Tox21 involved the testing of > 10.000 substances.

Tox21 involved the testing of > 10.000 substances using the automated robotic screening system housed at NCATS. During Phase II of testing, the assay is optimized, validated, and miniaturized into a 1536-well plate format, followed by the robotic validation. Robotic qHTS against the Tox21 10K library is then run, followed by data processing. The screening results are first shared by government partners, and then are made publicly accessible. Figure reproduced from (Attene-Ramos et al., 2013).


Table 1. List of webinar presentations held prior to the workshop.

Table 2. Regulatory statutes governing systemic toxicity data.

Regulatory statutes governing systemic toxicity data at the U.S. Consumer Product Safety Commission.

Table 3. Selected mechanisms of acute toxicity.1

This table provides an outline of the some of the known mechanisms involved in acute systemic toxicity along with prototypical initiators. In some cases, the exact molecular initiating event (MIE) isn't known. Examples of adverse outcome pathways (AOPs) under development in the OECD AOP Wiki are noted and can be found on the web: a) b) c)

Table 4. Acute systemic toxicity data are often used to set a variety of reference values.

Acute systemic toxicity data are often used to set a variety of reference values relevant to workers, consumers, or emergency response professionals.