Policies and Guidance for Implementation of Alternative Methods

To encourage adoption and use of NAMs, the 2018 ICCVAM Strategic Roadmap called on agencies to provide clear guidance on use and acceptance of data from these approaches. During 2020 and 2021, agencies issued guidance for use of NAMs in the areas of testing for skin sensitization, carcinogenicity, and developmental and reproductive toxicity, as well as for the testing of medical devices, vaccines, cancer drugs, pesticides, and consumer products.

Measuring Progress Toward Implementation of Alternative Methods in Toxicity Testing

In September 2019, the U.S. Government Accountability Office issued a report, “Animal Use in Research: Federal Agencies Should Assess and Report on Their Efforts to Develop and Promote Alternatives.” The report describes how the U.S. Department of Health and Human Services, USDA, and EPA ensure researchers consider the use of alternatives to animals and examines the steps the agencies have taken to facilitate the use of alternative research methods and to assess the effect of their efforts on animal use. The report recommended that ICCVAM establish a workgroup to develop metrics that ICCVAM member agencies could use to assess progress made toward reducing, refining, or replacing animal use in testing. Furthermore, the report recommended that such metrics be incorporated into ICCVAM Biennial Progress Reports. In response, ICCVAM established its Metrics Workgroup in early 2020. The workgroup included members from nine ICCVAM agencies. Its charge was to determine how agencies can best address the Government Accountability Office report’s recommendations within the context of the ICCVAM Authorization Act.

In March 2021, the ICCVAM Metrics Workgroup published “Measuring U.S. Federal Agency Progress Toward Implementation of Alternative Methods in Toxicity Testing,” which describes its findings and recommendations. The workgroup’s key finding was that no one set of metrics can be used by all ICCVAM member agencies to assess progress toward reducing, refining, or replacing animal use in testing. The workgroup instead recommended that each agency develop its own metrics that are relevant and practical to their unique situation. This document describes the recommendations of the ICCVAM Metrics Workgroup along with references and other materials that can be used to follow federal agency progress in promoting the use of alternative toxicological methods.

In response to the Metrics Workgroup’s recommendations, ICCVAM agencies have developed webpages to inform their stakeholders about progress on adoption of alternatives and reduction of animal use. Links to those pages are available on the NTP website. Activities to assess progress toward reducing, refining, or replacing animal use in testing are also described throughout this report.

Characteristics to Consider when Selecting a Positive Control Material for an In Vitro Assay

The use of in vitro assays to inform decision-making requires robust and reproducible results across studies, laboratories, and time. Experiments using positive control materials are integral to demonstrating the extent to which a measurement system is performing as expected. A paper by CPSC, NIEHS, and NIST scientists and collaborators (Petersen et al. 2021) reviewed 10 characteristics that should be considered when selecting a positive control material for an in vitro assay. These include: (1) the biological mechanism of action, (2) ease of preparation, (3) chemical purity, (4) verifiable physical properties, (5) stability, (6) ability to generate responses spanning the dynamic range of the assay, (7) technical or biological interference, (8) commercial availability, (9) user toxicity, and (10) disposability. The paper presented examples and a case study of the monocyte activation test to demonstrate the application of these characteristics for identification and selection of potential positive control materials. Because specific positive control materials are often written into testing standards for in vitro assays, selection of the positive control material based on these characteristics can aid in ensuring the long-term relevance and usability of these standards.

Draft Guidance on Carcinogenicity Testing

In an October 2021 Federal Register notice (86 FR 54982), FDA announced draft guidance for industry, “S1B(R1) Addendum to S1B Testing for Carcinogenicity of Pharmaceuticals.” The draft guidance expands the testing scheme for assessing human carcinogenic risk of small molecule pharmaceuticals. It introduces an integrative approach that provides specific weight-of-evidence criteria that inform whether a 2-year rat study adds value in completing a human carcinogenicity risk assessment. The draft guidance also adds a plasma exposure ratio-based approach for setting the high dose in the rasH2-Tg mouse model. FDA accepted comment on the draft guidance through December 2021.

Adoption of OECD Guideline on Defined Approaches for Skin Sensitization Testing

In June 2021, OECD issued Guideline 497, Defined Approaches on Skin Sensitisation. Drafted and sponsored by ICCVAM agency scientists and international partners, Guideline 497 is the first internationally harmonized guideline to describe a non-animal defined approach that can be used to replace an animal test to identify skin sensitizers. Validation against a curated set of human data indicates that this approach predicts human skin sensitization hazard better than the accepted animal test. NICEATM and ICCVAM are collaborating with stakeholder groups to convene events to raise awareness about the guideline and promote best practices for implementation of defined approaches to skin sensitization testing and assessment.

Draft Risk Assessment for Chlorothalonil

In a May 2021 Federal Register notice (86 FR 27593), EPA announced availability of draft human health and/or ecological risk assessments for several pesticides. EPA accepted public comment on the risk assessments through September 2021.

One of the pesticides covered by this announcement was the fungicide chlorothalonil, which was the subject of a case study to use an in vitro model to develop an inhalation risk assessment. The case study will be published in an OECD guidance document in 2022. Data from the in vitro model were used in conjunction with human dosimetry modeling to evaluate human inhalation risk, which is included in the document “Chlorothalonil: Revised Human Health Draft Risk Assessment for Registration Review.” This and other documents relevant to the chlorothalonil evaluation are available in a docket on Regulations.gov.

Proposed Guidance for Industry for Federal Hazardous Substances Act Testing

In a March 2021 Federal Register notice (86 FR 16704), CPSC requested comment on its “Proposed Guidance for Industry and Test Method Developers: CPSC Staff Evaluation of Alternative Test Methods and Integrated Testing Approaches and Data Generated from Such Methods to Support Federal Hazardous Substances Act (FHSA) Labeling Requirements.” CPSC has developed this guidance, building on its Animal Testing Policy, to assist stakeholders in determining what test methods are deemed reliable for determining compliance with the labeling requirements under the Federal Hazardous Substances Act. This includes clarification of CPSC informational requirements and a process for evaluating NAMs and IATA. CPSC accepted comments on the draft guidance through June 2021 and will release the final guidance in 2022.

EPA List of NAMs for TSCA Information Requirements

A 2016 update of the Toxic Substances Control Act (TSCA) required EPA to issue a list of methods and approaches that do not use vertebrate animals (i.e., NAMs) to develop new data or information required under TSCA. In February 2021, EPA updated the list of NAMs that the agency will consider for the purpose of satisfying information requirements under TSCA. Some updates reflected changes made to OECD test guidelines, while other updates considered guidance on acute systemic toxicity testing waivers issued by the EPA Office of Pesticide Programs and availability of a new expert system to predict carcinogenicity of organic chemicals, fibers, metals, and polymers.

The NAMs list is an element of EPA’s 2018 “Strategic Plan to Promote the Development and Implementation of Alternative Test Methods Within the TSCA Program.” Information about the NAMs list and the Strategic Plan is available on the EPA website.

Draft Risk Assessment for Isothiazolinones

In May 2020, EPA requested comment (85 FR 28944) on draft human health and ecological risk assessments for a group of antimicrobial chemicals known as isothiazolinones. EPA accepted comments on the draft risk assessments through November 2020.

Isothiazolinones are used in a variety of products including plastics, household cleaners, and laundry detergents. They frequently cause skin sensitization. EPA requested comment on the use of an in vitro and artificial neural network-based defined approach instead of using laboratory animal data to evaluate skin sensitization risks for products containing isothiazolinones. This is the first consideration of this type of defined approach in regulatory risk assessment.

The draft risk assessments rely heavily on work done by NIEHS and ICCVAM. The in vitro testing was conducted by the Toxicology Branch of the NIEHS Division of NTP (DNTP). NICEATM analyzed the in vitro data and ran the artificial neural network-based defined approach to provide quantitative potency predictions used to determine points of departure. The ICCVAM Skin Sensitization Expert Group reviewed the DNTP testing report and the NICEATM analyses before data were provided to EPA for development of the risk assessments.

Guidance on Nonclinical Evaluation of Immunotoxic Potential

In February 2020, FDA issued draft guidance on “Nonclinical Safety Evaluation of the Immunotoxic Potential of Drugs and Biologics.” This guidance supplements previously issued recommendations on nonclinical evaluations of immunotoxic potential and is intended to assist sponsors in such evaluations. The guidance includes several specific recommendations on assessing potential for dermal sensitization:

  • FDA no longer recommends that sponsors conduct the murine local lymph node assay to assess the sensitization potential of topical drug products due to the limitations of the assay.
  • As an alternative screen for skin sensitization for individual chemicals, FDA will consider a battery of in silico, in chemico, and in vitro studies that have been shown to adequately predict human skin sensitization with an accuracy similar to existing in vivo methods.
Guidance on In Vitro Drug Interaction Studies

In January 2020, FDA published “In Vitro Drug Interaction Studies – Cytochrome P450 Enzyme- and Transporter-mediated Drug Interactions Guidance for Industry.” This guidance is intended to help drug developers plan and evaluate studies to determine the drug-drug interaction potential of an investigational drug product. It focuses on in vitro approaches to evaluate the interaction potential between investigational drugs with cytochrome P450 enzymes and transporters, as well as how in vitro results can inform future clinical drug-drug interaction studies. The appendices of this guidance include factors to consider when choosing in vitro experimental systems, key issues regarding in vitro experimental conditions, and more detailed explanations regarding model-based drug-drug interaction prediction strategies.

Guidance on Reproductive and Developmental Toxicity Studies for Human Pharmaceuticals

In May 2021, FDA published “S5(R3) Detection of Reproductive and Developmental Toxicity for Human Pharmaceuticals - Guidance for Industry” This guidance is intended to help drug developers plan and evaluate nonclinical developmental and reproductive toxicity studies necessary to support human clinical trials and marketing authorization for pharmaceuticals. The 2021 revisions to this guidance describe testing strategies utilizing alternative assays for the assessment of malformations and embryofetal lethality. The guidance also provides basic principles that will assist in the development, qualification, and potential regulatory use of alternative assays for evaluating adverse effects on embryofetal development.

Amendment of Hazardous Materials Regulations to Consider In Vitro Methods for Classification of Corrosives

In August 2021, the Pipeline and Hazardous Materials Safety Administration (PHMSA) of the U.S. Department of Transportation published a Notice of Proposed Rulemaking (86 FR 43844) to amend certain sections of the Hazardous Materials Regulations (49 CFR parts 171 to 180). The purpose of the amendments is to maintain alignment with international regulations and standards relevant to the transportation of hazardous materials. The proposed amendments build on international collaborations of PHMSA with the United Nations to update the classification criteria for Class 8 (corrosive) substances to include in vitro non-animal test methods. The rulemaking incorporates the 2016 version of OECD Test No. 431, In Vitro Skin Corrosion: Reconstructed Human Epidermis (RHE) Test Method, used to assign a packing group to Class 8 materials. The rulemaking also provides an opportunity to assign certain Class 8 materials to a more conservative packing group in lieu of additional in vivo testing, reducing the need for animal testing to transport corrosive materials safely. The public comment period for the proposed rulemaking ended in October 2021 and the final rule is now being reviewed.