It has been promised for a hell of a long time, but are we now close to an era when a chemical’s toxic potential can be comprehensively defined without exploiting fellow sentient creatures. As well as this obvious ethical gain, alternative test methods to animal testing have the equally attractive potential to reduce the times and costs associated with safety testing.
Alternatives to Animal Lab Experiments
Techniques to replace, reduce and refine animal-based test methods (the “3Rs” principle) include read-across and New Approach Methodologies (NAMs) involving experimental work:
- in silico
- in chemico
- other in vitro
As opposed to laboratory animal studies focusing on endpoints like tissue damage, NAMs are being developed based on more of a mechanistic understanding of the pathways that might follow a chemical exposure to result in a particular adverse outcome in humans.
Advancements in Cosmetics and Chemical Regulation
This is all of particular relevance for cosmetics, for which animal toxicity testing is increasingly considered beyond the pale around the world. As of March 2013, for example, cosmetics tested on animals are no longer permitted to be marketed in the EU (EC, 2013) and, following Brexit, this restriction has been maintained in the UK (UK OPSS, 2023a,b). In the realm of industrial chemicals, the European Chemicals Agency (ECHA) has said that laboratory animal testing should be considered only as a “last resort”, when there is no other way to gather safety information (ECHA, 2023). Under REACH, in vitro test methods are now the default for determining skin and eye irritation, and skin sensitisation (ECHA, 2017). In addition, an analysis of REACH data up to 2022 found that, even for endpoints where laboratory animal testing is, or was, the standard, most registrants have used alternative methods to try to satisfy toxicological data requirements. The most common adaptation was read-across, followed by data waivers, weight of evidence (combining different alternative approaches), and in silico (Quantitative) Structure-Activity Relationships, or (Q)SARs (ECHA, 2023).
The Use of Read-Across & Threshold of Toxicological Concern (TTC) Strategies
Read-Across
The frequent use of read-across, where the properties of an untested chemical are predicted using available data on a related substance, or group of substances, is not just limited to industrial chemicals. While we at bibra have indeed put together numerous read-across strategies under REACH, we also use these principles in our evaluations for risk assessments in other sectors. This might involve identifying a structurally similar “analogue” chemical with test data, performing group evaluations of similar chemicals like fatty acids, or looking at the toxic potential of tested metabolites or precursors. As such, our desk-based literature searches have avoided the need for laboratory animal testing, even when no data are available on the exact substance of interest.
Threshold of Toxicological Concern (TTC)
The Threshold of Toxicological Concern (TTC) approach for risk assessment is based on similar assumptions, where several (typically very low) generic tolerable TTC exposures have been set for untested chemicals, based on the toxicological properties of a universe of tested chemicals. TTC-based assessments have, in our experience, also been particularly helpful for the evaluation of chemicals that have not been fully characterised, such as some extractables and leachables.
In Silico
In Silico (Q)SAR Assessments
Along the same lines, (Q)SAR-based assessments are driven by the idea that test data on chemicals with certain structural or physicochemical characteristics can be used to inform on the likely effects of untested chemicals with the same features. Various models and software are available for this in silico work, including the OECD QSAR Toolbox, which the Organisation for Economic Cooperation and Development developed in partnership with ECHA, Toxtree, DEREK and the FDA-favoured Leadscope. These can be invoked, for example, in evaluations of pharmaceutical impurities identifying “alerting structures” for mutagenicity, including a consideration of whether the same alerts are present (or not) in the structure of the active. For more complex endpoints such as repeated-dose toxicity, carcinogenicity, and reproductive/developmental toxicity, (Q)SAR models are currently only of limited value in isolation, but can be used to help put together, strengthen, and justify read-across approaches and weight-of-evidence arguments. No doubt, computational methods of data gap filling will be in the front of the Artificial Intelligence queue.
In Vitro & In Chemico Testing
Moving away from the office and into the laboratory, in vitro and in chemico testing can, of course, bring something important to the 3R party. While we have seen the results of in vitro testing in the field of genotoxicity for many years, with the Ames test first developed back in the 1970s, more recent alternative test successes have focused on local toxicity and sensitisation. About 50% of the studies conducted since 1990 for skin and eye irritation in the REACH database invoked an in vitro methodology. That number rises to 90% of those studies performed from 2019-2022 (ECHA, 2023). Developments from OECD have also been very helpful, notably the various Defined Approaches (DAs) and Integrated Approaches to Testing and Assessment (IATAs), as well as the Adverse Outcome Pathway (AOP) for skin sensitisation (OECD, 2012), and related guidelines for in vitro and in chemico (i.e., the Direct Peptide Reactivity Assay, or DPRA) test methods.
Industry-Wide Adoption of Alternatives to Animal Testing
Whilst the adoption of more “alternative” approaches to chemical safety testing has been much slower than the aficionados have long predicted, steady progress continues. At bibra, we have been using desk-based strategies such as read-across, TTC and (Q)SAR to minimise the need for animal testing for many years, and we are always on the lookout for key updates in the regulatory acceptance of new approaches and methodologies. If you are also that way inclined, you may find a free subscription to TRN very useful! If you would like to discuss this topic further, sign up for TRN, or have any questions on how we might be able to help you avoid or minimise laboratory animal testing, please get in touch and we’d be very happy to help.
References
EC (2013). European Commission. Communication from the Commission to the European Parliament and the Council on the animal testing and marketing ban and on the state of play in relation to alternative methods in the field of cosmetics. https://eur-lex.europa.eu/legal-content/EN/TXT/?uri=CELEX:52013DC0135
ECHA (2017). European Chemicals Agency. Guidance on Information Requirements and Chemical Safety Assessment. Chapter R.7a: Endpoint specific guidance. Version 6.0. July 2017. https://echa.europa.eu/documents/10162/13632/information_requirements_r7a_en.pdf/e4a2a18f-a2bd-4a04-ac6d-0ea425b2567f
ECHA (2023). European Chemicals Agency. The use of alternatives to testing on animals for the REACH Regulation. Fifth report under Article 117(3) of the REACH Regulation. June 2023. https://echa.europa.eu/documents/10162/23919267/230530_117_3_alternatives_test_animals_2023_en.pdf/9cfc291e-9baf-ffa2-466c-2bc2c6f06b8e?t=1685428213290
OECD (2012). Organisation for Economic Cooperation and Development. The Adverse Outcome Pathway for Skin Sensitisation Initiated by Covalent Binding to Proteins. Part 1: Scientific Evidence. Series on Testing and Assessment No.168. ENV/JM/MONO(2012)10/PART1. https://www.oecd-ilibrary.org/docserver/9789264221444-en.pdf?expires=1697031111&id=id&accname=guest&checksum=C8FA7CD1F0C248BF5AA69F5A55065535
UK OPSS (2023a). UK Office for Product Safety and Standards. Statutory guidance. Regulation 2009/1223 and the Cosmetic Products Enforcement Regulations 2013: Great Britain. Updated 4 May 2023. https://www.gov.uk/government/publications/cosmetic-products-enforcement-regulations-2013/regulation-20091223-and-the-cosmetic-products-enforcement-regulations-2013-great-britain
UK OPSS (2023b). UK Office for Product Safety and Standards. Statutory guidance. Regulation 2009/1223 and the Cosmetic Products Enforcement Regulations 2013: Northern Ireland. Updated 4 May 2023. https://www.gov.uk/government/publications/cosmetic-products-enforcement-regulations-2013/regulation-20091223-and-the-cosmetic-products-enforcement-regulations-2013-northern-ireland