Analysis of current reproductive toxicity assessments to reduce animal use

Applicant:Troy Seidle, MSc.
11 Brentwood Avenue                                                         
Kitchener, Ontario N2H 2C4

Under the EU REACH chemicals regulation, assessments of reproductive/developmental toxicity will be a key component for the risk assessment of chemicals. Recent impact studies have demonstrated that new toxicity testing for these endpoints using currently available methods will account for nearly 80% of all animals used (up to 18 million rats and rabbits) for fulfilling REACH information requirements. Moreover, there are currently no methods in sight that totally replace this kind of safety testing in animals in the short- to mid-term. Consequently, new testing strategies using reduction and refinement methods, as well as integrating partial replacement methods that have already been validated, must be developed and implemented without delay.

Reduction methods
The largest consumer of animals is the two-generation reproductive toxicity study, which uses up to 3,200 animals per substance tested. Two recent retrospective analyses of more than 350 two-generation studies have analyzed the added value of the second generation. It could be demonstrated that the breeding of a second generation affected neither the study’s “no-effect level” (NOAEL) nor critical effects, and consequently has no impact on regulatory decisions. ECVAM has therefore initiated a revision of the current OECD test guideline for one-generation studies in collaboration with the US Environmental Protection Agency (EPA) and the German Federal Institute for Risk Assessment (BfR). Additionally, a guidance document on the use of an “extended” one-generation study will be produced within this OECD project. The initial design for the extended 1-generation study was proposed by the HESI Task Force on Agricultural Chemical Safety Assessment (ACSA), and is currently being validated under the auspices of the OECD Validation Management Group for mammalian test methods (VMG-mam). The European Partnership for Alternative Approaches to Animal Testing (EPAA) is also an active participant in defining triggers/waiver for additional animal tests attached to the extended one-generation study, such as developmental toxicity, neurotoxicity, immunotoxicology, and endocrine parameters.

Replacement methods
Within the EU’s 6th Framework Programme, ECVAM has proposed an integrated project that is aiming to the integrate existing in vitro models and newly developed models into an “integrated testing strategy” (ITS) that will provide detailed information on the hazards of compounds to the mammalian reproductive cycle. The project named ReProTect has been funded since 2004 with 9 million   by the Commission and is starting to deliver novel tests for validation in the field of reproductive toxicology. It aims to develop a testing strategy using least animals. The testing strategy will not only decrease the number of animal tests in this area of toxicology, but should also provide more detailed information concerning toxicological mechanisms in different target tissues, thus providing valuable information for lead compound optimization as well as the hazard identification of substances. 

A key part of this project shall be an analysis of existing two-generation studies with the question: What would have happened if they had been stopped after one generation? This could reduce animal use from 3,200 to 1,400 rats per study. However, the validation of an extended one-generation study is complex, given that criteria for triggering and/or waiving additional study segments have yet to be clearly defined or validated. To this end, existing industry data provided through the EPAA shall be analysed and reports elaborated as needed. Additionally, detailed analyses of reproductive/developmental toxicants will be necessary in order to assemble meaningful batteries of in vitro tests within the ReProTect framework. This work will include prevalence analysis of sub-endpoints as well as understanding of toxicological mechanisms with a high prevalence. A pilot study of this nature has already been carried out by the candidate (attached). These activities will be carried out in collaboration with the head of ECVAM’s reproductive toxicity division and will require approximately two years.
This project will also involve regular participation in 3Rs-related European and international policy discussions, including participation in meetings, workshops, conferences, Commission inter-service and OECD consultations, etc. 

Hosting key area
ECVAM has offered to host this project in its reproductive toxicity division. A member of this division has been nominated by the EPPA working group on “Prioritisation, promotion and implementation of future research based on the application of the 3Rs” as an external expert in order to support the validation of the extended one-generation study. This division is also involved in an ECETOC group that is working to define triggering and waiving criteria for extensions to the one-generation study. In addition, members of ECVAM’s reproductive toxicity division are participating in an OECD expert group on the validation of the extended one-generation study, as well as in meetings of the OECD VMGs on mammalian and non-animal test methods and of the National Coordinators of the OECD Test Guidelines Programme. The scientific/technical management of ReProTect is also based in this division. In conclusion, ECVAM is able to provide the ideal working environment to implement this important project.