OECD TG 443 is the “Extended One-Generation Reproductive Toxicity Study” (EOGRTS) standard, i.e., the extended one-generation reproductive toxicity study, which is included in Section 4 (Health Effects) of the OECD Test Guidelines for the Testing of Chemicals. Adopted in 2011 and updated in 2018, the guideline is designed as a testing standard intended to replace the previous two-generation studies (TG 415 and TG 416) and offers a more flexible cohort structure as well as a more comprehensive list of endpoints.
The basic principle of the test is to administer the test substance to the parental generation (P) beginning before mating, to mate these animals, and to expose the F1 generation to the substance both in utero and through the mother’s milk. If necessary, the F1 offspring may also be given a direct dose after weaning. The study is typically conducted on rats.
The key innovation of the EOGRTS is its cohort-based design. The cohorts are activated or deactivated based on conditional triggers:
Cohort 1A (Reproduction and Development Cohort):
- Parameters Related to Mating, Fertility, and Conception
- Estrous cycle, sperm parameters
- Data on Pregnancy, Birth, and Lactation
- Postnatal Development in F1 Offspring (Body Weight, Viability)
- Anogenital Distance (AGD) and Number of Teats
- Pubertal Development (Vaginal Opening, Foreskin Separation)
- Thyroid hormone levels (T4, TSH)
- Weights and Histopathology of the Reproductive Organs and Related Organs
Cohort 1B (optional reproductive cohort):
- Additional cohort without extended histopathology, focusing on reproductive functions
- Used to enhance data reliability in certain regulatory contexts
Cohort 2A (Developmental Neurotoxicity – DNT):
- Motor activity, acoustic startle reflex (auditory startle reflex)
- Learning and memory tests (e.g., Morris water maze)
- Brain Weight and Neuropathological Examination
- Is activated only in the event of a structural alert or when triggered by previous data
Cohort 2B (optional DNT cohort):
- Behavioral Assessment Without Neuropathology
Cohort 3 (Developmental Immunotoxicity – DIT):
- T-cell-dependent antibody response (TDAR)
- Immunophenotyping
- Natural Killer Cell Activity
- Is activated only if there is evidence of immunotoxicity
This cohort structure is designed to significantly reduce animal use compared to two-generation studies (the literature reports a reduction of up to approximately 40%) while simultaneously expanding the scientific scope. The establishment of the cohorts is based on the structural properties of the test substance, previous toxicity data, and regulatory requirements.
The study design includes a negative control group, a solvent control group, and, if applicable, a positive control group.
Why is it performed?
OECD TG 443 is being conducted for the following scientific and regulatory purposes:
- Comprehensive Characterization of Reproductive and Developmental Toxicity: Holistic Assessment of Fertility, Pregnancy, Lactation, and F1 Development in a Single Study
- Study of Endocrine Effects: Assessment of endocrine activity based on sensitive endpoints such as AGD, number of nipples, timing of puberty, and thyroid hormone levels
- Assessment of Developmental Neurotoxicity (DNT): Evaluation of Neurodevelopmental Effects Using Behavioral and Neuropathological Endpoints
- Assessment of Developmental Immunotoxicity (DIT): Investigation of Immune System Development Using TDAR and Immunophenotyping
- Support with Regulatory Documentation: Use in REACH, Pesticide, Biocide, and Drug Safety Assessments
- Reducing the Need for Animal Testing (3Rs): Obtaining More Comprehensive Data Using Fewer Animals Compared to Two-Generation Studies
- "Weight-of-Evidence" Approach: A Contribution to Risk Assessment in Conjunction with Other Toxicity Data, Physicochemical Properties, and an Exposure Assessment
As part of our nano-testing services, we conduct studies in accordance with OECD Guideline TG 443 with scientific rigor, utilizing our accredited laboratory infrastructure and our quality management system. Our scope of services includes the following:
Study Design and Protocol Development
Preparation of a test protocol and a cohort design in accordance with OECD TG 443
Determination of the dosage range and the main test design based on the physicochemical properties of the test substance and regulatory requirements
Scientific Rationale for the Triggers for Cohort 2A (DNT) and Cohort 3 (DIT)
Specification of the negative control, the solvent control, and, if applicable, appropriate positive control groups; scientific justification for the animal species, route of administration, dosages, and duration of administration; compliance with requirements regarding ethical approvals and animal welfare