Points to Consider for Designing and Performing a Juvenile Animal Study (JAS)
By: Bassem Attalla, Bsc
Why Are Juvenile Animal Studies Needed?
Research shows inherent biological differences between the biology of adult and juvenile subjects. Juvenile subjects are not “smaller adults”. As subjects grow towards adulthood, their biological system matures and can react to therapeutic agents very differently at various stages of growth (neonates, infants, juveniles, or adolescents). The difference between a mature and an immature system (including the target organs) is a key component for consideration when evaluating therapeutic agents’ efficacy and toxicity. Additionally, therapeutic agents’ absorption, distribution and excretion differ according to subjects’ age.
Guidelines from various regulatory bodies such as the OECD, FDA, and EMA require that therapeutic agents intended for pediatric use or potentially affecting organs’ growth must undergo age-matched, nonclinical testing given the rapid evolution of some organs and body systems at young age.
Juvenile Animal Studies are Required When:
- Therapeutics agents are intended for pediatric use in the clinic.
- The therapeutic agent has a potential effect on organ or system’s growth.
- The therapeutic agent may affect the endocrine or neurological system.
Which Species Should be Selected for a Juvenile Animal Study?
Juvenile animal studies (JAS) can be performed using rodent or large animal species. Generally, only one species is required by regulators. Approximately 70% of nonclinical studies are performed using a rodent species, mainly rats, given their larger size compared to mice. Large animal species such as nonhuman primates or dogs may be used. However, a sound justification for their use, instead of rodents is generally expected.
At What Age Should Dose Administration Start?
Target organ(s) for the given therapeutic must be identified. Organs develop at different rates until they reach full maturity. Development stages in rats are shown in figure 1. Approximate postnatal development periods of key systems are shown in figure 2. Dose administration start should match the lowest target age in the clinic population, while taking into account the maturation process of the target organ(s). ITR Laboratories has developed capabilities to administer formulations starting Post-Natal Day 1 for parenteral routes and Post-Natal Day 4 for oral routes.
Considerations When Designing a Juvenile Animal Study:
- Study Design & Duration
- The study duration should cover both the period of clinical exposure and the relevant stages of target organ development.
- Animal Selection & Logistics
- Animal numbers must be sufficient to generate meaningful data while accounting for the fragility of developing organ systems.
- Multiple cohorts may be required due to the small size of young pups.
- JAS are logistically complex, involving challenges such as animal ordering schedules, handling and identification of small pups, managing litter dynamics (including dams), and conducting procedures like clinical observations and blood collection.
- Given this complexity, experienced CROs are critical to ensure proper execution and reliable data interpretation.
- Identification & Randomization
- Standard identification methods are often unsuitable for young pups due to their fragility. Alternative approaches, such as digit tattooing systems, can enable reliable identification of large groups.
- Randomization within litters presents unique challenges. A hybrid approach maintaining uncontaminated concurrent controls while ensuring representation of all dose levels within each litter can help reduce genetic bias.
- Sampling & Toxicokinetics
- Due to limited blood volume, toxicokinetic and other blood samples are typically collected terminally.
- Efficient scheduling of sampling procedures is essential for study success.
- Endpoints & Data Interpretation
- Certain standard toxicology endpoints (e.g., baseline ophthalmology, single-dose clinical pathology) may not be feasible due to animal fragility.
- Early-life variability can impact data interpretation, requiring experienced scientific oversight.
- Developmental Milestones
- Weaning typically occurs on Postnatal Day 21, after which pups are rehoused and food consumption measurements can begin.
- Necropsy Considerations
- Necropsy in very young pups can be challenging due to tissue fragility.
- For early timepoints, fixation of the entire carcass prior to tissue collection is recommended to preserve organ integrity.
Conclusion
Regulations require drugs intended for use on pediatric patients, or having the potential to affect the growth, to be tested nonclinically in age-matched patients.
Rats are generally the species of choice, but large animals such as dogs or non human primates may also be used, with adequate justification. The number of animals to be used, and the endpoints to be assessed must be carefully planned ahead of time, knowing that most blood samples can only be collected terminally and that some datapoints cannot be collected due to pups’ small size. CROs expertise in performing and interpreting endpoints from JAS is critical for sponsors to consider, since performing a JAS is logistically challenging.
Over the previous years, ITR has developed highly specialized techniques to administer formulations to rat pups as young as the day after parturition. This is equivalent to premature-born babies patients in the clinic, thus allowing therapeutic agents to be tested in age-matched rats as the youngest age possible. ITR also developed expertise to adequately identify and house pups, thus allowing to collect and interpret pertinent datapoints at a very young age. ITR additionally developed its own control reference data for pups at different ages for several key datapoints.
About the Author:
Bassem Attalla holds a B.Sc. in Microbiology and Immunology from the University of Montreal and brings over 20 years of experience in preclinical toxicology. His expertise spans general, inhalation, and safety pharmacology studies, with a strong focus on juvenile toxicology. As Senior Director, Scientific Advisory & Business Development at ITR Laboratories, he supports sponsors in designing and advancing nonclinical programs, including those for pediatric drug development.