Hexarelin and other drugs show human-like metabolism in a novel zebrafish water tank model

Studying drug metabolism is a critical, complex, and expensive part of pharmaceutical development. Traditional models, like rodent studies or in vitro assays, have limitations in cost, throughput, or physiological relevance. There's a persistent need for alternative models that are faster, cheaper, and can accurately predict human biotransformation pathways. The Zebrafish (Danio rerio) Water Tank (ZWT) model is being explored as one such alternative. This approach leverages the whole organism's metabolic machinery in a simple aquatic environment. The primary challenge is validating whether this unconventional model can replicate the complex metabolic pathways, including stereospecificity and the activity of specific enzyme systems like the Cytochrome P450 family (e.g., CYP2B6, CYP2C9), seen in mammals and humans for a diverse range of compounds.

The study investigated the Zebrafish Water Tank (ZWT) model using six experiments with four model compounds: sibutramine, selegiline, JWH-073, and the peptide hexarelin. For optimization, 18 fish were used per experiment with an experimental duration of 168 hours. The researchers used sibutramine to optimize ZWT conditions and compare its metabolism against human and mouse data. They also investigated the role of CYP2B6 using inhibition. Selegiline was used to test stereospecificity, and JWH-073 was used to evaluate cannabimimetic metabolism. Finally, hexarelin was introduced to the ZWT to assess the model's suitability for drugs with low stability.

The ZWT model successfully replicated key aspects of human metabolism for the tested compounds. After optimization, a minimum of 18 fish and 168 hours was required to produce a relevant panel of biotransformation products. While the same hydroxylated metabolites of sibutramine were found in ZWT, humans, and mice, there were differences in metabolite concentration ratios. Inhibition studies confirmed a relevant ZWT pathway for drug demethylation via CYP2B6. The model showed stereospecificity with selegiline, with no racemization observed. For JWH-073, the ZWT produced the same metabolites as humans, except for indol group hydroxylation, which was attributed to the absence of CYP2C9 orthologs in zebrafish. For the peptide hexarelin, the results were particularly notable.

As a result, hexarelin displays a very fast metabolization in ZWT conditions and all the metabolites described for human were observed in ZWT. This finding confirmed the model's utility even for compounds with low stability, demonstrating its potential breadth of application.