Zebrafish as complimentary model for drug discovery and chemical prioritization: an example of high-throughput screening method to validate teratogenic effects

ABSTRACT

Zebrafish has gained popularity in the field of safety pharmacology and ecotoxicology due to its unique biological qualities (small size, transparency and rapid development, highly similar to the mammalian) to such an extent that it is now considered a potential alternative to experimentation in higher vertebrates. In order to answer the increasing demand for a rapid, reliable and cost-effective methodology to detect toxicity of chemical substances, particularly in Human Risk Assessment, several laboratories across the world conducting zebrafish research (including ours) are acting together on different fronts to develop, standardize, harmonize and promote new test methods and relative guidance. With this purpose, we are participating in a large number of international initiatives as well as boosting our internal R&D to support and validate the model.

In this scenario, we present here a high-throughput screening system for validating the teratogenicity effects in zebrafish of 30 compounds, reported as teratogenic or non-teratogenic in humans. Zebrafish embryos are exposed at different concentrations from 6 to 96-hour post-fertilization and imaged with an automated capillary-based system. During the first phase, we perform a dose range finding assay in order to determine the Benchmark Dose 10 (BMD10) for each compound. The BMD10 is subsequently used to establish a narrower exposure range of concentrations: BMD10/2, BMD10, BMD10x2, BMDx4, and BMDx8 to determine and score morphological changes and functional abnormalities according to 15 physiological teratogenic endpoints.

Thanks to this new phenotypic approach, we are able to detect even subtle teratogenic effects, including those manifested at concentrations close to lethality. The study shows a clear dose-response teratogenic effects for those compounds reported as teratogens in humans, including thalidomide, usually reported as non-teratogen in zebrafish. The correlation between our results and the results previously published, both in zebrafish and in other higher vertebrates (including human), confirms the predictive potential of the model for high-throughput teratogenicity screening and supports its use as valid experimental alternative method.

Zebrafish as complimentary model for drug Discovery and chemical prioritization: an example of high-throughput screening method to validate teratogenic effects

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