Cisplatin alkylating activity in zebrafish causes resistance to chorionic degradation and inhibition of osteogenesis

Zebrafish have gained popularity as a model organism due to their rapid, external, and transparent development, high fecundity, and gene homology with higher vertebrate models and humans. Specifically, drug discovery has had high success in the implementation of zebrafish in studies for target discovery, efficacy, and toxicity. However, a major limitation of the zebrafish model is a dependence on waterborne exposure in order to maintain high throughput capabilities. Dose delivery can be impeded by a matrix of N-linked glycoproteins and other polypeptides called the chorion. This acelluar barrier is protective of the developing embryo, and thus new approaches for assessment have involved their removal. In these studies, we explored the chorionic interference of a well-characterized alkylating chemotherapeutic, cisplatin, known to accumulate in the chorion of zebrafish and cause delayed hatching. Our results indicated that increased exposure of cisplatin due to dechorionation did not alter morphological endpoints, although retained confinement reduced total body length and yolk utilization. Additionally, inhibition of osteogenesis visualized with Alizarian Red staining, was observable in dechorionated and non-dechorionated treatment groups. The chorions of cisplatin-treated embryos showed resistance to degradation unless treated with a pronase solution. This may be may be due to cisplatin covalently crosslinking which reinforces the structure. As such, the chorion may play an advantageous role in studies to determine alkylating activity of novel compounds. Furthermore, the expression of zebrafish hatching enzyme was not affected by cisplatin exposure. These studies demonstrate that not only was recapitulation of mechanistic activity supported in zebrafish, but highly relevant off-target toxicities observed in higher vertebrates were identified in zebrafish, regardless of chorionation. Experimental design in drug discovery should consider preliminary studies without dechorionation in order to determine dose impediment or off-target adducting.