Contract research organization Phylonix said this week that it has published a study showing that zebrafish, its primary drug-screening model, can be an “effective animal model for assessing human cancer cells at various stages of tumorigenesis.”
The research could be good news for Phylonix and other CROs – including Zygogen and Daniolabs – that use zebrafish as a drug-screening tool. But the relative scarcity of pharma input on the subject and hesitance on the part of US regulators to use the animal in pivotal toxicology studies indicates that shops hoping to commercialize the model organism for that end will continue to encounter challenges.
Traditionally, drug screening and preclinical development have been done at three levels – biochemical assays, cell-based assays, and small-animal testing. One of the most popular organisms for small-animal testing is the mouse because it provides data that are extremely representative of human physiology.
But pharmaceutical companies are wary of the time and money involved with small-animal testing, and would prefer to know sooner whether a drug will fail physiologically relevant tests.
This is one of the reasons that the use of cell-based assays has increased dramatically in pharma in recent years, but researchers are exploring other methods, too – namely, model organisms that fall somewhere in between rodents and cell-based assays because of their small size and simplicity.
These organisms include nematodes, fruit flies, and zebrafish, and of the three, zebrafish seem to have gained the most traction as a drug-screening model as several companies have sprouted up in an attempt to exploit the technology.
One of these is Phylonix, which offers zebrafish-based contract research to pharmaceutical and biotech companies. Based in Cambridge, Mass., Phylonix holds several patents related to the use of the fish in drug discovery, and maintains that its services are used by “dozens” of undisclosed biotechnology and pharmaceutical customers worldwide.
In the study, supported by a grant from the National Institute for Diabetes and Digestive and Kidney Disease, Phylonix scientists injected human melanoma, colorectal, and pancreatic cell lines into various sites in zebrafish, including the yolk sac, brain, and circulatory system.
Each of the cell lines “proliferated, migrated, formed masses, and stimulated angiogenesis” in the zebrafish, Phylonix said. The research is significant on at least three fronts, according to the company. First, the zebrafish embryos did not reject the human cells – typically a “major problem with other animal models,” the company said.
Second, the researchers confirmed via whole-mount immunostaining and histology that new zebrafish angiogenic vessels formed in and around the human cancer cell masses, which is similar to the way cancer progresses in humans. Third, zebrafish cells incorporated into the human cell masses, which indicated that cell signaling methods are highly conserved, the company said.
According to a statement from Phylonix, the research was published this week in the online version of the journal Angiogenesis. However, the article was not yet available on the journal’s website at the time this article was published. Phylonix did not respond to phone calls and emails before publication.
Other companies attempting to market zebrafish as a model drug-discovery organism include Atlanta-based Zygogen, which also offers contract research services; and UK-based CRO Daniolabs, which licensed specific technology from Zygogen for its services and internal drug-discovery program.
Zygogen has had two of its disease model assays approved for screening in the National Institutes of Health's Roadmap Initiative through a partnership with the Emory University Molecular Libraries Screening Center. It also has an ongoing collaboration with Sanofi-Aventis in the area of thrombosis.
“Zebrafish studies are unlikely to be accepted as pivotal safety studies by the FDA in the foreseeable future.”
According to Calum MacRae, an assistant professor of medicine at Harvard Medical School and a member of Zygogen’s scientific advisory board, the zebrafish will likely ultimately find its sweet spot as a tool for early-stage toxicology screening and similar studies, areas where most model organisms have seen little to no uptake.
“It took a while for the mouse to be accepted, and I think part of the drive toward the adoption of alternative model organisms is that the mouse still isn’t all it can be in pharmaceutical research,” MacRae said. “Pharmaceutical companies, I think, have used model organisms in the wrong way.”
MacRae added that pharma is starting to scratch the surface of using zebrafish as a screening tool, but none have moved it into the mainstream yet. “In pharma, the fish is starting to be seen as a disruptive technology compared with some of the other model organisms because it is cheaper and is capable of providing much higher throughput,” he said. “But it still has a ways to go.”
Underscoring MacRae’s point, it is difficult to find pharmaceutical companies using zebrafish for drug screening. Almost all scientific articles published on the subject have been authored by academic researchers or scientists from the aforementioned companies. Likewise, drug-discovery conferences rarely have pharmaceutical companies promoting the use of the organism.
According to Phylonix, zebrafish have several advantages as an alternative model for cancer research, including the fact that zebrafish are small and inexpensive to maintain; they are easily bred in large numbers; they require as few as 50 cells for transplant; they enable researchers to visualize cell migration in vivo due to their relative transparency; and they require small amounts of drug per experiment.
Abigail Jacobs, a researcher at the Center for Drug Evaluation and Research at the US Food and Drug Administration, wrote in a June review article in Expert Opinion on Drug Metabolism and Toxicology, that “when non-traditional species are used [as model organisms], there may be less historical information available and unique issues of their care, and differences in physiology and anatomy, and susceptibility to infection need to be understood.
“Non-mammalian test species, such as zebrafish and Caenorhabditis elegans may be used by drug sponsors in screening assays, but are not yet ready for use in pivotal toxicology studies because of the difficulty in extrapolating to mammalian species,” Jacobs wrote.
She declined to comment further this week, but reiterated in an e-mail that “zebrafish studies are unlikely to be accepted as pivotal safety studies by the FDA in the foreseeable future.”