01/13/2025 | News release | Distributed by Public on 01/13/2025 08:43
ReproTracker offers an alternative to animal testing for embryotoxicity
One of the major areas of safety testing during drug and chemical development is testing for teratogenicity, or the potential for a chemical to cause defects in fetal development. This potential was tragically realized in the 1950s when pregnant women were given the compound thalidomide for morning sickness. Animal testing had demonstrated that even very high doses had no ill effects on the subject, so researchers were confident in the drug's safety. It was not until a few years later, in 1961, that research was published indicating a link between birth defects and thalidomide.
Ever since, drug and chemical developers have been tasked by regulators with determining whether or not their compound could be teratogenic. For a drug, teratogenicity is not necessarily a death sentence, since the compound could still be sold with a warning against prescribing during pregnancy. Even thalidomide is still used today to treat multiple myeloma and leprosy. But for consumer goods like cosmetics, this would typically be a deal breaker.
For a long time, the only accepted methods for testing for teratogenicity were animal models. These are not only expensive and morally strict, but also not always correlated to the potential effects in humans. After all, rat fetal development is quite different from human.
Dutch biotech company Toxys has created an alternative using human induced pluripotent stem cells (hiPSCs), which they call ReproTracker. This in vitro assay uses human stem cells to simulate the early fetal development process.
"Actually, the development of this assay started by accident," said Giel Hendriks, CEO of Toxys. "The true story is that it started as a development for our genotoxicity testing platform, called ToxTracker."
The initial idea failed, since the differentiated cells stopped proliferating and were less sensitive to DNA damage, rendering them useless for testing compounds. However, a new idea emerged.
"We realized that if we differentiated stem cells into tissues, in this case liver, we could actually use that to predict early embryonic development defects. By following the whole differentiation process from stem cell to different mature cell types, that is where a lot of information is coming from," Hendriks said.
The protocols were already in place to differentiate mouse stem cells into heart and liver, so the research team tested compounds that were known to cause birth defects to see if the development of the stem cells was disrupted. In this system, the morphological development of the cells was tracked, and the expression of cell-specific biomarkers was measured to determine whether the cells developed normally. They found that known teratogenic compounds did indeed disrupt the differentiation of stem cells into heart, liver, or neural cells, demonstrating that the assay could be used to predict embryotoxicity.
Swapping mouse embryonic stem cells for human iPSCs was the next logical step. Toxys has been building the ReproTracker platform since at least 2017, expanding their validation database and refining their technique. At present, various validation studies of the ReproTracker assay indicate that the platform is 85% effective in predicting teratogenicity. Since the EU had banned in vivo cosmetics testing, ReproTracker is a popular choice for cosmetics developers, though it has also proven effective for chemical and drug developers as well.
The assay is continually refined by Toxys. Recently they moved to a smaller, 96-well plate format which uses 10 times less testing material. At a recent conference, they presented a poster on adding an additional cell layer to the assay looking at bone differentiation, which could predict skeletal malformations. They are also looking into refining their qualitative readouts, which offer more information than a quantitative "yes or no" testing outcome.
As the process becomes ever more refined, ReproTracker also has the potential to predict dosing levels for suspected teratogenic compounds. Advancements in assays like this are crucial to conforming to the 3R's of animal testing and represent a strong step forward toward replacing animals for developmental and reproductive toxicology studies.