Synthetic DNA Library of Animal Proteins an Industry First

Charles River Retrogenix® Cell Microarray is an in vitro tool that aims to help biopharma make smarter decisions about their candidates, and reduce the use of large animal models

Given how complex drugs are, the mechanics driving them can be surprisingly simple. In many cases, protein receptors embedded within the cell membrane receive signals from a drug outside the cell and react, triggering a cascade of cellular events. The goal for therapeutic developers is to find protein targets and drug candidates to these targets that are a efficacious, selective and safe.

Since our DNA contains instructions for tens of thousands of proteins, scientists have devised next-generation tools, like cell microarrays, to uncover important disease-relevant receptors coveted by drug developers, as well as a screening tool to identify off-target receptors that can could cause adverse reactions in the body.

Cellular microarrays, slides coated with DNA sequences and cells, allow the overexpression and representation of thousands of specific proteins in vitro, to test the binding characteristics of drugs under development. Until now, these synthetic DNA libraries have encoded for human proteins, but this month Charles River's Discovery site in High Peak, UK-which has more than 6,500 full-length human proteins individually over-expressed in human cells-launched the first-ever complimentary DNA (cDNA) library of 200 non-human proteins. The new development arms clients with species-specific protein binding information without the use of animal models and helps them de-risk candidates earlier on to make informed decisions about how to subsequently assess their candidates, says Mark Aspinall-O'Dea, PhD, Associate Director - Discovery at Charles River.

"As a synthetic tool, the Retrogenix® cell microarray helps you gather data that you would perhaps otherwise have found using an animal model, in terms of the way a molecule is going to behave, the interactions you're likely to see, and then explore those findings with in vitro tools rather than having to go in vivo," said Aspinall-O'Dea. "For instance, let's say we found an unlikely off-target event. Clients will probably do an in vitro study to evaluate whether that was a safety liability event or not, and then, may well decide not to do an in vivo test as a consequence."

The new tool also helps Charles River, and its client meet its 3Rs commitment of replacing, reducing and refining research animals. "Aside from making the drug discovery process more efficient, the cDNA encoding animal proteins also aligns with Charles River's 3Rs commitment by reducing the use of animals and replacing these with in vitro tests," noted Aspinall-O'Dea. "Whereas sponsors might have tested a promising candidate in an animal model further down the drug development pipeline, they can now use cell microarrays early on to de-risk a candidate rather than assuming they have to do an animal study because the regulators expect it."

"The bigger these libraries get the more exhaustive the coverage," said Aspinall-O'Dea. "A client could say, we are considering X, Y, Z animal models potentially in the future. Let us de-risk them on this platform and evaluate whether we see any known, or unknown interactions that could be problematic. It will help them pick the right model to go forward. And it may be that it helps them pick the right non-animal model to go forward. … We can start to feed the data into bioinformatics tools and AI in the future as well."

What are microarrays?

The study of miniaturized microarrays dates back to the mid-1990s when Patrick Brown and colleagues from Stanford University showed for the first time that the expression of many genes in a small sample could be quantitatively monitored in parallel by placing a whole yeast genome on a microarray. Today, microarrays are a common technique that allows researchers to investigate big biological questions; their use has made drug discovery faster and more efficient.

The process is exquisite. In a dish, human cells are seeded over the slides and grown up until the dish is completely covered with cells. Distinct clusters of cells become reverse transfected and overexpress the individual proteins encoded by the DNA plasmids beneath it. Test molecules are added and allowed to bind to the overexpressed receptor, and a detection reagent is able to elucidate both good targets and off-targets.

"What we effectively do is to use robotics to put that cDNA onto a slide, and then, we grow a cell line over the top of it, and it's the cell line that generates the desired protein in its native configuration," says Aspinall-O'Dea.

"The animal library currently focuses on cell surface proteins with the biggest differences to their equivalent human protein (90% match or below). Anything that is above a 90% match, and therefore very similar to the human equivalent protein, is not in the library, but over time the library could grow to accommodate that last 10 percent," said Aspinall-O'Dea.

"We kind of triaged it in such a way that we'll go for the biggest difference that's got the most tangible likelihood of causing a differential binding profile for a drug compared to the human equivalent, and then work back from there."

"Ultimately, our aim is to help drug developers make smarter, faster decisions to reduce the time and cost of bringing life changing drugs to market," says Aspinall-O'Dea. Being able to do this in a manner that reduces our dependence on animal models and helps us to meet our 3Rs is very important to Charles River, and the addition of non-human proteins to the Retrogenix® platform is a significant step on this journey.

Check out the Alternative Methods Advancement Project (AMAP) and learn more about the scientific and technological innovations Charles River is using to lead the industry into the next frontier of drug discovery and development.

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