(Editor’s Note: This breakthrough technology booted on a loan from NCBiotech originally appeared in Duke University Medical School’s online publication Magnify on Friday, June 12th. Used with permission.)
DURHAM – Life scientists love antibodies, not only because these little proteins help protect us all from pathogens, but because antibodies are also a very handy laboratory tool for identifying and labeling proteins of interest for their research.
If you’re trying to find something very tiny, you’ll need a small flag to mark it. It’s an antibody.
Like most life science researchers, Scott Soderling, chairman of Duke Cell Biology, relied on custom antibodies, molecules made to order by hundreds of different delivery laboratories that help scientists find specific proteins in cell cultures and living organisms and to mark.
“But there is a problem,” he explains in the small conference room next to his Nanaline Duke office. “50 percent of the antibodies on the market are junk. You are not specific. You can bind what you think they are binding, but then they bind to other things you don’t know about, or they do not bind to what you want to bind at all. “
Worse still, one batch of bespoke antibodies may not be the same as the last. “Suppose you have a perfect antibody that will bind exactly what you want and nothing else. And then you order the next lot and there’s another preparation from another animal and you’re back to zero. It doesn’t work. “
“It is assumed that these bad antibodies lead to a large part of the non-reproducible results,” says Söderling. “So it costs money, it costs time, and it costs credibility. This is a huge problem for science, both academia and industry. ”Part of the problem stems from the fact that custom manufacturing processes for antibodies date back to the 1970s, he says.
But Söderling has started a Duke spinout company that he hopes will solve the reliability problem. CasTag BioSciences is based on a technology developed in his laboratory that uses the genome editing tool CRISPR to mark interesting proteins in a completely new way.
An important research focus of Söderling was the identification of proteins in the synapses of the brain, the tiny gaps between nerve cells in which signals are transmitted and received. All this signal transmission is regulated by specific proteins. But identifying all these proteins in the synapse and interpreting what they are telling the cell is a huge problem in a very small space. Antibodies are an important tool, but the work has been frustrating and slow, in part because of the difficulty of working with custom antibodies.
When the news of the new gene editing technology called CRISPR spread about three years ago, Soderling and his team wanted to see if it would give them a better way to tag and visualize the hundreds and even thousands of proteins that make them up in the tiny synapse between neurons.
“We got the idea that CRISPR could be a really amazing tool to address the pressing problem of identifying and labeling these hundreds of proteins,” says Soderling. “What we came up with was a new modular way of essentially turning the labeling problem on its head.”
They use CRISPR to turn short sequences into a gene so that every protein it produces bears a tag they create, which is recognized by a known, reliable, and well-characterized antibody, rather than a custom antibody shot in the dark.
“These antibodies recognize a small section of amino acid sequences,” explains Söderling. “So we just take the DNA that codes for these amino acids – the handle – and we place this handle directly in the gene in vivo or in the cell,” says Söderling.
After the proof-of-concept experiments produced a nice protein marker in the mouse brain, Soderling looked at the pictures and said, “Okay, it’s huge.”
In fact, they named their new system HiUGE (homology-independent universal genome engineering), and it could actually be huge.
They call it plug-and-play biology because they can use just a few of their tags to address hundreds of unknown proteins and even incorporate multiple tags into a gene at the same time. According to Söderling, the system is modular and easy to use, which will enable semi-automated, high-throughput approaches to protein labeling.
For example, imagine a delivery truck driver slowly driving down the block after dark in a downpour and looking for house number 2345. Soderling and his team have a bright sign on every house with the number 2345 that says “Hey UPS! Over here!”
The HiUGE system is delivered to living cells by two adeno-associated viruses that work as a team, either in a shell or in an organism. A virus carries guide RNA that marks the point where CRISPR should cut the DNA and insert a new code. The second adeno-associated virus carries “the payload”, one or more tags they developed, which are now built into any protein that the gene subsequently produces.
The vectors, including a synthetic guide RNA and HiUGE tags, are agnostic or “homology-independent” as the name suggests. They don’t care what gene is around them. “We developed this guide RNA in such a way that it does not recognize anything in the genome of mice, humans, monkeys, cats or donkeys,” says Söderling.
It’s a clever way to explore the unknown.
Not only does this approach advance her own work, Söderling began to see that a quick, flexible, and more accurate method of labeling proteins could be a business opportunity as well. With a little research, he found that custom antibodies are a $ 2.4 billion market – again, with products that only work half the time advertised.
He reached out to Duke’s Office of Licensing and Ventures (OLV) to begin the patenting process and seek advice on how to start a business. “Then I had to find a way to run the business because I already have a great job.” In fact, he had just been appointed to the Chair of Cell Biology at the same time.
On the recommendation of the OLV, Soderling visited Biolabs North Carolina, a shared workspace in the Chesterfield Building in downtown Durham that rents individual wet lab benches monthly and provides all the basic equipment a startup needs, including refrigeration, gene PCR machines, centrifuges copy etc. He presented his idea to Biolabs and looked around.
The next day, Ed Field, president of BioLabs NC, called Soderling and asked if he had any help running the company. Field, a veteran startup, is now the CEO of CasTag. The company raised enough cash with a loan from the North Carolina Biotechnology Center to hire a recently graduated Fuqua Business School graduate as director of business development and a former postdoc for Soderling to run the lab part-time while he was after a job in the industry is looking for.
“We have a website. We have orders. We have customers. It runs and runs, ”says Söderling with a certain astonishment in his voice. His conference lectures on HiUGE and a July 1, 2019 paper in Neuron attracted some attention. Then the paper was republished as one of the magazine’s “Best of 2018-2019” which attracted even more attention.
And now they also have ideas for new products. “I hope that this will expand and become even bigger than just labeling proteins,” says Söderling.
“You know, North Carolina was an industrial nation back then,” says Soderling, a native of Tennessee. “I’d love to wake up one day and drive to downtown Durham and see one of the former manufacturing warehouses buzzing with people who make these reagents to ship all over the world. That is the dream. “
Durham-based academic research services company Research Square produced this 3 1/2 minute Vimeo video explaining CasTag BioSciences technology.
(c) North Carolina Biotechnology Center