Indee Lab’s CRISPR Delivery Technology for Affordable Immunotherapies

May 7, 2020 -- This article was contributed by Meenakshi Prabhune, Ph.D., Science Writer and Journalist at Synthego. Article source on Synthego’s CRISPR Cuts.

Ryan Pawell is the founder and CEO of Indee Labs, a Berkeley biotech. The mission of Indee Labs is to use microfluidic vortex shedding as a scalable method to deliver CRISPR reagents into cells for developing affordable cell and gene therapies in the future. In our previous blog post, we summarized the work from their latest publication on CRISPR-editing in T cells using microfluidic devices.

In this interview, we chatted with Ryan about the concept of using microfluidics for CRISPR delivery, the mission and ongoing projects at Indee Labs, and the convergence of engineering and biology for building scalable platforms.

Tune into the podcast interview or read the transcript below. The choice is yours! 

Meenakshi Prabhune (Minu): Ryan, let's start with an introduction about your educational background. Tell us how you got started.

Ryan Pawell: I got my Bachelor’s of Science in mechanical engineering at what I think is the best university in the world, UC Santa Barbara. Partway through, I took an internship at a now publicly traded medical device company started by three students. It was supposed to last one summer, but I stayed for almost 18 months.

From there, I read about stem cells and regenerative medicine along with the Stanford Biodesign book [about innovation in medical technologies], which said that disposable medical devices offered the best business opportunities for recurring revenue and high margins. So the general idea at the time was regenerative medicine based on cells would have a lot of impact in the future and microfluidic devices would be used to manufacture these therapies. That was in 2010.

Minu: Fast-forward to your current role, which is CEO at Indee Labs. Tell us a bit about what Indee Labs does and the inspiration behind the concept.

Ryan: Indee Labs is doing exactly what I was just talking about. We are developing a microfluidic device for processing cells. There’s a lot of things that you can do with cells when you want to process them. You can culture them, you can separate them, you can lyse them, or you can do gene delivery, where you put material, like CRISPR, inside the cells.

At Indee Labs we’re focused on the delivery element. The idea came about because I was working on a cell separation technology where we were pushing cells through these tiny, little post-arrays—like in a Plinko board, where big cells would go one way and the small cells would go the other way, so you could separate them. 

Then we wanted to figure out how to push large volumes of fluid through these post-arrays. So the question became what happens if we flow cells through a post-array really fast? The answer was that we poke tiny holes in the cell membrane, allowing for material like CRISPR to get in. (CRISPR didn’t actually exist at that time, so we used a membrane-impermeable dye.) Indee Labs has taken that technology, or that discovery, and developed it. Now we’re marketing it to Series A plus-funded biotechnology companies and to pharmaceutical companies. 

Minu: That is such an interesting concept. And what I find even more interesting is that your background is in engineering and then you switched over to biology. There are some similarities with our founder as well. But tell me what inspired you to move into biology and particularly into the area of gene therapy delivery.

Ryan: After undergrad, I went to Australia for a couple of weeks to go surfing, and I ended up staying about five years for grad school, where I studied these microfluidic devices for processing cells. Toward the end of grad school, I was accepted into a program called the New South Wales Health Medical Device Commercialization Training Program, which was basically a program funded by the state health department for professors and postdocs to get business training for medical device commercialization. I was the only student in my cohort. 

Shortly after completing that program, IndieBio made me an offer and gave me about 30 days to relocate from Sydney to San Francisco. Getting back to cell and gene therapy. At about the same time, there was a growing body of clinical literature on T-cell immunotherapies called CAR T showing unprecedented patient outcomes relative to the standard of care and for patients without any options. 

When determining how valuable a therapy will be, you look at the number of quality-adjusted life years added relative to survival with the current standard of care. Therapies need to be safe and effective to get FDA approval—but commercially viable cell therapies also need to add a significant number of quality-adjusted life years in order to qualify for reimbursement and coverage from health insurance companies.

So we had developed a skill set in processing cells with microfluidic devices, and then we needed a big area to focus on in cell and gene therapies. It turned out that CAR T, among cell and gene therapies, had the best patient outcomes relative to the standard of care. So we got excited about working in that area.

Minu: How would microfluidics be used in CRISPR delivery?

Ryan: Let’s take it back a step. Microfluidics is the manipulation of fluids with microscopic channels: these are tiny, little channels that are about 1/10th to 1/100th the thickness of a human hair. Think of it as similar to electrical circuit boards, but for fluids instead of copper wires and electricity. The reason we got into microfluidics for CRISPR delivery was because microfluidics lends itself to precisely processing cells. Cells are also about 1/10th to 1/100th the thickness of a human hair. For that reason, nobody’s going to have much success with ex vivo CRISPR delivery, using clunky instruments roughly equivalent to a drill press from the hardware store. It’s necessary to be on the same size scale.

Minu: We’ve been hearing a lot about gene therapies, and most of them are ex vivo. But there’s a lot of ongoing talk about developing in vivo gene therapies. Are these microfluidic devices, or is the concept in general, applicable to in vivo therapies as well?

Ryan: Our devices operate at eight to ten times atmospheric pressure, or about 120 PSI gauge pressure. You can’t pump up a human to those pressures in vivo. For that reason, in vivo therapies are outside of what Indee Labs does. Some complementary companies, like GenEdit, are working in this space, though, and I believe they’ve done pretty well. So in vivo is not for us, but other companies working on in vivo technologies are out there.

Minu: In CRISPR labs, researchers normally use either viral transduction methods or electroporation. Can you talk about the advantage of your method compared to these other delivery techniques? 

Ryan: Viral transduction and electroporation are the two industry standards, particularly for clinical and commercial workflows. Compared to them, though, our technology is simple and scalable, and we can rapidly process clinically relevant volumes of T-cells with high yields and minimal perturbation of the T-cell state. Our tiny 5-by-10-millimeter chip, which is about the size of your pinky nail, can process 30 million cells in less than 10 seconds.

And for viruses, if a pharma company wanted to develop their CAR T using viruses for delivery, they would be giving up hundreds of millions of dollars in royalties on therapeutic sales to invest the time in developing viruses for clinical trials and commercial manufacturing. Timescales can be 18 to 24 months and the viruses might not even work afterward. 

So we can see that developing viruses for gene delivery is really expensive. They take a long time to develop. Also, you can’t take them to scale, meaning that you can’t treat lots of patients with viral transduction. There are also a lot of hazards with viruses. We have a biosafety-2 lab in our facility. We can’t use viruses because it’s not BSL-3. 

Electroporation is another technology that people are using. The problem with electroporation is cells don’t like to be electrocuted. It changes the cell state. A lot of literature is showing that when you then put electroporated cells back in the body, they don't work as well as ones that are, let’s say, mechanically porated.

The other advantage of using microfluidics is its simplicity. Each new material component or reagent that you add to any process needs to be rigorously tested in order to get past the FDA. We keep things really simple. We use implant-grade materials, accredited designers, and Good Manufacturing Practices–grade reagents.

You also want to maximize the number of therapies you make per unit time per square foot of manufacturing space. Our devices are small. We can make thousands per day with existing manufacturing workflow. So from a comprehensive perspective, we are the ideal technology for developing these T cell immunotherapies. Both on the engineering and manufacturing side, as well as on the immunology and clinical side—because we have data showing the cell state is not perturbed or changed like it is with electroporation.

Minu: Is it becoming an essential feature to support scaling to combine engineering and biology? What would you say you've learned from working in these fields?

Ryan: I think everything will be multidisciplinary in the future. We’re somewhere between data science, engineering, and biology, and we recently got started with machine learning. Each field of study offers tools that help to complete a very complicated puzzle. For Synthego as well as Indee Labs, the more proficient your team is in these different fields, the more you can scale your productivity and solve complex problems.

 For us, as shown in our latest BioRxiv paper, we’re getting started on the data science and machine learning side of things. With each experiment, there are something like 150 different inputs or outputs that we measure, and a whole lot more that we should measure. It’s really difficult for me, or our team, to look at how the numbers change over time. We need a better way to determine which parameters are the most important or which ones we should not spend time on.

So when a company does have that proficiency across engineering, biology, and say, data science, it’s possible to maximize the value of each experiment by doing the “right” one, the most valuable one, first. I had say it will only get more interdisciplinary—that’s going to be a bigger and bigger requirement for companies like Synthego and Indee Labs in the future.

Minu: That makes a lot of sense. Now, since we’re talking about “big picture” things and I always like to have a fun question on the podcast, what does the day-to-day at Indee Labs look like for you? Putting aside the quarantine period, what would you normally do on day-to-day basis?

Ryan: I’m still somewhere between the business and the lab business side of things. And from time to time I’m involved with investor issues. In the lab, we do high-speed imaging experiments where we’re trying to take pictures of cells going through our chip with a camera that takes images at about 500,000 frames per second. Sometimes I’ll do a bit of coding, though it’s not my greatest strength. To some extent, I’m still involved with the design and optimization of the chips.

On the business side, I talk to as many pharma execs and biotech founders as possible to try to get our chips into their workflow. So it varies every day. It’s also always important to make time for your team. All our hands join in the science meetings, and we have open office hours, where anyone can talk about anything they want to talk about. So it’s really a little bit of everything.

Minu: Can you share with us the near-future plans for Indee Labs?

Ryan: The ultimate goal has always been to get these chips into the manufacturing workflows so we can help treat as many patients as possible. There’s a bit of ambiguity here still. But we recently received guidance from the FDA, stating that no new clinical trials are required to transition over to our technology, which is really exciting. We’ve started working with a major pharmaceutical company in our space that has some interesting plans to get our technology into their therapies. We may be involved in treating a lot of patients, and much sooner than we originally anticipated. So with these two developments, we could actually achieve our goals sooner than we’d expected, and it will require less work to get there, and no clinical trials. It’s a really great opportunity for me and the team at Indee Labs. 

Minu: It does sound absolutely exciting! We’ll look forward to hearing more about your work at Indee Labs and we’ll all stay tuned for updates.