Stepping on the Cas13: Carver Biosciences Uses CRISPR to Target RNA Viruses

Many of the world’s most common or deadly human pathogens are single-stranded RNA (ssRNA) viruses like Ebola and Zika. The world just spent two years at the mercy of an ssRNA virus—SARS-CoV-2—which doesn’t seem to be going anywhere. While there has been significant progress in developing protective measures against COVID and its seemingly forgotten fellow respiratory virus, the flu, with vaccines that work by helping build immunity, there seems to be no end in sight for many reasons. These include the steady emergence of new viral variants.

Not surprisingly, interest in applying CRISPR as both a diagnostic and a potential therapeutic against SARS-CoV-2 surged in the early months of the pandemic. Early research suggested that CRISPR-Cas9 effectors can inhibit the replication of ssRNA viruses with DNA intermediates in mammalian cells. While several studies have estimated that only 2.5% of those viruses have DNA intermediates that could be targeted using Cas9, RNA-targeting CRISPR effectors could offer a promising therapeutic alternative to ssRNA viruses.

Stealth startup Carver Biosciences has turned a different CRISPR RNA-cutting nuclease enzyme—Cas13—into an antiviral that can be programmed to detect and destroy RNA-based viruses in human cells. The technology development behind Carver is the brainchild of co-founder Cameron Myhrvold, PhD, Assistant Professor of Molecular Biology at Princeton University. If that name looks familiar, it should be: Cameron is the son of Nathan Myhrvold, former chief technology officer at Microsoft and now an award-winning author and molecular gastronomist.

During his postdoc with Pardis Sabeti at the Broad Institute, Myhrvold helped to harness Cas13’s programmable RNA-targeting activity to develop an end-to-end technology platform called Cas13-assisted restriction of viral expression and readout (CARVER). Myhrvold has since teamed up with CEO and Carver Bioscience co-founder Walter Strapps, PhD, who has worked at Merck, Intellia Therapeutics, and Gemini Therapeutics. This is Strapps’ first experience as a company CEO.

Together, Myhrvold and Strapps have their work cut out for them, beginning their journey of taking the CARVER technology from an in vitro proof of concept to a scalable clinical solution. GEN Edge spoke with both co-founders to discuss the steps they are taking to pave the way for Carver to go from stealth startup to gene-editing giant.

GEN Edge: What was the foundational science that drove the creation of Carver?

Myhrvold: One of the things that got me excited about Cas13 when I started my post-doc was this notion that we now have a CRISPR protein that can be targeted to RNA. In particular, RNA viruses cause so many of the infections that make us sick. About two-thirds of the viruses that infect humans are RNA viruses—there are hundreds of them— and almost all have no good treatment options or vaccines. We’ve made some progress in that regard, but only a few dozen have suitable treatments. I got excited by the notion that maybe we could use Cas13 to target then destroy the RNA of these viruses as they are replicating, which would represent a fundamentally new way of dealing with antiviral infection.

That launched an academic project to show that this works as a proof of concept. We did some experiments in tissue culture, trying to show that we can target multiple different RNA viruses using different spacer sequences. The critical thing for what we’re doing is relying on nucleic acid hybridization. Knowing the genome sequences of all these different viruses gives us a way of going after something totally different. We’re not targeting viral proteins or host factors—it’s focused on the genome sequence itself.

In many ways, we’re taking these CRISPR systems back to their roots. They evolved initially to go against RNA bacteriophages. That’s why I like the idea of repurposing them for viruses that can affect people. And we were successful, demonstrating in cell culture experiments several beneficial properties like efficacy and multiplexing capabilities, which is nice because we’re so concerned about the evolution of resistance. That’s not something we worry about here because we can simply re-target the CRISPR easily to go after different sequences.

Finally, we did some preliminary work looking for off-target effects because that’s something that everyone is concerned about in the CRISPR field. While we didn’t see evidence for a ton of that, we will continue evaluating that going forward. Fortunately, in this scenario, those would be effects just on the transcriptome and nothing permanent. Even if there was some modulation of gene expression, hopefully that would not be too harmful.

GEN Edge: How far along is Carver in turning this proof-of-concept in vitro work into clinical testing?

Strapps: I don’t know if there’s a formal definition of whether or not a company is in stealth. But we haven’t talked a lot out in the world about what we’re doing, which is why you can’t find out a lot of stuff. I just started as CEO of Carver about two months ago, and we hired our first employee about two and a half weeks ago! We’re very much in the beginning stages.

Before joining Carver, I was at Intellia Therapeutics, which was at a similar stage as we are now but in 2014 and 2015. We have this very interesting protein, and there are many directions you could go with that. The key is figuring out the therapeutic areas you can go after that will allow you to demonstrate that your technology is actually relevant.

RNA viruses are an excellent place for us to go while allowing us to demonstrate the platform’s utility against anything that uses RNA to replicate itself. You could envision going from viruses to somatic disease, directly targeting the RNA. But before you can dream that big, it’s important to demonstrate that you can do what you think you can do with the technology. That’s the advantage of going after viruses is for us. We can target those directly, and they’re transient.

We will target respiratory viruses because one of the most substantial issues with all of these technologies, whether it’s Cas-based or oligo-based, it all boils down to a problem of delivery. We have to be able to get the thing to the cell that it needs to be in. Going after respiratory viruses allows us to do that very directly. We can go straight into the lungs of people. That’s where the virus is, and we can target it directly. Doing that leads to any number of things, but we’ve got to start somewhere. The respiratory viruses are interesting because they’re accessible. Also, people over the last two years have become very familiar with the concept of respiratory viruses and the impact they can have on human life.

Myhrvold: This all started a while back when I gave a talk, and some venture capitalists in the audience were interested in my work. I had a series of conversations with them, resulting in seed funding from venture capital. That’s enough to support the work that we’re doing and is enough to get to this next milestone of using the platform in an in vivo system. Once we get there, we will raise a much larger round to take this to the next level. But we want to stay as focused as we can in these early stages. We’re trying to be light on our feet and nimble.

GEN Edge: What delivery system will Carver use to deliver the Cas13-based system?

Strapps: We don’t know for sure what we’re going to do. There’s a lot of work we need to do within this space. But our strong desire would be to use a non-viral delivery method. People are very familiar with adeno-associated virus (AAV) as a delivery system. But I think people at this point are also much more familiar with the lipid nanoparticles (LNPs). That’s what the mRNA vaccines from Pfizer-BioNTech and Moderna used, and it’s what Intellia used to do their liver directive programs for Cas9.

We’ll be looking at LNPs and other chemically formulated particles that we are more interested in for delivery than the virus-based methods. LNPs for respiratory viruses have lagged behind the liver in terms of delivery. However, over the last decade or so, a lot of work has gone into LNPs for delivery to the respiratory ecosystem.

GEN Edge: What are the immediate tasks for Carver to tackle to get off the ground?

Strapps: We’ve spent the last few months working scientifically to figure out what it is that we’re going to do. First, we have to recapitulate some of my co-founder’s work in his post-doc and then expand upon that. Most of the work he did used plasmids to express the components: the Cas13 and the guide RNAs (gRNA). That’s great for demonstrating the activity, but it’s not practical for a therapeutic. For a therapeutic, what you could do is to take those same components and turn the Cas13 into a messenger RNA analogous to the vaccines, chemically synthesize the gRNAs, and then formulate those things together into a particle that will then be delivered into the relevant cell that in an actual human being.

But before that, we have to do in vivo disease models. Employee number one will essentially be getting that up and going while figuring out how easy or hard that is to do. We’ve got to demonstrate that the Cas13 system can target the flu virus in vivo. A mouse is excellent for that introductory model system. Once we’ve shown that, we’ve got to move into a larger organism that is an accepted model within the field, the ferret. From there, we will have to tackle the scaling problem of making enough clean material. That isn’t often planned for in companies of our stage, but you’ve got to be ready for it. If you’re going to go into a human being, you need to be able to make a lot of material that is of high quality. That’s what we’ve got to do to prepare for clinical trials. A lot of work needs to go into each of those milestones.

There is a prodigious amount of work that has nothing to do with science in setting up a company! Getting space, employees, and capital is competitive. We know what we need to do so we can go out and make a convincing case for a Series A funding round. People need to know who we are when we go out there and start having those conversations. We are helped a lot by having the people backing us in our seed round. One of our advisors is Intellia’s [former] leader and founder, Nessan Bermingham.

There’s a critical mass in Boston, where we are based. Physically finding space to set up laboratories is difficult. It’s a very challenging environment for hiring people. It was always a challenge, even before the pandemic. Since the pandemic, it’s just become even that much greater. It’s an employee’s market. People have a lot of opportunities. We’ve got to spend our time figuring out how to do all that. Generally, the first employees you hire are your network employees who you’ve worked with before. That’s what we’ve done here. Employee #1 is someone that I’ve worked with extensively over the last decade or so. But it’s a lot of little logistic pieces to get things set up, like contracting and the ability to pay bills. That’s the nuts and bolts of setting up a company.