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Getting a Next-Generation Genome Editing Therapy for Sickle Cell Disease Back on Track

February 29, 2024

A serious adverse event in the first patient treated with an experimental genome editing therapy for sickle cell disease marked the beginning of the end for Graphite Bio. The company discontinued development of the treatment and eventually entered into a reverse merger with Lenz Therapeutics with a focus on improving vision. At the end of 2023, Kamau Therapeutics emerged from stealth following a strategic transaction with Graphite Bio that provided the new company with all of Graphite’s genome editing assets including next-generation platform technology and its lead program, a hematopoietic stem cell therapy engineered to restore adult hemoglobin by correcting a genetic mutation in people with sickle cell disease. We spoke to Matthew Porteus, co-founder of Graphite Bio and co-founder and CEO of Kamau Therapeutics, about the company’s genome editing technology, what’s now understood about the adverse event that occurred in the Graphite Bio clinical trial, and the development path forward for the therapy.

Daniel Levine: Matt, thanks for joining us.

Matt Porteus: Thank you very much for having me.

Daniel Levine: We’re going to talk about sickle cell disease, gene editing, and Kamau’s efforts to develop next generation gene editing technology. Sickle cell disease was the first disease to be diagnosed as a genetic condition. It’s been more than a hundred years since then, but it’s only recently that we’ve seen a surge of activity around the development of potentially significant therapies, including the recent approval of the first CRISPR therapy and a cell therapy. What’s happened to change the landscape for treatments and therapeutic development for the condition?

Matt Porteus: Yeah, thank you, it really is, and it’s such an important disease for the patients that have it, but also it’s been just a seminal disease for understanding of the interaction between genetics, human biology, disease, and as you mentioned, it’s been that way for decades. It’s been recognized for its genetics and the complicated interactions with the environment. But really we were waiting for the right technology to address this disease at the genetic basis so that we could address a genetic disease with a genetic therapy. And so, there were two approvals in December, which is super exciting for the field and for patients. One is based on using a lentiviral vector to add a gene to counteract the sickle cell gene, and that’s based on lentiviral delivery, which has been around for a couple decades. However, like all technologies, it’s been refined and optimized so it achieved the results. We wanted chiefly, the results that are beneficial to patients and justified its approval. But the other approval for Casgevy, or the exa-cel drug, was based on genome editing. And as you said, CRISPR Cas9. The amazing thing is CRISPR Cas 9 is an engineered nuclease [that] was first reported in a test tube in 2012, and to see a test tube discovery translated into a therapy in just over a decade is rather remarkable. Now, what I will say about the Casgevy approval is while using CRISPR Cas9 it is doing it in a way that builds on, again, tons of research and understanding of the biology of the disease, most notably, the recognition that if you couldn’t directly correct the sickle cell mutation, was there a way to use genome editing that could counteract the sickle hemoglobin, the pathologic sickle hemoglobin. From clinical research, it had been known that patients with sickle cell disease that had higher levels of fetal hemoglobin, that’s the hemoglobin that’s normally expressed before we’re born but normally gets turned off after we’re born. If some of that stays turned on, those patients who have higher levels have less severe disease. That led to the development of a drug called hydroxyurea, a 70, 80-year-old chemotherapy drug that works by upregulating fetal hemoglobin in ways we don’t still fully understand. And again, patients who take a small molecule drug, a chemotherapy drug every day have higher levels of fetal hemoglobin, then their disease is less severe. So if you couldn’t correct the mutation, and I hope later we’ll talk about how one day we’re going to directly correct the mutation, was there a way to use genome editing to sort of enhance this fetal hemoglobin effect? And for that, we turned to another line of research, which was understanding how fetal hemoglobin is turned off after birth, and using a combination of biochemistry, genomics, human genetics scientists identified that a transcription repressor called BC11A after birth would get turned on in red blood cells. It would bind to the fetal hemoglobin or gamma globin promoter and turn that off, allowing the beta globin or adult hemoglobin to get turned on. So now we have a proof of concept that fetal hemoglobin can significantly improve the course of disease. We have human genetics and biochemistry identifying a pathway that if you could interfere with it might keep fetal hemoglobin turned on at higher levels and for longer than what normally happens. And so, then people started using the tools of genome editing, think finger nucleases and CRISPR Cas nine to try to interfere with this BC11A pathway. And Casgevy is based on identifying that if you make a mutation in a specific regulatory region of the BC11A gene such that it no longer gets turned on in red blood cells, that results in developing red blood cells making more fetal hemoglobin. So, you’re essentially inhibiting an inhibitor in red blood cells to turn on fetal hemoglobin that compensates or blocks the pathologic effects of sickle hemoglobin. And again, that was shown in tens of patients to not only seem to be safe over a relatively short period of time, but highly effective. These patients have dramatically changed lives. Their daily lives, which were often dominated by worries or real pain, now the pain episodes are significantly less and in some patients even seemingly disappear, and that justified and it makes it very exciting to see this now as an approved drug for patients. So I realize I just rambled a bit, but I think the story is so neat that it weaves together so much to come to this really exciting new drug.

Daniel Levine: Let’s take a step back. For listeners not familiar with sickle cell disease, what is it?

Matt Porteus: Oh, sorry. Yeah, I jumped ahead. So sickle cell disease is a genetic disease. It’s what we call an autosomal recessive disease, meaning if you have most genes in our body, in our DNA, the DNA is the code for how cells behave. Most genes, we have two copies of them. This is a recessive disease, which means both copies have to have the sickle variant. The variation is in a protein called beta globin. Beta globin is a protein that’s part of a hemoglobin molecule and hemoglobin fills up red blood cells. And the role of hemoglobin is to take oxygen from our lungs to our tissues, so our brains and muscles and all the tissues of our body work properly in sickle cell disease. That hemoglobin molecule, which is basically near crystalline in terms of its high, high concentrations in red blood cells. When it unloads its oxygen to tissues like it’s supposed to do, it picks up oxygen and delivers oxygen just like adult hemoglobin. Hemoglobin A is what most of us carry. The term for the sickle hemoglobin is hemoglobin S. When a person with sickle cell disease, their red blood cells take up oxygen. delivers to tissues, and then after it delivers the oxygen, the hemoglobin molecule changes shape and that’s normal. But what’s not normal is when the sickle hemoglobin changes shape, it starts to polymerize, it binds to itself. And so instead of being in a liquid mass, it forms polymers and these polymers change the shape of the red blood cells so that they’re not able to get through the blood vessels very efficiently. In fact, can even cause blockages, what we call occlusions or vaso-occlusions, which then prevents other red blood cells from getting through and delivering oxygen to the tissues that need it. What this results in is that patients or people with sickle cell disease don’t get enough oxygen to their bones occasionally, and this causes these deep painful crises, really deep throbbing pain. And you listen to patients describe the pain, and as I said, they call it prolonged, very deep, persistent that can be unrelenting, can last for days or weeks, but it also can cause the inability to deliver oxygen to other tissues. And so you can get strokes, you can get crises in your lungs, so you’re not able to breathe very well. It can cause damage to your kidneys. So, the disease is based in red blood cells, but the manifestation of these abnormally behaving red blood cells is to cause the organs of the body to be damaged progressively and inevitably over time, the lifespan of people with sickle cell disease in the United States is in the median lifespan, so that 50 percent of people live longer and 50 percent of the people live shorter and is around the mid-forties. The lifespan for people with sickle cell disease in Africa, though, where most patients with sickle cell disease live is in the first decade of life because they do not have access to the medical therapies to help them get through these crises that I just described.

Daniel Levine: And what’s it actually like to live with this condition?

Matt Porteus: I can only report on what I’ve heard patients tell me, and there is some variability to this. So not every patient has the same experience or the same trajectory, but generally, first of all, patients when they’re feeling good, they are just like everyone else. They’re normal human beings, but when they have these crises, which can happen out of the blue, they can also be triggered by being a little dehydrated or getting a little cold or who knows why. They can develop these pains in their bones and it can be their thigh bones, their rib bones, their back, their hips. Sometimes the pain is manageable with say some Tylenol and ibuprofen. Sometimes it needs more stronger pain medicines, but they can still take the medicines at home. Sometimes the pain is so significant they need to go into the emergency room and get even stronger pain medicines. And occasionally the pain is so strong that not only do the stronger pain medicines in the emergency room not work, but they have to be admitted to the hospital to receive continuous high doses of pain medicines, opiates to just control their pain just so it’s tolerable and that the duration of these episodes can be highly variable from being very short where they don’t come to the attention to the medical system, as I said, to patients who can be in the hospital for weeks at a time.

Daniel Levine: In January 2023, Graphite Bio, which you co-founded, reported a significant adverse event in the first patient dosed with its experimental therapy for sickle cell disease. What happened?

Matt Porteus: Yeah, so let me step back a little bit. So, the approved drug in December using CRISPR Cas9, I described it upregulates fetal hemoglobin to counteract the sickle hemoglobin, but it doesn’t do anything directly to the mutation that causes sickle cell disease. We believe that in the long-term, a best therapy for sickle cell disease using gene editing is not to counteract by increasing fetal hemoglobin, but instead go directly to the root cause and change the sickle gene to the adult normal adult gene or the sickle adult gene using CRISPR Cas9 to start the process but using something called the homology directed repair to actually repair the mutation. And so, this was a program that was developed in my lab and then licensed to Graphite Bio and they treated a patient and the patient received cells that had been engineered where the mutation was corrected and then received her cells back as part of what we call an autologous, meaning the cells are her own or the patient’s own, bone marrow transplant. Now as part of a bone marrow transplant, and this is true with all of the sickle cell disease gene therapies and gene editing therapies, one has to give high doses of chemotherapy to get rid of the stem cells, the blood stem cells that are in the bone marrow. We want to make space for the new genetically engineered cells to take and engraft and make non sickling red blood cells. So that’s a shared feature of Lyfgenia that was approved, in Casgevy, and the graphite process, the drug’s called nula-cell. Now following that sort of chemotherapy and following the infusion of the patient’s own cells, there’s a period in which the bone marrow is not making blood cells, not making blood cells like white blood cells to fight infections or red blood cells to deliver oxygen or platelets to deal with clotting, but eventually the new cells will engraft and start making the new blood cells. What happened last January was that the patient was taking a prolonged period of time to make enough red blood cells and platelets on its own. The bone marrow wasn’t making enough of those cells and so the patient was still requiring transfusions. And so that condition was called pancytopenia, meaning multiple low blood counts. Now since, and as a public company, Graphite was required to report this event and it caused a lot of disappointment. Since that time, however, the patient’s blood counts have recovered and she’s no longer needing platelets and she’s no longer needing platelet transfusions or red blood cell transfusions and she has got blood counts that show that she no longer has sickle cell disease. And so the time it took for her to get there was longer than we had hoped and longer than what is going to be needed if we want to deliver this to hundreds or thousands of patients. But eventually she got to a very good place.

Daniel Levine: Kamau launched at the end of 2023 with assets from Graphite Bio. Kamau is taking a different approach. You talked a little about the homology directed repair. How different is the approach that Kamau is taking relative to what else we’ve seen in the area of gene editing for sickle cell disease?

Matt Porteus: Yeah. So yes, Kamau is taking over where Graphite left off because the patient has ended up in a good spot and because we’ve learned so much from her, the company and my co-founders, myself and our first employees are optimistic that with what we’ve learned and improved from the first patient, we can apply this homology directed repair process even more efficiently in the next patients. And Kamau is looking forward to implementing this improved process in the next patients in the next year or so. Sorry, I missed the second part of your question.

Daniel Levine: What advantages does this approach have over what else we’ve seen within the gene editing space?

Matt Porteus: Yeah, so there’s two advantages. One is clear. The clear advantage is that we remove the sickle hemoglobin from the blood by using HDR because yes, we add hemoglobin A, but we remove hemoglobin S. And so, the first patient on with this HDR approach with nula-cell has a hemoglobin S right now of less than 5 percent, so low it would not cause any signs or symptoms or manifestations of sickle cell disease. In Casgevy, the amount of residual sickle hemoglobin is ranging between 50 and 60 percent. So, there’s still a significant amount of sickle hemoglobin in the blood, which might continue to cause some problems including occasional pain crises or some residual hemolysis–hemolysis means the red blood cells being destroyed prematurely. So, we think, and when you talk to different people they have different opinions, but I think most people would agree that if one could achieve a very low level of hemoglobin S, that would be even better than just reducing it to 60 percent. The other potential advantage is that hemoglobin A is the natural hemoglobin that we carry as adults. Fetal hemoglobin is the hemoglobin that is normally only in fetuses. There are people who have a condition called hereditary persistence of fetal hemoglobin that do have higher levels of fetal hemoglobin and they seem to do well, but they’re very rare people and we don’t really know that it’ll be perfectly normal to have high levels of fetal hemoglobin for decades or as you get older. So that’s more of a theoretic concern and we’ll learn over time, but it’s clearly going to be better to have less hemoglobin S, better for your blood, better for your blood vessels, better for your tissues.

Daniel Levine: What does the nula-cel therapy consist of and what does it take to prepare it?

Matt Porteus: Yeah, so nula-cell is similar to some of the other gene therapies and gene editing therapies. The first thing is that the patient has to have their sickle cell disease calm down. Sickle cell disease because of all the processes I described, creates what we call an inflammatory state. There’s a level of activation, physiologic activation in the cells that we want to calm down. So, it takes a few months of giving patients what we call exchange transfusions where we replace the blood that they’re making with non-sickle cell blood to calm the sickle cell disease down. That then makes it safe to undergo the next step, which is to mobilize their stem cells out of their bone marrow into their blood. So, our normal blood stem cells live in the bone marrow, that’s where most of our blood is made, but the biologic property of blood stem cells or hematopoietic stem cells is that they will come out of the bone marrow, circulate in the blood, and then go back to the bone marrow. And we’ve learned that we can give a drug called plerixafor that will stimulate the blood stem cells to come out of the bone marrow and into the blood. And then this allows us to then connect the patient to what we call a pheresis machine where the blood is pulled out of the patient. It is processed in a way so that the red blood cells are given back, but the white blood cells and stem cells are put into a bag. So, we have mobilization to mobilize the stem cells out of the bone marrow into the blood and then pheresis to purify the stem cells in the blood into a bag. Now there’s a lot of other cells in that bag of blood. The next step is that pheresis product, as I said, is a mixture of a lot of different cells. In fact, the stem and progenitor cells that we want to engineer are just a small fraction of the total number of cells in that bag. So, the next step is that the CD34 cells, which is a protein that marks stem and progenitor cells, are selected for on a magnetic column. So now we have a concentrated preparation of cells that are stem and progenitor cells. This is common between all of the protocols—that all of them share this. It is this next step that the different engineering processes begin to differ. So, in nula-cel, these CD34 cells are grown basically in a bag or in a bioreactor for a couple days to get them prepared for the genome editing process for the HDR process.

They then have the CRISPR Cas9 molecule delivered into them via a process called electroporation, and then we deliver a sequence of DNA that serves as the template for the homology directed repair process. So, let me step back. The CRISPR Cas9 makes a break in the DNA right near this sickle cell variation and that break in the DNA triggers the cell to repair the break and it can either repair the break by just sticking the two ends back together, which can be both inaccurate and accurate, or it can repair the brake by homology directed repair. But to use homology directed repair, it needs to have an undamaged piece of DNA to use as a template. And so that’s what we provide along with the CRISPR Cas9. And then the cell spontaneously will fix the CRISPR Cas9 break by homology directed repair and convert the sickle hemoglobin gene to the adult hemoglobin gene. That process takes 24 hours and then that bioreactor full of cells, which is not a few cells, it’s hundreds of millions or over a billion cells, is then frozen in a safe way. And the product gets tested to make sure that what we want to occur has occurred, has occurred at the frequency that we think is good, that there hasn’t been something abnormal that’s occurred, making sure that we didn’t create a genetic change we didn’t want to see, as well as making sure that after this processing, the cells haven’t been contaminated in any way. This is what’s called quality control. If the product passes quality control, then it will be released to be infused into the patient. So that’s how nula-cell is manufactured. I can go into then what happens to nula-cell after that if you want.

Daniel Levine: Well please, what does happen and how does it work?

Matt Porteus: Yeah, so the next step is once the nula-cel drug product, the cell product. is cleared, having passed all the quality control tests, the doctor arranges for the patient to come into the hospital and the patient then gets chemotherapy. We use a drug called busulfan, which is a very strong chemotherapy drug that destroys the stem and progenitor cells in the bone marrow to create space for nula-cel or any other genetically engineered cell. Now the problem with busulfan is that while it destroys the bone marrow cells, it also can damage the lining of our mouth and our intestines causing mucositis. It can cause some other side effects because it’s chemotherapy. Nonetheless, this is the best way we know in 2024 about how to do the bone marrow transplant. We hope in the future, and I’m optimistic in the future, we’ll be able to replace busulfan with something that’s less harsh but has the same beneficial same effects. But once the patient has received the busulfan and the busulfan has been cleared from the body, now patients get the nula-cel cells. Now we call it a stem cell transplant and I am a pediatric stem cell transplant doctor, but we’re not surgeons. We can do our transplant in a very simple way, which is we infuse the cells into the vein. We don’t have to stick the cells straight back into the bone. We can put the cells into the bloodstream by just a transfusion. And the property of the cells is that they’ll go find the bone on their own, they’ll go find their natural home on their own, and so they’ll come out of the blood and into the bone and set up shop. Now it takes some time, like I said, for those cells to start setting up shop and making more of themselves and making more mature red blood cells. And during that time, the patient is at high risk for an infection and may need platelet and red blood cell transfusions. So during this time, while the new cells are in what we call engrafting, the patient needs to be in the hospital to be safe and they need to make sure they are monitored for infections and making sure they get enough nutrition and hydration and monitoring for any side effects from the chemotherapy, the busulfan. But eventually the new cells do take and white blood cells start to be made. And we know from, I guess we’re going on 50 years of bone marrow transplant now, that once the white blood cell level reaches a certain number that the patient is then safe to leave the hospital, the risk for getting an infection is much, much lower such that it’s safe enough for them to leave the hospital. So then they are able to leave the hospital. They usually then stay close by the hospital in case something turns up. But the longer they go without anything turning up and the stronger their bone marrow gets, they then can go home and then they get regular checkups. And then as they do better and better, the checkups are spread out more and more. So eventually they’re only seen every few months or every year. So that’s sort of the trajectory that a patient goes through.

Daniel Levine: And what’s known about the therapy from the work you’ve done to date?

Matt Porteus: What we know from pre-clinical—that is from the studies that we do before we treated a patient—is that the HDR process can be very efficient. We can get correction frequencies of 50 percent or even higher of the genes corrected. We know that when we correct it in the stem and progenitor cells and we make red blood cells in the laboratory, that they make red blood cells that don’t have sickle hemoglobin. And we know the cells when we put them, the human cells, when we ask can they engraft and can they show signs of functionality, they can do that in preclinical tests, but ultimately there’s only so much you can do before you have to test it in a patient. We also know that the process itself does not generate any changes in the genome that are particularly worrisome. So before, a really important part of evaluating any gene editing drug before you test it in patients is to do a variety of different assays or different ways of measuring safety. And the FDA has provided an overview or a guidance of the types of safety assays they want to see before they would say you’ve given enough evidence to begin testing in a patient. So we had gone through all of that before we treated the first patient. And then from the first patient we learned how quickly did the cells engraft and generate white blood cells. I discussed how it took longer than expected to generate red blood cells and platelets, but now they’re doing that and we learned how the overall process worked in the first patient and from that first patient have learned an enormous amount and are implementing changes in both how the number of cells we’ll make for the next patient. We’ve learned how to make the cells even more efficiently and we’ve learned how to provide the right medicines to the patient after they receive the cells. So we hope that the next patient will get the same effect, but it will be achieved in a much quicker timeline.

Daniel Levine: What’s the development path forward?

Matt Porteus: So, the development path forward is, as I said, we learned from the first patient that we needed to improve our manufacturing. That new manufacturing process is in hand. And now we need to transfer that manufacturing process to what we call a CDMO, our contract development and manufacturing organization. These are private organizations that specialize in making cells. That’s their job. And so we are in the process of showing that this improved version 2.0 process can be made in the CDMO. They will have to show that they can do this reproducibly and reliably. That data will be compiled and submitted to the FDA saying, we’ve changed our manufacturing and we have, here’s the data and science behind why this product shows improved properties pre-clinically, and we propose to use this new process to treat the next patient. And I’m optimistic and confident and having worked with the FDA now for several years that they will approve it. I also expect there’ll be some back and forth because that’s their job and we hope to achieve this approval of the new manufacturing process to treat the next patient in 2024, and we hope we will be able to treat the next patient with an improved nula-cel in the early part of 2025. Of course, the team and I are always working hard to do this as fast, but as safe and as effectively as possible.

Daniel Levine: How have the recent approvals affected your ability to garner interest in the development of this therapy? Has it helped? Has it dampened it?

Matt Porteus: It has been both, to be honest, and I think this is good, we have a new standard. And so I think where it’s helped is it has shown that a CRISPR drug can get through the developmental pipeline and be approved and have a real beneficial effect on patients. So that’s generated a lot of excitement where it’s, I won’t say hindered, but it’s set a bar where we have to show scientifically why we think nula-del would be the next generation, the next improved therapy beyond Casgevy. And that goes back to what we were talking about earlier, the idea of not just increasing fetal hemoglobin and leaving hemoglobin S behind, but replacing sickle hemoglobin with hemoglobin A. We believe that’s the next generation and improved CRISPR therapy for sickle cell disease.

Daniel Levine: How far will existing funding take you and what’s the plan for raising additional capital?

Matt Porteus:

Yeah, we’re in the midst of raising our capital right now, so I can’t provide any details at this point. Sorry about that. But what we’re targeting is to raise capital to get through the next handful of patients. We learned from other CRISPR drugs, including Casgevy when it was called exa-cel, that a small number of patients, a handful of patients or fewer, can generate the data. That then allows you to do two things. One is it allows you then to talk to the FDA about what would be the measurements you would want to see in patients to one day get approval, and it allows you to generate the excitement to raise the capital to do that next pivotal trial.

Daniel Levine: Matt Portus, co-founder and CEO of Kamau Therapeutics. Matt, thanks so much for your time today.

Matt Porteus: Thank you very much for having me, and I hope I have enough exciting data in the future that you’ll invite me back.

This transcript has been edited for clarity and readability.


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