Building a Pipeline of Therapies to Treat Rare Mineralization Disorders
September 23, 2022
ENPP1 deficiency is a rare mineralization disorder that leads to calcification of soft tissue. About half of newborns with the condition will die in the first year of life, while others will live well into adulthood. The condition can cause hearing loss, arterial calcification, and complications involving the heart and brain. There are currently no approved therapies for ENPP1 deficiency. Inozyme is developing a therapy for ENPP1 deficiency and other rare mineralization disorders. We spoke to Axel Bolte, co-founder and CEO of Inozyme, about ENPP1 deficiency, its lead experimental therapy to treat the condition, its work with Rady Children’s Institute for Genomic Medicine to improve the diagnosis of newborns.
Daniel Levine: Axel, thanks for joining us.
Axel Bolte: Thank you very much, Danny. It’s a pleasure to talk to you and thanks for having me on your podcast.
Daniel Levine: We’re going to talk about ENPP1 deficiency, Inozyme, and its efforts to develop therapies for rare mineralization disorders. You’re focused on both the ENPP1 and the ABCC6. These are two genes that when mutated cause deficiencies that cause mineralization. These are serious and deadly conditions. Let’s start with ENPP1 deficiency. What is it?
Axel Bolte: ENPP1 deficiency is, as the name implies, a deficiency in a protein as a result of many different mutations that can occur in the ENPP1 gene. The gene codes for a protein that fulfills a fundamental role in the body and actually is implicated in the mineralization process. And so, you have to think about how we as mammals—when I say we, I’m talking about all mammals which obviously as humans are a part of—started to build an internal skeleton as opposed to amphibious creatures and others that existed prior to the emergence of mammals, and the biology that is implicated in that has a key protein at the center. And that is the ENPP1 protein. And the role of the protein is to take ATP that is extracellular and convert it to a compound that is called PPi and is an inhibitor and regulator of mineralization. So, if I’m talking about an ENPP1 deficiency, a patient with that deficiency does not produce the ENPP1 protein and therefore is unable to properly mineralize. And so, that leads to pathology that leads, as you very correctly pointed out, to a very severe disease.
Daniel Levine: Well, how does this condition manifest itself and progress?
Axel Bolte: Absolutely. That’s the key question. As a result of the deficiency, we have low levels of pyrophosphate. Again, one of the other fascinating aspects of this whole biological pathway that actually speaks to the redundancy of nature is that extracellular ATP is very important—not intracellular, but extracellular, as we mostly know ATP to be an intracellular molecule. In this case, we’re talking about a different role for ATP. It’s a substrate of ENPP1, it’s cleaved, and two things are formed. One is PPi or pyrophosphate. The other is adenine monophosphate, and that is further converted to adenosine. So, we’re interested in the biological activity of these two molecules, pyrophosphate and adenosine. The deficiency leads to very profound effects. For example, the calcification is disturbed. So, what we need, and I think is something that I only thought about once I got involved in the company, we need mineralization to occur at very defined places, namely in our skeleton. We don’t want calcification in the body to occur suddenly in places where we don’t want calcification. If that does happen, the pathological event is basically what is called ectopic calcification. It’s when your soft tissues calcify, which is obviously undesired. Other places in the skeleton itself, we will see softness or dysplasias in the skeleton. So, the skeleton won’t grow correctly, the bones will be soft, et cetera. Indeed, in patients the initial presentation is predominantly as a result of ectopic calcification. So, newborns are forming calcification in places where it’s undesired, mostly in their vascular system and mostly in their cardiovascular system so the heart and their blood supply is severely impaired. We can talk about in more detail what happens when that occurs. But the other part is a longer term effect and that is, as I mentioned, a profound effect on the skeleton. So, you have a number of skeletal dysplasias, et cetera. We can talk about in more detail, but that’s the big picture. So, you have an acute initial phase that’s mostly driven by the ectopic calcification, the vascular calcification, and then you have a chronic phase that is probably predominantly driven by the skeletal effects, but it’s generally a disease that accumulates symptoms over time as a result of the lifelong enzyme deficiency.
Daniel Levine: How difficult a condition is this to diagnose and how is it generally diagnosed?
Axel Bolte: in rare disease, diagnosis is obviously key. As you know, the physicians oftentimes have never encountered the disease as a result of the rareness. So, this is obviously a challenge we face here. The disease is quite rare, currently one in 200,000 is the quoted live birth incidence. Although we have strong evidence that it’s much more frequent, leading to only couple of hundred, if not a couple of thousand patients in the major areas of North America and Europe and so forth. So, it will be very important to educate physicians to recognize the disease. Facilitating our effort, the disease is quite symptomatic. It usually presents shortly after birth. It puts infants into a fairly distressed state. Their organism is hypoxic and for example, the calcifications that I already described occurring in the wrong place are visible on imaging. For example, on ultrasound, in some cases, the disease is detected in utero pre-birth because there’s a strong echo on the ultrasound as a result of the calcification. So, we have a rare disease with a challenge to broaden understanding and recognition of the disease, but we do have a symptomatic disease. And if you put together the symptoms like calcification hypoxia and other effects such echogenicity on the major arteries or other symptoms that are detected by ultrasound, such as pericardial effusions, et cetera. As soon as you see those, it goes very fast into the direction of what is then at that stage called GACI. GACI stands for generalized arterial calcification of infancy, which is ENPP1 deficiency as it’s named in the infant population. Now to have the full diagnosis, of course, a genetic test will be required, and we are actually offering one in partnership with Prevention Genetics to improve the detection of the disease. But clinically, it’s a quite symptomatic disease and we’re investing a lot in disease education.
Daniel Levine: Do treatment options exist today, and what’s the prognosis for patients with the condition?
Axel Bolte: So, there aren’t any approved treatment options that basically address the underlying disease at this point. Symptomatic treatment is being performed. You will see the use, for example, of blood pressure lowering agents. You will see the use of, in some cases, bisphosphonates that partially block the calcification process. However, they have longer term toxic side effects. You can also use chelating agents such as sodium thiosulfate to chelate the calcium ions to try to reduce the calcification. These things don’t really address the underlying disease, obviously so they have ultimately limited effect. You can, in other cases when we’re talking about the adolescents with skeletal effects, try to administer calcitriol, vitamin D, and so forth to help with the, with the Ricketts that these patients develop, but none of these things address the disease. So, we would be a first- in-class therapy with our approach. The prognosis is quite poor for newborns. The natural history shows us that it’s about a 50 percent likelihood to die of this disease within the first let’s say six to 12 months of life. If you are lucky enough to belong to the 50 percent that survive, adulthood is reached and we have patients that reach quite significant age. It’s unknown if there’s a later mortality associated with the disease, the oldest patients we know are in their fifties, sixties, I would say. But in the infant phase, there’s high mortality.
Daniel Levine: You’re also working on a related condition, ABCC6 deficiency. How does that relate?
Axel Bolte: Yeah, great question. I described the extracellular ATP and the fundamental role of the control of mineralization through this pathway. And just upstream of the ENPP1 enzyme is a transport protein ABCC6, it’s a channel. It is believed to be a key transporter of intracellular ATP to the extracellular medium. So, because of the closeness in the biological pathway, there is a significant overlap between both pathologies ENPP1 and ABCC6 deficiency. In the case of ABCC6 deficiency, we see two distinct manifestations. One is affecting infants and the other one is later. It’s affecting late adolescence and early adulthood and is progressing then chronically. I will talk first about the infant presentation. That one is actually also called GACI type 2. I mentioned the infant population with ENPPP1 is called GACI. So, the name already implies that the medical community sees a strong overlap. Indeed, the symptomology is very similar: severe calcifications and the ensuing complications from that. But there is a much more common, more widespread, form of ABCC6 deficiency that is really much better known as pseudoxanthoma elasticum or PXE. The diagnosis in PXE is usually occurring in adolescence and early adulthood, initially relatively mild, but then accumulating effects progressively worsening disease through the third, fourth, and fifth decade of life, a slower progression. So, as opposed to ENPP1 deficiency, slower progressing and also affecting different tissues. It is not yet quite clear exactly which and why the tissues are differently affected, but in the case of ABCC6 deficiency PXC, the adult form, the initial presentation is in skin calcifications which are visible, easily detected by seeing that the skin changes. They can be very disfiguring and disturbing for patients, but they’re obviously not life threatening. There is a large proportion of patients that develop visual problems as as a result of the calcification of their retina.
Daniel Levine: For patients who survive infancy, what is it like to live with either of these conditions?
Axel Bolte: ENPP1 deficiency, as you go through the initial infant phase and survive to adolescence, the effect of the disease is strongly related to the impact it has on growth, on the changes in the body during adolescence. So I already talked about the effects on the skeleton. Obviously we know that in adolescence, the skeletal growth is the largest, and until we are about 16 or 17, we have what is called open growth plates, and the bone is elongating and so forth. So we see a strong effect on the development of the skeleton and the patients are usually short. They don’t grow, they have soft bones, they have bowing of bones. They have bone pain as well. They continue to accumulate calcification of other tissues, of course, that doesn’t go away. They can also accumulate intimal proliferation, which is a different pathological effect that is a consequence of growth of cells in vessels as a result of low levels of adenosine. I mentioned in the beginning that there are two molecules that we’re interested in, PPi that regulates calcification and adenosine that regulates intimal proliferation. So you will find patients with stents placed in them because they have blocked arteries as a result of cellular growth. And you can also find calcifications in the auditory system. Many patients can be strongly hearing impaired, etcetera. You can find calcifications in the brain. You can see strokes happening, So, it’s what we call a multi-organ disease where it varies between patients. Some of them develop more of one and some more the other, but quite a significant burden on these patients.
Daniel Levine: Inozyme is developing in INZ-701, an experimental therapy in development for the treatment of mineralization disorders of the circulatory system, bones, and kidneys. What is INZ-701 and how does it work?
Axel Bolte: Absolutely. So, the wild type ENPP1 enzyme is a phosphodiesterase. It is a membrane bound enzyme. It’s attached to the cell membrane. The active domain of the enzyme is at the extracellular face, facing outside. There the enzyme will grab an ATP molecule and cleave it to form AMP and PPi. And our co-founder our scientific founder, Dr. Braddock, from Yale University, engineered an initial drug candidate that consisted only of the catalytic domain, which is extracellular. He didn’t bother with the membrane domain and he took that and he fused it to an Fc fragment. An Fc fragment is a heart of a monoclonal antibody or of an antibody that is sometimes, in drug development, used to attach to a molecule where you want to extend the half-life, because our goal is to have the drug ultimately float around the circulatory system for as long as possible. It’s what’s called a fusion protein. So, the catalytic domain that cleaves the ATP is fused to an Fc; the only role of the Fc is extending the half-life. And that is actually the drug. And then afterwards when we formed the company, we tweaked it a little bit and improved it. And that is now what is INZ-701.
Daniel Levine: And what’s known about INZ-701 from studies that have been done to date?
Axel Bolte: So, we tested INZ-701in animals first and we were very pleased with very strong data that showed we can raise PPi or pyrophosphate rapidly in a significant way. And we also tested whether we can prevent the calcifications from setting in. We also investigated whether we could prevent the intimal proliferation from setting in. And once we had all that we moved forward and we went to the FDA. We developed a preclinical program. I will just say at this point, that drug has been very well tolerated and safe, and therefore we were allowed to go into humans. So, we are currently conducting two clinical trials, the first one in ENPP1 deficiency where in this first in human study we enroll nine subjects and we treat them for 32 days with INZ-701every three days, or twice a week and after 32 days, we assess PK and we measure PPi, the critical biomarker. Then the patients can roll over into what is called an open label extension for another 48 weeks, basically for a total of one year pretty much where we will be measuring clinical measurements, imaging x-rays, also physical function, patient reported outcomes, and other measurements. So, we are currently in the third cohort in this trial. So far, we’ve been very encouraged by the data we’ve seen. We announced significant rapid and sustained increases in PPi. These patients have about 80 to 90 percent lower PPi than a healthy volunteer and we were able to bring that PPi level right into the middle of the range that healthy volunteers show and so, we’re, at least from a biomarker [missing audio]. Also, I should say the safety profile of INZ-701in ENPP1 deficiency. We have a second clinical trial ongoing in our second indication, ABCC6 deficiency that is currently in the second dose cohort and that we have recently announced data from the first cohort, a very similar trial design as ENPP1 deficiency, nine subjects will be enrolled in total. We have seen rapid and significant increases in PPi after the injection and then we’ve seen levels of PPi that stayed in the lower end of the healthy volunteer range. We believe in that indication, we might need to go to a higher dose and that’s actually what’s going on right now and it will be followed by the third dose, which we expect to start sometime in the fall. So, we are very excited about both of these ongoing trials. We have interested and excited patients on the studies and, based on the results we gather, determine and design how the final studies will look that should then be submitted to the FDA for approval. Those will be initiated at the end of the year.
Daniel Levine: There are other non-genetic diseases of abnormal mineralization associated with low levels of PPi. Is the reason believe INZ-701 could benefit patients with those conditions and is there any plan for pursuing those indications?
Axel Bolte: Yeah, this is a great question, Danny. I will say that we believe that this is a unique opportunity in the sense that traditional ERT, or enzyme replacement, is really quite limited to a very particular enzyme deficiency, and therefore will always be relegated to just treating that ultra- rare or rare disease. We believe we have somewhat of a different situation here because of the mechanism of action. And as I mentioned already, we are really interested in the biological activity of pyrophosphate and adenosine. Now, both of these molecules are unavailable as drugs because they have very short half-lives. You couldn’t just administer pyrophosphate. You’d never be successful doing that. Neither could you administer adenosine. So, the way to get to those molecules and the way to get them to have their biological effect is to use INZ-701. And because there are basic effects that they have, this allows us to potentially go for indications that not necessarily have an enzyme deficiency, but for example, have low levels of pyrophosphate or adenosine. And indeed, there are a number of such conditions around. For example, calciphylaxis is a condition that affects patients with end-stage renal disease. It is a complication, a rare complication that can affect between 3 and 5% of patients with end-stage renal disease. That’s actually a pretty significant number and those patients have normal, we assume normal, levels of the ENPP1 enzyme. So they don’t technically have the deficiency. However, they have very low levels of PPi. And we recently presented that the ECTS Conference in Helsinki that the PPi levels in calciphylaxis patients are actually about 80 to 90% lower than in a healthy volunteer. And so, the simple concept would be to give those patients INZ-701and see whether you can raise PPi and therefore have an effect on the disease. Now, the symptoms of calciphylaxis look, in some aspect, very similar to ENPP1 deficiency in the sense that in small arteries, there is the similar type of calcification occurring. As we see in ENPP1 deficiencies, it’s happening interestingly in the small arteries and not in the large arteries, and because the small arteries don’t tolerate any blockage at all, neither through calcification, nor through intimal proliferation. They’re highly sensitive to being perturbed in their blood flow. And the small arteries have the key role to providing oxygen to the tissue directly, ultimately into the tissues. And if you have a calcification there, the tissue is not supplied with oxygen nutrients, and therefore will die off. And the severeness of calcification comes from the necrosis, as it’s called, of the tissue and the consequences that that has. We believe if we raise PPi and if we therefore prevent the calcification of the small arteries, we will be able to prevent this life threatening necrosis that occurs in calciphylaxis.
Daniel Levine: Inozyme is participating in the Rady Children’s Institute for Genomic Medicine’s BeginNGS Partnership to advance newborn screening. Can you describe the BeginNGS program and Inozyme’s role in it?
Axel Bolte: Absolutely. I’ll just say we have a number of activities related to raising the awareness of the disease and improving the diagnosis. I mentioned already our Prevention Genetics genetic testing program, but really we are very interested in increasing diagnosis and obviously identifying patients, and that is, in our mind, best achieved with the next generation technologies. Therefore, as the name already implies, BeginNGS actually has NGS in the word—next generation sequencing—to now provide a genetic confirmation. The goal is ultimately to expand this program to almost all newborns. So, the BeginNGS Consortium is building an ecosystem that will ultimately roll out the BeginNGS program across as many hospitals that in the end, almost every newborn in the United States will be screened with it. That’s the long term goal. Rady’s Children’s has profound experience in rapid genome sequencing. The turnaround time that they have is extremely fast. And that was one of the critical attributes, because as you can imagine in a critical disease, you don’t have time to wait for weeks and days for a result of a fully sequence genome and therefore from this kind of technology leadership, they became the driver. Parties involved in the BeginNGS program are AstraZeneca Rare Disease, which is Alexion, there’s a company called Travere Therapeutics, and there’s Inozyme. We are basically the founding members from the pharmaceutical side, but there are other providers that contribute various parts of the required technology. Sequencing technology is actually in partnership with Illumina, no surprise there, et cetera. So, we are incredibly excited about this program and to be part of it. We just started and we look forward to this hopefully being rolled out rapidly over the next couple of years.
Daniel Levine: And what do you ultimately hope comes from the partnership?
Axel Bolte: Well, obviously improved detection. I mean if you have a full genome sequence, you can detect many diseases. And if that is done in a newborn screening way that ultimately makes sure that all potential patients with the ENPP1 deficiency or any other rare disease will be identified early. Early identification is critical. Newborn screening exists now, but is an incredibly time consuming and slow process. Newborn screening is often misunderstood and will only be done in indications that have an approved therapy. Only once there’s a therapy approved can you start the addition to the newborn screening protocol and it’s actually a state by state thing. So, if we wanted to get our disease on newborn screening, we would be waiting, I don’t know, for another 20 years until that happened, or maybe a little bit less. But here, the idea is that the BeginNGS platform will leapfrog that because of the power of whole genome sequencing and just immediately make all the genomic information available and significantly improve early detection of these rare diseases across all rare diseases.
Daniel Levine: Axle Bolte, CEO of Inozyme Pharma. Axel, thanks so much for your time today.
Axel Bolte: My pleasure, Danny, it’s been a fun talking to you. I enjoyed it. Thank you.
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