Kelly M. Knee, PhD, Senior Principal Scientist, Rare Disease Researcher Unit, Pfizer
Sickle cell disease is a severe genetic disorder caused by a single point mutation on the β-chain of adult hemoglobin (Hb A), β6 Glu→Val (Hb S). Numerous small molecules which covalently bind to Hb S have been evaluated clinically, however, the molecules that have demonstrated clinical efficacy all carry a reactive aldehyde group. PFE-001 is a non-covalent molecule which binds selectively to Hb S, stabilizes the oxygenated state, leading to significant impacts on RBC sickling and hemolytic anemia in SCD model mice.
TRANSCRIPT
John:
We’d like to welcome you to the Sanguine Speaker Series webinar on PFE-001, a novel non-covalent modulator of hemoglobin improves anemia and reduces sickling in a mouse model of sickle cell disease presented by Kelly M. Knee, PhD. Kelly is a senior principal scientist at the rare disease research unit that Pfizer. But first, I would like to introduce Scott O’Donnell from Sanguine Biosciences who has a few words about Sanguine. Scott.
Scott O’Donnell:
Thank you, John. And thank you everyone for joining our virtual speaker series today, and thank you to Kelly Knee and Pfizer for speaking at the webinar. For anyone who does not know me, my name is Scott O’Donnell. I’m the senior account executive for Sanguine Biosciences and I handle most of the accounts in the Northeast and Midwest. Sanguine has worked with a lot of your companies, but for anyone who does not know us, Sanguine uses a direct to patient approach to procure minimally invasive samples directly from the patient’s home via our mobile phlebotomists. We have a database of over 30,000 patients spanning various therapeutic areas, and we can work with very stringent i.e. criteria, as well as handling the IRB documents, patient consenting and obtaining the patient’s medical records for you folks.
Scott O’Donnell:
One researcher we’ve worked with over the years is Kelly Knee who is the speaker for today’s topic. Kelly Knee, like John said, has a PhD and she is a senior principal scientist in the rare disease research unit at Pfizer. She’ll be talking today about sickle cell disease and Kelly, thank you very much for volunteering to present. And we all look forward to learning more about this topic. So without any further delay, I will hand the stage over to Kelly to begin her presentation.
Kelly Knee:
Hi everybody. Thanks everyone who’s here, to hear about PFE-001. This is our novel non-covalent modulator of hemoglobin that improves anemia and reduce the sickling in our mouse model with sickle cell disease. Sickle cell disease is a neglected healthcare burden in the world at large. It’s a well understood disease. The first observations of the disease were back in the 1910s and 1911. James Herrick observed sickle cells and a patient of his and that was actually the first report of this disease in the literature. Later on in the forties and fifties, the scientists identified both that this was really one of the first molecular diseases and that the cause of the mutation was this beta six glutamic acid availing mutation. However, once we figured all of that out, there was sort of a lull in the research and one thing that’s lag behind in this disease compared to other diseases that sort of have these initial genetic components, is there really wasn’t a lot of therapeutic options for sickle cell disease patients until the late nineties.
Kelly Knee:
One of the first attempts at a drug directly for sickle cell disease was Tucaresol that was in the early nineties. It wasn’t until the late nineties that the transgenic sickle cell mouse was developed. And also in the late nineties, the FDA actually approved a drug called Hydroxyurea for sickle cell disease. This is a repurposing of a drug that was actually synthesized first in the 1860s and was originally approved by the FDA for neoplastic diseases. Later, it was tested and found to be somewhat efficacious for sickle cell disease. And so, that really became the first drug that was approved for sickle cells. However, in the recent most 20 years, there’s been a great deal of additional interest in sickle cell disease. There’s been interest both in curing the disease outright by gene therapy. A Bluebird biotech has done a great deal of work on curing a sickle cell by correcting the gene error.
Kelly Knee:
And recently, there’s been some approvals of drugs directly for sickle cell that treat either the symptoms or the disease such as basal occlusive crisis. And that would be Endari and Crizanlizumab. And also, Voxelotor was approved in 2019. And that’s a drug that really treats the root cause of the disease. So, why are we so interested in this and why do we still need additional therapies in this space? Is really because the incidence of this disease keeps going up. So we see that the distribution, as you can see in the map on the left hand side of this panel, is it’s really centered in sort of areas that are in the developing world. And in the developing world, we see that the incidence of this disease is only going to get larger. There’s only going to be more people who have this disease.
Kelly Knee:
So really when we think about sickle cell, it’s less of a rare disease in the mode of something like a beta thalassemia or cystic fibrosis. And, it’s really more of a neglected disease. So, the pathology of the disease is quite varied. The hallmark of the disease of course, is the sickled cells. Those cells get integrated into, those cells get caught up in the circulatory system when they’re in this sickled shape. And they cause occlusions. And that causes a great deal of pain. It causes a condition called basal occlusive crisis, where you get recruitment of inflammatory cells to that same space and you get swelling and it’s quite excruciatingly painful. You get infarcts in various tissues and when the infarcts happen in your brain, they can lead to a stroke. The sickled cells themselves tend to lyse quite quickly. So patients who are chronically anemic, and if they get an infarct and occlusions in their lungs, they can get a syndrome called acute chest syndrome, which is quite dangerous and one of the number one causes of mortality among these patients, they can’t clear their lungs and they basically die of an acute pneumonia.
Kelly Knee:
Sickle retinopathy is another common manifestation of this disease, but really the hallmark of the disease is the sickle cells. The sickle cells cause all of the problems and really what problems happen to each patient is very dependent on the patients. But overall, the life expectancy for these patients is quite low compared to a healthy person in the developed world. The life expectancy for a sickle patient is 40 to 50 years. That’s slowly increasing, due to improvements in treatment in the developed world. In the developing world however, the news is not as good. The life expectancy for a sickle patient in the developing world is around 20 to 25 years. And in the developing world, sickle patients under five are the most vulnerable. The mortality rate in Africa for patients with sickle cell under five is very high. It’s actually the number one killer of children under five in sub-Saharan Africa.
Kelly Knee:
So, the root of the disease is a mutation in human hemoglobin. Almost everybody who’s ever taken any kind of biochemistry class has probably heard about hemoglobin. Adult hemoglobin, which we call Hb A, exists as a tetramer. It’s two alpha and two beta chains. And it really cycles between these two conformational states that are related to the actual function of hemoglobin. Hemoglobin’s main job is to carry oxygen from the lungs and deliver it out to the tissues. And so, it does that by binding the oxygen at the hemes, which are shown here in these yellow bond stick models. So, the oxygen would bind sort of right where that orange dot is. And the difference between a hemoglobin that has no oxygen found, which we called deoxyhemoglobin and the hemoglobin that has all the oxygen’s found, which we call oxyhemoglobin. The structural differences is quite profound.
Kelly Knee:
You can see that from the sort of structure of the deoxyhemoglobin, when it transitioned to oxy, you can see there’s a major confirmation, a rearrangement. You can see the center of that protein opens up quite a bit, and that conformational rearrangement actually plays into the manifestation of sickle cell, of the disease and facilitates the polymerization of the fibers. That’s the hallmark of the disease. The mutation is a single point mutation. It’s a glutamic acid availing substitution on the surface, it’s at the beta six position and that results in sickle hemoglobin, which we call Hb S. Other than that mutation, Hb S and Hb A are structurally identical. There’s nothing wrong with Hb S. It can carry oxygen just fine. What we have with it in the case of the mutation, the beta six Glu-Val, it’s really a gain of function mutation. So, the mechanism of sickle cell diseases is fairly well understood.
Kelly Knee:
And the polymerization that is the hallmark is driven by a series of hydrophobic interactions, only when hemoglobin is in the deoxygenated state. So, it’s really driven by the presence or absence of these hydrophobic patches on the surface of the molecule. So if you look at my sort of simplified cartoon, hemoglobin adult, normal human hemoglobin Hb A has no hydrophobic residues on the surface at all. So, it’s fine. Deoxygenated hemoglobin, because of that structure rearrangement I mentioned on the previous slide, a small hydrophobic pocket emerges, and that actually emerges on both of the beta chains. They don’t want to invade it to you. I’m only showing it on one beta chain for sort of clarity in this figure. That’s only present when the hemoglobin is in the deoxy state, because there’s only one hydrophobic region present, that doesn’t present much of a problem.
Kelly Knee:
Oxygenated hemoglobin S, which is the hemoglobin that has the mutation has a hydrophobic residue on the surface all the time. That’s the beta six, which is a little patch of hydrophobicity from the presence of that Valium instead of the glutamic acid. Also, kind of not a problem. The real problem comes in when we have deoxygenated Hb S. In that case, we have both this little corner pocket that was the result of the structural rearrangement. And we have one of these beta six patches. We have one of each on each beta chain and those pockets and patches can actually associate with the pocket or patch of another hemoglobin tetramer and the way that those are arranged as they sterically line up in such a way that they can form as long polymer. It’s a very straightforward sort of hydrophobic interaction that unfortunately has this terrible consequence of eventually what happens is the cells get packed with these fibers.
Kelly Knee:
Hemoglobin is at a very high concentration inside the cells. They get packed with these fibers and they adopt this format that we see the sickled shape, that’s sort of the visual representation of the disease. We know a lot about how these fibers form inside the cell. We know that the whole process starts with a few hemoglobin tetramers coming together into what we call a critical nucleus. And so, this process of coming together and coming apart and coming together is known as a critical nucleus formation. And the time that’s required for that is what we call the delay time. And what we know about it is that it’s extraordinarily concentration dependent. So, the higher the concentration, the faster that this nucleus is going to form. And once you have a critical nucleus, the formation of a whole polymer and actually additional polymers, which build off of that initial polymer goes very quickly.
Kelly Knee:
And at that point, it’s almost too late to stop the process. So, the idea is because nucleation occurs only in the deoxy state and we know that when cells get back to the lungs to be reoxygenation, this whole process resets. The fact that it’s highly concentration dependent and it only occurs in the deoxy state means that we can see a small change in concentration will then translate to a very large increase in the time that it takes to form this critical nucleus. And really, all we have to do at that point is beat the time required for the red cells to get to the lungs to reset the process. And, we would actually wipe out the formation of this critical nucleus. And if we could just prevent that nucleus from forming, we could prevent the polymer from forming and therefore we could prevent red blood cell sickling and ameliorate most of the disease pathology.
Kelly Knee:
So our idea is, and what we’ve done with PFE-001 is we’ve designed a molecule that stabilizes the oxygenated state of Hb S, which we know cannot polymerize and thus will not manifest the disease state. This is sadly not a new idea. I wish that I had thought of it myself, but I did not. Stabilizing the oxygenated state by a kinetic approach is something that’s been attempted a number of times, starting in the idea is very simple. You stabilize the oxygenated state, as I mentioned, you inhibit sickling. If you don’t have sickle cells anymore, you won’t have anemia. You won’t get these occlusive crises. You won’t end up with end organ damage. That’s also very prevalent in older sickle cell patients. And, you would basically cut off the chain of the disease and you would have what would be representative of a substantial improvement in the medical quality of these patient’s lives, as well as the overall quality of their lives.
Kelly Knee:
Pardon me? This has been tried before, starting in the mid seventies, a group of physicians tried extra corporal carbanilation. So in that case, they actually extracted some percentage of the blood from a sickle patient, around 20%. And they modified the hemoglobin by carbanilation, and then they reinfused it into the patients. And what they found was if they did this for a number of weeks, they did it for up to by about week three of this treatment, they saw a substantial improvement in anemia. They saw about an 80% reduction in sickle cell crisis. They saw about an 85% reduction in hospital crisis days. And these patients were profoundly improved in terms of the amount of time they were in some sort of crisis. They also saw complete healing of the chronic ulcers. So, this therapy was quite successful.
Kelly Knee:
The only problem was, the method of administration was extremely onerous and it was not really feasible for a great deal of the world that is impacted by this disease. So, that approach was eventually sort of abandoned. However, the idea remained and later on, it starting in the nineties, the reactive aldehyde compounds started to come to the front of the line. These are compounds that bind the hemoglobin alpha chains and stabilize the oxy state. And they do that by forming a shifts space with the end terminal veiling of the hemoglobin. One of the first attempts to go into the clinic was a compound called Tucaresol. This compound showed significant improvements in hemoglobin concentration and anemia.
Kelly Knee:
Unfortunately, we couldn’t get a better look at what else this drug might’ve been able to do because the trial was ended early due to safety concerns that were in fact linked to the reactive aldehyde. Recently, Voxelotor was just approved for the treatment of sickle cell disease. This is also a compound that has a reactive aldehyde. In the case of the phase three trial for Voxelotor, 59% of the subjects showed a one gram per deciliter increase in hemoglobin. That was their end point. So, they met their end point. They saw significant improvements and other markers of hemolytic anemia. The study wasn’t really powered for it, but they didn’t see much of a change in VOC. And as I mentioned, the compound was approved in late 2019.
Kelly Knee:
It was clear from the experience of Tucaresol and Voxelotor that it was possible to show improvements in the hemoglobin and other markers of anemia by using a small molecule. However, learning from the experience of Tucaresol, that reactive aldehyde does in some way, can in some way pose a safety risk. We decided that we would attempt to discover a non-covalent molecule that could do the same thing, that could bind the bind the hemoglobin, stabilize the oxy state. And what we came up with was PFE-001. It is, as I said, a non-covalent binder. It binds a di-topically. So, two molecules per hemoglobin tetramer. It stabilizes the oxygenated state and shifts the oxygen affinity. And, I’ll show you some data on the next few slides that speaks to that. And as of now, we see that it has a permissive safety profile for FIH trials that we’re planning on getting into later in the year.
Kelly Knee:
But before I get to the data, one of the things that we have learned both from the experience of carbanilation and from the experience of Voxelotor, is that when you’re stabilizing the oxygenated state, the coverage level is extremely critical for efficacy. So GBT’s preclinical data, which I’m showing on the left hand side of the slide show that mice that were dosed with Voxelotor, which at the time they were calling GBT440, showed quite a bit of variability in hemoglobin occupancy at the same dose. So you can see that the two groups of animals that were dosed, they saw occupant with the same dose. They saw occupancies between 11% and 40%. And there wasn’t a really great explanation for why that was, but this variability was quite apparent. And, that actually carried on through their human trials, into their phase three data I’m showing on the right hand side, the waterfall plot from their phase three paper, showing that they’re 59% of their patients who were taking Voxelotor at 1500 milligrams showed an increase of hemoglobin greater than or equal to one gram per deciliter.
Kelly Knee:
And at 900 milligrams, 38% showed an increase in hemoglobin of one gram per deciliter or greater. But what you can see from these waterfalls is that there’s a great deal of variability in how much one gram per deciliter, or more that they got. There’s patients who did very well and saw increases of upwards of three to four grams per deciliter. And then there were patients that just made it to one gram per deciliter, and this is all in the background of the same dose. And so that variability that was apparent early on, it still seems to be carrying through to their human data. And if we contrast that with the experience of carbanilation. In those studies, patients who were carbaniliated saw an average increase of two grams per deciliter, and they saw about 30 to 50% coverage of their red cells.
Kelly Knee:
And so, it’s very clear that the better your coverage, the better your response and the more robust your increase in hemoglobin, the more likely you are to have improvements in these other factors that are important such as a basal occlusive crisis. As of now, the clinical benefits of rising hemoglobin hadn’t really been established for Voxelotor or any of the other potential therapies. However, I know that they are in the middle of doing those studies to show that linkage. And, I’m sure we’ll be hearing from them shortly. So, back to PFE-001. We took this in a very holistic way. One thing Pfizer is very good at is screening small molecules. So, we started with an affinity selection mass spectrometry screen using purified human Hb S in the oxygenated state. One of the reasons that we chose ASMS over other options was that this particular method explicitly excludes covalent modifiers.
Kelly Knee:
So, we wouldn’t have been able to find a covalent modifier if we tried. And, we actually did put them in as tests and we never picked them up. So, all we were ever going to find from this method was going to be non-covalent modifiers of hemoglobin. So from that initial screening set that had around 750,000 molecules in it, we got around 350 hits. And from those hits, we synthesized about 2,500 additional molecules to optimize the sort of PK/PD properties. And what we ended up with as our clinical candidate was PFE-001. Up here in the top right is a crystal structure of PFE-001. You can see, as I mentioned, it’s a ditopic binder. So two molecules bind to each hemoglobin tetramer, and they actually interact with each other. We know that this compound preferentially partitions into red blood cells.
Kelly Knee:
It significantly stabilizes oxyhemoglobin in vitro and in vivo. And I’m showing on the bottom, right hand side, it shows coverage dependent inhibition of Hb S polymerization. So as we increase the coverage or the occupancy, we see an increase in the percent change of delayed time. At some point you change the delay time so much that there’s nothing left to change, and it goes to zero. So, we had very favorable in vitro data that suggested that this compound could be our guy, and it was time to bring it into an In vivo study. So that was our next step. So, we’re fortunate to have a really great mouse model. The town’s mouse has human hemoglobin in this human hemoglobin S, instead of mouse hemoglobin. And it recapitulates very well a number of the features of sickle cell disease in humans, in particular, the hemolytic anemia markers.
Kelly Knee:
And so, we see that this particular mouse has a very low hemoglobin relative to the control. And, that’s mimicked by what we see in human patients. Similarly, human patients have very high reticulocyte count as do the tiny towns mice, and they have very low red blood cell numbers. And that is also mimicked by the town’s mouse. And so, it’s a very good model for sickle cell. And, we thought it was the right choice for doing our next set of studies, in vivo. We started with some single doses to understand our RPK a little better. We started with doses of 10, 40, 220 and 490 mgs per kg. And you can see in the table on the top of the slide and what we saw was quite interesting as we increase the dose, we increase blood concentration and blood bonding.
Kelly Knee:
So, our pharmacokinetics are actually nonlinear. As we see these increases in blood binding, our clearance actually decreases. And so, that results in an increase of our half-life as we increase the dose. And, this half-life sort of increases until it plateaus with a concentration in the blood of around two to four millimolar. So it’s not forever, it’s not infinite. We see that it plateaus at around two to four millimolar, which is less than the total hemoglobin concentration. So, we don’t think that there’s a scenario where we could bind all the hemoglobin, even with these favorables of non-linearity of our pharmacokinetics. Once we’ve done our single dose, we were able to hit on a dose for a chronic study. And for that, we chose 200 mgs per kg, twice daily. This is an oral study, and we dose those animals for 15 days.
Kelly Knee:
And, what we saw was that PFE-001 achieved very high levels of exposure, around 4.2 millimolar by day five of dosing. And, we maintain that coverage throughout the duration of the study. And, we had originally chosen the 200 mgs per kg, twice daily as our dose in order to achieve around 20 to 30% target coverage. We were trying to sort of mimic what we had seen in previous articles and what, what they had seen in the carbanilation paper, and really what we got was exposure that approached 60% coverage. So, we got extraordinarily good coverage. And from a dose that’s not even at the top of the dosing that we did in our single dose study. So, that was excellent news for us. And what does all of that coverage get you?
Kelly Knee:
It gets you a very apparent stabilization of the hemoglobin oxy state in vivo. And so on the top left, I’m showing two curves. The green curve is PFE-001. These are examples from our treated animals and the blue curve is the vehicle. And you can see that there’s a large change in both the P50, which is the point at which 50% of the hemoglobin is oxygenated and 50% is deoxygenated, and the P20. And that’s the point where 20% is oxygenated and 80% is deoxygenated. You can see there’s a large gap. And when we quantify that across all of our animals, we saw that 001 shows a 53.7% reduction in P50. That’s on the top plot. And an 84.4% reduction in P20. And, these numbers really suggest that we’re seeing a significant modification of our Hb S and recall from the slides a few previous that we know that coverage is extraordinarily important to getting really good efficacy in these animals and presumably in patients.
Kelly Knee:
And so, we’re very excited by these results. We also saw a substantial reduction in red blood cell sickling. In these experiments, blood was drawn from either the vehicle animals or our PFE-001 treated animals, and it was subjected to hypoxia for four hours. And then, we stained and imaged and then did quantification of the sickled cells. And you can see in the vehicle, the images from the vehicle, which are on the left, there’s quite a few sickled cells. It’s very apparent that there’s a lot of sickling in those animals. And you can contrast that with the images on the right where we see PFE-001 treated animals. You can see a lot more healthy cells, far fewer sickled cells. And when we quantify that across all of our animals, we saw a 37.8% reduction in sickle cell.
Kelly Knee:
And, this is under a hypoxic exposure. It’s quite stringent compared to what actually would be going on in vivo. In addition to the impacts on oxygen affinity and red blood cell sickling, we saw that PFE-001 significantly impacted important markers of hemolytic anemia, and also impacted some markers of vascular inflammation. So at the end of our 15 day dosing with PFE-001, we saw a 42.4% increase in hemoglobin that’s shown at the top. You can see that our treated animals actually started to approach the wild type, black six levels of hemoglobin. And, the total amount of hemoglobin that was gained by these animals was about five grams per deciliter on average. And you can see the contrast between, it basically doubled the amount of hemoglobin that the control animals had. Other markers of hemolytic anemia also showed significant changes.
Kelly Knee:
And when we look at our red blood cell count, we saw an increase, a substantial increase in RBCs. And, that actually got us into the normal range for the black six animals, as opposed to the untreated animals, which sort of stayed very well. Similarly, reticular sites, which are our marker of sort of cell production. You can see, we had a very significant decrease in retakes, not quite to the level, not quite to the baseline of a healthy animal, but substantially improved over what we saw in the untreated animals. In addition to what we saw for hemolytic anemia, we tested a few of the markers of vascular inflammation. The study wasn’t really powered to do that. So this was sort of something we tried just to see if we could see anything. And in fact, we did. One of the things that we were very pleased to observe was that PFE-001 shows a small, but still significant statistically change in soluble VCAM, which is a marker of vascular information.
Kelly Knee:
So, we were hopeful that this is sort of pointing us in the direction of the fact that we have this very high coverage, and we have this improvement in hemolytic anemia is going to translate into improvement in the vascular inflammation that then would translate into improvements in people, in VOC and other inflammatory episodes, which would be sort of very clinically meaningful for these patients.
Kelly Knee:
So, that’s sort of the story of PFE-001. In conclusion, we have previous reports that suggest that compounds, which bind in the alpha one Valine pocket of hemoglobin are able to impact both hemolytic, anemia and VOC. If we can get that target occupancy, they really need a lot of horsepower in order to see those impacts on VOC. PFE-001, we hope has the muscle to do that. It’s a potent specific and efficacious non-covalent modifier of Hb S. It shows consistent pharmacological effects at the exposure levels that we saw. It decreases RBC sickling by about 37.8%. And, it positively impacts markers of hemolytic anemia, including a 42.4%, which is a five gram per deciliter increase in hemoglobin. And based on these strong in vivo data, we have plans to advance PFE-001 to the clinic. And I see, there was a question that when are we going to further studies or in the clinic?
Kelly Knee:
And the answer is now. We have a phase one first in human study planned. The design of that study is a single and multiple ascending dose of PFE-001 in healthy adults. It’s going to be a double blind study with sequential dose escalation. And really, what we’re looking for in that study is obviously safety. We’re going to look for adverse events, of course. We’re also going to look for changes in vitals, ECG and our lab test findings. Obviously, the other thing we’re really interested in with this molecule is pharmacokinetics after single and repeat dose. And then for our pharmacodynamic effect, we’re lucky enough that we can actually read out a PK and PD effect in our healthy volunteers, the change in oxygen affinities, that we’re going to look for modulations of P20 and P50 for that study. And then of course, we’re going to also look at some differences in food, plus or minus food, and a few different drug formulations is really the plan for that study.
Kelly Knee:
So, I’ll stop there and I’m happy to answer questions, which I think are going to come in over the chat and they’re already coming in. So, the age range of the study for this phase one will just be adults. We are in discussions about pediatrics, but we start with adults for our healthy volunteers study. I also see that somebody asks how long this research took. I think as of now, from sort of day one to where we are now was about, I would say five to six years of sort of intense bench research and subsequent to that, all of the other things that needed to go on with other groups and safety evaluations and pharmacogenetic, pharmacokinetics, and formulations and everything, and how many people are on my team, I’ll show you. I hope. I’m going to try, okay.
Kelly Knee:
So, this is the team. It’s quite large. There’s no way that something like this could be accomplished without the contributions of an enormous number of people. And, every single one of them made a huge contribution. You can see that this is a large collaboration across multiple departments at Pfizer. This is really a whole company effort, at least worldwide research and development. I think, there’s people on the team from every single part of WRD. And now that we’re getting ready to go to the clinic, we have even more people from different areas. So, this covered people from Cambridge, New York, Groton, Hanover, name a site, name a Pfizer site in the United States and we probably have someone from the team who’s based there. So, it’s a huge collaborative effort. So, I hope that answers everyone’s questions about how we did the research.
Kelly Knee:
How do we think we can differentiate from Voxelotor? I think we have some ideas about that, but we’re not at this point really prepared to discuss that without, we need to see the results of our phase one trial before we can really speculate on how we might differentiate from Voxelotor. Another question. Did we see improvement in organ damage in the in vivo studies? We did not because we didn’t look for that. We think that two weeks is probably not a sufficient amount of time to see organ studies. When and if we have another study with this molecule, we would take it out a little bit longer and see if we could see organ damage. How do we measure if any of this drug to hemoglobin? I can tell you that it is to the intact tetramer. We’re saving the method for our paper, but it’ll come out soon. But we do measure the intact tetramer. It’s not just a single chain. Yeah. So, the chain listed was for the intact tetramer. We’re doing a multiple dose study, so we haven’t been able to predict, we’re not predicting yet.
Kelly Knee:
We’re not saying what we’re predicting for our dose. That’s efficacious in patients. The question was, what dose do you predict will be efficacious in patients. We’re going to wait and see, and we’re not. The question is how long will this go through the clinic? I don’t think we know the answer to that yet.
Kelly Knee:
Okay. It seems like there’s no more questions. So again, thank you to everyone who participated and came to this talk. I appreciate it. One question, BID or QD? The study that we did in the animals was twice daily. As for in human studies, we aren’t talking about that quite yet. One more question. Do you plan to combine with other agents on the market? You know, everything’s on the table at this point, but we haven’t had any specific discussions about that. One more. The reduction in flint inflammation is very encouraging, any sense of what the driver is?
Kelly Knee:
Not really, no. Like I said, this was a sort of minor effects, so we didn’t have a great deal of opportunity to really look at it. Obviously the less hemolysis you have, the less lucky you are to have sickling or sorry, the less sickling you have, the less likely you are to have hemolysis. You’re not going to have free heme. You’re not going to have sickled cells sort of cutting up the circulatory system and all of those things are going to contribute to inflammatory responses. Okay. I’m being assured that that is it. So again, thank you very much to everyone who attended. I appreciate it. And, I hope everyone is staying healthy and is going to have a lovely rest of the day.
John:
Thank you, Kelly. And thank you all for joining Sanguine for our S3 webinar, PFE-001, a novel non-covalent modulator of hemoglobin improves anemia and reduces sickling in a mouse model of sickle cell disease. For a list of upcoming webinars, please visit researcher.sanguinebio.com. If you have a need for patient samples for your research, please visit the sanguinebio.com. Thank you again, and enjoy the rest of your day.