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What I want to present to you today is clinical part of our work at CellProthera.
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CellProthera is a French company located in the East of France, just at the border with Basel in Switzerland. The company was founded in 2008 by me and it's goal is to treat acute myocardial infarction patients using CD34 positive cells.
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As you know heart disease, more specifically heart attack is a leading cause of death worldwide and about 10% of cases will die within the one an hour and of the survivors about 20-25% will suffer severe infarction.
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And this leads to irreparable destruction of 1 to 2 billion myocardial cells and it's causing an irreversible secondary heart failure with a poor prognosis in the short-term. In this case 50% of the patient died within five years and all the patients died within seven years.
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And as you know, the heart is a totally differentiated organ with a very low possibility, very low capacity for self-regeneration.
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And there is no current treatment to prevent this fatal outcome, but we can slow it down.
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And for these reasons, cell therapy has generated an enormous hope, but achieved somewhat disappointing results.
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Depending on the type source of cells which were used, there were many clinical trials performed using bone marrow mononuclear cells.
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And you know, bone marrow nuclear cells contain very few stem cells.
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So most of them failed to prove interesting results.
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CD34 positive cells have been discovered by Curt Civin in 1984 and he developed a monoclonal antibody to mark the cells and to determine the cells, to qualify the cells, to quantify the cell.
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You know these cells have a big nucleus surrounded by cytoplasm and they are homogeneous but at the beginning, the CD34 positive cells were supposed to be only hematopoietic stem cells.
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When Curt Civin described it, he thought that it was only hematopoietic stem cells.
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And for many years it was the paradigm, CD34 positive cells are hematopoietic stem cells.
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This population was not absolutely homogeneous because there were more undifferentiated cells, a small part of more undifferentiated cells which are CD34 high they are more differentiated, more committed cells which are progenitors.
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which are CD34 positive, CD38 negative and positive.
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So there's a mix of these two categories of cells with an advantage for the more committed cells.
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But the revolution occurred in 1997 when the Japanese researcher, Asahara, showed that the CD34 antigen was also borne by other cells different to hematopoietic stem cells.
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It was borne by stem cells.
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So it was the first time that it was demonstrated that in fact the CD34 positive cells were not only hematopoietic stem cells, but can also be stem cells for other tissue.
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So starting from this paper published by Asahara, I decided in at the very beginning of the 2000s to try to use CD34 positive cells not for hematopoietic purpose, but for treatment of acute myocardial infarction .
7:05
And I launched pilots today, but with a very low number of patients, a small court of patients, it was only 7 patients.
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But these patients had very bad prognosis and they underwent CABG operation, but not through perfusing the ischemic area.
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And it was only a compassionate operation to not to prolong the life, but to make the life more easy before their deaths.
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And this time we harvested CD34 positive cells from the blood of these patients and we purified by immune-selection the CD34 positive and these cells were reinjected at the end of the CABG operation by epicardial injection.
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This is the shimmer of protocol, of course infarction.
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Then we mobilised the cells from the bone marrow into the blood by injection of G-CSF for five days.
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And on the 6th day, we performed leukapheresis to collect CD34 positive cells.
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And we sent the sample to the to the laboratory for immuno-selection.
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And at the end of the immuno-selection, the cells responded in human albumin and they were sent directly to the to the operating room and they were injected directly into the heart, into the ischemic lesion at the end of the CABG operation.
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And then after that we performed the PetScan for all these patients.
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And three of these patients were scheduled originally for a heart transplantation, but they lacked a donor.
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So they entered our protocol.
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And what we saw when performing the PetScan examination in these patients, this is the ischemic area, infarction area before cell injection.
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So you see there is no fixation of glucosis and six months after you have a fixation of glucosis in this area except at the apex.
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And when using another marker and glucosis, an ammonium marker which is a marker for vascularization, we also observed there was vascularization of this area.
10:15
From six months we obtained both regeneration of the cardiac tissue on the vascularization of the cardiac tissue.
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This is a second patient who should have undergone heart transplantation.
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He was a young patient, so he was 33-year-old, but he was an alcoholic and a smoker and he suffered a very severe myocardial infarction .
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You see, almost no fixation points before cell injection.
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And he was referred for heart transplantation immediately, but the transplanter didn't want to do that because he said, well, we have to transplant the heart and the maybe lung.
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And so he’s not a good patient for us.
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And he refused to try to find a donor.
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But at three months, he began to be a little bit better and that's at six months.
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We also observed the partial reparation of the tissue.
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If you compared this, this is at six months and this is at the beginning before cell injections.
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So we observed partial reparation.
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Of course, it was a very important myocardial infarction .
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But this patient is still alive, alive 18 years after the cell injection and any of these patients who were scheduled for heart transplantation, they did not receive heart transplantation.
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They avoided heart transplantation and two of three are still alive.
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One died last year after 21 years of follow up.
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This is the third patient who should have received heart transplantation.
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It also you can also see there is a complete regeneration of the tissue compared to before the cell injection.
12:31
We have long-term follow up here.
12:39
So one of the first patients failed to respond because they were not a good patient.
12:49
We collected the cells on the and we injected the cells eight years after the infarction, at this time we did not know who the good patients were.
13:01
And after that, of course, we reduce the time of injection to a maximum of six months.
13:15
And as I told you, we have recently updated the results and if you look at the six survivors, they have survived in good enough conditions for at least 12 years after cell therapy. when patients died from a stroke during the night, it was 84.
13:41
When patients died from a stroke during the night, it was 84. When we when we collected the cells and we infuse the Celsius was 73, so 72.
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And another patient has died from pancreatic cancer and another patient is now suffering from Alzheimer's disease.
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So we allow the patient to have another disease related with the age.
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This is a curve of infarction for this patient.
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And you see this one, this patient which was a good patient at the beginning suffered again of second infarction.
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But she's now 81-years-old.
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What are the key factors we have determined from ours today and from the publications in by other groups.
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Of course, the cell type for us is CD34 positive cells.
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What is important is the number of cells because if you consider severe infarctions, there were one to two billion cardiomyocyte cells which are destroyed.
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And if you inject only a few stem cells, maybe 100,000 or 1,000,000, it's not enough to repair to repair the heart.
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So you have to inject as many cells as possible.
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And the route of injection is also important because many clinical trials have used the intracoronary injections and that means that the cells were injected is into the unobstructed coronary artery.
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But the cells in this case are not retained by the lesion.
16:11
And they pass through the lesion and they are going to the spleen, to the bone marrow, to the liver, but they are not retained by the lesion.
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So what is the most important is to know to inject the cells directly into the heart, into the myocardium.
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And there are some catheters which have been developed to do that.
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And the timing of injection is also important because if you wait a too long time, the operation will fail.
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It’s better is to inject the cells between one or two months after the infarction, not too early, because if you inject one week after, the myocardium is too fragile.
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So where you have to wait for partial reconstruction of the ventricular wall but to work, the scars must still be inflammatory.
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Because if in this condition, the scar secrete chemokines and these chemokines will activate the injected cells.
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If you wait six months or more, the scar becomes fibrotic, it's sclerotic and it's not a good thing for selection which will die if you inject it in this area.
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So what we propose as mechanisms for CD34 positive stem cell cardiac regenerative medicine.
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First there is of course there is a cardiac lesion, the cardiac lesion, inflammatory cardiac lesion before secreting chemokines.
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Chemokines, it's an exocrine effect and these chemokines will stimulate the CD34 positive cells and from this time there will be two way of action.
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The first one is cell multiplication and then the differentiation and they will differentiate into cardiac progenitors, which finally will provide cardiomyocytes.
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And the other way is the secretion, the release of exosomes and growth factors which will have a paracrine effect and will favour the new vascularization.
19:18
So the first there is a tissue repair and then the vascularization using two way of action.
19:29
So the procedure we use for the pilots today was very difficult and demanding for the patient and it could not be performed everywhere but only in a select few specialised centres.
19:42
This would limit the access to the therapy to a small number of my patients and the CD34 positive cells used for cardiac repair, was a hematopathic indication and not considered as an ATMP.
20:11
So it's no bio drug, it's bio medicine and we have to prepare the cell graft in in GMP conditions and it's still an important point.
20:29
Of course, we are here in a meeting showing many possibilities for automation from the production of stem cells.
20:42
And it's very important because we are at the crossroads to pass crucial turning point to reach the market.
20:52
And that means that it needs a development of HCL tools allowing the good manufacturing practise, observance and reliable and stable production of cell grafts, which are considered as biotherapies ATMP.
21:11
Also, we have developed an automated device that would allow GMP stem cell expansion in vitro from whole blood samples withdrawn after G-CSF mobilisation to yield CD34 positive stem cells numbers at least equivalent to those collected during one leukapheresis procedure, thus avoiding the need for the latter.
21:34
This is the first machine we developed, in fact it was an experimental machine and we bought an incubator and we only performed the system for cell expansion and so on.
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And we use this machine which is called StemXpand.
22:09
We use this machine for the first two we I will present after that, but now we are developing a new one, much better performing and much more modern.
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We have a lot of bags, tubules and so on.
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Here the medium is contained in a cassette.
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And this cassette is like a pizza.
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You put a pizza in the in the oven and it cooks for 9 days.
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So this new machine, new automat can perform 5 grafts at the same time and this one only one,
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What we obtain with the incubator, we obtain an expansion of about 20 fold comparing to the number of cells contained in the sample collected before expansion.
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For example, if we start with 5 million CD34 positive cells put in culture, we obtain about between 50 and 100 million of cells.
23:36
And we have developed a potency test because they have the same potency as native CD34 positive cells with a significant correlation between ProtheraCytes and secreted VEGF concentration.
23:55
We have the recently presented the result of excellent Phase 1/2b study and ProtheraCytes cell therapy in acute myocardial infarction which was presented by cardiologists in Lisbon in May. This was this is a protocol with 44 patients randomised.
24:21
They received standard of care and stem cells and in the other arm it was this standard of care.
24:34
Well, this is a flow chart, but not very interesting. The baseline patient characteristic showed there was not any significant differences from one to another.
24:55
This is a route of injections.
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We use a catheter which is on the market and which is produced by a US company located in the Silicon Valley near Los Angeles.
25:15
And this catheter is introduced in the femoral artery pushed until the left ventricular.
25:24
And you see it can be oriented to be perpendicular to the to the wall.
25:31
And we at the end of the catheter there is a helical needle.
25:37
And in fact the cardiologist screw the needle into the myocardium.
25:48
Well, we have some problems with this injection because we had several tamponades.
25:57
But you know, it's a problem of formation of the cardiologist. Sometimes they inject the cells when said no, please do not inject the cells because the cardiac wall is too thin.
26:19
And of course they did it.
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So we have to correct that.
26:23
These are preliminary results, main results of efficacy endpoint.
26:28
We have observed significant reduction of the size of the infarction and more particularly when there was there was an obstruction of the micro vessels around the infected area.
26:49
We also observed it's not absolutely significant because it's 0.05, but there is a consistent trend of volume improvement of the heart after the cell injection.
27:07
And we have also strong reduction of the level of NT-pro-BNP which is also significant at three months.
27:18
We will see later if you look at the congruence of efficacy in points, all the end points are going in the good way.
27:30
I’ll remind you that it's only the follow up was only six months.
27:34
And in our pilots study we demonstrated that the improvement of the of the cardiac parameters occurred over two years.
27:50
At two years there was a plateau.
27:53
And if you look at the quality of life, there are also significant differences between the control group and the group receiving the cells in for most parameters.
28:09
So in conclusion, the regenerative therapy using ProtheraCytes injection after a post acute myocardial infarction emerge as an innovative strategy to prevent heart failure associated outcomes and the trans-endocardial ejection of ProtheraCytes was feasible, but it we may improve remedy.
28:35
It may improve remodeling as suggested by congruent directionally favourable changes in infarcted zone area, LV dimensions and NT-proBNP.
28:50
Of course, the risk mitigation strategy, we will announce procedure planning and operator training to address the endocardial injection learning curve, which may optimise safety.
29:06
And we also will reduce the volume of the of the cell suspension because we inject 15 millilitres of cell suspension and we will reduce at 10 millilitres. Thepotential long term 2 years remodelling effect of ProtheraCytes versus standard of care in the post AMI setting will be further evaluated in a plain large randomised control phase-3 trial.
29:30
We have also launched an observation study concerning the patients of the phase two to prolong the follow up for two years as we did during the pilots today.
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And we expect to improve the hard parameters.
29:51
Thank you for your attention.
