Empowering Change through Innovative Treatments of Sickle Cell Disease

15. Gene therapy in sickle cell disease – Dr Emmauel Payen

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Summary

This comprehensive transcript by Dr. Emmauel Payen, presented at The Royal College of Pathologists, explores the potential of gene therapy in treating sickle cell disease (SCD). Detailing advancements in scientific methods, Dr. Payen explains various gene therapy techniques, from using lentiviral vectors to CRISPR-Cas9, and their application in clinical trials. While noting the challenges and risks, such as myeloblative conditioning and potential genotoxicity, the presentation highlights promising outcomes shown by leveraging fetal hemoglobin and different genomic modifications. As new therapies await FDA approval, the talk underscores gene therapy's potential to revolutionize treatment for those suffering from SCD, emphasizing its transformative impact on patients' lives in severe health environments.

Highlights

Dr. Emmanuel Payen evaluates multiple gene therapy methods in-depth, offering fresh insights into treatment strategies for sickle cell disease. 🧪
Using lentiviral vectors and CRISPR-Cas9, researchers aim to correct genetic mutations and enhance hemoglobin production. 🔍
The remarkable success of fetal hemoglobin in clinical trials prevents sickling in red blood cells. 🔄
Gene therapy comes with challenges like potential genotoxicity, particularly at on-target and off-target sites. ⚠️
The future of sickle cell treatment might lie in cost-effective, accessible therapies to reach wider populations. 🌍

Key Takeaways

Gene therapy offers hope for sickle cell patients through innovative treatments like lentiviral vectors and CRISPR-Cas9. 🧬
The use of fetal hemoglobin is preferred in gene therapy to prevent polymer formation in sickle cell disease. 👶
Clinical trials show promising results but come with risks such as genotoxicity and on-target effects. 🔬
Patients have shown significant improvements, including the elimination of severe vaso-occlusive crises. 🚑
While promising, gene therapy remains expensive and complex, underscoring a need for broader accessibility. 💸

Overview

Dr. Emmauel Payen, with deep experience in gene therapy research, offers a thorough breakdown of advanced therapies for sickle cell disease. He discusses various strategies, emphasizing lentiviral vector gene addition and the editing potential of CRISPR-Cas9 technologies. Delving into technical aspects, Dr. Payen explores how these approaches could solve inherent challenges in treating hemoglobin disorders.

One compelling focus is on the utilization of fetal hemoglobin, which, due to its nature, reduces the polymer formation responsible for sickling in red blood cells. This breakthrough offers significant hope, as clinical trials indicate not only improved patient outcomes but also address severe complications like vaso-occlusive crises. However, these treatments also come with risks, such as genotoxicity, that researchers aim to minimize.

Despite promising advancements, Dr. Payen acknowledges the hurdles of cost and complexity that accompany gene therapy solutions. The need for developing economically viable and accessible treatments for low-income regions remains a critical area of focus. As the scientific community moves forward, the potential to change the lives of thousands living with SCD shines brightly, fostering a new era of medical innovation.

Chapters

00:00 - 06:00: Introduction and Background In the introduction, the speaker begins by greeting the audience and extends gratitude to the Royal College of Pathologists and the Ghana College of Physicians and Surgeons for the invitation to present. The topic of discussion is gene therapy for sickle cell disease. The speaker briefly introduces themselves before delving into the main topic.
06:00 - 12:00: Gene Therapy Approaches The chapter begins with an introduction by Emmanuel Paya, a researcher at the French Institute of Health and Medical Research. He has over 20 years of experience developing gene therapy programs for hemoglobin disorders. His career includes working with Professor Bazar at the Institute of Hematology of the Sunway Hospital in Paris and Professor Philippe Bush at the French Atomic Energy Commission, focusing on gene therapy and vector development. Additionally, Paya has served as a consultant in the field of genetics.
12:00 - 18:00: Lentiviral Vectors and Lovo-cel Trials The chapter titled 'Lentiviral Vectors and Lovo-cel Trials' discusses the development of an antiviral vector for the treatment of beta thalassemia and sickle cell disease. The presentation within this chapter provides a concise introduction to sickle cell disease, highlighting the importance of gene therapy. Various methods being explored for addressing sickle cell disease, many of which are applicable to beta thalassemia, are covered. The chapter also delves into clinical successes related to these treatments.
18:00 - 24:00: Gene Editing Technologies The chapter discusses the current status and potential risks associated with gene therapy products. It emphasizes the prevalence of sickle cell disease as the most common genetic disorder worldwide, with over 300,000 births annually. The chapter likely details how these risks are being addressed, and concludes with an update on gene therapy products either in clinical trials or awaiting a marketing decision after the submission of a biologics license application.
24:00 - 30:00: Clinical Trial Results and Safety Concerns The chapter discusses the results of clinical trials related to sickle cell disease and the associated safety concerns. It highlights the expected increase in the number of children with sickle cell disease due to decreased mortality rates in children. It notes that most affected children are born in Africa, where nearly half do not survive beyond the age of 10. In contrast, in high-income countries, patients often survive childhood and live up to 50 years of age.
30:00 - 39:00: Future Directions and Challenges The chapter discusses the large disparities in managing health issues, particularly those that can be attributed to genetics, early diagnosis, and education for caregivers and parents. It emphasizes the importance of access to care, such as vaccinations, prophylactic therapies, hydroxyurea, blood transfusions, and pain treatments. The availability of these resources is primarily found in countries with substantial financial resources and advanced healthcare systems.

15. Gene therapy in sickle cell disease – Dr Emmauel Payen Transcription

00:00 - 00:30 well good morning good afternoon everyone first of all I'd like to thanks the Royal College of Pathologists and the Ghana College of physician and surgeon for this kind of invitation the topic of my presentation uh will be gene therapy for sickle cell disease just a few words about myself as an
00:30 - 01:00 introduction I am Emmanuel Paya I am a researcher for the French Institute of Health and Medical Research I've been working for about 20 years to develop gene therapy programs for hemoglobin disorders first at The Institute of hematology of the Sunway Hospital in Paris with Professor here Bazar a specialist in hemoglobin disorders then at the site of the French atomic energy commission with Professor Philippe Bush a specialist in laundry room vectors I was also a consultant for genetics
01:00 - 01:30 Pharmaceuticals with which later became a brewed bio with whom would develop a antiviral Vector for bit atlassemia and sickle cell disease during this presentation I will give a very brief introduction to sickle cell disease and the value of gene therapy I'll spend some time to discuss the methods being considered for sickle cell disease most of which are also valid for betaalcemia I'll talk about clinical successes I'll
01:30 - 02:00 discuss the potential risk and how they are being addressed and I will conclude by indicating the status of the gene therapy product that I will have described to you either in clinical advice or awaiting a marketing decision following the feeling of biologics license agreement application as you know psycho cell disease is the most common genetic disease in the world with more than 300 000 births worldwide
02:00 - 02:30 per year and a significant increase is expected in the coming years due to the decrease in children mortality the majority of children with sickle cell disease or born in Africa and almost half of them die before the age of 10. India is the second most affected part of the World by the disease in high income countries patients mostly passed the childhood stage and leave up to 50 years of age or
02:30 - 03:00 even older this very large large disparity is related to many factors including genetic scanning early diagnosis education of post-caregiver and parents and of course access to care including vaccination prophylactic therapies use of hydroxyure access to blood transfusion and pain treatments in countries with sufficient Financial Resource as well as sophisticated
03:00 - 03:30 hospital and infrastructures allergenic transplantation is currently recommended for all serious severe Sickle Cell patients if they have a Geno identical and familiar donor this so far applies only to the severe patient which are those who have had a stroke or are likely to have one those who suffer from freaking painful dazzleclusive attacks and recurrent episode of acute syndrome
03:30 - 04:00 in this case the benefit or considered to outweigh the risk of mortality craft versus Oz disease infections of aircraft failure well an alternative to allergenic bone marrow transportation is gene therapy unlike in the case of allogenic Bond normal transplantation with cells or obtained from an angel identical intravenger gene therapy involves the
04:00 - 04:30 patient's own self that are genetically modified X Vivo before they are injected into the patient as with allogenic Transportation the cells are injected after my looplative conditioning the difficulty of gene therapy is twofold first the rate of modification that must be as high as possible to come close to the efficacy of allergenic transportation and second the effect on stem cell must be non-toxic both in the
04:30 - 05:00 shop and in the long term Gene correction of autologous cells has quite some advantage over allergenic transplantation because it avoids not only the risk of rejection by the recipient's immune cells but also and most importantly the main cause of morbidity and mortality that affect patients who are transplanted with allergenic cells which is craft versus all disease and all the risks associated with this disease
05:00 - 05:30 another very important issue with allergenic blood normal transplantation is the fact that less than 20 percent of sickle cell patients having an available intra-familiar donor therefore in theory at least if gene therapy effectivenet is recognized as well established gene therapy should allow to treat 100 of patching patients eligible for a mild relative treatment obviously it's not black and white there
05:30 - 06:00 are also a potential risk that are inherent to a metopotic stem cell gene therapy the risk associated with the myeloblation procure which is the same as photogenic transplantation since the recipient cell must be eradicated anyway it may even be a bit a little bit higher because it seems that the mild ablation must be as deep as possible and therefore the dose of the myoblative agent must be maximized for gene therapy there is a genotoxic toxic risk which is due to the manipulation of the genome whether by the insertion of their vector
06:00 - 06:30 or by the use of Mega nucleosis a risk of course that does not exist for allergenic transplantation finally there may be risk with regard to the quality of the injected stem cell or as they are manipulated in vitro and grown for some time a few days before they are injected back and therefore their long-term persistent may change as compared to the fresh cells that are injected during allergenic bone marrow transplant airflow transplantation
06:30 - 07:00 there is therefore a clear rational first to increase the number of passion we can have access to marrow transportation and second to reduce the immune risk but there are also additional risk in Iran to gene therapy that of course need to be measured and evaluated well the question then arises as to which hemoglobin is the best to replace SQL development in order to treat Sickle Cell patients with gene therapy would admit it be more effective to
07:00 - 07:30 express bitter or government Ruby Channel well the answer as you probably know is of course gamma globin in order to produce a fetal hemoglobin given its widely demonstrated impact on disease severity a high level of fetal hemogramine has been shown to improve improve the survival the risk of stroke pain occurrence of acute chest and home legal syrup Etc so clearly it is preferable to express the gamma-gobin channel fetal
07:30 - 08:00 hemoglobin rather than the beta gobit chain of adult hemoglobin well the reason for the advantage of feet on hemoglobin over audio hemoglobin is quite simple to understand for high regression of hemoglobinastic primer to occur the tetramer needs two things a valine in position six and in hydrophobic pocket hydrophobic valine accepting pocket in position 85 to 88 consisting of a
08:00 - 08:30 vanilla lining adrionine and a lesson in the non-mutated beta hn there is novel in but still the hydrophobic pocket is present here thus the heterodimer made of one bit is and one bit I can perfectly participate in the polymer formation as shown here on the contrary the gamma chain of fetal hemoglobin is neither the valin nor the hydrophobic pocket and therefore the heterodimer
08:30 - 09:00 cannot participate in the polymer formation under this condition the btis chain included in the heterodivers were excluded from the polymer formation therefore not only hbf prevent polymerization by lowering uh SQL hemoglobin concentration but also the presence of gamma globin prevents the beta s chance from participating in the polymer formation this is the reason for which it is preferable to express a
09:00 - 09:30 gamma globin chain than the b-diglobit chain to prevent polymer formation in patient with sickle cell disease so basically there are two main strategy for gene therapy we are shown in these two red squares one on the left is to dilute acetyl hemoglobin and to inhibit cycling the second that I did not discuss yet and which may be more complicated is to repair the bis mutation the valinate position 6. for
09:30 - 10:00 Edge strategies there are several technical possibilities the first technique the most advanced and the oldest one is Gene addition using antiviral vector their vectors can express a sales cycle cell inhibitory genes such as the gamma gene or chameric beta gamma globin chain that does not contain the volume in position 6 of course and that does not contain the hydrophobic acceptable pocket the lantival vector can also Express a
10:00 - 10:30 limiter of b11a for example an sh RNA targeting b11 why bc11 this is because this protein is one of the master Regulators of chemical engine expression in the retroexality is expressed and it elevates gamma globin gene expression so by inhibiting the expression of this 11a it is possible to the express the gamma globin genes in adult erykloid cells the second technology
10:30 - 11:00 which is much more recent as genome editing this technique can be used to either reactivate gene expression or to repair the genome in one case here the technique can be used to directly correct the causative mutation of SQL services and therefore the beta is gene in another case here it consists of reactivating the expression of the gamma
11:00 - 11:30 globin Gene there are basically two options one is to use the editing system to disrupt B17 expression in the retroid cells and that is again to stop the inhibition of the gamma group in gene expression in animal cells another possibility is to edit the genome close to the promoter region of the gamma globin Gene in order to mimic hpfh mutation which are known to reactivate hemoglobin gene expression we will speak
11:30 - 12:00 about all of these possibilities in the next slides slides let's first have a look on antiviral vectors which derive from HIV one the basic principle to make a vector from a virus is actually very simple it is a matter of eliminating all the HIV genes that are responsible for the replication of the virus and keeping only the CIS
12:00 - 12:30 acting elements here that are strictly necessary for the record conscription of the Arena genome in two double strand DNA and the integration of the verb of the viral DNA into the genomic DNA of the target cells when the vector is ready the objective is to maximize the transdiction efficiency in order to obtain the highest proportion of transducer as possible then the vector and the
12:30 - 13:00 transient or stably integrated in the DNA so that the cell progeny including the eritroid cells which are differentiated from the stem cell will also contain the transient if one has introduced a nerichoid promoter here to control the transient for example gamma globin Gene then third Gene will be expressed specifically in erythroid cells
13:00 - 13:30 the small difficulty is to produce lantival like particle if the vector does not contain any virology the principle is described on this slide and is based on the use of packaging cell lines into which you need to introduce several plasmid DNA the vector plasmid DNA of course that encode the RNA genome that must be packaged and all plasmid DNA containing Gene that Express
13:30 - 14:00 the protein which are required for production of the viral particles when all these plasmids are transfected this way the packaging cells then produce a functional Vector associating the the arena genome that contain the transient but also all the protein which are required to encapsulate the urine and genome into a viral-like particle that will then be able to enter the target cells and to deliver its genome into the
14:00 - 14:30 cells in case of hematopodic stem cell they envelope that is used is the is the one derived from the vesicular stomatitis virus the vsvg envelope the receptor of which is the LDL receptor so the particle first entered the cell Bay on those cytosis and then the the RNA genome is reverse transcribed in
14:30 - 15:00 double strand DNA by a various transcriptase the various transcriptase has been Incorporated in the viral particle um so it is very transcribed in double strand DNA that is then transported inside the nucleus where the integrase that is also transported by the viral particle allows integration of the topostron DNA into the genome the most important advantage of this method is the efficiency that can be
15:00 - 15:30 quite high with recent protocol Improvement that I've shown efficacy close to 90 hundred percent importantly and it was a concern many years ago but it's not the case anymore the production method in packaging cell precludes the production of replication competent antiviruses no replication competent antiverses have been detected so far in production Lots the disadvantage as you may know of the use of antiviral vectors is their
15:30 - 16:00 semi-random integration in the genome and this of course the potential risk associated with insertional mutagenesis but this is something that we will discuss a bit later um the Second Great series of tools dedicated to gene therapy and in particular to genome editing the one that has revolutionized the world of biotechnology in the last decade and now gene therapy comes from the discovery of the system that is well known as the
16:00 - 16:30 crispuka system this tool in fact is derived from the bacterial immune system that bacteria have developed to protect themselves against foreign DNA either plasmids or bacteriophage during an infection for examples bits of foreign DNA or integrate integrated into a Locus or a bacterial Locus that is called the crispr locus then this bit of DNA can undergo a transcription to form
16:30 - 17:00 crispr Airness together with the cast nucleus they form a complex that scan forehand DNA for the presence of the target sequence and when they encounter the compatible sequence soil homologous sequence the nuclease the cas9 nucleus here snip the DNA at a very specific location within the target sequence this is the way how bacteria can fight against a virus and plasmids
17:00 - 17:30 Jennifer Duna and Emmanuel charpentier were the first in 2012 to propose that this system can be adapted and used as a programmable programmable toolkit for genome editing in your career this system in fact is pretty simple as it only requires the presence in the cells of two components first the cas9 a protein the nucleus which is made of two
17:30 - 18:00 domain there are UVC and H and H domains each domain capable of cutting one DNA strand and math as well of the guide earning the guide RNA is a short synthetic RNA that is made of scaffold sequence here in Orange which is necessary for cast binding and 20 nucleotide spacers that Define the genomic Target to be cut by the case because nucleus
18:00 - 18:30 the sequence that is recognized by the complex is cut only if there is sufficient homology between the guide RNA and the target sequence it is therefore a very versatile tool since it is enough to change these 20 nucleotide here in the RNA sequence to modify one gene or another the only constraint is to meet two conditions the first is that this one team nucleotide Motif is unique compared
18:30 - 19:00 to the rest of the genome otherwise nucleus would cut at several position and that these 20 nucleotide Motif is followed by a small motif of a few nucleotide that is called the Pam for protospacer adjacent Motif the presence of which is necessary for the cast nucleus to bind DNA for cas9 for example the Palm sequence is ntg but there are other casts and castlan variants with different Motif if required
19:00 - 19:30 so what does the case broadcast system when it recognizes a specific DNA sequence in fact thanks to these two nucleus domain it cuts the DNA on the two strands and thus Max what is called a double strand DNA break once the ones a double strand DNA break is being generated what are what are the options for cell when this happened the DNA break can be
19:30 - 20:00 reapered by two distinct pathway either the error pawn nanomologous and joining pathway abbreviated as NH EJ or the precise homology directory purpose way abbreviated as HDR the nhj system repairs double strand Break by legating the free DNA hands but most of the time it also introduced
20:00 - 20:30 deletions or specific sequence at the joining Ace hence thus creating what are referred to as in-del mutations thus in this case if a gene is targeted it will be descriptive on the contrary in the presence of a DNA templates the homology directed repair pathway follows a directed correction strategy through a recombination with homologous adjacent sequence in this case the break is exactly Ripper
20:30 - 21:00 what is interesting is that if the DNA template like this one here is slightly different from the broken DNA here with the green nucleotide for example some new DNA information can be incorporated in the corrected DNA of course it's also possible to delete DNA sequences the homology directed Reaper is obviously advantages when the objective
21:00 - 21:30 is to precisely correct a mutation but the choice between the two Pathways for the cells is not straightforward and in fact it depends on many factors including the presence and the concentration of the donor the DNA donor template the lens of the homology arms or the donor DNA and most importantly the relative activities of the repair the repair system in the cells it depends on the cells in fact and it
21:30 - 22:00 depends mostly on the cell cycle in mammalian cells the nhg password is predominant furthermore HDR Works only during a late s and G2 phases of the cell cycle whereas NH EJ worked at any time thus if cells or in g0 or G1 phases of the cell cycle as it is the case for hccs the rate of homology directed
22:00 - 22:30 repair is quite weak as compared to nhcj another problem is the activation of p53 by the introduction of Diana DNA double strand break which is further increased in the presence of double strand DNA which may generate a lot of cell toxicity the addition of all these difficulties induced that the frequency of HD are
22:30 - 23:00 corrected Target even in the presence of a DNA template is never higher than 50 in any cell type or analogous and it's even worse in long-term hematopoid extensor due to the fact that they are mostly croissant so in fact in those cells even in the presence of DNA templates at most Target side the frequency of indel U2 in HHA is pretty high
23:00 - 23:30 anyway the Chris broadcast tool is really interesting because it is a modular and programmable system it is possible to cut at the desired position in the genome either to induce a cut and generate a series of cells with small insertion or deletion which will invalidate gene or maybe in the presence of a template to correct the mutation by introducing a new DNA fragment well this is the theory because in the reality it is little more complicated than that as
23:30 - 24:00 one can imagine they can be a cut at other sides that than the desired location which is called off Target and also they can be unexpected event at the desired site itself which is then called On Target genotoxicity whereas off-target events have been well described are quite predictable based on sequence homology within genomes and the
24:00 - 24:30 problem has been largely resolved by improving the Fidelity of caste protein On Target genotoxic events are probably still underestimated well let's start with off-target genotoxicity this can happen because the cas9 nucleus can cut DNA even if the homology is not perfect and especially if imperfections happens within 10 bases far from of the guide early
24:30 - 25:00 there are basically two situations if the on and off sides or on different chromosomes the generation of a double strand breakup at the offside can induce an invalidation of Gene but it can also induce chromosomal translocation between the two chromosomes targeted by the nucleus um also if the sides or on the same chromosome I mean the on and the offsides then cleavage at the both side
25:00 - 25:30 can result in deletion or chromosomal inversion of silver kilobases or several megabases the problem with gas9 is that this phenomena had been observed with the frequency of new of a few percent under the certain condition and some location which is obviously far too high this is the reason why this issue has called Andreas creatinity um an important thing is the fact that there are a large number of
25:30 - 26:00 tool and software that have been developed for minimizing and detecting off Target side in order to evaluate the risk of of Target and obligation something that has become clear also is the fact that the cas9 protein must be present in the cells at a limited consultation and for limited amount of time in order to limit its presence to the strict requested time so that the method used to deliver the potting which must be transient is really important
26:00 - 26:30 and finally as already stated in the previous slide protection against cast 9 cleavage at off Target side have been developed especially by using cast nucleus with enhanced specificity such as iPad class 1 HiFi cast 9 or cas9 hf1 it is likely that non-specific split still exist but it has been considerably reduced and is now considered considered
26:30 - 27:00 almost resolved well while off-target genotoxicity is almost part of a past issue and today part of the evaluation following the use of the crystal cast technology in ongoing Gene Clinic of highs the awareness of on-target genotoxicity is more recent On Target a genotoxicity concern every kind of unwanted rearrangement that may
27:00 - 27:30 happen at the Target site most of the time as I just described in the limitation or below 50 base pairs so that they only affect the target Gene and even a specific site within the target Gene which can be for example of a TNA element but sometime and that has been observed in a many cell type and at several loci much larger rearrangements can happen
27:30 - 28:00 these large-scale modification includes interstitial insertion or deletion interstitial copy natural loss of atherosygosity terminal dilation terminal copy natural loss of heterozygosity or even more catastrophic events such as chromotripsis and chromosomal loss has also been observed
28:00 - 28:30 most of these large-scale rearrangement however have been observed in immortalized cell line as they mostly occur in the absence of p53 which is obviously present in primary cells nevertheless copy not how laws of heterozygosity also called also called uniparental diesel me has been observed at the frequency of up to one percent in a metabolic projective cells this kind of loss of
28:30 - 29:00 heterozygosity and duplication of one of the parental chromosome is a genetic event that has been shown in various cancers including hematologic malignancy the mechanism by which they may contribute to the merogeneous is by duplicating a mutated oncogene or by deleting a functional copy of tumor superstition of course this danger must be put into perspective for the moment since to date the patients included in clinical hives
29:00 - 29:30 in which a double strand break was induced by the crispr castline system have not developed leukemia the most likely reason for that is that the severe Adverse Events that compromise genomic Integrity typically result in cell deaths due to p53 activation however and that is the most important risk factor the risk of a selective advantage and proliferation given to some of the cell is possible especially for those of the cells lacking p53 if
29:30 - 30:00 they exist in the pool of a patient cells this issue for sure will require further investigation and is a risk factor that has to be taken into account when including a patient in such a protocol so there is a risk it is still theoretical but an existing risk in using the crispr cast system to generate
30:00 - 30:30 doubles from break as a gene therapy tool in order to reduce this risk of on and off Target mutagenesis and ideas emerge which is to use the crispr cast system not to make a double strand break but and evaluate a genome or provide a DNA template but to provide a protein whose activity would be to modify a DNA sequence thanks to this id2 technology were born namely there is editing and Prime editing in the case of Base editing we still
30:30 - 31:00 have the two basic components of the crispr cast system which has which are the cast quality in Gray and the guide RNA to Target a specific sequence in the genome but in this particular case another protein domain a seated in the amines is used to cast line in order to convert ctd into a reading within nucleotide
31:00 - 31:30 four to eight of the portal spacer leading to a u g mismatch in principle in a Cell the ug base pair should be recognized as a mismatch by the cellular repair Machinery that would normally remove the oracle the cellular enzyme required for this U here to be removed is the irrational DNA glycosylase for this reason an extra
31:30 - 32:00 protein domain and Russell glycosal glycosylase inhibitor UGI has been added to cas9 in order to prevent the reversion of U and C another modification of the system is the mutation of cast 9 the d10a mutation so that only one of its two nucleus domain the H and H nucleus Cleaves only the complementary Stone which is the non-edited Upstream from the prime
32:00 - 32:30 sequence the sneaking strap step is quite important because it will favor the use of the U here as a template the user the U on the non-neakens run to replace the G either a as shown here by the DNA mismetric repair Machinery thus after repair and replication the GC uh desperate will have been converted into
32:30 - 33:00 an 80 bus pair a very similar system has been realized in order to convert 80 into GC I'm not going to describe that into Edge detail but basically the principle is quite the same the first system that convert G in a and c e t is called acetosis based editor or cbeer and the second system that convert T in C and A G is called an adenine base
33:00 - 33:30 editor or a B the scope of Base editing system is limited to six kind of nucleotide conversion between purine here a to G and g2e between pyrimidine c2t and t2c and also I did not talk about that but it's also possible to convert G to C and C to C
33:30 - 34:00 so all of the possible combination are shown here but as you can see most base transversion are not possible especially the conversion of a teaming into a medinine as it should be done to correct SQL sentences finally and I will stop here for the description of this tool this crispr cast derived tools another approach has
34:00 - 34:30 been described in 2019 by the group of David Liu in Cambridge and this a very interesting approach as the potential to write any kind of genetic information into a specified DNA site this system can be used to perform small insertions small deletion and all kinds of product mutation well it still rely on the crispr caste system but this time the cas9 protein is fused to reverse transcriptase
34:30 - 35:00 the crisp broadcast9 protein is mutated to become a new case that will cut the Palm strain only that time is a bit different from the previous system another important element in this system is the guide RNA of course that is still necessary to recognize the target sequence but in this system this guide RNA is also fused to what is called a prime editing RNA this Incorporated
35:00 - 35:30 Prime editing RNA sequence is made of a primary binding site and a template for the reverse transcriptase I'm not going to describe the mechanism in detail it would be too long but at the end this system results in the production of two kind of DNA either the non-edited water DNA or permanent modification that can be deleting insertion or bus based conversion of the target DNA sequence
35:30 - 36:00 I have tried here to summarize the property of these tools in order to compare the most important points to my opinion in the field of gene therapy first of all the efficacy of genetic modification it's now a very high with antiviral vector and after optimization it has been shown to reach level up to 90 it's also very high when the objective is to
36:00 - 36:30 use crispr cast system to induce energy probably quite a bit less for base editing and certainly still much lower for Prime editing and it's even probably less for homology mediated especially in long-term metabolic stem cell an advantage concerning the expression level very likely goes to the repair System since
36:30 - 37:00 in those case the expression level will be physiological whereas in the case of Gene Edition by antiviral vectors then the expression level will depend on the vector copy number and the position of the vector in the gene importantly the genotoxic risk may be quite different the risk is well established for an antiviral Vector
37:00 - 37:30 which is mainly the activation of oncogen possibly for the older system based on crispr cast technology most of the genotoxic risk will depend on the rate of double strand bricks although it is considerably reduced with the base and Prime editing system comparing system in which double strand break is deliberately inserted double strand breaks still happen with Prime and best editing system so that it may
37:30 - 38:00 still be an issue with base and Prime editing it is therefore very likely that with the high number of hematopoietic cells that are modified and injected into the patients some of the cell even with this single strand brake system will have chromosomal rearrangement this of course to be looked very closely during ongoing and future clinical trials finally a word about the cost the
38:00 - 38:30 production of wire Vector is is pretty expensive in fact it's difficult to produce the vectors um and quite more expensive than the production of RNA and reboniculated molecular protein complexes so in terms of cost there is likely an advantage to non-viral technology of course that will depend on the price to at which the the company will sell the product that may be
38:30 - 39:00 independent on their prediction price of course I'm now going to describe some important result of the Amazon River vector and the addicting technology but let's first start with click on drivers with long divide vectors and I choose on purpose only to describe the most advanced project which is derived from the use of this Vector the bb305 vector this one that we set up in our laboratory together with Google bio and that contain a modified version of the beta
39:00 - 39:30 Robin Gene it's a gene with the meditation at position 87 so that the throne in of the hydrophobic pocket that binds to the mutated violin is changed into equilibin according to experiment made a long time ago at vitro and we confirmed that this modification is start to be sufficient to transform the beta agrovention in the chain having the semi-libratory activity as the one of the camera grab Institute several clinical trials have been conducted with this Vector from 2007 in
39:30 - 40:00 France up to now this clinical trial have included passion with beta-lassemia and with sickle cell disease the product of two different names according to the Target population either bet itself or transfusion dependentalassemia where the lower self will obviously impression of a sickle cell disease there have been a number of Publications already and the purification of dealing with passion with sickle cell disease or underlying the result have been published in 2017
40:00 - 40:30 for the first SQL patient included in the HCB 205 study and in 2022 for most of the patients included in the htb2 OR CIS six steps the study for which we have the more impatient and the more result is the htb2 or 6 study which was initially a phase one trial but that has been modified in a phase one two lateral also
40:30 - 41:00 the protocol has changed quite a bit with time whereas the first two group the A and B group of passion at Bond marrow cell harvested for collection here the group C patient had a mobilization with plavixa for in order to get more cell to be transduced and Infused three other important chance have been introduced the conditioning that increased from group a to Group B and C the target concentration of a visual
41:00 - 41:30 fund was 16 000 micro mode minutes in group a but 20 000 income B and C the transduction efficacy which is the amount of Gene modification per saw in the dollar product was optimized for the preparation of tripodic probation in group b and crop C and finally in order to improve the quality of the collected cells before apheresis passion in Group B and C where
41:30 - 42:00 hyper transfused from two from two months before collection so a number of modification to try to improve the protocol between passion of group a and version of Group B and C have been made over the timer the result for group A and B have been disclosed at several congresses but they have been published only recently in 2023. on this graph you see in green here the proportion of
42:00 - 42:30 HBS and in rate the proportion of the therapeutic hemoglobin you can see that for group a patient the proportion of the therapeutic hemoglobin was pretty low between 7 and 28 whereas it was up to 35 54 for the two patient in op this is a mainly related to the percentage of transducer of course measured by the average Vector copy number person in the peripheral blood which was only about
42:30 - 43:00 0.1 in group a patient while it was 0.7 and even higher than two in one of the code B patients from the result in this early included patient it could therefore be concluded that the evolution of the protocol between group N would be the Improvement of the quality of the collector cell the higher proportion of transduced cells and the year higher intensity conditioning may have collectively led to a clear increase in the amount of transduced cells and thus the amount of
43:00 - 43:30 the therapeutic protein it was thus decided by the sponsor to include more passion to clearly establish the effectiveness of the treatment as I said before a modification in group C compared to what was done from passioning of A and B was the harvesting of the hematopoietic progenitor and stem cell for mobilized blood cell instead of bone marrow with the clear objective to harvest and eject morsel into the patients
43:30 - 44:00 the target number of patients in this group of this 2D was 50 edged between 12 and 50 with a history of severe basil crazy events not result with hydroxy URL and this trial the efficacy handpoint where the resolution of severe fossil creative events the production of modified hemoglobin at level belief to be therapeutic so higher or equal to 30 and a significant increase of total
44:00 - 44:30 hemoglobin for the first 35 patients included the mean number of two mobilization cycle were required to get the correct number of cells the median number of injected cd34 cells varied between 3 and 25 million per kilo and time to neutral field practice and diffusion was 20 and 36 Days you can nevertheless see that platelet recovery
44:30 - 45:00 could be quite long for some individuals for sure longer than expected it raised of course the question of to know whether the transduction protocol may have in some cases modified the short-term recovery capability of platelets as shown here on this slide modification of the protocol translated into a very strong Improvement in all the parameters studied compared to what had been
45:00 - 45:30 obtained in group a passion the number of vector copies was between 0.5 and 3 and even higher for two patients the quantity of the therapeutic hemoglobin was around 5 gram per deciliter and the proportion of the anti-cycling hemoglobin was greater than 40 in all patients there's all creative crisis or what bothers patient the most which is why their addiction or elimination of isoclusive Crisis is an essential
45:30 - 46:00 parameter this diagram here will present the number of vasocusive Crisis B4 and after treatment while the median number of Crisis was around three to four per year before the treatment no civil crisis was noted after the treatment nor any sign of acute cystanel which is likely to be a very important factor for the Improvement of the quality of life of life of a patients
46:00 - 46:30 crisis is a major issue intensity of the emotic anemia is closely associated with mortality so both part of the pathophysiology vasopressive events and hemolysis have to be addressed it is thus very important to observe here that increased therapeutic hemoglobin production in this slide was clearly associated with an intense reduction of all hemolysis marker
46:30 - 47:00 in general the safety profiles reflected perfectly the known side effect of prescription conditioning regime there has been no case of Vino occlusive liver disease craft failure or the presence of replication component antiviruses however two patients in group a a diet of acute myelody leukemia Patient 2 first was diagnosed as a matter of display dysplastic syndrome three years after lantiglobin infusion and progressed to acute myeloid leukemia that was fatal importantly no longer
47:00 - 47:30 Vector was detected in a blast cell indicating that leukemia was not the result of insertional limited Genesis patient one is also developed acid Melody leukemia in this patient the vector was detected in blood cell but it was inserted in Gin that is not known to be involved in leukemia furthermore expression studies and chromosomal version we are enormous studies within 10 megabase of flunking Vision did not show any abnormalities finally insertion in this Gene has been detected in most
47:30 - 48:00 patients of this style without any sign of hematological abnormality it was thus concluded that acute leukemia were not linked to insertion related Genesis the question then arise was these two patients develop accumulated leukemia while hope C patient did not and beta-lassemia patient also including clinical Heights since a period of time equivalent to group a passion did not develop leukemia so what was or were the cause of acute million leukemia in this position first
48:00 - 48:30 monosomi 7 was identified in the blood cell of these two patients and mono monosomi 7 is a type of chromosomal abnormality that is frequently detected in neoplasms secondary to exposure to alkylating engines such as specified second vendors a shift in these two patients were lower than the target dose suggesting the existence of persistent non-blatted and just non-corrected hematopoietic stem cell in these two patients
48:30 - 49:00 finally patient with sickle cell disease are known to have a significantly higher rate of hematological malignancy than the general population therefore malignancy were attributed to the Persistence of inability stem cells the use of person that may have compromised genomic stability of hematopoietic stem cells which may already be affected by years of chronic hypoxia chronic inflammation oxidative stress High substance and Vascular
49:00 - 49:30 damage all susceptible to increase the rate of hematological malignancy in sickle cell disease it was also attributed to the infusion of law corrected hematopoxic external number leading to the need for more replication cycle of persisting hematopoietic stem cell and finally to the preservation of a highly proliferative and inflammatory bone marrow due to poor disease Corrections all of this factor
49:30 - 50:00 in fact told this Spencer that the protocol had to change to be changed and that's why patience or change the protocol from group a to Group B and then to group C with increased abusilifying concentration higher number of infused cells and higher Constitution level and all of this Improvement were expected to lower the risk of mirrorless plastic sandal and acute maloid leukemia
50:00 - 50:30 one patient in group C died of cardiac arrest 20 months after gene therapy however this patient was severely affected by his disease before treatment treatment with 29 severe vasocusive even before gene therapy and an important history of pulmonary hypertension and Venous Thrombosis at autopsy significant cardiac abnormalities were detected it was therefore deduced that the patient had died from complication of his disease in the same group two patients one adult first and one adolescent a bit
50:30 - 51:00 later were suspected of myelodysplastic syndrome after they presented with erythroid dysplasia and low level of treason V8 however there were no clinical symptoms suggestive of malignancy no blood cell no driver mutation no clone abdominance no chromosomal abnormality they had low hemoglobin level but normal blood count otherwise therefore the both patient both diagnosis were amended to anemia very interestingly both patient and they were the only one in the cohort had two
51:00 - 51:30 alphaglobinine dilation it tasters proposed that Alpha globin thread together with robust beta Global Production may have contributed to anemia and disabled proposes following these cases this genotype was added to exclusion criteria for ongoing studies an important question of course is the durability of the treatment and so far it seems that the genetically modified cells or maintaining themselves this is well observed here in the study that
51:30 - 52:00 started a long time ago in particularly the group A and B patient for whom we observe the proportion of stable modified cell up to five years after gene therapy it has also been seen in beta atlassemia patients who were treated with a similar positive and a similar vector and here we can see the maintenance of the phenotype correction up to seven years after gene therapy to conclude that the one-time treatment this to normalization of laboratory
52:00 - 52:30 parameters and most importantly resolution of civil vasocusive event and reduction of hemolysis markers thanks to the wet production of anticycling hemoglobin the level of Which is higher than 40 if the vector copy number is 0.5 or higher an important information is that individuals carrying two alpha group engine deleted should not receive the treatment however in another try realized in France with the same Vector the result of which have been published in 2017 and 2022 patient one had one L5
52:30 - 53:00 machine missing and it has been treated successfully with normal immunity level and with no vasocus events several results confirmed the durability of the process group a patient and thalassemia patient of course longer term observation will be required to assess the risk of hematological marination to confirm whether optimization of the treatment protocol in group C compared to group a has reduced the risk of protestant Construction in methodological cancer
53:00 - 53:30 I did not present any other data using loving expressing better but there are other tries from going with some specificity either building the gamma globin Gene for example or another modified kind of beta globin the beta S3 and with full or reduced conditioning I just choose to focus on the more advanced expressing antiviral Vector that I just described let's now uh go to uh the reactivation
53:30 - 54:00 of the measurement letter of SQL so this is the fetal hemoglobin the most appropriate regulator for this as we already discussed is transcription translation factor b17a which is an inhibitor in ideal cells as shown here of the expression of gamma Gene thus if one little bit bc11a the gamma Gene can be expressed in a rate for itself a theoretical advantage of this system compared to the addition of multicyclone genes such as
54:00 - 54:30 beta 8087q or even the gamma globin Gene is that in this case not only the are the damaging active but also the expression of the arduled pythaglobin gene is inhibited because of the inviability of the locus control region to activate the occupation of the beta this may be an advantage and facilitate the increasing therapeutic hemoglobin proportion well one of the approach to inhibit this 11a is to use a longer Vector encoding
54:30 - 55:00 in artificial Mac or iron under the control of an erythroid-specific system here similar to that use for lower cell and this feasibility and safety study seven patients have been treated they were mobilized with preceptor and condition with busulframe the number of injected cells the target which fund or the median Vector copy number and the drug product and the duration of neutrophilian blood in the government were very similar to that seen in the HP g206 Group C study for lower cell
55:00 - 55:30 the vector copy number in peripheral blood cells and the total level of hemoglobin were also similar a vector copy number between 0.4 and 1.5 and the hemoglobin concentration around 11 gram per desolate importantly the fetal hemoglobin was higher than 20 and the proportion of air cells F cells increase from 14 at bus line before treatment to 70 after gene therapy
55:30 - 56:00 the three patient P2 P3 and p7 here will received regular exchange transfusion for secondary stroke of Lexis did not receive Rich cell transfusion or add a lower number of transfusions since engravement and still had no stronger since transplantation patient with a vasocusive events before gene therapy at no severe vasoc event after therapy only patient 4 who had
56:00 - 56:30 frequent hospitalization for pre-apism before a gene therapy had several episodes of cerebralism after gene therapy but no more episodes from 8 to 20 months a recent update at Ash in 2022 reported the result for 10 patients and indicated that one patient out of 10 failed with only a very low Vector carbonara among the others they confirmed the fetal hemoglobin production and the F cell proportion that were strictly maintained over time
56:30 - 57:00 between 20 and 40 for hbf and between 50 and 80 for air cells they also reported that one patient carrying a deletion of two alpha drug engine was cleared of his vasocusive sanctum while having a lower hemoglobin level than the other patient but importantly it did not it did not present a civil form of DC Metropolis as was shown for the two patients with two alpha Rubin Gene deletion treated with lower cell in the 206 study the result does provide by this 2D give the
57:00 - 57:30 validation that this 11a can be targeted by an sh or an emir to lead to successful immigrant fetal fetal hemoglobin injection it is not clear yet whether the fetal hemoglobin level detected in the treated passion will maintain protection from cycling and preventing complication of SCD as it was a phase one study the good news is that so far no leukemia nowadays were observed but of course longer follow-up would be needed to assess long-term effect which wished to focus on its Flagship
57:30 - 58:00 products the antiglobin lower cell recently withdrawal from this program nevertheless a phase 2 multi-side pride is now open under the responsibility of the Boston children another possibility to knock down this 11a is to Target a specific uh an insert element in the BSC in the bc118 together one binding site in a region of the genome that specifically control the expression of bc11 in
58:00 - 58:30 erector itself this binding site is located in the two of b11 vertex and crispr Therapeutics are designed and tested a crystal castling system to break this site so that the cell repair system and hdk introduce interpretation and disposition a clinical trial has started in 2018 and will include 45 patients with severe SCD the protocol for mobilization and myelobation using a high Target dose of pusulfame is similar to that use for lower cell from Pro
58:30 - 59:00 build first result published in 2021 were already convincing was patient having more than 40 feet on hemoglobin a proportion that was apparently stable up to almost two years after injection at least in the first patient the most recent data reported in 2022 we have shown that three the 31 patients who had been injected so far were free of vasopressive Crisis that has to be comparable of course to the number of severe event between two and nine per
59:00 - 59:30 year during the two years before injection the 31 patient included 11 who had been free of vasopressive classic of Crisis for at least 12 months after the last Red Cell infusion the minimoglobin level was 11 gram per deciliter and the mean fetal hemoglobin proportion was 40 most Adverse Events were considered to be correlated with Bissell fund conditioning the time for investment were similar to that scene for the multi-globin bb05 and the percentage of b11a edited allele was around 87
59:30 - 60:00 suggested a very high efficiency of the editing system in long-term and metoporetics in conclusion for excessile which is a crispr cas9 a product inducing that was from break the one-time treatment leads to Neon normalization of total hemoglobin level and proportional fetal hemoglobin comparable to what was obtained for lower cell for improved bio the resolution of other occlusive events is also on track the result concerning hemolysis markers have not yet been disclosed it will be important to know
60:00 - 60:30 them given the importance of this parameter on mortality but the eye level of edited cells make me believe that it will be as good as for lovosa no hematological malignancy I know this has been observed of course and intended off target editing and on target byproducts will have to receive considerable attention there is another clinical tie ongoing including 45 patients with transfusion dependent beta classifier and their results are also of major interest with
60:30 - 61:00 42 out of 44 patients will stop transfusion and as for sickle cell disease no death no malignation there are two other small phase three studies that will have started in 2022 that will include young children with tdt and SCD and all of their 114 patients will be enrolled in long-term follow-up City to evaluate the rate of malignancy a methodological disorders and mortality
61:00 - 61:30 um The Zing finger nucleus is a bit different system it's different than the crispr cast of course but they were among amongst the first genome editing tools applying in methopedic samsa they were made of Zing finger DNA binding domain that's specifically recognized DNA fused with an endonuclease domain of the fog one Extinction The Binding to specific Locus as Focus Pro Cast line result in the generation of a double strand break and the subsequent
61:30 - 62:00 activation of antigenous DNA repair mechanism have started the clinical study in 2018 the pre-season study which should include eight patients with server SQL services from 18 to 40 years as for most of the other studies the cell collection is met upon Plavix suffer injection and conditioning is performed using pucifer first let's discuss the percentage of
62:00 - 62:30 edited cells quite surprisingly the percentage of edited alleles measured by the proportion of bc11 L is carrying in the limitation fell sharply between what was measured in the injected product and what was measured in the patient cells suggesting a relatively low editing efficiency of a long-term long-term stem cell for unknown reason this is quite different to what was observed with XSL the product from crispr therapatics and
62:30 - 63:00 the reason is alone despite this quite a distant proportion of feed on hemoglobin could be measured of the other of 30 to 40 in at least three patients and less in the force subject furthermore the proportion of f cells were pretty high around 90 in the three patient with the highest fetal hemoglobin level interestingly the number of vessel occlusive crisis were considerably reduced with only one event here in the
63:00 - 63:30 patient with the lower level of fetal hemoglobin but no events in patients with 30 feet on hemoglobin or higher unfortunately no information was given on the amolysis marker but in my opinion there is a risk that the correction will be much worse than with lovo or exacyl because of the large number of uncorrected and edited cells that will most likely Disappear by abolysis it is thus difficult to conclude on the therapeutic value of this product even
63:30 - 64:00 though decent level of fetal hemoglobin were detected and strong reduction of vasopressive events were observed as well um another possibility to reactivate him fetal hemoglobin is not to play with expression of bc11 but to modify DNA elements um in the gamma globin parameter region which are known to reactivate gamma globinistic expression there are different limitations there are mutations such as the one shown in blue
64:00 - 64:30 here that are known to disrupt the burning side for transcription or repressors such as LF or B17 and there are also mutations such as the one shown in bone here that are known to create sites for transcriptional activator for example California tarawan or gather one so there are basically two possibilities either generate an Intel mutation by using the crisp broadcast nucleus and relying on nhg repair mechanism in order to imitate this represent binding site
64:30 - 65:00 or change a base using the crispr cast derived adenine base editor system that can change a t into a c or a into a g this is what editors the company has chosen to develop an editing system based on a slightly different nucleus than Casino derived from andobacteria the objective of their protocol is to generate in
65:00 - 65:30 their mutation and disrupt the motive responsible for a piece 11 and binding site within the two gamma globin parameters according to them the cast 12a protein has two advantages overcast9 first it introduced larger deletion and this has a higher impact on the ability of b7a to bind The Motif and thus a higher level of fetal developing production per cell and second it generates a higher frequency of genetic
65:30 - 66:00 mutation a clinical Tri called Ruby has recently started and should increase 20 patients between 18 and 50. no results have been published in any Journal of abstract yet but the press release was issued by the company in December last year giving encouraging result with two patients treated and the first patient already showing over 40 percent fetal hemoglobin after five months the results will of course have to be
66:00 - 66:30 confirmed and more importantly the level of genetic modification and the search for potential often on target effect and then since the nucleus Cuts Upstream of the two genes here hpg2 and hpg-1 it is likely that a significant fraction of cells contain DNA deletion between the two calcite leaving a single Hebrew Gene with the hpg-2 promoter fuse to hbg1g whereas the installation is expected to
66:30 - 67:00 give rise to higher fetal hemoglobin as well it is not clear what may be the kind of chromosomal modification that may occur at this site maybe as we discussed earlier a higher risk of copy natural loss of atherosygosity or terminal dilation this for sure will have to be looked at carefully foreign called otq923 by Novartis and intelia similar
67:00 - 67:30 to the one from edites but based on cast 9 this time and not chaos 12a the first click on visit have been published I've been recently discussed in December 2022. they showed that engravement went well as for the other trial but fetal hemoglobin production was apparently lower as you can see here than what was obtained with cast 12 a and the Adidas system this study was interesting anyway
67:30 - 68:00 as it showed an important proportion of the cell with this 5 KB fragment deletion as expected based on the prisms of one crispr cast Target side in each of the gamma grouping promoter well even so the result the results were not too bad for some non-disclosed reason novertis has decided not to pursue the development of this program perhaps due to the fact that the result were somewhat disappointing compared to those of vertex Bluebird and editors
68:00 - 68:30 OS with the idea of directly modifying the pointer of the decarima globin Gene but also with the objective of reducing the risk of chromosomal rearrangement associated with double strand breaks whether at the expected location or elsewhere beam Therapeutics has chosen the option of using base editor they are the first one to try this strategy in a clinical trial they have chosen to
68:30 - 69:00 modify the parameter of a gamma globin Gene by introducing introducing mutation known to stimulate their expression in particular by fixing the science fiction Factor Cal F1 so far we have only individual results showing the technique to work up to more than 50 80 percent of alleles in the metropolic progenerative source and high fetal hemoglobin production in erythroat cell the tire was just authorized to start and is expected to include 15
69:00 - 69:30 subject with a severe sickle cell disease well a solution that I did not talk about yet is the use of homologous a combination to directly modify the SQL mutation I didn't speak that much about this possibility because it was thought not to work very efficiently but any anyway graphic bio used the crispr cast 9 and the guide Arena to generate a dobos
69:30 - 70:00 front brake adjacent to the sequence to be modified and they use a collecting DNA template given through an AV Vector of the AV stereotype 6 that is known to enter into hematopoietic cells so that the cellular Machinery use it to repair the brake by a copy and paste method this method they have shown it to work in human cell transplanted in human immunodeficial mice and to improve a humanized mouse model of sickle cell disease and despite a several issues
70:00 - 70:30 such important issue to my opinion such as a drop in engraftment efficiency compared to what is obtained with non-created cell such as limited proportion of corrected hematopoietic stems are around 20 only and quite large proportion of On Target indel instead of homologous recombination graphic bill was granted authorization to start a phase one to clinical trial with the M2
70:30 - 71:00 enroll 15 SQL cell patients this authorization was based probably on the fact that 10 to 20 percent chimerism in allergenic transportation is sufficient to reach significant Improvement in SQL cell asantum the first patient was injected in August 2022 but unfortunately the company alerted the FDA of an ex unexpected adverse event depression in fact remained positopenic and needed to be transfused and treated with
71:00 - 71:30 cross-factored a few months after infusion the sponsor decided on its own to post the trial and said that it was focusing on the role of a heavy 16 stems of toxicity indeed V6 is very efficient to deliver your DNA for Amorous directed or combination in many cell type at the AV genome which is sensed by the DNA damage and inade immune response pathway they also altered the survival and self-renewal capacity of long-term hematopodic stem cells this this issue
71:30 - 72:00 may be part of the explanation as to why this first patient maintained low blood cell control requiring transfusion and gross vectors report the company has thus recently decided to stop the try one of the last possibility is to correct nucleotide causing the amino acid change not by homologous recombination but by using base editor unfortunately a conversion of the valley into the normal glutamic acid is not
72:00 - 72:30 possible using base editor as there is no bezilla that can convert timing into Anatomy however it is possible to convert timing into cytosine this action would convert the volume into an alanine the presence of which is observed in hemoglobin called G maccasar hemoglobin because it was discovered in the makassa city of Indonesia this hemoglobin does not polymerize and gives normal hematological parameters in plus heterozygous and homozygostate to that
72:30 - 73:00 so that conversion of HBS to HB makazar in cells should give rise to cells that we have similarly to Sickle Cell threat the beam therapeutic company which is already looking for best editing to create a calf one site in the two gamma globin monitor has also a program to develop this modification which is based on base editor and they plan to start another clinic called high where they
73:00 - 73:30 would try this possibility uh well the also obtain impressive result in vitro with more than 80 edited in metabolic cells and a high production level of the corrected hemoglobin we have no result of course for any kind of clinical trial you may have understood now there are many options to cure sickle cell disease by gene therapy launch over Vector crispr cast to describe business expression increase broadcast to
73:30 - 74:00 describe the target of piece 11a and other transcription repressor zinc finger nucleus as well even though they seem to be a bit less efficient than Chris broadcast technology and finally the crisp broadcast derived based editing technology correction using a DNA template and homologous directed the person to be more difficult to reach today in this presentation I have shown the result for SQL services but be aware that passion with bitter plasma are treatable by essentially analytical product this is the case for antiviral
74:00 - 74:30 vectors that explains the beta organoglobination or for crispr cast-based tool that M to reactivate the expression of the gamma group in genes so there have been the same kind of trial and result with beta Plus in your passion and finally the only gene therapy product approved today is beta cell for a beta glycemia patient in the US which is the rubric bio product and which shows exactly the same technology as that described for novocet betiser was already accepted in Europe in 2019 due to a lack of agreement on the reimbursement terms which Germany
74:30 - 75:00 England France Italy and Greece private bio unfortunately for the passion disengage itself from its activity and the European market even though the pipeline of flexible project for SQL Services was apparently overcrowded and it may be the impression that you have after this presentation it now seems a little less crowded two candidates indeed have recently dropped out of the race intelia Novartis or txu923 and graphic pure new lesser sangamo is now
75:00 - 75:30 looking for a new partner for phase 3 development but the project might be too far behind Chris praveatics exercise to draw merge interest and also agree with bio has decided not to support anymore the bb694 project to focus more on the development of global cell in total there are two complementary different products the oversell from grouped bio and excess cell from vertex and crispr Therapeutics one based on another non-verbal vector and the other one in crispr gas technology these two companies have very recently submitted
75:30 - 76:00 their product for FDA approval for Bluebird bio and also for EMA and UK approval for vedics of course follow-up studies over many years will be needed to confirm if this product are two one-time therapy and to what extent these two probably correct both vessel occlusive event and hemolysis the impact of this treatment on long-term correction of the disease are known quality of life will also have to be studied and compared to that measured 2.1 micro transplantation
76:00 - 76:30 one of the most important issue for sure today is patient safety and it will be important to understand long-term risk associated with these two therapies the rate of hematologic malignancy in the hbz206 group a patient was particularly High even though it was nothing to insertional metagenesis it will be very important to confirm that protocol modification was sufficient to reduce the risk also today clearly there is not enough experience with double strand brake technology will be essential to carefully follow the patient in the long
76:30 - 77:00 term this is why the recently developed base editing technology such as the one developed by therapeutic events so there is no data yet based on a single strand break could prove interesting even if it seems to come a bit late compared to that of vertex and bluebird Point only time will tell it the competition is much work on thanks to editors and beam Therapeutics given the setback of Betty cell in Europe for which Bluebird bio and the
77:00 - 77:30 institution responsible for reimbursement have not been able to agree on the price much to the display of a patients so here is the take on message we can say that hemoglobinopathies and beta globin Gene limitation have been having to stimulus for the search for gene therapy solution as you have seen there a considerable number of solutions for the treatment of sickle cell disease and hemoglobin disorders in general with gene therapy two gene therapy product today based on
77:30 - 78:00 very different technology or weight in marketing approval for sickle cell patient in 2023. the progress of crispr cast technology has has been meteoric but clinical height in this field have benefited from the experience gained in multiple vector class the next step for sure will be individual gene therapy the number of sickle cell patients and the presence in low-income countries will not load the majority of them to be treated by Hypersonic sophisticated and too
78:00 - 78:30 expensive techniques as they were tutor