128th ENMC International Workshop on ‘Preclinical optimization and Phase I/II Clinical Trials Using Antisense Oligonucleotides in Duchenne Muscular Dystrophy’ 22–24 October 2004, Naarden, The Netherlands
Article Outline
- 1. Comparison of the two ongoing projects
- 2. Strategies for the identification of effective AON
- 3. Different AON chemistry and carrier polymers
- 4. Efficacy and toxicity of AON in human: experience from other therapeutic trials
- 5. Dystrophin gene restoration therapy following direct injection of plasmid DNA: experience from the French trial
- 6. AON in DMD: selection of patients
- 7. Validation of AON effects in patients muscle cell cultures
- 8. Animal models for future developments on AON delivery and optimization
- 9. AON in DMD: discussion regarding the two protocols
- 10. AON in DMD: immunological aspects
- 11. AON in DMD ethical aspects and parental perspective
- 12. Conclusions
- Acknowledgements
- Workshop participants
- References
- Copyright
Twenty-six participants including parents, scientists, industry representatives and clinicians from Australia, Belgium, England, France, The Netherlands, and USA attended the 128th ENMC workshop on the topic of ‘Preclinical optimization and Phase I/II Clinical Trials Using Antisense Oligonucleotides in Duchenne Muscular Dystrophy’. The meeting was held in Naarden, The Netherlands, during the weekend of the 22nd–24th October 2004.
The aim of the meeting was for the two European consortia (one in The Netherlands/Belgium and the other in the UK) that are preparing for a clinical trial on antisense oligonucleotides (AON) in Duchenne Muscular Dystrophy (DMD) to meet and compare their respective protocols. In addition, the meeting was attended by experts on AON in DMD from other countries and by experts on the use of AON in other fields of medicine. Representatives of four companies, ISIS Pharmaceuticals, Prosensa, Afforce Healthcare and Transgene attended the workshop as well, together with a parents representative.
DMD is a common and severe form of muscular dystrophy, caused by intragenic mutations in the dystrophin gene. The majority of these mutations are out-of-frame deletions (and duplications); in-frame deletions/duplications characterize the milder allelic variant Becker muscular dystrophy (BMD).
Recent laboratory studies have shown that the addition of AON to cultured patient muscle cells, and their injection into muscles of a mouse model for the disease (the dystrophin deficient mdx mouse) can induce skipping of exons and restore the reading frame in these cell lines and animal models [1], [2], [3], [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14]. While the limitation of this approach is its temporary nature, and the need for the direct intramuscular injection of the AON, its efficacy in terms of restoring dystrophin expression and assembly of the dystrophin associated glycoprotein complex have improved considerably in the recent past. In the mdx mouse, this was also followed by improved functional properties of the dystrophic muscle [10]. Additional significant development will be necessary to improve the delivery aspects of AON before the antisense approach could be regarded as a realistic therapeutic option in DMD. Development of systemic delivery systems for AON are being pursued by a number of groups; in addition there is a wealth of expertise on the use of systemic delivery of AON in other pathologies. It has been calculated that approximately 70% of DMD patients with intragenic deletions could be theoretically helped by this approach. For a recent review of the literature see [7], [15].
1. Comparison of the two ongoing projects
The meeting started with a brief outline of the work planned by the two consortia. Regarding the UK project, funded by the Department of Health, Francesco Muntoni (London) explained that this comprises a preclinical and a clinical part. Regarding the preclinical studies, these involve testing novel AON targeted against exon–intron boundaries of exons 51 and 53; testing the efficacy of exon skipping in DMD patient myoblast cultures; in developing systemic delivery strategies. Regarding the clinical part, the work will focus initially on the characterization of the histological and muscle magnetic resonance imaging (MRI) of the state of preservation of a number of distal muscles (radial; tibialis anterior; extensor digitorum brevis) to decide the best target for the intramuscular injection. The age range of patients selected will likely be between the age of 14 and 18 and all patients studied will have previously had a muscle biopsy in which dystrophin expression has been studied. The specific deletion that will be targeted has not been decided yet, but it is likely to be either exon 51; or exon 53, or two groups of patients, one with a deletion which could be rescued by skipping exon 51 and the second group by exon 53. For a list of the out-of-frame deletions that could be targeted using a similar approach see Table 1. The plan is to administer a combination of AON and carrier polymer (similar to that used in recent animal studies) to three groups of patients. The first three patients will receive a relatively small dose of AON+polymer in the target muscle following direct injection. A muscle biopsy will be performed after 1
month. In the second group, the protocol will be identical, although a higher dose of AON will be used. In the third group, the best tolerated and most effective dosage will be used; however, after 1month from the first administration, a second AON administration will be performed. A muscle biopsy will be performed 1month after this second injection.
Table 1. Overview of the DMD-causing mutations, correctable by skipping one of the exons that will be targeted as part of the work performed in the two consortia
| Skippable exon | DMD patients potentially benefiting (exons deleted or duplicated) |
|---|---|
| 51 | 45–50; 47–50; 48–50; 49–50; 50; 52; 52–63 |
| 53 | 5–52; 47–52; 48–52; 49–52; 50–52; 52 |
The protocol from the Dutch/Belgian consortium, set up and managed by Prosensa BV (NL), was presented by Gert-Jan van Ommen (Leiden). Both the preclinical and clinical development is funded by Prosensa, the Dutch Government, Dutch Parent Project and UPPMD. The Consortium has already identified the most effective AON sequence, directed at an exon-internal, putative splicing enhancer element within exon 51. This AON will be administered to a group of four patients (age range comprised between 8 and 16 years, carrying out-of-frame deletions which would benefit from this approach (Table 1). The patients will be selected on the basis of a positive outcome, i.e. specific exon 51 skipping and dystrophin production, in cultured muscle cells isolated from muscle biopsies previously taken from these patients. The protocol involves administering a single dose AON (h51AON23) by multiple injections within a 1cm2 area of the tibialis anterior. A biopsy will be taken on day 28, for analysis on RNA and protein level. All biopsies are to be performed by the same investigator, using a well-established procedure involving a small-beaked conchotome. Several safety parameters will be evaluated during and after the treatment of the patients.
2. Strategies for the identification of effective AON
The next item discussed was the strategy that the different groups were following in order to identify effective AON. Judith van Deutekom (Leiden) described how AON were selected: two AONs were designed per exon, mainly targeting (partly) open structures, and, if possible, directed to purine-rich sequences that may represent putative splicing enhancer elements (ESE). While initially gel mobility shift assays were performed with the different AONs to determine which one binds with most favorable affinity, nowadays a software package (ESEfinder) is used to predict binding sites for the different splicing factors (SR proteins) in more detail. The more specific targeting of these sites typically yielded effective AONs in 67% of molecules tested. Only rarely the design of additional AONs was required. Following this strategy van Deutekom has already identified effective AONs for the skipping of 36 of the 79 dystrophin exons [13].
Ian Graham (London) explained the strategy he and George Dickson (London) have been following regarding exon 23 of the mdx dystrophin gene. In particular, he pointed out that their group has been successful using systematic constructed hybridization antisense arrays to identify open regions of the pre-mRNA [8], [14]. This strategy will now be used to identify AON directed against the exon–intron boundaries of exons 51 and 53.
Steve Wilton (Perth) illustrated his strategy for the identification of successful AON. He also has experience with targeting exon–intron boundaries, both of mdx mouse and of the human gene [3], [4], [16], [17]. In the human, Steve Wilton has mostly concentrated on skipping located in the 5′ deletion hot spot. He follows a similar strategy to the one initially used by van Deutekom, with a pre-screening using bandshift assays, followed by verification in muscle cultures. In addition to the predictive approach, he also uses an empirical approach, where overlapping AON against the exon–intron boundaries of interest are used in cultured cells to verify their efficacy.
3. Different AON chemistry and carrier polymers
A number of presentations outlined the relative efficacy and stability of different modifications to the AON backbone (Ian Graham, Aartsma-Rus, Steve Wilton) and a recent review on this can be found in Jason et al. [18]. Phosphorothioate AON have been extensively used in man with a wealth of general data related to its safety in the human, although fully 2-O-methylated PS AONs have not been studied yet. Regarding other backbone modifications such as, for example, morpholino or PNA AON [12], [19] some data on safety in the human are available [18]. Furthermore, in addition to the efficiency the backbone should have a high specificity (this is not the case for the LNA AON). While many participants are clearly interested in pursuing this aspect of preclinical research, the planned trials will likely use the 2-O-methylated phosphorothioated AON, unless data on a clear advantage of different modifications, previously tested in the human, are generated in the interim.
Regarding the use of carrier polymers, Terry Partridge (London) presented the collaborative work performed with Qi-Long Lu (London UK/Charlotte, USA) on the improved efficacy of distribution of transfection, in vivo but not in vitro, using the pluronic polymer F127. The data presented showed that with the use of this polymer, the number of fibres transfected following a single or repeated injections in the tibialis anterior of mdx mouse improved significantly, with more than 70% of the fibres eventually producing different levels of dystrophin [10]. Similar efficiency was also achieved in older and more fibrotic muscle. In this experiment there was restoration of the dystrophin-associated glycoprotein complex and of the force generated by the treated mdx muscle. However, only partial protection against the damage following eccentric exercise was achieved [10].
Recent data from the same investigators using AON plus the same pluronic polymer showed that the intravenous administration of these reagents in mdx mice was followed by low but detectable levels of skipping in a number of muscles, although not in all [20].
4. Efficacy and toxicity of AON in human: experience from other therapeutic trials
Art Levin, from ISIS Pharmaceuticals (USA), presented the preclinical and clinical data on safety and efficacy of AON from a number of human clinical trials. He explained that AON have on the whole a good safety profile from data available on human trials. Toxicity to AON can, however, be observed, and this can be both hybridization dependent, and hybridization independent. The first are typically secondary to an exaggerated pharmacological effect of the administered AON (and in this context none is expected for dystrophin); there is also the possibility of hybridization to unidentified targets, although this is extremely unlikely.
The more unpredictable toxicity arises from a hybridization independent mechanism. This can be due to specific motifs that have (currently) undefined receptors; or to specific motifs that define a receptor interaction. The best known is the immune reaction against the CpG motif, which is activated via the Toll-like receptor 9.
Other parameters relevant for the AON toxicity relate to their bio-distribution. Kidney has the highest concentration following either subcutaneous (SC) or intravenous (IV) administration. Following the administration of very high doses, it is possible to observe histological changes in proximal tubules in animal models. These changes are reversible. However, at the far lower doses used in clinical trials this does not appear to be of practical clinical significance and detailed studies in more than 1000 patients have failed to show any abnormality in renal functional analysis.
Liver is the organ with the second highest concentration of AON. In rodents, administration of AON often results in hepatic changes, with mononuclear infiltrates, Kupfer cell infiltrates and often elevation of ALT and AST. In the human there is usually no observed toxic liver effect; however, at very high doses some transient change in transaminase levels can be observed.
Regarding other tissues, splenomegaly can be often observed in rodents; however, this side effect has not been noticed in humans or primates.
Regarding the sequence specific toxicity, this is highly dependent on sequence. AON, especially those with phosphorothioate modification, stimulate immune responses if CpG motifs are present. CpG-dependent immune stimulation is Th1-dependent. Palindromic sequences can also be immune-stimulating and should also be avoided. There can also be other not fully predictable effects on immune response dependent on sequence recognition by receptors of innate immunity. In rodents, this is often a prominent side effect, but much less so in humans and primates.
On the whole clinical manifestations of the administration of AON in human clinical trials have been limited to the local side effects following SC injection (on the whole i.v. administration seems to be much better tolerated) and generalized side effects such as fever and chills that similar to the response to interferon administration, respond well to paracetamol. More than 4000 patients with different disorders have been treated so far using systemic delivery of first generation AON (phosphorothioate backbone), and approximately 500 following local administration. The typical dosage used ranged from 0.5mg/kg every other day for 1month in Crohn's disease, to 200mg twice weekly for 3 months in rheumatoid arthritis, to higher dosages of 2mg/kg day in other protocols dealing with malignancies.
Fewer patients (∼300) have been treated so far using new generation AON (uniform phosphorothioated backbone with flanking 2′ methoxyethoxy wing) delivered systemically at doses comprised between 0.5 and 9mg/kg per week for up to 3 weeks. It was noted that the local (intramuscular, i.m.) administration of a small dose of AON as proposed in both DMD trials should be safe. However, most of the expertise resides with AON with already well known modifications. It looks, therefore, realistic for the scope of these phase I trials, to only consider the use of known and tested chemical AON modifications.
Gerard Platenburg, from Prosensa (The Netherlands), clarified the various necessary steps to move a project from the pre-clinical phase to the clinical arena. He also explained various regulatory aspects related to clinical trials and in particular to the manufacturing of clinical grade AON for clinical trials. In particular, it was stressed that the same batch used for the toxicity analysis will be used for the clinical trials.
Finbarr Cotter (London) presented his experience over 10 years in human clinical trials using AON. He first reiterated some of the aspects related to the bio-distribution of AON (they do not cross the blood brain barrier, nor target the testis). He then described the rationale of a number of cancer trials in which he participated. In a number of cancers the anti-apoptotic molecule Bcl2 is abnormally up-regulated; Bcl2 has, therefore, been the target of AON aimed at its down-regulation [21], [22], [23], [24], [25].
He described the typical stages necessary to take a molecule from phases I to III trials. In phase I, trials in cancer patients in whom high Bcl2 levels were known to occur, a dose escalation at 100% increments is performed until some toxicity using multiple parameters is identified. The dose is then decreased to the immediately lower dose, at which no significant toxicity was observed.
Among side effects frequently observed, skin reactions are a common problem following subcutaneous (s.c.) administration. On the whole IV is better tolerated. For doses of phosphorothioate AON higher than 5mg/kg per day, thrombocytopenia and hyperglycemia were transiently observed. Relatively common but less severe observed side effects of high dosage i.v. or s.c. administration were transient leucopenia, neutropenia and lymphopenia, fever, asthenia and hypotension. Some of these flu-like symptoms respond well to the administration of paracetamol.
5. Dystrophin gene restoration therapy following direct injection of plasmid DNA: experience from the French trial
Serge Braun (Transgene, France) presented in detail the protocol design and optimization of the first gene therapy trial in DMD, performed in France following the collaborative efforts of his group at Transgene, the clinical team of the Institute de Myologie, Hôpital Pitié-Salpêtrière, Paris (Prof. Micheal Fardeau; Dr Normal Romero), funded by AFM. The scope of this trial was to assess the safety of the administration of a full-length dystrophin containing plasmid into a single muscle of a group of patients with DMD (three) or BMD (six). The results of this first study have been recently published [26]. Serge Braun explained how the protocol was developed, how long it took from the original preparation of the trial to its completion (5 years). Nine patients were sequentially enrolled in the study. All received one or two injections of dystrophin plasmid DNA into the radialis muscle and had a muscle biopsy taken 3 weeks after the last injection. The expression of dystrophin was studied using antibodies directed against the patients deleted dystrophin region, and nested RT-PCR studies. The plasmid DNA was seen at the injection site biopsies in 9/9 patients. Full-length dystrophin expression was restored in 6/9 of patients, although the percentage of labeled fibres was very low. Reassuringly, no side effects were observed; in particular no cellular or humoral anti-dystrophin response was detected. Following these preliminary but encouraging results, the French investigators are currently considering regional intravenous or arterial delivery of the same construct. Experiments in this direction are currently being performed in animal models [27], [28]. The preliminary data indicate this to be well tolerated and quite effective in the animal model. The next step will be to perform a phases I–II trial in patients with DMD, with the aim of regional plasmid DNA delivery to the forearm muscles.
6. AON in DMD: selection of patients
The discussion focused again on AON and dystrophin. Francesco Muntoni presented the figures of patients available in the UK Consortium with deletions who would benefit from skipping exons 51 or 53. He mentioned that in the UK there is a well established network of 14 pediatric neuromuscular centres working together towards the definition of a common assessment protocol that could be used towards future therapeutic trials (The North Star Network, funded by the Muscular Dystrophy Campaign) and that patients followed in any of these centres could be recruited to this phase I trial. He also noted that the choice of the 3′ end of the distal hot spot region as more likely to result in functional protein than that of other regions of the gene. Indeed, a number of reports of asymptomatic cases with deletions of exons 48–51; 48–53 [29]; 48 [30] and 48–51 and 50–53 [31], [32] are on record.
Ieke Ginjaar (Leiden) presented the cases currently followed among the various centres in The Netherlands and in Belgium, who could be recruited as part of the trial. The preferred choice is for patients who could benefit from skipping of exon 51, as these represent the most abundant cohort of DMD patients. The second option would be exon 46 which as a single-mutation cohort would be the largest. They also discussed that unexpectedly some mutations that ought to occur and be represented in the Leiden DMD database (http://www.dmd.nl/) apparently do not occur or are significantly underrepresented, or are identified incidentally in patients ascertained because of elevated serum CK. These are the deletions of 45–51; 50–51; 47–51, 48–51, 51–52. This further reinforces the view that this appears to be a well-dispensable region of the dystrophin protein and, therefore, exon skipping to induce some of the above mentioned scenarios ought to be very effective.
Steve Wilton presented his data on targeting patients with deletions towards the 5′ end of the gene. Some of these patients can also have a remarkably mild phenotype and in particular he mentioned a patient from Newcastle with a deletion of exons 3–9 who plays competitive sport. Steve Wilton already has very effective oligonucleotides tested in human muscle cell lines to induce skipping of exons 4; 8; 9; 15; 16; 19 and 20; 31; 33 and 35.
7. Validation of AON effects in patients muscle cell cultures
Jenny Morgan (London) and Annemieke Aartsma-Rus (Leiden) discussed the available techniques to test AON in either primary muscle cell cultures from patients with DMD or using fibroblast cell lines transfected with MyoD [33], [34] or treated with Galectin 1 [35], in order to force myogenic differentiation. It was noted that it would be important to test cells from each individual patient prior to recruitment to the trial to verify the efficacy of AON in that particular patient. This is important not only to verify that the target sequences are effectively accessible to the AON, but also to assess the ability of that particular patient to produce dystrophin following the transfection with AON. This is particularly important in view of the rare patients who carry more than one dystrophin gene mutation. Although this is a very rare occurrence, the recruitment of such a patient in the trial should be avoided.
8. Animal models for future developments on AON delivery and optimization
Judith van Deutekom (Leiden) presented the generation of a transgenic mouse model with the entire human dystrophin (hDMD) gene integrated on mouse chromosome 3. The advantage of this animal model is that the sequence of the tested AON is human, so any positive data can be immediately transferred to the human scenario [36]. In addition, there are sequence differences between the two species facilitating testing of the sequence-specificity of the AON. AON were administered following direct intramuscular injection with and without polyethylenimine. This compound induced some muscle necrosis and regeneration, which in fact increase the efficiency of exon skipping. Interestingly, it was possible to observe an exclusive skipping induction of the human and not mouse gene, suggesting that the observed effect is highly sequence specific [36].
Dominic Wells (London), presented the preliminary work related to the generation of a transgenic approach (the SpliceOmouse) to enable rapid screening for systemic delivery. This is based on bioluminescence, and takes advantage of luciferase expression to track expression in a variety of tissues. The transgene will contain the exons 22–24 of the mouse dystrophin gene, carrying the mdx mutation, and GFP/luciferase at the 3′ end, thus only allowing protein expression if exon 23 has been skipped. The crossing of these transgenic with mdx mice will give out multiple readouts over a chosen time course. It is envisaged that these two complementary mouse models will greatly facilitate the search for effective AON and their systemic delivery.
9. AON in DMD: discussion regarding the two protocols
During this part of the discussion, the two consortia presented the fine details of the protocols to be used. Similarities and dissimilarities were discussed in detail; Volker Straub; Kate Bushby; Francesco Muntoni; Ieke Ginjaar; Anneke van de Kooi; Natalie Goemans and Jan Verschuuren contributed to this part. In particular, the choice of a younger age group in the Dutch/Belgian protocol; the choice of a different muscle in the two studies (relatively spared in the late stage of the disease; immediately accessibility under the fascia); the range of dystrophic changes expected in distal muscles (extensor carpi radialis; tibialis anterior; extensor digitorum brevis); the use of muscle magnetic resonance imaging to verify the degree of skeletal muscle involvement; the pros and cons of open versus needle/conchotome muscle biopsy and of a general vs local anesthetic; the techniques of local injection were all discussed in detail. In addition, the technical aspects related to the muscle biopsy processing (general histology; RNA studies; protein studies) was also discussed. The possibility of using non-invasive methods in the future, such as contrast enhanced muscle MRI to look at muscled damage was also discussed by Volker Straub [37].
In order to take forward this helpful discussion it was decided to continue to exchange the results of future preparatory work that will be acquired by the two groups (such as data on preservation of distal muscle obtained from relatively old DMD patients undergoing other surgical procedures, for example). In addition it was agreed that it would be beneficial to use an identical technique for RNA and protein quantitation. The Dutch/Belgian consortium has decided to make the technique used for RNA quantitation available; the UK consortium is setting up an ELISA protein quantitation assay and will share it with the members of the other consortium. It was also decided that cross validation on the results obtained in the two consortia would be helpful (i.e. exchange of muscle biopsies from treated and untreated patients).
10. AON in DMD: immunological aspects
Jan Verschuuren (Leiden) presented the available evidence related to the possibility of inducing an immune response to the ‘new’ antigens in patients with DMD receiving AON. Indeed, these patients will not have produced full-length dystrophin and a theoretical concern is, therefore, that the restoration of dystrophin production might be accompanied by an autoimmune response.
However, Jan Verschuuren explained that one fundamental step of the development of tolerance is through the expression of relevant proteins in the thymus. In this context the shorter dystrophin isoform Dp71 is known to be expressed in the thymus. As Dp71 protein expression is not affected by the mid-rod deletions (only deletions with a 3′ breakpoint downstream to exon 63 will also affect Dp71 expression), it is anticipated that this will be sufficient to allow these patients to be tolerant to all the dystrophin isoforms. This phenomenon is known as bystander suppression and further references can be found in a recent review [38].
Nic Wells (London) also reported that studies in mdx mice suggest that revertant fibres and the expression of various other isoforms in other tissues than muscle can induce tolerance to dystrophin gene transfer [39], [40]. The administration of AON should in any event be expected to be less immunogenic than replacing whole length protein in patients who previously had large deletions.
11. AON in DMD ethical aspects and parental perspective
Michel Briejer (Afforce Healthcare, The Netherlands), summarized a number of regulatory and ethical aspects relevant to the conduct of clinical trials in the pediatric age ranged, with a special focus to the current legislation in The Netherlands. In particular, he noted that the classical phases I–III stages in clinical trials are more likely to be replaced by a cycle of confirmatory and exploratory studies, especially as far as trials in the pediatric age group are concerned. Another point of discussion was the new Clinical Trial Directive which will only be implemented early in 2005, while the existing national legislation is active in this interim period. He also pointed out that the criteria for approval in minors (less than 18) of non-therapeutic trials can only be approved if there is a negligible risk and there is no other patient population in which the proposed study could be performed. Parents or guardians need to provide consent but individuals in the age range 12–18 years are expected to co-sign the consent
Elizabeth Vroom (United Parent Project for Muscular Dystrophy, UPPMD) concluded the session with a parental perspective on the trial. She highlighted that while there is a lot of expectation on the announced trials, there is also a lot of confusion and that the availability of good quality dissemination material will be indispensable. She explained that for many DMD patients the concept of being made ‘better like a patient with BMD’ is not very clear, mostly because these children are likely not to appreciate the entire spectrum of BMD, but will rather know the severe end of the spectrum, edging towards DMD, as these are the patients they are likely to have met in clinics. She concluded by saying that while this is a very exciting time for the DMD community, it is very important not to be carried away by the enthusiasm and plan carefully each individual step so that any possible side effect or damage to children taking part in these studies is avoided. She finally explained that one genuine concern that many parents have is that there is effective and open collaboration between different scientists working in the field, so that scientific advances can be made more rapidly.
12. Conclusions
The members of the two consortia and the other participants to the workshop have agreed to create an International Consortium of AON in Neuromuscular Disorders. It was agreed that the International Consortium would share dissemination material for patients potentially interested in this experimental approach, and that this material will be made available in the ENMC web pages (http://www.enmc.org/workshops/reports.cfm?p=171). The agreed combined dissemination material relative to the AON in DMD trials is already available at ENMC, and this website will be kept up-to dated with a bulletin summarizing the progress in the two Consortia involved in the DMD-AON trials. The International ENMC Consortium is intended to function as an inclusive catalyst for future collaborative research and hence it is likely that in the future will expand to include other interested groups.
The two consortia also decided the possibility of cross-validation of the results obtained in the two studies regarding protein expression and RNA quantitation as this will help the parallel development of the two studies.
Also in the field of the mouse model studies collaborative further exploration will be explored, e.g. by generating SpliceOmouse models with larger dystrophin gene segments, based on the hDMD mouse.
As the remits of the International Consortium on AON is that of pursuing the research and development in this field, it is anticipated that further meetings of the consortium will be held in the future, both to discuss the results of the tolerability and success of the local AON administration, and the progress in the systemic delivery systems that could be used in future trials.
Acknowledgements
This Workshop was made possible thanks to the financial support of the European Neuromuscular Centre (ENMC) and ENMC main sponsors:
and ENMC associate member:
as well as:
Workshop participants
The Netherlands/Belgium Consortium
United Kingdom Consortium
Other participants
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PII: S0960-8966(05)00065-9
doi:10.1016/j.nmd.2005.02.007
© 2005 Elsevier B.V. All rights reserved.
