Duchenne muscular dystrophy Summary of important research results betw
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《中华医药杂志》英文版
Duchenne muscular dystrophy Summary of important research results between July and December 2006
Most of the new research results summarized here are based on the presentations at the conference of the British Parent Project in London on 21 October 2006，the remainder comes from publications and from many personal e-mail messages. Some of this new information is an update of my report about the annual meeting of the American Parent Project Muscular Dystrophy which took place in Cincinnati/Ohio in the USA from 13 to 16 July 2006. This report and all earlier ones can be seen and downloaded from my internet pages www.duchenne-research.com in English, German, and Spanish. The names of the researchers mentioned here are used without their titles, and, as is usual within the Duchenne community, often only their first names are used. Almost all are professors and all have either a PhD or an MD or both.
Clinical exon skipping trial in the Netherlands
Judith van Deutekom said in London, that the four participating Duchenne boys have received in their shin muscle, tibialis anterior, their single injection of the 2O-methyl AON against exon 51, that no side effects appeared in all four of them, and that biopsies were performed for two boys in which analyses were done for newly formed shortened dystrophin. Judith said that these preliminary results were “promising”, but that she was not allowed to say more.
At the end of her presentation, Judith said that we should be cautious because “we are not there yet”.Even if exon skipping works in one boy, this does not mean that it will be the same in others. Only a very small volume of one muscle was treated, and such a local treatment is not that what is needed. A systemic treatment is necessary, which would reach all muscles, and this is possible only by injections of the AONs into the blood circulation. Before this will be attempted next year-then for skipping of exon 46-the correct AON and the best dosage must be selected first. “But we will get there”, said Judith.
Clinical exon skipping trial in England
Francesco Muntoni directs the trial in England, prepared by the MDEX consortium, in which a morpholino AON for skipping exon 51 will be used with local injections into the small foot muscle extensor digitorum brevis. Francesco answered my questions on 18 December, that two of three permissions are now there, and as soon as the third is also obtained, the first boy will be recruited. This will possibly be in the first week of March 2007.
No preliminary results will be released. But after all data are analyzed, checked, and approved, they will be published. If the first trial is successful, the planned second study will start in the second half of 2007. Its results will be available in 2008. If everything progresses positively, the commercial manufacture of the AONs in large quantities and long-term clinical studies will be started.
Morpholinos do enter the heart
Up to now, the morpholino AONs could not be persuaded to migrate into the heart muscles of the mdx mice.
Im Talgrund 2, D-79874 Breitnau, Germany
Correspondence to Guenter Scheuerbrandt,Im Talgrund 2, D-79874 Breitnau, Germany
E-mail:gscheuerbrandt@t-online.de
This was a great disadvantage of the morpholinos to be used by the British, compared to the 2O-methyls, with which the Dutch are working and which cause exon skipping also in the heart of the mice. In London, Dominic Wells reported that the morpholinos do go into the hearts of the mice if one injects the contrasting agent Optison-micro albumin bubbles-together with the morpholinos and directs at the same time ultrasound from a diagnostic ultrasound apparatus directly at the heart. The ultrasound opens pores in the muscle cell membranes, the morpholinos can get through, and when the ultrasound is stopped, the pores close again. The exon skipping in the skeletal muscles is not disturbed by this procedure. It will take several years, until this simple technique can be clinically tested in children.
Exon skipping with gene transfer in France
I have already reported about the experiments of the research team around Luis García in Paris who transferred genes for the AONs with AAV vectors into the nuclei of muscle cells (www.duchenne-research.com). It was known that, with this technique, it was possible to skip for at least one year the exon 23 of mdx mice which contains a premature stop sign. Luis reported in London that this now works also in dystrophic dogs which are really sick and have about as many muscles as a small child. But they have a complex mutation which can only be repaired by a multiple skipping of exons 6, 7, and 8. And this worked anyway! However, the French researchers did not have enough of the viruses, loaded with the U7-RNA and the AON sequences, to treat the entire dog. Therefore, they blocked the blood circulation in one leg and injected into its veins within 15 minutes practically all their 100 trillion (1014!) prepared viruses. Some oedemas developed, but that was not bad, because then the viruses migrated without problems to where they were supposed to go. Quite a lot of new and shortened dystrophin appeared, which, however was not distributed evenly in the leg muscles, but which was there in amounts similar to what Becker patients have. After six months, it was still there! Now, a phase-I clinical trial with Duchenne boys is being prepared in which exon 51 will be skipped. Such a treatment with virus vectors will probably need immune suppression.
Luis and his colleagues are trying also to combine this treatment with a similar gene transfer in which the gene for myostatin will be blocked. This was already tried in mice, however with little success, because they regenerate their muscles too quickly. Now, it is being tried in dogs. The results will be reported next year.
Clinical gene-transfer trial in the USA
Scott McPhee of the company Asklepios in North Carolina, which together with Jerry Mendell in Columbus/Ohio has started a mini-dystrophin gene transfer study with AAV vectors, told me on 21 December that four of the six participating boys have now received their Biostrophin injections into their biceps muscles. The fourth boy belonged already to the second group of three boys who receive a high dose of vectors. No serious adverse side effects have been detected so far. By mid 2007, preliminary results will be available which will be published without delay, followed by the detailed final data not much later.
Ignoring premature stop signs with PTC124
Richard Finkel of the company. PTC in New Jersey presented in London details of the clinical trials with the compound PTC124. This novel drug, whose structure is kept secret, can be administered as tablets. In the now completed phase-Ⅱ clinical trial with 26 Duchenne boys, who have premature stop codons, and who received dosages of 18 mg/kg/day and 40 mg/kg/day of the drug, in some children who received the higher dose for 4 weeks, preliminary analyses detected the expected full-length dystrophin. In this study, as before in the phase-Ⅰ trial, no side effects appeared. But the level of the new dystrophin was still too low to produce a sufficient and reliable therapeutic effect. Therefore, another phase-Ⅱ trial will be started with 12 Duchenne boys, who will receive 80 mg/kg/day PTC124 to obtain a concentration of at least 10 microgram/ml in blood serum. The final data will be published in 2007. Then long-term studies lasting three to six months will be performed.
It is important to understand that, if all clinical trials proceed successfully, this kind of treatment will be a real cure, because the new dystrophin will have almost the normal length, it would not be a Becker dystrophin as is expected with the exon skipping technique. On the other hand, only those 10% to 15% of Duchenne boys can benefit from a PTC124 treatment who have a point mutation which led to a premature stop codon. Therefore, it is necessary to know the exact mutation of all Duchenne boys.
Mesoangioblasts “make lame dogs running again”
(such words were used for the headlines in some newspapers).
That was the effect of stem cells from blood vessel walls which really brought dramatic functional improvements in dystrophic dogs, as Giulio Cossu and co-workers in Milan reported in the Nature issue of 30 November. This publication and a commentary by Jeffrey Chamberlain can be obtained by e-mail from me. Giulio wrote to me on 9 December that he will send me, probably in February, information about his work to isolate these stem cells from human embryonic blood vessel tissue. In addition, he has found another efficacious drug, which he cannot discuss at the moment.
Upregulation of utrophin
Kay Davies of the University Oxford and Jon Tinsley of the company Vastox in Oxford reported in London, that, by automatically testing about 13,000 compounds, they have found 100 which can upregulate utrophin in cell cultures. Some of them could increase the utrophin level more than fourfold. They will be optimized further. The treatment of mdx mice with the compound VOX 185 could upregulate utrophin almost exclusively in muscles by acting on its A-promoter, and thus obtain a significant improvement of their function.
Other experiments with mice had shown already that a two to threefold increased utrophin level had caused significantly better therapeutic effects, and one hopes now to reach a five to sevenfold upregulation of utrophin with an optimized VOX 185 compound. Mid 2007, an effective potential drug should be ready which could be given as tablets for reaching all muscles. Then, all necessary clinical trials will be performed in the following years
Inhibition of myostatin
The company Wyeth Pharmaceuticals in the USA has now tested in a phase-Ⅰ/Ⅱ clinical trial the antibody MYO-39 in 108 adult muscle disease patients, among them some Becker patients. The trial is now complete, the results, which seem to be positive, will be published in the spring of 2007.
Myoblast transfer works again
At the end of September, Jacques Tremblay of Québec City in Canada sent me detailed information about his new experiments with the myoblast transfer technique, which should be better called “transfer of myogenic cells” to distinguish it from the machinations of Peter Law. In a clinical trial with 9 Duchenne patients, he could show that in 8 patients up to 26% muscle fibers with new normal dystrophin were created after the injection of normal myogenic cells from a relative. The cells were injected at a distance of only 1 to 2 mm from each other into a small area of the shin muscle.
At the moment, a second study is underway with injections into a muscle in which one would be able to demonstrate easily a functional improvement. This cell transfer would have some advantages, among them: The new dystrophin would have the normal length and be under the control of its normal control sequences, the positive effect would be long-term, immune problems could be avoided, the technique could be combined with a myostatin inhibition, and, most importantly, it could also help older Duchenne patients.
(Editor HOU)(Guenter Scheuerbrandt)
Most of the new research results summarized here are based on the presentations at the conference of the British Parent Project in London on 21 October 2006，the remainder comes from publications and from many personal e-mail messages. Some of this new information is an update of my report about the annual meeting of the American Parent Project Muscular Dystrophy which took place in Cincinnati/Ohio in the USA from 13 to 16 July 2006. This report and all earlier ones can be seen and downloaded from my internet pages www.duchenne-research.com in English, German, and Spanish. The names of the researchers mentioned here are used without their titles, and, as is usual within the Duchenne community, often only their first names are used. Almost all are professors and all have either a PhD or an MD or both.
Clinical exon skipping trial in the Netherlands
Judith van Deutekom said in London, that the four participating Duchenne boys have received in their shin muscle, tibialis anterior, their single injection of the 2O-methyl AON against exon 51, that no side effects appeared in all four of them, and that biopsies were performed for two boys in which analyses were done for newly formed shortened dystrophin. Judith said that these preliminary results were “promising”, but that she was not allowed to say more.
At the end of her presentation, Judith said that we should be cautious because “we are not there yet”.Even if exon skipping works in one boy, this does not mean that it will be the same in others. Only a very small volume of one muscle was treated, and such a local treatment is not that what is needed. A systemic treatment is necessary, which would reach all muscles, and this is possible only by injections of the AONs into the blood circulation. Before this will be attempted next year-then for skipping of exon 46-the correct AON and the best dosage must be selected first. “But we will get there”, said Judith.
Clinical exon skipping trial in England
Francesco Muntoni directs the trial in England, prepared by the MDEX consortium, in which a morpholino AON for skipping exon 51 will be used with local injections into the small foot muscle extensor digitorum brevis. Francesco answered my questions on 18 December, that two of three permissions are now there, and as soon as the third is also obtained, the first boy will be recruited. This will possibly be in the first week of March 2007.
No preliminary results will be released. But after all data are analyzed, checked, and approved, they will be published. If the first trial is successful, the planned second study will start in the second half of 2007. Its results will be available in 2008. If everything progresses positively, the commercial manufacture of the AONs in large quantities and long-term clinical studies will be started.
Morpholinos do enter the heart
Up to now, the morpholino AONs could not be persuaded to migrate into the heart muscles of the mdx mice.
Im Talgrund 2, D-79874 Breitnau, Germany
Correspondence to Guenter Scheuerbrandt,Im Talgrund 2, D-79874 Breitnau, Germany
E-mail:gscheuerbrandt@t-online.de
This was a great disadvantage of the morpholinos to be used by the British, compared to the 2O-methyls, with which the Dutch are working and which cause exon skipping also in the heart of the mice. In London, Dominic Wells reported that the morpholinos do go into the hearts of the mice if one injects the contrasting agent Optison-micro albumin bubbles-together with the morpholinos and directs at the same time ultrasound from a diagnostic ultrasound apparatus directly at the heart. The ultrasound opens pores in the muscle cell membranes, the morpholinos can get through, and when the ultrasound is stopped, the pores close again. The exon skipping in the skeletal muscles is not disturbed by this procedure. It will take several years, until this simple technique can be clinically tested in children.
Exon skipping with gene transfer in France
I have already reported about the experiments of the research team around Luis García in Paris who transferred genes for the AONs with AAV vectors into the nuclei of muscle cells (www.duchenne-research.com). It was known that, with this technique, it was possible to skip for at least one year the exon 23 of mdx mice which contains a premature stop sign. Luis reported in London that this now works also in dystrophic dogs which are really sick and have about as many muscles as a small child. But they have a complex mutation which can only be repaired by a multiple skipping of exons 6, 7, and 8. And this worked anyway! However, the French researchers did not have enough of the viruses, loaded with the U7-RNA and the AON sequences, to treat the entire dog. Therefore, they blocked the blood circulation in one leg and injected into its veins within 15 minutes practically all their 100 trillion (1014!) prepared viruses. Some oedemas developed, but that was not bad, because then the viruses migrated without problems to where they were supposed to go. Quite a lot of new and shortened dystrophin appeared, which, however was not distributed evenly in the leg muscles, but which was there in amounts similar to what Becker patients have. After six months, it was still there! Now, a phase-I clinical trial with Duchenne boys is being prepared in which exon 51 will be skipped. Such a treatment with virus vectors will probably need immune suppression.
Luis and his colleagues are trying also to combine this treatment with a similar gene transfer in which the gene for myostatin will be blocked. This was already tried in mice, however with little success, because they regenerate their muscles too quickly. Now, it is being tried in dogs. The results will be reported next year.
Clinical gene-transfer trial in the USA
Scott McPhee of the company Asklepios in North Carolina, which together with Jerry Mendell in Columbus/Ohio has started a mini-dystrophin gene transfer study with AAV vectors, told me on 21 December that four of the six participating boys have now received their Biostrophin injections into their biceps muscles. The fourth boy belonged already to the second group of three boys who receive a high dose of vectors. No serious adverse side effects have been detected so far. By mid 2007, preliminary results will be available which will be published without delay, followed by the detailed final data not much later.
Ignoring premature stop signs with PTC124
Richard Finkel of the company. PTC in New Jersey presented in London details of the clinical trials with the compound PTC124. This novel drug, whose structure is kept secret, can be administered as tablets. In the now completed phase-Ⅱ clinical trial with 26 Duchenne boys, who have premature stop codons, and who received dosages of 18 mg/kg/day and 40 mg/kg/day of the drug, in some children who received the higher dose for 4 weeks, preliminary analyses detected the expected full-length dystrophin. In this study, as before in the phase-Ⅰ trial, no side effects appeared. But the level of the new dystrophin was still too low to produce a sufficient and reliable therapeutic effect. Therefore, another phase-Ⅱ trial will be started with 12 Duchenne boys, who will receive 80 mg/kg/day PTC124 to obtain a concentration of at least 10 microgram/ml in blood serum. The final data will be published in 2007. Then long-term studies lasting three to six months will be performed.
It is important to understand that, if all clinical trials proceed successfully, this kind of treatment will be a real cure, because the new dystrophin will have almost the normal length, it would not be a Becker dystrophin as is expected with the exon skipping technique. On the other hand, only those 10% to 15% of Duchenne boys can benefit from a PTC124 treatment who have a point mutation which led to a premature stop codon. Therefore, it is necessary to know the exact mutation of all Duchenne boys.
Mesoangioblasts “make lame dogs running again”
(such words were used for the headlines in some newspapers).
That was the effect of stem cells from blood vessel walls which really brought dramatic functional improvements in dystrophic dogs, as Giulio Cossu and co-workers in Milan reported in the Nature issue of 30 November. This publication and a commentary by Jeffrey Chamberlain can be obtained by e-mail from me. Giulio wrote to me on 9 December that he will send me, probably in February, information about his work to isolate these stem cells from human embryonic blood vessel tissue. In addition, he has found another efficacious drug, which he cannot discuss at the moment.
Upregulation of utrophin
Kay Davies of the University Oxford and Jon Tinsley of the company Vastox in Oxford reported in London, that, by automatically testing about 13,000 compounds, they have found 100 which can upregulate utrophin in cell cultures. Some of them could increase the utrophin level more than fourfold. They will be optimized further. The treatment of mdx mice with the compound VOX 185 could upregulate utrophin almost exclusively in muscles by acting on its A-promoter, and thus obtain a significant improvement of their function.
Other experiments with mice had shown already that a two to threefold increased utrophin level had caused significantly better therapeutic effects, and one hopes now to reach a five to sevenfold upregulation of utrophin with an optimized VOX 185 compound. Mid 2007, an effective potential drug should be ready which could be given as tablets for reaching all muscles. Then, all necessary clinical trials will be performed in the following years
Inhibition of myostatin
The company Wyeth Pharmaceuticals in the USA has now tested in a phase-Ⅰ/Ⅱ clinical trial the antibody MYO-39 in 108 adult muscle disease patients, among them some Becker patients. The trial is now complete, the results, which seem to be positive, will be published in the spring of 2007.
Myoblast transfer works again
At the end of September, Jacques Tremblay of Québec City in Canada sent me detailed information about his new experiments with the myoblast transfer technique, which should be better called “transfer of myogenic cells” to distinguish it from the machinations of Peter Law. In a clinical trial with 9 Duchenne patients, he could show that in 8 patients up to 26% muscle fibers with new normal dystrophin were created after the injection of normal myogenic cells from a relative. The cells were injected at a distance of only 1 to 2 mm from each other into a small area of the shin muscle.
At the moment, a second study is underway with injections into a muscle in which one would be able to demonstrate easily a functional improvement. This cell transfer would have some advantages, among them: The new dystrophin would have the normal length and be under the control of its normal control sequences, the positive effect would be long-term, immune problems could be avoided, the technique could be combined with a myostatin inhibition, and, most importantly, it could also help older Duchenne patients.
(Editor HOU)(Guenter Scheuerbrandt)