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Systemic Administration of Antisense Therapy for Duchenne Dystrophy Found Safe, Possibly Effective


doi: 10.1097/01.NT.0000384109.18580.d7
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In two different studies, investigators injected an antisense molecule intravenous into boys with Duchenne muscular dystrophy, and found the approach was largely well tolerated.

TORONTO—Systemic delivery of antisense molecules is safe and increases dystrophin expression in boys with Duchenne muscular dystrophy (DMD), according to two studies from rival companies presented here at the AAN annual meeting in April.

Dose-dependent increase in protein production was confirmed by muscle biopsy in both studies, and preliminary efficacy results suggest treatment may have improved the walking speed in some patients.

In DMD, about 85 percent of patients have a deletion that creates an incorrect reading frame in the RNA and truncates the protein. The antisense molecule binds to pre-messenger RNA during RNA processing, causing the targeted exon to be skipped in the creation of the final messenger RNA. Removal of the chosen exon corrects the mutation not by removing the mutation, but by compensating for it, restoring the reading frame and allowing the shorter-length protein to be synthesized. Because most mutations occur in the repetitive middle region of dystrophin, and because dystrophin works adequately even without one or more of its repeated units, the resulting protein is fully functional. This shortened protein due to exon skipping is seen naturally with Becker muscular dystrophy where the genetic deletion is “in-frame.”



Both studies targeted the same exon of the dystrophin gene, exon 51, since skipping this one exon is predicted to be therapeutic in 13 percent of all boys with DMD. Both groups used an antisense molecule whose base sequence was complementary to the exon, and each modified the sugar-phosphate backbone of the molecule in order to prevent premature degradation of the antisense molecule. The exact nature of the modifications differed between the two groups.

Previous studies had shown that intramuscular injection generates dystrophin in the injected muscle. “But local injections to single muscles is not feasible in Duchenne muscular dystrophy,” because of the widespread nature of the muscle deficit, according to Francesco Muntoni, MD, professor of pediatric neurology at the University of College London Institute of Child Health and deputy director of the Medical Research Council Center for Neuromuscular Disease in London, and lead researcher on the study sponsored by AVI Biopharma.

To explore the potential of systemic delivery, Dr. Muntoni injected the same antisense molecule intravenously into 19 patients, ages 5 to 15 years, all of whom were ambulatory. The patients were divided into six cohorts, with the dose escalated from one cohort to the next. For safety purposes, each group began after all the patients in the previous cohort had received at least three doses without a problem. Each patient received up to 12 weekly intravenous infusions with a muscle biopsy before the study or during screen, and 17 had a biopsy two weeks after the last dose.

“The study was mainly a safety study. The treatment was remarkably benign...and very well tolerated,” Dr. Muntoni said. “We were not able to detect any safety signal in any patient.”

There was one case of a child with a mild pre-existing Duchenne related cardiomyopathy whose cardiac function worsened over the first few weeks, and the patient was withdrawn from treatment, “but we think this was unrelated to the medication, and he has remained stable since starting cardioprotective medication.”

Treatment did not lead to an increase of dystrophin in the two lowest-dose groups, but did at the higher doses. In the fourth group, the highest analyzed at the time of the presentation, 21 percent of muscle fibers were dystrophin-positive.

“I think it is probably safe to expect that the level of dystrophin will be higher [in cohorts 5 and 6] than what we have seen in cohorts 3 and 4, and I think it is again probably safe to conclude that we are at the level of dystrophin expression that could start to give some clinical benefit, some muscle protection to these patients, and I think that is encouraging,” Dr. Muntoni said. The study was not powered for clinical efficacy, and results of strength and endurance testing were not reported.

The second trial, sponsored by the biotechnology firm Prosensa, of Leiden, the Netherlands, enrolled 12 patients to receive weekly subcutaneous injections for five weeks, again separated into dose-escalation cohorts, with muscle biopsies at the beginning and end of treatment.

The results, presented by Nathalie Goemans, MD, a pediatric neurologist at the University of Leuven, Belgium, indicated that the drug was well tolerated at the highest dose, and that dystrophin expression increased in a dose-dependent manner. After the initial phase, which was double-blinded, all patients enrolled in an additional 12 weeks of treatment at the highest dose. At the end of this period, there was a variable but significant improvement in the six-minute walk distance, the first demonstration of clinical efficacy of antisense therapy in DMD. However, she cautioned, the small numbers of patients, the wide variability in baseline disability, and the open-label nature of the treatment means the results must be confirmed in a larger, controlled trial. Meanwhile, the open trial is continuing out to six months.

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“These results are very encouraging,” according to Charles Thornton, MD, professor of neurology at the University of Rochester Medical School, who was not involved with the study. “Systemic administration appears to be very well tolerated, and there were no major safety issues encountered. It's unclear what the maximally tolerated dose will be with either oligo chemistry,” he said, “and it is uncertain which of the two chemistries will work better.”

“A big question at this point is how much expression is needed and for how long, in order to see an improvement in muscle or a protection against further damage,” he continued. Although fibrosis may limit the extent of recovery, “it is still quite possible at least in theory that recovery may occur. But even failing that, slowing disease progression to any extent would be counted a major achievement for this disease.”



“In terms of treatments that are potentially close at hand, this [exon-skipping approach] would be the most exciting treatment for Duchenne muscular dystrophy that is currently in clinical trials,” Dr. Thornton said. Treatment of other mutations would likely follow if these preliminary results are supplemented by a conclusive demonstration of efficacy, he added.

One cautionary note, he noted, is that each antisense treatment must be tailored to the mutation of relatively small numbers of patients, one of the down-sides of personalized medicine compared to a one-size-fits-all treatment.

According to Judith van Deutekom, PhD, vice president for Drug Discovery at Prosensa, the company is also developing antisense molecules to target mutations not treated by exon 51 skipping. Their next goal, she said, is to go after exon 45, potentially therapeutic for 11 percent of patients, and exon 44, for 8 percent. Dr. Muntoni noted that targeting nine exons in all would cover approximately 83 percent of all patients.

Whether the US and European regulatory agencies will require testing each new antisense molecule as extensively as has been done for exon 51 “is a very relevant question,” Dr. van Deutekom said.

“By using the same class of chemistry, we hope it is not considered a different medicine, and we hope we can shortcut the toxicity program. It is probably going to be a long discussion” with the US FDA and the European Medicines Agency. “If they want to have companies develop treatments for patients with the more rare mutations, there is only one way,” namely, making the approval process shorter as more experience is gained with similar drugs.

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An alternative approach to overcoming the DMD reading frame error is to persuade the ribosome to read through nonsense mutations. This approach, using the oral drug ataluren, a benzoic acid derivative, has had mixed success in improving clinical performance in DMD.

The drug, formerly known as PTC124, increases dystrophin levels in both human and mouse muscle fibers, and improves muscle performance in mice. But according to results presented at the AAN annual meeting, and amplified in statements from its manufacturer, the drug improved walking time compared to placebo only in patients receiving low-dose, not high-dose, ataluren.

“But there is nothing about the ataluren trial results that would change our attitudes for the prospects of success for oligonucleotide-based antisense therapy,” said Charles Thornton, MD, professor of neurology at the University of Rochester Medical School, who was not involved with the study.

©2010 American Academy of Neurology