(From the March 2015 Issue of PediatricsOnline)
Exon skipping holds promise for converting Duchenne muscular dystrophy (DMD) to milder Becker muscular dystrophy (BMD). A new study led by researchers at Nationwide Children's Hospital suggests that precisely where doctors skip and splice gene segments in this kind of gene therapy could result in enormous clinical differences. They believe the work, published online in the journal Annals of Neurology in January 2015, can help guide therapy research.
Both forms of muscular dystrophy are the result of mutations in the gene that codes dystrophin, a protein essential to protecting muscle fibers. Patients who lack dystrophin have DMD and those who produce some usable form of the protein have BMD.
In healthy individuals, all 79 exons of the dystrophin gene complement their neighbors. They have end structures that link up like puzzle pieces, and when spliced together, they serve as the template for translation of the protein.
A deletion of one or more exons often results in these pieces no longer fitting together, in which case protein is not made. These are called out-of-frame deletions and result in the severe form of muscular dystrophy, DMD.
In contrast, sometimes the deletion still allows the remaining pieces to fit together. This is termed an in-frame deletion, in which the shorter chain of exons can still be translated into a partially functional protein. This results in the milder form of the disease, BMD.
One promising approach to treatment of DMD that has already reached clinical trials is directed at skipping more exons in patients with deletions in order to restore an assembled gene that can be translated.
“We are interested in whether the result of this approach is always beneficial,” says Kevin Flanigan, MD, a neurologist and principal investigator in the Center for Gene Therapy in The Research Institute at Nationwide Children’s. “We know that not all dystrophin proteins that result from this approach will be the same, because you’ll be missing different domains, which perhaps result in less stability or other differences.”
Dr. Flanigan, who is also an attending neurologist in Neuromuscular Disorders at Nationwide Children's and a faculty member of The Ohio State University College of Medicine, tested the idea that these varying proteins result in different degrees of severity of muscular dystrophy.
The researchers looked closely at records of patients who naturally have the equivalent of the in-frame deletions that would be expected to result from therapies intended to skip exon 45. The records are part of the United Dystrophinopathy Project database, which was created to help determine how different mutations in the dystrophin gene affect the clinical symptoms of DMD and BMD.
The researchers found that 97 percent of patients whose deletions ran from exon 45 to exons 47 or 48 had the Becker phenotype. Those whose deletion ran from exon 45 to 48 were walking into their 40s and those whose deletion stopped at 47 were walking well into their 30s.
Dr. Flanigan and his fellow researchers expected that patients whose deletion ran only from exon 45 to 46 would have the Becker phenotype. Surprisingly, all had the Duchenne phenotype. These patients were wheelchair-bound by age 12.
The scientists and doctors failed to find another possible deletion, running from exon 44 to 45, among the more than 900 records in the dystrophinopathy database.
The researchers suggest that this deletion hasn't been reported to the dystrophinopathy project because lack of the functioning pair produces no symptoms or symptoms so negligible the patients or their parents don't seek testing. Individuals with severe or deadly mutations would be better documented, they reason.
These results may have particular importance for patients with DMD due to a deletion of exon 45, which is the most common single exon deletion.
“Our conclusion is that skipping exons 45 and 46 is not likely to produce significant improvement,” Dr. Flanigan says. Instead, skipping exon 44 when 45 is deficient may be the best approach. “We predict that skipping 44 and 45 will likely result in much milder symptoms, but subsequent studies will have to prove it.”
Findlay AR, Wein N, Kaminoh Y, Taylor LE, Dunn DM, Mendell JR, King WM, Pestronk A, Florence JM, Mathews KD, Finkel RS, Swoboda KJ, Howard MT, Day JW, McDonald C, Nicolas A, Le Rumeur E, Weiss RB, Flanigan KM; for the United Dystrophinopathy Project. Clinical phenotypes as predictors of the outcome of skipping around DMD exon 45. Annals of Neurology. [Epub 2015, Jan. 21]