Genetic editing shows promise in Duchenne muscular dystrophy

Using a novel genetic ‘editing’ technique, Duke University biomedical engineers have been able to repair a defect responsible for one of the most common inherited disorders, Duchenne muscular dystrophy, in cell samples from Duchenne patients.

Instead of the common gene therapy approach of adding new genetic material to “override” the faulty gene, the Duke scientists have developed a way to change the existing mutated gene responsible for the disorder into a normally functioning gene. The Duke researchers believe their approach could be safer and more stable than current methods of gene therapy.

The researchers are now conducting further tests of this new approach in animal models of the disease.

Duchenne muscular dystrophy is a genetic disease affecting one in 3,600 newborn males. The genetic mutation is found on the X chromosome, of which males have only one copy. (Females, with two X chromosomes, presumably have at least one good copy of the gene.)

Patients with Duchenne muscular dystrophy cannot produce the protein known as dystrophin, which is essential in maintaining the structural integrity of muscle fibers. Over time, patients with the disorder suffer gradual muscle deterioration, which leads to paralysis and eventual death, usually by age 25.

“Conventional genetic approaches to treating the disease involve adding normal genes to compensate for the mutated genes,” said Charles Gersbach, assistant professor of biomedical engineering at Duke’s Pratt School of Engineering and Department of Orthopaedic Surgery and member of Duke’s Institute for Genome Sciences and Policy. “However, this can cause other unforeseen problems, or the beneficial effect does not always last very long.

Genetic editing shows promise in Duchenne muscular dystrophy “Our approach actually repairs the faulty gene, which is a lot simpler,” said David Ousterout, the Duke biomedical engineering graduate student in the Gersbach lab who led the work. “It finds the faulty gene, and fixes it so it can start producing a functional protein again.”

Duchenne muscular dystrophy, which generally affects males, is caused by a mutation in a gene called dystrophin. DMD causes generalized weakness and muscle wasting first affecting the muscles of the hips, pelvic area, thighs and shoulders. Calves often are enlarged. DMD eventually affects all voluntary muscles, and the heart and breathing muscles. A less severe variant is Becker muscular dystrophy (BMD).

The results of the Duke study were published online in Molecular Therapy, the journal of the American Society for Gene and Cell Therapy. The project was supported by the Hartwell Foundation, the March of Dimes Foundation and the National Institutes of Health.

The Duke experiments, which were carried out in cell samples from Duchenne muscular dystrophy patients, were made possible by using a new technology for building synthetic proteins known as transcription activator-like effector nucleases (TALENs), which are artificial enzymes that can be engineered to bind to and modify almost any gene sequence.

Duchenne muscular dystrophy is the most severe childhood form of muscular dystrophy caused by mutations in the gene responsible for dystrophin production. There is no cure, and treatment is limited to glucocorticoids that prolong ambulation and drugs to treat the cardiomyopathy. Multiple treatment strategies are under investigation and have shown promise for Duchenne muscular dystrophy. Use of molecular-based therapies that replace or correct the missing or nonfunctional dystrophin protein has gained momentum. These strategies include gene replacement with adeno-associated virus, exon skipping with antisense oligonucleotides, and mutation suppression with compounds that “read through” stop codon mutations. Other strategies include cell therapy and surrogate gene products to compensate for the loss of dystrophin. All of these approaches are discussed in this review, with particular emphasis on the most recent advances made in each therapeutic discipline. The advantages of each approach and challenges in translation are outlined in detail. Individually or in combination, all of these therapeutic strategies hold great promise for treatment of this devastating childhood disease.

These TALENs bind to the defective gene, and can correct the mutation to create a normally functioning gene.

Duchenne Muscular Dystrophy (DMD)

Mutations in a gene called dystrophin are responsible for the most common form of muscular dystrophy—Duchenne muscular dystrophy (Briguet 2008; Wang 2009; Muir 2009; Pilgram 2010). The dystrophin protein is responsible for maintaining muscle strength, so when the dystrophin gene is mutated in a way that prevents the dystrophin protein from being produced or functioning normally, muscles become weak (CDC 2012).

DMD occurs more frequently in young males, and accounts for approximately half of all muscular dystrophies (CDC 2012; Mayo Clinic 2012). In DMD, muscle weakness typically starts in the pelvis and legs, but can also occur in the arms, neck, and other regions of the body (PubMed Health 2013), while muscles of the face are normally spared. Calf muscles are also enlarged due to an accumulation of fatty tissue (NINDS 2011). People with DMD usually lose their ability to walk sometime between 7 and 13 years of age (CDC 2012), and they often die of respiratory failure before reaching age 40 as a result of damage to muscles that control breathing. About two-thirds of DMD cases run in families and one-third are caused by spontaneous mutations (NINDS 2011).

Females who carry the mutation usually do not display any symptoms, but about 8–10% of them will show some manifestation of the disease. When these symptoms do occur, they are typically more minor than the severe muscle weakness seen in males (Bushby 2005).

Signs and symptoms usually become evident when the child starts walking and may include (NINDS 2011; CDC 2012; Mayo Clinic 2012):

  Clumsiness and falling more often than other children of the same age
  A delay in walking
  Difficulty getting up from the sitting or lying position
  Difficulty running and jumping
  Walking on tip toes
  Large calf muscles
  A waddling gait
  A delay in using language
  Learning disabilities

Approximately 90% of patients with DMD die from cardiomyopathy (a chronic heart disease in which the heart muscle is thickened, abnormally enlarged, or stiffened) or muscular respiratory failure (Finsterer 2006). Endocrine (hormonal) problems also appear in DMD (as well as some other muscular dystrophies), and the glucocorticoid medications frequently used for treatment can have additional adverse effects on the hormonal system (Ashizawa 2011). Furthermore, some studies have reported that DMD patients have problems with blood clotting, which can complicate surgery (Morrison 2011).

“There is currently no effective treatment for this disease,” Gersbach said. “Patients usually are in a wheelchair by the age of ten and many die in their late teens or early twenties.”

Duchenne muscular dystrophy has been extensively studied by scientists, and it is believed that more than 60 percent of patients with this type of mutation can be treated with this novel genetic approach.

Genetic editing shows promise in Duchenne muscular dystrophy “Previous studies indicate that restoring the production of dystrophin proteins will be highly functional and alleviate disease symptoms when expressed in skeletal muscle tissue,” said Ousterout.

Similar approaches could be helpful in treating other genetic diseases where a few gene mutations are responsible, such as sickle cell disease, hemophilia, or other muscular dystrophies, Gersbach said.


Other members of the team were Duke’s Pablo Perez-Pinera, Pratiksha Thakore, Ami Kabadi, Matthew Brown, Xiaoxia Qin, and Olivier Fedrigo. Other participants were Vincent Mouly, Universite Pierre at Marie Curie, Paris, and Jacques Tremblay, Universite Laval, Quebec.

Citation: “Reading frame correction by targeted genome editing restores dystrophin expression in cells from Duchenne muscular dystrophy patients,” David Ousterout, et. al, Molecular Therapy, DOI 10.1038/mt.2013.111.


Richard Merritt

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Duke University

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