Dr Shivanee Shah
Muscular dystrophy is group of muscle disorders characterized by progressive muscle weakness, with Duchenne Muscular Dystrophy (DMD) being the most common. DMD is an X-linked recessive disorder primarily affecting males, with women being carriers. DMD is characterized by initial proximal muscle weakness of the pelvic girdle that eventually progresses to the shoulder girdle muscles. The onset of symptoms typically occurs between 3 to 5 years of age and by the age of 13-15 years, the child is generally wheelchair-bound. DMD is caused due to mutations in the dystrophingene which lies on the short arm of the X-chromosome (Xp21.2). The dystrophingene encodes the protein dystrophin, which plays a key role in maintaining the integrity of the cell membrane of skeletal and cardiac muscle cells. In the absence of dystrophin, muscle fibres disintegrate, resulting in the muscle weakness observed in DMD.
One such nine-and-a-half-year-old boy was brought to consult Dr. K. N. Shah at Lilawati Hospital and Research Centre, Mumbai. His previous history revealed that he was born to parents from a non-consanguineous marriage and had one female sibling. He was born via a normal, full-term pregnancy, and showed normal milestones up to 2 years of age. However, after two years, his parents noticed that his walking was delayed, though his social and cognitive milestones were normal. It was also reported that he was unable to climb stairs and his calf muscles had begun show pseudohypertrophy. As his condition slowly progressed, he was unable to get up from a sitting position without additional support to his knees, an observation that is commonly called the Gower’s sign and is classically observed in DMD children. Creatinine phosphokinase (CPK) levels, a marker of muscle injury, are consistently elevated in patients with Duchenne’s and even this child had a very high level (20,000 U/L) of CPK, strongly indicative of muscle dystrophy. However, Dr. Shah also cautions that elevated CPK levels cannot specifically confirm a diagnosis of DMD, since it is simply an indicator of muscle damage, and not indicative of DMD specifically.
While the gold standard of diagnosis for DMD is muscle biopsy followed by immunohistochemistry or western blot for dystrophin protein, new technological advances involve diagnosis via genetic testing. To appreciate how genetic testing can help diagnose DMD, it is first important to understand the dystrophingene at the genetic level and the types of mutations that have been identified in the recent past. Dystrophin gene is one of the largest known genes, consisting of 79 exons, many of which are repeating segments. Typically, mutations in this gene are either deletions or duplications of single or multiple exons.
A deletion or duplication of an exon can result in a change wherein dystrophin cannot be expressed at all. Such a mutation is called an ‘out-of-frame’ mutation. Alternatively, in some instances, even
though some exons maybe deleted or duplicated, the protein may still be expressed, albeit one that is not as effective as the wild-type one. Such mutations are called ‘in-frame’ deletions. Thus, the type of mutation can determine quantitative as well as qualitative changes in the expression of the dystrophin protein, resulting in different disorders. For instance, if the mutation results in the deletion of an exon due to which dystrophin cannot be expressed at all, the resulting dystrophy is DMD. However, if the mutation results in an exon being deleted, despite which dystrophin can be formed, then the resulting dystrophy is a milder version called Becker muscular dystrophy or BMD.
Since the progression of BMD is much slower than in DMD, and the treatment and management can vary depending on the particular muscle dystrophy, it is often critical to carry out a differential diagnosis by genetic testing.
Multiplex Ligation-Dependent Probe Amplification (MLPA) is a commonly employed genetic detection test, which can detect up to 70% of DMD positive cases. In the event of a negative MLPA result, clinicians still have the option of confirming the disease using next generation sequencing, which can pick up about 98-99% of the positive cases. However, muscle biopsy still remains the final confirmatory test. In this case, Dr. Shah proposed that MLPA be done, and the results revealed that exon 43 was deleted, which confirmed the clinical diagnosis of DMD. Genetic counselling would be important in case the parents wish to have another child, since prenatal genetic testing via MLPA at early stages of pregnancy can help determine whether the male child would have DMD. The prenatal tests can also help parents decide whether they should terminate such a pregnancy. Further, the parents of the boy were advised to get his sister also genetically tested to determine if she was a carrier for DMD mutation.
While currently there is no known cure for DMD, there is hope for the future. Several research studies have been carried out for different genetic therapy approaches. Exon skipping is a mutation-specific therapy in which a small piece of DNA can be introduced such that it will bind to a particular exon and encourage the cell to skip reading that exon, thereby effectively converting an out-of-frame mutation to an in-frame mutation. Such a therapy would allow DMD-type clinical conditions to mimic BMD-type clinical conditions, which are far more manageable and have better prognosis. However, till date, only a few specific therapies are approved. For Dr. Shah’s patient, there is no therapy for a deletion of exon 43. The only US FDA approved therapy is for deletions that can be corrected by skipping exon 51, involved in about 13-20% of DMD cases. However, research is ongoing, and it is likely that other, newer exon-skipping targets become available in the future.