Research part-funded by the Muscular Dystrophy Campaign led by Prof George Dickson has used exon skipping technology in mice to block the activity of myostatin, a protein that prevents muscles from growing bigger and stronger. Scientists think that by blocking the activity of myostatin, it might be possible to build up muscle size and strength in people with muscle disease. The team found that small pieces of DNA called antisense oligonucleotides (AOs) were able to inactivate myostatin in muscle cells grown in the laboratory. Injection of AOs into the bloodstream of healthy mice were able to increase the size of one of the three muscles that were analysed. This research has proven the principle that exon skipping technology could be used to control the activity of myostatin.
Myostatin is a protein naturally produced by the body. It is an inhibitor of muscle growth and, together with other proteins that promote muscle growth, it works to keep the size and strength of muscle within the normal range. Researchers have shown in animals that blocking the activity of myostatin causes the muscles to increase in strength and size. This approach is attractive because it may be a useful way to “bulk up” muscles in people with muscle disease, helping to increase their muscle strength.
Scientists have investigated a number of approaches to blocking myostatin activity, including a potential drug called ACE-031, a clinical for which was started in May. A gene therapy approach using a virus also recently gained funding for a clinical trial.
Prof Dickson decided to investigate using exon skipping to block myostatin activity following recent research that suggested this technology could be used to block the activity of certain genes. Exon skipping is currently in clinical trial for Duchenne muscular dystrophy in both the UK and Europe, where the AOs are being used to restore the production of the dystrophin protein. Initial results have been promising and the treatment has been well tolerated by the trial participants. There are two different chemistries of AOs being tested in these clinical trials.
Exon skipping works by masking part of a gene – an exon – so that the cell skips over it when the protein is being produced. In the case of exon skipping for Duchenne muscular dystrophy the AOs are designed to specifically mask the part of the dystrophin gene that contains a mistake, allowing a shortened but functional version of the protein to be made. The same technology can be applied to a healthy gene to disrupt the instructions and prevent protein production. In this new research AOs were designed to block the activity of the myostatin gene by skipping over an exon to disrupt the DNA code so that the cell no longer makes any myostatin protein.
The myostatin gene has three exons; Prof Dickson suggested that by making the cell skip over the second exon of the myostatin gene, it would no longer be able to read the instructions and so no myostatin protein would be produced. If no myostatin protein is produced, the muscles should be able to grow larger. To find out how this compares to the exon skipping in clinical trial for Duchenne muscular dystrophy see our FAQ.
The researchers designed and tested a number of AOs based on the different chemistries in clinical trial for Duchenne muscular dystrophy. They found that all the patches tested were able to induce exon skipping of myostatin in muscle cells grown in the laboratory, but to varying degrees. The cells treated with AOs multiplied in number more quickly.
The team then moved on to look at the effect of the AOs in healthy mice. They injected the AOs into the bloodstream of the mice and could detect myostatin exon skipping in the muscle. They measured the size of three different muscles in the leg and the size of one of them was found to be increased – the soleus muscle. Prof Dickson suggested that only one of the muscles increased in size because myostatin is particularly active in the soleus muscle so a greater difference could be seen when it was inactivated. To increase muscle size across the body the dose of AOs and frequency of administration might also need to be optimised.
The results of this study represent proof of principle, in the mouse, that exon skipping could be used to block myostatin activity. It is possible that this approach could be used for a range of neuromuscular conditions although it will have its limitations, because it does not mean that the genetic defect is repaired nor does it involve the addition of a healthy copy of the faulty gene into the muscle cells. This approach may work best in conjunction with other types of therapy so that both the primary cause of the muscle condition is treated and the muscle strength and size is built up.
Using exon skipping to block myostatin activity may have advantages over some of the other methods that are being developed to inhibit myostatin production. It is already known from recent trials for Duchenne muscular dystrophy, that AOs are well tolerated in humans. Some of the other approaches, such as gene therapy and the use of antibodies to block myostatin, could cause an immune reaction and thus prevent the therapy from acting or produce undesirable side effects for the patient.
These are preliminary experiments carried out in healthy mice and not mouse models of muscle disease, so further work will need to be done to determine how well these AOs might function in damaged muscle. It is clear that further work needs to be done to increase the efficiency of exon skipping to control myostatin throughout the muscles of the body. This does, however, represent a promising start.
Prof. George Dickson, said:
We are pleased and excited by these results which suggest that general muscle wasting and weakness which is associated with many neuromuscular diseases may be amenable to treatment by myostatin exon skipping. In the case of Duchenne muscular dystrophy, we envisage a cocktail of exon skipping “drugs” targeting both dystrophin and myostatin at the same time. Such an approach offers the prospect not only to rescue muscles from dystrophin deficiency but also to retrieve muscle size and strength.
Read more about exon skipping.
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