Genome editing in the spotlight for the treatment of muscle-wasting conditions

On 31 December 2015, molecular biologists were having a different kind of party; while the rest of the world was ‘seeing in’ 2016, they were celebrating the biggest breakthrough in genome editing so far.

Three research articles, published back to back in the prestigious journal Science showed that a genome editing technique called CRISPR/Cas9, which was hailed “2015 breakthrough of the year” by the journal, could be used to correct a genetic mutation in a fully-grown animal. What’s more, the mutation in question was the one causing Duchenne muscular dystrophy. Through this work, a muscle wasting condition came under the radar of the whole scientific community and the world.


Duchenne muscular dystrophy is caused by a mutation in the gene coding for a vital muscle protein called dystrophin. Dystrophin is essential to maintain muscle cell integrity and acts like a shock absorber at each muscle contraction. In its absence muscle cells get damaged and eventually die.

Different types of mutations can cause the dystrophin protein not to be produced by the cells. These can range from single letter changes in the DNA code to big chunks of the gene missing or extra bits of DNA being introduced. Sometimes these changes can cause the reading frame of the message contained in the gene to be disrupted and no protein to be made.

The mdx mouse is the most readily used animal model of Duchenne muscular dystrophy. As in some people with Duchenne muscular dystrophy, there is a single letter change in the dystrophin gene of the mdx mouse. This change introduces a premature stop signal to the message encoded by the gene. As a result, the message stops being read too early and no dystrophin protein can be produced.

CRISPR/Cas9 is a type of genome editing technique that can be used to make precise, targeted changes to the DNA of a living organism. It uses special DNA-cutting enzymes or ‘molecular scissors’ called Cas9. These are attached to specific molecules to guide them to the desired location in the DNA. The molecular scissors then cut the desired region and the cell repairs the cut in the DNA by joining the cut ends.

Unlike other therapeutic approaches that aim to address the underlying genetic cause of muscle-wasting conditions such as exon skipping or gene therapy using harmless viruses, genome editing offers permanent correction of the genetic mutation causing a condition.

CRISPR/Cas9 can be used to change the genes in adult cells as well as embryos. This can be done with the aim to answer scientific questions by turning certain genes off for example and studying their effect. It can also be used as a potential therapeutic approach to treat genetic conditions such as Duchenne muscular dystrophy.

Genetically modifying an embryo carries with it a lot of ethical considerations. In the UK, this is only allowed for research purposes on early embryos which can be grown in the laboratory for a maximum of 14 days only and with special permission from the Human Fertilisation and Embryology Authority (HFEA). Moreover even if it was allowed, using genome editing on embryos as a treatment would require an in vitro fertilisation (IVF) step. If an IVF step is to be used, pre-implementation genetic diagnosis is a viable approach allowing the selection of healthy embryos for implantation. Therefore the use of genome editing techniques in embryos to prevent the inheritance of muscle-wasting conditions such as Duchenne muscular dystrophy brings little advantage.

The three scientific articles published back to back on the last day of last year used the CRISPR/Cas9 technology to remove the part of the dystrophin gene that is faulty in the mdx mice.

Three independent groups of scientists in the US injected a harmless virus called adeno-associated virus (or AAV) containing the CRISPR/Cas9 system, into the muscle or the bloodstream of the mice. The system was designed in such a way that the molecular scissors could be guided to the exact location in the dystrophin gene where the fault was and cut it out. This way the information contained in the gene could be read properly and dystrophin protein could be produced. As part of the gene had been cut out, the resulting protein was shorter than normal but tests showed that it was still functional. The treated mice were significantly stronger than the untreated mice but still weaker than healthy mice.

One group of researchers also used the CRISPR/Cas9 system to remove the fault in the dystrophin gene in muscle stem cells that they obtained from the mdx mouse and grown in a petri dish in the laboratory. These cells, also called satellite cells, have the ability to turn into adult muscle cells. Using stem cells which have had their mutation repaired by genome editing could then be injected back into the body and replenish mature muscle tissue. With this approach the effect of CRISPR/cas9 should last longer because unlike adult muscle cells, satellite cells keep dividing and therefore passing on the repaired gene to daughter cells that will turn into new mature muscle cells.

Muscular Dystrophy UK is currently funding two research projects and one lectureship using genome editing to correct mutations in the dystrophin gene. All three projects are conducted on adult muscle cells. If the approach is successful, the technique could be the first therapy that offers permanent correction of these genetic mutations causing Duchenne muscular dystrophy in people. The technique could also be adapted for the treatment of other muscle-wasting conditions.

CRISPR/Cas9 is a tremendous technology with great potential to treat genetic conditions such as – but in no ways limited to – Duchenne muscular dystrophy and other muscle-wasting conditions.  The three Science articles provide proof of principle evidence that the technique can be used to repair a genetic mutation in a fully-grown animal (the mouse). They provide evidence supporting the efficacy of the approach. The authors are optimistic that it can be applied to people in the foreseeable future. However there is still more work to be done to optimise the design of the delivery system carrying the molecular scissors to the muscle. More work also needs to be done towards enhancing the safety and efficacy of the system before its full potential as a treatment can be achieved. For this reason, it is difficult to predict a time line to trials in humans.  In summary, it is a promising technique that has great therapeutic potential and will almost certainly be developed further by scientists in the future.

 

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