Gene Editing Tool Crispr-Cas9 Corrects Mutations In Muscular Dystrophy Patients

Researchers from the U.S. and Germany have used the gene editing tool CRISPR-Cas9 in a novel way to produce healthy heart muscle in Duchenne muscular dystrophy patients. The study is published in Science Advances.

Duchenne muscular dystrophy (DMD)

DMD is a progressive muscle wasting genetic disease which leads to degeneration of cardiac and skeletal muscles. Only boys are affected with DMD while girls can be asymptomatic carriers. The disease is caused by over 3000 different mutations in the dystrophin gene. Dystrophin is a protein that links the cytoskeleton of a muscle fiber to the surrounding extracellular matrix of muscle cells through the cell membrane. It maintains the integrity of the plasma membrane, acts as a shock absorber and prevents our muscles from breaking down as a result of wear and tear. Degeneration of the cardiac muscle, the muscle most affected by lack of dystrophin is the most common cause of death in DMD patients.

Mutations and their effect on dystrophin

The dystrophin gene is the largest gene in our body that can be roughly divided into three sections, the head, body, and tail. Mutations in the body are called internal mutations and can result in a partially functional protein. In this case, the mutation causes a milder form of muscular dystrophy called Beckers muscular dystrophy. However, if the mutations are in the head or tail exons, also called the “essential” regions of the gene, dystrophin is not produced and results in the severe and more lethal form of the disease called Duchenne muscular dystrophy. Without dystrophin, the patient’s muscle cells become leaky and eventually die. This causes degeneration of muscle tissue gradually spreading throughout the body. When the heart muscles degenerate, the condition is fatal.

CRISPR-Cas9 gene editing technology works by creating a nick at the desired spot on the genome where the surrounding gene sequence has to be edited or repaired. Mutations usually cluster themselves together in regions known as “hotspots.” The presence of 3000 odd mutations in DMD patients was making it difficult to use the CRISPR tool in DMD. To overcome this problem, a novel technique called “myoediting” was used along with CRISPR-Cas9 technology that can edit or repair entire clusters of mutations. It removes mutated sections of the dystrophin gene that cause DMD and restores normal function to cells with DMD mutations.

The researchers took cells from DMD patients and converted them to stem cells. The stem cells were then myoedited and reprogrammed to grow into heart muscle cells that were able to produce dystrophin protein. Thus when the cells made protein from the genes that underwent myoediting, the hotspots were ‘skipped-over;’ specifically, twelve mutation hotspots of Duchenne muscular dystrophy (DMD) were corrected to produce functional dystrophin protein. In the study, a tiny bit of heart muscle tissue was then grown, and the resultant tissue could beat and remained healthy.

While this approach cannot correct all DMD mutations, it may be a potential therapeutic for over 60% of the DMD afflicted population. Based on the type of mutation present myoediting can either correct the gene to produce a perfect dystrophin protein or a partially functional protein. If several internal exons are skipped, there is some abnormality in the dystrophin protein, and that would resemble Beckers muscular dystrophy. But, this too, is a significant relief for those with DMD. In case of insertions and duplications in the DMD gene, these can be perfectly corrected to restore a fully functional dystrophin protein.

References:

1. Correction of diverse muscular dystrophy mutations in human engineered heart muscle by single-site genome editing - (http://advances.sciencemag.org/content/4/1/eaap9004.full)
2. Long, C. et al. Correction of diverse muscular dystrophy mutations in human engineered heart muscle by single-site genome editing. Science Advances 4, eaap9004 (2018).

Source-Medindia