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Correct muscle stem cell mutations using new gene editing technology

New gene editing techniques can be used to correct mutations in muscle stem cells and pave the way for the first potential cell therapies for hereditary muscle disorders. The ECRC team, led by Professor Simone Spuler, published the findings in the journal JCI Insight.

Muscle stem cells allow our muscles to accumulate and regenerate throughout life through exercise. But if a particular muscle gene is mutated, the opposite happens. In patients with muscular dystrophy, skeletal muscle has already begun to weaken in childhood. Suddenly, these children are unable to run, play the piano, or climb stairs, and often rely on wheelchairs by the age of 15. Currently, there is no cure for this condition.

Currently, CRISPR-Cas9 technology can be used to access genetic mutations in these patients. We cared for more than 2,000 patients with muscle disorders at the Charite outpatient department and immediately realized the potential of new technology. “

Professor Simone Spuler, Director of Myology Laboratory, Experimental Clinical Research Center (ECRC)

Researchers soon began working with some of the affected families and now publish their results in journals. JCI Insight.. In the family studied, the parents were healthy and unaware that they had the mutated gene. All the children inherited a copy of the disease mutation from their parents.

Edited human muscle stem cells developed into mouse muscle fibers

The term “muscular dystrophy” is used to refer to about 50 different diseases. “They all follow the same course, but they are different because of mutations in different genes,” Spuler explains. “And even within the gene, different sites can be mutated.” Following genomic analysis of all patients, researchers chose one family for a particular form of the disease: limb band. Limb-girdle dystrophy 2D / R3 is relatively common, progresses rapidly, and has a suitable docking site for “gene scissors” for DNA mutations.

In this study, researchers took muscle tissue samples from a 10-year-old patient, isolated stem cells, proliferated them in vitro, and used base editing to replace base pairs at the site of mutation. The edited muscle stem cells were then injected into mouse muscle. This allows it to withstand foreign human cells. These proliferated in rodents and most developed into muscle fibers. “This was the first time we were able to show that it is possible to replace diseased muscle cells with healthy muscle cells,” says Spuler. After further testing, the repaired stem cells are reintroduced into the patient.

Base editing-sophisticated techniques

Base Editing is a new and highly sophisticated variant of the CRISPR-Cas9 gene editing tool. In the “classical” method, both strands of DNA are cleaved with these molecular scissors, but the Cas enzyme used for base editing simply cuts residual glucose from one base and attaches another. So the point where you want to create another base on the target. “This tool acts more like tweezers than scissors and is ideal for performing target point mutations in genes,” said Dr. Helena Escobar, a molecular biologist on Spuler’s team. “This is also a much safer method, as unwanted changes are so rare. In genetically repaired muscle stem cells, we did not see any misediting in unintended regions of the genome,” Escobar studies. The lead author of, who developed the technology for muscle cells.

Autologous cell therapy, which removes the patient’s own stem cells, edits them outside the body, and then injects them into the muscle, prevents the patient, who is already in a wheelchair, from walking again. “We can’t repair muscle that has already atrophied and replaced connective tissue,” Spuler emphasizes. Also, the number of cells that can be edited in vitro is limited. However, this study provides the first evidence that some previously incurable groups of illnesses can be treated and used to repair small muscle defects such as the flexor digitorum superficialis. I will.

One step closer to treatment

But this is just the first step. “The next milestone is to find a way to inject the base editor directly into the patient. Once inside the body, it” swims “for a while, edits all muscle stem cells, and then quickly breaks down again. The “” team wants to start the first test on the mouse model immediately. If this also works, in the future, newborns can be tested for the corresponding genetic mutation and curative therapy can be started when there are relatively few cells that need to be edited.

So what exactly does in-vivo treatment for muscular dystrophy look like? This is what scientists have been testing in animal models for some time using viral vectors. However, Helena Escobar explains that these vectors stay in the body for long periods of time, so the risk of misediting and toxic effects is too high. “Another method is an mRNA molecule that contains information for editors to synthesize tools in vivo,” says molecular biologists. “The mRNA breaks down so quickly in the body that the therapeutic enzyme can remain active for a short period of time.” Treatment can be repeated if desired. “I’m still not sure if this needs to be a treatment cycle that involves several applications.”

This treatment, unlike autologous cell therapy, means that not all patients need to be treated individually. For each form of muscle therapy, one “tool” is sufficient to treat muscle atrophy before major damage occurs. But for now, that’s a long way to go.


Journal reference:

Escobar, H. , et al. (2021) Base editing repairs SGCA mutations in human primary muscle stem cells. JCI Insight.

Correct muscle stem cell mutations using new gene editing technology

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