Gene therapy of myodystrophy

Gene therapy has put mice with myodystrophy on their feet

Polina Loseva, N+1

American scientists have tested whether it is possible to restore muscle function in mice in the late stages of myodystrophy. It turned out that one injection of a gene therapy drug is enough to make the animals walk longer and spend more time standing upright. In addition, the mice stopped dying — although they began to be treated at the moment when half of the animals had already died from the disease. The work was published in the journal Science Advances (Yonekawa et al., Large1 gene transfer in older myd mice with severe muscular dystrophy restores muscle function and greatly improves survival).

Muscular dystrophy, or myodystrophy, is a collective name for a group of genetic diseases in which muscle cells gradually die off. The most famous and widespread of them — Duchenne myodystrophy — is caused by a mutation in the dystrophin gene. This protein binds contractile fibers inside the muscle cell to the extracellular matrix and signaling proteins and transmits the contraction force to the surrounding tissue. Without it, the binding complex does not form, the cells work idly and receive distorted signals — and as a result they die.

This is the protein complex that binds contractile fibers (actin, bottom left) to the extracellular matrix (top). The complex includes, among others, dystrophin (red) and two dystroglycans (light blue). Nowak et al. / EMBO Reports, 2004

The rate at which muscle fibers stop working varies from dystrophy to dystrophy. Sometimes it happens quite slowly, and the disease causes only mild discomfort. But Duchenne myodystrophy, like some other dystrophies, develops rapidly and leads to progressive paralysis. As a result, patients often do not live to adulthood or maturity.

There is no treatment for myodystrophy yet. Scientists are trying to develop drugs based on gene therapy and the CRISPR/CAS system — and have already tested them on mice, pigs and dogs. Such drugs really restore the production of the necessary protein in the muscles. But it is unclear how much they can be applied to people. The fact is that people are usually diagnosed with myodystrophy quite late — by this time many muscle fibers have already been damaged, and it is unclear whether it is possible to return them to working capacity.

To find out, a group of researchers led by Kevin P. Campbell from The University of Iowa worked with mice of the myd line. They carry a mutation in the Large1 gene and suffer from dystroglycanopathy — another binding muscle protein, dystroglycan, does not work for them. To be more precise, an enzyme is not produced that attaches a carbohydrate "tail" to the dystroglycan, with which the protein clings to the extracellular matrix. This model, according to the authors of the work, is good because, firstly, dystroglycanopathy also occurs in humans. And secondly, in mice, such a disease manifests itself quite seriously: they have impaired mobility, they weigh less than usual, and half do not live up to a year (although the average life expectancy of a mouse is 2-3 years, depending on the line). Therefore, they can be used to check whether it is possible to restore the work of the muscles when they are already severely affected.

The authors of the work undertook to treat mice at the 35th week of life - at that moment half of their relatives had already died from dystrophy, that is, muscle damage should be quite serious. They were injected with an adenovirus vector with a "healthy" variant of the Large1 gene, and then began to monitor their activity and compare it with the control group, which did not receive any treatment.

The researchers calculated that after gene therapy, mice walk a much longer distance, being left to themselves, and spend more time standing upright and sniffing the walls of the enclosure. Animals from the control group lost the ability to stand on their hind legs during the first three months of the experiment. In addition, the strength of the muscle grip increased in mice after therapy, although it did not reach the level of a healthy animal.

This is how the physical activity of mice changed after therapy: they began to run longer (B) and spend more time sniffing the walls (C). Orange — control, mice without therapy, blue — mice after therapy. Here and below are the drawings from the article by Yonekawa et al.

The treatment also affected the appearance of the mice — they stopped losing weight. And despite the fact that they still, according to the researchers, looked skinny, and the thickness of their muscles did not reach normal values, they survived much better. At 60 weeks, when all the animals from the control group died (more precisely, they were euthanized for ethical reasons, since their condition was critical), only one mouse out of 14 died in the experimental group.

This is how the weight (J) and survival (K) of mice after therapy changed. Orange — control, mice without therapy, blue — mice after therapy.

The researchers checked that it was really the muscles: they colored skeletal muscle tissue and noticed that the fibers on average became thicker, and there was more dystroglycan in them. The same thing was noticed in the diaphragm, a muscle whose work is critical for patients with myodystrophy.

Dystroglycans beta (red) and alpha (green) in the muscles of healthy animals (left), patients with myodystrophy (center) and after therapy (right).

Unlike Duchenne myodystrophy, dystroglycanopathy affects the central nervous system as well. The Large1 gene works not only in muscles, but also in the brain. The authors measured its expression and found out that gene therapy does not help restore it to the desired level. This could have been expected, given that the adenovirus vector was injected intravenously into mice, and such drugs do not know how to overcome the blood-brain barrier and penetrate into the brain. However, this means that for the full treatment of dystroglycanopathy, it is necessary to figure out how to solve this problem too.

The authors of the article note that not all myodystrophy may be so successfully cured with the help of gene therapy. The Large1 gene is quite short, and easily fits into the adenovirus vector. The dystrophin gene is longer, so it is not possible to deliver it directly — and we have to deliver the CRISPR/Cas system instead, so that it edits the broken gene directly. Nevertheless, this experiment has shown that mammalian muscles are quite plastic and are able to recover at least partially if they are provided with sufficient amounts of the necessary protein in time.

Meanwhile, in humans, gene therapy is already being used to treat other groups of diseases. We wrote that it turned out to be at least safe for Fabry's disease (accumulation of lipids in different organs). And helped with epilepsy in Tay-Sachs disease. And in the trials of gene therapy for vision loss, it was suddenly discovered that the introduction of the drug into one eye improves the work of the other eye, too.

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