Target gene identified for therapies to combat muscular dystrophy

AGÊNCIA FAPESP/DICYT Researchers at the University of São Paulo’s Bioscience Institute (IB-USP) in Brazil have shown that a gene called Jagged1, or JAG1 for short, could be a target for the development of new approaches to treat Duchenne muscular dystrophy (DMD), a genetic disorder characterized by progressive muscle degeneration.

The research was carried out at the Human Genome and Stem-Cell Research Center (HUG-CELL), one of the Research, Innovation and Dissemination Centers (RIDCs) supported by FAPESP. The findings were published in the journal Cell.

“All the genetic therapies tested so far, with little success, have targeted the gene that codes for the protein dystrophin. We’re presenting a different approach, which opens up a range of new possibilities,” said Mayana Zatz, Full Professor of Genetics at IB-USP and Head of HUG-CELL.

Duchenne muscular dystrophy, Zatz explained, primarily affects males and is the most common and most rapidly progressing type of muscular dystrophy. It is caused by a mutation, usually inherited, in the gene that encodes dystrophin, a protein that is essential for muscle health and is entirely absent in DMD patients.

“Dystrophin maintains the integrity of the membrane surrounding muscle cells,” Zatz said. “When this protein is absent, the membrane becomes flaccid, so important proteins leak out of the muscle tissue and enter the bloodstream. Conversely, substances that should remain outside, such as calcium, are able to get in.”

The heart, diaphragm and skeletal muscles are affected. Difficulty with walking and running first appear in boys aged between 3 and 5. Patients are usually confined to a wheelchair by age 10-12.

“Without special care, patients don’t reach the age of 20. Nowadays, with assisted breathing, they may survive until they’re 40 or beyond,” Zatz said.

Over the past 15 years, researchers at HUG-CELL have performed experiments to extend our knowledge of DMD. They have studied animals such as Golden Retrievers born with a dystrophin gene mutation, which develop a clinical condition similar to human DMD. Most dystrophic dogs live for only two years or less.

“Some time ago, we identified a dog that totally lacked dystrophin yet presented with a much milder form of the disease,” Zatz said. “It survived for 11 years, considered normal for this breed, and left a descendant that inherited the mutation and is now nine years old.”

Ringo and Suflair, as the dogs were called, became the center of attention for the research group, especially Natássia Vieira, then a PhD student. In partnership with Sergio Verjovski-Almeida, a professor at the University of São Paulo’s Chemistry Institute (IQ-USP), Vieira compared gene expression in healthy dogs, dogs with severe muscular dystrophy, and the above two dogs with the milder form of the disease.

“We found some candidate genes, and thanks to a partnership with Professor Louis Kunkel and his team at Harvard Medical School, and Professor Kerstin Lindblad-Toh at the Broad Institute of MIT and Harvard in the United States, it was possible to combine the results with genetic data. We identified a region of the genome that’s associated with the benign clinical condition and found increased expression of JAG1, a gene in this region, in Ringo and Suflair. This could explain why they have a more benign form of the disease,” said Vieira, the first author of the article published in Cell.

Proof of concept

To confirm that alterations in JAG1 expression can indeed affect the severity of the disease, Vieira conducted experiments using a zebrafish model. The zebrafish (Danio rerio) shares approximately 70% of its genome with humans.

The experiments were performed in Prof. Kunkel’s laboratory. Prof. Kunkel discovered the dystrophin gene and is currently a researcher at Harvard Medical School.

“The zebrafish model also has a dystrophin gene mutation,” Zatz said. “As a result, its muscles are weak, and it can’t move. When we increased JAG1 expression in zebrafish without dystrophin, we found that 75% failed to develop the dystrophic phenotype.”

Overexpression was induced in the model by injecting embryos with JAG1 messenger RNA.

According to Zatz, the next step is to try to reproduce the experimental findings in a mouse model that lacks dystrophin and has a mutation in the gene that encodes the protein utrophin.

“These rodents typically survive for about four months,” Zatz said. “If we succeed in increasing their survival with this treatment, it will be a very positive sign.”

New strategy

According to Zatz, there is a milder form of the disease in humans, and this is known as Becker muscular dystrophy (BMD). Patients with BMD have a different mutation in the same gene: they produce dystrophin, but in smaller amounts than healthy individuals. The clinical course of BMD is less severe, and motor difficulty typically sets in after patients reach 20 years of age.

“The more dystrophin that is produced, the less severe the phenotype,” Zatz said. “For this reason, genetic therapies have hitherto targeted dystrophin. We’ve shown there can be a different approach. JAG1 may not be the answer, but a different protein on the same pathway may be identified in future studies.”

For Kunkel, one possibility could be the development of a drug that increases production of JAG1 and thereby improves patient well-being. “However,” he told Agência FAPESP, “I don’t believe genetic therapy focusing on JAG1 will replace genetic therapy focusing on the dystrophin gene. It might be a complement.”

Kunkel said he will continue collaborating with the Brazilian group and plans to test new drugs capable of boosting JAG1 expression.

“This project shows how interdisciplinary research really can be useful and move science forward,” said Lindblad-Toh.