Tag: muscular dystrophy

Categories : Diseases, Syndromes and Conditions GeneticsTags :

Scientists Snip Muscular Dystrophy Gene, Yielding Shorter but Now-functional Proteins

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CRISPR-Cas9 is a customisable tool that lets scientists cut and insert small pieces of DNA at precise areas along a DNA strand. This lets scientists study our genes in a specific, targeted way. Credit: Ernesto del Aguila III, National Human Genome Research Institute, NIH

The most common inherited muscular disorder and one of the most severe, Duchenne muscular dystrophy (DMD) results from mutations of the dystrophin gene. In the journal Stem Cell Reports, researchers used a dual CRISPR RNA method to restore dystrophin protein function in stem cells derived from DMD patients. By removing large sections of the dystrophin gene, the cells were able skip faulty or misaligned sections of the genetic code, yielding shortened but still functional proteins for a wide variety of mutation patterns associated with DMD.

“Dual CRISPR-Cas3 is a promising tool to induce a gigantic genomic deletion and restore dystrophin protein via multi-exon skipping induction,” says senior author Akitsu Hotta of Kyoto University. “We expect this study to enlighten new ways to treat DMD patients and other genetic disorders that require extensive deletions.”

Due to significant variations in the mutation patterns affecting the dystrophin gene, deleting a small section of the gene can only be used for a limited number of DMD patients. For example, the most common mono-exon skipping of exons 51, 53, and 45 can be applied to 13%, 8%, and 8% of DMD patients, respectively.

Multi-exon skipping (MES) has broad applicability to various DMD mutation patterns. By targeting the mutation hotspots in the dystrophin gene, MES from exon 45 to 55 was estimated to benefit more than 60% of DMD patients. Unfortunately, few techniques are available to induce a large deletion to cover the target exons spread over several hundred kilobases.

To overcome this hurdle, Hotta and his team used CRISPR-Cas3 to induce a deletion of up to 340 kilobases at the dystrophin exon 45-55 region in various DMD mutation patterns. Because it was rare to observe a deletion of more than a hundred kilobases using a single CRISPR RNA – which helps to locate the correct segment of DNA – the researchers used a pair of CRISPR RNAs inwardly sandwiching the target genomic region.

Limitations of the dual CRISPR RNA system include is variation in the deletion pattern, and the precise start and end points of the deletion cannot be fully controlled. This could be a drawback when a large but precise deletion is required. The study also did not demonstrate the functionality of the recovered dystrophin protein. Future research should aim to improve the overall genome editing efficiency of the Cas3 system.

“Our dual-Cas3 system might apply to future gene therapies once we’re able to deliver the dual-Cas3 components in vivo to skeletal muscle tissues safely and efficiently,” says Hotta. “The ability to induce several hundred kilobases of DNA deletion itself also has broad applicability for basic research when a large deletion is needed.”

Source: Science Daily

Categories : Metabolic DisordersTags :

Weekly Prednisone Could Reduce Obesity and Help in Muscular Dystrophy

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Source: Pixabay CCO

In a study on obese mice fed a high-fat diet, receiving prednisone once weekly improved their exercise endurance and strength, and their reduced weight. The study was published in the Journal of Experimental Medicine. A previous study had found that weekly prednisone was helpful in muscular dystrophy.

“These studies were done in mice. However, if these same pathways hold true in humans, then once-weekly prednisone could benefit obesity,” explained senior author Dr Elizabeth McNally.

“Daily prednisone is known to promote obesity and even metabolic syndrome – a disorder with elevated blood lipids and blood sugar and weight gain,” Dr McNally said. “So, these results, in which we intermittently ‘pulse’ the animals with once-weekly prednisone, are strikingly different. Obesity is a major problem, and the idea that once-weekly prednisone could promote nutrient uptake into muscle might be an approach to treating obesity.”

The once-weekly prednisone, a glucocorticoid steroid, promoted nutrient uptake into the muscles. The researchers also found these mice had increased adiponectin levels, an adipocyte-secreted hormone involved in protecting against diabetes and insulin resistance. Mice that were already obese from eating a high-fat diet were also found to benefit after once-weekly prednisone, experiencing increased strength, running capacity and lower blood glucose.

Most knowledge on steroids like prednisone is derived from studies of daily doses of prednisone

“We see a very different outcome when it is taken once a week,” Dr McNally said. “We need to fine tune dosing to figure out the right amount to make this work in humans, but knowing adiponectin might be one marker could provide a hint at determining what the right human dose is.”

Dr McNally described the weekly dose as “a bolus, or spike, of nutrients going into your muscle.”

“We think there is something special about promoting this spike of nutrients into muscle intermittently, and that it may be an efficient way to improve lean body mass,” she added.

“What is exciting to me about this work is the finding that a simple change in the dosing frequency can transform glucocorticoid drugs from inducers to preventers of obesity,” said corresponding author Mattia Quattrocelli. “Chronic once-daily intake of these drugs is known to promote obesity. Here we show that dosing the same type of drug intermittently – in this case, once weekly – reverses this effect, promotes muscle metabolism and energy expenditure, and curtails the metabolic stress induced by a fat-rich diet.”

Many patients take prednisone daily for different immune conditions, which has side effects including weight gain and even muscle atrophy with weakness. Investigators want to determine whether patients can get the same immune benefit with intermittent prednisone dosing, which could be much more beneficial to the muscle.

Dr McNally’s team previously found that intermittent prednisone administration was helpful for muscular dystrophy, showing once weekly prednisone improved strength, recently reporting that a pilot trial in humans with muscular dystrophy in which one weekly dose of prednisone improved lean mass.

McNally wants to identify biomarkers most critical to measure a beneficial response to prednisone.

“If we can determine how to choose the right dose of prednisone that minimises atrophy factors and maximises positive markers like adiponectin, then we can really personalise the dosing of prednisone,” she said.

The group also recently showed that weekly prednisone uses strikingly different molecular pathways to strengthening the muscle in male versus female mice, based on a new study just published in the Journal of Clinical Investigation by Isabella Salamone, a graduate student in Dr McNally’s lab.

The benefits of weekly prednisone are linked to circadian rhythms, according to another new study published in Science Advances. Human cortisol and steroid levels spike early in the morning before awakening.

“If you don’t give the drug at the right time of day, you don’t get the response,” Quattrocelli said. “In mice, we obtained good effects with intermittent prednisone in muscle mass and function when we dose them at the beginning of their daytime. Mice have a circadian rhythm inverted to us, as they generally sleep during the daytime and are active at night. This could mean that the optimal dosing time for humans during the day could be in the late afternoon/early evening, but this needs to be appropriately tested.”

Conducting these studies in mice is a major limitation, Dr McNally said.

“While we are encouraged by the pilot study in humans with muscular dystrophy, mouse muscles have more fast-twitch fibres than humans, and slow-twitch muscle could be different,” Dr McNally said. “More studies are needed to try to better understand whether these same mechanisms work in human muscles.”

Source: Northwestern University