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ӳ����ý researchers have developed a way to edit the genetic sequences at the root of Huntington’s disease and Friedreich’s ataxia.

The conditions are two of more than 40 severe neurological disorders caused by three-letter stretches of DNA that repeat consecutively. If longer than a certain threshold length, these sequences grow in length uncontrollably and lead to brain cell death in Huntington’s disease, and the breakdown of nerve fibers in Friedreich’s ataxia. There are no treatments that stop the progression of these diseases.

The ӳ����ý team has developed a new approach to prevent these DNA repeats from expanding. Using a technique called base editing, the team introduced single-letter changes into the middle of the repeated stretch of DNA, interrupting the sequence in patient cells and mouse models of Huntington’s disease and Friedreich’s ataxia. They found that the edited DNA tracts stayed the same in length or even became shorter over time.

The findings appeared recently in and come from the lab of gene-editing pioneer David Liu, the Richard Merkin Professor and director of the Merkin Institute of Transformative Technologies in Healthcare at the ӳ����ý. Liu is also an investigator with the Friedreich’s Ataxia Accelerator at ӳ����ý, a professor at Harvard University, and a Howard Hughes Medical Institute investigator. Mandana Arbab, a postdoctoral researcher in Liu’s lab when the study began who now holds the Lodish Family chair as an assistant professor at Boston Children’s Hospital, and Zaneta Matuszek, then a graduate student, are co-first authors on the study. Ricardo Mouro Pinto, a ӳ����ý associated scientist and an assistant professor at the Mass General Research Institute, is a co-corresponding author along with Liu.

More work will be needed to catalog the potential side effects of installing these edits in the genome, but the researchers say that the approach could be a valuable tool for understanding diseases caused by these kinds of DNA repeats.

“A lot more studies would be needed before we can know if disrupting these repeats with a base editor could be a viable therapeutic strategy to treat patients,” said Liu. “But being able to illuminate the biological consequences of interrupted repeats is a really useful and important milestone.”

Repeat interrupted

About one in 3,000 people has a disease caused by three-letter repeats in DNA. Individuals with the same disease can inherit different numbers of DNA repeats — people with more repeats generally experience symptoms sooner and more severely, and their diseases progress faster. By contrast, some patients have naturally occurring single-letter “interruptions” within these repeats, and have milder symptoms that develop later. These individuals are also less likely to pass their disease along to their children than people with uninterrupted repeats.

That gave Matuszek and Arbab an idea. If a gene-editing therapy could install an interruption that mimics those that occur naturally in some patients, it might stop the repeat from expanding and halt or slow down disease progression.

Matuszek and Arbab decided to use base editing, a tool developed by Liu’s lab in 2016 for making single-letter changes in DNA. They came up with a system that changed some G•C pairs to A•T pairs within a CAG repeat tract for Huntington’s disease. Another edited several A•T pairs to G•C pairs within the GAA repeats of Friedreich's ataxia.

When the team tested the editors in connective tissue cells derived from patients with the disorders, the number of repeats within treated cells stayed the same or even decreased over time — but untreated cells had more repeats than before.

To shuttle the base editors to specific cells in mice, the researchers packaged them into dual AAV9 vectors, an adeno-associated virus designed to deliver cargo to neurons. The base editors stabilized the repeat tracts in mouse models of Friedreich’s ataxia and Huntington’s disease.

“What’s really exciting is that we now have a tool to introduce interruptions in cell and animal models and study how they affect the biology of these diseases,” Matuszek said.

A new editing strategy

Liu cautions that because these repeat sequences occur elsewhere in the genome, the base editors can make edits in those parts of the genome, raising the possibility of unwanted side effects. But so far, his team has found that most off-target editing occurred in parts of the genome that are between genes or do not encode proteins, reducing the chance of unwanted side effects in people. The researchers plan to study these effects in specific cell populations and animal models that more faithfully mirror human disease.

However, the team thinks that a therapeutic strategy that introduces interruptions into repeat tracts could one day help treat Huntington’s, Friedreich’s ataxia, and other trinucleotide repeat disorders, because such changes in DNA occur naturally in people without disease or who only have mild symptoms.

“Not only does this study show for the first time that inducing interruptions has a profound stabilizing effect on repeats, but that the base-editing approach we’ve used can also be applied to study any of over a dozen repeat disorders,” Arbab said. “There’s still a lot of work to be done, but we’re hopeful that this approach could really accelerate therapeutic development for a lot of these diseases.”

In the meantime, the researchers are also developing a different approach using prime editing to replace disease-causing repeat tracts with a shorter, stable number of repeats all at once.