Just 11 years ago, the development of CRISPR genome editing launched a new era in biology and medicine, with new possibilities to treat genetic diseases. The first CRISPR-based cure, a treatment for sickle-cell disease, is expected to get FDA approval this year.

Now, an ambitious new initiative led by Cathleen Lutz, PhD, Vice President of the Rare Disease Translational Center at The Jackson Laboratory (JAX), in collaboration with David Liu, PhD, of the Broad Institute and other researchers and physicians at Massachusetts General Hospital, Boston Children’s Hospital, and UT Southwestern Medical Center, aims to use the most cutting-edge CRISPR tools to create treatments for Rett syndrome and three other rare neurological diseases: Freidreich’s ataxia, Huntington’s disease, and spinal muscular atrophy. A new grant from the National Institute of Neurological Disorders and Stroke provides $22.8 million over a five-year period to support this work.

“We work with a lot of patients, families, and communities, and see a tremendous unmet need,” says Lutz. “My interest in rare diseases really stems from the application of these gene-based medicines. They have incredible potential for the four diseases we’ve selected, each of which results from mutations in a single gene.”

Lutz first became aware of Rett syndrome about 15 years ago, when the demand for Rett model mice began to increase. To learn more about Rett syndrome, she attended a conference hosted by the Rett Syndrome Research Trust (RSRT), where she saw up close the struggle — and strength — of Rett families and became inspired by RSRT’s advocacy.

“Thirteen or 14 years ago, I found myself at a Rett syndrome meeting to learn more about the mouse models, and that's where I met Monica Coenraads. She was a pioneer in terms of patient-driven research. I could use so many adjectives of admiration and respect and tenacity to describe Monica. In the end, I just wanted to be able to help as much as we possibly could at JAX.”

Currently, RSRT is funding a program at JAX to create more Rett mouse models with the intent to aid biopharmaceutical companies with their Rett syndrome genetic medicine development programs.

The new grant aims to use the latest CRISPR-based genome-editing tools to correct disease-causing mutations in the central nervous system of individuals with these rare neurological diseases. While different academic and industry labs are working towards genomic treatments for Rett syndrome, they are up against a tough challenge. In girls and women with Rett syndrome, one copy of the gene MECP2 has a mutation in a critical region, leaving the individual with too little functional MECP2 protein. But adding it back, for example in the form of a gene therapy, needs to take into account that too much MECP2 is also harmful. Taysha Gene Therapies and Neurogene are addressing this challenge by incorporating biological feedback loops meant to regulate MECP2 protein levels.

But this “Goldilocks” problem of getting just the right amount of MECP2 — the amount that is naturally present when a person has two healthy copies of the gene — makes gene correction through genome editing an ideal approach for curing Rett syndrome.

“There's a gene dosage effect,” says Lutz. “Too much is a bad thing and too little is a bad thing. How do you solve this ‘Goldilocks’ problem? A lot of people have tried to make gene therapies where the amount of protein is self-regulated. Gene editing really takes that problem out of the equation.”

In the new project, researchers plan to use the latest CRISPR-based tools, called base editing and prime editing. Liu developed both base and prime editing as safer alternatives to conventional CRISPR. Conventional CRISPR-based editing changes DNA by first causing a break in the DNA double helix at a specific location. As the cell repairs the break, it creates the opportunity to rewrite sections of DNA near the break site.

Base and prime editing use CRISPR’s special ability to target specific locations in the genome but can make DNA changes without breaking both strands. In these methods, the CRISPR enzyme keeps its remarkable ability to target a specific location in the genome but is modified so that it only cuts one DNA strand. In base editing, another enzyme comes along for the ride and can change a single DNA base at the specified site. In prime editing, a “donor” genome sequence is provided along with the CRISPR proteins. The cell uses this donor sequence to copy over the same genomic region in the cell, resulting in swapping in the new sequence.

While conventional CRISPR has shown remarkable success in some clinical trials and the first CRISPR-based therapeutic is expected to get FDA approval this year, breaking the DNA double helix carries a risk of causing DNA rearrangements and other unwanted, and potentially harmful, changes at the break site. Because base and prime editing only cut one strand of DNA, instead of both, the risks of unwanted changes are reduced.

The research team will begin by creating base-editing treatments meant to correct some of the most common disease-causing mutations in individuals with Rett. They also have plans to create a prime-editing treatment that, because it could swap in a long sequence of DNA covering the “genomic hotspot” where disease-causing mutations are usually found, could be used to treat individuals with multiple different mutations.

The Liu lab has taken advantage of RSRT’s bioreposity and has received a number of patient fibroblasts and stem cell lines needed for research in neurons.

“Programmable nucleases excel at disrupting genes, but most disease-causing mutations require precise gene correction rather than gene disruption to best benefit patients,” says Liu. “Base editors use the targeting mechanism of programmable DNA-binding proteins like Cas9, but instead of cutting the DNA, they use enzymes to directly convert one target DNA base to another, thereby enabling precise gene correction of four of the most common types of point mutations that cause genetic diseases, without requiring cutting the DNA double helix. To enable precise gene correction beyond the changes that can be made by base editing, we developed prime editing. The hallmark of prime editing is remarkable versatility. Prime editing can mediate substitutions, deletions, or insertions of up to hundreds of base pairs in human cells without requiring double-stranded breaks or donor DNA templates."

The ultimate aim is to advance at least one candidate therapy for one of the four diseases through a successful investigational new drug (IND) application with the FDA within five years. With a successful IND, the treatment could begin to be tested in clinical trials. Research will continue on all disease aims, even if they don’t meet the five-year deadline.

"RSRT really invested a lot in animal models over the years, and we think it’s paid off for the entire Rett research community. This new grant is very ambitious, but we are going to hit it as hard as we can in terms of timelines.”

“David and I have have many projects together, and it's really complimentary in terms of strengths,” says Lutz. “He’s brilliant in terms of the editing strategies and developing editing technologies. And we have all of these mouse models, readouts and resources at JAX to move quickly. RSRT really invested a lot in animal models over the years, and we think it’s paid off for the entire Rett research community. This new grant is very ambitious, but we are going to hit it as hard as we can in terms of timelines.”

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