Gene Editing Strategy Could Treat Hundreds of Inherited Diseases More Effectively

Gene editing technologies developed in recent years offer the promise of repairing genetic mutations that cause disease. But putting these therapies to use typically requires creating a separate treatment tailored to each disease-causing mutation—an approach that becomes impractical given that a single gene can harbor dozens or even hundreds of different mutations.

Now, scientists at Gladstone Institutes and UC San Francisco (UCSF) have developed a new strategy that removes the mutated copy of a gene—or critical portions of it—from cells while leaving the healthy copy intact. The approach could be 20 to 40 times more efficient than personalizing individual therapies for each mutation, with just a few variations of the therapy treating nearly all patients with an inherited disorder.

In a new study published in Molecular Therapy, the researchers used their approach, called “haplotype editing,” to restore function to human nerve cells from patients with Charcot-Marie-Tooth (CMT) disease, a progressive neurological disorder that affects over 3 million people.

“With this approach, we don’t have to create an entirely new, customized gene editing therapy for every patient,” says Gladstone Senior Investigator Bruce Conklin, MD, co-senior author of the new study. “We’ve not only shown how effective it is for Charcot-Marie-Tooth disease, but also how it can be leveraged to treat hundreds or thousands of other genetic conditions.”

From One Mutation to Many

People have two copies of most genes—one inherited from their mother, the other from their father. Many genetic diseases happen when someone inherits one defective copy of a gene. Even if the other copy is healthy, the mutated version can disrupt the normal gene’s ability to do its job, like one bad ingredient spoiling an entire recipe.

The advent of gene editing technologies like CRISPR, which can make precise cuts and changes to DNA, has led many researchers to try developing therapies that fix this kind of disease-causing mutation. But for many diseases—including CMT—scaling this up to patients has been a colossal task.

That’s because CMT includes approximately 160 subtypes caused by mutations in more than 130 genes. One form of the disease studied by Gladstone scientists, called CMT2E, is caused by over 50 different mutations in a single gene. So, helping patients would involve developing and testing dozens of therapies to fix each of the different mutations that can cause the disease.

“If we had to create a separate gene editing therapy for every single mutation, we’d need 26 different treatments just to help half of Charcot-Marie-Tooth 2E patients,” says Luke Judge, MD, PhD, professor of pediatrics at UCSF Benioff Children’s Hospital and co-senior author of the study. “We realized we needed a different approach.”

Conklin and Judge wanted to take advantage of the fact that people can survive with only one copy of many genes, much like you can get by with one kidney. But to remove the mutated copy of a gene and leave behind the healthy one, the researchers had to find a way to differentiate the two. They homed in on common genetic variants called single nucleotide polymorphisms (SNPs) that are scattered throughout the human genome.

Like genetic “zip codes,” these variants are inherited in blocks of DNA called haplotypes. Because people typically inherit different haplotypes from each parent, these SNPs can serve as unique markers that distinguish the mutated chromosome from the healthy one.

The scientists used CRISPR to design pairs of molecular scissors that cut at these SNPs on either side of the mutated gene causing CMT2E, deleting the DNA between them—including the entire gene or critical portions of it. Because each pair of scissors targets specific SNP sequences, the approach selectively removes only the chromosome copy carrying the disease mutation, leaving the healthy copy intact.

Treating Charcot-Marie-Tooth and Beyond

The team tested their approach in stem cells from patients with two different CMT2E-causing mutations. By removing the DNA between sets of common SNPs, the researchers successfully deleted the mutant gene in both patient lines. And when these edited stem cells were matured into motor neurons—the type of brain cell impacted by the disease—the scientists observed dramatic improvements compared to neurons that carry the mutations.

Next, using genetic data from the 1000 Genomes Project, the researchers showed that just four haplotype-targeted therapies could potentially treat most CMT2E patients.

“We’re already applying our approach to other forms of Charcot-Marie-Tooth disease, inherited heart conditions, ALS, and retinal diseases.”

Luke Judge, MD, PhD

Each therapy targets a different pair of SNPs flanking the gene. Because different people have different SNP patterns, these four therapies together can cover 75 percent of the patient population—with each patient matched to whichever therapy targets SNPs near their mutated gene but not their healthy one.

“This efficiency comes from our shared evolutionary history,” Conklin explains. “Humans are a relatively young species—only about 1.5 million years old—so many SNPs from early human populations remain common today. We’re essentially taking advantage of that gift from human evolution.”

Patients receiving a haplotype editing therapy would need to have their DNA sequenced to determine which set of SNPs are near each of their copies of the mutated gene. But before the therapy can be tested in humans, it needs to be more thoroughly studied in mouse models of CMT2E—work that Conklin and Judge have already begun.

The group is also thinking ahead to how haplotype editing can be used to remove other disease genes that have many mutations, which has so far made them difficult to treat with standard gene editing techniques.

“We expect the conceptual framework we’ve established to be broadly applicable,” says Judge, who is also a visiting scientist at Gladstone. “We’re already applying our approach to other forms of Charcot-Marie-Tooth disease, inherited heart conditions, ALS, and retinal diseases.”