Researchers from MIT and Harvard have crafted a new tool to silence genes without editing their sequence and are using it to develop a treatment for prion disease, a fatal neurodegenerative condition caused by misfolded proteins.
Led by Sonia Vallabh of the Broad Institute and Jonathan Weissman of MIT’s Whitehead Institute, a team of scientists engineered a technology they call CHARM, which uses several tricks to recruit and then turn on a protein that can shut off DNA expression. CHARM is also small enough to fit into a viral delivery package called an AAV — one of the few ways to deliver a genetic medicine to the brain.
The researchers published their findings on Thursday in Science. CHARM represents a new method for epigenetic editing, a buzzy field in which hundreds of millions of dollars have been invested in hopes of developing one-time treatments that do not require changing the DNA code itself.
“CHARM is really a quantum leap over the previous epigenetic editors,” said Kiran Musunuru, a University of Pennsylvania cardiologist and gene editing researcher who was not involved in the study. “This really feels like something new that has much more translational potential.”
In an accompanying editorial, Musunuru said CHARM’s potential impact is comparable to next-generation gene editing techniques like base and prime editing.
In mice with prion disease, the treatment cut prion protein expression by about 80%, which is more than what’s been seen with previous approaches to reduce protein expression, according to the study authors.
But the scientists noted it will be years before CHARM is ready to enter human studies. For prion disease, the researchers plan to advance the technology using the NIH grant that funded the study, with a goal of having it ready for human trials by 2027.
Outside of prion disease, CHARM has been licensed by Chroma Medicine, a high-profile epigenetic editing company founded by gene editing luminaries, including Weissman. Chroma has been working on an earlier iteration of epigenetic editing called CRISPRoff that Weissman co-invented.
No epigenetic editing therapy has reached human studies yet. Chroma is one of several companies, including Tune Therapeutics and Epic Bio, trying to advance epigenetic editing to the clinic.
From CRISPRoff to CHARM
Wife-and-husband duo Vallabh and Eric Minikel lead the prion disease program at the Broad Institute. The two retrained to become scientists together after Vallabh’s mother passed away from genetic prion disease, and they learned that Vallabh also carries the mutation that puts her at very high risk of developing the fatal disease. At the Broad, they’ve been at the forefront of finding new potential treatments for prion disease — a very personal mission for them.
In May 2022, they reached out to Weissman, who had published CRISPRoff the previous year, asking whether it could potentially be used for prion disease. CRISPRoff tacks small chemical tags onto the DNA, which makes it inaccessible to the machinery that reads DNA and eventually turns the code into proteins.
But there were problems with CRISPRoff: It was too big to fit into an AAV, which is key for delivery to the brain, and it was toxic to cells.
To make a smaller epigenetic editor, the researchers turned to a much older, but more compact, gene editing tool called zinc finger proteins. But they still had to deal with toxicities from the chemical tagging method, known as methylation.
CRISPRoff borrows only the active part of a DNA-tagging enzyme that is typically inactive in our bodies. But the active portion by itself can be toxic when it’s over-expressed in cells.
“How can we take the methylation machinery that’s already in the cell?” Minikel asked.
Rather than using the enzyme itself, CHARM takes advantage of part of a specific protein that can recruit it. That’s linked to a histone tail, a gene regulator that can then activate the methylation enzyme.
“What ended up working was this sort of Frankenstein,” Minikel said of an epigenetic editing system. It used zinc fingers linked to the protein chunk plus the histone tail. (The researchers noted that instead of zinc finger proteins, the system can also use Cas9.)
When the epigenetic editor was delivered into mice, the researchers found it cut levels of both transcripts and proteins across the brain.
Self-silencing
Amusingly enough, CHARM ended up being too small. Weissman said that when they took the construct to Ben Deverman, the study group’s AAV expert, he asked them to add some filler so that random DNA wouldn’t get into the viral delivery package.
“Because it’s small, you now have room for lots of extra bells and whistles,” Weissman said, noting up to three epigenetic editors could fit into one AAV. So the scientists went ahead and addressed one more problem.
The epigenetic editors only need to be expressed for just enough time to turn off DNA expression. It’s meant to be a one-and-done approach. Any longer and they pose the risk of making unwelcome changes or could potentially trigger an unwanted immune reaction. The researchers developed an epigenetic editor that could turn off the other ones. It’s essentially a self-silencing program for after the intended DNA silencing changes are made.
In mice, they found that the prion gene silencing effect was durable to 13 weeks when the self-silencing CHARM was also used.
What’s next?
The development of a new epigenetic editor coincides with another promising finding in delivery. Deverman’s lab developed an AAV that can get through the blood-brain barrier in humans, which was described in Science last month.
The researchers are optimizing CHARM for the gene of the human prion protein and are working with Deverman to put that into one of the new AAVs. “The misfolding event [for prion disease] can be seeded almost anywhere in the brain, so we need to be able to deliver our CHARM therapeutic to as many neurons as possible,” Deverman said.
Chroma Medicine, which has the license for CHARM outside prion disease, is not disclosing where it plans to apply the technology. CEO Catherine Stehman-Breen pointed to categories where AAVs have shown potential in delivering genetic treatments (where “size matters”) as potential paths forward, including neurological, muscle and lung diseases.