A gene editing tool safely inserted large DNA segments in mice, pointing to a potential new route for genetic medicines.
The technique uses circular, single-stranded DNA rather than the usual double-stranded form of DNA, the molecule that carries genetic instructions.
The approach is known as INSTALL, short for “integration through nucleus-synthesised template addition of large lengths”, and was developed by a team led by Connor Tou and Benjamin Kleinstiver at Mass General Brigham.
The technique has proved capable of inserting DNA into the genomes of mice and lab-grown human cells.
Kleinstiver said the team is now working to use it in new medicines for liver diseases, blood disorders and metabolic conditions.
Kiran Musunuru, co-founder of Verve Therapeutics and a scientist at the Children’s Hospital of Philadelphia and the University of Pennsylvania, who was not involved in the study, described the work as addressing one of the major challenges in controlled DNA insertion.
Musunuru said: “This is beautiful work that addresses in an elegant way one of the big challenges with controlled DNA insertion into the genome.”
The key issue INSTALL is designed to address is the immune response to large chunks of double-stranded DNA, which are not normally found floating freely inside cells.
DNA in the cytoplasm, the area outside the nucleus, “usually means something is wrong”, such as a viral infection, Kleinstiver said.
When the cell detects DNA outside the nucleus, it launches an immune response that causes “downstream toxicity.”
To get around this, Tou and Kleinstiver took inspiration from certain bacteria and viruses that can naturally insert circular, single-stranded DNA into double-stranded genomes.
Tou said: “This was an exciting realisation. We then wondered whether these mechanisms could be recapitulated in human cells.”
The researchers worked with genome engineering specialist Full Circles Therapeutics to shape large chunks of DNA into circles and found they could slip through the cell undetected.
Because gene editing enzymes need double-stranded DNA to latch onto, they attached small snippets of double-stranded nucleotides, the building blocks of DNA, which were too small to trigger the immune system.
Kleinstiver said the technique allows large DNA strands to be inserted into the genome more precisely than is possible with viral vectors, the modified viruses often used to deliver gene therapies.
INSTALL instead uses a lipid nanoparticle, a tiny fat-based carrier, to deliver its cargo into cells.
Kleinstiver said: “The potential and upside of these tools is really high, because you could develop one editor that could treat lots of or all patients.”
Musunuru said techniques such as INSTALL could be especially useful for diseases that arise when large sections of a gene are missing.
He said: “That’s where gene replacement, as forecast by the work in this paper, will have a critical role to play.”
While noting that INSTALL “has a long way to go before it might be useful for human therapeutics”, Musunuru described it as “a promising step in the right direction.”
The gene editing field received a regulatory boost at the end of February with the official unveiling of the US Food and Drug Administration’s plausible mechanism pathway.
This is a new route for bespoke therapies designed for individual patients to gain agency approval.
Kleinstiver’s team, which provided the gene editing enzyme used in baby KJ’s custom treatment, is pursuing the new pathway to develop personalised gene editing therapies for patients with multisystemic smooth muscle dysfunction syndrome, an ultra-rare genetic disease.
Kleinstiver said: “We’ve developed a therapy for the most common mutation, but leveraging some of these principles might allow us to go and pursue therapies for some of the other mutations as well.”
He said that if INSTALL or a similar tool ever gained regulatory approval, it could make parts of the plausible mechanism pathway less necessary because treatments would not need to be tailored to each individual in the same way.
“We wouldn’t need 100 tools to treat 100 mutations,” Kleinstiver said. “We would have one tool to treat those 100 mutations.”
However, both Kleinstiver and Musunuru said that point remains a long way off.
Musunuru added: “There isn’t going to be a one-size-fits-all solution to all genetic diseases.”
