Gene repair becomes more reliable – NRC

Researchers from Harvard University and the Massachusetts Institute of Technology (USA) have found a way to send packages containing the latest Gene editingTechnology in rat cells in one piece. In this way, they were able to fix the genetic defect that caused the eyes of congenitally blind mice to respond “overly” to light stimuli. The Americans published their results on Monday in Natural biotechnology.

The new technology is called Main editing It is an alternative to what is called Crispr-cas9. This genetic “cut and paste” technique was discovered in 2012 and won a Nobel Prize in 2020. Using Crispr-cas9, you can make very precise cuts in DNA and then repair, add or turn off genes.

Despite the great promises of this technology, its application on the ground has been difficult until now. Just last November The first medical treatment based on Crispr-cas9 has been approved, in this case of the notorious sickle cell disease. One reason for this difficult application is the low success rate. The breaks made by the CRISPR protein in DNA They are often repaired very quickly by the body Errors occur relatively often.

More efficient variables

“Newer variants of gene editing are more efficient and reliable in this regard,” says Eva van Rooij, professor of molecular cardiology at UMC Utrecht. She was not involved in the new US study, but is working with these new gene-editing variants: Main editing And what is related to it Edit rule. Her group is investigating how it could be used as a treatment for hereditary heart disease. “Prime editing and base editing are two different types of Crispr-cas9, but they don't make a complete break in double-stranded DNA,” she says. “With prime editing, you can only correct one base, whereas with base editing you can also make more complex and extensive base changes.”

Because you're not doing a double break, there's less chance of errors, Van Rooij explains. Additionally, you can work more specifically with these two technologies than with a “regular” Crispr-cas9 machine. With both techniques, you can insert a package using the cut-and-paste technique into the host cell. You can do this by using a specially prepared virus, often an adenovirus, as a vehicle. But you can also do this using synthetic molecules called Virus-like particles. They are very similar to viruses and can therefore easily penetrate a cell – but you can tailor them in detail to your needs.

“This is what the Americans have done now,” says Van Rooij. “These are the same researchers who were the first in 2019 nature Published around the main edit. And In 2022 they showed That you can use these virus-like particles to edit the base. They have now adapted their molecules so that they are also suitable for primary editing. It's a challenge, because to do that you have to introduce a much larger particle into the cell.

This particle not only contains a protein that recognizes the specific defective DNA, but it also contains a protein that contains the information about the correct and desired DNA sequence. “The fact that there is a particle that can now effectively enter a cell is a great step forward.”

The Americans tested their approach in the laboratory using neurons from mice and human embryonic kidney cells. In this first cell line, prime editing was 80 times more effective than previously developed compounds; In a human cell line it is up to 170 times more effective.

The researchers then tested the new technique on young mice with a genetic eye defect. They had a mutation in the gene that codes for a protein in the retina that is important for transmitting electrical signals. Without this protein, young mice become visually impaired from three weeks old. In humans, such an anomaly leads to Leber's disease.

The researchers injected “charged” virus-like particles just below the retinas in five-week-old mice with this eye defect. After treatment, the researchers noted that the mice's retinas responded “much better” to light stimulation.

In another group of mice, the researchers did not inject its molecules into the eye, but into the cerebrospinal fluid. They did this to see if their technology would also have effects outside the eye. It turned out not to be the case: the particles penetrated – at will – the retinal cells just to do their work there.