Now, researchers at the Broad Institute of MIT and Harvard and the University of Iowa have developed a gene-editing approach that efficiently corrects the most common mutation that causes cystic fibrosis, found in 85 percent of patients. With further development, it could pave the way for treatments that are administered only once and have fewer side effects.

The new method, published today in Nature Biomedical Engineering, precisely and durably corrects the mutation in human lung cells, restoring cell function to levels similar to that of Trikafta. The approach is based on a technique called prime editing, which can make insertions, deletions, and substitutions up to hundreds of base pairs long in the genome with few unwanted byproducts. Prime editing was developed in 2019 by the lab of David Liu, who is the Richard Merkin Professor and director of the Merkin Institute of Transformative Technologies in Healthcare at the Broad, as well as a professor at Harvard University and a Howard Hughes Medical Institute investigator.

“We are hopeful that the use of prime editing to correct the predominant cause of cystic fibrosis might lead to a one-time, permanent treatment for this serious disease,” said Liu, the senior author on the study. “Developing a strategy to efficiently correct this challenging mutation also provided a blueprint for optimizing prime editing to precisely correct other mutations that cause devastating disorders.”

Postdoctoral researcher Alex Sousa and graduate student Colin Hemez, both from Liu’s lab, were first authors on the study.

Gene repair

Cystic fibrosis is caused by mutations in the CFTR gene that impair ion channels in the cell membrane that pump chloride out of cells. There are more than 2,000 known variants of the CFTR gene, 700 of which cause disease. The most common is a three base-pair CTT deletion that causes the ion channel protein to misfold and degrade.

Correcting the CTT deletion in CFTR has long been the goal of gene-editing therapies by labs including Liu’s, but most attempts have not been efficient enough to confer a therapeutic benefit, or use approaches such as CRISPR/Cas9 nuclease editing that generate double-stranded breaks in DNA, which can generate unwanted changes in the target gene and other locations in the genome.

Prime editing, a more flexible and controlled kind of gene editing that does not require double-stranded breaks, could help address this limitation. To more efficiently correct the CFTR mutation, Liu’s team combined six different enhancements to the technology. These included improving the prime editing guide RNAs that program prime editor proteins to find their target and to make the desired edit, as well as modifying the prime editor protein itself and other changes that make the target site more accessible. In combination, these refinements corrected about 60 percent of the CTT deletions in human lung cells and about 25 percent in cells taken directly from patient lungs and grown in a dish, an increase from previous methods that corrected less than 1 percent of the mutation in cells. The new approach also generated 3.5 times fewer unwanted insertions and deletions per edit than previous methods that use the Cas9 nuclease enzyme.

Next, researchers will need to develop ways to package and deliver the prime editing machinery to the airways in mice and ultimately humans. The team is hopeful that recent developments such as lipid nanoparticles that reach the lungs in mice may help expedite translation of this approach.