Researchers corrected mutations underlying cystic fibrosis (CF) in a three-dimensional (3D) cell model of the disorder, using a new form of gene editing.
This work serves as a proof-of-principle for the technique — called prime editing, seen as an improvement on the CRISPR/Cas9 gene editing tool — and raises the possibility of a future cure, according to the scientists.
The study, “Evaluating CRISPR-based prime editing for cancer modeling and CFTR repair in organoids,” was published in the journal Life Science Alliance.
Gene editing has emerged as a way to introduce highly targeted changes in DNA sequences to treat genetic disorders. Changes include inserting or deleting segments — even entire genes — and swapping disease-causing mutations embedded within a gene with their healthy counterparts.
One such gene is the cystic fibrosis transmembrane conductance regulator (CFTR), which encodes a protein of the same name that is essential for regulating the exchange of salts and water between cells and their environment. Mutations affecting the ability of the CFTR protein to work as it should lead to CF.
Most gene editing is performed using CRISPR/Cas9 (simply CRISPR for short). While highly effective, CRISPR also causes a number of off-target effects, which raises concerns over its use as a therapy.
Researchers at the Hubrecht Institute, in the Netherlands, working with improved version of CRISPR called prime editing, tested its potential once further optimized to “correct” CFTR mutations in human intestinal organoids — 3D “mini-organs” that better approximate the organ found in humans than do the more traditional, flat layer of cells in a Petri dish.
Although CF predominantly affects the lungs, CFTR also regulates fluid transport into the lumen, the cavity within hollow organs such as the intestines. This, the investigators argued, makes intestinal organoids a suitable model of the disorder.
Applying the technique to the CFTR gene in these organoids, the researchers observed that those treated by prime editing adopted the same features as healthy organoids, in response to a compound called forskolin, which stimulates fluid transport into the organoid lumen via CFTR.
“We applied prime editing to the mutations, after which the treated organoids demonstrated the same response as the healthy organoids: they became swollen. That provided us with proof that our technique worked and replaced the mutated DNA,” Maarten Geurts, a PhD candidate and the study’s first author, said in a press release.
The team found that prime editing had variable efficacy depending on organoid tissue type, but was safer than the more established CRISPR method, not causing genome-wide off-target changes.
“In our study, prime editing proves to be a safer technique than the conventional CRISPR/Cas9. It can build in a new piece of DNA without causing damage elsewhere in the DNA. That makes the technique promising for application in patients,” Geurts added.
This work brings the possibility of treating or even curing CF, and other diseases, through gene editing one step closer to clinical use, Geurts said. Work remains, however, to adapt prime editing for safe use in humans.
“Prime editing is a versatile tool that can be used for disease modeling and clinical repair of most types of disease-causing mutations in human adult stem cells,” the researchers wrote. “Yet, it will require further improvement to allow widespread use as a technique for mutational modeling and for gene repair.”