Cystic fibrosis (CF) is a common hereditary lethal disease caused by loss-of-function mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Various CFTR modulators have been approved, but these highly effective modulators (HEMTs) are only suitable for patients with at least one F508del allele or other responsive CFTR mutations, leaving many patients with (ultra-)rare CFTR mutations without treatment options. With advances in scientific research, gene therapy has emerged as a new opportunity for these mutations. Notably, the recently developed CRISPR-based system known as prime editing (PE) opens a new era for the treatment of genetic diseases. Prime editing can "rewrite" and correct mutations on patients' chromosomes in situ, providing new opportunities for treating monogenic diseases like CF.
In this study, researchers designed a prime editing strategy targeting the L227R and N1303K mutations in the CFTR gene using CRISPR-Cas9 technology. They constructed stable cell models expressing 3HA-L227R-CFTR and 3HA-N1303K-CFTR in HEK293T cells to evaluate the effects of prime editing. Using a developed DETECTOR machine learning algorithm to ensure efficiency and accuracy, the researchers further assessed gene and functional correction. The results showed editing efficiency as high as 25%, and the corrected CFTR protein exhibited significant restoration in glycosylation, localization, and ion channel function. These results were also validated in primary cell model experiments. Additionally, through whole-genome assessment analysis, no significant off-target editing events were found, demonstrating the high fidelity of prime editing, and the clinical relevance and safety assessments of the study were also validated. Overall, this study demonstrates the potential of prime editing technology in correcting CFTR gene mutations and restoring CFTR protein function, providing new ideas and methods for cystic fibrosis treatment.