Novel Cystic Fibrosis Ferret Model Enables Visualization of CFTR Expression Cells and Genetic CFTR Reactivation

Cystic fibrosis (CF) is caused by mutations in the <i>cystic fibrosis transmembrane conductance regulator</i> (<i>CFTR</i>). While gene therapy holds promise as a cure, the cell-type-specific heterogeneity of <i>CFTR</i> expression in the lung presents significant challenges. Current CF ferret models closely replicate the human disease phenotype but have limitations in studying functional complementation through cell-type-specific CFTR restoration. To address this, we developed a new transgenic ferret line, <i>CFTR</i>^{int1-eGFP(lsl)}, in which a Cre-recombinase (Cre)-excisable enhanced fluorescent protein (eGFP) reporter cassette is knocked in (KI) to intron 1 of the <i>CFTR</i> locus. Breeding this reporter line with <i>CFTR</i>^G551D CF ferret resulted in a novel CF model, <i>CFTR</i>^{int1-eGFP(lsl)/G551D}, with disease onset manageable via the administration of CFTR modulator VX770. In this study, we confirmed two key properties of the <i>CFTR</i>^{int1-eGFP(lsl)/G551D} CF ferrets: (1) cell-type-specific expression of the CFTR(N-24)-eGFP fusion protein, driven by the intrinsic <i>CFTR</i> promoter, in polarized epithelial cultures and selected tissues, and (2) functional reversion of the KI allele via Cre-mediated excision of the reporter cassette. This model provides a valuable tool for studying the effects of targeted CFTR reactivation in a cell-type-specific manner, which is crucial for enhancing our understanding of CFTR's roles in modulating airway clearance and innate immunity, and for identifying relevant cellular targets for CF gene therapy.