Abstract
Most of the ~2100 CFTR variants so far reported are very rare and still uncharacterized regarding their cystic fibrosis (CF) disease liability. Since some may respond to currently approved modulators, characterizing their defect and response to these drugs is essential. Here we aimed characterizing the defect associated with four rare missense (likely Class II) CFTR variants and assess their rescue by corrector drugs. We produced CFBE cell lines stably expressing CFTR with W57G, R560S, H1079P and Q1100P, assessed their effect upon CFTR expression and maturation and their rescue by VX-661/VX-445 correctors. Results were validated by forskolin-induced swelling assay (FIS) using intestinal organoids from individuals bearing these variants. Finally, knock-down (KD) of genes previously shown to rescue F508del-CFTR was assessed on these mutants. Results show that all the variants preclude the production of mature CFTR, confirming them as Class II mutations. None of the variants responded to VX-661 but the combination rescued H1079P- and Q1100P-CFTR. The KD of factors that correct F508del-CFTR retention only marginally rescued R560S- and H1079P-CFTR. Overall, data evidence that Class II mutations induce distinct molecular defects that are neither rescued by the same corrector compounds nor recognized by the same cellular machinery, thus requiring personalized drug discovery initiatives.
Highlights
IntroductionCystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene [3,4], which encodes a cAMP-regulated chloride (Cl-) and bicarbonate (HCO3-) channel expressed at the apical membrane of epithelial cells [5], that regulates other channels and transporters [6]
Cystic fibrosis (CF) is the most common lethal genetic recessive disorder amongCaucasians [1], affecting 1:2500–6000 newborns, depending on the geographic region [2].CF is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene [3,4], which encodes a cAMP-regulated chloride (Cl-) and bicarbonate (HCO3-) channel expressed at the apical membrane of epithelial cells [5], that regulates other channels and transporters [6]
Identification of small molecules that rescue F508del-CFTR defect resulted in the approval of three corrector drugs currently available for individuals with CF: VX-809, VX-661 or VX-445, all used in combination with VX-770, a gating potentiator [11,12,13]
Summary
CF is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene [3,4], which encodes a cAMP-regulated chloride (Cl-) and bicarbonate (HCO3-) channel expressed at the apical membrane of epithelial cells [5], that regulates other channels and transporters [6]. The most common mutation—F508del—is present in 80% of individuals with CF worldwide [8,9,10]. This mutant causes CFTR misfolding leading to endoplasmic reticulum (ER) retention by the ER quality control (ERQC), premature degradation and failure to reach the plasma membrane (PM). All variants result in abnormal Cl- secretion by epithelial cells and, according to the impact in CFTR, they may result in “classical” or “atypical”
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