Novel Correctors and Potentiators Enhance Functional Rescue of CFTR Nonsense Mutation Translational Readthrough.

Premature termination codons (PTCs) in cystic fibrosis transmembrane conductance regulator (CFTR) result in nonfunctional CFTR protein and are the proximate cause of ~11% of CF-causing alleles for which no treatments exist. CFTR corrector, lumacaftor and the potentiator ivacaftor improve CFTR function with terminal PTC mutations, and enhance the effect of readthrough agents. Novel correctors GLPG2222 (C1), GLPG3221 (C2), and potentiator GLPG1837 compare favorably to lumacaftor and ivacaftor in vitro. Here, we evaluated the effect of correctors C1a and C2a (derivatives of C1 and C2) and GLPG1837 alone or in combination with the readthrough compound G418, on CFTR function using heterologous FRT, genetically engineered 16HBE14o-, and primary human cells with PTC mutations. In FRT lines pretreated with G418, GLPG1837 elicited dose-dependent increases in CFTR activity that exceeded ivacaftor in FRT-W1282X and FRT-R1162X. A three-mechanism strategy consisting of G418, GLPG1837, and two correctors (C1a+C2a) yielded the greatest functional improvements in FRT and 16HBE14o- PTC variants, noting correction and potentiation without readthrough was sufficient to stimulate CFTR activity for W1282X. GLPG1837+C1a+C2a restored substantial function in G542X/F508del human bronchial epithelial cells and even more for W1282X/F508del, largely due to the corrector/potentiator effect with no additional benefit from G418. In G542X/R553X or R1162X/R1162X organoids, enhanced forskolin induced swelling (FIS) was observed with G418+GLPG1837+C1a+C2a although GLPG1837+C1a+C2a alone was sufficient to improve FIS in W1282X/W1282X organoids. Combination of CFTR correctors, potentiators and readthrough compounds augments the functional repair of CFTR nonsense mutations, indicating the potential for novel correctors and potentiators to restore function to truncated W1282X CFTR.