Partial rescue of F508del-cystic fibrosis transmembrane conductance regulator channel gating with modest improvement of protein processing, but not stability, by a dual-acting small molecule.

Abstract

BACKGROUND AND PURPOSE:

Rescue of F508del-cystic fibrosis (CF) transmembrane conductance regulator (CFTR), the most common CF mutation, requires small molecules that overcome protein processing, stability and channel gating defects. Here, we investigate F508del-CFTR rescue by CFFT-004, a small molecule designed to independently correct protein processing and channel gating defects.

EXPERIMENTAL APPROACH:

Using CFTR-expressing recombinant cells and CF patient-derived bronchial epithelial cells, we studied CFTR expression by Western blotting and channel gating and stability with the patch-clamp and Ussing chamber techniques.

KEY RESULTS:

Chronic treatment with CFFT-004 improved modestly F508del-CFTR processing, but not its plasma membrane stability. By contrast, CFFT-004 rescued F508del-CFTR channel gating better than C18, an analogue of the clinically used CFTR corrector lumacaftor. Subsequent acute addition of CFFT-004, but not C18, potentiated F508del-CFTR channel gating. However, CFFT-004 was without effect on A561E-CFTR, a CF mutation with a comparable mechanism of CFTR dysfunction as F508del-CFTR. To investigate the mechanism of action of CFFT-004, we used F508del-CFTR revertant mutations. Potentiation by CFFT-004 was unaffected by revertant mutations, but correction was abolished by the revertant mutation G550E. These data suggest that correction, but not potentiation, by CFFT-004 might involve nucleotide-binding domain 1 of CFTR.

CONCLUSIONS AND IMPLICATIONS:

CFFT-004 is a dual-acting small molecule with independent corrector and potentiator activities that partially rescues F508del-CFTR in recombinant cells and native airway epithelia. The limited efficacy and potency of CFFT-004 suggests that combinations of small molecules targeting different defects in F508del-CFTR might be a more effective therapeutic strategy than a single agent.

• Authors: Jia Liu, Hermann Bihler, Carlos M Farinha, Nikhil T Awatade, Ana M Romão, Dayna Mercadante, Yi Cheng, Isaac Musisi, Walailak Jantarajit, Yiting Wang, Zhiwei Cai, Margarida D Amaral, Martin Mense, David N Sheppard
• Journal: British journal of pharmacology (Br J Pharmacol) Vol. 175 Issue 7 Pg. 1017-1038 (04 2018) ISSN: 1476-5381 [Electronic] England
• PMID: 29318594 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: © 2018 The British Pharmacological Society.

Chemical References

• CFTR protein, human
• Cystic Fibrosis Transmembrane Conductance Regulator

Topics

• Animals
• Cell Line
• Cell Membrane (physiology)
• Cells, Cultured
• Cricetinae
• Cystic Fibrosis Transmembrane Conductance Regulator (physiology)
• Epithelial Cells (physiology)
• Humans
• Ion Channel Gating
• Mice
• Protein Processing, Post-Translational
• Protein Stability