PT  - JOURNAL ARTICLE
AU  - Wen-Ying Lin
AU  - Yoshiro Sohma
AU  - Tzyh-Chang Hwang
TI  - Synergistic Potentiation of CFTR gating by two chemically distinct potentiators, Ivacaftor (VX-770) and NPPB.
AID  - 10.1124/mol.116.104570
DP  - 2016 Jan 01
TA  - Molecular Pharmacology
PG  - mol.116.104570
4099  - http://molpharm.aspetjournals.org/content/early/2016/07/13/mol.116.104570.short
4100  - http://molpharm.aspetjournals.org/content/early/2016/07/13/mol.116.104570.full
AB  - Cystic Fibrosis (CF) is caused by loss-of-function mutations of the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene encoding a phosphorylation-activated, but ATP-gated chloride channel. Previous studies suggested that VX-770 (ivacaftor), a CFTR potentiator now used in clinics, increases the open probability (Po) of CFTR by shifting the gating conformational changes to favor the open channel configuration. Lately, another CFTR potentiator NPPB was reported to enhance CFTR activity through a modus operandi that exploits the ATP hydrolysis-driven, non-equilibrium gating mechanism unique to CFTR. Surprisingly however, NPPB increased the activity of non-hydrolytic G551D-CFTR, the third most common disease-associated mutation. Here, the mechanism of NPPB is investigated by assessing NPPB's interaction with well-studied VX-770. Interestingly, once G551D-CFTR was maximally potentiated by VX-770, NPPB further increases its activity. However, quantitative analysis of this drug-drug interaction suggests that this pharmacological synergism is not due to independent actions of NPPB and VX-770 on CFTR gating; instead, our data support a dependent mechanism involving two distinct binding sites. This latter idea is further supported by the observation that the locked-open time of a hydrolysis-deficient mutant K1250A was shortened by NPPB, but prolonged by VX-770. In addition, the effectiveness of NPPB, but not of VX-770, was greatly diminished in a mutant whose second nucleotide-binding domain was completely removed. Interpreting these results under the framework of current understanding of CFTR gating not only reveals insights into the mechanism of action for different CFTR potentiators, but also brings us one step forward to a more complete schematic for CFTR gating.