KCa3.1 potentiation stimulates Cl secretion in F508del and G551D CFTR-corrected primary human bronchial epithelial cells.

We previously identified potentiators of KCa3.1 (5,6-dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one; DCEBIO) that stimulate Cl secretion across human bronchial epithelial cells (HBEs) expressing wild-type (WT) cystic fibrosis transmembrane conductance regulator (CFTR). However, these compounds failed to stimulate Cl secretion in F508del CFTR HBEs. Drug discovery efforts identified CFTR potentiators (VX-770) and correctors (VX-445, VX-661) for cystic fibrosis (CF) disease-causing mutations, including F508del and G551D. Herein, we evaluated the effect of KCa3.1 potentiation on Cl equivalent current (I) across primary HBEs expressing WT, F508del, and G551D CFTR. Transepithelial impedance analysis was used to obtain estimates of apical (R) and basolateral membrane (BLM; R) resistances. In WT CFTR HBEs, DCEBIO stimulated I, which was increased by forskolin. Similarly, forskolin stimulated I, and this was increased by DCEBIO. The KCa3.1 blocker, TRAM-34 inhibited I. DCEBIO decreased R, whereas TRAM-34 increased R, consistent with BLM localization of KCa3.1. Following correction of F508del CFTR with VX-445 + VX-661, DCEBIO failed to stimulate I, although the subsequent addition of forskolin + VX-770 increased I. Importantly, following stimulation of I with forskolin + VX-770, DCEBIO induced a further significant increase in I. As above, DCEBIO reduced R, whereas TRAM-34 increased R, consistent with BLM localized KCa3.1. Finally, we assessed KCa3.1 potentiation on I in G551D/F508del CFTR HBEs in the absence or presence of VX-445 + VX-661. In both cases, DCEBIO failed to stimulate I. However, following stimulation with forskolin + VX-770, DCEBIO nearly doubled I. Our results demonstrate that following correction/potentiation of F508del and G551D CFTR, potentiation of KCa3.1 increases the Cl secretory response, suggesting this class of compounds may represent a novel means of further increasing Cl secretion across CF airway.