ABCMdb

PMID: 16150931

Pedemonte N, Diena T, Caci E, Nieddu E, Mazzei M, Ravazzolo R, Zegarra-Moran O, Galietta LJ
Antihypertensive 1,4-dihydropyridines as correctors of the cystic fibrosis transmembrane conductance regulator channel gating defect caused by cystic fibrosis mutations.
Mol Pharmacol. 2005 Dec;68(6):1736-46. Epub 2005 Sep 8., [PubMed]

Sentences

No. Mutations Sentence Comment

2 ABCC7 p.Gly551Asp Other mutations, like G551D, cause only a gating defect. Login to comment 8 ABCC7 p.Gly551Asp DHPs were also effective on the G551D-CFTR mutant by inducing a 16- to 45-fold increase of the CFTR Cl- currents. Login to comment 25 ABCC7 p.Gly551Asp ABCC7 p.Gly551Asp 6 Copyright (c) 2005 The American Society for Pharmacology and Experimental Therapeutics 15149/3064372 Mol Pharmacol 68:1736-1746, 2005 Printed in U.S.A. 1736 example of pure class III mutation is instead represented by G551D (glycine-to-aspartic acid change at position 551), which causes a severe impairment in CFTR channel activity (Gregory et al., 1991; Logan et al., 1994; Zegarra-Moran et al., 2002). Login to comment 27 ABCC7 p.Gly551Asp For example, genistein and other flavonoids, at high micromolar concentrations, strongly stimulate the activity of G551D and of ⌬F508 (Hwang et al., 1997; Illek et al., 1999; Zegarra-Moran et al., 2002). Login to comment 34 ABCC7 p.Gly551Asp We have found that the 1,4-dihydropyridines (DHPs) used to treat hypertension are able to stimulate the activity of ⌬F508- and G551D-CFTR mutants. Login to comment 39 ABCC7 p.Gly551Asp Fischer rat thyroid (FRT) cells, stably transfected with ⌬F508- or G551D-CFTR (Galietta et al., 2001b; Zegarra-Moran et al., 2002), were retransfected with the YFP-H148Q/I152L fluorescent protein, which allows optimal sensitivity for the detection of mutant CFTR activity (Galietta et al., 2001a). Login to comment 168 ABCC7 p.Gly551Asp DHPs were also tested as possible activators of the G551D mutant. Login to comment 170 ABCC7 p.Gly551Asp With forskolin alone, no increase of halide transport was detected in G551D cells with the fluorescence assay. Login to comment 187 ABCC7 p.Gly551Asp Dose-response relationships (Fig. 4B and Table 2) revealed that G551D activation required DHP concentrations higher than ⌬F508. Login to comment 188 ABCC7 p.Gly551Asp The potency order for the G551D mutant was the following: felodipine ϭ nicardipine ϭ nitrendipine Ͼ nifedipine ϭ nimodipine ϭ isradipine Ͼ niguldipine. Login to comment 189 ABCC7 p.Gly551Asp In Ussing chamber experiments, very small currents were activated after forskolin stimulation of G551D-CFTR cells. Login to comment 192 ABCC7 p.Gly551Asp It is interesting that the maximal response to DHPs like felodipine or isradipine was at least 2-fold larger than that to genistein, a known activator of the G551D mutant (Illek et al., 1999; Zegarra-Moran et al., 2002). Login to comment 204 ABCC7 p.Gly551Asp The S-(ϩ)-niguldipine had a modest activity also on G551D-CFTR, whereas the other stereoisomer was completely inactive. Login to comment 206 ABCC7 p.Gly551Asp Both compounds were effective on ⌬F508- and G551D-CFTR, the apparent affinity being lower for the latter mutant (Fig. 5). Login to comment 207 ABCC7 p.Gly551Asp BayK-8644 showed a modest but significant stereoselectivity for ⌬F508 (2-fold difference in Ka; p Ͻ 0.05) and no stereoselectivity at all for G551D. Login to comment 221 ABCC7 p.Gly551Asp The two different isradipine enantiomers showed different activity on the G551D mutant (Fig. 5). Login to comment 223 ABCC7 p.Gly551Asp Membrane patches from FRT cells expressing G551D-CFTR were excised in the inside-out configuration. Login to comment 227 ABCC7 p.Gly551Asp Under these conditions, G551D-CFTR channels had a very low activity (Po ϭ 0.021 Ϯ 0.012; n ϭ 3), despite the presence of the catalytic subunit of the protein kinase A to induce maximal phosphorylation (Fig. 6A). Login to comment 232 ABCC7 p.Gly551Asp Nasal epithelial cells from a patient carrying the G551D mutation (Fig. 7A) were largely unresponsive to maximal forskolin (20 ␮M), with an effect of only 0.05 to 0.2 ␮A/cm2 , in agreement with previous observations (Zegarra-Moran et al., 2002). Login to comment 246 ABCC7 p.Gly551Asp Activation of G551D-CFTR by DHPs. Login to comment 247 ABCC7 p.Gly551Asp A, representative traces from microplate reader experiments showing G551D-CFTR activity with saline alone, or forskolin (20 ␮M) with and without genistein (100 ␮M) or DHPs (20 ␮M). Login to comment 249 ABCC7 p.Gly551Asp C, representative Ussing chamber traces showing G551D-CFTR currents upon stimulation with forskolin (20 ␮M) and the indicated concentrations of genistein or DHPs. Login to comment 274 ABCC7 p.Gly551Asp An evidence in favor of this conclusion is represented by the differences of DHP potency for ⌬F508 and G551D mutants. Login to comment 276 ABCC7 p.Gly551Asp Conversely, for G551D, the Ka values of these three DHPs were comparable and significantly higher than that for ⌬F508. Login to comment 287 ABCC7 p.Gly551Asp The decreased channel activity of ⌬F508 and G551D mutants is considered to be the result of an intrinsic defect in CFTR protein function. Login to comment 289 ABCC7 p.Gly551Asp To this respect, it is interesting to TABLE 2 Activation properties of 1,4-dihydropyridines measured in FRT cells expressing G551D-CFTR Values for Vmax and Ka were obtained from experiments of the type shown in Fig. 4B and are the mean Ϯ S.E.M. of 5 to 10 experiments. Login to comment 323 ABCC7 p.Gly551Asp One classic example is G551D, which is a class III mutation characterized by a severe channel-open- Fig. 5. Login to comment 326 ABCC7 p.Gly551Asp Data (mean Ϯ S.E.M. of 5-10 experiments) were obtained from ⌬F508 (left) and G551D (right) using the YFP assay. Login to comment 330 ABCC7 p.Gly551Asp A, traces showing continuous recordings of currents from and inside-out membrane patch excised from a FRT cell expressing G551D-CFTR. Login to comment 335 ABCC7 p.Gly551Asp B, traces from the same experiment showing activation of G551D channels by the addition of 100 ␮M felodipine to the bath solution. Login to comment 338 ABCC7 p.Gly551Asp We have shown previously that genistein is an effective activator of the G551D mutant, with a level of functional correction close to 20% in FRT and nasal epithelial cells (Zegarra-Moran et al., 2002). Login to comment 339 ABCC7 p.Gly551Asp ABCC7 p.Gly551Asp Our present results show that the best DHP, felodipine, also stimulated Cl-secretion in G551D airway epithelial cells up to a level close to 20% of that measured in non-CF cells. Login to comment 342 ABCC7 p.Gly551Asp The different sensitivity of ⌬F508 and G551D mutants suggests that the DHP binding site resides in the NBDs. Login to comment