CFTR Potentiator Research Articles

CFTR chloride channel as a molecular target of anthraquinone compounds in herbal laxatives

To clarify whether CFTR is a molecular target of intestinal fluid secretion caused by the anthraquinone compounds from laxative herbal plants. A cell-based fluorescent assay to measure I(-) influx through CFTR chloride channel. A short-circuit current assay to measure transcellular Cl(-) current across single layer FRT cells and freshly isolated colon mucosa. A closed loop experiment to measure colon fluid secretion in vivo. Anthraquinone compounds rhein, aloe-emodin and 1,8-dihydroxyanthraquinone (DHAN) stimulated I(-) influx through CFTR chloride channel in a dose-dependent manner in the presence of physiological concentration of cAMP. In the short-circuit current assay, the three compound enhanced Cl(-) currents in epithelia formed by CFTR-expressing FRT cells with EC(50) values of 73 ± 1.4, 56 ± 1.7, and 50 ± 0.5 μmol/L, respectively, and Rhein also enhanced Cl(-) current in freshly isolated rat colonic mucosa with a similar potency. These effects were completely reversed by the CFTR selective blocker CFTR(inh)-172. In in vivo closed loop experiments, rhein 2 mmol/L stimulated colonic fluid accumulation that was largely blocked by CFTR(inh)-172. The anthraquinone compounds did not elevate cAMP level in cultured FRT cells and rat colonic mucosa, suggesting a direct effect on CFTR activity. Natural anthraquinone compounds in vegetable laxative drugs are CFTR potentiators that stimulated colonic chloride and fluid secretion. Thus CFTR chloride channel is a molecular target of vegetable laxative drugs.

Preventing Thermal Inactivation of ΔF508-CFTR Channel Activity by NBD1 Suppressor Mutations and by Constitutively Active Intracellular Loop Mutations

ΔF508 is a temperature sensitive CFTR mutant that causes cystic fibrosis by disrupting CFTR processing, trafficking and channel gating. To address the question of thermal stability of ΔF508-CFTR channel gating, we examined WT- and ΔF508-CFTR channel activity in excised patches at varying temperature (23 to 36°C) with or without exposure to CFTR correctors and potentiators. In addition, constructs that combined the ΔF508 mutation with intracellular loop mutations that promote ATP-independent channel gating and with NBD1 suppressor mutations that enhance ΔF508-CFTR biosynthetic maturation were examined under the same conditions.

The investigational CFTR potentiator, VX-770, potentiated multiple CFTR forms in vitro

The investigational CFTR potentiator, VX-770, potentiated multiple CFTR forms in vitro

How can the CFTR potentiator CBIQ activate the calcium activated K + channel KCa3.1?

How can the CFTR potentiator CBIQ activate the calcium activated K + channel KCa3.1?

Optimization of the NBD1 Site Improves the Function of G551D-CFTR Channels

CFTR chloride channels comprise two nucleotide binding domains (NBD1 and NBD2). Mutations (for example, G551D) that impair CFTR function result in the lethal genetic disease cystic fibrosis (CF). It's established that the opening and closing of CFTR are mainly controlled by ATP binding and hydrolysis respectively in NBD2 while ATP binding at NBD1 can modulate the stability of the open state. Using a non-hydrolytic ligand MgPPi, we locked open CFTR channels with mutations at NBD1. We found that two W401 mutations significantly increased the lock-open time (W401F: 72±3s; W401Y: 51±6s; WT: 27±2s). These gain-of-function mutations are unexpected since the W401-equivalent residue Y1219 in NBD2 can be effectively replaced by tryptophan but not phenylalanine. As the ATP molecule bound in NBD1 may interact with NBD2's signature sequence (LSHGH), we extended our study to this region. We found that reverting the histidine residue 1348 to the canonical glycine (H1348G) similarly stabilized the lock-open state (τ=67±8s). Once W401F, H1348G, or W401F/H1348G mutations were incorporated into G551D channels which no longer respond to ATP, the mutant channels become ATP-responsive with basal activity increased by ∼10-fold for W401F/G551D-, ∼4-fold for H1348G/5G551D-, and ∼25-fold for W401F/H1348G/G551D-channels. The increased channel activity is mainly due to an increased open-time. The results shown here suggest that NBD1 and NBD2 may employee different chemical mechanisms in binding ATP and that NBD1 can be a potential molecular target for developing CFTR potentiators for CF-related mutants. The effects of different nucleotides (for example, GTP and UTP) on NBD1 and NBD2 will be studied to gain a better understanding of the chemical mechanism underlying nucleotide-NBD interaction.

Improvement in ion transport biomarkers and spirometry with the investigational CFTR potentiator VX-770 in subjects with cystic fibrosis and the G551D-CFTR mutation

Hydrogen is considered as the most potential energy carrier in the near future and can be produced from nuclear energy by several means. Lead-based reactor is one of the advance reactors in hydrogen production. In this paper, a novel feasible system of lead cooled reactor coupled with solid oxide electrolysis cells (SOEC) is presented, which is named China LEAd-based Reactor for Hydrogen production (CLEAR-H). By taking advantage of the chemically inert and high density of lead–bismuth eutectic (LBE), the concept of LBE-water direct-contact steam generator is proposed to supply high temperature (up to 600 °C) and low pressure (0.1–0.2 MPa) steam for SOEC stack without additional decompression devices. The results show that 47 tons hydrogen and 3000 tons oxygen-enriched air are obtained per day, and the thermal-to-hydrogen efficiency can reach about 39%.

Synthesis of 4-thiophen-2′-yl-1,4-dihydropyridines as potentiators of the CFTR chloride channel

The gating of the CFTR chloride channel is altered by a group of mutations that cause cystic fibrosis. This gating defect may be corrected by small molecules called potentiators. Some 1,4-dihydropyridine (DHP) derivatives, bearing a thiophen-2-yl and a furanyl ring at the 4-position of the nucleus, were prepared and tested as CFTR potentiators. In particular, we evaluated the ability of novel DHPs to enhance the activity of the rescued ΔF508-CFTR as measured with a functional assay based on the halide-sensitive yellow fluorescent protein. Most DHPs showed an effect comparable to or better than that of the reference compound genistein. The potency was instead significantly improved, with some compounds, such as 3g, 3h, 3n, 4a, 4b, and 4d, having a half effective concentration in the submicromolar range. CoMFA analysis gave helpful suggestions to improve the activity of DHPs.

Rescue of CF airway epithelial cell function in vitro by a CFTR potentiator, VX-770

Rescue of CF airway epithelial cell function in vitro by a CFTR potentiator, VX-770

Stimulation of salivary secretion in vivo by CFTR potentiators in Cftr+/+ and Cftr−/− mice

Background Physiologically, salivary secretion is controlled by cholinergic and adrenergic pathways but the role of ionic channels in this process is not yet clearly understood. In cystic fibrosis (CF), most exocrine glands failed to response to β-adrenergic agonists. Methods To determine the implication of CFTR in this process, we measured in vivo the salivary secretion of Cftr +/+ and Cftr −/− mice in the presence of 2 water-soluble benzo[ c]quinolizinium derivatives; MPB-07 a potentiator of CFTR Cl − channel and MPB-05 an inactive analogue. We also used genistein and its vehicle ethanol to confirm the implication of CFTR in salivary secretion. Results We showed that subcutaneous injection of MPB-07 in the mice cheek enhanced in a dose dependent manner the isoprenaline-induced salivary secretion in Cftr +/+ but not in Cftr −/− mice. By contrast, MPB-05 did not activate the salivary secretion in Cftr +/+ mice. The CFTR activator genistein (50 μM) significantly potentiated the secretory response of Cftr +/+ mice whereas its vehicle, ethanol, had no effect. Conclusions These results show for the first time in vivo pharmacological stimulation of salivary secretion by a water-soluble CFTR potentiator, MPB-07 and by the isoflavone, ethanol-soluble genistein and suggest that this chloride channel plays an important role in salivary gland physiology.

A quantitative description of the activation and inhibition of CFTR by potentiators: Genistein

The CFTR, encoded by the gene mutated in cystic fibrosis (CF) patients, is responsible for cAMP dependent chloride transport in epithelia. Substances that activate CFTR have been suggested as possible CF therapy. Most substances investigated so far exert a dual effect on the CFTR: low concentrations stimulate CFTR, whereas higher concentrations inhibit CFTR. Besides, the CFTR phosphorylation level determines the apparent affinity of the drug. We have studied the properties of genistein, the well known CFTR potentiator, by measuring apical membrane current on epithelia formed by cells stably transfected with CFTR and stimulated with different concentrations of CPTcAMP. We propose a quantitative model to describe the activatory and inhibitory effect of genistein, accounting also for the cAMP dependent activation.