Click-based synthesis of triazolobithiazole ?F508-CFTR correctors for cystic fibrosis.
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ABSTRACT:
SUBMITTER: Donald MB
PROVIDER: S-EPMC3419814 | biostudies-literature | 2012 Sep
REPOSITORIES: biostudies-literature
Publications
Click-based synthesis of triazolobithiazole ΔF508-CFTR correctors for cystic fibrosis.
Donald Michael B MB Rodriguez Kevin X KX Shay Hannah H Phuan Puay-Wah PW Verkman A S AS Kurth Mark J MJ
Bioorganic & medicinal chemistry 20120706 17
Copper catalyzed azide-alkyne cycloaddition (CuAAC) chemistry is reported for the construction of previously unknown 5-(1H-1,2,3-triazol-1-yl)-4,5'-bithiazoles from 2-bromo-1-(thiazol-5-yl)ethanones. These novel triazolobithiazoles are shown to have cystic fibrosis (CF) corrector activity and, compared to the benchmark bithiazole CF corrector corr-4a, improved logP values (4.5 vs 5.96). ...[more]
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Project description:Deletion of phenylalanine 508 of the cystic fibrosis transmembrane conductance regulator (∆F508 CFTR) is the major cause of cystic fibrosis, one of the most common inherited childhood diseases. The mutated CFTR anion channel is not fully glycosylated and shows minimal activity in bronchial epithelial cells of patients with cystic fibrosis. Low temperature or inhibition of histone deacetylases can partly rescue ∆F508 CFTR cellular processing defects and function. A favourable change of ∆F508 CFTR protein-protein interactions was proposed as a mechanism of rescue; however, CFTR interactome dynamics during temperature shift and inhibition of histone deacetylases are unknown. Here we report the first comprehensive analysis of the CFTR and ∆F508 CFTR interactome and its dynamics during temperature shift and inhibition of histone deacetylases. By using a novel deep proteomic analysis method, we identify 638 individual high-confidence CFTR interactors and discover a ∆F508 deletion-specific interactome, which is extensively remodelled upon rescue. Detailed analysis of the interactome remodelling identifies key novel interactors, whose loss promote ∆F508 CFTR channel function in primary cystic fibrosis epithelia or which are critical for CFTR biogenesis. Our results demonstrate that global remodelling of ∆F508 CFTR interactions is crucial for rescue, and provide comprehensive insight into the molecular disease mechanisms of cystic fibrosis caused by deletion of F508.
| S-EPMC4826614 | biostudies-literature
Project description: Not available
| S-EPMC6513216 | biostudies-literature
Project description:Conformationally constrained bithiazoles were previously found to have improved efficacy over nonconstrained bithiazoles for correction of defective cellular processing of the ?F508 mutant cystic fibrosis transmembrane conductance regulator (CFTR) protein. In this study, two sets of constrained bithiazoles were designed, synthesized, and tested in vitro using ?F508-CFTR expressing epithelial cells. The SAR data demonstrated that modulating the constraining ring size between 7- versus 8-membered in these constrained bithiazole correctors did not significantly enhance their potency (IC50), but strongly affected maximum efficacy (Vmax), with constrained bithiazoles 9e and 10c increasing Vmax by 1.5-fold compared to benchmark bithiazole corr4a. The data suggest that the 7- and 8-membered constrained ring bithiazoles are similar in their ability to accommodate the requisite geometric constraints during protein binding.
| S-EPMC4136667 | biostudies-literature
Project description:Cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR). Newly developed "correctors" such as lumacaftor (VX-809) that improve CFTR maturation and trafficking and "potentiators" such as ivacaftor (VX-770) that enhance channel activity may provide important advances in CF therapy. Although VX-770 has demonstrated substantial clinical efficacy in the small subset of patients with a mutation (G551D) that affects only channel activity, a single compound is not sufficient to treat patients with the more common CFTR mutation, ?F508. Thus, patients with ?F508 will likely require treatment with both correctors and potentiators to achieve clinical benefit. However, whereas the effectiveness of acute treatment with this drug combination has been demonstrated in vitro, the impact of chronic therapy has not been established. In studies of human primary airway epithelial cells, we found that both acute and chronic treatment with VX-770 improved CFTR function in cells with the G551D mutation, consistent with clinical studies. In contrast, chronic VX-770 administration caused a dose-dependent reversal of VX-809-mediated CFTR correction in ?F508 homozygous cultures. This result reflected the destabilization of corrected ?F508 CFTR by VX-770, markedly increasing its turnover rate. Chronic VX-770 treatment also reduced mature wild-type CFTR levels and function. These findings demonstrate that chronic treatment with CFTR potentiators and correctors may have unexpected effects that cannot be predicted from short-term studies. Combining these drugs to maximize rescue of ?F508 CFTR may require changes in dosing and/or development of new potentiator compounds that do not interfere with CFTR stability.
| S-EPMC4272825 | biostudies-other
Project description:Cystic fibrosis (CF) is an autosomal recessive disease caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. The most common CF-associated mutation is ?F508, which deletes a phenylalanine in position 508. In vitro studies indicate that the resultant protein, CFTR-?F508, is misprocessed, although the in vivo consequences of this mutation remain uncertain. To better understand the effects of the ?F508 mutation in vivo, we produced CFTR(?F508/?F508) pigs. Our biochemical, immunocytochemical, and electrophysiological data on CFTR-?F508 in newborn pigs paralleled in vitro predictions. They also indicated that CFTR(?F508/?F508) airway epithelia retain a small residual CFTR conductance, with maximal stimulation producing ~6% of wild-type function. Cyclic adenosine 3',5'-monophosphate (cAMP) agonists were less potent at stimulating current in CFTR(?)(F508/)(?)(F508) epithelia, suggesting that quantitative tests of maximal anion current may overestimate transport under physiological conditions. Despite residual CFTR function, four older CFTR(?F508/?F508) pigs developed lung disease similar to human CF. These results suggest that this limited CFTR activity is insufficient to prevent lung or gastrointestinal disease in CF pigs. These data also suggest that studies of recombinant CFTR-?F508 misprocessing predict in vivo behavior, which validates its use in biochemical and drug discovery experiments. These findings help elucidate the molecular pathogenesis of the common CF mutation and will guide strategies for developing new therapeutics.
| S-EPMC3119077 | biostudies-literature
Project description:The deletion of Phe508 (?F508) in the first nucleotide binding domain (NBD1) of CFTR is the most common mutation associated with cystic fibrosis. The ?F508-CFTR mutant is recognized as improperly folded and targeted for proteasomal degradation. Based on molecular dynamics simulation results, we hypothesized that interaction between ?F508-NBD1 and housekeeping proteins prevents ?F508-CFTR delivery to the plasma membrane. Based on this assumption we applied structure-based virtual screening to identify new low-molecular-weight compounds that should bind to ?F508-NBD1 and act as protein-protein interaction inhibitors. Using different functional assays for CFTR activity, we demonstrated that in silico-selected compounds induced functional expression of ?F508-CFTR in transfected HeLa cells, human bronchial CF cells in primary culture, and in the nasal epithelium of homozygous ?F508-CFTR mice. The proposed compounds disrupt keratin8-?F508-CFTR interaction in ?F508-CFTR HeLa cells. Structural analysis of ?F508-NBD1 in the presence of these compounds suggests their binding to NBD1. We conclude that our strategy leads to the discovery of new compounds that are among the most potent correctors of ?F508-CFTR trafficking defect known to date.
| S-EPMC3799575 | biostudies-literature
Project description:Cystic fibrosis (CF) is a lethal genetic disease caused by the loss or dysfunction of the CF transmembrane conductance regulator (CFTR) channel. F508del is the most prevalent mutation of the CFTR gene and encodes a protein defective in folding and processing. VX-809 has been reported to facilitate the folding and trafficking of F508del-CFTR and augment its channel function. The mechanism of action of VX-809 has been poorly understood. In this study, we sought to answer a fundamental question underlying the mechanism of VX-809: does it bind CFTR directly in order to exert its action? We synthesized two VX-809 derivatives, ALK-809 and SUL-809, that possess an alkyne group and retain the rescue capacity of VX-809. By using Cu(I) -catalyzed click chemistry, we provide evidence that the VX-809 derivatives bind CFTR directly in vitro and in cells. Our findings will contribute to the elucidation of the mechanism of action of CFTR correctors and the design of more potent therapeutics to combat CF.
| S-EPMC4672727 | biostudies-literature
Project description:Cystic fibrosis (CF) is a genetic disease associated with the defective function of the cystic fibrosis transmembrane conductance regulator (CFTR) protein that causes obstructive disease and chronic bacterial infections in airway epithelia. The most prevalent CF-causing mutation, the deletion of phenylalanine at position 508 (F508del), leads to CFTR misfolding, trafficking defects and premature degradation. A number of correctors that are able to partially rescue F508del-CFTR processing defects have been identified. Clinical trials have demonstrated that, unfortunately, mono-therapy with the best correctors identified to date does not ameliorate lung function or sweat chloride concentration in homozygous F508del patients. Understanding the mechanisms exerted by currently available correctors to increase mutant F508del-CFTR expression is essential for the development of new CF-therapeutics. We investigated the activity of correctors on the mutant F508del and wild type (WT) CFTR to identify the protein domains whose expression is mostly affected by the action of correctors, and we investigated their mechanisms of action. We found that the four correctors under study, lumacaftor (VX809), the quinazoline derivative VX325, the bithiazole compound corr4a, and the new molecule tezacaftor (VX661), do not influence either the total expression or the maturation of the WT-CFTR transiently expressed in human embryonic kidney 293 (HEK293) cells. Contrarily, they significantly enhance the expression and the maturation of the full length F508del molecule. Three out of four correctors, VX809, VX661 and VX325, seem to specifically improve the expression and the maturation of the mutant CFTR N-half (M1N1, residues 1-633). By contrast, the CFTR C-half (M2N2, residues 837-1480) appears to be the region mainly affected by corr4a. VX809 was shown to stabilize both the WT- and F508del-CFTR N-half isoforms, while VX661 and VX325 demonstrated the ability to enhance the stability only of the mutant F508del polypeptide.
| S-EPMC6862496 | biostudies-literature
Project description:The ?F508 mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) gene is the most common cause of cystic fibrosis. The mutation disrupts biosynthetic processing, reduces channel opening rate, and decreases protein lifetime. In contrast to human CFTR (hCFTR)-?F508, mouse CFTR-?F508 is partially processed to the cell surface, although it exhibits a functional defect similar to hCFTR-?F508. To explore ?F508 abnormalities, we generated human-mouse chimeric channels. Substituting mouse nucleotide-binding domain-1 (mNBD1) into hCFTR partially rescued the ?F508-induced maturation defect, and substituting mouse membrane-spanning domain-2 or its intracellular loops (ICLs) into hCFTR prevented further ?F508-induced gating defects. The protective effect of the mouse ICLs was reverted by inserting mouse NBDs. Our results indicate that the ?F508 mutation affects maturation and gating via distinct regions of the protein; maturation of CFTR-?F508 depends on NBD1, and the ?F508-induced gating defect depends on the interaction between the membrane-spanning domain-2 ICLs and the NBDs. These appear to be distinct processes, because none of the chimeras repaired both defects. This distinction was exemplified by the I539T mutation, which improved CFTR-?F508 processing but worsened the gating defect. Our results, together with previous studies, suggest that many different NBD1 modifications improve CFTR-?F508 maturation and that the effect of modifications can be additive. Thus, it might be possible to enhance processing by targeting several different regions of the domain or by targeting a network of CFTR-associated proteins. Because no one modification corrected both maturation and gating, perhaps more than a single agent will be required to correct all CFTR-?F508 defects.
| S-EPMC3271874 | biostudies-other
Project description:Cystic fibrosis (CF) is a lethal autosomal recessive disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. The common ΔF508-CFTR mutation results in protein misfolding and proteasomal degradation. If ΔF508-CFTR trafficks to the cell surface, its anion channel function may be partially restored. Several in vitro strategies can partially correct ΔF508-CFTR trafficking and function, including low-temperature, small molecules, overexpression of miR-138, or knockdown of SIN3A. The challenge remains to translate such interventions into therapies and to understand their mechanisms. One approach for connecting such interventions to small molecule therapies that has previously succeeded for CF and other diseases is via mRNA expression profiling and iterative searches of small molecules with similar expression signatures. Here, we query the Library of Integrated Network-based Cellular Signatures using transcriptomic signatures from previously generated CF expression data, including RNAi- and low temperature-based rescue signatures. This LINCS in silico screen prioritized 135 small molecules that mimicked our rescue interventions based on their genomewide transcriptional perturbations. Functional screens of these small molecules identified eight compounds that partially restored ΔF508-CFTR function, as assessed by cAMP-activated chloride conductance. Of these, XL147 rescued ΔF508-CFTR function in primary CF airway epithelia, while also showing cooperativity when administered with C18. Improved CF corrector therapies are needed and this integrative drug prioritization approach offers a novel method to both identify small molecules that may rescue ΔF508-CFTR function and identify gene networks underlying such rescue.
| S-EPMC8846631 | biostudies-literature