Partial rescue of F508del‐cystic fibrosis transmembrane conductance regulator channel gating with modest improvement of protein processing, but not stability, by a dual‐acting small molecule

Liu, Jia; Bihler, Hermann; Farinha, Carlos M.; Awatade, Nikhil T; Romão, Ana Lúcia; Mercadante, Dayna; Cheng, Yi; Musisi, Isaac; Jantarajit, Walailak; Wang, Yiting; Cai, Zhiwei; Amaral, Margarida D.; Mense, Martin; Sheppard, David N.

doi:10.1111/bph.14141

Cited by 16 publications

(17 citation statements)

References 73 publications

(162 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…uncorrected F508del-CFTR blue star symbol vs. F508del/R1070W-CFTR red square symbol both compared to blue dotted line). Both results are consistent with patch-clamp records indicating a F508del/R1070W-CFTR PO comparable to that of WT-CFTR [48], but a much lower PO for temperature-corrected F508del-CFTR [45,46,48]. Figure 6 plots GCFTR as a function of  for the rare-mutation panel, giving an immediate representation of how severe a defect each mutation causes in biogenesis (distance from WT-CFTR on the x-axis) and/or in gating and permeation properties (vertical displacement from blue dotted line, which assumes ion-channel properties of baseline-activated WT-CFTR).…”

Section: Relationship Between Cftr Ion Channel Function and Membrane supporting

confidence: 90%

Fluorescence assay for simultaneous quantification of CFTR ion-channel function and plasma membrane proximity

Prins

Langron

Hastings

et al. 2020

Journal of Biological Chemistry

View full text Add to dashboard Cite

The cystic fibrosis transmembrane conductance regulator, CFTR, is a plasma membrane anion channel which plays a key role in controlling transepithelial fluid movement. Excessive activation results in intestinal fluid loss during secretory diarrhoeas, while CFTR mutations underlie cystic fibrosis (CF). Anion permeability depends both on how well CFTR channels work (permeation/gating) and on how many are present at the membrane. Recently, treatments with two drug classes targeting CFTR – one boosting ion-channel function (potentiators), the other increasing plasma membrane density (correctors) – have provided significant health benefits to CF patients. Here we present an image-based fluorescence assay that can rapidly and simultaneously estimate both CFTR ion-channel function and the protein’s proximity to the membrane. We monitor F508del-CFTR, the most common CF-causing variant, and confirm rescue by low temperature, CFTR-targeting drugs and second-site revertant mutation R1070W. In addition, we characterize a panel of 62 CF-causing mutations. Our measurements correlate well with published data (electrophysiology and biochemistry), further confirming validity of the assay. Finally, we profile effects of acute treatment with approved potentiator drug VX-770 on the rare-mutation panel. Mapping the potentiation profile on CFTR structures raises mechanistic hypotheses on drug action, suggesting that VX-770 might allow an open-channel conformation with an alternative arrangement of domain interfaces. The assay is a valuable tool for investigation of CFTR molecular mechanisms, allowing accurate inferences on gating/permeation. In addition, by providing a two-dimensional characterization of the CFTR protein, it could better inform development of single-drug and precision therapies addressing the root cause of CF disease.

show abstract

Section: Relationship Between Cftr Ion Channel Function and Membrane supporting

confidence: 90%

Fluorescence assay for simultaneous quantification of CFTR ion-channel function and plasma membrane proximity

Prins

Langron

Hastings

et al. 2020

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…4A, C; P , 0.05 for P o for both mutants). Because G509A/V510G mutations strikingly rescued DF508-CFTR processing and gating, the data suggest that they are revertant mutations similar to those previously described (e.g., [46][47][48][49][50][51]. These data further suggest that displacement of the G509 residue in the H3-H4 loop may be a pivotal defect that underlies functional abnormalities of DF508-CFTR.…”

Section: Ibi Prolongation Of Df508and Dy512-cftr Is Attenuated By Loosupporting

confidence: 73%

A defective flexible loop contributes to the processing and gating defects of the predominant cystic fibrosis‐causing mutation

et al. 2019

View full text Add to dashboard Cite

People with the genetic disease cystic fibrosis (CF) often carry a deletion mutation ∆F508 on the gene encoding the CF transmembrane conductance regulator (CFTR) Cl− channel. This mutation greatly reduces the CFTR maturation process and slows the channel opening rate. Here, we investigate whether residues near F508 contribute to these defects in ∆F508‐CFTR. Most deletion mutations, but not alanine substitutions, of individual residues from positions 503 to 513 impaired CFTR maturation. Interestingly, only protein processing of ∆Y512‐CFTR, like that of ∆F508‐CFTR, was greatly improved by low‐temperature culture at 27°C or small‐molecule corrector C18. The 2 mutant Cl− channels were equally slow to open, suggesting that they may share common structural flaws. Studies on the H3‐H4 loop that links residues F508 and Y512 demonstrate that G509A/V510G mutations, moving G5091 position backward in the loop, markedly enhanced ∆F508‐CFTR maturation and opening rate while promoting protein stability and persistence of the H3 helix in ∆F508 nucleotide‐binding domain 1. Moreover, V510A/S511A mutations noticeably increased ∆Y512‐CFTR maturation at 27°C and its opening rate. Thus, loop abnormalities may contribute to ∆F508‐ and ∆Y512‐CFTR defects. Importantly, correcting defects from G509 displacement in ∆F508‐CFTR may offer a new avenue for drug discovery and CF treatments.—Chen, X., Zhu, S., Zhenin, M., Xu, W., Bose, S. J., Wong, M. P.‐F., Leung, G. P. H., Senderowitz, H., Chen, J.‐H. A defective flexible loop contributes to the processing and gating defects of the predominant cystic fibrosis‐causing mutation. FASEB J. 33, 5126–5142 (2019). http://www.fasebj.org

show abstract

“…It has also recently been reported that long-term ivacaftor treatment enhances the turnover of rescued F508del-CFTR at the cell surface, and induces a significant decrease of cell surface stability ( Cholon et al, 2014 ; Veit et al, 2014 ), thus suggesting that identification of alternative dual-acting CFTR modulators may be a valuable perspective. Dual-acting small-molecules have been recently identified, which independently promote F508del-CFTR trafficking to the plasma membrane and boost its channel activity ( Pedemonte et al, 2011 ; Phuan et al, 2011 ; Liu et al, 2018 ).…”

Section: Discussionmentioning

confidence: 99%

Molecular Mechanism of Action of Trimethylangelicin Derivatives as CFTR Modulators

et al. 2018

View full text Add to dashboard Cite

The psoralen-related compound, 4,6,4′-trimethylangelicin (TMA) potentiates the cAMP/PKA-dependent activation of WT-CFTR and rescues F508del-CFTR-dependent chloride secretion in both primary and secondary airway cells homozygous for the F508del mutation. We recently demonstrated that TMA, like lumacaftor (VX-809), stabilizes the first membrane-spanning domain (MSD1) and enhances the interface between NBD1 and ICL4 (MSD2). TMA also demonstrated anti-inflammatory properties, via reduction of IL-8 expression, thus making TMA a promising agent for treatment of cystic fibrosis. Unfortunately, TMA was also found to display potential phototoxicity and mutagenicity, despite the fact that photo-reactivity is absent when the compound is not directly irradiated with UVA light. Due to concerns about these toxic effects, new TMA analogs, characterized by identical or better activity profiles and minimized or reduced side effects, were synthesized by modifying specific structural features on the TMA scaffold, thus generating compounds with no mutagenicity and phototoxicity. Among these compounds, we found TMA analogs which maintained the potentiation activity of CFTR in FRT-YFP-G551D cells. Nanomolar concentrations of these analogs significantly rescued F508del CFTR-dependent chloride efflux in FRT-YFP-F508del, HEK-293 and CF bronchial epithelial cells. We then investigated the ability of TMA analogs to enhance the stable expression of varying CFTR truncation mutants in HEK-293 cells, with the aim of studying the mechanism of their corrector activity. Not surprisingly, MSD1 was the smallest domain stabilized by TMA analogs, as previously observed for TMA. Moreover, we found that TMA analogs were not effective on F508del-CFTR protein which was already stabilized by a second-site mutation at the NBD1-ICL4 interface. Altogether, our findings demonstrate that these TMA analogs mediate correction by modifying MSD1 and indirectly stabilizing the interface between NBD1 and CL4.

show abstract

Partial rescue of F508del‐cystic fibrosis transmembrane conductance regulator channel gating with modest improvement of protein processing, but not stability, by a dual‐acting small molecule

Cited by 16 publications

References 73 publications

Fluorescence assay for simultaneous quantification of CFTR ion-channel function and plasma membrane proximity

Fluorescence assay for simultaneous quantification of CFTR ion-channel function and plasma membrane proximity

A defective flexible loop contributes to the processing and gating defects of the predominant cystic fibrosis‐causing mutation

Molecular Mechanism of Action of Trimethylangelicin Derivatives as CFTR Modulators

Contact Info

Product

Resources

About