Fibrosis Transmembrane Conductance Regulator Protein Research Articles

The Application of Bicarbonate Recovers the Chemical-Physical Properties of Airway Surface Liquid in Cystic Fibrosis Epithelia Models.

Simple SummaryCystic fibrosis (CF) is a multi-organ disease that affects the epithelia and exocrine glands, particularly the lungs, but also the pancreas, liver, kidneys, and intestines. Respiratory disease is the most common cause of death. Many of the problems in respiratory disease can be attributed to the viscous nature of the mucus. Indeed, thick mucus in the airways leads to reduced mucociliary clearance, chronic bacterial infections and inflammation followed by destruction of the lung parenchyma. The composition of the periciliary fluid (ASL) is regulated by the transport of water and ions through different ion channels and transporters distributed in a non-symmetrical way to the two parts of the epithelium. Mucus is the first line of defense against inhaled particles and protects the epithelium by trapping and eliminating harmful substances through the blink of the cells that push them towards the nose. In patients with CF the malfunction of the mutated CFTR causes a reduction in bicarbonate secretion with consequent reduction in the expansion of mucins and the formation of thick mucus. In this work we have shown that, in vitro, the administration of a solution containing bicarbonate ion would act not only on the osmotic component of the mucus, but directly on the expansion of the mucins by acting on its viscoelastic properties.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. Deletion of phenylalanine at position 508, p.F508del, the most frequent mutation among CF patients, causes a folding and traffic defect, resulting in a dramatic reduction in the CFTR expression. To investigate whether the direct application of bicarbonate could modify the properties of the airway surface liquid (ASL), we measured the micro-viscosity, fluid transport and pH of human bronchial epithelial cells monolayers. We have demonstrated that the treatment of a CF-epithelia with an iso-osmotic solution containing bicarbonate is capable of reducing both, the ASL viscosity and the apical fluid re-absorption. We suggest the possibility of design a supportive treatment based on topical application of bicarbonate, or any other alkaline buffer.

New insights into structure and function of bis-phosphinic acid derivatives and implications for CFTR modulation

C407 is a compound that corrects the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein carrying the p.Phe508del (F508del) mutation. We investigated the corrector effect of c407 and its derivatives on F508del-CFTR protein. Molecular docking and dynamics simulations combined with site-directed mutagenesis suggested that c407 stabilizes the F508del-Nucleotide Binding Domain 1 (NBD1) during the co-translational folding process by occupying the position of the p.Phe1068 side chain located at the fourth intracellular loop (ICL4). After CFTR domains assembly, c407 occupies the position of the missing p.Phe508 side chain. C407 alone or in combination with the F508del-CFTR corrector VX-809, increased CFTR activity in cell lines but not in primary respiratory cells carrying the F508del mutation. A structure-based approach resulted in the synthesis of an extended c407 analog G1, designed to improve the interaction with ICL4. G1 significantly increased CFTR activity and response to VX-809 in primary nasal cells of F508del homozygous patients. Our data demonstrate that in-silico optimized c407 derivative G1 acts by a mechanism different from the reference VX-809 corrector and provide insights into its possible molecular mode of action. These results pave the way for novel strategies aiming to optimize the flawed ICL4–NBD1 interface.

Assisting PNA transport through cystic fibrosis human airway epithelia with biodegradable hybrid lipid-polymer nanoparticles

Cystic fibrosis (CF) is characterized by an airway obstruction caused by a thick mucus due to a malfunctioning Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. The sticky mucus restricts drugs in reaching target cells limiting the efficiency of treatments. The development of new approaches to enhance drug delivery to the lungs represents CF treatment's main challenge. In this work, we report the production and characterization of hybrid core–shell nanoparticles (hNPs) comprising a PLGA core and a dipalmitoylphosphatidylcholine (DPPC) shell engineered for inhalation. We loaded hNPs with a 7-mer peptide nucleic acid (PNA) previously considered for its ability to modulate the post-transcriptional regulation of the CFTR gene. We also investigated the in vitro release kinetics of hNPs and their efficacy in PNA delivery across the human epithelial airway barrier using an ex vivo model based on human primary nasal epithelial cells (HNEC) from CF patients. Confocal analyses and hNPs transport assay demonstrated the ability of hNPs to overcome the mucus barrier and release their PNA cargo within the cytoplasm, where it can exert its biological function.

Can Targeting Iron Help in Combating Chronic Pseudomonas Infection? A Systematic Review.

Cystic fibrosis is an autosomal recessive disorder caused by a mutation in genes for cystic fibrosis transmembrane conductance regulator (CFTR) protein. CFTR gene is responsible for the production of sweat, digestive fluids, and mucus, and any mutation in this would lead to the thickening of these secretions. Cystic fibrosis is a multi-organ disorder, but 80% of patients suffer from respiratory problems due to chronic infections most commonly caused by Pseudomonas aeruginosa (P. aeruginosa). Eradication of these infections has become a challenge as P. aeruginosa has developed resistance to multiple antibiotics. In several studies, iron has been shown to play an integral role in biofilm formation, which is the predominant resistance mechanism used by P. aeruginosa to combat antibiotics. The increased iron content in cystic fibrosis patients' sputum samples explains their increased susceptibility to Pseudomonas infections. Hence in this review article, we have used the research data available on therapeutic agents that target iron as an adjuvant treatment for chronic Pseudomonas infection. We systematically screened three databases using focused words and Medical Subject Headings (MeSH) terms for relevant articles. Further, we applied the inclusion and exclusion criteria and performed a thorough quality appraisal. Thirty shortlisted relevant studies were meticulously reviewed. In our opinion, novel therapeutic approaches targeting iron such as iron chelators, gallium, and cefiderocol have potent anti-biofilm properties. Future studies and clinical trials using these approaches in the management of chronic Pseudomonas infection might help in decreasing morbidity and mortality in patients with cystic fibrosis. Exploring these approaches might also help to combat other resistant organisms whose survival is dependent on iron.

Genetics of CFTR and male infertility.

Cystic fibrosis (CF) is a rare autosomal-recessive disorder manifested as multisystem organ dysfunction. The cystic fibrosis transmembrane conductance regulator (CFTR) protein functions as an ion transporter on the epithelium of exocrine glands, regulating secretion viscosity. The CFTR gene, encoded on chromosome 7, is required for the production and trafficking of the intact and functional CFTR protein. Literally thousands of human CFTR allelic mutations have been identified, each with varying impact on protein quality and quantity. As a result, individuals harboring CFTR mutations present with a spectrum of symptoms ranging from CF to normal phenotypes. Those with loss of function but without full CF may present with CFTR-related disorders (CFTR-RDs) including male infertility, sinusitis, pancreatitis, atypical asthma and bronchitis. Studies have demonstrated associations between higher rates of CFTR mutations and oligospermia, epididymal obstruction, congenital bilateral absence of the vas deferens (CBAVD), and idiopathic ejaculatory duct obstruction (EDO). Genetic variants are detected in over three-quarters of men with CBAVD, the reproductive abnormality most classically associated with CFTR aberrations. Likewise, nearly all men with clinical CF will have CBAVD. Current guidelines from multiple groups recommend CFTR screening in all men with clinical CF or CBAVD though a consensus on the minimum number of variants for which to test is lacking. CFTR testing is not recommended as routine screening for men with other categories of infertility. While available CFTR panels include 30 to 96 of the most common variants, complete gene sequencing should be considered if there is a high index of suspicion in a high-risk couple (e.g., partner is CFTR mutation carrier). CF treatments to date have largely targeted end-organ complications. Novel CFTR-modulator treatments aim to directly target CFTR protein dysfunction, effectively circumventing downstream complications, and possibly preventing symptoms like vasal atresia at a young age. Future gene therapies may also hold promise in preventing or reversing genetic changes that lead to CF and CFTR-RD.

Managing Fungal Infections in Cystic Fibrosis Patients: Challenges in Clinical Practice.

Cystic Fibrosis (CF) is an autosomal recessive disease characterized by a mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) protein. Impairment of the CFTR protein in the respiratory tract results in the formation of thick mucus, development of inflammation, destruction of bronchial tissue, and development of bacterial or fungal infections over time. CF patients are commonly colonized and/or infected with fungal organisms, Candida albicans or Aspergillus fumigatus, with prevalence rates ranging from 5% to 78% in the literature. Risk factors for acquiring fungal organisms include older age, coinfection with Pseudomonas aeruginosa, prolonged use of oral and inhaled antibiotics, and lower forced expiratory volume (FEV1). There are limited data available to differentiate between contamination, colonization, and active infection. Furthermore, the pathogenicity of colonization is variable in the literature as some studies report a decline in lung function associated with fungal colonization whereas others showed no difference. Limited data are available for the eradication of fungal colonization and the treatment of active invasive aspergillosis in adult CF patients. In this review article, we discuss the challenges in clinical practice and current literature available for laboratory findings, clinical diagnosis, and treatment options for fungal infections in adult CF patients.

Chronic rhinosinusitis with nasal polyps is associated with impaired TMEM16A-mediated epithelial chloride secretion

Chronic rhinosinusitis with nasal polyps (CRSwNP) is one of the most common chronic disorders with limited therapeutic options. However, the pathogenesis of CRSwNP remains poorly understood. We sought to determine the role of abnormalities in nasal epithelial ion transport in primary epithelial cultures and patients with CRSwNP. We studied epithelial ion transport and transcript levels of the Cl- channels cystic fibrosis transmembrane conductance regulator and transmembrane protein 16A (TMEM16A) in human primary nasal epithelial cultures of patients with CRSwNP and healthy controls. Furthermore, we determined expression levels of proinflammatory cytokines that have been implicated in the regulation of epithelial ion channels (IL-1β, INF-γ, TNF-α, IL-13) and studied effects of the key TH2 signaling molecule IL-13 in CRSwNP and control nasal epithelial cultures. Finally, we measured invivo nasal potential difference to compare epithelial ion transport in patients with CRSwNP and controls. Bioelectric studies demonstrated that Ca2+-activated Cl- secretion was reduced in CRSwNP versus control nasal epithelial cultures. Transcript levels of IL-13 and the Ca2+-activated Cl- channel TMEM16A were increased in CRSwNP cultures. Stimulation with IL-13 increased TMEM16A expression further and restored Ca2+-activated Cl- secretion in CRSwNP cultures. Nasal potential difference measurements demonstrated reduced Ca2+-activated Cl- transport in patients with CRSwNP versus controls. This study demonstrates that TMEM16A-mediated Ca2+-activated Cl- secretion is reduced in primary nasal epithelial cultures and nasal epithelia of patients with CRSwNP. Our data suggest that the Ca2+-activated Cl- channel TMEM16A may be implicated in the pathogenesis and serve as a novel therapeutic target in patients with CRSwNP.

Dysfunctional Inflammation in Cystic Fibrosis Airways: From Mechanisms to Novel Therapeutic Approaches.

Cystic fibrosis (CF) is an inherited disorder caused by mutations in the gene encoding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein, an ATP-gated chloride channel expressed on the apical surface of airway epithelial cells. CFTR absence/dysfunction results in defective ion transport and subsequent airway surface liquid dehydration that severely compromise the airway microenvironment. Noxious agents and pathogens are entrapped inside the abnormally thick mucus layer and establish a highly inflammatory environment, ultimately leading to lung damage. Since chronic airway inflammation plays a crucial role in CF pathophysiology, several studies have investigated the mechanisms responsible for the altered inflammatory/immune response that, in turn, exacerbates the epithelial dysfunction and infection susceptibility in CF patients. In this review, we address the evidence for a critical role of dysfunctional inflammation in lung damage in CF and discuss current therapeutic approaches targeting this condition, as well as potential new treatments that have been developed recently. Traditional therapeutic strategies have shown several limitations and limited clinical benefits. Therefore, many efforts have been made to develop alternative treatments and novel therapeutic approaches, and recent findings have identified new molecules as potential anti-inflammatory agents that may exert beneficial effects in CF patients. Furthermore, the potential anti-inflammatory properties of CFTR modulators, a class of drugs that directly target the molecular defect of CF, also will be critically reviewed. Finally, we also will discuss the possible impact of SARS-CoV-2 infection on CF patients, with a major focus on the consequences that the viral infection could have on the persistent inflammation in these patients.

In silico drug repositioning on F508del-CFTR: A proof-of-concept study on the AIFA library.

Computational drug repositioning is of growing interest to academia and industry, for its ability to rapidly screen a huge number of candidates in silico (exploiting comprehensive drug datasets) together with reduced development cost and time. The potential of drug repositioning has not been fully evaluated yet for cystic fibrosis (CF), a disease mainly caused by deletion of Phe 508 (F508del) of the cystic fibrosis transmembrane conductance regulator (CFTR) protein. F508del-CFTR is thus withheld in the endoplasmic reticulum and rapidly degraded by the ubiquitin/proteasome system. CF is still a fatal disease. Nowadays, it is treatable by some CFTR-rescuing drugs, but new-generation drugs with stronger therapeutic benefits and fewer side effects are still awaited. In this manuscript we report about the results of a pilot computational drug repositioning screening in search of F508del-CFTR-targeted drugs performed on AIFA library by means of a dedicated computational pipeline and surface plasmon resonance binding assay to experimentally validate the computational findings.

Update on Respiratory Fungal Infections in Cystic Fibrosis Lung Disease and after Lung Transplantation.

Cystic fibrosis is the most common autosomal-recessive metabolic disease in the Western world. Impaired trans-membrane chloride transport via the cystic fibrosis transmembrane conductance regulator (CFTR) protein causes thickened body fluids. In the respiratory system, this leads to chronic suppurative cough and recurrent pulmonary infective exacerbations, resulting in progressive lung damage and respiratory failure. Whilst the impact of bacterial infections on CF lung disease has long been recognized, our understanding of pulmonary mycosis is less clear. The range and detection rates of fungal taxa isolated from CF airway samples are expanding, however, in the absence of consensus criteria and univocal treatment protocols for most respiratory fungal conditions, interpretation of laboratory reports and the decision to treat remain challenging. In this review, we give an overview on fungal airway infections in CF and CF-lung transplant recipients and focus on the most common fungal taxa detected in CF, Aspergillus fumigatus, Candida spp., Scedosporium apiospermum complex, Lomentospora species, and Exophiala dermatitidis, their clinical presentations, common treatments and prophylactic strategies, and clinical challenges from a physician’s point of view.

Corrector therapies (with or without potentiators) for people with cystic fibrosis with class II CFTR gene variants (most commonly F508del).

Corrector therapies (with or without potentiators) for people with cystic fibrosis with class II CFTR gene variants (most commonly F508del).

Factors influencing the acquisition and eradication of early Pseudomonas aeruginosa infection in cystic fibrosis

In recent years considerable improvements have been made in increasing the life expectancy of patients with cystic fibrosis. New highly effective modulator therapies targeting the underlying defect in the cystic fibrosis transmembrane conductance regulator protein are expected to enhance lifespan even further. However, chronic Pseudomonas aeruginosa pulmonary infections continue to threaten CF patient lung health and mortality rates. Early and aggressive antibiotic eradication therapies targeting P. aeruginosa are standard practice, but these eradication therapies fail in 10–40% of patients. The reasons for P. aeruginosa eradication failure remain unclear. Thus, this review summarizes the evidence to date for pseudomonal acquisition and eradication failure in the cystic fibrosis lung. A complex combination of host and bacterial factors are responsible for initial establishment of P. aeruginosa pulmonary infections. Moreover, host and pseudomonal factors, polymicrobial interactions, and antimicrobial limitations in relation to P. aeruginosa eradication therapy failure are summarized.

Effect of highly effective modulator therapy on quality of life in adults with cystic fibrosis.

Elexacaftor/tezacaftor/ivacaftor is a highly effective modulator that improves function of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, resulting in improved pulmonary function in patients with cystic fibrosis (CF). We hypothesize that improvements in lung function are associated with improvements in health-related quality of life and sinonasal health. The aim of this study is to measure the effect of elexacaftor/tezacaftor/ivacaftor on patient-reported sinonasal and overall quality of life, and to determine the relationship between changes in these 2 outcome measures. A prospective cohort study was conducted at an accredited adult CF care center. Participants completed the 22-item Sino-Nasal Outcome Test (SNOT-22) and the Cystic Fibrosis Questionnaire-Revised (CFQ-R), a validated patient-reported outcome metric for CF patients, at baseline and at 3 months after initiation of elexacaftor/tezacaftor/ivacaftor. Forty-three individuals completed the study. There was significant improvement in nearly all domains of the SNOT-22 and CFQ-R after 3 months of therapy. SNOT-22 improved from 34.8 to 24.4 (p = 0.000003). Mean baseline FEV-1 improved from 65% to 76% predicted (p = 0.0000005). The greatest effect was seen in those participants previously taking modulator therapy. Linear regression between the change in SNOT-22 individual domains and the CFQ-R respiratory domain revealed the strongest correlation between the extranasal domain score and the respiratory domain of the CFQ-R (R2 = 0.24). CF patients taking elexacaftor/tezacaftor/ivacaftor experience a significant improvement in both sinonasal and health-related quality of life.

Cystic fibrosis drug approved for patients aged 6-11years worked well in clinical practice.

Ivacaftor (IVA) and lumacaftor (LUM) modulate the cystic fibrosis (CF) transmembrane conductance regulator protein. IVA targets gating defects and LUM improves the conformational stability of the F508del mutation. This combined drug was approved for homozygous F508del CF patients aged 12 years or more in 2015 and was extended to children aged 6-11 in 2018.

G551D mutation impairs PKA-dependent activation of CFTR channel that can be restored by novel GOF mutations.

G551D is a major disease-associated gating mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) protein, an ATP- and phosphorylation-dependent chloride channel. G551D causes severe cystic fibrosis (CF) disease by disrupting ATP-dependent channel opening; however, whether G551D affects phosphorylation-dependent channel activation is unclear. Here, we use macropatch recording and Ussing chamber approaches to demonstrate that G551D impacts on phosphorylation-dependent activation of CFTR, and PKA-mediated phosphorylation regulates the interaction between the x-loop in nucleotide-binding domain 2 (NBD2) and cytosolic loop (CL) 1. We show that G551D not only disrupts ATP-dependent channel opening but also impairs phosphorylation-dependent channel activation by largely reducing PKA sensitivity consistent with the reciprocal relationship between channel opening/gating, ligand binding, and phosphorylation. Furthermore, we identified two novel GOF mutations: D1341R in the x-loop near the ATP-binding cassette signature motif in NBD2 and D173R in CL1, each of which strongly increased PKA sensitivity both in the wild-type (WT) background and when introduced into G551D-CFTR. When D1341R was combined with a second GOF mutation (e.g., K978C in CL3), we find that the double GOF mutation maximally increased G551D channel activity such that VX-770 had no further effect. We further show that a double charge-reversal mutation of D1341R/D173R-CFTR exhibited similar PKA sensitivity when compared with WT-CFTR. Together, our results suggest that charge repulsion between D173 and D1341 of WT-CFTR normally inhibits channel activation at low PKA activity by reducing PKA sensitivity, and negative allostery by the G551D is coupled to reduced PKA sensitivity of CFTR that can be restored by second GOF mutations.

Aerosolized lancovutide in adolescents (≥12 years) and adults with cystic fibrosis – a randomized trial

BackgroundLancovutide activates a chloride channel (TMEM-16A) other than the cystic fibrosis (CF) transmembrane conductance regulator protein and could benefit CF patients. MethodsIn this randomized, multi-center, double-blind, placebo-controlled, parallel-group trial 161 patients ≥12 years with a confirmed diagnosis of CF were randomized to either placebo (saline) or active drug in 3 different dosing schemes of 2.5mg inhaled lancovutide (once daily, every other day or twice a week) for eight weeks. The primary endpoint was the change in the forced expiratory volume in 1 second (FEV1) percent predicted. Secondary endpoints included further lung function parameters (FEV1 (absolute), functional vital capacity percent predicted, forced expiratory flow percent predicted, pulse oximetry), quality of life assessment, pulmonary exacerbations, hospitalization due to pulmonary exacerbations, time to first pulmonary exacerbation, duration of anti-inflammatory, mucolytic or antibiotic treatment, and safety. ResultsThere was no significant difference in the change in FEV1 percent predicted, quality of life, other lung function parameters, pulmonary exacerbations or requirement of additional treatment between groups. Overall, the inhalation of lancovutide was safe although a higher rate of adverse events, especially related to the respiratory system, occurred as compared to placebo. ConclusionsLancovutide did not improve FEV1 percent predicted when compared to placebo (NCT00671736).

Reduced Intestinal Inflammation With Lumacaftor/Ivacaftor in Adolescents With Cystic Fibrosis.

A chronic intestinal inflammation may occur in patients with cystic fibrosis (CF), while no therapeutic management is proposed. Although Lumacaftor/Ivacaftor is well-known to modulate the defective cystic fibrosis transmembrane conductance regulator (CFTR) protein in lungs, no data are available on the impact of this treatment on CF intestinal disorders. We, therefore, investigated the evolution of intestinal inflammation after initiation of Lumacaftor/Ivacaftor in CF adolescents (median of follow-up: 336 days [IQR: 278;435]). Median fecal calprotectin concentrations decreased significantly after Lumacaftor/Ivacaftor initiation (102 μg/g [IQR: 69-210]) compared with the baseline (713 μg/g (IQR:148-852), P = 0.001). To our knowledge, this study showed for the first time that CF-related intestinal inflammation is improved by Lumacaftor/Ivacaftor treatment.

The impact of ivacaftor on sinonasal pathology in S1251N-mediated cystic fibrosis patients.

ImportanceSinonasal symptoms in patients suffering from cystic fibrosis can negatively influence the quality of life and sinuses can be a niche for pathogens causing infection and inflammation leading to a decrease of lung function. Ivacaftor, a potentiator of the Cystic Fibrosis Transmembrane Conductance Regulator protein, has shown improvement in pulmonary function in cystic fibrosis patients with different forms of class III gating mutations. However, the effects of ivacaftor on sinonasal pathology have hardly been studied.ObjectiveTo determine the impact of ivacaftor therapy on sinonasal pathology in patients with cystic fibrosis with an S1251N mutation.DesignProspective observational mono-center cohort study, between June 2015 and December 2016.SettingA tertiary referral center in Utrecht, The Netherlands.ParticipantsEight patients with cystic fibrosis with an S1251N mutation, treated with the potentiator ivacaftor were investigated.ExposuresIvacaftor (Kalydeco, VX-770) therapy. Computed tomography imaging of paranasal sinuses. Nasal nitric oxide concentration measurements and nasal endoscopy.Main outcomes and measuresPrimary outcome is opacification of paranasal sinuses examined with computed tomography scan analysis and scaled by the modified Lund-Mackay score before and one year after treatment. Secondary outcomes are nasal nitric oxide concentration levels, sinonasal symptoms and nasal endoscopic findings before and approximately two months and in some cases one year after treatment.ResultsComputed tomography scan analysis showed a significant decrease in opacification of the majority of paranasal sinuses comparing the opacification score per paranasal sinus before and after one year of treatment with ivacaftor. Median nasal nitric oxide levels significantly improved from 220.00 (IQR:136.00–341.18) to 462.84 (IQR:233.17–636.25) (p = 0.017) parts per billion. Likewise, the majority of sinonasal symptoms and nasal endoscopic pathology decreased or resolved at two months after the use of ivacaftor.Conclusion and relevanceIvacaftor appears to improve sinonasal outcome parameters and thereby sinonasal health in patients with cystic fibrosis with an S1251N mutation.

Cytoskeleton regulators CAPZA2 and INF2 associate with CFTR to control its plasma membrane levels under EPAC1 activation.

Cystic Fibrosis (CF), the most common lethal autosomic recessive disorder among Caucasians, is caused by mutations in the gene encoding the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) protein, a cAMP-regulated chloride channel expressed at the apical surface of epithelial cells. Cyclic AMP regulates both CFTR channel gating through a protein kinase A (PKA)-dependent process and plasma membane (PM) stability through activation of the exchange protein directly activated by cAMP1 (EPAC1). This cAMP effector, when activated promotes the NHERF1:CFTR interaction leading to an increase in CFTR at the PM by decreasing its endocytosis. Here, we used protein interaction profiling and bioinformatic analysis to identify proteins that interact with CFTR under EPAC1 activation as possible regulators of this CFTR PM anchoring. We identified an enrichment in cytoskeleton related proteins among which we characterized CAPZA2 and INF2 as regulators of CFTR trafficking to the PM. We found that CAPZA2 promotes wt-CFTR trafficking under EPAC1 activation at the PM whereas reduction of INF2 levels leads to a similar trafficking promotion effect. These results suggest that CAPZA2 is a positive regulator and INF2 a negative one for the increase of CFTR at the PM after an increase of cAMP and concomitant EPAC1 activation. Identifying the specific interactions involving CFTR and elicited by EPAC1 activation provides novel insights into late CFTR trafficking, insertion and/or stabilization at the PM and highlighs new potential therapeutic targets to tackle CF disease.

Phosphorylation of the Chaperone-Like HspB5 Rescues Trafficking and Function of F508del-CFTR.

Cystic Fibrosis is a lethal monogenic autosomal recessive disease linked to mutations in Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) protein. The most frequent mutation is the deletion of phenylalanine at position 508 of the protein. This F508del-CFTR mutation leads to misfolded protein that is detected by the quality control machinery within the endoplasmic reticulum and targeted for destruction by the proteasome. Modulating quality control proteins as molecular chaperones is a promising strategy for attenuating the degradation and stabilizing the mutant CFTR at the plasma membrane. Among the molecular chaperones, the small heat shock protein HspB1 and HspB4 were shown to promote degradation of F508del-CFTR. Here, we investigated the impact of HspB5 expression and phosphorylation on transport to the plasma membrane, function and stability of F508del-CFTR. We show that a phosphomimetic form of HspB5 increases the transport to the plasma membrane, function and stability of F508del-CFTR. These activities are further enhanced in presence of therapeutic drugs currently used for the treatment of cystic fibrosis (VX-770/Ivacaftor, VX-770+VX-809/Orkambi). Overall, this study highlights the beneficial effects of a phosphorylated form of HspB5 on F508del-CFTR rescue and its therapeutic potential in cystic fibrosis.