Cell
Molecular mechanisms of CFTR chloride channel dysfunction in cystic fibrosis
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In vitro modulator responsiveness of 655 CFTR variants found in people with cystic fibrosis
2024, Journal of Cystic FibrosisIn 2017, the US Food and Drug Administration initiated expansion of drug labels for the treatment of cystic fibrosis (CF) to include CF transmembrane conductance regulator (CFTR) gene variants based on in vitro functional studies. This study aims to identify CFTR variants that result in increased chloride (Cl−) transport function by the CFTR protein after treatment with the CFTR modulator combination elexacaftor/tezacaftor/ivacaftor (ELX/TEZ/IVA). These data may benefit people with CF (pwCF) who are not currently eligible for modulator therapies.
Plasmid DNA encoding 655 CFTR variants and wild-type (WT) CFTR were transfected into Fisher Rat Thyroid cells that do not natively express CFTR. After 24 h of incubation with control or TEZ and ELX, and acute addition of IVA, CFTR function was assessed using the transepithelial current clamp conductance assay. Each variant's forskolin/cAMP-induced baseline Cl− transport activity, responsiveness to IVA alone, and responsiveness to the TEZ/ELX/IVA combination were measured in three different laboratories. Western blots were conducted to evaluate CFTR protein maturation and complement the functional data.
253 variants not currently approved for CFTR modulator therapy showed low baseline activity (<10 % of normal CFTR Cl− transport activity). For 152 of these variants, treatment with ELX/TEZ/IVA improved the Cl− transport activity by ≥10 % of normal CFTR function, which is suggestive of clinical benefit. ELX/TEZ/IVA increased CFTR function by ≥10 percentage points for an additional 140 unapproved variants with ≥10 % but <50 % of normal CFTR function at baseline. These findings significantly expand the number of rare CFTR variants for which ELX/TEZ/IVA treatment should result in clinical benefit.
Mutation targeted therapy in cystic fibrosis (CF) is still not eligible for all CF subjects, especially for cases carrying rare variants such as the CFTR genotype W57G/A234D (c.169T>G/c.701C>A). We performed in silico analysis of the effects of these variants on protein stability, which we functionally characterized using colonoids and reprogrammed nasal epithelial cells. The effect of mutations on cystic fibrosis transmembrane conductance regulator (CFTR) protein was analyzed by western blotting, forskolin-induced swelling (FIS), and Ussing chamber analysis. We detected a residual CFTR function that increases following treatment with the CFTR modulators VX661±VX445±VX770, correlates among models, and is associated with increased CFTR protein levels following treatment with CFTR correctors. In vivo treatment with VX770 reduced sweat chloride concentration to non-CF levels, increased the number of CFTR-dependent sweat droplets, and induced a 6% absolute increase in predicted FEV1% after 27 weeks of treatment indicating the relevance of theratyping with patient-derived cells in CF.
Multi-step synthesis of novel 2-methyl-3-carboxamide-4-quinolones and an enhanced sampling simulation method to identify potentiators for cystic fibrosis
2023, Materials Today ChemistryCystic fibrosis (CF) is a life-threatening hereditary disease caused by mutations in the CF trans membrane conductance regulator (CFTR) gene. The commonly utilized medication for CF is a potentiator that restores CFTR function by stabilizing the channel in open conformation. A multistep synthesis method has been demonstrated here for synthesizing 2-methyl-3-carboxamide-4-quinolones having potential activities on CF. The synthesized molecules were obtained in quantitative yield and purified using recrystallization techniques without column purification. An extensive computational analysis was carried out to access the potential of 2-methyl-3-carboxamide-4-quinolones as potentiators. The computational analysis using molecular docking, classical and membrane-embedded molecular dynamics simulations in different lipid environments, and MM-PBSA revealed that the molecule 4d formed a stable complex. The binding free energies from umbrella sampling simulations for standard molecules ivacaftor and GPLG1837 were −26.133 kJ/mol and −28.471 kJ/mol, respectively, while for 4d a lower free energy value was obtained (−37.852 kJ/mol), which further strengthened the candidature of 4d as a promising potentiator molecule. This research identified a hit molecule that could be utilized further in in-vitro and in-vivo studies to establish it as a potentiator against CF and also provided a suitable and efficient synthesis method for the pharmaceutical industries.
The future of cystic fibrosis treatment: from disease mechanisms to novel therapeutic approaches
2023, The LancetWith the 2019 breakthrough in the development of highly effective modulator therapy providing unprecedented clinical benefits for over 90% of patients with cystic fibrosis who are genetically eligible for treatment, this rare disease has become a front runner of transformative molecular therapy. This success is based on fundamental research, which led to the identification of the disease-causing CFTR gene and our subsequent understanding of the disease mechanisms underlying the pathogenesis of cystic fibrosis, working together with a continuously evolving clinical research and drug development pipeline. In this Series paper, we focus on advances since 2018, and remaining knowledge gaps in our understanding of the molecular mechanisms of CFTR dysfunction in the airway epithelium and their links to mucus dysfunction, impaired host defences, airway infection, and chronic inflammation of the lungs of people with cystic fibrosis. We review progress in (and the remaining obstacles to) pharmacological approaches to rescue CFTR function, and novel strategies for improved symptomatic therapies for cystic fibrosis, including how these might be applicable to common lung diseases, such as bronchiectasis and chronic obstructive pulmonary disease. Finally, we discuss the promise of genetic therapies and gene editing approaches to restore CFTR function in the lungs of all patients with cystic fibrosis independent of their CFTR genotype, and the unprecedented opportunities to transform cystic fibrosis from a fatal disease to a treatable and potentially curable one.
Cystic fibrosis
2023, Presse MedicaleCystic fibrosis (CF) is an autosomal recessive genetic disease caused by variants in the gene encoding for the cystic fibrosis transmembrane conductance regulator (CFTR) protein. CFTR dysfunction results in abnormal chloride and bicarbonate transport in epithelial cells, leading to a multiorgan disease dominated by respiratory and digestive manifestations. The respiratory disease, which is characterized by airway mucus plugging, chronic bacterial infection and progressive development of bronchiectasis, may lead to chronic respiratory failure, which is the main cause of premature death in people with CF. Over the past 50 years, major progress has been obtained by implementing multidisciplinary care, including nutritional support, airway clearance techniques and antibiotics in specialized CF centers. The past 10 years have further seen the progressive development of oral medications, called CFTR modulators, that partially restore ion transport and lead to a major improvement in clinical manifestations and lung function, presumably resulting in longer survival. Although an increasing proportion of people with CF are being treated with CFTR modulators, challenges remain regarding access to CFTR modulators due to their high cost, and their lack of marketing approval and/or effectiveness in people with rare CFTR variants. The anticipated increase in the number of adults with CF and their aging also challenge the current organization of CF care. The purpose of this review article is to describe current status and future perspective of CF disease and care.
Cystic fibrosis: Biological explorations and new therapies
2023, Option/Bio