Search Results (18)

Cystic fibrosis (CF), the most common hereditary lung disease in Caucasians, is caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR). We evaluated CFTR function using a newly developed Ussing chamber system, the Multi Trans Epithelial Current Clamp (MTECC), in an in vitro model of human airway epithelia. Air–liquid interface (ALI) cultures were established from nasal brushings of healthy controls (HC) and CF patients with biallelic CFTR variants. ALI layer thickness was similar between groups (HC: 62 ± 13 µm; CF: 55 ± 9 µm). Immunofluorescence showed apical CFTR expression in HC, but reduced or absent signal in CF cultures. MTECC enabled continuous measurement of transepithelial resistance (Rt), potential difference (PD), and conductance (G_t). G_t was significantly reduced in CF cultures compared to HC (0.825 ± 0.024 vs. −0.054 ± 0.016 mS/cm²), indicating impaired cAMP-inducible ion transport by CFTR. Treatment of CF cultures with elexacaftor, tezacaftor, and ivacaftor (Trikafta^®) increased G_t, reflecting partial restoration of CFTR function. These findings demonstrate the utility of MTECC in detecting functional differences in CFTR activity and support its use as a platform for evaluating CFTR-modulating therapies. Our model may contribute to the development of personalized treatment strategies for CF patients. Full article

Cystic fibrosis (CF) is a recessive genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) protein, a chloride and bicarbonate channel localized on the plasma membrane of epithelial cells. Over the last three decades, high-throughput screening assays have been extensively employed in identifying drugs that target specific defects arising from CFTR mutations. The two main categories of such compounds are potentiators, which enhance CFTR gating by increasing the channel’s open probability, and correctors, which improve CFTR protein folding and trafficking to the plasma membrane. In addition to these, other investigational molecules include amplifiers and stabilizers, which enhance the levels and the stability of CFTR on the cell surface, and read-through agents that promote the insertion of correct amino acids at premature termination codons. Currently, four CFTR modulators are clinically approved: the potentiator ivacaftor (VX-770), either as monotherapy or in combination with the correctors lumacaftor (VX-809), tezacaftor (VX-661), and elexacaftor (VX-445). Among these, the triple combination VX-445/VX-661/VX-770 (marketed as Trikafta^® in the US and Kaftrio^® in Europe) has emerged as the most effective CFTR modulator therapy to date, demonstrating significant clinical benefits in phase III trials for patients with at least one F508del CFTR allele. Despite these advancements, the mechanisms of action and binding sites of these modulators on CFTR have only recently begun to be elucidated. A deeper understanding of these mechanisms could provide essential insights for developing more potent and effective modulators, particularly in combination therapies. This narrative review delves into the mechanism of action, binding sites, and combinatorial effects of approved and investigational CFTR modulators, highlighting ongoing efforts to broaden therapeutic options for individuals with CF. Full article

The triple combination therapy for cystic fibrosis (CF), including elexacaftor, tezacaftor and ivacaftor (ETI or Trikafta), has been shown to improve lung function and reduce pulmonary exacerbations, thereby enhancing the quality of life for most CF patients. Recent findings suggest that both the individual components and ETI may have potential off-target effects, highlighting the need to understand how these modulators impact cellular physiology, particularly in cells that do not express CF transmembrane conductance regulator (CFTR). We used HEK293 cells, as a cell model not expressing the CFTR protein, to evaluate the effect of ETI and each of its components on autophagic machinery and on the Rab5/7 components of the Rab pathway. We firstly demonstrate that the single modulators Teza and Iva, and the combinations ET and ETI, increased ROS production in the absence of their target while decreasing it in cells expressing the CFTR ∆F508del. This increase in cellular stress was followed by an increase in the total level of polyubiquitinated proteins as well as the p62 level and LC3II/LC3I ratio. Furthermore, we found that ETI had the opposite effect on Rabs by increasing Rab5 levels while decreasing Rab7. Interestingly, these changes were abolished by the expression of mutated CFTR. Overall, our data suggest that in the absence of their target, both the individual modulators and ETI increased ROS production and halted both autophagic flux and plasma membrane protein recycling. Full article

Background: The most recent modulator combination, elexacaftor/tezacaftor/ivacaftor (Trikafta^®), has been shown to improve clinical outcomes in most patients with cystic fibrosis (PwCF). Unfortunately, the clinical benefits are sometimes variable; thus, improving our knowledge of the possible causes of this variability can help reduce it. Methods: Circulating mononuclear cells (CMCs) and plasma were collected from 16 PwCF (including those on Trikafta^® therapy) and 4 non-CF subjects. Cystic fibrosis transmembrane conductance regulator (CFTR) activity and matrix metalloprotease 9 (MMP9) expression were monitored before and after therapy, together with some clinical parameters. The relationship between MMP9 expression and the modulation of the extracellular-regulated 1/2 (ERK1/2) and nuclear factor-kB (NF-kB) pathways was also analyzed. Results: MMP9, markedly expressed in the CMCs and plasma of all the patients included in the study, was downregulated in the clinically responsive PwCF. In the non-responder, the MMP9 levels remained high. The modulation of MMP9 following treatment with Trikafta^® may be controlled by the NF-kB pathway. Conclusions: These data strongly suggest that MMP9 downregulation is a potential biomarker of therapy efficacy and that it could be useful in understanding the molecular events underlying the variable clinical responses of patients to Trikafta^®. This knowledge could be helpful for future studies of personalized medicine and thereby ensure improvements in individual responses to therapies. Full article

Elexacaftor/Tezacaftor/Ivacaftor (ELX/TEZ/IVA) is a new CFTR (Cystic Fibrosis Transmembrane Conductance Regulator) modulator treatment, used over the last few years, which has shown an improvement in different clinical outcomes in patients with cystic fibrosis (CF). The objective of this study was a systematic research of the literature on the efficacy and safety of this CFTR modulator on patients with CF. A search of Pubmed was conducted for randomized clinical trials and observational studies published from 2012 to September 2022. The included full manuscripts comprised nine clinical trials and 16 observational studies, whose participants were aged ≥12 years or were children 6–11 years old with at least one Phe508del mutation and/or advanced lung disease (ALD). These studies reported that ELX/TEZ/IVA has a significant positive effect on the lung function of patients with CF, by ameliorating parameters such as FEV₁, LCI, pulmonary exacerbations or sweat chloride concentration, increasing BMI and improving quality of their life. Its role in cystic fibrosis-related diabetes (CFRD) is not yet clear. It was found that this new CFTR modulator has an overall favorable safety profile, with mild to moderate adverse events. Further studies are needed for a deeper understanding of the impact of CFTR modulators on other CF manifestations, or the possibility of treating with ELX/TEZ/IVA CF patients with rare CFTR mutations. Full article

Cystic fibrosis (CF) is a potentially fatal monogenic disease that causes a progressive multisystemic pathology. Over the last decade, the introduction of CF transmembrane conductance regulator (CFTR) modulator drugs into clinical practice has profoundly modified the lives of many people with CF (PwCF) by targeting the fundamental cause of the disease. These drugs consist of the potentiator ivacaftor (VX-770) and the correctors lumacaftor (VX-809), tezacaftor (VX-661), and elexacaftor (VX-445). In particular, the triple combination of CFTR modulators composed of elexacaftor, tezacaftor, and ivacaftor (ETI) represents a life-changing therapy for the majority of PwCF worldwide. A growing number of clinical studies have demonstrated the safety and efficacy of ETI therapy in both short- and long-term (up to two years of follow-up to date) and its ability to significantly reduce pulmonary and gastrointestinal manifestations, sweat chloride concentration, exocrine pancreatic dysfunction, and infertility/subfertility, among other disease signs and symptoms. Nevertheless, ETI therapy-related adverse effects have also been reported, and close monitoring by a multidisciplinary healthcare team remains vital. This review aims to address and discuss the major therapeutic benefits and adverse effects reported by the clinical use of ETI therapy for PwCF. Full article

The new breakthrough cystic fibrosis (CF) drug combination of ivacaftor (IVA), tezacaftor (TEZ), and elexacaftor (ELX), namely “caftor” drugs, directly modulates the activity and trafficking of the defective CF transmembrane conductance regulator protein (CFTR) underlying the CF disease. The role of therapeutic drug monitoring (TDM) of caftor drugs in clinical settings has recently been established. The availability of reliable and robust analytical methods for the quantification of IVA, TEZ, and ELX is essential to support dose–concentration–effect studies. We have developed and validated a new liquid chromatography–tandem mass spectrometry (LC–MS/MS) for the rapid and simultaneous quantification of IVA, TEZ, and ELX from the plasma of CF patients. The method was based on a rapid extraction protocol from 50 μL human plasma and separation on a reversed-phase C-18 HPLC column after the addition of deuterated internal standards. Accurate analyte quantification using multiple reaction monitoring (MRM) detection was then obtained using a Thermofisher Quantiva triple-quadrupole MS coupled to an Ultimate 3000 UHPLC. The method has been validated following international (EMA) guidelines for bioanalytical method validation and has been tested on plasma samples from 62 CF patients treated with the three-drug combination IVA/TEZ/ELX, marketed as Kaftrio^® or Trikafta^®, in steady-state condition. The assay was linear over wide concentration ranges (0.008–12 mg/L) in plasma for IVA, TEZ, and ELX, suitable for a broad range of plasma concentrations, and accurate and reproducible in the absence of matrix effects. The stability of analytes for at least 30 days at room temperature could allow for cost-effective shipment and storage. On the same day of sample collection, a sweat test was evaluated for 26 associated patients’ samples, FEV1 (%) for 58, and BMI was calculated for 62. However, Spearman correlation showed no correlation between C_through plasma concentrations of analytes (IVA, TEZ, ELX) and sweat test, FEV1 (%), or BMI. Our method proved to be suitable for TDM and could be helpful in assessing dose–concentration–response correlations in larger studies. Full article

COPD is a lifestyle-related disease resulting from irreversible damage to respiratory tissues mostly due to chronic exposure to environmental pollutants, including cigarette smoke. Environmental pathogens and pollutants induce the acquired dysfunction of the CFTR Cl⁻ channel, which is invoked in COPD. Despite the increased incidence of CFTR polymorphism R75Q or M470V in COPD patients, the mechanism of how the CFTR variant affects COPD pathogenesis remains unclear. Here, we investigated the impact of CFTR polymorphisms (R75Q, M470V) on the CFTR function in airway epithelial cell models. While wild-type (WT) CFTR suppressed the proinflammatory cytokine production induced by COPD-related pathogens including pyocyanin (PYO), R75Q- or M470V-CFTR failed. Mechanistically, the R75Q- or M470V-CFTR fractional PM activity (FPMA) was significantly lower than WT-CFTR in the presence of PYO. Notably, the CF drug Trikafta corrected the PM expression of R75Q- or M470V-CFTR even upon PYO exposure and consequently suppressed the excessive IL-8 production. These results suggest that R75Q or M470V polymorphism impairs the CFTR function to suppress the excessive proinflammatory response to environmental pathogens associated with COPD. Moreover, Trikafta may be useful to prevent the COPD pathogenesis associated with acquired CFTR dysfunction. Full article

Mutations in CFTR cause misfolding and decreased or absent ion-channel function, resulting in the disease Cystic Fibrosis. Fortunately, a triple-modulator combination therapy (Trikafta) has been FDA-approved for 178 mutations, including all patients who have F508del on one allele. That so many CFTR mutants respond well to modulators developed for a single mutation is due to the nature of the folding process of this multidomain protein. We have addressed the question ‘What characterizes the exceptions: the mutants that functionally respond either not or extremely well’. A functional response is the product of the number of CFTR molecules on the cell surface, open probability, and conductivity of the CFTR chloride channel. By combining biosynthetic radiolabeling with protease-susceptibility assays, we have followed CF-causing mutants during the early and late stages of folding in the presence and absence of modulators. Most CFTR mutants showed typical biochemical responses for each modulator, such as a TMD1 conformational change or an increase in (cell-surface) stability, regardless of a functional response. These modulators thus should still be considered for hypo-responder genotypes. Understanding both biochemical and functional phenotypes of outlier mutations will boost our insights into CFTR folding and misfolding, and lead to improved therapeutic strategies. Full article

Cystic fibrosis (CF) is the most common monogenic disorder, caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Over the last 30 years, tremendous progress has been made in understanding the molecular basis of CF and the development of treatments that target the underlying defects in CF. Currently, a highly effective CFTR modulator treatment (Kalydeco™/Trikafta™) is available for 90% of people with CF. In this review, we will give an extensive overview of past and ongoing efforts in the development of therapies targeting the molecular defects in CF. We will discuss strategies targeting the CFTR protein (i.e., CFTR modulators such as correctors and potentiators), its cellular environment (i.e., proteostasis modulation, stabilization at the plasma membrane), the CFTR mRNA (i.e., amplifiers, nonsense mediated mRNA decay suppressors, translational readthrough inducing drugs) or the CFTR gene (gene therapies). Finally, we will focus on how these efforts can be applied to the 15% of people with CF for whom no causal therapy is available yet. Full article

Background: Modulator therapy represents a significant step forward in CF care and is expected to have a significant impact on the health and mortality of many individuals with CF. Studies have predominantly explored the physiological effects of modulator therapy on clinical outcomes, with little consideration of the individual lived experience of modulator therapy among adults with Cystic Fibrosis. Methods: To explore this, semi-structured interviews were conducted with 12 individuals currently taking Kaftrio, which were subsequently thematically analysed. Results: Three overarching themes were identified: (i) positive perception of Kaftrio, (ii) negative perception of Kaftrio, and (iii) the relationships with the clinical team. The experience of modulator therapy should be recognised as being unique to the individual, with perceptions of illness, self-identity, and outcomes strongly dictating the lived experience. Conclusions: There is a consensus that, while for many, the quality of life is evidently increased through the use of Kaftrio, this is not without its own challenges. This highlights the need for both individuals with CF and their clinical teams to learn to navigate this new disease landscape. Full article

Cystic Fibrosis (CF) is caused by mutations on the CF transmembrane conductance regulator (CFTR) gene and is associated with chronic infection and inflammation. Recently, it has been demonstrated that LPS-induced CFTR dysfunction in airway epithelial cells is due to an early oxidative stress. Dimethyl fumarate (DMF) is an approved anti-inflammatory and anti-oxidant drug for auto-immune and inflammatory diseases, but its role in the CF has never been investigated. In this study, we examined the effect of DMF on CF-related cytokines expression, ROS measurements and CFTR channel function. We found that DMF reduced the inflammatory response to LPS stimulation in both CF and non-CF bronchial epithelial cells, both as co-treatment and therapy, and restored LPS-mediated decrease of Trikafta™-mediated CFTR function in CF cells bearing the most common mutation, c.1521_1523delCTT (F508del). DMF also inhibited the inflammatory response induced by IL-1β/H₂O₂ and IL-1β/TNFα, mimicking the inflammatory status of CF patients. Finally, we also demonstrated that DMF exhibited an anti-oxidant effect on CF cells after different inflammatory stimulations. Since DMF is an approved drug, it could be further investigated as a novel anti-inflammatory molecule to ameliorate lung inflammation in CF and improve the CFTR modulators efficacy. Full article

The new CFTR modulator combination, elexacaftor/tezacaftor/ivacaftor (Trikafta) was approved by the FDA in October 2019 for treatment of Cystic Fibrosis in patients 6 years of age or older who have at least one F508del mutation in one allele and a minimal-function or another F508del mutation in the other allele. However, there is a group of patients, in addition to those with rare mutations, in which despite the presence of a F508del in one allele, it was not possible to identify any mutation in the other allele. To date, these patients are excluded from treatment with Trikafta in Italy, where the CF patients carrying F508del/unknown represent about 1.3% (71 patients) of the overall Italian CF patients. In this paper we show that the Trikafta treatment of nasal epithelial cells, derived from F508del/Unknown patients, results in a significant rescue of CFTR activity. Based on our findings, we think that the F508del/Unknown patients considered in this study could obtain clinical benefits from Trikafta treatment, and we strongly suggest their eligibility for this type of treatment. This study, adding further evidence in the literature, once again confirms the validity of functional studies on nasal cells in the cystic fibrosis theratyping and personalized medicine. Full article

Trikafta, a triple-combination drug, consisting of folding correctors VX-661 (tezacaftor), VX-445 (elexacaftor) and the gating potentiator VX-770 (ivacaftor) provided unprecedented clinical benefits for patients with the most common cystic fibrosis (CF) mutation, F508del. Trikafta indications were recently expanded to additional 177 mutations in the CF transmembrane conductance regulator (CFTR). To minimize life-long pharmacological and financial burden of drug administration, if possible, we determined the necessary and sufficient modulator combination that can achieve maximal benefit in preclinical setting for selected mutants. To this end, the biochemical and functional rescue of single corrector-responsive rare mutants were investigated in a bronchial epithelial cell line and patient-derived human primary nasal epithelia (HNE), respectively. The plasma membrane density of P67L-, L206W- or S549R-CFTR corrected by VX-661 or other type I correctors was moderately increased by VX-445. Short-circuit current measurements of HNE, however, uncovered that correction comparable to Trikafta was achieved for S549R-CFTR by VX-661 + VX-770 and for P67L- and L206W-CFTR by the VX-661 + VX-445 combination. Thus, introduction of a third modulator may not provide additional benefit for patients with a subset of rare CFTR missense mutations. These results also underscore that HNE, as a precision medicine model, enable the optimization of mutation-specific modulator combinations to maximize their efficacy and minimize life-long drug exposure of CF patients. Full article