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Insulin secretion is impaired in individuals with cystic fibrosis (CF), contributing to high rates of CF-related diabetes (CFRD) and substantially increasing disease burden. To develop improved therapies for CFRD, better knowledge of pancreatic pathology in CF is needed. Glucagon like peptide-1 (GLP-1) from islet α cells potentiates insulin secretion by binding GLP-1 receptors (GLP-1Rs) on β cells. We determined whether expression of GLP-1 and/or its signaling components are reduced in CFRD, thereby contributing to impaired insulin secretion. Immunohistochemistry of pancreas from humans with CFRD versus no-CF/no-diabetes revealed no difference in GLP-1 immunoreactivity per islet area, whereas GLP-1R immunoreactivity per islet area or per insulin-positive islet area was reduced in CFRD. Using spatial transcriptomics, we observed several differentially expressed α- and/or β-cell genes between CFRD and control pancreas. In CFRD, we found upregulation of α-cell PCSK1 which encodes the enzyme (PC1/3) that generates GLP-1, and downregulation of α-cell PCSK1N which inhibits PC1/3. Gene set enrichment analysis also revealed α and β cell-specific pathway dysregulation in CFRD. Together, our data suggest intra-islet GLP-1 is not limiting in CFRD, but its action may be restricted due to reduced GLP-1R protein levels. Thus, restoring β-cell GLP-1R protein expression may improve β-cell function in CFRD., (© 2024. The Author(s).)

Base editing could correct nonsense mutations that cause cystic fibrosis (CF), but clinical development is limited by the lack of delivery methods that efficiently breach the barriers presented by airway epithelia. Here, we present a novel amphiphilic shuttle peptide based on the previously reported S10 peptide that substantially improved base editor ribonucleoprotein (RNP) delivery. Studies of the S10 secondary structure revealed that the alpha-helix formed by the endosomal leakage domain (ELD), but not the cell penetrating peptide (CPP), was functionally important for delivery. By isolating and extending the ELD, we created a novel shuttle peptide, termed S237. While S237 achieved lower delivery of green fluorescent protein, it outperformed S10 at Cas9 RNP delivery to cultured human airway epithelial cells and to pig airway epithelia in vivo, possibly due to its lower net charge. In well-differentiated primary human airway epithelial cell cultures, S237 achieved a 4.6-fold increase in base editor RNP delivery, correcting up to 9.4% of the cystic fibrosis transmembrane conductance regulator (CFTR) R553X allele and restoring CFTR channel function close to non-CF levels. These findings deepen the understanding of peptide-mediated delivery and offer a translational approach for base editor RNP delivery for CF airway disease., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Cystic fibrosis related diabetes (CFRD), the main co-morbidity in cystic fibrosis (CF), is associated with higher rates of lung function decline. We hypothesize that airway epithelial barrier function is impaired in CF and is further exacerbated under hyperglycemia, worsening pulmonary outcomes. Using 16HBE cells, we studied the effects of hyperglycemia in airway epithelial barrier function. Results show increased paracellular dye flux in CF cells in response to insulin under hyperglycemia. Gene expression experiments identified claudin-4 (CLDN4) as a key tight junction protein dysregulated in CF cells. CLDN4 protein localization by confocal microscopy showed that CLDN4 was tightly localized at tight junctions in WT cells, which did not change under hyperglycemia. ln contrast, CLDN4 was less well-localized in CF cells at normal glucose and localization was worsened under hyperglycemia. Treatment with highly effective modulator compounds (ETI) reversed this trend, and CFTR rescue was not affected by insulin or hyperglycemia. Bulk RNA sequencing showed differences in transcriptional responses in CF compared to WT cells under normal or high glucose, highlighting promising targets for future investigation. One of these targets is protein tyrosine phosphatase receptor type G (PTPRG), which has been previously found to play a role in defective Akt signaling and insulin resistance., (© 2024. The Author(s).)

Background/objectives: Cystic fibrosis (CF) is one of the most common autosomal-recessive disorders worldwide. The incidence of CF depends on the prevalence of cystic fibrosis transmembrane conductance regulator gene ( CFTR ) mutations in the population, which is determined by genetic diversity and ethnicity., Methods: The search for the causes of mutations in the transmembrane conductance regulator gene (CFTR) was carried out using targeted next-generation sequencing (NGS) on the Illumina platform in patients with cystic fibrosis from the Republic of Bashkortostan (Russia), taking into account the ethnic structure of the sample., Results: A total of 35 distinct causal variants were found in 139 cases from 129 families. Five (F508del, E92K, 3849+10kbC>T, CFTRdele2.3, L138ins) explain 78.7% of identified CF causal alleles. Variants N13103K and 394delTT were found in four families each. Variants 2143delT, S1196X, W1282X, Y84X, G194R, and 1525-1G>A, as well as the two previously described complex alleles-c. [S466X; R1070Q] and str.[G509D;E217G]-were found in two or three families each. Twenty additional variants occurred only once. Variant c.3883_3888dup has not been described previously. Thus, regional and ethnic features were identified in the spectrum of frequencies of pathogenic variants of the CFTR gene in the three major sub-groups of patients-Russians, Tatars, and Bashkirs., Conclusions: Taking into account these results, highlighting the genetic specificity of the region, a more efficient search for CFTR mutations in patients can be performed. In particular it is possible to choose certain test kits for quick and effective genetic screening before use of NGS sequencing.

The cotranslational misfolding of the cystic fibrosis transmembrane conductance regulator chloride channel (CFTR) plays a central role in the molecular basis of CF. The misfolding of the most common CF variant (ΔF508) remodels both the translational regulation and quality control of CFTR. Nevertheless, it is unclear how the misassembly of the nascent polypeptide may directly influence the activity of the translation machinery. In this work, we identify a structural motif within the CFTR transcript that stimulates efficient -1 ribosomal frameshifting and triggers the premature termination of translation. Though this motif does not appear to impact the interactome of wild-type CFTR, silent mutations that disrupt this RNA structure alter the association of nascent ΔF508 CFTR with numerous translation and quality control proteins. Moreover, disrupting this RNA structure enhances the functional gating of the ΔF508 CFTR channel at the plasma membrane and its pharmacological rescue by the CFTR modulators contained in the CF drug Trikafta. The effects of the RNA structure on ΔF508 CFTR appear to be attenuated in the absence of the ER membrane protein complex, which was previously found to modulate ribosome collisions during "preemptive quality control" of a misfolded CFTR homolog. Together, our results reveal that ribosomal frameshifting selectively modulates the assembly, function, and pharmacological rescue of a misfolded CFTR variant. These findings suggest that interactions between the nascent chain, quality control machinery, and ribosome may dynamically modulate ribosomal frameshifting in order to tune the processivity of translation in response to cotranslational misfolding., Competing Interests: Competing interests statement:The authors declare no competing interest.

To identify modifier loci underlying variation in body mass index (BMI) in persons with cystic fibrosis (pwCF), we performed a genome-wide association study (GWAS). Utilizing longitudinal height and weight data, along with demographic information and covariates from 4,393 pwCF, we calculated AvgBMIz representing the average of per-quarter BMI Z scores. The GWAS incorporated 9.8M single nucleotide polymorphisms (SNPs) with a minor allele frequency (MAF) > 0.005 extracted from whole-genome sequencing (WGS) of each study subject. We observed genome-wide significant association with a variant in FTO (FaT mass and Obesity-associated gene; rs28567725; p value = 1.21e-08; MAF = 0.41, β = 0.106; n = 4,393 individuals) and a variant within ADAMTS5 (A Disintegrin And Metalloproteinase with ThromboSpondin motifs 5; rs162500; p value = 2.11e-10; MAF = 0.005, β = -0.768; n = 4,085 pancreatic-insufficient individuals). Notably, BMI-associated variants in ADAMTS5 occur on a haplotype that is much more common in African (AFR, MAF = 0.183) than European (EUR, MAF = 0.006) populations (1000 Genomes project). A polygenic risk score (PRS) calculated using 924 SNPs (excluding 17 in FTO) showed significant association with AvgBMIz (p value = 2.2e-16; r 2  = 0.03). Association between variants in FTO and the PRS correlation reveals similarities in the genetic architecture of BMI in CF and the general population. Inclusion of Black individuals in whom the single-gene disorder CF is much less common but genomic diversity is greater facilitated detection of association with variants that are in LD with functional SNPs in ADAMTS5. Our results illustrate the importance of population diversity, particularly when attempting to identify variants that manifest only under certain physiologic conditions., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Since January 2020, neonatal screening for cystic fibrosis (CF-NBS) has been implemented in the Wallonia-Brussels Federation. It's based on the immunoreactive trypsin (IRT1) assay between day 2 and day 4, associated with a 12 CFTR pathogenic variants analysis and with an IRT control on day 21. The aim of this study is to evaluate the performance of our CF-NBS in Liège according to the quality criteria defined by the European Cystic Fibrosis Society's working group on neonatal screening. After four years, 58.762 newborns have been screened. Nineteen children with cystic fibrosis were diagnosed : 14 by NBS, 3 following a meconium ileus, 1 by family history and 1 false negative diagnosed on clinical basis. Furthermore, 39 healthy carriers and 2 uncertain diagnosis (CFSPID) were identified. The sensitivity of CF NBS is 93,3 % (target ≥ 95 %), the positive predictive value (PPV) 17,7 % (target ≥ 30 %). Increasing the TIR1 threshold by 0,1 in 0,1 from P99 to P99,5, would be associated with a lower sensitivity and a non-significant improvement of PPV. A national assessment of CF NBS needs to be carried out.

Increased intestinal permeability is a manifestation of cystic fibrosis (CF) in people with CF (pwCF) and in CF mouse models. CF transmembrane conductance regulator knockout (Cftr KO) mouse intestine exhibits increased proliferation and Wnt/β-catenin signaling relative to wild-type mice (WT). Since the Rho GTPase Cdc42 plays a central role in intestinal epithelial proliferation and tight junction remodeling, we hypothesized that Cdc42 may be altered in the Cftr KO crypts. Immunofluorescence showed distinct tight junction localization of Cdc42 in Cftr KO fresh crypts and enteroids, the latter indicating an epithelial-autonomous feature. Quantitative PCR and immunoblots revealed similar expression of Cdc42 in the Cftr KO crypts/enteroids relative to WT, whereas pulldown assays showed increased GTP-bound (active) Cdc42 in proportion to total Cdc42 in Cftr KO enteroids. Cdc42 activity in the Cftr KO and WT enteroids could be reduced by inhibition of the Wnt transducer Disheveled. With the use of a dye permeability assay, Cftr KO enteroids exhibited increased paracellular permeability to 3 kDa dextran relative to WT. Leak permeability and Cdc42 tight junction localization were reduced to a greater extent by inhibition of Wnt/β-catenin signaling with endo-IWR1 in Cftr KO relative to WT enteroids. Increased proliferation or inhibition of Cdc42 activity with ML141 in WT enteroids had no effect on permeability. In contrast, inhibition of Cdc42 with ML141 increased permeability to both 3 kDa dextran and tight junction impermeant 500 kDa dextran in Cftr KO enteroids. These data suggest that increased constitutive Cdc42 activity may alter the stability of paracellular permeability in Cftr KO crypt epithelium. NEW & NOTEWORTHY Increased tight junction localization and GTP-bound activity of the Rho GTPase Cdc42 was identified in small intestinal crypts and enteroids of cystic fibrosis (CF) transmembrane conductance regulator knockout (Cftr KO) mice. The increase in epithelial Cdc42 activity was associated with increased Wnt signaling. Paracellular flux of an uncharged solute (3 kDa dextran) in Cftr KO enteroids indicated a moderate leak permeability under basal conditions that was strongly exacerbated by Cdc42 inhibition. These findings suggest increased activity of Cdc42 in the Cftr KO intestine underlies alterations in intestinal permeability.

Context.—: Existing targeted cystic fibrosis screening assays miss important pathogenic CFTR variants in the ethnically diverse US population., Objective.—: To evaluate the analytic performance of a multiplex polymerase chain reaction (PCR)/capillary electrophoresis (CE) CFTR assay panel that simultaneously interrogates primary pathogenic variants of different ethnic/ancestral groups., Design.—: Performance characteristic assessment and variant coverage comparison of the panel with a focus on ethnicity-specific CFTR variants were performed. Sample DNA was primarily from whole blood or cell lines. Detection of CFTR carriers was compared across several commercially available CFTR kits and recommended variant sets based on panel content., Results.—: The panel interrogated 65 pathogenic CFTR variants representing 92% coverage from a recent genomic sequencing survey of the US population, including 4 variants with top 5 frequency in African or Asian populations not reflected in other targeted panels. In simulation studies, the panel represented 95% of carriers across the global population, resulting in a 6.9% to 19.0% higher carrier detection rate compared with 10 targeted panels or variant sets. Precision and sensitivity/specificity were 100% concordant. Multisite sample-level genotyping accuracy was 99.2%. Across PCR and CE instruments, sample-level genotyping accuracy was 97.1%, with greater than 99% agreement for all variant-level metrics., Conclusions.—: The CFTR assay achieves 92% or higher coverage of CFTR variants in diverse populations and provides improved pan-ethnic coverage of minority subgroups of the US populace. The assay can be completed within 5 hours from DNA sample to genotype, and performance data exceed acceptance criteria for analytic metrics. This assay panel content may help address gaps in ancestry-specific CFTR genotypes while providing a streamlined procedure with rapidly generated results., Competing Interests: Hall, Milligan, Kelnar, Hallmark, Ashton, Parker, Filipovic-Sadic, and Latham are employees of Asuragen and report owning company stock/options. The other authors have no relevant financial interest in the products or companies described in this article., (© 2024 College of American Pathologists.)

Sweat chloride concentration, a diagnostic feature in cystic fibrosis (CF), reflects CF transmembrane conductance regulator (CFTR) activity. CFTR modulator therapies, especially elexacaftor/tezacaftor/ivacaftor (ETI), has improved CF outcomes. We report nationwide, real-world data on sweat chloride concentration in people with CF (pwCF) with and without modulator therapies. All Danish pwCF with a minimum of one F508del allele were included. Sweat chloride measurements were stratified by genotype and modulator treatment. Differences were assessed using mixed-effects models. We included 977 sweat chloride measurements from 430 pwCF, 71% of which were F508del homozygous. Heterozygous and homozygous ETI-treated pwCF had an estimated mean sweat chloride concentration of 43 mmol/L (95% confidence interval: 39; 48) and 43 mmol/L (39; 47), respectively-48% and 59% lower than those without treatment. High variation in concentrations remained regardless of treatment status. Despite ETI treatment, 27% heterozygous and 23% homozygous pwCF had elevated concentrations (≥60 mmol/L). These real-world data confirm a substantial decrease in sweat chloride concentration during modulator treatment, especially ETI, where mean concentrations halved. However, large variation remained, including persistently high concentrations. These findings emphasize the potential of sweat chloride concentration as a treatment response biomarker and the need to explore its heterogeneity and relationship with clinical outcomes., (© 2024 The Author(s). APMIS published by John Wiley & Sons Ltd on behalf of Scandinavian Societies for Pathology, Medical Microbiology and Immunology.)

Cystic fibrosis-related diabetes (CFRD), the most common comorbidity in cystic fibrosis (CF), leads to increased mortality by accelerating the decline in lung function. Scnn1b- Tg transgenic mice overexpressing the epithelial sodium channel β subunit exhibit spontaneous CF-like lung disease, including airway mucus obstruction and chronic inflammation. Here, we established a chronic CFRD-like model using Scnn1b- Tg mice made diabetic by injection of streptozotocin (STZ). In Ussing chamber recordings of the trachea, Scnn1b- Tg mice exhibited larger amiloride-sensitive currents and forskolin-activated currents, without a difference in adenosine triphosphate (ATP)-activated currents compared with wild-type (WT) littermates. Both diabetic WT (WT-D) and diabetic Scnn1b- Tg ( Scnn1b -Tg-D) mice on the same genetic background exhibited substantially elevated blood glucose at 8 wk; glucose levels also were elevated in bronchoalveolar lavage fluid (BALF). Bulk lung RNA-seq data showed significant differences between WT-D and Scnn1b -Tg-D mice. Neutrophil counts in BALF were substantially increased in Scnn1b- Tg-D lungs compared with controls ( Scnn1b -Tg-con) and compared with WT-D lungs. Lung histology data showed enhanced parenchymal destruction, alveolar wall thickening, and neutrophilic infiltration in Scnn1b- Tg-D mice compared with WT-D mice, consistent with the development of a spontaneous lung infection. We intranasally administered Pseudomonas aeruginosa to induce lung infection in these mice for 24 h, which led to severe lung leukocytic infiltration and an increase in pro-inflammatory cytokine levels in the BALF. In summary, we established a chronic CFRD-like lung mouse model using the Scnn1b -Tg mice. The model can be used for future studies toward understanding the mechanisms underlying the lung pathophysiology associated with CFRD and developing novel therapeutics. NEW & NOTEWORTHY We established a chronic CFRD-like mouse model using the Scnn1b -Tg transgenic mice overexpressing the epithelial sodium channel β subunit made diabetic by injection of streptozotocin. The results underscore the urgent need to develop novel therapeutics for CF lung disease.

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene result in cystic fibrosis, a multiorgan disease characterized by aberrant epithelial cell fluid secretion. Recent studies describe a connection between CFTR malfunctioning and a heightened susceptibility to colorectal cancer (CRC). Here, we outline these links and suggest directions for further studies., Competing Interests: Declaration of interests J.M.B. is an inventor on patents related to the FIS-assay and received financial royalties from 2017 onward. J.M.B reports receiving research grants and consultancy fees from various industries, including Vertex Pharmaceuticals, Proteostasis Therapeutics, Eloxx Pharmaceuticals, Teva Pharmaceutical Industries, and Galapagos outside the submitted work. G.H.K. reports research grants from Vertex Pharmaceuticals, GSK, TEVA, Ubbo Emmius Foundation, EU, Lung Foundation Netherlands (Money to institution), outside the submitted work. All other authors have nothing to disclose., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Mucocele formation in dogs is a unique and enigmatic muco-obstructive disease of the gallbladder caused by the amassment of abnormal mucus that bears striking pathological similarity to cystic fibrosis. We investigated the role of cystic fibrosis transmembrane conductance regulatory protein (CFTR) in the pathogenesis of this disease. The location and frequency of disease-associated variants in the coding region of CFTR were compared using whole genome sequence data from 2,642 dogs representing breeds at low-risk, high-risk, or with confirmed disease. Expression, localization, and ion transport activity of CFTR were quantified in control and mucocele gallbladders by NanoString, Western blotting, immunofluorescence imaging, and studies in Ussing chambers. Our results establish a significant loss of CFTR-dependent anion secretion by mucocele gallbladder mucosa. A significantly lower quantity of CFTR protein was demonstrated relative to E-cadherin in mucocele compared with control gallbladder mucosa. Immunofluorescence identified CFTR along the apical membrane of epithelial cells in control gallbladders but not in mucocele gallbladder epithelium. Decreases in mRNA copy number for CFTR were accompanied by decreases in mRNA for the Cl - /[Formula: see text] exchanger SLC26A3 , K + channels ( KCNQ1 , KCNN4 ), and vasoactive intestinal polypeptide receptor ( VIPR1 ), which suggest a driving force for change in secretory function of gallbladder epithelial cells in the pathogenesis of mucocele formation. There were no significant differences in CFTR gene variant frequency, type, or predicted impact comparing low-risk, high-risk, and definitively diagnosed groups of dogs. This study describes a unique, naturally occurring muco-obstructive disease of the canine gallbladder, with uncanny similarity to cystic fibrosis, and driven by the underlying failure of CFTR function. NEW & NOTEWORTHY Cystic fibrosis transmembrane conductance regulatory protein (CFTR) genomic variants and expression of mRNA, protein, and electrogenic anion secretory activity of CFTR were characterized in dog gallbladder. Acquired inhibition of CFTR expression by gallbladder epithelium was identified as underpinning a naturally occurring muco-obstructive disease of the dog gallbladder that bears striking pathological similarity to animal models of cystic fibrosis.

Cystic fibrosis (CF) is a genetic disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator ( CFTR ) gene, resulting in defective chloride ion channels. This leads to thick, dehydrated mucus that severely disrupts mucociliary clearance in the respiratory system and triggers infection that eventually is the cause of death of CF patients. Current therapeutic strategies primarily focus on restoring CFTR function, blocking epithelial sodium channels to prevent mucus dehydration, or directly targeting mucus to reduce its viscosity. Among the ion channels expressed in ciliated bronchial epithelial cells, the transient receptor potential vanilloid 4 (TRPV4) channel emerges as a significant channel in CF pathogenesis. Activation of TRPV4 channels affects the regulation of airway surface liquid by modulating sodium absorption and intracellular calcium levels, which indirectly influences CFTR activity. TRPV4 is also involved in the regulatory volume decrease (RVD) process and enhances inflammatory responses in CF patients. Here, we combine current findings on TRPV4 channel modulation as a promising therapeutic approach for CF. Although limited studies have directly explored TRPV4 in CF, emerging evidence indicates that TRPV4 activation can significantly impact key pathological processes in the disease. Further investigation into TRPV4 modulators could lead to innovative treatments that alleviate severe respiratory complications and improve outcomes for CF patients.

Background: Cystic Fibrosis (CF) is a monogenic disease caused by mutations in the gene coding the Cystic Fibrosis Transmembrane Regulator (CFTR) protein, but its overall physio-pathology cannot be solely explained by the loss of the CFTR chloride channel function. Indeed, CFTR belongs to a yet not fully deciphered network of proteins participating in various signalling pathways., Methods: We propose a systems biology approach to study how the absence of the CFTR protein at the membrane leads to perturbation of these pathways, resulting in a panel of deleterious CF cellular phenotypes., Results: Based on publicly available transcriptomic datasets, we built and analyzed a CF network that recapitulates signalling dysregulations. The CF network topology and its resulting phenotypes were found to be consistent with CF pathology., Conclusion: Analysis of the network topology highlighted a few proteins that may initiate the propagation of dysregulations, those that trigger CF cellular phenotypes, and suggested several candidate therapeutic targets. Although our research is focused on CF, the global approach proposed in the present paper could also be followed to study other rare monogenic diseases., (© 2024. The Author(s).)

Complex alleles of the CFTR gene complicate the diagnosis of cystic fibrosis (CF), the classification of its pathogenic variants, affect the clinical picture of the disease and can affect the efficiency of targeted drugs. The total frequency of complex allele [L467F;F508del] in the Russian population of patients with CF is 0.74%, and in patients with the F508del/F508del genotype, its frequency reaches 8%. This article presents multi-faceted study of the complex allele [L467F;F508del] in a cohort of patients with genotypes [L467F;F508del]/class I (c.3532_3535dup, c.1766+2T>C, W1310X, 712-1G>T), and data for a unique patient with the genotype [L467F;F508del]/[L467F;F508del]. Using the intestinal current measurement method, it was demonstrated the absence of CFTR function for [L467F;F508del]/class I and [L467F;F508del]/[L467F;F508del] genotypes. In intestinal organoids, it was shown that [L467F;F508del] in combination with class I variants and in the homozygotes abolishes the efficacy of both two-component (ivacaftor+lumacaftor; ivacaftor+tezacaftor) and three-component (ivacaftor+tezacaftor+elexacaftor) targeted drugs. When prescribing ivacaftor+tezacaftor+elexacaftor to three patients, they did not have a clinical effect after 6-12 months.

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.

Cystic fibrosis (CF) is a genetic disease caused by variants in the gene encoding for the CF transmembrane conductance regulator (CFTR) protein, a chloride and bicarbonate channel. CFTR dysfunction results in a multiorgan disease with the main clinical features being exocrine pancreatic insufficiency and diffuse bronchiectasis with chronic airway infection leading to respiratory failure and premature death. Over the past decades, major progress has been made by implementing multidisciplinary care, including nutritional support, airway clearance techniques and antibiotics in specialised CF centres. The past decade has further seen the progressive development of oral medications, called CFTR modulators, for which around 80% of people with CF are genetically eligible in Europe. CFTR modulators partially restore ion transport and lead to a rapid and major improvement in clinical manifestations and lung function, presumably resulting in longer survival. CFTR modulators have been game-changing in the care of people with CF. However, many questions remain unanswered, such as the long-term effects of CFTR modulators, especially when treatment is started very early in life, or the new CF-related disease emerging due to CFTR modulators. Moreover, severe complications of CF, such as diabetes or cirrhosis, are not reversed on CFTR modulators and around 20% of people with CF bear CFTR variants leading to a CFTR protein that is unresponsive to CFTR modulators. Challenges also arise in adapting CF care to a changing disease. In this review article, we highlight the new questions and challenges emerging from this revolution in CF care., Competing Interests: Conflict of interest: I. Fajac reports grants from AbbVie, Bayer, Boehringer Ingelheim, Insmed, GSK, Vertex Pharmaceuticals and Zambon; consulting fees from AbbVie, Boehringer Ingelheim, Genvade, Kither Biotech and Vertex Pharmaceuticals; lecture honoraria from Vertex Pharmaceuticals; and a leadership role as President of the European Cystic Fibrosis Society, outside the submitted work. P-R. Burgel reports grants from Vertex Pharmaceuticals and GSK; consulting fees from AstraZeneca, Chiesi, GSK, Insmed, Vertex, Viatris and Zambon; and travel support from AstraZeneca and Chiesi, outside the submitted work. C. Martin reports lecture honoraria from AstraZeneca, Chiesi and Zambon; travel support from Chiesi, Sanofi and Zambon; and advisory board participation with Vertex, Zambon and GSK, outside the submitted work., (Copyright ©The authors 2024.)

Since screening for cystic fibrosis (CF) was incorporated into the newborn screening program, the number of recognised variants in the CF transmembrane conductance regulator (CFTR ) gene has significantly increased. This has led to the discovery of combinations of gene variants with an uncertain prognosis. One outcome is the designation of 'cystic fibrosis screen positive inconclusive diagnosis' (CFSPID). While the majority of these children are expected to be unaffected by their CFTR variants, a small proportion have been seen to develop symptoms or increasing sweat chloride levels over time, which may reflect dysfunction of the CFTR protein.As the number of children with CFSPID increases, paediatricians and those working in primary care are more likely to encounter them in their practice. It is important that professionals have an understanding of CFSPID: what it is and, importantly, what it is not (ie, they do not have CF). In this article, we hope to explore this using some example cases, illustrating the ways in which these children may present symptomatically and how to manage them., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. No commercial re-use. See rights and permissions. Published by BMJ.)

Two cystic fibrosis (CF) rat models, one carrying the common Phe508del mutation and the other a nonsense cystic fibrosis transmembrane conductance regulator (CFTR) mutation (knockout) were previously characterised. Although relevant CFTR mRNA reductions were present in the lung, no overt CF lung disease was observed. This study used flexiVent lung mechanic assessment and regional ventilation assessment via X-ray velocimetry (XV) functional imaging to assess the lung phenotype in both models. To determine the sensitivity of XV regional ventilation imaging, the effect of a localised physical obstruction (delivery of agar beads to part of the lungs) on lung ventilation was examined. At baseline, Phe508del and knockout CF rats had a lower inspiratory capacity, total respiratory system compliance, and static compliance than wildtype rats. Following agar bead delivery all XV ventilation parameters were altered, with substantial increases in poorly ventilated regions and ventilation heterogeneity. XV ventilation maps accurately identified locations of bead-induced airflow changes. Despite unremarkable lung histopathology, this study indicated that CF rats display altered respiratory mechanics, with CF rats needing to exert additional effort to expand and deflate their lungs due to increased stiffness. This study demonstrated the utility of XV imaging providing spatial lung ventilation information., (© 2024. The Author(s).)

Background: Predictions based on patient-derived materials of CFTR modulators efficacy have been performed lately in patient-derived cells, extending FDA-approved drugs for CF patients harboring rare variants. Here we developed intestinal organoids from subjects carrying S737F- and T465N-CFTR in trans with null alleles to evaluate their functional impact on CFTR protein function and their restoration upon CFTR modulator treatment. The characterization of S737F-CFTR was performed in two subjects recently assessed in nasal epithelial cells but not in colonoids., Results: Our functional analysis (Ussing chamber) confirmed that S737F-CFTR is a mild variant with residual function as investigated in colonoids of patients with S737F/Dele22-24 and S737F/W1282X genotypes. An increase of current upon Elexacaftor/Tezacaftor/Ivacaftor (ETI) treatment was recorded for the former genotype. T465N is a poorly characterized missense variant that strongly impacts CFTR function, as almost no CFTR-mediated anion secretion was registered for T465N/Q39X colonoids. ETI treatment substantially improved CFTR-mediated anion secretion and increased the rescue of mature CFTR expression compared to either untreated colonoids or to dual CFTR modulator therapies., Conclusions: Our study confirms the presence of a residual function of the S737F variant and its limited response to CFTR modulators while predicting for the first time the potential clinical benefit of Trikafta® for patients carrying the rare T465N variant., (© 2024. The Author(s).)

A fundamental challenge for cystic fibrosis (CF) gene therapy is ensuring sufficient transduction of airway epithelia to achieve therapeutic correction. Hypertonic saline (HTS) is frequently administered to people with CF to enhance mucus clearance. HTS transiently disrupts epithelial cell tight junctions, but its ability to improve gene transfer has not been investigated. Here, we asked if increasing the concentration of NaCl enhances the transduction efficiency of three gene therapy vectors: adenovirus, AAV, and lentiviral vectors. Vectors formulated with 3-7% NaCl exhibited markedly increased transduction for all three platforms, leading to anion channel correction in primary cultures of human CF epithelial cells and enhanced gene transfer in mouse and pig airways in vivo. The mechanism of transduction enhancement involved tonicity but not osmolarity or pH. Formulating vectors with a high ionic strength solution is a simple strategy to greatly enhance efficacy and immediately improve preclinical or clinical applications., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Cystic fibrosis (CF) is inherited by CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations. A variety of mutations have been identified in the CFTR gene that may be associated with cystic fibrosis, and these mutations demonstrate extensive molecular genetic heterogeneity in this disease. Little is known about the molecular mechanism by which mutations affect CFTR function, and only a minority of mutations have been characterized by functional studies. There has been an increase in the number of complex alleles. This may partly explain the difficulty in establishing genotype-phenotype correlations and complicate genetic counseling and diagnosis in some cases. Therefore, the identification of complex alleles has several important implications for recessive disorders. This will facilitate diagnosis; improve judgements concerning prognosis, and enable appropriate genetic counselling for affected families. This review describes the complex cystic fibrosis allele to better understand the contribution of this allele in the wide phenotypic variability of cystic fibrosis disease. It occurs in the complex allele that the second cis mutation can modulate the effects of the first mutation or vice versa. The phenotypic variability between CF or CFTR-RD (CFTR related disease) patients may be due to several factors, including different genetic and environmental backgrounds. It is important to determine the allele complex so that optimal treatment can be established.

The CFTR modulator Trikafta has markedly improved lung disease for Cystic Fibrosis (CF) patients carrying the common delta F508 (F508del-CFTR) CFTR mutation. F508del-CFTR results in an apical trafficking defect and loss of function in CFTR-expressing epithelial cells. However, Trikafta has not resulted in improved gastrointestinal function in CF patients. A humanized mouse model of F508del-CFTR was recently generated to evaluate CFTR modulators and other compounds to treat human F508del-CFTR CF intestinal disease. Short-term (4 h) treatment of rats with Dexamethasone (Dex) potently activates serum glucocorticoid kinase 1 (SGK1) and increases CFTR apical traffic and ion transport in the native intestine. This study examined CFTR localization and ion transport in intestinal segments from humanized F508del-CFTR mice following treatment with Dex in the presence/absence of Trikafta. Dex treatment improved apical CFTR localization and function but was inconsistent along intestinal segments. Combined treatment with Dex and Trikafta was superior to Dex alone but inconsistently improved CFTR localization and function. These data suggest further optimization of humanized CF mouse models will be necessary to test the efficacy of compounds to treat human CF intestinal disease., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Nadia Ameen reports financial support was provided by Cystic Fibrosis Foundation. Nadia Ameen reports financial support was provided by National Institutes of Health. Nadia Ameen reports a relationship with Cystic Fibrosis Foundation that includes: funding grants. Nadia Ameen reports a relationship with National Institutes of Health that includes: funding grants. Nadia Ameen has patent N/A pending to N/A. The authors have expressed no conflicts of interest, and the manuscript has not been published previously nor is being considered concurrently by another publication. All authors have read and approved it. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: We recently demonstrated that elexacaftor/tezacaftor/ivacaftor (ETI) improves the lung clearance index (LCI) and abnormalities in lung morphology detected by magnetic resonance imaging (MRI) in adolescent and adult patients with cystic fibrosis (CF). However, real-world data on the effect of ETI on these sensitive outcomes of lung structure and function in school-age children with CF have not been reported. The aim of this study was therefore to examine the effect of ETI on the LCI and the lung MRI score in children aged 6-11 years with CF and one or two F508del alleles., Methods: This prospective, observational, multicentre, post-approval study assessed the longitudinal LCI up to 12 months and the lung MRI score before and 3 months after initiation of ETI., Results: A total of 107 children with CF including 40 heterozygous for F508del and a minimal function mutation (F/MF) and 67 homozygous for F508del (F/F) were enrolled in this study. Treatment with ETI improved the median (interquartile range (IQR)) LCI in F/MF (-1.0 (-2.0- -0.1); p<0.01) and F/F children (-0.8 (-1.9- -0.2); p<0.001) from 3 months onwards. Further, ETI improved the median (IQR) MRI global score in F/MF (-4.0 (-9.0-0.0); p<0.01) and F/F children (-3.5 (-7.3- -0.8); p<0.001)., Conclusions: ETI improves early abnormalities in lung ventilation and morphology in school-age children with CF and at least one F508del allele in a real-world setting. Our results support early initiation of ETI to reduce or even prevent lung disease progression in school-age children with CF., Competing Interests: Conflicts of interest: M. Stahl, S.Y. Graeber and S. Thee are participants of the Berlin Institute of Health (BIH)-Charité Clinician Scientist Program, and J. Röhmel is participant of the Case Analysis and Decision Support (CADS) program funded by Charité – Universitätsmedizin Berlin and the BIH. M. Stahl reports an Independent Research Innovation Award and honoraria for lectures and participation in advisory boards, all by Vertex Pharmaceuticals Incorporated, outside of the submitted work; she is Chairman of the German CF Research Council (FGM), Treasurer of the German Society of Paediatric Pulmonology (GPP) and was Secretary of the Group CF of the Paediatric Assembly of the ERS. M. Dohna is a participant of the Ellen-Schmidt Habilitationsförderung funded by the Hannover Medical School. S.Y. Graeber reports grants from the German CF Foundation and Vertex Pharmaceuticals Incorporated, and honoraria from Chiesi GmbH and Vertex Pharmaceuticals Incorporated for lectures and participation in advisory boards, outside of the submitted work. O. Sommerburg reports grants and honoraria from Vertex Pharmaceuticals Incorporated for lectures, outside of the submitted work. S.T. Pallenberg is a member of the Else-Kröner Forschungskolleg TITUS. A. Voskrebenzev reports a grant and honoraria for lectures from Siemens Healthineers, outside of the submitted work, holds a patent for a method of quantitative magnetic resonance lung imaging (Voskrebenzev, Gutberlet, Vogel-Claussen; number EP3107066, US-2016-0367200-Al 22.12.2016), and is a stockholder and CEO of BioVisioneers GmbH. K. Schütz reports payments for attending meetings and/or travel from Vertex Pharmaceuticals Incorporated, outside of the submitted work. G. Hansen reports receipt of consultation fees from Sanofi GmbH, outside of the submitted work. F. Doellinger reports payment or honoraria for lectures, presentations, manuscript writing or educational events from Bayer, Bayer Vital, Berlin-Chemie Menarini, Boehringer Ingelheim and Chiesi GmbH, payment for expert testimony from Calyx, and support for attending meetings from Bayer. E. Steinke reports grants from Berlin Institute of Health at Charité Berlin, and payment or honoraria for lectures, presentations, manuscript writing or educational events from Vertex Pharmaceuticals Incorporated. S. Thee reports honoraria for lectures and payment for attending meetings and/or travel from Vertex Pharmaceuticals Incorporated and Viatris, outside of the submitted work. J. Röhmel reports honoraria for lectures from Vertex Pharmaceuticals Incorporated, outside the submitted work; additionally, he is work package leader in BEAT-PCD (ERS-CRC). M.O. Wielpütz reports a grant from Vertex Pharmaceuticals Incorporated, and receipt of consulting fees and honoraria for lectures from Vertex Pharmaceuticals Incorporated and Boehringer Ingelheim, outside of the submitted work. L. Naehrlich reports receipt of fees for a data quality project of the German CF Registry. He is the medical lead of the German CF Registry, the pharmacovigilance study manager of the European Cystic Fibrosis Society Patient Registry and part of the Trial Steering Committee for CF STORM. He also reports grants from the German Center for Lung Research, Vertex Pharmaceuticals and Mukoviszidose Institute, and receipt of medical writing services from Articulate Science. J. Vogel-Claussen reports grants from BMBF, Siemens Healthineers, AstraZeneca, Boehringer Ingelheim and GSK, royalties or licenses from Siemens Healthineers, receipt of consulting fees from AstraZeneca, honoraria for lectures from Siemens Healthineers, AstraZeneca, Boehringer Ingelheim, GSK, Roche, Coreline Soft and Bayer, payments for attending meetings and/or travel from Vertex Pharmaceuticals Incorporated, Bayer, GSK and AstraZeneca, and holds a patent for a method of quantitative magnetic resonance lung imaging (Voskrebenzev, Gutberlet, Vogel-Claussen; number EP3107066, US-2016-0367200-Al 22.12.2016). B. Tümmler reports support for the present study from Bundesministerium für Forschung und Technologie, grants from the German Research Foundation (DFG; CRC 900; Excellence cluster “RESIST”), consultancy fees from Helmholtz Institut für Infektionsforschung, payment or honoraria for lectures, presentations, manuscript writing or educational events from Vertex Pharmaceutical (Germany) Incorporated, participation on a data and safety monitoring board or advisory board with Vertex Pharmaceuticals Incorporated, and leadership roles with Christiane Herzog Stiftung and the Microbiome/Metagenome Group of the German Center for Lung Research (DZL). M.A. Mall reports grants from the German Research Foundation (DFG; SFB-TR 84, and project 450557679) and the German Innovation Fund (01NVF19008), outside of the submitted work. Additionally, he reports receipt of consulting fees from AbbVie, Antabio, Arrowhead, Boehringer Ingelheim, Enterprise Therapeutics, Kither Biotec, Prieris, Recode, Santhera, Splisense and Vertex Pharmaceuticals Incorporated, of honoraria for lectures from Vertex Pharmaceuticals Incorporated and participation in advisory boards from AbbVie, Antabio, Arrowhead, Boehringer Ingelheim, Enterprise Therapeutics, Kither Biotec, Pari and Vertex Pharmaceuticals Incorporated, and of payment for travel from Vertex Pharmaceuticals Incorporated and Boehringer Ingelheim, all outside of the submitted work. He is a Fellow of ERS (FERS). A-M. Dittrich reports support for the present study from the German Center for Lung Research (DZL), Vertex Pharmaceuticals Incorporated and European Cystic Fibrosis Society Clinical Trial Network (ECFS-CTN), grants from Vertex Pharmaceuticals Incorporated, ECFS-CTN, DFG and Christiane Herzog Stiftung, consultancy fees from the c4c consortium, GSK and European Cystic Fibrosis Society. The remaining authors have no potential conflicts of interest to disclose., (Copyright ©The authors 2024.)

Just over thirty years is the span of a generation. It is also the time that has passed since the discovery of the gene responsible for cystic fibrosis. Today, it is safe to say that this discovery has revolutionized our understanding, research perspectives, and management of this disease, which was, thirty years ago, a pediatric condition with a grim prognosis. The aim of this review is to present the advances that science and medicine have brought to our understanding of the pathophysiology of the disease and its management, which in many ways, epitomizes modern molecular genetic research. Since the discovery of the cystic fibrosis transmembrane conductance regulator (CFTR) gene in 1989, modeling the CFTR protein, deciphering its function as an ion channel, and identifying its molecular partners have led to numerous therapeutic advances. The most significant advancement in this field has been the discovery of protein modulators that can target its membrane localization and chloride channel activity. However, further progress is needed to ensure that all patients can benefit from a therapy tailored to their mutations, with the primary challenge being the development of treatments for mutations leading to a complete absence of the protein. The present review delves into the history of the multifaceted world of CF, covering main historical facts, current landscape, clinical management, emerging therapies, patient perspectives, and the importance of ongoing research, bridging science and medicine in the fight against the disease.

Background & Aims: Cystic fibrosis (CF) is considered a multisystemic disorder in which CF-associated liver disease (CFLD) is the third most common cause of mortality. Currently, no effective treatment is available for CFLD because its pathophysiology is still unclear. Interestingly, CFLD exhibits identical vascular characteristics as non-cirrhotic portal hypertension, recently classified as porto-sinusoidal vascular disorders (PSVD)., Methods: Since endothelial cells (ECs) are an important component in PSVD, we performed single-cell RNA sequencing (scRNA-seq) on four explant livers from CFLD patients to identify differential endothelial characteristics which could contribute to the disease. We comprehensively characterized the endothelial compartment and compared it with publicly available scRNA-seq datasets from cirrhotic and healthy livers. Key gene signatures were validated ex vivo on patient tissues., Results: We found that ECs from CF liver explants are more closely related to healthy than cirrhotic patients. In CF patients we also discovered a distinct population of liver sinusoidal ECs-coined CF LSECs-upregulating genes involved in the complement cascade and coagulation. Finally, our immunostainings further validated the predominant periportal location of CF LSECs., Conclusions: Our work showed novel aspects of human liver ECs at the single-cell level thereby supporting endothelial involvement in CFLD, and reinforcing the hypothesis that ECs could be a driver of PSVD. Therefore, considering the vascular compartment in CF and CFLD may help developing new therapeutic approaches for these diseases., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Lung function is a key outcome used in the evaluation of disease progression in cystic fibrosis. The variability of individual lung function measurements over time (within-individual variability) has been shown to predict subsequent lung function changes. Nevertheless, the association between within-individual lung function variability and demographic and genetic covariates has not been quantified., Methods: We performed a longitudinal analysis of data from a cohort of 7099 adults with cystic fibrosis (between 18 and 49 years old) from the UK cystic fibrosis registry, containing annual review data between 1996 and 2020. A mixed-effects location-scale model is used to quantify mean FEV 1 (forced expiratory volume in 1 s) trajectories and FEV 1 within-individual variability as a function of sex, age at annual review, diagnosis after first year of life, homozygous F508 genotype and birth cohort., Results: Mean FEV 1 decreased with age and lung function variability showed a near-quadratic trend by age. Males showed higher FEV 1 mean and variability than females across the whole age range. Earlier diagnosis and homozygous F508 genotype were also associated with higher FEV 1 mean and variability. Individuals who died during follow-up showed on average higher lung function variability than those who survived., Conclusions: Key variables known to be linked with mean lung function in cystic fibrosis are also associated with an individual's lung function variability. This work opens new avenues to understand the role played by lung function variability in disease progression and its utility in predicting key outcomes such as mortality., Competing Interests: Declaration of competing interest JKB has received research funding for unrelated work from F. Hoffmann-La Roche Ltd., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Objectives: Aim of this study was to identify risk factors for a progression to cystic fibrosis (CF) in individuals detected as CF Screening Positive, Inconclusive Diagnosis (CFSPID)., Methods: This is a systematic review through literature databases (2015-2023). Blood immunoreactive trypsinogen (b-IRT) values, CFTR genotype, sweat chloride (SC) values, isolation of Pseudomonas aeruginosa (Pa) from respiratory samples, Lung Clearance Index (LCI) values in CFSPIDs who converted to CF (CFSPID > CF) and age at CF transition were assessed., Results: Percentage of CFSPID > CF varies from 5.3 % to 44 %. Presence of one CF-causing CFTR variant in trans with a variant with variable clinical consequences (VVCC), an initial SC ≥ 40 mmol/L, an increase of SC > 2.5 mmol/L/year and recurrent isolation of pseudomonas aeruginosa (Pa) from airway samples could allow identification of subjects at risk of progression to CF., Conclusions: CFSPIDs with CF causing variant/VVCC genotype and first SC in the higher borderline range may require more frequent and prolonged clinical follow-up., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Access to elexacaftor/tezacaftor/ivacaftor (ETI) for people with cystic fibrosis (PwCF) without a F508del variant is limited due to lack of clinical data supporting efficacy., Methods: In this systematic review and meta-analysis, we examined patient-level data from studies reporting the clinical response to ETI for PwCF with non-F508del CFTR variants. We searched electronic data sources including Embase, MEDLINE, and CENTRAL from January 1st, 2019 to May 14th, 2024., Findings: Our search results identified 4,795 studies and 20 met the eligibility criteria. 120 of 164 (73 %) individuals had a positive clinical response to ETI, defined by a sweat chloride (SwCl) decrease of ≥10 mmol/L or percent-predicted FEV1 (ppFEV1) improvement of ≥5 %. 51 unique ETI-responsive variants were represented across these 120 individuals and 27 of these variants (53 %) have not been previously approved by the U.S. FDA. For variants with at least 10 individuals treated with ETI to date, a consistent positive clinical response was observed for N1303K and G85E. For N1303K (n = 48), the median increase in ppFEV1 was 16 % (IQR: 8 %, 29 %), with a median decrease in SwCl of -9 (IQR: -4, -22) mmol/L. For G85E (n = 16), the median increase in ppFEV1 was 13.5 % (IQR: 8 %, 19 %) with a median decrease in SwCl of -46 (IQR: -39, -66) mmol/L., Conclusion: Additional ETI-responsive variants were identified following a comprehensive review of ETI clinical use in PwCF without F508del. This data can be used by the CF community in efforts to expand the labelled indications or to help advocate for off-label ETI reimbursement., Competing Interests: Declaration of competing interest BSQ has participated in Vertex Pharmaceuticals sponsored clinical trials as site PI and has received honoraria for speaking engagements organized by Vertex Pharmaceuticals., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: With advancements in CF drug development, people with cystic fibrosis (PwCF) now take a median of seven medications daily, increasing treatment complexity, risk of drug therapy problems (DTPs), and interference with treatment goals. Given that some of these DTPs can be prevented with preemptive pharmacogenetic testing, the overall goal of this study was to test the clinical utility of a multi-gene pharmacogenetics (PGx) panel in potentially reducing DTPs in PwCF., Methods: A population based retrospective study of patients with CF was conducted at the University of Utah Health Care System. The patients were genotyped for CYP450 enzymes using a pharmacogenomic assay, and their drug utilization information was obtained retrospectively. This pharmacogenomic information was combined with clinical guidelines to predict the number of actionable PGx interventions in this patient cohort., Results: A total of 52 patients were included in this study. In the patient sample, a minimum of one order of actionable PGx medication was observed in 75 % of the cases. Results revealed that 4.2 treatment modifications per 10 patients can be enabled with the help of a PGx intervention in this patient population. Additionally, our findings suggest that polymorphisms in CYP2D6 and CYP2C19 are most likely to be the primary contributors to DTP's within PwCF., Conclusion: This study provides evidence that the PGx panel has the potential to help alleviate the clinical burden of DTPs in PwCF and can assist in informing pharmacotherapy recommendations. Future research should validate these findings and evaluate which subgroups of PwCF would most benefit from pharmacogenetic testing., Competing Interests: Declaration of competing interest Anindit Chhibber reports no potential conflict of interest. Hollyann Jacobs reports no potential conflict of interest. Alexandre Hikiji Watanabe reports no potential conflict of interest. Aditi Kharat reports no potential conflict of interest. Sakil M. Syeed reports no potential conflict of interest. Catherine M. Sherwin reports no potential conflict of interest. Nathorn Chaiyakunapruk reports no potential conflict of interest. Joseph Biskupiak reports no potential conflict of interest. Venkata K. Yellepeddi reports no potential conflict of interest. Diana Brixner reports no potential conflict of interest. David C. Young reports no potential conflict of interest., (Copyright © 2024 European Cystic Fibrosis Society. Published by Elsevier B.V. All rights reserved.)

Purpose of Review: To provide a comprehensive overview of the underlying genetic defects of pulmonary (vascular) diseases and novel treatment avenues., Recent Findings: Pulmonary arterial hypertension (PAH) is the prime example of a pulmonary vascular disease, which can be caused by genetic mutations in some patients. Germline mutations in the BMPR2 gene and further genes lead to vessel remodelling, increase of pulmonary vascular resistance and onset of heritable PAH. The PAH genes with the highest evidence and strategies for genetic testing and counselling have been assessed and evaluated in 2023 by international expert consortia. Moreover, first treatment options have just arisen targeting the molecular basis of PAH., Summary: Apart from PAH, this review touches on the underlying genetic causes of further lung diseases including alpha 1 antitrypsin deficiency, cystic fibrosis, familial pulmonary fibrosis and lymphangioleiomyomatosis. We point out the main disease genes, the underlying pathomechanisms and novel therapies trying not only to relieve symptoms but to treat the molecular causes of the diseases., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Expanded carrier screening (ECS) is a genetic screening test carried out by analysing a blood sample. This screen can be used to detect whether the individual unknowingly carries gene variants associated with common genetic conditions, such as cystic fibrosis, that may be passed on to their children. It is typically performed in reproductive medicine for those who are considering having a family either naturally or via fertility treatment. Many donor sperm and egg banks, particularly in the USA and Europe, also perform blanket ECS testing on all their prospective sperm and egg donors. ECS is not currently routine practice in the UK, but a growing number of patients are requesting it before treatment. All of us carry gene variants of some sort that may cause autosomal recessive disease in their children if their partner or donor also carry a variant in the same gene. An autosomal recessive disease means two copies of an abnormal gene must be present in order for the disease or trait (such as cystic fibrosis or sickle cell disease) to develop. One copy of the variant means the person is a carrier but does not have the condition. Two copies, i.e. from the mother and father, means the child has a 25% chance of having the genetic disease. Carrying a gene variant does not mean that the individual would necessarily have any symptoms of the disease or any features of the condition. Genetic tests for specific conditions are currently available either before or during pregnancy for prospective parents who have a family or personal history of a genetic condition, or for those from ethnic backgrounds where certain conditions - such as haemoglobinopathies (blood disorders) - are common, prompting referral to a clinical genetics department. Expanded carrier screens may test for more than 100 genetic conditions. The list of conditions screened for is called a panel. Common panels are 250 or 600 genes. Not all expanded carrier screens that are available analyse the same genes. Some may test for genes that do not cause serious disease, or cause diseases that occur in later life; others test for genes that cause severe conditions in childhood. There is no agreement as to which panel of genes should be tested for in an ECS. Understanding the screening that is being offered, and the meaning of any results, is complicated and requires support from appropriately trained professionals to best inform the prospective parent or parents., (© 2024 Royal College of Obstetricians and Gynaecologists.)

Cystic fibrosis (CF) is caused by mutations in the gene encoding the CF transmembrane conductance regulator (CFTR) protein. Although CF affects multiple organs, lung disease is the main cause of morbidity and mortality, and gene therapy is expected to provide a mutation-agnostic option for treatment. SP-101 is a recombinant adeno-associated virus (AAV) gene therapy vector carrying a human CFTR minigene, hCFTRΔR , and is being investigated as an inhalation treatment for people with CF. To further understand SP-101 activity, in vitro studies were performed in human airway epithelia (HAE) derived from multiple CF and non-CF donors. SP-101 restored CFTR-mediated chloride conductance, measured via Ussing chamber assay, at a multiplicity of infection (MOI) as low as 5E2 in the presence of doxorubicin, a small molecule known to augment AAV transduction. Functional correction of CF HAE increased with increasing MOI and doxorubicin concentration and correlated with increasing cell-associated vector genomes and hCFTRΔR mRNA expression. Tropism studies using a fluorescent reporter vector and single-cell mRNA sequencing of SP-101-mediated hCFTRΔR mRNA demonstrated broad expression in all cell types after apical transduction, including secretory, ciliated, and basal cells. In summary, SP-101, particularly in combination with doxorubicin, shows promise for a novel CF treatment strategy and strongly supports continued development.

Cystic fibrosis (CF) is a serious genetic disease caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Approved small molecule therapies benefit the majority of people with CF (pwCF), but unfortunately not all. Gene addition offers a mutation agnostic treatment option for all pwCF. SP-101 is an adeno-associated virus gene therapy vector (AAV2.5T) that has been optimized for efficient human airway cell transduction, and that contains a functional and regulated shortened human CFTR minigene ( hCFTRΔR ) with a small synthetic promoter/enhancer. To understand SP-101 airway distribution, activity, and the associated immune response, in vivo studies were performed in wild-type and CF ferrets. After single dose inhaled delivery of SP-101, followed by single dose inhaled doxorubicin (an AAV transduction augmenter) or saline, SP-101 vector genomes were detected throughout the respiratory tract. hCFTRΔR mRNA expression was highest in ferrets also receiving doxorubicin and persisted for the duration of the study (13 weeks). Pre-existing mucus in the CF ferrets did not present a barrier to effective transduction. Binding and neutralizing antibodies to the AAV2.5T capsid were observed regardless of doxorubicin exposure. Only a portion of ferrets exhibited a weak T-cell response to AAV2.5T and no T-cell response was seen against hCFTRΔR. These data strongly support the continued development of inhaled SP-101, followed by inhaled doxorubicin, for the treatment of CF.

Background: Microbial dysbiosis has been linked to cystic fibrosis (CF); however, the composition of gut microbiota in adult CF patients in relation to severity of CF transmembrane conductance regulator (CFTR) gene mutation and nutritional status have not yet been explored. Study aimed to assess the gut microbiota composition in adults with CF, and its relationship with the severity of CFTR mutations, and BMI., Methods: Gut microbiota of 41 adults with CF, and 26 non-CF controls were compared using whole 16S rRNA gene sequencing. Differences in the microbial community between groups of patients classified according to the severity of CFTR mutations, and BMI were assessed. The alpha diversity, beta diversity, and taxa abundance were identified to reflect gut microbiota composition., Results: Results showed a significant decrease in alpha diversity of bacterial communities in CF compared to non-CF group, but no significant difference between the CF groups distinguished by the severity of CFTR mutations. However, more severe mutations were associated with the higher relative abundance of Bacteroides and Streptococcus and the lower relative abundance of Faecalibacterium and Blautia. Undernourished CF patients showed significantly lower alpha diversity compared to non-CF group and CF patients with BMI within the norm. Significant differences in the structure of the gut microbiota between CF and non-CF groups, as well as between BMI groups were also found., Conclusions: Our research indicates that CF is associated with alterations in gut microbiota in adults. Additionally, in adult CF patients, the composition of the gut microbiota is also related to BMI., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Gene therapy for CF has concentrated on targeting the lung. Here we took a different approach by injecting into the cephalic vein and spraying into the trachea of G551D, CF ferrets either AAV1 or 6 containing Δ27-264-CFTR, a truncated version of CFTR. Treatment with the potentiator VX-770 was halted for 7 days before instillation to induce a disease phenotype. Indeed, all ferrets were pancreas-insufficient when they entered the study. Four ferrets (three receiving AAV1 and one AAV6) were necropsied 48 days after vector delivery, and four (three receiving AAV6, one AAV1) were euthanized or died prior to the planned necropsy. AAV1 or AAV6 vector genomes, mRNA expression, and CFTR protein were detected in all tracheal and lung samples and in the liver, pancreas, and ileum of the treated ferrets. Surface and basal airway cells, pancreatic and bile ducts, and ileal crypts and villi were successfully transduced. Obstruction of the airways accompanied by pulmonary hemorrhaging, plugged pancreatic and bile ducts as well as mucous plugs in the ileum were noticed in untreated but absent from transduced ferrets necropsied at 48 days. Transduction of G551D ferrets suggests that a combination of systemic and airway application may be the preferred route of delivery for CF., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Hepatobiliary complications of Cystic Fibrosis (CF) constitute a significant burden for persons with CF of all ages, with advanced CF liver disease in particular representing a leading cause of mortality. The causes of the heterogeneity of clinical manifestations, ranging from steatosis to focal biliary cholestasis and biliary strictures, are poorly understood and likely reflect a variety of environmental and disease-modifying factors in the setting of underlying CFTR mutations. This review summarizes the current understanding of the pathophysiology of hepatobiliary manifestations of CF, and discusses emerging disease models and therapeutic approaches that hold promise to impact this important yet incompletely addressed aspect of CF care., (© 2024 Wiley Periodicals LLC.)

Cystic fibrosis-related hepatobiliary involvement (CFHBI) is a term used to describe a spectrum of hepatobiliary involvement ranging from a transient elevation of transaminase levels to advanced cystic fibrosis-associated liver disease (aCFLD). While CFHBI is common among people with cystic fibrosis (PwCF), aCFLD is rare impacting only approximately 5%-10% of the CF population. After respiratory/cardiorespiratory issues and transplant-related complications, aCFLD is now the 4th leading cause of mortality among PwCF. Additionally, aCFLD is an independent predictor of all-cause mortality and is associated with significant morbidity. Despite this recognition, our ability to predict those patients at greatest risk for aCFLD, identify early aCFLD, and monitor the incremental progression of CFHBI is lacking. Here, we review the strengths and weaknesses of the common biomarkers and imaging modalities used in the evaluation and monitoring of CFHBI, as well as the current understanding of genetic modifiers related to aCFLD., (© 2024 Wiley Periodicals LLC.)

Cystic fibrosis (CF) is characterized by chronic airway inflammation and premature aging. The link with leukocyte telomere length (LTL) as a marker of biological aging is unclear. We studied disease severity and LTL in 168 CF patients of which 85 patients had a second retrospective LTL assessment. A higher FEV 1 was associated with longer LTL, with a stronger effect in men (5.08% longer LTL) compared to women (0.41% longer LTL). A higher FEV 1 /FVC ratio was associated with 7.05% ( P =0.017) longer LTL in men. CF asthma, as defined by the treatment with inhaled corticosteroids, was associated with -6.65% shorter LTL ( P =0.028). Men homozygous for the ΔF508 genotype showed a -10.48% ( P =0.026) shorter LTL compared to heterozygotes. A genotype-specific non-linear association between LTL shortening and chronological age was observed. Stronger age-related LTL shortening was observed in patients homozygous for the ΔF508 genotype ( P -interaction= 0.044). This work showed that disease severity in CF patients negatively influences LTL, with slightly more pronounced effects in men. The homozygous genotype for ΔF508 may play a role in LTL attrition in CF patients. Understanding factors in CF patients that accelerate biological aging provides insights into mechanisms that can extend the overall life quality in CF-diseased.

Nonsense mutations account for >10% of human genetic disorders, including cystic fibrosis, Alagille syndrome, and Duchenne muscular dystrophy. A nonsense mutation results in the expression of a truncated protein, and therapeutic strategies aim to restore full-length protein expression. Most strategies under development, including small-molecule aminoglycosides, suppressor tRNAs, or the targeted degradation of termination factors, lack mRNA target selectivity and may poorly differentiate between nonsense and normal stop codons, resulting in off-target translation errors. Here, we demonstrate that antisense oligonucleotides can stimulate readthrough of disease-causing nonsense codons, resulting in high yields of full-length protein in mammalian cellular lysate. Readthrough efficiency depends on the sequence context near the stop codon and on the precise targeting position of an oligonucleotide, whose interaction with mRNA inhibits peptide release to promote readthrough. Readthrough-inducing antisense oligonucleotides (R-ASOs) enhance the potency of non-specific readthrough agents, including aminoglycoside G418 and suppressor tRNA, enabling a path toward target-specific readthrough of nonsense mutations in CFTR, JAG1, DMD, BRCA1 and other mutant genes. Finally, through systematic chemical engineering, we identify heavily modified fully functional R-ASO variants, enabling future therapeutic development., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Cystic fibrosis is a rare genetic disease caused by mutations in CFTR, the gene encoding cystic fibrosis transmembrane conductance regulator (CFTR). The discovery of CFTR in 1989 has enabled the unravelling of disease mechanisms and, more recently, the development of CFTR-directed therapeutics that target the underlying molecular defect. The CFTR protein functions as an ion channel that is crucial for correct ion and fluid transport across epithelial cells lining the airways and other organs. Consequently, CFTR dysfunction causes a complex multi-organ disease but, to date, most of the morbidity and mortality in people with cystic fibrosis is due to muco-obstructive lung disease. Cystic fibrosis care has long been limited to treating symptoms using nutritional support, airway clearance techniques and antibiotics to suppress airway infection. The widespread implementation of newborn screening for cystic fibrosis and the introduction of a highly effective triple combination CFTR modulator therapy that has unprecedented clinical benefits in up to 90% of genetically eligible people with cystic fibrosis has fundamentally changed the therapeutic landscape and improved prognosis. However, people with cystic fibrosis who are not eligible based on their CFTR genotype or who live in countries where they do not have access to this breakthrough therapy remain with a high unmet medical need., (© 2024. Springer Nature Limited.)