Cystic Fibrosis Transmembrane Conductance Regulator Mutations Research Articles

Therapeutic applications of curcumin nanomedicine formulations in cystic fibrosis.

Medicinal applications of turmeric-derived curcumin have been known to mankind for long ages. Its potential in managing "cystic fibrosis" has also been evaluated. This autosomal recessive genetic disease is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) which involves an impaired secretion of chloride ions and leads to hypersecretion of thick and sticky mucus and serious complications including airway obstruction, chronic lung infection, and inflammatory reactions. This narrative review aims to highlight the available evidence for the efficacy of curcumin nanoformulations in its potential treatment of cystic fibrosis. Recent research has shown that curcumin acts on the localized mutant CFTR ion channel at the plasma membrane. Preclinical studies have also shown that curcumin nanoformulations have promising effects in the treatment of cystic fibrosis. In this context, the purpose of this narrative review is to highlight the general bioactivity of curcumin, the types of formulations and related studies, thus opening new therapeutic perspectives for CF.

Emerging Concepts in Defective Macrophage Phagocytosis in Cystic Fibrosis.

Cystic fibrosis (CF) is caused by mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Chronic inflammation and decline in lung function are major reasons for morbidity in CF. Mutant CFTR expressed in phagocytic cells such as macrophages contributes to persistent infection, inflammation, and lung disease in CF. Macrophages play a central role in innate immunity by eliminating pathogenic microbes by a process called phagocytosis. Phagocytosis is required for tissue homeostasis, balancing inflammation, and crosstalk with the adaptive immune system for antigen presentation. This review focused on (1) current understandings of the signaling underlying phagocytic mechanisms; (2) existing evidence for phagocytic dysregulation in CF; and (3) the emerging role of CFTR modulators in influencing CF phagocytic function. Alterations in CF macrophages from receptor initiation to phagosome formation are linked to disease progression in CF. A deeper understanding of macrophages in the context of CFTR and phagocytosis proteins at each step of phagosome formation might contribute to the new therapeutic development of dysregulated innate immunity in CF. Therefore, the review also indicates future areas of research in the context of CFTR and macrophages.

P-554 Impact of genotype and phenotype on ICSI outcomes of cystic fibrosis (CF) men patients: a cohort study about 107 ICSI from 1999 to 2019

Abstract Study question Is there an impact of CFTR (cystic fibrosis transmembrane-conductance regulator) gene mutation type and/or general health condition of men with CF and the ICSI outcomes? Summary answer Neither genetic severity nor clinical severity seems to have a negative impact on the biological or clinical ICSI issues for cystic fibrosis men patients. What is known already CFTR protein regulates electrolyte and fluid transport in many tissues with exocrine function, including male reproductive tract. Mutation of CFTR gene causes CF, which affects the function of several organs, and impairs male fertility. CF is generally associated to an obstructive azoospermia because of bilateral absence of vas deferens and seminal vesicles degeneration. CFTR protein is detected in human fetus at the early developmental stages and highly expressed in testis and epididymis. CFTR provides establishment of specific fluid environment for germ cell differentiation and maturation. According to literature, it seems likely that CFTR mutations affect sperm quality and embryo development. Study design, size, duration This cohort study was conducted in the Assisted Reproduction Center of an university hospital on 52 patients, for whom a CF has been diagnosed and monitored in the same hospital. The CF diagnosis was based on CFTR genetic tests and clinical symptoms. All men were azoospermic and underwent on a microscopic epididymal sperm aspiration (MESA) and/or a testicular sperm extraction (TESE). 52 couples underwent in total 107 ICSI with frozen sperm between 1999 and 2019. Participants/materials, setting, methods Clinical data- one year preceding surgical sperm collection and freeze - related to the severity of the CF were collected: respiratory spirometry data, Pseudomonas aeruginosa colonization, number of antibiotic treatment, BMI and Pancreatic insufficiency, as well as CFTR mutations. Linear regression tests (fisher and χ²) were carried out to establish or not correlation between biological and clinical ICSI outcomes and on the other hand genotype and/ or each clinical parameter. Main results and the role of chance The mean age of patients at sperm retrieval was 31.3 years [21-55]. Mean BMI was 21.3. Patients with severe genotype represented 67.8% and likely (67.3%) with external pancreatic deficiency. P.aeroginosa colonization was revealed in 30% of cases and 45.6% patients received at least one IV antibiotic treatment. Respiratory function has been impaired in 18 cases, with a maximal expiratory volume per second < 40%. Spermatozoa were found and frozen for all patients after surgical sperm retrieval (20 MESA and 32 MESA + TESE). The mean number of epididymal motile progressive spermatozoa was satisfactory (10.2±18.83 million). Women mean age was 30 years at ICSI and female infertility was associated in 15.4%. Fertilization and cleavage rates were 66% both. Mean number of transferred and/or frozen embryos per attempt was 3. Implantation rate was 20%. A total of 50 pregnancies and 44 live births were obtained (35.5% of cumulative clinical pregnancy rate per ICSI and 29.1% of live birth rate per transfer). Fertilization rate, absence of pregnancy or absence of live birth were not affected by the genetic severity (p = 0.63, p = 0.46 and p = 0.16 respectively). Likewise, and after several univariate analyses there was no statistically significant correlation between clinical severity and ICSI outcomes. Limitations, reasons for caution Practices of assisted reproduction have constantly evolved during the study period, this concerns the controlled ovarian stimulation protocols, IVF techniques, equipment and culture media used. Our study needs to be completed by a multivariate analyse Wider implications of the findings Regarding to better quality of life of CF patients nowadays and to our results- with no significant correlation between genetic and/or clinical severity and ICSI outcomes, surgical sperm collection can be proposed only in case of a conceptional project or before transplantation and immunosuppressive agents in order to perform ICSI. Trial registration number for non-clinical trials

Effectiveness and Safety of Cystic Fibrosis Transmembrane Conductance Regulator Modulators in Children With Cystic Fibrosis: A Meta-Analysis.

Background and AimCystic fibrosis (CF) is a genetic disease that is difficult to treat and caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) protein. Small molecules have been used to treat the symptom caused by CFTR mutations by restoring CFTR protein function. However, the data on children with CF are scarce. This meta-analysis aimed to evaluate the effectiveness and safety of this therapy in children diagnosed with CF.Materials and MethodsRelevant studies were identified through searching medical databases before April 1, 2022. The primary outcomes of ppFEV1, lung clearance index2.5 (LCI2.5), sweat chloride concentration (SwCI), and Cystic Fibrosis Questionnaire-Revised (CFQ-R) score were pooled and analyzed. The secondary outcomes were nutritional status (weight, BMI, stature, and their z-score) and adverse events under therapy.ResultsA total of twelve studies were included. Compared with the placebo group, the pooled outcome of the ppFEV1, LCI2.5, SwCI, and CFQ-R score were improved by 7.91 {[95% confidence interval (CI), 3.71–12.12], –1.00 (95% CI, –1.38 to –0.63), –35.22 (95% CI, –55.51 to –14.92), and 4.45 (95% CI, 2.31–6.59), respectively}. Compared with the placebo group, the pooled result of the change in weight was improved by 1.53 (95% CI, 0.42–2.63). All the aforementioned results were also improved in single-arm studies. No clear differences in adverse events were found between CFTR modulator therapy and the placebo group.ConclusionCFTR modulators could improve multiaspect function in children with CF and result in comparable adverse events.

Cirrhosis and Portal Hypertension in Compound Heterozygous People With Cystic Fibrosis Harboring One F508del CFTR Gene Mutation

Cirrhosis and Portal Hypertension in Compound Heterozygous People With Cystic Fibrosis Harboring One F508del CFTR Gene Mutation

1451-P: Peri-Islet Fibrosis in Cystic Fibrosis Is Associated with Altered Alpha-Cell Phenotype

Cystic fibrosis (CF) is caused by CF transmembrane conductance regulator mutations leading to impaired epithelial cell anion transport and thickened pancreatic exocrine secretions. CF-related diabetes commonly ensues through impact of pancreatic exocrine pathology on islet cell mass and function although specific mechanisms remain unclear. Paraffin-embedded blocks obtained post mortem from pancreatic body in 9 donors with CF (6 (67%) male; age: 0-27 years) were studied. Following haematoxylin and eosin staining, CF changes were scored as early, advanced or end-stage and islet microenvironment was evaluated for presence of surrounding fibrosis and adipocyte replacement. Four donors were categorised as ‘early’ pathology with 0 (1) % (median (IQR)) of islets surrounded by uninterrupted fibrosis and 0 (0) % by adipocyte replacement; 1 donor was categorised as ‘advanced’ with 100% fibrosis, 0% adipocyte replacement; and 4 donors were categorised as ‘end-stage’ with 65 (46) % fibrosis, 12 (32) % adipocyte replacement. Presence of glucagon/vimentin co-expressing cells was demonstrated by immunofluorescence staining / confocal microscopy in 1 (25%) donor with ‘early’ CF pathology and in all with more advanced exocrine changes. All islets with altered alpha-cell phenotype had peri-islet fibrosis. In pancreata with <50% fibrosis (11±17% (mean±SD) ; n=6) altered alpha-cell phenotype was seen in 11±14% of islets vs. pancreata with >50% fibrosis (93±7%; n=3) with altered alpha-cell phenotype in 65±10% islets (p<0.001) . Analysis of a chronic pancreatitis resection with incrementally increasing fibrosis from head to tail showed parallel increase in islet proportion with altered alpha-cell phenotype (head: 28%; body: 80%; tail: 78%) . We have demonstrated that peri-islet fibrosis is associated with aberrant islet glucagon/vimentin co-expression providing further evidence for exocrine-endocrine signalling in human pancreatic pathology. Disclosure N.Kattner: None. Y.Al-selwi: None. D.Tiniakos: None. G.Klöppel: None. J.A.Shaw: None. Funding Cystic Fibrosis Trust (SRC 019)

828-P: Preservation of ß-Cell Function in Pancreatic Insufficient Cystic Fibrosis (CF) with Highly Effective CFTR Modulator Therapy

CF related diabetes is common with pancreatic insufficient CF (PI-CF) . Highly effective modulator therapy enhances aberrant CF transmembrane conductance regulator (CFTR) function and improves pulmonary and nutritional status. Ivacaftor therapy for indicated CFTR mutations enhanced insulin secretion to glucose-potentiated arginine (GPA) testing; however the effects of elexacaftor/tezacaftor/ivacaftor (ETI; Trikafta™) on β-cell function have not been assessed. We retrospectively compared changes in GPA measures performed on two visits in individuals with PI-CF without diabetes and who were 1) initiated on ETI after the baseline visit (ETI group) vs. 2) not treated with CFTR modulator therapy (controls) . ETI participants (mean±SD age 27±9 years) and 8 matched controls were followed up after a median (IQR) 5.8 (3.7-6.7) and 2.7 (1.9-3.0) years, respectively (p=0.001) , with ETI initiation 1.1 (0.2-1.2) years before the follow-up visit. Weight increased in both groups, but by more in the ETI group (p<0.01) . No differences in lung function or HbA1c were observed; however, improvement in oral glucose tolerance test 2-hour glucose was observed in the ETI group (p<0.05) that was different by trend from the change seen in controls (p=0.09) . Acute insulin and C-peptide responses to glucose-potentiated arginine deteriorated in controls (p<0.05) but not in the ETI group, while C-peptide changes differed between groups (p<0.05) . A trend towards lower acute proinsulin response was seen in the ETI group (p=0.07) with the change different between groups (p<0.05) . Deterioration in the proinsulin secretory ratio was observed in the control (p<0.05) but not the ETI group (between group change, p=0.01) . ETI therapy appears to preserve β-cell function in CF through effects on glucose-dependent insulin secretion and proinsulin processing. Further work should determine whether early intervention with ETI can prevent deterioration of glucose tolerance in PI-CF. Disclosure A.Flatt: None. M.R.Rickels: Advisory Panel; Sernova, Corp., Vertex Pharmaceuticals Incorporated, Zealand Pharma A/S, Consultant; L-Nutra Inc. A.Kelly: Research Support; vTv Therapeutics. S.Sheikh: None. A.J.Peleckis: None. K.Gallagher: None. P.Alvarado: None. D.Hadjiliadis: Advisory Panel; AstraZeneca, Research Support; Mylan N.V. D.Stefanovski: None. R.J.Gallop: None. R.C.Rubenstein: None. Funding Public Health Service research grants (RDK97830; K23 DK107937; UL1 TR001878) and National Institutes of Health (P30 DK19525) .

Losartan ameliorates TGF-β1-induced CFTR dysfunction and improves correction by cystic fibrosis modulator therapies.

Highly effective modulator therapies dramatically improve the prognosis for those with cystic fibrosis (CF). The triple combination of elexacaftor, tezacaftor, and ivacaftor (ETI) benefits many, but not all, of those with the most common F508del mutation in the CF transmembrane conductance regulator (CFTR). Here, we showed that poor sweat chloride concentration responses and lung function improvements upon initiation of ETI were associated with elevated levels of active TGF-β1 in the upper airway. Furthermore, TGF-β1 impaired the function of ETI-corrected F508del-CFTR, thereby increasing airway surface liquid (ASL) absorption rates and inducing mucus hyperconcentration in primary CF bronchial epithelial cells in vitro. TGF-β1 not only decreased CFTR mRNA, but was also associated with increases in the mRNA expression of TNFA and COX2 and TNF-α protein. Losartan improved TGF-β1–mediated inhibition of ETI-corrected F508del-CFTR function and reduced TNFA and COX2 mRNA and TNF-α protein expression. This likely occurred by improving correction of mutant CFTR rather than increasing its mRNA (without an effect on potentiation), thereby reversing the negative effects of TGF-β1 and improving ASL hydration in the CF airway epithelium in vitro. Importantly, these effects were independent of type 1 angiotensin II receptor inhibition.

Functional evaluation of the cystic fibrosis transmembrane conductance regulator in the endocervix†.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an apical membrane chloride/bicarbonate ion channel in epithelial cells. Mutations in CFTR cause cystic fibrosis, a disease characterized by thickened mucus secretions and is associated with subfertility and infertility. CFTR function has been well characterized in vitro and in vivo in airway and other epithelia studies. However, little is known about CFTR function in the cervix in health and its contribution to cyclic regulation of fertility from endocervical mucus changes. Contributing to this research gap is the lack of information on the effect of sex steroid hormones on CFTR expression in cervical epithelial cells across the menstrual cycle. Herein, we demonstrate the hormonal regulation of CFTR expression in endocervical cells both in vitro and in vivo, and that conditionally reprogrammed endocervical epithelial cells can be used to interrogate CFTR ion channel function. CFTR activity was demonstrated in vitro using electrophysiological methods and functionally inhibited by the CFTR-specific inhibitors inh-172 and GlyH-101. We also report that CFTR expression is increased by estradiol in the macaque cervix both in vitro and in vivo in Rhesus macaques treated with artificial menstrual cycles. Estrogen upregulation of CFTR is blocked in vivo by cotreatment with progesterone. Our findings provide the most comprehensive evidence to date that steroid hormones drive changes in CFTR expression. These data are integral to understanding the role of CFTR as a fertility regulator in the endocervix.

PD09-04 INCREASED FREQUENCY OF CYSTIC FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR GENE CARRIER MUTATIONS IN NON-CBAVD INFERTILE MALES

You have accessJournal of UrologyCME1 May 2022PD09-04 INCREASED FREQUENCY OF CYSTIC FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATOR GENE CARRIER MUTATIONS IN NON-CBAVD INFERTILE MALES Thomas Garcia, J. Sam Fisher, Kenny Kim, and Larry Lipshultz Thomas GarciaThomas Garcia More articles by this author , J. Sam FisherJ. Sam Fisher More articles by this author , Kenny KimKenny Kim More articles by this author , and Larry LipshultzLarry Lipshultz More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002535.04AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Homozygous damaging mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene is the primary cause of congenital bilateral absence of the vas deferens (CBAVD). In addition, a high incidence of heterozygous damaging mutations in CFTR have long been implicated in infertility in non-CBAVD males. In the present study we sought to identify novel carrier mutations significantly associated with male infertility from a panel of 62 genes causing common congenital disorders including cystic fibrosis. METHODS: Over the most recent 18-month period, our center performed expanded carrier screening on 165 men receiving a variety of urological treatments including treatment for male infertility, hypogonadism, erectile dysfunction, hematuria, Peyronie's disease, scrotal mass, varicocele, vasectomy, and vasovasostomy. Of these 165 men, 59 were being treated for infertility with full semen analysis reports confirming abnormal semen parameters in 72% of these men (azoospermia, oligozoospermia, asthenozoospermia, or oligoasthenozoospermia). RESULTS: Of the 62 genes tested in 165 men, heterozygous carrier mutations were detected in 12 genes in 57 men (Table 1). No published association between these gene mutations and male infertility could be found for any of the genes except for CFTR. For CFTR, we found a significant 2.3-fold (P <0.05) increase in the number of infertile men with CFTR mutations (9 out of 59 men, or 15.3% of men) relative to the number of other men (men treated for other conditions) with CFTR mutations (7 out of 106 men, or 6.6% of men). None of the azoospermic men with CFTR mutations had congenital bilateral absence of the vas deferens. CONCLUSIONS: Our results are consistent with, but more robust than those of previous studies due to the increased number of mutations tested for in our study. CFTR function has been shown in the epididymis, CFTR protein is present in developing germ cells and sperm, and its function contributes to events required for capacitation in the female reproductive tract. In conclusion, these data demonstrate that mutations in CFTR, even heterozygous mutations, are a likely cause of non-CBAVD male factor infertility in at least a subset of men affected by CFTR mutations. Source of Funding: None © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e178 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Thomas Garcia More articles by this author J. Sam Fisher More articles by this author Kenny Kim More articles by this author Larry Lipshultz More articles by this author Expand All Advertisement PDF DownloadLoading ...

CYSTIC FIBROSIS SCENARIO IN INDIA

Abnormal transport of chloride ion in the epithelial cells is caused by a autosomal recessive monogenic condition known as cystic fibrosis (CF). It belongs to the rare genetic disease in India. Persistent coughing with phlegm, pneumonia, bronchitis, bulky stool and hard bowl movement are most symptoms of the disease. Mutation in CFTR (cystic fibrosis transmembrane conductance regulator) gene present on chromosome 7 having 230 kb nucleotides with 23 exons leads to development of disease. Determination of sweat electrolyte is considered as optimal diagnostic method. Results of previous studies have shown that cystic fibrosis increase the sodium and chloride concentrations. Mutations like ΔF508, G542X, R553X, N130K and 621+1 (G →T) are most common in CF patients in India. Among them ΔF508 is most severe and predominant mutation. Reports have shown as high as 56% frequency of ΔF508 in Indian patients. CF can be treated with anti-inflammatory drugs, CFTR modulators and combination therapies. Early screening can be an effective strategy for early diagnosis and treatment of cystic fibrosis.

CFTR protein quantification as a cystic fibrosis diagnostic biomarker in dried blood spots using multiple reaction monitoring tandem mass spectrometry

The cystic fibrosis transmembrane conductance regulator (CFTR) is a chloride channel found on the apical surface of epithelial cells in the airway and gastrointestinal tract. A mutation in the CFTR protein is responsible for developing cystic fibrosis (CF) disease. Therefore, circulating CFTR protein could be a promising biomarker of CF disease.Multiple methodological challenges are associated with CF's available diagnostic and screening methods, such as low specificity and potential false discovery rate, mainly for ethnic groups whose CFTR mutations are not covered in the mutation panels.Herein, we have developed an absolute quantification (AQUA) method based on two CFTR signature peptides (SPs). A liquid chromatography-tandem spectrometry (LC-MS/MS) method in multiple reaction monitoring (MRM) mode (MRM transitions 1168.90 > 85.929 and 707.19 > 85.93 of SP1 and SP2, respectively) enabled the accurate quantification of CFTR protein in a dried blood spot (DBS). The method was validated successfully based on international guidelines in terms of signal linearity, precision (within-run CV 3.37–8.54%; between-run CV 5.15–11.06% for the selected SPs), and accuracy (within-run 93.4–105.59%; between-run 97.45–103.28% for the selected SPs). The level of soluble CFTR protein was evaluated as a potential biomarker for CF using patients (n = 39) and healthy controls (n = 30), were found to be in CF patients lower than controls. For instant, the level of signature peptide 1 (SP1) was 2.09 ± 0.55 nM, 68.77 ± 1.40 nM in CF patients compared to Ctrl, respectively; p < 0.0001.This study is the first to report CFTR levels in DBS using signature peptides by LC-MS/MS as a diagnostic marker for CF. The receiver operating characteristic (ROC) for CFTR SP1 and SP2 showed a significant area under the curves (AUC) 0.7714 (99% CI, p < 0.0001), and 0.8234 (99% CI, p < 0.0001), respectively. The presented MRM method provides a highly specific and sensitive approach to CFTR quantification in a DBS and could be applied in CF screening.

Molecular Dynamics and Theratyping in Airway and Gut Organoids Reveal R352Q-CFTR Conductance Defect

A significant challenge to making targeted cystic fibrosis transmembrane conductance regulator (CFTR) modulator therapies accessible to all individuals with cystic fibrosis (CF) are many mutations in the CFTR gene that can cause CF, most of which remain uncharacterized. Here, we characterized the structural and functional defects of the rare CFTR mutation R352Q, with a potential role contributing to intrapore chloride ion permeation, in patient-derived cell models of the airway and gut. CFTR function in differentiated nasal epithelial cultures and matched intestinal organoids was assessed using an ion transport assay and forskolin-induced swelling assay, respectively. CFTR potentiators (VX-770, GLPG1837, and VX-445) and correctors (VX-809, VX-445, with or without VX-661) were tested. Data from R352Q-CFTR were compared with data of 20 participants with mutations with known impact on CFTR function. R352Q-CFTR has residual CFTR function that was restored to functional CFTR activity by CFTR potentiators but not the corrector. Molecular dynamics simulations of R352Q-CFTR were carried out, which indicated the presence of a chloride conductance defect, with little evidence supporting a gating defect. The combination approach of in vitro patient-derived cell models and in silico molecular dynamics simulations to characterize rare CFTR mutations can improve the specificity and sensitivity of modulator response predictions and aid in their translational use for CF precision medicine.

Occurrence of Cystic Fibrosis Transmembrane Conductance Regulator Gene Mutations in Patients with Allergic Bronchopulmonary Aspergillosis Complicating Asthma.

Whether cystic fibrosis transmembrane conductance regulator (CFTR) gene mutations contribute to the high prevalence of allergic bronchopulmonary aspergillosis (ABPA) in India remains unknown. We aimed to evaluate the occurrence of CFTR mutations in subjects with ABPA complicating asthma. We sequenced the CFTR gene using genomic DNA from blood on the Illumina NextSeq500 platform. Before undertaking zygosity analysis by genome analysis toolkit, the known or novel single nucleotide polymorphisms (SNPs) and indels were called. For rigorous analysis, we included only high-quality SNPs (scores > 500) and coverage ranging from 30 × 150x. We included 18, 12, and eight adult participants of ABPA, asthma, and healthy controls, respectively. The frequency of SNPs was higher in asthmatic subjects than ABPA or healthy controls, albeit not statistically significant (9/12 [75%] vs. 11/18 [61.1%] vs. 3/8 [37.5%], p = 0.24). Of the 38 subjects, 23 yielded 50 variants (healthy controls [n = 5], ABPA [n = 22], asthma [n = 23]) corresponding to six SNPs not previously linked with ABPA. Of these, four SNPs (rs213950, rs200735475, rs1800113, and rs1800136) were catalogued in the NCBI database. We identified two novel SNPs (chr7:117250703, chr7:117282655) in four (ABPA [n = 1], asthma [n = 3]) subjects without corresponding reference SNP. Most SNPs (85.5%) were heterozygous. The frequency of SNPs was higher in ABPA subjects with high-attenuation mucus (52.2%) and bronchiectasis (39.1%) than serological ABPA (8.7%). Our study suggests the role of CFTR mutations in the pathogenesis of ABPA. The SNPs in the CFTR gene may contribute to disease severity in ABPA. Larger studies are required to confirm our findings.

Distinct proteostasis states drive pharmacologic chaperone susceptibility for cystic fibrosis transmembrane conductance regulator misfolding mutants.

Pharmacological chaperones represent a class of therapeutic compounds for treating protein misfolding diseases. One of the most prominent examples is the FDA-approved pharmacological chaperone lumacaftor (VX-809), which has transformed cystic fibrosis (CF) therapy. CF is a fatal disease caused by mutations in the CF transmembrane conductance regulator (CFTR). VX-809 corrects folding of F508del CFTR, the most common patient mutation, yet F508del exhibits only mild VX-809 response. In contrast, rarer mutations P67L and L206W are hyperresponsive to VX-809, while G85E is nonresponsive. Despite the clinical success of VX-809, the mechanistic origin for the distinct susceptibility of mutants remains unclear. Here we use interactomics to characterize the impact of VX-809 on proteostasis interactions of P67L and L206W and compare these with F508del and G85E. We determine that hyperresponsive mutations P67L and L206W exhibit decreased interactions with proteasomal and autophagy degradation machinery compared with F508del and G85E. We then show inhibiting the proteasome attenuates P67L and L206W VX-809 response. Our data suggest a previously unidentified but required role for protein degradation in VX-809 correction. Furthermore, we present an approach for identifying proteostasis characteristics of mutant-specific therapeutic response to pharmacological chaperones.

Seamless Gene Correction in the Human Cystic Fibrosis Transmembrane Conductance Regulator Locus by Vector Replacement and Vector Insertion Events.

Background: Gene correction via homology directed repair (HDR) in patient-derived induced pluripotent stem (iPS) cells for regenerative medicine are becoming a more realistic approach to develop personalized and mutation-specific therapeutic strategies due to current developments in gene editing and iPSC technology. Cystic fibrosis (CF) is the most common inherited disease in the Caucasian population, caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. Since CF causes significant multi-organ damage and with over 2,000 reported CFTR mutations, CF patients could be one prominent population benefiting from gene and cell therapies. When considering gene-editing techniques for clinical applications, seamless gene corrections of the responsible mutations, restoring native “wildtype” DNA sequence without remnants of drug selectable markers or unwanted DNA sequence changes, would be the most desirable approach. Result: The studies reported here describe the seamless correction of the W1282X CFTR mutation using CRISPR/Cas9 nickases (Cas9n) in iPS cells derived from a CF patient homozygous for the W1282X Class I CFTR mutation. In addition to the expected HDR vector replacement product, we discovered another class of HDR products resulting from vector insertion events that created partial duplications of the CFTR exon 23 region. These vector insertion events were removed via intrachromosomal homologous recombination (IHR) enhanced by double nicking with CRISPR/Cas9n which resulted in the seamless correction of CFTR exon 23 in CF-iPS cells. Conclusion: We show here the removal of the drug resistance cassette and generation of seamless gene corrected cell lines by two independent processes: by treatment with the PiggyBac (PB) transposase in vector replacements or by IHR between the tandemly duplicated CFTR gene sequences.

CFTR Modulator Therapy for Rare CFTR Mutants

Cystic fibrosis (CF), the most common genetic disease among the Caucasian population, is caused by mutations in the gene encoding for the CF transmembrane conductance regulator (CFTR), a chloride epithelial channel whose dysfunction results in severe airway obstruction and inflammation, eventually leading to respiratory failure. The discovery of the CFTR gene in 1989 provided new insights into the basic genetic defect of CF and allowed the study of potential therapies targeting the aberrant protein. In recent years, the approval of “CFTR modulators”, the first molecules designed to selectively target the underlying molecular defects caused by specific CF-causing mutations, marked the beginning of a new era in CF treatment. These drugs have been demonstrated to significantly improve lung function and ameliorate the quality of life of many patients, especially those bearing the most common CFTR mutatant F508del. However, a substantial portion of CF subjects, accounting for ~20% of the European CF population, carry rare CFTR mutations and are still not eligible for CFTR modulator therapy, partly due to our limited understanding of the molecular defects associated with these genetic alterations. Thus, the implementation of models to study the phenotype of these rare CFTR mutations and their response to currently approved drugs, as well as to compounds under research and clinical development, is of key importance. The purpose of this review is to summarize the current knowledge on the potential of CFTR modulators in rescuing the function of rare CF-causing CFTR variants, focusing on both investigational and clinically approved molecules.

CFTR-mediated monocyte/macrophage dysfunction revealed by cystic fibrosis proband-parent comparisons.

Cystic fibrosis (CF) is an inherited disorder caused by biallelic mutations of the CF transmembrane conductance regulator (CFTR) gene. Converging evidence suggests that CF carriers with only 1 defective CFTR copy are at increased risk for CF-related conditions and pulmonary infections, but the molecular mechanisms underpinning this effect remain unknown. We performed transcriptomic profiling of peripheral blood mononuclear cells (PBMCs) of CF child-parent trios (proband, father, and mother) and healthy control (HC) PBMCs or THP-1 cells incubated with the plasma of these participants. Transcriptomic analyses revealed suppression of cytokine-enriched immune-related genes (IL-1β, CXCL8, CREM), implicating lipopolysaccharide tolerance in innate immune cells (monocytes) of CF probands and their parents. These data suggest that a homozygous as well as a heterozygous CFTR mutation can modulate the immune/inflammatory system. This conclusion is further supported by the finding of lower numbers of circulating monocytes in CF probands and their parents, compared with HCs, and the abundance of mononuclear phagocyte subsets, which correlated with Pseudomonas aeruginosa infection, lung disease severity, and CF progression in the probands. This study provides insight into demonstrated CFTR-related innate immune dysfunction in individuals with CF and carriers of a CFTR mutation that may serve as a target for personalized therapy.

Efficacy and Safety of Triple Combination Cystic Fibrosis Transmembrane Conductance Regulator Modulators in Patients With Cystic Fibrosis: A Meta-Analysis of Randomized Controlled Trials

Background: Cystic fibrosis is a rare, recessive, progressive genetic disease caused by dysfunction of the cystic fibrosis transmembrane conductance regulator (CFTR) protein. Small molecules have recently been developed to treat the molecular consequences of CFTR mutations and restore CFTR protein function. However, the data on triple combination therapy (mainly from Vertex Pharmaceuticals, which is most tested in clinical trials) are limited. This meta-analysis was aimed to assess the efficacy and safety of this therapy according to different mutation genotypes and comparators. Methods: Relevant publications were identified through searching several medical databases before 31 December 2021. The primary outcomes of ppFEV1, sweat chloride concentration and Cystic Fibrosis Questionnaire-Revised (CFQ-R) score were pooled and analyzed. The secondary outcomes were adverse events in triple combination therapy. Results: Six randomized controlled trials were eligible for analysis. The total outcome of the ppFEV1 change was higher with triple combination therapy than triple placebo or active control (mean difference, MD, 13.6% and 8.74%, respectively). The pooled result of sweat chloride concentrations with triple combination therapy was lower than that of triple placebo or active control (MD, −44.13 and −39.26, respectively). The pooled estimate of the CFQ-R score was higher with triple combination therapy than triple placebo or active control (MD, 19.8% and 14.63%, respectively). No clear differences in adverse events were found between triple combination therapy and the control (placebo or active control). Conclusion: CFTR modulators in triple combination achieve better clinical results than placebo and active control, and result in comparable adverse events. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42021293402, identifier PROSPERO 2021 CRD42021293402.

CFTR modulator use and risk of nontuberculous mycobacteria positivity in cystic fibrosis, 2011-2018.

BackgroundPeople with cystic fibrosis are at increased risk of pulmonary nontuberculous mycobacteria (NTM) disease. Cystic fibrosis transmembrane conductance regulator (CFTR) modulators are associated with reduced lung infection with pathogens like Pseudomonas aeruginosa and Staphylococcus aureus. This association has not been studied with NTM.MethodsUsing encounter-level data from the US Cystic Fibrosis Foundation Patient Registry from 2011 to 2018, we identified individuals aged >12 years with one or more NTM-negative sputum culture and information on receipt of ivacaftor therapy. We used a Cox proportional hazards model to assess the relationship between CFTR modulator usage (any and monotherapy versus combination therapy) and NTM sputum culture positivity, controlling for sex, least severe class of CFTR mutation, receipt of chronic macrolides, age, body mass index and percentage predicted forced expiratory volume.ResultsOut of 25 987 unique individuals, 17 403 individuals met inclusion criteria. During follow-up, 42% of individuals received CFTR modulator therapy, and 23% had incident NTM. The median (interquartile range) time to event was 6.1 (4.0–7.3) years for those ever receiving CFTR modulators compared to 4.0 (1.6–6.5) years in those never receiving CFTR modulators. CFTR modulator use was associated with a significantly reduced hazard of NTM culture positivity (hazard ratio (HR) 0.88, 95% CI 0.79–0.97); there was no significant difference in the hazard between those receiving ivacaftor monotherapy versus combination therapy (combination HR 1.01, 95% CI 0.79–1.23).ConclusionsCFTR modulator therapy is associated with a decreased risk of NTM positivity in individuals with cystic fibrosis.