Cystic Fibrosis Transmembrane Conductance Regulator Alleles Research Articles

Gain- and Loss-of-Function CFTR Alleles Are Associated with COVID-19 Clinical Outcomes.

Carriers of single pathogenic variants of the CFTR (cystic fibrosis transmembrane conductance regulator) gene have a higher risk of severe COVID-19 and 14-day death. The machine learning post-Mendelian model pinpointed CFTR as a bidirectional modulator of COVID-19 outcomes. Here, we demonstrate that the rare complex allele [G576V;R668C] is associated with a milder disease via a gain-of-function mechanism. Conversely, CFTR ultra-rare alleles with reduced function are associated with disease severity either alone (dominant disorder) or with another hypomorphic allele in the second chromosome (recessive disorder) with a global residual CFTR activity between 50 to 91%. Furthermore, we characterized novel CFTR complex alleles, including [A238V;F508del], [R74W;D1270N;V201M], [I1027T;F508del], [I506V;D1168G], and simple alleles, including R347C, F1052V, Y625N, I328V, K68E, A309D, A252T, G542*, V562I, R1066H, I506V, I807M, which lead to a reduced CFTR function and thus, to more severe COVID-19. In conclusion, CFTR genetic analysis is an important tool in identifying patients at risk of severe COVID-19.

Mutation profiling of the c.1521_1523delCTT (p.Phe508del, F508del) cystic fibrosis transmembrane conductance regulator allele using haplotype-resolved long-read next generation sequencing.

Current approaches to characterize the mutational profile of the cystic fibrosis transmembrane conductance regulator (CFTR) gene are based on targeted mutation analysis or whole gene studies derived from short-read next generation sequencing (NGS). However, these methods lack phasing capability which, in certain scenarios, can provide clinically valuable information. In the present work, we performed near-full length CFTR using Single-Molecule Real-Time Sequencing to produce haplotype-resolved data from both homozygous and heterozygous individuals for mutation c.1521_1523delCTT (p.Phe508del, F508del). This approach utilizes target enrichment of the CFTR gene using biotinylated probes, facilitates multiplexing samples in the same sequencing run, and utilizes fully-automated bioinformatics pipelines for error correction and variant calling. We show a remarkable conservation of F508del haplotype, consistent with the single gene founder effect, as well as diverse mutational profiles in non-F508del alleles. By the same method, 105 single nucleotide polymorphisms (SNPs) exhibiting invariant linkage to F508del CFTR (which better define the founder haplotype) were identified. High level homology between F508del sequences derived from heterozygotes, and those obtained from homozygous individuals, demonstrate accuracy of this method to produce haplotype resolved sequencing. The studies provide a new diagnostic technology for detailed analysis of complex CFTR alleles linked to disease severity.

Triple Therapy for Cystic Fibrosis Phe508del–Gating and –Residual Function Genotypes

BackgroundElexacaftor–tezacaftor–ivacaftor is a small-molecule cystic fibrosis transmembrane conductance regulator (CFTR) modulator regimen shown to be efficacious in patients with at least one Phe508del allele, which indicates that this combination can modulate a single Phe508del allele. In patients whose other CFTR allele contains a gating or residual function mutation that is already effectively treated with previous CFTR modulators (ivacaftor or tezacaftor–ivacaftor), the potential for additional benefit from restoring Phe508del CFTR protein function is unclear.MethodsWe conducted a phase 3, double-blind, randomized, active-controlled trial involving patients 12 years of age or older with cystic fibrosis and Phe508del–gating or Phe508del–residual function genotypes. After a 4-week run-in period with ivacaftor or tezacaftor–ivacaftor, patients were randomly assigned to receive elexacaftor–tezacaftor–ivacaftor or active control for 8 weeks. The primary end point was the absolute change in the percentage of predicted forced expiratory volume in 1 second (FEV1) from baseline through week 8 in the elexacaftor–tezacaftor–ivacaftor group.ResultsAfter the run-in period, 132 patients received elexacaftor–tezacaftor–ivacaftor and 126 received active control. Elexacaftor–tezacaftor–ivacaftor resulted in a percentage of predicted FEV1 that was higher by 3.7 percentage points (95% confidence interval [CI], 2.8 to 4.6) relative to baseline and higher by 3.5 percentage points (95% CI, 2.2 to 4.7) relative to active control and a sweat chloride concentration that was lower by 22.3 mmol per liter (95% CI, 20.2 to 24.5) relative to baseline and lower by 23.1 mmol per liter (95% CI, 20.1 to 26.1) relative to active control (P<0.001 for all comparisons). The change from baseline in the Cystic Fibrosis Questionnaire–Revised respiratory domain score (range, 0 to 100, with higher scores indicating better quality of life) with elexacaftor–tezacaftor–ivacaftor was 10.3 points (95% CI, 8.0 to 12.7) and with active control was 1.6 points (95% CI, −0.8 to 4.1). The incidence of adverse events was similar in the two groups; adverse events led to treatment discontinuation in one patient (elevated aminotransferase level) in the elexacaftor–tezacaftor–ivacaftor group and in two patients (anxiety or depression and pulmonary exacerbation) in the active control group.ConclusionsElexacaftor–tezacaftor–ivacaftor was efficacious and safe in patients with Phe508del–gating or Phe508del–residual function genotypes and conferred additional benefit relative to previous CFTR modulators. (Funded by Vertex Pharmaceuticals; VX18-445-104 ClinicalTrials.gov number, NCT04058353.)

Two Years of Newborn Screening for Cystic Fibrosis in North Macedonia: First Experience.

There is a widely accepted consensus on the benefits of newborn screening (NBS) for cystic fibrosis (CF) in terms of reduced disease severity, improved quality of life, lower treatment burden, and reduced costs. More and more countries in the world are introducing NBS for CF as a national preventive health program. Newborn screening for CF was introduced in the Republic of North Macedonia (RNM) in April, 2019, after a pilot study of 6 months in 2018. A two-step immunoreactive trysinogen (IRT-IRT) algorithm is performed, and then a sweat test for confirmation/exclusion of the CF diagnosis when the IRT values were both over the cutoff (70.0 and 45.0 ng/mL, respectively). In cases with confirmed diagnosis of CF (a sweat chloride concentration >60.0 mmol/L) or with intermediate sweat test results (a sweat chloride concentration of between 30.0 and 59.0 mmol/L), CF transmembrane conductance regulator (CFTR) mutation analysis is performed. By the end of 2020, over a period of 27 months, including the pilot study period, a total number of 43,139 newborns were screened for CF. Seventeen (0.039%) newborns were diagnosed with CF. In all newly discovered CF cases by screening, the diagnosis was confirmed by determination of the CFTR mutations. The most common CFTR mutation, F508del, was found with an overall incidence of 70.6%. Other more frequent mutations were G542X (11.8%) and N1303K (5.9%). Four mutations were found in one CFTR allele each: G1349D, G126D, 457TAT>G and CFTRdupexon22, with the last one being newly discovered with unknown consequences. An incredibly large difference was found in the incidence of the disease between the Macedonian and Albanian neonatal population, with almost four time higher prevalence among Albanians (1:4530 vs. 1:1284).

The Effect of Synonymous Single-Nucleotide Polymorphisms on an Atypical Cystic Fibrosis Clinical Presentation

Synonymous single nucleotide polymorphisms (sSNPs), which change a nucleotide, but not the encoded amino acid, are perceived as neutral to protein function and thus, classified as benign. We report a patient who was diagnosed with cystic fibrosis (CF) at an advanced age and presented very mild CF symptoms. The sequencing of the whole cystic fibrosis transmembrane conductance regulator (CFTR) gene locus revealed that the patient lacks known CF-causing mutations. We found a homozygous sSNP (c.1584G>A) at the end of exon 11 in the CFTR gene. Using sensitive molecular methods, we report that the c.1584G>A sSNP causes cognate exon skipping and retention of a sequence from the downstream intron, both of which, however, occur at a relatively low frequency. In addition, we found two other sSNPs (c.2562T>G (p.Thr854=) and c.4389G>A (p.Gln1463=)), for which the patient is also homozygous. These two sSNPs stabilize the CFTR protein expression, compensating, at least in part, for the c.1584G>A-triggered inefficient splicing. Our data highlight the importance of considering sSNPs when assessing the effect(s) of complex CFTR alleles. sSNPs may epistatically modulate mRNA and protein expression levels and consequently influence disease phenotype and progression.

Cystic fibrosis transmembrane conductance regulator-related male infertility: Relevance of genetic testing & counselling in Indian population.

Background & objectives:Due to limited information available on the frequency and spectrum of cystic fibrosis (CF) transmembrane conductance regulator (CFTR) gene mutations in congenital bilateral absence of vas deferens (CBAVD) in Indian population, it is difficult to provide accurate genetic counselling to couples. The present study was undertaken to investigate the spectrum and frequency of CFTR gene mutations in Indian men with CBAVD and to determine the female CF carrier status.Methods:Direct DNA sequencing of the CFTR gene was carried out in eighty CBAVD men, their female partners and fifty controls from the general population. Pathological significance of the identified novel CFTR gene variants was carried out using in silico tools. Appropriate genetic counselling was provided to the couples prior to intracytoplasmic sperm injection (ICSI).Results:A significant association was observed for CFTR gene variants in Indian CBAVD men versus controls (odds ratio: 12.1; 95% confidence interval: 4.8-30.4; P<0.0001). A total of 20 CFTR gene variants were identified in 53 CBAVD men. Eight novel missense CFTR gene variants (L214V, A238P, E379V, L578I, F587L, L926W, R1325K and R1453Q); two novel splice-site gene variants (c.1-30C>G and IVS1+2T>G) and ten previously reported mutations (R75Q, c.1210-12[5], F508del, A309G, R334W, I444T, R668C, R709X, A1285V and Q1352H) were detected in CBAVD men. The novel and reported CFTR gene mutations were L926W (2.5%, P=0.26), R1453Q (2.5%, P=0.26), F508del (8.75%, P=0.03) and c.1210-12[5] (42.5%, P=0.002). A total of 13 (16.2%) female partners were found to be a CF carrier. Nine couples had a risk of transmitting mutant CFTR allele to the offspring.Interpretation & conclusions:The heterogeneous spectrum of CFTR gene in Indian population suggests the necessity of screening CBAVD men and female partners for accurate genetic counselling prior to undergoing ICSI.

Human airway epithelial cells investigated by atomic force microscopy: A hint to cystic fibrosis epithelial pathology

The pathophysiology of cystic fibrosis (CF) airway disease stems from mutations in the CF Transmembrane Conductance Regulator (CFTR) gene, leading to a chronic respiratory disease. Actin cytoskeleton is disorganized in CF airway epithelial cells, likely contributing to the CF-associated basic defects, i.e. defective chloride secretion and sodium/fluid hypersorption. In this work, we aimed to find whether this alteration could be pointed out by means of Atomic Force Microscopy (AFM) investigation, as roughness and Young's elastic module. Moreover, we also sought to determine whether disorganization of actin cytoskeleton is linked to hypersoption of apical fluid. Not only CFBE41o- (CFBE) cells, immortalized airway epithelial cells homozygous for the F508del CFTR allele, showed a different morphology in comparison with 16HBE14o- (16HBE) epithelial cells, wild-type for CFTR, but also they displayed a lack of stress fibers, suggestive of a disorganized actin cytoskeleton. AFM measurements showed that CFBE cells presented a higher membrane roughness and decreased rigidity as compared with 16HBE cells. CFBE overexpressing wtCFTR became more elongated than the parental CFBE cell line and presented actin stress fibers. CFBE cells absorbed more fluid from the apical compartment. Study of fluid absorption with the F-actin-depolymerizing agent Latrunculin B demonstrated that actin cytoskeletal disorganization increased fluid absorption, an effect observed at higher magnitude in 16HBE than in CFBE cells. For the first time, we demonstrate that actin cytoskeleton disorganization is reflected by AFM parameters in CF airway epithelial cells. Our data also strongly suggest that the lack of stress fibers is involved in at least one of the early step in CF pathophysiology at the levels of the airways, i.e. fluid hypersorption.

From CFTR biology toward combinatorial pharmacotherapy: expanded classification of cystic fibrosis mutations.

More than 2000 mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) have been described that confer a range of molecular cell biological and functional phenotypes. Most of these mutations lead to compromised anion conductance at the apical plasma membrane of secretory epithelia and cause cystic fibrosis (CF) with variable disease severity. Based on the molecular phenotypic complexity of CFTR mutants and their susceptibility to pharmacotherapy, it has been recognized that mutations may impose combinatorial defects in CFTR channel biology. This notion led to the conclusion that the combination of pharmacotherapies addressing single defects (e.g., transcription, translation, folding, and/or gating) may show improved clinical benefit over available low-efficacy monotherapies. Indeed, recent phase 3 clinical trials combining ivacaftor (a gating potentiator) and lumacaftor (a folding corrector) have proven efficacious in CF patients harboring the most common mutation (deletion of residue F508, ΔF508, or Phe508del). This drug combination was recently approved by the U.S. Food and Drug Administration for patients homozygous for ΔF508. Emerging studies of the structural, cell biological, and functional defects caused by rare mutations provide a new framework that reveals a mixture of deficiencies in different CFTR alleles. Establishment of a set of combinatorial categories of the previously defined basic defects in CF alleles will aid the design of even more efficacious therapeutic interventions for CF patients.

185. Targeted Correction and Restored Function of CFTR Gene in Cystic Fibrosis Induced Pluripotent Stem Cells

Recently developed reprogramming and genome editing technologies make possible the derivation of corrected patient-specific pluripotent stem cell sources – potentially useful for the development of new therapeutic approaches. The primary defect in Cystic Fibrosis (CF), an autosomal recessive disorder, is the regulation of epithelial chloride transport by a chloride channel protein encoded by the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Recurrent pulmonary infections are responsible for 80 to 90% of the deaths in CF patients. Starting with skin fibroblasts from patients diagnosed with CF, we have derived and characterized induced pluripotent stem cell (iPSC) lines. We then utilized zinc finger nucleases (ZFNs), designed to target the endogenous CFTR gene, to mediate correction of the inherited genetic mutation in these patient-derived lines via homology directed repair (HDR). We observed an exquisitely sensitive, homology-dependent preference for targeting one CFTR allele vs. the other. Differentiation for a total of 19 days in a protocol designed to derive anterior foregut endoderm, subsequently up-regulated expression of NKX2-1, SOX9, TP63, FOXP2, FOXA2, and CFTR, suggesting commitment of at least a sub-population of cells within the endodermal culture to a lung epithelial cell fate.Once differentiated, mutant CF iPSCs yielded neither mature CFTR protein nor CFTR-specific chloride channel activity (as assayed in Ussing chamber experiments) – whereas corrected CF iPSCs and the normal control WA09 hESCs yielded mature CFTR protein and CFTR-specific chloride channel activity.In vitro differentiation of the mutant CF iPSCs into lung epithelial cells and tissue, controlled for by the parallel differentiation of the otherwise isogenic corrected CF iPSCs, may provide a valuable tool for drug screening and examining the functional consequence of mutant CFTR expression. Furthermore, corrected CF iPSCs present a potential source of patient-specific cells capable, in vitro, of differentiation into various lung stem/progenitor cells – either for transplantation of autologous lung cells or for seeding de-vitalized lung scaffolds ex vivo to generate autologous lungs.

Flavonoids and flavonoid-rich natural extracts inhibit cytokine release in cystic fibrosis bronchial epithelial cells by regulating NF-kB pathway

Cystic fibrosis (CF) is a genetic life-shortening condition in Caucasians. Despite being a multi-organ disease, CF is classically diagnosed by symptoms of acute/chronic respiratory disease, with persistent pulmonary infections (Amaral, 2015). CF is caused by mutations in the gene encoding cystic fibrosis transmembrane conductance regulator (CFTR) protein. Inheritance of mutant CFTR alleles results in surface liquid depletion and defective mucociliary clearance leading to pulmonary failure. Defects in CFTR perturb the regulation of many intracellular signaling pathways including the NF-kB pathway causing excessive production of pro-inflammatory mediators. Current CF therapies are directed to delay CF lung damage by restoring CFTR function and controlling abnormal inflammation. However, only few anti-inflammatory drugs are effective for CF treatments (mainly oral corticosteroids and ibuprofen), these drugs have limited beneficial effects in presence of considerable side effects. Flavonoids have been reported as promising anti-inflammatory drugs and some of them seem to act as CFTR direct activators (Amaral, 2015). From this respect, herbal remedies or plant bioactive molecules may be of great interest. To this aim, we tested the anti-inflammatory activity of apolar extracts from the roots of three Peonia species (Paeoniaceae family), namely P. rockii P. ostii and P. lactiflora , on CFTR ΔF508/ΔF508 CuFi1 cells and normal counterpart. The effects of the extracts on intrinsic as well as TNFα-induced inflammation were evaluated by determining IL-8, IL-6 and RANTES production. Further- more, to study the direct effect of the extracts on NF-kB activation, Human Embryonic Kidney cells were used in transient transfection of NF-kB reporter plasmid and NF-kB activity and cytokine productions were also evaluated. Results showed a significant anti-inflammatory activity of all three Peonia extracts with the P. lactiflora being the most effective. Furthermore, we also tested the anti-inflammatory potential of the pure flavonoid naringin in the same model systems. We found that naringin was able to reduce cytokine release through inhibiting the key enzymes of the NF-kB and MAPK/ ERK pathways. Interestingly, preliminary results on spray dried pharmaceutical formulations of this molecule, show that naringin co-sprayed with leucine improves pharmacological activity of the flavonoid neat raw drug.

WS11.6 Interpretation of nasal potential difference (nPD) measurements in difficult cases of possible cystic fibrosis and the role of published equations

Objectives: Norway introduced newborn screening for Cystic Fibrosis (CF) in March 2012 based on a three tier immunoreactive trypsinogen (IRT)/DNA/DNA protocol. The cystic fibrosis transmembrane conductance regulator (CFTR) mutation spectrum of Norwegian CF patients as well as mutations previously not seen in the population were included in the mutation panels. We present the results from the first 22 months of nationwide CF screening in Norway. Methods: IRT was measured using GSPTM (Perkin Elmer) and samples above 60 ng/ml were included in 2nd tier testing. This involved Luminex-based analysis of 71 mutations and Sanger sequencing of a local common mutation. If only one mutation was found, 3rd tier testing included sequencing of an additional panel of clinically rare alleles found in Norwegian CF patients. Infants carrying two mutations were reported for diagnostic follow-up. Results: At the end of 2013, 111 648 samples had been screened for CF and 875 children (0.78%) had tested positive in primary IRT screening. 2nd tier DNA assessments revealed 22 samples carrying two CFTR mutations. 3rd tier DNA testing of 86 samples disclosed one child with two mutations. Based on the 23 reported children the most frequent alleles were p.R117H (32.6%) and p.F508del (30.4%). In addition, five infants where no mutation was found were reported due to very high IRT values (>400 ng/ml). Conclusion: We found fewer children with a clear CF genotype than expected, and the CFTR mutations and allele frequencies were different compared to clinical material. This suggests reviewing the IRT cut-off level and continued reporting of p.R117H variants as well as extending the 2nd tier mutation panel. WS11.6 Interpretation of nasal potential difference (nPD) measurements in difficult cases of possible cystic fibrosis and the role of published equations R. Pabary1,2, M. Waller2, K. Harman2, I. Ya-Tung2, D. Bilton2,3, N.J. Simmonds2,3, E.W. Alton2, J.C. Davies1,2. 1Royal Brompton & Harefield NHS Foundation Trust, Paediatric Respiratory Medicine, London, United Kingdom; 2Imperial College, National Heart and Lung Institute, London, United Kingdom; 3Royal Brompton & Harefield NHS Foundation Trust, Adult Cystic Fibrosis Unit, London, United Kingdom

WS11.5 Newborn screening for cystic fibrosis in Norway

Objectives: Norway introduced newborn screening for Cystic Fibrosis (CF) in March 2012 based on a three tier immunoreactive trypsinogen (IRT)/DNA/DNA protocol. The cystic fibrosis transmembrane conductance regulator (CFTR) mutation spectrum of Norwegian CF patients as well as mutations previously not seen in the population were included in the mutation panels. We present the results from the first 22 months of nationwide CF screening in Norway. Methods: IRT was measured using GSPTM (Perkin Elmer) and samples above 60 ng/ml were included in 2nd tier testing. This involved Luminex-based analysis of 71 mutations and Sanger sequencing of a local common mutation. If only one mutation was found, 3rd tier testing included sequencing of an additional panel of clinically rare alleles found in Norwegian CF patients. Infants carrying two mutations were reported for diagnostic follow-up. Results: At the end of 2013, 111 648 samples had been screened for CF and 875 children (0.78%) had tested positive in primary IRT screening. 2nd tier DNA assessments revealed 22 samples carrying two CFTR mutations. 3rd tier DNA testing of 86 samples disclosed one child with two mutations. Based on the 23 reported children the most frequent alleles were p.R117H (32.6%) and p.F508del (30.4%). In addition, five infants where no mutation was found were reported due to very high IRT values (>400 ng/ml). Conclusion: We found fewer children with a clear CF genotype than expected, and the CFTR mutations and allele frequencies were different compared to clinical material. This suggests reviewing the IRT cut-off level and continued reporting of p.R117H variants as well as extending the 2nd tier mutation panel. WS11.6 Interpretation of nasal potential difference (nPD) measurements in difficult cases of possible cystic fibrosis and the role of published equations R. Pabary1,2, M. Waller2, K. Harman2, I. Ya-Tung2, D. Bilton2,3, N.J. Simmonds2,3, E.W. Alton2, J.C. Davies1,2. 1Royal Brompton & Harefield NHS Foundation Trust, Paediatric Respiratory Medicine, London, United Kingdom; 2Imperial College, National Heart and Lung Institute, London, United Kingdom; 3Royal Brompton & Harefield NHS Foundation Trust, Adult Cystic Fibrosis Unit, London, United Kingdom

Cystic Fibrosis Transmembrane Regulator Correctors and Potentiators

Cystic fibrosis (CF) is caused by loss-of-function mutations in the CF transmembrane conductance regulator (CFTR) protein, a cAMP-regulated anion channel expressed primarily at the apical plasma membrane of secretory epithelia. Nearly 2000 mutations in the CFTR gene have been identified that cause disease by impairing its translation, cellular processing, and/or chloride channel gating. The fundamental premise of CFTR corrector and potentiator therapy for CF is that addressing the underlying defects in the cellular processing and chloride channel function of CF-causing mutant CFTR alleles will result in clinical benefit by addressing the basic defect underlying CF. Correctors are principally targeted at F508del cellular misprocessing, whereas potentiators are intended to restore cAMP-dependent chloride channel activity to mutant CFTRs at the cell surface. This article reviews the discovery of CFTR potentiators and correctors, what is known regarding their mechanistic basis, and encouraging results achieved in clinical testing.

Report on the p.Ser489X (p.Ser489*) CFTR mutation, a variant with severe associated phenotype and high prevalence in a Quebec French-Canadian cystic fibrosis patient population

PurposeThis study reports on the phenotype of cystic fibrosis patients identified to be carriers of the p.Ser489X (p.Ser489*; c.1466C>A) cystic fibrosis transmembrane conductance regulator (CFTR) mutation, a variant rarely described in the cystic fibrosis literature, as well as on its allelic frequency in a French-Canadian cystic fibrosis patient cohort. MethodsReported phenotypes and allelic frequency of this variant were collected based on the data from a large French-Canadian cystic fibrosis patient cohort. ResultsCystic fibrosis patients found to carry the p.Ser489X variant generally presented with classic gastrointestinal manifestations of this condition in infancy. The allelic frequency of this variant was calculated to be 0.7% for this population. ConclusionThe p.Ser489X CFTR variant is a severe disease-causing CFTR allele that is relatively frequent in the French-Canadian cystic fibrosis patient population, warranting its inclusion into CFTR molecular testing panel for this population.Genet Med 2012:14(10):883–886

Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) in Chinese patients with congenital bilateral absence of vas deferens

BackgroundGenetic testing of the cystic fibrosis transmembrane conductance (CFTR) gene is currently performed in patients with congenital bilateral absence of vas deferens (CBAVD). This study was conducted to investigate the role of mutations in the CFTR gene in CBAVD-dependent male infertility. Methods73 Chinese patients diagnosed with CBAVD were studied. The entire coding regions and splice sites of 27 exons of the CFTR gene were sequenced in 146 chromosomes from the 73 CBAVD patients. Screening was carried out using PCR, gel electrophoresis and DNA sequencing to identify novel variants of the entire coding regions and boundaries of the 27 exons. ResultsFive novel nonsynonymous mutations, three novel splice site mutations and one deletion were identified by sequencing. Apart from the novel variants, we also found 19 previously reported mutations and polymorphism sites. Thirty-four patients (46.57%) had the 5T variant (6 homozygous and 28 heterozygous) and in two of them it was not associated with any detectable mutation of the CFTR gene. All potential pathogenic mutations are not contained in the 1000 Genome Project database. In total, the present study identified 30 potential pathogenic variations in the CFTR gene, 9 of which had not previously been described. ConclusionsMost patients with CBAVD have mutations in the CFTR gene. A mild genotype with one or two mild or variable mutations was observed in all the patients. These findings improve our understanding of the distribution of CFTR alleles in CBAVD patients and will facilitate the development of more sensitive CFTR mutation screening.

Poly Thymidine Polymorphism and Cystic Fibrosis in a Non-Caucasian Population

Background:Cystic fibrosis is a monogenic recessive disorder found predominantly in Caucasian population. This disease arises from mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. In this study we consider poly T polymorphism c.1210-12T[5], c.1210-12T[7], c.1210-12T[9] (T5, T7, T9) in the intron 8 of CFTR gene in normal individuals and cystic fibrosis patients in the north of Iran.Material and methods:40 CF patients and 40 normal individuals were screened for poly T polymorphism in intron 8 of CFTR gene using Reverse Dot Blot method which was also used to detect p.Phe508del among CF patients.Results:T7allele is the most prevalent in both normal and CF patients. Its abundance is approximately 75%. T9and T5represent approximately 20% and 5% of alleles respectively. T7/ T7genotype is the most present in both normal and CF patients with 72.5% and 60% prevalence respectively. p.Phe508del was present in 13 CFTR alleles belonging to 7 patients with either homozygote T9/ T9, T7/ T7or compound heterozygote T7/ T9genotypes.Conclusion:Contrary to the Caucasians, T7allele is more frequent in Northern Iranian CF patients. The presence of p.Phe508del and T7allele in the same framework is reported for the first time in this part of the world. Further investigations of other populations will help to understand whether p.Phe508del arose by selection pressure in this part of the world or was imported from European countries. The abundance of T5, T7, T9alleles indicates that this polymorphism can be used as one of the informative markers for detection of normal and mutant alleles in prenatal diagnosis or carrier assessment in families with previous history of the disease in regions with high degree of CFTR mutation heterogeneity.

Poly thymidine polymorphism and cystic fibrosis in a non-Caucasian population.

Background: Cystic fibrosis is a monogenic recessive disorder found predominantly in Caucasian population. This disease arises from mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. In this study we consider poly T polymorphism c.1210-12T[5], c.1210-12T[7], c.1210-12T[9] (T5, T7, T9) in the intron 8 of CFTR gene in normal individuals and cystic fibrosis patients in the north of Iran. Material and methods: 40 CF patients and 40 normal individuals were screened for poly T polymorphism in intron 8 of CFTR gene using Reverse Dot Blot method which was also used to detect p.Phe508del among CF patients. Results: T7 allele is the most prevalent in both normal and CF patients. Its abundance is approximately 75%. T9 and T5 represent approximately 20% and 5% of alleles respectively. T7/ T7 genotype is the most present in both normal and CF patients with 72.5% and 60% prevalence respectively. p.Phe508del was present in 13 CFTR alleles belonging to 7 patients with either homozygote T9/ T9, T7/ T7 or compound heterozygote T7/ T9 genotypes. Conclusion: Contrary to the Caucasians, T7 allele is more frequent in Northern Iranian CF patients. The presence of p.Phe508del and T7 allele in the same framework is reported for the first time in this part of the world. Further investigations of other populations will help to understand whether p.Phe508del arose by selection pressure in this part of the world or was imported from European countries. The abundance of T5, T7, T9 alleles indicates that this polymorphism can be used as one of the informative markers for detection of normal and mutant alleles in prenatal diagnosis or carrier assessment in families with previous history of the disease in regions with high degree of CFTR mutation heterogeneity.

Personalized medicine for cystic fibrosis: the next generation.

The field of personalized medicine, based on genetic information, represents a tremendous but largely unfulfilled opportunity for a future generation of therapies [1]. While better clinical outcomes have been realized using predictive biomarkers in some forms of cancer [2], the influence of genetic information on the treatment of most other human diseases has been limited. However, in the case of the inherited genetic disease cystic fibrosis (CF), pre-emptive possibilities have recently been uncovered. In particular, an investigational drug known as VX-770, has demonstrated great promise in clinical trials in patients with a specific cystic fibrosis transmembrane conductance regulator (CFTR) allele selected for clinical investigation based on data from in vitro studies [3,4]. This article highlights some of the opportunities and challenges associated with developing novel genotype-directed therapies for CF.

The Novel CFTR Mutation A457P in a Male with a Delayed Diagnosis of Cystic Fibrosis

Cystic fibrosis (CF) is an autosomal recessive disease that may be caused by more than 1000 different mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. We describe the case of a CF patient who was initially diagnosed at 16 years of age after presenting with mild respiratory compromise and pancreatic sufficiency. When genetic testing was first performed using a CF mutation panel, only a single F508del CFTR allele was identified. We subsequently performed testing, which revealed a previously unreported mutation: A457P (p.Ala457Pro, c.1369G>C). The patient's clinical course through adulthood is described, and genotype-phenotype correlation is discussed. The A457P mutation appears to confer a relatively mild phenotype, as is usually observed with CFTR class IV–VI defects. With the advent of more comprehensive and widely available genetic testing techniques, identification of CF genotypes in patients with milder disease variants may help stratify patients for targeted therapy and prevent late complications of the disease.

Genotype-phenotype correlation in cystic fibrosis patients bearing [H939R;H949L] allele

Cystic fibrosis (CF) is caused by CFTR (cystic fibrosis transmembrane conductance regulator) gene mutations. We ascertained five patients with a novel complex CFTR allele, with two mutations, H939R and H949L, inherited in cis in the same exon of CFTR gene, and one different mutation per patient inherited in trans in a wide population of 289 Caucasian CF subjects from South Italy. The genotype-phenotype relationship in patients bearing this complex allele was investigated. The two associated mutations were related to classical severe CF phenotypes.