Assays of CFTR Function In Vitro, Ex Vivo and In Vivo.

Cystic fibrosis, a multi-organ genetic disease, is characterized by abnormal function of the cystic fibrosis transmembrane conductance regulator (CFTR) protein, a chloride channel at the apical membrane of several epithelia. In recent years, therapeutic strategies have been developed to correct the CFTR defect. To evaluate CFTR function at baseline for diagnosis, or the efficacy of CFTR-restoring therapy, reliable tests are needed to measure CFTR function, in vitro, ex vivo and in vivo. In vitro techniques either directly or indirectly measure ion fluxes; direct measurement of ion fluxes and quenching of fluorescence in cell-based assays, change in transmembrane voltage or current in patch clamp or Ussing chamber, swelling of CFTR-containing organoids by secondary water influx upon CFTR activation. Several cell or tissue types can be used. Ex vivo and in vivo assays similarly evaluate current (intestinal current measurement) and membrane potential differences (nasal potential difference), on tissues from individual patients. In the sweat test, the most frequently used in vivo evaluation of CFTR function, chloride concentration or stimulated sweat rate can be directly measured. Here, we will describe the currently available bio-assays for quantitative evaluation of CFTR function, their indications, advantages and disadvantages, and correlation with clinical outcome measures.

Ataluren (PTC124) induces cystic fibrosis transmembrane conductance regulator protein expression and activity in children with nonsense mutation cystic fibrosis.

RATIONALE: Nonsense (premature stop codon) mutations in mRNA for the cystic fibrosis transmembrane conductance regulator (CFTR) cause cystic fibrosis (CF) in approximately 10% of patients. Ataluren (PTC124) is an oral drug that permits ribosomes to readthrough premature stop codons in mRNA to produce functional protein. OBJECTIVES: To evaluate ataluren activity, safety, and pharmacokinetics in children with nonsense mutation CF. METHODS: Patients were assessed in two 28-day cycles, comprising 14 days on and 14 days off ataluren. Patients took ataluren three times per day (morning, midday, and evening) with randomization to the order of receiving a lower dose (4, 4, and 8 mg/kg) and a higher dose (10, 10, and 20 mg/kg) in the two cycles. MEASUREMENTS AND MAIN RESULTS: The study enrolled 30 patients (16 male and 14 female, ages 6 through 18 yr) with a nonsense mutation in at least one allele of the CFTR gene, a classical CF phenotype, and abnormal baseline nasal epithelial chloride transport. Ataluren induced a nasal chloride transport response (at least a -5-mV improvement) or hyperpolarization (value more electrically negative than -5 mV) in 50% and 47% of patients, respectively, with more hyperpolarizations at the higher dose. Improvements were seen in seven of nine nonsense mutation genotypes represented. Ataluren significantly increased the proportion of nasal epithelial cells expressing apical full-length CFTR protein. Adverse events and laboratory abnormalities were infrequent and usually mild. Ataluren pharmacokinetics were similar to those in adults. CONCLUSIONS: In children with nonsense mutation CF, ataluren can induce functional CFTR production and is well tolerated.

Clericuzio type poikiloderma with neutropenia is distinct from Rothmund-Thomson syndrome.

Two siblings from a consanguineous family presented with a poikiloderma of limbs and face, plantar keratoderma, and toenail pachyonychia. Neutropenia and neutrophil dysfunction with impairment of the respiratory burst and bacterial killing resulted in frequent respiratory tract infections. A bronchocentric granulomatous pneumonia was a fatal complication. The clinical presentation is consistent with Clericuzio type poikiloderma with neutropenia. Literature review identified several additional probable patients. Genetic linkage analysis excluded the locus of the RECQL4 gene, mutations in which have been described in some patients with the Rothmund-Thomson poikiloderma syndrome. This report confirms the clinical and genetic identity of the Clericuzio type of poikiloderma with neutropenia syndrome.

Monitoring of haemoglobin oxygen saturation in healthy infants using a new generation pulse oximeter which takes motion artifacts into account.

UNLABELLED: The aim of this study was to establish normal values for overnight oxygen saturation (SpO2) in healthy term infants using an oximeter which takes into account motion artifacts and to compare these to normal values collected with a previous generation oximeter not correcting for motion artifacts. We recorded overnight SpO2 in 26 term, healthy infants (median age 136 days, range 6-364 days) in the home environment using the Nellcor Symphony N 3000 pulse oximeter with an averaging time of 3 s. A sample rate of 5 s was chosen. Motion artifacts were excluded from the analysis. Data were compared with those from a previous study, using the same inclusion and exclusion criteria with the Oxford Medilog. Median (quartiles) SpO2 was 98% (97%-99%). Median percentage of study time below SpO2 94% was 0.2% (0.1%-0.7%); median percentage of study time below SpO2 90% was 0.0% (0.0%-0.01%). Median SpO2with the Oxford oximeter was 97% (96%-98%); percentage of study time below SpO2 94% was 8% (2%-14%); percentage of study time below SpO2 90% was 2% (0%-4%). These data were compared with the Nellcor Symphony data: differences in median SpO2 were significant ( P<0.05); differences in percentage of time below SpO2 94% and 90% were also statistically significant ( P<0.001). CONCLUSION: we established normal values of oxygen saturation in healthy term infants using the Nellcor Symphony 3000 pulse oximeter. Care should be taken in interpreting values obtained with different types of pulse oximeters.