rhDNase Treatment Research Articles

Targeted DNase treatment of obstructive lung disease: a pilot randomised controlled trial

BackgroundSputum extracellular DNA(eDNA) is associated with disease severity in asthma and COPD and therefore emerging as a potential therapeutic target.ObjectiveTo investigate the effect of 10 days of recombinant human DNase(rhDNase) treatment of eDNA-high asthma and COPD, on sputum eDNA levels, neutrophil-related inflammation, lung function and symptoms.MethodsAdults with asthma (n=80) or COPD (n=66) were screened for the presence of high (>20 µg·mL−1) sputum eDNA and those eligible [n=18 asthma, n=17 COPD] were randomised to a two-period crossover controlled trial consisting of daily nebulised rhDNase (2.5 mg/2.5 mL) or placebo (5 mL 0.9%saline) for 10 days, with a 2-week washout period. The primary outcome was sputum eDNA, and secondary outcomes included sputum neutrophil extracellular trap (NET)-related biomarkers, inflammatory cell counts, lung function and respiratory symptoms.ResultsAt screening, high eDNA was associated with significantly higher sputum total cell count, sputum colour score, and inflammation (HNP1-3, LL-37 and IL-1β) in both asthma and COPD compared to low eDNA groups. In asthma, participants with high eDNA were older, and had poorer lung function and asthma control compared to low eDNA. Administration of nebulised rhDNase significantly reduced sputum eDNA levels in both asthma (median(q1,q3) Pre: 48.4 (22.1,74.1); Post: 17.0 (5.0,31.0) µg·mL−1; p=0.022) and COPD (median(q1,q3) Pre: 39.3 (36.7,55.6); Post: 25.4 (11.3,38.6) µg·mL−1; p=0.044) compared to placebo. Symptoms, lung function and NET biomarkers remained unchanged. In asthma, there was a reduction in banded blood neutrophils (3.2(0,7.7) to 0.0 (0.0,1.5); p=0.044).ConclusionTargeted rhDNase treatment for 10 days effectively reduced sputum eDNA in eDNA-high asthma and COPD.

Nebulization of PEGylated recombinant human deoxyribonuclease I using vibrating membrane nebulizers: A technical feasibility study

Recombinant human deoxyribonuclease I (rhDNase, Pulmozyme®) is the most frequently used mucolytic agent for the symptomatic treatment of cystic fibrosis (CF) lung disease. Conjugation of rhDNase to polyethylene glycol (PEG) has been shown to greatly prolong its residence time in the lungs and improve its therapeutic efficacy in mice. To present an added value over current rhDNase treatment, PEGylated rhDNase needs to be efficiently and less frequently administrated by aerosolization and possibly at higher concentrations than existing rhDNase. In this study, the effects of PEGylation on the thermodynamic stability of rhDNase was investigated using linear 20 kDa, linear 30 kDa and 2-armed 40 kDa PEGs. The suitability of PEG30-rhDNase to electrohydrodynamic atomization (electrospraying) as well as the feasibility of using two vibrating mesh nebulizers, the optimized eFlow® Technology nebulizer (eFlow) and Innospire Go, at varying protein concentrations were investigated. PEGylation was shown to destabilize rhDNase upon chemical-induced denaturation and ethanol exposure. Yet, PEG30-rhDNase was stable enough to withstand aerosolization stresses using the eFlow and Innospire Go nebulizers even at higher concentrations (5 mg of protein per ml) than conventional rhDNase formulation (1 mg/ml). High aerosol output (up to 1.5 ml per min) and excellent aerosol characteristics (up to 83% fine particle fraction) were achieved while preserving protein integrity and enzymatic activity. This work demonstrates the technical feasibility of PEG-rhDNase nebulization with advanced vibrating membrane nebulizers, encouraging further pharmaceutical and clinical developments of a long-acting PEGylated alternative to rhDNase for treating patients with CF.

Inhaled Dry Powder Mannitol Treatment in Pediatric Patients with Cystic Fibrosis: Evaluation of Clinical Data in a Real-World Setting.

Background: Cystic fibrosis (CF) is a genetic disorder, in which defective clearance of airway secretions leads to progressive lung function loss. Inhaled mannitol is used to increase sputum and mucociliary clearance. There are little data from real-world studies on the effectiveness of mannitol in children. Our objective was to evaluate the spirometry and clinical results of mannitol in pediatric patients. Methods: We retrospectively reviewed the records of 30 children and adolescents with CF receiving inhaled mannitol who were already on recombinant human deoxyribonuclease (rhDNase) treatment. The change in forced expiratory volume in 1 second (FEV1) from baseline at 2-4 months was the primary outcome. Secondary measures were other spirometry results, body mass index (BMI), hospital admissions, sputum characteristics, and positive bacterial colonization. Results: Compared to baseline, we found significant improvement in percent predicted FEV1 at 2-4 months of treatment; 84.50 (58.00-99.00) vs. 96.00 (66.00-106.00) (P = 0.0007). The absolute change in FEV1 was +11.5% at 2-4 months, +6.5% at 5-7 months, and +4% at 8-12 months. Also, significant improvements in other spirometry results were observed. Adolescents had significantly lower FEV1 results, but the improvement in their lung function was sustained for a more extended period than children. Mannitol provided easier sputum removal, increased sputum volume, significant decline in hospitalizations, and significantly fewer patients with positive sputum cultures. A significant increase in BMI at 8-12 months was observed. Cough was the most frequent adverse effect. Conclusion: In a real-world setting, our results demonstrated that adding mannitol to rhDNase therapy is tolerable in pediatric patients with CF and may provide improved spirometry and clinical outcomes. In addition, our results showed that mannitol provided recovery in overall lung function at 2-4 months, which was sustained up to 12 months together with improved BMI, easier sputum removal, and a decline in bacterial colonization and hospital admissions. However, cough was the most frequent side effect.

Neutrophil Extracellular Traps Induce Organ Damage during Experimental and Clinical Sepsis.

Organ dysfunction is a major concern in sepsis pathophysiology and contributes to its high mortality rate. Neutrophil extracellular traps (NETs) have been implicated in endothelial damage and take part in the pathogenesis of organ dysfunction in several conditions. NETs also have an important role in counteracting invading microorganisms during infection. The aim of this study was to evaluate systemic NETs formation, their participation in host bacterial clearance and their contribution to organ dysfunction in sepsis. C57Bl/6 mice were subjected to endotoxic shock or a polymicrobial sepsis model induced by cecal ligation and puncture (CLP). The involvement of cf-DNA/NETs in the physiopathology of sepsis was evaluated through NETs degradation by rhDNase. This treatment was also associated with a broad-spectrum antibiotic treatment (ertapenem) in mice after CLP. CLP or endotoxin administration induced a significant increase in the serum concentrations of NETs. The increase in CLP-induced NETs was sustained over a period of 3 to 24 h after surgery in mice and was not inhibited by the antibiotic treatment. Systemic rhDNase treatment reduced serum NETs and increased the bacterial load in non-antibiotic-treated septic mice. rhDNase plus antibiotics attenuated sepsis-induced organ damage and improved the survival rate. The correlation between the presence of NETs in peripheral blood and organ dysfunction was evaluated in 31 septic patients. Higher cf-DNA concentrations were detected in septic patients in comparison with healthy controls, and levels were correlated with sepsis severity and organ dysfunction. In conclusion, cf-DNA/NETs are formed during sepsis and are associated with sepsis severity. In the experimental setting, the degradation of NETs by rhDNase attenuates organ damage only when combined with antibiotics, confirming that NETs take part in sepsis pathogenesis. Altogether, our results suggest that NETs are important for host bacterial control and are relevant actors in the pathogenesis of sepsis.

Nebulized rhDNase therapy in newborn infants with atelectasis

Gonderilme tarihi: 05.11.2013 Kabul tarihi: 10.01.2014 Abstract Purpose:To see the efficacy in atelectasis and clinical effectiveness of rhDNase in newborns unresponsive to conventional therapy. Material and methods: Eight newborns (7 premature) with atelectasis in the lungs were evaluated retrospectively. All patients were given rhDNase 2.5 mg twice daily with nebulizer for up to 3 days after conventional therapies. Results:The birth weight and gestational age of the patients ranged respectively from 500 g to 3000 g and 25 weeks to 41 weeks. When atelectasis developed, six of eight patients were mechanically ventilated. Infants received a dosage of 2.5 mg rhDNase every 12 hours with nebulizer for 3 days. Six of 8 patients showed almost complete improvement after the rhDNase treatment. One patient who did not respond to the treatment had an underlying hypoxic ischemic encephalopathy and complicated pneumonia. This patient died during the follow up. The other patient gave partial response to treatment. Of 6 mechanically ventilated patients 3 patients extubated in 3 days, one patient extubated after 9 days and two patients could not be extubated. Atelectasis relapsed in 6 patients. After second 3-day course of treatment by a nebulizer, recovery was seen in 3 days. There was no side effect due to rhDNase treatment. Conclusion:We suggest that rhDNase may be administered as an alternative treatment to the newborn infants with atelectasis in their lung. Especially, if there was no underlying disease, and if they did not respond to conventional treatment for atelectasis. More studies should be done with larger study groups.

Exercise is a potent stimulus for enhancing circulating DNase activity

ObjectivesTo elucidate cell free DNA (cfDNA) clearance kinetics following an acute bout of high intensity exercise by measuring circulating DNase activity reduction (AR). Design and methodsSerum cfDNA concentration and DNase-AR were measured prior to and post (immediately post, 7 and 30min post) an acute bout of rowing exercise until exhaustion. ResultsSerum cfDNA concentration was significantly (P≤.001) elevated immediately post (2.5-fold) and 7min post exercise (2.3-fold) with a return close to baseline at 30min post exercise (1.5-fold). The rise in cfDNA was accompanied by a concomitant, significant (P≤.001) decrease in serum DNase-AR from 15.1% prior to exercise to 3.1% AR at cessation of the exercise test and 7min post exercise (3.9% AR). DNase-AR returned close to baseline at 30min post exercise (5.2% AR). ConclusionsA single bout of high intensity exercise is a potent stimulus for enhancing circulating DNase activity in healthy people. Acute exercise may therefore be considered as a non-pharmacological stimulus to trigger DNase activity.This finding may be relevant for pathological conditions associated with increased cfDNA concentrations like cystic fibrosis, where pharmacological recombinant human DNase (rhDNase) treatment has been successfully used to improve patients' health and physical function.

The use of rhDNAse in severely ill, non-intubated adult asthmatics refractory to bronchodilators: A pilot study

Mucous plugging is associated with fatal asthma and may have a causative role for non-fatal cases of severe acute asthma. However, mucolytic agents have not been found effective in reversing the obstruction of acute asthma. We test the hypothesis that rhDNAse, an agent that reduces viscoelasticity of sputum in patients with cystic fibrosis, has a therapeutic role in acute asthma. Symptomatic asthmatics aged 18-55 years presenting to an Emergency Department with an FEV(1) < 60% predicted after 2 nebulized albuterol and ipratropium treatments were included. Patients were randomized into one of three nebulized rhDNAse treatment groups of 2.5, 5.0 or 7.5 mg, or placebo. Standardized bronchodilator therapy was continued throughout the protocol and the FEV(1) at 6 h was the primary study endpoint. 50 patients were enrolled. There were no significant differences in FEV(1)% predicted between the rhDNAse and placebo patients at any of the post-randomization time points. The dose of rhDNAse administered did not influence response. In a post-hoc stratification, patients with the lowest pre-randomization FEV(1) tended to improve more from rhDNAse, particularly at times 60 and 120 min post-randomization. In this pilot study rhDNAse did not cause clinical improvement among severely ill adults refractory to standardized care. The observed trend to higher FEV(1) among the most severely obstructed patients is an exploratory finding that may warrant further study. This clinical trial was registered as NCT00169962 under the name "Study of Pulmozyme to Treat Severe Asthma Episodes".

Safety of recombinant human deoxyribonuclease as a rescue treatment for persistent atelectasis in newborns

BACKGROUND AND OBJECTIVEPulmonary problems are vitally important in newborns. Increased intense and mucoid secretions may lead to atelectasis, pulmonary infections, respiratory distress, prolonged mechanical ventilation or even death. The aim of this study was to evaluate the safety of recombinant human deoxyribonuclease (rhDNase) in the management of persistent atelectasis in term and preterm newborns, unresponsive to the conventional treatment.DESIGN AND SETTINGProspective study of patients admitted to a general community setting of a neonatal intensive care unit between December 2007 and December 2009.PATIENTS AND METHODSThe study included 22 patients (12 premature and 10 term) who were admitted to the neonatal intensive care unit because of respiratory distress and developed atelectasis, and were unresponsive to conventional treatment. Nebulized rhDNase was administered to all patients at a dose of 1 mg/m2 twice daily for 3 days. In patients who did not respond to 3 days of treatment, endotracheal rhDNase was administered at a dose of 1 mg/m2. We assessed the clinical (respiratory rate and oxygen requirement) and radiologic responses (chest radiographic score), recurrence of atelectasis, the need for a repetitive treatment, and mortality rate.RESULTSA clinical and radiologic improvement of atelectasis was observed in 18 of 22 patients following 3 days of nebulized rhDNase treatment. Atelectasis relapsed in 4 patients. Following the administration of combined endotracheal and nebulized rhDNase treatment, an improvement of atelectasis was noted in all four recurrent cases. No adverse events were observed in patients because of the rhDNase treatment.CONCLUSIONSrhDNase treatment is a safe option and may be used as an effective method for the management of persistent atelectasis in newborns, which is resistant to other conventional treatment methods.

Depletion of neutrophil extracellular traps in vivo results in hypersusceptibility to polymicrobial sepsis in mice

IntroductionAlthough the formation of neutrophil (PMN) extracellular traps (NETs) has been detected during infection and sepsis, their role in vivo is still unclear. This study was performed in order to evaluate the influence of NETs depletion by administration of recombinant human (rh)DNase on bacterial spreading, PMN tissue infiltration and inflammatory response in a mouse model of polymicrobial sepsis.MethodsIn a prospective controlled double-armed animal trial, polymicrobial sepsis was induced by cecal ligation and puncture (CLP). After CLP, mice were treated with rhDNase or phosphate buffered saline, respectively. Survival, colony forming unit (CFU) counts in the peritoneal cavity, lung, liver and blood were determined. PMN and platelet counts, IL-6 and circulating free (cf)-DNA/NETs levels were monitored. PMN infiltration, as well as organ damage, was analyzed histologically in the lungs and liver. Capability and capacity of PMN to form NETs were determined over time.Resultscf-DNA/NETs were found to be significantly increased 6, 24, and 48 hours after CLP when compared to the levels determined in sham and naïve mice. Peak levels after 24 hours were correlated to enhanced capacity of bone marrow-derived PMN to form NETs after ex vivo stimulation with phorbol-12-myristate-13-acetate at the same time. rhDNase treatment of mice resulted in a significant reduction of cf-DNA/NETs levels 24 hours after CLP (P < 0.001). Although overall survival was not affected by rhDNase treatment, median survival after 24 hours was significantly lower when compared with the CLP group (P < 0.01). In mice receiving rhDNase treatment, CFU counts in the lung (P < 0.001) and peritoneal cavity (P < 0.05), as well as serum IL-6 levels (P < 0.001), were found to be already increased six hours after CLP. Additionally, enhanced PMN infiltration and tissue damage in the lungs and liver were found after 24 hours. In contrast, CFU counts in mice without rhDNase treatment increased later but more strongly 24 hours after CLP (P < 0.001). Similarly, serum IL-6 levels peaked after 24 hours (P < 0.01).ConclusionsThis study shows, for the first time, that depletion of NETs by rhDNase administration impedes the early immune response and aggravates the pathology that follows polymicrobial sepsis in vivo.

Modeling gap times between recurrent events by marginal rate function

Gap times between recurrent events are often encountered in longitudinal follow-up studies related to medical science, biostatistics, econometrics, reliability, criminology, demography, and other areas. There have been many models to fit such data, such as proportional hazards (PH) model and additive hazards (AH) model, among others. Standard partial likelihood can be employed to draw their statistical inference. The inference from a direct PH or AH assumption on the gap times, however, is less intuitive and straightforward than marginal rate models–which are often preferred by practitioners due to their more direct interpretations for identifying risk factors. In addition, the existing models have not adequately considered zero-recurrence subjects often encountered in recurrent event data. To overcome these shortcomings, we propose an alternative gap time model using an additive marginal rate function that accounts for zero-recurrence subjects. Local profile-likelihood is applied to estimate the model attributes, and the asymptotic properties of the estimators are established as well. The performance of the proposed estimators is evaluated by a simulation study. The proposed model is applied to analyze a set of data on pulmonary exacerbations and rhDNase treatment.

Nebulized hypertonic saline and recombinant human DNase in the treatment of pulmonary atelectasis in newborns

The aim of this study was to compare and evaluate the efficacy of nebulized 3% hypertonic saline (HS) and recombinant human DNase (rhDNase) treatment for resolution of persistent atelectasis in newborns. Forty newborns (38 preterms) who did not respond to conventional treatment were enrolled to receive either nebulized 3% HS solution (n = 20) or rhDNase (n = 20) between September 2007 and March 2008. Clinical parameters, oxygen saturation and radiological response (chest X-ray scoring) were analyzed before and after administration of 3% HS or rhDNase. The patients of the nebulized 3% HS solution group improved better chest X-ray scores parameters than the patients of the rhDNase group: chest X-ray scores were 5.1 ± 1.9 vs 4.8 ± 1.7 before treatment and 1.0 ± 0.8 vs 2.1 ± 1.4 after treatment (P < 0.001). Resolution time of atelectasis did not differ between the two groups after whole treatment but the percentage of atelectasis resolution after 3 days treatment were 90% (18/20) in the 3% HS group and 70% (14/20) in the rhDNase group. The patients in the 3% HS group improved better also in clinical parameters in comparison to the rhDNase treatment. The difference of oxygen saturation before and after the treatment was 4.6 ± 0.8 in 3% HS group in comparison to 2.6 ± 0.1 in the rhDNase group (P < 0.05). All serum sodium levels were normal in two groups before and after the treatment modalities. This is the first study on the usefulness of nebulized 3% hypertonic saline solution in treating newborns with pulmonary atelectasis. In addition, 3% HS solution was a more effective therapeutic option on the basis of clinical and radiological improvement compared to rhDNase treatment in newborns with pulmonary atelectasis.

DNA Quantification and Fragmentation in Sputum after Inhalation of Recombinant Human Deoxyribonuclease

Background: Inhaled rhDNase may improve sputum viscosity and mucociliary clearance by cleavage of extracellular DNA derived for instance from dead leukocytes in purulent, highly viscous patient sputum. Methods: Here we established a method to quantify rhDNase-mediated DNA fragmentation in sputum using gel electrophoresis. Sputum of Pseudomonas aeruginosa colonized cystic fibrosis (CF) patients with (CF+) or without (CF-) rhDNase treatment or mechanically ventilated non-CF patients receiving rhDNase (non-CF+) or not (non-CF-) was analyzed. DNA measurements from T-lymphocytes served as controls. Absolute DNA content and the relative quantity within eight molecular mass ranges (12000 to 200 bp) was determined by gel electrophoresis and densitometric analysis. Results: Geometric mean sputum DNA concentrations were 0.41 mg/dl for CF- (n=54), 0.78 mg/dl for CF+ (n=60), 0.053 mg/dl for non-CF- (n=41) and 0.049 mg/dl for non-CF+ (n=28). Treatment with rhDNase resulted in fragmentation of DNA that was quantified by separation and densitometric analysis of the DNA on agarose gels. The new analysis method permits analysis of DNA cleavage with high accuracy. Conclusion: This new monitoring method facilitates DNA quantification and in vitro monitoring of rhDNase in sputum.

Effect of aerosolized rhDNase (Pulmozyme®) on pulmonary colonization in patients with cystic fibrosis

Aerosolized recombinant human deoxyribonuclease (rhDNase I (Pulmozyme)) has previously been shown to increase pulmonary function and reduce exacerbations of respiratory symptoms in cystic fibrosis (CF) patients with moderate to severe reduction in pulmonary function. To analyse whether aerosolized Pulmozyme could reduce the number of bacterial infections in the lower respiratory airways of CF patients without chronic pulmonary infection. Patients were randomized either to aerosolized Pulmozyme 2(1/2) mg once daily or to no rhDNase treatment. The study period was 1 y, and the study was blinded for the Department of Clinical Microbiology. Overall, the number of positive cultures was significantly higher in the untreated group (82%) compared with the treated group (72%) (p<0.05). The most striking difference was found for Staphylococcus aureus, with a prevalence of 30% in the untreated group compared with 16% in the treated group (chi2 test, p<0.0001). Pulmonary function (FEV1) in the treated group showed a significant increase of 7.3% compared to 0.9% in the untreated group (p<0.05). Long-term DNase treatment was beneficial to CF patients without chronic lower respiratory tract infection, leading to reduced demand for antibiotics and to improved lung function.

Effect of Treatment with Dornase Alpha on Airway Inflammation in Patients with Cystic Fibrosis

Recombinant human deoxyribonuclease (rhDNase) has been shown to improve lung function and reduce the number of pulmonary exacerbations in patients with cystic fibrosis (CF), but its long-term effect on airway inflammation remains unknown. In this study, we used bronchoalveolar lavage (BAL) to investigate the long-term effect of rhDNase on inflammation in patients with CF having mild lung disease. A total of 105 patients with CF (> or =5 years of age) having normal lung function were randomized to receive rhDNase (2.5 mg/day) or no rhDNase. Patients with a normal percentage of neutrophils in BAL fluid at baseline were not randomized and served as the control group. The percentage of neutrophils in the pooled BAL sample was similar in both randomized groups at baseline. A significant increase in neutrophils was observed over the 3-year study period in both untreated patients and control subjects, whereas neutrophils remained unchanged in patients treated with rhDNase. Elastase activities and interleukin-8 concentrations also increased in untreated patients and remained stable in patients on rhDNase. We conclude that in patients with CF, an increase in neutrophilic airway inflammation is found that is positively influenced by rhDNase treatment.

Efficacy of Human Recombinant DNase in Pediatric Patients with Cystic Fibrosis

Background Most respiratory complications in cystic fibrosis (CF) arise from abnormally viscid mucus, and rhDNase has shown to be effective in enhancing mucous clearance. We explored the responses to rhDNase in a Mexican population of CF patients. Methods Patients with CF received aerosolized rhDNase (2.5 mg daily) during 3 months, followed by daily aerosolized placebo during 3 months. Results A total of 21 CF patients entered the study (11.1 ± 0.5 years of age, mean ± SEM, 10 girls): 15 patients (71%) had basal forced vital capacity (FVC) higher than the 70% predicted value, and the remainder of the patients had an FVC of between 30 and 70%. As a group, rhDNase progressively increased the forced expiratory flow at 1 sec (FEV 1) as well as the FVC, reaching statistical significance ( p <0.005) at the end of the third month of treatment. Sputum production and difficulty to expectorate or to breathe also improved during the rhDNase treatment period ( p <0.05 to p <0.001). All these changes progressively decreased to basal values after 3 months with aerosolized placebo. Adverse reactions were almost null, with a sole patient reporting dysphonia. Conclusions Aerosolized rhDNase was effective in progressively improving respiratory function and symptoms in most CF patients.

STATISTICAL ANALYSIS OF THE LONG-TERM EFFECTS OF RECOMBINANT HUMAN DEOXYRIBONUCLEASE ON PULMONARY FUNCTION IN CYSTIC FIBROSIS PATIENTS

The problem of evaluating the long-term effects of recombinant human deoxyribonuclease (rhDNase) on forced expiratory volume in one second (FEV1) in cystic fibrosis (CF) patients was considered. A two-stage mixed effects model, incorporating relevant predictive variables, captured the diverse patterns of decline of FEV1 for patients with different demographic characteristics. Based on the results of modeling the dropout process, it is clear that the probability of early dropout was closely related to patient's responsiveness to rhDNase treatment. Failure to consider the existence of informative censoring severely biased the estimates of the rate of decline and affected the interpretation of the results.

Effects of sputum oscillations and rhDNase in vitro: a combined approach to treat cystic fibrosis lung disease.

Mucolytic treatment with rhDNase is part of the current therapy for cystic fibrosis (CF) lung disease. The Flutter valve, a device for enhancing airway mucus clearance, has recently been approved for use in CF patients. Exhalation through the Flutter valve leads to oscillations of expiratory airflow, improving mucus viscoelasticity and stimulating clearance. The goal of our in vitro study was to evaluate the individual and combined effects of Flutter valve oscillations and rhDNase treatment on the viscoelastic (rheological) properties of CF sputum. Sputum specimens were collected from 19 CF patients and subjected to the following protocols: 1) baseline sample with no treatment applied; 2) application of oscillations generated by airflow through the Flutter valve; 3) incubation at 37 degrees C for 30 min with 10% vol/wt rhDNase (Pulmozyme) to achieve a final concentration of 2.5 microg/mL (approximately 100 nM); 4) combination of Flutter valve oscillations and 10% vol/wt normal saline (0.9% NaCl); 5) combination of Flutter valve oscillations and 10% vol/wt rhDNase at 2.5 microg/mL final concentration. For each protocol, the mucus rigidity index (log G* at 1 rad/s) was measured at baseline and at 30 min. Values are presented as mean+/-SEM. The cough clearability index (CCI) was computed from measurements of mucus viscoelasticity, based on relationships established in model studies. Flutter valve treatment alone did not result in a significant reduction in the rigidity of CF sputum (2.24+/-0.13 vs. 2.11+/-0.13, P=0.19), nor did rhDNase (2.5 microg/mL) alone, although we have previously shown (Pediatr. Pulmonol. 1995; 20:78) that both of these treatments reduce sputum spinnability, which is more sensitive to molecular weight reduction. In comparison to individual treatments, combined treatment with Flutter valve oscillations and rhDNase significantly reduced the mucus rigidity to 1.85+/-0.19 from 2.24+/-0.13 (P< 0.001), consequently increasing the predicted clearability of the sputum (from 1.09+/-0.26 to 1.83+/-0.48, P=0.012). These in vitro results suggest that a combination of biochemical treatment (e.g., DNase) and mechanical oscillation may have a better therapeutic potential for mucus clearance in CF lung disease.

Improvement of cystic fibrosis airway mucus transportability by recombinant human DNase is related to changes in phospholipid profile.

The aim of this study was to test whether changes in mucus surface properties by rhDNase treatment could be related to an increased recovery of phospholipids. Purulent sputa from 18 patients with cystic fibrosis (CF) were incubated with either rhDNase (4 microg/ml) or control excipient. The incubation of mucus samples with rhDNase induced a significant increase (p < 0.002) in the sol phase proportion (33.7 +/- 24.0%) compared with that obtained with excipient (12.6 +/- 12.4%). Phospholipids were recovered in significantly (p < 0.05) greater amounts from both mucus gel and sol phases after incubation with rhDNase. The phosphatidylglycerol content of mucus sol phase was significantly increased by rhDNase (p < 0.03), as well as the mucus gel phase surface properties and transport by ciliary activity and by cough (p < 0.05). The improvement of mucus gel surface properties and transport capacity by ciliary activity were significantly related to the increased recovery of phosphatidylglycerol (r = -0.74, p < 0.03 and r = 0.94, p < 0.05, respectively). These results suggest that rhDNase is able to increase the free water content and alter the phospholipid profile of mucus, with a related improvement in CF mucus transportability.

Aerosolized rhDNase in cystic fibrosis: effect on leucocyte proteases in sputum

In cystic fibrosis (CF), large amounts of free leucocyte proteases are present in bronchial secretions, contributing to progressive lung damage. Recombinant, human deoxyribonuclease (rhDNase) is a new therapeutic agent that decreases sputum viscosity. However, deoxyribonuclease has been shown, in vitro, to release cationic enzymes from complexes with deoxyribonucleic acid (DNA). The present study was conducted to assess this effect in vivo. Free human leucocyte elastase (HLE), human leucocyte cathepsin G (HCG), total chemotactic activity, and interleukin-8 (IL-8) were determined in sputum from eight patients before, during and after rhDNase treatment. After 15 days of treatment, HLE activity increased by 81+/-44% (NS), and HCG by 189+/-70% (p<0.05). One week after stopping a 4-6 months treatment, HLE activity decreased by 35+/-18% (p<0.05), and HCG by 43+/-11% (p<0.05). Sputum bacterial density, chemotactic activity, and IL-8 concentration did not change. Thus, treatment with rhDNase can indeed increase the activity of HLE and HCG in the bronchial secretions of CF patients, and this effect is still detectable after several months of treatment. If this can be shown to be clinically relevant, combination therapy of recombinant human deoxyribonuclease with protease inhibitors should be considered as an approach to the problem.

Effect of rhDNase on airflow obstruction and mucociliary clearance in cystic fibrosis.

We tested the hypothesis that recombinant human deoxyribonuclease 1 (rhDNase) reduces airflow obstruction and improves mucociliary clearance in patients with cystic fibrosis (CF), and that improvements seen in FEV1 and FVC after rhDNase treatment are independent of chest physical therapy (CPT). CF patients inhaled placebo (10 patients) or 2.5 mg rhDNAse aerosol (10 patients) twice a day for six consecutive days. Compared with baseline, there were no statistically significant differences between the two study groups by Day 6 for indices of airflow obstruction obtained from gamma-camera images of the right lung following inhalation of 99mTc aerosol, or for mucociliary clearance or the rate of clearance of the radioaerosol, quantified over a 6-h period. By Day 6, FEV1 and FVC were significantly higher in the rhDNase-treated group than in the placebo group, increasing by an average of 9.4 +/- 3.5% and 12.7 +/- 2.6%, respectively, as compared with a decrease of 1.8 +/- 1.7% and an increase of 0.4 +/- 1.1%, respectively (p < 0.05). There was no significant change in the FEV1/FVC ratio on Day 6 (0.68 +/- 0.05) compared with baseline (0.70 +/- 0.05) in the rhDNase group. On Day 6, FEV1 and FVC decreased after CPT in both study groups, but the decreases were not significant. Our results indicate that aerosolized rhDNase improves FEV1 and FVC independent of CPT. We were unable to demonstrate that rhDNase reduces airflow obstruction or improves mucociliary clearance.