Microbiological changes observed over 48 weeks of treatment with inhaled liposomal ciprofloxacin in individuals with non-cystic fibrosis bronchiectasis and chronic Pseudomonas aeruginosa lung infection.

OBJECTIVES: Non-cystic fibrosis bronchiectasis (NCFBE) with Pseudomonas aeruginosa has been associated with increased pulmonary exacerbation (PEx) and mortality risk. European Respiratory Society guidelines conditionally recommend inhaled antimicrobials for persons with NCFBE, P aeruginosa and three or more PEx/year. We report microbiological results of two randomized, 48-week placebo-controlled trials of ARD-3150 (inhaled liposomal ciprofloxacin) in individuals with NCFBE with P aeruginosa and PEx history [Lancet Respir Med 2019;7:213-26]. METHODS: Respiratory secretions from 582 participants receiving up to six 28-day on/off treatment cycles were analysed for sputum P. aeruginosa, Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus and Escherichia coli densities, P. aeruginosa susceptibilities to ciprofloxacin and nine other antimicrobials, and prevalence of other bacterial opportunists. Associations between PEx risk and sputum density, antimicrobial susceptibility and opportunist prevalence changes were studied. RESULTS: Sputum P. aeruginosa density reductions from baseline after ARD-3150 treatments ranged from 1.77 (95% CI 2.13-1.40) versus 0.54 (95% CI 0.89-0.19) log10 CFU/g for placebo (second period) to 2.07 (95% CI 2.45-1.69) versus 0.70 (95% CI 1.11-0.29) log10 CFU/g for placebo (fourth period) with only modest correlation between density reduction magnitude and PEx benefit. ARD-3150 (but not placebo) treatment was associated with increased P. aeruginosa ciprofloxacin MIC but not emergence of other bacterial opportunists across the study; ciprofloxacin MIC50 increased from 0.5 to 1 mg/L, MIC90 increased from 4 to 16 mg/L. Other antimicrobial MIC were mostly unaffected. CONCLUSION: Microbiological changes over 48 weeks of ARD-3150 treatment appear modest. Ciprofloxacin susceptibility (but not other antimicrobial susceptibility) decreases were observed that did not appear to preclude PEx risk reduction benefit.

Ask the experts: the benefits and challenges of pulmonary drug delivery.

The field of pulmonary drug delivery is ever progressing with technological advances in inhaler device design, the development of increasingly sophisticated techniques in targeted delivery and new opportunities in drug formulation, largely as a product of the rapidly advancing area of nanotechnology. Though pulmonary administration offers numerous advantages in terms of both local and systemic drug delivery, the translation of inhaled drugs from bench-to-bedside presents an ongoing challenge. Hannah Coaker, Assistant Commissioning Editor, spoke to six experts and discussed their motivations for becoming involved in the field, major obstacles in aerosol drug development and the evolving area of pulmonary drug delivery.

The ascent of pulmonary drug delivery.

The origins of inhalation therapy can be traced back to the early civilizations but this route of administration was relatively uncommon until recently. Direct delivery of drugs to the lung by inhalation for the treatment of respiratory disease grew rapidly in the second half of the 20th century as a result of the availability of effective asthma drugs in convenient, portable delivery systems. In the search for non-invasive delivery of biologics, it was discovered that the large highly absorptive surface area of the lung could be used for systemic delivery of proteins such as insulin. New delivery systems with efficiency and reproducibility to match the high cost and therapeutic constraints of biologics are currently in late stage clinical trials. Even small molecular weight drugs previously administered by injection are tested via the inhalation route either to provide non-invasively rapid onset of action, or to improve the therapeutic ratio for drugs acting in the lung. Gene therapy of pulmonary disease is still in its infancy but could provide valuable solutions to currently unmet medical needs. The beginning of the new millennium is therefore likely to witness development of many valuable therapeutic products delivered by inhalation.

Coarse spray delivery to a localized region of the pulmonary airways for gene therapy.

Targeting adenoviral vectors for cystic fibrosis gene therapy to the human airways with minimal exposure to alveoli would avoid adverse reactions and maximize response. At present, to deliver gene therapy vectors, large volumes of fluid are instilled or nebulized as aerosols. Either approach would likely cause alveolar exposure and increases the potential for side effects. We describe a coarse spray delivery device that precisely and reproducibly delivers the viral vector to the human airways to treat a small region of the airways for clinical trials. An endoscopic washing pipe (Olympus) that can be inserted into the channel of a bronchoscope was used. To minimize the escape of the therapeutic material downstream from the site of administration, we restricted the volume delivered to <150 microl (to prevent bulk flow), and used large droplets. Their high velocity further enhanced the probability of impaction in the vicinity of the nozzle. A pneumatic dosing system (Kahnetics) was used to reproducibly deliver the spray. The droplet size distribution was determined by laser diffraction and confirmed by cascade impaction: 190-microm volume median diameter with 1% mass <10 microm. The localization of the spray was studied in hollow cast models of human airways. 99mTc-sulfur colloid was used as a radiolabeled marker for these studies. Localization of the deposited spray was determined by scintigraphy and by measuring the radioactivity exiting the terminal airways. In the lung casts the spray was localized to one or two generations over an approximately 2-cm2 area. We conclude that delivery of large droplet sprays limits exposure to a few generations and may be useful in topical gene delivery clinical trials.

Pulmonary insulin administration using the AERx system: physiological and physicochemical factors influencing insulin effectiveness in healthy fasting subjects.

BACKGROUND: Orally inhaled insulin may provide a convenient and effective therapy for prandial glucose control in patients with diabetes. This study evaluated the influence of formulation pH and concentration and different respiratory maneuvers on pharmacokinetic and pharmacodynamic properties of inhaled insulin. METHODS: Three, open-label crossover studies in a total of 23 healthy subjects were conducted in which the safety, pharmacokinetics, and pharmacodynamics of insulin inhalation were compared to subcutaneous (SC) injection into the abdomen of commercially available regular insulin. A novel, aerosol generating system (AERx Diabetes Management System, Aradigm Corporation, Hayward, CA) was used to deliver aqueous insulin bolus aerosols to the lower respiratory tract from formulations at pH 3.5 or 7.4 and concentrations of U250 (250 U/mL) or U500 (500 U/mL). RESULTS: Time to maximum insulin concentration in serum (Tmax) after SC dosing occurred approximately 50-60 minutes with the time to minimum plasma glucose concentration (i.e., maximum hypoglycemic effect), (TGmin), occurring later, at around 100-120 minutes. In contrast, pulmonary delivery led to a significantly earlier Tmax (7-20 minutes) and TGmin (60-70 minutes), parameters that were shown to be largely unaffected by changing the pH or concentration of the insulin. However, investigation of changes in inhaled volume (achieved by different programming of the AERx system) for administration of the same sized aerosol bolus revealed significant effects. Significantly slower absorption and time to peak hypoglycemic activity occurred when aerosol delivery of insulin occurred during a shallow (approximately 40% vital capacity) as opposed to a deep (approximately 80% vital capacity) inspiration. In addition, it was shown that serum concentration of insulin increased immediately after a series of forced expiraratory maneuvers 30 minutes after inhaled delivery. CONCLUSIONS: Pulmonary delivery of aqueous bolus aerosols of insulin in healthy subjects resulted in rapid absorption with an associated hypoglycemic effect quicker than is achieved after subcutaneous dosing of regular insulin. Inhaled insulin pharmacokinetics and pharmacodynamics were independent of formulation variables (pH, concentration) but affected by certain respiratory maneuvers.

Airway epithelial CFTR mRNA expression in cystic fibrosis patients after repetitive administration of a recombinant adenovirus.

We sought to evaluate the ability of an E1(-), E3(-) adenovirus (Ad) vector (Ad(GV)CFTR.10) to transfer the normal human cystic fibrosis transmembrane conductance regulator (CFTR) cDNA to the airway epithelium of individuals with cystic fibrosis (CF). We administered Ad(GV)CFTR.10 at doses of 3 x 10(6) to 2 x 10(9) plaque-forming units over 9 months by endobronchial spray to 7 pairs of individuals with CF. Each 3-month cycle, we measured vector-derived versus endogenous CFTR mRNA in airway epithelial cells prior to therapy, as well as 3 and 30 days after therapy. The data demonstrate that (a) this strategy appears to be safe; (b) after the first administration, vector-derived CFTR cDNA expression in the CF airway epithelium is dose-dependent, with greater than 5% endogenous CFTR mRNA levels at the higher vector doses; (c) expression is transient, lasting less than 30 days; (d) expression can be achieved with a second administration, but only at intermediate doses, and no expression is observed with the third administration; and (e) the progressive lack of expression with repetitive administration does not closely correlate with induction of systemic anti-Ad neutralizing antibodies. The major advantage of an Ad vector is that it can deliver sufficient levels of CFTR cDNA to the airway epithelium so that CFTR expression protects the lungs from the respiratory manifestations of CF. However, this impressive level of expression is linked to the challenging fact that expression is limited in time. Although this can be initially overcome by repetitive administration, unknown mechanisms eventually limit this strategy, and further repetitive administration does not lead to repetitive expression.

Development of a mathematical model for the water distribution in freeze-dried solids.

PURPOSE: Development of a mathematical model to provide information about the amount of water associated with a protein and an excipient in a lyophilized product. METHODS: The moisture content of the product and the mass fraction of each component were used to derive a model for the calculation of the mass of water associating with each component. The model was applied to lyophilized formulations of rhDNase containing various amounts of mannitol or lactose. The total water content was investigated by thermogravimetry, crystalline properties by X-ray powder diffraction and water uptake behaviour using a moisture microbalance system. RESULTS: Calculations based on the model suggest that in a lyophilized rhDNase-mannitol formulation where the sugar is crystalline, most of the water is taken up by the protein. However, in the lyophilized rhDNase-lactose formulation where the sugar is amorphous, water is taken up by both the sugar and protein to a comparative extent. At high relative humidities when the amorphous sugar undergoes crystallization, the model can accommodate such a change by allowing for the formation of an additional crystalline phase. CONCLUSIONS: The rhDNase-sugar formulations show excellent conformity to the model which provides quantitative information about the distribution of water in the lyophilized binary protein-excipient products.

Inhalation delivery systems with compliance and disease management capabilities.

Non-compliance with prescribed medication is a major reason for poor therapeutic outcomes, leading to unnecessary contributions to healthcare costs. Poor technique in self-administration of inhalation therapy is a special type of non-compliance associated with this route of administration. However, pulmonary drug delivery has fundamental advantages for therapy of diseases of the respiratory tract because it is site-directed. The lung is also a promising portal for drug delivery into the systemic circulation. Incorporation of microprocessors into pulmonary drug delivery systems facilitates sophisticated compliance management of chronic diseases such as asthma and diabetes. Microprocessor-assisted systems afford control of patients' administration technique during the therapeutic inhalation event, thus leading to efficient and reproducible regional deposition of the inhaled drug or diagnostic agent. SmartMist is a hand-held asthma disease management device that aids patients to use optimally metered dose inhalers. It also measures pulmonary lung function and provides a long term downloadable electronic record of the therapeutic and diagnostic events. The AERx pulmonary delivery system utilizes similar microprocessor capabilities; however, it employs a novel means of generating aqueous aerosols from unit dose packages, thus providing a broad inhalation technology base for delivery of a wide variety of therapeutic and diagnostic agents into the respiratory tract, and via the lung into the systemic circulation.

Solid state characterization of spray-dried powders of recombinant human deoxyribonuclease (rhDNase).

rhDNase is a recombinant human protein approved as an aqueous solution for human use by inhalation. To study the feasibility of preparation of dry powders for inhalation, spray-dried powders of pure rhDNase and co-spray-dried mixtures of rhDNase with an excipient approved for inhalation products, lactose, were prepared. Both types of powders were initially amorphous. The lactose, however, was found to crystallize after exposure to a humid environment. The crystals in the powder were identified as the alpha-monohydrate polymorph of lactose by hot-stage optical and scanning electron microscopy, differential scanning calorimetry and thermogravimetry, FTIR spectroscopy, and X-ray powder diffraction. Moisture sorption isotherms indicated that crystallization occurred at high relative humidities (70-85%), depending on the temperature of the environment (5-40 degreesC). The practical implications for the manufacturing and storage of protein powders for inhalation are discussed.

Mathematical modeling of deposition and disposition of drugs administered via the nose.

This articles reviews the mathematical models of deposition and disposition of drugs administered into the human nasal cavity for systemic activity. The modeling of the disposition kinetics includes drug release from carriers, translocation within the nasal cavity and into the gastrointestinal tract, drug decomposition, and drug absorption during the transit through the nose and the gastrointestinal tract. The uses of such mathematical models for design and analysis of nasal delivery systems are illustrated, with particular reference to the new therapeutic materials and excipients.

A new method for bronchial-provocation testing in asthmatic subjects using a dry powder of mannitol.

We developed a bronchial provocation test (BPT) with a dry powder preparation of mannitol. The mannitol was inhaled from gelatin capsules containing 5, 10, 20, or 40 mg to a cumulative dose of 635 mg, and was delivered via an inhalator, Halermatic, or Dinkihaler device. We studied the airway sensitivity to inhaled mannitol, the repeatability of the response, and the recovery after challenge in 43 asthmatic subjects 18 to 39 yr of age who had a 20% decrease in FEV1 in response to inhaling a 4.5% NaCl. We compared this with the airway response to methacholine in 25 subjects. The geometric mean (GM) for the dose of dry mannitol required to reduce the FEV1 by 15% of the baseline value (PD15) was 64 mg, with a 95% confidence interval (CI) of 45 to 91. Subjects responsive to mannitol had a PD20 to metacholine of < 7.8 mumol, with a GM of 0.7 mumol (CI: 0.4 to 1.2). For the first of two challenges to mannitol the PD15 was 59 mg (CI: 36 to 97) and for the second the PD15 was 58 mg (CI: 35 to 94) p = 0.91 (n = 23). Spontaneous recovery to within 5% of baseline occurred within 60 min and within 10 min after 0.5 mg terbutaline sulfate was inhaled. Arterial oxygen saturation (SaO2) remained at 93% or above during mannitol challenge. Subjects tolerated the inhalation of the mannitol well. A dry powder preparation of mannitol may be suitable to develop for bronchial provocation testing.

Spray dried powders and powder blends of recombinant human deoxyribonuclease (rhDNase) for aerosol delivery.

PURPOSE: We have used rhDNase to investigate the feasibility of developing a dry protein powder aerosol for inhalation delivery. METHODS: Powders of rhDNase alone and with sodium chloride were prepared by spray drying. Powder blends were obtained by mixing (tumbling and sieving) pure rhDNase powder with 'carrier' materials (lactose, mannitol or sodium chloride). The weight percent of drug in the blends was between 5 and 70%. The particle size distributions and crystallinity of the spray dried powders were obtained by laser diffraction and X-ray powder diffraction, respectively. Particle morphology was examined by scanning electron microscopy. The ability of the powders and powder blends to be dispersed into respirable aerosols was measured using a Rotahaler connected to a multistage liquid impinger operating at 60 L/min. RESULTS: Pure rhDNase powder was quite cohesive with a fine particle fraction (FPF or "respirable fraction': % wt. of particles < 7 microns in the aerosol cloud) of about 20%. When particles also contained NaCl, the powders were dispersed better to form aerosols. A linear relationship was observed between the NaCl content and FPF for a similar primary size (approximately 3 microns volume median diameter) of particles. The particle morphology of these powders varied systematically with the salt content. For the blends, SEM revealed a monolayer-like adhesion of the fine drug particles to the carriers at drug contents > or = 50% wt. An overall 2-fold increase in FPF of rhDNase in the aerosol cloud was obtained for all the blends compared to the pure drug aerosols. CONCLUSIONS: The aerosol properties of spray dried rhDNase powders can be controlled by incorporation of a suitable excipient, such as NaCl, and its relative proportion. Coarse carriers can also enhance the performance of rhDNase dry powder aerosols.

The effect of inhaling a dry powder of sodium chloride on the airways of asthmatic subjects.

Wet aerosols of 4.5% sodium chloride (NaCl) are often used to assess the bronchial responsiveness associated with asthma. We questioned whether dry NaCl could be used as an alternative. Dry powder NaCl was inhaled from capsules containing either 5, 10, 20 or 40 mg to a cumulative dose of 635 mg. The powder was delivered via an Inhalator or Halermatic. The airway sensitivity to the dry and wet NaCl was compared in 24 patients with asthma aged 19-39 yrs. All subjects responded to both preparations and the geometric mean (95% confidence intervals) for the provocative dose of NaCl causing forced expiratory volume in one second (FEV1) to fall 20% from baseline (PD[20,NaCl]) for dry NaCl was 103 mg (68-157) versus 172 mg (102-292), p<0.03 for the wet NaCl. The response to dry NaCl was reproducible and on repeat challenge the PD20 was 108 mg (75-153). The mean maximum fall in FEV1 was approximately 25% on each of the two test days. Spontaneous recovery occurred within 60 min after challenge with dry NaCl and within 5 min after bronchodilator. There were no serious side-effects requiring medical attention, however some patients coughed on inhalation of the 40 mg dose and three gagged. Arterial oxygen saturation remained within normal limits. We conclude that a suitably prepared dry powder of sodium chloride could potentially replace wet sodium chloride to assess bronchial responsiveness in patients with asthma, but further studies are required to establish the long-term stability of the dry powder preparation.

Regional deposition of inhaled hygroscopic aerosols: in vivo SPECT compared with mathematical modeling.

The regional deposition patterns of inhaled hygroscopic aerosols obtained in vivo in the studies of Phipps et al. (P. R. Phipps, I. Gonda, D. L. Bailey, P. Borham, G. Bautovich, and S. D. Anderson. Am. Rev. Respir. Dis. 139: 1516, 1989; and P. R. Phipps, I. Gonda, S. D. Anderson, D. L. Bailey, and G. Bautovich. Eur. Respir. J. 7: 1474-1482, 1994) and Chan et al. (H.-K. Chan, P. R. Phipps, I. Gonda, P. Cook, R. Fulton, I. Young, and G. Bautovich. Eur. Respir. J. 7: 1483-1489, 1994) by using single-photon-emission computerized tomography (SPECT) are compared with the regional deposition predicted by the hygroscopic lung deposition model of Finlay and Stapleton (W. H. Finlay and K. W. Stapleton. J. Aerosol Sci. 26: 655-670, 1995). Three pairs of saline aerosols are considered: isotonic with small [2.6-microns mass median aerodynamic diameter (MMAD), geometric standard deviation (GSD) 1.4] vs. large (5.5-microns MMAD, GSD 1.7) droplets; hypotonic (0.3% NaCl) vs. hypertonic (4.5% NaCl) with 3.7- to 3.8-microns MMAD (GSD 1.4), and hypotonic vs. hypertonic (3.7- to 3.8-microns MMAD, GSD 1.5-1.8) with reduced number of droplets per cubic centimeter. For each of the three pairs of aerosols, no significant difference (P > 0.05) was found between the in vivo and computational results for either the mean value or the variance of the difference in peripheral to central deposition. Thus it appears that theoretical calculations can be used to predict the pattern of lung deposition of hygroscopic aerosols in populations of normal subjects.

Effects of additives on heat denaturation of rhDNase in solutions.

PURPOSE: To study the thermal stability of recombinant human deoxyribonuclease I (rhDNase) in aqueous solutions. METHODS: Differential scanning calorimetry (DSC) was used to measure the denaturation or melting temperature (T(m)) and enthalpy (H(m)) of rhDNase. The effects of denaturants (guanidine HCl and urea) and additives (mainly divalent cations and disaccharides) were investigated at pH 6-7. RESULTS: The T(m) and H(m) of rhDNase in pure water were measured as 67.4 degrees C and 18.0 J/g respectively, values typical of globular proteins. The melting peak disappeared on re-running the sample after cooling to room temperature, indicating that the thermal denaturation was irreversible. The latter was due to the occurrence of aggregation accompanying the unfolding process of rhDNase. Size exclusion chromatography indicated that during heat denaturation, rhDNase formed soluble high molecular weight aggregates with a molecular size >300kD estimated by the void volume. Of particular interest are the divalent cations: Ca(2+) stabilizes rhDNase against thermal denaturation and elevates T(m) and H(m) while Mg(2+), Mn(2+) and Zn(2+) destabilize it. Sugars also stabilize rhDNase. As expected, denaturants destabilize the protein and lower the T(m) and H(m). All destabilization of rhDNase can be prevented by adding Ca(2+) to the solutions. CONCLUSIONS: CaCl(2) and sugars were found to stabilize rhDNase against thermal denaturation while divalent cations, urea and guanidine HCl destabilize the protein. The effects could be explained by a mixture of mechanisms. For Ca(2+) the protective effect is believed to be due to an ordering of the rhDNase structure in its native state, and by prevention of breaking of a disulfide bridge, thus making it less susceptible to unfold under thermal stress.

Mucociliary clearance during and after isocapnic hyperventilation with dry air in the presence of frusemide.

We have previously shown that mucociliary clearance (MCC) decreased during and increased after isocapnic hyperventilation (ISH) with dry air, both in asthmatic and healthy subjects. Inhaled frusemide, an inhibitor of the Na+/K+/2Cl- and NaCl co-transporters on the basolateral membrane of the epithelial cell, prevents the airway narrowing provoked by ISH with dry air. The co-transport system controls epithelial cell volume and chloride secretion and, thus, frusemide has the potential to modify the rate of recovery of periciliary fluid volume during and after ISH with dry air, and hence affect MCC. Frusemide also blocks mediator release from mast cells, which may also modify the increase in MCC after ISH. Eleven asthmatic and 11 healthy subjects inhaled frusemide (35.7 +/- 0.44 mg) or its vehicle, from a Fisoneb ultrasonic nebulizer 30 min before ISH with dry air, on two separate occasions. MCC was measured using 99mTc-sulphur colloid and a gamma camera. Frusemide, compared to its vehicle, did not affect MCC during or 45 min after ISH. However, in the presence of frusemide, the onset of the increase of MCC after ISH was significantly delayed for approximately 10 min in the whole right lung (p < 0.002) and central region (p < 0.01) in the asthmatic but not in the healthy subjects. These findings could be explained by frusemide delaying the recovery of the periciliary fluid volume after ISH with dry air and/or interfering with the stimulus that causes the increase in MCC in the asthmatic subjects after ISH.

Inhalation of hypertonic saline aerosol enhances mucociliary clearance in asthmatic and healthy subjects.

Hyperosmolarity of the airway surface liquid (ASL) has been proposed as the stimulus for hyperpnoea-induced asthma. We found previously that mucociliary clearance (MCC) was increased after isocapnic hyperventilation (ISH) with dry air, and we proposed that the increase related to transient hyperosmolarity of the ASL. We investigated the effect of increasing the osmolarity of the ASL on MCC, by administering an aerosol of concentrated salt solution. MCC was measured using 99mTc-sulphur colloid, gamma camera and computer analysis in 12 asthmatic and 10 healthy subjects on three separate days, involving administration of each of the following: 1) ultrasonically nebulized 14.4% saline; 2) ultrasonically nebulized 0.9% saline; and 3) no aerosol intervention (control). The (mean +/- SD) volume of nebulized 14.4% saline was 2.2 +/- 1.2 mL for asthmatics and 3.2 +/- 0.7 mL for healthy subjects. This volume was delivered over a period of 5.4 +/- 1.3 and 6.4 +/- 0.7 min for asthmatic and healthy subjects, respectively. The airway response to 14.4% saline was assessed on a separate visit and the fall in forced expiratory volume in one second (FEV1) was 22 +/- 4% in the asthmatic and 3 +/- 2% in the healthy subjects. Compared to the MCC with the 0.9% saline and control, the hypertonic aerosol increased MCC in both groups. In asthmatic subjects, MCC of the whole right lung in 1 h was 68 +/- 10% with 14.4% saline vs 44 +/- 14% with 0.9% saline and 39 +/- 13% with control. In healthy subjects, MCC of the whole right lung in 1 h was 53 +/- 12% with 14.4% saline vs 41 +/- 15% with 0.9% saline and 36 +/- 13% with control. We conclude that an increase in osmolarity of the airway surface liquid increases mucociliary clearance both in asthmatic and healthy subjects. These findings are in keeping with our previous suggestion that the increase in mucociliary clearance after isotonic hyperventilation with dry air is due to a transient hyperosmolarity of the airway surface liquid.