Search

Background and purpose Human pharmacokinetic studies of lung-targeted drugs are typically limited to measurements of systemic plasma concentrations, which provide no direct information on lung target-site concentrations. We aimed to evaluate lung pharmacokinetics of commonly prescribed drugs by sampling different lung compartments after inhalation and oral administration. Experimental approach Healthy volunteers received single, sequential doses of either inhaled salbutamol, salmeterol, and fluticasone propionate (n=12), or oral salbutamol and propranolol (n=6). Each participant underwent bronchoscopies and gave breath samples for analysis of particles in exhaled air at two time points after drug administration (1 and 6, 2 and 9, 3 and 12, or 4 and 18 hours). Lung samples were taken via bronchosorption, bronchial brush, mucosal biopsy, and bronchoalveolar lavage during each bronchoscopy. Blood samples were taken during the 24 hours after administration. Pharmacokinetic profiles were generated by combining data from multiple individuals, covering all sample timings. Key results Pharmacokinetic profiles were obtained for each drug in lung epithelial lining fluid, lung tissue, and plasma. Inhalation of salbutamol resulted in approximately 100-fold higher concentrations in lung versus plasma. Salmeterol and fluticasone concentration ratios in lung versus plasma were higher still. Bronchosorption- and bronchoalveolar-lavage-generated profiles of inhaled drugs in epithelial lining fluid were comparable, whereas, for oral drugs, epithelial-lining-fluid concentrations appeared to be overestimated in bronchoalveolar-lavage-generated profiles. Conclusion and implications Combining pharmacokinetic data derived from multiple individuals and techniques sampling different lung compartments enabled generation of pharmacokinetic profiles for evaluation of lung targeting after inhaled and oral drug delivery.

After inhaled drug administration, pharmacokinetic (PK) data is typically limited to plasma concentrations and provides no direct information on lung target site levels. This study aims to increase understanding of regional lung PK of selected drugs by taking samples from various lung compartments using innovative sampling methods. Eighteen healthy volunteers were enrolled to either receive single doses of orally inhaled salbutamol (200µg), salmeterol (50µg) and fluticasone propionate (500µg) (n=12) or single oral doses of salbutamol (4 mg) and propranolol (40 mg) (n=6). Two bronchoscopies were performed in each subject. During each bronchoscopy 2 samples of bronchosorption, bronchial brush, mucosal biopsy and one bronchoalveolar lavage (BAL) were performed at different pre-specified times (1, 2, 3, 4, 6, 9, 12, 18 hours) after the drug administration. PK profiles of the model drugs were obtained for concentrations in human lung epithelial lining fluid and lung tissue, enabling insight into the spatial lung distribution of drugs after inhaled and oral administration (Fig. 1). Innovative study design with direct sampling from different compartments in lung provided high quality PK profiles in human lung and demonstrated extent of lung targeting following inhaled drug delivery.

Background Acute Respiratory Distress Syndrome (ARDS) is associated with increased pulmonaryvascular permeability. In the lung, transient receptor potential vanilloid 4 (TRPV4), a Ca2+-permeable cation channel, is a regulator of endothelial permeability and pulmonary edema. We performed a Phase I, placebocontrolled, doubleblind, randomized, parallel group, proofofmechanism study to investigate the effects of TRPV4 channel blocker, GSK2798745, on pulmonaryvascular barrier permeability using a model of lipopolysaccharide (LPS)induced lung inflammation. Methods Healthy participants were randomized 1:1 to receive 2 single doses of GSK2798745 or placebo, 12 hours apart. Two hours after the first dose, participants underwent bronchoscopy and segmental LPS instillation. Total protein concentration and neutrophil counts were measured in bronchoalveolar lavage (BAL) samples collected before and 24 hours after LPS challenge, as markers of barrier permeability and inflammation, respectively. The primary endpoint was baseline adjusted total protein concentration in BAL at 24 hours after LPS challenge. A Bayesian framework was used to estimate the posterior probability of any percentage reduction (GSK2798745 relative to placebo). Safety endpoints included the incidence of adverse events (AEs), vital signs, 12-lead electrocardiogram, clinical laboratory and haematological evaluations, and spirometry. Results Forty-seven participants were dosed and 45 completed the study (22 on GSK2798745 and 23 on placebo). Overall, GSK2798745 was well tolerated. Small reductions in mean baseline adjusted BAL total protein (~ 9%) and neutrophils (~ 7%) in the LPS-challenged segment were observed in the GSK2798745 group compared with the placebo group; however, the reductions did not meet pre-specified success criteria of at least a 95% posterior probability that the percentage reduction in the mean 24hours post LPS BAL total protein level (GSK2798745 relative to placebo) exceeded zero. Median plasma concentrations of GSK2798745 were predicted to inhibit TRPV4 on lung vascular endothelial cells by ~ 7085% during the 24 hours after LPS challenge; median ureacorrected BAL concentrations of GSK2798745 were 3.0 to 8.7fold higher than those in plasma. Conclusions GSK2798745 did not affect segmental LPS-induced elevation of BAL total protein or neutrophils, despite blood and lung exposures that were predicted to be efficacious.

BACKGROUND: Asthma is a heterogeneous disease in which there is a differential response to asthma treatments. This heterogeneity needs to be evaluated so that a personalised management approach can be provided.OBJECTIVES: We stratified patients with moderate-to-severe asthma based on clinico-physiological parameters and performed an -omics analysis of sputum.METHODS: Partition-around-medoid clustering was applied to a training set of 266 asthma participants from the European U-BIOPRED adult cohort using 8 pre-specified clinic-physiological variables. This was repeated in a separate validation set of 152 asthmatics. The clusters were compared based on sputum proteomic and transcriptomic data.RESULTS: Four reproducible and stable clusters of asthmatics were identified. The training set cluster T1 consists of well-controlled moderate-to-severe asthmatics, while cluster T2 is a group of late-onset severe asthmatics with history of smoking and chronic airflow obstruction. Cluster T3 is similar to cluster T2 in terms of chronic airflow obstruction but is composed of non-smokers. Cluster T4 is predominantly composed of obese female uncontrolled severe asthmatics with increased exacerbations, but with normal lung function. The validation set exhibited similar clusters, demonstrating reproducibility of the classification. There were significant differences in sputum proteomics and transcriptomics between the clusters. The severe asthma clusters, T2, T3 and T4, had higher sputum eosinophilia than T1 with no differences in sputum neutrophil counts, exhaled nitric oxide and serum IgE levels.CONCLUSION: Clustering based on clinico-physiological parameters yielded 4 stable and reproducible clusters that associate with different pathobiological pathways.CLINICAL IMPLICATIONS: The definition of four distinct clusters of asthma linked to different pathobiological pathways provides a better template for the phenotyping and personalised treatment of severe asthma, where high unmet needs remain.CAPSULE SUMMARY: Unsupervised clustering of asthma on clinical features alone has led to the definition of four phenotypes. Sputum 'omics' analysis has revealed different biological pathways pointing towards potential new treatments.

There is a need to develop novel noninvasive imaging biomarkers that help to evaluate antiinflammatory asthma treatments.To investigate whether the extent of the segmental lung edema measured noninvasively using turbo-inversion recovery-magnitude magnetic resonance imaging (TIRM MRI) corresponds to the severity of the regional allergic reaction determined by the percentage of eosinophils in bronchoalveolar lavage fluid (BAL) 24 hours after segmental allergen challenge in patients with asthma compared with normal control subjects.Eleven volunteers with allergic asthma and five healthy volunteers underwent segmental challenges with different allergen doses by two bronchoscopies 24 hours apart. They had lung MRI at baseline and 6 and 24 hours after segmental challenge. MRI TIRM scores were correlated with the eosinophilic response at 24 hours.In patients with asthma, there were significant differences of eosinophil percentages in BAL at 24 hours from segments given standard-dose, low-dose, or no allergen (saline) (P0.001). Correspondingly significant differences between the TIRM score in allergen standard-dose, low-dose, and saline-treated segments were observed at 24 hours post-challenge (P0.001). With increasing TIRM score at 24 hours the percent eosinophils per segment 24 hours post-challenge also increased accordingly (P0.001). There was interobserver agreement for TIRM score grading (kappa = 0.72 for 24-h time point).The MRI-based noninvasive TIRM score is a promising biomarker for the noninvasive detection of the inflammatory response after segmental allergen challenge in patients with asthma and may serve to monitor the therapeutic effectiveness of novel antiinflammatory drugs in future human trials.

To evaluate oxygen-enhanced T1-mapping magnetic resonance (MR) imaging as a noninvasive method for visualization and quantification of regional inflammation after segmental allergen challenge in asthmatic patients compared with control subjects.After institutional review board approval, nine asthmatic and four healthy individuals gave written informed consent. MR imaging (1.5 T) was performed by using an inversion-recovery snapshot fast low-angle shot sequence before (0 hours) and 6 hours and 24 hours after segmental allergen challenge by using either normal- or low-dose allergen or saline. The volume of lung tissue with increased relaxation times was determined by using a threshold-based method. As a biomarker for oxygen transfer from the lungs into the blood, the oxygen transfer function ( OTF oxygen transfer function ) was calculated. After the third MR imaging examination, eosinophils in bronchoalveolar lavage fluid were counted. Differences between times and segments were analyzed with nonparametric Wilcoxon matched-pairs test and Spearman correlation.In lung segments treated with the standard dose of allergen, the OTF oxygen transfer function was decreased at 6 hours in asthmatic patients, compared with saline-treated segments (P = .0078). In asthmatic patients at 24 hours, the volume over threshold was significantly increased in normal allergen dose-treated segments compared with saline-treated segments (P = .004). In corresponding lung segments, the volume over threshold at 24 hours in the asthmatic group showed a positive correlation (r = 0.65, P = .0001) and the OTF oxygen transfer function at 6 hours showed an inverse correlation (r = -0.67, P = .0001) with the percentage of eosinophils in the bronchoalveolar lavage fluid.OTF oxygen transfer function and volume over threshold are noninvasive MR imaging-derived parameters to visualize and quantify the regional allergic reaction after segmental endobronchial allergen challenge.

RATIONALE AND OBJECTIVES: Detection of a systemic hemodynamic response in patients suffering from allergic asthma after segmental endobronchial allergen challenge using phase-contrast magnetic resonance imaging (MRI). MATERIALS AND METHODS: Nine asthma patients and four healthy volunteers were examined using MRI (1.5T) before (0 hour), 6 hours, and 24 hours after segmental allergen challenge. Two-dimensional phase-contrast MRI measurements were performed in the aorta (AO) and in the pulmonary artery (PA). In addition, short-axis balanced steady state free precession cardiac cine MRI was performed. Maximum systolic flow, maximum flow acceleration, acceleration volume, acceleration time, distensibility, ejection fraction, stroke volume, end-systolic/diastolic volume, cardiac mass, heart rate (HR), and cardiac output (CO) were determined. Spirometry and bronchoalveolar lavage were also performed. RESULTS: In patients with asthma, maximal systolic flow and maximal flow acceleration increased 6 hours after provocation in the AO (112.3% and 118.9%, respectively) and PA (113.9% and 116.0%, respectively) compared to baseline (100%, P < .05). HR and CO increased significantly at 6 hours (115% and 118%, respectively) compared to baseline (100%, P = .003). In healthy subjects, almost all MRI-derived hemodynamic parameters did not significantly change at 6 hours and were significantly lower than baseline values at 24 hours (P < .02). Twenty-four hours after allergen challenge, all MRI-derived flow parameters were significantly lower in the control group compared to the asthma group (P < .05). HR, CO, and cardiac function parameters measured at 24 hours showed no significant difference comparing the two groups (P > .05). CONCLUSIONS: In asthmatic patients, MRI-derived hemodynamic parameters using phase-contrast MRI are slightly altered after segmental allergen provocation compared to normal controls indicating a mild systemic reaction to local allergen challenge.

Rationale: Segmental endotoxin (LPS) challenge induces neutrophilic airway inflammation in healthy subjects. It allows the precise administration of LPS to a specific lung region and the use of a vehicle challenged segment within the same individual as a control. When compared to whole lung LPS challenge, it targets more peripheral lung regions, requires only a fraction of the LPS dose and has been shown to respond to treatment with a PDE4 inhibitor (Hohlfeld et al. PPT 2008). We assessed the reproducibility of systemic biomarkers in response to local pulmonary LPS challenge and analysed the relationship of serum biomarkers assessed at seven different time points after LPS challenge with the extent of neutrophilic airway inflammation detected 6 and 24 hours following the LPS challenge. Methods: Two cohorts with 10 subjects each underwent 2 segmental LPS challenges within 4 weeks. The inflammatory response was evaluated in BAL fluid at 6 (cohort 1) and 24 hours (cohort 2) both in the LPS and in the saline challenged segments, as well as in serum prior to and at 1, 3, 5.5, 8, 10, 23.5 and 26 hours post LPS challenge. Results: For the baseline values (prior to LPS challenge) the correlation coefficients (n=20 subjects) for the comparison between visits were >0.8 for CC16, MIP1ß, MPO, SP-D, 0.73 for lactoferrin and 0.68 for TNF-. A repeatable increase in serum concentration up to 5.5h following LPS challenge compared to baseline was observed for CC16 (1, 3, 5.5h), IL-6 (5.5h), lactoferrin (5.5h), MCP-1 (5.5h), MIP1ß (3, 5.5 h) and TNF- (5.5 h) (repeated measures ANOVA). No significant correlations were observed between 6 h or the 24 h BAL neutrophil counts and serum collected at the different time points. Conclusion: Serum CC16, IL-6, lactoferrin, MCP-1, and MIP1ß show a repeatable response to segmental LPS challenge, but individual serum biomarkers were not related to the cellular inflammatory response observed in BAL after LPS challenge. Despite reproducibility, serum biomarker assessment following segmental LPS challenge seem to be of limited added value to monitor the local pulmonary response.

Rationale Alveolar Macrophages (AM) play an important role in the innate and adaptive immune response of the lung. During pulmonary infection and allergic airway inflammation AM can be activated and polarized. The alternatively activated phenotype (aaM) is induced by interleukin (IL)-4, a cytokine which is increased during the allergic inflammation. However, the functional role of aaM during the allergic airway response is poorly understood. Therefore, we characterized aaM with regard to the adaptive immune response. Methods Macrophages and dendritic cells (DCs) were generated from human peripheral blood monocytes of atopic donors and aaM were induced in the presence of IL-4. DCs, M and aaM were stimulated with allergen and co-cultured with autologous CD4+ T-lymphocytes. Additionally human AM were isolated from bronchoalveolar lavage (BAL) following segmental allergen provocation and co-cultured with autologous CD4+ T-lymphocytes in the presence or absence of allergen. T-lymphocyte proliferation was assessed by 3H-thymidine incorporation and alternative activation of human macrophages was further characterized by flow cytometric analysis of co-stimulatory surface markers (HLA-DR, CD86 and CD206). Results Unlike DCs, allergen-stimulated M induced no proliferation of antigen-specific CD4+ T-lymphocytes. However, aaM exhibited enhanced antigen presenting capacities, inducing a significant increase of T-lymphocyte proliferation (n=7, p

Segmental challenge with allergen or endotoxin is a powerful tool to study mechanisms of airway inflammation and to test for efficacy of anti-inflammatory compounds. The assessment of airway inflammation by bronchoalveolar lavage (BAL) requires a bronchoscopy. Moreover, repetitive procedures to study inflammation over time are not possible. Noninvasive assessment of airway inflammation by magnetic resonance imaging (MRI) might add to the diagnostic armamentarium and might allow for sequential assessment of inflammation. 11 Patients with mild intermittent allergic asthma with positive responses to inhaled allergen (either grass mix or house dust mite) underwent segmental allergen challenges. After a baseline (B) BAL, four segments were instilled with either saline (S: right upper lobe), a low dose of allergen (LA: medial segment of the right middle lobe), or a high dose of allergen in two segments of contralateral lungs (HA: lateral segment of the right middle lobe and a lingular segment). MRI at 1.5T (Avanto, Siemens, Germany) was acquired 6 hours and 24 hours after segmental challenge using different sequences (TrueFISP, VIBE, T2HASTE, and T2-TIRM). Introduction

Background The aim of this study was to test whether repeatable biomarkers collected from serum, bronchoalveolar lavage (BAL) and sputum of healthy smokers and smokers with COPD would have a prognostic value with respect to the decline in lung function over a 5 year period. Methods In 2006/2007 we had repeatedly collected serum, BAL and sputum of 23 healthy smokers and 24 smokers with COPD (GOLD II) and analysed a panel of more than 100 different parameters. In 2012 we reinvited these subjects to assess the change in lung function to enable the investigation of the potential prognostic value of the 2006/2007 markers and to determine the long-term repeatability of selected blood and serum markers. In this follow-up study we performed body-plethysmography, a blood gas analysis and collected blood and urine samples. The change in lung function was compared with 67 markers from BAL, sputum, serum and whole blood that were shown in the 2006/2007 assessment to be repeatable over a 6 week period. Results We were able to recruit 13 (54%) smokers with COPD and 11 (48%) former healthy smokers that participated in the 2006/2007 study. The decline in lung function was larger in COPD smokers; five of them changed to GOLD III, one to GOLD IV. Two healthy smokers changed to GOLD I. Blood cells, serum von Willebrand factor and alpha-1-antitrypsin showed a good repeatability over 5 years. In COPD smokers a weak correlation between 2006/2007 sputum markers of neutrophilic inflammation and the 5 year change in FEV1/FVC was found. Conclusions Our data suggests that inter-individual and group differences are maintained over a five year period. Despite the large panel of markers available for this analysis, a potential prognostic value appears to exist only for some sputum inflammatory markers. If these data can be confirmed in larger COPD cohorts, it would emphasize the value of sputum markers in clinical trials and support the assumption that an anti-inflammatory treatment can have long term benefits in COPD.