Segmental instillation of lipopolysaccharide (LPS) by bronchoscopy safely induces transient airway inflammation in human lungs. This model enables investigation of pulmonary inflammatory mechanisms as well as pharmacodynamic analysis of investigational drugs. The aim of this work was to describe the transcriptomic profile of human segmental LPS challenge with contextualization to major respiratory diseases. Pre-challenge bronchoalveolar lavage (BAL) fluid and biopsies were sampled from 28 smoking, healthy participants, followed by segmental instillation of LPS and saline as control. Twenty-four hours post instillation, BAL and biopsies were collected from challenged lung segments. Total RNA of cells from BAL and biopsy samples were sequenced and analysed for differentially expressed genes (DEGs). After challenge with LPS compared with saline, 6316 DEGs were upregulated and 241 were downregulated in BAL, but only one DEG was downregulated in biopsy samples. Upregulated DEGs in BAL were related to molecular functions such as “Inflammatory response” or “chemokine receptor activity”, and upregulated pro-inflammatory pathways such as “Wnt-"/“Ras-"/“JAK-STAT” “-signaling pathway”. Furthermore, the segmental LPS challenge model resembled aspects of the five most prevalent respiratory diseases chronic obstructive pulmonary disease (COPD), asthma, pneumonia, tuberculosis and lung cancer and featured similarities with acute exacerbations in COPD (AECOPD) and community-acquired pneumonia. Overall, our study provides extensive information about the transcriptomic profile from BAL cells and mucosal biopsies following LPS challenge in healthy smokers. It expands the knowledge about the LPS challenge model providing potential overlap with respiratory diseases in general and infection-triggered respiratory insults such as AECOPD in particular.

Background Bronchiectasis is characterised by uncontrolled neutrophil serine protease (NSP) activity. Cathepsin C (CatC; dipeptidyl peptidase 1) activates NSPs during neutrophil maturation. CatC inhibitors can potentially reduce neutrophil-mediated lung damage. This phase II, randomised, double-blind, placebo-controlled trial (AIRLEAF (R); clinicaltrials.gov identifier NCT05238675) evaluated efficacy, safety and optimal dosing of BI 1291583, a novel, reversible CatC inhibitor, in adults with bronchiectasis. Methods In total, 322 participants were randomised (2:1:1:2) to receive one of three oral doses of BI 1291583 (1 mg/2.5 mg/5 mg) or placebo for 24-48 weeks. A multiple comparison procedure and modelling approach was used to demonstrate a nonflat dose-response curve based on the time to first pulmonary exacerbation up to week 48. In addition, efficacy of individual BI 1291583 doses was evaluated based on the frequency of exacerbations, severe exacerbations (fatal or leading to hospitalisation and/or intravenous antibiotic administration), lung function and quality of life. Results A significant dose-dependent benefit of BI 1291583 over placebo was established based on time to first exacerbation (shape: maximum effect curve 1; adjusted p=0.0448). Treatment with BI 1291583 5 mg and 2.5 mg numerically reduced the risk of an exacerbation compared with placebo (hazard ratio (95% CI) 0.71 (0.48 to 1.05) and 0.66 (0.40 to 1.08), respectively; both p>0.05). BI 1291583 2.5 mg showed numerically better efficacy compared with 5 mg across several end-points; 1 mg was similar to placebo. The safety profile of BI 1291583 was similar to placebo. Conclusion Treatment with BI 1291583 resulted in a reduction in the risk of experiencing an exacerbation in adults with bronchiectasis.

Abstract Background FOOTPRINTS® is a prospective, longitudinal, 3-year study assessing the association between biomarkers of inflammation/lung tissue destruction and chronic obstructive pulmonary disease (COPD) severity and progression in ex-smokers with mild-to-severe COPD. Here, we present baseline characteristics and select biomarkers of study subjects. Methods The methodology of FOOTPRINTS® has been published previously. The study population included ex-smokers with a range of COPD severities (Global Initiative for Chronic Obstructive Lung Disease [GOLD] stages 1–3), ex-smokers with COPD and alpha-1-antitrypsin deficiency (A1ATD) and a control group of ex-smokers without airflow limitation (EwAL). At study entry, data were collected for: demographics, disease characteristics, history of comorbidities and COPD exacerbations, symptoms, lung function and volume, exercise capacity, soluble biomarkers, and quantitative and qualitative computed tomography. Baseline data are presented with descriptive statistical comparisons for soluble biomarkers in the individual GOLD and A1ATD groups versus EwAL. Results In total, 463 subjects were enrolled. The per-protocol set comprised 456 subjects, mostly male (64.5%). The mean (standard deviation) age was 60.7 (6.9) years. At baseline, increasing pulmonary symptoms, worse lung function, increased residual volume, reduced diffusing capacity of the lung for carbon monoxide (DLco) and greater prevalence of centrilobular emphysema were observed with increasing disease severity amongst GOLD 1–3 subjects. Subjects with A1ATD (n = 19) had similar lung function parameters to GOLD 2–3 subjects, a high residual volume comparable to GOLD 3 subjects, and similar air trapping to GOLD 2 subjects. Compared with EwAL (n = 61), subjects with A1ATD had worse lung function, increased residual volume, reduced DLco, and a greater prevalence of confluent or advanced destructive emphysema. The soluble inflammatory biomarkers white blood cell count, fibrinogen, high-sensitivity C-reactive protein and plasma surfactant protein were higher in GOLD 1–3 groups than in the EwAL group. Interleukin-6 was expressed less often in EwAL subjects compared with subjects in the GOLD and A1ATD groups. Soluble receptor for advanced glycation end product was lowest in GOLD 3 subjects, indicative of more severe emphysema. Conclusions These findings provide context for upcoming results from FOOTPRINTS®, which aims to establish correlations between biomarkers and disease progression in a representative COPD population. Trial registration number: NCT02719184, study start date 13/04/2016.

Background: Bradykinin 1 receptor (B1R) signalling pathways may be involved in the inflammatory pathophysiology of chronic obstructive pulmonary disease (COPD). B1R signalling is induced by inflammatory stimuli or tissue injury and leads to activation and increased migration of pro-inflammatory cells. Lipopolysaccharide (LPS) lung challenge in man is an experimental method of exploring inflammation in the lung whereby interference in these pathways can help to assess pharmacologic interventions in COPD. BI 1026706, a potent B1R antagonist, was hypothesized to reduce the inflammatory activity after segmental lipopolysaccharide (LPS) challenge in humans due to decreased pulmonary cell influx. Methods: In a monocentric, randomized, double-blind, placebo-controlled, parallel-group, phase I trial, 57 healthy, smoking subjects were treated for 28 days with either oral BI 1026706 100 mg bid or placebo. At day 21, turbo-inversion recovery magnitude magnetic resonance imaging (TIRM MRI) was performed. On the last day of treatment, pre-challenge bronchoalveolar lavage fluid (BAL) and biopsies were sampled, followed by segmental LPS challenge (40 endotoxin units/kg body weight) and saline control instillation in different lung lobes. Twentyfour hours later, TIRM MRI was performed, then BAL and biopsies were collected from the challenged segments. In BAL samples, cells were differentiated for neutrophil numbers as the primary endpoint. Other endpoints included assessment of safety, biomarkers in BAL (e.g. interleukin-8 [IL-8], albumin and total protein), B1R expression in lung biopsies and TIRM score by MRI as a measure for the extent of pulmonary oedema. Results: After LPS, but not after saline, high numbers of inflammatory cells, predominantly neutrophils were observed in the airways. IL-8, albumin and total protein were also increased in BAL samples after LPS challenge as compared with saline control. There were no significant differences in cells or other biomarkers from BAL in volunteers treated with BI 1026706 compared with those treated with placebo. Unexpectedly, neutrophil numbers in BAL were 30% higher and MRI-derived extent of oedema was significantly higher with BI 1026706 treatment compared with placebo, 24 h after LPS challenge. Adverse events were mainly mild to moderate and not different between treatment groups. Conclusions: Treatment with BI 1026706 for four weeks was safe and well-tolerated in healthy smoking subjects. BI 1026706 100 mg bid did not provide evidence for anti-inflammatory effects in the human bronchial LPS challenge model. Trial registration: The study was registered on January 14, 2016 at ClinicalTrials.gov (NCT02657408).

New therapies are needed to prevent exacerbations, improve quality of life and slow disease progression in bronchiectasis. Inhibition of cathepsin C (CatC) activity has the potential to decrease activation of neutrophil-derived serine proteases in patients with bronchiectasis, thereby reducing airway inflammation, improving symptoms, reducing exacerbations and preventing further airway damage. Here we present the design of a phase 2 trial (Airleaf™; NCT05238675) assessing the efficacy and safety of a novel CatC inhibitor, BI 1291583, in adult patients with bronchiectasis. This multinational, randomised, double-blind, placebo-controlled, parallel-group, dose-finding study has a screening period of at least 6 weeks, a treatment period of 24–48 weeks and a follow-up period of 4 weeks. ∼240 adults with bronchiectasis of multiple aetiologies will be randomised to placebo once daily, or BI 1291583 1 mg once daily, 2.5 mg once daily or 5 mg once daily in a 2:1:1:2 ratio, stratified by Pseudomonas aeruginosa infection and maintenance use of macrolides. The primary efficacy objective is to evaluate the dose–response relationship for the three oral doses of BI 1291583 versus placebo on time to first pulmonary exacerbation up to Week 48 (the primary end-point). Efficacy will be assessed using exacerbations, patient-reported outcomes, measures of symptoms, sputum neutrophil elastase activity and pulmonary function testing. Safety assessment will include adverse event reporting, physical examination, monitoring of vital signs, safety laboratory parameters, 12-lead electrocardiogram, and periodontal and dermatological assessments. If efficacy and safety are demonstrated, results will support further investigation of BI 1291583 in phase 3 trials.

BackgroundDespite the availability of COVID-19 vaccinations, there remains a need to investigate treatments to reduce the risk or severity of potentially fatal complications of COVID-19, such as acute respiratory distress syndrome (ARDS). This study evaluated the efficacy and safety of the transient receptor potential channel C6 (TRPC6) inhibitor, BI 764198, in reducing the risk and/or severity of ARDS in patients hospitalised for COVID-19 and requiring non-invasive, supplemental oxygen support (oxygen by mask or nasal prongs, oxygen by non-invasive ventilation or high-flow nasal oxygen).MethodsMulticentre, double-blind, randomised phase II trial comparing once-daily oral BI 764198 (n=65) with placebo (n=64) for 28 days (+2-month follow-up). Primary endpoint: proportion of patients alive and free of mechanical ventilation at day 29. Secondary endpoints: proportion of patients alive and discharged without oxygen (day 29); occurrence of either in-hospital mortality, intensive care unit admission or mechanical ventilation (day 29); time to first response (clinical improvement/recovery); ventilator-free days (day 29); and mortality (days 15, 29, 60 and 90).ResultsNo difference was observed for the primary endpoint: BI 764198 (83.1%) versus placebo (87.5%) (estimated risk difference –5.39%; 95% CI –16.08 to 5.30; p=0.323). For secondary endpoints, a longer time to first response (rate ratio 0.67; 95% CI 0.46 to 0.99; p=0.045) and longer hospitalisation (+3.41 days; 95% CI 0.49 to 6.34; p=0.023) for BI 764198 versus placebo was observed; no other significant differences were observed. On-treatment adverse events were similar between trial arms and more fatal events were reported for BI 764198 (n=7) versus placebo (n=2). Treatment was stopped early based on an interim observation of a lack of efficacy and an imbalance of fatal events (Data Monitoring Committee recommendation).ConclusionsTRPC6 inhibition was not effective in reducing the risk and/or severity of ARDS in patients with COVID-19 requiring non-invasive, supplemental oxygen support.Trial registration numberNCT04604184.

Background: Uncontrolled neutrophil serine proteases, activated by cathepsin C (CatC), are key drivers of inflammation in bronchiectasis. BI 1291583, a novel CatC inhibitor, will be investigated in two Phase 2 trials in non-cystic fibrosis bronchiectasis (NCFBE; Airleaf™) and cystic fibrosis bronchiectasis (CFBE; Clairafly™). Here, we outline the design of a Phase 2 rollover study (Clairleaf™) assessing long-term safety and efficacy of BI 1291583. Methods: Clairleaf™ will include ~220 patients who have completed the parent trials Airleaf™ or Clairafly™. Part A of Clairleaf™ is a randomised, double-blind study assessing efficacy and safety of three doses of BI 1291583. Patients treated with BI 1291583 in a parent trial will stay on the same dose. Those who received placebo will be randomised to one of the three BI 1291583 doses (1:1:1). The duration of Part A will depend on completion of the parent trials and will likely be no longer than 1 year for the first patients entering this trial. After Part A, patients may continue to Part B where they will receive the preferred Phase 3 dose of BI 1291583 (open-label) to assess long-term safety. The length of Part B will depend on the occurrence or outcome of Phase 3. Results: The primary endpoint is the occurrence of treatment-emergent adverse events during Part A and B. Secondary endpoints are time to first pulmonary exacerbation and rate of pulmonary exacerbations. The trial will have been submitted to health authorities by the time of presentation and is expected to begin in the second half of 2023. Conclusions: This study will help in understanding the long-term safety and efficacy of BI 1291583 in patients with CFBE and NCFBE.

Autoimmune pulmonary alveolar proteinosis (aPAP) is a rare lung disorder involving production of autoantibodies against endogenous granulocyte–macrophage colony-stimulating factor (GM-CSF). This study aimed to identify biomarkers that could be used to monitor for aPAP, particularly in patients treated with anti-GM-CSF antibodies. This was an exploratory, prospective, observational, single-center study. Pre-specified biomarkers were evaluated between baseline and Day 120 in serum/plasma, whole blood, sputum and exhaled breath condensate from patients with aPAP, healthy volunteers, and patients with chronic obstructive pulmonary disease (COPD) and asthma (not treated with anti-GM-CSF and with no evidence of aPAP). Pulmonary function tests were also performed. Overall, 144 individuals were enrolled (aPAP: n = 34, healthy volunteers: n = 24, COPD: n = 40 and asthma: n = 46). Plasma GM-CSF levels were lower, and Krebs von den Lungen 6 and GM-CSF autoantibody ranges were higher, in patients with aPAP compared with other populations. Surfactant proteins-A and -D, lactate dehydrogenase and carcinoembryonic antigen ranges partially or completely overlapped across populations. Most plasma biomarkers showed high sensitivity and specificity for detection of aPAP; GM-CSF and GM-CSF autoantibody concentrations demonstrated equivalent sensitivity for differentiating aPAP. In addition to characteristic GM-CSF autoantibodies, assessment of plasma GM-CSF may identify individuals at risk of developing aPAP. Trial registration: EudraCT, 2012-003475-19. Registered 23 July 2012— https://eudract.ema.europa.eu/ .

Background Inhibition of the epithelial sodium channel (ENaC) in cystic fibrosis (CF) airways provides a mutation-agnostic approach that could improve mucociliary clearance in all CF patients. BI 1265162 is an ENaC inhibitor with demonstrated pre-clinical efficacy and safety already demonstrated in humans. Objective We present results from BALANCE-CF (TM) 1, a phase II, placebo-controlled, randomised, double-blind study of four dose levels of BI 1265162 versus placebo for 4 weeks on top of standard of care in adults and adolescents with CF. Results Initially, 28 randomised subjects (BI 1265162 200 mu g twice daily n=14, placebo twice daily n=14) were assessed at an interim futility analysis. Compared with placebo, numerical changes of -0.8% (95% CI -6.6 to 4.9%) in percentage predicted forced expiratory volume in 1s (ppFEV(1)) and +2.1 units (95% CI -2.4 to 6.5 units) in lung clearance index (LCI) were observed in the active group, meeting a pre-defined stopping rule; accordingly, the study was terminated. Recruitment had continued during the interim analysis and pending results; 24 patients were added across three dose levels and placebo. The final results including these patients (+1.5% ppFEV(1), 200 mu g twice-daily dose versus placebo) were not supportive of relevant clinical effect. Furthermore, LCI change was not supportive, although interpretation was limited due to insufficient traces meeting quality criteria. A 9.4-point improvement in the Cystic Fibrosis Questionnaire - Revised Respiratory Domain was observed in the 200 mu g twice daily dose group versus placebo. BI 1265162 up to 200 mu g twice daily was safe and well-tolerated. Pharmacokinetics were similar to those in healthy volunteers. Conclusion BI 1265162 was safe, but did not demonstrate a potential for clinical benefit. Development has been terminated.

Introduction A better understanding is needed of the different phenotypes that exist for patients with chronic obstructive pulmonary disease (COPD), their relationship with the pathogenesis of COPD and how they may affect disease progression. Biomarkers, including those associated with emphysema, may assist in characterising patients and in predicting and monitoring the course of disease. The FOOTPRINTS study (study 352.2069) aims to identify biomarkers associated with emphysema, over a 3-year period.Methods and analysis The FOOTPRINTS study is a prospective, longitudinal, multinational (12 countries), multicentre (51 sites) biomarker study, which has enrolled a total of 463 ex-smokers, including subjects without airflow limitation (as defined by the 2015 Global Initiative for Chronic Obstructive Lung Disease (GOLD) strategy report), patients with COPD across the GOLD stages 1–3 and patients with COPD and alpha1-antitrypsin deficiency. The study has an observational period lasting 156 weeks that includes seven site visits and additional phone interviews. Biomarkers in blood and sputum, imaging data (CT and magnetic resonance), clinical parameters, medical events of special interest and safety are being assessed at regular visits. Disease progression based on biomarker values and COPD phenotypes are being assessed using multivariate statistical prediction models.Ethics and dissemination The study protocol was approved by the authorities and ethics committees/institutional review boards of the respective institutions where applicable, which included study sites in Belgium, Canada, Denmark, Finland, Germany, Japan, Korea, Poland, Spain, Sweden, UK and USA; written informed consent has been obtained from all study participants. Ethics committee approval was obtained for all participating sites prior to enrolment of the study participants. The study results will be reported in peer-reviewed publications.Trial registration number NCT02719184.

BACKGROUND:Epithelial sodium channel (ENaC) is an important regulator of airway surface liquid volume; ENaC is hyperactivated in cystic fibrosis (CF). ENaC inhibition is a potential therapeutic target for CF. Here, we report in vitro and in vivo results for BI 1265162, an inhaled ENaC inhibitor currently in Phase II clinical development, administered via the Respimat® Soft Mist™ inhaler.METHODS:In vitro inhibition of sodium ion (Na+) transport by BI 1265162 was tested in mouse renal collecting duct cells (M1) and human bronchial epithelial cells (NCI-H441); inhibition of water transport was measured using M1 cells. In vivo inhibition of liquid absorption from rat airway epithelium and acceleration of mucociliary clearance (MCC) in sheep lungs were assessed. Fully differentiated normal and CF human epithelium was used to measure the effect of BI 1265162 with or without ivacaftor and lumacaftor on water transport and MCC.RESULTS:BI 1265162 dose-dependently inhibited Na+ transport and decreased water resorption in cell line models. BI 1265162 reduced liquid absorption in rat lungs and increased MCC in sheep. No effects on renal function were seen in the animal models. BI 1265162 alone and in combination with CF transmembrane conductance regulator (CFTR) modulators decreased water transport and increased MCC in both normal and CF airway human epithelial models and also increased the effects of CFTR modulators in CF epithelium to reach the effect size seen in healthy epithelium with ivacaftor/lumacaftor alone.CONCLUSION:These results demonstrate the potential of BI 1265162 as a mutation agnostic, ENaC-inhibitor-based therapy for CF.

Inhibition of the epithelial sodium channel (ENaC) represents an important, mutation-agnostic therapeutic approach to restore airway surface liquid in patients with cystic fibrosis (CF). A phase II trial of the ENaC inhibitor BI 1265162, inhaled via the Respimat® Soft Mist™ inhaler, in patients aged ≥12 years with CF is being conducted to assess the efficacy and safety of BI 1265162, on top of standard CF treatment (www.clinicaltrials.gov identifier NCT04059094). BALANCE-CF™ 1 is a multinational, randomised, double-blind, placebo-controlled, parallel-group, dose-ranging trial consisting of 2 weeks’ screening, 4 weeks’ randomised treatment and 1 week follow-up. 98 patients, including ≥21 adolescents, will be randomised. First, 28 patients will be allocated to the highest dose of BI 1265162 (200 µg twice daily) or placebo in a 1:1 ratio. The remaining 70 patients will be allocated to one of five treatment arms (200 µg, 100 µg, 50 µg, 20 µg or placebo twice daily), with a final distribution ratio of 2:1:1:1:2. Recruitment and randomisation will begin with adult patients. An independent data monitoring committee will review safety data to advise on inclusion of adolescents and study continuation. A futility analysis will be conducted after 28 patients to prevent exposure of further patients in case of insufficient evidence of clinical efficacy. The design ensures that potential for effect is assessed ahead of wider enrolment, allowing investigation of a dose–response effect with minimal patient numbers. The results will increase understanding of efficacy, safety and optimal dosing of the inhaled ENaC inhibitor BI 1265162 in adults and adolescents with CF.

Background: Methacholine challenges have been used in clinical trials to assess therapeutic effects and potential adverse reactions of interventions on pulmonary function in a sensitive population, such as in subjects with asthma. Here, we evaluate the variability of the methacholine challenge recovery model, and compare the results obtained for both incremental and bolus challenge methods. Methods: The extent, time course and variability of change in forced expiratory volume in 1 s (FEV1) following repeated methacholine challenges in subjects with mild asthma were investigated in an open-label, four-period, fixed-sequence, two-method, replicate crossover study. At Visits 1 and 2, subjects underwent an incremental challenge using doubling doses of methacholine until a >= 20% decrease in FEV1 was observed; at Visits 3 and 4, subjects underwent a bolus challenge, inhaling a single dose of methacholine calculated from the cumulative dose established during Visit 1. Results: A total of 19 subjects were included in the study. Both the mean FEV1 area under the curve (FEV1 AUC(0-tz)) and mean maximum reductions in FEV1 (absolute and relative) 120 min post-challenge values were higher for the incremental challenges than the bolus challenges, with no reported difference between repetitions of the same methodology. FEV1 AUC(0-tz) decrease 120 min post challenge demonstrated an intra-subject coefficient of variation (CV) of 47.2% (incremental) and 78.3% (bolus), suggesting considerable between-visit variability. The mean absolute, and similarly relative, maximum reductions in FEV1 compared with post-diluent baseline values demonstrated lower intra-subject variability (incremental 21.16%, bolus 40.67%) than the FEV1 AUC(0-tz)-based endpoint. There was a trend towards faster recovery following the bolus challenge than with the incremental challenge. The provocative dose of methacholine inducing a >= 20% decrease in FEV1 resulted in a between-group mean difference of 27.20% in the incremental challenge periods, with a high intra-subject CV of 80.64%, demonstrating considerable variability. Conclusion: Maximum reduction in FEV1 had the lowest variability. There was little difference between repetitions of the same methodology, as indicated by overlapping confidence intervals. There was a trend towards faster recovery following bolus challenge than with the incremental challenge. The results of this trial could be of value when designing future clinical trials using the methacholine challenge methodology.