A novel quinoline with airway relaxant effects and anti-inflammatory properties.

BACKGROUND: In chronic pulmonary diseases characterized by inflammation and airway obstruction, such as asthma and COPD, there are unmet needs for improved treatment. Quinolines is a group of small heterocyclic compounds that have a broad range of pharmacological properties. Here, we investigated the airway relaxant and anti-inflammatory properties of a novel quinoline (RCD405). METHODS: The airway relaxant effect of RCD405 was examined in isolated airways from humans, dogs, rats and mice. Murine models of ovalbumin (OVA)-induced allergic asthma and LPS-induced airway inflammation were used to study the effects in vivo. RCD405 (10 mg/kg) or, for comparisons in selected studies, budesonide (3 mg/kg), were administered intratracheally 1 h prior to each challenge. Airway responsiveness was determined using methacholine provocation. Immune cell recruitment to bronchi was measured using flow cytometry and histological analyses were applied to investigate cell influx and goblet cell hyperplasia of the airways. Furthermore, production of cytokines and chemokines was measured using a multiplex immunoassay. The expression levels of asthma-related genes in murine lung tissue were determined by PCR. The involvement of NF-κB and metabolic activity was measured in the human monocytic cell line THP-1. RESULTS: RCD405 demonstrated a relaxant effect on carbachol precontracted airways in all four species investigated (potency ranking: human = rat > dog = mouse). The OVA-specific IgE and airway hyperresponsiveness (AHR) were significantly reduced by intratracheal treatment with RCD405, while no significant changes were observed for budesonide. In addition, administration of RCD405 to mice significantly decreased the expression of proinflammatory cytokines and chemokines as well as recruitment of immune cells to the lungs in both OVA- and LPS-induced airway inflammation, with a similar effect as for budesonide (in the OVA-model). However, the effect on gene expression of Il-4, IL-5 and Il-13 was more pronounced for RCD405 as compared to budesonide. Finally, in vitro, RCD405 reduced the LPS-induced NF-κB activation and by itself reduced cellular metabolism. CONCLUSIONS: RCD405 has airway relaxant effects, and it reduces AHR as well as airway inflammation in the models used, suggesting that it could be a clinically relevant compound to treat inflammatory airway diseases. Possible targets of this compound are complexes of mitochondrial oxidative phosphorylation, resulting in decreased metabolic activity of targeted cells as well as through pathways associated to NF-κB. However, further studies are needed to elucidate the mode of action.

Macrophage expressed tartrate-resistant acid phosphatase 5 promotes pulmonary fibrosis progression.

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disorder involving scarring of pulmonary tissue and a subsequent decrease in respiratory capacity, ultimately resulting in death. Tartrate resistant acid phosphatase 5 (ACP5) plays a role in IPF but the exact mechanisms are yet to be elucidated. In this study, we have utilized various perturbations of the bleomycin mouse model of IPF including genetic knockout, RANKL inhibition, and macrophage adoptive transfer to further understand ACP5's role in pulmonary fibrosis. Genetic ablation of Acp5 decreased immune cell recruitment to the lungs and reduced the levels of hydroxyproline (reflecting extracellular matrix-production) as well as histological damage. Additionally, gene expression profiling of murine lung tissue revealed downregulation of genes including Ccl13, Mmp13, and Il-1α that encodes proteins specifically related to immune cell recruitment and macrophage/fibroblast interactions. Furthermore, antibody-based neutralization of RANKL, an important inducer of Acp5 expression, reduced immune cell recruitment but did not decrease fibrotic lung development. Adoptive transfer of Acp5-/- bone marrow-derived monocyte (BMDM) macrophages 7 or 14 days after bleomycin administration resulted in reductions of cytokine production and decreased levels of lung damage, compared to adoptive transfer of WT control macrophages. Taken together, the data presented in this study suggest that macrophage derived ACP5 plays an important role in development of pulmonary fibrosis and could present a tractable target for therapeutic intervention in IPF.

High prevalence of interstitial lung abnormalities in middle-aged never-smokers.

Background: Interstitial lung abnormalities (ILA) are incidental findings on chest computed tomography (CT). These patterns can present at an early stage of fibrotic lung disease. Our aim was to estimate the prevalence of ILA in the Swedish population, in particular in never-smokers, and find out its association with demographics, comorbidities and symptoms. Methods: Participants were recruited to the Swedish CArdioPulmonary BioImage Study (SCAPIS), a population-based survey including men and women aged 50-64 years performed at six university hospitals in Sweden. CT scan, spirometry and questionnaires were performed. ILA were defined as cysts, ground-glass opacities, reticular abnormality, bronchiectasis and honeycombing. Findings: Out of 29 521 participants, 14 487 were never-smokers and 14 380 were men. In the whole population, 2870 (9.7%) had ILA of which 134 (0.5%) were fibrotic. In never-smokers, the prevalence was 7.9% of which 0.3% were fibrotic. In the whole population, age, smoking history, chronic bronchitis, cancer, coronary artery calcium score and high-sensitive C-reactive protein were associated with ILA. Both ILA and fibrotic ILA were associated with restrictive spirometric pattern and impaired diffusing capacity of the lung for carbon monoxide. However, individuals with ILA did not report more symptoms compared with individuals without ILA. Interpretation: ILA are common in a middle-aged Swedish population including never-smokers. ILA may be at risk of being underdiagnosed among never-smokers since they are not a target for screening.

Substrate-specific binding of 8-oxoguanine DNA glycosylase 1 (OGG1) reprograms mucosal adaptations to chronic airway injury.

Recent advances have uncovered the non-random distribution of 7, 8-dihydro-8-oxoguanine (8-oxoGua) induced by reactive oxygen species, which is believed to have epigenetic effects. Its cognate repair protein, 8-oxoguanine DNA glycosylase 1 (OGG1), reads oxidative substrates and participates in transcriptional initiation. When redox signaling is activated in small airway epithelial cells, the DNA repair function of OGG1 is repurposed to transmit acute inflammatory signals accompanied by cell state transitions and modification of the extracellular matrix. Epithelial-mesenchymal and epithelial-immune interactions act cooperatively to establish a local niche that instructs the mucosal immune landscape. If the transitional cell state governed by OGG1 remains responsive to inflammatory mediators instead of differentiation, the collateral damage provides positive feedback to inflammation, ascribing inflammatory remodeling to one of the drivers in chronic pathologies. In this review, we discuss the substrate-specific read through OGG1 has evolved in regulating the innate immune response, controlling adaptations of the airway to environmental and inflammatory injury, with a focus on the reader function of OGG1 in initiation and progression of epithelial to mesenchymal transitions in chronic pulmonary disease.

8-Oxoguanine targeted by 8-oxoguanine DNA glycosylase 1 (OGG1) is central to fibrogenic gene activation upon lung injury.

Reactive oxygen species (ROS) are implicated in epithelial cell-state transition and deposition of extracellular matrix upon airway injury. Of the many cellular targets of ROS, oxidative DNA modification is a major driving signal. However, the role of oxidative DNA damage in modulation profibrotic processes has not been fully delineated. Herein, we report that oxidative DNA base lesions, 8-oxoG, complexed with 8-oxoguanine DNA glycosylase 1 (OGG1) functions as a pioneer factor, contributing to transcriptional reprogramming within airway epithelial cells. We show that TGFß1-induced ROS increased 8-oxoG levels in open chromatin, dynamically reconfigure the chromatin state. OGG1 complexed with 8-oxoG recruits transcription factors, including phosphorylated SMAD3, to pro-fibrotic gene promoters thereby facilitating gene activation. Moreover, 8-oxoG levels are elevated in lungs of mice subjected to TGFß1-induced injury. Pharmacologic targeting of OGG1 with the selective small molecule inhibitor of 8-oxoG binding, TH5487, abrogates fibrotic gene expression and remodeling in this model. Collectively, our study implicates that 8-oxoG substrate-specific binding by OGG1 is a central modulator of transcriptional regulation in response to tissue repair.

Trends in Treatments With Disease-Specific and Interfering Drugs in Patients With Hereditary Angioedema in Sweden.

BACKGROUND: Hereditary angioedema (HAE) is caused by low levels of or defects in C1 inhibitor. Although disease activity may be modified by prophylaxis, emergency treatment, treatment for comorbidities, and oral contraceptives, the extent of their use is unclear. OBJECTIVE: To investigate trends in the use of disease-specific and interfering drugs in patients with HAE compared with the general population in Sweden. METHODS: In a nationwide, longitudinal study, 239 patients with HAE and 2 383 controls were compared with the Prescribed Drug Register (2005-2019). These data reflect rates of dispensed prescriptions from pharmacies in Sweden. RESULTS: Attenuated androgens were used by approximately 10% of patients with HAE. The number of individuals treated with prophylactic plasma-derived C1 inhibitor increased during this period to reach almost 25% in men and 35% in women in 2019. Tranexamic acid was prescribed to 5% to 15% of patients, primarily children and young adults. Rates of prescriptions for icatibant, an emergency medication, showed a steady increase since its introduction in 2010, in particular among middle-aged women, suggesting poorly controlled disease. The use of diuretics, calcium channel blockers, and gestagens was more common in patients with HAE than in controls, whereas angiotensin-converting enzyme inhibitors were rarely collected. CONCLUSIONS: Despite concerns regarding side effects, approximately 10% of patients with HAE received attenuated androgens for long-term prophylaxis. The common use of emergency medication also suggests poorly controlled disease in many patients, highlighting the need for increased focus on prophylactic treatment.

Tartrate resistant acid phosphatase 5 (TRAP5) mediates immune cell recruitment in a murine model of pulmonary bacterial infection.

Introduction: During airway infection, upregulation of proinflammatory cytokines and subsequent immune cell recruitment is essential to mitigate bacterial infection. Conversely, during prolonged and non-resolving airway inflammation, neutrophils contribute to tissue damage and remodeling. This occurs during diseases including cystic fibrosis (CF) and COPD where bacterial pathogens, not least Pseudomonas aeruginosa, contribute to disease progression through long-lasting infections. Tartrate-resistant acid phosphatase (TRAP) 5 is a metalloenzyme expressed by alveolar macrophages and one of its target substrates is the phosphoglycoprotein osteopontin (OPN). Methods: We used a knockout mouse strain (Trap5-/-) and BALB/c-Tg (Rela-luc)31Xen mice paired with siRNA administration or functional protein add-back to elucidate the role of Trap5 during bacterial infection. In a series of experiments, Trap5-/- and wild-type control mice received intratracheal administration of P.aerugniosa (Xen41) or LPS, with mice monitored using intravital imaging (IVIS). In addition, multiplex cytokine immunoassays, flow cytometry, multispectral analyses, histological staining were performed. Results: In this study, we found that Trap5-/- mice had impaired clearance of P. aeruginosa airway infection and reduced recruitment of immune cells (i.e. neutrophils and inflammatory macrophages). Trap5 knockdown using siRNA resulted in a decreased activation of the proinflammatory transcription factor NF-κB in reporter mice and a subsequent decrease of proinflammatory gene expression. Add-back experiments of enzymatically active TRAP5 to Trap5-/- mice restored immune cell recruitment and bacterial killing. In human CF lung tissue, TRAP5 of alveolar macrophages was detected in proximity to OPN to a higher degree than in normal lung tissue, indicating possible interactions. Discussion: Taken together, the findings of this study suggest a key role for TRAP5 in modulating airway inflammation. This could have bearing in diseases such as CF and COPD where excessive neutrophilic inflammation could be targeted by pharmacological inhibitors of TRAP5.

Pharmacological OGG1 inhibition decreases murine allergic airway inflammation.

Background and aim: Allergic asthma is a complex inflammatory disease involving type 2 innate lymphoid cells, type 2 T helper cells, macrophages, and eosinophils. The disease is characterized by wheezing, dyspnea, coughing, chest tightness and variable airflow limitation for which there is no cure and is symptomatically treated with inhaled corticosteroids and ß2-agonists. Molecular mechanisms underlying its complex pathogenesis are not fully understood. However, 8-oxoguanine DNA glycosylase-1 (OGG1), a DNA repair protein may play a central role, as OGG1 deficiency decreases both innate and allergic inflammation. Methods: Using a murine ovalbumin (OVA) model of allergic airway inflammation we assessed the utility of an inhibitor of OGG1 (TH5487) in this disease context. Cytokines and chemokines, promoting immune cell recruitment were measured using a 23-multiplex assay and Western blotting. Additionally, immune cell recruitment to bronchi was measured using flow cytometry. Histological analyses and immunofluorescent staining were used to confirm immune cell influx and goblet cell hyperplasia of the airways. A PCR array was used to assess asthma-related genes in murine lung tissue following TH5487 treatment. Finally, airway hyperresponsiveness was determined using in vivo lung function measurement. Results: In this study, administration of TH5487 to mice with OVA-induced allergic airway inflammation significantly decreased goblet cell hyperplasia and mucus production. TH5487 treatment also decreased levels of activated NF-κB and expression of proinflammatory cytokines and chemokines resulting in significantly lower recruitment of eosinophils and other immune cells to the lungs. Gene expression profiling of asthma and allergy-related proteins after TH5487 treatment revealed differences in several important regulators, including down regulation of Tnfrsf4, Arg1, Ccl12 and Ccl11, and upregulation of the negative regulator of type 2 inflammation, Bcl6. Furthermore, the gene Clca1 was upregulated following TH5487 treatment, which should be explored further due to its ambiguous role in allergic asthma. In addition, the OVA-induced airway hyperresponsiveness was significantly reduced by TH5487 treatment. Conclusion: Taken together, the data presented in this study suggest OGG1 as a clinically relevant pharmacological target for the treatment of allergic inflammation.

Zoledronic Acid Targeting of the Mevalonate Pathway Causes Reduced Cell Recruitment and Attenuates Pulmonary Fibrosis.

Background and aim: Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease causing irreparable scarring of lung tissue, with most patients succumbing rapidly after diagnosis. The mevalonate pathway, which is involved in the regulation of cell proliferation, survival, and motility, is targeted by the bisphosphonate zoledronic acid (ZA). The aim of this study was to assess the antifibrotic effects of ZA and to elucidate the mechanisms by which potential IPF treatment occurs. Methods: A series of in vitro and in vivo models were employed to identify the therapeutic potential of ZA in treating IPF. In vitro transwell assays were used to assess the ability of ZA to reduce fibrotic-related immune cell recruitment. Farnesyl diphosphate synthase (FDPS) was screened as a potential antifibrotic target using a bleomycin mouse model. FDPS-targeting siRNA and ZA were administered to mice following the onset of experimentally-induced lung fibrosis. Downstream analyses were conducted on murine lung tissues and lung fluids including 23-plex cytokine array, flow cytometry, histology, Western blotting, immunofluorescent staining, and PCR analysis. Results: In vitro administration of ZA reduced myofibroblast transition and blocked NF-κB signaling in macrophages leading to impaired immune cell recruitment in a transwell assay. FDPS-targeting siRNA administration significantly attenuated profibrotic cytokine production and lung damage in a murine lung fibrosis model. Furthermore, ZA treatment of mice with bleomycin-induced lung damage displayed decreased cytokine levels in the BALF, plasma, and lung tissue, resulting in less histologically visible fibrotic scarring. Bleomycin-induced upregulation of the ZA target, FDPS, was reduced in lung tissue and fibroblasts upon ZA treatment. Confirmatory increases in FDPS immunoreactivity was seen in human IPF resected lung samples compared to control tissue indicating potential translational value of the approach. Additionally, ZA polarized macrophages towards a less profibrotic phenotype contributing to decreased IPF pathogenesis. Conclusion: This study highlights ZA as an expedient and efficacious treatment option against IPF in a clinical setting.

The effect of the COVID-19 pandemic on severe asthma care in Europe: will care change for good?

Background: The coronavirus disease 2019 (COVID-19) pandemic has put pressure on healthcare services, forcing the reorganisation of traditional care pathways. We investigated how physicians taking care of severe asthma patients in Europe reorganised care, and how these changes affected patient satisfaction, asthma control and future care. Methods: In this European-wide cross-sectional study, patient surveys were sent to patients with a physician-diagnosis of severe asthma, and physician surveys to severe asthma specialists between November 2020 and May 2021. Results: 1101 patients and 268 physicians from 16 European countries contributed to the study. Common physician-reported changes in severe asthma care included use of video/phone consultations (46%), reduced availability of physicians (43%) and change to home-administered biologics (38%). Change to phone/video consultations was reported in 45% of patients, of whom 79% were satisfied or very satisfied with this change. Of 709 patients on biologics, 24% experienced changes in biologic care, of whom 92% were changed to home-administered biologics and of these 62% were satisfied or very satisfied with this change. Only 2% reported worsening asthma symptoms associated with changes in biologic care. Many physicians expect continued implementation of video/phone consultations (41%) and home administration of biologics (52%). Conclusions: Change to video/phone consultations and home administration of biologics was common in severe asthma care during the COVID-19 pandemic and was associated with high satisfaction levels in most but not all cases. Many physicians expect these changes to continue in future severe asthma care, though satisfaction levels may change after the pandemic.

Comorbidities in hereditary angioedema-A population-based cohort study.

BACKGROUND: In hereditary angioedema (HAE), low levels (type 1) or defect in function (type 2) of the serine-protease inhibitor C1 Inhibitor protein results in activation of the classical pathway of the complement system as well as the contact system. Here, we investigated the risk of comorbidities in HAE. METHODS: Individuals with HAE (n = 239; identified through a physician made diagnosis) and a control cohort from the general population (n = 2383; matched for age, gender, and county of residence) were compared with the Swedish inpatient, cause of death, cancer, and prescription registers. Conditional logistic regression was used to analyze the data. RESULTS: Increased risk of cardiovascular disease (odds ratio [OR] 1.83; 95% confidence interval [CI] 1.32-2.54), including arterial (OR 6.74; 95% CI 1.89-24.06) and venous thromboembolic disease (OR 4.20; 95% CI 2.42-7.23) as well as hypertension (OR 1.64; 95% CI 1.12-2.39) was seen in HAE. There was also an increased number of individuals diagnosed with hyperlipidemia (OR 2.01; 95% CI 1.16-3.50) among HAE patients. Furthermore, the risk of autoimmune disease was increased (OR 1.65; 95% CI 1.15-2.35) being particularly pronounced for systemic lupus erythematosus (OR 71.87; 95% CI 8.80-586.7). The risk of having two or more autoimmune diseases was also higher among HAE patients (p = 0.017). In contrast, the risk of cancer was not increased. Data from the prescription register revealed higher prescription rates of drugs against hypertension, hypothyroidism, and hyperlipidemia among HAE patients. CONCLUSIONS: The results warrant for awareness and prevention of comorbid conditions, in particular, thromboembolic and autoimmune diseases in HAE. Future prophylactic interventions may modify these risks.

Citrullination of extracellular histone H3.1 reduces antibacterial activity and exacerbates its proteolytic degradation.

BACKGROUND: Cystic fibrosis (CF), involves excessive airway accumulation of neutrophils, often in parallel with severe infection caused by Pseudomonas aeruginosa. Free histones are known to possess bactericidal properties, but the degree of antibacterial activity exerted on specific lung-based pathogens is largely unknown. Neutrophils have a high content of peptidyl deiminase 4 (PADI4), which citrullinate cationic peptidyl-arginines. In histone H3.1, several positions in the NH2-terminal tail are subject to citrullination. METHODS: Full-length and segmented histone subunit H3.1 was investigated for bactericidal activity towards P. aeruginosa (strain PAO1). PADI4-induced citrullination of histone H3.1 was assessed for antibacterial activity towards P. aeruginosa. Next, the effect of neutrophil elastase (NE)-mediated proteolysis of histone H3.1 was investigated. Finally, PADI4, H3.1, and citrullinated H3.1 were examined in healthy control and CF patient lung tissues. RESULTS: Full-length histone H3.1 and sections of the histone H3.1 tail, displayed bactericidal activity towards P. aeruginosa. These antibacterial effects were reduced following citrullination by PADI4 or proteolysis by NE. Interestingly, citrullination of histone H3.1 exacerbated NE-mediated degradation. In CF lung tissue, citrullinated histone H3.1 and PADI4 immunoreactivity was abundant. Degraded histone H3.1 was detected in the sputum of CF patients but was absent in the sputum of healthy controls. CONCLUSIONS: Citrullination impairs the antibacterial activity of histone H3.1 and exacerbates its proteolytic degradation by NE. Citrullination is likely to play an important role during resolution of acute inflammation. However, in chronic inflammation akin to CF, citrullination may dampen host defense and promote pathogen survival, as exemplified by P. aeruginosa.