Clinical and transcriptomic features of persistent exacerbation-prone severe asthma in U-BIOPRED cohort

Background: Exacerbation-prone asthma is a feature of severe disease. Yet, the basis for its persistency remains unclear. Objectives: To determine the clinical and transcriptomic features of the frequent-exacerbator (FE) and of persistent FEs (PFE) in U-BIOPRED cohort. Methods: We compared features of FE (≥2 exacerbations in past year) to infrequent exacerbators (IE, <2 exacerbations) and of PFE with repeat ≥2 exacerbations during the following year to persistent IE (PIE). Transcriptomic data in blood, bronchial and nasal epithelial brushings, bronchial biopsies and sputum cells were analysed by gene set variation analysis for 103 gene signatures. Results: Of 317 patients, 62.4 % were FE of whom 63.6% were PFE, while 37.6% were IE of whom 61.3% were PIE. Using multivariate analysis, FE was associated with short-acting beta-agonist use, sinusitis and daily oral corticosteroid use, while PFE with eczema, short-acting beta-agonist use and asthma control index. CEA Cell Adhesion Molecule 5 (CEACAM5) was the only differentially-expressed transcript in bronchial biopsies between PE and IE. There were no differentially-expressed genes in the other 4 compartments. There were higher expression scores for Type 2 , T-helper type-17 and Type 1 pathway signatures together with those associated with viral infections in bronchial biopsies from FE compared to IE, while higher expression scores of Type 2, Type 1 and steroid insensitivity pathway signatures in bronchial biopsies of PFE compared to PIE. Conclusion: FE group and its PFE subgroup are associated with poor asthma control while expressing higher Type 1 and Type 2 activation pathways compared to IE and PIE, respectively

Sputum proteomics and airway cell transcripts of current and ex-smokers with severe asthma in U-BIOPRED: an exploratory analysis

Background: Severe asthma patients with a significant smoking history have airflow obstruction with reported neutrophilia. We hypothesise that multi-omic analysis will enable the definition of smoking and ex-smoking severe asthma molecular phenotypes. Methods: The U-BIOPRED severe asthma patients containing current-smokers (CSA), exsmokers (ESA), non-smokers (NSA) and healthy non-smokers (NH) was examined. Blood and sputum cell counts, fractional exhaled nitric oxide and spirometry were obtained. Exploratory proteomic analysis of sputum supernatants and transcriptomic analysis of bronchial brushings, biopsies and sputum cells was performed. Results: Colony stimulating factor (CSF)2 protein levels were increased in CSA sputum supernatants with azurocidin 1, neutrophil elastase and CXCL8 upregulated in ESA. Phagocytosis and innate immune pathways were associated with neutrophilic inflammation in ESA. Gene Set Variation Analysis of bronchial epithelial cell transcriptome from CSA showed enrichment of xenobiotic metabolism, oxidative stress and endoplasmic reticulum stress compared to other groups. CXCL5 and matrix metallopeptidase 12 genes were upregulated in ESA and the epithelial protective genes, mucin 2 and cystatin SN, were downregulated. Conclusion: Despite little difference in clinical characteristics, CSA were distinguishable from ESA subjects at the sputum proteomic level with CSA having increased CSF2 expression and ESA patients showed sustained loss of epithelial barrier processes

Severe asthma exists despite suppressed tissue inflammation: findings of the U-BIOPRED study

The U-BIOPRED study is a multicentre European study aimed at a better understanding of severe asthma. It included three steroid-treated adult asthma groups (severe nonsmokers (SAn group), severe current/ex-smokers (SAs/ex group) and those with mild–moderate disease (MMA group)) and healthy controls (HC group). The aim of this cross-sectional, bronchoscopy substudy was to compare bronchial immunopathology between these groups. In 158 participants, bronchial biopsies and bronchial epithelial brushings were collected for immunopathologic and transcriptomic analysis. Immunohistochemical analysis of glycol methacrylate resin-embedded biopsies showed there were more mast cells in submucosa of the HC group (33.6 mm⁻ ²) compared with both severe asthma groups (SAn: 17.4 mm⁻ ², p<0.001; SAs/ex: 22.2 mm⁻ ², p=0.01) and with the MMA group (21.2 mm⁻ ², p=0.01). The number of CD4+ lymphocytes was decreased in the SAs/ex group (4.7 mm⁻ ²) compared with the SAn (11.6 mm⁻ ², p=0.002), MMA (10.1 mm⁻ ², p=0.008) and HC (10.6 mm⁻ ², p<0.001) groups. No other differences were observed. Affymetrix microarray analysis identified seven probe sets in the bronchial brushing samples that had a positive relationship with submucosal eosinophils. These mapped to COX-2 (cyclo-oxygenase-2), ADAM-7 (disintegrin and metalloproteinase domain-containing protein 7), SLCO1A2 (solute carrier organic anion transporter family member 1A2), TMEFF2 (transmembrane protein with epidermal growth factor like and two follistatin like domains 2) and TRPM-1 (transient receptor potential cation channel subfamily M member 1); the remaining two are unnamed. We conclude that in nonsmoking and smoking patients on currently recommended therapy, severe asthma exists despite suppressed tissue inflammation within the proximal airway wall

Epithelial IL-6 trans-signaling defines a new asthma phenotype with increased airway inflammation

Background: Although several studies link high levels of IL-6 and soluble IL-6 receptor (sIL-6R) to asthma severity and decreased lung function, the role of IL-6 trans-signaling (IL-6TS) in asthmatic patients is unclear. Objective: We sought to explore the association between epithelial IL-6TS pathway activation and molecular and clinical phenotypes in asthmatic patients. Methods: An IL-6TS gene signature obtained from air-liquid interface cultures of human bronchial epithelial cells stimulated with IL-6 and sIL-6R was used to stratify lung epithelial transcriptomic data (Unbiased Biomarkers in Prediction of Respiratory Disease Outcomes [U-BIOPRED] cohorts) by means of hierarchical clustering. IL-6TS-specific protein markers were used to stratify sputum biomarker data (Wessex cohort). Molecular phenotyping was based on transcriptional profiling of epithelial brushings, pathway analysis, and immunohistochemical analysis of bronchial biopsy specimens. Results: Activation of IL-6TS in air-liquid interface cultures reduced epithelial integrity and induced a specific gene signature enriched in genes associated with airway remodeling. The IL-6TS signature identified a subset of patients with IL-6TS-high asthma with increased epithelial expression of IL-6TS-inducible genes in the absence of systemic inflammation. The IL-6TS-high subset had an overrepresentation of frequent exacerbators, blood eosinophilia, and submucosal infiltration of T cells and macrophages. In bronchial brushings Toll-like receptor pathway genes were upregulated, whereas expression of cell junction genes was reduced. Sputum sIL-6R and IL-6 levels correlated with sputum markers of remodeling and innate immune activation, in particular YKL-40, matrix metalloproteinase 3, macrophage inflammatory protein 1 beta, IL-8, and IL-1 beta. Conclusions: Local lung epithelial IL-6TS activation in the absence of type 2 airway inflammation defines a novel subset of asthmatic patients and might drive airway inflammation and epithelial dysfunction in these patients.Peer reviewe

Epithelial IL-6 trans-signaling defines a new asthma phenotype with increased airway inflammation

Background: Although several studies link high levels of IL-6 and soluble IL-6 receptor (sIL-6R) with asthma severity and decreased lung function, the role of IL-6 trans-signaling (IL-6TS) in asthma is unclear. Objective: To explore the association between epithelial IL-6TS pathway activation and molecular and clinical phenotypes in asthma. Methods: An IL-6TS gene signature, obtained from air-liquid interface (ALI) cultures of human bronchial epithelial cells stimulated with IL-6 and sIL-6R, was used to stratify lung epithelium transcriptomic data (U-BIOPRED cohorts) by hierarchical clustering. IL-6TS-specific protein markers were used to stratify sputum biomarker data (Wessex cohort). Molecular phenotyping was based on transcriptional profiling of epithelial brushings, pathway analysis and immunohistochemical analysis of bronchial biopsies. Results: Activation of IL-6TS in ALI cultures reduced epithelial integrity and induced a specific gene signature enriched in genes associated with airway remodeling. The IL-6TS signature identified a subset of IL-6TS. High asthma patients with increased epithelial expression of IL-6TS inducible genes in absence of systemic inflammation. The IL-6TS High subset had an overrepresentation of frequent exacerbators, blood eosinophilia, and submucosal infiltration of T cells and macrophages. In bronchial brushings, TLR pathway genes were up-regulated while the expression of tight junction genes was reduced. Sputum sIL-6R and IL-6 levels correlated with sputum markers of remodeling and innate immune activation, in particular YKL-40, MMP3, MIP-1β, IL-8 and IL-1β. Conclusions: Local lung epithelial IL-6TS activation in absence of type 2 airway inflammation defines a novel subset of asthmatics and may drive airway inflammation and epithelial dysfunction in these patients

Selective inhibition of prostaglandin D-2 biosynthesis in human mast cells to overcome need for multiple receptor antagonists : Biochemical consequences

Background The major mast cell prostanoid PGD(2) is targeted for therapy of asthma and other diseases, because the biological actions include bronchoconstriction, vasodilation and regulation of immune cells mediated by three different receptors. It is not known if the alternative to selectively inhibit the biosynthesis of PGD(2) affects release of other prostanoids in human mast cells. Objectives To determine the biochemical consequences of inhibition of the hematopoietic prostaglandin D synthase (hPGDS) PGD(2) in human mast cells. Methods Four human mast cell models, LAD2, cord blood derived mast cells (CBMC), peripheral blood derived mast cells (PBMC) and human lung mast cells (HLMC), were activated by anti-IgE or ionophore A23187. Prostanoids were measured by UPLC-MS/MS. Results All mast cells almost exclusively released PGD(2) when activated by anti-IgE or A23187. The biosynthesis was in all four cell types entirely initiated by COX-1. When pharmacologic inhibition of hPGDS abolished formation of PGD(2), PGE(2) was detected and release of TXA(2) increased. Conversely, when the thromboxane synthase was inhibited, levels of PGD(2) increased. Adding exogenous PGH(2) confirmed predominant conversion to PGD(2) under control conditions, and increased levels of TXB2 and PGE(2) when hPGDS was inhibited. However, PGE(2) was formed by non-enzymatic degradation. Conclusions Inhibition of hPGDS effectively blocks mast cell dependent PGD(2) formation. The inhibition was associated with redirected use of the intermediate PGH(2) and shunting into biosynthesis of TXA(2). However, the levels of TXA(2) did not reach those of PGD(2) in naive cells. It remains to determine if this diversion occurs in vivo and has clinical relevance

Nordic consensus statement on the systematic assessment and management of possible severe asthma in adults

Although a minority of asthma patients suffer from severe asthma, they represent a major clinical challenge in terms of poor symptom control despite high-dose treatment, risk of exacerbations, and side effects. Novel biological treatments may benefit patients with severe asthma, but are expensive, and are only effective in appropriately targeted patients. In some patients, symptoms are driven by other factors than asthma, and all patients with suspected severe asthma ('difficult asthma') should undergo systematic assessment, in order to differentiate between true severe asthma, and 'difficult-to-treat' patients, in whom poor control is related to factors such as poor adherence or co-morbidities. The Nordic Consensus Statement on severe asthma was developed by the Nordic Severe Asthma Network, consisting of members from Norway, Sweden, Finland, Denmark, Iceland and Estonia, including representatives from the respective national respiratory scientific societies with the aim to provide an overview and recommendations regarding the diagnosis, systematic assessment and management of severe asthma. Furthermore, the Consensus Statement proposes recommendations for the organization of severe asthma management in primary, secondary, and tertiary care.Peer reviewe

Inhaled allergen bronchoprovocation tests

<p>The allergen bronchoprovocation test is a long-standing exacerbation model of allergic asthma that can induce several clinical and pathophysiologic features of asthma in sensitized subjects. Standardized allergen challenge is primarily a research tool, and when properly conducted by qualified and experienced investigators, it is safe and highly reproducible. In combination with validated airway sampling and sensitive detection techniques, allergen challenge allows the study of several features of the physiology of mainly T(H)2 cell-driven asthma in relation to the kinetics of the underlying airway pathology occurring during the allergen-induced late response. Furthermore, given the small within-subject variability in allergen-induced airway responses, allergen challenge offers an adequate disease model for the evaluation of new (targeted) controller therapies for asthma in a limited number of subjects. In proof-of-efficacy studies thus far, allergen challenge showed a fair positive predicted value and an excellent negative predictive value for the actual clinical efficacy of new antiasthma therapies, underscoring its important role in early drug development. In this review we provide recommendations on challenge methods, response measurements, sample size, safety, and harmonization for future applications.</p>