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Background: The most recognizable phenotype of severe asthma comprises people who are blood eosinophil and FeNO-high, driven by type-2 (T2) cytokine biology which responds to targeted biological therapies. However, in many people with severe asthma, these T2 biomarkers are suppressed but poorly controlled asthma persists. The mechanisms driving asthma in the absence of T2 biology are poorly understood. Objectives: To explore airway pathology in T2 biomarker-high and -low severe asthma. Methods: T2 biomarker-high severe asthma (T2-high, n=17) was compared to biomarker-intermediate (T2-intermediate, n=21) and biomarker-low (T2-low, n=20) severe asthma, and healthy controls (n=28). Bronchoscopy samples were processed for immunohistochemistry, and sputum for cytokines, PGD2 and LTE4 measurements. Results: Tissue eosinophil, neutrophil and mast cell counts were similar across severe asthma phenotypes and not increased when compared to healthy controls. In contrast, the remodeling features of airway smooth muscle mass and MUC5AC expression were increased in all asthma groups compared to health, but similar across asthma subgroups. Submucosal glands were increased in T2-intermediate and T2-low asthma. In spite of similar tissue cellular inflammation, sputum IL-4, IL-5, and CCL26 were increased in T2-high versus T2-low asthma, and several further T2-associated cytokines, PGD2 and LTE4, were increased in T2-high and T2-intermediate asthma compared to healthy controls. Conclusions: Eosinophilic tissue inflammation within proximal airways is suppressed in T2 biomarker-high and T2-low severe asthma, but inflammatory and structural cell activation is present, with sputum T2-associated cytokines highest in T2 biomarker-high patients. Airway remodeling persists, and may be important for residual disease expression beyond eosinophilic exacerbations.

Chronic rhinosinusitis with nasal polyps (CRSwNP) is generally associated with eosinophilic tissue infiltration linked to type 2 inflammation and characterized by elevated levels of interleukin (IL)-5 and other type 2 inflammatory mediators. Although distinct and overlapping contributions of eosinophils and IL-5 to CRSwNP pathology are still being explored, they are both known to play an important role in NP inflammation. Eosinophils secrete numerous type 2 inflammatory mediators including granule proteins, enzymes, cytokines, chemokines, growth factors, lipids, and oxidative products. IL-5 is critical for the differentiation, migration, activation, and survival of eosinophils but is also implicated in the biological functions of mast cells, basophils, innate lymphoid cells, B cells, and epithelial cells. Results from clinical trials of therapeutics that target type 2 inflammatory mediators (including but not limited to anti-IL-5, anti-immunoglobulin-E, and anti-IL-4/13) may provide further evidence of how eosinophils and IL-5 contribute to CRSwNP. Finally, the association between eosinophilia/elevated IL-5 and greater rates of NP recurrence after endoscopic sinus surgery (ESS) suggests that these mediators may have utility as biomarkers of NP recurrence in diagnosing and assessing the severity of CRSwNP. This review provides an overview of eosinophil and IL-5 biology and explores the literature regarding the role of these mediators in CRSwNP pathogenesis and NP recurrence following ESS. Based on current published evidence, we suggest that although eosinophils play a key role in CRSwNP pathophysiology, IL-5, a cytokine that activates these cells, also represents a pertinent and effective treatment target in patients with CRSwNP.

International audience; Eosinophils play a homeostatic role in the body's immune responses. These cells are involved in combating some parasitic, bacterial, and viral infections and certain cancers and have pathologic roles in diseases including asthma, chronic rhinosinusitis with nasal polyps, eosinophilic gastrointestinal disorders, and hypereosinophilic syndromes. Treatment of eosinophilic diseases has traditionally been through nonspecific eosinophil attenuation by use of glucocorticoids. However, several novel biologic therapies targeting eosinophil maturation factors, such as interleukin (IL)-5 and the IL-5 receptor or IL-4/IL-13, have recently been approved for clinical use. Despite the success of biologic therapies, some patients with eosinophilic inflammatory disease may not achieve adequate symptom control, underlining the need to further investigate the contribution of patient characteristics, such as comorbidities and other processes, in driving ongoing disease activity. New research has shown that eosinophils are also involved in several homeostatic processes, including metabolism, tissue remodeling and development, neuronal regulation, epithelial and microbiome regulation, and immunoregulation, indicating that these cells may play a crucial role in metabolic regulation and organ function in healthy humans. Consequently, further investigation is needed into the homeostatic roles of eosinophils and eosinophil-mediated processes across different tissues and their varied microenvironments. Such work may provide important insights into the role of eosinophils not only under disease conditions but also in health. This narrative review synthesizes relevant publications retrieved from PubMed informed by author expertise to provide new insights into the diverse roles of eosinophils in health and disease, with particular emphasis on the implications for current and future development of eosinophil-targeted therapies.

Background Sputum cytology is currently the gold standard to evaluate cellular inflammation in the airways and phenotyping patients with airways diseases. Sputum eosinophil proportions have been used to guide treatment for moderate to severe asthma. Furthermore, raised sputum neutrophils are associated with poor disease control and impaired lung function in both asthma and COPD and small airways disease in cystic fibrosis. However, induced-sputum analysis is subjective and resource heavy, requiring dedicated specialist processing and assessment; this limits its utility in most clinical settings. Indirect blood eosinophil measures have been adopted in clinical care. However, there are currently no good peripheral blood biomarkers of airway neutrophils. A resource-light sputum processing approach could thus help integrate induced sputum more readily into routine clinical care. New mechanical disruption (MD) methods can rapidly obtain viable single cell suspensions from sputum samples. Aims The aim of this study was to compare MD sputum processing to traditional methods for cell viability, granulocyte proportions and sputum cytokine analysis. Methods Sputum plugs were split and processed using traditional methods and the MD method, and samples were then compared. Results The MD method produced a homogeneous cell suspension in 62 s; 70 min faster than the standard method used. No significant difference was seen between the cell viability (p = 0.09), or the concentration of eosinophils (p = 0.83), neutrophils (p = 0.99) or interleukin-8 (p = 0.86) using MD. Conclusion This cost-effective method of sputum processing could provide a more pragmatic, sustainable means of directly monitoring the airway milieu. Therefore, we recommend this method be taken forward for further investigation.

Background: IL-13 is considered an archetypal T2 cytokine central to the clinical disease expression of asthma. The IL-13 response genes, which are upregulated in central airway bronchial epithelial of asthma patients, can be normalized by high-dose inhaled steroid therapy in severe asthma. However, this is not the case within the peripheral airways. We have sought to further understand IL-13 in the peripheral airways in severe asthma through bronchoalveolar analysis. Methods: bronchoalveolar lavage samples were collected from 203 asthmatic and healthy volunteers, including 78 with severe asthma. Inflammatory mediators were measured using a multiple cytokine immunoassay platform. This analysis was replicated in a further 59 volunteers, in whom 16S rRNA analysis of BAL samples was undertaken by terminal restriction fragment length polymorphism. Results: severe asthma patients with high BAL IL-13, despite treatment with high-dose inhaled corticosteroids, had more severe lung function and significantly higher BAL neutrophil percentages, but not BAL eosinophils than those with normal BAL-13 concentrations. This finding was replicated in the second cohort, which further associated BAL IL-13 and neutrophilia with a greater abundance of potentially pathogenic bacteria in the peripheral airways. Conclusion: our findings demonstrate a steroid unresponsive source of IL-13 that is associated with BAL neutrophilia and bacterial dysbiosis in severe asthma. Our findings highlight the biological complexity of severe asthma and the importance of a greater understanding of the innate and adaptive immune responses in the peripheral airways in this disease.

Background: Asthma is a complex disease with multiple phenotypes that may differ in disease pathobiology and treatment response. IL33 single nucleotide polymorphisms (SNPs) have been reproducibly associated with asthma. IL33 levels are elevated in sputum and bronchial biopsies of patients with asthma. The functional consequences of IL33 asthma SNPs remain unknown. Objective: This study sought to determine whether IL33 SNPs associate with asthma-related phenotypes and with IL33 expression in lung or bronchial epithelium. This study investigated the effect of increased IL33 expression on human bronchial epithelial cell (HBEC) function. Methods: Association between IL33 SNPs (Chr9: 5,815,786-6,657,983) and asthma phenotypes (Lifelines/DAG [Dutch Asthma GWAS]/GASP [Genetics of Asthma Severity & Phenotypes] cohorts) and between SNPs and expression (lung tissue, bronchial brushes, HBECs) was done using regression modeling. Lentiviral overexpression was used to study IL33 effects on HBECs. Results: We found that 161 SNPs spanning the IL33 region associated with 1 or more asthma phenotypes after correction for multiple testing. We report a main independent signal tagged by rs992969 associating with blood eosinophil levels, asthma, and eosinophilic asthma. A second, independent signal tagged by rs4008366 presented modest association with eosinophilic asthma. Neither signal associated with FEV 1, FEV 1/forced vital capacity, atopy, and age of asthma onset. The 2 IL33 signals are expression quantitative loci in bronchial brushes and cultured HBECs, but not in lung tissue. IL33 overexpression in vitro resulted in reduced viability and reactive oxygen species–capturing of HBECs, without influencing epithelial cell count, metabolic activity, or barrier function. Conclusions: We identify IL33 as an epithelial susceptibility gene for eosinophilia and asthma, provide mechanistic insight, and implicate targeting of the IL33 pathway specifically in eosinophilic asthma.

Background Mepolizumab (100 mg delivered s.c. every 4 weeks) is indicated for add-on maintenance treatment for patients with severe eosinophilic asthma. Mepolizumab has been shown to reduce exacerbations and the requirement for daily oral corticosteroids, and improve asthma control and symptoms. However, data on the durability of the response to mepolizumab during dosing periods are limited. The aim of this study was to investigate the efficacy profile in patients with severe eosinophilic asthma over the 4-weekly dosing period for various fixed mepolizumab doses. Methods This was a post hoc analysis of data from the phase IIb/III DREAM study. Patients ≥12 years of age with severe eosinophilic asthma were randomised (1:1:1:1) to receive intravenous mepolizumab 75 mg (equivalent to 100 mg s.c.), 250 mg, 750 mg or placebo, plus standard of care, every 4 weeks for 52 weeks. The number of exacerbations and eDiary data (peak expiratory flow, rescue medication use and symptom scores) from two periods in each 4-weekly dosing interval (days 1–14 and 15–28) over the 52-week treatment period were analysed. Findings eDiary data and the proportion of patients experiencing ≥1 exacerbation were similar during the first and second 2 weeks of a dosing period across all mepolizumab doses. Interpretation These results demonstrate that the response to mepolizumab is sustained over the 4-weekly dosing period with no differences across a 10-fold dose range and supports the use of the current mepolizumab dosing regimen in patients with severe eosinophilic asthma., Post hoc analysis of data from DREAM demonstrated a sustained response to mepolizumab across the 4-weekly dosing period, suggesting therapeutic benefit is maintained between each mepolizumab dose following long-term treatment https://bit.ly/3843lH6

Eosinophils have a broad range of functions, both homeostatic and pathological, mediated through an array of cell surface receptors and specific secretory granules that promote interactions with their microenvironment. Eosinophil development, differentiation, activation, survival and recruitment are closely regulated by a number of type 2 cytokines, including interleukin (IL)-5, the key driver of eosinophilopoiesis. Evidence shows that type 2 inflammation, driven mainly by interleukin (IL)-4, IL-5 and IL-13, plays an important role in the pathophysiology of eosinophilic airway diseases, including asthma, chronic rhinosinusitis with nasal polyps, eosinophilic granulomatosis with polyangiitis and hypereosinophilic syndrome. Several biologic therapies have been developed to suppress type 2 inflammation, namely mepolizumab, reslizumab, benralizumab, dupilumab, omalizumab and tezepelumab. While these therapies have been associated with clinical benefits in a range of eosinophilic diseases, their development has highlighted several challenges and directions for future research. These include the need for further information on disease progression and identification of treatable traits, including clinical characteristics or biomarkers that will improve the prediction of treatment response. The Nordic countries have a long tradition of collaboration using patient registries and Nordic asthma registries provide unique opportunities to address these research questions. One example of such a registry is the NORdic Dataset for aSThmA Research (NORDSTAR), a longitudinal population-based dataset containing all 3.3 million individuals with asthma from four Nordic countries (Denmark, Finland, Norway and Sweden). Large-scale, real-world registry data such as those from Nordic countries may provide important information regarding the progression of eosinophilic asthma, in addition to clinical characteristics or biomarkers that could allow targeted treatment and ensure optimal patient outcomes. publishedVersion

Background: Interleukin 5, an archetypal type 2 cytokine, is fundamental to eosinophilic airways disease, considered to signal the bone marrow to promote eosinophil progenitor maturation and increase levels of eosinophils in the circulation. Aim: To investigate the relationship between airway IL-5 concentrations and inflammatory phenotypes of severe asthma. Methods: Healthy volunteers and biologic naive severe asthma patients were clinically characterised and had blood and induced sputum samples collected. Sputum was analysed for differential cell count and supernatant (PBS processed) proteins, measured by single plex ELISA. Eosinophilic asthma was defined as sputum eosinophils >2%. Between group comparisons were assessed by Kruskal Wallis and relationships between variables by spearman rank correlations. Results: 17 healthy controls, 53 non-eosinophilic and 26 eosinophilic patients with severe asthma participated in the study. In severe asthma, there were modest correlations between sputum IL-5 and sputum eosinophils (r = 0.519, p Discussion: Despite high levels of steroid treatment, there is persistent IL-5 generation within the airways of severe asthma.

Asthma is a heterogeneous disease with multiple clinical presentations (phenotypes). Neutrophilic asthma is characterised by increased sputum neutrophils and generally has a poor response to corticosteroids and limited other therapeutic options. Neutrophilia originates from different factors, including the defective resolution of inflammation or bacterial infections. An association between airway bacterial imbalance (disturbance) and the neutrophilic phenotype has been reported, suggesting that airway microbiota composition is involved in neutrophilic asthma. Rather than being a separate entity, neutrophilic asthma may be in part, an alliance between innate immunity and microbiota composition that prompts protective mechanisms against invading pathogens.

Introduction: Maintenance OCS use in severe asthma (SA) is associated with significant side-effects and has the potential to mask the systemic type-2 inflammatory biomarker signal required for initiation of biologic therapies. Method: WSAC is a large published cohort of 342 well characterised SA patients fulfilling the ATS/ERS 2014 definition of SA undertaken prior to the widespread use of biologic treatments. We compared the clinical, physiological & biological characteristics of patients requiring & not requiring maintenance OCS. Results: Discussion: Maintenance OCS use is associated with a significantly greater burden of comorbidities, poorer quality of life & disease control with more frequent exacerbations & ITU admissions. Despite this no significant differences were identified in lung function, atopic status, inflammatory cell counts or the other extensive biological measures assessed in the cohort. 44% of patients on maintenance OCS showed no biomarker signal of T2 inflammation. 77% of those on maintenance OCS with a sputum eos ≥2% had a blood eos count ≥0.3x109/L whilst only 60% had a FeNO ≥25ppb (43% FeNO ≥50ppb) suggesting blood eos remains a useful biomarker of T2 inflammation.

Introduction: Increasing corticosteroids (CS) to control asthma symptoms and exacerbations has potential for inappropriately high and difficult to down-titrate CS doses. Aims and Objectives: We tested whether a T2 biomarker-based strategy (BS) to guide CS reduction (factoring fractional exhaled nitric oxide (FeNO), blood eosinophils, serum periostin) vs. a symptom-based control would see fewer exacerbations and better asthma control and lung function. Methods: In this randomised (4:1/BS:control), single-blind parallel group trial, patients (pts) with severe asthma and FeNO Results: In the intention-to-treat population (240 BS vs. 61 control pts), the proportion of BS pts on lower CS dose at Wk48 was 28.4% vs. 18.5% in the controls (adjusted OR: 1.71; 95%CI 0.80, 3.63; p=0.165). In the per protocol (PP) population (n=121), significantly more pts had lower CS doses at Wk48 in the BS vs. the control group (30.7% vs. 5.0% (OR: 11.48; 95%CI 1.35, 97.83; p=0.026). Failure to follow treatment advice was the main reason for ITT and PP population differences. There was no difference in secondary outcomes between study arms. Conclusion: Biomarker based CS adjustment did not result in a greater proportion of pts reducing CS vs. control, probably because many pts did not follow treatment advice. The biomarker strategy seemed beneficial in pts where symptoms and T2 biomarker profile were discordant.

Background: In asthmatic patients, airway eosinophilia identifies a phenotype of patients that suffer from recurrent exacerbations and respond to biologic therapies. Direct measurement, through sputum induction, is impractical as part of routine clinical care so blood eosinophil counts are commonly used as a surrogate marker. We sought to assess the degree of concordance between these measures in our cohort of severe asthma patients. Methods: WATCH patients with severe asthma had blood tests, FeNO measurement and sputum induction performed on the same day. Spearman’s rank correlation coefficients and classification tables were calculated between sputum eosinophil, blood eosinophil, serum IgE and FeNO measures. Prediction of sputum eosinophilia (>3%) was assessed using receiver operating curves. Results: 203 patients, treated with high dose inhaled corticosteroids, were included in the analysis. Using commonly used cut-offs for blood eosinophilia (>300 cells/μL) and sputum eosinophilia (>3%), measurements were concordant in 138 patient (73.0%). 22 patients (11.6%) demonstrated blood eosinophilia alone and 29 patients (15.3%) sputum eosinophilia alone. Conclusions: We find that in a proportion of patients with severe asthma have sputum eosinophilia not recognised by blood. A better understanding of the nature of these airway eosinophils and the basis for the lack of apparent systemic IL5 signalling would help understand if these are a potentially biologically overlooked population or represent a different disease endotype.

Introduction: Depression is a well-recognised comorbidity in SA associated with poorer disease control & quality of life (QOL) but it remains unclear whether this is the consequence of altered pathobiology. Method: We compared the clinical, physiological & biological characteristics of patients with & without depression (defined by HADS depression score ≥8) within WSAC, a large published cohort of well characterised patients fulfilling the ATS/ERS 2014 definition of SA. Results: Discussion: SA patients with depression report poorer disease control & QOL associated with a greater burden of co-morbidities including obesity, rhinosinusitis & reflux. However, no statistically significant differences were identified in their other clinical or physiological characteristics. Inflammatory phenotypes did not differ between the groups but those with depression were found to have significantly higher serum hs-CRP & sputum IL-6 levels (not seen when subdivided for BMI). There is a well-recognised association between inflammation & depression with elevated serum IL-6 levels previously reported. Our findings raise the possibility that depression influences the pathobiology of asthma & thus reflects a targetable trait.

Background In all chronic airway diseases, the dynamics of airway function are influenced by underlying airway inflammation and bronchial hyperresponsiveness along with limitations in reversibility owing to airway and lung remodeling as well as mucous plugging. The relative contribution of each component translates into specific clinical patterns of symptoms, quality of life, exacerbation risk, and treatment success. Objective We aimed to evaluate whether subgrouping of patients with obstructive airway diseases according to patterns of fluctuation in lung function allows identification of specific phenotypes with distinct clinical characteristics. Methods We applied the novel method of fluctuation-based clustering (FBC) to twice-daily FEV1 measurements recorded over a 1-year period in a mixed group of 134 adults with mild-to-moderate asthma, severe asthma, or chronic obstructive pulmonary disease from the European BIOAIR cohort. Results Independently of clinical diagnosis, FBC divided patients into 4 fluctuation-based clusters with progressively increasing alterations in lung function that corresponded to patterns of increasing clinical severity, risk of exacerbation, and lower quality of life. Clusters of patients with airway disease with significantly elevated levels of biomarkers relating to remodeling (osteonectin) and cellular senescence (plasminogen activator inhibitor-1), accompanied by a loss of airway reversibility, pulmonary hyperinflation, and loss of diffusion capacity, were identified. The 4 clusters generated were stable over time and revealed no differences in levels of markers of type 2 inflammation (blood eosinophils and periostin). Conclusion FBC-based phenotyping provides another level of information that is complementary to clinical diagnosis and unrelated to eosinophilic inflammation, which could identify patients who may benefit from specific treatment strategies or closer monitoring.

SummaryBackgroundPatients with severe asthma may have a greater risk of dying from COVID-19 disease caused by SARS-CoV-2 virus. Angiotensin converting enzyme 2 (ACE2) receptor and enzyme proteases, transmembrane protease, serine 2 (TMPRSS2) and furin are needed for the attachment and invasion of the virus into host cells. We determined whether their expression in the airways of severe asthma patients is increased.MethodWe examined the microarray mRNA expression of ACE2, TMPRSS2 and furin in the sputum, bronchial brush and bronchial biopsies of participants in the European U-BIOPRED cohort.ResultsACE2 and furin sputum gene expression was significantly increased in severe non-smoking asthma compared to mild-moderate asthma and healthy volunteers. By contrast, TMPRSS2 expression in bronchial biopsy and bronchial brushings was increased in severe smoking and ex-smoking asthmatics, and so was furin expression in bronchial brushings. Several clinical parameters including male gender, oral steroid use and nasal polyps were positively associated with ACE2, TMPRSS2 and furin expression levels. There was a higher expression of ACE2 and furin in the sputum neutrophilic molecular phenotype with inflammasome activation compared to the eosinophilic Type2-high or paucigranulocytic phenotypes. The enrichment score of the IL-13-Type2 gene signature was positively correlated with ACE2, TMPRSS2 and furin levels.ConclusionThese key determinants of virus entry into the lungs may contribute to the poorer outcomes from COVID-19 disease in patients with severe asthma.“take home” messageIn severe asthma, gene expression of ACE, TMPRSS2 and furin are elevated compared to mild-moderate asthma and healthy volunteers, particularly in neutrophilic asthma. This might explain the increased risk of death in severe asthma afflicted with COVID19.

The IL1RL1 (ST2) gene locus is robustly associated with asthma; however, the contribution of single nucleotide polymorphisms (SNPs) in this locus to specific asthma subtypes and the functional mechanisms underlying these associations remain to be defined. We tested for association between IL1RL1 region SNPs and characteristics of asthma as defined by clinical and immunological measures and addressed functional effects of these genetic variants in lung tissue and airway epithelium. Utilizing 4 independent cohorts (Lifelines, Dutch Asthma GWAS [DAG], Genetics of Asthma Severity and Phenotypes [GASP], and Manchester Asthma and Allergy Study [MAAS]) and resequencing data, we identified 3 key signals associated with asthma features. Investigations in lung tissue and primary bronchial epithelial cells identified context-dependent relationships between the signals and IL1RL1 mRNA and soluble protein expression. This was also observed for asthma-associated IL1RL1 nonsynonymous coding TIR domain SNPs. Bronchial epithelial cell cultures from asthma patients, exposed to exacerbation-relevant stimulations, revealed modulatory effects for all 4 signals on IL1RL1 mRNA and/ or protein expression, suggesting SNP-environment interactions. The IL1RL1 TIR signaling domain haplotype affected IL-33-driven NF-κB signaling, while not interfering with TLR signaling. In summary, we identify that IL1RL1 genetic signals potentially contribute to severe and eosinophilic phenotypes in asthma, as well as provide initial mechanistic insight, including genetic regulation of IL1RL1 isoform expression and receptor signaling.

Asthma arises from the complex interplay of inflammatory pathways in diverse cell types and tissues. We sought to undertake a comprehensive transcriptomic assessment of the epithelium and airway T cells that remain understudied in asthma and investigate interactions between multiple cells and tissues. Epithelial brushings and flow-sorted CD3+ T cells from sputum and BAL were obtained from healthy subjects (n = 19) and patients with asthma (mild, moderate, and severe asthma; n = 46). Gene expression was assessed using Affymetrix HT HG-U133+ PM GeneChips, and results were validated by real-time quantitative PCR. In the epithelium, IL-13 response genes (POSTN, SERPINB2, and CLCA1), mast cell mediators (CPA3 and TPSAB1), inducible nitric oxide synthase, and cystatins (CST1, CST2, and CST4) were upregulated in mild asthma, but, except for cystatins, were suppressed by corticosteroids in moderate asthma. In severe asthma—with predominantly neutrophilic phenotype—several distinct processes were upregulated, including neutrophilia (TCN1 and MMP9), mucins, and oxidative stress responses. The majority of the disease signature was evident in sputum T cells in severe asthma, where 267 genes were differentially regulated compared with health, highlighting compartmentalization of inflammation. This signature included IL-17–inducible chemokines (CXCL1, CXCL2, CXCL3, IL8, and CSF3) and chemoattractants for neutrophils (IL8, CCL3, and LGALS3), T cells, and monocytes. A protein interaction network in severe asthma highlighted signatures of responses to bacterial infections across tissues (CEACAM5, CD14, and TLR2), including Toll-like receptor signaling. In conclusion, the activation of innate immune pathways in the airways suggests that activated T cells may be driving neutrophilic inflammation and steroid-insensitive IL-17 response in severe asthma.

Introduction Ameliorer le controle de l’asthme est un objectif essentiel de la prise en charge de l’asthme. L’effet du mepolizumab sur le controle de l’asthme evalue par l’Asthma Control Questionnaire-5(ACQ-5) a ete rapporte dans les essais cliniques, mais les donnees en vraie vie sont limitees. Methodes REALITI-A, etude prospective en vraie vie, multicentrique, internationale, a enrole des patients avec un asthme severe a eosinophiles (ASE), ayant initie un traitement par mepolizumab sur 51 centres dans 7 pays. Une periode de 12 mois de donnees precedant l’inclusion etait necessaire pour decrire les donnees demographiques des participants, le fardeau de l’ASE et les traitements. Le controle de l’asthme a ete evalue par l’ACQ-5. Les scores a l’initiation ont ete compares aux scores a 3,6,9 et 12 mois. Les changements du controle depuis l’initiation et le taux de participants ayant une difference minimale importante [DMI] depuis l’initiation de 0,5 unites ont ete evalues. Des modeles mixtes de mesures repetees ont ete utilises pour l’analyse. Resultats Les donnees initiales etaient disponibles pour 368 patients qui ont complete un suivi de 12 mois ( Fig. 1 ). Dix-neuf pour cent (n = 70/368) des patients ont arrete le mepolizumab dans l’annee suivant l’initiation ; les raisons les plus frequentes etaient : la decision du patient (7 % ; n = 27/368), la discretion de l’investigateur (4 % ; n = 14/368), le manque d’efficacite (4 % ; n = 13/368). A l’inclusion, les patients avaient un score moyen ACQ-5 en moindres carres (IC95 %) de 2,99 (2,85 ; 3,13). L’initiation du mepolizumab a ete associee a une amelioration du controle de l’asthme, avec des moyennes MC (IC95 %) du score ACQ-5 a 3,6,9 et 12 mois de 1,81 (1,66 ; 1,96), 1,73 (1,56 ; 1,89), 1,84 (1,62 ; 2,05), et 1,84 (1,63 ; 2,05). Dans les 3 mois, 65 % (152/234) des patients ont obtenu une DMI, passant a 71 % (84/119) apres 12 mois. Les ameliorations ont ete observees dans chaque tranche de taux d’eosinophiles sanguins a l’inclusion, avec 56 % (10/18), 60 % (9/15), et 75 % (62/83) des patients avec des taux d’eosinophiles sanguins Conclusion Cette etude prospective en vie reelle confirme l’efficacite du mepolizumab chez les patients avec un ASE en termes d’amelioration du controle de l’asthme a 3 mois, avec des ameliorations significatives du score ACQ-5. Ces resultats montrent un benefice rapide et significatif du controle de l’asthme avec le mepolizumab en vraie vie, coherent avec les resultats des essais cliniques controles.

Objectives There is limited information about the prevalence of rhinosinusitis in the UK community. This study aims to identify its prevalence and investigate any association with demographic variables. The secondary aims are to determine the degree of impairment, impact on quality of life and any costs incurred by patients. Design We used a modified version (MSNOT-20) of a quality of life instrument; the sinonasal outcome test -20 (SNOT-20), in a small and successful pilot project. It was then used in a community-based survey and a second phase 6-months later to test repeatability. Nasal examination and comparison of its quality of life section with other health-related quality of life tools occurred in the second phase. Setting and Participants The questionnaire was administered by post to 2,000 Farnborough (UK) residents, selected through stratified randomisation. The relation of an abnormal MSNOT-20 score with hay fever, asthma, smoking, food allergy, work productivity and social limitation were also analysed. Main Outcome and Results The response rate was 79.8%; over thirty per cent of the community suffer from upper respiratory tract symptoms with impact on multiple domains of quality of life including emotional, financial costs and loss of days at work. The MSNOT-20 provided a more sensitive assessment of health related quality of life than the Short Form-36 questionnaire. Conclusion Rhinosinusitis is prevalent in the Farnborough community and associated with significant morbidity and impairment on quality of life. The MSNOT-20 is a useful disease-specific quality of life tool in rhinosinusitis. This article is protected by copyright. All rights reserved.

Introduction: Asthmatics have been reported to have airway microbial dysbiosis [Chung JACI 2017]. We aimed to detect severe asthma phenotypes using induced sputum microbiome profiles and to assess cluster-wise stability after 12-18 months of prospective follow-up. Methods: Induced sputum samples were collected from a subset of the U-BIOPRED adult severe asthmatics. 16s rRNA sequencing and metagenomics were used to characterize the sputum microbiome. Unsupervised hierarchical clustering was performed by Bray-Curtis β-diversity measure of 16s microbiome data. The clustering was validated using partition around medoids, topological data analysis, consensus cluster distribution and bootstrapping. Sputum samples collected after 12-18 months were used to assess patient cluster migration. Results: Analysis of sputum samples of 100 severe asthmatics (median age: 55 years, 42% male) revealed two microbiome-driven clusters. The clusters were significantly different in: age of asthma onset, residential location, smoking status, percentage of sputum neutrophils, and spirometry (p-values Conclusion: Induced sputum microbiome identified two distinct severe asthma phenotypes. The relative stability of microbiome-driven phenotypic clusters indicates its suitability for diagnostic and therapeutic precision medicine approaches.

Background: Common cold (rhinovirus, RV) infections are the most frequent trigger of asthma exacerbations. Asthma is a common chronic inflammatory disease of the airways. Asthmatics show interferon-deficient responses to RV; however, the mechanisms underlying increased RV-driven symptoms leading to exacerbations are inadequately understood. Aim: To determine the role of RNA processing in antiviral immunity in asthma. Methods: We applied Frac-seq (subcellular fractionation and RNA-seq) in primary bronchial epithelial cells (BECs) from asthmatics and healthy controls. Frac-seq determines which cytoplasmic mRNAs bind to polyribosomes, the cellular translational machinery. We employed RNA immunoprecipitation (RIP) and siRNA knock-down experiments in bronchial epithelial cells (BEAS-2B) infected with RV1C. Results: Frac-seq data showed decreased translation of UPF1 (upstream frameshift 1) in asthma BECs. UPF1 translational levels negatively correlated with reversibility (r=-0.7235 P=0.0067). UPF1 is the main mediator of nonsense mediated decay (NMD), an RNA surveillance pathway that degrades potentially deleterious mRNAs (e.g. mRNAs with premature termination codons) and postulated to regulate 10-30% of all mRNAs. Emetine (a translation inhibitor that leads to upregulation of NMD targets) resulted in increased RV RNA levels. RIP showed that UPF1 directly binds RV RNA. Depletion of UPF1 in bronchial epithelial cells resulted in deficient RV-induced interferon (beta and lambda) and genome-wide changes in antiviral responses. Conclusions: Our data suggest that decreased UPF1 levels in asthma BECs contribute to impaired antiviral immunity and RV-induced exacerbations in asthma.

Background: Bronchial Thermoplasty (BT) reduces exacerbations and improves asthma quality of life questionnaire (AQLQ) scores in asthma [1]. Studies have shown reductions in airway smooth muscle (ASM) mass and reticular basement membrane (RBM) thickness after BT, but are limited by small sample size. Aims: To investigate airway remodeling responses to BT, and compare these to clinical outcomes. Methods: Pre- and post-BT data was pooled for 99 patients at 7 centres. ASM mass and RBM thickness were measured on bronchial biopsies, and compared to clinical outcomes. Results: Mean (SD) age was 48.4 years (11.7) and BMI was 29.0 kg/m2 (6.4). Baseline FEV1 was 68.8% predicted (23.1) and blood eosinophils were 0.25x10-9/L (0.26). 59 of 99 patients were taking maintenance oral Prednisolone (mean dose 25.7mg). In response to BT, mean (SD) annual exacerbation rate decreased from 6.4 (5.5) to 1.1 (1.7) (p= Conclusion: ASM mass and RBM thickness reduce rapidly following BT, and effects persist beyond 1 year. The observed improvements in airway remodeling parameters are not related to improvements in clinical outcomes. Other bronchial mucosal changes or mechanisms may play a role in the clinical response to BT. Reference: 1) Castro et al, AJRCCM. 2010;181(2):116-24

Background: The aim of this study was to evaluate the role of serum and sputum YKL-40 measurements as biomarkers in severe asthma phenotypes. Material and Methods: 243 adult subjects (48.9±14.2 years) from the Wessex Severe Asthma Cohort (WSAC), spirometry according to BTS/SIGN guidelines; sputum induction with inhaled hypertonic saline in 99 subjects. Serum and sputum levels of YKL-40 were determined by ELISA (Quidel corporation, San Diego, California, USA). Statistical analysis used SPSS (version 22) ; statistical significance at p Results: We observed correlation between markers of airway neutrophilia and sputum YKL-40 levels. Sputum YKL-40 was strongly correlated with both sputum MPO (Spearman’s rho 0.733; p≤0.001) and sputum absolute neutrophil count (Spearman’s rho 0.663; p≤0.001), but not serum levels of YKL-40. Sputum (p=0,025), but not serum(p=0,753), YKL-40 levels were also significantly different across the inflammatory cell subtypes in sputum. When comparing YKL-40 levels in serum and sputum between SARP clusters, serum ( p=0.002), but not sputum (p=0.081) YKL-40 levels were significantly different between phenotypes (namely differentiating clusters 1 and 2 from 3, 4 and 5). Conclusions: It is thus apparent that serum YKL-40 and sputum YKL-40 are reflecting different processes within the airways. Serum YKL-40 might reflect a more systemic inflammation opposed to sputum YKL-40 that might reflect more local: airway inflammation.

Background: Mepolizumab (100 mg delivered subcutaneously every 4 weeks) is approved for use in patients with severe eosinophilic asthma (SEA) and has been shown to reduce exacerbations and improve asthma symptoms vs placebo. There are limited data on the durability of the response to mepolizumab between doses. Aim: To investigate the efficacy profile in patients with SEA over the 4-week dosing period for various fixed doses of mepolizumab. Methods: This was a post hoc analysis of data from the Phase IIb DREAM trial. Patients with SEA ≥12 years of age were randomised (1:1:1:1) to receive mepolizumab 75 mg (equivalent to 100 mg SC), 250 mg or 750 mg, or placebo intravenously (IV) every 4 weeks for 52 weeks. Mean daily peak expiratory flow (PEF) and symptom scores recorded as diary data were compared between days 1–14 and 15–28 days (periods 1 and 2, respectively) in each successive 4-weekly dosing period. Results: All three doses of mepolizumab showed a similar effect on PEF and symptoms during periods 1 and 2. The number of patients whose symptoms increased by ≥1 point in period 2 vs period 1 was similar to placebo. Conclusions: These results demonstrate the durability of the response to mepolizumab between doses and that mepolizumab 75 mg IV has a similar efficacy profile over the 4-weekly dosing period compared with higher doses. Further analysis is needed to determine whether other outcomes show a similar trend. Funding: GSK [MEA112997;NCT01000506].

Rationale: Severe asthma is a heterogeneous disease with different inflammatory phenotypes based on blood or sputum measures. Increasingly blood eosinophils (Eos) are used to direct therapy choice but there is limited information how stable inflammatory measures in sputum and blood are over time and how they relate. Method: Sputum was successfully induced in 48 severe asthma patients at 2 timepoints within 2 years. At each timepoint a full blood count was completed. Patients were at least 4-weeks clear of exacerbation and considered stable. The percentage of granulocytes were reported and the stability at these two timepoints was assessed using Spearman’s rho. Results: In this cohort the sputum Eos and blood Eos significantly correlated between the two timepoints (sputum r=0.739,p= Discussion: These results demonstrate that airway neutrophilic inflammation is more dynamic than eosinophilic inflammation and poorly reflected in peripheral blood. A greater understanding of the underlying factors responsible for airway neutrophilic changes is needed if progress is to be made in addressing this severe asthma phenotype.

Background: Common cold (rhinovirus, RV) infections are the most frequent trigger of asthma exacerbations. Asthma is a common chronic inflammatory disease of the airways. Asthmatics show interferon-deficient responses to RV; however, the mechanisms underlying increased RV-driven symptoms leading to exacerbations are inadequately understood. Aim: To determine the role of RNA processing in antiviral immunity in asthma. Methods: We applied Frac-seq (subcellular fractionation and RNA-seq) in primary bronchial epithelial cells (BECs) from asthmatics and healthy controls. Frac-seq determines which cytoplasmic mRNAs bind to polyribosomes, the cellular translational machinery. We employed RNA immunoprecipitation (RIP) and siRNA knock-down experiments in bronchial epithelial cells (BEAS-2B) infected with RV1C. Results: Frac-seq data showed decreased translation of UPF1 (upstream frameshift 1) in asthma BECs. UPF1 translational levels negatively correlated with reversibility (r=-0.7235 P=0.0067). UPF1 is the main mediator of nonsense mediated decay (NMD), an RNA surveillance pathway that degrades potentially deleterious mRNAs (e.g. mRNAs with premature termination codons) and postulated to regulate 10-30% of all mRNAs. Emetine (a translation inhibitor that leads to upregulation of NMD targets) resulted in increased RV RNA levels. RIP showed that UPF1 directly binds RV RNA. Depletion of UPF1 in bronchial epithelial cells resulted in deficient RV-induced interferon (beta and lambda) and genome-wide changes in antiviral responses. Conclusions: Our data suggest that decreased UPF1 levels in asthma BECs contribute to impaired antiviral immunity and RV-induced exacerbations in asthma.

The search for biomarkers that can guide precision medicine in asthma, particularly those that can be translated to the clinic, has seen recent interest in exhaled volatile organic compounds (VOCs). Given the number of studies reporting “breathomics” findings and its growing integration in clinical trials, we performed a systematic review of the literature to summarise current evidence and understanding of breathomics technology in asthma.A PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses)-oriented systematic search was performed (CRD42017084145) of MEDLINE, Embase and the Cochrane databases to search for any reports that assessed exhaled VOCs in adult asthma patients, using the following terms (asthma AND (volatile organic compounds AND exhaled) OR breathomics).Two authors independently determined the eligibility of 2957 unique records, of which 66 underwent full-text review. Data extraction and risk of bias assessment was performed on the 22 studies deemed to fulfil the search criteria. The studies are described in terms of methodology and the evidence narratively summarised under the following clinical headings: diagnostics, phenotyping, treatment stratification, treatment monitoring and exacerbation prediction/assessment.Our review found that most studies were designed to assess diagnostic potential rather than focus on underlying biology or treatable traits. Results are generally limited by a lack of methodological standardisation and external validation and by insufficiently powered studies, but there is consistency across the literature that exhaled VOCs are sensitive to underlying inflammation. Modern studies are applying robust breath analysis workflows to large multi-centre study designs, which should unlock the full potential of measurement of exhaled volatile organic compounds in airways diseases such as asthma.

Objective: Whilst the exact cause of chronic rhinosinusitis (CRS) remains elusive, it is clear that both inflammation and remodelling are key disease processes. Environmental fungi have been linked to airway inflammation in CRS; however, their role in the pathogenesis of this condition remains controversial. The current consensus suggests that whilst fungi may not be directly causative, it is likely that CRS patients have deficits in their innate and potentially acquired immunity, which in turn may modify their ability to react to fungi. This study used a nasal polyp explant tissue stimulation model to study the inflammatory and remodelling responses related to challenge with common airborne fungal species. Methods: Ex vivo nasal polyp tissue from six well phenotyped CRSwNP patients undergoing functional endoscopic sinus surgery was stimulated with 1, 10 and 100 μg/ml of Alternaria alternata, Aspergillus niger, Cladosporium sphaerospermum and Penicillium notatum and compared with unchallenged polyp tissue as control. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of pro-inflammatory cytokines interleukin-6 (IL-6), granulocyte macrophage colony stimulating factor (GM-CSF) and tumour necrosis factor-α (TNF-α); and pro-remodelling cytokines transforming growth factor-b1 (TGF-b1), and basic fibroblast growth factor (bFGF) in the polyp supernatant. Results: Aspergillus niger stimulation increased pro-inflammatory cytokines TNF-α, GM-CSF and IL-6 whilst having little effect on the remodelling cytokines bFGF and TGF-b1. In contrast, stimulation with Cladosporium sphaerospermum, Alternaria alternata and Penicillium notatum reduced pro-inflammatory cytokines TNF-α and IL-6, but induced a dose-dependent increase in remodelling cytokines TGF-b1 and bFGF. Conclusions: This study shows that common airborne fungi induce species-specific effects on the upper airway inflammatory and remodelling responses. These findings provide further immunological evidence of a disease-modifying role for fungi in CRS.