Matthew Richardson

The relationship between biomarkers of fungal allergy and lung damage in asthma.

Immunological biomarkers are the key to the diagnosis of allergic bronchopulmonary aspergillosis (ABPA) and fungal sensitisation, but how these relate to clinically relevant outcomes is unclear.

A modular transcriptional signature identifies phenotypic heterogeneity of human tuberculosis infection

Whole blood transcriptional signatures distinguishing active tuberculosis patients from asymptomatic latently infected individuals exist. Consensus has not been achieved regarding the optimal reduced gene sets as diagnostic biomarkers that also achieve discrimination from other diseases. Here we show a blood transcriptional signature of active tuberculosis using RNA-Seq, confirming microarray results, that discriminates active tuberculosis from latently infected and healthy individuals, validating this signature in an independent cohort. Using an advanced modular approach, we utilise the information from the entire transcriptome, which includes overabundance of type I interferon-inducible genes and underabundance of IFNG and TBX21, to develop a signature that discriminates active tuberculosis patients from latently infected individuals or those with acute viral and bacterial infections. We suggest that methods targeting gene selection across multiple discriminant modules can improve the development of diagnostic biomarkers with improved performance. Finally, utilising the modular approach, we demonstrate dynamic heterogeneity in a longitudinal study of recent tuberculosis contacts.Mass screening diagnostics for Mycobacterium tuberculosis exist, but criticisms exist regarding the sensitivity and specificity of these tools. Here the authors use RNA-Seq and a modular bioinformatics approach using data from their own cohorts and meta-analysis of published cohorts to create a reduced signature for detection of tuberculosis that does not detect other diseases.

Prospective observational cohort study of symptom control prediction in paediatric asthma by using the Royal College of Physicians three questions

The Royal College of Physicians three questions (RCP3Q) is widely used for assessing asthma control within primary care in the UK, despite limited evidence in children. This study compared the RCP3Q as a tool for assessing asthma control in children (5–16 years) against the validated Asthma Control Test (ACT), Childhood Asthma Control Test (C-ACT), and Mini-Paediatric Quality of Life Questionnaire (MiniPAQLQ). We conducted a prospective observational cohort study involving children from eight primary care practices in Leicestershire. Children with doctor diagnosed asthma, or receiving regular asthma medication, were invited to participate. A total of 319 participants completed the MiniPAQLQ and the C-ACT/ACT questionnaires, before RCP3Q responses were collected as part of their routine asthma review conducted immediately afterwards. RCP3Q sensitivity for detecting uncontrolled asthma ranged from 43–60% and specificity from 80–82%. Using an RCP3Q score ≥2 to predict uncontrolled asthma and an RCP3Q score of zero to predict well-controlled asthma resulted in 10% of participants misclassified as uncontrolled and 8% as well-controlled, respectively. Using an RCP3Q threshold score of ≥1 resulted in 25% of participants being misclassified as uncontrolled. Our data suggests limited utility of the RCP3Q to assess asthma control in children. Alternative indicators of asthma control, such as the validated Asthma Control Test and the Children’s Asthma Control Test should be considered instead.Childhood asthma: importance of validated control testsValidated asthma control tests should be used to assess children rather than the ‘three questions’ survey recently developed by the Royal College of Physicians. The UK-based organisation developed the RCP3Q as a practical, rapid way of assessing asthma control in primary care. However, the RCP3Q was never comprehensively trialed for use with children. Erol Gaillard and co-workers at the University of Leicester compared the RCP3Q with three validated tests and questionnaires to determine its efficacy in assessing patients aged 5 to 16. 319 child patients completed the validated tests and their RCP3Q responses were collected immediately afterwards on the same day. In comparison with validated tests, the RCP3Q varied in its accuracy depending on the threshold scores selected. A threshold score of 1 resulted in 25 per cent of participants being misclassified with uncontrolled asthma.

Airway pathological heterogeneity in asthma: Visualization of disease microclusters using topological data analysis

Background: Asthma is a complex chronic disease underpinned by pathological changes within the airway wall. How variations in structural airway pathology and cellular inflammation contribute to the expression and severity of asthma are poorly understood. Objectives: Therefore we evaluated pathological heterogeneity using topological data analysis (TDA) with the aim of visualizing disease clusters and microclusters. Methods: A discovery population of 202 adult patients (142 asthmatic patients and 60 healthy subjects) and an external replication population (59 patients with severe asthma) were evaluated. Pathology and gene expression were examined in bronchial biopsy samples. TDA was applied by using pathological variables alone to create pathology‐driven visual networks. Results: In the discovery cohort TDA identified 4 groups/networks with multiple microclusters/regions of interest that were masked by group‐level statistics. Specifically, TDA group 1 consisted of a high proportion of healthy subjects, with a microcluster representing a topological continuum connecting healthy subjects to patients with mild‐to‐moderate asthma. Three additional TDA groups with moderate‐to‐severe asthma (Airway Smooth MuscleHigh, Reticular Basement MembraneHigh, and RemodelingLow groups) were identified and contained numerous microclusters with varying pathological and clinical features. Mutually exclusive TH2 and TH17 tissue gene expression signatures were identified in all pathological groups. Discovery and external replication applied to the severe asthma subgroup identified only highly similar “pathological data shapes” through analyses of persistent homology. Conclusions: We have identified and replicated novel pathological phenotypes of asthma using TDA. Our methodology is applicable to other complex chronic diseases. Graphical abstract: Figure. No caption available.

Particles in exhaled air (PExA): non-invasive phenotyping of small airways disease in adult asthma

RATIONALE Asthma is often characterised by inflammation, damage and dysfunction of the small airways, but no standardised biomarkers are available. OBJECTIVES Using a novel approach-particles in exhaled air (PExA)-we sought to (a) sample and analyse abundant protein biomarkers: surfactant protein A (SPA) and albumin in adult asthmatic and healthy patients and (b) relate protein concentrations with physiological markers using phenotyping. METHODS 83 adult asthmatics and 21 healthy volunteers were recruited from a discovery cohort in Leicester, UK, and 32 adult asthmatics as replication cohort from Sweden. Markers of airways closure/small airways dysfunction were evaluated using forced vital capacity, impulse oscillometry and multiple breath washout. SPA/albumin from PEx (PExA sample) were analysed using ELISA and corrected for acquired particle mass. Topological data analysis (TDA) was applied to small airway physiology and PExA protein data to identify phenotypes. RESULTS PExA manoeuvres were feasible, including severe asthmatic subjects. TDA identified a clinically important phenotype of asthmatic patients with multiple physiological markers of peripheral airway dysfunction, and significantly lower levels of both SPA and albumin. CONCLUSION We report that the PExA method is feasible across the spectrum of asthma severity and could be used to identify small airway disease phenotypes.

S63 Do the royal college of physicians ‘three questions’ predict symptom control in paediatric asthma?

Introduction and Objectives The UK Quality Outcomes Framework (QOF) rewards primary-care practices for completing the Royal College of Physicians “Three Questions” (RCP3Q) score for all patients listed on their asthma register. Almost no validation data currently exists, however, to support its use in children. This study aimed to investigate the performance of the RCP3Q to predict asthma control in children, by comparing it with the validated Asthma Control Test (ACT) or Childhood Asthma Control Test (C-ACT). Methods This was a prospective, observational study involving 8 primary-care practices. Children aged 5–16 on the QOF asthma register and/or receiving asthma medication were invited to self-complete the ACT (age 12–16, n=96) or C-ACT (age 5–11, n=223) questionnaire immediately prior to a primary-care asthma review, where responses to the RCP3Q were collected. RCP3Q scores were compared with ACT or C-ACT data to assess performance of the RCP3Q in predicting asthma control. The RCP3Q scoring system is summarised in figure 1. Results Questionnaire and RCP3Q data was completed for 319 participants. RCP3Q scores correlated moderately with C-ACT and ACT data (Spearman’s rho −0.49 and −0.52 respectively, p<0.001). A RCP3Q score of ≥2 predicted uncontrolled asthma (C-ACT or ACT ≤19) with a sensitivity of 57% and specificity of 81%. A lower threshold RCP3Q score of ≥1 gave a specificity of 55%, resulting in a high false positive rate. A RCP3Q score of 0 predicted well-controlled asthma (C-ACT or ACT ≥20) with a sensitivity of 55% and specificity of 81%. Using thresholds of RCP3Q≥2 for uncontrolled asthma and RCP3Q=0 for good control resulted in 25% participants unclassified (RCP3Q=1) and 18% of participants scoring 0, 2 or 3 incorrectly classified. Binary logistic regression showed that individual positive answers to RCP questions 1 and 2, but not 3, significantly increased the likelihood of uncontrolled asthma. Conclusions Our data in ≥300 participants does not support use of the RCP3Q to classify asthma control in children. Our findings support current BTS/SIGN guidelines, which recommend use of validated asthma control questionnaires, such as C-ACT, when conducting a paediatric asthma review. Abstract S63 Figure 1

P207 Ambient air pollution and admissions to hospital with exacerbations of asthma

Background Air pollution has been linked to increased morbidity and mortality associated with a number of chronic health conditions including asthma. We hypothesised that levels of ambient air pollution would be related to the number of hospital admissions with exacerbations of asthma. Methods Data on asthma admissions to a large acute NHS trust in the East Midlands were extracted using discharge diagnosis codes over a five-year period from April 2011 to March 2016. Ambient air pollution levels during this period were obtained from the website of the Department for Environment, Food and Rural Affairs. These data were based on a single monitoring station located in the centre of the City which recorded hourly readings of ozone, nitrogen oxides and small particulate matter (PM2.5). Daily mean data were utilised in the analysis. The effect of ambient air pollution on daily admissions for asthma was assessed using a generalised linear model in R version 3.3.1. Independent variables in the model were time, Ozone, NO, NOx and PM2.5. Fourier terms to capture any long-term trend or seasonality where also included in the model. Delayed exposure effects where investigated by fitting separate constrained lag models. Results During the period from April 2011 to March 2016 there were 4204 admissions due to asthma exacerbations (71% female, mean age 48 years). Admission numbers increased progressively from 726 in 2011/12 to 1043 in 2015/16. Admissions were most frequent in the month of December (2.9 per day) and least frequent in August (1.5 per day). None of Ozone, NO, NOx and PM2.5 or their lagged terms were found to be independent predictors of daily admissions for asthma. Conclusion Ambient air pollution measured at a single fixed monitoring station within a city is not able to predict the frequency of admissions due to asthma. Further research on the health effects of air pollution should make use of more detailed assessments of city-wide exposure, for instance by utilising satellite imaging and geospatial mapping.

P131 Outdoor fungal spore levels, lung function and symptoms in patients with asthma and aspergillus sensitisation

Background IgE sensitisation to Aspergillus fumigatus is seen in a significant proportion of patients with refractory asthma. The EVITA3 study recently showed that three months’ treatment with voriconazole did not improve asthma-related outcomes in this patient group. It is not known whether daily variations in outdoor fungal spore levels are associated with concomitant fluctuations in symptoms and lung function in patients with Aspergillus-associated asthma. Methods Participants in the EVITA3 study kept daily diaries of peak expiratory flow (PEF) and asthma symptoms during their follow-up period. These diary records were retrospectively analysed together with contemporaneous fungal spore levels measured locally, in those patients (n = 36) who consented to the secondary use of their clinical and research data. Participants also underwent skin-prick tests for Aspergillus, Alternaria, Cladosporium, Penicillium and Botrytis. For each participant, cross-correlation was used to investigate the relationship between PEF and local spore counts of Aspergillus/Penicillium, Alternaria, Cladosporium, Botrytis, Sporobolomyces, Tilletiopsis, and Didymella. Group-level relationships were investigated using linear mixed models for PEF and generalised estimating equations for daily symptom scores, with participants stratified by skin prick test status. The analyses were performed with the exposure and outcome measured on the same day (lag 0), and with the exposure lagged by 1 day with respect to the outcome (lag 1). Results The analysis cohort comprised 20 men and 16 women with a mean (standard deviation) age of 60 (8) years. No significant or consistent relationships were observed between fungal spore counts and either PEF or self-reported symptom scores, regardless of skin prick test status and lag time between exposure and outcome. In a linear mixed model, the effect size of total fungal spore count on morning PEF was negligible (−0.000011, p = 0.343 for lag 0; −0.000002, p = 0.847 for lag 1). Conclusion In this retrospective analysis we found no evidence of a significant link between fungal spore counts and either PEF or symptoms in patients with Aspergillus-associated asthma. Further research is required to confirm this result in a prospective study and to identify whether aeroallergen levels relate to other important asthma outcomes such as exacerbations.

S7 Airway Pathological Phenotypes and their Clinical Utility in Adult Asthma

Background Airway remodelling and cellular inflammation are well recognised pathological features of asthma. However the relationship between asthma phenotype, treatment intensity and pathology is poorly understood. Objectives We performed a study of common pathological features in adult asthmatic bronchial biopsies to identify (i) whether discrete ‘pathological phenotypes/subtypes’ exist and (ii) their clinical utility. Methods 202 patients (142 asthma and 60 healthy volunteers) were recruited. Patients underwent bronchoscopy and endobronchial biopsy. Bronchial biopsies were evaluated for eleven common features of asthma pathology. Standard biostatistical analyses including a range of cluster analyses and machine learning were applied to pathological features alone to evaluate our objectives. Results Three distinct immunopathological clusters were identified and characterised by distinct biopsy features of cellular inflammation and remodelling. Specifically, i) late onset severe eosinophilic asthma [cluster 1] with evidence of reticular basement membrane thickening, increased epithelial area and vascular remodelling, ii) milder late onset asthma [cluster 2] with few features of remodelling and iii), an early onset atopic eosinophilic asthma [cluster 3] with features of Th2 high asthma, increased airway smooth muscle (ASM) mass, increased mast cells within the ASM and a mixed granulocytic submucosal inflammation. Pre bronchodilator FEV1 and decline (in a subset) differed across the clusters. Pathological features did not add value to the clinical prediction of asthma. Conclusion We have identified three novel pathological clusters of asthma with differing features of airway remodelling, cellular inflammation and airway function. Asthma may be characterised by variable pathological phenotypes warranting further evaluation in larger population studies.