Exacerbations in Severe Asthma Patients: Mechanisms and Biomarkers
Phase: NA
Conditions: Asthma
Sponsor: Imperial College London

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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Official title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
NCT02660853 
Document date: 08/05/2013 
 
Background 
         Exacerbations are important events in asthma and represent an important burden for the 
patient. The risk of experiencing an exacerbation is greatest in the patient with severe asthma, who 
already experiences daily symptoms, impaired lung function and adverse effects from corticosteroid 
therapy despite using maximal amounts of anti-asthma therapies particularly corticosteroids(1;2). In 
the SARP severe asthma cohort, 40% of patients required one or more emergency department visits, 
compared to 20% and 15% in moderate and mild asthma groups respectively(3);  for those using >3 
oral corticosteroid bursts in the past year, this was 54% in severe, 13% in moderate and 5% in mild. In 
the UK BTS cohort, up to 89% of severe asthma patients required one or more courses of high dose 
prednisolone in the previous year(4). Exacerbations are occurring even while on maximal doses of 
therapy.  
 
       According to the ERS/ATS Task Force on Asthma Control and Exacerbations(5), the definition of an 
asthma exacerbation is based on deterioration of symptoms that needs urgent action with the need 
to use of systemic corticosteroids defining severe exacerbations, and those without the needing for 
corticosteroids or increasing corticosteroids defining moderate exacerbations. However, these 
definitions while useful in clinical trials are not adequate for health care professionals and for patients, 
where a prospective definition is needed. There has been relatively few studies examining the 
circumstances of an exacerbation. In patients with mild-to-moderate asthma, exacerbations were 
characterized by a gradual fall in peak expiratory flows (PEF) over several days, followed by more rapid 
changes over 2 to 3 days with a parallel increase in symptoms and rescue beta-agonist use(6), which 
was unaffected by the use of inhaled corticosteroids or of a long-acting beta-agonist. Very little is 
known about the course and severity of these exacerbations in severe asthma. We hypothesise that 
the causes of an exacerbation in severe asthma may be different than those in non-severe asthma.  
 
Causes of exacerbations 
          Viral respiratory tract infections have been recognised as triggers for exacerbations of asthma 
with rhinoviruses being the most common virus(7). Bacteria have also been associated with 
exacerbations including atypical Mycoplasma pneumonia and Chlamydiae pneumonia, in 27% of 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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exacerbations(8). In stable patients with severe asthma from RBH, ~50% of sputum cultures were 
positive for bacteria, and this was more likely in those with an exacerbation in the previous year (9). 
Microbiome studies of the lower airways of asthma have indicated a  more diverse microbial 
population compared to non-asthma subjects(10;11). In collaboration with Professors Cookson and 
Moffatt (NHLI), we performed microbiome studies on sputum samples and bronchial brushings in 
patients with severe asthma and found that Streptococcus operational taxonomic units were more 
abundant in severe asthma sputum samples than in non-severe asthmatics and healthy subjects(12). 
The potential role of bacterial infections during exacerbations is supported by the observation that a 
macrolide antibiotic provided a small but significant benefit in accelerating the recovery period (13). 
It is possible that bacterial infections may play a greater part in the exacerbation observed in patients 
with severe asthma. However viral infections may also be important.  We have recently shown that 
blood CD8+ T cells from severe asthma were markedly activated in terms of expression of T cell 
activation markers and of MAP kinases, JAK/STAT and the transcription factor, NF-κB (14), with one 
potential explanation being the activation from viruses. Viral infections may also increase the chance 
of a bacterial reactivation, as has been shown in virus-induced COPD exacerbations (15). 
 
         Exacerbations in severe asthma may also be related to the deranged innate immune responses 
to infective agents. A reduced interferon response to rhinovirus  infections of the asthmatic airways 
has been described (16), allowing  for increased viral replication leading to an increased inflammatory 
response. In severe asthmatic, we have found a reduction in the ability of the alveolar macrophage 
and of the monocyte-derived macrophage (MDM) in phagocytosing beads coated with Staph aureus  
and Haem influenzae (unpublished; Fig 1). Rhinoviruses could also impair phagocytosis of bacteria by 
macrophages(17). A defective response in the phagocytosis of apoptotic cells (‘efferocytosis’) of 
macrophages  from severe asthma and from neutrophilic asthma has also been described(18;19). The 
impaired phagocytosis has been associated with an intracellular imbalance of glutathione sulphide 
homeostasis, together with a reduction in histone deacetylase activity (20).  In adult asthmatics, an 
exacerbation is characterised by a neutrophilic inflammation in sputum (21), and rhinovirus-induced 
asthma exacerbations is  also accompanied by a neutrophilic response (22), although rhinovirus has 
also been associated with an activation of Th-2 mechanisms.   The high Th2 phenotype characterised 
by eosinophilic asthma is highly prone to exacerbations (23).  The potential for an interaction between 
viral infections, allergen exposure and atopic sensitisation as conferring the greatest risk has been 
proposed(24).  
 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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        Viral and bacterial infections can both contribute to oxidative stress and have also been 
associated with the generation of reactive oxygen species through the induction of inflammation, with 
down-regulation of antioxidant defences. Elevated levels of 8-isoprostanes have been measured in 
sputum infected with bacteria(25). Oxidative stress occurs after allergen challenge, with a rapid loss 
of superoxide dismutase activity(26). Lower airway levels of GSH with increased GSSG in 
bronchoalveolar  lavage  fluid has been reported in asthmatic children with elevated levels of 
malondialdehyde, 8-isoprostanes and H2O2(20). Increased oxidant stress has been seen as a constant 
feature of severe asthma(27). During an exacerbation, there may be a further augmentation of 
oxidative stress that may in turn contribute to airway smooth muscle contraction, inflammation and 
corticosteroid insensitivity(28). We have demonstrated the presence of CS insensitivity in 
macrophages and airway smooth muscle cells from patients with severe asthma(29;30).  This may 
contribute to the lack of effect of corticosteroids in controlling the deterioration in asthma. Finally, 
activation of MAP kinases, JAK/STAT and the transcription factor, NF-κB, in blood CD8+ T-cells of 
patients with severe asthma may contribute to greater risk of exacerbations(14).  
 
Heterogeneity and Biomarkers of exacerbations 
        Because of the different potential causes associated with exacerbations in patients with severe 
asthma, it is likely that there are different phenotypes. This is important to determine because more 
specifically-targeted treatments for exacerbations may lead to better resolution without the need to 
use or increase corticosteroid use.  There are also no biomarkers identified for the detection or for 
quantifying the severity of an exacerbation, and that would also be valuable in patients with severe 
asthma who are already taking medication that could influence the levels of biomarkers. One potential 
improvement may result from an earlier detection of the exacerbation and therefore a biomarker for 
an earlier detection of exacerbation in severe asthma would be useful, particularly for initiation of  
treatments.  
 
Hypothesis and objectives 
      We hypothesise that (i) exacerbations in severe asthma are causally heterogeneous and (ii) 
represent a corticosteroid-insensitive event caused by an excess of oxidants and inflammation 
resulting  from respiratory viral and bacterial agents.   
      The objectives of this project are to:  
(i) 
determine the symptoms and lung function characteristics and course of  asthma exacerbations  
in patients with severe asthma  
(ii) evaluate the role of infective agents (respiratory viruses and bacteria)  

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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(iii) define the biomarkers in exhaled breath, sputum and in blood  
(iv) study the presence of oxidative stress, phagocytic capacity of macrophages and corticosteroid 
insensitivity.  
      Major outcomes of this study will be (i) to assess the feasibility of diagnosing exacerbations earlier 
(ii) to determine the heterogeneity of the presentation and course of exacerbations (iii) to identify 
exacerbation phenotypes in terms of causes and (iv) to look for potential targets for treatment.   
 
Study design 
    We have a database of 300 current patients with severe asthma defined according to ATS criteria(1) 
with  exacerbation rate of 1.5± 2.8 (SD)/year. To achieve 1.7 exacerbations after 16  weeks will require 
36 patients at 90% power and 5% significance. Adjusting for a possible dropout of 30% would require 
56 patients. Therefore, 60 patients will be recruited. To be on the safe side, we will enrol  patients 
with a history of ≥3 exacerbations/year for a prospective observational study.  
 
    At entry Visit 1, we will enrol and characterise these patients. They will be observed over 12 months 
during which the number of exacerbations will be recorded on the basis of objective measures with 
evaluation of ACQ and daily morning and evening PEF. At the earliest onset of each exacerbation, the 
patient will be requested to contact by phone the Asthma BRU Unit.  For one of these events, patients 
will be asked to attend the laboratory for a series of tests at Visit 2 similar to Visit 1. Visit 3 will be 
planned within 14 days after Visit 2, when the patient has recovered.  
 
In 30 of the 60 patients, we will request for an additional attendance of a subsequent exacerbation 
(Visit 4) with follow-up (Visit 5). For other exacerbations not studied, the patient will be asked to keep 
a detailed diary record of symptoms with severity scoring and spirometric and PEF measurements 
(Exacerbation Diary) over a period of 2 weeks after the onset of exacerbation. 
 
Patients will have their exacerbations treated in the usual way. The only exception will be the use of 
antibiotics which will be withheld and initiated (if indicated) after the patients has been through Visit 
2. Those who have to be hospitalised will not be studied, and only those who can attend the Clinical 
Research Unit will be studied. The more severe exacerbations will therefore be missed but will be 
recorded in the patient’s diary.  
 
Tests to be performed Visits 1, 2 and 3: 
1. Diary of symptom scores, use of beta-agonist reliever and other treatments for 2 weeks 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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2. PEFR and spirometry twice daily for 2 weeks 
3. Markers of systemic inflammation in blood: CRP, IL-8, IL-6, together with Meso-Scale-
Discovery (MSD) platform for other cytokines. 
4. Markers of oxidative stress in blood, exhaled breath condensate (EBC) and urine: 
malondialdehyde (MDA) and 8-isoprostanes (31). 
5. Nitric oxide (NO) levels in exhaled breath measured twice daily for 2 weeks using a portable 
hand-held NO meter (NObreath). EBC for pH & free iron (32) 
6. Induced or spontaneous sputum for inflammatory cells and assays of cytokines in 
supernatants using Meso-Scale-Discovery (MSD) platform, bacteriological culture and for 
microbiome analysis. H2S which is elevated in asthma in relation to sputum neutrophil counts 
(33)will also be measured. If spontaneous sputum is not available, sputum will be induced 
using ultrasonic nebulization of isotonic saline (34), which is safe in patients during an 
exacerbation (35).   
7. Sputum for bacteriological culture and colony forming units. 
8. PCR for respiratory viruses from nasopharyngeal swabs and sputum RNA and reverse 
transcription performed for respiratory viruses including rhinovirus, influenza virus types A 
and B, parainfluenza viruses 1 to 3, coronaviruses and respiratory syncytial virus types A and 
B , and Chlamydia pneumonia, and Mycoplasma pneumonia using real-time ‘TaqMan’ assays 
in collaboration with Professor Sebastian Johnston, NHLI. 
9. Sputum microbiome: 16S rRNA PCR and pyrosequencing will be performed on sputum with 
amplification of the V3-V5 regions (36) in collaboration with Professors Cookson and Moffatt, 
NHLI.  
10. Studies on blood cells: 
(i) 
MDM’s phagocytosis of fluorescently-labelled Pseudomonas aeruginosa and 
Staphylococcus aureus (Professor Louise Donnelly, NHLI) on macrophage-derived 
monocytes as described previously (37) 
(ii) 
Transcriptomic analysis of blood CD4+ and CD8+ T-cells and monocytes, as previously 
described (14) 
(iii) 
Studies of corticosteroid insensitivity in peripheral blood mononuclear cells and in 
whole blood (Dr Pank Bhavsar, NHLI) (38) 
 
Definition of exacerbation and monitoring 
           Because these patients with severe asthma are usually well-experienced ‘exacerbators’, they 
will be asked to ‘diagnose’ their own exacerbations. Each has an individualised way of recognising an 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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exacerbation. We will discuss with each patient t their way of recognising a deterioration and try and 
work out whether earlier signal warning is possible. One of our aims is to find out what the objective 
measures of deterioration are.  We will also ask patients to record their symptoms and lung function 
as soon as they feel the onset of an exacerbation, since exacerbations are recognised by the patient 
as events that are ‘clinically identified by being outside the patient's usual range of day-to-day asthma 
variation’(5). They will also contact the Study Coordinator (Nurse or Clinical Research Fellow) to 
determine if they can attend for Visit 2. The patient will receive or administer treatments for the 
exacerbation as usual without interference from the Research team. Except for the exacerbation that 
will be studied (Visit 2 or Visit 4), the patient will be requested to refrain from starting any antibiotic 
therapies, which will be started (if this has been prescribed) as soon as Visit 2 studies have been 
completed. 
 
Data analysis 
1. The daily variation of symptoms and lung function during the exacerbation visits (2 and 4) will be 
analysed, and related to the changes observed in the various parameters collected at exacerbation. 
Comparison of these biomarkers to those at Visit 1 and at the recovery visit will allow for 
ascertainment of biomarkers that may be associated with exacerbations. Bacterial- or virus-
associated, or eosinophil- or neutrophil-associated exacerbation will be defined.  
2. Unbiased mathematical methods such as factor and cluster analyses will be used to identify factors 
at exacerbations and classify groups of similar characteristics looking at all the data collected 
including the transcriptomic and cytokine profile data. The analysis will be performed by the 
Bioinformatics group led by Prof Yike Guo of Discovery Science at Imperial College. 
3. The stability of the exacerbation profile will be examined within the 30 patients who have had a 
repeat exacerbation visit. We could also look at this small group as a validation data set.  
 
Timelines  and milestones 
Oct 2013-Dec 2013: Setting up of study, identification of patients, setting up methods. 
Jan 2014-Dec 2014: Recruitment of 30 patients, and follow up to Dec 2014  
Jan 2015-Dec 2015: Recruitment of 30 patients, and follow up to Dec 2015  
Jan 2016-Sept 2016: Completion of assays, analysis of data, biomarker validation 
 
Future plans of research 
1.        Information obtained from this study will allow us to plan better any future therapeutic studies 
on asthma exacerbations and will also provide us with an idea of targets for treatments for 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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particular types of exacerbations. Therefore, we plan to use the new information gathered to plan 
for future studies (eg the number of subjects needed for a therapeutic study).  
        One target of therapy that we are seeking confirmation for is that of oxidant stress. Currently 
there are no good antioxidants, but N-acetylcysteine or other similar medications  is available and 
could be tested as treatment for exacerbations. We will also test drugs not only for treating 
exacerbations, but also for reducing exacerbation rate. This may include approaches to reduce 
corticosteroid insensitivity since the recurrence of exacerbations in patients taking high dose 
corticosteroids is a reflection of corticosteroid insensitivity. p38 MAPK inhibitors may be such a 
drug (39).  
2.  There will be a need for validation of some of the identified biomarkers for each exacerbation 
phenotype, and this will be done in multicentre studies in collaboration with other Centres within 
the BTS Severe Asthma Group.  
3. This research will also provide information about the best tools to monitor exacerbations in severe 
asthma patients. This will be examined in newly-recruited patients to our cohort. 
 
 

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Reference List 
 
 
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(4)  Gibeon D, Batuwita R, et al. Obesity associated severe asthma represents a distinct clinical 
phenotype Chest , -In press. 2012.  
 
 
(5)  Reddel HK, Taylor DR, Bateman ED, et al. An official American Thoracic Society/European 
Respiratory Society statement: asthma control and exacerbations. Am J Respir Crit Care 
Med 2009;180(1):59-99. 
 
(6)  Tattersfield AE, Postma DS, et al. Exacerbations of asthma: a descriptive study of 425 
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1999;160(2):594-9. 
 
(7)  Papadopoulos NG, Christodoulou I, et al. Viruses and bacteria in acute asthma 
exacerbations--a GA(2) LEN-DARE systematic review. Allergy 2011;66(4):458-68. 
 
(8)  Cazzola M, Matera MG, Rossi F. Bronchial hyperresponsiveness and bacterial respiratory 
infections. Clin Ther 1991 January;13(1):157-71. 
 
(9)  Zhang Q, Illing R, et al. Bacteria in sputum of stable severe asthma and increased airway 
wall thickness. Respir Res 2012;13:35. 
 (10)  Hilty M, Burke C, Pedro H, t al. Disordered microbial communities in asthmatic airways. 
PLoS One 2010;5(1):e8578. 
 (11)  Huang YJ, Nelson CE, Brodie EL et al. Airway microbiota and bronchial 
hyperresponsiveness in patients with suboptimally controlled asthma. J Allergy Clin 
Immunol 2011 February;127(2):372-81. 
 (12)  Zhang, Cox M, et al. Altered airway miucrobiome in severe asthma. Am.J Respir.Crit Care 
Med. 185, A6717. 2012.  
 
 (13)  Johnston SL, Blasi F, Black PN, Martin RJ, Farrell DJ, Nieman RB. The effect of telithromycin 
in acute exacerbations of asthma. N Engl J Med 2006 April 13;354(15):1589-600. 
 (14)  Tsitsiou E, Williams AE, et al. Transcriptome analysis shows activation of circulating CD8+ T 
cells in patients with severe asthma. J Allergy Clin Immunol 2012 January;129(1):95-103. 

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 (15)  Mallia P, Footitt J, et al. Rhinovirus infection induces degradation of antimicrobial peptides 
and secondary bacterial infection in COPD. Am J Respir Crit Care Med 2012 December 
1;186(11):1117-24. 
 (16)  Wark PA, Johnston SL, et al. Asthmatic bronchial epithelial cells have a deficient innate 
immune response to infection with rhinovirus. J Exp Med 2005 March 21;201(6):937-47. 
 (17)  Oliver BG, Lim S, et al. Rhinovirus exposure impairs immune responses to bacterial 
products in human alveolar macrophages. Thorax 2008 June;63(6):519-25. 
 (18)  Huynh ML, Malcolm KC, et al. Defective Apoptotic Cell Phagocytosis Attenuates PGE2 and 
15-HETE in Severe Asthma Alveolar Macrophages. Am J Respir Crit Care Med 2005; 
172:972-9. 
 (19)  Simpson JL, Gibson PG, et al. Impaired macrophage phagocytosis in non-eosinophilic 
asthma. Clin Exp Allergy 2013 January;43(1):29-35. 
 (20)  Fitzpatrick AM, Teague WG, Burwell L, Brown MS, Brown LA. Glutathione oxidation is 
associated with airway macrophage functional impairment in children with severe asthma. 
Pediatr Res 2011 February;69(2):154-9. 
 (21)  Wang F, He XY, et al. Different inflammatory phenotypes in adults and children with acute 
asthma. Eur Respir J 2011 September;38(3):567-74. 
 (22)  Wark PA, Johnston SL, Moric I, Simpson JL, Hensley MJ, Gibson PG. Neutrophil 
degranulation and cell lysis is associated with clinical severity in virus-induced asthma. Eur 
Respir J 2002 January;19(1):68-75. 
 (23)  Woodruff PG, Modrek B, et al. T-helper type 2-driven inflammation defines major 
subphenotypes of asthma. Am J Respir Crit Care Med 2009 September 1;180(5):388-95. 
 (24)  Green RM, Custovic A, Sanderson G, Hunter J, Johnston SL, Woodcock A. Synergism 
between allergens and viruses and risk of hospital admission with asthma: case-control 
study. BMJ 2002 March 30;324(7340):763. 
 (25)  Wood LG, Simpson JL, Hansbro PM, Gibson PG. Potentially pathogenic bacteria cultured 
from the sputum of stable asthmatics are associated with increased 8-isoprostane and 
airway neutrophilia. Free Radic Res 2010 February;44(2):146-54. 
 (26)  Comhair SA, Xu W, et al. Superoxide dismutase inactivation in pathophysiology of 
asthmatic airway remodeling and reactivity. Am J Pathol 2005 March;166(3):663-74. 
 (27)  Comhair SA, Xu W, et al. Superoxide dismutase inactivation in pathophysiology of 
asthmatic airway remodeling and reactivity. Am J Pathol 2005 March;166(3):663-74. 
 (28)  Chung KF, Marwick JA. Molecular mechanisms of oxidative stress in airways and lungs with 
reference to asthma and chronic obstructive pulmonary disease. Ann N Y Acad Sci 2010 
August;1203:85-91. 
 (29)  Bhavsar P, Hew M, , et al. Relative corticosteroid insensitivity of alveolar macrophages in 
severe asthma compared with non-severe asthma. Thorax 2008 September;63(9):784-90. 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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 (30)  Chang PJ, Bhavsar PK, Michaeloudes C, Khorasani N, Chung KF. Corticosteroid insensitivity 
of chemokine expression in airway smooth muscle of patients with severe asthma. J 
Allergy Clin Immunol 2012 September 1. 
 (31)  Gong J, Zhu T, et al. Malondialdehyde in exhaled breath condensate and urine as a 
biomarker of air pollution induced oxidative stress. J Expo Sci Environ Epidemiol 2013 
January 16. 
 (32)  Mumby S, Chung KF, McCreanor JE, Moloney ED, Griffiths MJ, Quinlan GJ. Pro-oxidant iron 
in exhaled breath condensate: a potential excretory mechanism. Respir Med 2011 
September;105(9):1290-5. 
 (33)  Saito J, Zhang Q, et al. Sputum hydrogen sulfide as a novel biomarker of obstructive 
neutrophilic asthma. J Allergy Clin Immunol 2013;131(1):232-4. 
 (34)  Macedo P, Hew M, et al. Inflammatory biomarkers in airways of patients with severe 
asthma compared with non-severe asthma. Clin Exp Allergy 2009;39(11):1668-76. 
 (35)  Wark PA, Simpson JL, Hensley MJ, Gibson PG. Safety of sputum induction with isotonic 
saline in adults with acute severe asthma. Clin Exp Allergy 2001 November;31(11):1745-53. 
 (36)  Sim K, Cox MJ, et al. Improved detection of bifidobacteria with optimised 16S rRNA-gene 
based pyrosequencing. PLoS One 2012;7(3):e32543. 
 (37)  Taylor AE, Finney-Hayward TK, et al. Defective macrophage phagocytosis of bacteria in 
COPD. Eur Respir J 2010 May;35(5):1039-47. 
 (38)  Hew M, Bhavsar P, et al. Relative corticosteroid insensitivity of peripheral blood 
mononuclear cells in severe asthma. Am J Respir Crit Care Med 2006 July 15;174(2):134-41. 
 (39)  Chung KF. p38 mitogen-activated protein kinase pathways in asthma and COPD. Chest 
2011 June;139(6):1470-9. 
 
 
 
 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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Figure:  
Phagocytosis of fluorescently-labelled Haemophilus influenzae and Staphyloccoccus aureus by 
monocyte-derived macrophages (MDM; top panels) and by alveolar macrophages (AM; lower panels) 
from normal subjects (Normal) and from non-severe asthmatics (NSA) and severe asthmatics (SA). 
There was no difference in phagocytosis of fluorescently-labelled beads not coated with bacteria 
(Panel A). However, there was significantly lower phagocytosis of beads coated with Haemophilus 
influenzae and Staphyloccoccus aureus by MDMs from severe asthma patients (Panel B). For 
macrophages, there was a reduction of phagocytosis of Haemophilus influenzae and Staphyloccoccus 
aureus by AMs from both asthma groups, although this was only significant for Staphyloccoccus 
aureus phagocytosis by AMs from severe asthma (Panel C). * p<0.05; **p<0.01.  
 
 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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Patient Information and Informed Consent Form 
Exacerbations in severe asthma patients: mechanisms and biomarkers 
Asthma: Version 1.1 21 June 2013 
 
INTRODUCTION 
    We would like to invite you to take part in a research study examining exacerbations 
in severe asthma. Before you decide whether to take part, it’s important to understand 
why the research is being done and what it involves.  Please take time to read this 
information sheet.  Talk to your family, friends, doctor or nurse if you wish. If anything 
is not clear and you require more information before you decide whether or not to take 
part in the study, please telephone the study team on 0207 351 8051.  Additionally, 
you may find it useful to visit the INVOLVE website (http://www.invo.org.uk/) which has 
information for patients about research in the NHS. 
 
1. 
WHAT IS THE PURPOSE OF THE STUDY? 
Asthma and other wheezing illnesses affect one in ten adults in the UK.  Asthma is a 
condition that affects the airways – the small tubes that carry air in and out of the lungs.  
The main aim of this research is to better understand the exacerbations (attacks) that 
occur in severe asthma.  Severe asthmatics have difficulty in controlling their disease, 
despite good medical care and taking all the medicines that usually work well in 
asthma. Having an exacerbation is quite common in patients with severe asthma, and 
it is one of the events that patients with severe asthma are usually very concerned 
about. We need to understand better what happens during an exacerbation of asthma. 
We would first like to find out whether there are certain viruses and bacteria that are 
associated with the exacerbation, and secondly we would like to find out what sort of 
inflammation is occurring during the exacerbation by measuring various proteins in the 
blood and in the phlegm from the lungs. We also wish to find out whether the type of 
exacerbation is different from one person to another and we will also find out whether 
there something that can give an early warning that an exacerbation will be happening 
in the near future. This research will give us some insight into how to obtain better 
treatments for the exacerbation and also to prevent them from occurring. 
2. 
WHY HAVE I BEEN INVITED TO PARTICIPATE? 
You have been invited to participate because you have severe asthma and you are a 
patient attending our clinic at the Royal Brompton Hospital.   
3. 
DO I HAVE TO TAKE PART? 
Participation in this study is entirely voluntary. It is up to you to decide whether or not 
to take part. If you do take part, you will be given this information sheet to keep and 
be asked to sign a consent form.  You are free to withdraw from the study at any time 
and without giving a reason. This will not in any way affect the standard of care you 
receive. The information sheet and consent form for the study have been approved by 
an Ethics Committee (EC).  
4. 
WHAT WILL HAPPEN TO ME IF I TAKE PART? 
If you decide that you want to take part in this research study, we will ask you to 
participate in a variety of tests, which will look at different aspects of your asthma.  
The entire study takes approximately 12 months and includes between 3 to 5 visits 
to the Asthma lab at the Royal Brompton Hospital.  As soon as you experience an 
asthma attack (exacerbation), you will be asked  you to contact us, the study team. 
We will then arrange whether you will be able and if so, when you can come to the 
study centre to be assessed. 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
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Details of the tests to be performed are attached at the end of this document. 
• Visit 1: Screening visit  – The Nurse or member of the study team will 
explain the study in detail.  If you have any questions the nurse or study 
staff will be able to answer them.  If you are satisfied with the answers and 
would like to take part in the study you will be asked to sign a consent 
form.  
A number of tests will then be carried out. This visit will last for up to 3 hours. During 
this visit we would like to do the following: 
Medical history, Physical examination,Questionnaires, Blood test ,Urine , Throat swab, 
Nasal wash and nasal brush, Lung function testing , Forced oscillation technique , 
Collecting breath condensates,  Sputum induction, Exhaled Nitric Oxide, Methacholine 
challenge test, Diary card  
 
• Visit 2, Exacerbation visit, which will take up to 3 hours 
Exacerbation visit – this should take place within the first 6 month from the time of 
visit 1. This visit will take place at the Asthma lab at the Royal Brompton Hospital; 
you will not be required to stay overnight, unless there is any reason for you to need 
further treatment in hospital. You will receive treatment as usual for your 
exacerbation. We will arrange transport for you to attend our lab. 
At this visit we would like to do the following: 
Physical examination, Medication history, Lung function testing , Forced oscillation 
technique , Blood sample , Urine sample , Nasopharyngeal swab, Sputum induction , 
Exhaled Nitric Oxide , Exhaled Breath Condensate  
 
• 
Visit 3, Exacerbation recovery visit at 14 days after the exacerbation 
visit, which will take up to 3 hours. 
We will repeat the same tests as at Visit 2. 
 
• 
Visit 4, Second Exacerbation visit (Similar to Visit 2) –  
This is optional as we are looking for half of the subjects undergoing this. The 
same tests will be performed as above.  This visit will take place over not more than 
2-3 hours). You will not be required to stay in hospital overnight.   
 
• Visit 5, Second recovery Exacerbation visit (Similar to Visit 3)- This is 
optional and will only be done if you have gone through Visit 4. The same 
tests will be performed again. 
5.     WILL EXPENSES BE PAID? 
Reasonable time and travel expenses you may have incurred as a result of 
participation in this study will be reimbursed.  
6. WHAT ARE THE ALTERNATIVES FOR DIAGNOSIS AND OR TREATMENTS? 
Regardless of whether you join this study, you will be treated with the current best 
treatment. This is particularly with respect to your treatment for exacerbations. This 
should be treated as usual  
7.  WHAT ARE THE SIDE EFFECTS OF ANY TREATMENT RECEIVED WHEN 
TAKING PART? 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
14 
 
This study does not include a new or experimental drug. You will continue with your 
prescribed asthma medication and your usual  treatments for your exacerbations.  
8. WHAT ARE THE POSSIBLE BENEFITS OF TAKING PART IN THIS STUDY? 
We cannot promise the study will help you but the information we get from this study 
may help improve the treatment and monitoring of people with asthma, and particularly 
for treating exacerbations of asthma.  
9. WHAT IF THERE IS A PROBLEM? 
Imperial College London holds insurance policies which apply to this study.  If you 
experience serious and enduring harm or injury as a result of taking part in this study, 
you may be eligible to claim compensation without having to prove that Imperial 
College is at fault.  This does not affect your legal rights to seek compensation. 
  
If you are harmed due to someone’s negligence, then you may have grounds for a 
legal action.  Regardless of this, if you wish to complain, or have any concerns about 
any aspect of the way you have been treated during the course of this study then you 
should immediately inform the Investigator Professor K F Chung at 0207 351 8995 
(Secretary: Nicola Bell).  The normal National Health Service complaint complaints 
mechanisms are also available to you.  If you are still not satisfied with the response, 
you may contact the Imperial AHSC Joint Research Office.   
10. WILL TAKING PART IN THIS STUDY BE KEPT CONFIDENTIAL? 
All information which is collected about you during the course of the study will be 
treated confidentially. All study data related to you will be coded. This means that you 
will be given a code number that will be used to identify you and information about you 
without having to use your name, medical record number, or other common identifiers. 
If you give your consent to participate in this study, you also give your consent for 
access to your medical file.  
We do need to let your GP and hospital consultant know you are taking part in this 
study. If any clinically relevant information is obtained as a result of your involvement 
in the study we will discuss this with you. Your permission will be sought to share this 
information with the relevant doctor involved in your care.  
All samples that are provided by you will be uniquely identified with a sample identifier 
and your code. If you change your mind about participating in the research, the code 
and link to your identity will allow us to locate your samples and destroy them so they 
cannot be used for further research.  
11. WHAT WILL MY STUDY DATA ARE USED FOR? 
Your study data will be used for research into asthma and may be used in research 
related to the development of pharmaceutical products, diagnostics or medical aids. 
The handling of your study data will be in accordance with applicable Data Protection 
law(s).   
12. WHAT WILL HAPPEN TO THE SAMPLES I GIVE? 
The samples you provide will be examined within the first 2 years of their collection. 
Any remaining samples will be stored for up to 20 years, at special facilities that are 
designed to store samples safely and securely. No information will be stored on the 
sample labels that may be used to identify you. Any movement and storage of samples 
will be carried out in accordance with the Human Tissue Act 2004. At the end of the 
storage time any remaining samples will be destroyed. 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
15 
 
13. 
WHAT WILL HAPPEN IF I DO NOT WANT TO CARRY ON WITH THE 
STUDY? 
You can decide to withdraw from the study at any time. Refusal to take part or if you 
withdraw after giving your consent will have no consequences for your present or 
future treatment by your doctor. Information collected may still be used. If you ask, we 
will destroy any stored samples that can still be identified as yours.This will not impact 
on the treatment and care that you are or will be receiving in any way. 
14. WHAT WILL HAPPEN TO THE RESULTS OF THE RESEARCH STUDY? 
The result of the study will be considered for publication or presentation at (scientific) 
symposia or congresses. You will not be identified in any report/publication or 
presentations of the study. During the study and at the end of the study all participants 
will be informed on the general but not individual results.  
You have the right to request information about your study data be provided to you 
with a copy of them.  You also have the right to request that any inaccuracies in such 
data be corrected.  If you wish to make a request, then please contact your clinic at 
Royal Brompton Hospital. 
15.   WHO IS ORGANISING AND FUNDING THE RESEARCH?  
The research is being organised under Imperial College London and under the 
Biomedical Research Unit of the Royal Brompton Hospital. We have applied to Asthma 
UK to fund this study. 
16. CONTACT DETAILS & SOURCE FOR ADDITIONAL INFORMATION. 
For the duration of the study, you will be under the care of Professor Chung. If at any 
time between your visits you feel that any of your symptoms are causing problems, or 
you have any questions during the study, please contact your study team. The 
telephone number to reach the study team is  0207 351 8051 (ask for Mr Joao Rocha, 
the Study Nurse). 
 
 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
16 
 
APPENDIX: 
Visit details: 
Visit 1 – Screening visit  – The Nurse or member of the study team will explain the 
study in detail.  If you have any questions the nurse or study staff will be able to 
answer them.  If you are satisfied with the answers and would like to take part in the 
study you will be asked to sign a consent form.  
A number of tests will then be carried out. This visit will last for up to 3 hours. During 
this visit we would like to do the following: 
• Medical history - you will be asked about your medical history including 
respiratory and non respiratory related history, smoking, allergies and what 
medications you use.  
• Physical examination – we will perform assessments such as record your 
height, weight, blood pressure and listen to your chest  
• Questionnaires: 
• Blood test – we would like to obtain a sample (10 teaspoons, 50 mls) of 
your blood to keep for measurement of various markers. Some of the 
blood will be used to measure how cells in the blood can engulf particles 
and bacteria.  We will separate various cells in the blood to study them in 
the test-tube. We will extract genes from the blood cells to examine the 
genes expressed.  Obtaining the blood may cause some local discomfort 
and bruising, rarely there is a risk of irritation of the nerves. 
• Urine - we would like to collect a sample of your urine. We will use this to 
check for indicators of asthma and exposure to cigarette smoke.  
• Throat swab: This will be taken by using swab to scrape the back of the 
throat. This will be used to detect any bacteria. 
• Nasal wash and nasal brush: This will be done by push some salt water into 
one nostril and collecting the return back, Then one side of the nose will be 
gently scraped with a tiny brush to retrieve some cells from the wall of the 
nose. These will also be used to isolate any bacteria. 
• Lung function testing – we will check your breathing with spirometry. It is a 
test to measure the amount of air you have in your lungs by breathing in 
and how well you can push the air back out by blowing it hard into a tube.  
If your asthma diagnosis has not been confirmed within 12 months of this 
visit we will repeat the spirometry (‘reversibility’ test) before and after you 
inhale salbutamol. This is to measure how much your airways respond to 
salbutamol. Spirometry may cause you to cough or make your chest feel 
tight. Salbutamol will be available if this occurs.  Salbutamol relaxes 
muscles in the air passages of the lungs. It helps to keep the airways 
open, making it easier to breathe. Salbutamol may make you feel a little 
shaky and increase your heart rate. 
• Forced oscillation technique - this test is used to measure lung movement 
during normal breathing. You will be asked to breathe normally through a 
mouthpiece into a tube during which time measurements are made. This 
will be repeated 3 times. 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
17 
 
• Collecting breath condensates. We will collect a sample of water condensate 
from the air you breathe out. We will use this to measure various markers. 
This is done by btreathing normally onto a tube for approx 10 min. There is 
no discomfort associated with this test. 
• Sputum induction - you will be asked to inhale a salty mist of different 
concentrations through a nebulizer and then asked to cough into a plastic 
container.  This process will be repeated on the second day.  Your lung 
function will be monitored. You may experience an unpleasant salty taste 
in your mouth, nausea or a sore throat. The salty mist may cause you to 
wheeze. You will be given salbutamol before the test to prevent the 
likelihood of wheezing and if necessary further salbutamol can be given if 
wheezing develops. This will be used to measure the inflammation in your 
airways. The sputum sample will also be used to culture fro bacteria and 
will also be used to detect bacterial and viral genetic material. 
• Exhaled Nitric Oxide -Nitric oxide is a gas present in everyone’s breath. 
You will be asked to exhale into a mouthpiece, breathing out at different 
speeds. By doing this the nitrogen content of your breath will be measured 
by a computer.  This will measure the inflammation in your airways. 
If you are one of those asked to take part because of your asthma, but who has not 
had your asthma diagnosis confirmed in the last 12 months we will want to carry out 
another test on a day separate from the previous assessments. 
• Methacholine challenge test - before we carry out this test we will ask 
women of child bearing age to confirm that they are not pregnant.  If there 
is any doubt a pregnancy test will be carried out.   You will be asked to inhale 
a mist that contains different concentrations of methacholine. The mist is 
produced by a device called a nebulizer and you inhale the mist through a 
mouthpiece. Before the test begins and after each period of inhalation you 
will be asked to blow forcefully into a spirometer.  This test is used to 
determine how responsive (or irritable) your airways are and to determine 
the severity of any asthma.  The inhalation of aerosols may be associated 
with mild shortness of breath, cough, chest tightness, wheezing, chest 
soreness or headache. Many patients do not have any symptoms at all. 
Symptoms (if they occur) are mild, last only a few minutes, and disappear 
following the inhalation of a bronchodilator medication such as salbutamol.  
• We will discuss with you your experience of your previous exacerbations 
and what warning you get before the onset of an exacerbation.We will 
discuss with you what would be an early warning for you to start recording 
your symptoms and peak flow measurements at home on a machine that 
we will provide you. We will ask you to call us as soon as possible when you 
feell an exacerbation is impending. We will ask you to take or seek treatment 
for your exacerbation as you would usually do. We do not wish you to 
withhold treatments for your exacerbation.  
• Diary card – we would like you to collect information on your asthma for 
the next two weeks. We will give you a machine to use twice daily to 
record your peak flow and spirometry tests. You will be given instructions 
as to how to use the machine at home. 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
18 
 
• If your asthma worsens and you have an ‘exacerbation’, we will want you 
to record information as soon as possible for 4 weeks.  We will explain to 
you what you need to record and when to do this. We will then review the 
information you have recorded with you at your next follow-up visit.  
 
Visit 2 - Exacerbation visit – this should take place within the first 6 month from the 
time of visit 1. This visit will take place at the Asthma lab at the Royal Brompton 
Hospital; you will not be required to stay overnight, unless there is any reason for 
you to need further treatrment in hospital. You will receive treatment as usual for 
your exacerbation. We will arrange transport for you to attend our lab. 
At this visit we would like to do the following: 
• Physical examination – the same as the Screening Visit.  
• Medication history- we will ask you about any other medicines you are 
taking.  
• Lung function testing - We will test your breathing as before, but will not 
need to confirm your asthma diagnosis.  
• Forced oscillation technique - this test is used to measure lung movement 
during normal breathing. You will be asked to breathe normally through a 
mouthpiece into a tube during which time measurements are made. This 
will be repeated 3 times. 
• Blood sample – we would like to obtain a sample of up to 50 ml (10 
teaspoons) of blood for assessing markers of inflammation. By markers we 
mean substances that include molecules called proteins, lipids and 
messenger RNA. Messenger RNA can help us understand how genes 
function. These and other molecules can affect disease processes (in this 
case asthma) and how patients like you respond to medicines. In addition, 
we may need to use a small amount of the blood to look at a specific 
substance (IgE) that can be increased if you are allergic to certain things, if 
not already done in the 3 months before screening. 
• Urine sample- we will ask you to provide a urine sample (about 40 ml) to 
look for substances that are indicators of asthma.  
• Sputum induction - you will be asked to inhale a salty mist of different 
concentrations through a nebulizer and then asked to cough into a plastic 
container.  This process will be repeated on the second day.  Your lung 
function will be monitored. You may experience an unpleasant salty taste 
in your mouth, nausea or a sore throat. The salty mist may cause you to 
wheeze. You will be given salbutamol before the test to prevent the 
likelihood of wheezing and if necessary further salbutamol can be given if 
wheezing develops. This will be used to measure the inflammation in your 
airways. 
• Exhaled Nitric Oxide -Nitric oxide is a gas present in everyone’s breath. 
You will be asked to exhale into a mouthpiece, breathing out at different 
speeds. By doing this the nitrogen content of your breath will be measured 
by a computer.  This will measure the inflammation in your airways. 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
19 
 
• Exhaled Breath Condensate – you will be asked to breathe into a cold 
tube. Due to the warmth of your breath a liquid will be produced, called a 
‘condensate’. This can then be analysed to determine the inflammation in 
your airways.  
• Nasopharyngeal swabs – We will take a mild scrape of the back of the 
throat in order to look for bacteria.  
• If any of the tests prove to be too uncomfortable for you, we will not 
proceed with the test. 
 
Visit 3 – Recovery exacerbation visit – after 14-21 days after Visit 2.  Most of the 
assessments will be over 3 hours. During this visit we would like to do the following:  
• Questionnaires – these will include asthma questionnaires, to check for any 
changes since the Baseline Visit.  
• Physical examination – the same as previous visits.  
• Asthma and medication history- since the last visit. 
• Lung function testing – the same tests as carried out at the Baseline Visit, 
including the ‘reversibility’ test.  
• Diary card –we will review the information you have recorded.  
• Pregnancy test- if appropriate  
If you have attended the ‘Repeatability’ Visit you will not be required to attend this 
visit. 
 
Visit 4 - Second Exacerbation visit  (Similar to Visit 2)– This is optional as we are 
looking for half of the subjects undergoing this. The same tests will be performed as 
above.  This visit will take place over not more than 3 hours). You will not be required 
to stay in hospital overnight.   
 
Visit 5-Second recovery Exacerbation visit (Similar to Visit 3)- This is optional 
and will only be done if you have gone through Visit 4. The same tests will be 
performed again. 
 
 

Title: Exacerbation in severe asthma patients: mechanisms and biomarkers 
 
 
20 
 
CONSENT FORM: 
 
Title of Study: Exacerbations in severe asthma patients: mechanisms and biomarkers 
 
Name of Researcher:  
 
Subject identification number: 
Please initial relevant box 
Yes      
  No 
 
1. I have received verbal information about this study and confirm that I have 
read and understand the information sheet for the above study. I have had the 
opportunity to consider the information, ask questions and have had these 
answered satisfactorily. 
 
2. I understand that my participation is voluntary and that I am free to withdraw at 
any time, without giving any reason, without my medical care or legal rights 
being affected. 
 
3. I understand that relevant sections of my medical notes and data collected 
during the study may be looked at by responsible individuals from Imperial 
College, the NHS Trust or from regulatory authorities where it is relevant to my 
taking part in this research. I give permission for these individuals to have 
access to my records. 
 
  
4. I agree to my hospital Consultant and GP being informed of my participation in 
the study. 
 
5. I understand that the samples/data may be used for commercial development, by 
the partners (academics and pharmaceutical industry) and with agreed third 
parties without financial or other benefit to me, for the investigation and treatment 
of medical conditions, potentially leading to new preventive measures for such 
conditions in keeping with the gift nature of my sample. 
 
6.  I agree to take part in the Study 
 
7. I have received a copy of this information sheet and informed consent 
 
________________________             _______________            _____________________________ 
Name of subject                                 Date                                   Signature                    
 
 
_________________________         ________________       ___________________________ 
Name of person taking consent 
(if different from researcher) 
   Date                                 Signature 
 
_________________________ 
________________ 
____________________ 
Researcher 
   Date 
Signature 
When completed, 1 for patient; 1 for researcher site file; 1 (original) to be kept in medical notes