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Rhinovirus-Induced Asthma Exacerbations: How that · PDF fileDecreased IFN-ßmRNA...
Transcript of Rhinovirus-Induced Asthma Exacerbations: How that · PDF fileDecreased IFN-ßmRNA...
Rhinovirus-Induced Asthma Exacerbations:How that Happens and Implications for
Treatment
Larry Borish, M.D.Professor of Medicine and Microbiology
University of Virginia Health SystemCharlottesville, VA
DISCLOSURES
• ContractedResearch*– CumberlandPharmaceuticals
• Consultant;uncompensated– Allakos
• CompensatedDSMB– PluriStem
*allfundsgototheUniversityofVirginia
LEARNINGOBJECTIVES
• ExaminetheroleofbystanderallergicreactionsinthepathogenesisofRV-inducedasthmaexacerbations.
• DescribetheroleofaberrantinnateimmuneresponsesinRV-associatedasthmaexacerbations
• Clarifytherapeuticimplicationsfortreating andpreventing asthmaexacerbations
Acknowledgements
• Heymann LabPeter HeymannJoshua L KennedyHolly CarperDeb Murphy
• Braciale LabTom BracialeMartha Spano
• Borish/Steinke LabJohn W SteinkeJulie NegriMadison RamsdenTheresa Altherr
• Woodfolk LabJudith A WoodfolkLyndsey Muehling
• Turner LabRon TurnerDeborah Thacker
September (and May) Epidemics of Asthma Exacerbations in Children and Adolescents in North America
Johnston et al. J Allergy Clin Immunol 2005 Feb; 115 : 130-8.
Number of hospitalizations of children age 5 to 15 yo by week of the year in Ontario
0 10 20 30 40 50 60
Coronavirus
Enterovirus
Rhinovirus
Influenza A, B
Metapneumovirus
RSV A, B
Parainfluenza 1, 2, 3
Adenovirus
Percent of Patients Who Tested Positive for Each Virus
Wheeze
Control
p<0.001
Children (age ≥3) and Adolescents Hospitalized for Asthma in Charlottesville, VA:
Heymann, P et al JACI 2004;114:239-47
AsthmaControl
and this occurs despite there being no compelling evidence that RV infections only occur in September (and May)• RV infections occur year round• i.e., this cannot just be explained by kids returning
to school
Seasonal incidence of respiratory viral infections in hospitalized patients
Johnston S et al. Am J. Respir Crit Care Med. 1996;154:654-60.
• Evidence of an “allergic” reaction:– eosinophilia, high ECP, high leukotrienes
• High total IgE– allergic sensitization (atopy)
Zambrano, J., et al. J Allergy Clin. Immunol. 2003;111:1008-16
TOTA
L SE
RU
M IG
E [I
U/m
L]
2
8
32
128
512
2048
8192
WHEEZEn=79
CONTROLn=77
ASTHMAn=33
CONTROLn=34
ASTHMAn=20
CONTROLn=21
< 3 YEARS OLD 3 - 9 YEARS OLD 10 - 18 YEARS OLD
128
377
42
401
33
Asthma exacerbations link to a highly allergic phenotype
Asthma exacerbations link to a highly allergic phenotype
• Evidence of an “allergic” reaction:– eosinophilia, high ECP, high leukotrienes
• High total IgE– allergic sensitization (atopy)
Zambrano, J., et al. J Allergy Clin. Immunol. 2003;111:1008-16
– Hypothesis:– Autumn exacerbation requires RV AND:
• sensitivity to ragweed, alternaria, dermatophagoides– Spring exacerbation requires RV AND:
• sensitivity to grass
So, if this were true what would we observe in an environment where allergen exposure is perennial?
• Hypothesis:If the allergen exposure is perennial, and RV infections occur perennially then exacerbations will occur perennially without a seasonal “spike”
CostaRicaERStudyHospitalNacionaldeNiños,SanJosé
(ChildrenAges7to12years)
• February enrollmentwhenchildrenstartschool• October enrollment• Enrollmentincluded:
96childrentreatedforacuteasthmaexacerbations,and126non-asthmaticcontrols65childrenwhorequiredtreatmentforasthmawithintheprevious12months(i.e.,“stableasthma”atthetimeofenrollment)
Soto-Quiros,Metal.J.AllergyClin Immuol 2012;129:1499-1505
0%
20%
40%
60%
80%
100%
Current Wheeze Stable Asthma ControlN= 44 51 35 29 57 69
80%
73%
14% 14%10% 9%
p<0.001
p<0.001
February October February October February October
% C
hild
ren
with
Pos
itive
PCR
for
RV,
HEV
, or R
SV
ChildrenwithPositiveTestsforViralInfection(qPCR)
N=445135295769
*
*p<0.03forthepercentageofRVpositivetestsinFebandOct
RhinovirusRSVEnterovirus
Exacerbating asthmatics
Soto-Quiros,Metal.J.AllergyClin Immuol 2012;129:1499-1505
Wheeze Stable Asthma Control0
100
200
300
400
500
Tota
l IgE
Total IgE levels in Costa Rica
482
332
86
p=0.1
p<0.001
p<0.001
AsthmaExacerbation
(n=95)
StableAsthma(n=64)
Controls(n=123)
Soto-Quiros,Metal.J.AllergyClin Immuol 2012;129:1499-1505
D. pter
Blomia
B. germ
P. amer
Ascaris
Alternari
a
Aspergillu
sDog Cat
Bahia gr
0.1
1
10
100
1000
<0.35x 9 x 9 x 44 x 58 x 53 x 88 x 81 x 69 x 77 x 82
Tite
r of I
gE (I
U/m
l) 25
12
1.8 1.4 1.62.4
0.91.3
1.8 1.4
93 90 53 38 42 4 12 25 16 11% Positive:
*GMvalues(inblue)includeonlychildrenwhosetiterswere≥0.35IU/ml
Costa Rica: Titers of allergen-specific IgE antibody in children and adolescents having an asthma exacerbation:
Soto-Quiros,Metal.J.AllergyClin Immuol 2012;129:1499-1505
IgE to Mite (IU/ml)
Prob
abili
ty o
f Cur
rent
Whe
eze
1 10 100 1000
0.0
0.2
0.4
0.6
0.8
1.0
IgE to Mite (IU/ml)1 10 100 1000
0.0
0.2
0.4
0.6
0.8
1.0
RV Negative RV PositiveA B
Probability of Asthma Exacerbation Based on Titers of IgE ab to D. pteronyssinus and Tests for Rhinovirus (Costa Rica)
Soto-Quiros,Metal.J.AllergyClin Immuol 2012;129:1499-1505
Conclusions, part I
• Most asthma exacerbations in children and adolescents occur in association with RV infection
• This reflects concomitant presence of allergic sensitization to aerollergens expressed at the time of the infection
Two Schools of Thought• The seed= the virus
– Three virus clades• A (ICAM/LDL), B (ICAM/LDL) and C (Cadherin-related family
member 3)– Asthmagenic* Rhinovirus (Strain C?)
Clades/ SpeciesHRV-A(75serotypes)HRV-B(25serotypes)HRV-C(60genotypes)
Order Family GenusPicornavirinae Picornaviridae Enterovirus
Jacobs,SEetal.ClinicalMicrobiologyReviews2013;26:135-62
RV Serotypes and Frequency of Inducing of Inducing Asthma Exacerbations
Lee,W-M,etal.Am.J.Respir Crit.CareMed.2012;186:886-91
*this could reflect either increased inherent virulence or altered immune responseMSI – moderate to severe illness
Viral AnalysisAsthma (n=68)
Acute Rhinitis (n=32)
Control (n=14)
qPCR for RV, % positive 57.1* 56.2* 7.1RV-A, % positive 50 59 0RV-C, % positive 50 35 100RV-B, % positive 0 6Other viruses, % positive 21 6
*p<0.01 compared to control
Soto-Quiros, M et al. J. Allergy Clin Immuol 2012;129:1499-1505
RV-C
• Unique binding receptor is cadherin-related family member 3 (CDHR3)– Previously linked to increased susceptibility to severe
asthma exacerbations in GWAS study,(in contrast to RV-A/-B which bind ICAM-1 or LDLR)
• Identified and grown using sinus epithelial explants– Could sinus infection and “sinobronchial” reflexes
trigger the exacerbation?
CDHR3 expression in RV-C unsusceptible and susceptible cell lines
Bochkov YA et al. Proc Nat Acad Sci (USA) 2015
Computed Tomographic Study of the Common Cold
Gwaltney JMJr.,etal.NEngl JMed1994;330:25-30
Day 4 of illness Day 13
Two Schools of Thought• The seed= the virus
– Three virus clades• A (ICAM/LDL), B (ICAM/LDL) and C (Cadherin-related family
member 3)– Asthmagenic* Rhinovirus (Strain C?)
• The soil= the asthmatic– Innate immune deficiency in asthmatics
• Inability to control virusà increased viral load + exacerbations
*this could reflect either increased inherent virulence or altered immune response
Decreased IFN-ß mRNA expression from bronchial epithelial cells in response to RV infection
Wark PAetal.JExp Med. 2005;201:937-47
•with similar results shown for type III IFNs and IL-15
•and linked to higher viral loads
RVmayreplicatebetterinthelungsofasthmatics
Mosser et al, J Infect Dis. 185:734-743, 2002
Bronchus
• Infection(worsenedinfection)ofthelowerairway– inassociationwithreducedinnateimmuneresponse(typeIIFN,typeIIIIFN,andIL-15)
ButdoesRVevenneedtoinfectthelowerairwaytodriveanasthmaexacerbation?
RV
HumoralMechanismsofRV-inducedupperrespiratoryrhinitisdrivinglowerrespiratoryasthmaexacerbations
another consideration might be that RV-C uniquely infects the sinuses and acute sinusitis drives the asthma exacerbation
• Morerecentstudieshavenotconfirmedsignificantdifferencesininnateimmuneresponsivenessinthelungsofasthmatics
• andpre-treatmentwithIFN-ß wasassociatedwithlotsofSEswithoutrobustevidenceformuchbenefit
DoesRVreplicatebetterinthenoseofasthmatics?
however, this isn’t necessarily comparing viral load on the same day post-infectiontherefore, need to do viral challenge approach
RVqPCRforRVinNasalWashesfromChildrenwithAsthmaExacerbations,AcuteRhinitis,andControlsintheER
Kennedy,JLetal.Am.J.Respir.Crit.CareMed.2014;189:532-9.
Asthma Exacerbation Acute Rhinitis Controln=28 n=32 n=14
Viral loads in Controls and “Low” and “High” IgE Asthmatics:
Kennedy J et al. Am J Respir Crit Care Med 2014; 189:532-9
Despite similar viral loads, symptoms are worse in allergic asthmatics (and last longer):
Study Day
Upp
er R
espi
rato
ry T
ract
Sym
ptom
s
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
0
1
2
3
4
5
6
7ControlLow IgEHigh IgE
ControlLow IgEHigh IgE
ControlLow IgEHigh IgE
Innate Immune Response
Adaptive Immune Response
Alternative(complementary)hypothesis:Coulddiminishedinnateimmuneresponsealterthepathological responsetoRV?
• RVinfection– inhealthysubjects– producesnodiscerniblepathologyinnasalbiopsies:– Nopathology– Noidentifiableinfected/necroticcells
AbsenceofCytopathologyinRVRhinitis:AirwayEpithelialCellsEfficientlyEngulfApoptotic
EpithelialCells
Development of a flow cytometry based apoptotic cell phagocytosis assay:
Juncadella, I et al. Nature. 2013;27:367-71
andthisengulfmentisassociatedwithgenerationofanti-inflammatorymediatorsTGF-ß andPgE2*(andrequiresengagementofPSreceptors)
Juncadella, I et al. Nature. 2013;27:367-71
*and IL-10
CoulddiminishedinnateimmuneresponsealterthepathologicalresponsetoRV?
• normal innateimmuneresponse– apoptosis ofRVinfectedcells– enhanced IL-10/PgE2/TGF-ß – noinflammation
• diminished innateimmuneresponse– necrosis ofRVinfectedcells– reduced IL-10/PgE2/TGF-ß –inflammation– and,possibly,withIL-33secretion
Conclusions, part II
• RV-induced asthma exacerbations do not appear to reflect differences in viral load– or, by extension, do not reflect differences in innate
anti-viral immune responses– (but this could be associated with conversion of
apoptosis to necrosis)– (or, alternatively, perhaps this is limited to the lower
airway)
Buthowwouldanyofthisexplaintheenhanced“allergy”requirement?
i.e., it’s the soil
(there may be “asthmagenic” viruses but it is not their inherent virulence -- but the nature of the immune response they generate that is the problem)
and if asthmatics have such an awful innate capacity to respond to viruses, why aren’t they have horrific responses to influenza*, adenovirus, etc., etc., etc.?
Two Schools of Thought• The seed= the virus
– Three virus clades• A (ICAM/LDL), B (ICAM/LDL) and C (Cadherin-related family
member 3)– Asthmagenic Rhinovirus (Strain C?)
• The soil= the asthmatic– Innate immune deficiency in asthmatics
• Inability to control virusà increased viral load + exacerbations– Robust immune response with viral infection
• And this immune response is being directed to:– bystander allergens
Sohowdoallergicimmuneresponsesgetgenerated?
• Ingeneticallypredisposedindividualstheredevelopsakeycentralroleforepithelial-derived:– IL-25– IL-33– TSLP
IL-33
TSLP
DC
Th0 Th2
Epithelial damage
Allergen
Microbes
IL-25
IL-4IL-5IL-9IL-13
Exceptthatepitheliumdoesn’tactuallyneedThelper2lymphocytestogeneratetheTh2cytokinemilieu
IL-33
TSLP
Epithelial damage
Allergen
Microbes
IL-25
ILC2IL-5IL-13
IL-4IL-5IL-9IL-13
Mast Cell
DC
Th0 Th2
Andonceepithelialcellsareepigeneticallydifferentiatedtoproducethesecytokines,they– andtheirdaughtercells– constitutivelyandpermanentlyretainthattendency
IL-33
TSLP
IL-25
ILC2IL-5IL-13
IL-4IL-5IL-9IL-13
Mast Cell
Andthisepigenetic“differentiation”oftheepitheliumincludesprogrammingtosecretecytokines/chemokinesthatdriveeosinophilia(whichalsomakeTh2cytokines)
IL-33
TSLP
IL-25
ILC2IL-5IL-13
IL-4IL-5IL-9IL-13
Mast Cell
GM-CSF, CCL5, CCL11, CCL24, CCL26
Eosinophil
IL-4,IL-13
Methods• Primaryepithelialcellcultures
– InfectedwithRV39(3TCID50/mL)at33°C• Placedin37°Cincubatorandharvestedcellsat2,4,24,and48hours.
– EvaluatemRNAandproteinexpressionofdifferentcytokinesandchemokines
DD
CT
Nor
mal
ized
tob-
actin
p<0.01
2 4 24 48Hours
DD
CT
Nor
mal
ized
tob-
actin
2 4 24 48
Hours
n=6 (Asthmatics)n=9 (Controls)
DD
CT
Nor
mal
ized
tob-
actin
2 4 24 48
Hours
IL-25 TSLP
IL-33
Epigenetically-programmedrapidupregulationofIL-25/IL-33/TSLPmRNAfromRV-InfectedRespiratoryEpithelium
Andinourviralinfectionmodelsthereisa“suggestion”ofasystemicTh2signatureimmuneresponse:SerumtotalIgE(bydayofstudy)
*increased IgE in 9/10 subjects
Local IgE Production in the Nose of Allergic and Non-Allergic Rhinitis Patients
• Quantification of allergen-specific and total IgE in the circulation and in both nasal secretions / interstitial nasal fluid• The concept being that “diffusion” of IgE
would be allergen non-specific • and as such a ratio of specific:total IgE in the
nares compared to the circulation >1 would signify local production
34%*
19%**
0% 0% 0%
5%
10%
15%
20%
25%
30%
35%
40%
LocalDPIgE LocalBTIgE
Allergic Non-Allergic
0
20
40
60Pe
rcen
tage
AllergenDP BT
- no virus- RV/HEV+
Local IgE Production in the Nose of Allergic Rhinitis Patients With and Without
RV Infection
Despite similar viral loads, symptoms are worse in allergic asthmatics:
Study Day
Upp
er R
espi
rato
ry T
ract
Sym
ptom
s
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
0
1
2
3
4
5
6
7ControlLow IgEHigh IgE
ControlLow IgEHigh IgE
ControlLow IgEHigh IgE
Innate Immune Response
Adaptive Immune Response
Reflecting an exacerbated and prolonged allergic state?
so, is IgE important to RV-induced asthma exacerbations?
and will targeting IgE-mediated bystander allergic responses protect against RV-induced exacerbations?
Antihistamines / Intranasal CCS – never studiedAllergy Immunotherapy – never studiedOmalizumab?
Omalizumab in Allergen-Exacerbated Asthma:Reduction in Exacerbations
Soler M et al. Eur Respir J. 2001;18:254-61; Busse W et al. J Allergy Clin Immunol. 2001;108:184-90; Milgrom H et al. Pediatrics 2001;108:E36.
Includingexacerbationsduringtherhinovirus-associatedMay– Septemberasthmaepidemics
Seasonal Variation in Frequency of Exacerbations(width of the bands represents the 95% CI)
Busse, WW et al. New Engl J Med 2011; 364:2011
Conclusions, part III• RV-mediated asthma exacerbations in children and adolescents are
dependent upon an enhanced IgE-driven immune response to bystander allergens– this may be attenuated by treating the allergies:
• Intranasal CCS / Anithistamines – unstudied• IT – perhaps• Omalizumab
• Proof of this concept will require a randomized controlled trial of omalizumab in patients undergoing experimental RV challenge:– in progress