Mechanisms of sensitization, disease development and desensitization:towards novel approaches for prevention and therapy
Ronald van ReeAcademic Medical Center, Amsterdam
Immunology: B-cells and antibodies
Immunotherapy: mechanisms and monitoring
Atopic march: eczema → asthma → rhinitis → asthma?
Priorities for allergy/asthma research
Exposure to allergen results in:
no response?
protective response (“active tolerance”)?
sensitization?
+/- symptoms?
Interaction between allergen and IgE is atthe basis of allergy and (extrinsic) asthma.
How is the process of B-cell activity towards or away fromallergic inflammation regulated?
B-cells and antibodies
Ig?B-cell
DC
Th1 Th2
naïveTh
IgMB-cell
Th1
Th2
no allergy
IL4/IL13IFNγTh1
Th2
allergy
IgEB-cell
genetic factorsexogenous factors
(environment incl. allergens infections, diet, lifestyle)
Ig?B-cell
DC
Th1 Th2
naïveTh
IgMB-cell
Th1
Th2
no allergy
IL4/IL13IFNγTh1
Th2
allergy
IgEB-cell
genetic factorsexogenous factors
(environment incl. allergens infections, diet, lifestyle)
Scheme of a simple Th1-Th2 disbalance to distinguishbetween allergic and non-allergic has actually been
dismissed as being too simple.
IgG/A?B-cell
DC
Th1and/orTreg
Th2
naïveTh
IgMB-cell
Th1 Th2
no allergy
IL4/IL13IFNγTh1
Th2
allergy
IgEB-cell
genetic factorsexogenous factors
(environment incl. allergens infections, diet, lifestyle)
Treg
no antibody response?IgG? IgA?
?
healthy non-atopic?
IL10TGFβ
IgG/A?B-cell
DC
Th1and/orTreg
Th2
naïveTh
IgMB-cell
Th1 Th2
no allergy
IL4/IL13IFNγTh1
Th2
allergy
IgEB-cell
genetic factorsexogenous factors
(environment incl. allergens infections, diet, lifestyle)
Treg
no antibody response?IgG? IgA?
?
healthy non-atopic?
IL10TGFβ
Questions and Discrepancies
• healthy immune response against allergens: Treg/IL10/TGFβ or Th1/IFNγ or both?• do B-cells of non-atopic individuals ignore allergens or produce IgG/IgA antibodies?• Is this different for low-exposure allergens (e.g. pollen and mite) and high- exposure allergens (e.g. cat, bee venom and occupational allergens)?
IgG1B-cell
DC
Th2
naïveTh
IgMB-cell
no allergy
IL4/IL13
IL10
Th1
Th2
allergy
IgEB-cell
genetic factorsexogenous factors
timing/dose/context of allergen exposure
modTh2
IgG4B-cell
Th1
modTh2
IgE?B-cell
healthy atopic (or atopic treated by immunotherapy) and non-atopic?
IgG1B-cell
DC
Th2
naïveTh
IgMB-cell
no allergy
IL4/IL13
IL10
Th1
Th2
allergy
IgEB-cell
genetic factorsexogenous factors
timing/dose/context of allergen exposure
modTh2
IgG4B-cell
Th1
modTh2
IgE?B-cell
healthy atopic (or atopic treated by immunotherapy) and non-atopic?
Questions and Discrepancies• is a modified Th2 response in atopic individuals IgG4 without IgE or with (little/harmless) IgE?• is there an early-life window of opportunity for inducing the protective modified Th2?• why are IgG4 responses to allergens higher in atopics than in non-atopics?• atopic: predisposition to produce IgE and high IgG4? non-atopic: no tendency to develop IgE and only low IgG4?• atopic background seems to be less important for IgE/IgG4 production in case of high exposures: cats, bee venom, occupational allergens, parasites.
Increased exposure to cat allergen
Allergy (sensitization)IgG (exposure / protection)
Lancet 2001 ; Tom Platts-Mills et al.
High early exposure to cat allergen is protective
Why are IgE responses always so low compared to IgG responses?
Half-life of IgE is very short but this can not explain the 1000-folddifference in serum titers.
A major cause most likely is the poor generation of memory B-cellsfor IgE caused by inefficient processing of mRNA for membrane IgE.
Circulating IgE is derived from long-lived plasma cells hiding in survivalniches like the bone-marrow and inflammatory sites.
Two situations:• low allergen exposure (e.g. pollen/mite) i.e. a weak Th2 response that will not effectively induce a germinal center necessary for induction of memory B-cells
• high allergen exposure (e.g. cat, bee venom) i.e. a strong Th2 response that will a generate mature germinal center facilitating induction of memory
Low exposure situation: no memory only plasma cells
plasma cells
Atopics respond with all three but IgG is not protective under these conditionsNon-atopics only make a little IgG / are hypo-responsive overall compared toatopics.
Fro
m:
Aal
bers
e R
C e
t al
.J
Alle
rgy
Clin
Im
mun
ol.
2004
May
;113
(5):
983
-6.
Poor expression of membrane form of IgEfavors apoptosis resulting in poor memory
High exposure situation: memory generation for IgG but not for IgE
plasma cells
This also reflects the situation during allergen-specific immunotherapy
Same specif
icity
Fro
m:
Aal
bers
e R
C e
t al
.J
Alle
rgy
Clin
Im
mun
ol.
2004
May
;113
(5):
983
-6.
For primary prevention it is of the greatest importance toinvestigate the dose-response relation between allergenand (the quality) of the immune response.
The window of opportunity is of great importance.
A high-dose protective effect as observed for cat hasso far not been found for house dust mite. For foodallergens this is even more debated.
The outcome has very significant public health impact.
“Promote cats and peanut butter sandwiches early on or not?”
What is needed to study the dose-response relationbetween allergen exposure and (the quality of) theimmune response?
• Analysis of (existing) birth cohort samples for IgG and IgA responses. In most cases only IgE has been measured. Multiple time points in first year of life are required. Multiplex systems now allow in detailed analysis with limited sample.
• Analysis of (existing) birth cohort samples for mRNA profiling
• In vitro cellular analyses to study the process of immune skewing, preferably in an allergen specific fashion.
• Mouse models that go beyond the ovalbumin model by using real allergens.
plasmaB-cellmemory
B-cell
Breg
immatureB-cell
exogenous factors
(environment incl. allergens infections, diet, lifestyle)
genetic factors
Regulatory B-cells, a missing link?
CD1dhiCD5+
major source of IL10promote Treg development
In several mouse models of auto-immune diseases an anti-inflammatory (protective)role has been established for regulatoryB-cells. Do they play a protective role inallergy/asthma. If so, how can they bepromoted?
The quality of IgE antibodies
Do they always translate into clinical allergy? No
There are several examples of IgE responsesthat are without clinical relevance, i.e. the onesobserved during parasite infections, the onesdirected to plant glycans and the ones observedduring successful immunotherapy.
Tregs or Th1 cells can not explain the lack of biological activity.
The explanation can also not (always) simply be found ina protective effect of IgG4 (“blocking antibodies”).
How is the biological activity of IgE assessed?
lactic acid treatment:removal of IgE
incubation with serum: sensitization with IgE
basophils fromnon-atopic donor
stripped basophils fromnon-atopic donor
basophils re-sensitizedwith IgE under study
Stripped basophil protocol
specific IgE
0
5
10
15
20
25
30
35
40
me
an
(S
E)
of
an
ti-m
ite I
gE
in I
U/m
l
Gabonese Dutch
Gabonese (pink) vs Dutch (green) (n=16) (n=7)
0
20
40
60
80
0.1 10 1000
ng/ml
% h
ista
min
e r
ele
ase
(S
E)
a-IgE
mite
a-IgE
mite
Allergen-specific IgE from parasite-infected children lacks activity.This is not explained by high allergen-specific IgG4 titers
What explains the poor biological activity?
0
20
40
60
0.0001 0.01 1 100 10000
concentration (ug/ml)
% h
ista
min
rele
ase transgenic human
lactoferrin
native humanlactoferrin
rPhl p 5
IgE antibodies against plant N-glycans have the samepoor biological activity
An explanation needs to be found why some IgE antibodies arepoor inducers of mediator release, i.e. without clinical relevance.
What is different about these type of IgE antibodies?
Lower affinity? Something else?
Is this also observed during immunotherapy where IgE antibodiesare persistent but skin reactivity clearly decreases.Are these IgE antibodies the same as before the start of therapyor are they (partly) newly formed IgE antibodies.
B-cells and antibodies have not received the attentionthey deserve compared to T-cells. They do howeverproduce the antibodies that cause the disease!!
Reasons to study IgE/IgG/IgA antibody responses:
• They are a good read-out of the nature of the immune response• It needs to be established when/why they are clinically relevant, i.e. either disease-inducing (IgE) or protective (IgG/IgA).• Is induction of IgG/IgA before IgE a protection against future sensitization?• What is the difference between IgE from direct (μ to ε) and from indirect (μ to ε via γ4)?
Reasons for studying B-cells:
• What determines the quality of IgE antibodies?• What is the role of Bregs in allergy/asthma?• How could we target persistent plasma B-cells for IgE?
Immunotherapy: mechanism and monitoring
Many of the same questions need to be answered.
Blocking IgG4 antibodies?What is the difference between IgE before and after therapy?Th1 skewing?Tregs?Or perhaps Bregs?
Expression profiling / proteomics for biomarker search
Investigator-initiated trials needed. Companies are too small toset up trials that include in depth immunological analyses. Betterinsight in mechanism is a prerequisite for improvement of therapy.
Primary outcome (natural exposure) is an ill-controlled disaster!Towards pollen chamber monitoring in season.