Mechanisms of sensitization, disease development and desensitization:

1
Mechanisms of sensitization, disease development
and desensitization towards novel approaches for
prevention and therapy Ronald van Ree Academic
Medical Center, Amsterdam
2
Priorities for allergy/asthma research

• Immunology B-cells and antibodies
• Immunotherapy mechanisms and monitoring
• Atopic march eczema ? asthma ? rhinitis ?
  asthma?

3
B-cells and antibodies
Interaction between allergen and IgE is at the
basis of allergy and (extrinsic) asthma.
Exposure to allergen results in no
response? protective response (active
tolerance)? sensitization? /- symptoms?
How is the process of B-cell activity towards or
away from allergic inflammation regulated?
4
genetic factors
exogenous factors (environment incl. allergens
infections, diet, lifestyle)
DC
Th2
Th1
IFN?
IL4/IL13
Th1
Th2
allergy
no allergy
5
genetic factors
exogenous factors (environment incl. allergens
infections, diet, lifestyle)
Scheme of a simple Th1-Th2 disbalance to
distinguish between allergic and non-allergic has
actually been dismissed as being too simple.
DC
Th2
Th1
IFN?
IL4/IL13
Th1
Th2
allergy
no allergy
6
genetic factors
exogenous factors (environment incl. allergens
infections, diet, lifestyle)
DC
Treg
Th1
Th2
Th1
?
IFN?
IL4/IL13
Th1 and/or Treg
Th2
IL10 TGFß
no antibody response? IgG? IgA?
allergy
no allergy
healthy non-atopic?
7
genetic factors

• Questions and Discrepancies
• healthy immune response against allergens
  Treg/IL10/TGFß or Th1/IFN? 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)?

exogenous factors (environment incl. allergens
infections, diet, lifestyle)
DC
Treg
Th1
Th2
Th1
?
IFN?
IL4/IL13
Th1 and/or Treg
Th2
IL10 TGFß
no antibody response? IgG? IgA?
allergy
no allergy
healthy non-atopic?
8
genetic factors
exogenous factors timing/dose/context of
allergen exposure
DC
Th1
modTh2
Th1
IL4/IL13
Th2
modTh2
IL10
allergy
no allergy
healthy atopic (or atopic treated by
immunotherapy) and non-atopic?
9

• 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.

genetic factors
exogenous factors timing/dose/context of
allergen exposure
DC
Th1
modTh2
Th1
IL4/IL13
Th2
modTh2
IL10
allergy
no allergy
healthy atopic (or atopic treated by
immunotherapy) and non-atopic?
10
High early exposure to cat allergen is protective
11

• 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-fold
• difference in serum titers.
• A major cause most likely is the poor generation
  of memory B-cells
• for IgE caused by inefficient processing of mRNA
  for membrane IgE.
• Circulating IgE is derived from long-lived plasma
  cells hiding in survival
• niches 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

12
Low exposure situation no memory only plasma
cells
plasma cells
From Aalberse RC et al. J Allergy Clin Immunol.
2004 May113(5)983-6.
Atopics respond with all three but IgG is not
protective under these conditions Non-atopics
only make a little IgG / are hypo-responsive
overall compared to atopics.
13
High exposure situation memory generation for
IgG but not for IgE
Same specificity
plasma cells
From Aalberse RC et al. J Allergy Clin Immunol.
2004 May113(5)983-6.
Poor expression of membrane form of IgE favors
apoptosis resulting in poor memory
This also reflects the situation during
allergen-specific immunotherapy
14
For primary prevention it is of the greatest
importance to investigate the dose-response
relation between allergen and (the quality) of
the immune response. The window of opportunity
is of great importance. A high-dose protective
effect as observed for cat has so far not been
found for house dust mite. For food allergens
this is even more debated. The outcome has very
significant public health impact. Promote cats
and peanut butter sandwiches early on or not?
15

• What is needed to study the dose-response
  relation
• between allergen exposure and (the quality of)
  the
• immune 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.

16
Regulatory B-cells, a missing link?
exogenous factors (environment incl. allergens
infections, diet, lifestyle)
genetic factors
CD1dhiCD5
In several mouse models of auto-immune diseases
an anti-inflammatory (protective) role has been
established for regulatory B-cells. Do they play
a protective role in allergy/asthma. If so, how
can they be promoted?
major source of IL10 promote Treg development
17
The quality of IgE antibodies Do they always
translate into clinical allergy? No There are
several examples of IgE responses that are
without clinical relevance, i.e. the
ones observed during parasite infections, the
ones directed to plant glycans and the ones
observed during successful immunotherapy. Tregs
or Th1 cells can not explain the lack of
biological activity. The explanation can also
not (always) simply be found in a protective
effect of IgG4 (blocking antibodies). How is
the biological activity of IgE assessed?
18
Stripped basophil protocol
basophils from non-atopic donor
stripped basophils from non-atopic donor
basophils re-sensitized with IgE under study
lactic acid treatment removal of IgE
incubation with serum sensitization with IgE
19
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?
20
IgE antibodies against plant N-glycans have the
same poor biological activity
An explanation needs to be found why some IgE
antibodies are poor inducers of mediator release,
i.e. without clinical relevance.
21
What is different about these type of IgE
antibodies? Lower affinity? Something else? Is
this also observed during immunotherapy where IgE
antibodies are persistent but skin reactivity
clearly decreases. Are these IgE antibodies the
same as before the start of therapy or are they
(partly) newly formed IgE antibodies.
22

• B-cells and antibodies have not received the
  attention
• they deserve compared to T-cells. They do however
• produce 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 e) and from
• indirect (µ to e via ?4)?
• 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?

23
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 to set up trials that
include in depth immunological analyses.
Better insight in mechanism is a prerequisite for
improvement of therapy. Primary outcome (natural
exposure) is an ill-controlled disaster! Towards
pollen chamber monitoring in season.