Stephen R DurhamSection Allergy and Clinical Immunology

NHLI, Imperial College

and Royal Brompton Hospital London

Mechanisms of subcutaneous allergen immunotherapy

WAO Cancun Dec 4th 2011

TGF-

IgA

(-)

IL-10

IgG4

(-)

IFN

IgG

Th1

Immunotherapy(high dose Ag)

Treg

Natural exposure (low dose Ag) + IgE

Eosinophil

IL-4

IL-5

IgE

AllergyAPC Th2

B cellIL-13

(Adapted from Robinson DS, Larche ML and Durham SR J Clin Invest 2004; 114: 1389-97 )

Mechanism of tolerance induced by allergen immunotherapy

J Allergy Clin Immunol 2008; 121: 1467-72

0

30

60

90

120

1 15 29 12 26 10 24 7 216 20 3 17 1 15 29 12 4 18 1 15 29 13 27 3 17

2000PollenCount/m 3

MAY JUNE JULY AUGUST

Before 2 years

MAY JUNE JULY AUGUST MAY JUNE JULY AUGUST

Baseline Treatmentn=44 n=37

Bx Bx

Immunotherapy

Placebo

Grass pollen immunotherapy: Local changes in nasal mucosa

Wilson DR et al., J Allergy Clin immunol 2001;107:971-76

% re

ducti

on fr

om b

asel

ine

(199

8/19

96)

0

-50

-100

P = .01

P = .007

Immunotherapy group

Placebo group

Symptoms Medication

Grass pollen immunotherapy for seasonal rhinitis/asthma

Wilson DR et al., J Allergy Clin immunol 2001;107:971-76

EG2+ eosinophils in the nasal mucosa

Before Peak

IT

0

20

40

60

80

100

EG

2+c

ell

s/m

m 2

Before Peak

Placebo Controls

p=0.004p=0.03

p=0.04

(105)(178)

Non-atopic

Wilson DR et al., J Allergy Clin immunol 2001;107:971-76

Eosinophils in the nasal mucosa

0

10

20

30

40To

tal

sym

pto

m s

core

(2

day

s)

0 10 20 30 40

EG2 +

cells / mm 2

peak season

r=0.74p<0.001

(178;51)

(75;35)

Eosinophils in nasal mucosa correlate with total symptom scores

Pre Post

Placebo

0

50

100

150

200

c-K

it+

mas

t ce

lls/

mm

2

Pre Post

IT Controls After IT

p=0.0000 p=0.03

p=0.001

(286)

p=0.1

p=0.004

(238)(215)

C-kit+ mast cells in the nasal mucosa

Nouri-Aria KT, Pilette C et al, J Allergy Clin Immunol 2005

ImmunotherapyHayfever Controls

0

10

20

30

Fo

xp

3+C

D25

+

p=0.03p=0.03ce

lls/m

m2

Im m unotherapyHayfever Controls

0

10

20

30

Fo

xp

3+C

D4

+

p=0.04

(36.7)

ce

lls/m

m2

p=0.7

Foxp3+CD25+ Foxp3+CD4+

Grass pollen IT increases Foxp3+ T cells in the nasal mucosa

(a)Blocking peptide

TonsilCD25Foxp3Tonsil

HF IT NC HF IT NC

(b)

CD4Foxp3 Nose

J Immunol 2004; 172: 3252-59

Pre-treatment

Posttreatment

Peak season

0

5

10

15

IL-1

0 m

RN

A+

cel

ls /m

m2

Pre-treatment

Posttreatment

Peak season

Posttreatment

Out ofseason

Normalcontrols

(18) (28)

Placebo Immunotherapy

p=0.03

IL-10 mRNA+ cells

IL-10 protein+ cells

IL-10

Grass pollen immunotherapy: IL-10 in the nasal mucosa

Nouri-Aria K et al J Immunol. 2004; 172:3252-9

0

10

20

30

40

Se

rum

Ig

G4 (A

rbit

rary

Un

its

)

Pre ITFeb-Mar

'96

Post ITMay Jun

'98

Pre ITMay-Jun

'96

Post ITOct'98

A.

p=0.000 p=0.000

Allergen-specific IgG4

Wachholz P et al J Allergy Clin Immunol 2003; 112, 915-922.

J Immunol 2007; 178: 4658-66

Pre-treatment

Posttreatment

Peak season

0

5

10

15

20

TG

F

mR

NA

+ c

ells

/mm

2

Pre-treatment

Posttreatment

Peak season

Posttreatment

Out ofseason

Normalcontrols

(73)(70)

Placebo Immunotherapy

p=0.05

(53)

(30)

TGF-

Grass pollen immunotherapy: TGF- in the nasal mucosa and serum IgA2

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

Before IT After IT

0

20

40

60

80

100

out in in outPh

l p5

IgA2

P= 0.0001

CD3+ cells CD68+ cells

0

20

40

%T

GF

-b

ex

pre

ss

ing

ce

lls

0

5

10

% c

ells

ex

pre

ss

ing

TG

F-

Percentage of TGF- expressing

Percentage of CD3+ cells

CD3+ and CD68+ cells

and CD68+ which are TGF-+

(A)

(B)

CD3 CD68

Colocalization of TGF-β to CD3+ and CD68+ cells

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

TGF-β Phl p5/IgA2

(x200) (x400)

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

Grass pollen immunotherapy: TGFb-positive and IgA-positive grass pollen-specific cells

in the nasal mucosa

0

4

8

12

0 50 100 150 200

(53.1)(73.1)

r=0.61p=0.009

Serum Phl p 5 IgA2 (a.u.)

TG

F-

mR

NA

(ce

lls/

mm

2 )

12

(53.1)(73.1)

A

B Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

Grass pollen immunotherapy:Local nasal TGF-beta+ cells correlate with

serum grass allergen-specific IgA

Post immunotherapy serum fractionation: Phl p-IgA1, Phl p-IgA2 and Phl p-IgG

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

Post-immunotherapy serum IgG fractions but not IgA fractions do inhibit IgE-FAB

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

Phl-p induced IL-10 from monocytes passively sensitised with post-IT serum fractions

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

Post immunotherapy serum IgA fractions induce IL-10 synthesis and release from monocytes

Pillette C, Nouri-Aria K et al J Immunol 2007; 178:4658-66.

IL-10: intracellular staining IL-10: RT-PCRIL-10: ELISA

J Allergy Clin Immunol 2008; 121: 1120-5

IL-10 induction and suppression of skin LPR precedes IgG inhibitory antibody activity

IL-10 IgG4

Skin LPR Skin EPR

J Allergy Clin Immunol. 2011 J AllergyJ Allergy 2011; 127: 509-516

Longterm tolerance after grass pollen immunotherapy: Study design

James LK, Shamji MH et al, J Allergy 2011; 127: 509-516

Longterm tolerance after grass pollen immunotherapy: combined scores and serum FAB

James LK, Shamji MH et al, J Allergy 2011; 127: 509-516

Longterm tolerance after grass pollen immunotherapy: serum IgG1 and IgG4

James LK, Shamji MH et al, J Allergy 2011; 127: 509-516

Relation between serum FAB, IgG4 and combined Sx/Rx scores

James LK, Shamji MH et al, J Allergy 2011; 127: 509-516

TGF-

IgA

(-)

IL-10

IgG4

(-)

IFN

IgG

Th1

Immunotherapy(high dose Ag)

Treg

Natural exposure (low dose Ag) + IgE

Eosinophil

IL-4

IL-5

IgE

AllergyAPC Th2

B cellIL-13

(Adapted from Robinson DS, Larche ML and Durham SR J Clin Invest 2004; 114: 1389-97 )

Mechanism of tolerance induced by allergen immunotherapy

Allergy and Clinical Immunology, Imperial College and Royal Brompton Hospital, London, UK

M H Shamji K T Nouri-Aria S J Till

J N Francis C Pilette M Makatsori

S M Walker L K James G Rotiroti

D R Wilson M R Jacobson G W Scadding

G Pajno P Wachholz M Calderon