The role of T cells in the pathogenesis - ankararomatoloji.com fileThe role of T cells in the pathogenesis of rheumatic disease (….. with a focus on RA) “T cells in health and

The role of T cells in the pathogenesis of rheumatic disease

(….. with a focus on RA)

“T cells in health and rheumatic diseases” - Sheraton Hotel, Ankara, 13 March 2009

Andrew P. Cope MD PhDArthritis Research Campaign Professor of Rheumatology

Academic Department of RheumatologyDivision of Immunology, Infection and Inflammatory Diseases

King’s College School of MedicineKing’s College London

[email protected]

ANKARA ROMATOLOJİ SEMPOZYUMU “Sağlıkta ve Romatizmal Hastalıklarda T Hücreleri”13 Mart 2009 Sheraton Otel, Ankara

A working model of disease pathogenesis for RA

progression from UA to RA Healthy

population

little joint destruction

RA

much joint destruction

environmental triggers

genes

immune

response nodisease

auto-antibodies arthritis

onset

diagnosisof RA

non-specific arthritis

spontaneousremission

genotype serotype phenotype

remission


Susceptible host

Onset of arthritis

Chronicdisease

initiation persistence

tobaccosmoke

trauma

infection

checkpoints defining the evolution and progression of RA

↑ innateimmunity

initiation factors

adaptive i.r. to modified self

antigens

pathways perturbing

immuneregulation

effector cellsmediating

persistence

THERAPY

Checkpoints defining the immunobiology of RA

1

2

pathways regulating

jointintegrity

4

3 5


Overview of the Lecture

1. What triggers the adaptive immune response in RA?


What is the molecular basis of adaptive immunity?Thymic selection

a repertoire ofself-reactive

T cells

positive selection

cTECself

ligand

DPThy

TCR

MHCII

CD4/8

Lymphocytes are referential to self-ligands!

Maintenance of T cellsin the periphery

TCR “tickling”of self-reactive

T cells

APCmultipleligands

T cellclonotypicreceptor

dendritic cell


Immune recognition of “infectious non-self”(as opposed to “non-infectious self”)

DC

T

TCRCD28

MHCIICD80/86Signal 1 “constitutive”Signal 2

“constitutive”

“inducible”

How are costimulatory molecules induced?

pathogen derivedpeptide antigen


TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

Lipoproteins(bacteria,

mycoplasma,mycobacteria)

dsRNA(viruses)

LPShsp

Flagellin(bacteria)

ssRNA(viruses) synethetic

compounds

ssRNA CpG DNA Factors fromuropathogenic

bacteria

?Exogenous ligands

Pathogen Associated Molecular Patterns (PAMPs) and Pattern Recognition Receptors (PRRs)

TLR1/6 TLR2 TLR4 TLR5 TLR7 TLR8 TLR9 TLR10 TLR11TLR3

DANGER SIGNALS


Immune recognition of “infectious non-self”(as opposed to “non-infectious self”)

DC

T

TCRCD28

MHCIICD80/86Signal 1 “constitutive”Signal 2

“constitutive”

“inducible”

How are costimulatory molecules induced?

pathogen derivedpeptide antigen

TLR signal


TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

TIR

Lipoproteins(bacteria,

mycoplasma,mycobacteria)

dsRNA(viruses)

LPShsp

Flagellin(bacteria)

ssRNA(viruses) synethetic

compounds

ssRNA CpG DNA Factors fromuropathogenic

bacteria

?Exogenous ligands

Pathogen Associated Molecular Patterns (PAMPs) and Pattern Recognition Receptors (PRRs)

TLR1/6 TLR2 TLR4 TLR5 TLR7 TLR8 TLR9 TLR10 TLR11TLR3

Endogenous ligands

Hsp60,70HMGB1

Gp96HA, HS

FN

chromatin-IgGcomplexes

dsRNA ssRNAssRNA DNAnecroticcells

TLR signal transduction pathways

adapted from O’Neill, 2006


Inhibition of TLR dependent production of inflammatory mediators

Sacre et al, 2007 Am J Path 170: 518-525

Uninfec

ted

Adβ-gal

AdMyD

88dn

AdMald

n

AdIκBα

0

500

1000

1500

2000

2500

TNFα

(pg/

ml)

Uninfec

ted

Adβ-gal

AdMyD

88dn

AdMald

n

AdIκBα

0

1000

2000

3000

4000

MM

P-3

(ng/

ml)

Uninfec

ted

Adβ-gal

AdMyD

88dn

AdMald

n

AdIκBα

0.0

1.5

3.0

4.5

MM

P-13

(ng/

ml)




2. What do T cells see?


P4Asp-

β71Lys+

collagen II (1168-1180)N-terminus

C-terminus

Non-associated DRB1*0402β86 Valβ67 Ileβ70 Aspβ71 Glu Pocket 4}

DRα chainα-helix

DRβ chainα-helix

Disease associated DRB1*0401β86 Glyβ67 Leuβ70 Glnβ71 Lys Pocket 4}

Proposed hierarchy of HLA-DRB1 alleles indetermining disease severity in RA

(from Weyand et al, 1995)

*0401/*0401

*0401/*0404

*0401/*01

*0404/*01

*0401/X

*0404/X

*01/*01 nodular vasculitis

*01/X seropositive erosive disease

X/Xnon-erosive erosive

seronegative diseaseANKARA ROMATOLOJİ SEMPOZYUMU “Sağlıkta ve Romatizmal Hastalıklarda T Hücreleri”

13 Mart 2009 Sheraton Otel, Ankara

HCgp39 in IFA

DRαβ1*0401 DRαβ1*0402

Defining HLA-DR4 restricted T cell responses to cartilage antigens in transgenic mice lacking mouse MHC class II

harvest draining LN @ 10d

evaluate antigen & peptide

specific responses

fuse with TCRneg

thymoma asfusion partner

clone hybridomas


DRαβ1*0401 and DRαβ1*0402 present completely distinct sets of HCgp-39 peptide epitopes to CD4+ T cells

0

10

20

30

40

50

60

% a

ll re

spon

ding

hyb

rids

22 28 34 40 46 52 58 64 70 76 82 88 94 100

106

112

118

124

130

136

142

148

154

160

166

172

178

184

190

196

202

208

214

220

226

232

238

244

250

256

262

268

274

280

286

292

298

304

310

316

322

328

334

340

346

352

358

364

368

HCgp-39 peptide

DRαβ 1*0402 (n = 151)

DRαβ1*0401 (n = 250)

Cope et al, 1999 Arthritis Rheum 42:1497-1507


The immune response is different (1) a.

0

5000

10000

15000

20000

25000

30000

[3H

] thy

mid

ine

inco

rpor

atio

n (c

pm)

*0401 *0402DRB1*04 subtype

SI - 3.4 SI - 3.6

HCgp-39

medium

0

4000

8000

12000

16000

IFNγ

(pg/

ml)

*0401 *0402DRB1*04 subtype

b.

I-d pool

peptide pool

HCgp-39



0

4000

8000

12000

16000

IFNγ

(pg/

ml)

*0401 (C-line)

*0401(3C4)

*0402

DRB1*04 subtype

0

200

400

600

800

1000

1200

TN

F (p

g/m

l)

*0401 (C-line)

*0401 (3C4)

*0402

DRB1*04 subtype

c. IFNγ d. TNFα


The immune response is different (2)


DC

Teff

TCRCD28

MHC class II

B disease specificautoantibodies

antigen experiencedeffector T cells

Defining the key autoantigenic determinants


What have RA-specific autoantibodies taught us?

reactivity to“modified”

self

“stress”



self

“stress”

citrullinatedproteins

arginine → citrulline“neoepitopes”




self

“stress”

inflammation

infiltration

PAD2/4


mΦ, PMN

vimentin, fibrin, histones, α-enolase

deiminationCa2+

Schellekens, Vossenaar, Tak, van Venrooij et alinflammation specific




self

“stress”

inflammation

infiltration

PAD2/4


mΦ, PMN

PAD4 geneticpolymorphism

↑ mRNAstability

deiminationCa2+

Schellekens, Vossenaar, Tak, van Venrooij et alinflammation specific

Suzuki et al, Nat Gen 2003





self

“stress”

inflammation

infiltration

PAD2/4


HLA class II

T cellresponse

B cellresponse

mΦ, PMN

PAD4 geneticpolymorphism

↑ mRNAstability

synovialplasma cells

anti-CCP

TH

IgG1

inflammation specific disease specific

deiminationCa2+

Suzuki et al, Nat Gen 2003

?



P4Asp-

β71Lys+

collagen II (1168-1180)N-terminus

C-terminus

Non-associated DRB1*0402β86 Valβ67 Ileβ70 Aspβ71 Glu Pocket 4}

DRα chainα-helix

DRβ chainα-helix

Disease associated DRB1*0401β86 Glyβ67 Leuβ70 Glnβ71 Lys Pocket 4}

Arg not favouredCit permissiveat P4




3. Have any immunogenetic studies been informative?


DC

Teff

TCRCD28

modified“signal output”

Functional impact

Thymic selection

Cell survival

Th differentiation

Migration

Effector responses

Immune regulation

Gene polymorphism

HLA-DRB1

CIITA

PADI4

PTPN22

PD1

CTLA4

CD25

STAT4

“signal input”






4. What do RA T cells look like?


Prediction:

1.Activated phenotype2.Features of prior antigen exposure3.Extensive proliferative activity4.Expansions of selected (high affinity) clones5.Reactive to tissue specific (cartilage, synovium, bone)

antigens6. Features of distinct Th lineage differentiation7. Intrinsic survival advantage8. Potent migratory capacity9. Impaired regulatory function


RAsynovial

T cell

RAsynovial

T cell

senescence

memory effector

migration

antigen recognition

effectorresponse

TCR

TCRζdim

costimulation CD28

OX40

ICOSL

CD28null

CD45RO,CD45RBdim

CD44,CD69,CD62Llo

CCR7lo

VLA-1,VLA-4

LFA-1

CCR4,CCR5,CCR6, CXCR3,CXCR4,CXCR5

NKG2D

CX3CR1

CD25neg

CD27neg

CD40Lneg

IL-4RTNF+

IFNγ+

IL-17+

IL-10+

RANKL

H202+

mTNF

LTα1β2

IL-6R

IL-23R

IL-15R

TNFR

Cope AP 2008 Arthritis Res Ther

The reality:

Bcl-2lo

Bcl-XLlo





4. What do RA T cells look like?

5. Multiple pathways of T cell effector function


The TCR/CD3 antigen receptor complex

TCR/CD3 complex

TCRζbright

TCRζdim

CD3εTC

Rζ


TCRζdim cells are enriched for effector Th cell subsets

Zhang et al, Blood 2007

Data.027

100 101 102 103 104SSC-WIL-17

TCR

ζ

CD4+

TCRζdim

%IF

Nγ

expr

essi

ng c

ells

TCRζdimTCRζbright

P < 0.0005

0

10

20

30

40

50

60

70(A) (B)

(stim: PMA and ionomycin)


Regulatory CD4+ T cell subsets reside in the TCRζbright population%

IL-1

0 ex

pres

sing

cel

ls

TCRζdimTCRζbright

P < 0.0005

Zhang et al, Blood 2007

(A)

0

10

20

30

40

50

0

10

20

30

40

50

60

70

80

90

100

%Fo

xp3+

cells

(B)

TCRζdimTCRζbright

P < 0.0001


healthy donorPBL

transwell

EC

gelatin

TCRζbright

TCRζdim

TNF

Migration of TCRζdim T cells across activated endotheliumlower chamberupper chamber

CD3

TCR

ζ

88.5%

11.5%

25%

75%

bright dim bright dim bright dim0

20

40

60

80

100

% c

ells

mig

ratin

g

TCRζ expressing subset CD3 CD4 CD8

p < 0.008 p < 0.035 p > 0.16

Zhang et al, Blood 2007ANKARA ROMATOLOJİ SEMPOZYUMU “Sağlıkta ve Romatizmal Hastalıklarda T Hücreleri”


- +--

- - - - -- - - - -

+ ++

+- - -

+

--0

500

1000

1500 IFNγ IL-10

Accessory signals are intact in TCRζdim T cells

in collaboration with Fionula Brennan TCRζdim cells generated by stimulation with IL-2, IL-6 and TNFα

cyto

kine

pro

duct

ion

(pg/

ml)

TCRζdim cells plus: CHO/vector:CHO/CD80:CHO/CD86:


0

50

100

150

200

250

300m

onoc

ytes 3:

1

5:1

7:1

T cells:monocytes

TNFα

(pg/

ml)

(TCRζdim)(TCRζbright) mon

ocyt

es

T ce

lls

mon

ocyt

es+

LPS 3:1

5:1

7:10

100

200

300

4004000

5000

T cells:monocytes

ζbright ζdim

Analysis of contact dependent effector responses


0

10

20

30

40

50

60

70

80

CD3ε

TCR

ζ

TCRζdim T cells are enriched at sites of inflammation

SF

PB SF SM

CD3+ T cells

% T

CR

ζdim

T ce

lls

PB

Zhang et al, Blood 2007ANKARA ROMATOLOJİ SEMPOZYUMU “Sağlıkta ve Romatizmal Hastalıklarda T Hücreleri”


% C

D3+ T

CR

ζdim

cells

PB SF SM 0

10

20

30

40

50

60

70

80

TNF blockade

effector T cells

TNF blockade

• memory, activated• CCR/CXCR profile• integrins• senescence• Ag engagement

PHENOTYPE

Persistence of TCRζdim effector T cells after infliximab therapy

circulating TCRζdim

synovial tissue

high disease activity

circulating TCRζdim

synovial tissue

low disease activity

CD

3TCRζ

11.1% 88.9%

TCRζ

45.4% 54.6%

KEY MESSAGES

1. The phenotype and function of PB and synovial T cells points to a key role for inflammation and aging in the pathogenic process.

2. Expression of costimulatory molecules can be exploited for therapeutic process.

3. Antigen determinants for T cells remain poorly define. Extendedautoantibody profiling is likely to provide insights in the future.

4. T cell effector pathways are complex. The precise role of Th1, Th2 and Th17 cells requires further investigation.

5. Deeper understanding of the basis for defective immune regulation is urgently required.

6. Genome wide association studies have provided new clues to the perturbations of adaptive immunity in chronic inflammatory autoimmune diseases such as RA.


FLS

ACUTE PHASIC CHRONIC counter-

regulatorynetworks

TLR inhibitors IL-4, IL-10, TGFβ, IL-13, L-11, IL-2 IL-1Ra, sTNF-R, IL-10, IL-18bp, OPG, adiponectin

transmigration of monocytesPMN, mast cells and NK cells

tissue damage,TLR ligands

TLR

IFNs +survivalfactors

1, TNF, IL-6, IL-22PGE2, bFGF, PDGFEGF, VEGF, TGFβ

COX, LOX

CXCL5/8/9/10/12/13,CCL2/3/5/20/21

CX3CL1

LTβR

activation of endothelium,↑ adhesion molecules

INITIATION ANTIGEN MODE INFLAMMATION MODE

B

Teff

MΦ FLS

Adipo

TCRζdimCD28null

Treg

IL-2deficiency

+ IL-6

IL-1, IL-6, TNFIL-15, IL-32GM-CSFoncostatin MM-CSF, VEGFChemokinesadipokinesHMGB1IL-10

tissue response

adipokines

osteoclasts chrondrocyte

TNFM-CSF

OPG

RANKL

IL-17+

MMPADAMADAM

TNFIL-1

onco-statin M

PGE2

features of inflammation and repair

DC

Teff

TCR

CD28

B

Treg

CD40L

CD40

IL-2

EARLY LATE IL-2, IL-4, IFNγIL-13, TGFβ IL-17IL-15 RANTESGM-CSF TNF

LTα/βIL-10

IL-12, IL-23IL-15, IL-18

IL-6IL-10chemo-kines

AutoAb

BLySAPRIL

TGFβIL-10

TGFβIL-10

CD80/86

Credits ………

TCRζ genetics

Claire Gorman

Andrew RussellTim Vyse

mRNA stability

Andy Clark (3’UTR)

TCR signalling

Joanna ClarkZhuoli ZhangPia IsomäkiKarolina AleksiyadisNina Panesar

Federica Marelli-BergClaudia MonacoEnrico Ammirati

Alex AnnenkovYuti Chernajovsky

SKG mouse studies

Xiang WuOli SomenziTharsana Tharmalingam

Shimon Sakaguchi

FundingWellcome TrustarcMRCEU


Documents

The role of T cells in the pathogenesis - ankararomatoloji.com fileThe role of T cells in the pathogenesis of rheumatic disease (….. with a focus on RA) “T cells in health and