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Costimulation and the immune response
• There are positive and negative second signals• Costimulation regulates naive and activated cells• Expanding complexity of T cell costimulation
B7:CD28 family TNF:TNFR family CD2 family
+-
The B7/CD28 superfamily provides T cell costimulatio n
The B7/CD28 superfamily provides T cell costimulatio n
Comparing PD-1 with other CD28 Family Members
late lethalityhyperactivation
early lethalityhyperactivation
↓↓↓↓T cell activation
Knockout
PD-L1/PD-L2B7-1/B7-2B7-1/B7-2Ligand
ITIM / ITSMITIM-like domainNo ITIMSignaling
inducibleinducibleconstitutiveKinetics
T, BT cellsT cellsExpression
PD-1CTLA-4CD28
PD-L1 is broadly expressed in tissues and lymphoid tissues, while PD-L2 is expressed only on APCs
PD-L1
Parenchymal
Placenta, eye, vascular endothelium,
pancreatic islets
None
Hematopoietic
B cells, T cells, DCs, MΦs
DCs, MΦsPD-L2
TCR/CD3
MY
PP
PY
YYYY
CD28M
YP
PP
YYYYY
MY
PP
PY
YYYY P
Proximal signaling kinases
Transcriptional activation
Y
Y
SHP-2PD-1
Inhibition of antigen receptor
signaling
MHC
PD-LB7
PD-1 inhibits antigen receptor signaling
PD-1 is negative regulator of T cell activation
CFSE
PD
-1
Gated on CD8+ cells
• PD-1 is upregulated upon activation of lymphocytes.
• PD-1-/- mice develop spontaneous autoimmunity.
• PD-1 signaling can affect cytokine production and cell
cycle progression.
Lymphoid organs
NaiveCD4+
NaiveCD8+
Antigen Recognition
Activation/Clonal
Expansion
IL-2
IL-2
Differentiation
Effectorfunctions
Effector CD4 +
Peripheral tissues
Activation of B cells, macrophages, etc.
Memory CD4 +
Effector CD8 +
Killing of target cells, macrophage activation
Memory CD8 +
PD-1 in the priming and effector phase of the immune response
PD-L have overlapping roles in naïve T cell priming by DCs
WT DCsPD-L1-/- DCsPD-L2-/- DCsPD-L1/L2-/- DCs
104 naïve DO11.10 T cells were stimulated 1:1 with BM derived DCs
4
8
12
16
1 2 3 4 5
ng/m
l
DAY
0.2
0.4
0.6
0.8
1
1 2 3 4 5DAY
IFN-γγγγIL-2
DCs
OT-I
RIP-ovahi pancreas
Do naïve CD8 T cells that lack PD-1 display tolerance to peripheral self-antigens?
Ovalbumin-specific CD8+ T cells
PD-1-/- OT-I T cells are increased in the pancreatic LN and traffic into the pancreas
WT OT-I
PD-1-/- OT-I
0
1
2
3
4
3 5 7 14
0
1
2
3
4
Cal
cula
ted
# O
T-I
(x1
04 )
3 5 7 14Days post-transfer
ILN
Days post-transfer
PLN
0
1
2
3
3 5 7 14
pancreas
Days post-transfer
*
*
* *
* = p<0.05
Increased effector cells and infiltration of tissue s by PD-1 -/- OT-I T cells
Pancreas
WT PD-1-/-
7 days after transfer of 5 x 106 PD-1-/-OT-I or OT-I T cells into RIP-ovahi recipient
PLN
%D
ivid
ed
ILN
WT OT-I
PD-1-/- OT-I
0
20
40
60
80
3 5 7 14
*
0
10
20
30
40
50
3 5 7 14
Lymphoid organs
NaiveCD4+
NaiveCD8+
Antigen Recognition
Activation/Clonal
Expansion
IL-2
IL-2
Differentiation
Effectorfunctions
Effector CD4 +
Peripheral tissues
Activation of B cells, macrophages, etc.
Memory CD4 +
Effector CD8 +
Killing of target cells, macrophage activation
Memory CD8 +
PD-1 in the priming and effector phase of the immune response
Naïve PD-1-/- OT-I T cells induce diabetes in RIP-ova hi mice
Adoptive transfer of of 5 x 106 purified OT-I CD8+ cells (intravenous)
0
20
40
60
80
100%
Dia
betic
3 6 9 12 15 18 21 24 27 30
OTI (n=5)
PD-1-/- OTI (n=4)
Draining Lymph Node
Target tissue
Auto-antigen
Migration to target tissue
Pancreas
T cell mediated autoimmune destruction
Pathogenic T cell survival in target tissue
Activation of autoreactive T cell
by APC
PD-1-/-
Is PD-1 exerting a cell-intrinsic effect on cross-p resentation?
WT
PD-1 exerts a cell-intrinsic effect on cell divisio n
CFSE
Thy
1.2
ILN
WT OT-I
PD-1-/- OT-I
51±3
49±3
pancreas
83±4
17±4
PLN
66±3
34±3
OT-I only
OT-I from OT-I + PD-1-/- OT-I
0
20
40
60
%D
ivid
ed
ILN PLN
NOD mouse model of autoimmune diabetes
• Non-obese diabetic (NOD) mice are an in-bred strain that develop autoimmune T-cell mediated insulin-dependent diabetes mellitus (IDDM).
• Autoimmunity in the NOD mice has been shown to be under complex heritable polygenic control.
• The development of autoimmunity in the NOD mouse is also influenced by diet and environmental factors.
• Early onset of insulitis in PD-L1/PD-L2-/- NOD mice• Increased severity of insulitis in PD-L1/PD-L2-/- NOD mice• Increased number of T cells in pancreatic LN• Increased cytokine production by T cells from pancreatic LN• Early onset diabetes phenotype is found in PD-L1-/- but not
PD-L2-/- NOD mice
Week
WTNOD maleWT NOD femalePD-L1/PD-L2-/- NOD malePD-L1/PD-L2-/- NOD female
0
25
50
75
100
% D
iabe
tic
n=9 n=16
n=12
n=13
3 6 9 12 15 18 21 24 27 30 33 36 39
Does PD-L1/PD-L2 expression protect islets from T cell mediated autoimmunity?
Week
107 T cells from non-diabetic wild type NOD mice were transferred into wild type or PD-L1/L2 -/- NOD SCID mice.
PD-L1/PD-L2 expression on non-lymphoid cells inhibits autoimmune diabetes
T cells
WT→WT NOD SCID
WT→PD-L1/L2-/- NOD SCIDn=5
n=6
%D
iabe
tic
0
20
40
60
80
100
2 4 6 8 10 12 14
p=0.0006
0
20
40
60
80
100
%D
iabe
tic
7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22
WT NOD SCIDPD-L1/PD-L2-/- NOD SCIDBDC2.5 CD4+
Effectors
p=0.02
PD-L1/PD-L2 expression can regulate CD4 + effector T cell function
2 x 105 BDC2.5 CD4+CD25-CD62L+ T cells from BDC2.5 WT NOD mice were transferred into WT NOD SCID or PD-L1/PD-L2-/- NOD SCID mice.
n=9n=9
Days
% o
f CD
4+ce
lls
0
4
8
12
16 PLN*
*
*
WT�
KO
WT�
WT
IFN-γ+W
T�
KO
WT�
WT
TNF-α+
IFN-γ+
WT�
KO
WT�
WT
TNF-α+
PD-L1/PD-L2 expression can limit CD4 + T celleffector cytokine production
2 x 105 BDC2.5 CD4+CD25-CD62L+ T cells from BDC2.5 WT NOD mice were transferred into WT NOD SCID or PD-L1/PD-L2-/- NOD SCID mice. Recipients were sacrificed 7 days after transfer.
*p<0.05
Can PD-L1/PD-L2 expression on antigen presenting ce lls protect against rapid induction of diabetes ?
PD-L1
PD-L1
CD11c+
CD11c+
Adoptive transfer prediabetic T cells into BM chimeras
PD-L1/PD-L2 expression on APCs, but not on parenchymal cells
No PD-L1/PD-L2 expression on APCs or parenchymal cells
A
B
Non-diabetic WT NOD
T cells
PD-L1/PD-L2-/- NOD SCID BM→PD-L1/PD-L2-/- NOD SCID
CD11c+
B
WT NOD SCID BM→PD-L1/PD-L2-/- NOD SCID
PD-L1
CD11c+A
WT→CHIMERA AWT→CHIMERA B%
Dia
betic
T cells
0
20
40
60
80
100
2 4 6 8 10 12
n=5 n=9
Week
p=0.31
PD-L1/PD-L2 expression on antigen presenting cells does not protect against rapid induction of diabetes
PD-L1 expression on islets protects against self-re active T cells
PD-L1 on islets inhibits self-reactive T cells
PancreasPancreatic
LN
Self-Ag
Self-reactive T cells
PD-L1
Draining Lymph Node
Target tissue
Auto-antigen
Migration to target tissue
Pancreas
T cell mediated autoimmune destruction
Pathogenic T cell survival in target tissue
Activation of autoreactive T cell
by APC
PD-1
PD-1 can function at two checkpoints
PD-1
Draining Lymph Node
Target tissue
Auto-antigen
Migration to target tissue
Pancreas
T cell mediated autoimmune destruction
Pathogenic T cell survival in target tissue
Activation of autoreactive T cell
by APC
PD-1
PD-L1
PD-1 delivers stronger inhibition of effector cells
A strong association between PD-1:PD-L and cancer
Breast cancerGhebeh et al, Neoplasia 8:190Ghebeh et al, Int J Canc 121:751
Esophageal cancerOhigashi et al, Clin Canc Res 11:2947
Gastric carcinoma Sun et al, Tissue Antigens 69:19Wu et al, Acta Histochem 108:19
Glioma Parsa et al, Nat Med 13:84
LymphomaDorfman et al, Am J Surg Path 30:802Yang et al, Blood 107:3639
Non-small cell lung cancerKonishi et al, Clin Canc Res 10:5094
Oral squamous cell carcinomaTsushima et al, Oral Oncol 42:268
Ovarian cancerHamanishi et al, Proc Natl Acad Sci104:3360
Pancreatic cancer Nomi et al, Clin Canc Res 13:2151
Renal cell carcinoma Thompson et al, Clin Canc Res 13:1757Krambeck et al, Clin Canc Res 13:1749Thompson et al, Canc Res 66:3381Thompson et al, Cancer 104:2084Thompson et al , Urology 66:10
Urothelial cancer Nakanishi et al, Canc Immunol Immunother 56:1173Inman et al, Cancer 109:1499
PD-1:PD-L1 and cancer
Draining Lymph Node
Tumor antigen
Auto-antigen
Migration to target tissue
Tumor
T cell mediated tumor destruction
Tumor infiltrating lymphocyte (TIL)
Activation of autoreactive T cell
by APC
PD-1
PD-1:PD-L1 and cancer
Draining Lymph Node
Tumor antigen
Auto-antigen
Migration to target tissue
T cell mediated tumor destruction
Activation of autoreactive T cell
by APC
PD-1
Tumor
Tumor infiltrating lymphocyte (TIL)
PD-1 expression on TIL
PD-L1 expression on
tumor cells
Can PD-1:PD-L1 blockade invigorate the anti-tumor immune response?
QuickTime™ and aTIFF (Uncompressed) decompressor
are needed to see this picture.
Tumor clearance
Tumor tolerance
Tumor Ag-specific
lymphocyte
Tumor Ag
The promise of PD-1:PD-L1 blockade
• Increased anti-tumor T cell responsesBlank et al, Int J Cancer 119:317
• Increased tumor clearanceIwai et al, Proc Nat’l Acad Sci 99:12293Iwai et al, Int Immunol 17:133
• Increased efficacy of other anti-tumor therapiesHirano et al, Cancer Res 65:1089Nomi et al, Clin Cancer Res 13:2157
B7-1 also can bind and signal through PD-L1
• B7-1 interacts more strongly with PD-L1 (1.7 µM) than with CD28 (4 µM) but less strongly than with CTLA-4 (0.2 µM).
• B7-1:PD-L1 binding site overlaps with B7-1:CTLA-4 and PD-L1:PD-1 interfaces.
• Multiple classes of mAbs: single vs. dual blockers of interactions.
• B7-1:PD-L1 interaction bidirectionally inhibits T cell proliferation and cytokine production.
• Increased significance of B7-1 and PD-L1 on T cells.
Butte, Keir et al, Immunity (in press)
Lee AlbackerManish Butte
Yvette LatchmanSpencer LiangArlene Sharpe
Harvard Medical School
Gordon FreemanDana Farber Cancer Institute
Indira GuleriaMohamed Sayegh
Brigham and Women’s Hospital
Andi QipoMaria Koulmanda
Massachusetts General Hospital
QuickTime™ and aTIFF (Uncompressed) decompressor
are needed to see this picture.
This work was supported by the Cancer Research Institute