Transcript Tumor immunology Advanced immunology Fall, 2001, 3

Tumor immunology
BC Yang
1
Expected life-span
100
60
10
Cancer/ Neuronal disorder
0 year
Infection/Development
Immune capability
2010
衛生署資料，台灣人十大死因
2
排名
年度
88
89
90
91
92
93
94
95
96
1
惡性腫瘤
135.32
142.23
147.68
152.88
156.01
160.54
163.8
166.5
175.9
2
腦血管疾病
57.39
60.1
58.82
53.46
54.98
54.48
57.8
55.2
56.2
3
心臟疾病
51.33
47.56
49.25
50.93
52.23
56.79
57.1
53.8
56.7
4
糖尿病
40.99
42.6
40.79
39.26
44.38
40.58
46.2
42.5
44.6
5
事故傷害
58.88
47.4
42.58
37.79
36.3
37.33
36.8
35.1
31.1
6
慢性病跟肝硬化
23.53
23.32
23.45
21.35
22.98
23.63
24.7
22.1
22.5
7
肺炎
18.2
14.88
16.77
20.17
22.6
24.44
25.0
23.6
25.7
8
腎病徵候群及腎病性病
15.78
17.45
18.15
18.55
19.08
20.67
21.2
20.6
22.2
9
自殺
10.36
11.14
12.45
13.59
14.16
15.31
18.8
19.3
17.2
10
高血壓性疾病
8.43
--
7.9
8.67
8.17
7.97
8.3
8.0
8.6
98年 癌症、心臟疾病（不含高血壓）、腦血管疾病、肺炎、糖尿病、事故傷害、慢性下呼吸道疾病、
慢性肝病及肝硬化，自殺、腎臟疾病。
1960 Nobel prize in physiology or Medicine
for discovery of acquired immunological tolerance
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THE JOURNAL OF EXPERIMENTAL BIOLOGY 207 (23): 4013-4014
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THE BIRTH OF TRANSPLANTATION IMMUNOLOGY: THE
BILLINGHAM–MEDAWAR EXPERIMENTS AT BIRMINGHAM
UNIVERSITY AND UNIVERSITY COLLEGE LONDON
There are very few scientific papers
that continue to have an impact on
research more than 50 years after
their initial publication. A paper
published in J. Exp. Biol. in 1951 by
Rupert Billingham and Peter Medawar
is one such paper.
(The technique of free skin grafting in mammals)
Evidence for active host defense against cancer
5
 80 years of immunotherapy. (Currie GA,
1972, Br. J. Cancer 26: 141)
 A critique of the evidence for active host
defense against cancer, based on
personal studies of 27 murine tumors of
spontaneous origin. (Hewitt HB, et al. 1976, Br.
J. Cancer 33:241)
Circumvent Evidence
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 The high frequency of cancers in
immunosuppressed patients
Extremes of age
 Immunosuppressive drugs

 Tumors that are infiltrated by T cells
have an improved prognosis
 Spontaneous regression occurs

Melanoma, breast, lung cancers, etc
 Circulating tumor antibodies
 MHC-I down-regulation
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The ways to prove effective
tumor immunity：
Immune surveillance, animal model
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Full immunity needed to fight cancer
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CD4+ T cells contribute to the remodeling of the
microenvironment required for sustained tumor regression upon
oncogene inactivation (Cancer Cell, 2010, 18:485-498)
Full immunity needed to fight cancer
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A fight between immune cells and cancer
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But, sometimes we lose
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THE GREAT
ESCAPE:
IMMUNE
EVASION
VERSUS TUMOR
PROGRESSION
有些事只有在變動的狀況才看得見事物的本質
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Difficulties due to study model
14
It is a dynamic process
15
death
survival
Tumor formation of Ras-over-expression cells in BALB/c
Immune selection in the development
of cancer: no two tumors are alike
16
Fey MF & Tobler A 1996
 Initiation,
proliferation
diversification
 Microevolution,
selection of
immune
resistance
 Immune escape
and unchecked
proliferation
Use of tumor cell lines
17
 Commonly derived from advanced
tumors
 Retain the genetic instability and lose
the ability of adaptations
 introduced in large numbers (more than
106 cells)
Successful immunization for cancer
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Immune therapy (T cell-based)
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 Recombinant and synthetic vaccination
 Cytokine treatments (IL-2; GM-CSF; IFN)
 Cellular therapy with tumor-specific CTL
 Engineered macrophages (Th1…?)
 Antigen-pulsed macrophages or dendritic
cells
Tumor associated antigens
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 Tumor-specific shared antigens: restricted
in expression to tumors and immune privilege sites.
 Tissue-specific differentiation antigens:
tyrosinase. (melanoma)
 Tumor-specific antigens: mutated,
tranlocated genes.
 Ubiquitous antigens with over-expression
in tumors.
Rosenberg SA. 1999, Immunity 10:281
Peptide epitopes for melanoma
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Nicholaas P, et al. 1999, Curr Opin Oncol 11:50
Tumor and activated T cells
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Two major pathways for TCL: Fas-mediated and perforrin-mediated
HLA class I molecules
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 Antigen presentation for T
cells, cytotoxic
 Inhibitory signals for NK
cells
Immune escape/erosion
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




Making invisible
Resistance to death
Repelling
Active suppression
….
An overview
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Examples
26
 Down-regulation of
the class I
presentation pathway
 Resistance to killing
 Antigen-specific
mechanisms (Treg?)
 Global mechanisms
Antigen-specific mechanisms
27
 Tumor antigen-loss variants
Loss of the melanoma tumor-associated
antigen in patients with recurrent metastatic
melanoma (J Clin Invest 1996, 98:1633)
 Tolerance
B cell tumors expressing class II induced a rapid
tolerance of cognate CD4 T cell carrying a
transgenic TCR. (PNAS, 1998, 95:1178)
Down regulation of the MHC class
presentation pathway
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 Downregulation of MHC class I expression is
frequently seen in human tumors.
 Loss of MHC-I as a mechanism for tumor escape
from CTL-mediated elimination (longitudinal study of
melanoma patients)
 Five major HLA altered phenotypes found in
tumor tissues (Human Immunol. 2000, 61:65)
The five altered phenotypes
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Normal
1. Total loss
2. Haplotype loss
3. Locus loss
4. Allelic loss
5. Compound
phenotype
A1A2B8B35Cw7Cw4
A1B8Cw7
A1A2B8B35
A2B8B35Cw7Cw4
A1
(Human Immunol. 2000, 61:65)
Resistance to killing
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Cytotoxic T cells
 Defective Fas pathway
 Resistance to Granzyme B
Innate immunity

Fas-L and Neutrophil
Nature 407, 789 5(2000)
Fas signal
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Caspase 3
Death substrates
Apoptosis
Apaf-1
Caspase 9
Key person in Fas signal study
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Shigekazu Nagata
Peter H. Krammer
For a man who spends most of his time thinking
about death, Shigekazu Nagata is remarkably
upbeat. For over a decade, he has been making a
name for himself with research on apoptosis, the
mechanism of programmed cell death, and
during that time he has watched the field come
alive.
Nature Medicine 7, 759 (2001)
BC Yang 2008/2/20
Loss sensitivity to Fas-mediated apoptosis
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 FLICE-like inhibitory proteins
 Bcl-2, Bcl-XL
 defected sphingomyelinase
activation (ceramide)
 decoy receptor 3 (DcR3), soluble
Fas
Fas-L+-melanoma cells are relatively
resistant to killing of neutrophils
34
B16F10
Chen YL, et al J. Immunol. 171:1183-1192.
Global Mechanisms
35
 TGF-beta
 IL-10
 Growth in immune privilege sites
 Mucin production: interfering
intercellular adhesion
 Fas-L? (Fas counterattack)
 Tumor extracellular matrix
TGF-b signaling in tumor signaling and cancer
progression
36
IL-10 (Th3
or Treg)
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Tumors or other cells in environments
Immune cells are there! But, in peripheral
stroma area (cell cuffing)
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NPC Done by 張逸如 (1998)
Hematoxylin/eosin staining
CD3+ cells
Shaping T cell functions by extracellular matrix.
How can tumor cells highjack immune cells?
Extracellular matrix shapes Fas-mediated apoptosis.
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

Protective: U-118, Huh-7, HeLa, A549,
NIH3T3
Non-protective: MCF-7, HepG2
Su CC, et al. 2007 Phosphatidylinositol 3-Kinase/Akt activation by
integrin-tumor matrix interaction suppresses Fas-mediated apoptosis in
T cells. J Immunol 179:4589-4597.
Coculture with tumor cells inhibits the
Fas-mediated apoptosis.
40
24 h
48 h
60.0
Apoptosis (%)
50.0
40.0
30.0
20.0
10.0
0.0
J
CH-11
U118V
U118V
+CH-11
U118R
U118R
+CH-11
HeLa
HeLa
+CH-11
Huh-7
Huh-7
+CH-11
A549
A549
+CH-11
Direct cell contact is required.
41
Apoptosis program is inhibited.
42
Caspase 3
Death substrates
Apoptosis
Apaf-1
Caspase 9
Fas-mediated death is not suppressed by components of
MAPK, NFkB, PKA, HSP70.
43
Jurkat
Jurkat + CH-11
Jurkat + U-118MG
Jurkat + U-118MG + CH-11
60
Apoptosis (%)
50
40
30
20
10
0
None
Z-vade
(10μm)
SP600125
(10μm)
PD98059
(10μm)
SB202190
(5μm)
SN-50
(10μm)
KT5720
(2.5μm)
HSP70
(10μm)
What will happen when Fas-stimulated immune cells
resist to die?
44
Tissue environment ?
Tumor
Fas-L
Fas
Cytokines ?
Immune
cells
T cells in tumor microenvironment
45
Jurkat and Molt-4 cells
were cultured alone
(lane 1) or in the
presence of U118(V),
U118(R), U373(V), or
U373(R) (lanes 2, 3, 4,
and 5, respectively) for
24 h.
林亨楷
J Immunol, 2003, 171: 3947–3954.
Regulatory T cells: the third man
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Shimon Sakaguchi (2000) Regulatory T Cells Key Controllers of
Immunologic Self-Tolerance Cell 101: 455-458
A very dynamic interactions
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 Tumor microenviroment
 Individual properties
A FasL mystery
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1. Tissue dependant
2. Reverse signaling?
3. Other than death-triggering
Joe O’connell, 2002, Fas ligand and the fate of antitumorur
cytotoxic T lymphocytes. Immunology 105:263-266.
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In vivo consequences of Fas-L expression by tumors
(using over-expression system)
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
Enhanced
rejection

Renca, MH134,
L5178Y, B16BL6, CT26*
Note: *:syngenic, nude, SCID
Delayed
rejection
CT26#
B16-F10
#:
allogenic, lpr
Ref: Lejeune FJ et al., 1998, Curr. Op. Immunol.
Distinct structure of tumor mass
51
Control
 Glioma in Nude
mice
 May be Fas-L
associated?
 Why TIL in
Fas-LR
particular sites?
 Penetration of tumor
by immune cells
B16-F10 (melanoma) in B6 mice
52
Control
Fas-LRibozyme
Kang, S. M. et al. 1997. Fas ligand expression in islets of Langerhans does not
confer immune privilege and instead targets them for rapid destruction.
Nature Med. 3:738.
53
Granulocytes mediates the Fas-L-associated apoptosis during
lung metastasis of melanoma that determines the metastatic
behavior (B16F10 in C57BL/6) Br J Cancer, 87: 359 (2002)
i.v. 5 x 105 cells /mouse; Observed at day 14; (bar = 1 cm)
The structure of FasL
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FasL reverse signaling
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 Suzuki I. and Fink PJ.. Maximal
Proliferation of Cytotoxic T lymphocytes
Requires Reverse Signaling Through Fas
Ligand. J. Exp. Med., 187:123, 1998.
 Boursalian TE, Fink PJ, Mutation in Fas
Ligand Impairs Maturation of
Thymocytes Bearing Moderate Affinity T
Cell Receptors, J. Exp. Med., 198, 349,
2003.
Pamela J. Fink, h.D.
 Sun M, Ames KT, Suzuki I,. Fink PJ, The
Cytoplasmic Domain of Fas Ligand
Costimulates TCR Signals, J. Immunol.,
177, 1481, 2006.
Department of Immunology
University of Washington
 Fas ligand (FasL/CD95L) is best known for its role in delivering apoptotic
signals through its receptor, Fas (APO-1/CD95). In this study, we present
evidence that FasL has a second role as a signaling receptor. …….(abstract)
56
Suzuki I. and Fink PJ. (1998) J. Exp. Med., 187:123.
Depressed proliferation and cell
recovery of FasL-deficient CTLs
relative to FasL+ CTLs over the
course of an MLC. (a and b)
[3H]TdR uptake and viable cell
counts over days 1-5 from longterm CTL lines from B6, B6.lpr,
and B6.gld mice cultured with
allogeneic H-2k splenocytes. (c)
CTL responders were purified
CD8+ T cells derived from B6
and B6.gld mice cultured with H2bm1 splenocytes. (d) [3H]TdR
uptake by CTL responders used
in a, cultured with C3H.gld
splenocytes.
Suzuki I. and Fink PJ. (1998) J. Exp. Med., 187:123.
57
 Fas ligand (FasL/CD95L) is best
known for its role in delivering
apoptotic signals through its
receptor, Fas (APO-1/CD95). In this
study, we present evidence that
FasL has a second role as a
signaling receptor. …….(abstract)
Ectopic expression of FasL in NIH3T3
58
(A). Full-length and truncated FasL. Adapted from Orlinick JR, et al. J Biol Chem
1997, 272:32221-29. Jodo S, et al. J Immunol 2005, 174:4470-4474. FasL-△70:
deleting the N-terminal 2-70 aa; FasL-△33: deleting the N-terminal 2-33 aa.
Full-length FasL suppresses lung metastasis, while truncated
FasL promotes tumor formation.
59
A
B
(A). Nude mice were injected with 5x105 cells of NIH3T3 cells carrying FasL as indicated via
tail vein. After 4 weeks, spontaneous transformed NIH3T3 tumor nodules formed in the lung.
Ling sections were stained with H/E. Tumor nodules show dark purple stain. (B). FasL
constructs were tranfected into B16F10 cells and injected into B6 mice. After 2 weeks, tumor
nodules on the lung surface were counted. N1: B16F10-transfected with plasmid vector,
Robozyme: B16F10-transfected with FasL-ribozyme plasmid.
FasL forms complex with Met and activate authentic
Met signaling pathway
60
林煥晴
A new paradigm for FasL signaling
61
林煥晴
Alternative paradigm
62
 A tumor is a local growth of abnormal
tissue consisting of genetic-altered
transformed cells and a number of other
cell types and connective tissue components
characteristic of each tumor type.
 No host, as a tissue, no fighting.
Seljelid R et al 1999, Anticancer Res 19:4809
Tumor Stroma
63
 Fibroblasts
 Macrophages
 mast cells
 Endothelium
 lymphoid cells (T, B, granulocytes, NK cells)
 intercellular substance; extracellular matrix
Effects of ECM on the tumor microenviroment:
Collagen and tumor fibroblast
64
Ctrl.
Fibroblast attraction
shCol
林若曦
Black squares: tumor cells; Round: lymphocytes;
Oval: macrophages; small circles: mast cells
65
 A. tumor as abnormal
growth of transformed
cells.
A
 B. Tumor as malignant
tissue.
B
 C. Tumor hijacks
macrophage to direct
growth.
C
Seljelid R. 1997, Scan J Immunol 46:437