Download Tumor suppressor genes(TSGs)

Tumor pathogenesis
 Oncogenes
 Tumor suppressor genes
 Invasion and Metastasis
Introduction
 Carcinogensis is multistep process involving the multiple
genetic changes including the activation of cooperating
oncogenes and the inactivation of tumor suppressors in
somatic cells
2
 Usually, a single oncogene is not enough to turn a normal cell
into a cancer cell, and many mutations in a number of different
genes may be required to make a cell cancerous.
肿瘤的信号转导通路调控异常
Figure 2. Intracellular Signaling Networks Regulate the Operations of the Cancer Cell. An elaborate integrated
circuit operates within normal cells and is reprogrammed to regulate hallmark capabilities within cancer cells.
Separate subcircuits, depicted here in differently colored fields, are specialized to orchestrate the various
capabilities. At one level, this depiction is simplistic, as there is considerable crosstalk between such subcircuits.
In addition, because each cancer cell is exposed to a complex mixture of signals from its microenvironment, each
of these subcircuits is connected with signals originating from other cells in the tumor microenvironment, as
outlined in Figure 5. (Hanahan D, Weinberg RA. Hallmarks of Cancer: The Next Generation. Cell 2011, 144:646)
Michael R. Stratton. Exploring the Genomes of Cancer Cells: Progress and Promise. Science 331, 1553 (2011).
Michael R. Stratton. Exploring the Genomes of Cancer Cells: Progress and Promise. Science 331, 1553 (2011).
Oncogene
Concept:
An oncogene is a gene that when mutated or
expressed at abnormally-high levels contributes to
converting a normal cell into a cancer cell.
 Cellular oncogene (c-onc)：
--- proto-oncogene （proto-onc）：in normal physiologic
version
--- Oncogene：altered in cancer
 Viral oncogene （v-onc）
Fuctions of proto-oncogenes
Proto-oncogenes have been identified at all levels of the
various signal transduction cascades that control
cell growth, proliferation and differentiation:




extracellular proteins function as growth factors,
membrane proteins as cell surface receptors
cellular proteins that relay signals
proteins in nucleus, which activate the transcription and
promote the cell cycle
This signaling process involves a series of steps that:



begin from the extracellular environment to cell membrane;
involve a host of intermediaries in the cytoplasm;
end in the nucleus with the activation of transcription factors
that help to move the cell through its growth cycle.
Classification of proto-oncogenes
 Growth factors, e.g. V-sis, PDGF-b, int-2
 Receptor Tyrosine Kinases, e.g. Her-2/neu/ erbb2,
 Membrane Associated Non-Receptor Tyrosine Kinases,





e.g. src, Lck
G-Protein Coupled Receptors e.g. Mas
Membrane Associated G-Proteins , e.g. Ras
Serine/Threonine Kinases e.g. Raf
Nuclear DNA-Binding/Transcription Factors, e.g. myc, fos
Others
Apoptosis regulators, e.g. Bcl-2,
Regulators of cell cycle, e.g. Cyclin D1, CDK4
Mechanisms of Oncogene Activation
1. Gene amplification, e.g. myc, CCND1
2. Point mutation, e.g. ras,
3. Chromosomal rearrangement or translocation
the transcriptional activation of proto-onc.
 the creation of fusion genes, e.g. abl-bcr
4. Viral insertion activation, e.g. c-Myc

Translocation
Amplification
CHROMOSOMAL REARRANGEMENTS OR TRANSLOCATIONS
Neoplasm
Translocation
Proto-oncogene
Burkitt lymphoma
t(8;14) 80% of cases
t(8;22) 15% of cases
t(2;8)
5% of cases
c-myc1
Chronic myelogenous
leukemia
t(9;22) 90-95% of cases
bcr-abl2
Acute lymphocytic
Leukemia
t(9;22) 10-15% of cases
bcr-abl2
1c-myc
is translocated to the IgG locus, which results in its activated expression
2bcr-abl
fusion protein is produced, which results in a constitutively active abl kinase
15
GENE AMPLIFICATION
Oncogene
Amplification
Source of tumor
c-myc
~20-fold
leukemia and lung carcinoma
N-myc
5-1,000-fold
neuroblastoma
retinoblastoma
L-myc
10-20-fold
small-cell lung cancer
c-abl
~5-fold
c-myb
5-10-fold
acute myeloid leukemia
colon carcinoma
c-erbB
~30-fold
epidermoid carcinoma
K-ras
4-20-fold
30-60-fold
colon carcinoma
adrenocortical carcinoma
chronic myoloid leukemia
16
Mechanisms of action of oncogenes
Ras
Locates on chromosome 11, codes for a protein with
GTPase activity
relays signals by acting as a switch: When receptors on the
cell surface are stimulated, Ras is switched on and transduces
signals that tell the cell to grow. If the cell-surface receptor is not
stimulated, Ras is not activated and so the pathway that results in cell
growth is not initiated.
mutated in about 30% of human cancers so that it is
permanently switched on, telling the cell to grow
regardless of whether receptors on the cell surface are
activated or not.
Her2/neu/erbB-2
This gene was discovered by three different
groups. That is why it has three different
names.
It is a member of EGFR superfamily, also be
a receptor tyrosine kinases
Dr. Slamon (UCLA) described the role of
Her2/neu in breast cancer and ovarian
cancer.
Overexpression, amplification, rare
translocations
No ligand is known
Ras relays signals from the cell
surface receptors to the nucleus
Ras relays signals by acting as a switch
Prospect
A breakthrough for our understanding of the molecular
and genetic basis of cancer
Provided important knowledge concerning the regulation of
normal cell proliferation, differentiation, and programed cell
death.
The identification of oncogene abnormalities has provided
tools for the molecular diagnosis and monitoring of cancer.
Oncogenes represent potential targets for new types of
cancer therapies.
Tumor suppressor genes
Concept:
genes that sustain loss-of function mutations
in the development of cancer
TSGs
Transcriptional factor: p53, WT1,
Direct transcription regulator: Rb, APC
Inhibitor of cell cylcle kinase: p16INK4A, p19ARF,
Cell structural components: NF2
Phosphatase: PTEN
Potential mediator of mRNA processing: VHL
Components involved in DNA repair: MSH2,
MLH1, BRCA1, p53
TUMOR SUPPRESSOR GENES
Disorders in which gene is affected
Gene (locus)
Function
Familial
Sporadic
DCC (18q)
cell surface
interactions
unknown
colorectal
cancer
WT1 (11p)
transcription
Wilm’s tumor
lung cancer
Rb1 (13q)
transcription
retinoblastoma
small-cell lung
carcinoma
p53 (17p)
transcription
Li-Fraumeni
syndrome
breast, colon,
& lung cancer
BRCA1(17q)
transcriptional
breast cancer
breast/ovarian
tumors
BRCA2 (13q)
regulator/DNA repair
Mechanism for the inactivation of TSGs
1. Mutation: point mutation or frameshift mutation, p53
2. Deletion: LOH (loss of heterozygosity) or homozygous
deletion, Rb
3. Viral oncoprotein inactivation: p53, Rb
4. Promoter hypermethylation, histone modification
changes: p16
Rb function
Rb regulates G1/S transition
Rb inactivation by
viral oncoprotein
KNUDSON TWO HIT HYPOTHESIS IN SPORADIC CASES
Normal
Cells
RB
RB
RB
RB
RB
Mutation
RB
LOH
Tumor cells
Inactivation of a tumor
suppressor gene
requires two somatic
mutations.
29
P53
Function as gatekeeper
Inactivation of p53 in cancer
•LOH on 17p13 in a number of tumors
Bax
•Point mutation on exon 5-8 “hot-spot”
(Dominant negative mutation)
•MDM2 negative regulation
• viral-oncogene products inactivation
Invasion and Metastasis
Stepwise Malignant Progression of Cancer
The process of metastasis
consists of sequential linked steps
1.
2.
3.
4.
Growth at primary site and angiogenesis
Tumor cell invasion
Lymphatic and hematogenous metastasis
Growth at secondary site and angiogenesis
Mechanisms involved in tumor cell invasion
1.Loss of cell-to cell cohesive forces
2. Secretion of ECM-degrading enzymes
3. Active Locomotion
4. Tumor angiogenesis
5. Metastasis-related genes
5. Metastasis-enhancing Genes:
Oncogenes,CD44, Integrinβ1, CEA, MMP2, u-PA, etc
1. Loss of cell-to cell cohesive forces:
Cell adhesion molecules (CAMs）：
细胞粘附分子:介导细胞之间或细胞与ECM之间的选择性粘附。
• E-cadherin: Expression↓
Loss of cell-cell adhesion，Increased cell motility
• Integrins： Expression↓→↑
• Immunoglobin superfamily：NCAM, VCAM-1,CEA, DCC, etc
• Selectins
• CD44 variants
2. Secretion of ECM-degrading enzymes
• Matrix Metalloproteinases (MMPs)：～20
• Tissue inhibitors of metalloproteinases (TIMPs)： ～4
• Plasminogen Activators (PAs) :urokinase-type, tissue-type PA
• PA inhibitors (PAIs): ～3
Metastasis-associated
proteinases
Cell invasion of the extracellular matrix
The MMP family (at least 23 members)
(1) Classification
基质金属蛋白酶
间质胶原酶（Interstitial Collagenase ），如MMP-1、MMP-5、
MMP-8、 MMP-13等，作用底物主要为间质胶原 Ⅰ、Ⅱ、Ⅲ、Ⅶ、Ⅹ
型胶原，但不能降解明胶和Ⅳ型胶原。
明胶酶(Gelatinase )又称Ⅳ型胶原酶，如MMP-2、 MMP- 9，作用底
物主要是Ⅳ型胶原和明胶，还可以降解Ⅵ、Ⅶ、Ⅷ和Ⅹ型胶原，但
不能降解间质胶原。
基质溶解素（Stromelysin）,如MMP-3、 MMP-7、 MMP-10 和MMP-11
等，作用底物主要是基质中的蛋白多糖和糖蛋白，如纤维连接蛋白
（FN）、层黏连蛋白(LN)等。此外，基质溶解素对胶原的作用不同
于间质胶原酶间质和胶原酶，他们能降解Ⅳ、Ⅴ、Ⅷ、Ⅹ型胶原的
非螺旋区及Ⅰ型胶原的氨基末端。
膜型金属蛋白酶（Membrane-type MMPs, MT-MMPs ）,目前已发现四
种，包括MT1-MMP、MT2-MMP、MT3-MMP和MT4-MMP。MT-MMPs主要定位
于肿瘤细胞及其基质成纤维细胞的细胞膜上，是MMP的受体，也是
MMP的激活剂，还可以降解Ⅰ、Ⅱ、Ⅳ型胶原和FN，其表达受刀豆蛋
白、癌基因等因素的调解。
MMPs share the following common characteristics:
TIMP:
TIMPs play a key role in maintaining the balance between
ECM deposition and its degradation by binding tightly to and
regulating MMP actions
Four isoforms: TIMP 1-4
uPA¯uPAR-initiated signal transduction and consequences
Plasminogen/Plasmin System
3. Active Locomotion
• E- cadherin
• Growth factors and receptors,
• Autocrine motility factor (AMF),
• Autotaxin (ATX),
• Cytoskeletal proteins
• ECM components (laminin, LN, etc）
4. Tumor angiogenesis factors (TAFs)：angiogenin, etc
Inhibitors：angiostatin, etc
Models of Tumour Angiogenesis
Endogenous angiogenesis inhibitors
5. Metastasis-enhancing Genes:
Oncogenes, CD44, Integrinβ1, CEA, MMP2, u-PA,
etc
Metastasis Promoting Genes - I
Gene
Tissue Site
Function
ARM-1
Lymphoma
Promotes adhesion of tumor cells to the
endothelium
ATX
Breast, Liver, Lung, Melanoma,
Teratocarcinoma
cytoskeletal reorganization and motility; Gprotein coupled receptor activation
CD44
Multiple sites
cell-cell interactions; activates HGF/c-Met
pathway
Cox2
Breast, Colorectal, Gastric
Prostaglandin synthase; induces VEGF
Cyr61
Breast
Mediates adhesion; Erb-B2/3/4 pathway
Ezrin
Liver, Ovary, Pancreas, Prostate, Membrane-cytoskeletal linker; RHO and RAC
Uterus
interactions
HMG-I(Y)
Breast, Cervical, Colorectal,
Prostate, Skin, Thyroid, Uterus
Regulated by EGF and MMP-9
Laminin-5
Multiple sites
EGF and TGF-a induce expression of laminin
subunits; cell adhesion, motility
c-Met
Multiple sites
Activated by HGF; Modulates Ras and PI3
kinase
Metastasis Promoting Genes - II
Gene
Tissue Site
Function
MTA1
Breast, Cervix, Melanoma,
Ovary
Neucleosome remodeling; histone
deacetylase complex
Oncostatin M
Lung
Activates PKA-dependent pathway
PP2A
Not determined
Activated by p38/MAPK; inhibits MEK1,
MEK2, and MMP-1
RAGE
Gastric, Lung, Pancreatic,
Renal
transmembrane receptor; activates p21,
MAPKs, NF-6B, cdc42/rac
S100A4
Breast, Colorectal, Prostate
Calcium-binding protein; activates c-erbB-2
S100A9
Colon, Gastric, Skin
Calcium-binding protein; Modulates Mac-1
integrin receptor through G-protein
Semaphorins
Gastric, Leukemia, Lung, Skin
cell-cell interactions; Receptor crosstalk with
c-Met binding semaphorin receptor, plexin
Thymosin-b15
Prostate
actin binding; motility
Wnt-5a
Breast, Colon, Lung,
Melanoma, Pancreas, Prostate
PKC activation with associated changes in
cytoskeleton, cell adhesion, and motility
6. Metastasis-Suppressor Genes Identified
nm23, KAI 1, TIMPs, E-cadherin, Kiss, etc
Modified from JNCI 2000; 92:1717
The nm23 gene family
The first metastasis suppressor gene identified was nm23
Eight members of the human nm23 family have been reported and are
found in multiple subcellular compartments.
Biochemical functions
nm23 proteins posses multiple biochemical
functions
1. Interaction with numerous proteins
Tiam1, Ras, cytoskeletal protein
2. A NDPKinase activity
3. DNA nuclease
4. Serine or histidine protein kinase
inhibition of the Map kinase pathway and correlated
with motility suppression.
Nucleoside diphosphate kinase (NDPKinase) activity
The nm23-H1 gene product has been identified as the NDPKA isoform
The nm23-H2 gene product has been identified as the NDPKB isoform
NDPKs: catalyze the phosphorylation of nucleoside diphosphates to
the corresponding nucleoside triphosphates, mainly at the expense of
the ATP synthesized through oxidative phosphorylation
KAI1 / CD 82
Names : KAI1 / CD82, (C33, R2, IA4)
Gender : Transmembrane Glycoprotein
Ligands ? Signal Pathways : ?
Particularity
Biological Function :
motility
invasiveness
cell-cell interactions
Member of the tetraspanin or transmembrane 4 superfamily (TM4SF)
Contains an internalization sequence at its C-terminus (YSKV)
Current Address :
Cell membrane (lymphocytes, epithelial cells)
Lysosomes
Vesicles
KAI1 / CD 82 and Cancer
Correlations
High level of KAI1/CD82 is a good
prognosis factor or associated with
low grade histology :
prostate
pancreas
colon
lung
carcinoma
KAI1/CD82 expression is inversely
related to the metastatic potential :
Experimental Data
Transfection of tetraspanin reduces
metastatic potential
melanoma
prostate
breast
B16
MDA-MB-435 *
AT6.1, AT6.3
MDA-MB-231
(from Boucheix & Rubinstein , 2001
prostate
lung carcinoma
colon
hepatoma
breast
lung (non-small-cell carcinoma)
bladder cancer
ovary
melanoma
Loss of KAI-1 Expression in Prostate Cancer
KAI-1 Functions
+ (Promotes)
- (Inhibits)
Cell Aggregability
Cell Adhesion
Invasion
Motility
Metastasis
Metastasis Suppressor Pathways
C
D
H
1
K
A
I
1
b-Catenin
E
G
F
R
I
G
F
R
1
F
A
S
M
E
T
G
P
R
5
4
R
T
K
CRSP3
RAS
VAV
RhoGDI2
PI3K
Rho
Rac
RAF
KSR
PTEN
ASK1
NME1
MKK4
MKK7
KISS1
PLC
TXN
MEK
AKT
TXNIP
MKK6
ERK
MKK3
JNK
Apoptosis
Inflammation
Differentiation
Survival
NFkB
HIF
BAD
p38
Apoptosis
Cytokines
Survival
Angiogenesis
Motility
Invasion
Growth
Differentiation
Metastasis Facts
Up to 70% of patients with invasive cancer have overt or occult
metastases at diagnosis.
Acquisition of the invasive and metastatic phenotype is an early
event in cancer progression.
Millions of tumor cells are shed daily into the circulation.
Less than 0.01% of circulating tumor cells successfully initiate a
metastatic focus.
Angiogenesis is a ubiquitous and early event that is necessary
for and promotes metastatic dissemination.
Invasion and angiogenesis use the same signal transduction
programs and gene expression cassettes.
Circulating tumor cells can be detected in patients who do not
develop overt metastatic disease.
Metastases may be as susceptible to anti- cancer therapy as
their primary tumors?
Metastasis Therapeutic Targets and Agents
A. Targeted Therapeutics
Target
Example Agents
Effects
Growth factors C225 (anti-EGFR)
Block growth factor signaling
Tyrphostins (anti-RTK)
Cell adhesion Anti-avb3 (Vitaxin)
Blocks endothelial cell
avb3 peptidomimetics
interaction with matrix may
regulate MP activation
Proteolysis
MMPIs uPAR-I
Blocks degradation of matrix,
blocks activation of proteases,
growth factors
Motility
Taxanes
Blockade of microtubule
cycling
B. Signal Inhibitors: Blockade of signals necessary for angiogenesis ,
invasion, and metastasis
Agent
Target
CAI
Calcium influx
Squalamine
Inhibits NHE-3
PI3K inhibitors
MAPK inhibitors
Activity
Inhibits adhesion, motility, angiogenesis
Anti-angiogenic
Inhibit motility, proliferation, promote
Inhibit invasion, proliferation
Tumor
Microenvironment
Figure 4. The Cells of the Tumor Microenvironment.
(Upper) An assemblage of distinct cell types constitutes most solid tumors. Both the parenchyma and stroma of
tumors contain distinct cell types and subtypes that collectively enable tumor growth and progression. Notably, the
immune inflammatory cells present in tumors can include both tumor-promoting as well as tumor-killing
subclasses.
(Lower) The distinctive microenvironments of tumors. The multiple stromal cell types create a succession of tumor
microenvironments that change as tumors invade normal tissue and thereafter seed and colonize distant tissues.
The abundance, histologic organization, and phenotypic characteristics of the stromal cell types, as well as of the
extracellular matrix (hatched background), evolve during progression, thereby enabling primary, invasive, and then
Figure 5. Signaling Interactions in
the Tumor Microenvironment
during Malignant Progression
(Upper) The assembly and
collective contributions of the
assorted cell types constituting
the tumor microenvironment are
orchestrated and maintained by
reciprocal heterotypic signaling
interactions, of which only a few
are illustrated.
(Lower) The intracellular signalin
depicted in the upper panel withi
the tumor microenvironment is no
static but instead changes during
tumor progression as
a result of reciprocal signaling
interactions between cancer cells
of the parenchyma and stromal
cells that convey the increasingly
aggressive phenotypes that
underlie growth, invasion, and
metastatic dissemination.
Importantly, the predisposition to
spawn metastatic lesions can
begin early, being influenced by
the
differentiation program of the
normal cell-of-origin or by
initiating oncogenic lesions.
The diagnosis and therapy of cancer
Figure 6. Therapeutic Targeting of the Hallmarks of Cancer.
Drugs that interfere with each of the acquired capabilities necessary for tumor growth and progression have
been developed and are in clinical trials or in some cases approved for clinical use in treating certain forms
of human cancer. Additionally, the investigational drugs are being developed to target each of the
enabling characteristics and emerging hallmarks depicted in Figure 3, which also hold promise as cancer
therapeutics. The drugs listed are but illustrative examples; there is a deep pipeline of candidate drugs with
different molecular targets and modes of action in development for most of these hallmarks.
(Hanahan D, Weinberg RA. Hallmarks of Cancer: The Next Generation. Cell 2011, 144:646).
Three lines
of accomplishments are converging to
enable clinical researchers to investigate
therapies that target the molecular and
genetic abnormalities detected in an
individual
patient’s cancer: (i) We have identified
most of the few hundred genes that
are mutated or abnormally expressed in
human cancers. (ii) Genome sequencing
technology will soon enable us to screen
the mutations in a human cancer biopsy
in less than a week, at a cost of a few
thousand
dollars. Abnormalities in cellular
proteins and RNA can already be detected
in that time interval and at a reasonable
cost. (iii) Pharmaceutical and biotechnology
companies have more than 800 new
experimental drugs and biological agents
in the pipeline that are designed to target
the products of abnormal genes found in
human cancers