hallmarks of cancer - university of south carolinaww2.biol.sc.edu/~bergerlab/guest lectures...
TRANSCRIPT
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Hallmarks of Cancer
Troy A. Baudino, Ph.D.Department Cell and Developmental Biology
University of South Carolina School of MedicineColumbia, South Carolina 29209
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What is Normal?Dependence on growth factors
–Cell and tissue specific signals –Loss of these signals leads to apoptosis
•Anchorage dependant proliferation–Requires interaction of transmembrane proteins (integrins) with components of the ECM
•Contact inhibition–Contact with other cells inhibits proliferation and movement
•Limited proliferative capacity–Normal somatic cells have a limited number of divisions before entering senescence
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Acquired Capabilities of Cancer
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• Originally coined as a genetic term to describe any gene capable of causing cancer
-Later came tumor suppressors – loss of functiongenes that can cause cancer
• Oncogenes refers to genes that contribute to cancer in a gain-of-function manner
• Proto-oncogenes are the normal genes
Hallmark #1: Self-Sufficiency in Growth Signals : Oncogenes
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Tumor-Suppressor Genes Vs Oncogenes
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Routes to the Genesis of an Oncogene
•Translocation•Over-expression
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Myc Regulates Proteins Involved in Cell Cycle Progression
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• Tumor suppressor genes are altered by inactivating mutations and this can lead to cancer
– Point mutations– Deletion of regions of chromosomes– LOH– Altered methylation of the promotor
Hallmark #2: Insensitivity to Negative Signals : Tumor Suppressor Genes
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•Retinoblastoma gene (Rb)– Germline mutations in Rb and one acquiredsomatic mutation
Classic Tumor Suppressor Genes
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The Rb Pathway and the Cell Cycle
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• p53 gene– 60-70% of all cancers have a p53 mutation
• Loss of both alleles or a dominant negative allele
Classic Tumor Suppressor Genes
An important consequence of p53 loss is a sharply elevated mutation rate, as well as genetic instability. Both are due to the inability to arrest the cell cycle in response to DNA damage
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p53 Mutation or Loss Plays a Dominant Role in Cancer
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• Apoptosis is programmed cell death-Apoptosis is programmed cell death, in which morphological nuclear changes and DNA fragmentation occur. The nuclear changes are nuclear blebbing to form micronuclei, and condensation of chromatin. Other features include loss of intercellular contact, vacuolation, and a relative conservation of cellular organelles.
-Necrosis, the other major type of cell death occurs as a result of cytoplasmic membrane damage, ion release and osmotic lysis with little nuclear or DNA damage until relatively late in the process.
• Damaged cells are effectively removed by this mechanism-This is a mechanism by which cells that have oncogenic mutations are removed-Is a critical defense against cancer
Hallmark #3: Evasion of Apoptosis
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Programmed Cell Death is an Essential Aspect of Animal Life
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•Death receptors transmit signals leading to apoptosis-FAS ligand and FAS receptor-TNF-α and TNF-αR1-Decoy receptors that don’t signal can promote survival
• Intracellular proteins that monitor DNA damage-p53
•Pro-survival factors-Bcl-2 family of proteins
Evasion of Apoptosis
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Bcl-XL
Bcl-2
Bax
0 6 12 244-HT (hrs)
wild-type
Myc Suppresses Bcl-XLExpression in Pre-B Cells
MycERHsp
+ 4-HT
MycERMax
CACGTG
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Myc Suppresses Bcl-2 or Bcl-XL Expression in a Cell Context-Specific Fashion
WT Eu WT Eu
Bcl-2
Bcl-XL
ß-Actin
Mature B Immature B
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ApoptosisApoptosis
c-Myc
Bcl-X Bcl-2
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• Cells have a finite lifespan and limited ability to replicate-Due to chromosome shortening-Ends of chromosomes are called telomeres (hexamerrepeats - TTAGGG)
• Hayflick limit : approximately 50-80 doublings- Cells reach replicative senescence
• Inactivating pRb or p53 extends lifespan 30 doublings
• Rare mutations lead to immortalization-Activation of telomerase
Hallmark #4: Acquisition of Limitless Proliferative Capacity
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•All tumors require a blood supply to grow to a significant size
• Pro-angiogenic factors such as VEGF, FGF1 and FGF2 are activated in tumors and signal endothelial cell proliferation and growth of blood vessels.
Hallmark #5: Sustained Angiogenesis
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Angiogenesis andVascular Endothelial Cells
Vascular endothelial cells
Blood vessel
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Angiogenesis
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Angiogenesis and Regulatory Proteins
Rare cell division
Blood vessel
Inhibitors low
Activators high
Inhibitors high
Activators low
Frequent cell division
Concentration of Angiogenesis Inhibitors
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Anti-Angiogenic Factors
Angiostatin HeparinasesCD59 complement fragment IL-12Endostatin Interferon-α,-β,-γFibronectin fragment TIMPsInterferon inducible protein-10Platlet factor 4 RetinoidsProlactin (16kDa fragment) TSP-1TGF-β Vasculostatin
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Pro-Angiogenic Factors
Angiogenin PlGFAngiopoietin-1 PDGF-BBDel-1 ProliferinaFGF TGF-αbFGF TGF-βFollistatin TNF-αGranulocyte-CSF HGFIL-8 VEGFIL-6
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Angiogenesis: A Delicate Balance
VEGFbFGF
IL-8Tsp-1Endostatin
Angiostatin Ang1
Angiogenesis
VEGFbFGF
IL-8
Tsp-1EndostatinAngiostatin
Ang1
Angiogenesis
X
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The Angiogenesis Signaling Cascade
Transcription
Cancer cell
VEGF (or bFGF)
Endothelial CellReceptor
Proteins stimulate new endothelial
cell growth
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Association between VEGF and angiogenesis, malignancy and metastasis has been demonstrated
•Many pro- and anti-angiogenic cellular factors regulate angiogenesis
•Research suggests that VEGF is a potent and predominant factor in this process
Angiogenesis and VEGF in Cancer
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Angiogenesis
Invasion
J Folkman 1974IJ Fidler 2002
Progression of Cancer
150-200µInitation
Promotion
Diffusion
Perfusion
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What Is Tumor Angiogenesis?
Blood vessel
Tumor that can grow and spreadSmall localized tumor
Signaling molecule
(i.e. VEGF)
Angiogenesis
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Angiogenesis and Cancer
AngiogenesisVessel dilation
New TheoryOld Theory
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•Abnormal architecture: tumor vasculature is highly disorganized -Vessels are tortuous and dilated with uneven diameter, excessive branching and shunts
•High vascular permeability: walls have numerous ‘openings’ (endothelial fenestrae, vesicles and transcellular holes), widened inter-endothelial junctions, and a discontinuous or absent BM
Structure and Function of Tumor Vessels
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McDonald & Choyke Nat Med 2003
Normal vs Tumor VasculatureDisorganized and Numerous
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Hallmark #6: Tissue Invasion and Metastasis
• Cell-Extracellular Matrix Interactions are altered– Changes in structural proteins (e.g. integrins)– Changes in enzymes (signaling enzymes, proteases, etc.)– Not as relevant to hematologic tumors
• Result: Increased Migration and invasion (critical steps in metastasis)
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Tissue Invasion and Metastasis
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c-Myc and its role in angiogenesis and tumorigenesis
E2F1 and its role in Myc-induced lymphomagenesis
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CANCER
Homeostasis
Proliferation
ProliferationDifferentiation
Differentiation
ApoptosisApoptosis
ApoptosisApoptosis
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Burkitt’s Lymphomas, other human lymphoid malignancies
Translocation
AmplificationAML, Lung, Breast, Colon, Brain, & Prostate cancers
Overexpression Breast, Ovarian, & Colon carcinomas
• 70% of all cancers have deregulated Myc
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ZipHLHb
c-Myc
TAD
NLS
MB1 MB21 439
L-Myc
N-Myc1 462ZipHLHb
1
TAD
Repression
ZipHLHb 364MB1
MB1 MB2
MB2
Structure and Function of Myc Family Members
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c-Myc Max
CACGTG
c-Myc Max
Relief ofRepression
Transactivation
Mad Max
CACGTG
XTransrepression
Mad
CACGTG
Transcriptional Regulation by Myc
Mnt
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Myc ARF Mdm2 p53
Apoptosis
The c-Myc-ARF-p53 Pathway
CellCycleArrest
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Both ARF and p53 Mediate Myc-InducedApoptosis in B-Cells
Time (hrs)
% V
iabi
lity
ARF-/-;p53-/-
24181260
20
40
60
80
100
WT
p53-/-ARF-/-
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The Eμ-myc Transgenic Mouse
PeripheralPre-B cells
2 Weeks
BM Pre-&B-cells
2 Months
Clonal B-cellLymphomaDEATH
3-5 Months
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Eμ-myc Lymphomas Express High Levelsof ODC & VEGF & are Highly Vascularized
Eμ-
myc
WT
WT
VEGF
β-actin
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The ARF-p53 Pathway is ConstitutivelyActive in Eμ-myc Bone Marrow
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100
Weeks
% S
urvi
val
The ARF-Mdm2-p53 Pathway Harnesses Myc-Induced Lymphomagenesis
20
40
60
80
5 15 25 35 45 55
ARF+/+ (n=31)24% ARF deletes28% mutant p53
ARF+/- (n=85)80% ARF deletesNo p53 mutants
ARF-/- (n=20)No p53 Mutants
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Myc ARF Mdm2 p53
Apoptosis
The c-Myc-ARF-p53 Pathway
CellCycleArrest
E2f1
Leone et al. 2001
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20
40
60
80
100
0 3 6 12 24
% V
iabi
lity
WTE2f1-/-
WT/Myc-ER
E2f1-/- /Myc-ER
Time (hrs)
Myc-ER
Myc-Induced Apoptosis is Independent of E2f1 in MEFs
MycERHsp
+ 4-HT MycERMax
CACGTG
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Myc Activates the ARF-p53 Pathway Independent of E2f1
E2f1
ARF
p53
p21
E2f1+/+ E2f1-/-
0 6 12 24 48+4-HT (hr) 0 6 12 24 48
Myc-ER
β-actin
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0
20
40
60
80
100
0 8 16 24 32 40 48
% S
urvi
val
Weeks
E2f1+/+ (n=22)
E2f1+/- (n=48)E2f1-/- (n=25)
E2f1 Loss Impairs Myc-Induced Lymphomagenesis
P<0.001
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Expression of p53, p19Arf and E2F1 in Eu-myc lymphomas
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The ARF-p53 Pathway Is FrequentlyInactivated in Myc-Induced Tumors
0
10
20
30
40
50
60
Wild type E2f1+/-;-/-
p53 mutation ARF deleteMdm2 Overexpression
Tum
or %
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4
6
8
10
12
% su
b-G
1
+/+ +/- -/-E2f1 +/+Eμ-myc
Myc-Induced Death is Independent of E2f1
Ex vivo
0
5
10
15
20
Apo
ptot
ic C
ells
/mm
3
+/+ +/- -/-E2f1 +/+Eμ-myc
25
In vivo
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Myc ARF Mdm2 p53
ApoptosisE2f1
The c-Myc-ARF-p53 Pathway
XCell Growth?
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E2f1 Loss Impairs Myc-Induced B cell Growth Ex vivo
8
12
14
16
18
% S
pha
se
+/- -/-E2f1 +/+Eμ-myc
10
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Spleen
0
4
8
12
+/+ +/- -/-
WTEμ-myc
Bone Marrow
0
10
20
30
40
50
+/+ +/- -/-
% S
-pha
se
E2f1
E2f1 Loss Decreases Myc-Induced Cell GrowthIn B Cells In Vivo
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•Loss of E2f1 impairs Myc-induced lymphomagenesis
•E2f1 loss is impairing Myc-induced cell growth, butnot Myc-induced apoptosis
Myc p27 Cell Growth
β-actin
+4-HT (hr)
p27Kip1
Myc-ER0 6 12 24
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p27 Loss Enhances Myc-Induced Lymphomagenesis
Martins and Berns, EMBOJ 21:3739-48 2002
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E2f1 loss Blocks Myc-MediatedSuppression of p27Kip1
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E2f1 is Required for Myc-Induced Downregulation of p27Kip1
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Myc-Mediated Proliferation and Lymphomagenesis, But Not Apoptosis Is Compromised by E2f1 Loss
Myc ARF Mdm2 p53
ApoptosisE2f1 p27 Cell Growth
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Acknowledgements
•Jennifer Old
•Chuck Sherr
•Martine Roussel