1. Tumor biology Lecture 12: Optional! Chapter 20 1205-1265
Cell biology 2014 (updated 18/2, 4/7 -13 & 1/1 -14)
Lecture 12:
Tumor biology
Cancer: latin word for crayfish
Will develop cancer
Will die of cancer
Cell Biology interactive  media  ”video” or ”interactive”

2. Basic tumor nomenclature
Benign tumor
Malignant tumor = cancer
Metastasis forming cell
(primary killer)
Carcinoma: derived from epithelial cells (90% of all cancers)
Sarcoma: derived from connective or muscle tissue
Leukemia: derived from hematopoietic cells (BM and blood)
Lymphoma: derived from lymphocytes (lymph nods)

3. Different views on cancer biology
Old fasion view
A heterotypic cell biology view
Non-autonomous
heterogeneous
cancer cells
Immune cells
Autonomous
cancer cells
Other cells
Endothelial cells
In vitro propagated cell lines can only rarely be established from tumor biopsies  tumor cells depends on their specific surrounding

4. Progenitors of the same clone, but still a heterogeneous tumor
Tumor progression
A malignant tumor does not arise from a single genetic change;
many changes are required to produce a life threatening cancer
Tumor progression is defined as the acquisition of permanent
changes in characteristics of selected subpopulations of the tumor
Progenitors of the same clone, but still a heterogeneous tumor

5. Onc TS Definitions: oncogenes and tumor suppressors
An oncogene is a gene that when mutated, or
overexpressed, contributes to converting a normal cell into
a tumor cell (constitutive activity dominant phenotype)
Bcl-2
Ras
point mutation
overexpression
A tumor suppressor-gene is a gene whose loss, or
inactivation, contributes to converting a normal cell into
a tumor cell (recessive phenotype) TS
CKI
p53 Rb
Inactivating point mutations or loss of the entire gene
(germ line mutation in one allele and/or acquired somatic mutations)

6. The normal stability of the genome makes
cancer development statistically improbable
Tumors acquire the capability to rapidly accumulate genetic changes by e.g., the following mechanisms:
1. Microsatellite INstability (MIN): Point mutations
Common causes: defective DNA mismatch repair genes
2. Chromosomal INstability (CIN): Aneuploidy
Common causes: aberrant centrosome numbers
defective spindle regulatory proteins
defective checkpoint control
3. Chromosome breaks and translocations
Common causes: eroded telomeres
DNA breaks

7. Chromosome separation,
Common cause of gene loss and amplification
DNA
strand
break
DNA
duplication
End
fusion
Chromosome separation,
novel breaks
Gene loss TS
DNA break
Telomere
Gene amplification
Onc

8. The six hallmarks of cancer – A cell biology perspective
1. Self-sufficiency in proliferative signals
2. Insensitivity to anti-growth signals
3. Evading cell death (apoptosis)
4. Limitless replicative potential
Make new
blood vessel!
5. Sustained angiogenesis
6. Metastasis capability
Adopted from Hanahan & Weinberg, Cell 2000

9. Tumor progression - molecular mechanisms
To be able to turn into a malignant tumor, each of the six
hallmarks has to be fulfilled
This is done by changing the level/activity of various proteins
Only one protein per pathway needs to be changed!
For example, a single protein in a mitogenic signaling pathway: G1 G1
myc
myc
Even if two tumors would belong to the same diagnostic group, they still have a unique combination of genetic alterations

10. Onc Onc Onc TS 1. Self-sufficiency in proliferative signals
Mitogenic signaling (growth promoting signals)
Cdk G1
Onc
Onc
The retinoblastoma pathway TS Rb
Production of
S cyclin
Cdk S
E2F
Initiation of
replication
DNA
replisome P
Cdc6
DNA
replisome
Mcm P
ORC
Production of DNA
replisome components

11. 1. Cell type specific mitogenic pathways
Cells from different tissues express distinct sets of
growth factor receptors and signaling proteins
Cell type B
Cell type C
Cell type A
Major mitogen
signaling pathway:
RTK
Wnt
Hedgehog
Alterations
in tumors: RTK signaling
Wnt signaling
Hedgehog signaling

12. Onc Onc TS Onc TS TS Onc TS Onc Onc Onc Onc Onc Onc Onc Onc
1. Aberrant proliferative signals in tumors
XGF
Wnt
Hedgehog
Onc
RTK
Onc
Frizzled TS
Patched
Ras
Onc
Smoothened
Dishevelled TS TS
GSK-3b
Axin
APC
Raf
Onc
Fused
Erk
SuFu TS
b-catenin
Onc
myc
Onc
myc G1
Onc
Gli
Gli
Onc
Onc
Onc
myc
Onc G1
Onc G1

13. TS TS Onc 2. Insensitivity to anti-growth signals Mitogen signaling
p15
TGF-b
Cdk G1
p21
p16 TS
The retinoblastoma pathway TS
HPV E7
viral
Onc Rb
Cdk S
E2F
All CDKI dependent growth arrest signaling pathways acts upstream of Rb. P
Cdc6
DNA
replisome P
ORC

14. Onc TS TS TS Onc 3. Evading cell death (apoptosis) Ligand p53 Death
Survival factor signaling
Ligand
Onc
p53 TS
Death
receptor
BH3
only TS
Adaptor
Cyt. C
Bcl-2
Onc
Bax TS
Caspase 8
Caspase 9
Caspase 3
Apoptosis

15. Onc Onc TS Onc 3. Survival factor signaling PI-3 K or GPCR or RTK P P
PTEN + G1
Onc TS
Onc
PKB/Akt
elF4E
Bad
Cell growth
Apoptosis

16. 4. Limitless replicative potential
Telomeres: stretches of repetitive DNA at the chromosome ends
that can form a protective loop structure
Chromosome lacking
telomeres will trigger
a p53 dependent cell
cycle block 5´ 5´ 3´ 3´
Complementarity due to the
repetitive sequence T A T G 5´
-GGGTTAGGGTTAGGGTTA G G
Telomerase, usually
not expressed in
somatic cells
AUCCCAAU 3´
-CCCAATCC
To maintain telomere length tumor cells can re-start expression
of telomerase. An alternative mechanism employs enzymes
that are involved in DNA recombination

17. 5. Sustained angiogenesis
Make new
blood vessel!
5. Sustained angiogenesis
Blood
vessel
< 100 mm
Endothelial cell
Diffusion of O2
and nutrients
Too long
I am suffocating!
Let’s express VEGF
VEGF: Vascular endothelial
growth factor
Anim angiogenesis

18. Onc TS 5. Vascular Endothelial Growth Factor - VEGF Ras 4.
Ras dependent signaling
can increase expression
levels of HIF-1
Onc 1.
Constitutively
produced
in all tissues
HIF-1
VEGF
VEGF gene 2.
Constitutively
degraded via
pVHL, unless…
pVHL 3. TS
HIF-1
The Von Hippel–Lindau tumor suppressor protein is encoded by the VHL gene and when inactivated is associated with Von Hippel–Lindau disease.
Von Hippel–Lindau syndrome (VHL) is a dominantly inherited hereditary cancer syndrome predisposing to a variety of malignant and benign tumors of the eye, brain, spinal cord, kidney, pancreas, and adrenal glands. A germline mutation of this gene is the basis of familial inheritance of VHL syndrome. Individuals with VHL syndrome inherit one mutation in the VHL protein that causes the protein's normal function to be lost or altered. Over time, sporadic mutation in the second copy of the VHL protein can lead to carcinomas.
…hypoxia
(low O2) Ub
HIF-1: Hypoxia induced factor
pVHL: von Hippel–Lindau syndrome (hereditary cancer) is caused by a germline mutation in the VHL gene
Proteosome

19. 5. Angiogenic factors affecting endothelial cells
Activators
Inhibitors
p53
VEGF
Thrombospondin-1
Loss of p loss of
angiogenisis inhibition
Tumor with active p53
Tumor without active p53
No angiogenesis
Angiogenesis

20. Onc TS TS 5. Summary: regulation of angiogenesis Ras p53 HIF-1 pVHL
Thrombo-
spondin-1
Avastinä
VEGF
Angiogenesis

21. 6. Metastasis capability
Metastasis, the ability of cancer cells to migrate, results from
multiple mutation events 1.
Basal
lamina nm 2. 3. 4. 1.
Loss of cell-cell adhesion 2.
Loss of hemidesmosomes 3.
Proteolytic degradation of the ECM 4.
Migration through the ECM

22. TS 6. Example of loss of cell-cell adhesion
Loss of E-cadherin is an important step in generating daughter tumors (metastasis) in carcinomas
Benign tumor
Malignant tumor
Tumor
progression TS
Migration, resettlement
and further proliferation
Loss of E-cadherin
decreased cell adhesion
Metastasis

23. 6. Penetration of basal lamina
Collagen IV fibril 1. 2. 3.
Laminin
Reprogramming / de-differentiation of cells: 1.
Loss of hemidesmosomes/laminin receptor (integrin) 2.
Expression of collagenase 3.
Cytoskeletal changes
 Epithelial–mesenchymal transition (EMT)

24. 6. Making it through the connective tissue
Cell secretes proteases to clear
a path through the ECM
Blood vessel

25. 6. Sites of metastasis – blood flow
Blood flow pattern determine the metastasis pattern in most
case (~70%)
Capillary of
the lung
Tumor cell entering
the blood system
Lung metastasis
Capillary of
the liver
Stomach or intestinal
tumor cell entering
the blood system
Liver metastasis

26. 6. Sites of metastasis – microenvironment
”Seed-soil” pattern determine the metastasis pattern in other
cases (~30%)
Capillary of
the lung
Prostate tumor
cell entering
the blood system
No lung metastasis
due to non-
favorable ”climate”
Adjacent bone cells
produce specific
factors needed for
tumor cell growth X X
Capillary of
a bone

27. Tumor progression: Familial adenomatous polyposis
Alberts et al. Fig 3. 2. X 5. X X X X 1. X 4. 6. 1.
Self-sufficiency in
proliferative signals 2.
Insensitivity to
Anti-growth signals 4.
Limitless replicative potential 5.
Sustained angiogenesis 3.
Evading cell death 6.
Metastasis capability

28. TS Step I. Starting point of familial adenomatous polyposis
By chance loss of the intact APC allele!
APC
APC TS
Wnt
GSK-3b
Axin
APC
b-catenin
MMP7 G1
Self-sufficiency in
proliferative signals
Chromosomal instability
Metastasis
capability
Note dual action of APC: Hallmarks 1 & 6

29. proliferative signals
Step II. Progression of colon carcinoma
Loss of SMAD4
Oncogenic mutation in RAS
XGF
TGF b -
Ras
Smad 4 TS
Onc
VEGF G1
p15
Self-sufficiency in
proliferative signals
Insensitivity to
anti-growth signals
Sustained
angiogenesis
Hallmarks 1, 2 & 5

30. proliferation signals
Step III. Progression of colon carcinoma
Loss of p53
DNA damage
p53
Aberrant/incomplete
proliferation signals TS
Thrombo-
spondin-1
p21
Bax
PUMA
Bcl-2
Insensitivity to
anti-growth signals
Sustained
angiogenesis
Evading cell death
Hallmarks 2, 3 & 5

31. Limitless replicative
Step IV. Progression of colon carcinoma
Expression of telomerase
Loss of E-CADHERIN
AUCCCAAU
Limitless replicative
potential TS
Metastasis
capability
The End
Hallmarks 4 & 6

32. Fulfilling the hallmarks of cancer in colon cancer
APC
Ras
1. Self-sufficiency in proliferative signals
p53
2. Insensitivity to anti-growth signals
Smad 4
p53
3. Evading cell death (apoptosis)
Telomerase
4. Limitless replicative potential
AUCCCAAU
Ras
5. Sustained angiogenesis
p53
APC
6. Metastasis capability
E cadherin
Fulfilling hallmarks 1 – 6 within a life
time depends on genomic instability

33. Breast cancer in Sweden
6,623 new cases in 2002 (early onset)
Incidence/year ~115 per 100,000
Mortality/year ~ 35 per 100,000

34. The TNM system for clinical staging
Tumor/ Node/ Metastasis:
T, clinical/mammographic evaluation of tumor (0-4).
N, clinical evaluation of regional lymph nodes (0-3).
M, distant metastases (0, 1).
Stage 0: Tis N M0
Stage I: T1 N M0
Stage 2: T1-3 N M0
Stage 3: T1-4 N M0
Stage 4: T1-4 N M1
(is: in situ  well encapsulated)

35. Stage and prognosis

36. Routine prognostic and predictive factors
TNM (Tumor/Node/Metastasis)
Histologic type and grade (as judge by the appearance under the microscope)
Molecular markers: Ki-67, estrogen and progesteron receptors, and ERBB2 (EGF receptor).

37. Decision tree: breast cancer treatment at NUS
T: clinical/mammographic evaluation
N: regional lymph nodes
M: distant metastases
Non-specific
Mix of cytostatic drugs, e.g., FEC
(5-FU, epirubicin, cyklofosfamid)
or SBG mix
Irradiation therapy
Specific
TAM: Tamoxifen (anti-estrogen)
A temporary cure!
(3-60 (?) years)

38. Future goals of (molecular) diagnostics
Early detection
Accurate prognosis
Good prediction (of therapy response)
Reveal molecular therapy targets