HALLMARKS OF CANCER

1. Hallmarks of
Cancer:
The Next
Generation
Members:
Tay Zheng Yi U2004973
Rithana Gunasegaran U2004885
Ikhmal Daniel 17205531/2
Ranesaa Nambiar U2004815
Lim Khang Ning U2004924

2. Sustaining
Proliferative
Signaling
i.

3. Fundamental Traits of Cancer Cells
● Cancer cells exhibit a
fundamental trait that involves
the deregulation of
growth-promoting signals,
allowing cancer cells to control
their own destiny.
● Cancer cells have the ability to
sustain chronic proliferation

4. Normal Tissue Regulation
● Balanced Growth Signals
○ Control the production and
release of growth-promoting
signals
● Growth Factors and Receptors
○ Growth factors→bind to
cell-surface receptors→emit
growth promoting signals
○ Receptors→ typically contain
intracellular tyrosine kinase
domains, initiating downstream
signaling

5. Normal Tissue Regulation
● Intracellular Signaling Pathways
○ Regulate cell cycle progression and growth
● Maintain Homeostasis
○ crucial for maintaining tissue architecture and function
○ Precise control of growth signals is essential for the overall
health and function of tissues

6. Deregulation in Cancer Cells
● Cancer cells exhibit a profound deregulation of
growth signals, a fundamental trait
● Growth Factor Deregulation
○ Cancer cells often hijack the binding of
growth receptors to cell-surface receptors,
become insensitive to normal regulatory
cues
● Receptor Hyperresponsiveness
○ Cancer cells have the ability to increase the
levels of receptor proteins on their cell
surface.
○ This elevation makes these receptors
hyperresponsive, meaning they become
more sensitive and responsive to external
signals

7. Deregulation in Cancer Cells
● Intracellular Signaling Pathways
○ Cancer cells may sustain proliferative signaling by constitutively
activating downstream pathways
Example: Ras Signaling Pathway
Activation of Ras pathway repeats the regulatory instructions
related to cell cycle progression, growth, and other cellular
functions

8. Autocrine
stimulation
Produce growth factor ligands themselves, to which they can respond
via the expression of cognate receptors
Stimulation of
Stroma
Cancer cells may send signals to stimulate normal cells within the
supporting tumor-associated stroma, which reciprocate by supplying
the cancer cells with various growth factors
Receptor
Deregulation
● Receptor signaling deregulated by elevating the levels of receptor
proteins displayed at the cancer cell surface
● Structural alterations in the receptor molecules that facilitate
ligand-independent signalling
Alternative Ways of Proliferative Signaling in
Cancer

9. Somatic Mutations Activate
Additional Downstream
Pathways
01

10. ● Mutation causes significant changes in an enzyme at the start of a signal
transduction pathway, it will disrupt the entire signal pathway.
● OR protein involved in downstream of a signal cascade.
Genetic Influences on Signalling
Somatic mutations in human tumors that predict
constitutive activation of signaling circuits usually
triggered by activated growth factor receptors.

11. 70% of human melanomas
contain activating mutations
affecting the structure of the
B-Raf protein, resulting in
constitutive signaling through
the Raf to mitogen activated
protein (MAP)-kinase pathway
Roberts & Der (2007).

12. Disruptions of
Negative-Feedback
Mechanisms that
Attenuate Proliferative
Signaling
02

13. ● Negative feedback loops normally operate to dampen various types of
signaling and thereby ensure homeostatic regulation of the flux of signals
coursing through the intracellular circuitry.
● Defects in these feedback mechanisms are capable of enhancing
proliferative signaling.
● Compromised negative-feedback loops in signaling pathways prove to be
widespread among human cancer cells and serve as an important means
by which these cells can achieve proliferative independence.
Feedback Loops and Proliferative Signaling
Importance of negative-feedback loops in signaling

14. PIP3
Growth Factor
Receptor Tyrosine Kinase
—---------------------------------------------------------------------------------------------------------------------
Akt
PIP2
● Cell Survival
● Apoptosis Inhibition
Example - PI3K-Akt pathway
PI3 kinase
PTEN
phosphatase

15. Drug Resistance and Signaling
Compromised/Disruption of negative-feedback loops may lead to adaptive
resistance towards drugs targeting mitogenic signalling.
Negative feedback
mechanisms limit extend of
pathway activation to prevent
excessive cell growth.
Overreaction of
Targeted Pathway
● Overactivation when
drug is introduced
● Cell can upregulate
components of the
pathway -cells less
sensitive to drug’s
inhibitory effects.
Negative feedback
mechanisms ensure alternative
pathway is suppressed when
primary mitogenic pathway is
active.
Compensatory
Pathway activation
● Cells activate
compensatory pathways
response to drug-induced
inhibition of primary
pathway.
● Able to proliferate even
the presence of drug
Mitogenic signalling pathway
can regulate cell survival
mechanisms.
Enhanced Survival
Mechanisms
● Activation of survival
pathways
● Cells resist
drug-induced cell death
● More resistant to
apoptosis even drug
inhibits cell growth.

16. Ras
Mitogenic Signal
Receptor
Transduction
Cascade
—------------------------------------------------------------------------------
Response
Raf
MAPK
SOS
PI3
Kinase
Cell growth Survival
Recalling…

17. Excessive Proliferative
Signaling Can Trigger
Cell Senescence
03

18. Counteracting Responses
Excessive Proliferative Signaling and Cell Senescence
Signal
Receptor
Oncoproteins
—------------------------------------------------------------------------------
RAF
Response
MAPK
MYC
RAS Early studies showed an image that
the increase expressed of
oncogenes and growth factor
(signals) would result in increased
cancer cell proliferation and tumor
growth
Cell growth
Cell growth
Cell growth
Cell growth
Cell growth

19. Counteracting Responses
Excessive Proliferative Signaling and Cell Senescence
Signal
Receptor
Oncoproteins
—------------------------------------------------------------------------------—-----------------------------------
RAF
Response
MAPK
MYC
RAS
Senescence/
Apoptosis
RAF
MAPK
MYC
RAS
Avoid
Senescence
Cells that
expressing high
levels of
Oncoproteins may
enter senescence
state
Cells expressing
low levels of this
protein may avoid
senescence and
proliferate.

20. Morphological features of Senescence:
Enlarged Cytoplasm
Expression of the senescence-induced
β-galactosidase enzyme
Absence of
proliferation markers

21. Counteracting Responses
Excessive Proliferative Signaling and Cell Senescence
Paradoxical
responses
Intrinsic cellular defense
mechanisms to
eliminate cells
experiencing excessive
signaling.
Alternative
Some cancer cells
adapt high levels of
oncogenic signaling by
disabling senescence-
or apoptosis-inducing
circuitry
Potential
compromises in
cancer cells
Excessive signalling -
maximal mitogenic
stimulation or avoidance of
antiproliferative defenses?

22. Evading Growth
Suppressors
ii.

23. Introduction to Growth Suppression
● Cancer cells must evade growth suppressors.
● Tumor suppressor genes play a crucial role in regulating cell
proliferation.
● The two prototypical tumor suppressors: RB and TP53.

24. Cellular proliferation/
growth
Cellular quiescence/
death
Stimulatory
factors
Inhibitory
factors
Perfectly balanced
(one-to-one balance)

25. Cellular proliferation/growth Cellular quiescence/death
Stimulatory
factors
Inhibitory
factors
Sustained
Proliferative
Signalling predominates
Hallmark of cancer

26. Cellular proliferation/growth Cellular quiescence/death
Stimulatory
factors
Inhibitory
factors
Sustained
Proliferative
Signalling
Evasion of
growth
suppressors

27. ● Tumor suppressors negatively regulate cell proliferation.
● Many tumor suppressors inactivate in animal or human
cancers.
● Validated as bona fide tumor suppressors through gain- or
loss-of-function experiments in mice
Role of Tumor Suppressors
"Bona fide tumor" refers to a genuine or authentic tumor. It
distinguishes actual cancerous growths from benign
growths or other conditions that may resemble tumors but
are not, in fact, cancer. Researchers and healthcare
professionals use the term "bona fide tumor" to emphasize
the true cancerous nature of a growth or lesion when
making diagnoses or conducting studies related to cancer
(Roberts et al., 2002).

28. ● Tumor suppressors negatively regulate cell proliferation.
● Many tumor suppressors inactivate in animal or human cancers.
Role of Tumor Suppressors
Cellular
proliferation/growth
Cellular
quiescence/death
Tumor
Suppressor
Proto-oncogenes

29. RB Protein as a tumor suppressor
The RB protein integrates signals from diverse sources.
Decides whether a cell should proceed through its
growth-and-division cycle (Burkhart and Sage, 2008; Deshpande et al., 2005; Sherr and
McCormick, 2002).
Missing RB function which are the critical gatekeeper of cell-cycle
progression allows persistent cell proliferation.

30. Recap: E2F role
recruits
Translates into proteins
for cell cycle progression

31. RB protein
Rb stops the DNA transcription
machinery from being able to function No transcription
occurs
Cyclin D binds to CDK4/6 forming
a complex that binds to Rb The complex
phosphorylates Rb
Rb freed from E2F Now, RNA
Polymerase able to
perform its function
of translating DNA

32. P53 protein senses and repairs the genotoxic stress
Oxidative
stress
Hypoxia
DNA
Damage
Nutrient
deprivation
Oncogenes
expression
Telomere
attrition
Halts the Cell Cycle
Begins a number of different mechanisms to
try and repair the genotoxic stress
Autophagy Senescence Apoptosis Migration DNA repair

33. Functional Redundancy in Tumor Suppression
● TP53 and RB are key suppressors of proliferation.
● Evidence indicates they function within a larger network with
functional redundancy.
● Example: Chimeric mice with RB-null cells.
● Contrary to expectations, RB null cells lead to relatively normal tissue
development.
● Only observed neoplasia late in life: pituitary tumors (Lipinski and Jacks, 1999).
● Implication: TP53 and RB are part of a complex regulatory network
that prevents uncontrolled cell proliferation.

34. Mechanisms of
Contact Inhibition and
Its Evasion
01

35. Mechanism of contact
inhibition
1) NF2 and Merlin : Gatekeepers
2) LKB1 : Tissue integrity maintainer

36. 1) NF2 and Merlin : Gatekeepers of Contact Inhibition
Contact Inhibition
Phenomenon that regulates the growth and division of cells in
response to physical environment & contact with neighbouring cells
NF2
(gene)
A tumor suppressor genes that
encodes the protein known as
“Merlin” or (or schwannomin)
Merlin
(protein)
A member of ERM (ezrin,
radixin, moexin) family of
membrane : cytoskeletal linker
protein
Loss of Merlin can trigger human
neurofibromatosis :
genetic disorder → tumours → nerve tissues
Neurofibromatosis Type 2 (NF2). Image retrieved from
https://www.youtube.com/watch?v=DbuwmzuYzS4
Benign
tumour

37. How Merlin orchestrates contact inhibition?
Cell-surface
adhesion
molecules
Cadherin
EGF
receptor
Transmembrane
receptor tyrosine
kinases
1. Merlin strengthens the adhesivity
of cadherin-mediated cell-to-cell
attachments
2. Merlin limits growth factor
receptors ability to efficiently
emit mitogenic signals by
sequestering them
(Carroll, 2011)

38. (Carroll, 2011)
Schematic demonstrate the cycling of Merlin between its active
(dephosphorylated) and inactive (phosphorylated) state
*Note that different intramembranous
receptors can inactivate or activate Merlin
Activating Merlin
Inactivating Merlin
Intramembranous receptor → Cadherins, CD44 → Myosin
Phosphatase Targeting Protein 1 (MYPT1) → Merlin (activated)
RTK complex
pKA
P21-activated
kinase (Pak)
cAMP
Rac1
P
X Merlin

39. 2) LKB1 : Maintaining Tissue Integrity
A tumor suppressor genes that
normally functioned as the
suppressor of inappropriate
proliferation
Organizes epithelial
structures
Maintain tissue
integrity
Loss of LKB1 :
Causes malignancies in human
Suppression :
● Epithelial cells become
susceptible to MyC-induced
transformation
● Epithelial integrity is
destabilized
Upregulation :
● Overrule the mitogenic
effects of MyC
oncogene in the
organized, quiescent
cells
Liver Kinase B1
(LKB1)
Role of LKB1 in overriding mitogenic effects of oncogenes

40. Suppression :
● Epithelial cells become susceptible
to MyC-induced transformation
● Continue on proliferating
● Epithelial integrity is destabilized
Upregulation :
● Overrule the mitogenic effects
of MyC oncogene in the
organized, quiescent cells
Liver Kinase B1
(LKB1)
The Myc/Max/Mad network is a critical regulatory
pathway that plays a fundamental role in controlling
the transcription of genes in various cell growth,
proliferation, differentiation, and apoptosis.
Cell Proliferation:
● c-Myc/Max complexes promote the
expression of genes that drive cell
proliferation.
● An increase in c-Myc activity can
lead to uncontrolled cell growth and
is often associated with cancer.

41. Corruption of the
TGF-β Pathway
Promotes Malignancy
02

42. TGF-β Pathway: Antiproliferative Effects
TGF-β (cytokine)
Antiproliferative effects
Cell Cycle Arrest:
● TGF-β can induce cell cycle
arrest (specifically at the G1 → S
phase)
● This arrest is achieved by
inhibiting (CDKs).
● Or promoting the expression of
cell cycle inhibitors like p15 and
p21.
Inhibition of Mitogenic Signaling:
TGF-β interferes with the signaling
pathways that promote cell proliferation :
● mitogen-activated protein kinase
(MAPK)
● phosphoinositide 3-kinase (PI3K)/Akt
pathways.
By doing so, it limits the responsiveness of
cells to proliferative signals.

43. TGF-β Pathway: Dual role in cancer
Redirection of TGF-β signaling in late tumors → EMT
Epithelial - to - mesenchymal transition
● EMT = cellular program where epithelial cells (normally tightly connected and organized)
transform into mesenchymal-like cells.
● This condition is achieved when the TGF-β is altered / corrupted and cancer cells manage
develop mechanisms to evade its tumor-suppressive effects.
● TGF-β is now redirected away from becomes tumor suppressor and acquires new
role to activate EMT
TGF-β in normal cells TGF-β in late-stage
tumor
- primarily acts as
tumor suppressor
- plays a role in
regulating cell growth
and homeostasis
- Activates EMT
(Di Gregorio et al., 2020)

44. Comparison in cell behavior (general)
Aspects of comparison Normal epithelial cells
(non-cancerous) under TGF-β
late-stage mesenchymal cells under
TGF-β
Cell shape ● Tightly packed, polygonal or cuboidal ● Spindle-shaped and elongated
Cell-cell adhesion ● Strongly adherent, forming tight
junctions and adherens junctions
● Lack cell junctions, reduced cell-cell
adhesion
Cell polarity ● Exhibit distinct apical-basal polarity ● Lack of apical-basal polarity (more
uniform)
Extracellular Matrix
Interaction (ECM)
● N/A ● Producing components of the ECM
(collagen or fibronectin) for invasive
properties
Motility ● N/A ● Motile and can change shape
(Di Gregorio et al., 2020)

45. Traits associated with high-grade malignancy conferred by EMT.
Trait Explanation
Increased Motility ● Enable them to more freely and invade surrounding
tissues and blood vessels.
Enhanced Extracellular Matrix Remodeling ● Mesenchymal-like cells can more effectively degrade
the extracellular matrix.
● Making it easier for them to invade nearby tissues and
reach the bloodstream for potential metastasis.
Stem Cell-Like Properties ● EMT can confer stem cell-like properties on cancer
cells.
● Acquires greater adaptability.
Heterogeneity ● EMT lead to increased in heterogeneity because cells
undergo transition within tumor.
● Different subpopulation required different treatment
options

46. SUPPLEMENTARY
SLIDES

47. References
1. Fouad, Y. A., & Aanei, C. (2017). Revisiting the hallmarks of cancer. American journal of cancer research, 7(5),
1016–1036.
2. https://blogs.scientificamerican.com/guest-blog/hallmarks-of-cancer-1-self-sufficiency-in-growth-signals/
3. Hanahan, D., & Weinberg, R. A. (2000). The hallmarks of cancer. Cell, 100(1), 57-70.
https://doi.org/10.1016/s0092-8674(00)81683-9
4. Marescal, O., & Cheeseman, I. M. (2020). Cellular Mechanisms and Regulation of Quiescence. Developmental
Cell, 55(3), 259-271. https://doi.org/https://doi.org/10.1016/j.devcel.2020.09.029
5. Sun, S., & Gresham, D. (2021). Cellular quiescence in budding yeast. Yeast, 38(1), 12-29.
https://doi.org/https://doi.org/10.1002/yea.3545
6. Weinberg, R. A. (1995). The retinoblastoma protein and cell cycle control. Cell, 81(3), 323-330.
https://doi.org/10.1016/0092-8674(95)90385-2
7. Carroll, S. L. (2012). Molecular mechanisms promoting the pathogenesis of Schwann cell neoplasms. Acta
Neuropathol, 123(3), 321-348. https://doi.org/10.1007/s00401-011-0928-6

48. TGFβ in Antigrowth Signaling
TGFβ (cytokine)
Preventing pRb phosphorylation
Blocking G1 phase progression
3 ways on how
TGFβ operates
1) Inhibition of pRb Phosphorylation
2) Disruption of pRb pathway in cancer
3) Effects on p15INK4B and p21
* a prerequisite for cell cycle
progression from G1 to S phase
* liberation of E2F → proliferation
*renders cell insensitive to
antigrowth factor
* inhibit CDK complex for pRb
phosphorylation

49. TGFβ in Antigrowth Signaling
3) Effects on p15INK4B and p21
TGFβ
p15INK4B
p21
CDK4
CDK6
CDK
kinases
or
induce
Cyclin-dependent
kinase inhibitor
+
pRb
phosphorylation

50. Evasion of Differentiation Signals
Normal tissue
● In normal tissues, antiproliferative signals maintain cellular
quiescence and tissue homeostasis.
● Cells have mechanisms to permanently enter postmitotic,
differentiated states as part of growth control.
Tumor tissue
● Dedicated to continue proliferating.
● Must find a way to avoid “Terminal differentiation”
● Terminal differentiation : process by which a cell becomes
highly specialized and attains its final mature form, typically
losing its ability to further divide.
Key mechanism :
Overexpression of c-Myc oncogene
Regulator / proto-oncogene
● Code for TF

51. The role of c-Myc overexpression in impairing
differentiation.
● During normal development, Myc's
growth-stimulating action, in
association with Max, can be
counterbalanced by alternative
complexes of Max with Mad
transcription factors.
● These Mad-Max complexes induce
differentiation-inducing signals.
● However, in many tumors, c-Myc is
overexpressed, disrupting this
balance and favoring Myc-Max
complexes. This shift impairs
differentiation and promotes
continued growth.

52. Inactivation of the APC/β-Catenin Pathway
Context : Colon cancer
Normal tissue
● In a normal state, pRb remains
hypophosphorylated.
● Cell proliferation is inhibited by
sequestering E2F transcription factors,
which control the expression of genes
needed for progression into the S phase.
Tumor tissue
● The pRb pathway can be disrupted.
● This disruption liberates E2Fs, allowing
cell proliferation and rendering cells
insensitive to antigrowth signals.
● Inactivation of the APC/β-catenin pathway can
block the egress of enterocytes in the colonic
crypts into a differentiated, postmitotic state.
● This means that colon cells lose the ability to
differentiate into their specialized,
non-dividing forms, contributing to
uncontrolled growth.
Importance :
Investigating and targeting these pathways may lead to
therapeutic strategies for preventing or treating cancer
by restoring normal growth control mechanisms.
● pRb = remains hypophosphorylated
● E2F = sequestered = no proliferation
Inactivation of pRb pathway can block differentiation

53. Summing Up:
● Cancer development involves evading antigrowth and differentiation signals.
● Normal tissues employ signals to maintain cellular quiescence, homeostasis, and
differentiation.
● Antigrowth signals prevent excessive proliferation through quiescence or
differentiation.
● Disruption of the pRb pathway liberates E2Fs, leading to uncontrolled cell growth.
● TGFβ prevents pRb phosphorylation, inhibiting G1 phase progression.
● Inactivation of differentiation pathways, like APC/B-catenin, blocks differentiation in
colon cells.
● c-Myc overexpression impairs differentiation, favoring cell growth.
● Understanding these mechanisms is critical in cancer development and may guide
therapies to restore normal growth control mechanisms.