Cancer Lecturenotes 2014 Teacher

Property of National Junior College
H2 Biology Lecture Notes
Name:
Date:
CORE
SYLLABUS
CANCER
Your syllabus requires you to:

Cellular Functions
(A1n) Explain how uncontrolled cell division can result in cancer, and identify causative factors
(e.g. genetic, chemical carcinogens, radiation, loss of immunity) which can increase the
chances of cancerous growth. (Knowledge that dysregulation of checkpoints of cell
division can result in uncontrolled cell division and cancer is required, but detail of the
mechanism is not required.)
Organisation and Control of Prokaryotic and Eukaryotic Genome
(A4i)
Describe the functions of common proto-oncogenes and tumour suppressor genes

(limited to ras and p53) and explain how loss of function mutation and gain of function
mutation can contribute to cancer.
(A4j) Describe the development of cancer as a multi-step process.
June 2014
References:
1.
2.
3.
4.
5.
Biology by Campbell and Reece

Molecular Biology of the Cell by Alberts, Johnson, Lewis, Raff, Roberts and Walter
Biology by Raven, Johnson, Losos and Singer
Genetics: From Genes to Genomes by Hartwell, Hood, Goldberg, Reynolds, Silver and
Veres.
Molecular Cell Biology by Lodish, Berk, Zipursky, Matsudaira, Baltimore and Darnell
Table of Contents
1. Introduction: How uncontrolled cell division results in cancer .........................................3
1.1
2.
Characteristics of cancer cells ................................................................................................................ 3
The cell cycle ........................................................................................................................................4

2.1
2.2
2.3
Phases of the cell cycle and how it is being regulated: .................................................................. 5

Regulation of the cell cycle ...................................................................................................................... 6
The cyclin control system ........................................................................................................................ 8
3. Characteristics of Cancer Cell ....................................................................................................... 11

4.
Mutations of certain genes which can cause Cancer .......................................................... 12

4.2
4.3
4.4
4.5
4.6
4.7
Gain of function mutations in proto-oncogenes ...........................................................................14

How do gain of function mutation occur? (3 possible ways) ...................................................16
An example of a proto-oncogene: RAS gene ....................................................................................18
Loss of function mutations in tumour suppressor genes, P53 .................................................20
How do loss of function mutation occurs? (3 possible ways) ..................................................21
An example of tumour suppressor gene: p53 gene .....................................................................22
5.
Development of Cancer is a Multi-Step Process: .................................................................. 23
6.
Causative factors and cancer ...................................................................................................... 26
June 2014
1. Introduction: How uncontrolled cell division results in cancer
Cancer can occur in almost any tissue, and happens when a cell divides more frequently
than normal cell type it descended from.
It starts when a cell escapes the mechanisms that normally regulate the cell cycle and
begins to divide in an uncontrolled and invasive way.
The result is a mass of cancer cells, called a primary tumour, which constantly expands in
size.Tumours can be differentiated into two types, (1) benign, and (2)malignant.
A benign tumour consists of slow growing mass of cells and usually will not spread.
A malignant tumour, on the other hand, consists of fast growing mass of cells that may be
carried to other parts of the body via the blood or lymphatic system (FYI: It is another
system in our body that helps to remove interstitial fluid, transport white blood cells,
removes and transport fatty acids etc) and give rise to secondary tumours at distant
sites. When this happens, we say metastasis (spread of cancer cells) has occurred.
Shown below, on the left, is a primary tumour developing from epithelial cells that line the
interior surface of a human lung. As the primary tumour expands in size, it invades
surrounding tissues, eventually penetrating lymphatic and blood vessels, which carry
metastatic cancer cells throughout the body, where they lodge and grow, forming
secondary tumours.
1.1
Characteristics of cancer cells
Cancer cells never stop dividing. They are virtually immortal until the body in which
they reside dies.
Disorganized behavior and independence from surrounding normal tissues.
Permanent loss of cell differentiation.
Expression of special markers on their surface.
Over the last few decades, much progress has been achieved in cancer research using
molecular biological techniques.
We now know that cancer is a genetic disorder of somatic tissue , often involving
mutations of genes encoding proteins that regulate the cell cycle.
June 2014
2.
The cell cycle
As cancer is essentially a disease of cell division, the knowledge of the cell cycle and how it is
being regulated is crucial in helping us understand the development of cancer. Lets study how
the cell cycle is regulated in normal cells first.
The Cell cycle can be divided into three main stages:
Interphase (consisting of G1, S, G2 phases);
Mitosis Phase (M phase) when nuclear division occurs and
Cytokinesis phase where cytoplasmic cleavage occurs.
Different cells have different duration in a cell cycle (NB: cell cycle refers to the time taken for a
cell to complete one cell division.)
The events in a cell cycle are controlled by the interaction of 2 groups of proteins. Cyclins and
Cdk (Cyclin dependent kinases). Cells also receive protein signals (growth factors) that
affect cell division.
Cell cycle is regulated by various checkpoints. A
cell cycle checkpoint is a critical control point
where the stop and go-ahead signals can regulate
the cell cycle.
These checkpoints ensure that the cell is

only allowed to proceed to the next phase
only if it has properly completed the previous
phase.
These checkpoints monitor DNA

replication, DNA damage and
Chromosome-to-spindle tubule
attachments.
There are three major Checkpoints in the

cell cycle, and are found in G1, G2, and M
phase.
June 2014
2.1
Phases of the cell cycle and how it is being regulated

Each cell cycle consists of 5 phases G1, S, G2,
M and C. Of which, G1, S, and G2 are often
collectively referred to as the interphase of the
cell cycle.
Raven and Johnson (1999)
Events Within Cell During Cell division

(Recall from cell division)
G1
S
synthesis
G1 is a primary growth phase of the cell cycle. For many organisms, this
encompasses the major portion of the cells life span.
S is the phase in which the cell synthesizes a replica of its genome

(i.e. undergoes DNA replication).
G2
G2 is the second growth phase of the cell cycle, in which preparations are made for
genomic separation. It is also during this phase that mitochondria and other organelles
replicate.
M
Mitosis
M is the phase in which nuclear division occurs. It is traditionally subdivided into 4

stages prophase, metaphase, anaphase, and telophase, although mitosis is a
continuous process.
C
cytokinesis
C is the phase of the cell cycle when the cytoplasm divides, where there is equal
distribution of organelles and cytoplasm creating two daughter cells
Different organisms or tissues take different lengths of time to complete 1 cell cycle. Most of
the variation in the length of the cell cycle lies in the G1 phase.
Cells often pause in G1 before DNA replication and enter a resting state called GO phase.
They may remain in GO phase for days to years before resuming cell division.
At any given time, most of the cells in an animals body are in GO phase
(i.e. quiescent). Some, such as muscle cells and nerve cells, remain there permanently (i.e.
do not divide at all in a mature human). Others, such as liver cells, can resume G1 phase in
response to factors released during injury.
Typically, a rapidly dividing mammalian cell completes its cell cycle in about
24 hours. During the cycle, growth occurs throughout the G1 and G2 phases (sometimes
referred to as gap phases, as they separate S from M), as well as during the S phase. The M
phase takes only about 1 hour.
June 2014
2.2
Regulation of the cell cycle

Key checkpoints (as shown in the diagram on the previous page) control the cell cycle in
eukaryotes:
the G1 checkpoint (Are mitogens present? Is the cell size big enough?)
the G2 checkpoint (Have all the genes been replicated?)
the M checkpoint (Are all the chromosomes aligned properly at metaphase?)
(NB: Mitogens are chemical susbtances that encourages a cell to commence cell division, triggering mitosis.)
A set of proteins sensitive to the condition of the cell interact at the checkpoints to trigger the
next events in the cell cycle. Two key types of proteins participate in this interaction:
Cyclin dependent kinases (Cdks)
Cyclins (proteins that get their name from their cyclically fluctuating concentrations in the
cell).
Note that there may be different types of cyclins responsible for regulation of each of the
cell cycle checkpoints (e.g. see table on next page. CyclinD and CyclinE are involved in
regulating the G1 checkpoint. However, CyclinB is responsible for regulating the G2
checkpoint.)
However, the same group of Cdks are involved in all different cell cycle checkpoints.
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Stage
G1 / S
Phase
G2 / M
Phase
M
Check
Point /
Spindle
Check
Point
Description
1. For many cells, the G1 checkpoint
seems to be the most important.
2. G1 is a primary point where the cell
decides to divide or not.
3. G1 ensures that cell is sufficiently
large (minimum cell size is reached)
to divide, and has sufficient nutrients
to support its resulting daughter cells.
4. If a cell receives a go-ahead signal, it
will usually continue and complete the
cell cycle.
5. If cell does not receive the go-ahead
signal, it will exit the cell cycle and
switch to a non-dividing state called
G0.
Factors affecting / Regulation
External factors such as

growth factors and internal
factors such as nutritional
state e.g. starvation or lack
of growth factors can stop
the cell cycle at this stage.
Genome must be intact.
Any damage to DNA can
halt the cell cycle at this
point.
Types of
cyclin
involved
(FYI)
CyclinD
CyclinE
(Also
called G1
cyclins)
NB: Most cells in human body

are in the G0 phase.
1. This
checkpoint
assesses
the
success of DNA replication and
ensures that DNA replication in S
phase is completed before DNA
segregation in M phase begins.
2. Entry into M Phase (mitosis) is
prevented unless all genes have
been replicated.
3. Such quality control is necessary to
safeguard the integrity of the
genome. Genome must not be
damaged.
MPF = M Phase
Promoting Factor must be
present
Cyclin B
(Also
called
mitotic
cyclin)
All chromosomes must be

present and the tensions
between opposite poles are
important.
Cyclin B
1. Just like the G2 checkpoint, the M

checkpoint serves to safeguard the
integrity of the genome.
2. Ensures that all chromosomes are
attached to the spindle tubules in
preparation for anaphase.
3. In the event of defective spindle
assembly
(e.g.
presence
of
unattached kinetochores), the cell
cycle will be arrested at metaphase.
4. Transit into anaphase is prevented
until all chromosomes are aligned
properly at metaphase.
5. The
possibility
of
aneuploidy
(abnormal number of chromosomes
in daughter cells) occurring is thus
ruled out.
June 2014
2.3
The cyclin control system
Cdks (cyclin-dependent kinases) are enzymes (protein kinases) that activate or

inactivate their substrate proteins by phosphorylating (adding phosphate groups to)
them. (Cdks phosphorylate other proteins using inorganic phosphate groups that are not
yet bound to any other molecules)
In a growing cell, Cdks are present at a constant concentration, but most of the time,
they are in an inactive form.
For a Cdk-cyclin complex to be active, it must be:

(1) attached to a cyclin,
(2) itself not phosphorylated, and
(3) not bound to its inhibitor.
There are several types of cyclins, each type appearing and accumulating on cue at
each phase of the cell cycle, associating with the Cdks to carry out the phosphorylation
reactions, then disappearing (broken down) to make way for the succeeding set of
cyclins (of a different type).
The cycle of precisely timed cyclin appearances, accumulations and disappearances is
the result of two mechanisms:
1. Gene regulation that turns on and off the synthesis of particular cyclins
2. Regulated protein degradation that removes the cyclins
The cyclin portion of a Cdk-cyclin complex determines the target proteins that the
complex will phosphorylate; the Cdk portion of the complex performs the actual
phosphorylation
June 2014
The activity of a Cdk rises and falls with changes in the concentration of its cyclin partner.
Here MPF (the first cyclin-Cdk complex that was discovered) shall be used as an
example:
MPF (maturation-promoting factor / M-phase promoting factor) is the Cdk-cyclin

complex formed by Cdk and Mitotic cyclin which regulates the passage of the cell
through the G2 checkpoint as well as the M checkpoint (at metapase).
Campbell and Reece (2008)
Campbell and Reece (2008)
June 2014
As the cell cycle passes through the G1 and G2 checkpoints, Cdk becomes associated with
different cyclins and, as a result, activates different cellular processes. At the completion of
each phase, the cyclins are degraded, bringing Cdk activity to a halt until the next set of
cyclins appears.
June 2014
10
3. Characteristics of cancer cell

1. Loss of cell cycle control: Cell division is a carefully controlled process. Whether a cell
divides or stops dividing or differentiates depends upon signals from the surrounding cells. A
cancer cell simply ceases to respond correctly to these signals and divide uncontrollable and
more frequent than the cells which they arose.
2. Loss of cell death: Normal cells die when they do not receive any growth factors, or when
exposed to agents such as toxins or X-rays that damage them. The normal cells undergo
programmed cell death, thus preventing such cells from proliferating when they should not and
eliminates badly damaged cells. Most cancer cells are much more resistant than normal cells
to programmed cell death.
3. Immortality due to activated Telomerase gene activity: A cancer cell line divides indefinitely,
and has limitless potential. Most cells have a limit of 60-80 division before they cease to divide
again. The length of repetitive telomeres sequences at the end of human chromosomes
determines the number of times a cell can divide. Most normal cells (except for rare stem cells)
stop dividing and die spontaneously after a certain number of cell divisions. Each division
shortens the telomeres sequence. Cancer cell have activated telomerase gene, which
produces telomerases that lengthen the telomeres, allowing the cells to keep dividing
indefinitely! Most normal human somatic cells do not express the enzyme telomerase and this
lack of telomerase expression prevents them from replicating the repeated sequences in the
telomeres at the ends of their chromosomes. As a result, after a certain number of cell
divisions, telomeres shorten to the point where they contribute to cell senescence and death.
4. Lack contact inhibition: Cancer cells do not stop growing when they crowd around other cells
and are insensitive to growth-inhibitory signals. Normal cells placed in a container divide to
form a single layer. Cancer cells will pile up on one another. This piling up of cells leads to
formation of a tumour.
5. Less differentiated and less adherent: Cancer cell is less specialized than normal cell types
around it. It is also rounder than normal cells because it does not adhere to surrounding
normal cells as strongly as other normal cells would do.
6. Ability to induce local blood vessel formation (angiogenesis): Once the adult human body
has developed, new blood vessels do not normally form except to heal a wound. However for
tumour cells, once the mass of cancer (tumour) reaches a certain size, the interior cancer cells
respond to the oxygen poor environment by secreting a protein that stimulates nearby
capillaries to extend towards the tumour. The new vessels serve as supply lines through
which the tumour cells metastsize.
7. Ability to metastasize: Normal cells stay within defined boundaries. Cancer cells have the
ability to migrate and move to new parts of the patients body.
8. Exhibit Genomic Instability: Due to faster rate of cell division and a breakdown in cell cycle
check points. Normal cells have elaborate system to repair DNA damage. These systems that
repair DNA damages do not function correctly in cancer cells. Thus mutations accumulate
faster in cancer cells than normal cells.
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11
4.
Mutations of certain genes which can cause cancer
Recall that cancer is a disease of unrestrained cell proliferation caused by damage in

genes regulating the cell division cycle.
The cell cycle is regulated by a sophisticated group of proteins, which include growth
factors, their receptors, and the intracellular molecules of signalling pathways.
An organism/person can develop cancer should mutations occur in the genes

encoding these proteins.
There are two types of mutations that lead to cancer:

1. Gain of function mutations in proto-oncogenes
2. Loss of function mutations in tumour supressor genes
(NB: A third category includes mutations in DNA repair genes that allow mutations to persist unfixed and to
accumulate.)
When such mutations inactivates tumour-suppressor genes or active oncogenes

(the mutated form of proto-oncogenes), cancer results.
More than 100 oncogenes have been discovered, which cause cancer when they
are inappropriately activated and more than 30 tumour supressor genes whose
deletion or inactivation causes cancer.
These cancer-causing mutations can be random and spontaneous mutations,
caused by environmental influences e.g. chemical carcinogens, X-rays, and
certain viruses.
June 2014
12
The mutations can come about as a result of changes in nucleotide sequence

(gene mutations). Sometimes even due to single base substitutions, or as a result
of large scale changes in chromosomes (chromosomal mutations). The diagram
below shows how chromosomal mutations can occur.
June 2014
13
4.1
Gain of function mutations in proto-oncogenes
Proto-oncogenes are genes whose normal products (see diagram below for
examples) stimulate cell division.
In normal cells, proto-oncogenes code for proteins that send a signal to nucleus to
stimulate cell division.
The proteins they encode are under cellular control as they can be switched on or off to
serve the needs of the body. [Common proteins from proto-oncogene are circled below]
The conversion of proto-oncogenes to oncogenes is a Gain-of-function mutation

because the cells with the mutant form of the protein have gained a new function not
present in cells with the normal gene.
The conversion of proto-oncogenes to oncogenes is also deemed a dominant mutation.

Most oncogenes arises from dominant mutations; a single copy of the oncogene is
sufficient for the over expression of the growth trait. (Recall definition of dominant allele)
Raven and
Johnson (1999)
Mutations that make these proteins hyperactive and uncontrolled will cause the cells
that contain them to proliferate excessively.
Such gain-of-function mutations in the proto-oncogenes will make the latter become
cancer-causing genes called oncogenes. (Greek onco-, tumour)
In general, an oncogene arises from a genetic change that leads to an increase either in
the amount of the proto-oncogenes protein product or in the intrinsic activity of
each protein molecule i.e. how long it can stay functional in a cells cytoplasm.
June 2014
14
Table of Proto-oncogene and Oncogenes:

Types of Classification of
Proto-oncogenes for Genes
coding
(a)
Growth factors
(Normal cells growing in culture
will not divide unless they are
stimulated by one or more
growth factors present in the
culture.)
(b)
Growth factor receptor

proteins (are typically
transmembrane proteins, which
growth factors bind to. E.g.
tyrosine kinase)
Examples
Effect of Mutation
(i.e. Becomes Oncogenes)
Platelet derived growth factor

(PDGF) - a protein produced by
blood platelets that stimulates the
proliferation of connective tissue
cells, e.g. at the site of injury, for
wound healing
Gliomas are brain tumors that

have the characteristics of glial
cells. Glial cells are specialized
cells whose normal job is to
maintain the function and
interactions of neurons. In one
form of this cancer, there is an
over expression of the growth
factor and the receptor in
undifferentiated cells.
E.g. erbB - Receptor for

epidermal growth factor.
In the mutated form, the

receptor tyrosine kinase can
dimerise without the presence of
the growth factor i.e. it is
constitutively active
AbI tyrosine kinase (ABL gene)

This enzyme functions in the
nucleus as part of the normal
signaling pathway that causes
cells with damaged DNA to self
destruct by apoptosis.
In chronic myelogenous
leukemia (CML), mutation
results in the formation of an
abnormal version of Abl tyrosine
kinase that stays in the
cytoplasm and therefore cannot
trigger apoptosis.
(To be learnt in greater detail in

SH2)
(c)
Intracellular protein kinases

(These are enzymes that add
phosphate groups to target
proteins, thereby modifying
their functions.)
Note that in this case, the

mutation enhances the survival
of these cells rather than their
proliferation.
(d)**
G Protein
Ras protein (By ras gene) This

protein is associated with the
inner surface of the plasma
membrane. GTP and GDP
regulate its activity.
The mutated gene produced a

protein that retains the GTP and
thereby maintaining the protein
in an active stage. It will thus
continue to send stimulatory
signal to the rest of the signal
transduction pathway.
(e)
Transcription factors
Myc transcription factors (by

MYC gene)
Increased expression of the

MYC gene will lead to
overproduction of the Myc
protein, resulting in cell
proliferation.
Myc protein stimulates the

transcription of genes required for
cell proliferation.
* Required for H2 syllabus
June 2014
15
4.1.1 How do gain of function mutation occur? (3 possible ways)

1. Movement of DNA within the genome due to Translocation
If a translocated proto-oncogene ends up near an especially active promoter

(Refers to a promoter which will bind with greater affinity to RNA Polymerase,
leading to a higher transcription efficiency), its transcription may increase,
making it an oncogene.
Due to DNA translocation, a more active promoter might be placed near a protooncogene, which will lead to a higher gene expression of the Proto-oncogene.
2. Gene Amplification of a proto-oncogene
Errors in DNA replication may produce extra gene copies, which can lead to
overproduction of the protein. Amplification of the proto-oncogene has been
associated with the onset of cancer.
The amount of the proto-oncogenes protein product increases when the number of
copies of a proto-oncogene in the cell increases.
Process of Gene Amplification: This process involves the selective replication of a

region of a chromosome. The process can be repeated many times, making
many copies of that particular region.
The genes within the amplified portion of the chromosome can each be transcribed
to produce the normal protein. Hence resulting in the production of a large amount
of the proteins.
While this process is not seen in normal cells, it occurs quite often in cancer
cells. If an oncogene is included in the amplified region, then the resulting
over expression of that gene can lead to deregulated cell growth.
3. Point mutations within a control element or the proto-oncogene itself
June 2014
Point mutations in the promoter or the enhancer that controls a proto-oncogene

may result in an increased expression of the proto-oncogene.
Point mutations in the proto-oncogene itself may give rise to a protein product
that is more active or more resistant to degradation than the normal protein.
E.g. RAS Oncogene (See Next Page for detailed function of RAS Oncogene)
16
ENRICHMENT
Gene Amplification Example 1: Example of this includes the amplification of the myc oncogene
in a wide range of tumors and the amplification of the ErbB-2 or HER-2/neu oncogene in breast
and ovarian cancer. (HER = human epidermal growth receptor)
Gene Amplification Example 2: Gene amplification and Drug resistance'

Gene amplification also contributes to one of the biggest problems in cancer treatment:
drug resistance. Drug resistant tumours can continue to grow and spread even in the presence
of chemotherapy drugs. A gene commonly involved is called MDR for multiple drug resistance.
The protein product of this gene acts as a pump located in the membrane of cells. It is capable
of selectively ejecting molecules from the cell, including chemotherapy drugs. This removal
renders the drugs ineffective.
June 2014
17
4.1.2 An example of a proto-oncogene: RAS gene
Ras gene codes for a signal protein called RAS protein
The normal function of the Ras protein is to participate in the cell signalling process in
response to growth factors (signal molecules) in the normal stimulation of the cell cycle
The cell signaling process that Ras protein participates in (ref. to steps in diagram below):
o
(1) A growth factor binds to (2) its receptor in the plasma membrane.
The signal is relayed to (3) a G protein called Ras. Like all G proteins, Ras is
active when GTP is bound to it.
Active Ras passes the signal to (4) a series of protein kinases. [Kinases are
enzymes which add a phosphate group to another molecule]
(5) The last kinase activates a transcription factor that turns on one or more
genes encoding proteins that stimulate the cell cycle.
NB: If a mutation makes Ras protein or any other component in this pathway abnormally
active, or increases the amount of that protein, excessive cell division and cancer may
result.
Result of gain of function mutation in Ras

gene.
June 2014
18
[Diagram showing detailed mechanism of activating Ras protein.]
The ras proto-oncogene is located on human chromosome 11. (FYI)
Ras proto-oncogene codes for a G Protein, which is involved in kinase signaling

pathways that control the transcription of genes, which then regulate cell growth and
differentiation.
Under normal conditions, the signaling pathway will not operate unless triggered by
appropriate growth factor binding to its receptor (shown as signal molecule above). This
binding triggers a GTP molecule to binds to the ras protein, making it 'active'.
Active ras protein then passes on the signal to a series of cytoplasmic kinases, which in
turn activate transcription factors that turn on genes for proteins that stimulate the
cell cycle. To turn the pathway "off", the ras protein has an intrinsic GTPase function that
serves to hydrolyze the GTP molecule after some time (to form GDP), causing the Ras
protein to revert back to the inactive state.
A point mutation in the RAS gene resulting in a substitution from guanine to thymine is
frequently associated with cancer. This simple change results in glycine at amino acid
number 12 being substituted with a valine. This dramatically changes the function of the
G-protein encoded by the ras gene.
The mutation does not allow the release of GTP, and the ras oncoprotein is
continuously active. Because the signal delivered by the ras oncoprotein is
continuously delivered, this causes the signaling pathway to remain in the "on"
position, leading to uncontrolled and excessive cell growth and prolife ration,
which is a major step in tumour formation.
June 2014
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4.2
Loss of function mutations in tumour suppressor genes, P53
Tumour suppressor genes are genes whose normal protein products inhibit cell
division when necessary to prevent uncontrolled cell proliferation.
Tumour suppressor proteins, (e.g. p53 proteins) are normally involved in the activation
of other genes that synthesize proteins involved in repair of damaged DNA, a function that
prevents the cell from accumulating cancer-causing mutations. Tumour suppressor
proteins may also trigger cell death if DNA damage is beyond repair. [see diagram below]
Proteins coded by tumour suppressor genes normally function as cell's brakes that restraint
cell growth and prevent tumour formation.
Mutations in these genes result in cells that do not show normal inhibition of cell growth
and division.
Raven and Johnson (1999)
Other tumour suppressor proteins control the adhesion of cells to each other or to the
extracellular matrix. Their actions ensure proper cell anchorage, which is crucial in normal
tissues.
Any loss of function mutation that leads to a decrease either in the amount of the
tumour suppressors protein product or in the normal activity of a tumour suppressor
protein may contribute to the onset of cancer due to the absence of suppression.
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4.2.1 How do loss of function mutation occurs? (3 possible ways)

1. A point mutation in the gene that leads to a truncated (due to nonsense mutation) or
misfolded protein that is no longer able to function normally,
2. A chromosomal mutation resulting in the deletion of the normal functioning gene
resulting in a "loss of heterozygosity" or LOH.
(Loss of heterozygosity (LOH) in a cell represents the loss of normal function of one allele of a
gene in which the other allele was already inactivated). Hence the person will have 2 non-functional
Tumour suppressor alleles.
3. Mutations may also occur in the promoter regions. Promoter regions control how often, and
when the gene is transcribed. A mutation in the promoter region can result in a decrease or
absence of the tumour suppressor protein in the cell.
4. Mutations can also occur in the protein coding region of the gene causing the protein to be
more susceptible to degradation. If the tumour suppressor proteins are being degraded at a
higher than normal rate, they will not be able to perform their functions as tumour suppressors
since the duration which they remain in the cytoplasm is not greatly shortened.
Unlike oncogenes, tumor suppressor genes generally follow the 'two-hit hypothesis', which
implies that both alleles that code for a particular gene must be affected (mutated or
removed) before an effect is manifested. This is due to the fact that if only one allele for
the gene is damaged, the second can still produce the correct protein.
This is a "loss of function" mutation (ability to inhibit cell growth).
The mutation is also usually recessive. This means that the trait is not expressed unless
both copies of the normal gene in the cell are mutated.
[Question: How is it that both copies of the gene can be mutated?]
How is it that both copies of the gene can become mutated? In some cases, the first mutation is
already present in a germ line cell (egg or sperm); thus, all the cells in the individual inherit it.
Because the mutation is recessive, the trait is not expressed. Later on in life, a mutation occurs in
the second copy of the gene in a somatic cell. In that cell both copies are mutated and the cell
develops uncontrolled growth.
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4.2.2
An example of tumour suppressor gene: p53 gene
p53 gene codes for a p53 protein which is a transcription factor. The p53 transcription
factor activates the expression of DNA repair genes as well as the expression of a Cdk
inhibitor known as p21.
p21 in turn stops the cell cycle at G 1 phase allowing time for the cell to repair the damaged
DNA before it initiates DNA synthesis
Thus in normal cells expressing normal p53 protein, the exposure to radiation, chemical
mutagens, or UV light can activate p53 protein which will:
1. cause the inhibition of cell cycle
2. activate DNA repair genes
3. stimulate programmed cell death (apoptosis) if DNA damage is beyond repair
The following mechanism activates the p53 protein activity:

1. DNA damage is an intracellular signal
2. The signal is passed via protein kinases
3. The final protein kinase causes the activation of a transcription factor, p53. Active p53
promotes transcription of the gene encoding a protein that inhibits the cell cycle. The
resulting suppresion of cell division ensures that the damaged DNA is not replicated.
Result of loss of function mutation in p53

gene
If p53 is damaged or mutated, it will
remain in the inactive state allowing DNA
damage to accumulate within a cell, thus
increasing the risk for cancer formation.
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22
5.
Development of cancer is a multi-step process
Cells control proliferation at several checkpoints and all these controls must be inactivated for
a cancer to be initiated. Development of cancer is therefore a multi-step process as it
involved an accumulation of an estimated 4 to 6 independent mutations in key regulatory
genes.
Most human cancers develop over many decades of time. The need to inactivate several
regulatory genes almost certainly explains why most cancers occur in people over 40 yrs
age.
More than one somatic mutation is generally needed to produce all the changes
characteristic of a full-fledged cancer cell.
In another words, cancer results from an accumulation of mutations.
About 4- 6 mutations must occur at the DNA level for a cell to become fully cancerous.
These usually include the appearance of at least 1 active oncogene and the mutation
or loss of several tumour suppressor genes.
Furthermore, since mutant tumour suppressor alleles are usually recessive, in most
cases mutations must knock out both alleles in a cells genome to block tumour
suppression. (Most oncogenes, on the other hand, behave as dominant alleles.)
Finally, in many malignant tumours, the gene for telomerase is activated. This enzyme
prevents the shortening of chromosome ends during DNA replication. Production of
telomerase in cancer cells removes a natural limit on the number of times the cells can
divide.
Since mutations occur throughout life, therefore the longer we live, the more likely we are
to develop cancer.
Using Colorectal Cancer as a model to support the multi-hit hypothesis for the formation of cancer
(see diagram below) :
The multi-step model of cancer development is well supported by studies of one of the
best understood types of human cancer colorectal cancer.
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23
Numerous changes must happen at the DNA level for a cell to become cancerous. These
usually include the appearance of al least one active oncogene and mutation or loss of
several tumour suppressor genes.
Since the tumour suppressor mutations are usually recessive, mutations must knock out
both copies in a cells genome to block the tumour suppression.
Like most cancers, colorectal cancer develops gradually. The first sign is often a polyp, a
small benign growth in the colon lining (known as polyp).
The cells of the polyp look normal, they divide unusually to become malignant, invading
other tissues.
The development of a malignant tumour is paralleled by a gradual accumulation of
mutations that convert proto-oncogenes to oncogenes and knock-out tumour
suppressor genes.
A ras oncogene and a mutated p53 tumour suppressor gene as well as other
tumour suppressor genes are often involved.
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The multi-step model of cancer development
Multiple Mutations Transform a

Normal Colon Epithelial Cell into a
Cancer Cell
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6.
Causative factors of cancer
Genetic, chemical carcinogens, radiation, loss of immunity can increase the chances of
cancerous growth. All these are contributing factors to cancer as they may increase the number
of mutations in any given cell.
1. Genetics (Genetic pre-disposition)

The inheritance of mutated proto-oncogenes/tumour suppressor genes increases chances of
cancer development (e.g. inherited defects in DNA repair genes such as BRCA1, BRCA2 and p53
higher chance of breast cancer development
Recall from previous section that cancer is a multi-step process that requires 4-6 mutations
for a cell to become fully cancerous. The mutations usually include the appearance of at
least 1 active oncogene (via gain of function mutation) and the mutation or loss of
several tumour suppressor genes (via loss of function mutations).
Thus any inherited mutations in these genes would increase the chances of the cell
eventually becoming fully cancerous after additional mutations.
2. Chemical
To date, many hundreds of synthetic chemicals have been shown capable of causing
cancer in laboratory animals. Among them are acrylamide, asbestos, benzene,
trichloroethylene, and vinyl chloride.
Chemicals, many of which have been historically linked to the workplace, have been
successfully limited through public health efforts, because they have been associated
with a variety of cancers. Examples of common chemicals that fall in this category are:
o
benzene (myelogenous leukaemia)
arsenic containing pesticides (lung cancer)
polychlorinated biphenyls (liver and skin cancers)
mineral oils (skin cancer)
Asbestos - lung cancer and mesothelioma (is a rare form of cancer that develops
from the protective lining that covers many of the body's internal organs, the
mesothelium. It is usually caused by exposure to asbestos.
One common feature among all chemical carcinogens (cancer-causing chemicals) is

the capability to produce mutations in DNA.
The reason why smokers have high risk of getting lung cancer lies in the fact that
smoking places the mutagens found in the cigarette smoke in direct contact with their
lung tissues.
In general, exposure to mutagens increases the chance of gain of function mutations and
loss of function mutations occurring in proto-oncogenes and tumour suppressor genes
respectively. When these happen, cancer results.
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3. Radiation
Radiation exposure increases the chance of cancer development

UV-B rays from the sun can damage DNA and is associated with more than 90% of skin
cancers, including melanomas
Radon (formed as part of the normal radioactive decay chain of uranium) has been
associated with lung cancer among those who work in mines; general levels of radon have
not posed a significant cancer threat
Radio frequency electromagnetic radiation from mobile phones or microwave ovens has
not been linked to cancer.)
Nuclear radiation is of sufficient energy to ionise molecules and is therefore carcinogenic.
4. Loss of immunity
Cells with cancer-causing potential arise constantly in the body but the immune system
normally discovers and destroys them
Cell-mediated immunity involves TC (cytotoxic T cells), NK (natural killer) & macrophages,

antibodies.
It appears that these cells recognize abnormal or foreign surface markers on tumor cells
Immune system fails in detecting cancer cells as cancer cells

o
may not be immunogenic enough (not able to cause the immune system to
perceive it as foreign enough)
may retain self-markers and not be recognized by the surveillance system
In other cases the tumor antigen may have mutated to escape detection
Enrichment: Immunity and Cancer

The immune system provides one of the body's main defences against cancer. When normal cells turn into cancer cells,
some of the antigens on their surface change. These new or altered antigens flag immune defenders, including cytotoxic
T cells, natural killer cells, and macrophages.
According to one theory, patrolling cells of the immune system provide continuing body wide surveillance, spying out
and eliminating cells that undergo malignant transformation. Tumors develop when the surveillance system breaks down
or is overwhelmed. Some tumors may elude the immune defenses by hiding or disguising their tumor antigens.
Alternatively, tumors may survive by encouraging the production of suppressor T cells; these T cells act as the tumor's
allies, blocking cytotoxic T cells that would normally attack it.
Blood tests show that people can develop antibodies to many types of tumor antigens (although the antibodies may not
actually be effective in fighting the tumor). Skin testing has demonstrated that tumors provoke cellular immunity as well.
Other efforts to attack cancer through the immune system centre on stimulating or replenishing the patient's immune
responses with substances known as biological response modifiers. Among these are interferons (now obtained through
genetic engineering) and interleukins.
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27
5. Other factors that can increase the chance of cancer development

i.
Viral infection
Some viruses carry oncogenes whose products cause transformation of host cells into
cancer cells
Viral genome may be inserted into regulatory sites e.g. after hosts promoter sequence.
Human papilloma virus (HPV) that causes cervical cancer
Epstein-Barr virus Burkitts lymphoma infection by hepatitis B and C viruses higher

chance of liver cancer development
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ii.
Bacterial infection
E.g. infection by Helicobacter pylori bacterium
This enhances production of free radicals in the cells more mutations higher chance
of stomach cancer development
iii.
iv.
Age
older person accumulates more mutations higher chance of cancer development
Lifestyle
v.
e.g. smoker / alcoholic higher chance of cancer development

Diet
e.g. high consumption of saturated animal fats higher chance of cancer development
June 2014
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Enrichment: Therapeutic Targets for the treatment of cancer

Modulation of the Cell Cycle
The disruption of normal cell cycle regulation, which is the hallmark of cancer, presents numerous
opportunities for targeting checkpoint controls to develop new therapeutic strategies for this
disease. Such strategies include induction of checkpoint arrest leading to apoptosis (programmed
cell death), arrest of proliferating cells in stages of the cell cycle, which may sensitize them to
treatment with other therapeutic agents such as radiation, and targeting of therapies toward
specific regulatory components of the cell cycle.
In Table 2, several classes of chemotherapeutic agents and their mechanisms of action are listed
along with information regarding their effects on the cell cycle. One of the most established
chemotherapeutic approaches is the induction of DNA damage and subsequent induction of
apoptosis.
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Your Deposits into Cancer Vocabulary Bank Account

(you may use the space provided beside the term to write down the definition from this set of notes or from
other reference books)
Keyword
Definition
Cancer
Mutation
Metastasis
Regulatory proteins
Tumour
Supressor
genes
Protooncogenes
Oncogenes
Loss-of-function mutation
Gain-of-function mutation
Apoptosis
Angiogensis
Ras gene
Rb gene
p53
June 2014
31
2014 National Junior College

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H2 Biology Lecture Notes

Your syllabus requires you to:

Describe the functions of common proto-oncogenes and tumour suppressor genes

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H2 Biology Lecture Notes

Biology by Campbell and Reece

Characteristics of cancer cells ................................................................................................................ 3

The cell cycle ........................................................................................................................................4

Phases of the cell cycle and how it is being regulated: .................................................................. 5

3. Characteristics of Cancer Cell ....................................................................................................... 11

Mutations of certain genes which can cause Cancer .......................................................... 12

Gain of function mutations in proto-oncogenes ...........................................................................14

Development of Cancer is a Multi-Step Process: .................................................................. 23

Causative factors and cancer ...................................................................................................... 26

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H2 Biology Lecture Notes

1. Introduction: How uncontrolled cell division results in cancer

Characteristics of cancer cells

Disorganized behavior and independence from surrounding normal tissues.

Permanent loss of cell differentiation.

Expression of special markers on their surface.

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H2 Biology Lecture Notes

The cell cycle

Interphase (consisting of G1, S, G2 phases);

Mitosis Phase (M phase) when nuclear division occurs and

Cytokinesis phase where cytoplasmic cleavage occurs.

These checkpoints ensure that the cell is

These checkpoints monitor DNA

There are three major Checkpoints in the

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H2 Biology Lecture Notes

Phases of the cell cycle and how it is being regulated

Raven and Johnson (1999)

Events Within Cell During Cell division

S is the phase in which the cell synthesizes a replica of its genome

M is the phase in which nuclear division occurs. It is traditionally subdivided into 4

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H2 Biology Lecture Notes

Regulation of the cell cycle

the G2 checkpoint (Have all the genes been replicated?)

the M checkpoint (Are all the chromosomes aligned properly at metaphase?)

Cyclin dependent kinases (Cdks)

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H2 Biology Lecture Notes

Factors affecting / Regulation

External factors such as

NB: Most cells in human body

All chromosomes must be

1. Just like the G2 checkpoint, the M

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H2 Biology Lecture Notes

The cyclin control system

Cdks (cyclin-dependent kinases) are enzymes (protein kinases) that activate or

For a Cdk-cyclin complex to be active, it must be:

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H2 Biology Lecture Notes

MPF (maturation-promoting factor / M-phase promoting factor) is the Cdk-cyclin

Campbell and Reece (2008)

Campbell and Reece (2008)

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H2 Biology Lecture Notes

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