cancer lecturenotes 2014 teacher

Property of National Junior College H2 Biology Lecture Notes

June 2014 1

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.


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References:

1. “Biology” by Campbell and Reece

2. “Molecular Biology of the Cell” by Alberts, Johnson, Lewis, Raff, Roberts and Walter

3. “Biology” by Raven, Johnson, Losos and Singer

4. “Genetics: From Genes to Genomes” by Hartwell, Hood, Goldberg, Reynolds, Silver and Veres.

5. “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 Characteristics of cancer cells ................................................................................................................ 3

2. The cell cycle ........................................................................................................................................ 4 2.1 Phases of the cell cycle and how it is being regulated: .................................................................. 5 2.2 Regulation of the cell cycle ...................................................................................................................... 6 2.3 The cyclin control system ........................................................................................................................ 8

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

4. Mutations of certain genes which can cause Cancer .......................................................... 12 4.2 Gain of function mutations in proto-oncogenes ........................................................................... 14 4.3 How do gain of function mutation occur? (3 possible ways) ................................................... 16 4.4 An example of a proto-oncogene: RAS gene .................................................................................... 18 4.5 Loss of function mutations in tumour suppressor genes, P53 ................................................. 20 4.6 How do loss of function mutation occurs? (3 possible ways) .................................................. 21 4.7 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


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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.


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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. Let’s study how the cell cycle is regulated in normal cells first.

The Cell cycle can be divided into three main stages:

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

o Mitosis Phase (M phase) when nuclear division occurs and

o 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.

o These checkpoints ensure that the cell is only allowed to proceed to the next phase only if it has properly completed the previous phase.

o These checkpoints monitor DNA replication, DNA damage and Chromosome-to-spindle tubule attachments.

o There are three major Checkpoints in the cell cycle, and are found in G1, G2, and M phase.


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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.

Events Within Cell During Cell division (Recall from cell division)

G1

G1 is a primary growth phase of the cell cycle. For many organisms, this encompasses the major portion of the cell’s life span.

S

synthesis

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 animal’s 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.

Raven and Johnson (1999)


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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

Description

Factors affecting / Regulation

Types of cyclin

involved (FYI)

G1 / S

Phase

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.

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.

NB: Most cells in human body are in the G0 phase.

CyclinD CyclinE (Also called G1 cyclins)

G2 / M 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)

M Check Point / Spindle Check Point

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.

All chromosomes must be present and the tensions between opposite poles are important.

Cyclin B


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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


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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)


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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.


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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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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.


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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.


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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)

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-oncogene’s protein product or in the intrinsic activity of each protein molecule i.e. how long it can stay functional in a cell’s cytoplasm.



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Table of Proto-oncogene and Oncogenes:

Types of Classification of Proto-oncogenes for Genes coding

Examples Effect of Mutation

(i.e. Becomes Oncogenes)

(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.)

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.

(b) Growth factor receptor proteins (are typically transmembrane proteins, which growth factors bind to. E.g. tyrosine kinase)

(To be learnt in greater detail in SH2)

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

(c) Intracellular protein kinases (These are enzymes that add phosphate groups to target proteins, thereby modifying their functions.)

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.

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)

Myc protein stimulates the transcription of genes required for cell proliferation.

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

* Required for H2 syllabus


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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 proto-oncogene, 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-oncogene’s 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

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)


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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.


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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.

o The signal is relayed to (3) a G protein called Ras. Like all G proteins, Ras is active when GTP is bound to it.

o Active Ras passes the signal to (4) a series of protein kinases. [Kinases are enzymes which add a phosphate group to another molecule]

o (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.


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[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 proliferation, which is a major step in tumour formation.


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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.

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 suppressor’s 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 G1 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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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 cell’s 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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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 cell’s 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)

o arsenic containing pesticides (lung cancer)

o polychlorinated biphenyls (liver and skin cancers)

o mineral oils (skin cancer)

o 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.

http://en.wikipedia.org/wiki/Cancer

http://en.wikipedia.org/wiki/Mesothelium

http://en.wikipedia.org/wiki/Asbestos


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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)

o may retain self-markers and not be recognized by the surveillance system

o 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.

http://thyroid.about.com/library/immune/blimm35.htm#Biological response modifiers


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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 host’s promoter sequence.

Human papilloma virus (HPV) that causes cervical cancer

Epstein-Barr virus – Burkitt’s 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. Age

older person accumulates more mutations higher chance of cancer development

iv. Lifestyle

e.g. smoker / alcoholic higher chance of cancer development

v. Diet

e.g. high consumption of saturated animal fats higher chance of cancer development


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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

Proto-

oncogenes

Oncogenes

Loss-of-function mutation

Gain-of-function mutation

Apoptosis

Angiogensis

Ras gene

Rb gene

p53


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cancer lecturenotes 2014 teacher

Documents

cell cycle

molecular cell biology

development of cancer

core syllabus cancer

loss of function mutations

molecular biology

gain of function mutations

tumour suppressor genes