Cancer Notes 2010

St Andrew’s Junior College Cellular Functions / Cancer

2010 H2 Biology 1

Cellular Functions/ Organisation & Control of Prokaryotic & Eukaryotic Genomes

Molecular Biology of Cancer & Multistep Model of Cancer Development

Essential Reading: Campbell, N.A., and Reece, J.B. (2008). Biology (8th Edition). Chapter 12 pp 242 – 243; Chapter 18 pp. 373-377 Objectives: 1n) Explain how uncontrolled cell division can result in cancer, and identify factors which can increase the chances of

cancerous growth (knowledge that dysregulation of checkpoints of cell division can lead to uncontrolled cell division and cancer is required but details of mechanisms are not required).

4(j). Describe the development of cancer as a multistep process. 4(i) Describe the functions of common proto-oncogenes and tumour suppressor genes e.g. ras and p53 genes. 4(i) Describe how oncogenes are formed through gain of function mutations in proto-oncogenes and loss of function

mutations in tumour suppressor genes. CANCER

• Clinically, cancer is defined as a family of a large number of different complex diseases. • Cancers vary in their ages of onset, growth rates, prognoses and responsiveness to treatments. • Despite their differences, all cancers exhibit common characteristics at the molecular level that

unite them as a family. • Cancers are a result of uncontrolled cell division (mitosis).

TUMOURS • When a cell loses genetic control over cell growth (due to mutations), it can result in a tumour. • Tumour = a mass or lump of cells, with an inherited

capacity for autonomous, uncontrolled growth, resulting from uncontrolled cell division.

• 2 types of tumours:

Benign tumours o Surrounded by a connective tissue capsule

and thus isolated (eg, moles, warts).

o Cells are not cancerous but conversion to cancerous cells is possible.

o Benign tumours can be removed surgically or killed by radiation, usually eliminating any further cancer development at that site.

o More typical compared to malignant tumours.


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Malignant tumour o Made up of cancer cells o Connective tissue capsule breaks down and cancer cells send out signals for the

production of a new blood vessels (angiogenesis) at the tumour site. This is because blood supply allows removal of metabolic waste and supply of

nutrients which allow for rapid growth. Angiogenesis is also required for the spread of cancer.

o These cells can also:

(i) invade and damage surrounding tissues (invasion); (ii) separate from the original tumour and penetrate blood and lymph vessels

of the circulatory system, thus spreading to other parts of the body and proliferate to form new tumours (metastasis).

Metastasis is the spreading of cancer cells to locations distant from their original site. Usually

surgery is performed to remove the tumour, followed by radiation and chemotherapy.

(Figure of malignant tumour showcasing invasion and metastasis)


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PROPERTIES OF CANCER CELLS

• Contain oncogenes (mutated version of proto-oncogenes; more details will be covered under CANCER DEVELOPMENT)

• Exhibit abnormal cell growth and division (cell proliferation).

• Do not undergo apoptosis i.e. programmed cell death.

• Do not become specialized i.e. remain undifferentiated.

• Can stimulate growth of blood vessels towards itself (angiogenesis)

• Can invade surrounding tissues or metastasize to other tissues.

• Lack of control by cell cycle checkpoints. • Do not show density-dependent inhibition and anchorage dependence. DENSITY-DEPENDENT INHIBITION • Phenomenon in which normal cells stop dividing because of insufficient nutrients • When a normal cell population reaches a certain density, the availability of nutrients becomes

insufficient to allow continued cell growth and division. • Cancer cells can divide well beyond a single layer to give a clump of overlapping cells. ANCHORAGE DEPENDENCE • In order for most animal cells to divide, they must be attached to a substratum e.g. extra-cellular

matrix • Anchorage is signalled to the cell cycle control system via pathways involving plasma membrane

proteins and elements of the cytoskeleton linked to them.

(Figure showing density-dependent inhibition and anchorage dependence in (a) normal cells and (b) cancer cells)

(Figure showing angiogenesis)


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CELL CYCLE CHECKPOINTS • The cell cycle consists of 3 major checkpoints (G1, G2 and M phase checkpoints). • Checkpoints are control points whereby stop and go-ahead signals can regulate the cell cycle. • G1 phase checkpoint seems to be the most

important for most cells. o If go-ahead signal is received:

cell will usually complete the G1, S, G2 and M phases

and cell divides. o If go-ahead signal is NOT received:

cell will exit cell cycle, switching into a non-dividing state known as the G0 phase.

DIFFERENCES BETWEEN NORMAL CELLS AND CANCER CELLS Normal Cells Cancer Cells

• Have proto-oncogenes, whose functions are to promote the normal growth and division of cells

• Have oncogenes, which are mutated forms of proto-oncogenes. When oncogenes are switched on, cells divide excessively, resulting in abnormal cell proliferation.

• Show programmed cell death. They divide for a certain number of times then stop dividing

• Do not show programmed cell death. They can divide indefinitely

• Exhibit density-dependent inhibition. • Do not exhibit density-dependent inhibition. • Exhibit anchorage dependence • Do not exhibit anchorage dependence • Differentiate properly to become specialised

cells, e.g. nerve cells, liver cells etc• Fail to differentiate properly

• Regulation by cell cycle checkpoints • Lack of regulation by cell cycle checkpoints


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FACTORS THAT INCREASE THE CHANCE OF CANCER

• Cancer develops gradually as a result of a complex interaction of factors related to environment, lifestyle, and heredity.

1) Age

• Chances of developing cancers increase with age. Cancer results from mutations in certain genes and since each mutation is a rare event, most cancers develop later in life.

2) Genetic Factors

• Certain cancers run in families. Eg. colon cancer. Mutated genes may be passed from one generation to the next.

3) Carcinogens

• Agents are capable of causing cancer, such as radiation and some types of chemicals. • Examples are UV light (possible cause of skin cancer) and X-rays (cause of leukaemia and skin

cancer). • X-rays cause ionisation of molecules and the highly reactive ions stimulate mitosis and destroy cells. • Chemical carcinogens include cigarette smoke which contains 4000 different chemicals, many of

which are responsible for causing lung cancer.

4) Viruses

• Some viruses can cause genetic changes in cells, increasing their likelihood to become cancerous. • Possible ways are

(i) the viral DNA integrates into the host cell DNA (cause the cells to proliferate), (ii) changing the host cell’s surface interactions (loss of density-dependent inhibition), (iii) switching on DNA replicating machinery of the host (continued host cell division) (iv) providing oncogenes.

• Eg. The human papilloma virus causes cancer of the cervix and the Epstein-Barr virus is associated with Burkitt’s lymphoma.


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INDICATIONS THAT CANCER IS A MULTISTEP PROCESS REQUIRING MULTIPLE MUTATIONS • Cancer is a multistep process: develop in progressive steps from mildly aberrant cells (cells that

deviate from the norm) to increasingly tumourigenic and malignant (accumulation of mutations) • Therefore a single mutation is not sufficient to transform a normal cell into a malignant cell. • Indication 1: Chances of getting cancer

increases with age, indicating that cancer develops from the accumulation of several mutagenic events in a single cell.

• Up to ten independent mutations, occurring

randomly and with a low probability, are necessary before a cell is transformed into a malignant cancer cell.

• Indication 2: Delay that occurs between

exposure to carcinogens and the appearance of cancer. • E.g. an incubation period of 5 to 8 years

separated exposure of people to the radiation of the atomic explosions at Hiroshima and Nagasaki and the onset of leukaemia.

MULTISTEP MODEL OF CANCER DEVELOPMENT: COLORECTAL CANCER • Colorectal cancer, like most cancers, develops gradually. • First sign is a small, benign growth (polyp) with fast dividing cells in the colon lining. • The cells in the polyp may look normal, but they are dividing unusually frequently. • The tumour grows and may eventually 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 - knock out tumour suppressor genes.

(Multi-step model of colorectal cancer showing an accumulation of mutations in critical genes involved in cancer development (i.e. tumour suppressor genes & oncogenes)).


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PROTO-ONCOGENES • Proto-oncogenes are genes whose products promote cell growth and division, and have essential

function in normal cells. • They do this by encoding:

1. transcription factors that stimulate expression of other genes 2. signal transduction molecules that stimulate cell division e.g. ras gene codes for Ras protein 3. cell cycle regulators that move the cell through the cell cycle

• Proto-oncogene products may be located in the plasma membrane, cytoplasm, or nucleus • Activities of proto-oncogene products are controlled in various ways e.g. regulation in the

transcriptional, translational and protein modification levels. • When cells become quiescent (non-dividing) and cease division, expression of most proto-

oncogene products are repressed. ras PROTO-ONCOGENES • The ras gene family encodes signal transduction molecules (Ras proteins) that are associated

with the cell membrane. • Ras proteins normally transmit signals from the cell membrane to the nucleus, stimulating the cell

to divide in response to external growth factors; thus, regulate cell growth & division. • Ras proteins cycle between an inactive and an active state by binding either GDP or GTP

respectively Signal Transduction Pathway mediated by Ras 1. When a cell encounters a growth factor, the

growth factor binds to the receptor on the cell membrane, resulting in autophosphorylation of the cytoplasmic portion of the growth factor receptor

2. Recruitment of nucleotide exchange factors (proteins) to the plasma membrane cause Ras to release GDP in exchange for GTP, thereby becoming activated.

3. The activated, GTP-bound Ras sends signals through protein phosphorylation cascades in the cytoplasm.

4. Activation of nuclear transcriptional factors stimulates expression of genes whose products activate or repress gene transcription, driving the quiescent cell into the cell cycle (stimulates the cell cycle).

5. Once Ras has sent its signals to the nucleus, it hydrolyses GTP to GDP and becomes inactive; hence Ras also known as GTPase.

Nucleotide exchange factor


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GENETIC CHANGES CAUSE CONVERSION OF PROTO-ONCOGENES TO ONCOGENES • When a proto-oncogene is mutated or aberrantly expressed, and contributes to the development

of cancer, they are known as oncogenes. • Oncogenes = cancer-causing genes • Proto-oncogenes may be:

- Mutated, resulting in a protein product that is continually in an “on” state (constitutively active1) which constantly stimulates the cell to divide.

- Aberrantly expressed, resulting in high levels of the protein product which constantly stimulate the cell to divide; proto-oncogenes are either over-expressed or unable to be transcriptionally repressed at the correct time.

• Specific genetic alterations can convert normal proto-oncogenes to abnormal oncogenes:

1. Chromosomal translocations 2. Gene amplification 3. Viral integration 4. Point mutation

1. CHROMOSOMAL TRANSLOCATION (i) Cancer cells are frequently found to contain

chromosomes that have broken and rejoined incorrectly. (ii) If a translocated proto-oncogene ends up near an

especially active promoter, or other control element, its transcription may increase, making it an oncogene.

(iii) Movement of transposable elements may also place a more active promoter near a proto-oncogene hence increasing its expression.

2. GENE AMPLIFICATION (i) An abnormal increase in the copy number2 of a

proto-oncogene. (ii) For example, the myc gene, which codes for a transcription factor, has been amplified in

human leukemias, breast, stomach, lung, and colon carcinomas3.

1 Constitutive – continually / at a constant rate 2 Copy number - number of copies of a gene within a cell's genome 3 Carcinoma – malignant cancer

Over-expression

Constitutive active


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3. VIRAL INTEGRATION When a virus integrates into the chromosome, it may enhance the expression of nearby proto-oncogenes.

4. POINT MUTATION (i) Within the control element: change in the nucleotide sequence in the promoter or enhancer

causing an increased expression of proto-oncogene or changes in the silencer resulting in the inability to repress gene expression.

(ii) Within the gene: change in the coding sequence resulting in a protein that is more active or

more resistant to degradation than the normal protein (constitutively activated protein).


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GAIN OF FUNCTION MUTATIONS IN ras PROTO-ONCOGENE • One of the most frequently mutated proto-oncogenes in human tumours comes from the ras gene

family. • Point mutation in ras proto-oncogene converts it to ras oncogene encoding for abnormal Ras

protein • A comparison of amino acid sequences of Ras proteins from normal and cancer cells shows that

oncogenic Ras proteins have single amino acid substitutions at either position 12 or 61.

• Changes in the structure of the Ras protein:

1. Prevent Ras protein from hydrolysing GTP to GDP 2. Freezes the Ras protein in a permanent “on” conformation

• Abnormal Ras protein is constitutively activated even in the absence of a growth factor; resulting in excessive cell division

(Action of ras oncogene on over-stimulation of cell cycle)


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TUMOUR –SUPPRESSOR GENES • Besides proto-oncogenes, cells also contain tumour-suppressor genes whose normal products

inhibit cell division during DNA damage and help prevent uncontrolled cell growth by - regulating cell cycle checkpoints - initiating the process of apoptosis (programmed cell death)

• The normal proteins encoded by tumour-suppressor genes - are components of cell-signalling pathways that inhibit the cell cycle. - stop progress through the cell cycle in response to DNA damage, or growth-

suppression signals from the extra-cellular environment - control the adhesion of cells to each other or to the extra-cellular matrix

p53 TUMOUR SUPPRESSOR GENE • The most frequently mutated tumour suppressor gene in human cancers is the p53 gene. • p53 gene encodes a nuclear protein that acts as a transcription factor that represses or stimulates

transcription of more than 50 different genes. • When not needed, p53 protein is normally bound to another protein called Mdm2. • Mdm 2 prevents phosphorylations and acetylations that convert the p53 protein from an inactive to

an active form • Normally the p53 protein is continuously synthesised but is rapidly degraded so it is present in

cells at low amounts. • A primary role of the p53 protein is to determine if a cell has incurred DNA damage. • DNA damage can be caused by several events:

- chemical damage to DNA - double-stranded breaks in DNA induced by ionising radiation - presence of DNA-repair intermediates generated by exposure of cells to ultraviolet

light • DNA damage results in

- Increase in p53 protein phosphorylation and acetylation - Increase in p53 protein stability

• Hence, rapid increase in the nuclear levels of activated p53 protein • Activated p53 protein initiates three different responses to DNA damage through activation of

genes: 1. Activate genes that promote DNA repair.

• To prevent accumulation of mutations that activates oncogenes or inactivate tumour suppressor genes.

2. Activate genes that arrest cell division and may generally repress other genes that required for

cell division. • To allow more time for the cell to repair its DNA. • To avoid producing two mutant daughter cells.

3. Activate genes that promote apoptosis (programmed cell death).

• An active process that causes cell shrinkage, chromatin condensation, and DNA degradation.

• Each of these responses is accomplished by p53 acting as transcription factor that stimulates or represses the expression of genes involved in each of these responses E.g. stimulate genes that promote gene repair, while repress genes of cell division

High levels of activated p53


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GENETIC CHANGES IN TUMOUR SUPPRESSOR GENES • When tumour suppressor genes are mutated or inactivated, cells are unable to

- respond normally to cell cycle checkpoints - undergo programmed cell death if DNA damage is extensive.

• This leads to further increase in mutations, and the inability to leave the cell cycle when the cell should be quiescent.

- stimulating growth through the absence of suppressor LOSS OF FUNCTION MUTATIONS IN p53 TUMOUR SUPPRESSOR GENE • Cells lacking functional p53 are unable to arrest at cell cycle checkpoints or to enter apoptosis in

response to DNA damage. • As a result, they move unchecked through the cell cycle, regardless of the condition of the cell’s

DNA (mutated) • Mutations & chromosomal aberrations that lead to cancer accumulate; hence cells lacking p53

have high mutation rates.

(Action of p53 tumour-suppressor gene on inhibition of cell cycle)


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CHANGES AT THE DNA LEVEL (FOR A FULLY CANCEROUS CELL) (i) Appearance of at least one active oncogene.

- Most oncogenes behave as dominant alleles; only need one mutated allele to produce abnormal protein.

- Gain of function mutation in ras proto-oncogene. (ii) Mutation or Loss of function of several tumour suppressor genes.

- Mutant tumour suppressor alleles are usually recessive; mutations must knock out both alleles in a cell’s genome to produce abnormal protein (unblock tumour suppression).

- Loss of function mutation in p53 tumour suppressor gene

(iii) In many malignant tumours, the gene for telomerase is activated, preventing the shortening of

chromosome ends during DNA replication – number of times the cell can divide is unlimited.

Loss of function uncontrolled cell proliferation

Documents

Cancer Notes 2010