11 - cell cycle and cancer I

Question 1

How many people will develop cancer?

Answer 1

1 in 2

Question 2

Why has cancer overtaken heart disease as the leading cause of death in UK?

Answer 2

1. A disease of the aging population

2. Better treatment for heart disease; lower mortality

Question 3

Cancer: the facts and figures

Answer 3

Lung cancer - most common cancer (both sexes)
Prostate cancer - second biggest killer in men
Breast cancer - second biggest killer in women

Question 4

cancer

Answer 4

a serious disease that is caused when cells in the body grow in a way that is uncontrolled and not normal, killing normal cells and often causing death

Question 5

Is cancer a single disease?

Answer 5

NO- it is a complex family of diseases.
There are as many forms of cancer as there are types of cell in the body - cell cycle regulation can go wrong in all cells - proliferation. >200 different cell types!

Question 6

Carcinoma

Answer 6

cancers arising from epithelial cells (surface cells - e.g. lining of gut, skin, cells lining airways of the lungs)
- constitute 80-90% of all cancers -
why? - these cells are exposed to the environment - carcinogens

Question 7

Sarcoma

Answer 7

• cancers of connective and supportive tissues e.g. bone cancer, muscle – rare 1%

Question 8

Myeloma

Answer 8

cancers of the plasma cells of the bone marrow - antibody producing cells - secondary infections (pneumonia and pyelonephritis (urinary tract
infection)

Question 9

Lymphoma

Answer 9

solid tumours of the lymphatic system
lymph glands, lymph nodes or in organs - tonsils, spleen, thymus
– formed from maturing WBC

Question 10

Leukemia

Answer 10

• ‘blood’ cancers - more specifically precursor blood cells in bone marrow
  – circulating white or red blood cells.
  Excess of immature cells - don’t function - anaemia - suppressed immunity

Question 11

Mixed classifications

Answer 11

teratocarcinoma –
cancers originating in germ cells and stem cells – therefore encompasses a range of cancers – testicular, ovarian, even placental

Question 12

Normal cells in culture

Answer 12

1. Anchorage dependent growth - no attachment no growth
2. Density dependent growth - stop growing when confluent - signals from other cells- will re-grow to fill gaps but then stop again

Question 13

Cancerous cells in culture

Answer 13

1. No anchorage dependence - seldom anchor to base of flasks but grow anyway - generally have rounded appearance
2. No density dependence - growth not controlled by other cells - instead of a monolayer they just continue growing on top of each other

Question 14

Other factors contributing to the abnormal proliferation of cancerous cells

Answer 14

1. Immortality
2. Reduced reliance on growth factors produced by other cells
3. Increased production of growth factors
4. Changes in cell membrane structure and function

Question 15

1. Immortality

Answer 15

• Normal diploid (body) cells - limited life expectancy in culture e.g. human
fibroblasts 50 to 60 population doublings - viability then decreases (Hayflick limit)
• Cancerous cells - indefinite number of population doublings
• Life expectancy of normal cells related to shortening of chromosomal telomeres (buffer zone at end of chromosome). Cancerous cells are able to maintain telomere length - telomerase
• HeLa cells - cultured from cervical carcinoma from Henrietta Lacks - died from cancer in 1951 Cells still used in labs around the world today

Question 16

2. Reduced reliance on growth factors produced by other cells

Answer 16

External growth factors required for progression through G1 cell cycle checkpoint

• e.g. 3T3 fibroblasts - normal cells only grow in culture media containing certain growth factors.
• Transform these cells (turn them into cancerous cells) by viral infection (e.g. SV40, Rous sarcoma virus)
• grow happily on a basal media lacking the same growth factors

Question 17

3. Increased production of growth factors

Answer 17

In addition to being less reliant on growth factors produced by other cells, cancerous cells themselves may over produce growth factors in order to promote growth

• increased expression of growth factors
• increased ‘shedding’ of growth factors

Question 18

4. Changes in cell membrane structure and function

Answer 18

Cell surface / plasma membrane is a strong determinant of cellular ‘social’
behaviour;
- e.g. communication, cell movement, adherence, access to nutrients, recognition by the immune system
- glycolipids, glycoproteins, proteoglycans, mucins

Question 19

Basic stages of cancer disease progression

Answer 19

1. Initiation
2. Clonal expansion
3. Primary tumour
4. Secondary mutations
5. Malignancy
6. Invasion
7. Metastasis

Question 20

1. Initiation

Answer 20

Single cell undergoes a single mutation - confers a growth advantage which causes it to lose some of its growth control

Question 21

2. Clonal expansion

Answer 21

Proliferation begins - mutated cell divides quicker than surrounding cells to form a cluster of ‘clones’
- disease is monoclonal

Question 22

3. primary tumour

Answer 22

The cancer remains in situ (i.e. not moved from site of original mutation).
Tumour benign - not invaded surrounding tissues - surgery possible

Question 23

4. Secondary mutation

Answer 23

Secondary mutations provide a new phenotype with a selective advantage

Question 24

5. malignant cancer

Answer 24

Following the secondary mutation the cells lose contacts with their neighbours

• become invasive
• secrete proteases to breakdown the extracellular matrix holding cells in place
• risk of metastasis

Question 25

6. invasion of lymph and/or blood vessels

Answer 25

First stages of metastasis - cancer cells have low adherence - easy to break off main tumour and enter vessels

Question 26

7. Metastatic tumour

Answer 26

Cell from original tumour in lung has now entered a vessel and emerged at the other end to form a new tumour in another organ

Question 27

Characteristics of malignant tumour cells

Answer 27

1. Show excessive proliferation 2. Unusual number of chromosomes (aneuploidy) - e.g. HeLa (cervical carcinoma) cells have 82 chromosomes instead of 46; multiple copies of some chromosomes 3. 'Deranged' metabolism - e.g. increased demands for nutrients and use aerobic glycolysis for ATP generation (Warburg Effect) 4. Reduced attachments to neighbouring cells - enables spread to other tissues 5. Invasive phenotype: Detaches from original tumour and enter blood stream and lymph system 6. Proliferate in other parts of the body - METASTASIS

Question 28

Carcinogens:

Answer 28

Cigarette smoke- mutagenic chemicals Sunlight - UV radiation Viruses - insert DNA into genome e.g. human papillomavirus (HPV) cervical cancer

Question 29

DNA mutations:

Answer 29

Mutations in genes encoding proteins which regulate cell division

Question 30

Proto-oncogenes become oncogenes in a number of ways

Answer 30

in each case, there is an excess of growth-stimulating activity Translocation/transposition: New promoter means gene transcribed more efficiently - more protein Gene amplification: Gene replicated - more total protein Point mutation within a control element: e.g. mutation in existing promoter - more efficient transcription Point mutation within gene: Changes the protein itself - more growth promoting?

Question 31

Looking for oncogenes

Answer 31

1. Extract DNA from human tumor cells - contains mutated oncogene 2. Tumour DNA 'transfected' (introduced) into mouse cells (3T3). Cells which have taken up the oncogene proliferate and for dense foci 3. Cells in the foci may have taken up other bits of the human DNA in addition to the oncogene. 4. DNA from 'focus' is reintroduced into fresh mouse (3T3) cells to dilute out human DNA other than the oncogene 5. Extract mouse genomic DNA from cells - now contains the human oncogene 6. Fragment DNA (restriction digest) & introduce into bacterial virus - PHAGE LIBRARY

Question 32

How do we know which phage contains the oncogene?

Answer 32

1. Add phages to a plate of bacteria - at the right dilution each single phage will kill the bacteria - empty spot on plate 2. 'Blot' plate onto filter paper. Human DNA contains Alu sequences (mouse does not) - detect the human oncogene DNA using a probe against Alu

Question 33

Oncogenes identified in this way were found to code for components of signalling pathways

Answer 33

1. Growth factor binds to a receptor at the cell surface 2. Receptor is also a protein kinase which autophosphorylates itself when bound to growth factor 3. Change in receptor phosphorylation detected by Ras When Ras detects phosphorylated vector it binds to GTP which then activates Ras 4. Activated (GTP bound) Ras signals to Mitogen activated protein kinases (MAPK) operating in a cascade (one phosphorylates and activates another) 5. Cascade ends when the final kinase phosphorylates a transcription factor in the nucleus - in the case of cancers the activated transcription factors enhance the expression of genes controlling the cell cycle e.g. cyclins

Question 34

Ras

Answer 34

a 'small G protein' (small and bind guanine nucleotides)

Question 35

Transcription factors

Answer 35

regulate gene expression

Question 36

The Ras oncogene

Answer 36

1. Discovered when DNA from a bladder tumour was used to transform mouse 3T3 cells 2. The oncogene differs from the proto-gene by only a single nucleotide - different amino acid in protein (G12V) - causes Ras to remain permanently bound to GTP – permanently active! 3. Stimulatory signal to nucleus never switched off - excessive production of cyclins - uncontrollable growth! 4. Explains why cancer cells don't require external growth factors like normal cells