Edmead Lec 3

Question 1

Tumour suppressor genes:
Normally function to _____ growth
_____, loss of function mutations predispose to cancer formation (need loss of function in ____ alleles) unlike oncogenes which are normally ____ mutations.

Answer 1

Restrict
Recessive
2
Dominant

Question 2

Sporadic mutations are in the ______

Germline mutations are _____

Answer 2

Individual

Inherited

Question 3

Cancers tend to have inactivating mutations in 1 or more ____ _____ genes.
Deletions/____ mutations results in no protein or a protein with altered functions (_______).

Sequence deletions or ____ mutations can mean _____ termination occurs resulting in a short non functional protein which is usually ______ in cells meaning a loss of protein. This can be due to a ____ chance which can introduce a stop codon.

Answer 3

Tumour suppressor 
Point
Dysfunctional 
Point 
Premature 
Degraded 
Base

Question 4

Main classes of tumour suppressor genes:

1) Growth/development suppressors e.g. ____, patched1
2) Cell cycle ____ proteins e.g. ___, ___
3) Cell cycle inhibitors e.g. CDKI, ___
4) Inducers of apoptosis e.g. ___, ____
5) DNA repair enzymes e.g. Xeroderma pigmentosa
6) Developmental pathways e.g. patched, Hh pathway

Answer 4

TGFB (inhibitory signals dampen down cell growth)
Checkpoint e.g. pRb, p53
CDKI, P16
Bax, P53 (to kill off cells with mutations)

Question 5

BRCA1/2 are tumour suppressor genes:
Predispose to ____ cancer - argument for screening
Mutated genes produce _____ proteins causing LOF
Involved in homologous recombination and ____ _____ break repair so maintain integrity of genome but if defective then the genome is ________ and lead to chromosomal rearrangement and mutations.

Answer 5

Breast
Truncated
Double strand break repair
Destabilised

Question 6

Retinoblastoma (Rb protein is a tumour suppressor)
40% or retinoblastoma is inherited as inheriting one mutant copy of the gene means if the other allele is mutated then cancer occurs.
Rb protein regulates cell cycle by inhibiting __ to _ phase transition.
Rb indirectly regulates transcription for specific gene expression that affects cell ____ and ______.
Rb binds and modulates transcription factor ___ and chromatin remodelling enzyme.

Answer 6

G1 to S phase
Proliferation and differentiation
E2F

Question 7

pRb regulates the ___ ___.
It interacts with ___ and _____ to prevent the transition of __ to _ phase.
In absence of growth signals, pRb is ____phosphorylated and binds both ___ and ___ to sequester ___ and block its transactivation domain which prevents it interacting with general transcription factors. ____ inhibit the expression of ___ target genes e.g. Cyclin E, A and CDK2 which are needed for cycle cycle progression.

Answer 7

Cell cycle
E2F and HDACs
G1 to S phase
Hypophosphorylated 
E2F and HDACs to sequester E2F 
HDACs 
E2F

Question 8

Cell cycle is regulated by signals e.g. growth factor stimulation or upregulation of critical proteins e.g. ____ D which activates ____ which then phosphorylates ___. In it’s resting state ____ is unphosphorylated and sequesters ___ but _____ D and _____ phosphorylate ___ and change its conformation and ___ is then released and can stimulate genes and proteins so continuation into _ phase from __ occurs.

Cell cycle then continues and cell growth and proliferation can occur.

Answer 8

Cyclin D 
CDK4
pRb
pRb
E2F
Cyclin D and CDK4
pRb 
E2F  
S
G1

Question 9

P53 evolved to prevent _____ development.
It is usually present in ___ levels in cells and is bound to an inhibitor ____ (which sequesters the P53).
If a stress signal is received e.g. mutation, broken DNA, Infection, cytokine stress then the ____ is degraded and the P53 is freed up and activated.

Answer 9

Tumour
Low
MDM2
MDM2

Question 10

Free P53 is ____ when unbound and is _____ quickly. But it is ____ by binding damaged ___.

Answer 10

Unstable
Degraded
Stabilised
DNA

Question 11

P53 is a ______ factor which enters the ____ of cells to find damaged ____, bind to it and ____ itself to stop itself being _____. Once bound to ____ it undergoes it’s role as a _______ regulator and can switch on genes

Answer 11

Transcription 
Nucleus 
DNA
Stabilise
Degraded
DNA
Transcriptional

Question 12

P53 binds as a _____. So requires _ molecules of P53 to associate together to act as a ______ factor to switch on gene ______.

Answer 12

Tetramer
4
Transcriptional
Transcription

Question 13

Name 4 types of stress that can activate P53:

What does P53 do as a result of this? (5)

Answer 13

DNA damage
Hypoxia/anoxia
Loss of survival signals
Oncogenes

Cell cycle arrest - upreg CDKs e.g. P16
Differentiation of cells - less proliferation
DNA repair - activate DNA repair enzymes
Senescence - prolonged rest, repair and growth occurs
Apoptosis - upreg bax if damage too great to repair

Also metabolic effects e.g. increase autophagy and decrease glycolysis.

Question 14

P53 regulates the expression of:
___and___ - cyclin dependent kinase inhibitors
____ (inhibitor of P53 action)
___ (pro apoptotic)

Answer 14

P21 and P16
MDM2
BAX

Question 15

Mutations of P53 occur in over __% of human cancers:
Start to acquire mutations as cells don’t die by _____
Mutant P53 proteins are more ___ and block binding sites of functional P53
Mutations often occur in ___ binding region of P53 so can’t bind DNA and promote gene _____.

Answer 15

50% 
Apoptosis 
Stable 
DNA 
Transcription

Question 16

Mutant P53 can interfere with WT P53 action:
If _ mutated P53 allele then P53 protein is still made but __% made is mutated that doesn’t bind ___.
Need _ molecules of P53 to make an active transcription factor but if __% P53 made is mutated then chance of incorporating one mutated P53 molecule is high and ____ will be dysfunctional.

E.g. in Li Fraumeni syndrome where born with mutated P53

Answer 16

1 
50% 
DNA 
4
50 
Tetramer

Question 17

____ sequesters P53 in healthy cells and if damaged DNA is present then P53 is ____ to activate it and ____ releases the P53. It then binds as a ___ to damaged DNA and causes transcription. BUT if one P53 in ____ is mutated then it can’t cause transcription and can’t cause apoptosis or halt cell cycle.

Answer 17

MDM2
Phosphorylated 
MDM2 
Tetramer 
Tetramer

Question 18

P53 mutations:
In __% lung cancers there are mutations in P53
Mutagens in cigarette smoke cause G to _ ___ mutations in DNA binding regions of P53.
Hence the apoptotic pathway cannot occur as ___ not transcribed etc…

In liver cancer aflatoxin (____ metabolite) leads to G to _ transversion in DNA binding regions.

Answer 18

60
G to T transversions (point mutations)
BAX

Fungal
G to T transversion

Question 19

P53 structure:
N terminus on ___ and C terminus on ___
____ binding region on left
___ virus binds P53 and inhibits its actions - leads to cervical cancer
The _______ domain is on the right
The DNA sequence specific binding domain is in the centre - needed as it’s a ____ factor.

Answer 19

Left 
Right 
MDM2
HPV 
Tetramerisation 
Transcription factor

Question 20

P53 status:
P53 is rarely mutated in cervial cancer but the polymorphism affects susceptibility to HPV mediated ____ - screening would be costly and time consuming so all girls are now vaccinated against HPV

Answer 20

Degradation

Question 21

Chemotherapeutic agents and radiation rely on inducing ______ for cytotoxic effects e.g. by causing x linking.
BUT if P53 is mutated then the _____ pathway cannot be activated so cytotoxic agents don’t work. So lack of P53 action often makes tumour cells ______ to cytotoxic agents through lack of functioning apoptotic pathway.

Research is focussed on restoring ____ in tumour cells so apoptotic pathway is functional and then can use agents to promote apoptosis!

Answer 21

Apoptosis
Apoptotic
Resistant
P53

Question 22

P53 as target for therapy:
A____ is used in ______ transfer of P53 gene into tumour cells.
It doesn’t replace genes that are there but swamps cells with functional P53 genes to make more good protein.
Restores the probability of having functional P53 in the ____ complex.
Used in trials for NSCLC (Swisher et al 2003) and LABC - shown to reduce tumour size and slows growth.
These trials often involved using A____ as an ____ to other chemotherapeutic agents.
This is because A_____ restores P53 function but other mutations in the cancer cell are likely to be present so need ways of targeting these.
E.g. in the LABC study - ______ and docetaxel were used as chemotherapeutic agents)

A____ was shown to be relatively well ____ in trials and multiple daily dosing regimes were feasible.

Answer 22

Advexin 
Adenoviral transfer 
Tetramer 
Advexin 
Adjuvant 
Advexin 
Doxorubicin 
Advexin 
Tolerated