Edmead Lec 3 Flashcards
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.
Restrict
Recessive
2
Dominant
Sporadic mutations are in the ______
Germline mutations are _____
Individual
Inherited
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.
Tumour suppressor Point Dysfunctional Point Premature Degraded Base
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
TGFB (inhibitory signals dampen down cell growth)
Checkpoint e.g. pRb, p53
CDKI, P16
Bax, P53 (to kill off cells with mutations)
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.
Breast
Truncated
Double strand break repair
Destabilised
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.
G1 to S phase
Proliferation and differentiation
E2F
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.
Cell cycle E2F and HDACs G1 to S phase Hypophosphorylated E2F and HDACs to sequester E2F HDACs E2F
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.
Cyclin D CDK4 pRb pRb E2F Cyclin D and CDK4 pRb E2F S G1
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.
Tumour
Low
MDM2
MDM2
Free P53 is ____ when unbound and is _____ quickly. But it is ____ by binding damaged ___.
Unstable
Degraded
Stabilised
DNA
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
Transcription Nucleus DNA Stabilise Degraded DNA Transcriptional
P53 binds as a _____. So requires _ molecules of P53 to associate together to act as a ______ factor to switch on gene ______.
Tetramer
4
Transcriptional
Transcription
Name 4 types of stress that can activate P53:
What does P53 do as a result of this? (5)
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.
P53 regulates the expression of:
___and___ - cyclin dependent kinase inhibitors
____ (inhibitor of P53 action)
___ (pro apoptotic)
P21 and P16
MDM2
BAX
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 _____.
50% Apoptosis Stable DNA Transcription
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
1 50% DNA 4 50 Tetramer
____ 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.
MDM2 Phosphorylated MDM2 Tetramer Tetramer
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.
60
G to T transversions (point mutations)
BAX
Fungal
G to T transversion
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.
Left Right MDM2 HPV Tetramerisation Transcription factor
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
Degradation
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!
Apoptosis
Apoptotic
Resistant
P53
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.
Advexin Adenoviral transfer Tetramer Advexin Adjuvant Advexin Doxorubicin Advexin Tolerated
