Cancer Flashcards
What is a tumor suppressor gene
Tumor suppressors normally function to control cell growth and proliferation LOF mutations contribute to the abnormal proliferation of cancer cells
What is an oncogene
• A gene that normally is involved in controlling cellular proliferation.
• When altered/over-activated, oncogenes can help transform normal cells into tumour cells by promoting uncontrolled cell growth.
Associated with GOF mutations
What are the main categories of TSG
Gate Keeper TSGs- control cell cycle progression e.g. TP53, cyclins and CDKs. e.g. mutation of TP53 (normally inhibits cell cycle progression) is mutated in 50% of tumours
Caretaker TSGs- maintain the fidelity of the genome by repairing damage e.g BRCA1/2 (HR), MSH2, MSH6, MLH1, PMS2 (MMR), MUTYH (BER) etc
How do TSGs restrain cell growth
- Inhibit the cell cycle (cyclins, CDKs, RB, TP53, APC)
- Apoptosis (TP53)
- Repair damage
Give 2 examples of TSGs
RB1
TP53
What is the Knudson hypothesis?
The Knudson hypothesis is derived from the genetic mechanisms underlying RB1 (first TSG discovered)
Explained the differences between hereditary (early onset bilateral retinoblastoma with risk of cancer in other tissues) and sporadic RB (usually unilateral, later onset).
2 hit hypothesis in which both alleles of a TSG must be lost to develop cancer. In hereditary cancer one copy of the gene is already KO so only a single additional acquired mutation is required resulting in earlier onset. this results in variable penetrate and apparently AD inheritance
Describe Retinoblastoma
early onset (<5yrs) aggressive childhood cancer of the eye. Characterised by whitening of the pupil.
Can be unilateral (usually sporadic) or bilateral (usually familial + increased risk of soft tissue and bone cancers)
Mutation spectrum includes SNVs, CNVs, SVs and hypermethylation of the RB1 promoter (10%). 60-70% display LOH of 1 allele with a mutation in the other allele. truncating mutations and deletions associated with almost complete penetrance whereas missense and splice site can have reduced penetrance and variable expressivity
What is the role of the RB1 gene
RB1 is found at 13q14
Key role in G1/S phase cell cycle checkpoint
nuclear phosphoprotein which is involved in cell cycle progression. When it is unphosphorlylated it bind E2F transcription factor preventing it entering the nucleus to activate transcription of target genes= cell cycle repression
When phosphorylated by cylcin D CDK 4/6 it dissociates from E2F which can then activate transcription of target genes (cylin E) and the cell can progress from G1 to S phase.
Describe p53’s role as a TSG
second TSG to be discovered
mutated in 50% of tumours
essential in multiple signalling pathways associated with cell cycle, apoptosis and DNA repair
How does p53 contribute to cell cycle control
p53 can act as a transcription factor to activate the transcription of genes associated with cell cycle arrest and apoptosis.
In normal cells it is bound to MDM2 which retains it in the cytoplasm. (MDM2 required phosphorylation to migrate to the nucleus and bind to p53 and cause it to migrate to the cytoplasm) in the cytoplasm p53 is degraded by the ubiquitin/pretoeosom pathway.
in response to stress p53 is phosphorylated and acetylated = it can dissociate from MDM2 and activate transcription of genes e.g. PUMA which controls apoptosis.
Methods for loss of p53 function
- mutations to upstream genes e.g. ATM or CHCK2
- mutations in p53 (~50% of tumours)
p53 mutations can be LOF or GOF
-mutations in gene that act downstream of p53 e.g. PTEN (germline mutations associated with Cowdens syndrome)
Describe the role of CDKN2A in the cell cycle.
CDKN2a encodes 2 unrelated proteins:
p16 INK4a- this inhibit CDK4/6 keeping RB1 dephosphorylated and bound to E2F = inhibits cell cycle
p14ARF- destabilises the interaction between MDM2 and p53 = p53 active resulting in cell cycle arrest
Germline mutations in CDKN2a is associated with malignant melanoma (penetrance depends on age and sun exposure)
Describe 2 examples of miRNAs that can function as TSGs
- Let-7
normally expressed in differentiated tissues but lost in NSCLC
negatively regulates cell cycle oncogenes and exogenous application to human lung caner cell reduces proliferation - miR-34 family
Lost in lung cancer
expression activated by p53 and associated with apoptosis
Name 3 additional TSGs, their molecular function and associated cancer susceptibility syndrome
- BRCA1/2- DSB repairr and HR (HBOC)
- Lynch syndrome- MMR (CRC)
- APC- negative regulator of b-catenin. It is part of the B-catenin complex and targets it for ubiquitin mediated degradation. (FAP)
What are the 5 broad categories of oncogenes?
- secreted growth factors
- growth factor receptors
- Signal transducers
- Inhibitors of apoptosis
- Transcription factors
Give an example of a secreted growth factor acting as an oncogene
secreted growth factors can act as an oncogene due to the constitutive activation of a growth factor gene e.g. normal wnt/b-catenin signalling is involved in embryonic development whereas over activation of the pathway is involved in many cancers including breast cancer.
Give an example of a growth factor receptor acting as an oncogene
e.g. EGFR and RET
EGFR receptor is constitutively expressed in NSCLC
- activating mutations occur in exons 18, 19 and 12. The gene encodes a RTK and mutation results in constitutive activation and over activation of downstream pathways
- Activating mutations can result in dependency for the cancerous cell on aberrant EGFR signalling
- Can be targeted by specific anti-EGFR therapies
- mutations can occur in the ATP binding pocket which reduce the affinity for ATP and increase the sensitivity to the RTK which competes with ATP for binding.
- Resistance mutations can occur in the catalytic domain which weaken the interaction between the inhibitor and its target
Give an example of a secreted signal transducer acting as an oncogene
PI3KA
- calls 1 signal transducer
- composed of a heterodimer of a catalytic and regulatory subunit and results in phosphorylation of phosphatidyl inositol lipids.
- plays a role in cell motabilism, motility and cell cycle regulation
- transmits signals from RTKs and GPCRs
- 13% have PI3KA mutations with a hotspot in the kinase domain
- RTK phosphorylates and activates PI3KA
- PI3KA phosphorlyates the inositol ring of PIP2 and converts it to PIP3
(PIP3 can also be directly activated by RAS by biding to the catalytic domain) - PIP3 is active and can directly bind and activated proteins with a pleckstrin homology domain e.g. PDPK1 and AKT which results in cell growth and proliferation.
PTEN tightly regulate PIP3 in normal cells by dephosphorylation to the inactive PIP2
Give an example of a an oncogene which inhibits apoptosis
BCL2
cytoplasmic protein which localises to the mitochondria and inhibits apoptosis.
overexpressed in most follicular lymphoma
t(14;18) BCL2-IGH rearangement results in BCL2 being under the control of the IGH locus
Give an example of a transcription factor acting as an oncogene
EWS-FLI1 rearrangement in Ewings sarcomma
found in soft tissue and bone cancers
results in a fusion between the FLI1 gene resulting in an aberrant transcription factor = aberrant regulation of growth and cell proliferation
What are the main mutation mechanisms for activating an oncogene (GOF)
Point mutations- can constiutively activate a signalling pathway e.g. mutation to the regulatory domain or dimerisation domain. Results in hyperactivation of a protein that is expressed in normal amounts
Amplification- overexpression of a non-mutated gene leading to excess protein e.g. Her-2 in BC or MYCN in NB
Translocation to produce a novel fusion gene- aberrant or dysregulated function. e.g. BCR-ABL in CML
Translocation into a transcriptionally active region e.g. Burkitts Lymphoma. t(8;14)(q24;q32) is seen in 75% of patients and results in juxtaposition of the MYC oncogene with an immunoglobulin (IG) locus and consequently, MYC is brought under transcriptional control of the IG locus at the same time as losing its own. Other Ig locus rearrangements are also common in cancer.
Local DNA rearangements- fusion genes can be created by inversion or deletion of the intervening region between a gene e.g. Inv(16) in AML
Insertional mutagenesis- Hepatitis B in hepatocellular carcinoma. Viral oncogenes insert near cellular genes such as MYC and aberrantly activate it to initiate unchecked cellular proliferation.
what is tumour mutation burden (TMB) testing?
it is a measurement of the total number of non synonymous mutations in the tumour exome
What is the clinical utility of TMB?
can be used as a biomarker for immunotherapy, especially for the use of immune checkpoint inhibitors.
how does TMB influence immune checkpoint inhibitors?
somatic mutations can result in the expression of neoantigens and the chance is greater the higher the TMB
The neoantigens can activate the proliferation of T-cells which act to kill the cancer cells and such tumours can be targeted with immune checkpoint inhibitors-
How do checkpoint inhibitors work
these work by blocking the binding of checkpoint proteins to their partners
prevent the off signal from being sent so T-cells kill the cancer cells.
ICIs show variable efficicy and biomarkers help stratfiy patients that will respond
- MSI high tumours also show good response to ICIs as there is a high TMB
high TMB correlates with a good response to ICIs and increased survival in some cancers including NSCLC
response is not consistent across different cancer types and there is no established threshold or methodology for TMB
What tests can be used to determine TMB?
WES/WGS panels- WES considered best but not as cheap or fast as targeted panels- targeted panels are used to exptrapolate the number of somatic coding mutations observed in the targeted genomic space to the no. that would be observed across the whole genome. tumour percentage, qualit and seq depth all influence the number of somatic mutations detected.
standardization is required to fully assess and implement TMB as a biomarker in the UK
what does ‘actionable’ mean in terms of variant interp
variant can be used a biomarker fot he patients disease
what is a biomarker?
a marker of disease that can provide info that is useful for the diagnosis, prognosis or treatment of a patient
What are the current UL guidelines for somatic variant interpretation
ACMG and ACGS BPG for variant classification state that a different set of interpretation guidelines is needed for somatic variants
2017 American association for molecular pathology released guidance and a working group has been established in the UK
what are the differences between somatic and germline variant interpretation
constitutional-
homogeneous, allele fraction 0.5 or 1 (unless mosaic), low AF can generally be discounted (be wary of mosaic),
family studies are possible
Is the variant causative of phenotype?
germ line- tumour heterogeneity variable allele fraction low AF- genuine or seq artifact cannot use linkage Is the variant a driver of tumourigenesis, useful for tumour classification, have prognostic implications, target-able with a drug?
describe the difference stages of the cells cycle
G1= growth 1. RNA and proteins are synthesised (not DNA). each chromosome exits as a double helix
S- DNA synthesis- each chromosome is now present as sister chromatids
G2- cell continues to grow
M- mitosis- cell stops growing and there is nuclear division (miosis) followed by cellular division (cytokinesis) to produce to daughter cells.
Go= senescence the cell has left the cell cycle and stopped dividing. the cell may re-join the cell cycle in response to specific signals.
How id the cell cycle controlled?
cell cycle checkpoints
