Flashcards in Cancer genomics Deck (128):
What are the 2 next gene sequencing technologies likely to come into mainstream use?
Both are too expensive atm (but will get competitive soon). They can read up to 10kb but make mistakes. This is in contrast to Illumina which can read 150 bases but is more accurate. Long reads with mistakes would help string sequences together.
How does exome sequencing work?
Break up genomic DNA, hybridise with bits you're interested (e.g. exome) and pull down. Sequence these bits. Lose some information but is cheaper.
How does Illumina sequencing work?
Error rate is 1/100 or 1/1000 depending on location in genome. Do paired end sequencing (sequence from both ends) of 250-500bp fragments of DNA. Get approx. 150bp in from each end. Then align to a reference.
What are the problems with sequence alignment in Illumina?
If there are too many repeats/errors in the sequence it can't be lined up
If there are many mutations it won't align perfectly - go for where it is closest but may be in wrong place or won't align at all.
If there are translocations, they will be found if one end of the fragment aligns with chromosome a and the other aligns with chromosome b. Have to allow for mismatches however as there is a high error rate in this process - misalignment (where there is homology in reference genome on another chromosome) is as common as a translocation. Can also get translocations when repairing fragments if they join with a piece from another chromosome.
LOTS of noise in translocations, need to sequence many fragments
How can misalignments in Illumina sequencing be verified?
Can't do by Sanger as is too expensive. Look at agreement between labs using the same data -- 80-90% agreement for single base mutations; 30-40% for INDELs. Due to use of different software.
Artefacts of the sequencing can lead to findings of 'translocations' in cancer which are false.
How can cancer mutations be found by non-genome methods?
Historically looked in retroviruses that cause tumours in animals. If retroviruses are selected for efficiency, many had acquired an oncogene which could be confirmed by sequencing the virus
Can transfect oncogenes into cells and look for phenotype
How can cancer mutations be found by genome-wide methods?
Hereditary predisposition (map, find location, find tumour w/ deletion and sequence) - helped find tumour suppressors
Cytogenetics (oncogenes in translocations)
Losses by cytogenetics and loss of heterozygosity (looking for common deletions in cancer)
Sanger sequencing screens (e.g. MAPK screen found BRAF)
What are the advantages and disadvantages of using cytogenetics to look for chromosome translocations?
Good in leukaemia as there are few translocations
Not good in epithelial cancers where there are too many translocations. Improved a bit with FISH but still don't know what parts of the chromomse have swapped so can't tell significance.
Also can't pick up small translocations e.g. TMPRSS2-ERG - TMPRSS2 gives promoter, ERG is a transcription factor. Is a small gene fusion
What has CGH arrays taught us about cancer genomics?
Comparative genome hybridisation arrays. Able to count the number of copies of a genome region in cancer (deletions and amplifications) - informs about copy number variants. Normal DNA is dyed green, tumour DNA is dyed red, they are hybridised and a signal is looked for. Doesn't take to gene level - looking at 100kb areas at best. Has been superseded by whole genome sequencing - count reads at each location.
What have Sanger sequencing screens taught us about cancer genomics?
PCR exon by exon and sequence and look for statistically significant results. Can look for candidate screens e.g. hypothesis beta-catenin could be an oncogene and go look for mutations. Also, screens for the MAPK pathway have been done.
What is the significance of TTC28?
Has a mobile element in an intron and therefore is translocated all over the genome, with the same sequence with in 4kb. This is found a lot in cancers as non-LTR LINEs can be activated. LINE is copied and takes part of TTC28 with it.
How does LINE1 move around the genome?
mRNA is transcribed and translated to make reverse transcriptase. Get DNA copied from the mRNA. In the case of TTC28, the mRNA extends into TTC28 and is therefore carried when it is reverse transcribed. L1 is polymorphic and not unique so can't align and is therefore not detected.
What is the current knowledge of cancer genetics?
Point mutations in exons - high reliability, 50% sensitivity
Indels in exons - less than 50% reliability
Small mutations in non-coding sequences (promoters, enhancers) - little known
Structural rearrangements - few from sequencing, well known ones for cytogenetics/CGH
Mobile element insertions - only just discovered
Epigenetics - poor
What are the different levels of transcription regulation?
DNA state (histone code, DNA methylation, chromatin structure)
Protein post translational modifications
RNA Pol II mediated gene expression (near promoters)
Describe the histone modifications found at enhancers and promoters?
Are distinct from each other
Enhancers: mono/dimethylation of H3K4 (not tri, required for enhancer function)
Promoters: trimethylation of H3K4 for activation. Also acetylation. Methylation at K9 and K27 is repressive.
How can enhancers be identified?
Tend to be in euchromatin. Can be identified using DNase I hypersensitivity. Embryonic stem cells have ~3% of the genome open (could be an enhancer). These close as cells differentiate.
What is special about nuclear receptors?
Are the only transcription factors that can be constituently switched on by a ligand.
Describe the structure of nuclear receptors
Have a variable section at the N terminus, then a DNA binding domain consisting of 2 zinc finger domains, then a ligand binding domain.
How can nuclear receptors be drugged?
The ligand binding domain is unique and therefore druggable
How do nuclear receptors act in cancer?
Oestrogen-ER causes breast cancer (75%)
Androgen-AR causes prostate cancer (100%)
Are key targets in therapy and provide models to learn about transcription and cell growth.
How do cofactors work with nuclear receptors?
A very complex interaction involving many co-factors. Varies between cell type - not all co-factors function in all cells. Can edit histone modifications e.g. CBBP/p300 is tethered to the nuclear receptor by SRC-1 and others and they can acetylate histones. Cofactors are commonly mutated in cancer.
How can the binding points for ER be found in an unbiased way?
Use Illumina to find the DNA bound to the transcription factor. Pull down ER and sequence, remove noise and look. Is an unbiased method that is reproducible. Found that most binding sites were in the middle of nowhere.
What did the discovery of where ER binds tell us?
Found that ER mostly bound in the middle of nowhere. Thought to not be random as DNA was DNase I hypersensitive and had histone markers (H3K4me1/2). Also seemed to have a cis regulatory element (the oestrogen responsive element or the forkhead TF binding motif. for a forkhead protein).
What is FoxA1?
Is a forkhead box protein. Is a pioneer factor that binds forkhead motifs at tightly wound DNA (not heterochromatin) and opens it up to allow other transcription factors to bind.
How was it found which ER binding site activated which gene?
Hypothesised chromatin looping. Did chromosome conformation capture and ChIA-PET - fix with formaldehyde, cut with restriction enzymes and religate. Then pull down and sequence. Found flexible loops covering the whole genome, but no inter chromosome interactions. Found that anchor genes are brought to a hub of ER and FoxA1 and those looped out are silenced. TFF1 and TFF2 are activated by oestrogen and ER binds the promoter (one of the only examples)
What is GRO-seq?
Take all transcriptions, biotinylate and pull down then sequence. Is an unbiased way of looking at all transcripts and found that intervening DNA between genes is super active
How did GRO-seq inform us about ER activity?
27% of whole genome is transcribed when ER is active. Includes enhancer RNAs along with ncRNAs, antisense transcripts, divergent transcripts etc
What do eRNAs do?
Thought to aid chromatin looping - forming bridges between loops. If they are silenced get loss of cell growth and abnormal gene expression.
How is the TMPRSS2-ERG fusion associated with nuclear receptors?
TMPRSS2 is a gene controlled by the androgen receptor. When this fusion takes place, the TMPRSS2 promoter is fused to an oncogenic transcription factor (ERG) and expression is activated by androgen. There are also other examples of fusions of ETS factor and AR target genes
How can DNA properties be drugged in cancer?
5-aza-cytodine is a DNA demethylation agent (need to control dosage)
G-quadruplexes may be able to be targeted by inhibitors
How can transcription/pioneer factors be drugged in cancer?
Could regulate other transcription factors as a decoy or block protein-protein interfaces (hard to get into the cell)
How can co-factors and associated proteins that are mutated in cancer be drugged?
AIB1 is a breast co-factor where a drug has been found
IBET inhibit BRD4 co-factors
What factors increase the risk of breast cancer?
Factors that increase oestrogen exposure
More periods in your life (early menarche, late menopause)
No children (lowers oestrogen)
Hormone replacement therapy
High BMI (oestrogen metabolised from fat)
What is the normal function for nuclear receptors?
Required for tissue development e.g. ER mediates cell growth for mammary gland development, AR mediates cell growth for prostate development, glucocorticoids, progesterone, prolactin etc are all important for normal physiology.
How does ER cause cell growth in breast cancer?
Oestrogen diffuses in and binds ER dimers in the cytoplasm. These then translocate to the nucleus where they bind DNA and up regulate transcription. AR works in exactly the same way for prostate cancer
How does Tamoxifen affect ER activity?
Can bind ER dimers instead of oestrogen and promote translocation into the nucleus. ER dimers bind DNA but transcription isn't up regulated (is not active.
How does aromatase affect ER activity?
Oestrogen is metabolised from precursors by aromatase. If aromatase is inhibited, it limits the amount of oestrogen that can form (increases hot flushes etc)
How does anti androgens affect AR activity?
Bind AR dimers in the cytoplasm and allow translation to nucleus. AR dimers sit on DNA but don't up regulate transcription - are inactive. Examples of new effective agents are abiraterone, enzalutamide.
What did we learn from mapping AR binding sites?
Loads of AR binding events, like ER, in prostate cancer cells; very few promoters bound.
Forkhead motifs were also enriched
GATA motifs were enriched
How does blocking FoxA1 binding affect ER binding?
No ER binding at all; no gene expression, no cell growth
How does blocking FoxA1 binding affect AR binding?
No AR binding at the original sites, but AR goes and binds new sites. Cell still stops growing, so the movement has no effect. FoxA1 and FoxA2 is present in prostate (only FoxA1 in breast)
How is FoxA1 associated with breast cancer outcome?
FoxA1 is found in all ER+ breast cancers. Can be found in all primary tumours and in metastases.
ER can move around the genome, dictated by FoxA1. This has an influence on cell fate and survival rate.
How does drug resistance arise in ER brest cancer?
Phosphorylation of ER, ER mutations to prevent drug binding (in ligand binding domain). Often found in metastasis.
Why is FoxA1 a good drug target?
Is absolutely necessary for ER+ cancers, even in drug resistant cells. Also needed in AR+ cancers and castrate resistant prostate cancer.
Can we drug FoxA1?
Maybe. Has a restricted tissue expression profile
How do prostate cancers become drug resistant?
Select splicing variants with different ligand binding
What are molecular apocrine breast cancers?
Are ER negative but have other hallmarks of ER positive cancers. AR can substitute for ER. Have different peaks in a Chip-seq to prostate cancer, but is very similar to ER breast cancer. AR can substitute for ER due to FoxA1 which dictates binding points - AR can go to ER binding points. Binding points have a higher affinity for ER, but if there is no ER get AR to drive genes.
AR is in all ER+ breast cancers but silenced (unless no ER)
How can molecular apocrine cancers be treated?
If women don't respond to tamoxifen, can give same drugs as prostate cancer which work well. However, if give these drugs to an ER+ cancer, it makes the cancer worse.
How does progesterone interact with the ER pathway?
PR blocks ER binding and activity, leading to 'good' transcriptional outcomes. 20% of ER+ cancers lose PR. This occurs through a strong interaction of PR with ER and sequestering of ER. Suggested that progesterone is protective - strong correlation between levels of progesterone and age of diagnosis (as decrease get increase).
What is a new way of treating ER+ cancers?
Giving progestins with ER inhibitors such as tamoxifen seems to be a potent way of treating cancer. This is in clinical trails now.
What is the problem with treating cancers with progestins?
In mice, found that progestins encouraged proliferation in mouse mammary glands. This was extrapolated to say that progestins are pro-proliferative in human cancers when in fact the opposite is true. Also have data from HRT which showed that progestins gave a small increase in proliferation - due to a slight lack of specificity to PR (hits other nuclear receptors too). In France, use of progesterone (as opposed to progestins) has given protection from cancer
What machineries modify chromatin?
Histone modification/histone variants
ncRNA and relation with expression (can interact with transcription factors)
What is EZH2?
Part of the polychrome repressive complex 2 (PRC2) which works to silence gene expression. Is an enzyme with a SET domain that adds methyl groups to H2K27 (up to 3 methylations) which switches off transcription.
How can EZH2 be mutated in cancer?
In non-Hodgkin lymphoma: In SET domain, Y641A creates more di/trimethylation. Therefore is an attractive target for drugs - GSK126 inhibits growth of cancers with this mutation.
Also get inactivating mutations in some cancers e.t. T-cell acute lymphoblastic leukaemia
How does 5' methylation of cytosine alter gene expression?
Can recruit CpG DNA binding proteins
Could block DNA binding proteins
Could change DNA charge and alter the architecture
How is DNA methylation affected in cancers?
Disturbed in 75% of cancers. Global hypomethylation with some hypermethylation at specific loci to silence certain genes e.g. TP53.
How does epigenetics have an effect on genomic instability?
Changes in DNA methylation gives genomic instability, potentially due to recombination or lack of repression of transposable elements
Where is DNA hypomethylated in cancer?
Oncogenes, DNA satellites, repetitive elements.
How can DNA methylation be used as a diagnostic tool?
Can be used to predict prognosis. Observe hypomethylation of LINE elements and can be used to guide drug targets by looking at methylation of certain genes. Can see DNA hypomethylation in bodily fluids - could be used to diagnose cancers
What external factors influence DNA methylation?
Age - genome tends to become hypomethylated but CpG islands are hypermethlated (same as in cancer, could be linked to increase cancer incidence)
Diet - S-adenosyl methione is substrate for methylation, gained through folate and methionine pathway
Environment - arsenic (hypomethylates Ras gene) and cadmium (global hypomethylation by DNMT1 inactivation) associated with cancer
How can DNA methylation be detected?
Bisulphide addition. Convert mC to uracil, sequence and compare. Can't distinguish between mC and hmC which is part of the removal mechanism - could influence how correct the literature is
How is DNA methylation involved in ovarian cancer?
Hypermethylation of tumour suppressor genes e.g. those involved in DNA mismatch repair (e.g. BRCA1, hMLH1)
Hypomethylation of LINE elements (correlates with advanced cancer)
Hypomethylation of TUBB3 - a determinant of taxane resistance
What histone modifications are associated with cancer?
H3K4me2/3 - activating
H3K27me3 at silenced genes
How does EZH2 expression affect expression of E cadherin?
EZH2 is super active in cancers and adds the histone modification H3K27 to silence genes. This occurs at E cadherin. E cadherin normally mediate cell-cell adhesion and keeps cells together and prevents moving. If it is inhibited, this allows metastasis. Found increase in cell movement in a cell invasion assay when EZH2 was over expressed and E-cadherin was silenced.
How are BET proteins involved inn cancer?
Bind acetylation on histones, involved activation and transcription. In rare leukaemia's, BET proteins are misplaced and get aberrant activation
How can BET proteins be inhibited?
JQ1 is a drug which binds in the pocket to block target binding. However has a short half life and causes male sterility.
I-BET762 is more promising and can also be used in solid tumours
What is Vorinostat?
A drug against HDACs. Works on the cancer stem cell hypothesis - the cancer transcriptome is rewritten to look like a stem cell. In aggressive cancers get a shift in gene expression profile along with change of DNA methylation and histone markers.
What is GSKJ4?
A drug which restores methylation at H3K27. Used to treat diffuse intrinsic pontine glioma in which there is hypomethylation of H3K27 leading to a more accessible chromatin state and aberrant gene expression. Treatment with GSKJ4 restores methylation and causes tumour shrinkage.
How do mutations of ncRNAs affect cancer?
Deletion of 13q14 region encodes miR-15/16 which regulate apoptosis. Found in chronic lymphocytic leukaemia
Amplification of miR-17-92 found in diffuse large B cell lymphoma patients
CNV in some miRNAs confers chemo/radioresistance
Also have epigenetic control and processing defects
What does mi-R10b do in cancer?
Is involved in breast cancer metastasis
What does miR-31 do in cancer?
Deregulated in breast cancer, get leakage into the blood stream. Could be used as a diagnostic marker and prognosis
What is the difference between siRNAs and miRNAs?
siRNAs need a perfect match (1 gene target)
miRNAs don't need a perfect match and can therefore have many gene targets and wide spread effects. May have a seed sequence.
What does miR-132 do in cancer?
Promoter of miR-132 is hypermethylated in prostate cancer. Silencing of miR-132 leads to increased cell death and increased invasion and migration. Targets HB-EGF and TALIN2. Is silenced by EZH2 in ovarian cancer.
Also get a metabolic switch and increased GLUT1 expression when silenced.
What is HOTAIR?
A long non coding RNA. Interacts with PRC2; is necessary for PRC2 occupancy at H3K27me3 at the HoxD locus and MYC in breast cancer.
What is MEG3?
A lncRNA. Stimulates interaction between JARID2 and PRC2 component EZH2 leading to recruitment of EZH2 to chromatin at the imprinted locus Dlk1-Dio3 and silencing.
Is a tumour suppressor - is associated with MDM2 expression and p53 accumulation.
What is MALAT1?
A lncRNA which can act as an oncogene. Is associated with metastasis in lung cancer and angiogenesis. Is up regulated indirectly through N-myc.
What is miR-101?
A microRNA that inhibits expression and function of EZH2. Expression of miR-101 decreases during cancer progression as EZH2 activity increases.
What is miR-31?
An miRNA which is repressed through EZH2 activity. Is lost in adult T cell leukaemia. Is lost n prostate cancer through promoter hypermethylation.
What are germ cell tumours (GCTs)?
Tumours that arise from primordial germ cells. Include testicular, ovarian, CNS and other locations.
Describe the histological heterogeneity of GCTs
Many different histologies, due to different pathways down the differentiation lines.
What miRNAs are unregulated in GCTs?
miR371-373 and miR-302/367. Found in both childhood and adult tumours. Have identical seed regions (nucleotides 2-7 which determine mRNA binding - rest don't have to be perfectly complimentary), so down regulate similar targets.
How can miRNAs be identified as passengers or drivers?
Look for a seed region - can look for in down regulated miRNAs then see if it is present in mRNAs that are down regulated. See what the mRNAs code for - signalling/biological processes.
In mouse embryos, high level of miRNAs are found, including miRNAs found in cancer. These are down regulated during differentiation as they can be oncogenic
How can miRNA be used as a biomarker for GCTs?
At the moment use levels of serum proteins - different for different GCTs and have varying levels. Therefore many CT scans are needed (expensive, radiation stress). Instead can look for circulating miRNA as these are less likely to be degraded than mRNAs (which are v long). GCTs have a low mutation rate (especially testicular), so looking for tumour DNA isn't good.
What are the advantages for using miRNAs as a biomarker?
Tumour cells release exosomes into the microenvironment and blood. miRNA levels in the serum are very stable and can be left at RT for a day with no change, and freeze-thawed. In a case study, found that miR-372 levels correlated with the protein marker normally used. Also correlation between miR-372 levels and tumour size has been found. Trials with 4 miRNAs have found high specificity and sensitivity - looks promising! Can track relapses as well
How can miRNA be used in diagnosis of brain tumours?
If the tumour is marker negative, would have to do a biopsy to determine if it malignant - this can upset sodium levels. Found that miRNA can be used as a marker and that it is proportional to tumour size. Even when levels have been normalised to tumour size still see significant increase.
How can miRNA be used in differentiating between 2 pituitary thickening diseases?
LCH or malignant GCT. If diameter is too small for a biopsy, can look at miRNA levels (if no other markers). Have found that if there are high miRNA levels can keep tracking the tumour and see growth - a neurosurgeon would then go in to biopsy/remove.
Why is it important to improve diagnosis of GCT?
Still high cure rates with delayed diagnosis, but has more chance of becoming metastatic and can affect memory and behaviour
What are the pitfalls in using miRNA to look for GCT?
Serum volume is dependent on size - if a small tumour in a big person have to look very carefully. May be better to look at spinal fluid
How can miRNA be used in diagnosis of other tumours that GCT?
Unclear. Many things can effect the results such as tube used to collect the sample, method of collection, same normalisation, spin speeds (aberrant results at high spin speeds). In GCT studies all of these have been the same
When does genomic recombination occur in normal development?
Meiosis for progeny fitness and evolution
What are the types of genomic instability?
Chromosomal instability (CIN) - structural (deletions, duplications, translocations) or numerical (aneuploidy/polyploidy)
Micro satellite instability - changes in number of repeats, can get double strand breaks
Base pair mutations
What causes genomic instability?
Failure of DNA repair
Failure of checkpoints
Failure of chromosomal segregation
What has Lynch syndrome taught us about genomic instability in cancer?
Is hereditary non-polyposis colon cancer. Found micro satellite instability and tracked this to mutations in mismatch repair genes. Same mutations are found in 15% of sporadic cancers
What has HBOC taught us about genomic instability in cancers?
Hereditary breast and ovarian cancer. Found mutation in BRCA1 and BRCA2, involved in DNA damage response and DNA repair (homologous recombination repair)
What do BRCA1 and BRCA2 do?
BRCA1 is part of the sensor complex for double strand breaks
BRCA2 is part of the repair complex, may be a scaffold protein.
If homologous recombination is defective get more non-homologous end joining which is much less stable.
How does homologous recombination work to repair double strand breaks?
Break is degraded to get an overhang. Single strand invades the other chromosome and anneals, get DNA replication induced by the break.
How do we know genomic instability in cancers isn't caused by rapid cycling?
Different tumours have different patterns of genomic instability - suggests there are specific mutations in certain repair pathways.
What mutations are found that cause failure of DNA repair?
In hereditary cancers (Lynch syndrome - MIN; HBOC - BRCA)
Very little found in sporadic cancers - mutations not often found in targeted studies. Suggested that a full mutation would be too much instability - there could be more subtle changes on an epigenetic/gene expression level.
What mutations are found that cause failure of checkpoints?
p53 deletions as a DNA damage checkpoint - suggests that this could cause genomic instability. However, still get precancerous lesions with genomic instability and normal p53.
What is oncogene induced DNA damage?
Suggestion that activated oncogenes and growth pathways could lead to genomic instability through DNA replication stress. Mechanism is unknown.
How do failures in chromosome segregation cause genomic instability?
If there are problems organising the spindle (e.g. multiple centrosomes) could get CIN and cancer. In 2010 found that 25% of genome is affected by arm/whole chromosome somatic copy number alterations.
If get incorrect segregation get polyploidy/aneuploidy
What is euploidy?
Normal number of chromosomes within a cell for a species
What is aneuploidy?
Having an abnormal chromosome number
What is polyploidy?
Having one or more extra sets of chromosome
How can cells gain or lose chromosomes?
Mitotic checkpoint defects - no division of sister chromatids at anaphase
Cohesion defects - premature loss of sister chromatid cohesion
Merotelic attachments - is increased if microtubule depolymerising kinesis 13 is inactivated (found in CIN)
Multipolar mitotic spindles
How do multipolar mitotic spindles cause genomic instability?
Can get division into more than 2 cells - cells tend to die by apoptosis as not enough chromosomes to survive
Or get clustering of 2 MTOCs at one of the poles and minor chromosome missegregation. Get more lagging chromosomes as there are more merotelic attachments.
How do extra centrosomes form?
Centrosome amplification (more than 1 daughter centrosome to each mother)
Fragmented pericentriolar matrix can nucleate microtubules and form more poles
Loss of centriole cohesion
Over expression o fPLK4 leading to increased number of centrosomes at subsequent divisions
How can cells become tetraploid?
Cell-cell fusion (viruses)
Mitotic slippage (prolonged arrest)
Endoreduplication (in megakaryocytic differentiation)
If there are 4 centrosomes in a tetraploid cell, get more lagging chromosomes than if only 2.
What are micronuclei?
Formed by mis-segregated chromosomes. Either persist as cell divides or are reincorporated into the nuclei. Undergo asynchronous DNA replication resulting in DNA damage and chromosome fragmentation - may explain chromothripis observations in some cancer cells
What is endoreduplication?
In which the cellular DNA is replicated but mitosis doesn't occur. Could be due to persistent DNA damage signalling such as telomere damage.
How can endoreduplication arise?
Combined loss of p53 and Rb can lead to endoreplication and polyploidy and proliferation
How can prolonged mitosis lead to DNA damage?
Depletion of the anti-apoptotic protein MCL1 causes activation of DNase CAD which cleaves decondensed DNA loops. Sister chromatid cohesion is gradually lost which promotes premature separation and merotelic attachment. These processes can lead to chromosome breaks.
How can lagging chromatin cause cytokinesis failure and DNA damage?
Can get stuck in bridge and causes abscission failure and bridge stabilisation
Can cause regression of the furrow and tetraploidy
Can get double strand break and chromatin cleavage
How is abscission during cytokinesis prevented in the presence of lagging chromatin?
Aurora B controls the activity of ESCRT-III to delay abscission in the presence of DNA
What is the evidence for aneuploidy facilitating tumourigenesis?
SAC-deficient mice with high levels of aneuploidy show an increase in carcinogen induced tumours. Also, over expression of some mitotic components can promote tumourigenesis in these mice.
What is the evidence that aneuploidy has no link to tumourigenesis?
Tumours only form in a fraction of aneuploid animals and humans - no direct correlation. SAC-deficient mice have high levels of aneuploidy but no increase in spontaneous tumourigenesis.
What is the evidence that aneuploidy suppresses tumourigenesis?
If there are mutations in tumour suppressor genes (p19, Rb, Apc) alongside aneuploidy get suppression of tumourigenesis
Does aneuploidy cause tumorigenesis?
Probably no. Thought that it may facilitate tumour development in the presence of other insults though. Can also suppress tumourigenesis in some circumstances - thought to be due to levels of genomic instability being higher than the threshold compatible with cell viability.
What is the evidence for polyploidy facilitating tumorigenesis?
in p53 deficient tetraploid mice mammary epithelial cells get CIN (structural and numerical). These cause tumours in nude mice when transplanted in
A midbody kelch protein for cytokinesis is associated with Hodgkins lymphoma and binucleate cells
In the progression from Barretts oesophagus to adenocarcinoma can do a normal Vogelstein model OR lose p53 then get genome doubling
Tetraploid cells display more CIN
What is the chromosomal translocation associated with Hodgkins lymphoma?
Reciprocal between chromosome 2 and 3, break point in the middle of the KLHDC8B gene - depletions in this gene have shown to give failure to cytokinesis and binucleate cells
How can oesophageal adenocarcinoma form from Barretts oesophagus?
Comparing cells before and after tumourigenesis. Either see a traditional Vogelstein method in which oncogenes accumulate OR a loss of p53 and then gene doubling events.
What is the theory as to why polyploidy may facilitate tumourigenesis?
Can cope with more mitotic errors and chromosomal instability. Can also cope better with chemotherapeutic drugs (seen in tetraploid cells in culture)
Is associated with poor prognosis in colon cancers
What is the evidence for supernumerary centrosomes facilitating carcinogenesis?
Centrosome amplification by over expressing SAK/Pik4 promotes tumourigenesis in drosophila
Mutation of genes required for centrosome formation can promote tumorigenesis
Centrosome amplification (if with an inducible promoter) can promote tumorigenesis in the presence or absence of p53
Centrosome amplification can induce oncogene like cellular invasion
What is the evidence against supernumerary centrosomes facilitating carcinogenesis
2 contradicting papers - one found that over expression of Plk4 didn't cause tumours even with lack of p53 or treatment with carcinogens. Other found that induced Plk4 over expression did promote tumorigenesis with or without p53. Second author used an inducible promoter - potentially suggesting that a bit of extra centrosomes are tumour permissive but too much is detrimental.
How was it shown that centrosome amplification by over expression of SAK/PIk4l promotes tumorigenesis in drosophila?
Neuroblasts with extra centrosomes form bipolar spindles during mitosis
Higher levels of aneuploidy in SAK over expressed cells
Failure to divide asymmetrically in neuroblasts, suggesting that tumour formation could be caused by impaired asymmetric cel division and stem cell over proliferation
How was it shown that mutation of genes required for centrosome formation can promote tumorigenesis in drosophila?
SAK or dsas-4 mutations promoted tumorigenesis in brain tissue - not in other somatic tissues though in which cells were dividing symmetrically. Perhaps need impaired asymmetric cell division to see this - get asymmetric cell division in the stem cells of the brain
Cells have chromosomal instability (aneuploidy)
How can centrosome amplification induce oncogene like cellular invasion?
Activates Rac-1 and disrupts cell-cell adhesion