MOD 1 Cancer Bio Flashcards Preview

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Flashcards in MOD 1 Cancer Bio Deck (58):

Carcinogenesis causes

Physical (Ionising Radiation, Asbestos) – Radiation affects cells by directly damaging DNA

Chemical (Cigarette Smoke, Industrial Dyes, ETOH, burnt food) – often found bound to DNA, and likely to damage DNA directly

- HPV16/18 - cervical cancer
- HHV8 (Human Herpes Virus 8) - Kaposi’s sarcoma
- EBV – variety of different malignancies associated
- HCV – Hepatocellular carcinoma

Chronic Inflammation E.g. H pylori and GI cancer


What is a Mutagen?

Any chemical that has the ability to cause mutations in DNA


Classification of cancer causing viruses?

Cancer causing viruses can be either acutely transforming or slowly transforming:
o Acutely transforming means the virus carries an oncogene which is immediately activated once inside the cell – e.g. HPV virus
o Slowly transforming means the virus genome is inserted near a proto-oncogene, and the virus promotor region causes over-expression of that proto-oncogene – e.g. HCV


Apoptosis: Intrinsic

Caspase 9
Cell stress --> Cyt C into cytoplasm
Apoptosome complex: Apaf1, Procaspase 9, ATP, Cyt C
Forms heptamer - each Apaf1 can bind 1 procaspase 9
Therefore 7 procaspase 9 molecules cleaved and activated



Regulates Intrinsic pathway:
Inhibits dimerisation of procaspase 9


BCL-2 family

Regulates Intrinsic pathway by altering mit membrane permeability
Pro-apoptotic: BH3 only, Bax, Bak
Ant-apoptotic: Bcl-2-like


Apoptosis: Extrinsic

Caspase 8
Two adaptor proteins:
- FADD (positive regulator) --> Binds to Fas receptor, Procaspase 8 binds --> DISC

- FLIP (negative regulator) - Cellular Flice-like inhibitory protein

N.b. Receptor mediated pathway soon activates intrinsic pathway anyway via activated Bid that enhances Cyt C release


Other mechanisms of cell death

Necrosis – Death due to injury by Toxins or Pathogens; Associated with Immune response and occurs in regulated, genetic manner, regulated e.g. by RIPK1 and RIPK3

Autophagy – Catabolic, Self-degradation Process

Mitotic Catastrophe (Triggered by Aberrant Mitosis)

Anoikis – “Homelessness” - Response to adherent cells due to loss of Cell Cell/Matrix Interactions

Entosis – Engulfed cells which are degraded by the Lysosome

Pyroptosis – Caspase-1 Dependent Cell Death


Role of EMT transcription factors?

- Loss of Apico-basal polarity
- Enable the conversion from an epithelial to a spindle-like/fibroblastic morphology
- Directly inhibit expression of adherins junctions and tight junctions
- Increase expression of matrix degrading enzymes (MMPs)
- Increased motility
- Heightened resistance to apoptosis


What drives tumour metastasis?

Genomic Instability:
o Point mutations and chromosomal aberration leads to karyotype abnormalities and abnormal gene products
o Dysregulation of multiple downstream biological events is involved in tumour initiation and progression

However, not sufficient to explain rapid and reversible ability to differentiate...

Non-Genomic Instability:
o Phenotypic changes not genetic in origin
o Response to regulatory/biochemical perturbations (e.g. growth factors, hypoxia, chemotherapy, tumour microenvironment) – switches between cellular states
o Activation of transcription factors or signalling pathways
o RNA processing and translational control
o Post-transcriptional and post-translational modification

Non-genetic plasticity also occurs in somatic cells; Clonally identical but trans-differentiation which occurs in developmental pathways determines differentiation.
- Allows reactivation of developmental signal in non-development contexts i.e. cancer


Mesenchymal vs Amoeboid migration

- Occurs in Dense ECM
- Actin Polymerisation and formation of Stress Fibres, Focal Adhesion Complexes, Secretion of MMP
-Formation of the Invadopodia

Examples: Fibrosarcoma, glioblastoma, epithelial cancers

- Occurs in Sparse ECM
- Cortical Actomyosin Contraction
- Use of Hydrostatic Pressure

Examples: Lymphoma, small-cell lung cancer, small-cell prostate cancer


What is anoikis resistance

Anoikis – programmed cell death induced upon cell detachment from ECM
o Resistance to anoikis occurs through Integrin switching and activation of anti-apoptotic signalling pathways (AKT/PI3K)
This therefore allows for survival of circulating tumour cells


Soil and Seed vs Mechanical Entrapment

- Cells can only metastasise only in location biochemically and physiologically favourable for implantation and growth
- Metastasis formation is due to mechanical and circulatory factors; First organ encountered by CTC would be site of greatest tumour cell arrest and formation of Metastatic colonies


Pharm control of metastases

- Prevention formation of Focal Adhesion Complex – FAK Inhibitors
- Preventing Epithelial Mesenchymal Transformation – c-Met Inhibitors
- Blocking ECM interactions – MMP Inhibitors
- Anti-Angiogenic Therapy – Bevacizumab (=Avastin, Anti VEGF Monoclonal ab);
o NB: Complete eradication of tumour vasculature prevents chemotherapy and radiotherapy from working


Limitations of current anti-metastases therapies

Current therapy fails to account for potential molecular and proliferative differences between different sub-populations of tumour cells
Could explain why majority of treatment fails in metastatic disease
“Selection” for resistant cells; Relapse is increasingly resistant
Concept of cancer stem cells:
o Indefinite proliferation
o Tumourigenic population
o Able to self-renew and differentiate into cells with non-stem cell characteristics


Erlotinib - effect and relevance?


- TK inhibitor against EGFR receptor
- Initial response to the drug was disappointing
o However a small subset of patients (10 to 20%) with non small cell lung cancer (NSCLC) responded well to EGFR kinase inhibitors
o Those patients who responded well had an oncogenic mutation of the EGF receptor


Oestrogens as risk factors for breast cancer - examples?

- HRT and OCP increase risk
- Plasma levels of oestradiol in post-menopausal women are significantly correlated with breast cancer risk
- It is the lifetime of exposure to oestrogens that are particularly important:
o Early age of onset of menarche (<11 y/o)
o Late age to menopause (>55)
o Late first full-time pregnancy
o Nulliparity vs multiparity
o Breast feeding is protective – associated with reduced levels of oestrogen
o Obesity in post-menopausal women


SERM - example and MOA?

E.g. Tamoxifen – bind competitively to ER, with tissue specific effects


SERD - example and MOA

E.g. Fulvestrant – bind competitively to ER to promote ER degradation


ATLAS study findings

For women with ER+ BrCa, continuing Tamoxifen for 10 years rather than stopping at 5 provides reduction in reoccurance and mortality
10 years of therapy can halve BrCa mortality during the second decade after diagnosis


How is oestrogen signalling targeted in hormonal BrCa therapy?

Lowering oestrogen levels:
- Ovarian oblation
- LHRH agonists

- Aromatase inhibitors


Ant-oestrogen therapies - MOA?

The ER receptor is a transcription factor
o It is recruited to particular regions of the genome
Conformational change occurs to the ER when oestrogen binds, permitting the recruitment of regulatory regions – enhancers, promoters of target genes
There is recruitment of co-regulatory molecules called co-activators
All of these processes act to permit transcription of genes that drive cell survival and proliferation


Resistance to hormonal therapies

De novo: Tumour independent of a requirement for oestrogen despite being ER+
- Usually, the ER itself is NOT lost.
- Growth promotion of initially oestrogen-dependent tumours by other mitogenic pathways
- Ligand-independent signalling - EGF activates ER receptor indirectly - it binds a TK at cell surface!
- ER post-translational modifications
- Co-activators/Co-repressors


Big difference between Primary and Metastatic BrCa?

Metastatic BrCa often exhibit ER mutants (active without presence of oestrogen)


Hormone therapy side effects/toxicity

- Tamoxifen is associated with an increased risk of Endometrial cancer
o This is because it acts like an oestrogen in the uterus
o Also causes weight gain and increased risk DVT
- Aromatase inhibitors:
o Associated with osteoporosis (oestrogen is required for bone maintenance)
o Joint discomfort
o Menopausal symptoms
- Anti-androgens
o Osteoporosis
o Impotence



- Small molecule inhibitor of the P450 enzyme CYP17A1
- This enzyme is required for both adrenal and intratumoral de novo biosynthesis of androgen hormones


Anti-androgen therapies

- GnRH agonists to reduce the androgen supply to the prostate from the testes
- Surgical/chemical castration
- Anti-androgens – Flutamide/Bicalutamide
New therapy: 17,20 Lyase inhibitor – Abiraterone acetate


Structural unit of chromatin?

Nucleosome, consisting of a segment of DNA wound in sequence around eight histone protein cores ( 2 each of H2A, H2B, H3, H4)
- Packaging and positioning of nucleosome determines gene expression!


CpG islands - what are they, when are they methylated

Regions of DNA around the promoter rich in CpG dinucleotides
Unmethylated in normal cells: Gene - ON
Methylation associated with gene silencing e.g. X-chromosome inactivation
Can become methylated in tumour cells --> suppression of gene expression e.g. loss of p53

Catalysed by DNA methyltransferase enzyme


CpG non-islands

Scattered throughout the genome
Methylated in normal cells, become unmethylated in cancer
- Its there as a protective mechanism to prevent mutations and therefore genomic instability


Histone modifications

Acetylation associated with open conformation
Methylation associated with condensed histone conformation


Epigenetic therapies

DNA methyltransferase (DNMT) inhibitors
- Approved for MDS, CMML and AML
- Approved in the US for treatment of all FAB classifications of MDS
These agents are incorporated into DNA/RNA
They sequester the DNA methyltransferases

Histone Deacetylase (HDAC) inhibitors
Vorinostat and Romidepsin
- Approval in the US for advanced cutaneous T-cell lymphomas
- Sadly a very rare cancer but these drugs show remarkable effects

Lack of specificity!


Normal stem cells vs CSC similarities?

- Asymmetric division:
o Self renewal
♣ Tissue-specific normal stem cells must self-renew throughout the lifetime of the organism to maintain specific organs
♣ Cancer stem cells undergo self-renewal to maintain tumour growth i.e. infinite growth potential
o Differentiation into phenotypically diverse mature cell types
♣ Give rise to a heterogeneous population of cells that compose the organ or the tumour but lack the ability for unlimited proliferation (hierarchical arrangement of cells)
- Resistant to DNA damage
o Over-expression of drug resistance genes
- Regulated by similar pathways
o Pathways that regulate self-renewal in normal stem cells are often deregulated in cancer stem cells


Clonal Evolution hypothesis

All cells within tumour functionally equivalent
Spatial and temporal tumour heterogeneity explained by selection:
Multiple mutations occur in different tumour cells
When a mutation arises that gives that lineage a growth advantage, a selection sweep occurs --> that lineage becomes dominant
- Exacerbated by the use of chemoradiation


CSC model

Only a small subset of cells within the tumour possess replicative mortality and therefore drive its growth
These cells are responsible for generation of all non-tumourigenic tumour cells
Heterogeneity attributed to the ability of the CSC to differentiate into phenotypically diverse subtypes


Combined model of cancer heterogeneity

Initially, growth and tumour survival driven by a specific subtype of CSC
Subsequent CSC mutation/epigenetic change (may be from external source) produces a new lineage that is more aggressive and takes over as the dominant cell type


CSC theory - cancer resistance

Would be expected that tumour recurrence after chemo is a result of CSC as they are the only cells able to induce tumour growth
- CSC would be expected to be chemoresistant cells as they (like normal stem cells) posses greater levels of drug resistant gene expression
- Theory limited by chemoresistance relapse as it would be expected that CSCs would regenerate the chemosensitive population


Knoechel et al 2014

NOTCH inhibitors used to treat ALL
Small population of resistant cells remained - this resistance could be reversed by epigenetic modulation
Suggestion that CSCs exist in an epigenetically poised state and may be driven by external factors to take on a resistance state by switching on drug resistance genes


miRNA biosynthesis

Transcribed via RNA polymerase II - hairpin structure (Pri-miRNA transcript)
Picked up by Pasha-Drosha complex in nucleus - cleaves hairpin structure --> pre-miRNA
Chaperoned into cytoplasm via RAN-GTP-exportin 5
Picked up by Dicer - processed to mature miRNA (can be from primary or complementary strand)
Assembled on RISC complex (RNA induced silencing complex)
Seed region binds mRNA
- Can be perfect fit (degradation) or non-perfect (translational repression)


TP53 & miRNA

Master regulator of miRNA expression
Expression of p53 in a cell leads to mass expression of a host of miRNAs
These miRNAs act to silence genes such as:
ZEB 1/2


C-MYC and miRNA

c-MYC directly involved in transcriptional silencing of multiple miRNAs
Silences miRNAs that act to inhibit:
- BCL2


miRNA examples in CLL

miR-15 (tumour suppressive miRNA) normally suppresses BCL2. Mutated/epigenetically silenced in CLL --> loss of apoptosis signalling

miR-92 (oncogenic miRNA) amplified in CLL
--> decreased expression E2F1
--> increased angiogenesis


miRNAs as biomarkers

Diagnosis, patient strat, prognosis
- Very stable in clinical preparations
- Secreted in multiple body fluids
BUT yet to reach clinical practice:
- Expensive as must be validated against histopathological biopsy which takes time and money


miRNAs in cancer treatment

Fire & Mello 2006 - nobel prize RNA interference

Kota et al. 2009
miR-26 downregulated in HCC mouse model
Reinstated it which prevented disease progression


Limitations to miRNA therapies

Transcript instability
Non-targeted biodistribution


DNA polymerase I

DNA repair


Types of DNA damage

1. Covalent base modification
- E.g. deamination of C --> U
2. Mismatch of normal bases due to failure of DNA proofreading
- E.g. U (RNA only) inserted instead of a C
3. Breaks in the DNA backbone
- Commonly caused by ionising radiation
4. Covalent crosslinking between bases
- Same DNA strand - intrastrand
- Opposite strand - interstrand
- Commonly caused by UV damage (crosslinking between pyriminidines on the same strand


Types of DNA repair

Direct reversal
- E.g. C-->T involves methylation and then deamination
- Repaired by glycosylamines that replace the base without damaging the backbone
- E.g. MGMT (methylguanine-DNA methyltransferase)
- Removes methyl groups. BUT wasteful as can only be used once, must be resynthesised repeatedly

Excision repair


BER enzymes

DNA glycosylases removes damaged base
Backbone cut by endonuclease
Replacement put in place via DNA polymerase beta
Nick sealed by DNA ligase


NER enzymes

DNA polymerase stops following recognition of a mutation - acts as a platform for DNA repair protein recruitment
GG-NER - Utilises XPGs to detect mutations across the entire genome

Both use TFIIH complex that unwinds the area being removed
DNA polymerases delta and epsilon responsible for the DNA synthesis to replace the damaged section
DNA ligase seals the nick



Mutations in these genes leads to HNPCC
Uses DNA polymerases delta and epsilon (from NER)


Cockayne's syndrome

CSA/CSB mutations
These gene products associated with RNA polyermase II and repair mutations that the RNA polymerase II recognises during transcription (it recruits CSA/B)


BRCA genes

Responsible for repairing DSBs
PARP (poly ADP ribose polymerase) - responsible for SSB repair. If SSBs not recognised then DSBs can occur when the DNA is replicated.
Therefore PARP inhibs induce DSBs which cannot be fixed by BRCA mutated Br Ca cells and therefore they undergo apoptosis


Platinum compounds: Mechanism of action

Chemically inert
Activated once in the cell - Cl groups replaced with water molecules
Active form binds N7 sites on purine bases
Forms platinum-DNA crosslinks within DNA
GpG intra-strand, ApG intrastrand
DNA adduct formation/damage leads to upregulation of P53 --> upregulation of pro-apoptotic mediators --> apoptosis
Pt also leads to ROS accumulation --> loss of integrity of mitochondrial outer membrane --> intrinsic apoptotic pathway


Pre-target resistance mechanisms

Pharmacokinetics: Renal/hepatic clearance, tumour vascularity
Drug transport:
ABC transport protein over-expression (MDR1).
CTR1 under-expression - prevents cisplatin entering cell
Drug inactivation:
Increased levels reduced Glutathione in platinum resistant cell lines suggests it is involved in platinum in-activation


On-target resistance

Target alteration:
Mutations in the target protein stop the chemo agent from binding
- Mutation in kinase domain of BCR-ABL prevents imatinib binding but preserves its oncogenic function

DNA repair pathway dysregulation
- Gain of function in NER --> enhanced ability to repair platinum adducts --> resistance to platinum therapies
- MMR loss of function mutations: --> MMR usually recognises Pt adducts and pushes cell through apoptosis pathway. Loss of function of this pathway --> failure of apoptosis --> resistance


Post-target resistance

p53 mutation --> less transcription of Puma & Noxa, two pro-apoptotic mediators. Less p53 = no apoptosis

Overexpression of inhibitors of intrinsic pathway e.g. survivin, XIAP --> failure of apoptosis therefore resistance

Platinum therapy increases phosphorylation of pro-apoptotic Bad, inactivating it
--> example of how Pt therapy itself leads to resistance formation

Overexpression of cell survival pathway mediators
- AKT1/2 overexpression correlates with Paclitaxel resistance in ovarian cancer
- Equally loss of PTEN is associated with platinum resistance


Two models of the evolution of drug resistance in ovarian cancer

1. Assumes somatic mutations that give rise to resistance are the sole result of genomic instability that occurs following long-term platinum exposure

2. These resistant cells actually exist prior to chemotherapy - chemo treatment just selects them for survival (similar to CSC hypothesis)