12-16: Cancer

Question 1

What is the fundamental difference between benign and malignant neoplasms?

Answer 1

Benign neoplasms are non-cancerous, and are just a collection of abnormal cells

Malignant neoplasms are cancerous cells that spread to adjacent tissues, forming secondary tumours

Question 2

State the stages of development for a cancerous tumour

Answer 2

Normal Epithelium
-> Hyperplastic Epithelium
-> Dysplastic Epithelium
-> Benign Neoplasm
-> Malignant Neoplasm
-> Metastasis

Question 3

Name some of the types of cancer based on types of tissue they arise from

Answer 3

• Carcinomas (from epithelium)
• Sarcomas (from Mesodermal tissues)
• Lymphoma and Leukaemia (from blood, bone marrow and lymphoid)
• Melanomas (from Neuroectoderm)
• Many others that are difficult to classify

Question 4

What is meant by a carcinoma in situ?

Answer 4

It is an epithelial neoplasm that has progressed beyond being benign (i.e. it has become more dysplastic) but has not yet spread to a secondary tissue (metastasis)

Question 5

What are the three main groups of aetiological factors for cancer mentioned (and what are aetiological factors)?

Answer 5

Aetiological Factors - show a STRONG association between exposure and incidence, so can be considered causes

• Chemical carcinogens (e.g., Coal Tar -> Lung/Skin Cancers)
• Physical carcinogens (e.g., UV -> Skin Cancer)
• Viruses (e.g., Epstein-Barr virus, Burkitt’s Lymphoma)

Question 6

Name some common Risk Factors of cancer that are not considered aetiological factors

Answer 6

Occupation (e.g., asbestos mining)
Reproductive history (e.g., number and age of pregnancies)
Diet (e.g., high fat, red meat, processed food)
Lifestyle (e.g., smoking, drinking, sunbathing)
Family History (cancer incidence CAN show Mendelian or Polygenic inheritance)

These risk factors, and others, can increase our exposure to aetiological agents, or otherwise exacerbate disease progression.

Question 7

Are significant variations in cancer incidence between countries mainly explained by environmental or genetic differences?

Answer 7

Environmental - for example, Japanese people who have moved to Hawaii have cancer incidence more similar to the average in Hawaii than the average in Japan

Question 8

What is the Ames Test?

Answer 8

A test to determine if a compound is MUTAGENIC (which many carcinogens are)

In the Ames test, the compound is mixed with homogenised rat liver (so that, if the compound IS mutagenic, it will be activated by the liver enzymes)

Then, the mixture is added to bacteria which are unable to grow without added histidine

The number of bacterial colonies that grow reflects the number of mutations, as any growing bacteria must have undergone a mutation allowing them to grow without histidine

Question 9

Name the extremely potent mutagen and carcinogen produced by Aspergillus mould

Answer 9

Aflatoxin

Question 10

Describe the rodent experiment that demonstrates the interplay between carcinogens and promoters in carcinogenesis

Answer 10

DMBA is a carcinogen, but addition of it alone did not immediately induce cancer in mice

Once an irritant (TPA) was added, papillomas formed, then carcinomas - induced by inflammation

Question 11

What is meant by “Tumour Promoters”?

Answer 11

Agents that promote tumour growth but are NOT MUTAGENIC (e.g., asbestos) - they stimulate the growth of mutated cells

They can be endogenous or exogenous GFs, or toxic compounds that induce compensatory proliferation. Many also induce inflammation

Question 12

Describe the different approaches that independently confirmed the role of genetic mutations in cancer

Answer 12

1. Cloning of oncogenes by cellular transformation assays (inducing cancer-like properties in vitro)
2. Positional Cloning of Familial Cancer Genes (Linkage Analysis)
3. Sequencing of cancer genomes, then engineering animal genomes to contain such mutations
4. Recombinant animal models (especially mice)

Question 13

What are the two broad classes of cancer-causing genes (and what are the differences between them)?

Answer 13

Oncogenes and Tumour Suppressor Genes

Oncogenes:
- Act dominantly
- Can be mutated by amplification, GOFs, translocations, etc.
- Include Ras, Myc, Src

TSGs:
- Act recessively
- Can be mutated by deletion, nonsense mutations, LOFs, etc.
- Include p53, Rb, APC, PTEN

Question 14

Give an example of how a translocation can activate an oncogene and lead to cancer

Answer 14

Philadelphia Chromosome (found in all Chronic Myeloid Lymphoma cells):

Translocation between chromosomes 9 and 22 results in a fusion protein between the BCR and ABL genes, thus removing the regulatory region from the Abl kinase (GoF), leading to excessive proliferation

Question 15

Name and explain the small molecule that can be used to treat Chronic Myeloid Lymphoma

Answer 15

Imatinib (TM Gleevec) is a small inhibitor of Abl, and can thus prevent the excessive proliferation seen in CML

Question 16

What is meant by “two-hit inactivation” of TSGs (and what was the example used in Lecture 12)?

Answer 16

There are two copies of each TSG, and BOTH copies must be inactivated to cause tumorigenesis

For example, a mutation in one Rb gene doesn’t lead to a phenotypic change, but mitotic recombination can result in Loss of Heterozygosity, leading to Retinoblastoma

Question 17

Why is tumorigenesis referred to in Lecture 12 as a “multi-step process,” what are some of these steps, and what does this mean for the type of cells that can become cancerous?

Answer 17

Each stage of cancer development (e.g., initiated, pre-cancer, cancer) requires some “distinct event” in order to progress - these events don’t happen often, which is why cancers are rare in youth

Many different mutations can drive tumour progression, some appear to be more essential or common than others:
- Loss of APC is often detected early in progression, and seems to be quite important for tumorigenesis
- Later events are more variable, and include activation of oncogenes, inactivation of TSGs, and epigenetic changes such as DNA hypomethylation

Statistical modelling suggests 6-7 mutagenic events are normally required for cancer, and these events occur every 10-15 years on average - this means a target cell in neoplasia must either be long lived enough to accumulate mutations over 40+ years (i.e. be a stem cell) or must have a fundamental shift in longevity

Question 18

Where can mutations in oncogenes and TSGs come from?

Answer 18

1. DNA Damage (e.g., mutagens, ROS, errors in replication, defective repair)
2. Defective chromosome maintenance and segregation (defective mitosis or erosion of telomeres)

Question 19

Name some of the DNA repair pathways that form part of our natural defences against neoplasia

Answer 19

Mismatch Repair Pathway (MMR)
Nucleotide Excision Repair Pathway (NER)
Base Excision Repair Pathway (BER)
Homologous Repair (HR)
Non-Homologous End Joining (NHEJ)

Germline mutations in genes encoding such DNA damage repair/signalling molecules give rise to inherited cancer susceptibility

Question 20

How can aneuploidy contribute to cancer?

Answer 20

Aneuploidy is a common trait in CML - and other cancer - cells

It contributes by deleting TSGs and/or producing GoFs in oncogenes

Question 21

Explain how “Darwin-like evolution” may drive cancer progression

Answer 21

In this model, mutations conferring a Growth Advantage in a single neoplastic cell are selected, allowing expansion of this cell’s lineage, until it dominates the neoplasm (this is the first clonal expansion)
-> Multiple rounds of clonal expansion (as other mutations required to pass through a bottleneck)

Question 22

BONUS: Name all 10 Hallmarks of Cancer (according to the 2011 update)

Answer 22

1. Sustaining Proliferative Signalling
2. Evading Growth Suppressors
3. Avoiding Immune Destruction
4. Enabling Replicative Immortality
5. Tumor-Promoting Inflammation
6. Activating Invasion and Metastasis
7. Inducing angiogenesis
8. Genome Instability and Mutation
9. Resisting Cell Death
10. Deregulating Cellular Energetics

There are rational therapies that aim to target each of these (some more promising than others)

Question 23

Summarise the Key Concepts of normal proliferation

Answer 23

Proliferation of normal cells is TIGHTLY CONTROLLED by positive (proto-oncogenes) and negative (TSGs) regulators:

• Normal cells ONLY divide when they receive an extracellular signal - Growth Factors (Mitogens)
• Mitogenic Signalling activates protein translation, and biosynthetic pathways involved in energy, anabolic metabolism, organelle production, etc. -> this triggers entry into the cell cycle
• The cycle is negatively regulated by Checkpoint Controls that “police” progress in cell growth, DNA replication, and chromosome segregation
• Strong cell-cell contacts (and other readouts of intact tissues) inhibit proliferation

Question 24

Describe in general terms how an extracellular GF (mitogen) induces a response in the cell

Answer 24

The mitogen binds to a cell membrane RECEPTOR

The signal is propagated across the PM and amplified by a cascade of SECONDARY MESSENGERS

The signal reaches the nucleus (TFs) where it promotes a program of gene expression that leads to proliferation

Question 25

Name and Describe the largest family of GF receptors

Answer 25

Receptor Tyrosine Kinases (RTKs) - all share similarity in their catalytic Tyrosine Kinase domain, but Ectodomains are highly variable, allowing them to respond to a range of ligands RTKs can be grouped into subfamilies based on sequence and structural homology (e.g., some ectodomains have Cys-rich domains, some have immunoglobulin-like domains, some have fibonectin type 3-like domains, etc.)

Question 26

Describe specifically how Growth Factor molecules lead to a signal being propagated across the Plasma Membrane (and how this can be deregulated)

Answer 26

Binding of GF molecules induces homo- or hetero-Dimerisation of RTKs (often aided by the GF ligands themselves ALSO being dimers) Dimerisation leads to conformational changes, allowing the TK domains to TRANSPHOSPHORYLATE each other (e.g., by removing an activation loop) Transphosphorylation creates docking sites for adaptor proteins with SH2 domains which often seed formation of multi-protein complexes and propagate the signal DEREGULATION: loss of the extracellular binding domain, or mutations in the cytoplasmic domain, can lead to ligand-independent firing of RTKs

Question 27

How does oncogenic activation of p21 Ras allow cells to proliferate even in the absence of mitogenic signals (and which specific mutations can cause this)?

Answer 27

A mutation in Ras that prevents hydrolysis by GAPs leads to constitutively active Ras, as it is always in the active, GTP-bound form Ras then constitutively activates multiple downstream signalling cascades which reprogram gene expression to allow cells to grow and divide: MAPKKK (Raf) -> MAPKK (MEK) -> MAPK (Erk1/2) -> many downstream proteins P13K -> PIP3 -> Akt and RhoGEFs (together, stimulate cell growth and inhibit apoptsis) Ral-GEF -> ... -> Cdc42 (Filo) and Rac (Lamellipodia) Note that these cascades can also be independently targeted by mutations, but Ras activation is notable for activating ALL pathways simultaneously. Many such common oncogenes and TSGs are hubs at the "crossroads" of multiple signal inputs and outputs SPECIFIC MUTATIONS: All 3 forms of Mammalian Ras (K, N and H) are frequently mutated in cancers, but especially K. Gly12 is a mutation hotspot, as it forms part of a GTP-binding pocket. Substitution of Gly12 with Valine prevents GTP hydrolysis and locks Ras in an active form

Question 28

Describe the MAPK (or Raf) signalling pathway, and state which components have been implicated as oncogenes or TSGs

Answer 28

MAPKKK (Raf)* is activated by Ras*, downstream of RTKs MAPKKK -> MAPKK (MEK) MAPKK (MEK) -> MAPK (ERK) ERK -> Myc* and also Elk1 Elk1 promotes transcription of c-fos* which combines with Jun* to form the TF dimer AP-1 and promote further gene expression

Question 29

Explain how mitogens and the MAPK pathway relate to the cell cycle

Answer 29

The majority of body cells are not dividing at any given moment, and are in G0 (either terminally differentiated, or stem/blast cells that are poised to divide but awaiting a mitogenic stimulus) Mitogenic signalling via GF -> RTK -> Ras -> Raf -> MEK -> ERK goads G0 cells out of quiescence and into the cell division cycle

Question 30

How is progression through the cell cycle "policed"?

Answer 30

It is policed at several Checkpoints by the TSGs pRb and p53: 1: pRb at the R point in late G1 - beyond which, the cell is committed to completing a division cycle and is no longer responsive to mitogens or TGF-ß [The R point is deregulated in most or all cancer cells] 2: p53 at the first DNA damage checkpoint, end of G1 (entrance into S is blocked if genome damaged) 3: p53 at second DNA damage checkpoint (DNA replication is halted if genome damaged) 4: p53 at end of G2 (entrance into M is blocked if DNA replication is not completed) Also, anaphase is blocked if chromatids are not properly assembled on the spindle

Question 31

What catalyses transitions through the cell cycle?

Answer 31

Cyclin-CDK dimer complexes: CDKs are Ser/Thr kinases which, stimulated by cyclins (which also recognise substrates), catalyse progression from one stage to the next CDK levels remain constant, but Cyclins are degraded by the proteasome (irreversible!) and created de novo at each turn of the cell cycle Mitogens initiate the cascade in the first place by inducing Cyclin D expression

Question 32

Which cyclins/CDKs are active at each stage of the cell cycle?

Answer 32

G1: Cyclin D, CDK4/6 At R point in G1: Cyclin E, CDK2 From start to mid-S-phase: Cyclin A, CDK2 From mid-S to end of G2: Cyclin A, CDK1/CDC2 M-Phase: Cyclin B, CDK1/CDC2

Question 33

Describe the role of pRb in regulating the R point, and what happens to it during the cell cycle

Answer 33

pRb is initially unphosphorylated, and binds both the Transcription Factor E2F, and Histone Deacetylase (HDAC), thus inhibiting E2F pRb is initially hypo-phosphorylated in G1 by CyclinD/CDK4/6 (mitogen-dependent at Cyclin D expression required) Eventually, pRb becomes HYPER-phosphorylated, and releases E2F, which binds Histone Acetylase and can now activate transcription of genes that lead to cyclin E production and progression into S-phase Once past the R point, serial phosphorylation by later Cyclin-CDK complexes (e.g., E-CDK2) -> positive feedback NOTE: Most or All Cancers involve deregulation of this restriction point, achieved through mutation or epigenetics, often amplifying cyclin D or deleting Rb

Question 34

Give an overview of rationale therapy for cancer based on suppressing mitogenic signalling

Answer 34

Traditionally, therapies targeted proteins downstream of oncogenic mutation. However, increasingly, drugs are being developed which directly target the mutated oncogene - MoAbs (e.g., Herceptin) target the Ectodomains of GFRs - Small molecule inhibitors (e.g., anti-Her1/2/4) target TK domains - Other small molecules have been developed to target mTOR, Raf, MEK, and other kinases in the cascade CDK inhibitors have been proposed, but are controversial as pan-CDKi's were quite toxic. CDK4/6-specific inhibitors have been approved

Question 35

Define "Anoikis" and explain its significance in the context of cancer

Answer 35

Anoikis is when a cell undergoes apoptosis due to detachment from its matrix and consequent lack of contact signals This is a major barrier for any potential metastasis to overcome

Question 36

What are the three groups of Bcl2-family proteins, and what is the role of each in apoptosis?

Answer 36

Anti-apoptotic Bcl2s (e.g., Bcl2, Bcl-XL, Bcl-W) contain the BH1-4 domains and a transmembrane domain - these proteins inhibit pro-apoptotic Bcl2s PRO-apoptotic Bcl2s (e.g., Bax, Bak, Bok) lack the BH4 domain, and promote apoptosis by oligomerising to form the MOMP complex, which forms large, non-selective pores in the OMM BH3-only proteins (e.g., Bid, Bim, Bik, Bad, Noxa, Puma) lack all homologous domains except the BH3 domain, and regulate the other two classes of Bcl2s, either by activating pro-apoptotics, or by inhibiting pro-survival factors The various BH3-only proteins are activated in response to a range of factors, e.g., DNA damage, lack of GFs, anoikis, chemo/radiotherapy, oncogene activation, etc. Many BH3-only proteins are sequestered in inactive forms until activated, or require proteolytic cleavage

Question 37

How was the first protein in the Bcl2 family discovered and named?

Answer 37

Bcl2 - B-cell Lymphoma 2 This pro-survival factor is an oncogene first discovered in follicular B-cell lymphoma, in which a translocation, t(14;18) brings it under the control of the ubiquitous IgG promoter, thus upregulating its expression This isn't inherently tumour-producing, but overexpression of Bcl2 co-operates with overexpression of oncogenes such as Myc in lymphoma formation

Question 38

Describe the role of P13K in survival signalling

Answer 38

P13K is recruited (via its SH2 domain + Ras) to the PM by IGFR in response to RTK activation by IGF Now activated, P13K phosphorylates PIP2 to generate the second messenger PIP3, which is recognised by, and activates, Akt/PKB Akt/PKB is a ser/thr kinase which is involved in many signalling pathways, including inactivation of Bad by phosphorylation of its BH3 domain (which leads to sequestration by 14-3-3) When no GF is present, Akt cannot phosphorylate Bad, so it is free to associate with mitochondria-associated Bcl2-family proteins and promote MOMP P13K is an oncogene and its often mutated in cancers

Question 39

Describe the general significance of p53, and the most common mutations thereof

Answer 39

p53 is a ubiquitous TSG, and is the most frequently mutated gene in cancers Most p53 mutations are missense mutations in its DNA-binding domain, which prevent it from carrying out its TF function

Question 40

Explain the role of p53 as a tumour suppressor

Answer 40

It is a critical transducer of abnormality signals, as it integrates many stressful stimuli (e.g., UV/ionising radiation, oncogene signalling, hypoxia, blockage of transcription, lack of nucleotides, etc.), then induces a number of cellular responses: - Cell Cycle Arrest (this buys time to repair DNA, then return to proliferation, or, if more severe, senescence or apoptosis) - DNA Repair - Blockage of angiogenesis - Apoptosis

Question 41

What are the five main classes of genes regulated by p53?

Answer 41

Growth arrest genes (mainly p21) Regulators of apoptosis, (e.g., Bcl2-family, APAF1, NF-kB, PTEN, Fas) DNA Repair genes (e.g., Siah-1, 14-3-3) Anti-angiogenic proteins (TSP-1) -> Also p53 antagonist MDM2 (negative feedback)

Question 42

What regulates p53 levels in the cell?

Answer 42

p53 is very dynamically made - it is normally degraded by the proteasome, and is only upregulated in response to a specific stimulus MDM2 is a downstream target of p53, and is also an E3 ligase which poly-ubiquitinates p53 - in healthy cells, this negative feedback keeps p53 low, but interference allows [p53] to rise quickly For example, phosphorylation of p53 by damage sensors such as ATM blocks MDM2 association and prevents degradation of p53

Question 43

Give three examples of downstream pathways by which p53 activation leads to apoptosis

Answer 43

1. p53 upregulates Fas -> Casp8 and Casp10 -> ECs -> Apoptosome 2. p53 upregulates Noxa/Puma -> lower threshold for MOMP -> pores in OMM -> Cyt c released -> apoptosome 3. p53 upregulates IGFBP-3 -> binds IGF-1/2 and prevents activation of IGF-1R (so less P13K -> Akt -> phosphorylation of Bad)

Question 44

Explain the significance of p53 and the loss of its function

Answer 44

p53 acts as a single key "genome guardian," making it a weak link in the line of defense against neoplasia, as the loss of this one gene greatly increases the chance of a cell becoming neoplastically transformed LOSS of p53 makes cells resistant to: - Loss of survival signalling - Hypoxia - Oncogene activation - Chemo/radiotherapy All of these would normally induce apoptosis via p53 Many cancer therapies deliberately damage DNA in order to activate p53, but this is ineffective if p53 function is already lost

Question 45

What are the three main (highlighted) types of aberrations seen in cancer cells that increase resistance to apoptosis?

Answer 45

- Altered survival factor signalling (e.g., P13K, Akt, PTN, IGFBP mutations etc.) - Inactivation of p53 - Changes in levels of Bcl2s

Question 46

What are the two (mentioned) molecules that aim to treat cancer by targeting resistance to apoptosis?

Answer 46

ABT-263 - a small BH3-mimetic which competes with BH3 proteins for binding to certain anti-apoptotic Bcl2 proteins, thus inhibiting pro-survival factors and promoting apoptosis Nutlin-2 binds the p53-binding pocket of MDM2, thus inhibiting p53 degradation and promoting apoptosis

Question 47

What is senescence, why does it occur, and what can allow cells to bypass it?

Answer 47

Senescence is an irreversible phenomenon, in which many cellular processes deteriorate as the cell ages (after around 60 doublings for fibroblasts) After a cell has doubled many times, p53 and Rb are activated, shutting down the cell cycle, and there are certain morphological markers, e.g., larger, flatter cells, and upregulation of acidic ß-galactosidase and cyclin inhibitors Senescence is a physiological defence against neoplasia. However, inactivating p53 and pRb (e.g., by the LT oncogene) allows cells to bypass senescence and keep dividing indefinitely

Question 48

Briefly describe what the telomeres are, and the role they play in senescence

Answer 48

Telomeres are TTAGGG tandem repeats found on the ends of linear chromosomes - they are the solution to the End Replication Problem (without them, the ends of the chromosomes could not be replicated as the RNA primer could not anneal) Telomeres get shorter with each DNA replication - this is why senescence is eventually necessary; if it is prevented, CRISIS will eventually occur

Question 49

Name the two essential telomeric protein complexes, and briefly describe what they do

Answer 49

Shelterin and CST: Shelterin regulates formation of the T (Telomere) Loop, restricting telomerase access to the 3' tail. When the telomere opens up during S-phase, then telomerase can bind to the 3' tail and add telomeric repeats. CST then inhibits telomerase to prevent excessive extension

Question 50

What is meant by "Crisis" in the context of cancer cells?

Answer 50

Cells that avoid senescence via mutations hit a second "roadblock," triggered by telomere erosion: Eventually, the telomeres become too short, and the genome becomes increasingly unstable as chromosomes undergo breakage-fusion-bridge events and structural rearrangement This genome instability is known as crisis Many cancer cells undergo crisis, but THEN reactivate the telomeres, meaning they become immortal AND ALSO live on with the legacy of an aneuploid genome, giving them a proliferative advantage over cells that reactivated telomeres WITHOUT undergoing crisis

Question 51

Describe the two classes of Cancer Therapies that target telomerase activity

Answer 51

Telomerase activity is a validated therapeutic target: 1. Small molecule hTERT inhibitors (several molecules are being developed that inhibit the hTERT promoter, which is often mutated to allow telomerase reactivation) 2. Antisense therapy (GRN163L is a covalently modified antisense oligonucleotide, which has base complementarity with the telomerase RNA template, and thus inhibits telomere extension)

Question 52

How does circulation limit tumour growth, and what do tumours do to overcome this?

Answer 52

Tumours require access to circulation - cells more than around 100µm from a capillary suffer from hypoxia and lactate build up -> necrosis Tumours must create new vasculature to grow - this is a physiological barrier they must overcome

Question 53

How does hypoxia affect gene expression in tumours?

Answer 53

Hypoxia induces stabilisation of the TF HIF-1alpha (which would normally be hydroxylated, ubiquitinated by pVHL, and degraded) This TF then targets genes such as VEGF to induce new blood supply, and other targets which allow the cell to shift to anaerobic respiration

Question 54

Describe the process of angiogenesis

Answer 54

New blood vessels arise from pre-existing capillaries Firstly, pericytes detach and vessels dilate. Then, the BM and ECM are degraded by MMPs, allowing endothelial cells to migrate into the perivascular space, towards angiogenic stimuli produced by tumour cells (e.g., VEGF) Then, endothelial cells proliferate, loosely follow each other, guided by pericytes, and adhere to each other to create a lumen (accompanied by BM formation and pericyte attachment)

Question 55

What features of tumour vasculature are different to normal vasculature

Answer 55

Blood vessels in tumours are leaky due to fenestrations in endothelial cells This raises the hydrostatic pressure within the interstices of tumours (and thus inhibits the distribution of chemotherapy) Tumour vasculature is also more chaotic and less organised

Question 56

What regulates the Angiogenic Switch? (I.e. whether endothelial cells are in a quiescent or angiogenic state)

Answer 56

Imbalance between angiogenesis activators and inhibitors: ACTIVATORS are mainly RTKs such as VEGF, or factors which upregulate VEGF (e.g., EGF, LPA) Many inhibitors (e.g., angiostatin, endostatin) are derived from larger proteins that have no effect on angiogenesis Note: angiogenesis factors can come from inflammatory cells (e.g., masts and macrophages), and the ECM, not just from cancer cells

Question 57

Summarise the potential of angiogenesis inhibitors in treatment of cancer

Answer 57

There is proof of principle: Some inhibit the angiogenesis signalling cascade (e.g., interferon-alpha, anti-VEGF Ab) Some target endothelial cells directly (e.g., EMD promotes apoptosis thereof) Some block the ability of Endo's to break down the ECM (e.g., Marimistat and other drugs that block MMPs to prevent migration)

Question 58

What is one (mentioned) reason why drugs that target angiogenesis may be less effective than other classes of cancer therapies?

Answer 58

There are alternatives to angiogenesis that tumours can use instead (e.g., Sprouting angiogenesis and Vasculogenesis)

Question 59

What is the Warburg Effect?

Answer 59

The Warburg Effect refers to the altered metabolism of tumour cells: Cancer cells show increased conversion of glucose into lactic acid (i.e. fermentation) even in the presence of oxygen - Aerobic Glycolysis Simultaneously, MORE glucose is consumed (e.g., by upregulating glucose transporters), and more is diverted into biosynthetic reactions

Question 60

Explain the significance of Glutaminolysis in tumour metabolism

Answer 60

Glutamine can be a source of Carbon, ATP and Nitrogen, making it essential for tumour growth and cellular transformation Glutaminolysis allows conversion of glutamine to glutamate, aspartate, CO2, pyruvate, alanine, lactate, and the TCA Intermediate CITRATE (this makes it an example of anaplerosis) Deamination of glutamine releases an amine group, which can then be converted to nucleotides, or used in production of other amino acids, etc.

Question 61

What are the advantages to cancer cells of their altered metabolism (i.e. the Warburg Effect and Glutaminolysis)

Answer 61

- They are better adapted to fluctuating oxygen levels - Less harmful ROS generated by OxPhos - Metabolites can be redirected to biosynthesis without affecting Energy - Lactic acid can be used by normoxic cells to regnerate pyruvate (symbiosis); Lactate can also suppress immune cells

Question 62

What causes the reprogramming of intermediary metabolism in cancer cells?

Answer 62

Oncoproteins and TSPs regulate expression and activity of MANY ENZYMES that regulate metabolic flux - Cancer cells overexpress PKM2 (different splicing to PKM1), induced by the Myc oncogene (PMK2 low activity reduces conversion of PEP to pyruvate, allowing metabolites upstream of pyruvate to accumulate and be channelled into biosynthetic pathways) - Many other enzymes downstream of oncogenes and TSGs contribute to altered metabolism - Some are potential therapy targets, e.g., PDK, MCT4, LDH, HK2, IDH, etc etc. However, no drugs have yet balanced the effects on cancer cells and normal cells

Question 63

State the major events of metastasis

Answer 63

1. Detach from original location and breach BM 2. Migrate through stroma 3. Cross endothelial layer 4. Travel through blood/lymph vessel 5. Extravasate into distal tissue 6. Establish a new colony of cells

Question 64

What cellular change are required for a malignant cell to show Local Invasiveness?

Answer 64

Reduced cell-cell adhesion (down-regulation of E-cadherin) Proteolytic degradation of BM (invadopodia) Acquisition of motile phenotype: -> Adhesion to stromal ECM (altered integrin expression) -> Cytoskeletal reorganisation -> Propulsive force (actomyosin + actin pol) -> Proteolytic degradation of stromal ECM (MMPs)

Question 65

What is the difference between the transformation of cancer cells to acquire motility/invasiveness, and the similar transformation that occurs physiologically in embryogenesis and wound healing?

Answer 65

In the physiological examples, although the transformation is similar, it is tightly regulated, whereas in neoplasia, it is deregulated

Question 66

What is the observable difference in cultured cells (e.g., MDCK cells) between WT and Ras mutant cells?

Answer 66

WT cells grow in colonies due to strong cell-cell adhesions Ras mutant cells form pseudopodia and crawl around the dish - if a GF is added as well, the cells migrate and form branches through the matrix

Question 67

What is the role of invadopodia in invasiveness of cancer cells?

Answer 67

Invadopodia are actin-rich protrusions, which focus the proteolytic power of MMPs to degrade the BM, and then expand behind, to widen the gaps and allow the cell to squeeze through

Question 68

Describe the role of altered integrin adhesion in malignancy, and what happens if integrin adhesion is knocked down in tumour cells

Answer 68

Malignant cells use integrins to adhere to the ECM, and use collagen fibres like "vines" to migrate Blocking integrin attachment (e.g., using RGD as a competitive inhibitor) does not prevent the primary tumour from forming, but does prevent metastsis

Question 69

Explain why cytoskeletal re-organisation is necessary for cell migration and invasion - and name the family that regulates this

Answer 69

Cells must undergo the Epithelial-Mesenchymal Transition, and form Lamellipodia at the leading edge, Filopodia to sense their environment, and Stress Fibres to retract the rear Rho GTPase Family Proteins orchestrate this reorganisation, and are themselves regulated by GEFS and GAPs, and also via sequestration by RhoGDI

Question 70

How can the cytoskeletal re-organisation by Rho, Rac and Cdc42 be visualised?

Answer 70

Staining actin and vinculin (which shows areas of focal contact)

Question 71

Describe the role of Proteases in Invasiveness

Answer 71

Various classes of proteases (including 4 classes of Endoproteinases, and some amino- and carboxy-peptidases) contribute to ECM degradation Some are integral membrane proteins (bound by GPI-anchor), others are bound to other membrane proteins Proteinases generally increase efficiency of migration and limit collateral damage - specificity is important, as the goal is not to degrade ALL of the ECM, but only the parts that are a physical barrier to migration

Question 72

What are the challenges of Intravasation for a tumour cell?

Answer 72

- Must initially degrade blood vessel BM - Must resist anoikis, shear stress AND predation by immune cells when in circulation (most perish here)

Question 73

What happens to tumour cells that survive in the circulation?

Answer 73

They are usually trapped at the next capillary junction Platelets then form a microembolism, then the cancer cell denudes the endothelium and binds to the BM Cancer cells begin to proliferate within the lumen, and eventually erupt into surrounding tissue

Question 74

What are Micrometastases and why are they significant?

Answer 74

Most cells colonising a distal site form DORMANT tumours - micrometastases (dormancy not fully understood, but partly due to limited blood supply) Dormant but Significant: If a patient already has micrometastases when a clinically significant tumour is removed, their survival chances are lower

Question 75

Do cancers show a preference for specific organs as the site of metastasis?

Answer 75

Some show no preference, but many show a strong predilection for certain organs (e.g., melanoma in lungs, colorectal in liver, prostate in bones) (Several theories as to why)

Question 76

Why do some types of cancer (e.g., melanomas) show a strong preference for certain organs as the site of metastasis?

Answer 76

Initially, this was explained by the pattern of blood flow around the body (e.g., colorectal cancers would be first taken up the hepatic portal to the liver) However, this can't be the whole story, as some cancer types show "tropism" for certain tissues The "Seed and Soil" Hypothesis states that only certain tissues have similar tropic signals to the primary site and support growth Also, chemokines drive a HOMING response, triggering chemotactic and invasive responses (e.g., lung tissue expresses a lot of CXL12 - the ligand for the CXCR4 receptor expressed in breast cancer cells) - such signalling could activate pathways such as MAPK, polymerisation of actin, etc. Also, metastatic cells create a METASTATIC NICHE to establish a tumour - due to expression of matrix components, GFs and chemokines at the site of metastasis, circulating cells are attracted to these sites where they find an optimal environment to settle and proliferate, forming a secondary tumour

Question 77

Describe the range of therapies that aim to target the invasion metastasis cascade

Answer 77

Some drugs could be effective prior to dissemination of micrometastases, but it is clinically uncommon to be able to target this early Some aim to inhibit the growth of already disseminated micrometastases, but these drugs tend to be toxic, making research difficult (it is also hard to tell whether they have killed micrometastases that were too small to detect in the first place) Drugs that can induce regression of already macroscopic metastases (e.g., biphosphonates, SD-208, Desmosumab) may be the most clinically useful