1. Self-sufficiency in growth signals
2. insensitivity to growth-inhibitory signals
3. Tissue invasion and metastasis capability
4. Limitless replicative potential
5. Sustained angiogenesis
6. Evasion of programmed cell death (apoptosis)

Diffusable growth factors

Extracellular matrix components

Cell-to-cell adhesion/interactions

A gene that has experienced a gain of function mutation that promotes cell proliferation instead of apoptosis (dominant)

--

An oncogene is a gene that, when mutated or expressed at high levels, helps turn a normal cell into a tumor cell.

Many cells normally undergo a programmed form of death (apoptosis). Activated oncogenes can cause those cells to survive and proliferate instead. Most oncogenes require an additional step, such as mutations in another gene, or environmental factors, such as viral infection, to cause cancer. Since the 1970s, dozens of oncogenes have been identified in human cancer. Many cancer drugs target those DNA sequences and their products.

Loss of function mutations in genes that restrain cell proliferation or guard the genome (recessive)

--

A tumor suppressor gene, or antioncogene, is a gene that protects a cell from one step on the path to cancer. When this gene ismutated to cause a loss or reduction in its function, the cell can progress to cancer, usually in combination with other genetic changes.

ErbB2/HER/NEU

RAS

ErbB2/HER2/Neu

• What is
• What happens during overexpression
• What extends the life of breast cancer patients w/over-expression of Her2

• Receptor tyrosine kinase of the epidermal growth factor receptor (EGFR) family
• Overexpression of receptor makes much more sensitive to ligand
• Herceptin is a monoclonal antibody shown to extend the life of breast cancer patients that over-express Her2

RAS

• What is
• Mutation creates what and does what
• RAS is altered in __% of all human cancers

• RAS is a small GTPase; an intracellular switch that can be fixed in the "on" position; RAS normally cycles between active and inactive forms
• Mutation creates structural alterations that generate extended firing periods
• 25%

GEF and GAP

Disruption of cell membranes

Structural filaments broken down

Chromosomes degraded

Debris is phagocytosed

Survival Signals (IGFs, interleukins)

Death signals (FAS, TNF-

alpha)

DNA damage

Overexpressed oncogenes

Hypoxia

Tumour angiogenesis

• What protein regulates expression of genes involved in angiogenesis
• Hydroxylated when?

HIF regulates expression of genes involved in angiogenesis

• Under nl conditions, HIF is hydroxylated
• During hypoxia, unhydroxylated HIF is stabilised and can bind coactivators, allowing for cellular division

• Bcl-2 family of proteins
• loss of p53 function

CdK_i

• Expression regulated by __; instigated by; resulting in ____

• TGF ligands bind to a type II receptor
• This recruits and binds a type I receptor
• Type I receptor phosphorylates receptor-regulated SMADs which can now bind the coSMAD SMAD4.  
• R-SMAD/coSMAD complexes accumulate in the nucleus where they act as txption factors and participate in the regulation of target gene expression

• Wildtype type II TGF-beta receptor and TGF-beta --> S/T kinase --> SMAD3 binds to SMAD4 --> Expression of p15^ink4b and p21^clp1

1. Down-regulation or mutation of receptor
2. Loss of SMAD4
3. Deletion of p15 CDKi
4. Mutation of CDK4
5. Overexpression of Myc (v-myc)

• Unphosphorylated = G0 and early G4 (activity on)
• Hypophosphorylated = pre-restriction point G1 (activity on)
• Hyperphosphorylated = post-restriction point (activity off)

Def- a DNA tumour virus encodes proteins that inactivates Rb or tags it for protolysis

Ex- F1A (adenovirus); T-ag (SV40), E7 (HPV)

Genetic repair

Fidelity is normally very high, and rare mutations are usually repaired, but...(3)

1. Repair is not 100%, and becomes less precise with age
2. Repair pathway genetics may be mutated
3. Massive carcinogen exposure

Incorporation of errors during DNA synthesis:

• When DNA polymerase is present:
• When proofreading is present:
• When mismatch repair is present:

• 10^5
• 10^7
• 10^9

1. Incorporation of errors during DNA synthesis
2. Ntt decay
3. DNA damage
4. Pyrimidine dimers

Depurination (mammalian cells sustain more than 10,000 abasic sites per day; def-A & G

Depyrimidination

Spontaneous deamination

1. Oxidative damage (endogenous)
2. Chemicals
3. Radiation (UV, xrays, etc)

1. Polycyclic aromatic hydrocarbons (lung epithelium)
2. Ultraviolet light (epidermis)
3. Aflatoxin (liver)

Polycyclic aromatic hydrocarons

• Source
• Damage
• Effect

• Smoke/environment
• DNA adducts
• G to T

UV Light

• Source
• Damage
• Effect

• Sun
• Pyrimidine dimers
• Point mutation

Reactive oxygen species (ROS)

• Source
• Damage
• Effect

• Normal metabolism/x-rays
• abasic sites/ss and ds breaks/cross-links
• Point mutation/translocation

Aflatoxin

• Source
• Damage
• Effect

• Diet (peanuts)
• DNA adducts
• G to T

Heterocyclic amines

• Source
• Damage
• Effect

• Diet (cooked meat)
• DNA adducts
• G to T

1. Mismatch
2. Ntt excision repair
3. Homology-based repair (HR)
4. Non-homologous end joining (NHEJ)

1. Binding of mismatch proof-reading proteins
2. DNA scanning detects nick in new DNA strand
3. Strand removal
4. Repair DNA synthesis

Examples of mismatch repair genes mutated in cancer cells

Mutation causes what

Ntt excision repair

• Def
• Steps
• Example of dx of repair system

• Repairs bulky DNA adducts and pyrimidine dimers by a multi-step process
• Detection (distortion); excision (endonuclease); DNA synthesis (delta or epsion); ligation

Triggers of dbl-strand break repair (HR & NHEF)

(3)

• Reactive oxidation species
• DNA topisomerase inhibitor (chemo)
• "replicative stress"

• Melt helix of nl chromosome in the appropriate region
• Invasion by the free end of severed strand by DNA polymerase
• Extension of severed strand by DNA poymerase
• Re-joining of the severed strand with DNA from the other side of break

• Dbl strand break
• Resection of single strands by exonuclease
• DNA strands brought together; possible limited base pairing between them
• Strands filled in; joined by ligation
• Double helix reconstruction

Gene: BRCA1/2

Affected pathway:

Gene: p53 and CHK2

Affected pathway:

Dx/syndrome:

Damage detection and signalling

Li-Fraumenti

Gene: BLM (helicase)

Affected pathway:

Dx/syndrome:

DNA replication

Solid tumour

Gene: MMR genes

Affected pathway:

Dx/Syndrome:

Mismatch repair

Colon cancer (HNPCC)

Gene: XP group of genes

Affected pathway:

Dx/syndrome:

NER

Xeroderma pigmentosum

• Entrance to "S" phase is block when:
• Entrance to "M" is blocked if:
• Entrance to anaphase is blocked if:

• Genome is damaged
• DNA replication is not complete
• Chromosomes are not assembled on the mitotic spindle

• Cdk2 (preventing entry to S)
• PCNA (halting DNA replication already taking place)
• Other targets? (Ciz1; Cip1-Interacting zinc finger protein)

PCNA

• What is
• What does
• What does p21 do?

• DNA polymerase accessory factor - 'sliding clamp'
• Attracts DNA repair factors to arrest DNA replication forks
• p21 inhibits the DNA replication activity of PCNA but leaves its repair activity intact

Damage = ds breaks

Damage detection = sensor molecules

Damage signaling = transducer molecules

Damage response = mediator molecules

Damage repair = effector molecules

• PIKK = Pi3 kinase family are check point proteins
• Recruited to sites of damage by partner proteins, where they phosphorylate downstream targets

Chk2

• Activated by ___ at sites of ___
• homodimerization/autophosphorylation

• Activated by ATM at sites of double strand breaks
• Phosphorylation by ATM promotes homodimerization and autophosphorylation of Chk2
• Resulting structural alteration allows Chk2 to phosphorylate other substrates
• Spreads signal throughout the nucleus

Gene: ATM

Affected pathway:

Dx/syndrome:

Gene: Ataxia Talengectasia Mutated

Affected pathway: Response to DS breaks

Dx: Ataxia talengectasia (lymphomas)

NHEJ

HR is cell-cycle regulated (G2)

Loss of BRCA1/2 confers susceptibility to cancer; germ-line mutation in either causes cancer in:

• __% of familial breast cancer
• __% of ovarian cancer

• 50% breast
• 70% ovarian

• At an early stage, incipient tumour cells experience 'oncogenic stress'
• They respond by activating protective pathways (arrest or die)
• Mutations compromising these pathways (including defects in ATM, Chk2, p53) might allow cell proliferation and survival
• This compromises the checkpoints and results in increased genomic instability and tumour progression
• IN OTHER WORDS: chronic activation leads to deactivation and then to hyper mutation

1. Disruption of ntt pools (mutation of synthetic pathways or nutrient deprivation)
2. DNA damage or protein obstructions that blocks replication from progressing
3. DNA secondary structure that affects unwinding
4. Altered expression of replication factors (eg, cyclin E)
5. Passage of DNA replication forks turns SS breaks into DS breaks

Activated oncogenes

Unscheduled replication

Aberrent replication structures

_____

___ or ___

_____

_____

1. Cell cycle arrest
2. DNA repair
3. Apoptosis

p53 mutation typically is what kind of mutation?

Results in what phenotype?

Post-translational Modification - Phosphorylation

• Response to
• Which molecule

• Phosphorylation
• ATM

• Stress
• p300

• Stress
• SET9

• Stress
• PIAS

• Basal
• MDM2

1. Rapid
2. Reversible
3. Requires less energy
4. stress dependent
5. sensitive to the magnitude of stress

MDM2

• Def
• Does what
• MDM2-/-
• MDM2-/- ; p53-/-

• Def = murine double minute clone 2
• Keeps p53 under control
• Embronically lethal
• Viable

Activation of p53 function without DNA damage

• Happens why
• Critical p53 residues are
• Bonds where
• Druggable target?

• Inhibition of MDM2-p53 interaction
• Phe-19, Trp-23, and Leu-26
• Hydrophobic pocket on MDM2
• Yes.  Strategy would be used in tumours that: retain wild-type p53 (~50% of tumours) and overexpression of MDM2 (~7% of tumours)

Sirtuins

• What are
• Example in humans
• Does what

• NAD+ dependent protein deacetylases
• Resveratrol in red wine
• Increases lipid metabolism, improves muscle function, inhibits alzheimers

MMP

• Def
• What happens during cancer
• Expression in cancer often is associated with

• Family of proteases that collectively can degrade virtually all components of the ECM
• Upregulated in almost every type of cancer
• Poor survival

1. Tumour growth
2. Angiogenesis
3. Invasion
4. Intravasation and extravasation

1. Some of the oncogenes, implicated in breast cancer development (such as myc and ras) can induce expression of MMP-2 and MMP-9 in breast cancer cell lines
2. Overexpression of MMP-3 in transgenic mice causes malignant mammary gland tumours
3. High levels of MMP-2 and MMP-11 correlate with poor prognosis in breast cancer patients

MT1-MMP

• First identified where; did what; located where
• MMP2 were identified where

• Lung carcinoma cells; acted to degrade ECM proteins and activate proMMP-2; located at the leading edge of migrating cells facilitating ECM degradation
• Lung, gastric, colon, liver, breast, bladder, ovarian, cervical, and brain tumours

• Angiogenesis; as the primary tumour grows it reaches a critical size where it needs to generate its own bld supply to meet its metabolic needs
• Intravasation; tumour cells use this new bld supply to enter circulation
• Extravasation; tumour cells travel in the blood to distant sites where they leave circulation
• Secondary tumour forms

1. Tumour cells release angiogenic factors to surrounding tissue
2. Activates specific genes in that tissue to encourage growth of new blood vessels

VEGF (vascular endothelial growth factor)

FGFs (Fibroblast growth factor)

TGFs (Transforming growth factors)

Angiostatin

• Does what genetically
• Phenotypic results

• Downregulates VEGF expression
• Increases EC and tumour cell apoptosis; inhibits EC proliferation; decreases blood vessel density

• Blocks binding of VEGF with its receptor
• Inhibits EC proliferation; reduces EC migration

Soil and Seed

• Def
• What cancers form bone metastases

• Cancer cells often display a selective pattern of metastasis; cells released from primary tumours often home to proferred target issues
• Prostate and breast cancers

S&S Hypothesis

• By whom
• Did what
• Challenged?

• Stephen Paget
• Analysed 900 autopsy records for patients with different primary tumours; identified non-random metastatic tissue distributition.  "Seed" "Soil"
• James Ewing; claimed that circulatory patterns and mechanical influences were suficient to explain organ-specific metastasis
• Current evidence supports a role for both

Soil and Seed - Mechanical factors

• Def
• Capillaries

• Blood flow will initially direct cancer cells released from the primary tumour; first pass organ = the first organ encountered that lies downstream of the primary tuour in the circulatory system
• Capillary vessel diameter (3-8 microm) allows passage of RBCs (~7 microm); cancer cells (~20 microm) will arrest in capillary bed by size restriction; Will also embolise with platelets and bld cells

1. The prevailing model of metastasis suggests that a minority of tumour cells with metastatic potential escape form distant metastases
2. In humans, breast cancer expression profiles of the primary tumour can predict disease outcome.  The capacity to metastise is displayed by the whole tumour-cell population. A poor-prognosis signature strongly predicts the development of metastasses, incontrast to the good=prognosis signatures
3. A variant model for the primary tumours that have high metastatic capacity and displays the poor-prognosis signature.  Within this group, subpopulations of cells also display a tissue-specific expression profile predicting the site of metastasis

Targeted metastasis of primary breast cancer cells

• Cancer vs. nl cels express large amount of
• These do what

• CXCR4 chemokine receptor
• Chemokines (such as CXCL12) recognise these receptor are released by certain organs such as bone marrow, liver, and lung (kidney = low)
• Breast cancer cells with elevated expression of IL-11, MMP-1, CXCR4, CTGF (connective tissue growth factor) and osteopontin are associated with high bone metastatic potential in vivo
• Breast cancer populations that did not express any of these genes did not generate bone metastases

• Osteolytic - resulting from breast/lung primaries
• Osteoblastic - resulting from prostate primaries

Osteolytic bone metastases RANK/RANKL

How it works

• Production of PTH or PTHrP, IL-1, IL-6, and IL-11 by tumour cells stimulates RANKL expression
• Some of these factors (for example, PTHrP) also decreases the production of OPG
• Signalling through RANK activates transcription factors such as AP1 and NF-kappaB leading to the differentiation of osteoclast progenitors into mature osteoclasts

Osteoblastic bone mets in prostate ca

Tumour cells release ___ to promote osteoblast activity and bone formation

FGF - Fibroblast growth factor

BMP - Bone morphogenic proteins

PDGF - Platelet-derived GF

TGF-beta - Transforming GF-beta

Prostate

Proteases, such as ___ and __ are induced by activators, such as _____

prostate-specific antigen and plasmin

urokinase-type plasminogen activator (uPA)

·      Proteases, such as prostate-specific antigen and plasmin, are induced by activators, such as urokinase-type plasminogen activator (uPA)

These proteases:

• Activate latent TGF-beta
• Release insulin-like growth factor (IGF) from inhibitory binding proteins (IGFBPs)
• Inactivate the osteolytic factor PTHrP

• Interactions between tumour cells and osteoclasts cause not only osteoclast activation and bone destruction, but also aggressive growth of the tumour
• Bone resorption releases bone-derived growth factors, including TGF-beta and IGFI, and raises extracellular [Ca]
• The GFs bind to receptors on the tumour-cell surface and activate signalling through pathways that involve SMAD and mitogen-activated protein kinase (MAPK)
• Extracellular Ca binds and activates Ca pump. Signalling through these pathways promotes tumour-cell proliferation and production of PTHrP
• Other cytokines might be involved, such as IL-1, IL-6, IL-11, and IL-18

How many different types of cancer/

What accounts for >50% of all new cases

>200

Breast, lung, colorectal, and prostate

Epithelial differentiation varies in (3)

This reflects differences in:

1. Susceptibility to cancer
2. Risk factors
3. Molecular genetic pathways

Intestine

• Cell types
• Function
• High turnover rate?
• Exposed to carcinogens how?

• Non-stratified, "simple," columnar epithelium
• Absorptive, mucus-secreting fx
• Yes; cells shed from surface (anoikis); Replaced from proliferative pool; irreversible loss of proliferative capacity during differentiation
• Diet or gut pathogens

Urinary bladder

• Lined with what
• Proliferation how
• Rate of apoptosis

• Urothelium (specialised-barrier epithelium)
• Proliferation "switch:" mitotically-quiescent but high regenerative capacity in response to injury/infection
• Low rate of apoptosis

Drugs/metabolites

Environment

Industrial

70% superficial papillary dx

20% invasive dx

10% carcinoma in situ

superficial papillary dx

• Rate of recurrence
• Rate of progression
• Predictive markers of progression?

• High
• Low (<5% Ta & ~30% T1)
• None

Invasive disease

• Outcome

Carcinoma in situ

• Def
• Rate of progression

• An early form of carcinoma defined by the absence of invasion of surrounding tissues.  IOW, neoplastic cells proliferate their nl habitat
• High rate of progression

Bladder ca

• Dx made on
• Bladder cancer is staged according to what
• Low grade
• High grade

• histopathology
• according to depth of invasion/spread
• well differentiated, good prognosis
• poorly differentiated (anaplastic), poor prognosis

• Mouse uroplakin II gene promoter to target expression of p53DN (dominant negative) mutant in urothelium of transgenic mice
• p53DN binds and stabilises endogenous wilt-type p53 -- induces nuclear abnormality, hyperplasia, and occasionally dysplasia, not carcinoma
• Transgenic mouse with activated Ha-Ras induced urothelial hyperplasia
• Concurrent expression of p53DN and Ha-Ras induced urothelial hyperplasia but failed to accelerate tumour formaiton
• Crossing activated Ha-ras transgenic with p53-/- mice to express activated Ha-ras in the absence of p53 results in early-onset bladder tumours (low-grade papillary or high-grade)

• p53 deficiency predisposes the urothelium to hyperproliferation, but is insufficient for bladder tumorigenesis
• Reduction in p53 dosage (eg, transgenic mice expression p53DN or heterozygous p53 knockouts) does not synergise with Ha-ras to induce bladder tumours; and
• Complete loss of p53 is a prerequisite for collaborating with activated Ha-ras to promote bladder tumorigenesis

1. Progressive telomere shortening limits the replicative potential of human tells
2. Human ca cells must acquire capacity to maintain telomeres (ie, through telomerase activation)
3. In the lab mouse, telomeres are 4-6x longer than humans and telomerase is expressed more widely
4. In mouse tumorigenesis there is no selection pressure to acquire replicative potential

1. Repapitulates original tumour behaviour in immunocompromised mice and 3d models
2. Immortal
3. uncloned - represents tumour biodiversity

1. Highly selective - minority of cancers 'take'
2. Selective pressures from long-term adaptation to culture
3. Have acquired multiple genetic changes
4. May not be useful background to investigate candidate ca genes
5. Beware of contamination (other cell lines, micoplasma spp.)

Cross-contamination by cell-lines

• Extensive contamination arose due to
• Estimated __% of cell lines cross-contaminated
• Why is this a problem? (4)
• How to detect/eradicate problem?

• Widely distributed between labs; poor practice and unawareness of problem; HeLa - robust cell line that competes out other cells in a mixed culture
• 20%
• 20% of publications use cross-cont. cell lines; flawed research and wasted funding;anthithesis of fundamental scientific principles; loss of scientific cred
• Karyotyping, HLA typing, and isozyme profiling; Short-tandem-repeats analysis (PCR multiplex amplification of polymorphic loci and separation on a gell - unique profile obtained for that DNA sample source); Use product sizes to generate series of numbers as a barcode

Mycoplasma spp

• What is
• Reported in __% of cell lines
• spreads how

• small, self-replicating prokaryote that lacks a cell wall and has the ability to cyto-absorb onto host cells
• 15-35%
• Spreads between cell lines, through media and reagents, or oporator droplet infection during culture

• Microbiological culture
• PCR
• DNA staining for extrnuclear DNA (bis-benzimide)

1. Altered DNA, RNA, and protein synthesis
2. Induction of chromosomes abberations
3. Altered cell membrane composition
4. Altered cell activation and cytokine expression
5. Influence on signal transduction
6. Altered growth characteristics

1. Obtain cell cultures from reliable suppliers
2. Good cell culture practice
3. Use 0.2 micron filters
4. Implement routine testing

E-cadherin

• Does what
• Absent where
• Inactivation of E-cadherin in NHU cells does what

• Acts like a zipper to hold neighbouring cells together
• Absent from most invasive cancers
• Overexpressed chimeric protein; extracellular domain of MHC class I with transmembrane and intracellular domains of E-cadherin; LEADS to proliferation, migration, and invasion

Experimental models (mouse vs. human)

• Rodent systems provide...
• Human systems...
• Nl human cell models are useful because...

• Rodent systems provide insight, but may not translate directly to man
• Human systems: no single model appropriate for all cancers; incomplete understanding of nl biology
• Nl human cell models are useful because: Insight into nl reg. pathways in tissue-specific background; role of specific pathways or genes in cancer

Stochastic Model of Carcinogenesis

• Def
• Theory states that

• Every cell within a tumour is potentially tumour-initiating
• Theory states that it is impossible to predict which kind of cells will be T-IC

• Differentiated cells which no longer capable of division (cardiac muscle, neurons, short life span and continually replaced)
• Proliferative cells (amplifying cells)
• Stem cells (all adult tissue classically demonstrated in digestive tract, skin, blood; divide to produce daughter cells that either differentiate or remain as stem cells)

Blood cell lineage

• All bld cells develop from where
• Immediately differentiated?

• Pluripotent stem cell in the bone marrow
• Precursors of differentiated cells undergo several rounds of cell division as they mature (transit amplifying cells); proliferation ceases at terminal stages of differentiation)

• A cell that has the ability to divide (self-replicate) for indefinite periods -- throughout lifetime of organism
• Can differentiate to the many different cell types that make up the organism (under the right conditions or given the right signal)

Amplifying cells

• What are
• Daughters committed to differentiation go through...

• Stem cells in many tissues divide only rarely (disruption of quiesence leads to premature exhaustion of the stem cell pool; can cause haematological failure under stress conditions)
• Gives rise to amplifying cells
• Daughters committed to differentiation go through a limited series of rapid division
• Each stem cell division gives rise to 8 terminally differentiated progeny

Cancer stem cells

• Are all cells equal in cancer?
• __ tumour cells needed to initiate tumour growth
• Clonogenic?
• All cells are tumourigenic?

• No!
• 10^6 tumour cells needed to initiate tumour growth
• Small subset of tumour cells are clonogenic, both in vitro and in vivo
• Tumours are heterogeneous and only a rare sub-set of cells are tumourigenic
• Model predicts that a distinct phenotype will be T-IC, whilst the bulk of the tumour will be non T-IC

Stem cells are long-lived cells; would require fewer mutations;quiescence

Early progenitors

Must acquire ability to self-renew

• TMPRSS2:ERG is an initiating mutation in 60% of prostate cancers and is present in the stem cell population

• Twin studies: the initiating mutation (AML10ETO fusion) has a foetal origin and occurs in a single long-lived cell. Further work showed that this cell was a stem cell
• Initiating mutation does not cause leukaemia outright but is present in foetal blood
• Dx of leukemia: 2-10 years after initiating mutation
• Patients in remission are AM1-ETO+

Page 54 - Cell sort and test function

Page 55 - Knock-out/knock-in mouse colon cancer

1. Quiescence (most cytotoxic drugs target proliferative cells; stem cells are insensitive to drug)
2. Target not expressed
3. Radioresistance (activation of DNA damage response)
4. ABC transporters (efflux drugs)
5. Express anti-apoptotic genes (bcl-2)

Androgen response hormonal therapy

• Treatment in prostate cancer

• Hormonal therapy has been used for treatment of advanced prostate cancer
• ~80% of prostate cancers initially respond to hormonal therapy; more than half of the responders gradually become resistant to this therapy
• Androgen-responsive to androgen-unresponsive state
• ***Prostate CSCs do not express the androgen receptor***

• Cancer Genome Project: (Cancer Gene Census; COSMIC; CGP Cancer Cell Line Project; CGP Copy Number Analysis in Cancer)
• Cancer Genome Anatomy Project: (SAGE; Chromosome analysis)

1. Potential to initiate tumours in vivo
2. Self-renewel
3. Capacity to differentiate into phenotypes typical of pt's tumour
4. Quienscence/slow-cycling

1. Established cancer cell lines
2. Carcinogen treated animals
3. Xenogenic transplantation (subq)
4. Transgenic mice (genetic modification of normal cells in vivo)

Advantages and Disadvantages of:

 Established ca cell lines

Adv: Retains characteristics of originating tumour

Dis: Genotypic/phenotypic drift due to long-term adaptation to culture

Adv: Provides info in initiation/promotion stages of cancer development

Dis: Neoplastic transformation pathways in animals are not directly relevant to man

Adv: may provide additional info over cell culture as forms 3D tumour in an in vivo environment eg, angiogenesis

Dis: Host interactions may not be relevant -- cross-species, tissue specific

Adv: Investigate specific tumour development eg, by combining tissue-specific promoter with cancer-predisposing genetic modification

Dis: Neoplastic transformation pathways in animals are not directly relevant to man