Cancer Flashcards
What is a Metaplasia?
Reversible chane in a tissue where one type of adult cells is replaced with another adult cell type
- usually epithelium
- can be physiological, is normally functionla (e.g. reflux)
What is Dysplasia?
an abnormal pattern of growth in which some of the cellular and architectural features of malignancy are present
•pre-invasive stage with intact basement membrane
–> no invasion meaning good chances if revmoved!
What is a Neoplasm?
An abnormal, autonomous proliferation of cells unresponsive to normal growth control mechanisms
What is a Tumor?
A neoplasm that forms a mass
What are the characteristical changes in dysplasia?
- loss of architectural orientation
- loss in uniformity of individual cells
- nuclei: hyperchromatic, enlarged –> increased nucleus: cytoplasm ratio
- mitotic figures: abundant, abnormal, in placeswhere not usually found
What are the characteristics of a benign tumor?
-
Non-invasive
- not into other tissues
- no metastisis
- usually well-differentiated –> characteristics of tissue of origin
- encapsuled
- slow growing
- normal mitosis
What are the cases in which a benign tumor can be fatal?
- When it sits at the wron spot
- brain, meningines, pituitary
- When it secretes something dangerous
- insulinoma
- pituitary adenoma
- gets infected
- bleeds or ruptures
- causes torsion of organs (e.g. ovaria cyst)
What is a Metastisis?
•A metastasis is a discontinuous growing colony of tumour cells, at some distance from the primary cancer
What are the characteristics of a malignant
- invade surrounding tissues
- spread to distant sites
- no capsule
- well to poorly differentiated –> can’t see characteristics of tissue of origin
- rapidly growing
- abnormal mitoses
How would you call a benign epithelial tumor of surface epithelium (e.g. skin, bladder)
Papilloma
What is a papilloma?
A benign tumor of surface epithelium (e.g. skin, bladder)
How would you call a benign tumor of glandular epithelium?
Adenoma
What is an adenoma?
A benign tumor of glandular epithelium
What is a carcinoma?
•A malignant tumour derived from epithelium
How do you call a malignant tumour derived from epithelium?
Carcinoma
Name some examples of different form of carcinomas
- squamous cell carcinoma
- adenocarcinoma,
- transitional cell carcinoma
- basal cell carcinoma
–> dependant on the tissue they derive from
How would you call a benign tumor of soft tissue?
E.g. lipoma, osteoma
–> ending oma, dependant on the tissue type
What is an osteoma?
Benign tumor derived from bone
How would you call a malignant tumour derived from connective tissue (mesenchymal) cells?
Sarcoma
What is a sarcoma?
a malignant tumour derived from connective tissue (mesenchymal) cells
Name differnet types of maglignant soft tisseue tumors in
- Fat
- bone
- cartilage
- Fat
- Liposarcoma
- Bone
- Osteosarcoma
- Cartilage
- Chondrosarcoma
How would you call malignant tumors of
- striate muscle
- smooth muscle
- Nerve sheath•
- Muscle
- striated = RhabdomyoSARCOMA,
- smooth = LeiomyoSARCOMA
- Nerve sheath = Malignant Peripheral Nerve Sheath Tumour
What is Leukemia?
A malignant tumor of bone marrow
(can usually be seen in blood cells )
How would you call a malignant tumor of bone marrow?
Leukaemia
What is a Lymphoma?
A malignant tumour of lymphocytes (usually) in lymph nodes
–> can normally be seen in the tissue
How would you call a malignant tumour of lymphocytes (usually) in lymph nodes?
Lymphoma
What is a teratoma?
A tumor derived from germ cells –> Can differentiate into any adult cell type
How would you call a tumor of germ cells that can normally differentiate into any type of adult cell?
A Teratoma
Are teratomas usually benign or malignant?
- In Males: almost always malignant
- In Females: Most are benign
What is a HAMARTOMA?
- localised overgrowth of cells and tissues native to the organ.
- cells are mature but architecturally abnormal
- –> When just looking at the cell, they look completely normal but are just organised in a different way
- common in children, and should stop growing when they do,
How do you call localised overgrowth of cells and tissues native to the organ, just with abnormal architecture?
Hamartoma
What is a Anaplastic tumor?
When there is no differentiation seen and no origin cell type can be determined
What is the grade of a tumor?
The level of differentiation (high grade= low differentiation)
What is the stage of a tumor?
Which Criteria are used to determine it?
Stage of a tumor is how far the tumor has spread
–> TNM
- Tumor
- Node
- Metastisis
Explain the relationship between grade and stage
- High grade tumors tend to be high stage
- BUT: stage is the most impoertant factor when it comes to prognosis!
List five morphological features that allow assessment of the differentiation of a tumor
- loss of architectural orientation
- loss in uniformity of individual cells
- nuclei: hyperchromatic, enlarged
- mitotic figures: abundant, abnormal, in places where not usually found
- Evidence of normal function still present production of:
keratin,
mucin
bile
hormones
Which factors influence the speed of cell devision?
- embryonic vs. adult cell
- complexity of systems (e.g. rapid devision in yeast cells)
- Necessitiy for renewal (intestinal epithelium vs. hepatocytes)
- state of differentiation (e.g. neurons/cardiomyocytes that don’t replicate)
- Tumor cells ?? –> different mechanisms
What happens when the appropriate regulation of cell devision goes wrong?
Which factors might lead to it?
Cancer might develop that
- mutation in TSG/oncogenes (–> + aneupleudy)
- chormosome instability (loss/gain of chromosome)
- abnormal mitosis –> mistake in cell cylce regulatory proteins
- Contact inhibition of growth –> cells don’t stop growing when they reach the edges of a space
What is the cell cycle?
Orderly sequence of events in which a cell duplicates its contents and divides in two
What are the different phases in the cell cycle of an eucariotic cell?
- M Mitosis
- Interphase
- G0
- cell cycle machinery dismantled –> cell fulfils normal function
- G1
- phase (Gap) - Decision point: are specific structures (e.g. centromeres) duplicated?
- S
- DNA dublication
- G2
- phase (Gap) - Decision point
- check new DNA for mistakes
- G0
Explain the structure of a centrosome
Consists of two centrioles (barrels of nine triplet microtubules
What happens during the S-phase of the cell cycle?
- DNA replication
- Replication of organelles (e.g. golgi, mitochondria, centromeres)
- Increased protein synthesis
Explain the life cycle and replication of the centrosomes
Split in G1 and are fully replicated at the end of S-phase
What happens during the prophase of mitosis?
- condensation of DNA
- centrosomes migrate to opposite sides in cell
- spindels start to form
Explain the formation of the mitotic spindles
- ASTERS form (radial micturtubule arrays) around each centromer –> Microtubule organisation centre)
- Tadial arrays meet
- Polar micrutubules form (= radial microtubules that meet in middle)
What happens during the early prometaphase in the cell cycle?
- Chromosomes aligned at equator of the spindle
In the early:
- break down of nuclear membrane
- spindles are largely complete
- attach of chromosomes to the spindles via kinetochores at the centromer region of chromosome
What is the function of the centrosome?
- microtubule organizing center (MTOC) (also in interphase!!)
- mitotic spindle
What happens during Anaphase A in mitosis?
- Breakdown cohesin (protein that holds togehter sister chromatids)
- Microtubules get shorter
- Daughter chromosomes pulled toward opposite spindle poles
What happens during Anaphase B?
- Daughter chromosomes migrate towards poles
- Spindle poles (centrosomes) migrate apart
- so cytokinesis does not happen too close to chormosomes and they have enough space
What happens during telophase of mitosis?
Chromosomes arrive at spindles
Nuclear envelope resembels
Assembly of contractile ring (for cytokinesis), made of actin and myosin
What are the different factors required in the Spindle Assembly checkpoint?
(aka Mitotic Checkpoint)
- CENP-E (signals when not attached)
- BUB protein kinases (signals when attached)
BUBs dissociate from kinetochore when chromosomes are properly attached to the spindle
Explain the different things that can go wrong at the Spindle assembly checkpoint
Missatachmen of spindles to kinetochore might lead to aneupleudy
- Meroteilic attachment (2 differnt spindels attach to the same chromatid)
- one spindle attaches to both sister chromatids of one chromosomes
How might alterations in the cell cycle lead to aneupleudy?
- errors in DNA or centrosome replication
- might lead to multipolar spindles
- or in aberrant (irrtümliche) citokinesis
Explaint the exploitation of chromosome-missegregation as an anti-cancer treatment
- Inhibition of Checkpoint kinase (CHKE1 and CHKE2)
- Normally : Serine threonine kinase activation holds cells in G2 phase until all is ready
- inhibits attachment-error-correction mechanism
- inhibition leads to untimely cell transition to mitosis –> not ready yet because there is still an error
- Normally : Serine threonine kinase activation holds cells in G2 phase until all is ready
- Taxanes and vinca alkaloids
- Alters microtubule dynamics
- Produces unattached kinetochores
- Causes long-term mitotic arrest.
What are the different check points in the cell that drive/ control cell proliferation?
How is that important in cancer?
- G1: Checkpoint in G1 (Growth Factor dependant)
- initiate progession
- G2: Damage of DNA
- M: Sister chromatid alignment
All these checkpoints can be altered by tumors!!
Explain the concept of De-regulation of cell cycle during tumorigenesis
Tumors can inhibit entering into G0 phase where the cell just does its nomal function and promote proliveration
Summarise the intracellular signaling cascade that takes place during aktivation of cell proliveration by a growth factor
They set of the signal cascade:
- Response to extracellular factors
- Signal amplification
- Signal integration
- Modulation by other pathways
- Regulation of divergent responses
Explain the process of a growth factor binding to a growth factor receptor (RPTK)
In present of a (dimeric) ligand–> growth factors
- Receptors form dimers
- auto-crossphorylation causes activation of receptor
- (phosphorylated AA in receptor)
What happens during the late prometaphase
- Microtubule from opposite pole is captured by sister kinetochore
- Chromosomes attached to each pole congress to the middle
- Chromosome slides rapidly towards center along microtubules
Explain the process of phosphorylation
Exchange of Hydroxy-group for a phosphate group (ATP dependant, katalysed by Kinase)
Only 3 AA can be phosphorylated:
- Serine
- Threonine
- Tyrosine
What does the auto-phosphorylation in the tyrosine kinase domain of a growth factor receptor cause?
It causes receptor activation leading to
- kinase cascade
- binding of adaptor proteins
Explain the protein kinase cascade
An activated kinase will often activate further kinases
- will lead to signal amplification
- and gives the opportunity for better regulation because
- Phosphorylation in reversible (by a phosphatase) means that individual kinases can be inactivated
What happes during the anaphase in mitosis?
- Paired chromosomes separate to form two daughter chromosomes
- Cohesin holds sister chromatids together
- There is a anaphase A+ B
What happens at the transition out of metaphase before Anaphase can start?
Explain its mechanism
Spindle Assembly checkpoint
- are all chromosomes connected hthe kinetochore?
- When kinetochores are connected they stop signaling
- At the checkpoint it is waited, until there is no more signaling (i.e. all chormonsomes are connected) until Anaphase can start
What happens during Metaphase of Mitosis?
Starts once MT are attached to Kinetochores
- align at equator of spindles
What is the difference in chromosome movement between Prometaphase and Metaphase?
In Prometaphase: try to bring a bit of order into the chromosomes + tries to get it at the right angle (picture)
In Metaphase: Alignment at eqator of spindle (begins once Microtubule are attached)
Explain the process of Cytokinesis
Actin myosin ring between two cells forms and contracts
- new membrane is inserted
- midbody remains (connection between the two daughter cells)
- also midbody gets seperated
What if something goes wrong during the cell cycle? e.g Cell is not big enough or DNA damage
- Cell cylce arrest at checkpoints (G1+ spindle checkpoint)
- can be temporarily and resolved e.g. with DNA repair
- Apoptosis
- if DNA damage to great, chromosomal abnormalities or toxic agents
What are the physiological triggers that tell the cell to enter the cell cycle and devide?
Most cells in body are in G0 phase and fulfil their normal functions:
Exit from G0 highly regulated - requires growth factors and intracellular signalling cascades
Name examples of two growth factors
Which receptor do they bind to?
Epidermal growth factor (EGF)
Platelet-derived growth factor (PDGF)
–> both bind to Receptor Protein Tyrosine Kinase (RPTK)
How can phosphorylation of an Amino-Acid lead to altered protein function?
The added phosphate group (negatively charged) can alter protein function by:
- causing a change in shape (conformation) leading to change in activity (+ve or –ve)
- creating a docking site for another protein
What is An Amphiletic Microtubule attachment?
normal (centrosome-kinetochore1, centrosome-kinetochore2) pulled to 2 ends
What is an Syntelic attachment of microtubule to the chromosome?
Same Centrosome
2 microtubules from 1 centrosome connect to both kinetochores, pulled to one end – one cell has duplication, other has one less chromosome
What is Merotelic attachment of Mictotubule to the Chormosome?
Ripped
2 microtubules (each from different centrosome) connect to 1 kinetochore, ripped apart, both sister cells with one less chromosome
What is a Monotelic attachment of Micturubule to Chromosome?
•one microtubule attached to one kinetochore (no microtubule attached to other Kinetochore)
In which cell cycle phase does the Mitotic Checkpoint Happen?
Between Meta and Anaphase
–> Checks for Attachment of Telomeres
What is the most vulnerable phase of the cell cycle?
Why?
Mitosis because
- Cells are more easily killed (irradiation, heat shock, chemicals)
- DNA damage can not be repaired
- Gene transcription silenced
What happens to protein synthesis in S-phase of the Cell cycle?
initiation of translation and elongation increased; capacity is also increased
Which proteins stop processing in the cell cycle if damages/ errors are present at Checkpoints?
- Checkpoint kinase (CHKE1 and CHKE2) – When kinase signals it holds the cell and stops transition into next phase
- inhibition leads to untimely cell transition to mitosis
- exploited by Vinca Alkaloids and Taxanes
What are the four main modalities for cancer treatment?
- Surgery
- Chemotherapy
- Radiotherapy
- Immunotherapy
Which type of genetic mutations might cause cancer?
- Chromosome translocation
- Gene amplification (copy number variation)
- Point mutations within promoter or enhancer regions of genes
- Deletions or insertions
- Epigenetic alterations to gene expression
- Mutations Can be inherited
What are the overall characteristics of systemically administerd chemotherapeutic agents?
- IV or PO administration
- Non “targeted” – affects all rapidly dividing cells in the body –> causes many of the side-effects
How do you call a chemotherapeutic agent administerd post-surgery?
adjuvant
How do you call a chemotherapeutic agent administered pre-operatively?
Neoajuvant
What is the MOA of Akylating Agents?
Interfere with DNA synthesis
- Add alkyl groups to guanine residues in DNA
- Cross-link (intra, inter, DNA-protein) DNA strands and prevents DNA from uncoiling at replication
- Trigger apoptosis (via checkpoint pathway)
- Encourage miss-pairing - oncogenic
What is the MOA of pseudo-alkylating agents?
Add platinum to guanine in DNA, otherwise causes the same effect as Alkylating agents
- Cross-link (intra, inter, DNA-protein) DNA strands and prevents DNA from uncoiling at replication
- Trigger apoptosis (via checkpoint pathway)
- Encourage miss-pairing - oncogenic
Which drug class do
Chlorambucil, cyclophosphamide, dacarbazine, temozolomide
belong to?
Alkylating agents
Which drug class to
carboplatin, cisplatin, oxaliplatin
belong to?
Pseudo-alkylating agents (add platinum group)
What are the side-effect of alkylating and pseudoalkylating agents?
- hair loss (not carboplatin),
- nephrotoxicity,
- neurotoxicity,
- ototoxicity (platinums)
- nausea, vomiting
- diarrhoea
- immunosuppression
- tiredness
Explain the MOA of Anti-metabolites
Interfere with DNA synthesis by
- Masquerade as purine or pyrimidine residues leading to
- inhibition of DNA synthesis,
- DNA double strand breaks
- apoptosis
Which drug class do
methotrexate, 6-mercaptopurine, decarbazine and fludarabine, 5-fluorouracil, capecitabine, gemcitabine
belong to?
Anti-metabolites
Explain the MOA of Anthracyclines
- Inhibit transcription and replication by intercalating (i.e. inserting between) nucleotides within the DNA/RNA strand
- thereby: inhibit Tropoisomerase II
- Also block DNA repair - mutagenic
- Create free oxygen radiacals that damage DNA and cell membrane
*
What are the side-effects of anti-metabolites?
- Hair loss (alopecia) – not 5FU or capecitabine
- Bone marrow suppression causing anaemia, neutropenia and thrombocytopenia
- Increased risk of neutropenic sepsis (and death) or bleeding
- Nausea and vomiting
- Mucositis and diarrhoea
- Palmar-plantar erythrodysesthesia (PPE)
- Fatigue
Which drug class do
doxorubicin, epirubicin
belong to?
Anthracyclines
What are the side effects of Anthracyclines?
- Cardiac toxicity (arrythmias, heart failure) – probably due to damage induced by free radicals
- Alopecia= spot hair loss
- Neutropenia
- Nausea and Vomiting
- Fatigue
- Skin changes
- Red urine (doxorubicin “the red devil”)
Explain the MOA of Vinca Alkaloids and taxanes
Microtubule inhibiting drugs–> leading to mitotic arrest in mitosis by
- inhibiting assembly (vinca alkaloids)
- disassembly (taxanes) of mitotic microtubules
Which drug type are the micrutubule targeting drugs?
Vinka alkaloids and taxanes
What are the side-effects of vinka alkaloids and taxanes?
- Nerve damage: peripheral neuropathy, autonomic neuropathy
- Hair loss
- Nausea
- Vomiting
- Bone marrow suppression (neutropenia, anaemia etc)
- Arthralgia (joint pain)
- Allergy
Explain the MOA o fTopoisomerase inhibitors
Topoisomerase is responsible for adding/removing supercoils during DNA replication + protect the free ends of DNA from aberrant( abweichenden, verwirrenden) recombination events
–> alter binding of the complex to DNA and allow permanent DNA breaks
Which class of drug do
Topotecan, irinotecan, etoposide
belong to?
Topoisormerase inhibitors
Topotecan and irinotecan (topo I)
and etoposide (topoII)
What are the side effects of Topoisomerase inhibitors?
- (irinotecan): Acute cholinergic type syndrome – diarrhoea, abdominal cramps and diaphoresis (sweating). Therefore given with atropine
- Hair loss
- Nausea, vomiting
- Fatigue
- Bone marrow suppression
•
•
Through which mechanims might cancer cells develop resistance against the treatment?
- Upregulation of DNA repair mechanisms and DNA damage is repaired
- DNA adducts replaced by Base Excision repair (using PARP) –> specific repair mechanism
- Drug effluxed from the cell by ATP-binding cassette (ABC) transporters
What are the main characteristics of cancer cells?
(Hallmarks of cancer)
- Self –sufficient
- Insensitive to anti-growth signals
- Anti-apoptotic
- Pro-invasive and metastatic
- Pro-angiogenic
- Non-senescent
- Dysregulated metabolism
- Evades the immune system
- Unstable DNA
- Inflammation
Explain the role of receptor over-expression in cancer
Over-expression of some receptors (expecially growth factor receptors) can be seen in many cancers:
- HER2 – amplified and over-expressed in 25% breast cancer
- EGFR – over-expressed in breast and colorectal cancer
- PDGFR- glioma (brain cancer)
Explain the relationship with overexpression of growth factor receptor ligands and cancer
Vascular Endothelial Growth Factor is overexpressed:
prostate cancer, kidney cancer, breast cancer
–> leading to increased Kinase cascade and signal amplification
–> angiogenisis in cancer
What is the MOA of a dual kinase inhibitor?
Inhibit 2 protein kinases to prevent cancer cells switching to other pathways (which needs another kinase)
But: are also more toxic
Explain the role of Constitutive (ligand independent) receptor activation in cancer
EGFR (lung cancer) epidermal growth factor receptor
FGFR (head and neck cancers, myeloma)
Leading to. Kinase cascade and signal amplification
What is the advantage of monoclonal cancers therapeutically?
A single “wire cutting”, inhibition of cellular pathway might be enough to kill the cell
But: only works for some cancers, other might regulate other pathways up
What do normal cells require in order to grow and poliferate?
How is that different from cancer cells?
Normall cells need growth factors to proliferate
Cancer cells don’t need them/ develop strategies to get higher stimmulation
Explain the MOA of small molecule inhibitors
It is a type of targeted cancer treatment
- bind to kinase domain of tyrosine kinase
- can also bind to intracellular kinase pathways
- block autophosphorilation and downstream regulation
- e.g. Gleevac
What is the general target of monochlonal antibodies in the treatment of cancer?
They are normally designed to target + inhibit growth factor receptors
Explain the MOA of the use of monochlonal antibodies in cancer treatment
They are designes to stop the
- Groth Factor receptor system
- neutralise the ligand
- inhibit dimerisation
- cause internilisation
- might induce phagocytosis + cytolysis
What are the types of targeted cancer treatment?
- Monoclonal antibodies
- Small molecule inhibitors
What is the overall principle of targeted cancer treatment?
Normally: targeted treatment acts on receptors and modulates cancer halmarks
- VEGF alters blood flow to tumor
- AKT inhibitors block reisistance to apoptosis
–> without toxicity of other treatments observed
Explain the use of anti-sense oligonucleiotides in cancer treatment
- Single Stranded, DNA like molecole (17-22 nucleotides long)
- Binds to target mRNA hinders translation of proteins by break down of mRNA
Explain the use of RNA interference in cancer treatment
- SS complementary RNA
- (lagging behind oligosense nucleotides)
What are the main disadvantages of targetet therapy of cancer?
Resistance
What are the main resistance mechanism in targeted cancer therapy?
- Mutations in ATP-binding domain
- Intrinsic resistance
- Different gene mutations/ that might not make the targeted receptor/pathway responsible
- Intragenic mutations
- Upregulation of downstream or parallel pathways
What are the endings used to classify different antibodies (e.g. used in anti-cancer treatment)
- -momab (derived from mouse antibodies)
- -ximab (chimeric) e.g cetuximab
- -zumab (humanised) e.g. bevacizumab trastuzumab
- -mumab (fully human) e.g. panitumumab
Explain the MOA of glivec
In which type of disease is it used?
Glivec is a small molecule inhibitor and targets the ATP binding region within the kinase domain, inhibiting kinase activity of ABL1
–> Used in the treatment of specific form of CML
Explain the use of anti-sense oligonucleiotides in cancer treatment
- Single Stranded, DNA like molecole (17-22 nucleotides long)
- Binds to target mRNA hinders translation of proteins by break down of mRNA
Explain the use of RNA interference in cancer treatment
- SS complementary RNA
- (lagging behind oligosense nucleotides)
Briefly Summarise the overall events that happen from Growth factor bindin to receptor to change in gene expressions that trigger mitosis
- Mitogenic Growth Factor binds to Growth Factor receptor
- Dimerisation of Receptor causes activation of tyrosine kinase
- Tyrosine Kinase recruits receptor and adaptor proteins (e.g G-protein RAS)
- Adaptor Proteins set off Kinase cascase
- KInase cascade causes change in gene expression and activates mitotic proteins
What is the result of phosphorylation of the intracellular tyrosine caused by dimerisation of the Growth factor receptor after Growth factor binding?
Phosphorylation of thyrosine caused by dimerisation of the growth factor receptor provides docking sites for adapter proteins
What are tha characteristics of adaptor proteins and what is their function?
They tend to facilitate binding between different proteins instead of having an enzymatic function itself (but can have enzymatic function) –> bring proteins together (protein-protein interactions)
Thereofre they are
- modular –> many protein-binding molecules and domains (structural and functional units that can be seen in many other molecules)
What are adaptor proteins?
Proteins that bind to the phosphorylated Thyrosine Kinase in at the receptor membrane and assist in further signal transduction
Explain the role of Grb2 in the activation of RAS
The exchange factor SOS binds to the SH3 domain of Grb2 and is activated by it
- SOS exchanges GDP bound to RAS to GTP and thereby activates it
How does RAS get activated and inactivated?
Activation
- GDP is replaced by GTP (done by exchange factors e.g. SOS)
Inactivation
- by GAP (GTPase activating protiens) that turn GTP to GDP again
What kind of protein is Grb2?
What is its function?
Growth factor receptor-bound protein 2 is an adaptor protein
Explain the structure and function of Gbr2
Is an adaptor protein–> brings proteins together
- It has three domains
- SH2 domain bindin to the phosphorylated tyrosine of GFR
- 2 SH3 domains bindig to other molecules
- proline-rich regions of other molecules
What is the Name of the different tyrosine Kinases that are activated in the ERK cascade?
- Kinase
- RAF
- Kinase
- MEK
- Kinase
- ERK
Explain the involvement of the ERK cascade in the formation of cancer
B-Raf is an oncogene - mutationally activated in melanomas
What does activation of the ERK cascase (or other kinase cascades) lead to?
- Change in protein activity
- Change in gene expression
What are Cdks?
When are they present in the cell?
How are they regulated?
Cyclin-dependent kinases
- always present in human cells but
- only activated with cyclins and phosphorylation
What are cyclins?
When are they present in the cell?
How are they regulated?
Cyclins
- only present at specific times in cell (often: mitosis)
- regulated at the level of expression
- synthesised and then quickly degraded
What is the function of cyclins?
To bind to Cdks and activate them
- quickly degraded so good as tightly control funciton in cell
What is the gross function of activated Cdks?
They phosphorylate proteins (on Serine or Threonine) to drive cell cycle progression
- during mitosis
- to initiate mitosis
Explain the activation and regulation of Cdks activity
Regulated in 3 steps
- Cyclin binding
- Activating Phosphorylation by CAK (cdk activating kinase)
- Removal of inactivating phosphorylation by cdc25
Explain how differnet cycline and Cdks are present in the cell cycle
At each step of the cell cycle different Cdks and cyclins are present and needed
- During Mitosis: M-Cdk (Cdk1+ Cyclin B)
- During G1: G1/S Cdk (Cdk2 + Cyclin E)
- During S: S-Cdk (Cdk2+Cyclin A)
How does Growh factor binding lead to the initiation of the cell cycle?
- Leading to set of of the ERK cascase
- Expression of c-Myc (transcriptionfactor)
- stimmulates transcription of other factors –> cyclin D –> Cdk 4/6 Complex initiates cell cycle
How is the expression of cyclin/Cdk reguates?
- By transcriptionfactors, such as c-Myc
- Cdks also stimmulate synthesis of genes required for the next phase
- e.g. Cdk4/6+ Cyclin D stimmulate expression of Cyclin E
Explain how Cdks can regulate gene expression
Example: inactivation of Rb protein by Cdk 4/6+ Cyclin D
- Normally: Rb protein inhibits the transcription Factor E2F
- Rb protein gets progressively inhibited by Cdk4/6+ Cyclin D
- –> activation (disinhibition) of E2F
- transcription of further molecules (e.g. cyclin E)
What is the Rb protein?
The Retinoblastom protein is a tumor supressor gene that normally inhibits E2F (transcription factor that drived cell cycle)
Explain the regulation of E2F and Rb protein throughout the cell cycle
E2F is more and more upregulated throughout the cell cycle due to progressive inhibition of Rb protein (via multiple phosphorylation)
What are possible oncogenic mutations in the RAS gene?
Why are they oncogenic?
Mutations can lead to permanent onswitch of RAS –> no inactivation possible
- e.g. mutation prevents GAP binding –> no inactivation
- e..g Mutation prevents GTP hydrolysis
How can Cdks be inactivated?
- Unbinding + degradatio of cyclins
- Cdk inhibitors (CKI)
What is the effects of an activated RAS protein?
it activates a protein kinase cascade
more specifically: Extracellular signal-regulated kinase (ERK) cascade
What is the name of the intracellular kinase cascades that induce mitosis?
- Generally
- ACtivated by RAS
- Gneral Name= Mitogen-activated protein kinase (MAPK) cascades
- RAS activates (or extracellular binding) Extracellular signal-regulated kinase (ERK) cascade
What can happen in mutations to CKIs?
May result in cancer
Which cdk/cyclin complex activates the transisiton from G0 to G1?
How is it activated?
- Cdk 4/6 + cyclin D initiate transtion from G0 to 1
Activated via
- cyclin D expression is regulated via c-Myc
What are Genes regulated by the transcription factor EGF2?
Proto-oncogenes
- c-Myc
- N-Myc
Cell cycle genes
- cyclins A+E
- Cdk4, Cdk2
- E2F genes
- Rb protein
Many more+ DNA synthesis genes
What are the two families of CKIs?
CKI = Cdk imhibitors
- INK4 family
- in G1 phase
- inhibit CDK4/6 by displacing cyclin D
- CIP/KIP family (e.g. p27KIP1)
- S phase
- inhibit all Cdks
What needs to happen to the CKIs to allow cell cycle regulation?
Must be degraded + gene supression to allow cell cyclie progression
How do EGFR/HER2 lead to cancer?
They are often over-expressed in cancer and lead to increased cell proliferation by binding to Growth factor recetor
Why can DNA easily be damaged?
because the DNA bases are planar carbon rings –> easily chemically activated and then
- be reactive, react with other molecules
- are chemically similar –> can be transfered into one another
What is Deamination in the process of modification of an AA?
What does it lead to?
The loss of one amine group of a DNA base (often converted to keto group)
- can lead to mutation via conversion of
- cytosin to uracil (essentially thymine)
- adenine to hypoxanthine
- guanine to xanthine
- and 5-methyl cytosine to thymine.
What are “other chemical modificatios” of DNA?
What do they lead to?
Many reacgtions, often induced by free radicals or hyper-reactive oxygen molecules (byproducts of normal motabolism or can be produced by ionising radioation)
Leading to
- formation of double-bonds
- methylation, alkylation of DNA bases
- adduct formation
What is photodamage?
How does it change the DNA?
intra-strandal change by UV light being absorbed by the DNA bases
- UV light activates and causes
- pyrimindien dimer (often thyamine dimers)
- driver in skin cancer
What are the different modifications/damages that can occur in DNA change?
- Deamination
- other chemical modification
- methylation
- alkylation
- adduct formation
- photodamage
What is the consequence a Base-mismatch in the DNA?
There will be a bulge in DNA –> will be used to find the site of damage and repair of the DNA
How does Radiodamage modifies the DNA
It can sometimes break the phospho-diester bonds–> leading to gap in DNA
- also used to find the site of damage and repair
Overall spoken: What causes DNA damage?
- Carcinogens
- dietary
- lifestyle
- environmental
- occupational
- medical
- endogenous
- Radiation
- ionizing
- solar
- cosmic
Explain how polycyclic aromatic hydrocarbons might lead to DNA damage?
Where are they found?
- Common environmental pollutants
- Formed from combustion of fossil fuels
- Formed from combustion of tobacco
E.g. Benzo(a) pyrene (B(a)P)
- itself it nor carcinogenic but gets activated by metabolism (highly carciongenic)
- Forms DNA adduct when activated
Name an example of a food-born carcinogen and how it could cause cancer
Aflatoxin B1– >formed by fungi on grains and peanuts
- metabolism converts it into carcinogen
- activation–> highly reactive
- adduct formation
Why is DNA damage problematic?
Damage can lead to mutation –> mutations can lead to cancer
- damaging DNA is an important strategy in cancer therapy (might lead to apoptosis)
Why is DNA repair so important?
What are its physiological capacities?
There is a lot of DNA damage going on all the time needs to be repaired to not cause cancer
–> normally: cell has a lot higher capcity to repair than damage happens
Explain adduct formation in DNA
Reactive proteins can bind to DNA (or DNA gets activated and forms Adduct itself) and alter its chemical structure by binding to it
How does UV radioation cause cancer?
UV radiation might induce
- Pyrimidine (thymine) dimers
- mutation
- melanoma
Explain how ionizin radioation can cause DNA damage
- Generates free radicals in cells
- Includes oxygen free radicals
- super oxide radical: O2•
- hydroxyl radical: HO•
- Includes oxygen free radicals
- Possess unpaired electrons
- electrophilic and therefore seek out electron-rich DNA
Explain the effects of free radicals on the cell
They attack the DNA and cause:
- Double and single strand breaks
- Apurinic & apyrimidinic sites
- Base modifications
- ring-opened guanine & adenine
- thymine & cytosine glycols
- 8-hydroxyadenine & 8-hydroxyguanine (mutagenic)
When would Base-excision repair take place?
How does it waork?
BER repairs damages base with intace phosphate diester bonds
- DNA glycosylase removed wrong DNA
- AP endonuclease cuts phospho-diester bond
- Polymerase fills the gap with corect DNA
- Ligase binds the DNA pieces
When is Nucleotide excision repair used?
Explain its principle
Used when there is damages Base with damages phosphate-diester bond (e..g in removal of large adducts)
- Xeroderma pigmentosum proteins (XP proteins) assemble at the damage
- Endonuclease cuts edges of diester bonds that need to be removed (?) –> larger area around damage
- Helicase cuts off large DNA piece
- DNA polymerase synthesises new DNA
- DNA ligase binds them
When does DNA mismatch repair occur?
scrutinisation of DNA for apposed bases that do not paired properly
- only during replication/ new synthesation of DNA
How is DNA mismatch recognised?
By wrong Watson-crick base paring–> DNA forms bulges that can be recpgnised
Which factors activate p53?
What are the effects of p53 activation?
Upregulation of cellular pathways leading to
inhibits progression of cells with damaged DNA from S-phase, later produced CKI-21
- In mild and moderate stress
- antioxidant defence
- DNA repair
- growth arrest
- senescense
- In severe cellular stress
- apoptosis
What are the consequences carcinogen-DNA damge?
- Normal repair–> normal gene and function is restored
- Apoptosis –> damage too severe
- Incorrect repair+ altered primary structure
- Fixed mutations leading to
- Aberrant protiens (abweichend)
- If that is in TSG/oncogenes: carcinogenesis
- Fixed mutations leading to
How do you determine, wether a substance is a carcinogen or not?
- just looking at the chemical strucutre: is it likely to be a carcinogen?
- AMES test (in vitro) –> bacteria
- Chromosomal damge (in vitro), mammalian cells
- Micro-nuclei around DNA formed (in vitro), mammalian cells
- Murine Bone Marrow Micronucleus assay (in vivo)
Explain the AMES test in testing for Carcinogens
- A bacterium that cannot produce histidin is cultured in a histidine free environment
- Chemical to be tested + metabolite enzymes are added –> converstion to active metabolite?
- Given to bacteria –> see if they can cultivate
- If yes: mutation has occured and bacteria make histidin –> carciongenic
Why do you look at micronuclei in mamalian cultivated cells when testing for a carcinogen?
- Binucleate cells assessed for presence of micronuclei
- Can stain the kinetochore proteins to determine if chemical treatment caused clastgenicity(chromosomal breakage) or aneuploidy (chromosomal loss)
–> It shows severe chormosomal damage
Explain the Murine Bone Marrow Micronucleus Assay in testing for carciongens
Treat animals with chemical and examine bone marrow cells or peripheral blood erythrocytes for micronuclei
What is an apurinic/ Apyrimidinic site?
It is a site in DNA that has no base (mosltly due to DNA damage)
Explain the process of DNA mismatch repair
- Recognition of the wrong base (using old base as a template)
- Cutting out the wrong DNA
- can be nucleotide-excision repair (NER) or base-excision repair (BER)
- Polymerase restores DNA and fills Base in
Can only occur during replication!
Explain the process of direct DNA repair
Direct Repair involves the reversal or simple removal of the damage by the use of proteins which carry out specific enzymatic reactions
E.g.
- Photolyases repair thymine dimers.
- O6 methylguanine-DNA methyltransferase (MGMT) directly reverses some simple alkylation adducts.
What are the two different types of excision repair of DNA ?
- Base excision repair (BER)
- Nucleotide excision repair (NER)
When might a mutation in NER lead to cancer?
•Xeroderma Pigmentosum
- •severe light sensitivity
- •severe pigmentation irregularities
- •early onset of skin cancer at high incidence
- •elevated frequency of other forms of cancer
- •frequent neurological defects
–> no repair made by NER possible
- also other conditions that are often linked to cognitive impairment/neurological developmental issues
What is the role of p53 to controll cellular repair pathways?
p53 is a TSG that is normallly supressed by MDM2
- in cellular stress: p53 gets activated (trasncription factor) and acitvates DNA repair pathways
Which kind of DNA damages are repaired by BER?
- Oxidized bases: 8-oxoguanine, 2,6-diamino-4-hydroxy-5-formamidopyrimidine (FapyG, FapyA)
- Alkylated bases: 3-methyladenine, 7-methylguanosine
- Deaminated bases:
- Uracil
- single-strand breaks
Which kind of DNA damages are repaired by NER?
Damages that are formed by UV light (dusruption of helical structure)
- adducts (e.g. thymine dimers)
- *
What are the resons why apoptosis takes place?
- Harmful cells (e.g. cells with viral infection, DNA damage).
- Developmentally defective cells (e.g. B lymphocytes expressing antibodies against self antigens).
- Excess / unnecessary cells (embryonic development: brain to eliminate excess neurons; liver regeneration; sculpting of digits and organs).
- Obsolete cells (e.g. mammary epithelium at the end of lactation)
–> Can be Exploited therapeutically - Chemotherapeutic killing of cells
What are the characteristics of necrosisi?
- unregulated cell death
- normally due to trauma
- there is cellular disruption
- associated with inflammation (due to release of intracellular contents)
What are the characteristics of apoptosis?
- programmed, regulated cell death
- controlled dissembly of contents without membrane disruption
- no inflammation
- ATP dependant
How many different types of cell death are ther?
Many many types of cell death, ranging from Necrosis on the one end to Apoptosis on the other hand –> many are in the middle and have characteristics of both
Explain the process of necrosis
- Plasma membrane becomes permeable
- Cell swelling and rupture of cellular membranes
- Release of proteases and intracellular contents leading to autodigestion and dissolution of the cell
- Localised inflammation
What are the two phases of Apoptosis?
What are theri characteristics?
- Latent phase
- apoptosis pahtways activated but no morphological change in cell
- Execution phase
- morphologican changes and destruction of cell
Explain the processes that take place during the execution phase of apoptosis?
Example of epithelial cells
- loss of microvilli and intracellular junctions
- cell shrink
- loss of plasma membrane assymetry (important in normal function, can be seen e.g. by phosphatidylserine lipid appearing on surface)
- Chromatin and nuclear condensation
- DNA fragmentation
- Formation of memrane bleps
- Fragmentation into membrane-enclosed bodies (get phagocytosed by macrophages etc)
- no release of cell contents
What happens in the latent phaese of apoptosis?
death pathways are activated, but cells appear morphologically the same
What happens to the DNA during apoptosis?
•Chromatin and nuclear condensation
• DNA fragmentation
Explain the characteristics of Apoptosis-like PCD (programmed cell death)
some, but not all, features of apoptosis. E.g. Display of phagocytic recognition molecules before plasma membrane lysis
Explain the characteristics of Necrosis-like programmed cell death (PCD)
Variable features of apoptosis and necrosis before cell lysis present; “Aborted apoptosis”
What two types of caspades are there?
What is their characteristic and function?
There are two types
- Initiator caspases
- get activated directly by intrinsic/extrinsic pathways
- Effector caspases
- get activated by initiator caspases, set off futher routes
What are the protien domains that all caspases have in common?
What are the additional domapins the different initiator capsades have?
What is their function?
All have in common: p20 and p10 domain
- Caspase 9+2 also have a CARD domain (caspase recruitement domain –> will localise the caspases at specific sites in the cell)
- Caspase 10+8 also have a DED domain (Ceath effector domain) –> binds to adaptor protein and involved in activation of the capsase
What is meant by the term of caspase maturation?
Explain the process
It is the activation of a procaspase to the active form
- Pro-domain and SS domain of caspase are cleaved (proteolysis)
- Dimer formation of one Large (L) and one small(2 chain)
- 2 dimers come together to form the active heterotetramer, consisting of 2L2S
What are the different concepts by which the effector caspases can execute the apoptotic program?
(what do they do to proteins?)
- Inactivate by proteolysis (cleavage of proteins)
- single proteins
- protein complexes
- Activate enzymes by cleavage
- direct activation
- indirect by destruction of inhibitory molecules
Explain the processe that happens after trimerisation of the death receptor at the example of FAS
- adaptor Protein (FADD) will bind to FAS via the Death domain (they both have)
What happens after FADD binds to FAS via the DD?
This will lead to an recruitement of caspase 8 that binds to FADD via the DED (both have it)
and will lead to oligomerisation that activated caspase
Explain how oligomerisation of caspases lead to their activation
- Oligodimerisation leads to conformational change of proteins
- some caspases need transcleavage to be activated
Concept of Transcleavage:
- FAS is a trimeric receptor
- 3 FADD molecules with 3 DED bind to it
- 1 caspase (8?) binds to each of them–> 3 caspases
- Crosscleave each other leading to acitvation and
- Release of activtive initiator caspase 8 (tetramer) –> at least 2 caspases required
Explainn how FLIP inactivates the apoptotic pathway
It inhibits the activation of caspases competitevely
- Binds to the DED ends of FADD
- but has no proteolytic funtion–> no caspase tetramer formation
Explain the caspase cascase (which caspase activated which
Which proteins/facotrs are required in the formation of teh apoptosome?
Apaf-1 (Apoptotic activating factor-1),
Cytochrome c,
ATP,
procaspase 9
Explain the formation and structure of the apoptosome
- Apaf-1 (Apoptotic activating factor-1) come together at the CARD domain and form a heptamer (7)
- Outside: there is the WD-40 domain that interactis with cytochrome c
- When cytochrome c bound–> caspase 9 (several, each Apaf-1 can bind one) bind to CARD of apoptosome and gets activated (oligomerisation and trans-cleavage)
- Caspase 3 recruitement
- Activation leading to caspase cascade acivation
How are the intrinsic and extrinsic pathways of apoptosis initiation connected?
- When Extrinsic pathway is activated:
- Bid protein gets activated by caspase 8 and triggers mitochrondrial release of mitochrondrial proteins
- –> activation of intrinsic pathway
What are proteins from the Bcl-2 family?
Proteins that are involved in regulation and modulation of apoptosis (can be pro-apoptotic and anti-apoptotic)
- differnet groups/proteins with different domains but all: BH3 as dimerisation motif
Explain the role of Phosphatidylinositol 3’-kinase (PI3’-K) in the cell survival
Phosphatidylinositol 3’-kinase (PI3’-K) is a lipid kinase that promotes cell survival
- gets activated by Growth factor receptor
- converts PIP2 into PIP3
- PIP3 Activates protein kinase PKB/Akt which is anti-apoptotic
Explain the relationship between the PKB/Akt pathway and the Bcl-2 proteins
- PKB/Akt are activated via the GF/PI3’-K pathway during growth factor binding
- they inactivate BAD
- which then means, that the heterodimers on the mitochondrium are inactive and and don’t cause activation of the intrinsic pathway
Explain how the absence of GF might lead to apoptosis
If there is no GF –> no activation of PKB/Akt (PI3’-K, GF pathways)
- no inhibition of Bad
- displaces Bax/BAc from Bcl-2/-xL binding site (all bind together via BH3)
- Bax/Bac are now released and form the Bax/Bac Pore -> Apoptosis via activation of the intrinsic pathway
Explain the role of PTEN in the regulation of apoptosis
PTEN can induce apoptosis by inhibiting the phosphorylation of PIP2 into PIP3 (and thereby the pro survival(phosphoinositide 3 kinase (PI3K) ) signaling)
- disinhibition of apoptosis
How do you call different forms of new vascular formation?
How do they differ?
- Vasculargenesis
- in emberyological development –> from bone marrow progenitor cells
- Ateriogenesis
- collateral growth
- Antiogenesis
- sprouting of blood vessels, important in adult life and tumor development
Explain the role of Hypoxia in Angiogenesis
Hypoxia is a powerful stimulus that triggers antiogenesis
- In presence of oxygen: HIF release is inhibited by pVHL: Von Hippel–Lindau tumor
- Hypoxia causes release of HIF (hypoxia-inducible transcription factor)
- –> HIF triggers VEGF release and other Growth factors
Explain the release of VEGF?
Often released due to Hypoxia:
In presence of oxygen: HIF release is inhibited by pVHL: Von Hippel–Lindau tumor
Hypoxia causes release of HIF (hypoxia-inducible transcription factor)
–> HIF triggers VEGF release and other Growth factors
What are the stalk cells?
How do they get activated?
Stalk cells are the cells next to the endothelia tip cells
- get activated via the Notch system
- Notch ligand on tip cell activates the Notch receptor on stalk cell
- this sets of the production of notch intracellular domain (NICD)
- NICD is brought to nucleus where it activates transcription factors RBP-J
Explain the cellular process of the tip cell selection
- In stable blood vessels, Dll4 and Notch signalling maintain quiescence
- VEGF increases expression of Dll4 (Tip Cell)
- –> Dll4 drives Notch signalling, which inhibits expression of VEGFR2 in the adjacent cell
- Tip-cell phenotype by Dll4-expressing tip cells acquire a motile, invasive and sprouting phenotype
- Adjacent cells forms Stalk cells
How do tip cells move and lead the path to angiogenesis?
They navigate in response to guidance signals
Adhere to Extracellular Matrix to migrate
Stalk cells behind the tip cell proliferate, elongate and form a lumen, and sprouts of tip cells fuse to establish a perfused neovessel.
Explain the role of platelets in angiogenesis
They are both: pro-angiogenic and anti-angiogenic and involved in physiolgical and pathological angiogenesis
What happens during stabelisation and quiescence of newly formed vessels?
- Lumen formation allows perfusion of neovessels (possible after fusion of neighboring branches)
- Stabelisation of new vessel via
- re-establishing junctions
- deposition of basement membrane
- maturation of pericytes
- production of vascular maintenance signals
Explain the role and regulation of tight junctions and adherence junctions in endothelial cells in angiogenesis
Very important to restore connections and Barrier formations during stabelisation of neovessel:
Mainly regulated via VE-cadherin
- Constitutively expressed at junctions
- mediates adhesion between endothelial cells and intracellular signalling
- Controls contact inhibition of cell growth
- Promotes survival of EC
Explain the role of Ang-1 in angiogenesis
How is it released?
Ang-1 is an agonist of the Tie 2 receptor, importnat in stabelising of neovessels
- when bindin to it
- promotes vessel stability
- reduces inflammatory gene expression
Released by the pericytes
Explain the role and release of Ang-2 in angiogenesis
It is released upon inflammatory stimmuli and antagonises the Tie2 receptor
- blocks the effects of Ang-1:
- increases vascular instability
- promotes VEGF dependant angiogenesis
Summarise the process of angiogenesis
What is meant by the term the angeiogenic switch?
What happes there?
It is the initiation of angiogenesis in a tumor cell
- mostly a hypoxia induced release of angiogenic factors leading to
- angiogenesis ( perivascular detachment and vessel dilation (b), followed by angiogenic sprouting (c), new vessel formation and maturation, and the recruitment of perivascular cells (d))
What is the role of Cance-associated Fibroblasts in angiogenesis of cancer cells?
secrete extracellular matrix; pro-angiogenic growth factors,
What are the potential mechanisms of resistance of tumor cells agaisnt anti-VEGF therapy?
- Other hypoxia induced growth factors might be produced by tumor
- Tumours vessels maybe less sensitive to VEGF inhibition due to vessel lining by tumour cells or endothelial cells derived from tumours
- Tumour cells that recruit pericytes maybe less responsive to VEGF therapy
What is the aim for anti-angiogenic therapies in cancer treatment?
Why?
If there is too much anti- angiogenesis
- formation of resistance
- damage of healthy vasculature
- no adequate delivery of chemotherapy/drugs to cancer cells
So aim for: anti-angiogenic therapy that normalises vasculature
- reduces hypoxia
- Increase efficacy of conventional therapies
What is vascular mimicry?
It is the ability of a cancer to form vessesl-like channels in cancer cells (not via signaling of body cells)
–> agressive and poor prognosis!
What are the potential mechanisms of resistance of tumor cells agaisnt anti-VEGF therapy?
- Other hypoxia induced growth factors might be produced by tumor
- Tumours vessels maybe less sensitive to VEGF inhibition due to vessel lining by tumour cells or endothelial cells derived from tumours
- Tumour cells that recruit pericytes maybe less responsive to VEGF therapy
What is the aim for anti-angiogenic therapies in cancer treatment?
Why?
If there is too much anti- angiogenesis
- formation of resistance
- damage of healthy vasculature
- no adequate delivery of chemotherapy/drugs to cancer cells
So aim for: anti-angiogenic therapy that normalises vasculature
- reduces hypoxia
- Increase efficacy of conventional therapies
What is vascular mimicry?
It is the ability of a cancer to form vessesl-like channels in cancer cells (not via signaling of body cells)
–> agressive and poor prognosis!
Explain the different steps in tumor Progession
When do benign Tumors turn to malignant tumors?
- Homeostasis (normal cells)
- Genetic alteration
- Hyper-proliferation
–> Benign
- De-differentiation
- dissasembly of cell-cell contacts
- loss of polarity
- Invasion
- increase motility
- cleavage of ECM proteins and basemsent membranes –> Metastisis
What are the two main types of tumor cell migration?
How do they differ?
-
Individual cell migration
- Ameoboid cells
- Mesenchymal (single) cells
- Mesenchymal(chain) cells
-
Collective cell migration
- Cluster cohorts
- Multicellular strands/sheaths
What factors are needed in Cluser/multicellular cell migration of tumors?
- Integrins and proteases to cleave basement membrane
- Gap junctions and cadherins for coordination of movements
What are the characteristics of tumor cell metastisis?
They mimic normal morpholigical movements
- as seen in angiogenesis
- branching morphogenseis like in mamillary gland
- etc.
What are focal adhesions?
They are connections between the cell and the extracellular matrix through integrins
- Extracellularly: integrins bind to extracellular ligands
- Intracellularly: multiproteins assemble (Plaque) and are connected to the actin cytoskeleton
What is the role of focal adhesions?
Important in signaling of movement
- anchor the cell
- Signaling: inform the cell about the composition of the ECM –> influence behaviour of cell
What are filopodia?
Finger like projections of cell membrane made of actin filaments, involved in cell motility
Explain the structure of Fillopodia
They are organised in
- straight, crosslinked actin filaments
- fomed by
- Polerymerisation
- Bundling of actin filaments
- Crosslinking of the filaments
What are Lamellipodia?
Shee-like pertrusions of cell membrane, rich in actin filaments needed for cell motility
*
What are the four steps that repetitively happen in cell movement?
- Extention
- Lamellipodia and Fillopodia grow into direction of movement
- Adhesion
- New adhesions (focal adhesions) with ECM+ Basement membrane found
- Translocation
- Cell body is moves in the direction of the movement, via contraction of actin filaments on the side opposite of movement
- De-adhesion of old adhesion
How does the overall cell shape change in a migrating cell compated to a normal cell?
It will become a polerised (in shape) cell
What is the first and rate limiting step in actin assembly?
What happens durin this step
Nucleation
- Attachment of actin to the inner cell membrane
- Arp proteins form a complex and bind actin monomers to create a nucleated actin filament
- Actin monomers bind to Arp-complex
Which proteins are important in the nucleation of actin filaments (other than G actin itself?)
Arp complexes
After initial formation of a actin trimer, what is the next step in actin filament formation?
Which protein is neede for it?
Elongation
- Controlled by
- +: profilin
- inhibited by thymosin
What happens after the Elongation of actin monomers to form an actin filament?
Capping
- further elongation inhibited
- done by proteins like: (also Arp protein involved)
After capping of Actin filaments, what is the next step?
Why is it importantn?
Severing
- enables quicker break down and reassembly of actin filaments (because chunks of actin are available, not jut monomeres)
What ar the possible fates of single severed actin filaments?
- De-polymerisation and monomer recylcling
- Annealing to other parts
- Elongation (at (+)Barbed-end with profilin actin filaments)
What is the next step after formation of a fully functional indivudual actin filament?
Cross-linking and bundling of strands
- functional dependant, many proteins can cross link e.g.
- Bundling
- Buckling if needed for contraction
Explain the role and the protein involved in actin branching
Branching forms a stable network of actin filaments
–> needed in formation for Lemelipodia
- 70° angle used
Mediated by the Arp complex
What happens during the Gel-Sol transition of actin filaments?
Why does it happen?
A process of severing
- a few actin filaments are cleaved to allow movement (sol-phase) from steady gel phase
Which cytoskeletal changes occur during Extention of the cell during movement?
Cell polymerisation is needed and all processes happen to deassambly/re-assembly actin filaments(e..g for lamellae protrusion)
- Disassembly
- Nucleation
- Branching
- Severing
- Capping
- Bundling
Which cytoskeletal changes occur durin lamellae portrusion?
Polymerization, disassembly, branching, capping
Which cytoskeletal processes occur during Filopodia bulding?
- Polimerization
- Bundling
- Crosslinkin
Which proteins regulate the cytoskeleton?
Name indivudual ones that controll
Overall: Rho proteins
- Folopodia: Cdc42
- Lamellipodia: Rac
- Stress fibres: Rho
How do Rho proteins get activated?
activated by
- receptor tyrosine kinase
- adhesion receptors
- signal transduction pathways.
–> Cause GTP binding and therefore acivation (quick indactivation via phosphorylation)
How do Rho proteins controll the cytoskeleton?
- They set of a set of intracellular messenger systems and activate actin bindin proteins
- Lead to actin polerization/organisation
- Arp2/3 in nucleation
- Cofilin, Profilin
- etc.
Explain the role of Rac in cell migration and movement
Needed in the
- Adhesion
- and Extention phase of cell migration
- controlles lamellipodia and therefore needed for
- focal adhesion assembly
- actin polimerizsation and branching
Explain the role of Rho (specific protein, not family) in cell migration and movement
Controlls stress fibres therefore needed in
- Adhesion
- Translocation
- De-adhstion of cell
Via
- tension and contraction of filamnents
Explain the role of ECM in cell phenotype
The composition of the ECM has a crucial role in determenin the cells organisation and phenotype
How do cells “sense” the composition of the ECM?
Via binding of integrins
- Special ECM receptors that can bind to many different ECM molecules
- Link to cytoplasm
- Contunitnity between cytoplasm and external environment
Explain the structure of Integrins
Are compsed of
- alpha and ß subunit
- have a head binding to ECM molecules
- and a tail/Leg which has the membrane and cytoplasmic domains
How do integrins sense different ECM compositions?
There are different combiations of the alpha and beta subunits
- Each recognises different short AA sequences of ECM molecules
- They can be specific to one ECM molecule or are found in more than one molecule
Explain the structure of focal adhesions
Integrins cluster togetheer to then form focal adhesions (sometimes can also form hemidesmosomes)
- involved in signal transduction
*
Explain ther role of Intrgrins during “outide in” -signal regulation
Integrins can undergo confirmational change and therefore change ability to
- bind ligands
- change the signaling pathway
How can integrins sense the mechanical properties of the surrounding?
Via the amoun of force that is generated at a focal adhesion depends on
- both the force generated by the cytoskeleton (F cell)
- and stiffness of the ECM
What is “inside-out” integrin signalling?
An intracellular signal (e.g. as response to a homone binding etc.) can
- change the integrin affinity for the ECM
- used e.g. in inflammation, blood clotting
Explain the mechanism of anchorage dependance
- growth factor receptors and integrin signalling complexes can each activate identical signalling pathways (e.g. MAPK)
- Alone: this activation is weak and/or transient
- •Both needed for strong proliferative signal!
Explain the concept of Contact-induced spreading of epithelial cells
When epitelial cells meet:
- form stable junctions/cell-cell adhesions
- leading to cell spreading to form stable mono-layers
Explain the intracellular mechanism how cell-cell adhesions influence proliferation
Cell-cell junctions are calcium dependant: low Ca2+= low juncitons= high proliferation
- Without cell-cell junction
- high MAPK activity
- low p27KIP1 (CKI)
- promote proliferation
- With cell junctions
- MAPK inhibited
- inhibit proliferation
What is the mediator molecule that might be responeible for the link of cell-cell adhesions and proliferation
Thought to be mediated by ß-cadenin (link between calcium dependant junction and further intracellular signaling)
Explaint the role and regulation of ß-catenin in cell proliferation
ß-catenin is bound to cell-cell junction molecules
- When there is no junction–> ß-catenenin is free in cytoplasms
- Binds to LEF-1 and promotes gene transcription and cell proliferation
- Gets inactivated by APC complex
Explain the formation and Mechanism of Adenomatous polyposis coli (APC)
It is a form of inhierited disease forming uncontrolled colorectal polyps
- Due to a disfunciton of the APC gene that cannot degrade ß-catenin
- No degradation of ß-catenin –> uncontrolled proliferation
What are the different steps in metastisis formation?
- Cells must break away from the primary tumour
- travel to a blood or lymph vessel
- enter the vessel
- lodge at a distant site
- leave the vessel,ultimately
- establish a secondary tumour
What do primary carcinomal cells alter/have to metastisise?
- Reduction of cell-cell adhesions (e.g. cadherin levels reduced)
- Primary cells must be motile
- degradation of ECM must take place; matrix metaloproteinase (MMP) levels increased in order to migrate through basal lamina and interstitial ECM
- the degree of carcinoma cell-cell adhesion is an indicator of how differentiated the primary tumour is, and indicates its invasiveness and the prognosis
What is Blebbing?
Happens when Cells are detached from ECM
- membrane swells into several spherical bubbels
Can end
- apoptotic
- Non-apoptotic = cell locomotion, cell division and stresses
What are the different steps in metastisis formation?
- Cells must break away from the primary tumour
- travel to a blood or lymph vessel
- enter the vessel
- lodge at a distant site
- leave the vessel,ultimately
- establish a secondary tumour
What do primary carcinomal cells alter/have to metastisise?
- Reduction of cell-cell adhesions (e.g. cadherin levels reduced)
- Primary cells must be motile
- degradation of ECM must take place; matrix metaloproteinase (MMP) levels increased in order to migrate through basal lamina and interstitial ECM
- the degree of carcinoma cell-cell adhesion is an indicator of how differentiated the primary tumour is, and indicates its invasiveness and the prognosis
What is Blebbing?
Happens when Cells are detached from ECM
- membrane swells into several spherical bubbels
Can end
- apoptotic
- Non-apoptotic = cell locomotion, cell division and stresses
What are the 6 original hallmarks of cancer?
SARCOMA
S – Self-sufficiency of growth signaling
A – Apoptosis evasion
R – Resistance to anti-growth factor signaling
CO – Continuous replication (Immortality)
M – Metastasis
A – Angiogenesis
What are the 4 additional Hallmarks of cancer?
What are proto-oncogenes?
Genes that code for essential proteins involved in maintenance of cell growth, division and differentiation
What kind of mutations normally occur in proto-oncognes that convert them into oncogenes?
A single mutation can be enough:
- point mutation
- deletion
- Gene-amplification
- Chromosomal translocations / Insertional mutagenes
- E.g. Philadelphia chromosome – > hyperactiveness of transcribed gene
- E.g. Burkitts lymphoma –> stonger enhancer in front of gene increase normal protein levels
What kind of mutation is the phildelphia chromosome?
How does it lead to the formation of cancer?
It is a chromosomal translocation resulting in
- via the BCR-ABL gene fusion product
- BCL causes permanent activation of the ABL-gene product
- ABL is important in proliferation
explain how tumour suppressor genes have been discovered through heritable malignancies
Via the Retinoblastoma gene, development of the 2 hip hypothesis
- Each cell has 2 copies of a TSG
- Need loss of both copies for Cancer
- If one is already damaged –> can be inherited and only one hit is required to develop malignangy, leading to
- Family history of related cancers.
- Unusually early age of onset.
- Bilateral tumours in paired organs.
- Synchronous or successive tumours.
- Tumours in different organ systems in same individual.
- Mutation inherited through the germline.
At the example of colorectal cancer, explain how gene mutations can lead to cancer
When does p53 get activated?
It gets activated at cellular damage
- cellular stress
- hypoxia
- nitric oxide
- oxidative stress
- Damage
- oncogene activation
- Double strand breaks
- Telomere erosion
How does p53 get activated?
Via inhibiton of MDM2
- Normally: MDM2 inhibits p53 activtiy
- When activated: sets off repair mechanisms
Explain the function of PTEN and its role in carciogenesis
It is a Tumor supressor gene
- PTEN can induce apoptosis by inhibiting the phosphorylation of PIP2 into PIP3 (and thereby the pro survival signaling)
Explain the trend of cancer incidence rates in western countries
Overall: it used to increase, now further followed by a plateau of cases in many countries (might even decrease in some wetern countries)!
What is the role of infections in cancer?
16% cases of cancer likely caused by infectious agents worldwide
- 25% in Africa
- <10% in Europe (1 in 33 in UK)
What are the 5 cancers with the highes incidence in women worldwide?
- Breast cancer
- Colorectal cancer
- Lung
- Cervix Uteri
- Thyroid
What are the cancers with the highes mortality in women worldwide?
- Breast
- Lung
- Colorectal
- Cervical
- Stomach
What are the 5 cancers with the highest incidence among men worlwide?
- Lung
- Prostate
- Colorectal
- Stomach
- Liver
What are the 5 cancers most men die from worldwide?
- Lung
- Liver
- Prostate
- Colorectal
- Stomach
What are the 5 leading cancers in countries with high HDI? (incidence)?
- Breast
- Colorectal
- Lung
- Prostate
- Bladder
What are the 5 leading cancers in countries with high HDI? (mortality)
- Lung
- Colorectal
- Pancreas
- Breast
- Stomach
How do you call the pre-cancerous state in “breast cancer”?
What are its characteristics?
It is called “carcinoma in situ”
- pre-invasive
- often: the characterisitcs of individual cells are normal but they have an abnormal growth pattern
Picture: carcinomal im situ of luminal epitheilal cells
What are the differnet forms of cancer that can develop from a mamillary carcinoma in situ?
What is the most common one?
Carcinoma in situ can go on and develop differnt types of cancer– >originate in terminal duct lobular unit
- Lubular carcinoma (10-15%)
-
Infiltrating Ductal Carcinoma, not otherwise specified
- 80%!
- Medullary carcinoma, rare, agressive
How do you asses if a Mamma Ca is oestrogen positive or negative?
How many carcinomas show each characteristic?
It is done by histological staining with antibodies against human Estrogen Receptor (ER)
- 80% of IDCs (infiltrating ductal carcinomas) are ER positive!
Explain the location, activation and effect of the Estrogen Receptor and ER activation
It is an steroid (intracellular) receptor
- Estrogen binds to it and releases hsp90
- Causing dimerisation of ER
- Travel into cell and alter DNA transcription
Which molecules get expressed via the activation of an Estrogen Receptor?
Normally upregulates pro- proliferative and anti-apoptotic factors
- upregulation of Progesterone Receptor (PR)
- Cyclin D1
- c-myc
- TGF-a
What is the main rationale behind endocrine therapy in breast cancer?
What are the different approaches used?
RAtionale: To reduce Estrogen and therefore growth and proliferation of the tumor
- In pre-menopausal women: ovarian supression
- Blocking estrogen production by enzymatic inhibition
- Inhibiting estrogen responses
Explain the process of hormonal supression in the treatment of breast cancer
LHRH Agonist used to supress the production of LH/FSH
- overstimmulation of LHRH-receptor leads to its downregulation –> less/no response to LHRH
- reduced Estrogen production
What are the main side-effects of the use of Tamoxifen?
- It increases the riks of thrombosis/embolisms
- It stimmulates growh of the endometrium –> endrometrial cancer
- increased frequency of cataracts
What is the main issue with endocrine treatments in breast cancer?
Resistance
- expecially in metastatic disease, all patients become resistant
- But still express ER and are driven by estrogen –> switch from tamoxifen to aromatase inhibitors
–> Use endocrine treatments as long as possible but after that: additional treatment required
What are the different types of adenomas that can occur in the colon?
What are their differences?
- Tubular (>75% tubular)
- Normally Pedunculated –> harder to invade into submucosa and mucosa
- Tubulovillous (25- 50% villous)
- mixed type)
- Villous ( > 50% villous)
- normally sessile –> easier to invade into submucosa and mucularis
Explain the microscopic structures of tubular adenomas
- Columnar cells with nuclear enlargement, elongation, multilayering and loss of polarity
- Increased proliferative activity
- Reduced differentiation
- Complexity/disorganisation of architecture
What are the microscopic changes and characteristics of villous adenomas?
- Mucinous cells with nuclear enlargement, elongation, multilayering and loss of polarity
- Exophytic, frond-like extensions –> finger like
- Rarely may have hypersecretory function and result in excess mucus discharge and hypokalemia
Explain the adenoma-carcinoma sequence in the formation of Colorectal cancer
AK 53: 1: APC, 2nd K-ras, 3rd. P53
- First hit of TSG (can be inherited or or aquired)
- E.g. APC, MSH2 (missmatch repair)
- nothing happens
- 2nd hit of TSG
- APC, MSH2, ß-catenin
- nothing happens
- Protooncogene mutation
- e.g. K-Ras
- adenoma formation
- Homozygous loss of additional cancer supressor
- P53, LOH
- Carcinoma
- Irregulated control and loss of many genes
Why is APC Especially Important in Colon Cancer?
Because it is the gene that turns off ongoing proliferation in the crypts
–> in colon proliferation is needed all the time!
What are the macroscopic features of Colorectal Cancer?
Small carcinomas may be present within larger polypoid adenomas, pedunculated or sessile
What is the distribution of colorectal carcinomals in the gut
What is the Dukes Classification?
A way of staging Colorectal carcinomas
A-C with increasing severity
- Dukes A - growth limited to mucosa/submucosa
- nodes negative
Dukes B - growth into or beyond muscpropria
- nodes negative
Dukes C1 - nodes positive
- apical LN negative
Dukes C2 - apical LN positive
What is the site disease in Leukaemia?
Leukaemia is “cancer of the blood”, more specifically
- bone marrow disease (not all patients have abnormal cells in the blood)
- it is cancer in lymphoid/myeloid stem cells
What are the different types of cells that can be affected in leukaemia?
-
Pluripotent haematopoietic Stem Cell
- would lead to mixed phenotype leukaemia
-
Myeloid Stem Cell
- Myeloid leukaemia
-
Lymphoid Stem Cell
- Lmphatic leukaemias
- Pre B Lymphocyte
- Pro T Lymphocyte
Explain and summarise the development of leukaemias
Leukaemias need
-
multiple mutations in a single myelod/lympoid stem cell
- leading to a potency of cell for increased proliferation, differentiation, or cell survival
- Leading to expansion of the leukaemic clone
What are symptoms of ALL?
Resulting from accumulation of abnormal cells (+inflitration of tissues) leading to:
- Bone pain
- Hepatomegaly
- Splenomegaly
- Lymphadenopathy
- Thymic enlargement
- Testicular enlargement
And Resulting from crowding out of normal cells (ANT symptoms)
- Anaemia casuing Fatigue, lethargy, pallor, breathlessness
- Neutropenia: Fever and other features of infection
- Thrombocytopenia: Bruising, petechiae, bleeding
What are haematological findings in ALL?
- Leucocytosis with lymphoblasts in the blood
- Anaemia (normocytic, normochromic)
- Neutropenia
- Thrombocytopenia
- Replacement of normal bone marrow cells by lymphoblasts
Explain the importance of immunophenotyping in ALL
It is important to determine the cause of the Leukaemia
- B cell? T cell?
- Done by flourescent analysis of cells (binding to CD factors of cells)
Recall the gross anatomy of the skin
- Epidermis
- Dermis
- separated by BM
- Hypodermis
Name a skin cancer that is lymphocyte derived
Mycosis fungoides
cutaneous T-cell lymphoma
What are the possible outcomes to photodamaged cells?
- Apoptosis (e.g. in sunburn)
- DNA repair
- Carcinogenesis
Explain the effect of UV light on the immune system
Influence immunity (UV ligh supresses immmune system)
- Via:Change of immunoregulatory genes in epidermis
- Depleting Langerhans cells in the epidermis
- reduce immunocompetence and surveilance
- used therapeutically e.g. in psoriasis (schuppenflechte)
–> Further increase in cancer risk
What does the host response to UV light depend on?
Depeinging on different genetic influences, expecially skin phototype
Where do melanocytes sit?
What do they do?
They ususally sit at the basement membrane in the epidermis
- produce melanin as reaction to UV light exposure
What is the prognosis of someone with malignant melanoma?
Depending on the stage of the disease and spreading
- measurement of granular layer to bottom of layer
- <1mm: thin
- 1-4 intermediate
- >4mm thick
but overall:
possibility of metastasis
What is a Lentigo maligna?
How would it usually look like?
A melanoma in situ
- proliferation of melanocytes within the epidermis
- non-invasive –> no risk of metastisis
Presentation
- Irregular shape
- Light & dark brown colours
- Size usually >2.0 cm
What are the characteristics of superficial spreading malignant melanoma?
- Lateral proliferation of malignant melanocytes
- Invade basement membrane
- Risk of metastasis
Can be diagnosed via ABCD(E)
- Asymmetry
- Border irregular
- Colour variation (dark, brown)
- Diameter >0.7mm and increasing
- Erythema
What are the characteristics of a nodular malignant melanoma?
•Vertical proliferation of malignant melanocytes
(no previous horizontal growth)
Risk of metastasis –> worst prognosis and most agressive form of melanoma
What is a basal cell carcinoma?
What are its chracteristics, causes and usuall sites of occurence?
- Malignant tumour arising from basal layer of epidermis
- Caused
- sun exposure
- Genetics
- Caused
- Characteristics
- Slow growing
- Invades tissue, but does not metastasise
- Common on face
- offen appear “parly”: greyish, red colour with shiny and dialated capillary vessels on top
