Cell signalling and cancer

Question 1

What kind of signalling molecules can bind to intracellular receptors?

Answer 1

Small, hydrophobic molecules that can pass through the membrane e.g. steroids / hormones

Question 2

What kind of signalling molecules can bind to extracellular receptors?

Answer 2

Hydrophilic proteins e.g. insulin

Question 3

What happens once a molecule has bound to an extracellular receptor?

Answer 3

It induces an intracellular signal transduction to alter target proteins for metabolism, gene expression and structure.

Question 4

What is a receptor Tyrosine Kinase?

Answer 4

Enzyme-coupled receptor on the cell surface, that spans the membrane once.
The extracellular domain may vary
The intracellular domain is a cytoplasmic TK.

Question 5

What does TK do?

Answer 5

TK phosphorylates tyrosine (Tyr) residues by catalysing the transfer of a terminal phosphate group from ATP onto self ser, Thr or Tyr.

Question 6

Give 3 examples of signalling proteins that will activate RTK?

Answer 6

EGF, Insulin, PDGF

Question 7

What pathway is used by RTK?

Answer 7

MAP kinases pathway (mitogen activated)

PI3-kinase pathway (Phosphatidylinositol)

Question 8

What does TK activity result in?

Answer 8

Either activates or inhibits the target protein.

Question 9

Is phosphate addition reversible?

Answer 9

Yes with phosphatases

Question 10

What happens when a signalling molecule binds to RTK?

Answer 10

Induces receptor DIMERISATION. The TK domains then cross-phosphorylate each other at multiple points for AUTOPHOSPHORYLATION.

Question 11

What does the phosphorylation of Tyr residues allow?

Answer 11

Creates a high affinity binding site for cytoplasmic proteins such as:
Grb-2 for MAP kinase pathway
PI3-kinase

Question 12

What induces the MAP kinase pathway? What does the pathway result in?

Answer 12

GF

GROWTH and PROLIFERATION

Question 13

Give the overall order of the proteins involved in MAP kinase pathway.

Answer 13

Signalling molecule —> RTK —> Grb-2 —> Ras —> Raf —> Mek —> Erk = Cell GROWTH and PROLIFERATION

Question 14

What is Grb-2?

Answer 14

A GF receptor bound protein. It has a SH2 domain that can recognise phosphorylated Tyr residues on RTK. The SH3 domain binds to Sos through its proline rich motif.

Question 15

What results from the formation of receptor - Grb2 - Sos complex?

Answer 15

Recruits and activates Ras.

Question 16

What is Ras?

Answer 16

A G protein with intrinsic GTPase activity. It is an oncogene attached to the plasma membrane.

Question 17

What is Sos?

Answer 17

GEF - guanine nucleotide exchange factor that activates Ras by stimulating it to exchange GDP for GTP.

Question 18

What inactivates Ras?

Answer 18

GAPs - GTPase activating proteins stimulate Ras’ intrinsic activity to hydrolyse GTP to GDP.

Question 19

What happens after Ras has been activated?

Answer 19

It activates Raf by changing its formation. Raf is a kinase that goes on to activate Mek and Erk.

Question 20

What are Raf, Mek and Erk?

Answer 20

They are serine 3 kinases also referred to as MAP kinases.

Question 21

What is the function of Erk?

Answer 21

Erk phosphorylates target proteins such as kinases, gene regulatory proteins and TF in the nucleus to induce changes in cell behaviour.

Question 22

Which gene regulatory proteins are activated by Erk?

Answer 22

c-Myc, cyclin D, c-Myb, c-fos, c-jun

Question 23

How is MAP kinase pathway switched off and why is it important to do so?

Answer 23

Removal of extracellular signal.
Protein tyrosine PHOSPHATASES switch off RTK.
GAPs inactivate Ras.
Dephosphorylation of target proteins by phosphatases.

Allows tight control to prevent constitutive activation.

Question 24

Which genes involved in this pathway are oncogenes that predispose to cancer?

Answer 24

RTKs, Ras, jun, fos, Myc

Question 25

What is the result in a cell from the activation of PI3 kinase pathway?

Answer 25

Cell SURVIVAL, GROWTH and PROLIFERATION.

Question 26

Give an overview in order of the proteins involved in the PI3 kinase pathway.

Answer 26

Signalling molecule ---> RTK ---> PI3 kinase ---> PIP3 ---> PDK1 ---> Akt ---> Survival or mTor ---> Growth and Proliferation

Question 27

How is PI3 kinase activated and what is its function?

Answer 27

By binding to phosphorylated Tyr residues on RTKs. It catalyses the phosphorylation of PIP2 into PIP3.

Question 28

What is PIP3?

Answer 28

PI(3,4,5)P3 is a phospholipid in cell membranes. It produces a high affinity docking site for PDK1 and Akt.

Question 29

Why must both PDK1 and Akt bind to PIP3?

Answer 29

PDK1 phosphorylates Akt.

Question 30

What is the function of Akt when active?

Answer 30

It dissociates from the membrane to phosphorylate BAD = release of anti-apoptotic molecules for survival. Targets mTOR.

Question 31

What are the two types of mTOR and their functions?

Answer 31

Complex 1 = stimulates growth by promoting ribosome production and protein synthesis. Inhibits protein degradation. Complex 2 = Stimulates cell survival and growth by helping PDK1 and activate Akt.

Question 32

How is PI3 kinase pathway regulated?

Answer 32

Tyrosine PHOSPHATASES will switch off RTK and dephosphorylate target proteins. pTEN is a tumour suppresspor inositol phosphatase that removes phosphate from PIP3 so it cannot act as a docking site.

Question 33

What is HER2?

Answer 33

An oncogenic protein of the EGF family. It is an orphan receptor with no known ligand and so responds to ligand of EGFR.

Question 34

What happens to EGFR1 when it recieves a GF signal?

Answer 34

Homodimerises with another EGFR1.

Question 35

What happens to HER2 when it recieves a GF signal?

Answer 35

It heterodimerises with EGFR1 for growth and survival through PI3 kinase and MAP kinase pathway.

Question 36

What happens if HER2 is overexpressed?

Answer 36

Overexpression due to gene amplification or mutation, allows HER2 to HOMOdimerise when in excess and signal in the ABSENCE of a GF. This causes constitutive signalling and abnormal growth.

Question 37

In what cancer is HER2 overexpressed?

Answer 37

Breast Cancer

Question 38

What therapy is used to target HER2 constitutive signalling in cancer?

Answer 38

Anti-HER2 antibody e.g. Transtuzumab, Herceptin. | Causes HER2 internalisation and degradation and ADCC (antibody dependent cellular cytotoxicity).

Question 39

In what kind of cancers are anti-HER2 antibodies effective?

Answer 39

In cancers that are HER2 +ve.

Question 40

How is the HER2 antibody produced to minimise immune response on administration?

Answer 40

Monoclonal Ig produced in mice but humanised by switching the sequences.

Question 41

How does the Philadelphia chromosome form?

Answer 41

Through translocation of chromosome 22 and 9. It rejoins at the sites of BCR on 22 and Abl on 9.

Question 42

What does the Philadelphia chromosome result in?

Answer 42

Creates a hybrid gene that encodes for a BCR-ABL fusion/chimeric protein.

Question 43

What is the normal function of Abl gene?

Answer 43

Cytoplasmic TK for survival and growth

Question 44

What happens to Abl when in the chimeric protein?

Answer 44

The N-terminus of Abl is substituted for BCR sequences to constitutively activate Abl TK activity. This stimulates inappropriate proliferation haematopoietic precursor cells and prevents apoptosis.

Question 45

What type of cancer is the BCR-Abl mutation associated with?

Answer 45

Chronic Myeloid Leukaemia - Uncontrolled proliferation of haematopoietic stem cells leading to excessive WBCs accumulating in the blood stream.

Question 46

What treatment is used in CML?

Answer 46

Abl Kinase inhibitor e.g. Imatinib. Blocks ATP binding pocket on the TK domain of BCR-Abl to prevent phoshorylation of proteins.

Question 47

Why do only 80% of patients respond to Imatinib?

Answer 47

Pts in acute phase CML with show an initial response but the relapse as a result of resistance and second mutations in the TK domain to prevent the drug from binding in the pocket.