EGF signalling

Question 1

Define intracrine signalling

Answer 1

This is when a cell signals to itself. The signal never leaves the cell.

Question 2

Define autocrine signalling.

Answer 2

This is when the cell signals to itself, but the signal leaves the cell to cause effect.

Question 3

Define juxtacrine signalling.

Answer 3

This is when a cell signals to immediate cells.

Question 4

Define paracrine signalling.

Answer 4

This is when a cell signals to the same tissue but a different cell.

Question 5

Define endocrine signalling.

Answer 5

This is when a signal travels to complete different parts of the body where receptors can be found.

Question 6

How is EGF ligand produced?

Answer 6

EGF is a prototype ligand, aka it is synthesised as a large molecule, then undergoes proteolytic cleavage to produce the mature protein.

Question 7

What are the receptors that bind EGF family ligands?

Answer 7

There is EGFR, HER-2, HER-3, HER-4. These are all cell surface receptors with intrinsic ligand-dependent kinase activity.

Question 8

How does ligand binding result in receptor activation and signal transduction?

Answer 8

1. EGF binds in the ligand binding domain 1 and 2, and cross links them.
2. The dimerisation domain in the cysteine-rich domain can then associate with the DD of another receptor.
3. Receptor dimerisation then occurs, forming homo- or hetero-dimer.
4. Kinase becomes activation, forming an asymmetric conformation.
5. This allows the availability of the activation loop (due to leucine position changing), allowing ATP to further phosphorylate tyrosine kinase domain.
6. Adaptor proteins are recruited, e.g. GRB2 and SOS.
7. SOS interacts with Ras-GDP, activating it.
8. Ras-GDP become active, forms Ras-GTP.
9. Ras-GTP activate Raf.
10. Raf activates MAPK signalling which promotes cell proliferation and survival.

Question 9

How is the expression of EGFR linked to cancer?

Answer 9

Increased expression of this receptor could lead to worse prognosis, decreased overall survival and decreased disease-free survival, as well as increased metastases in NSCLC, breast cancer, and CRC and head and neck cancer (Perez et al, 2001).

Question 10

What does the -inib suffix imply?

Answer 10

It is a small molecule inhibitor

Question 11

Name examples of tyrosine kinase inhibitors used to target EGF signalling. Are they effective?

Answer 11

Gefitinib - this is an inhibitor of EGFR TK. It prevents the activation of signal transduction by inhibiting ATP. Originally, it was ineffective for NSCLC with an increased expression of EGFR. However, they discovered that it was effective in patients with a mutated EGFR TK domain. Therefore they allowed patient stratification, and now the drug is given as a first line therapy in women, non-smokers, Asian origin patients in over 60 countries.
Lapatinib works the same but is a dual inhibitor - it inhibits EGFR and HER2 TKs (HER-2-HER-3 heterodimers produce the most potent response, therefore this aims to target this signal also)

Question 12

Name examples of monoclonal antibodies used for targeting EGF signalling. Are they effective?

Answer 12

Trastuzumab is a humanised monoclonal antibodies against the HER-2 receptor. It binds to the cysteine rich domain-2 to prevent dimerisation. It also promotes antibody dependent cell mediated cytotoxicity of the receptor. However there are cardiotoxic effects associated with its use. This drug actually caused 39% reduction in death rates when given for a 1-year duration. Only patients with a HER2+ status are eligible to receive this drug - patient stratification. Response rate when given to grade 3+ patients was 35% in comparison to 0% when given to grade 1 (Baselga et al., 2001).

Question 13

How does resistance to Trastuzumab occur?

Answer 13

The receptor may mutate, or there may be mutations in Ras/Raf, causing an activated signal downstream. It is imperative to try and prevent this resistance, therefore often combination therapies are used, e.g. Trastuzumab + Pertuzumab + Docetaxel has showed increased overall survival and increased progression free survival.

Question 14

What is Kadcyla? Why were there issues initially around its licensing?

Answer 14

Kadcyla is a drug otherwise known as Trastuzumab-emantasine. It is an antibody with a cytotoxic drug conjugate added. This ensures the antibody delivers the cytotoxic drug specifically to the tumour. Results were very promising however, its full list price was estimated at >£90,000, therefore the financial cost was too high for healthcare system. Later, it was discovered that its QALY was £166,000 which is well above the normal threshold, therefore Roche agreed a new commercial access arrangement for NHS.

Question 15

What is the structure of the EGFR?

Answer 15

Contains:

1. Ligand binding domain 1
2. Cysteine rich domain 1
3. Ligand binding domain 2
4. Cysteine rich domain 2

Transmembrane domain

5. Juxtamembrane domain
6. Tyrosine kinase domain
7. Signal transduction domain

Question 16

How can resistance occur to EGF therapeutics?

Answer 16

Can acquire mutations in the EGFR domain, e.g. EGFR ectodomain mutation S492R confers resistance to Cetuximab used to treat colorectal cancer.

Question 17

What solutions are available for Cetuximab resistance?

Answer 17

Can instead give panitumumab, nature article showed that this was successful (Montagut et al., 2012)

Question 18

What targeted treatments have been implemented utilising ‘magic bullet approach’ for combatting resistance?

Answer 18

Kadcyla!! Demonstrated decreased side effects and increased overall survival from 15.8 months with physicians choice of standard treatment, to 22.7 months (p = 0.0007) in HER-2 BC.