Signaling through Tyrosine Kinasses

Question 1

What is RTK?

What is EGFR?

Answer 1

Receptor Tyrosine Kinase
Epidermal growth factor receptor

[EGFR is a member of the class of RTKs.

Question 2

Explain molecular mechanism of stimulation of ras GTPase by RTKs

Answer 2

• Tyrosine Phosphorylation of receptor causes binding by SH2-domain-containing proteins including the adaptor protein Grb2, which binds a Ras GEF called Sos. Proximity of Sos with membrane-bound Ras results in guanine nucleotide exchange.

Question 3

Describe mechanism of action of two main classes of RTK-targeted anti-cancer agents (antibodies and TKI’s)

Answer 3

• primary role of antibodies is to block ligand binding to the receptor. TKIs inhibit catalytic activity (usually) by binding in substrate- binding site of the kinase.

Question 4

List tumor cell characteristics that predict clinical response to EGFR

Answer 4

-targeted therapeutics-response to EGFR TKI correlated with receptor mutations that may “activate” the receptor, EGFR amplification or overexpression as determined by FISH or immunohistochemistry.

Question 5

Describe mechanism of resistance to TKI’s.

Answer 5

Acquired resistance- second site mutations in EGFR arising or selected in patients who initially benefit from therapy but then acquire resistance and disease progression. These mutations block inhibitor binding to the kinase active site. May be able to design new inhibitors to avoid this problem. Activation of other receptors like Met or ErbB2. Combine inhibitors or make dual specificity inhibitors?

Primary resistance- if the tumor has a Ras mutation inhibiting the receptor further up the pathway will not be effective

Question 6

Describe mechanism of receptor tyrosine kinase activation.

Answer 6

Ligand binding drives dimerization, which activates catalytic activity of the kinase resulting in Tyrosine autophosphorylation at specific sites.

Question 7

Approximately how many protein kinases are there? Of these, how many are Tryosine kinases?

Answer 7

~500 Human Protein kinases

~60 Receptor Tyrosine Kinases (RTKs) in ~15 families: e.g. PDGFR, VEGFR, Tie, IGFR, EGFR etc.

Question 8

RTKs can ____ or _____ to activate signaling

Answer 8

homodimerize or heterodimerize

Question 9

Guanine Nucleotide Exchange Factors (GEFs) ____ monomeric GTPases by stimulating the release of guanosine diphosphate (GDP) to allow binding of guanosine triphosphate (GTP).

Answer 9

activate

Question 10

GTPase-activating proteins (GAPs) ____ signaling by inducing GTP hydrolysis.

Answer 10

terminate

Question 11

In general, GEFs turn on signaling by _____, whereas GAPs terminate signaling by ______.

Answer 11

catalyzing the exchange from G-protein-bound GDP to GTP; inducing GTP hydrolysis

Question 12

Sos is a _____, contains a Proline-rich region that binds to the Grb2 SH3 domain.

Answer 12

Ras GEF

Question 13

EGFR Inhibitor blocks _____

Answer 13

ATP binding

Question 14

Anti-EGFR (cetuximab) blocks the ____

Answer 14

ligand binding site

Question 15

Anti-EGFR antibodies block ligand binding and _____

Answer 15

prevent receptor dimerization

Question 16

Gefitinib blocks _____- and thus stops downstream signaling

Answer 16

EGFR kinase activity

Question 17

Secondary EGFR mutations are selected in tumors that progress after treatment with TKI. New mutation (T790M) found in tumors that arise after an initial beneficial response to______

Answer 17

gefitinib

Question 18

1. Describe mechanism of receptor tyrosine kinase (RTK) activation.
2. Explain molecular mechanism of stimulation of ras GTPase by RTKs.
3. Describe mechanism of action of two main classes of RTK-targeted anti-cancer agents (antibodies and TKI’s).
4. List tumor cell characteristics that predict clinical response to EGFR-targeted therapeutics.
5. Describe mechanism of resistance to TKI’s such as EGFR inhibitors.

Answer 18

x

Question 19

Why is EGFR of clinical interest?

Answer 19

EGFR is a therapeutic target in cancer

1) it is overexpressed in tumors: breast, lung, glioblastoma, head & neck, bladder, colorectal, ovarian, prostate.
2) Mutated leading to constitutive activation in glioblastoma.
3) Increased EGFR correlates with poorer clinical outcome in breast, lung, head & neck.
4) Increased receptor associated with increased production of ligands- autocrine stimulatory pathway.