Receptor Tyrosine Kinases

Question 1

Kinases are involved in?

Answer 1

Cell growth, motility, metabolism, survival differentiation

Question 2

Where do RTK’s exist?

Answer 2

at the PM

Question 3

Structure of RTK?

Answer 3

Extracell, TM, and intracell domain which has the tyrosine kinase activity

Question 4

Ligand binding does what?

Answer 4

Causes dimerization, which activates catalytic activity of the kinase. Get autophosphorylation at specific sites (tyrosine residues)
Can have homo or heterodimerzation.
SH2 domains recognize this and can phosphorylate bound protein, get signal cascade

Question 5

Ras

Answer 5

GTPase. active with GTP.
Activation regulated by GTP exchange factors and GTPase activating proteins.
Can be switched “on” or “off” (GAP/GEF)
Act as switches that bind downstream effectors and localize them to the membrane to activate them.
Could hydrolyze its own GTP but too slow (GAP catalyzes)

Question 6

SOS

Answer 6

A GEF that stimulates Ras activation. Binds Sh3 and forces release of GDP.
Recruiting SOS to the receptor brings it close to Ras, which is at the cell membrane.

Question 7

Grb2

Answer 7

Grb2 is an adaptor protein. Has SH2 (binds phospho-try containing peptides)
Has SH3 domain that binds pro-containing peptides. SOS binds SH3.

Question 8

Ras activation

Answer 8

All you really need is translocation of SOS to PM (don’t even need RTK signaling).

Question 9

GEF

Answer 9

insert and inhibit interaction with gamma phosphate, GTP enters.

Question 10

Steps of Ras

Answer 10

1. ) Tyrosine phosphorylation on receptor
2. ) SH2 domain of GRB2 binds (binds phosphorylated kinases)
3. ) Binds Ras GEF (SOS) via SH3 domain
4. ) Proximity of SOS with Ras results in G exchange (need to be at membrane)
5. ) Activates Ras
6. ) Ras phosphorylates several protein kinases

Question 11

EGFR overexpression

Answer 11

more receptors than normal, they spontaneously dimerize indep. of ligands (normally ligand dep).

Question 12

Mabs and EGFR

Answer 12

they bind EGFR and prevent activation.
Cetuximab blocks ligand binding site and prevents receptor dimerization, ligand dependent activation of the receptor.
Extracellular action.

Question 13

EGFR small TKI’s

Answer 13

Erlotinib and gefitinib. Against 1 to a few RTK’s.
TKI’s inhibit the catalytic activity by binding in substrate binding site of the kinase, small mlcs that can get inserted into cells.
PREVENT ATP BINDING>

Question 14

Positive EGFR to TKI response?

Answer 14

Correlated with receptor mutations that may activate the receptor.
EGFR amplification of overexpression (seen on FISH or IHC).
pts more sensitive to lower conc of drug

Question 15

TKI resistance?

Answer 15

Pts get add’l mutations in cytoplasmic domain, 2nd mutation leads to resistance.
Threonine to methionine (gatekeeper mutation).
Increased binding ATP or decreased binding of inhibitor.
Can try other drugs, activate other RTKs

Question 16

Ras and EGFR

Answer 16

if pt has RAS mutation, inhibiting receptor further up pathway… drugs will not work.