Wk 15: Tyrosine kinase inhibitors

Question 1

What was the first generation drug?

Answer 1

• Imatinib
• Inhibit: Bcr-abl, C-kit, PDGF-R, ABL2 + DDR1 tyrosine kinase

Question 2

What are the different resistance that can happen against protein kinase inhibitors?

Answer 2

Target dependent resistance:
- Target point mutation

• Target amplification/overexpression

Target independent resistance:
- Dec drug uptake

• Inc drug efflux
• Upreg of alternate pathway

Question 3

What are the 4 mutations that account for 60% of resistant mutants?

Answer 3

• Thr-315-Ile (gatekeeper mutation)
• Tyr-253-Phe (changes in VDW)
• Glu-255-Lys/Val (inside ATP binding pocket)
• Met-351-Thr (aE-helix of C-lobe)

Question 4

What interactions can occur that may inhibit response?

Answer 4

• Caution w/ food/med impacting CYP3A4, if can’t avoid dec dose
• Drugs that raise gastric pH (PPI, antacids): agent = pH dependent solubility, oral + bioavailability compromised due to inc gastric pH

Question 5

What are the targets of protein kinase?

Answer 5

• Linked to phenotype
• Upstream MAP-kinase pathway
• Single kinase (mono-specific)
• Multiple kinase (polyspecific)

Question 6

What are protein kinase?

Answer 6

• Phosphorylated residues on protein (serine/threonine)
• Exists in multicellular organisms

Question 7

What form are protein kinase usually in?

Answer 7

Inactive:
- Blocked by intrasteric control/pseudo substrates

• Turned on by interaction w/ messenger or phosphorylation

Question 8

What is the DFG-in conformation?

Answer 8

• Conserved DFG motif (Mg positioning loop)
• Asp200 coordinates Mg at active site
• Phe201 packs into hydrophobic pocket creating hydrophobic regulatory spine

Question 9

What is DFG-out conformation?

Answer 9

• Inactivation occurs when Phe201 moves out hydrophobic pocket
• Disrupts orientation of Asp200 + sterically blocks ATP binding site
• Asp200 no longer coordinates Mg

Question 10

What are the different types of protein kinase inhibitors?

Answer 10

• Type 1: Competitive inhibitor, active, DFG in
• Type 1 1/2: Competitive inhibitor, inactive, DFG in
• Type 2: Competitive inhibitor, inactive, DFG out
• Type 3: Allosteric inhibitor near ATP
• Type 4: Allosteric inhibitor away from ATP

Question 11

What are the structure-activity relationship of type 1 + 2 inhibitors?

Answer 11

• Heteroaromatic ring competes at adenine-binding domain of hinge region
• Lipophilic + H bonding occupy allosteric + hydrophobic pockets adjacent to adenine binding domain
• Polar groups extend into phosphate binding region

Question 12

What are the structure-activity relationship of type 3 inhibitors?

Answer 12

• Bind in allosteric pocket adjacent adenine-binding site
• Don’t compete w/ ATP
• Hydrophobic, pie stacking + H bond groups = affinity + potency
• Electrostatic interaction btw inhibitor + ATP terminal phosphate of bound ATP + charged residue in catalytic site

Question 13

What are the structure-activity relationship of type 4 inhibitors?

Answer 13

• Electrophilic
• Contain michael acceptor forming irreversible covalent bond w/ Cys797 flanking ATP binding site

Question 14

What is the MOA of tyrosine kinase?

Answer 14

• ATP binds to kinase active site
• Prod ADP + phosphate unit
• Activate residue

Question 15

Why is introduction of a phosphate able to cause change in conformation?

Answer 15

• OH of residue is less able to form H bonds
• Phosphate ionised at pH 7.4 + introduces ionic interactions

Question 16

What are the components of the tyrosine kinase receptor?

Answer 16

• Extracellular domain
• Transmembrane domain
• Cytoplasmic domain (contains tyrosine kinase function triggering intracellular response)

Question 17

What activates the tyrosine kinase receptor?

Answer 17

• Dimers
• Once it binds, extracellular component moves across membrane surface + docks w/ 2nd monomeric receptor

Question 18

In the absence of a ligand, how does the tyrosine kinase receptor exists as?

Answer 18

Monomeric receptor

Question 19

What happens in receptor-ligand binding?

Answer 19

• Receptor dimerisation
• Leads to reorganisation of intracellular domain, results in activation of tyrosine kinase

Question 20

What is the key component of dimerisation?

Answer 20

Dimerisation arm - β-hairpin in EGFR extracellular domain - makes extensive contacts w/ binding region in other EGFR of dimer

Question 21

What do mAbs target?

Answer 21

GF + GFr outside cell membrane

Question 22

What do nibs target?

Answer 22

Target intracellular process:

• RTK phosphorylation
• Secondary messenger pathway

Question 23

Which 4 approved monoclonal antibodies target members of the Erb-B family?

Answer 23

• EGFR: cetuximab
• EGFR: panitumumab
• HER2: pertuzumab
• HER2: trastuzumab

Question 24

What is the mechanism of action of trastuzumab?

Answer 24

• Activation of antibody-dependent cellular cytotoxicity: bind to HER2 flag cells for destruction + attracts lymphocytes inducing apoptosis
• Prevention of HER2 formation: juxtamembrane region prone to proteolytic cleavage resulting in p95HER2
• Bond to extracellular domain preventing down stream activation that induces proliferation + angiogenesis - activates p27

Question 25

What is the structure of trastuzumab emtansine?

Answer 25

• Trastuzumab w/ DM1 (cytotoxic maytansinoid)
• Each trastuzumab linked to 0-8 DM1 molecule via lysine

Question 26

What is the mechanism of action of cetuximab?

Answer 26

• Chimeric mAb
• Binds to extracellular EGFR + turns off RAS
• Competitively blocks receptor associated tyrosine kinase activity

Question 27

What is the mechanism of action of irreversible protein kinase inhibitors?

Answer 27

• Form covalent bond to cysteine residue in kinase active site
• Use a,b-unsaturated Michael acceptor functionality

Question 28

What are the examples of irreversible protein kinase inhibitors?

Answer 28

Afatinib + ibrutinib