Platelet Aggregation Inhibitors MAHMOUD KHATTAB, Ph.D.

The components of a platelet

Platelets and the Mechanisms of Arterial Thrombosis Platelet adhesion and aggregation occur at the site of plaque rupture at luminal exposed subendothelium ADHESION occurs in response to collagen or vWf followed by a cascade of further platelets recruitment Activated platelets exert pro-coagulant effects and the soluble coagulation cascade is activated Fibrin strands and erythrocytes predominate within the lumen of the vessel and downstream in the body and tail of the thrombus

Platelet Aggregation Activated platelets undergo three consecutive processes (a) shape change, (b) secretion of platelet granular contents (ADP, fibrinogen & 5-HT) and finally (c) platelet aggregation The final common pathway in platelet aggregation is cross-linking of the activated GP IIb/IIIa receptor (integrin αIIbβ3) with circulating adhesive arginine-glycine-asparagine (R-G-D) sequence macromolecules, predominantly fibrinogen and von Willebrand factor There is ~50,000-80,000 GP IIb/IIIa receptors on the surface of each platelet

Platelet Aggregation GP IIb/IIIa receptors undergo inside-out (low-high affinity) signalling in order to bind to vWf/fibrinogen Main stimuli for full platelet aggregation include: Collagen, ADP, thromboxane A2 (TXA2), & thrombin All except for collagen act through G-protein coupled receptors activating two transductions Phospholipase C Phospholipase A2 Stored ADP & de novo synthesized TXA2 act as positive feedback mediators

The stimulatory pathway for platelet secretion TXA2 is produced from AA via 3-steps enzymatic process; PLA2-COX-1/TXA2 synthase ADP activates Gi-coupled P2Y12 receptors TXA2 & thrombin activate Gq-PLC-coupled TXA2 & thrombin receptors respectively Ultimately, GP IIb/IIIa receptors undergo inside-out signalling bringing platelet aggregation

Platelet Activation Activation of membrane-bound G proteins catalyzes the intracellular signaling system, predominantly inositol biphosphate (IP3) and diacyl glycerol (DAG) IP3 causes the release of intracellular calcium from the SR & dense granules, phosphorylation of the myosin light chain, and allows the platelet contractile apparatus to perform platelet shape changes DAG activates PKC and in turn the activation of the surface integrin glycoprotein (GP) IIb/IIIa receptor occurs Activated receptor undergoes a conformational change and binds to the arginine-glycine-asparagine (R-G-D) sequence on circulating macromolecules mainly fibrinogen & vWf, and is responsible for cross-linking platelets together TXA2 provides amplification of the aggregation response & a significant vasoconstrictor effect

Increased platelet activity implicated in post-MI reocclusion and ischemia Several lines of evidence implicate increased platelet activity in reocclusion and recurrent ischemia following acute MI A, Autopsy data indicate that patients who died after acute MI are several times more likely to have a platelet-rich thrombus occluding the coronary infarct-related coronary artery if they had received thrombolytic therapy than if they had been treated without it B, The production of thromboxane B2 (TXB2), the stable metabolite of TXA2, is markedly increased, indicating that platelet activation is stimulated in the early minutes after thrombolytic therapy is given (The administration of oral aspirin is able to block this reaction) C, Platelet aggregability following thrombolysis with either streptokinase or tissue-type plasminogen activator is increased, as measured by responses to either thrombin or ADP added to platelet-rich plasma

ASPIRIN IN ACUTE ISCHEMIC SYNDROMES Low dose of aspirin inhibit TXA2 production by inhibting irreversibly COX-1 acetylation (at serine-530) After oral intake, platelets are exposed to relatively high concentration of ASA in the portal circulation Antiplatelet activity is achieved at low dose of ASA than that required for analgesia or endothelial PGI2 production?

Aspirin Antiplatelet Efficacy 1- Dose Inhibition of 90% to 95% of TXA2 levels is needed to abolish in vitro platelet aggregation Given over a period of days, this level of inhibition can be achieved with doses as low as 20 mg In volunteer subjects, a dose of 80 mg can inhibit aggregation as soon as 15 minutes after ingestion; in the unstable patient, however, in whom absorption is more likely to be a problem and in whom circulating platelets are more likely to be activated Most authorities recommend initial therapy with a dose of 160 mg (one half-tablet) to 325 mg (one adult tablet) The aspirin should be crushed or chewed to facilitate absorption. The frequency of gastrointestinal complications appears to be dose-dependent

Aspirin Antiplatelet Efficacy A, Efficacy of aspirin in patients with unstable angina Studies have demonstrated reductions in morbid ischemic events in patients with unstable angina following the ingestion of aspirin using varying doses of aspirin and varying periods between the onset of symptoms and treatment with aspirin B. Efficacy of aspirin in patients following acute MI Analysis of 10 trials of antiplatelet agents (mainly aspirin) for secondary prophylaxis of vascular events following acute MI, the Antiplatelet Trialists Collaboration reported striking reductions in nonfatal MI and nonfatal stroke in patients treated chronically with antiplatelet therapy (n=18,441) as well as a highly significant reduction in cardiovascular mortality Similar reductions were seen for patients receiving antiplatelet therapy following stroke. There were no differences in effect between varying doses of aspirin

II- Glycoprotein IIb/IIIa Receptor Antagonists 1- Glycoprotein IIb/IIIa murine-derived 7E3 Fab monoclonal antibody (Abciximab) Abciximab is composed of 7E3 Fab fragments a murine-derived (m) monoclonal antibody directed against glycoprotein GPIIb/IIIa. Mechanism: The m7E3 Fab binds selectively to the glycoprotein GPIIb/IIIa receptors inhibiting platelets binding to fibrinogen and von Willebrand factor, and consequently inhibiting platelet aggregation Clinical Efficacy: In acute MI patients, when the m7E3 Fab antibody was administered 3, 6, or 15 hours after beginning treatment with rt-PA, the incidence of recurrent ischemia was lower than expected

II- Glycoprotein IIb/IIIa Receptor Antagonists 1- Glycoprotein IIb/IIIa murine-derived 7E3 Fab monoclonal antibody (Abciximab) Administration and therapeutic use: Abciximab is administered intravenously as an adjuvant to angioplasty surgery for the prevention of ischemic complications of angioplasty Heparin or aspirin is given with abciximab Blockade of glycoprotein IIb/IIIa receptors following a bolus injection of c7E3 Fab (0.25 mg/kg): Although the plasma half-life of this monoclonal antibody is only several minutes, a potent biologic effect can be measured 2 hours after the injection In patients undergoing elective angioplasty, at this point, more than 80% of receptors are bound by the antibody (and therefore unable to participate in aggregation). At 6 hours, between 60% and 70% of receptors are bound, and at 24 hours, 50% are bound. In patients undergoing thrombolysis for acute MI, the return toward baseline occurs more rapidly. A continuous infusion leads to sustained blockade of the receptor

II- Glycoprotein IIb/IIIa Receptor Antagonists 2- Synthetic arginine-glycine-aspartic acid (R-G-D) sequence mimetics Eptifibatide (peptidergic) and tirofiban (non-peptiic) are synthetic mimetics of the R-G-D sequence of fibrinogen Hence, they block the binding of fibrinogen to glycoprotein GPIIb/IIIa receptors They are given intravenously for the reduction of thrombotic complications during coronary angioplasty Clinical trials showed reductions in incidence of death and non-fatal MI in response to the use of both agents

III- Thromboxane Antagonists Ridogrel is a combined thromboxane synthase inhibitor and thromboxane A2 (TXA2) receptor antagonist, orally active It has no effect on the vascular production of prostacyclin but cyclic endoperoxides (PGH2) may increase In the Ridogrel Aspirin Perfusion Trial (RAPT), patients received either ridogrel or aspirin (initially given IV then orally) + IV streptokinase for acute MI In both groups of patients inhibition of TXA2 was markedly inhibited, but recurrent ischemic events (angina, reinfarction, and ischemic stroke) were observed less frequently among the ridogrel-treated patients

IV- Platelet ADP Receptor Antagonists (Thienopyridines) Ticlopidine & Clopidogrel They inhibit irreversibly ADP binding to receptors Prevent ADP-mediated activation of the glycoprotein GPIIb/IIIa & ultimately inhibiting platelet aggregation No effect on PGs synthesis In the small percentage of patients who are truly aspirin-intolerant, ticlopidine should be considered as an alternative form of therapy

Ticlopidine & Clopidogrel Adverse Effects Ticlopidine is associated with more side effects than ticlopidine Nausea, dyspepsia, diarrhea (20% of patients) Hemorrahge (5%) Leukopenia in 1% of patients (most serious), check WBC in the first 3 months of treatment Ticlopidine, but not, clopidogrel can lead to fatal neutopenia Thrombotic thrombocytopenic purpura may occur with both agents

Other Antiplatelet Agents Dipyridamole, inhibits phosphodiesterase activity and inhibit adenosine uptake It is weak antiplatelet vasodilator If used, should be combined with aspirin Cliostazol is another PD inhibitor used for intermittent claudication

Antiplatelet Drugs

Overview of Antiplatelets