Pharmacology of emerging novel platelet inhibitors

Prasugrel (CS-747; LY-640315) 

Prasugrel is a third-generation oral thienopyridine that is chemically distinct from clopidogrel (Figure 1). Like clopidogrel, prasugrel is a specific, irreversible antagonist of the platelet adenosine 5′-diphosphate (ADP) P2Y12 receptor (Figure 2). Prasugrel is a prodrug that must be converted to an active metabolite (R-138727) by the hepatic cytochrome P450 (CYP) system to exert antiplatelet activity (Figure 3). It is rapidly absorbed and metabolized, with a median time for achieving the maximal concentration of its active metabolite in the circulation of about 30 minutes.1, 2 The mean elimination half-life of the prasugrel active metabolite is 3.7 hours, and renal excretion (approximately 70%) is the major route for elimination of prasugrel metabolites in humans.3 An important difference between the metabolism of prasugrel and that of clopidogrel is that a significant portion of the administered dose of clopidogrel is deactivated in the early stages of its metabolism, resulting in less availability of the active metabolite. In a 2-step biotransformation, prasugrel is rapidly hydrolyzed by carboxylesterases to a thiolactone (R-95913), which is then converted to the prasugrel active metabolite (R-138727) by multiple CYP isotypes, primarily CYP3A and CYP2B6.4 In contrast, a 2-step process that is mainly CYP3A4-dependent catalyzes the conversion of clopidogrel to 2-oxo-clopidogrel, which is then transformed to the clopidogrel active metabolite.5 Esterases primarily catalyze clopidogrel to an inactive carboxylic acid derivative (SR26334), whereas the prasugrel active metabolite R-138727 is metabolized to inactive metabolites through S-methylation. The active metabolites of both prasugrel and clopidogrel covalently bind to the active site of the P2Y12 receptor in a similar manner through a disulfide linkage.

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Figure 1. Chemical structures of antiplatelet agents. Shown here are the chemical structures of 6 established and emerging antiplatelet agents. Ticlopidine, clopidogrel, and prasugrel are first, second, and third-generation thienopyridines, respectively, that irreversibly inhibit the P2Y12 receptor. AZD6140 is a cyclopentyltriazolopyrimidine that reversibly inhibits the P2Y12 receptor. Cangrelor is an ATP analog that reversibly inhibits the P2Y12 receptor. SCH 530348 is a TRA that reversibly inhibits PAR-1.

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Figure 2. Molecular targets of established and emerging antiplatelet therapies. Thrombin binds to its cognate receptor, PAR-1, on platelets, which leads to shape change, granule secretion (including the extracellular release of ADP, which interacts with its platelet receptors to augment thrombin activity), thromboxane A2 (TXA2) generation, and activation of the glycoprotein (GP) IIb/IIIa receptor, resulting in sustained platelet aggregation. Protease-activated receptor 1, a 7-transmembrane serpentine receptor, is linked to Gq-and Gi-coupled signaling pathways that mediate platelet function. SCH 530348 is a selective PAR-1 receptor antagonist that abrogates thrombin-induced platelet activation and aggregation. Aspirin irreversibly inhibits cyclooxygenase (COX)-1, which catalyzes the production of TXA2, a potent platelet aggregator, generated by platelets activated by thrombin and other agonists (eg, ADP). Adenosine 5′-diphosphate binds to its 7-transmembrane serpentine receptors, P2Y1 and P2Y12, to activate platelets. P2Y1 is coupled to Gq and G12, and Gq is linked to a signaling pathway involving phospholipase C (PLC) activation, resulting in a rise in the intracellular calcium concentration ([Ca+2]i) and protein kinase C (PKC) activation, leading to GP IIb/IIIa activation and transient platelet aggregation. The P2Y1-coupled G12 signaling pathway mediates platelet shape change. P2Y12 is linked to Gi-coupled signaling cascades associated with adenylcyclase down-regulation and decreased cyclic-3′,5′-monophosphate (cAMP) production, which mediates GP IIb/IIIa receptor activation, leading to sustained platelet aggregation and a pathway that triggers augmentation of granule secretion and TXA2 generation and GP IIb/IIIa receptor activation. Adenosine 5′-triphosphate activates the P2X1 receptor, which triggers calcium movement associated with platelet shape change. The prodrugs clopidogrel and prasugrel are metabolized by the hepatic CYP system to active metabolites that are selective P2Y12 receptor antagonists that block ADP-induced platelet activation. AZD6140, a reversible P2Y12 antagonist, and cangrelor, an ATP analog, are selective P2Y12 receptor antagonists that block ADP-stimulated platelet aggregation. The binding of fibrinogen to activated GP IIb/IIIa receptors on adjacent platelets is the final common pathway for platelet aggregation and thrombus formation. Intravenous abciximab, eptifibatide, and tirofiban are selective GP IIb/IIIa receptor antagonists that block the GP IIb/IIIa receptor–fibrinogen interaction and thereby mitigate platelet aggregation.

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Figure 3. Primary metabolic pathways for prasugrel and clopidogrel. A significant portion (approximately 85%) of the administered dose of clopidogrel is deactivated in the early stages of its metabolism by esterases to a carboxylic acid derivative, SR26334, resulting in less availability of the active metabolite. A 2-step process that is mainly CYP3A4-dependent catalyzes the conversion of clopidogrel to 2-oxo-clopidogrel, which is then transformed to the clopidogrel active metabolite. Prasugrel is rapidly hydrolyzed by carboxyesterases to a lactone, R-95913, which is then converted to its active metabolite, R-138727, by multiple CYP isotypes. Thus, there is a more efficient metabolic conversion of prasugrel into its active metabolite, which yields concentrations 10-fold to 100-fold higher than that of clopidogrel. The active metabolites of both prasugrel and clopidogrel covalently bind to the active site of the P2Y12 receptor in a similar manner through a disulfide linkage.

As a consequence of the difference in the metabolic pathways for prasugrel and clopidogrel (Figure 3), prasugrel produces higher concentrations of its active metabolite than does clopidogrel.3 Consistent with these data, preclinical studies have shown that prasugrel is an orally active antiplatelet agent that is a more potent inhibitor of platelet aggregation on a milligram per kilogram basis, with a faster onset of action than that of clopidogrel.6 Prasugrel prevents thrombus formation dose-dependently in rats and prolongs bleeding time, a property also exhibited by clopidogrel and ticlopidine.7

Ticagrelor (AZD6140) 

AZD6140 or ticagrelor is the first of a new class of orally active antiplatelet agents, the cyclopentyltriazolopyrimidines (Figure 1). Similar to the thienopyridines, AZD6140 inhibits the prothrombotic effects of ADP by blocking the platelet P2Y12 receptor (Figure 2). This adenosine 5′-triphosphate (ATP) analog evolved through the chemical modification of ATP to yield cangrelor (see section below), an intravenously administered P2Y12 receptor antagonist, followed by β,γ-methylene substitutions of the ester linkages in the triphosphate group.8 It is notable that in contrast to AZD6140, which is relatively resistant to enzymatic degradation, the naturally occurring ATP is not suitable as an antiplatelet agent because it is rapidly inactivated in vivo by ectonucleotidases.8 Unlike the thienopyridines, AZD6140 binds to the P2Y12 receptor in a reversible fashion. AZD6140 confers nearly complete inhibition of ADP-induced platelet aggregation and is highly specific for the P2Y12 receptor with insignificant affinity for other P2 receptors.8 Furthermore, AZD6140 does not require metabolic activation for activity.

AR-C124910XX, an active metabolite of AZD6140, has been identified in the circulation; it is about as potent as the parent molecule with respect to blocking the P2Y12 receptor and is thought to contribute to the antiplatelet effect of the parent molecule.9 Peak plasma levels of AZD6140 are reached between 1.5 and 3 hours after treatment, with steady state reached after 2 to 3 days. The mean elimination half-life is 6 to 12 hours, independent of dose.9

Cangrelor (ARC-69931MX) 

Cangrelor is a member of an investigational novel chemical class of ATP analogs undergoing late-stage clinical development for intravenous use during coronary procedures (Figure 1). This compound is a high-affinity, reversible antagonist for the P2Y12 receptor that causes a pronounced, virtually total inhibition of ADP-induced platelet aggregation (Figure 2). Cangrelor, like AZD6140, is a chemically modified ATP derivative that is stable to enzymatic degradation. It does not require metabolic conversion to an active metabolite and hence is immediately active after infusion.10 The human pharmacokinetics of cangrelor is favorable, showing it to achieve rapid steady-state concentrations with a clearance of 50 L/h and a short half-life of 2.6 to 3.3 minutes.10

Animal studies with therapeutic doses of cangrelor have shown the inhibition of ex vivo ADP-induced platelet aggregation associated with antithrombotic effects without a significant prolongation of bleeding times.11, 12, 13, 14, 15 These preclinical models have demonstrated a separation between increase in bleeding times and platelet inhibition for cangrelor and a rapid return of normal platelet function after discontinuation of treatment.11, 12, 13 Studies in canine models have demonstrated that cangrelor successfully prevents thrombus formation.11, 12 Cangrelor also reduced the size and number of emboli in a thromboembolic model in rabbits14 and, when used in conjunction with concurrent fibrinolytic therapy, reduced the dose of fibrinolytic needed to achieve arterial patency in a canine thrombus model.15 Overall, the results of these preclinical studies imply that cangrelor has the potential to prevent thrombus formation and reduce emboli formation as well as restore arterial patency and improve blood flow in combination with fibrinolytic agents.

SCH 530348 (thrombin receptor antagonist) 

Thrombin is a major agonist of platelet activation and plays a seminal role in blood coagulation. Thus, the selective interference of thrombin-induced platelet activation, but not fibrinogen/fibrin-associated blood coagulation, may constitute an effective and safe form of antiplatelet therapy for cardiovascular diseases and PCI. SCH 530348 is the first of a new class of agents called thrombin receptor antagonists (TRAs) (Figure 1). Thrombin activates platelets and smooth muscle cells by the proteolytic activation of a cell surface Gp/Gi-coupled protease-activated receptor (PAR)-1, also known as the thrombin receptor, which is present in various cell types, including platelets (Figure 2). There are 4 known PAR subtypes, and activation of human platelets by thrombin is mediated by PAR-1 and PAR-4, with PAR-1 proposed as the principal thrombin receptor in human platelets.16, 17, 18, 19, 20 Thrombin binds to the hirudinlike extracellular amino terminal domain of PAR-1 and cleaves between Arg41 and Ser42, which results in the binding of the cleaved receptor to the neighboring thrombin receptor and subsequent cell activation.16 SCH 530348 blocks the platelet PAR-1 receptor to which thrombin binds (Figure 2), thus inhibiting thrombin-induced activation and aggregation of platelets. Because a TRA does not inhibit the ability of thrombin to catalyze the production of fibrin, agents in this class may produce fewer hemorrhagic side effects than would conventional anticoagulants.

Other emerging antiplatelet agents 

Two other antiplatelet agents in the early stages of clinical development also represent promising platelet inhibition therapies. PRT060128 (PRT128) is a direct-acting, reversible P2Y12 inhibitor available in both oral and intravenous formulations. In a randomized, double-blind, placebo-controlled trial in which single intravenous doses of PRT128, 1 to 40 mg, were administered for 20 minutes to 5 groups of 8 healthy subjects (6 receiving the active drug and 2 receiving placebo), PRT128 yielded dose-dependent, complete inhibition of ADP-induced platelet aggregation with maximum platelet inhibition observed at the first time point measured (20 minutes).21, 22 When combined with aspirin, PRT128 had a synergistic inhibitory effect on collagen-induced platelet aggregation.21 PRT128 also promoted thrombus destabilization and inhibited thrombus growth starting at a dose of 30 mg. All doses of PRT128 were well tolerated, with no serious adverse events observed.

ERASE-MI (Early Rapid Reversal of Platelet Thrombosis With Intravenous PRT060128 Before PCI to Optimize Reperfusion in Acute MI) is a phase II clinical trial of the intravenous formulation of PRT128 that has been initiated in patients experiencing an ST-segment elevation myocardial infarction. The trial will enroll approximately 200 patients across North America and the European Union.

E5555 is a novel protease PAR-1 antagonist that is currently in phase II testing. In studies on platelet-rich plasma derived from healthy volunteers, E5555 inhibited the release or expression of the inflammatory markers that have been linked to a high risk of events in patients with ACS, including the release of sCD40L and interleukin 6 and the expression of P selectin.23 Two randomized, double-blind, placebo-controlled phase II clinical trials of E5555 called LANCELOT (Lessons from Antagonizing the Cellular Effects of Thrombin) have been initiated to evaluate the safety and tolerability of this drug and to assess its effects on platelet aggregation inhibition, endovascular inflammatory processes, and the incidence of major adverse cardiovascular events in patients with coronary artery disease and non–ST-segment elevation ACS. The anticipated enrollment for each study is about 600 patients.

Because both PRT128 and E5555 are in the early stages of clinical development, they will not be discussed in greater detail in this supplement.

Conclusions 

There are few currently available alternative therapies for patients who have poor responsiveness to clopidogrel. The pharmacologic profiles of the 3 novel P2Y12 receptor antagonists and the TRA described here suggest that they may have clinical advantages compared with the currently available antiplatelet agents. Current clinical data in support of this conclusion will be further described in other contributions in this supplement.