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Drugs affecting coagulation
Pharmacology
Drugs affecting coagulation
First-generation agents Streptokinase is a non-enzymatic protein produced by β- haemolytic streptococci. It activates the fibrinolytic system indirectly by forming a 1:1 stoichometric complex with plasmin ogen, thus activating plasminogen into plasmin. Urokinase is a trypsin-like serine protease composed of two polypeptide chains connected by a disulphide bridge. It activates plasminogen directly, converting it to active plasmin. Anistreplase (anisoylated plasminogen streptokinase activa tor complex, Eminase) is a purified human plasminogen. It is a bacterial acylated streptokinase complex, which when given leads to deacylation, thus activating the streptokinaseproactivator complex. It is given by rapid intravenous injection and has enhanced clot selectivity. It has more activity at clotassociated plasminogen than at free blood plasminogen, thus its thrombolytic activity is greater.
Sian Marsh Balraj Appadu
Abstract Heparin and warfarin have been in clinical use for more than 50 years. However, the limitations of these traditional anticoagulants have prompted the development of new drugs. In the past 15 years new agents with improved safety profile and greater ease of use that target almost every step of the coagulation cascade have been developed. These include factor Xa inhibitors and direct thrombin inhibitors. The mechanism of action of these new anticoagulants and also the ‘older’ agents are reviewed in this article.
Second-generation agents Tissue plasminogen activator (t-PA) – native t-PA is a serine protease that consists of one polypeptide chain containing 527 amino acids. In the plasma, this molecule is converted to a two-chain activator linked by one disulphide bond by cleavage. It is a poor enzyme in the absence of fibrin, but fibrin strikingly enhances the activation rate of plasminogen. This unique property is explained by an increased affinity of fibrin-bound t-PA for plasminogen without significant influence on the catalytic efficiency of the enzyme. Fibrin essentially increases the local plasminogen concentration by creating an additional interaction between t-PA and its substrate. The high affinity of t-PA for plasminogen in the presence of fibrin thus allows efficient activation of the fibrin clot, whereas no efficient plasminogen activation by t-PA occurs in the plasma.
Keywords direct thrombin inhibitors; fondaparinux; glycoprotein IIb/IIIa antagonists; heparin; warfarin
Coagulation is a major defence mechanism against bleeding. After injury to the vessel wall, tissue factor is exposed on the surface of the damaged endothelium. The interaction between tissue factor and factor VII activates the coagulation cascade, which produces thrombin and culminates in the formation of an insoluble clot (Figure 1). Thrombin is central to the clotting process because it converts soluble fibrinogen to fibrin, activates factors V, VIII and XI (which generates more thrombin) and stimulates platelets. The coagulation cascade is regulated by natural anticoagulants, such as tissue factor pathway inhibitor (TFPI), the protein C and protein S systems, and antithrombin; all of which help to restrict the formation of a haemostatic plug at the site of injury.
Third-generation agents Reteplase (r-PA) or recombinant plasminogen activator is a deletion mutant that contains the kringle-2 and protease domains of the parent t-PA molecule. It has a prolonged half-life (18 minutes) and is given in two abbreviated intravenous infusions (lasting 2 minutes), 30 minutes apart. It is approved in the UK for the treatment of acute myocardial infarction. Lanoplase (n-PA) is a deletion and point mutant of wildtype t-PA. The deletion of the finger and epidermal growth factor domains and a point mutation within the kringle-1 domain contribute to the molecule’s long circulating half-life (30–45 minutes). In clinical trials n-PA was found to have a thrombolytic activity equivalent to r-PA, albeit with a higher incidence of bleeding.
Thrombolytic agents Thrombolytic therapy uses the vascular system’s native thromboresistant properties by accelerating and amplifying the conversion of an inactive precursor, plasminogen, to the active enzyme, plasmin. In turn, plasmin hydrolyses the fibrin clot matrix, leading to dissolution (lysis), thus restoring vital blood flow to the organs.
Direct thrombin inhibitors Thrombin-inhibiting drugs can block the action of thrombin by binding to three domains: the active or catalytic site and exosites 1 and 2, located near the active site. Exosite 1 acts as a dock for fibrin and exosite 2 serves as the heparin-binding domain. Bivalent inhibitors such as hirudin and bivalirudin block thrombin at the active site and exosite 1, and univalent inhibitors such as argatroban and melagatran (and its oral precursor, ximelagatran) bind only to the active site. By reducing the thrombin-mediated activation of platelets, these inhibitors also have an antiplatelet effect.
Sian Marsh, MBBS, FRCA, is a fourth-year Special Registrar in Anaesthetics on the East Anglian rotation. She qualified from St George’s Hospital Medical School, London in 1997. After basic training in many specialties in Queensland, Australia, she completed basic anaesthetic SHO training in Derbyshire NHS Trust Hospitals. Balraj Appadu, MD, FRCA, is a Consultant in Anaesthesia and Intensive Care at the Peterborough Hospitals NHS Foundation Trust. He graduated from the University of St Etienne, France in 1986. His main interests are basic sciences as applied to anaesthesia, intensive care, blood transfusion and haematology.
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Pharmacology
Overview of coagulation pathways, and selected antithrombotic agents Extrinsic pathway activation Factor VII Tissue thromboplastin Coagulation factors affected by warfarin
Activated factor X inhibitors Indirect • UFH • LMWH • Fondaparinux • Idraparinux Direct • Razaxaban
Factor X
Activated factor X Intrinsic pathway activation Factor IX Factor VIII
Prothrombin
Thrombin inhibitors Indirect • UFH Direct • Hirudin • Argatroban • Ximelagatran • Others
Prothrombinase complex Activated factor X Activated factor V Ca2+, phospholipid Thrombin
Fibrinogen
Soluble fibrin
Fibrin (clot)
Figure 1
Because direct thrombin inhibitors do not bind to plasma roteins they produce a more predictable response than heparin, p and should be more effective than low molecular weight heparin (LMWH) because they inhibit fibrin-bound thrombin. The pharmacokinetics profile of various direct thrombin inhibitors is shown in Table 1. These agents have a predominant renal clearance, and drugs such as hirudin and melagatran are likely to accumulate in patients with impaired renal function. Argatroban is predominantly cleared by hepatic metabolism and requires dose adjustment in patients with hepatic dysfunction. It seems that the use of aspirin does not influence the plasma
concentration of direct thrombin inhibitors. These inhibitors are mainly used in acute coronary syndromes with or without percutaneous coronary intervention and heparin-induced thrombocytopenia (HIT). The monitoring of treatment with these agents has not been clearly established. The usefulness of the activated partial thromboplastin time (APTT) is limited by its poor linearity and reproducibility. The ecarin clotting time gives a better reflection of the actual plasma concentration of direct thrombin inhibitors, and should be used in patients at a high risk of bleeding as there are no antidotes for rapidly reversing the effects of these agents.
Main properties and pharmacokinetic characteristics of direct thrombin inhibitors Product
Route of administration
Plasma half-life (min)
Main site of clearance
Dosage
Argatroban Lepirudin
Intravenous Intravenous
39–51 20–30
Liver Kidney
Bivalirudin
Intravenous
24
Kidney, liver
Ximelagatran Melagatran
Oral Intravenous Subcutaneous Oral
180–300 120–180 – 720
Kidney Kidney – Kidney
2 mg/kg/h 0.4 mg/kg bolus 0.1–0.15 mg/kg/h 1 mg/kg bolus 2.5 mg/kg/h for 4 h 0.2 mg/kg to 20 h 24–36 mg twice daily – – –
Dabigatran Table 1
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as new platelets are added to the circulation. Aspirin also inhibits COX-2. A single dose of 160 mg almost completely inhibits platelet COX-1. The same effect can be achieved by daily doses of 30–50 mg of aspirin for 7–10 days. Aspirin is rapidly deacetylated in the liver. However, platelet COX-1 can be inhibited by low-dose aspirin in the portal circulation, before aspirin enters the systemic circulation, thus sparing endothelial cell COX-1 and PGI2 synthesis. Aspirin significantly prolongs the bleeding time of individuals, although not necessarily into the abnormal range. Adverse events that result from aspirin therapy comprise bleeding and gastrointestinal toxicity (heartburn, indigestion, nausea and vomiting, and gastric ulcer). Aspirin has been used in primary and secondary disease prevention in patients with atherotic vascular disease.
Platelet antagonists Platelets play a critical role in normal haemostasis by adhering to sites of injury. However, platelets adhere and ultimately aggregate to abnormal vascular surfaces via surface membrane glycoprotein receptors that can be expressed in greater numbers with platelet activation. These glycoproteins have been used as potential targets for therapy. Glycoprotein IIb/IIIa inhibitors glycoprotein (GP) IIb/IIIA receptors represent the final common pathway to platelet aggregation and clot formation. GP IIb/IIIa is a member of the integrin family of receptors consisting of α and β subunits. Bleeding complications represent a significant limitation to the effectiveness of GP IIb/IIIa inhibitors, with most bleeding occurring at the vascular access sites during interventional procedures. Thrombocytopenia is an uncommon, rarely fatal, but nonetheless worrisome complication of GP IIb/IIIa inhibitors. GP IIb/IIIa inhibitors are increasingly being used in patients with unstable angina or nonST segment elevation myocardial infarction undergoing percutaneous coronary interventions. GP IIb/IIIa inhibitors have been shown to reduce the risk of death, myocardial infarction and the need for revascularization therapy, with these effects persisting after 1 year. Three GP IIb/IIIa inhibitors are available for clinical use in the UK. Abcimixab (47615 Da) is the Fab fragment of the chimeric human-murine monoclonal antibody, produced by continuous perfusion in mammalian cell culture. Abcimixab binding is non-specific. Following intravenous bolus administration, free plasma concentration of abcimixab decreases rapidly, with an initial half-life of less than 10 minutes and a second-phase halflife of 30 minutes, which is probably related to the high-affinity binding to the GP IIb/IIIa receptor. Platelet function recovers within 48 hours. However, abcimixab remains in the circulation for up to 10 days in the platelet-bound state. This property has an important clinical implication in situations where rapid abcimixab reversal is needed. Platelet transfusion will be effective in increasing the pool of circulating platelets and their available GP IIb/IIIa receptors. Abcimixab will redistribute over the entire pool, thereby diluting the drug’s antiplatelet effect. Eptifibatide (835 Da) is a specific inhibitor of GP IIb/IIIa receptor and has a low affinity. The cyclic structure of eptifibatide makes it more resistant to plasma proteases. Plasma clearance is mainly renal, and the dose should be adjusted for patients with renal impairment. The elimination half-life is 2–3 hours. Tirofibran (495 Da) is a non-immunogenic, non-peptide small molecule, which has the same affinity to the GP IIb/IIIa receptor as well as similar pharmacokinetic properties as eptifibatide. Its inhibitory effect rapidly decreases after discontinuation of the drug. Elimination of tirofibran is mainly renal, with a half-life similar to eptifibatide.
Inhibitors of ADP-mediated platelet activation Thienopyridines – ticlopidine and clopidogrel are thienopyridine derivatives that impair platelet function by irreversibly antagonizing the receptor for ADP. Platelets of patients given ticlopidine or clopidogrel are unable to aggregate ex vivo in response to ADP. Neither ticlopidine nor clopidogrel inhibit platelet function in vitro at clinically effective doses. Rather, unidentified hepatic metabolites seem to be inhibitory agents. Therefore, it is necessary to administer these drugs for 3–5 days to maximally inhibit platelet aggregation, and their effects persist for up to 10 days after they are withdrawn. The inhibition of platelet aggregation by ticlopidine and clopidogrel is concentration dependent. Ticlopidine has a number of potentially serious side effects: 2% of patients develop reversible granulocytopenia usually within the first 2 months of starting therapy. Further, cases of aplastic anaemia have been reported. Other toxicity includes diarrhoea in up to 20% of patients, nausea, cholestatic jaundice and skin rashes. A number of reports have associated ticlopidine use with thrombotic thrombocytic pupura (TTP) within one month of starting treatment. Ticlopidine has been used for the secondary prevention of thrombosis in patients with established vascular diseases. It is often used with aspirin to prevent thrombosis and vascular events in patients who have had coronary stenting. Clopidogrel is a structural analogue of ticlopidine with an identical mechanism of action. However, it does not have the side effects of ticlopidine such as TTP or granulocytopenia. Thus, it is increasingly being used instead of ticlopidine to prevent thrombosis in patients receiving coronary stents. Phosphodiesterase inhibitors: dipyridamole is a phospho diesterase inhibitor that has been used as an antiplatelet agent, normally with either warfarin or aspirin. However, there is very little evidence to suggest that dipyridamole is an effective phosphodiesterase inhibitor at plasma concentrations following oral administration. The combination of dipyridamole and aspirin has been shown to be more effective in the secondary prevention of stroke. Cilostazol, a quinolone derivative, is a specific inhibitor of prostaglandin E3, the most abundant form of phosphodiesterase isoform present in platelets. Besides inhibiting platelet function, cilostazol is a vasodilator and has been used for the treatment of intermittent claudication due to peripheral vascular disease. Side
Cyclooxygenase inhibitors Aspirin irreversibly inactivates the enzyme cyclooxygenase- 1 (COX-1). The antiplatelet effects of aspirin represent a balance between inhibition of thromboxane A2 by platelets and prostaglandin I2 (PGI2) synthesis by endothelial cells. COX- 1 is expressed by many tissues, including platelets, the gastric mucosa and endothelial cells. Platelets cannot replace acetylated COX-1, thus after aspirin is stopped, COX-1 activity returns only
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effects include headache, diarrhoea, palpitations and dizziness. It is contraindicated in patients with congestive heart disease.
metabolized by microsomal liver enzymes, including cytochrome p450, and its breakdown products are excreted as glucuronide conjugates in urine. Oral anticoagulants are used in the prophylaxis and treatment of deep vein thrombosis, pulmonary embolism, atrial fibrillation with a risk of embolization, and to prevent the deposition of thrombi on prosthetic heart valves. There is considerable variability in the response of patients to warfarin. Its effectiveness is influenced by age, genetic factors, diet, liver disease, and drug interactions. It is therefore essential to monitor warfarin therapy, comparing the prothrombin time with a control international standard to produce an international normalized ratio (INR). The target INR ranges between 2.0 and 4.0 depending on the clinical situation. The adverse effects of warfarin include haemorrhage, hypersensitivity, skin rashes, alopecia, and purpura. Warfarin can readily cross the placenta causing congenital malformations, haemorrhage, and intrauterine death. If urgent reversal of warfarin is needed, treatment includes infusion of fresh frozen plasma or prothrombin complex concentrate. Recombinant factor VIIa can also be used if severe bleeding occurs.
Activated Factor X Inhibitors Fondaparinux is a synthetic analogue of the antithrombinbinding pentasaccharide sequence that mediates the anti coagulant activity of heparin and LMWH. Fondaparinux binds antithrombin and enhances its reactivity with factor Xa; however, it is a small molecule which is too short to bridge antithrombin to thrombin, thus fondaparinux has no effect on the rate of thrombin inhibition. Fondaparinux exhibits complete bioavailability after subcutaneous injections, and with a plasma half-life of 17 hours, it can be administered once daily. This agent is excreted unchanged in the urine and is contraindicated in patients with renal insufficiency. Fondaparinux does not cause HIT because it does not bind to platelet factor 4 (PF4) to form the heparin/PF4 complexes, which are antigenic targets for antibodies that cause the condition. There is no antidote to fondaparinux. If uncontrolled bleeding occurs, recombinant factor VIIa may be effective. Fondaparinux has been evaluated for thromboprophylaxis in orthopaedic, medical and surgical patients, initial treatment of venous thromboembolism, and acute coronary syndromes.
Heparins Unfractionated heparin (UFH) is a glycosaminoglycan. It consists of a group of mucopolysaccharides and has an average molecular weight of 15,000 Da. Commercial preparations are obtained from bovine lung or porcine intestine. The anticoagulant activity of UFH depends on a specific pentasaccharide, present in about 30% of heparin molecules, which binds to antithrombin-III (AT- III). Heparin binding produces a conformational change in the reactive site of the AT-III molecule. This enhances the action of AT-III in combining and neutralizing factors VIIa, IXa, XIa, XIIa, and thrombin, to form stable complexes. Heparin then dissociates from the AT-III complex and can be reused. Thrombin (IIa) and factor Xa are most responsive to inhibition. Inactivation of thrombin inhibits fibrin formation and the activation of factor V and VII. Heparin and LMWHs also induce a vascular endothelial cell TFPI that reduces the procoagulant activity of tissue factor VII complex, which contributes to their antithrombotic action. Heparin may inhibit von Willebranddependent platelet function, and in high doses can inhibit platelet aggregation to prolong bleeding time. Heparin is a large polar molecule, which has difficulty crossing cell membranes. It is ineffective orally, must be given intravenously or subcutaneously and does not easily cross the placenta or blood–brain barrier. Once in the circulation, heparin binds to plasma proteins that can reduce its activity at low concentrations. Clearance initially occurs by a rapid saturatable mechanism, involving binding to endothelial cells and macrophages, and a slow first-order mechanism due to renal clearance. The half-life and intensity of the effects of heparin rise disproportionately with increasing dose. The effects of heparin can be monitored by measuring the APTT, where plasma, phospholipids and calcium are added to kaolin. The APTT is sensitive to the effects of heparin on thrombin, factor Xa and IXa. Therapeutic levels are generally 1.5 to 2.5 times that of the control. High doses of heparin can affect the prothrombin time (PT), but to a lesser extent than its effect on the APTT.
Idraparinux is a more negatively charged derivative of fondaparinux. It binds antithrombin with higher affinity than fondaparinux. Thus, idraparinux has a terminal plasma half-life of around 80 hours, similar to that of thrombin. Consequently, it can be given subcutaneously once a week. However, idraparinux has been shown to produce dose-dependent bleeding. In a phase II trial, patients given a dose of 5 mg had a significantly higher risk of severe bleeding, but patients given a dose of 2.5 mg had less bleeding than patients receiving warfarin.1 Razaxaban is a synthetic, non-peptidic, orally active factor Xa inhibitor. It has been evaluated in a phase II trial of patients undergoing elective knee arthroplasty. However, the risk of bleeding in the razaxaban patient group was significantly high and the trial was prematurely stopped. Oral anticoagulants Warfarin sodium is a 4-hydroxycoumarin compound and is now the standard oral anticoagulant. Warfarin inhibits the synthesis of factors dependent on vitamin K (factors II, VII, IX, and X, protein C, protein S). Carboxylation of the glutamate groups in the inactive precursor proteins is required for the synthesis of active coagulation factors. This process is coupled with the oxidation and reduction of vitamin K. Warfarin inhibits epoxide reductase, the enzyme required for continual production of the reduced, biologically active form of vitamin K, and therefore prevents activation of precursors. Circulating factors are not affected by warfarin and its effects are delayed until previously synthesized factors are consumed. Factor VII decreases rapidly (during less than 24 hours). However, factor II has a longer halflife, which means that the therapeutic effects of warfarin may take 48–72 hours to develop. Warfarin is rapidly absorbed from the gastrointestinal tract. Approximately 97% of warfarin is bound to albumin, resulting in partly restricted diffusion across membranes. Warfarin is
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Comparison of low molecular weight heparins Product
Preparation
Mean molecular weight (range, Da)
Half-life (min)
Xa:IIa ratio
Bioavailability (%)
Enoxaparin Dalteparin Nadroparin Tinzaparin Ardeparin Reviparin Heparin
Benzylation, alkaline hydrolysis Nitrous oxide depolymerization Nitrous acid depolymerization Heparinase digestion Peroxidase cleavage Nitrous acid digestion –
4500 (3000–8000) 5000 (2000–9000) 4500 (2000–8000) 4500 (3000–6000) 6000 (2000–15,000) 4600 (3000–7000) 11400
129–180 119–139 132–162 111 200 180 60
2.7:1 2.0:1 3.2:1 1.9:1 2.0:1 1.9:1 1.0:1
90–92 87 98 90 92 90 30 (range 10–40)
Table 2
The most important adverse effect of heparin is bleeding, which is treated by stopping the drug and administering prota mine, a strong basic drug that forms an inactive complex with heparin. Thrombocytopenia can also occur, either at an early stage or with long-term treatment. HIT can cause venous and arterial thrombosis, a fall in platelets or skin lesions. Hyper sensitivity, osteoporosis, and alopecia have also been reported.
molecular weight heparinoid consisting of a mixture of heparan sulphate (84%), dermatan sulphate (12%) and small amounts of chondroitin sulphate (4%), and its antithrombotic activity is well established. Its pharmacological effect is exerted primarily by inhibiting factors Xa and IIa at a ratio greater than heparin, with a minimal effect on platelet function. Danaparoid exhibits low cross-reactivity with heparin-induced antibodies when compared with heparin or LMWHs, thereby making it an excellent choice for the management of HIT. It has excellent bioavailability following subcutaneous injection. Danaparoid has little effect on routine coagulation tests (APTT, PT and thrombin time). Patients with elevated serum creatinine should be monitored carefully. Danaparoid has been found to be effective in the treatment of HIT and is frequently used for this purpose. However, the information accompanying this drug does not mention its use for the treatment of this condition. ◆
Low molecular weight heparins: standard heparin can be fractionated either by chemical or enzymatic depolymerization to produce preparations with a lower molecular weight (4000–6000 Da) (Table 2). LMWHs produce their anticoagulant effect by binding and neutralizing factor Xa. They have minimal inhibitory effects on thrombin because compared with UFH, the smaller fragments of LMWHs are unable to bind AT-III and thrombin simultan eously. This bridging process is less critical for the inactivation of factor Xa. Different preparations vary in their ratio of anti-Xa to antithrombin activity. LMWHs are well absorbed when given subcutaneously and there is little binding to circulating and cellular proteins. Clearance occurs by the renal route and biological half-life is prolonged in patients with renal failure. LMWHs have a longer plasma halflife and a better bioavailability at low doses than UFH. They have a more predictable dose–response relationship than UFH, and the dose is calculated on the basis of patient weight. Monitoring is rarely required except in rare cases where dosage may be difficult (e.g. morbid obesity or renal failure). There is no effect on APTT, and it is necessary to measure plasma anti- Xa activity, ideally 4 hours after injection. Therapeutic benefits of LMWHs seem to be in the reduction of mortality in unstable angina, and in surgical thromboprophylaxis. There is still a risk of haemorrhage with LMWHs, but they have less of an effect on platelet aggregation, and a lower risk of thrombocytopenia than UFH. Protamine has little effect on reversing LMWHs because its main mechanism of action is in neutralizing antithrombin activity.
Reference 1 PERSIST Investigators: a novel long-acting synthetic factor Xa inhibitor (SanOrg34006) to replace warfarin for secondary prevention in deep vein thrombosis: a Phase II evaluation. J Thromb Haemost 2004; 2: 47–53. (Erratum in: Blood 2002; 100: 82a).
Further reading Agnelli G, Becattini C. New anticoagulants. Semi Thromb Hemost 2006; 32: 793–802. Bates SM, Weitz J. The status of new anticoagulants. Br J Haematol 2006; 134: 3–19. Hirsch J, Warkentin TE, Shaughnessy SG, et al. Heparin and low molecular weight heparin: mechanism of action, pharmacokinetics, dosing, monitoring, efficacy, and safety. Chest 2001; 119: 64S–94S. Kam PCA, Egan MK. Platelet glycoprotein IIb/IIIa antagonists: pharmacology and clinical developments. Anesthesiology 2002; 96: 1237–49. Vandermeulen E. Anaesthesia and new antithrombotic drugs. Curr Opin Anaesthesiol 2005; 18: 353–9. Weitz JI, Hirsch J. New anticoagulant drugs. Chest 2001; 119: 95S– 107S.
Danaparoid sodium: danaparoid sodium (Orgaran) is a heparinoid glycosaminoglycuronan antithrombotic agent approved in the UK for the prophylaxis of postoperative deep vein throm bosis, which may lead to pulmonary embolism in patients undergoing elective hip replacement surgery. Danaparoid is a low
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Drugs affecting coagulation
First-generation agents Streptokinase is a non-enzymatic protein produced by β- haemolytic streptococci. It activates the fibrinolytic system indirectly by forming a 1:1 stoichometric complex with plasmin ogen, thus activating plasminogen into plasmin. Urokinase is a trypsin-like serine protease composed of two polypeptide chains connected by a disulphide bridge. It activates plasminogen directly, converting it to active plasmin. Anistreplase (anisoylated plasminogen streptokinase activa tor complex, Eminase) is a purified human plasminogen. It is a bacterial acylated streptokinase complex, which when given leads to deacylation, thus activating the streptokinaseproactivator complex. It is given by rapid intravenous injection and has enhanced clot selectivity. It has more activity at clotassociated plasminogen than at free blood plasminogen, thus its thrombolytic activity is greater.
Sian Marsh Balraj Appadu
Abstract Heparin and warfarin have been in clinical use for more than 50 years. However, the limitations of these traditional anticoagulants have prompted the development of new drugs. In the past 15 years new agents with improved safety profile and greater ease of use that target almost every step of the coagulation cascade have been developed. These include factor Xa inhibitors and direct thrombin inhibitors. The mechanism of action of these new anticoagulants and also the ‘older’ agents are reviewed in this article.
Second-generation agents Tissue plasminogen activator (t-PA) – native t-PA is a serine protease that consists of one polypeptide chain containing 527 amino acids. In the plasma, this molecule is converted to a two-chain activator linked by one disulphide bond by cleavage. It is a poor enzyme in the absence of fibrin, but fibrin strikingly enhances the activation rate of plasminogen. This unique property is explained by an increased affinity of fibrin-bound t-PA for plasminogen without significant influence on the catalytic efficiency of the enzyme. Fibrin essentially increases the local plasminogen concentration by creating an additional interaction between t-PA and its substrate. The high affinity of t-PA for plasminogen in the presence of fibrin thus allows efficient activation of the fibrin clot, whereas no efficient plasminogen activation by t-PA occurs in the plasma.
Keywords direct thrombin inhibitors; fondaparinux; glycoprotein IIb/IIIa antagonists; heparin; warfarin
Coagulation is a major defence mechanism against bleeding. After injury to the vessel wall, tissue factor is exposed on the surface of the damaged endothelium. The interaction between tissue factor and factor VII activates the coagulation cascade, which produces thrombin and culminates in the formation of an insoluble clot (Figure 1). Thrombin is central to the clotting process because it converts soluble fibrinogen to fibrin, activates factors V, VIII and XI (which generates more thrombin) and stimulates platelets. The coagulation cascade is regulated by natural anticoagulants, such as tissue factor pathway inhibitor (TFPI), the protein C and protein S systems, and antithrombin; all of which help to restrict the formation of a haemostatic plug at the site of injury.
Third-generation agents Reteplase (r-PA) or recombinant plasminogen activator is a deletion mutant that contains the kringle-2 and protease domains of the parent t-PA molecule. It has a prolonged half-life (18 minutes) and is given in two abbreviated intravenous infusions (lasting 2 minutes), 30 minutes apart. It is approved in the UK for the treatment of acute myocardial infarction. Lanoplase (n-PA) is a deletion and point mutant of wildtype t-PA. The deletion of the finger and epidermal growth factor domains and a point mutation within the kringle-1 domain contribute to the molecule’s long circulating half-life (30–45 minutes). In clinical trials n-PA was found to have a thrombolytic activity equivalent to r-PA, albeit with a higher incidence of bleeding.
Thrombolytic agents Thrombolytic therapy uses the vascular system’s native thromboresistant properties by accelerating and amplifying the conversion of an inactive precursor, plasminogen, to the active enzyme, plasmin. In turn, plasmin hydrolyses the fibrin clot matrix, leading to dissolution (lysis), thus restoring vital blood flow to the organs.
Direct thrombin inhibitors Thrombin-inhibiting drugs can block the action of thrombin by binding to three domains: the active or catalytic site and exosites 1 and 2, located near the active site. Exosite 1 acts as a dock for fibrin and exosite 2 serves as the heparin-binding domain. Bivalent inhibitors such as hirudin and bivalirudin block thrombin at the active site and exosite 1, and univalent inhibitors such as argatroban and melagatran (and its oral precursor, ximelagatran) bind only to the active site. By reducing the thrombin-mediated activation of platelets, these inhibitors also have an antiplatelet effect.
Sian Marsh, MBBS, FRCA, is a fourth-year Special Registrar in Anaesthetics on the East Anglian rotation. She qualified from St George’s Hospital Medical School, London in 1997. After basic training in many specialties in Queensland, Australia, she completed basic anaesthetic SHO training in Derbyshire NHS Trust Hospitals. Balraj Appadu, MD, FRCA, is a Consultant in Anaesthesia and Intensive Care at the Peterborough Hospitals NHS Foundation Trust. He graduated from the University of St Etienne, France in 1986. His main interests are basic sciences as applied to anaesthesia, intensive care, blood transfusion and haematology.
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© 2007 Published by Elsevier Ltd.
Pharmacology
Overview of coagulation pathways, and selected antithrombotic agents Extrinsic pathway activation Factor VII Tissue thromboplastin Coagulation factors affected by warfarin
Activated factor X inhibitors Indirect • UFH • LMWH • Fondaparinux • Idraparinux Direct • Razaxaban
Factor X
Activated factor X Intrinsic pathway activation Factor IX Factor VIII
Prothrombin
Thrombin inhibitors Indirect • UFH Direct • Hirudin • Argatroban • Ximelagatran • Others
Prothrombinase complex Activated factor X Activated factor V Ca2+, phospholipid Thrombin
Fibrinogen
Soluble fibrin
Fibrin (clot)
Figure 1
Because direct thrombin inhibitors do not bind to plasma roteins they produce a more predictable response than heparin, p and should be more effective than low molecular weight heparin (LMWH) because they inhibit fibrin-bound thrombin. The pharmacokinetics profile of various direct thrombin inhibitors is shown in Table 1. These agents have a predominant renal clearance, and drugs such as hirudin and melagatran are likely to accumulate in patients with impaired renal function. Argatroban is predominantly cleared by hepatic metabolism and requires dose adjustment in patients with hepatic dysfunction. It seems that the use of aspirin does not influence the plasma
concentration of direct thrombin inhibitors. These inhibitors are mainly used in acute coronary syndromes with or without percutaneous coronary intervention and heparin-induced thrombocytopenia (HIT). The monitoring of treatment with these agents has not been clearly established. The usefulness of the activated partial thromboplastin time (APTT) is limited by its poor linearity and reproducibility. The ecarin clotting time gives a better reflection of the actual plasma concentration of direct thrombin inhibitors, and should be used in patients at a high risk of bleeding as there are no antidotes for rapidly reversing the effects of these agents.
Main properties and pharmacokinetic characteristics of direct thrombin inhibitors Product
Route of administration
Plasma half-life (min)
Main site of clearance
Dosage
Argatroban Lepirudin
Intravenous Intravenous
39–51 20–30
Liver Kidney
Bivalirudin
Intravenous
24
Kidney, liver
Ximelagatran Melagatran
Oral Intravenous Subcutaneous Oral
180–300 120–180 – 720
Kidney Kidney – Kidney
2 mg/kg/h 0.4 mg/kg bolus 0.1–0.15 mg/kg/h 1 mg/kg bolus 2.5 mg/kg/h for 4 h 0.2 mg/kg to 20 h 24–36 mg twice daily – – –
Dabigatran Table 1
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as new platelets are added to the circulation. Aspirin also inhibits COX-2. A single dose of 160 mg almost completely inhibits platelet COX-1. The same effect can be achieved by daily doses of 30–50 mg of aspirin for 7–10 days. Aspirin is rapidly deacetylated in the liver. However, platelet COX-1 can be inhibited by low-dose aspirin in the portal circulation, before aspirin enters the systemic circulation, thus sparing endothelial cell COX-1 and PGI2 synthesis. Aspirin significantly prolongs the bleeding time of individuals, although not necessarily into the abnormal range. Adverse events that result from aspirin therapy comprise bleeding and gastrointestinal toxicity (heartburn, indigestion, nausea and vomiting, and gastric ulcer). Aspirin has been used in primary and secondary disease prevention in patients with atherotic vascular disease.
Platelet antagonists Platelets play a critical role in normal haemostasis by adhering to sites of injury. However, platelets adhere and ultimately aggregate to abnormal vascular surfaces via surface membrane glycoprotein receptors that can be expressed in greater numbers with platelet activation. These glycoproteins have been used as potential targets for therapy. Glycoprotein IIb/IIIa inhibitors glycoprotein (GP) IIb/IIIA receptors represent the final common pathway to platelet aggregation and clot formation. GP IIb/IIIa is a member of the integrin family of receptors consisting of α and β subunits. Bleeding complications represent a significant limitation to the effectiveness of GP IIb/IIIa inhibitors, with most bleeding occurring at the vascular access sites during interventional procedures. Thrombocytopenia is an uncommon, rarely fatal, but nonetheless worrisome complication of GP IIb/IIIa inhibitors. GP IIb/IIIa inhibitors are increasingly being used in patients with unstable angina or nonST segment elevation myocardial infarction undergoing percutaneous coronary interventions. GP IIb/IIIa inhibitors have been shown to reduce the risk of death, myocardial infarction and the need for revascularization therapy, with these effects persisting after 1 year. Three GP IIb/IIIa inhibitors are available for clinical use in the UK. Abcimixab (47615 Da) is the Fab fragment of the chimeric human-murine monoclonal antibody, produced by continuous perfusion in mammalian cell culture. Abcimixab binding is non-specific. Following intravenous bolus administration, free plasma concentration of abcimixab decreases rapidly, with an initial half-life of less than 10 minutes and a second-phase halflife of 30 minutes, which is probably related to the high-affinity binding to the GP IIb/IIIa receptor. Platelet function recovers within 48 hours. However, abcimixab remains in the circulation for up to 10 days in the platelet-bound state. This property has an important clinical implication in situations where rapid abcimixab reversal is needed. Platelet transfusion will be effective in increasing the pool of circulating platelets and their available GP IIb/IIIa receptors. Abcimixab will redistribute over the entire pool, thereby diluting the drug’s antiplatelet effect. Eptifibatide (835 Da) is a specific inhibitor of GP IIb/IIIa receptor and has a low affinity. The cyclic structure of eptifibatide makes it more resistant to plasma proteases. Plasma clearance is mainly renal, and the dose should be adjusted for patients with renal impairment. The elimination half-life is 2–3 hours. Tirofibran (495 Da) is a non-immunogenic, non-peptide small molecule, which has the same affinity to the GP IIb/IIIa receptor as well as similar pharmacokinetic properties as eptifibatide. Its inhibitory effect rapidly decreases after discontinuation of the drug. Elimination of tirofibran is mainly renal, with a half-life similar to eptifibatide.
Inhibitors of ADP-mediated platelet activation Thienopyridines – ticlopidine and clopidogrel are thienopyridine derivatives that impair platelet function by irreversibly antagonizing the receptor for ADP. Platelets of patients given ticlopidine or clopidogrel are unable to aggregate ex vivo in response to ADP. Neither ticlopidine nor clopidogrel inhibit platelet function in vitro at clinically effective doses. Rather, unidentified hepatic metabolites seem to be inhibitory agents. Therefore, it is necessary to administer these drugs for 3–5 days to maximally inhibit platelet aggregation, and their effects persist for up to 10 days after they are withdrawn. The inhibition of platelet aggregation by ticlopidine and clopidogrel is concentration dependent. Ticlopidine has a number of potentially serious side effects: 2% of patients develop reversible granulocytopenia usually within the first 2 months of starting therapy. Further, cases of aplastic anaemia have been reported. Other toxicity includes diarrhoea in up to 20% of patients, nausea, cholestatic jaundice and skin rashes. A number of reports have associated ticlopidine use with thrombotic thrombocytic pupura (TTP) within one month of starting treatment. Ticlopidine has been used for the secondary prevention of thrombosis in patients with established vascular diseases. It is often used with aspirin to prevent thrombosis and vascular events in patients who have had coronary stenting. Clopidogrel is a structural analogue of ticlopidine with an identical mechanism of action. However, it does not have the side effects of ticlopidine such as TTP or granulocytopenia. Thus, it is increasingly being used instead of ticlopidine to prevent thrombosis in patients receiving coronary stents. Phosphodiesterase inhibitors: dipyridamole is a phospho diesterase inhibitor that has been used as an antiplatelet agent, normally with either warfarin or aspirin. However, there is very little evidence to suggest that dipyridamole is an effective phosphodiesterase inhibitor at plasma concentrations following oral administration. The combination of dipyridamole and aspirin has been shown to be more effective in the secondary prevention of stroke. Cilostazol, a quinolone derivative, is a specific inhibitor of prostaglandin E3, the most abundant form of phosphodiesterase isoform present in platelets. Besides inhibiting platelet function, cilostazol is a vasodilator and has been used for the treatment of intermittent claudication due to peripheral vascular disease. Side
Cyclooxygenase inhibitors Aspirin irreversibly inactivates the enzyme cyclooxygenase- 1 (COX-1). The antiplatelet effects of aspirin represent a balance between inhibition of thromboxane A2 by platelets and prostaglandin I2 (PGI2) synthesis by endothelial cells. COX- 1 is expressed by many tissues, including platelets, the gastric mucosa and endothelial cells. Platelets cannot replace acetylated COX-1, thus after aspirin is stopped, COX-1 activity returns only
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effects include headache, diarrhoea, palpitations and dizziness. It is contraindicated in patients with congestive heart disease.
metabolized by microsomal liver enzymes, including cytochrome p450, and its breakdown products are excreted as glucuronide conjugates in urine. Oral anticoagulants are used in the prophylaxis and treatment of deep vein thrombosis, pulmonary embolism, atrial fibrillation with a risk of embolization, and to prevent the deposition of thrombi on prosthetic heart valves. There is considerable variability in the response of patients to warfarin. Its effectiveness is influenced by age, genetic factors, diet, liver disease, and drug interactions. It is therefore essential to monitor warfarin therapy, comparing the prothrombin time with a control international standard to produce an international normalized ratio (INR). The target INR ranges between 2.0 and 4.0 depending on the clinical situation. The adverse effects of warfarin include haemorrhage, hypersensitivity, skin rashes, alopecia, and purpura. Warfarin can readily cross the placenta causing congenital malformations, haemorrhage, and intrauterine death. If urgent reversal of warfarin is needed, treatment includes infusion of fresh frozen plasma or prothrombin complex concentrate. Recombinant factor VIIa can also be used if severe bleeding occurs.
Activated Factor X Inhibitors Fondaparinux is a synthetic analogue of the antithrombinbinding pentasaccharide sequence that mediates the anti coagulant activity of heparin and LMWH. Fondaparinux binds antithrombin and enhances its reactivity with factor Xa; however, it is a small molecule which is too short to bridge antithrombin to thrombin, thus fondaparinux has no effect on the rate of thrombin inhibition. Fondaparinux exhibits complete bioavailability after subcutaneous injections, and with a plasma half-life of 17 hours, it can be administered once daily. This agent is excreted unchanged in the urine and is contraindicated in patients with renal insufficiency. Fondaparinux does not cause HIT because it does not bind to platelet factor 4 (PF4) to form the heparin/PF4 complexes, which are antigenic targets for antibodies that cause the condition. There is no antidote to fondaparinux. If uncontrolled bleeding occurs, recombinant factor VIIa may be effective. Fondaparinux has been evaluated for thromboprophylaxis in orthopaedic, medical and surgical patients, initial treatment of venous thromboembolism, and acute coronary syndromes.
Heparins Unfractionated heparin (UFH) is a glycosaminoglycan. It consists of a group of mucopolysaccharides and has an average molecular weight of 15,000 Da. Commercial preparations are obtained from bovine lung or porcine intestine. The anticoagulant activity of UFH depends on a specific pentasaccharide, present in about 30% of heparin molecules, which binds to antithrombin-III (AT- III). Heparin binding produces a conformational change in the reactive site of the AT-III molecule. This enhances the action of AT-III in combining and neutralizing factors VIIa, IXa, XIa, XIIa, and thrombin, to form stable complexes. Heparin then dissociates from the AT-III complex and can be reused. Thrombin (IIa) and factor Xa are most responsive to inhibition. Inactivation of thrombin inhibits fibrin formation and the activation of factor V and VII. Heparin and LMWHs also induce a vascular endothelial cell TFPI that reduces the procoagulant activity of tissue factor VII complex, which contributes to their antithrombotic action. Heparin may inhibit von Willebranddependent platelet function, and in high doses can inhibit platelet aggregation to prolong bleeding time. Heparin is a large polar molecule, which has difficulty crossing cell membranes. It is ineffective orally, must be given intravenously or subcutaneously and does not easily cross the placenta or blood–brain barrier. Once in the circulation, heparin binds to plasma proteins that can reduce its activity at low concentrations. Clearance initially occurs by a rapid saturatable mechanism, involving binding to endothelial cells and macrophages, and a slow first-order mechanism due to renal clearance. The half-life and intensity of the effects of heparin rise disproportionately with increasing dose. The effects of heparin can be monitored by measuring the APTT, where plasma, phospholipids and calcium are added to kaolin. The APTT is sensitive to the effects of heparin on thrombin, factor Xa and IXa. Therapeutic levels are generally 1.5 to 2.5 times that of the control. High doses of heparin can affect the prothrombin time (PT), but to a lesser extent than its effect on the APTT.
Idraparinux is a more negatively charged derivative of fondaparinux. It binds antithrombin with higher affinity than fondaparinux. Thus, idraparinux has a terminal plasma half-life of around 80 hours, similar to that of thrombin. Consequently, it can be given subcutaneously once a week. However, idraparinux has been shown to produce dose-dependent bleeding. In a phase II trial, patients given a dose of 5 mg had a significantly higher risk of severe bleeding, but patients given a dose of 2.5 mg had less bleeding than patients receiving warfarin.1 Razaxaban is a synthetic, non-peptidic, orally active factor Xa inhibitor. It has been evaluated in a phase II trial of patients undergoing elective knee arthroplasty. However, the risk of bleeding in the razaxaban patient group was significantly high and the trial was prematurely stopped. Oral anticoagulants Warfarin sodium is a 4-hydroxycoumarin compound and is now the standard oral anticoagulant. Warfarin inhibits the synthesis of factors dependent on vitamin K (factors II, VII, IX, and X, protein C, protein S). Carboxylation of the glutamate groups in the inactive precursor proteins is required for the synthesis of active coagulation factors. This process is coupled with the oxidation and reduction of vitamin K. Warfarin inhibits epoxide reductase, the enzyme required for continual production of the reduced, biologically active form of vitamin K, and therefore prevents activation of precursors. Circulating factors are not affected by warfarin and its effects are delayed until previously synthesized factors are consumed. Factor VII decreases rapidly (during less than 24 hours). However, factor II has a longer halflife, which means that the therapeutic effects of warfarin may take 48–72 hours to develop. Warfarin is rapidly absorbed from the gastrointestinal tract. Approximately 97% of warfarin is bound to albumin, resulting in partly restricted diffusion across membranes. Warfarin is
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Comparison of low molecular weight heparins Product
Preparation
Mean molecular weight (range, Da)
Half-life (min)
Xa:IIa ratio
Bioavailability (%)
Enoxaparin Dalteparin Nadroparin Tinzaparin Ardeparin Reviparin Heparin
Benzylation, alkaline hydrolysis Nitrous oxide depolymerization Nitrous acid depolymerization Heparinase digestion Peroxidase cleavage Nitrous acid digestion –
4500 (3000–8000) 5000 (2000–9000) 4500 (2000–8000) 4500 (3000–6000) 6000 (2000–15,000) 4600 (3000–7000) 11400
129–180 119–139 132–162 111 200 180 60
2.7:1 2.0:1 3.2:1 1.9:1 2.0:1 1.9:1 1.0:1
90–92 87 98 90 92 90 30 (range 10–40)
Table 2
The most important adverse effect of heparin is bleeding, which is treated by stopping the drug and administering prota mine, a strong basic drug that forms an inactive complex with heparin. Thrombocytopenia can also occur, either at an early stage or with long-term treatment. HIT can cause venous and arterial thrombosis, a fall in platelets or skin lesions. Hyper sensitivity, osteoporosis, and alopecia have also been reported.
molecular weight heparinoid consisting of a mixture of heparan sulphate (84%), dermatan sulphate (12%) and small amounts of chondroitin sulphate (4%), and its antithrombotic activity is well established. Its pharmacological effect is exerted primarily by inhibiting factors Xa and IIa at a ratio greater than heparin, with a minimal effect on platelet function. Danaparoid exhibits low cross-reactivity with heparin-induced antibodies when compared with heparin or LMWHs, thereby making it an excellent choice for the management of HIT. It has excellent bioavailability following subcutaneous injection. Danaparoid has little effect on routine coagulation tests (APTT, PT and thrombin time). Patients with elevated serum creatinine should be monitored carefully. Danaparoid has been found to be effective in the treatment of HIT and is frequently used for this purpose. However, the information accompanying this drug does not mention its use for the treatment of this condition. ◆
Low molecular weight heparins: standard heparin can be fractionated either by chemical or enzymatic depolymerization to produce preparations with a lower molecular weight (4000–6000 Da) (Table 2). LMWHs produce their anticoagulant effect by binding and neutralizing factor Xa. They have minimal inhibitory effects on thrombin because compared with UFH, the smaller fragments of LMWHs are unable to bind AT-III and thrombin simultan eously. This bridging process is less critical for the inactivation of factor Xa. Different preparations vary in their ratio of anti-Xa to antithrombin activity. LMWHs are well absorbed when given subcutaneously and there is little binding to circulating and cellular proteins. Clearance occurs by the renal route and biological half-life is prolonged in patients with renal failure. LMWHs have a longer plasma halflife and a better bioavailability at low doses than UFH. They have a more predictable dose–response relationship than UFH, and the dose is calculated on the basis of patient weight. Monitoring is rarely required except in rare cases where dosage may be difficult (e.g. morbid obesity or renal failure). There is no effect on APTT, and it is necessary to measure plasma anti- Xa activity, ideally 4 hours after injection. Therapeutic benefits of LMWHs seem to be in the reduction of mortality in unstable angina, and in surgical thromboprophylaxis. There is still a risk of haemorrhage with LMWHs, but they have less of an effect on platelet aggregation, and a lower risk of thrombocytopenia than UFH. Protamine has little effect on reversing LMWHs because its main mechanism of action is in neutralizing antithrombin activity.
Reference 1 PERSIST Investigators: a novel long-acting synthetic factor Xa inhibitor (SanOrg34006) to replace warfarin for secondary prevention in deep vein thrombosis: a Phase II evaluation. J Thromb Haemost 2004; 2: 47–53. (Erratum in: Blood 2002; 100: 82a).
Further reading Agnelli G, Becattini C. New anticoagulants. Semi Thromb Hemost 2006; 32: 793–802. Bates SM, Weitz J. The status of new anticoagulants. Br J Haematol 2006; 134: 3–19. Hirsch J, Warkentin TE, Shaughnessy SG, et al. Heparin and low molecular weight heparin: mechanism of action, pharmacokinetics, dosing, monitoring, efficacy, and safety. Chest 2001; 119: 64S–94S. Kam PCA, Egan MK. Platelet glycoprotein IIb/IIIa antagonists: pharmacology and clinical developments. Anesthesiology 2002; 96: 1237–49. Vandermeulen E. Anaesthesia and new antithrombotic drugs. Curr Opin Anaesthesiol 2005; 18: 353–9. Weitz JI, Hirsch J. New anticoagulant drugs. Chest 2001; 119: 95S– 107S.
Danaparoid sodium: danaparoid sodium (Orgaran) is a heparinoid glycosaminoglycuronan antithrombotic agent approved in the UK for the prophylaxis of postoperative deep vein throm bosis, which may lead to pulmonary embolism in patients undergoing elective hip replacement surgery. Danaparoid is a low
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