CHAPTER 134 FIBRINOLYTIC THERAPY
CHAPTER 134 FIBRINOLYTIC THERAPY
Currently Available Agents
Monitoring Thrombolytic Therapy
Clinical Thrombolytic Regimens
Shortcomings of Thrombolytic Therapy
Adjunctive Therapy for Fibrinolysis
Fibrinolytic therapy is designed to facilitate thrombolysis and thus decrease the ischemic damage produced by thrombotic events. Currently available therapeutic agents all essentially impart proteolytic activity to the inactive plasma zymogen plasminogen. These agents either bind to and induce a conformational change in plasminogen, imparting plasmin-like protease activity to the molecule, or proteolytically cleave plasminogen into plasmin directly. Streptokinase and staphylokinase are bacterial products that can now be synthesized by recombinant DNA technology and impart proteolytic activity to plasminogen. Tissue-type plasminogen activator and urokinase-type plasminogen activators are endogenous plasminogen activators that directly activate plasminogen by converting it into plasmin. Newer agents, including reteplase, TNK-t-PA, and bat-PA, have been developed to enhance fibrinolytic specificity and the rate of lysis. This chapter will focus on the pharmacology of currently available fibrinolytic agents, provide an overview of their clinical applications in specific thrombotic disorders, and review the use of adjunctive antithrombotic therapies with thrombolytic agents in these disorders.
Acronyms and abbreviations that appear in this chapter include: APSAC, anisoylated plasminogen-streptokinase activator complex; CRE, cyclic AMP-responsive element; HMW-tcu-PA, high molecular weight two-chain u-PA; LMW-tcu-PA, low molecular weight two-chain u-PA; PAI, plasmin activator inhibitor; PPACK, d-Phe-Pro-Arg-chloromethylketone; scu-PA, single-chain u-PA; TNK-t-PA, T103N,N117Q, KHRR(296–299)AAAA; t-PA, tissue-type plasminogen activator; t-PA-DFEK1, reteplase; u-PA, urokinase-type plasminogen activator; u-PAR, u-PA receptor.
CURRENTLY AVAILABLE AGENTS
Streptokinase was first isolated in 19331 and subsequently demonstrated to possess fibrinolytic activity in vivo in animals and humans in the 1940s and 1950s.2,3 A product of Lance field group C b-hemolytic streptococci, streptokinase is a single-chain polypeptide with a molecular mass of 47 kDa. Despite its amino-terminal sequence homology to trypsinlike serine proteases, streptokinase is devoid of an active site serine residue and is thus incapable of proteolytic or amidolytic activity.4
Streptokinase itself has no enzymatic activity; instead plasmin-like activity is conferred upon the streptokinase-plasminogen complex without plasminogen proteolysis (see Chap. 116). Figure 134-1 illustrates the sequence of reactions that accounts for the activity and metabolism of streptokinase and the streptokinase-plasminogen complex. Stoichiometric amounts of streptokinase combine with plasminogen to form a streptokinase-plasminogen complex that possesses plasmin-like activity. This complex then reacts with uncomplexed plasminogen to convert it to plasmin directly. In addition, the streptokinase-plasminogen complex undergoes proteolytic cleavage in solution at the Arg560-Val561 and the Lys77-Lys78 bonds of plasminogen. The complex is then itself subjected to progressive degradation into smaller fragments, resulting in the loss of plasminogen activator activity.5,6 and 7 The activation of plasminogen by streptokinase-plasminogen obeys Michaelis-Menten kinetics8 with a Km of 1.4 µM and a kcat of 21.8 s–1. The streptokinase-plasminogen complex activates partially cleaved plasminogen (Lys-plasminogen) approximately fivefold more effectively on a molar basis than intact plasminogen (Glu-plasminogen). The relative catalytic efficiency of the streptokinase-plasminogen complex depends upon whether Glu-plasminogen or Lys-plasminogen is in the complex.
FIGURE 134-1 Sequential reactions of streptokinase and plasminogen.
Proteolytic cleavage of both streptokinase and plasminogen in the streptokinase-plasminogen complex leads to the formation of the streptokinase-plasmin complex. The streptokinase-plasmin complex also forms when streptokinase is exposed to plasmin, and it also obeys Michaelis-Menten kinetics8 with a Km of 1.1 µM and a kcat of 16.6 s–1. As with streptokinase-plasminogen, the streptokinase-plasmin complex is approximately fivefold more efficient with Lys-plasminogen as a substrate than with Glu-plasminogen.
There are three important comparative differences between streptokinase-plasmin and free plasmin.9,10 and 11 First, the streptokinase-plasmin complex has twice the catalytic efficiency of plasmin. Second, streptokinase-plasmin can directly convert plasminogen to plasmin, while free plasmin is only efficient at converting Glu-plasminogen to Lys-plasminogen. Third, the important plasmin inhibitors, a2-antiplasmin and a2-macroglobulin, are comparatively poor inhibitors of the complex.
Fibrinogen, fibrin, and the fibrin(ogen) degradation products, fragments D and E, enhance the activity of streptokinase significantly, with an overall order of efficacy of fibrin>fibrinogen>fragment D>fragment E.9,10 Fibrin increases the catalytic efficiency of streptokinase-plasminogen-mediated activation of plasminogen 6.5-fold, while fibrinogen only induces a twofold increase in catalytic efficiency.
Pharmacology When administered in vivo to an individual bearing a thrombus, streptokinase binds to circulating plasminogen to form the streptokinase-plasminogen complex. Conversion of streptokinase-plasminogen to streptokinase-plasmin then ensues by one complex cleaving plasminogen in a second complex or by plasmin, produced by the complex acting on plasminogen, complexing with streptokinase. Fibrin binding of the streptokinase-plasmin(ogen) complex is mediated by the “kringle” domains of plasmin,5 in particular kringles 1 and 4. Owing to the lack of inhibition of streptokinase-plasmin by a2-antiplasmin,11 this complex can activate clot-bound plasminogen directly. The subsequent generation of plasmin achieves two opposing ends. First, local degradation of fibrin leads to the elaboration of soluble fibrin(ogen) degradation products in the vicinity of the thrombus, which act to augment thrombolysis by enhancing streptokinase-plasmin activity. Second, the degradation of the streptokinase-plasmin(ogen) complex by plasmin leads to progressive attenuation of plasmin activity. In addition, circulating streptokinase-plasmin(ogen) acts upon circulating plasminogen, leading to systemic plasminemia, the proteolytic cleavage of fibrinogen into its degradation products, and the genesis of a systemic lytic state. Profound plasminemia may also lead to reduced concentrations of plasminogen within the thrombus by reequilibration, and this redistribution to plasma may attenuate rates of lysis within the thrombus (“plasminogen steal”).
Pharmacokinetic analysis shows that streptokinase is eliminated from plasma with a monophasic half-life of 18 to 30 min.12 The biologic half-life (i.e., the half-life of the lytic effects of the plasminogen activator), by contrast, is more prolonged, ranging from 82 to 184 min.13,14
Clinical Application Since it was first used over 40 years ago, streptokinase has been widely employed in the treatment of a variety of thrombotic disorders (Table 134-1),15,16,17,18,19,20 and 21 including venous thrombosis, pulmonary embolism, peripheral arterial thrombosis, cerebral embolism, thrombotic ureteral obstruction, and acute myocardial infarction. The agent has proved to be effective for lysing culprit thrombi in the majority of these disorders and to improve clinical outcomes in many. Its overall safety is somewhat limited by bleeding complications and, owing to its bacterial origin, allergic reactions including fever, hypotension, urticarial eruption, and bronchospasm. The incidence of these side effects is relatively low, with hemorrhagic complications occurring in approximately 5 percent of patients, 20 percent of which are major bleeding events; allergic reactions occurring in <1 percent of patients; and hypotensive responses requiring drug therapy occurring in 6 to 8 percent of patients. Owing to its bacterial nature, streptokinase induces antibody production in most individuals, and this may be important in patients with recent streptococcal infection. Within 3 or 4 days of administration, the level of neutralizing antibodies becomes sufficiently great to inactivate standard therapeutic doses of streptokinase. These neutralizing antibody titers persist in up to 80 percent of individuals up to 1 year following therapy and in approximately 50 percent of individuals at 2 to 4 years. For this reason, reuse of streptokinase has limited application. If reuse is contemplated, neutralizing antibody titers should first be quantified in order to ascertain the appropriate dosing strategy.
TABLE 134-1 DISORDERS TREATED WITH FIBRINOLYTIC THERAPY15
ANISOYLATED PLASMINOGEN-STREPTOKINASE ACTIVATOR COMPLEX
Biochemistry The pronounced catalytic efficiency of the streptokinase-plasmin(ogen) complex is somewhat limited by its rapid inactivation and clearance from the blood circulation. In an effort to overcome these shortcomings, a chemically modified derivative of streptokinase was developed in which the activator is noncovalently associated with plasminogen and the active site of plasminogen is, in turn, protected by covalent modification with a p-anisoyl group22 (Fig. 134-2). This compound, termed APSAC (anisoylated plasminogen-streptokinase activator complex), binds fibrin through the plasminogen kringle domains and has a significantly prolonged half-life (94 min).
FIGURE 134-2 Structure of APSAC (SK, streptokinase; PLG, plasminogen).
The combined molecular mass of plasminogen and streptokinase in APSAC is 131 kDa. Specific acylation of the active site Ser740 in plasminogen is achieved by the use of p-amidinophenyl-p’-anisate hydrochloride, a so-called inverse acylating agent. In the process of acylation the cationic amidino group forms an ionic bond with Asp734 at the catalytic center of plasminogen, positioning the p-anisoyl group near the active site serine, Ser740. An acyl transfer reaction then leads to the p-anisoylation of the catalytic center of the enzyme.
Pharmacology Several important pharmacological properties are conferred on streptokinase-plasminogen as a result of p-anisoylation. APSAC can be administered rapidly as a single bolus without producing significant hypotension23; by contrast, bolus administration of streptokinase frequently leads to hypotension.24 In addition, compared with streptokinase, APSAC has greater plasma stability, possesses a longer circulating half-life, and has greater lytic potency.
After bolus administration, deacylation of APSAC commences immediately, leading to the generation of the active streptokinase-plasminogen complex. The rate of deacylation follows first-order kinetics; in whole blood, the half-life for the process is 104 min.25 The plasma half-life of APSAC is approximately 90 min, significantly longer than that of streptokinase.26 The precise mechanism by which the half-life is prolonged is unclear but may be a consequence of slower hepatic clearance, less neutralization by plasma inhibitors, or relative resistance to proteolytic degradation in plasma.
Clinical Applications APSAC has been shown to be efficacious in acute myocardial infarction and is probably equivalent to streptokinase (see Table 134-1). Its specific benefit is ease of administration; however, hemorrhagic complications appear to be greater than with streptokinase or tissue-type plasminogen activator27 (discussed under “Endogenous Agents” below). Bolus administration of this compound has not proven to be a critical element in the choice of agent at the current time.
Side effects of APSAC are similar to those for streptokinase, with hemorrhagic complications being of greatest concern. Again, the potential for allergic reactions remains an issue, with urticarial eruption, bronchospasm, erythema, anaphylaxis, and angioedema reported. In addition, neutralizing antibodies have been detected in patients treated with this agent.
UROKINASE-TYPE PLASMINOGEN ACTIVATORS
Molecular Biology and Biochemistry The human urokinase-type plasminogen activator (u-PA) gene is located on chromosome 10, spans 6.4 kb, and contains 11 exons.28,29 and 30 Transcription factors AP1 and AP2 may be involved in promoter modulation of u-PA expression.30 Functional analysis of the promoter region of the human u-PA gene indicates the presence of a potential enhancer element approximately 2 kb upstream of the transcription start site as well as potential negative regulatory elements approximately 1.5 kb upstream.31,32
The mature gene product, single-chain u-PA (scu-PA), is a 54-kDa glycoprotein28 isolated from urine, plasma, and conditioned cell culture media33,34 and is synthesized in endothelial cells. scu-PA is converted to the high molecular weight two-chain derivative (HMW-tcu-PA) by limited hydrolysis of the Lys158-Ile159 bond by plasmin and kallikrein.35 The resulting amino-terminal light chain contains 158 amino acid residues, and the carboxy-terminal heavy chain 254 amino acid residues; light and heavy chains are linked by a single disulfide bond (Cys148-Cys279).36,37 The serine protease active site is located in the heavy chain.29 The molecule also contains an epidermal growth factor–like domain and a kringle-type domain, both encoded by exons III to VI.28
HMW-tcu-PA can be converted to a low molecular weight form (LMW-tcu-PA) in which the kringle domain is removed by cleavage at Lys135-Lys136. Both the HMW-tcu-PA and this smaller 33-kDa cleavage product are used clinically.
The enzymology of u-PA is complex. scu-PA has little intrinsic plasminogen activator activity38,39; according to the best estimates using plasmin-resistant scu-PA mutants, the catalytic efficiency is approximately 0.00015 µM–1s–1, representing approximately 0.1 to 0.5 percent that of HMW-tcu-PA.40 scu-PA is also resistant to active site inhibitors of HMW-tcu-PA. Thus, scu-PA meets the criteria for defining it as a true zymogen.
Plasminogen activation by HMW-tcu-PA follows classical Michaelis-Menten kinetics. With Glu-plasminogen as a substrate, the Km is 50 µM and the kcat is 1.0 s–1.41 With Lys-plasminogen, the catalytic efficiency has been variably reported to be increased three- to tenfold over that for Glu-plasminogen.42 Fibrin, fibrinogen, and fibrinogen degradation fragments D and E, as well as e-aminocarboxylic acids such as e-aminocaproic acid or tranexamic acid, enhance the catalytic efficiency of activation of Glu-plasminogen by approximately fivefold.39,43,44 and 45 This improved catalytic efficiency is a consequence of a conformational change in Glu-plasminogen induced by the binding of these agents to lysine-binding sites in the zymogen, which leads to Glu-plasminogen adopting a structure similar to that of the Lys-plasminogen. As expected from this mechanism, the rate of activation of Lys-plasminogen is not affected by fibrin(ogen) or its degradation products. Importantly, scu-PA and HMW-tcu-PA do not bind to fibrin; hence, the fibrin-specificity of HMW-tcu-PA is likely to be a consequence of the enhanced catalytic efficiency of activation of fibrin-bound Glu-plasminogen that results from this fibrin-induced confor-mational change in the zymogen.45,46 and 47 The cell-surface receptor for u-PA, u-PAR, serves to enhance the catalytic efficiency of u-PA up to twentyfold.48
Pharmacology The pharmacokinetics of u-PA is best characterized by a two-compartment model involving intravascular and interstitial compartments. In humans, scu-PA and HMW-tcu-PA are cleared differently, with initial and terminal half-lives of 69 and 27 min, respectively, for scu-PA and of 12 and 61 min for HMW-tcu-PA.49 Studies with active-site-blocked enzymes suggest that the plasma half-life is a property of the protein itself and not a consequence of inactivation by plasma inhibitors50; clearance is also not dependent on glycosylation.
Clinical Applications The u-PAs have been used successfully to treat a myriad of thrombotic disorders (see Table 134-1). These include venous thrombosis, pulmonary embolism, peripheral arterial thrombosis, cerebrovascular thrombosis, unstable angina, acute myocardial infarction, and acute reocclusion following percutaneous transluminal angioplasty.
TISSUE-TYPE PLASMINOGEN ACTIVATOR
Molecular Biology and Biochemistry Tissue-type plasminogen activator (t-PA) is a 68-kDa serine protease, the cDNA for which was cloned and sequenced in 1983.51,52 The human t-PA gene is on chromosome 8,53,54 and approximately 36 kb of the genomic sequence have been determined.55 The t-PA gene is complex and consists of 14 exons, with the intron-exon organization suggestive of the principle of exon shuffling in the evolution and assembly of the complete molecule. The proximal promoter sequences contain the typical TATA and CAAT sequences, and potential recognition sites for transcription factors have been identified [e.g., cyclic AMP-responsive element (CRE), AP1, NF1, SP1, and AP2], which may function in the regulation of gene expression.56,57
t-PA is synthesized by endothelial cells as a single-chain polypeptide.58,59 and 60 The mature protein contains a single free cysteine, Cys83, and 17 disulfide bonds. Plasmin, kallikrein, and factor Xa are all able to cleave Arg275-Ile276 to produce the two-chain form of t-PA61; when compared with its single-chain precursor, the two-chain form has a lower catalytic efficiency in the absence of fibrin but an equivalent catalytic efficiency in the presence of fibrin.62
There are five distinct domains in the t-PA molecule (Fig. 134-3); from amino to carboxy terminus they include a fibronectin finger-like domain, an epidermal growth factor–like domain, two kringle-type domains, and a serine protease domain. Specific domains account for distinctive properties of the parent molecule, including fibrin binding, plasma clearance, cell surface binding, and fibrin stimulation of plasminogen activation; knowledge of the structure-function relationships among these domains has been provided by analyses of deletion and inclusion mutants, as well as chimeric molecules.
FIGURE 134-3 Structure of t-PA (F, fibronectin finger-like domain; E, epidermal growth factor–like domain; K1, kringle 1 domain; K2, kringle 2 domain; S, serine protease domain).
Plasminogen activation by the single-chain form of t-PA follows Michaelis-Menten kinetics, with a Km of approximately 65 µM and a kcat of 0.05 s–1.63 Fibrin increases the catalytic efficiency of the molecule several hundredfold and appears to be a consequence of direct binding by t-PA (Km, 0.1 µM) to the E domain of fibrinogen through its fibronectin finger-like domain and the second kringle-type domain,64,65 as well as to higher-ordered polymeric structures in the mature fibrin clot.66 Fibrin-induced enhancement of t-PA activity appears to be a consequence of the induction of a conformational change in t-PA, in plasminogen, or both that promotes the interaction of t-PA with plasminogen on the fibrin surface.67 Surface receptors on endothelial cells,68 monocytes,69 and platelets70 also serve to enhance the catalytic efficiency of t-PA (see Chap. 116).
Pharmacology Single- and two-chain t-PAs are cleared from plasma in a manner that is best approximated by a two-compartment model.71 Single-chain t-PA has an initial half-life of 4.1 min and a terminal half-life of 46 min, while the two-chain form has an initial half-life of 5.2 min and a terminal half-life of 46 min. Clearance of single- and two-chain t-PA occurs in the liver, where specific hepatic receptors mediate the uptake of the molecule from plasma. The dependence of t-PA clearance on hepatic blood flow and function emphasizes the need for cautious use of this plasminogen activator in patients with intrinsic liver disease or reduced hepatic blood flow, such as in patients with significant congestive heart failure.
Clinical Applications t-PA has been used successfully in a variety of thrombotic disorders (see Table 134-1), including venous thrombosis, pulmonary embolism, unstable angina, acute myocardial infarction, cerebrovascular thrombosis, and thrombosed artificial cardiac valves. The relative fibrin specificity of t-PA compared with other currently available plasminogen activators may not be as important as once believed. This point is particularly relevant in the treatment of thrombotic arterial occlusive disorders requiring rapid lysis to minimize tissue injury; typically, the dose of t-PA required for adequate rapidity of lysis is sufficiently high that systemic plasminemia and a lytic state are achieved. The lytic state produced by t-PA is, however, rarely as pronounced as it is with streptokinase or two-chain urokinase.
NEWER PLASMINOGEN ACTIVATORS
Staphylokinase Staphylokinase, a 15.5-kDa protein produced by Staphylococcus aureus, was shown to have profibrinolytic properties more than 40 years ago.72 Its mechanism of action is similar to that of streptokinase73 in that staphylokinase also forms a 1:1 stoichiometric complex with plasminogen which, after conversion, forms plasmin.74 In contrast with streptokinase, however, it can be inhibited by a2-antiplasmin, and fibrin binding prevents inactivation by this protease inhibitor.74 In addition, staphylokinase is much more effective at lysing platelet-rich or retracted thrombi than is streptokinase.75
Plasminogen activation by staphylokinase follows Michaelis-Menten kinetics. With Glu-plasminogen as the substrate, the Km is 7 µM and the kcat is 1.5 s–1. Fibrin binding enhances the activity of this bacterial product approximately fourfold, primarily as a consequence of binding to plasminogen’s kringle domains.
Staphylokinase is a highly efficient fibrinolytic agent, producing high rates of clot lysis without significantly influencing plasma fibrinogen, plasminogen, or a2-antiplasmin levels. Owing to its bacterial origin, however, staphylokinase is antigenic when used in humans with neutralizing antibodies detectable 2 weeks following therapy.76 Recent data suggest that modification of two of its three immunodominant epitopes can attenuate antigenicity without altering its potency.77 In a preliminary clinical trial of patients with acute myocardial infarction,76 10 or 20 mg of staphylokinase administered over 30 min was shown to be equally efficacious and safe as weight-adjusted t-PA at restoring coronary vascular patency without depleting fibrinogen.
Mutant Tissue-Type Plasminogen Activators Numerous attempts have been made to improve the efficacy and safety of plasminogen activators by altering the structure of the enzyme using recombinant DNA techniques. One deletion mutant, t-PADDFEK1, or reteplase, contains only the kringle 2 and serine protease domains and demonstrates fibrin binding and plasminogen activation,78 with a prolonged serum half-life of 58 min.79 The efficacy and safety of this recently approved agent was best demonstrated in a recent large trial in which it was found to restore vessel patency more effectively than t-PA in patients with acute myocardial infarction; however, this benefit did not translate into a mortality benefit at 30 days, with both agents producing equivalent outcomes for this endpoint.80
Another t-PA mutant, TNK-t-PA, was designed to improve half-life, fibrin specificity, and resistance to inhibition by plasminogen activator inhibitor type I. Its name is derived from threonine and asparagine substitutions at positions 103 and 117, respectively, and four alanine substitutions at positions 296–299. Results of an early clinical trial confirm that the half-life of TNK-t-PA is prolonged (17 min), with equivalent efficacy as t-PA at restoring vascular patency without fibrinogenolysis.81
Bat-PA The vampire bat Desmodus rotundus secretes plasminogen activators in its saliva; the cDNAs of four of these have been cloned and sequenced, and recombinant forms have been expressed.82 One isoform of this group of enzymes has been tested in animal models and found to be somewhat more potent than t-PA and also more fibrin-selective.83
Other Chemical Conjugates, Mutants, and Chimeras Given the shortcomings of currently available thrombolytic agents,84,85 many attempts have been made to engineer an optimal agent using recombinant DNA technologies. Owing to an inability to define the “ideal” plasminogen activator (e.g., short versus long half-life, fibrin-selective versus relatively nonselective), none of the molecules designed thus far has shown significant improvement in hard clinical endpoints over “wild type” molecules.86,87
MONITORING THROMBOLYTIC THERAPY
In the early trials of thrombolytic therapy, investigators were careful to monitor a variety of coagulation tests as markers of thrombolytic efficacy and as possible predictors of clinical efficacy.88 Analysis of these data, provided by many trials over the past 10 years, fails to show any distinct utility of these measurements. Prothrombin time, activated partial thromboplastin time, and thrombin time all increase with fibrinolytic therapy; fibrinogen, plasminogen, and a2-antiplasmin are all consumed; and fibrin(ogen) degradation products are produced. None of these parameters has been shown to correlate with clinical efficacy, and only reductions in fibrinogen below 100 mg/dl (SI units) appear to be associated with an increased hemorrhagic risk. Thus, routine, repeated monitoring of these parameters is not indicated in the typical patient receiving thrombolytic therapy. One may, however, consider selected measurements to identify patients with undiagnosed hemorrhagic diatheses prior to administration of the fibrinolytic agent; to identify patients who fail to lyse as a consequence of (antibody-mediated) neutralization of the agent (seen infrequently with streptokinase use); or to monitor the adequacy of heparin therapy used adjunctively following the administration of the plasminogen activator (see “Clinical Thrombolytic Regimens”). To assess the adequacy of therapy, evidence for the elaboration of a systemic lytic state should be obtained, such as a reduction in fibrinogen, production of fibrin(ogen) degradation products, or depletion of plasminogen or a2-antiplasmin.
CLINICAL THROMBOLYTIC REGIMENS
Although a variety of regimens have been used to administer thrombolytic therapy over the past 30 years, abundant data acquired through large clinical trials clearly show the efficacy and relative safety of specific dosing schedules in patients with acute myocardial infarction, deep venous thrombosis, and pulmonary embolism. These commonly accepted regimens are listed in Table 134-2. Fibrinolytic therapy has proven to be quite efficacious in a variety of arterial and venous thrombotic disorders as indicated in Table 134-1, and therapy has been shown to be particularly beneficial in patients with acute myocardial infarction,18 peripheral arterial occlusion,20 acute cerebrovascular thrombosis,21 deep venous thrombosis, 16 and pulmonary embolism.17
TABLE 134-2 THERAPEUTIC THROMBOLYTIC REGIMENS
Evidence from numerous trials over the past 15 years clearly indicates that fibrinolytic therapy with t-PA or streptokinase improves mortality and residual left ventricular function; this benefit is time-limited, and the earlier one achieves recanalization of the occluded infarct-related artery the greater the benefit.18 An analysis of the largest trials of thrombolytic therapy indicate that approximately 12 lives are saved for every 1000 patients treated for acute myocardial infarction. Current practice involves the timely administration of t-PA or streptokinase to patients with acute myocardial infarction who present within 6 h of the onset of chest pain; those presenting within 6 to 12 h of the onset of the infarction may also receive some benefit from fibrinolytic therapy, but the advantages are somewhat outweighed by the disadvantages that are a consequence of the increased hemorrhagic risk of therapy in these patients. Absolute contraindications to fibrinolytic therapy include active internal bleeding, hemorrhagic stroke, nonhemorrhagic stroke within the past year, intracranial neoplasm, and suspected aortic dissection; relative contraindications to therapy include prolonged cardiopulmonary resuscitation, severe hypertension, trauma within the past 4 weeks, surgery within the past 3 weeks, a history of bleeding diathesis, pregnancy, and active peptic ulcer disease. As will be discussed in more detail in “Adjunctive Therapy for Fibrinolysis,” it is important to recognize that when treating patients with acute myocardial infarction the adjunctive use of aspirin (325 mg chewed immediately, then daily thereafter) and of full-dose intravenous heparin (administered immediately with the infusion of t-PA or begun 2 to 4 h following infusion of streptokinase) are strongly advised. While it is clearly the case that intravenous heparin increases the rate of bleeding complications approximately twofold, the efficacy of this agent in maintaining patency of an occluded vessel is believed to outweigh the hemorrhagic risk.
u-PA has been shown to be effective in restoring patency in thrombosed peripheral arteries and does so in almost three-quarters of patients.20 This restoration of blood flow is accompanied by a reduction in rates of both amputation and surgical revascularization.
Most recently, the benefits of fibrinolytic therapy with t-PA for patients with cerebrovascular thrombosis or embolism have been demonstrated.21 Patients treated within 3 h of the onset of stroke with t-PA were shown to have a clear improvement in 3-month neurological status and disability compared to those treated with placebo. This benefit is tempered somewhat by the risk of intracerebral hemorrhage, although in the final analysis the benefits outweighed the risks in this population. In addition, the benefits persisted up to 24 months following therapy. Of course, patients with a hemorrhagic stroke must be excluded at the time of presentation, mandating rapid assessment by computed tomography or magnetic resonance imaging in all patients considered for therapy.
The benefits of fibrinolytic therapy in patients with venous thromboembolic disease is less dramatic than for arterial disorders. While treatment with streptokinase, u-PA, or t-PA can lyse deep venous thrombi or pulmonary emboli quite effectively, the clinical benefits over conventional therapy with heparin and warfarin have been marginal.16,17 Only in patients with pulmonary emboli sufficiently massive to evoke right ventricular dysfunction should fibrinolytic therapy be routinely administered; however, even among this group of patients no convincing data are yet available to support an overall benefit in mortality or residual cardiopulmonary morbidity.
SHORTCOMINGS OF THROMBOLYTIC THERAPY
There are three principal shortcomings of thrombolytic therapy as it is currently used: hemorrhage, delays in times to lysis, and reocclusion. The mechanism(s) underlying these adverse outcomes of therapy are complex and require an understanding of the diverse biochemical consequences of pharmacological plasminogen activation.89,90 and 91 Plasmin has the well-known antithrombotic effects of (1) direct fibrinolysis, (2) fibrinogenolysis leading to a reduction in blood viscosity and improvement in blood flow, (3) production of fibrin(ogen) degradation products which themselves are antithrombotic by virtue of their ability to inhibit fibrin polymerization and platelet aggregation, and (4) direct inhibition of platelet aggregation. These effects not only promote thrombolysis and vascular patency but also are at least in part responsible for the hemorrhagic complications of thrombolytic therapy. However, the actual risk of serious hemorrhage, even with the concurrent use of the antithrombotic agents described below, is in the range of 1 percent,92,93 an incidence that must be critically compared with the serious outcomes that may result from withholding such therapy in selected patients.
Paradoxical prothrombotic effects of plasmin have also been appreciated, and these may account at least in part for the failure of thrombolytic therapy to achieve rapid lysis and prevent reocclusion.90,91 Plasmin at relatively high concentrations (achievable during thrombolytic therapy) can activate platelets directly.94 In addition, plasmin can lead to factor X activation and prothrombinase activity.95,96 These two important effects of therapeutic thrombolysis, coupled with the continued exposure of the prothrombotic components of the fissured atheroma to flowing blood once thrombus is lysed, probably account for the adverse thrombotic events accompanying thrombolysis. In the current thrombolytic era these complications exceed hemorrhagic complications; for example, up to 25 percent reocclusion rates are noted for arterial thrombotic disorders following initially successful thrombolysis with t-PA.97
ADJUNCTIVE THERAPY FOR FIBRINOLYSIS
The complications of hemorrhage, failure to lyse, and acute reocclusion following successful lysis exemplify the complexity of mechanisms active during pharmacological fibrinolysis and provide a basis for the current approaches to combined, adjunctive therapy. Hemorrhage is prevented by limiting infusions of plasminogen activators to relatively brief periods (minutes for APSAC and up to 3 h for t-PA). By comparison with older lengthy infusions of streptokinase or urokinase for deep venous thrombosis or pulmonary embolism (24- to 72-h infusions), brief infusions are equally efficacious and associated with far less bleeding. Minimizing invasive procedures also greatly reduces hemorrhagic complications. When bleeding does occur, cessation of infusion of the plasminogen activator and mechanical compression (when possible) are the primary treatment options. In extreme cases the use of fresh-frozen plasma, platelet transfusions, or e-aminocaproic acid should be considered. Recombinant plasminogen activator inhibitor (PAI)-1 and desmopressin may, in the future, also have a role in the treatment of hemorrhage in selected patients.
The use of antiplatelet agents such as aspirin, antiplatelet prostaglandins, or glycoprotein IIb/IIIa antagonists and of thrombin inhibitors such as heparin or hirudin greatly attenuates the thrombotic complications of thrombolytic therapy. Their combined use has been associated with increases in the vascular patency rates of thrombolytic regimens for arterial disorders in conjunction with a reduction in markers of thrombin activity (such as fibrinopeptide A) and of platelet activation. Conventional agents include aspirin and heparin. More recently, platelet glycoprotein IIb/IIIa antagonists, chief among which is the monoclonal antibody abciximab, have been shown to be superb adjunctive antiplatelet agents for patients with acute myocardial infarction treated with fibrinolytic therapy101 or for patients with acute coronary syndromes undergoing percutaneous transluminal coronary angioplasty.102 A host of other nonantibody glycoprotein IIb/IIIa antagonists (e.g., tirofiban, lamifiban, xemilofiban) are currently undergoing clinical evaluation and have proven to be useful adjunctive therapies in preliminary trials. However, owing to their short half-lives compared with abciximab, rebound platelet activation may occur at trough concentrations of these agents, potentially leading to recurrent thrombosis. Other experimental agents that have yet to be approved as adjunctive therapies but do provide synergism with plasminogen activators in the restoration and maintenance of vascular patency in animal models and early human trials include iloprost, prostaglandin E1, thromboxane receptor antagonist and thromboxane synthase inhibitors, serotonin receptor antagonists, argatroban, activated recombinant protein C, low molecular weight heparins (e.g., enoxaparin, fraxiparin, dalteparin), D-Phe-Pro-Arg-chloromethylketone (PPACK), and the direct thrombin inhibitors hirudin and bivalirudin (Hirulog). Inasmuch as these agents can provide synergistic antithrombotic effects, they can, as well, evoke synergistic hemorrhagic complications. Clearly, in each case the antithrombotic benefit of these adjunctive agents must be weighed carefully against the hemorrhagic risk. Table 134-398,99,100,101,102 and 103 contains a listing of the currently used adjunctive therapies and newer, experimental therapies currently undergoing clinical evaluation.
TABLE 134-3 ADJUNCTIVE ANTITHROMBOTIC AGENTS USED WITH FIBRINOLYTICS
Tillett WS, Garner RL: The fibrinolytic activity of hemolytic streptococci. J Exp Med 58:483, 1933.
Tillett WS, Sherry S: The effect in patients of streptococcal fibrinolysis (streptokinase) and streptococcal deoxyribonuclease on fibrinous, purulent and sanguinous pleural exudations. J Clin Invest 38:1627, 1949.
Fletcher AP, Sherry S, Alkjaersig N, et al: The maintenance of a sustained thrombolytic state in man: II. Clinical observations on patients with myocardial infarction and other thromboembolic disorders. J Clin Invest 38:1111, 1959.
Jackson KW, Tang J: Complete amino acid sequence of streptokinase and its homology with serine proteases. Biochemistry 21:6620, 1982.
Reddy KN, Markus G: Mechanism of activation of human plasminogen by streptokinase. Presence of an active center in streptokinase-plasminogen complex. J Biol Chem 247:1683, 1972.
Summaria L, Wohl RC, Boreisha IG, Robbins KC: A virgin enzyme derived from human plasminogen. Specific cleavage of the arginyl-560 valyl peptide bond in the diisopropoxyphosphinyl virgin enzyme by plasminogen activators. Biochemistry 21:2056, 1982.
Siefring GE Jr, Castellino FJ: Interaction of streptokinase with plasminogen. Isolation and characterization of a streptokinase degradation product. J Biol Chem 251:3913, 1976.
Wohl RC, Summaria L, Arzadon L, Robbins KC: Steady state kinetics of activation of human and bovine plasminogens by streptokinase and its equimolar complexes with various activated forms of human plasminogen. J Biol Chem 253:1402, 1978.
Chibber BA, Morris JP, Castellino FJ: Effects of human fibrinogen and its cleavage products on activation of human plasminogen by streptokinase. Biochemistry 24:3429, 1985.
Takada A, Takada Y: Kinetic analyses of potentiation of plasminogen activation by streptokinase in the presence of fibrin or its degradation products. Haemostasis 17:1, 1987.
Wiman B: On the reaction of plasmin or the plasmin-streptokinase complex with aprotinin or a-antiplasmin. Thromb Res 17:143, 1980.
Mentzer RL, Budzynski AZ, Sherry S: High-dose brief-duration intravenous infusion of streptokinase in acute myocardial infarction: description of effects in circulation. Am J Cardiol 57:1220, 1986.
Grierson DS, Bjornsson TD: Pharmacokinetics of streptokinase in patients based on amidolytic activator complex activity. Clin Pharmacol Ther 41:304, 1987.
Col JJ, Col-De Beys CM, Renkin JP, Lavenne-Pardonge EM, Bachy JL, Moriau MH: Pharmacokinetics, thrombolytic efficacy and hemorrhagic risk of different streptokinase regimens in heparin-treated acute myocardial infarction. Am J Cardiol 63:1185, 1989.
Benedict CR, Mueller S, Anderson HV, Willerson JT: Thrombolytic therapy: a state of the art review. Hosp Pract 27:61, 1992.
Hyers TM, Hull RD, Weg JG: Antithrombotic therapy for venous thromboembolic disease. Chest 102:408S, 1992.
Goldhaber SZ: Evolving concepts in thrombolytic therapy for pulmonary embolism. Chest 101(suppl 4):183S, 1992.
Collins R, Peto B, Baigent C, Sleight P: Drug therapy: aspirin, heparin, and fibrinolytic therapy in suspected acute myocardial infarction. N Engl J Med 336:846, 1997.
Waters D, Lam J, Theroux P: Newer concepts in the treatment of unstable angina. Am J Cardiol 68:34C, 1991.
Clagett GP, Graor RA, Salzman EW: Antithrombotic therapy for peripheral arterial occlusive disease. Chest 102(suppl 4):516S, 1992.
Anonymous: Tissue plasminogen activator for acute ischemic stroke. The National Institute of Neurological Disorders and Stroke rt-PA Study Group. N Engl J Med 333:1581, 1995.
Smith RA, Dupe RJ, English PD, Green J: Fibrinolysis with acyl-enzymes: a new approach to thrombolytic therapy. Nature 290:505, 1981.
Green J, Dupe RJ, Smith RAG, Harris GS, English PD: Comparison of the hypotensive effects of streptokinase (human)-plasmin activator complex and BRL 26921 (p-anisoylated streptokinase-plasminogen activator complex) in the dog after high-dose bolus administration. Thromb Res 36:29, 1984.
Ferres H: Preclinical pharmacological evaluation of anisoylated plasminogen streptokinase activator complex. Drugs 33(suppl 3):33, 1987.
Ferres H, Hibbs M, Smith RA: Deacylation studies in vitro on anisoylated plasminogen streptokinase activator complex. Drugs 33(suppl 3):80, 1987.
Gemmill JD, Hogg KJ, Rae AP, et al: A comparison of the pharmacokinetic properties of streptokinase and anistreplase in acute myocardial infarction. Br J Clin Pharmacol 31:143, 1988.
O’Connor CM, Meese R, Carney R, et al: A randomized trial of intravenous heparin in conjunction with antistreplase (anisoylated plasminogen streptokinase activator complex) in acute myocardial infarction: the Duke University Clinical Cardiology Study (DUCCS) 1. J Am Coll Cardiol 23:11, 1994.
Holmes WE, Pernica D, Blaber M, et al: Cloning and expression of the gene for prourokinase in E. coli. Biotechnology 3:923, 1985.
Riccio A, Grimaldi G, Verde P, Sebastio G, Boast S, Blasi F: The human urokinase-plasminogen activator gene and its promoter. Nucleic Acids Res 13:2759, 1985.
Verde P, Stoppelli MP, Galeffi P, Di Norcera P, Blasi P: Identification and primary sequence of an unspliced human urokinase poly(A)exp(+) RNA. Proc Natl Acad Sci USA 81:4727, 1984.
Verde P, Boast S, Franze A, Robbiati F, Blasi F: An upstream enhancer and a negative element in the 5′-flanking region of the human urokinase plasminogen activator gene. Nucleic Acid Res 16:10699, 1988.
Van Hinsbergh VW, van den Berg EA, Fiers W, Dooijewaard G: Tumor necrosis factor induces the production of urokinase-type plasminogen activator by human endothelial cells. Blood 75:1991, 1990.
Hussain S, Gurewich V, Lipinski B: Purification and characterization of a single-chain high-molecular-weight form of urokinase from human urine. Arch Biochem Biophys 220:31, 1983.
Wun TC, Schleuning WD, Reich E: Isolation and characterization of urokinase from human plasma. J Biol Chem 257:3276, 1982.
Wun TC, Ossowski L, Reich E: A proenzyme form of human urokinase. J Biol Chem 257:7262, 1982.
Gunzler W, Steffens G, Otting F, et al: The primary structure of high molecular mass urokinase from human urine. The complete amino acid sequence of the A chain. Hoppe Seyler’s Z Physiol Chem 363:1155, 1982.
Gunzler WA, Steffens GJ, Otting F, Buse G, Flohe L: Structural relationship between high and low molecular mass urokinase. Hoppe Seyler’s Z Physiol Chem 363:133, 1982.
Ellis V, Scully MF, Kakkar VV: Plasminogen activation by single-chain urokinase in functional isolation. A kinetic study. J Biol Chem 262:14998, 1987.
Pannell R, Gurewich V: Activation of plasminogen by single-chain urokinase or by two-chain urokinase—a demonstration that single-chain urokinase has a low catalytic activity (pro-urokinase). Blood 69:22, 1987.
Nelles L, Lijnen HR, Collen D, Holmes WE: Characterization of recombinant human single chain urokinase-type plasminogen activator mutants produced by site-specific mutagenesis of lys-158. J Biol Chem 262:5682, 1987.
Collen D, Zamarron C, Lijnen HR, Hoylaerts M: Activation of plasminogen by pro-urokinase: II. Kinetics. J Biol Chem 261:1259, 1986.
Christensen U, Mullertz S: Kinetic studies on the urokinase catalyzed conversion of NH2-terminal lysine plasminogen to plasmin. Biochem Biophys Acta 480:275, 1977.
Takada A, Watahiki Y, Takada Y: Release of the N-terminal peptides from Glu-plasminogen by plasmin in the presence of fibrin. Thromb Res 41:819, 1986.
Lucas MA, Straight DL, Fretto LJ, McKee PA: The effects of fibrinogen and its cleavage products on the kinetics of plasminogen activation by urokinase and subsequent plasmin activity. J Biol Chem 258:12171, 1983.
Markus G, Priore RL, Wissler FC: The binding of tranexamic acid to native (glu) and modified (lys) plasminogen and its effect on conformation. J Biol Chem 254:1121, 1979.
Vali Z, Patthy L: The fibrin binding site of human plasminogen: arginines 32 and 34 are essential for fibrin affinity of the kringle 1 domain. J Biol Chem 259:13690, 1984.
Violand BN, Sodetz JM, Castellino FJ: The effect of epsilon-amino caproic acid on the gross conformation of plasminogen and plasmin. Arch Biochem Biophys 170:300, 1975.
Ellis V, Behrendt N, Dano N: Plasminogen activation by receptor-bound urokinase. A kinetic study with both cell-associated and isolated receptor. J Biol Chem 266:12752, 1991.
Kohler M, Sen S, Miyashita C, et al: Half-life of single-chain urokinase-type plasminogen activator (scu-PA) and two-chain urokinase-type plasminogen activator (tcu-PA) in patients with acute myocardial infarction. Thromb Res 62:75, 1991.
Collen D, De Cock F, Lijnen HR: Biological and thrombolytic properties of proenzyme and active forms of human urokinase-II. Turnover of natural and recombinant urokinase in rabbits and squirrel monkeys. Thromb Haemost 52:24, 1984.
Pennica D, Holmes WE, Kohr WJ, et al: Cloning and expression of human tissue-type plasminogen activator cDNA in E. coli. Nature 301:214, 1983.
Edlund T, Ny T, Ranby M, et al: Isolation of cDNA sequences coding for a part of human tissue plasminogen activator. Proc Natl Acad Sci USA 80:349, 1983.
Verheijen JH, Visse R, Wijnen J TH, Chang GT, Kluft C, Meerg Khan P: Assignment of the human tissue-type plasminogen activator gene (PLAT) to chromosome 8. Hum Genet 72:153, 1986.
Rajput B, Degen SF, Reich E, et al: Chromosomal locations of human tissue plasminogen activator and urokinase genes. Science 230:672, 1985.
Degen SF, Rajput B, Reich E: The human tissue plasminogen activator gene. J Biol Chem 261:6972, 1986.
Kooistra T, Bosma PJ, Toet K, et al: Role of protein kinase C and cyclic adenosine monophosphate in the regulation of tissue-type plasminogen activator, plasminogen activator inhibitor-1, and platelet-derived growth factor mRNA levels in human endothelial cells. Possible involvement in proto-oncogenes c-jun and c-fos. Arterioscler Thromb 11:1042, 1991.
Medcalf RL, Ruegg M, Schleuning WD: A DNA motif related to the cAMP-responsive element and an exon-located activator protein-2 binding site in the human tissue-type plasminogen activator gene promoter cooperate in basal expression and convey activation by phorbol ester and cAMP. J Biol Chem 265:14618, 1990.
Rijken DC, Wijngaards G, Zaal-de Jong M, Welbergen J: Purification and partial characterization of plasminogen activator from human uterine tissue. Biochem Biophys Acta 580:140, 1979.
Levine EG, Loskutoff DJ: Cultured bovine endothelial cells produce both urokinase and tissue-type plasminogen activators. J Cell Biol 94:631, 1982.
Pohl G, Einarssan M, Nilsson B, Svensson S: The size heterogeneity in melanoma tissue plasminogen activator is caused by carbohydrate differences. Thromb Res 50:163, 1988.
Ichinose A, Kisiel W, Fukikawa K: Proteolytic activation of tissue plasminogen activator by plasmin and tissue enzymes. FEBS Lett 175:412, 1984.
Loscalzo J: Structural and kinetic comparison of recombinant human single- and two-chain tissue plasminogen activator. J Clin Invest 82:1391, 1988.
Hoylaerts M, Rijken DL, Lijnen HR, Collen D: Kinetics of the activation of plasminogen by human tissue plasminogen activator. Role of fibrin. J Biol Chem 257:2912, 1982.
Nieuwenhuizen W, Voskuilen M, Vermond A, Hoegee-de Nobel B, Traas DW: The influence of fibrin(ogen) fragments on the kinetic parameters of the tissue-type plasminogen-activator-mediated activation of different forms of plasminogen. Eur J Biochem 174:163, 1988.
Suenson E, Bjerrum P, Holm A, et al: The role of fragment X polymers in the fibrin enhancement of tissue plasminogen activator-catalyzed plasmin formation. J Biol Chem 265:22228, 1990.
Rijken DL, Groeneveld E: Isolation and functional characterization of the heavy and light chains of human tissue-type plasminogen activator. J Biol Chem 261:3098, 1986.
Suensen E, Petersen LC: Fibrin and plasminogen structures essential to stimulation of plasmin formation by tissue-type plasminogen activator. Biochem Biophys Acta 870:510, 1986.
Hajjar KA, Hamel NM, Harpel PC, Nachman RL: Binding of tissue plasminogen activator to cultured human endothelial cells. J Clin Invest 80:1712, 1987.
Felez J, Chanquia CJ, Leven EG, Miles LA, Plow EF. Binding of tissue plasminogen activator to human monocytes and monocytoid cells. Blood 78:2318, 1991.
Stricker RB, Wang D, Shiu DT, Reyes PT, Shuman MA: Activation of plasminogen by tissue plasminogen activator on normal and thrombasthenic platelets: effects on surface proteins and aggregation. Blood 68:275, 1986.
Verstraete M, Su CA, Tanswell P, Feuerer W, Collen D: Pharmacokinetics and effects on fibrinolytic and coagulation parameters of two doses of recombinant tissue-type plasminogen activator in healthy volunteers. Thromb Haemost 56:1, 1986.
Lack CH: A product from S. aureus that is fibrinolytic. Nature 161:559, 1948.
Collen D: Staphylokinase: a potent, uniquely fibrin-selective thrombolytic agent. Nat Med 4:279, 1998.
Lijnen HR, Van Hoef B, De Cock F, et al: On the mechanism of fibrin-specific plasminogen activation by staphylokinase. J Biol Chem 266:11826, 1991.
Suehiro A, Tsujioka H, Yoshimoto H, Ueda M, Higasa S, Kakishita E: Enhancing effect of platelets on staphylokinase-mediated clot lysis and plasminogen activation. Thromb Res 80:135, 1995.
Vanderschueren S, Dens J, Kerbinchai P, et al: Randomized coronary patency trial of double-bolus recombinant staphylokinase versus front-loaded alteplase in acute myocardial infarction. Am Heart J 134:213, 1997.
Collen D, Bernaerts R, Declerck P, et al: Recombinant staphylokinase variants with altered immunoreactivity: I. Construction and characterization. Circulation 94:197, 1996.
Burck PJ, Berg DH, Warrick MW, et al: Characterization of a modified human tissue plasminogen activator comprising a kringle-2 and a protease domain. J Biol Chem 265:5170, 1990.
Jackson CV, Crowe VG, Craft TJ, et al: Thrombolytic activity of a novel plasminogen activator, LY210825, compared with recombinant tissue-type plasminogen activator in a canine model of coronary artery thrombosis. Circulation 82:930, 1990.
Anonymous. A comparison of reteplase with alteplase for acute myocardial infarction. The Global Use of Strategies to Open Occluded Arteries (GUSTO III) Investigators. N Engl J Med 337:1118, 1998.
Cannon CP, McCabe CH, Gibson CM, et al: TNK-tissue plasminogen activator in acute myocardial infarction. Results of the Thrombolysis in Myocardial Infarction (TIMI) 10A dose-ranging trial. Circulation 95:351, 1997.
Kratzschmar J, Haendler B, Langer G, et al: The plasminogen activator family from the salivary gland of the vampire bat Desmodus rotundus: cloning and expression. Gene 105:229, 1991.
Witt W, Baldus B, Bringmann P, Cashion L, Donner P, Schleuning WD: Thrombolytic properties of Desmodus rotundus (vampire bat) salivary plasminogen activator in experimental pulmonary embolism in rats. Blood 79:1213, 1992.
Granger CB, Califf RM, Topol EJ: Thrombolytic therapy for acute myocardial infarction. Drugs 44:293, 1992.
Cairns JA, Fuster V, Kennedy JW: Coronary thrombolysis. Chest 102:482S, 1992.
Haber E, Quertermous T, Matsueda GR, Runge MS: Innovative approaches to plasminogen activator therapy. Science 243:51, 1989.
Vaughan DE, Loscalzo J: New directions in thrombolytic therapy: Molecular mutants and biochemical conjugates. Trans Cardiovasc Med 1:36, 1991.
Bovill EG, Becker R, Tracy RP: Monitoring thrombolytic therapy. Prog Cardiovasc Dis 34:279, 1992.
Loscalzo J: Thrombolysis in the management of acute myocardial infarction and unstable angina. Drugs 37:191, 1989.
Coller BS: Platelets and thrombolytic therapy. N Engl J Med 322:33, 1990.
Shebuski RJ: Principles underlying the use of conjunctive agents with plasminogen activators. Ann NY Acad Sci 667:382, 1992.
Sherry S, Marder VJ: Thrombosis, fibrinolysis, and thrombolytic therapy: a perspective. Prog Cardiovasc Dis 34:89, 1991.
Califf RM, Topol EJ, George BS, Boswick JM, Abbottsmith C, Sigmon KN, et al: Hemorrhagic complications associated with the use of intravenous tissue plasminogen activator in treatment of acute myocardial infarction. Am J Med 85:353, 1988.
Pasche B, Loscalzo J: Platelets and fibrinolysis. Platelets 2:125, 1991.
Eisenberg PR, Sherman LA, Jaffe AS: Paradoxic elevation of fibrinopeptide A after streptokinase: evidence for continued thrombosis despite intense fibrinolysis. J Am Coll Cardiol 10:527, 1987.
Owen J, Friedman KD, Grossman BA, Wilkins C, Berke AD, Powers ER: Thrombolytic therapy with tissue plasminogen activator or streptokinase induces transient thrombin activity. Blood 72:616, 1988.
Anonymous: Comparison of invasive and conservative strategies after treatment with intravenous tissue-type plasminogen activator in acute myocardial infarction (TIMI) phase II. N Engl J Med 320:618, 1989.
Roux S, Christeller S, Luediu E: Effects of aspirin on coronary reocclusion and recurrent ischemia after thrombolysis: A meta-analysis. J Am Coll Cardiol 19:671, 1992.
Hirsh J: Heparin and low-molecular-weight heparins. Cor Art Dis 3:990, 1992.
Ezratty A, Loscalzo J: New approaches to antiplatelet therapy. Blood Coag Fibrinol 2:317, 1991.
Coller BS: Inhibitors of platelet glycoprotein IIb/IIIa receptor as conjunctive therapy for coronary artery thrombolysis. Cor Art Dis 3:1016, 1992.
Anonymous: Platelet glycoprotein IIb/IIIa receptor blockade and low-dose heparin during percutaneous coronary revascularization. The EPILOG Investigators. N Engl J Med 336:1989, 1997.
Scharfstein J, Loscalzo J: The molecular biology of antithrombotic therapy. Hosp Pract 27:41, 1992.
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Ernest Beutler, Marshall A. Lichtman, Barry S. Coller, Thomas J. Kipps, and Uri Seligsohn