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Fibrinolysis with recombinant single-chain urokinase- type plasminogen activator (saruplase) in acute myocardial infarction
Fibrinolyse met recombinant single-chain urokinase-type plasminogen activator (saruplase) bij het acute myocardinfarct
Proefschrift ter verkrijging van de graad van doctor aan de Universiteit Nijmegen op gezag van de Rector magnificus Prof. Dr T.J.M, van Els en volgens het besluit van het College van Decanen.
De openbare verdediging zal plaats vinden op vrijdag 18 juni 1999 om 11.00 uur door
Herman Rudolf Michels
Geboren te Paramaribo in 1945. Promotor: Prof. Dr. F.W. Α. Verheugt
Co-promotor: Prof. Dr. F.W.H.M. Bär (Universiteit Maasstricht)
Referent: Prof. Dr. D. Collen (Katholieke Universiteit Leuven)
Manuscriptcommissie: Prof. Dr. P. Smits (voorzitter) Prof. Dr. J.H.J. Ruijs Prof. Dr. T. de Witte Publication of this thesis was financially supported by the Cardiac Therapy Research Institute Eindhoven, 'CATHREINE'. 1/w»i:
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Some returns on investment are impossible to measure
(N. Rockefeller) Contents
Chapter 1 11
General introduction
Chapter 2 15
Saruplase in acute myocardial infarction. Frits W. Bär, Frank Vermeer, Rolf Michels, Jean Boland, Jürgen Meyer, Gwyn Hopkins, Hannes Barth, Wolfgang A. Giinzler. J Thromb Thrombol 1995; 2:195-204. Reprinted with permission from the Journal of Thrombosis and Thrombolysis. ©Kluwer Academic Publishers, Boston.
Chapter 3 35
A double-blind multicenter comparison of the efficacy and safety of saruplase and urokinase in the treatment of acute myocardial infarction: report of the SUTAMI study group. Rolf Michels, Hans Hoffmann, Jürgen Windeler, Hannes Barth, and Gwyn Hopkins on behalf of the SUTAMI investigators. J Thromb Thrombol 1995; 2: 117-124. Reprinted with permission from the Journal of Thrombosis and Thrombolysis. ©Kluwer Academic Publishers, Boston.
Chapter 4 55
Hemostatic changes after thrombolytic therapy with saruplase (unglycosylated single-chain urokinase-type plasminogen activator) and urokinase (two-chain urokinase-type plasminogen activator). H.R. Michels, J.J.M.L. Hoffmann, J. Windeler, G.R. Hopkins. Blood Coag and Fibrinolysis 1996; 7: 766-771. Reprinted with permission from Blood Coagulation & Fibrinolysis. ©1996, Rapid Science Publishers Chapter 5 69
Plasma markers of thrombin activity during coronary thrombolytic therapy with saruplase or urokinase: no prediction of reinfarction. J.J.M.L. Hoffmann, H.R. Michels, J. Windeler, W.A. Günzler. Fibrinolysis 1993; 7: 330-334. Reprinted with permission from Fibrinolysis. © Longman Group, UK Ltd.
Chapter 6 83
Comparison of saruplase and alteplase in acute myocardial infarction. Frits W. Bär, Jürgen Meyer, Frank Vermeer, Rolf Michels, Bernard Charbonnier, Klaus Haerten, Martin Spiecker, Carlos Macaya, Michel Hanssen, Magda Heras, Jean P. Boland, Marie-Claude Morice, Francis G. Dunn, Rainer Uebis, Christian Hamm, Oded Ayzenberg, Gerhard Strupp, Adrie Withagen, Werner Klein, Jürgen Windeler, Gwyn Hopkins, Hannes Barth, Michael von Fisenne, for the SESAM study group. Am J Cardiol 1997; 79: 727-732. Reprinted with permission from the American Journal of Cardiology. © 1997 by Excerpta Medica, Inc.
Chapter 7 99
Randomized, double-blind study comparing saruplase with streptokinase therapy in acute myocardial infarction: the COMPASS equivalence trial. Ulrich Tebbe, Rolf Michels, Jennifer Adgey, Jean Boland, Avi Caspi, Bernard Charbonnier, Jürgen Windeler, Hannes Barth, Robert Groves, Gwyn R. Hopkins, Wiliam Fennell, Amadeo Betriu, Mikhial Ruda, Johannes Mlzoch, for the COMPASS trial of saruplase and streptokinase (COMPASS) investigators. J Am Coll Cardiol 1998; 31: 487-493. Reprinted with permission from the American Society of Cardiology. © 1998 by the American College of Cardiology. Chapter 8 119
Pharmacokinetics and hemostatic effects of saruplase in patients with acute myocardial infarction. Comparison of infusion, single bolus and split-bolus administration. H.R. Michels, J.J.M.L. Hoffmann, F.W. H. M. Bär. Submitted for publication..
Chapter 9. Summary and conclusions 137
Fibrinolysis with recombinant single-chain urokinase-type plasminogen activator (saruplase) in acute myocardial infarction. Summary of the results of clinical trials. H.R. Michels, F.W.H.M. Bär, F.W.A. Verheugt. Submitted for publication.
Chapter 10 159
Samenvatting en conclusies.
Dankbetuiging 163
Curriculum vitae 165 10 INTRODUCTION
Chapter 1
General introduction
Rapid and sustained coronary reperfusion and the prevention of (sub)acute reocclusion, bleeding complications and allergic reactions are the main objectives of coronary thrombolysis for acute myocardial infarction.With currently used intravenous thrombolytic agents the time to reperfusion is usually greater than 45 min and (sub)acute reocclusion is not infrequent. Current thrombolytics result in, at best, a maximum patency of 75% at 90 min after the start of infusion. The most commonly used agent, streptokinase reaches this plateau only hours later. Streptokinase is a nonphysiological, non-fibrin-selective fibrinolytic agent. The physiological fibrin-selective recombinant tissue-type plasminogen activator (rt-PA) has shown a difference to streptokinase in terms of efficacy of early reperfusion. The risk of reocclusion probably varies with different thrombolytic agents, and may be related to the extent of systemic lysis (i.e. plasmin generation), fibrinogen depletion, increase of (fibrinogen) degradation products, alteration of platelet activity and blood viscosity and the severity and geometry of residual stenosis. Bleeding complications occur in approximately 10-15% of patients. There is a trend towards more hemorrhagic strokes with the more potent fibrin-selective agents. This trend may partially counteract the benefit of more rapid reperfusion. Nonphysiological fibrinolytic agents consist of proteins which are foreign to the human body and may generate antibodies and cause allergic reactions. Shortly after streptokinase administration the anti-streptokinase titer rises up to 100 times the baseline value and remains high for over 6 months. Since antibodies may affect thrombolytic efficacy, streptokinase should not be re-administered during this time. "Time is myocardium": storage, preparation and administration of medication should be as simple as possible (preferably as a bolus). Better drugs are generally more expensive, however, greater efficacy should also be cost-effective. In summary, a fibrinolytic agent should ideally show very early maximum reperfusion, little reocclusion, no excess of bleeding, and no antibody formation. Moreover preparation and administration should be simple, and the drug should be inexpensive. None of the currently available agents fulfills all of these criteria, and whilst rt-PA so far appears the most satisfactory it is not exactly inexpensive, and is not recommended to be
11 CHAPTER 1 administered as a bolus. The focus is therefore on effective, physiological, fibrin-selective, not-too-expensive newcomers in the field of fibrinolytic therapy, that possess none of the disadvantages of the currently commercially available fibrinolytic agents. Saruplase is examined in the light of these criteria. A detailed evaluation of the newer mutants of wild- type t-PA, such as lanetoplase, reteplase, and TNK-t-PA, is beyond the scope of this review, the aim of which will be restricted to discussing recombinant unglycosylated single-chain urokinase-type plasminogen activator (saruplase) in the context of fibrinolytic medications commercially available in Europe. Of the latter only streptokinase and rt- PA have gained widespread popularity in clinical practice. The human enzyme urokinase or two-chain urokinase-type plasminogen activator (tcu- PA), is an effective but expensive fibrinolytic agent that has been registered for acute myocardial infarction in some, but not all European countries. In this thesis the studies that compare saruplase (given as a bolus of 20 mg intravenously followed by an infusion of 60 mg over 1 hour) with streptokinase, urokinase and rt-PA will be presented and discussed. In addition the pharmacokinetics and hemostatic effects of single bolus and split-bolus administration of saruplase will be presented and discussed. No comparisons of saruplase with anistreplase, currently the only fibrinolytic agent available for a single bolus administration, or with reteplase, recently registered for double bolus administration, are available. Anistreplase, or anisoylated plasminogen-streptokinase-activator complex (APSAC), may have some benefits over streptokinase, but it not only has all the shortcomings of that agent, and its high price has generally operated against its widespread use. The efficacy and safety of reteplase and rt-PA are quite comparable. APSAC and the newer mutants of rt-PA will be mentioned only when relevant to the point under discussion. Chapter 2 presents the background and biochemistry of saruplase, and the results of the trials up to 1995, that have compared saruplase with streptokinase, urokinase and alteplase. Chapter 3 presents the SUT AMI study, a comparison of the efficacy and safety of saruplase and urokinase in the treatment of acute myocardial infarction. Chapter 4 presents a sub-study of the SUT AMI study, that concentrates on the hemostatic changes after thrombolytic therapy with saruplase and urokinase. Chapter 5 presents another sub-study of the SUTAMI study, that concentrates on possible plasma markers of reinfarction.
12 INTRODUCTION
Chapter 6 presents the results of the SESAM study, a comparison of saruplase and alteplase in acute myocardial infarction. Chapter 7 presents the results of the COMPASS trial , an equivalence study, that compared saruplase with streptokinase in acute myocardial infarction. Chapter 8 presents a comparison of the pharmacokinetics and hemostatic effects of saruplase in acute myocardial infarction, when given as infusion, single bolus or split-bolus. Chapter 9 gives a summary and conclusions from the preceding chapters. Chapter 10 contains the abbreviated summary and conclusions in Dutch. The thesis concludes with the acknowledgments (in Dutch) and a curriculum vitae (in English)
13 14 CHAPTER 2
Saruplase in Myocardial Infarction
Frits W. Bär,' Frank Vermeer,1 Rolf'Michels,2 Jean Boland,3 Jürgen Meyer,4 Gwyn Hopkins,5 Hannes Barth, ' Wolfgang A. Grünzier5
'Department of Cardiology, University Hospital Maastricht, Maastricht, the Netherlands, department of Cardiology, Catharina Hospital Eindhoven, Eindhoven, the Netherlands, 'Department of Cardiology, CHR Citadelle Liège, Liège Belgium, ^Department of Cardiology, Universitätsklinik Johannes Gutenberg, Mainz, Germany, 5Grunenthal GmbH, Aachen, Germany
Abstract. Saruplase is an unglucosylated single-chain recombinant urokmase-type plasminogen activator Dose finding studies in patients with acute myocardial infarction indicated that a dose of 80 mg of saruplase, given as a bolus of 20 mg and iv infusion of 60 mg in one hour, led to excellent patency figures Saruplase is most effective when combined with a bolus of 5000 IU heparin followed by an iv heparin infusion for at least 24 hours When saruplase is compared to other thrombolytic agents (streptokinase, alteplase, urokinase), it becomes apparent that its profile is excellent harly patency rates are at least comparable to alteplase Further reocclusion rates of saruplase after one day are lower than those of streptokinase and alteplase Patency rates 24 - 72 hours after start of medication are comparable between saruplase and urokinase The large database in over 6000 patients shows that saruplase, in comparison to the other thrombolytic agents, is safe Its bleeding complication rate is significantly lower than streptokinase, and a trend to lower in-hospital mortality is observed when compared to urokinase Summarizing, when comparing to the presently available thrombolytic agents, saruplase is a fast acting, effective and safe thrombolytic agent
Key Words. Myocardial infarction, thrombolytic therapy, patency, safety
Myocardial infarction is a major cause of death in the industrialized world. Over the last few years major advances have been made both in the prevention and treatment of acute myocardial infarction Autopsy data and studies with thrombolytic agents in patients with myocardial infarction have demonstrated that thrombosis usually plays a crucial role in the occurrence
15 Chapter 2 of a sudden occlusion of coronary arteries. It had been shown that thrombolytic therapy is capable of reopening the majority of these occluded vessels, thereby limiting infarct size, morbidity, and mortality. As expected, reperfusion therapy is most effective if it is given early, and when it is infused in patients with extensive myocardial infarction [1-3]. Streptokinase and urokinase were the first available drugs in this class [2,4]. Later, as other agents became available, it became apparent that the profiles of the different thrombolytics were not identical [5,6]. For example, alteplase treatment was shown to result in earlier reperfusion than streptokinase treatment [7 - 9]. In addition, these and other studies suggested a greater reocclusion rate for alteplase than for streptokinase during the following few days [10]. This at least partially explains why in large-scale trials no major difference in clinical outcome was found between the drugs [11,12]. Differences in the safety profiles of the above-mentioned agents were minor, with the exception of hypotension and the formation of antibodies in patients treated with streptokinase [2] or anistreplase [13-15]. In recent years, the new thrombolytic agent saruplase has been tested in patients with acute myocardial infarction. In this paper we report on its efficacy and safety.
Profile: Saruplase
Saruplase is the international nonproprietary name for unglycosylated human-type high molecular weight single-chain urokinase-type plasminogen activator produced by recombinant gene technology in E. coli bacteria. Saruplase has the same amino acid sequence as natural scu-PA but, in contrast, lacks a carbohydrate side chain. With saruplase, effective and clot-specific lysis has been observed both in vitro and in vivo. Animal experiments with artificially embolized coronary vessels have shown that the intravenous infusion of saruplase is able to cause fast recanalization without systemic fibrinogenolysis. These results were unexpected since saruplase, in contrast to t-PA, does not bind to fibrin, and it resembles a zymogen in that it is neither inactivated by plasma inhibitors nor shows significant enzymatic activity when assayed with a urokinase peptide substrate in vitro. Its fibrin-specific action may be explained as follows. Saruplase is unable to activate the circulating form of plasminogen in plasma, whereas it rapidly and selectively cleaves the plasminogen form bound to the fibrin of a blood clot without necessarily
16 Saruplase ¡η AMI being converted into its two-chain urokinase derivative. This explanation means that active plasmin is generated selectively at the site of the clot. Since traces of plasmin rapidly convert saruplase to its active two-chain (scu-PA) derivative, a positive feedback on the plasmin generation occurs at the site of the clot that amplifies the local lytic potency, whereas the local proteinase inhibitors are relatively ineffective. When washed out into the circulation, plasmin and the two-chain urokinases derived from saruplase are rapidly inactivated by plasma inhibitors, for example, a2-antiplasmin and plasminogen activator inhibitors. Consequently, the fibrinolytic activity of saruplase is primarily limited to the area of the fibrin clot, while the systemic plasminogen activation is inhibited. However, as this inhibition depends on the endogenous inhibitor capacity, systemic plasmin activity and its sequelae can appear during thrombolysis with saruplase, especially when high doses are emloyed. This explains why systemic effects, for example, o.2- antiplasmin consumption and fibrinogen breakdown, are observed during the course of lysis with saruplase in animal experiments and clinical studies.
Conversions of Saruplase in Plasma
Although scu-PA is able to trigger the dissolution of thrombi on its own, it achieves its full plasminogen-activating activity only after proteolytic conversion (Figure 1). Accordingly, saruplase is cleaved by proteases such as plasmin or kallikrein at position 158 into its two-chain derivative (Figure 2). The aminoterminal A chain contains the EGF-like and kringle domains, and is linked by a disulfide bridge to the В chain, which represents the fully active serine protease domain. While scu-PA is virtually amidolytically inactive, chromogenic peptide substrates are rapidly cleaved by tcu-PA. Another conversion of saruplase is indicated in Figures 1 and 2. The high molecular weight form is converted into the low molecular weight form by cleavage of the Lys 134 - Lys 135 bond. Since no disulfide bond bridges the domains to the serine protease domain, the low molecular weight u-PA forms lack EGF-like and kringle domains (Figure 2).
17 Chapter 2
Fig. 1. The amino acid sequence of saruplase in single letter code. Arrows indicate peptide cleavage sites that convert saruplase into its active two-chain high molecular mass (HMW tcu-PA) derivative (at Lys158) and into a low, molecular mass (LMW tcu-PA) derivative (at Lys138). Asterisks mark active site amino acids.
Experimental Experience with Saruplase In animals several studies have been performed indicating that saruplase is a potent thrombolytic agent [16-19]. These results stimulated further work in humans, which will be discussed.
Clinical Studies with Saruplase in Patients with Myocardial Infarction
Dose-finding studies Two studies [20,21] were done to determine the adequate dose regimen of saruplase in patients with acute myocardial infarction. In the first dose- finding trial [20], eight patients were treated with 40 mg saruplase (10 mg iv bolus plus 30 mg iv infusion over 60 minutes), and nine patients received 70
18 Saruplase in AMI mg saruplase (10 mg iv bolus and 60 mg iv infusion over 60 minutes). This reperfusion study showed higher patencies of the 70 mg dose: at 90 minutes six of nine patients had TIMI grade 3 flow and one patient had TIMI grade 2 flow, whereas patients treated with 40 mg saruplase showed complete reperfusion in three of eight patients and partial reperfusion in another three patients. In the second dose-finding reperfusion study [21], patients were treated with either 40 mg saruplase (10 mg iv bolus plus 30 mg iv infusion over 60 minutes, η = 12) or 80 mg saruplase (20 mg iv bolus plus 60 mg iv infusion over 60 minutes, η = 12). At 90 minutes 4 of 12 patients in the 40 mg group had reperfusion (TIMI 2 or 3) compared with 10 of 12 patients in the 80 mg group. As expected, disturbance of the hemostatic system was most pronounced in the patients treated with 80 mg of saruplase. From the outcome of these two studies, it was decided that future trials would use the 80 mg dosing regime of a 20 mg iv bolus plus 60 mg iv infusion over 60 minutes.
LMW soj-PA (activible)
HMW IQJ-PA inhibitor complex (inactive)
Plaemln proteinase Inhibitors e.g. PAM
LMW leu-ΡΑ (active) LMW Icu-PA inhibitor complex (inactive)
Fig. 2. Derivatives of saruplase potentially generated in plasma in vivo.
19 Chapter 2
Interaction Studies
Two interaction studies [22,23] have been performed. In the first investigation the effect of a heparin bolus administered prior to saruplase was studied. The second trial addressed the effect of the concomitant administration of the prostacyclin analogue taprostene to saruplase.
Bolus of heparin From animal experience it was concluded that conjunctive heparin was able to enhance the efficacy of saruplase [24,25]. To test this finding in humans, a placebo-controlled study was conducted in 118 acute myocardial infarction patients [22]. In 56 patients a single bolus of 5000 IU heparin was given prior to thrombolytic therapy, and 62 patients received a placebo for heparin (control group). Starting 30 minutes after completion of the saruplase infusion, all patients had a 5 day uninterrupted infusion of heparin titrated against the APTT. Angiography, performed between 6 and 12 hours after the start of therapy, showed a significantly higher patency (TIMI 2 or 3) in the prelysis heparin group: 78.6% versus 56.5% in the control group (p = 0.01). Safety data showed a trend toward a better outcome in the prelysis heparin group: In-hospital mortality was 5.4% versus 14.5% in the control group. Because of the results of this study, it was strongly recommended that saruplase always should be preceded by an iv bolus of 5000 IU of heparin.
Taprostene study Taprostene is a prostacyclin analogue that inhibits platelet aggregation [26 - 29]. After successful thrombolysis, the exposed ruptured plaque promotes platelet aggregation. Therefore it was thought that taprostene might be a useful adjuvant to thrombolytic therapy. In a dose-finding study [23] performed in 80 patients, three dose levels of taprostene were compared with placebo. No significant difference was found in patency when comparing placebo with the three dose levels, and thus taprostene cotherapy probably does not enhance lysis.
20 Sarupi ase in AMI
Comparison of Saruplase with Other Thrombolytic Agents
Saruplase and streptokinase In the PRIMI trial [30], 80 mg iv saruplase was compared with 1.5 million iv streptokinase over 60 minutes in 401 patients with a first myocardial infarction. The first important finding was that saruplase had significantly higher patency rates (TIMI 2 or 3) than streptokinase at 60 minutes after the start of therapy: 71.8% versus 48.0% (p< 0.001). The difference was less pronounced at 90 minutes, 71.2% and 63.9%, respectively (p = 0.15) (Figure 3).
PRIMI patency rate (%)
100 A 88 D saruplase 85 D streptokinase 80 72 71 I 64 60 4Θ
40
20
0 ш 60 min 90 mm 24 to 36 h
Fig. 3. Comparison of patency rates (TIMI 2 or 3) at 60 minutes, 90 minutes, and 24-36 hours in patients with acute myocardial infarction treated with saruplase (20 mg iv bolus 60 mg as 1 hour infusion; η = 198) or streptokinase (1 5 million units infusion over 1 hour; η = 203). At 60 minutes patency rates are significantly higher in the saruplase group than in the streptokinase group (p< 0 001). Thereafter differences do not reach statistical significance.
21 Chapter 2
PRIMI Fibrinogen concentration (median; g/L)
A D saruplase D streptokinase
baseline 30 min 60 mm 90 mm 6 to 12 h
Fig. 4. Fibrinogen concentrations in the PRIMI trial (saruplase vs. Streptokinase). After treatment, fibrinogen fall was significantly lower at all given time points (p < 0.0001) in the streptokinase group than in the saruplase group.
Interestingly, of those patients with patent arteries at 90 minutes in whom no further intervention was done, angiographic reocclusion the following day was only 1% for the saruplase patients compared with 4% for the streptokinase patients. The third important finding of this trial was that saruplase led to fewer bleeding complications (14.7%) than streptokinase (24.6%) (p< 0.001). Intracranial bleeding rates were comparable (Table 1). Streptokinase also had a more marked effect on the hemostatic system (Figure 4). Other safety data indicated low in-hospital and 1 year mortality rates for both groups, with fewer episodes of hypotension in the immediate infusion period in saruplase patients.
22 Sarupíase in AMI
Table 1. Adverse event data of the PRIMI trial comparing saruplase with streptokinase in patients with acute myocardial infarction
Adverse events Saruplase Streptokinase (n = 198) (n = 208)
Reinfarction 7.1 % 5.9 % Reocclusion 5.0 % 4.4 % Mortality Hospital 3.5 % 4.9 % 1 year 6.3 % 5.6 % Bleeding Total 14.1% 24.6 % Severe 4.0 % 11.3% Stroke Ischemic 0.5 % 0.5% Hemorrhagic 1.0% 0.5%
23 Chapter 2
Saruplase and urokinase The SUT AMI trial [31,32] was a randomized study in 543 patients treated with either 80 mg saruplase or 3.0 million urokinase in 60 minutes. This study started before the outcome of the heparin bolus study [22] was available, and no pretreatment with a heparin bolus was given. Patency rates (TIMI 2 or 8) were determined 24 - 72 hours after the start of medication and were comparable: 75.4% and 74.2%, respectively. Safety data showed a trend toward more hemorrhagic strokes in the saruplase (1.1%) versus the urokinase (0%) group (p = 0.25). However, in-hospital mortality had the opposite trend: saruplase 4.4% and urokinase 8.1% (p = 0.08) (Table 2).
Table 2. Adverse event data of the SUTAM1 study comparing saruplase with urokinase in patients with acute myocardial infarction
Adverse events Saruplase Streptokinase (n =198) (n = 208) Reinfarction 6.9 % 4.7 % Mortality Hospital 4.4 % 8.1 % 1 year 7.5 % 11.7% Bleeding Total 10.7 % 10.7 % Severe 3.3 % 2.6 % Stroke Ischemic 1.5% 0.7 % Hemorrhagic 1.1 % 0.0 %
Saruplase and alteplase The SAT trial [33] was a pilot study primarily examining the effects of saruplase and alteplase on the hemostatic system. Fifty-two acute myocardial infarction patients were randomly treated with saruplase or alteplase (10 mg iv bolus plus iv infusion of 50 mg over 60 minutes and iv infusion of 40 mg over 120 minutes). Again, in this early trial no preceding heparin bolus was given before study medication. Saruplase had a consistent and significantly greater effect on concentrations of fibrinogen, fibrin(ogen) degradation products, plasminogen, a2-antiplasmin, and D-dimer than alteplase (Figure
24 Sa rupiase in AMI
5). Bleeding complications, however, were more frequent and more severe in the alteplase group. Also patency 24 - 72 hours after the start of treatment was in favor of saruplase: 88% had open arteries compared with only 61% of the patients treated with alteplase.
The SESAM trial [34] was a confirmatory patency study. The questions to be answered in this study were based on the findings of the PRIMI and SAT trials [30,33]. In the former study, saruplase had been shown to act faster than streptokinase, with higher patency rates at 60 minutes after the start of medication. Further, the number of patients who had an angiographic reocclusion after successful thrombolysis with saruplase was extremely low (1%). Based on the literature [7,9], it was thought that alteplase would have an early effect on patency that was comparable with saruplase but that reocclusion would be greater. Two hundred and thirty-six patients were treated with saruplase and 237 patients with alteplase. Medication regimens were the same as in the SAT pilot study [33], except for an iv bolus of 5000 Ш of heparin before the start of the study medication followed by a heparin infusion. To investigate the speed of reperfusion, the first angiogram was made at 45 minutes after the start of medication. Saruplase had a trend toward a higher patency (TIMI 2 or 3) at 45 minutes (74.6%) than alteplase (68.9%) (p = 0.22). Over the next 45 minutes, the difference gradually disappeared: At 60 minutes patency rates were 79.9% and 75.3%, and at 90 minutes they were 79.9% and 81.4% (Figure 6). Interestingly, the patency data for alteplase were at least as good as those seen with the accelerated therapy infusion regimen used in the angiography section ofthat GUSTO trial [12].
25 Chapter 2
SAT fibrinogen concentration (%)
„ I/ —-·-/•' - - . - /- - ---/ -— -7' baseline 1 h after end of infusion end of infusion start of infusion t+1 h saruplase +12 to 18 h t+3 h alteplase
Fig. 5. Comparison of fibrogen concentrations in patients with acute myocardial infarction treated with saruplase (20 mg iv bolus, 60 mg as 1 hour infusion; η = 24) or alteplase (10 mg iv bolus, infusion of 50 mg in 60 minutes and 40 mg in 120 minutes; η = 28). After the start of treatment, fibrinogen concentrations are significantly lower in the alteplase group at all given time points (p < 0.0001).
Angiography was repeated at 24 - 40 hours and again showed a low reocclusion rate (determined by the need for rescue percutaneous transluminal coronary angioplasty (PTCA) during the first day or angiography at 24 - 40 hours) for saruplase (1.2%) in patients who had successful thrombolysis without further intervention. The surprising finding in this study was, however, that alteplase also had an unexpectedly low reocclusion rate of 2.4%. The safety data in the two study groups were comparable and not unusual. Therefore these will not be discussed in detail (Table 3).
26 Saruplase in AMI
SESAM patency rate (%)
100 D saruplase 94 92 94 94 D alteplase
80 75 2 75 69
60
40
20
Ша 45 min 60 min 90 min after intervention 24 to 40 h Fig. 6. Comparison of patency rates at 45 minutes, 60 minutes, last view of first catheterization, and 24 - 40 hours m patients with acute myocardial infarction treated with saruplase (20 mg iv bolus 60 as 1 hour infusion; η = 236), or alteplase (10 mg iv bolus, infusion of 50 mg over 60 minutes and 40 mg over 120 minutes, η = 237). Note that saruplase might act slightly faster than alteplase, however, at none of the given time points are differences significant. Table 3. Adverse event data of the SESAM trial comparing saruplase with alteplase in patients with acute myocardial infarction
Adverse events Saruplase Alteplase (n = 236) (n = 237)
Reinfarction 4.2 % 4.2 % Mortuality, hospital 4.7 % 3.8% Bleeding Total 42.8 % 39.7 % Severe 9.3 % 8.4 % Stroke Ischemic 0.8 % 1.3% Hemorrhagic 0.8% 0.8 %
27 Chapter 2
Safety study with saruplase The open-label PASS trial [35] was performed to assess the applicability and safety of saruplase in the daily routine of treatment of patients with acute myocardial infarction. In total, 1698 patients were included. Concomitant medication, including aspirin and heparin, was given according to the local routine. Adverse event data are shown in Table 4. The in-hospital mortality in this rather unselected patient population was low, 5.4%. Major bleeding in need of blood transfusion occurred in 1.2% of patients. Hemorrhagic and ischemic stroke rates were 0.5% and 0.4%, respectively. Reinfarction during admission was diagnosed in only 2.9% of patients. A coronary angiography was performed within 72 hours in 347 of 1698 patients (20.4%) and revealed a patency rate (TIMI 2 or 3) of 70.0%.
Table 4. Adverse event rates in 1698 myocardial infarction patients of the open-label PASS study with saruplase
η % In-hospital mortality 92 5Λ Total bleedings 84 4.9 Major bleedings 20 1.2 Cerebrovascular 15 0.9 accident Hemorrhagic 8 0.5 Ischemic 7 0.4 Reinfarcion 50 2.9
Discussion
The findings of the present studies with saruplase in acute myocardial infarction show good efficacy and safety profiles.
28 Saruplase in AMI
Patency and reocclusion Figure 7 shows a hypothetical curve of patency rates in time for saruplase, alteplase, and streptokinase using data of the PRIMI and the SESAM studies [30,34]. It is expected from earlier reperfusion studies [36,37] that patency at the start of thrombolytic therapy is between 15% and 20%.
100
40-
20-
0-1—ι 1 1 1 1 1 1—' 0 15 30 45 60 90 24 hrs
Fig. 7. Hypothetical patency (TIMI 2 or 3) curves in time are drawn after treatment with saruplase, streptokinase, and alteplase. At the start of thrombolytic agent therapy patency is set at 15%. Noninvasive ECG parameters suggest that reperfusion starts after 30 minutes. Saruplase has the fastest reperfusion rates, and streptokinase has the slowest rates. After 24 hours patency data are comparable. Data have been retrieved from the PRIMI and the SESAM trials (30,34).
Angiographic data from these two trials indicate that saruplase is probably a faster acting drug when compared with streptokinase, and it might also be slightly faster than alteplase. We also used (nonpublished) noninvasive parameters, such as the ECG and reduction of chest pain, to help to draw the lines indicating patency rates. The advantages of a fast-acting drug are clear. Reperfusion is achieved earlier, thereby limiting infarct size. In addition, the decision to perform rescue PTCA in case of failed thrombolysis can also be made earlier. The number of patients with reocclusion in the angiographic PRIMI and SESAM studies is low [30,34]. Saruplase is probably more effective in preventing
29 Chapter 2 reocclusion after successful thrombolysis than alteplase and streptokinase. It is likely that heparin and aspirin contribute positively to the excellent outcome seen with three drugs.
Safety The data, especially from the open-label PASS study [35], indicate an excellent safety profile for saruplase. Due to selection bias, this outcome might be too optimistic. However, when directly comparing saruplase with other thrombolytic agents in general, a comparable safety profile is found. Streptokinase seems to result in more bleeding complications. There does not appear to be any important difference between saruplase and alteplase. When compared with urokinase, saruplase had a higher incidence of strokes, but mortality was lower in saruplase patients. However, the rather high frequency of strokes in the SUT AMI trial [31,32] was not found in other saruplase studies. A direct comparison of saruplase with anistreplase is not available. Other studies suggest that anistreplase safety data are comparable with streptokinase [36,38]. Overall, it seems reasonable to presume that saruplase is at least as safe as the other presently available thrombolytic agents.
Hemostatic effects The effect of saruplase of the hemostatic parameters shows a profile lying between that of streptokinase and alteplase. Although saruplase has a more profound effect on the hemostatic system than alteplase, this has not been reflected in a difference in the incidence of bleeding.
Conclusions
When compared with the presently available thrombolytic agents, saruplase is a fast-acting, effective, and safe thrombolytic agent
Acknowledgment
We would like to thank Mrs. Margriet Muytjens for typing this manuscript.
30 Sa ru piase in AMI
References
1. Bär FW, Vermeer F, de Zwaan С, Ramentol M, Braat S, Simoons ML. Value of admission electrocardiogram in predicting outcome of thrombolytic therapy in acute myocardial infarction. Am J Cardiol 1987;59:6 - 13. 2. Gruppo Italiano per lo Studio della Streptochinase nell' Infarto Miocardio (GISSI). Effectiveness of intravenous thrombolytic treatment in acute myocardial infarction. Lancet 1986;1:397-402. 3. Vermeer F, Simoons M, Bär FW, Tijssen JGP, Domburg v. RT, Serruys PW. Which patients benefit most from early thrombolytic therapy with intracoronary streptokinase. Circulation 1986;74:1379-1389. 4. Mathey DG, Schofer J, Sheehan FH, Becher H, Tilsen V, Dodge HT. Intravenous urokinase in acute myocardial infarction. Am J Cardiol 1985;55:878-882. 5. The TIMI Study Group. The thrombolysis in myocardial infarction (TIMI) trial. N Engl J Med 1985;312:932-936. 6. Van de Werf F, Arnold AER. Intravenous tissue plasminogen activator and size of infarct, left ventricular function, and survival in acute myocardial infarction. Br Med J1988; 297: 1374-1379. 7. Chesebro JH, Knatterud G, Roberts R, Borer Y, Cohen AS, Dalen J. Thrombolysis in myocardial infarction (TIMI) trial, phase I. A comparison between intravenous tissue plasminogen activator and intravenous streptokinase. Circulation 1987;76:142 -154. 8. Verstraete M, Bernard R, Bory M, Brown RW, Collen D, de Bono DP. Randomized trial of intravenous recombinant tissue-type plasminogen activator versus intravenous streptokinase in acute myocardial infarction. Report from the European Cooperative Study Group for Recombinant Tissue- type Plasminogen Activator. Lancet 1985;1:842 - 847. 9. Simoons ML, Arnold AER, Betriu A, de Bono DP, Col J, Dougherty FC. Thrombolysis with tissue plasminogen activator in acute myocardial infarction: No additional benefit from immediate percutaneous coronary angioplasty. Lancet 1988;2:197-203. 10. Ohman EM, Califf RM, Topoi EJ, Candela R, Abbottsmith С, Ellis S. Consequences of reocclusion after successful reperfusion therapy in acute myocardial infarction. Circulation 1990;82:781 -791. 11. ISIS 2 (Second International Study of Infarct Survival) Collaborative Group. Randomized trial of intravenous streptokinase, oral aspirin, both, or neither among 17,187 cases of suspected acute myocardial infarction: ISIS-2. Lancet 1988;2:349-360. 12. The GUSTO Angiographic Investigators. The effects of tissue plasminogen activator, streptokinase, or both on coronary artery patency, ventricular function,
31 Chapter 2
and survival after acute myocardial infarction N Engl J Med 1993; 329: 1615 - 1623. 13. Relik-van Wely L, Visser RF, Van der Pol J, Bartholomeus I, Couvee JF, Drowst H. Angiographically assessed coronary arterial patency and reocclusion in patients with acute myocardial infarction treated with anistreplase: Results of the anistreplase reocclusion multicentre study (ARMS). Arn. J Cardiol 1991;68:296-300. 14. Bonnier JJRM, Visser RF, Klomps HC, Hoffman JJML. El Deeb MF, El Gamal MIH. Comparison of intravenous anisoylated plasminogen streptokinase activator complex and intracoronary streptokinase in acute myocardial infarction. Am J Cardiol 1988;62:25 - 30. 15. Marder VJ, Rothbarth RL, Fitzpatrick PG, Francis CW. Rapid lysis of coronary artery thrombi with anisoylated plasminogen streptokinase activator complex: Treatment by bolus intravenous injection. Ann Intern Med 1986;104: 304-310. 16. Hanbücken FW, Schneider J, Giinzler WA, Friderichs E, Giertz H, Flohe-L. Selective fibrinolytic activity of recombinant non-glycosylated human pro- urokinase (single-chain urokinase-type plasminogen activator) from bacteria. Arzneim Forsch. Drug Res 1989;37:993- 997. 17. Söhngen W, Mickelson JK, Simpson PJ, Lucchesi BR. Recombinant single- chain urokinase plasminogen activator (rscu-PA) induces thrombolysis and systemic fibrinolysis in a canine model of coronary artery thrombosis. Thromb Res 1988; 51:63-74. 18. Van de Werf F, Jang IK, Collen D. Thrombolysis with recombinant human single-chain urokinase-type plasminogen activator (rscu-PA): Dose-response in dogs with coronary artery thrombosis. J Cardiovasc Pharmacol 1987;9:91 - 93. 19. Flameng W, Vanhaecke J, Stump DC, Van de Werf F, Holmes W, Giinzler WA. Coronary thrombolysis by intravenous infusion of recombinant single chain urokinase-type plasminogen activator or recombinant urokinse in baboons: Effect on regional blood flow, infarct size and hemostatis. J Am Coll Cardiol 1986;8:118-124. 20. Van der Werf F, Vanhaecke J, De Geest H, Verstraete M, Collen D. Coronary thrombolysis with recombinant single chain urokinase-type plasminogen activator in patients with acute myocardial infarction. Circulation 1986;74:1066 -1070. 21. Diefenbach C, Erbel R, Pop T, Mathey D, Schofer Y, Hamm С. Recombinant single-chain urokinase-type plasminogen activator during acute myocardial infarction. Am J Cardiol 1988;61:966 - 970. 22. Tebbe U, Windeier J, Boesl J, Hoffman H, Wojcik J, Ashmawy M. Thrombolysis with recombinant unglucosylated Single-Chain Urokinase-Type Plasminogen activator (Saruplase) in acute myocardial infarction: Influence of heparin on early patency rate (AIMITS Study) J Am Coll Cardiol 1995;26:365 - 373.
32 Sarupiase in AMI
23. Bär FW, Meyer J, Michels R, Uebis R, Gange S, Barth H. The effect of taprostene in patients with acute myocardial infarction treated with thrombolytics: Results of the START study. Eur Heart J 1993;14:1118-1126. 24. Schneider J. Interactions of saruplase with acetylsalicylic acid, heparin, glyceryl trinitrate, tranexamic acid and aprotinin in a rabbit pulmonary thrombosis model. Arzneim Forsch Drug Res 1990;40(II): 1180 - 1184. 25. Schneider J. Heparin and the thrombin inhibitor argatroban enhance fibrinolysis by infused or bolus-injected saruplase (r-scu-PA) in rabbit femoral artery thrombosis. Thromb Res 1991;64:677 - 689. 26. Barth H, Lintz W, Michel G, Osterloh G, Seipp U, Flohe-L. Inhibition of platelet aggregation by intravenous administration of the biochemically stable prostacyclin analogue CG4203 in man. Naunyn Schmiedebergs Arch. Pharmacol 1983;(Suppl 324). 27. Groves R, Schneider J, Flohé L. Cooperative action of the prostacyclin analogue taprostene and the fibrinolytic prourokinase (r-scu-PA) in experimental artery thrombosis. In: Schrör К, Sinzinler M, eds. Prostaglandins in Clinical Research: New York: Alan R. Liss 1989:603- 607. 28. Flohé L, Böhlke H, Frankus E, Kim SMA, Gintz W, Goschen G. Designing prostacyclin analogues. Drug Res 1983;33: 1240. 29. Kaliman J, Fitscha P, Barth H, Sinzinger H. Clinical and platelet effects of CG 4203, a stable PGI2, analogue. In: Schör К, Sinzinger M, eds. Abstracts of the 6th International Conference on Prostaglandins and Related Compounds, Florence, Italy, June 3-6, New York: Alan R. Liss, 1986, ρ 443. 30. PRIMI Study Group. Randomized double-blind trial of recombinant pro urokinase against streptokinase in acute myocardial infarction. Lancet 1989;1:863-886. 31. Hoffmann JJML, Vijgen M, Nieuwenhuizen A. Comparison of the specificity of four fibrinogen assays during thrombolytic therapy. Fibrinolysis 1990;4:121 - 123. 32. Michels R, Hoffmann H, Windier J, Barth H, Hopkins G· A double-blind multicentre comparison of the efficacy and safety of saruplase and urokinase in the treatment of acute myocardial infarction. Report of the SUT AMI study group. J Thrombosis and Thrombolysis 1995;2:117 - 124. 33. The Belgian Saruplase Alteplase Trial Group. Effects of alteplase and saruplase on hemostatic variables: A single- blind, randomized trial in patients with acute myocardial infarction. CorArtDis 1991;2:349-355. 34. The SESAM Investigators. Early patency and reocclusion in acute myocardial infarction. A comparison between the thrombolytic agents saruplase and alteplase. Results of the SESAM trial. J Am Coll Cardiol 1994;Feb:345 (abstract). 35. Vermeer F, Bär FW, Windeler J, Schenkel W. Saruplase, a new fibrin specific thrombolytic agent. Final results of the PASS study (1698 patients) (abstract). Circulation 1993; 88:292.
33 Chapter 2
36. Hatzler GO, Rutherford BD, McConahay DR, Johnson WL, McCallister BD, Gura GM. Percutaneous transluminal coronary angiography with and without thrombolytic therapy for treatment of acute myocardial infarction. Am Heart J 1983; 106:965-973. 37. Simoons ML, Serruys PW, VD Brand M, Fioretti P. Thrombolysis in acute myocardial infarction: Limitation of infarct size and improved survival. J Am Coll Cardiol 1986;7: 717 - 728. 38. Hogg KJ, Gemmili JD, Burns JM, Gibson WK, Rae AP, Dunn G. Angiographic patency study of anistreplase versus streptokinase in acute myocardial infarction. Lancet 1990; 335:254-258.
34 CHAPTER 3
A Double-Blind Multicenter Comparison of the Efficacy and Safety of Saruplase and Urokinase in the Treatment of Acute Myocardial Infarction: Report of the SUTAMI Study Group
Rolf Michels,1 Hans Hoffmann,' Jürgen Windeler,2 Hannes Barth,3 and Gwyn Hopkins3 on Behalf of the SUTAMI Investigators
'Department of Cardiology, Catharina Hospital, Eindhoven, The Netherlands, 'Department of Biostatics, Ruhr University, Bochum, Germany, Grunenthal GmbH, Department of Clinical Research, Aachen, Germany
Abstract. Background Urokinase or two-chain urokinase-type plasminogen activator has been shown to be effective in the treatment of acute myocardial infarction Its parent molecule, single-chain urokinase-type plasminogen activator (scu-PA), unlike urokinase, can selectively activate fibrin- bound plasminogen The induced clot lysis is amplified by plasmin-tnggered conversion of scu-PA to urokinase and by further plasmin generation The aim of our study was to compare the efficacy and safety of recombinant unglycosylated scu-PA, or saruplase, and urokinase at doses considered optimal inpatients with acute myocardial infarction withm 6 hours of onset of pain Methods and results In a double-blind trial 543 patients were randomized to saruplase (20 mg bolus + 60 mg/hr) or urokinase (I 5 million unit bolus + 15 million umts/hr) Primary endpoint The patency rates at 24- 72 hours were 75 4% (95% CI 70 3-80 5%) for saruplase and 74 2% (95% CI 69 0-79 4%, Ρ = 0 77) for urokinase Secondary endpoint The incidence of bleeding events in both groups was 10 7% There were three hemorrhagic strokes in the saruplase group (ns) Other efficacy and safety evaluations Apart from the generation of more fibrinogen degradation products under saruplase, the changes in hemostatic parameters did not differ Hospital mortality was 4 4% for saruplase and 8 l%for urokinase This nonsignificant difference was maintained for 1 year Conclusion The efficacy and safety of saruplase and urokinase m the regimens used are very similar
Key Words, saruplase, urokinase, myocardial infarction
35 Chapter 3
Early myocardial infarction trials, where two-chain urokinase-type plasminogen activator (urokinase) was given over a long period of time, had disappointing results [1 - 3]. Later trials with an intravenous bolus of 2 million units demonstrated the drug to be safe and effective in achieving patency [4] and in improving early cardiac performance [5]. More recently an intravenous bolus of 1.5 million units, followed by an infusion of 1.5 million units in 1 hour, resulted in a patency similar to that of t-PA (alteplase) and appeared equally safe despite causing more hemostatic changes [6]. In 1978 Bernik postulated the presence of an inactive proenzyme (UK precursor) that can be activated to urokinase by a number of enzymes [7]. This precursor, now identified as single-chain urokinase- type plasminogen activator (scu-PA), can be activated by plasmin to full proteolytic activity [8,9]. Later it was shown to be fibrin specific and to be able to lyse clots without causing much systemic fibrinogenolysis [10]. In contrast to t-PA, its fibrin specificity is not the result of direct fibrin binding [10 - 13]. Scu-PA has intrinsic plasminogen activation potential, which in vivo is focused on the plasminogen bound to the fibrin clot [13 - 18]. The plasmin formed locally splits scu-PA into two-chain urokinase. The focal conversion further amplifies plasmin generation and thereby supports the lytic effect. Clinical studies confirmed that thrombolysis can be achieved without substantial fibrinogenolysis in vivo [17]. Experiments indicated that unglycosylated recombinant scu-PA (ΓΝΝ saruplase) could be given as an intravenous bolus, followed by a 1 hour infusion [19]. Two dose-finding trials indicated that the best reperfusion results were to be expected with a 20 mg bolus and a 60 mg infusion [20-22]. This dose caused more systemic lysis but no more bleeding complications. The PRIMI study, a double-blind comparison of saruplase and streptokinase, demonstrated a higher patency for saruplase at 60 minutes, with less hemostatic disturbances [23]. It then became interesting to compare urokinase to its unglycosylated parent single-chain molecule, and so the SUTAMI trial was designed to compare the safety and efficacy of saruplase and urokinase in the treatment of acute myocardial infarction.
36 Report of the SUT AMI study group
Methods
Study design The study was prospective, randomized, double-blind, double dummy, parallel group, and multicenter in design. The primary endpoint was the patency of the infarct-related artery 24 - 72 hours after the start of therapy. The secondary endpoint was the incidence of bleeding complications. Other efficacy and safety evaluations were hemostatic changes, mortality rate, reinfarction rate, angina score, and heart failure. A subsidiary angiographic endpoint was ejection fraction.
Patients Inclusion criteria consisted of typical chest pain for at least 30 minutes, ST-segment elevation of at least 0.3 mV in two or more precordial leads, or ST-segment elevation of at least 0.2 mV in two or more frontal plane leads, start of therapy within 6 hours onset of chest pain, age 20 - 75 years, and informed consent. Patients with a previous infarction, longer than 6 months previously, and not in the same region, could also be included. The exclusion criteria were the generally accepted contraindications for thrombolytic therapy [24]. The protocol was approved by the institutional ethics committees, and by the Central Ethics and Advisory Board. The start of experimental medication was taken as irrevocable entry into the trial.
Investigatory study medication Saruplase is the full-length unglycosylated human recombinant single- chain urokinase-type plasminogen activator obtained from genetically engineered Escherichia coli [25,26]. Saruplase and placebo to saruplase were prepared by Grünenthal GmbH, Aachen, Germany, and were provided in vials containing 10 mg of lyophilized saruplase plus stabilizing ingredients, or the stabilizing ingredients alone. Urokinase and placebo to urokinase were supplied by Behringwerke AG, Marburg, Germany, and were provided in vials containing 600,000 Ш lyophilized urokinase plus stabilizing ingredients, or the stabilizing ingredients alone.
37 Chapter 3
Treatment After informed consent all patients started with a nitroglycerin infusion of 3 mg/hr unless contraindicated. No heparin was given before thrombolysis. First urokinase, 1.5 million IU/25 ml, or its placebo was given over 5 minutes. Then saruplase, 20 mg/20 ml, or its placebo was given over 2 minutes, immediately followed by a parallel, double infusion of saruplase (60 mg/1 hr) and placebo to urokinase, or urokinase (1.5 million Ш/1 hr) and placebo to saruplase. Thirty minutes after the end of thrombolytic therapy, an intravenous heparin infusion was started and continued until control angiography between 24 and 72 hours. The initial dose of 15 IU/kg/hr was titrated against the activated partial thromboplastin time (APTT) to obtain a value of 1.5 - 2.5 times the control. Other medication was given if necessary.
Electrocardiography The patients were included in the study on the basis of a 12-lead ECG recorded immediately after admission. The admission ECGs were assessed centrally to verify the inclusion criteria.
Angiography Ventriculography and coronary angiography were performed 24 - 72 hours after the start of thrombolytic therapy. Ventriculography was performed first. The presumptive infarct-related artery was to be visualized last. The films were assessed by the local investigator and centrally by a team of experienced coronary angiographers, who were blinded to treatment assignment. The central assessment was compared with the assessment of the investigator. If consensus could not be reached, two independent experts made the final decision. The coronary angiograms were scored segment by segment according to the code of the National Heart, Lung and Blood Institute, Coronary Artery Surgery Study (CASS) [27]. Patency was classified according to the definitions in the Thrombolysis In Myocardial Infarction (TIMI) study [28]. TIMI grades 2 and 3 were classified as open vessels.
Bleeding complications and other adverse events Questioning on the presence of adverse events was at fixed time points during the hospital stay and at follow-up. Blood pressure and heart rate were recorded on admission; at 30, 60, and 120 minutes after the start of thrombolytic therapy; and at discharge. Special questioning was
38 Report of the SUT AMI study group made for bleeding, stroke, heart failure, arrhythmias, postinfarct angina, and reinfarction. Bleeding complications were classified as severe when active measures were required to stop the bleeding, such as the use of aprotinin, surgical procedures, or a transfusion of > 500 ml whole blood or blood products. A neurologist was to decide whether a stroke was hemorrhagic or ischemic.
Blood sampling Blood was sampled through an indwelling 19 gauge needle exclusively for this purpose, in the arm opposite to that used for study medication administration. Hematology and chemistry were analyzed on admission and daily for 3 days at the local laboratory. Cardiac enzymes were determined on admission, twice daily for the first 3 days and once daily for the next 2 days, at local laboratories.
Hemostatic parameters Blood samples for the analysis of fibrinogen, fibrinogen degradation products, plasminogen, and o^.antiplasmin concentration were taken on admission and at 1, 2, 6 - 12, 24 - 36 hours (before angiography), and 60 - 72 hours (after angiography). A special kit was provided for uniform handling. At each time point, two 9 ml samples were collected. One sample was drawn into citrate, and the other into citrate supplemented with aprotinin. The samples were placed on melting ice and were spun as soon as possible. The plasma samples were stored at - 20°C until transfer to the central laboratory, where they were stored at - 70°C; each sample was thawed only once. Fibrinogen was determined by the Clauss method; fibrinogen degradation products were measured by ELISA and a2-antiplasmin and antithrombin III by functional, proteinase quenching capacities. The presence of antibodies against saruplase and E. coli protein, a trace contaminant of saruplase, was examined by the sensitive ELISA technique in citrated plasma samples from blood taken at day 7 - 14 by comparing results with pretreatment samples.
Follow-up Follow-up was scheduled at 3, 6, and 12 months. Angina was graded using the Canadian Cardiovascular Society scoring system [29]. Heart
39 Chapter 3 failure was graded using the criteria of the New York Heart Association [30].
Statistics The sample size was based on a presumptive patency of 63% for urokinase. For a clinically relevant difference of 12% to be detected, based on Fisher's exact test with α at 0.05 (two sided) and β at 0.2, 498 patients were required. Data were collected and analyzed at the independent Centre for Statistics (Bochum). Errors were discussed at steering committee meetings. The primary endpoint analysis was performed on an intent-to-treat basis. Therefore all patients who received experimental medication were included in the data analysis. Patients who did not have the investigatory angiogram within the scheduled time frame were regarded as "closed vessel" patients, as were all patients who had coronary interventions before the scheduled angiogram. Patency data were analyzed by the use of Fisher's exact test. For safety surveillance, mortality and major bleeding events were analyzed sequentially, using pairs of patients within each center. Dichotomous variables were analyzed by the chi-square test, except stroke incidence, for which Fisher's exact test was used because of low patient numbers. The unpaired t-test was used for the comparison of continuous variables, given here as the mean and standard deviation unless otherwise stated. Analysis of variance with repeated measures was performed for the comparison of hemostatic parameters, given here as median concentrations. All p-values given are two sided. The Central Ethics and Advisory Board met for a semiblinded interim analysis after inclusion of 200 patients. There was no indication for prematurely stopping the study.
Results
Protocol compliance and baseline characteristics Informed consent was obtained in 557 patients from 23 centers in four European countries, but therapy was not started in 14 because of premature death (2), exclusion criteria (9), technical problems (2), and ECG normalization (1). Five hundred and forty-three patients entered the trial between July 1988 and February 1990, and 531 met all
40 Report of the SUTAMI study group inclusion criteria. Because of the intent-to-treat design, all 543patients who received experimental medication were included in the data analysis.
Baseline characteristics Two hundred and seventy-two patients received treatment with saruplase, and 271 received urokinase. The two groups were similar with respect to age, sex, height, weight, blood pressure, and time between onset of symptoms and start of infusion but were different with respect to infarct site (anterior infarction 40.4% for saruplase and 50.2% for urokinase; ρ = 0.02) and pulse rate (72.3 ± 14.8 beats/min for saruplase and 76.1 ± 17.6 beats/min for urokinase, ρ < 0.01).
Table 1. Baseline characteristics before randomization Saruplase Urokinase 90% confídence limits of difference
Number of patients (n) 272 271 Age (yr) 58.6 ±9.5 59.0 ±9.7 -1.75;0.96 Sex, male (%) 82.7 81.5 -4.20;6.60 Height (cm) 172.8 ± 8.4 173.1 ±8.1 -1.47;0.87 Weight (kg) 76.8 ± 11.1 76.9 ±12.1 -1.74;1.54 Anterior MI (%) 40.4 50.2 -16.8;-2.8 Systolic BP (mm Hg) 135.2123.5 134.5 ±24.5 -2.70;4.10 Diastolic BP (mm Hg) 83.9 ±14.1 83.6 ±15.2 -1.77;2.37 Pulse rate (/min) 72.3 ± 14.8 76.1 ±17.6 -6.10;-1.50 Time to admission 1:52 ± 1:10 1:51 ±1:00 -0.08;0.10 (hr:min) Time to infusion (hnmin) 2:46 ±1:15 2:44 ±1:07 -0.08;0.12 FgDP (all samples, mg/L) 0.52 ±0.59 0.68 ±1.14 -0.29;-0.03
FgDP =fibrinogen degradatio n products; Ml = myocardial infarction; BP = blood pressure. Where applicable values are given as means ± standard deviations.
41 Chapter 3
Also the baseline fibrinogen degradation products differed slightly (saruplase 0.52 ± 0.59 mg/1 and urokinase 0.68 +· 1.14 mg/l, ρ = 0.047). Myocardial infarction was confirmed by central ECG verification in 93.6% of the patients and by enzyme findings in 97.8%. Only one patient had neither ECG nor enzyme confirmation.
Angiography The investigatory angiogram was performed in 96.0% (261/272) of patients in the saruplase group and in 92.6% (251/271) of patients in the urokinase group. The reasons for no control angiography are listed in Table 2. The median time to angiogram was 45:13 hours for saruplase and 44:30 hours for urokinase. The angiograms were outside the time window set for analysis in three saruplase and in four urokinase patients. There was no difference in the patency of the infarct-related artery, which was open in 205 of 272 saruplase patients (75.4%, 95% confidence limits 70.3 - 80.5%) and in 201 of 271 urokinase patients (74.2%, 95% confidence limits 69.0 - 79.4%; ρ = 0.77). Table 2. Reasons for no control angiogram at 24 - 72 hours
Saruplase (n) Urokinase (n) Death 2 13 Cardiac problems 3 2 Cerebral bleeding 2 0 Intervening angiogram 3 1 Intervening CABG 1 0 Other 0 4 Total 11 * 20
CABG = coronary artery bypass graft.
Per protocol analysis (only timely investigatory angiograms) also showed no difference: 205 of 258 saruplase patients (79.5%, 95% confidence limits 74.5 84.4%), and 201 of 247 urokinase patients (81.4%, 76.5 - 86.2%; ρ = 0.65) had an open, infarct-related artery.
42 Report of the SUTAMI study group
Mean ejection fraction was 55.2 ± 12.3% for saruplase and 53.4 ± 12.1% for urokinase (p = 0.09). Ejection fraction was 55.3 ± 12.1% for open vessels compared with 49.9 ± 11.9% for closed vessels (p < 0.001).
Bleeding events Twenty-nine patients in each group (10.7%) had one or more bleeding events (Table 3). Including hemorrhagic strokes (three patients), bleeding was classified as severe in nine cases with saruplase and in seven cases with urokinase.
Table 3. Bleeding events recorded in hospital before discharge Saruplase Urokinase All Severe All Severe Puncture site bleed 15 4 15 3 Hematuria 1 0 2 0 Hematemesis 4 1 5 1 Melena 1 0 3 1 Hemorrhagic stroke 3 3 0 0 Other 7 1 7 2 Total events (n) 31 9 32 7 Patients with events {iU%) } 29(10.7) 29(10.7)
More than one event per patient is possible.
Strokes All three hemorrhagic strokes (1.1%, ρ = 0.25) occurred in saruplase patients, and as a consequence two of these patients died. The diagnosis was confirmed by computed tomography (CT) scan in two patients. All three occurred in elderly men (66, 68, and 70 years) and within 48 hours (16, 29, and 43 hours). Six nonfatal ischemic strokes were reported, with four (1.5%) in the saruplase group (one a mild transient ischemic attack). Two (0.7%) ischemic strokes were reported in the urokinase group (p = 0.69), one of which was fatal. The diagnosis of ischemic stroke was confirmed by CT scan in three patients.
43 Chapter 3
Mortality Thirty-four patients died during the hospital phase (6.3%). Hospital mortality was higher (8.1% vs. 4.4%, ρ = 0.08) and earlier (median 2 days vs. 9 days) for urokinase patients. In the saruplase group eight patients died of cardiac causes, two from hemorrhagic stroke, one of vascular renal failure, and one of sepsis. In the urokinase group 21 patients died of cardiac causes and 1 from an ischemic cerebrovascular infarction. Cardiogenic shock occurred in 9 (3.3%) saruplase patients and was fatal in 4, whereas of the 13 (4.8%) urokinase patients with cardiogenic shock, 11 died.
Reinfarction Reinfarction was documented in 32 patients: 19 (7.0%) in the saruplase group and 13 (4.8%) in the urokinase group (p = 0.28) after 4 days (median). Reinfarction was fatal in two saruplase patients and four urokinase patients.
Postinfarct unstable angina Postinfarct unstable angina was observed in 83 patients: 36 (13.2%) in the saruplase group and 47 (17.3%) in the urokinase group.
Clinical events Saruplase patients had more episodes of transient hypotension (31.6% vs. 21.4%, ρ < 0.01). This difference was already present during the thrombolytic infusion (22.1% vs. 13.7%); however, no hypotension due to allergy was reported in the saruplase group. The need for balloon angioplasty or bypass surgery did not differ between the groups. All clinical events are summarized in Table 4.
Hemostatic parameters The time from the start of infusion to blood sampling was almost identical for the two treatment groups. Only complete sample series, available in 467 patients (86%), are reported here (Figure 1). All concentrations given are median values. The hemostatic changes caused by saruplase and urokinase were virtually identical, except for the fibrinogen degradation products. Fibrinogen dropped after
44 Report of the SUT AMI study group saruplase from 3.00 g/1 before treatment (0 hours) to a minimum of 0.35 g/1 at 2 hours (urokinase: 3.20 g/1 and 0.44 g/1).
Table 4. Clinical events before hospital discharge Sarupllas e Urokinase
00 (%) (n) (%) Ρ
Bleeding events (pts) 29 10.7 29 10.7 >0.50 Hemorrhagic stroke 3 1.1 - - 0.25 Ischemic stroke 4 1.5 2 0.7 >0.50 Mortality 12 4.4 22 8.1 0.08 Cardiogenic shock 9 3.3 13 4.8 0.40 Reinfarction 19 7.0 14 5.2 0.47 Unstable angina 36 13.2 47 17.3 0.19 Transient hypotension 86 31.6 58 21.4 <0.01 PTCA 31 11.4 29 10.7 >0.50 CABG 10 3.7 13 4.8 >0.50 Allergic reactions - - 2 0.7 0.25
Transient hypotension is defined as systolic blood pressure < 90 mmHg. CABG = coronary artery bypass graft; PTCA = percutaneous transluminal coronary angioplasty.
Recovery to baseline did not occur until 36-72 hours. Global fibrinogen was higher in patients with closed vessels than in patients with open vessels (p < 0.01). There was a difference in fibrinogen at 1 hour between patients with bleeding events and those without bleeding (0.32 ± 0.28 g/land 0.64 ± 0.67 g/1, respectively; ρ < 0.03); this was not related to the type of thrombolytic drug used. Fibrinogen degradation products rose markedly after saruplase from 0.43 mg/1 at 0 hours to 160 mg/1 at 2 hours and were still elevated to 0.67 mg/L at 36 - 72 hours, whereas the increase after urokinase (from 0.45 mg/1 to 89.0 mg/1) was significantly smaller (p < 0.001). A greater variation of the fibrinogen degradation products was seen for saruplase (25% and 75% quintiles at 1 hour: 24 mg/1 and 353 mg/1) when compared with urokinase (44 mg/1 and 190 mg/1).
45 Chapter 3
Oo-antiplasmin was not detectable at 1 and 2 hours after either saruplase or urokinase. Full recovery was observed at 36 - 72 hours. Plasminogen fell from 0.97 U/ml at 0 hours to a minimum of 0.32 U/ml at 2 hours for saruplase (urokinase 0.97 U/ml and 0.34 U/ml), and recovery to baseline had not taken place at 36 - 72 hours (saruplase 0.87 U/ml, urokinase 0.89 U/ml).
Antibodies No antibody against saruplase was detected in the samples of 241 saruplase-treated (or of 232 urokinase-treated patients) that could be analyzed. Antibodies against E. coli protein were also not detected in 19 randomly selected samples from saruplase treated patients.
Fig. 1. Fibrinogen degradation products (median, mg/L)
0 1 2 6-12 24-36 36-72 h
—·—sarLplase 0 43 0 43 0 43 0 43 0 43 0 43 --o—LTokinase 0 45 0 45 0 45 0 45 0 45 0 45
Fig.1. Analysis of variance for the fibrinogen degradation product concentrations.
46 Report of the SUTAMI study group
Table 5. One year follow-up, depicting total mortality, and infarction rate after discharge Saruplase Urokinase
(n) (%) (n) (%) Ρ
Total mortality 20 7.5 31 11.7 0.1 Infarction after 16 6.3 17 7.1 >0.5 discharge
One year follow-up Four hundred and eighty patients attended the 12 month follow-up (Table 5). Thirty-four patients had already died in hospital, and 18 more (saruplase η = 8, urokinase η = 10) died during follow-up. Eleven patients (saruplase η = 5, urokinase η = 6) were lost to follow-up. Total mortality at 12 months was 7.5% (20/267) for saruplase and 11.7% (31 of 265, ρ = 0.10) for urokinase (one more patient died shortly after 12 months). The 12 month reinfarction rate after discharge was 6.3% for saruplase and 7.1% for urokinase, and four of these reinfarctions (saruplase η = 1, urokinase η = 3) were fatal. At 12 months, 31.7% of patients reported angina, and 14.2% reported heart failure. No difference was observed between the two treatment groups.
Discussion
The primary endpoint, patency at 24 - 72 hours, was not different for the two drugs. The patency rate for saruplase (75.4%) was in the range predicted at the start of the study, but urokinase showed a higher patency (74.2%) than expected from earlier trials (4,6). Further subdivision of patency in TIMI 2 and TIMI 3 does not alter the results. Only 11 patients on saruplase and 12 on urokinase showed TIMI 2 flow. With hindsight the assumption that late patency, like early patency, may differ between plasminogen activators was wrong. However, at the time of the conception of this trial, the "catch up phenomenon" [31] was not yet fully appreciated. At that time the clinical rule of conduct, to shun early angiography, was based on the
47 Chapter 3 outcome of three separate trials, which showed no benefit of early invasive strategy [32 - 34]. By following this course we have missed the opportunity to show the relevance of the intrinsic fibrin-specific activity of saruplase [16]. However, the dose-finding trials did show a short mean time delay to reperfusion [20 - 21], and the PRIMI trial demonstrated a maximum reperfusion plateau within 60 minutes [23]. The incidence of bleeding complications was low and identical (10.7%) in both treatment groups. As early catheterization contributes to bleeding complications [32 - 34], it is no surprise that we found a lower incidence in comparison with trials with early angiography [23]. In our study three patients (1.1%) had a hemorrhagic stroke with saruplase: A comparable incidence was found in the PRIMI trial (1.0%). In an open label study with saruplase in 1698 patients, the occurrence of hemorrhagic stroke was 0.5%, not different from t-PA and anistreplase [35 - 37]. The hemostatic changes were quite similar; only the degree of generation of fibrinogen degradation products differed significantly. This substantial fibrinogen degradation was not seen in animal experiments [17 - 19,38,39] but has been found in other clinical trials [23,40] and may be due to conversion of saruplase to a two-chain urokinase-type plasminogen activator, as has been confirmed in a recent pharmacokinetic study that shows conversion of approximately 30% of 80 mg saruplase [41]. We found a nonsignificant but conspicuous difference for total mortality. Exploratory subgroup analysis, which was not prespecified, of the cardiac mortality showed eight cardiac deaths in the saruplase group and 21 in the urokinase group (p = 0.01). Cardiac mortality was associated with a high pretreatment pulse rate (p < 0.01) and with anterior infarction (p = 0.05). The asymmetrical distribution of anterior infarctions between the two treatment groups does not help explain the difference in the mortality, as the mortality rates for anterior infarction reflect the overall rates: 5.5% (6 of 110) for saruplase and 11.0% (15 of 136) for urokinase. Intent-to-treat analysis showed a patency of 82.7% (91 of 110) for anterior infarctions in the saruplase group compared with 73.5% (100 of 136) in urokinase patients. These mortality differences are likely to be artifacts of the methodology, but they might indicate earlier maximal reperfusion, a point that we failed to prove by study design, but a hypothesis that deserves further investigation.
48 Report of the SUT AMI study group
Conclusions
The efficacy and safety profiles of 80 mg saruplase and 3 million units urokinase are very similar. Saruplase causes no antibody formation. Saruplase generates more fibrinogen degradation products. Like other fibrin-specific agents, saruplase treatment appears efficacious at dose levels that induce substantial systemic lysis. In this trial a better survival outweighs a higher hemorrhagic stroke rate. Further investigation should concentrate on reperfusion or early patency, especially in large anterior infarctions and other high-risk subsets, where rapid reperfusion has the greatest advantage.
Acknowledgments
We thank Anne Hoi and Monique van den Broek for their secretarial contribution to the preparation of the manuscript This study was sponsored by Grunenthal GmbH, the manufacturer of saruplase, and was cosponsored by Behringwerke, the manufacturer of urokinase.
Study Organization
Participating centers. Universitätskrankenhaus Eppendorf, Hamburg (W. Bleifeldt, Chr. Hamm); Evangelisches Krankenhaus, Oberhausen (R. Oberheiden); Robert-Bosch-Krankenhaus GmbH, Stuttgart (P. Heimburg); Marienhospital, Wesel (K. Haerten); Krankenhaus der Stadt Wien/Lainz, Wien (W. Enenkel); Kardiologische Universitätsklinik, Wien (P. Probst); Rudolf- Stiftung, Wien (C. Stollberger); Middelheim Ziekenhuis, Antwerpen (R. Ranquin); Hospital de la Citadelle, Liège (J. Boland); Sint Lucas Ziekenhuis, Amsterdam (I. Hellemans); Onze Lieve Vrouwe Gasthuis, Amsterdam (F. Kiemeneij); R. de Graaf Ziekenhuis, Delft (A. Withagen); Refaja Ziekenhuis, Dordrecht (J. Ten Kate); St. Joseph Ziekenhuis, Veldhoven (A. Bosma); Diaconessenhuis, Eindhoven (L. Relik-van Wely); St. Annaziekenhuis, Geldrop (W. van Ekelen); Ziekenhuis Tjongerschans, Heerenveen (G. Jochemsen); St. Lambertos Ziekenhuis, Helmond-Deurne (P. Bendermacher); Maria Ziekenhuis, Tilburg (P. Lindner); St. Elisabeth Ziekenhuis, Tilburg (P. Zijnen); St. Jans Gasthuis, Weert (Η. Klomps); Sophia Ziekenhuis, Zwolle (R. Enthoven); St. Franciscusziekenhuis, Roosendaal (R. Bos)
49 Chapter 3
Study Chairman. R. Michels, Catharina Hospital (Eindhoven)
Central Ethics and Advisory Board. J. Meyer, Mainz (Chairman); F. Vermeer, Maastricht; J. Bonnier, Eindhoven; R. Vosveld, Eindhoven; E. Lesaffrc, Antwerpen; Ζ. Suyker, Tilburg
Center for Statistics. H. J. Trampisch, J. Windeier, M. Vieil, Bochum
Angiographie Core Laboratory. R. Michels (Eindhoven)
Second Angiographie Core Laboratory. D. Mathey, J. Schofer (Hamburg)
Hemostatic Core Laboratory. H. Hoffmann (Eindhoven)
Antibody Test Laboratory. H. Beier, Grünenthal GmbH (Aachen)
Data and Plausibility Check. F. Vermeer (Maastricht)
50 Report of the SUT AMI study group
References
1. A European Collaborative Study. Controlled trial of urokinase in myocardial infarction. Lancet 1975;2:624 - 626. 2. Gormsen J, Tidstrom B, Fedderson C, et al. Biochemical evaluation of low dose urokinase in acute myocardial infarction. A double-blind study. Acta Med Scand 1973;194: 191-198. 3. Brochier M, Raynaud R, Planiol T, et al. The treatment of myocardial infarction and impending myocardial infarction with urokinase: Randomized study of 120 patients. Arch Mal Coeur 1975;68:563-569. 4. Mathey D, Schofer J, Sheehan F, et al. Intravenous urokinase in acute myocardial infarction. Am J Cardiol 1985;55: 878-882. 5. Penco M, Fedele F, Agati L, et al. Effects of systemic treatment with urokinase (UK) on left ventricular function, (abstr). Eur Heart J 1987;8(Suppl2):24. 6. Neuhaus K, Tebbe U, Gottwick M, et al. Intravenous recombinant tissue plasminogen activator (rt-PA) and urokinase in acute myocardial infarction: Results of the German Activator Urokinase Study (GAUS). J Am Coll Cardiol 1988;12:581-587. 7. Bemik M, Oiler P. Increased plasminogen activator (urokinase) in tissue culture after fibrin deposition. J Clin Invest 1973;52:823 - 834. 8. Wun T, Ossowoski L, Reich E. A proenzyme form of urokinase. J Biol Ckem 1982;257:7262-7268. 9. Gurewich V, Pannell R. Fibrin-specificity and efficacy of proteolysis induced by single-chain urokinase in plasma. Thromb Haemost 1983;50:321. 10. Gurewich V, Pannell R, Louie S, et al. Effective and fibrin-specific clot lysis by a zymogen precursor form of urokinase (pro-urokinase). J Clin Invest \Ш\1Ъ:Ш\-Ш9. 11. Gurewich V, Pannell R, Greenlee R, et al. The fibrin specificity of single- chain urokinase is not dependent of fibrin binding. Thromb Haemost 1983;50:386. 12. Stump D, Thienpont M, Collen D. Urokinase-related proteins in human urine. JDiolChem 1986;261:1267-1273. 13. Lijnen H, Zamarron С, Blaber M, et al. Activation of plasminogen by Pro- urokinase, I. Mechanism. J Biol Chem 1986;261:1253-1258. 14. Gurewich V. The sequential, complementary and synergistic activation of fibrinbound plasminogen by tissue plasminogen activator and pro- urokinase. Fibrinolysis 1989;3: 59-66. 15. Gurewich V. Fibrinolytic properties of single-chain urokinase plasminogen activator and how they complement those of tissue plasminogen activator.
51 Chapter 3
In: Haber E, Braunwald E, eds. Thrombolysis. Basic Contributions and Clinical Progress. St. Louis, MO: Mosby Year Book, 1991:51-60. 16. Pannell R, Gurewich V. Pro-urokinase - a study of its stability in plasma and of a mechanism for its selective fibrinolytic effect. Blood 1986;67:1215-1223. 17. Collen D, Stassen J, Blaber M, et al. Biological and thrombolytic properties of proenzyme and active forms of human urokinase. III. Thrombolytic properties of natural and recombinant urokinase in rabbits with experimental jugular vein thrombosis. Thromb Haemost 1984;52:27 - 30. 18. Hanbücken F, Schneider J, Günzler W, et al. Selective fibrinolytic activity of recombinant human pro-urokinase (single-chain urokinasetype plasminogen activator) from bacteria. Drug Res 1987;37:993 - 997. 19. Flameng W, Vanhaecke J, Stump D, et al. Coronary thrombolysis by intravenous infusion of recombinant single-chain.urokinase-type plasminogen activator or recombinant urokinase in baboons: Effect on regional blood flow, infarct size and hemostasis. J Am Coll Cardiol 1986;8:118-124. 20. Van de Werf F, Vanhaecke J, De Geest H, et al. Coronary thrombolysis with recombinant single-chain urokinase-type plasminogen activator in patients with acute myocardial infarction. Circulation 1986;74:1066-1070. 21. Van de Werf F, Nobuhara M, Collen D. Coronary thrombolysis with human single-chain urokinase-type plasminogen activator (pro-urokinase) in patients with acute myocardial infarction. Ann Intern Med 1986;104:345-348. 22. Diefenbach C, Erbel R, Pop T, et al. Recombinant single-chain urokinase- type plasminogen activator during acute myocardial infarction. Am J Cardiol 1988;61:966-970. 23. Primi Trial Study Group. Randomized double-blind trial of recombinant pro-urokinase in acute myocardial infarction. Lancet 1989;1:863 - 868. 24. Genton E. Fibrinolytic therapy. In: van de Loo J, Prentice C, Beller F, eds. The Thrombotic Disorders. Stuttgart: Schattauer, 1983:192. 25. Holmes W, Pérmica D, Blaber M, et al. Cloning and expression of the gene for pro-urokinase in Escherichia coli. Bio- technology 1985;3:923 - 929. 26. Flohé L. Single-chain urokinase-type plasminogen activators: New hopes for clot-specific lysis. Eur Heart J1985;6: 905-908. 27. The principal investigators of CASS and their associates. The National Heart, Lung and Blood Institute Coronary Artery Surgery Study (CASS). Circulation 1981;63(Suppl I):I1-I8. 28. Chesebro J, Knatterud G, Roberts R, et al. Thrombolysis in Myocardial Infarction (TIMI) trial. Phase I: A comparison between intravenous tissue
52 Report of the SUTAMI study group
plasminogen activator and intravenous streptokinase. Circulation 1987;76:142-154. 29. Campeau L. Grading of angina pectoris. Circulation 1976; 54:522. 30. Criteria Committee, New York Heart Association. In: Diseases of the Heart and Blood Vessels. Nomenclature and Criteria for Diagnosis, 6th ed. Boston: Little, Brown, 1964:114. 31. Granger C, Califf R, Topoi E. Thrombolytic therapy for acute myocardial infarction. A review. Drugs 1992;44: 293-325. 32. Topol E, Califf R, George B, et al. A randomized trial of immediate versus delayed elective angioplasty after intravenous tissue plasminogen activator in acute myocardial infarction. N Engl J Med 1987;317:581 - 588. 33. Simoons M, Arnold A, Betriu A, et al. Thrombolysis with tissue plasminogen activator in acute myocardial infarction: No additional benefit from immediate percutaneous coronary angioplasty. Lancet 1988;1:197-203. 34. Rogers W, Bairn D, Gore J, et al. Comparison of immediate invasive, delayed invasive and conservative strategies after tissue-type plasminogen activator. Result of the Thrombolysis in Myocardial Infarction (TIMI) Phase II-A Trial. Circulation 1990;81:1457-1476. 35. Vermeer F, Bär F, Windeler J, et al. Saruplase, a new Fibrin Specific Thrombolytic agent; Final Results of the Pass Study (1698 patients) (abstr). Circulation 1993; 88:1560. 36. ISIS-3(Third International Study of Infarct Survival) Collaborative Group: A randomized comparison of streptokinase versus tissue plasminogen activator versus anistreplase and of aspirin plus heparin versus aspirin alone among 41,229 cases of suspected acute myocardial infarction. ¿аисеП992;339:753-770. 37. Erlemeier H, Zangemeister W, Burmester L, et al. Bleeding after thrombolysis in acute myocardial infarction. Eur Heart J\989;10:16 - 23. 38. Van de Werf F, Jang I, Collen D. Thrombolysis with recombinant human single-chain urokinase-type plasminogen activator (rscu-PA): Dose- response in dogs with coronary artery thrombosis. J Cardiovasc Pharmacol 1987;9:91 - 93. 39. Soehngen W, Mickelson J, Simpson PJ, et al. Recombinant single-chain urokinase-type plasminogen activator (rscu- PA) induces thrombolysis and systemic fibrinolysis in a canine model of coronary artery thrombosis. ThrombRes 1988;51:63-74. 40. Gulba D, Fisher K, Bartheis M, et al. Low dose urokinase preactivated natural рго-urokinase for thrombolysis in acute myocardial infarction. Am J Cardiol 1989;63:1025-1031. 41. Koster R, Cohen A, Hopkins G, et al. Thrombolysis with saruplase: Pharmacokinetics and conversion to urokinase- type plasminogen activator in acute myocardial infarction. J Am Coll Cardiol 1992;19:131A.
53 54 CHAPTER 4
Hemostatic changes after thrombolytic therapy with saruplase (unglycosylated single-chain urokinase-type plasminogen activator) and urokinase (two-chain urokinase-type plasminogen activator)
H. R. Michels, J. J. M. L. Hoffmann, J. Windeier and G. R. Hopkins
H, R Michels, Department of Cardiology, Catharina Hospital, Eindhoven, the Netherlands, J J M L Hoffman, Department of Clinical Laboratories, Hemostasis Division, Catharina Hospital, Eindhoven J Windeier, Department of Medical Informatics and Biomathics, Ruhr University, Bochum, Germany G R Hopkins, Department of Clinical Research, Aachen, Germany
Abstract: Urokinase, or two-chain urokinase-type plasminogen activator (tcu-PA), is an effective thrombolytic agent Its single-chain precursor (scu-PA), unlike tcu-PA, is able to selectively activate fibrin-bound plasminogen The induced clot lysis is amplified by plasmin-triggered conversion of scu-PA to tcu-PA The aim of our study was to compare the hemostatic effects of recombinant unglycosylated scu- PA, INN saruplase, and urokinase, at doses considered optimal (80 mg saruplase and 3 million units urokinase) in patients with acute myocardial infarction, within 6 h of onset of pain Complete sample sets from more than 230 patients in each treatment group have been analyzed The median hemostatic changes caused by saruplase and urokinase were virtually identical indicating that saruplase is converted early in vivo to its active tcu-PA derivative with the dose regimen used Fibrinogen degradation products were higher for saruplase with a maximum at 2 h They rose markedly after saruplase from 0 43 mg/l at 0 h to a maximum of 160 mg/l at 2 h, whereas the increase after urokinase (from 0 45 mg/l to 89 0 mg/l) was significantly smaller (P < 0 001)
Key words: Saruplase, urokinase, hemostatic parameters.
Introduction
Two-chain urokinase-type plasminogen activator (tcu-PA), or urokinase, is an effective thrombolytic agent. In patients with an acute myocardial infarction an intravenous bolus of 1.5 million units, followed by an
55 Chapter 4 infusion of 1.5 million units in 1 h, results in patency rates similar to those obtained with t-PA (alteplase).1 In contrast to alteplase, urokinase is not considered to be 'fibrin-specific', urokinase causes more systemic plasminogen activation than alteplase. However, despite causing more hemostatic changes, urokinase appears to be equally safe, and the systemic activation of plasminogen, rather than being a drawback, may help explain the low rate of reocclusion following urokinase. ',2 Single-chain precursor plasminogen activator (scu-PA) was initially described as pro-urokinase, a proenzyme, which could be activated by plasmin to the two chain form (tcu-PA) for full proteolytic activity.3,4 Later scu-PA was shown to be able to lyse clots without causing much systemic fibrinogenolysis. 5 Unlike t- PA, its fibrin specificity is not the result of direct fibrin binding,5'8 as scu- PA has an intrinsic plasminogen activation potential which in vivo is focused on the fibrin-bound plasminogen.8"12 The plasmin which is formed locally splits scu-PA into active tcu-PA. This focal conversion amplifies the plasmin generation, thereby supporting and enhancing the lytic effect. Eventually the conversion causes dissipation of plasminogen activation into the systemic circulation. Although thrombolysis can be achieved without substantial fibrinogenolysis in vivo,3,12'1'1 there is a concentration- dependent fibrinogenolysis in clinical studies which have assessed early and consistent patency in acute myocardial infarction.15'17 The degradation of fibrinogen in plasma is indicative of the extent of systemic conversion of scu-PA to tcu-PA. Saruplase is a full-length unglycosylated human recombinant scu-PA obtained from genetically engineered Es- cherichia coli.1819 Favourable reperfusion was achieved with saruplase given as a 20 mg bolus and 60 mg infusion for 60 min.15"17 The SUTAMI trial was designed to compare the efficacy and safety of saruplase and urokinase.20 This report communicates the changes in hemostatics observed in the plasma of patients enrolled in the study, during and after thrombolytic treatment.
56 Hemostatic changes after saruplase and urokinase
Materials and methods
Patients The study was a prospective, randomized, double-blind, parallel group, multicenter design. The sample size was based on the primary endpoint: the patency of the infarct related artery 24 - 72 h after start of therapy. The hemostatic changes were a secondary endpoint. Patients between 20 and 75 years, presenting within 6 h of the onset of chest pain, were included into the trial if they met the usual criteria.21 Saruplase and placebo to saruplase were prepared by Grünenthal GmbH, Aachen, Germany, and provided in vials containing 10 mg of lyophilized saruplase plus stabilizing ingredients, or the stabilizing ingredients only. Urokinase and placebo to urokinase were supplied by Behringwerke AG, Marburg, Ger many, and provided in vials containing 600 000 IU lyophilized urokinase plus stabilizing ingredients, or the stabilizing ingredients only. After informed consent all patients were started on a nitroglycerin infusion of 3 mg/h, unless contra- indicated. No heparin was given before thrombolysis. First urokinase, 1.5 million IU/25 ml, or its placebo was given over 5 min. Then saruplase, 20 mg/20 ml or its placebo was given over 2 min, immediately followed by a parallel double infusion of saruplase (60 mg/h) and placebo to urokinase, or urokinase (1.5 million IU/h) and placebo to saruplase. Thirty minutes after the end of thrombolytic therapy, an intravenous heparin infusion was started and continued until control angiography between 24 and 72 h. The initial heparin dose (15 IU/kg/h) was titrated against the activated partial thromboplastin time (APTT), to obtain a value of 1.5 - 2.5 times the control. Other medication was given if necessary. Blood was sampled through an indwelling 19-gauge needle exclusively for this purpose, from the arm opposite to that used for study medication administration. Blood samples for the analysis of fibrinogen, fibrinogen degradation products, fibrin degradation products, plasminogen and ot2-antiplasmin concentration were collected on admission and at 1, 2, 6 - 12, 24 - 36 h (before angiography), and 60 - 72 h (after angiography). A special kit was provided for uniform handling. At each time point, the first 2 ml of blood was discarded and two 9 ml samples were collected. One sample was drawn into citrate (0.01 M final concentration), the other into citrate
57 Chapter 4 supplemented with aprotinin (250 KIU/ml blood) for the instant inhibition of in vitro plasmin activity. The samples were placed on melting ice, and spun as soon as possible. Aliquots of the plasma were taken and stored at - 20°C until transfer to the central laboratory where they were stored at - 70°C; each sample was thawed only once.
Assay methods The plasma samples containing aprotinin were used for determining fibrinogen, fibrinogen degradation products (FgDP), fibrin degradation products (FbDP) and antithrombin III activity. Plasminogen and 04- antiplasmin activity assays were performed using citrate plasma samples without aprotinin, because aprotinin interferes with these assays.22 Fibrinogen was determined using the clotting rate method of Clauss. FgDP and FbDP were measured by ELISA, based on specific monoclonal antibodies (Fibrinostika kits, Organon Teknika, Boxtel, the 23 24 Netherlands). ' Antithrombin III, plasminogen and a2-antiplasmin were measured using a chromogenic substrate assay (Kabi/Chromogenix, Amsterdam, the Netherlands). Data were collected and analysed at an independent centre for statistics. Errors were discussed at Steering Committee meetings. Analysis of variance with repeated measures was performed for the comparison of hemostatic parameters. To adjust for the skewed distributions log-transformed values were used. Median values are presented in the tables and figures. All given Ρ values are two-sided.
Results
The time differences from the start of the infusion to the blood sampling time points was almost identical between the two treatment groups. Five hundred and forty-three patients were recruited in the SUTAMI study, but in 34 patients all the plasma samples were missing, and there were 311 (1.7%) individual measurements missing from the samples of a further 34 patients. Therefore complete sample recovery was 92.1%. For methodological reasons only complete sample series were included in the analysis of variance for these variables and only patients with a complete record are commented upon here. Despite these restrictions sample sets of more than 230 patients in each group were evaluated. The distribution of completely missing and partially missing samples was even between the
58 Hemostatic changes after saruplase and urokinase saruplase and urokinase groups. There was a similar fall in fibrinogen in both the saruplase and the urokinase treatment group (Table 1, Figure 1), with a tendency towards a greater spread of the individual patient's results in the saruplase group for samples at 1 h, 2 h and 6 - 12 h.
Table 1. Fibrinogen concentration (g/l, median values)
Time Saruplase (n = 234) Urokinase (n = 233)
Admission 3.00 3.20 1 h 0.48 0.48 2h 0.35 0.44 6- 12 h 0.46 0.54 24-36 h 1.80 1.70 36-72 h 3.55 3.50 g/L
5
4
3
2
-1 _i_ admission 1 h 2h 6-12 h 24-36 h 36-72 h
Figure 1. Distribution of fibrinogen levels. (Note, the spread bars represent the standard deviation which, as a mathematical representation of the variation of the mean, can go under zero)
For example at the 1 h sample the 25% and 75% quantiles were 0.10 g/l and 0.86 g/l in the saruplase group and 0.31 g/l and 0.70 g/l in the
59 Chapter 4
urokinase treatment group. This trend towards greater differential distribution of fibrinogen levels was not statistically significant. Full recovery of the fibrinogen concentrations to baseline values did not occur until the last sample at 36 - 72 h. In both of the treatment groups the concentration of fibrinogen degradation products rose markedly after therapy with the thrombolytic medication (Table 2, Figure 2).
Table 2. Fibrinogen degradation products (mg/l, median values)
Time Saruplase (n = 234) Urokinase (n = 233)
Admission 0.43 0.45 1 h 103.50 90.00 2 h 160.00 89.00 6-12 h 52.00 41.50 24-36 h 3.80 3.00 36-72 h 0.67 0.58
There was a tendency for a greater variation in the concentration of the fibrinogen degradation products in the saruplase treatment group. For example at the 1 h sample the 25% and 75% quantiles were 24 mg/l and 353 mg/l in the saruplase group and 44 mg/l and 190 mg/l in the urokinase group.
At the last sample (36 - 72 h) there was still a tendency for the median concentration of fibrinogen degradation products to be elevated. Treatment with saruplase leads to a higher concentration of fibrinogen degradation products over the whole time period than does urokinase (P < 0.001). Notwithstanding this difference in FgDP concentration, the incidence of bleeding was identical in both treatment groups (10.7%).20 The concentration of the fibrin degradation product, D-dimer, rose after the thrombolytic treatment.
60 Hemostatic changes after saruplase and urokinase
mg/L 600
saruplase 500 • ι urokinase ! 400
300
200 )
.t Τ 100 t
0
-100 I I I I , ad mission 1 h 2 τ 6-12 h 24-36 h 36-72 h
Figure 2. Distribution of the levels of fibrinogen degradation products
No difference was found between the two treatment groups (Table 3), and unlike the situation seen with the fibrinogen degradation products, there was no tendency for a greater spread of D-dimer concentrations in the saruplase treatment group. The changes in concentration of a2-antiplasmin did not differ between the two treatment groups.
Table 3. Fibrinogen degradation products, D-dimer (mg/l, median values)
Time Saruplase (n = 238) Urokinase (n = 234)
Admission 0.36 0.39 lh 13.00 14.00 2h 17.00 16.00 6-12h 7.85 7 90 24-36 h 1.20 0.96 36-72 h 0.47 0.44
At 1 h and 2 h following the start of thrombolytic therapy with either saruplase or urokinase the concentration of 0C2-antiplasm was below the limit of quantification in the majority of the patients (Table 4). The concentration of plasminogen fell after therapy with the thrombolytic
61 Chapter 4 medications and full recovery of the median concentrations to the baseline values had not taken place until the last sample time at 36 - 72 h (Table 5). No difference was found between the two treatment groups. The concentration of antithrombin III showed a slight fall during the measurement sequence and the analysis of variance did not indicate any difference between the two treatment groups (Table 6).
Table 4. a2-antiplasmin (U/l, median values)
Time Saruplase (n = 238) Urokinase (n = 237) , . . 068 (Ш AdmissioA n 1 h 0.00 0.00 2 h 0.00 0.02 6-12 h 0.29 0.29 24-36 h 0.52 0.50 36-72 h 0.71 0.69
Table 5. a2-antiplasmin (U/l, median values)
Time Saruplase (n = 238) Urokinase (n = 237) . , . . (λ97 0^97 Admission 1 h 0.33 0.35 2 h 0.32 0.34 6-12 h 0.40 0.41 24-36 h 0.61 0.64 36-72 h 0.87 0.89
62 Hemostatic changes after saruplase and urokinase
Table 6. Antithrombin III (U/ml, median values)
Time Saruplase (n = 238) Urokinase (n = 237) 096 Ö94 Admission lh 0.91 0.91 2h 0.92 0.93 6-12h 0.91 0.92 24-36h 0.83 0.86 36-72 h 0.84 0.85
Discussion
The finding that saruplase influences the hemostatic system to a similar extent as urokinase apparently contradicts results of animal experiments 512"14 but is in line with clinical experience.25"27 The difference between early experiments and clinical experience could be explained by a different milieu, type of clot used in in vitro studies, and by the different dosing schemes needed in clinical practice for acute thrombolysis. At doses used ъ С IT 1¿ ΊΟ in recent clinical trials, " ' the intrinsic clot selectivity of saruplase is completely masked. A pharmacokinetic study 26 demonstrated the conversion of approximately 28% of saruplase into its active tcu-PA degradation product and the hemostatic effects observed in our study confirm the concept of early and substantial conversion of saruplase by the systemically generated plasmin. There were higher values for fibrinogen degradation products with saruplase than with urokinase, with a tendency towards greater interindividual variation. Apparently contradicting this finding were the similar effects on fibrinogen, cc2-antiplasmin and plasminogen. However, a decrease in fibrinogen measured by a clotting rate test does not necessarily correlate directly to fibrinogen degradation product values, and low borderline values of a.2-antiplasmin and plasminogen are possibly less discriminating than increased fibrinogen degradation product values. Despite possible alternative causes e.g. divergent fibrinogen degradation product patterns or plasma sample matrices, the most straightforward explanation for the different values is the different plasminogen activating mechanism of saruplase and urokinase. Whereas urokinase is gradually inhibited in plasma, scu-PA is
63 Chapter 4
not quenched by inhibitors" and rapidly sets free its plasminogen activating activity (i.e. tcu-PA) in a positive feedback during treatment. This results in a more rapid generation of plasmin and, consequently, fibrinogen degradation products in the saruplase group. The effects measured refer to processes in plasma which do not necessarily reflect those at the clot site. Therefore, the observation of systemic plasminogen activation does not disprove a clot selective action of saruplase which might still be favorable for the rapid initiation of clot lysis. The local enhancing effect caused by the conversion of scu-PA to tcu-PA at the site of the clot is considered to contribute significantly to the efficacy of saruplase, and the increased generation of fibrinogen degradation products might contribute to reduce the risk of reocclusion. This is in accordance with the clinical observation that early maximum reperfusion and a low rate of reocclusion can be obtained by treatment with saruplase.28 We conclude that after treatment of patients with an acute myocardial infarction with suitable dose regimens of saruplase and urokinase the effect on the hemostatic system was essentially similar. Higher plasma fibrinogen degradation values were found after treatment with saruplase and are explained by different mechanisms of action of uroki- nase and saruplase.
Acknowledgements - We thank Anne Hoi and Monique van den Broek for their secretarial assistance. The study was sponsored by Grünenthal GmbH, the manufacturer of saruplase, and was co-sponsored by Behringwerke, the manufacturer of urokinase.
64 Hemostatic changes after saruplase and urokinase
References
1. Neuhaus K, Tebbe U, Gottwick M, et al. Intravenous recombinant tissue plasminogen activator (rt-PA) and urokinase in acute myocardial infarction: results of the German Activator Urokinase Study (GAUS). J Am Coll Cardiol 1988;12:581-587. 2. Mathey D, Schofer J, Sheehan F, Becher H, Tilsner V, Dodge HT. Intravenous urokinase in acute myocardial infarction. Am J Cardiol 1985; 55: 878 - 882. 3. Wun T, Ossowoski L, Reich E. A proenzyme form of urokinase. J Biol Chem 1982;257:7262-7268. 4. Gurewich V, Pannell R. Fibrin-specificity and efficacy of proteolysis induced by single-chain urokinase in plasma. Thromb Haemostasis 1983; 50: 321. 5. Gurewich V, Pannell R, Louie S, Kelley P, Suddith RK, Greenlee R. Effective and fibrin-specific clot lysis by a zymogen precursor form of urokinase (pro- urokinase). J Clin Invest 1984; 73: 1731 - 1739. 6. Gurewich V, Pannell R, Greenlee R, Suddith R. The fibrin specificity of single-chain urokinase is not dependent of fibrin binding. Thromb Haemostasis 1983; 50: 386. 7. Stump D, Thienpont M, Collen D. Urokinase-related proteins in human urine. J Biol Chem 1986; 261: 1267-1273. 8. Lijnen H, Zamarron С, Blaber M, Winkler ME, Collen D. Activation of plasminogen by pro-urokinase, I. Mechanism. J Biol Chem 1986; 261: 1253 - 1258. 9. Gurewich V. The sequential, complementary and synergistic activation of fibrin-bound plasminogen by tissue plasminogen activator and pro-urokinase. Fibrinolysis 1989; 3: 59 - 66. 10. Gurewich V. Fibrinolytic properties, of single-chain urokinase plasminogen activator and how they complement those of tissue plasminogen activator. In: Haber E, Braunwald E, eds. Thrombolysis. Basic contributions and clinical progress. Mosby Year Book, 1991: 51-60. 11. Pannell R, Gurewich V. Pro-urokinase - a study of its stability in plasma and of a mechanism for its selective fibrinolytic effect. Blood 1986; 67: 1215 - 1223. 12. Collen D, Stassen J, Blaber M, Winkler M, Verstraete M. Biological and thrombolytic properties of proenzyme and active forms of human urokinase. III. Thrombolytic properties of natural and recombinant urokinase in rabbits with experimental jugular vein thrombosis. Thromb Haemostasis 1984; 52: 27 -30. 13. Hanbücken F, Schneider J, Gilnzler WA, Friderichs E, Giertz H, Flohè L. Selective fibrinolytic activity of recombinant human pro-urokinase (single- chain urokinasetype plasminogen activator) from bacteria. Drug Res 1987; 37: 993-997.
65 Chapter 4
14. Flameng W, Vanhaecke J, Stump D, et al. Coronary thrombolysis by intravenous infusion of recombinant single-chain urokinase-type plasminogen activator or recombinant urokinase in baboons: effect on regional blood flow, infarct size and hemostasis. J Am Coll Cardiol 1986; 8: 118 - 124. 15. Van de Werf F, Vanhaecke J, De Geest H, Verstraete M, Collen D. Coronary thrombolysis with recombinant single-chain urokinase-type plasminogen activator in patients with acute myocardial infarction. Circulation 1986; 74: 1066-1070. 16. Van de Werf F, Nobuhara M, Collen D. Coronary thrombolysis with human single-chain urokinase-type plasminogen activator (pro-urokinase) in patients with acute myocardial infarction. Ann Intern Med 1986; 104: 345-348. 17. Diefenbach C, Erbel R, Pop, et al. Recombinant single-chain urokinase-type plasminogen activator during acute myocardial infarction. Am J Cardiol 1988; 61: 966-970. 18. Holmes W, Pennica D, Blaber M, et al. Cloning and expression of the gene for pro-urokinase in Escherichia coli. Biotechnology 1985; 3: 923 - 929. 19. Flohé L. Recombinant human pro-urokinase (non- glycosylated). Drugs of the Future 1986; 11: 851 -852. 20. Michels R, Hoffmann H, Windeier J, Barth H, Hopkins G. A double-blind multicenter comparison of the efficacy and safety of saruplase and urokinase in the treatment of acute myocardial infarction: report of the SUTAMI Group. J Thromb Thrombolysis 1995; 2: 117-124. 21. Genton E. Fibrinolytic therapy. In: van de Loo J, Prentice C, Beller F, eds. The Thrombotic Disorders. Stuttgart, New York: Schattauer, 1983: 192. 22. Hoffmann JJML, Vijgen M. Prevention of in vitro fibrinogenolysis during laboratory monitoring of thrombolytic therapy with streptokinase or APSAC. Blood Coag Fibrinol 1991; 2: 279-284. 23. Koppert PW, Hoegee-de Nobel E, Nieuwenhuizen W. A monoclonal antibody-based enzyme immunoassay for fibrin degradation products in plasma. ThrombHaemostasis 1988; 59: 310-315. 24. Koppert PW, Kuipers W, Hoegee-de Nobel B, Brommer EJP, Koopman J, Nieuwenhuizen W. A quantitative enzyme immunoassay for primary fibrinogenolysis products in plasma. Thromb Haemostasis 1987; 57: 25-28. 25. Ostermann H, Schmitz-Huebner U, Windeier J, Bär F, Meyer J, van de Loo J. Rate of fibrinogen breakdown related to coronary patency and bleeding complications in patients with thrombolysis in acute myocardial infarction. Result from the PRIMI trail. Eur Heart J 1992; 13: 1225 - 1232. 26. Koster RW, Cohen AF, Hopkins GR, Beier H, Günzler WA, van der Wouw PA. Pharmacokinetics and pharmacodynamics of saruplase, an unglycosylated single-chain urokinase-type plasminogen activator, in patients with acute myocardial infarction. Thromb Haemostasis 1994; 71: 740 - 744.
66 Hemostatic changes aftei saruplase and urokinase
27 The Belgian Saruplase Alteplase Trial Group Effects of alteplase and saruplase on hemostatic variables a single-blind, randomized trial in patients with acute myocardial infarction Coronary Artery Disease 1991, 2: 349-355 28 Primi Trial Study Group Randomized double-blind trial of recombinant pro- urokmase m acute myocardial infarction Lancet 1989, 1(8643) 863 - 868
67 68 CHAPTER 5
Plasma Markers of Thrombin Activity During Coronary Thrombolytic Therapy with Saruplase or Urokinase: No Prediction of Reinfarction
J. J. M. L. Hoffmann, H. R. Michels, J. Windeier, W. A. Günzler
J. J. M. L. Hoffmann, Department of Clinical Laboratories, Haemostasis Division, Catharma Hospital, Eindhoven, H. R. Michels, Department of Cardiology, Catharma Hospital, The Netherlands J. Windeier, Department of Medical Informatics and Biomathematics, Ruhr University, Bochum, Germany, W. A. Günzler, Grunenthal GmbH, Department of Clinical Research, Aachen, Germany
SUMMARY. One of the principal problems associated with thrombolytic therapy ¡s rethrombosis of vessels which were initially patent Although platelets as well as coagulation activation have been implicated in rethrombosis, the specific mechanisms leading to this complication are still unclear Available evidence is limited to smaller studies using the current thrombolytic agents Here we report on the multicentre SUTAMI trial comparing recombinant saruplase and urokinase in 543 patients with acute myocardial infarction, in 33 of whom early reinfarction was documented Plasma from these patients and 33 matched patients without reinfarction was investigated for thrombin-antithrombin 111 complex and prothrombin activation fragments 1+2 as markers of activated coagulation, during 72h after starting the lytic therapy Both drugs caused considerable systemic degradation of fibrinogen and the degree of systemic lysis was very similar The median concentrations of both thrombin-antithrombin III complex and prothrombin fragments 1+2 significantly increased 3- to 6-fold after the therapy, indicating extensive activation of the coagulation system Following heparin administration, both parameters returned towards normal in most patients At no time points studied was there any significant difference m these coagulation parameters between the patients with and those without reinfarction In contrast to other findings, thrombin- antithrombin 111 complex concentration was not a useful indicator of reinfarction in the patients studied and neither was the concentration of prothrombin activation fragments 1 +2
Early thrombolytic treatment of coronary thrombosis has been proven to significantly reduce the mortality of patients with acute myocardial infarction (AMI).1"3 Rapid and sustained restoration of the patency of the infarct-related arteries is necessary for salvaging viable myocardial tissue,
69 Chapter 5
but rethrombosis of initially patent vessels can occur in up to 30% of the patients. The mechanisms leading to reocclusion have not yet been fully elucidated, but there is growing evidence that activation of the coagulation system and of platelets might cause ongoing thrombosis, which exceeds the fibrinolytic activity induced by the thrombolytic agents.4 This is supported by clinical and experimental observations that adjunctive therapy with anticoagulants or anti-platelet drugs accelerates or enhances thrombolysis. 4'5 Moreover, increased thrombin activity has been shown during thrombolytic therapy using sensitive techniques.6"9
Coagulation activation can reliably be demonstrated by determining one of the activation-specific substances in plasma, for which accurate and sensitive assays are available. Fibrinopeptide-A (FpA) is a small fibrinogen fragment, which is released when thrombin converts fibrinogen into fibrin. Thrombin-antithrombin III complex (TAT) is rapidly formed in the circulation between active thrombin and its natural inhibitor antithrombin III. Prothrombin fragments 1 and 2 (F|+2) are split off by factor Xa when it activates prothrombin. Because all these molecules are rapidly eliminated from the circulation, they are well suited as sensitive markers of in vivo activation of the coagulation system. One group of investigators reported that an elevated plasma concentration of TAT during thrombolytic therapy was highly predictive of coronary reocclusion within 24 to 36h.9 These results are based on observations in only 13 patients with reocclusion and they have not yet been confirmed by others. This prompted us to perform the present study, which is part of a clinical trial comparing the efficacy and safety of saruplase (recombinant single- chain, non-glycosylated human pro- urokinase) and urokinase in patients with AMI. The aim of our investigation was to document the course of TAT and Fi+2 as markers of in vivo thrombin activity during thrombolytic therapy with saruplase or urokinase, in patients who had clinical signs of reinfarction and in an equal number of matched patients, who experienced an uneventful clinical course. In addition, we examined whether TAT and F|+2 could be used as prognostic indicators of reinfarction after thrombolysis in this patient group.
70 Plasma markers of thromboactivity
PATIENTS AND METHODS
Patients and Treatment The patients were selected from a multicentre, randomised, double-blind trial comparing saruplase and urokinase in the treatment of acute myocardial infarction (SUTAMI).12 Patients between 20 and 75 years presenting within 6h after the onset of chest pain indicative of AMI, were included into this trial if they met the usual criteria. They were randomised to treatment with either saruplase or urokinase. Saruplase was the full- length, non-glycosylated, single chain form of human urokinase-type plasminogen activator (pro-urokinase), produced by recombinant DNA technology in E. coli (Grünenthal GmbH, Aachen, Germany) and urokinase was purified from human urine (Behringwerke AG, Marburg, Germany). Saruplase was given as a 20mg bolus, followed by 60mg as an intravenous infusion over lh. Urokinase was administered by intravenous infusion, first 1.5MU in 5 min and then 1.5MU over lh. Each regimen included a placebo medication for the other arm of the study (double- dummy technique). In both groups, heparin was started 30 min after completion of the active drug infusion (thus, 90 min after starting the thrombolytic therapy) by continuous infusion of 15 U/kg/h and the dose was adapted according to the activated partial thromboplastin time. Heparin was continued until the coronary angiography, 24 to 72h after start of the therapy. In-hospital reinfarction was assessed using clinical and electrocardiographic signs before discharge, 7 to 14 days after admission.
Blood Collection Before administering the thrombolytic medication and after lh (end of infusion), 2h (after 30 min heparin infusion), 6 to 12h (during heparin), 24 - 36h (prior to angiography) and 60 - 72h (after angiography), blood was taken for haemostatic assays. At each time point, two 9ml samples were collected in plastic syringes using a 19 gauge needle. One sample was drawn into 1ml citrate (0.01 M final concentration) and the other one into lml citrate supplemented with aprotinin (250 KIU/ml blood) in order to instantly inhibit in vitro plasmin activity. The samples were immediately placed in melting ice and centrifuged as soon as possible for at least 10 min at 2000g. The plasma was pipetted off, divided into three aliquots and frozen at - 20°C until transfer to the central laboratory. There, the samples were stored at - 70°C; each sample was thawed only once.
71 Chapter 5
Assays TAT and Fi+2 were determined in duplicate in aprotinin-citrate plasma, using commercially obtained ELISA kits (Enzygnost®; Behringwerke AG, Marburg, Germany), as described before.10'11 In our hands, the lower limit of detection of the TAT assay was 0.7 μg/l, the normal range <4.0 μg/l and the inter-assay coefficient of variation (CV) 10.0% at 9.1 μβ/1 (n=47). For Fi+2 these data were: detection limit 0.02 nmol/1, normal range 0.4 to 1.2 nmol/l and CV 10.9% (mean 0.45 nmol/1; n=20).
Statistics For each of the patients with reinfarction, a matching patient without reinfarction was selected on the basis of successively: treatment group, sex, infarct localisation, body weight and age. The matching for the first four items was complete; the median difference in age within a pair was 3 years (range 0-21 years). Median and interquartile ranges of TAT and F 1+2 values at each of the six time points were calculated. Groups of patients with and without reinfarction were compared using analysis of variance in a repeated measures model; this analysis was restricted to patients with complete data (n=54). The values of F]+2 were log-transformed prior to analysis. Ρ values <0.05 were considered significant. After it had been verified that the type of thrombolytic treatment had no significant effect on the TAT and F]+2 results, we pooled the data in order to form two groups with and without reinfarction, respectively.
RESULTS In the SUTAMI study as a whole, the pretreatment fibrinogen concentration was normal; there was no difference between the two treatment groups (mean±SD: 3.16±0.73 for saruplase and 3.22±0.84 g/1 for urokinase). After 2h, fibrinogen had substantially decreased and it was closely comparable in the patients treated with saruplase (0.48±0.57 g/1) and urokinase (0.48±0.39 g/1). Similarly, plasminogen and 0C2-antiplasmin decreased, but again without significant differences between the two groups.12 Reinfarction was documented in 33 subjects (8.1% of all patients); 19 had been treated with saruplase and 14 with urokinase. The age of the patients with reinfarction (mean±SD: 60.3+9.1 years) was not significantly
72 Plasma markers of thromboactivity different from those without reinfarction (58.8+9.6 years) and neither were there differences with respect to sex (79% males), location of infarction (42% anterior) and body weight (80±12kg).
TAT (pg/L) 60
6-12 24-36 60-72 time (h) Fig. 1. Course of median TAT concentration in patients with (· solid line) and without (O, dotted line) reinfarction. Vertical lines represent interquartile range and the shaded area is the normal range.
TAT During Thrombolysis The median concentration of TAT and the 25 to 75% range at each of the six time points is given in Figure 1 for the group of patients with reinfarction as well as those without. At admission, the TAT concentration was elevated in 41/54 (76%) of the patients. After Ih, just after completing the thrombolytic infusion, TAT had further increased in 39 of the patients and in 15 it was slightly lower than before thrombolysis, but above the normal range in all patients. At 2h, when the patients were on heparin for 30 min, TAT decreased significantly in the majority of patients (72%) and at 6 to 12h it had decreased further, but was still elevated in nearly all patients. The decrease continued in most patients during the infusion of heparin. However, there were 19 patients in whom TAT increased in either of the two samples during the heparinisation period and a single patient
73 Chapter 5
displayed a persistent increase in TAT under heparin. After 24 to 36h, before the coronary angiography, TAT was still above the upper limit of normal in 50% of the patients (Fig. 1). After the angiogram had been made, TAT increased again in 39 patients and remained normal in only 12 patients. There was no significant difference in TAT between the patients with reinfarction and those without over the entire study period (P=0.38 by analysis of variance), as illustrated in Figure 1.
F1+2 During Thrombolysis
Figure 2 shows the course of the Fi+2 concentration (median and interquartile range) during the treatment period. At presentation, 17 of the
patients (31%) had a F|+2 concentration above the upper limit of normal. After thrombolysis, Fi+2 increased in 47 patients and decreased in 7, who all had an increased F1+2 at admission. Shortly after starting the heparin
infusion, at 2h, median Fi+2, changed only negligibly, but it had normalised in more than half the patients by 6 to 12h and continued to decrease thereafter (Fig. 2). The decrease was continuous in 37 patients, whereas in the other 17 either of the two samples had a higher value than the previous one. Before angiography, the F1+2 concentration was within the normal range in 30 patients and only slightly higher in another 12 patients. After angiography, F1+2 increased again in the vast majority of patients (86%): in 29 it remained within normal limits whereas 25 patients had elevated F]+2 . Like for TAT, there was no significant difference in Fi+2 concentration between the patients with and without reinfarction (P=0.33).
TAT and F1+2 as Indicators of Reinfarction Risk When using a cut-off value for TAT of 6 μ^Ι,9 neither the values obtained at lh, nor those at 2h did reliably indicate reinfarction. At lh, the relative risk for reinfarction associated with high TAT was 0.963 (95% confidence interval: 0.89 - 1.04). The sensitivity of increased TAT for reinfarction was 93% and the specificity 0% (Table).
Similarly, as shown in the Table, Fi+2 at lh did not indicate reinfarction when 2.0 птоІЛ was used as a cut-off value: relative risk 1.04 (95% CI: 0.77 - 1.39), sensitivity 76% and specificity 25%. Also at the other time points, neither Fi+2 nor TAT were of value as indicators of the outcome in terms of reinfarction (data not shown).
74 Plasma markers of thromboactivity
prothrombin fragment Fus (nmol/L) 5-1 —
6-12 24 - 36 60 - 72 time (h)
Fig. 2. Course of median F1+2 concentration in patients with (·, solid line) and without (O, dotted line) reinfarction. Vertical lines represent interquartile range and the shaded area is the normal range.
Table 1. Contingency tables of TAT and F1+2 concentrations at 1h according to reinfarction
TAT 4+2
<6.0 ug/L > 6.0 ug/L <2 nmol/L >2 nmol/L Reinfarction 1 26 6 21 No reinfarction 0 28 7 21
75 Chapter 5
DISCUSSION
There is accumulating evidence that thrombolytic therapy is associated with generation of thrombin activity.6'9 Increased FpA in plasma has been demonstrated during treatment with streptokinase6,7 as well as tissue-type plasminogen activator, 7'8'13 so the type of thrombolytic agent probably plays no role in this phenomenon. In theory, FpA may not be specific for thrombin activity, since plasmin and tissue-type plasminogen activator can cleave small peptides, containing the FpA sequence, off fibrinogen.8,14 In vitro experiments have confirmed that plasmin will increase the FpA concentration in platelet-rich plasma, but Winters et al have shown that this effect is mediated by thrombin, since it can be markedly suppressed by thrombin inhibitors.15 In addition there is supplementary evidence for the involvement of thrombin activation in thrombolysis. Concomitant hirudin and heparin could accelerate and enhance the lysis of thrombi by tissue- type plasminogen activator and prevent reocclusion in animal models or in vitro,' "' while in clinical studies heparin increased the patency rate after alteplase treatment.' Technical requirements for blood collection and assay of FpA are rather stringent, contrary to two novel assays, which measure other aspects 10,u of thrombin activation, namely TAT and Fi+2 At present, data on the clinical usefulness of TAT and Fi+2 during thrombolytic therapy are still scanty and unconfirmed.919 This inspired us to perform the study described here. Although the protocol of the SUTAMI trial included only a single coronary angiography and thus did not allow us to record reocclusion, there were 33 patients in whom reinfarction was diagnosed on the basis of clinical and electrocardiographic data. Since reinfarction after thrombolysis may be regarded as the clinical analogue of angiographically proven reocclusion, we studied the course of TAT and Fi+2 in these patients and their value for indicating reinfarction. We found that activated thrombin is present in vivo at admission in AMI patients and that thrombolytic therapy amplifies the generation of thrombin, which confirms previous reports.6"9,18
The course of TAT and F1+2 in our patients showed a marked increase until the heparin infusion was started (between 1 and 2 h). This is in close agreement with the results of Munkvad et al,19 who used a similar scheme of heparinisation after rtPA. In our patients the median peak
76 Plasma markers of thromboactivity
concentrations of TAT and F]+2 were 1.5-2 times higher, which might reflect the more pronounced fibrinogenolysis in our patients as compared with their study.l9The results of Gulba et al are quite different, however. These authors described an immediate decrease in TAT in those patients with sustained patency, whereas a significant increase in mean TAT was observed only in patients with non-successful lysis or those with reocclusion.9 It should be noted, however, that Gulba and co-workers started the infusion of heparin already before the thrombolytic agent was given, while we began heparin only 30 min afer completing the thrombolytic infusion. Moreover, the design of the studies was different: Gulba et al9 conducted a single-centre study, while we recruited our patients in a multi-centre trial. This may have induced variations in blood collection and sample processing, resulting in less consistent results. Therefore, the conclusions drawn by Gulba et al9 may not be applicable during routine clinical thrombolysis, since under such circumstances we found that TAT and F1+2 concentrations were not indicative of reinfarction (Table).
A conspicuous difference in the courses of TAT and F]+2 became apparent: the median TAT decreased immediately after starting heparin administration, but it lasted until 24 - 36h before it was within the normal range in 50% of the patients (Fig. 1). Median FH2 , on the contrary, did hardly decrease shortly after the heparin infusion was started, but it normalised much faster than TAT. The quicker initial decrease in TAT as compared to F]+2 can be explained by the difference in half-life (about 3 min and 90 min, respectively), but this does not account for the fact that Fi+2 normalised more rapidly than TAT. The same discrepancy has been noted previously during heparin treatment of deep venous thrombosis,20 but remains as yet unexplained. It is intriguing to dwell on the mechanisms involved in the activation of coagulation during thrombolytic therapy. It is known that fibrin-bound thrombin, which is enzymatically active and is inaccessible to heparin- antithrombin III complex, 21 can be released upon plasmin-mediated degradation of fibrin.22 While this might explain the rise in TAT, it does not account for the increase in F]+2 , which represents newly activated thrombin rather than release of already active thrombin. Alternatively, it has been shown that a partially lysed thrombus is highly thrombogenic itself and that further dissolution of the clot in the coronary artery might re-expose the underlying vessel wall injury, which provides an ongoing thrombogenic stimulus.23 Recently it has been suggested that lysis of fibrin
77 Chapter 5
exposes procoagulant activity on endothelial cells and thus causes coagulation activation not only at coronary sites, but on the vascular intima in the systemic circulation.19 Finally, activation of factor X or prothrombin by circulating free plasmin might also provide an explanation for the results observed here. It is still too early to favour either of these possible mechanisms. In conclusion, we have demonstrated that during thrombolytic therapy with saruplase and urokinase, which caused a very similar degree of systemic plasminaemia, the coagulation system is extensively activated and that this increased in vivo thrombin generation can be suppressed with heparin. It proved impossible to get an indication of early reinfarction from the concentrations of TAT and Fi+2 measured during thrombolytic therapy, at least when utilising the current scheme of heparin administration after the thrombolytic drug. Although TAT may be of some value in concomitant thrombolytic and heparin therapy, we can not yet recommend the TAT and Fi+2 assays for prognostic use during clinical thrombolysis.
A CKNO WLED GEMENTS: The authors are indebted to Mrs M. Vijgen for her expert technical assistance, to all cardiologists participating in the SUTAMI trial and to Grünenthal GmbH, Aachen, Germany for financial support.
78 Plasma markers of thromboactivity
REFERENCES
1. Gruppo Italiano per lo studio della streptochinasi nell'infarto miocardico (GISSI). Long-term effects of intravenous thrombolysis in acute myocardial infarction: final report of the GISSI study. Lancet 1987; ii: 871 -874. 2. AIMS trial study group. Effect of intravenous APSAC on mortality after acute myocardial infarction: preliminary report of a placebo- controlled clinical trial. Lancet 1988; i: 545-549. 3. Wilcox R G, von der Lippe G, Olsson С G, Jensen G, Skene A M, Hampton J R for the ASSET study group. Trial of tissue plasminogen activator for mortality reduction in acute myocardial infarction. Anglo- Scandinavian study of early thrombolysis (ASSET). Lancet 1988; ii: 525-530. 4. Sobel Β E. Thrombolysis in the treatment of acute myocardial infarction. In: Fuster V, Verstraete M, eds. Thrombosis in cardiovascular disorders. Philadelphia: WB Saunders Co. 1992: 289 - 326. 5. Becker R C, Gore J M. Adjuvant antiplatelet strategies in coronary thrombolysis. Circulation 1991; 83: 1115 - 1117. 6. Eisenberg Ρ R, Sherman L A, Jaffe A S. Paradoxic elevation of fibrinopeptide A after streptokinase: evidence for continued thrombosis despite intense fibrinolysis. J Am Coll Cardiol 1987; 10: 527 - 529. 7. Owen J, Friedman К D, Grossman В A, Wilkins C, Berke A D, Powers E R. Thrombolytic therapy with tissue plasminogen activator or streptokinase induces transient thrombin activity. Blood 1988; 72: 616-620. 8. Rapold H J, Kuemmerli H, Weiss M, Baur H, Haeberli A. Monitoring of fibrin generation during thrombolytic therapy of acute myocardial infarction with recombinant tissue-type plasminogen activator. Circulation 1989; 79: 980-989. 9. Gulba D C, Bartheis M, Westhoff-Bleck M, et al. Increased thrombin levels during thrombolytic therapy in acute myocardial infarction: relevance for the success of therapy. Circulation 1991; 83: 937 - 944. 10. Pelzer H, Schwartz A, Heimburger N. Determination of human thrombin-antithrombin III complex in plasma with an enzyme-linked immunosorbent assay. Thromb Haemost 1988; 59: 101-106.
79 Chapter 5
11. Pelzer H, Schwartz A, Stüber W. Determination of human prothrombin activation fragment 1+2 in plasma with an antibody against a synthetic peptide. Thromb Haemost 1991; 65: 153-159. 12. Michels H R, Withagen A, Bosma A, et al for the SUTAMI study group. SUTAMI: double-blind comparison of saruplase and urokinase in acute myocardial infarction. Neth J Cardiol 1991; 4: 254 (abstract). 13. Rapold H J, de Bono D, Arnold A E R, et al. Plasma fibrinopeptide A levels in patients with acute myocardial infarction treated with alteplase. Correlation with concomitant heparin, coronary artery patency and recurrent ischemia. Circulation 1992; 85: 928-934. 14. Weitz J I, Cruickshank Μ К, Thong В, et al. Human tissue-type plasminogen activator releases fibrinopeptides A and В from fibrinogen. J Clin Invest 1988; 82: 1700-1707. 15. Winters К J, Santoro S A, Miletich J P, Eisenberg Ρ R. Relative importance of thrombin compared with plasmin-mediated platelet activation in response to plasminogen activation with streptokinase. Circulation 1991; 84: 1552-1560. 16. Mirshahi M, Soria J, Soria С, et al. Evaluation of the inhibition by heparin and hirudin of coagulation activation during r-t-PA-induced thrombolysis. Blood 1989; 74: 1025 - 1030. 17. Rapold H J, Lu H R, Wu Ζ, Nijs Η, Collen D. Requirement of heparin for arterial and venous thrombolysis with recombinant tissue-type plasminogen activator. Blood 1991; 77: 1020- 1024. 18. Haskel E J, Prager N A, Sobel Β E, Abendschein D R. Relative efficacy of antithrombin compared with antiplatelet agents in accelerating coronary thrombolysis and preventing early reocclusion. Circulation 1991; 83: 1048-1056. 19. Munkvad S, Gram J, Jespersen J. Possible role of vascular intima for generation of coagulant activity in patients undergoing coronary thrombolysis with recombinant tissue-type plasminogen activator. A randomized, placebo-controlled study. Scand J Clin Lab Invest 1991; 51:581-590. 20. Estivals M, Pelzer H, Sie Ρ, Pichón J, Boccalon H, Boneu В. Prothrombin fragment 1+2, thrombin-antithrombin HI complexes and D-dimers in acute deep vein thrombosis: effects of heparin treatment. Br J Haematol 1991; 78: 421 -424. 21. Weitz J I, Hudoba M, Massel D, Maraganore J, Hirsh J. Clot-bound thrombin is protected from inhibition by heparin-antithrombin III but
80 Plasma markers of thromboactivity
is susceptible to inactivation by antithrombin Ill-independent inhibitors. J Clin Invest 1990; 86: 385-391. 22. Francis С W, Markham R E Jr, Barlow G H, Florack Τ M, Dobrzynski D M, Marder V J. Thrombin activity of fibrin thrombi and soluble plasmic derivatives. J Lab Clin Med 1983; 102: 220-230. 23. Badimon L, Badimon J J, Fuster V. Pathogenesis of thrombosis. In: Fuster V, Verstraete M, eds. Thrombosis in cardiovascular disorders. Philadelphia: WB Saunders Co. 1992: 17-39.
81 82 CHAPTER 6
Comparison of Saruplase and Alteplase in Acute Myocardial Infarction
Frits W. Bär, MD, Jürgen Meyer, MD, Frank Vermeer, MD, Rolf Michels, MD, Bernard Charbonnier, MD, Klaus Haerten, MD, Marfìn Spiecker, MD, Carlos Macaya, MD, Michel Haussen, MD, Magda Heras, MD, Jean P. Boland, MD, Marie-Claude Morice, MD, Francis G. Dunn, MD, Rainer Uebis, MD, Christian Hamm, MD, Oded Ayzenberg, MD, Gerhard Strupp, MD, Adrie J. Withagen, MD, Werner Klein, MD, Jürgen Windeler, PhD, Gwyn Hopkins, MD, Hannes Barth, MD, and Michael J.M. von Fisenne MSc, for the SESAM Study Group*f
From the Department of Cardiology, University Hospital Maastricht, Maastricht, The Netherlands This study was supported by Grunenthal GmbH, Aachen, Germany *See Appendix for participating centers tStudy in Europe with Saruplase and Alteplase in Myocardial Infarction
Abstract: Four hundred seventy-three patients with acute myocardial infarction (AMI) were treated with either saruplase (80 mg/hour, η = 236) or alteplase (100 mg every 3 hours, η =237) Comedication included heparin and acetyhahcylic acid Angiography was performed at 45 and 60 minuter after the start of thrombolytic therapy When flow was insufficient, angiography was repeated at 90 minutes Coronary angioplasty was then performed if Thrombolysis In Myocardial Infarction (TIMI) trial 0 to 1 flow was seen Control angiography was at 24 to 40 hours Baseline characteristics were similar Angiography showed comparable and remarkably high early patency rates (TIMI 2 or 3 flow) in both treatment groups at 45 minutes, 74 6% versus 68 9% (p = 0 22), and at 60 minutes 79 9% versus 75 3% (p = 0 26) Patency rates at 90 minutes before additional interventions were also comparable (79 9% and 8 14%) Angiographic reocclusion rates were not significantly different 1 2% vei sus 2 4% (ρ - 0 68) After rescue angioplasty, angiographic reocclusion rates of 22 0% and 15 0% were observed Safety data were similar for both groups Thus, (1) early patency rates were high for saruplase and alteplase treatment, (2) reocclusion rates for both drugs were remarkably low, and (3) complication rates were similar Thus, saruplase seems to be as safe and effective as alteplase
83 Chapter 6
Previous experience with the new thrombolytic agent saruplase indicated that it is superior to streptokinase in the speed of eliciting reopening of the occluded artery.1 It seemed valuable to compare saruplase with alteplase tissue plasminogen activator, which is considered by many as the best thrombolytic agent currently on the market. Until now, only 1 small study directly compared saruplase with alteplase.2 Patency rates of the Thrombolysis In Myocardial Infarction (TIMI) trials 2 and 3 at 24 to 72 hours after the start of thrombolytic therapy appeared higher in the saruplase group: 71% compared with 54% in the alteplase group (p = 0.3). Once an occluded coronary artery has been reopened, it should remain open. In the Pro-urokinase in Myocardial Infarction (PRIMI) trial,1 angiographic reocclusion of patients treated with saruplase with no additional interventions was remarkably low (0.9%), whereas reocclusion rates of 10% to 15% for alteplase are reported.3"5 In the present randomized multicenter study, a direct comparison between saruplase and alteplase was done to study reocclusion as the primary end point within 24 to 40 hours after the start of thrombolytic therapy. In addition, the speed of reperfusion was evaluated at the earliest feasible time point after initiation of therapy (45 minutes). Other end points were patency rates at 60 minutes and 90 minutes, and safety data.
METHODS
Study design and patient entry criteria: The Study in Europe with Saruplase and Alteplase in Myocardial infarction (SESAM) trial was a randomized, double- blind study using a double-dummy technique comparing unglycosylated recombinant single-chain urokinase-type plasminogen activator (saruplase) with recombinant plasminogen activator (alteplase) in 17 European hospitals. Except for 1 center, which included only 4 patients, all hospitals had the opportunity to perform rescue angioplasty if indicated. Saruplase and matching placebo were prepared by Grünenthal GmbH, Aachen, Germany, and alteplase and matching placebo were purchased from Thomae AG, Biberach, Germany, and delivered by Grünenthal GmbH. Inclusion criteria for the study were: patients aged <70 years and suspected of having acute myocardial infarction with chest pain lasting for >30 minutes and therapy delayed to within 6 hours. In case of
84 Report of SESAM study group recurrent myocardial infarction, <6 months should have elapsed; in addition, the location should in a different area. The electrocardiographic criteria were: ST-segment elevation >0.1 mV in >2 frontal plane leads (I, II, III, aVL, and VF) or ST-segment elevation >0.2 mV in >2 precordial leads (Vito б). Patients with an electrocardiogram that made the presence of myocardial infarction difficult to confirm (e.g., left bundle branch block or Wolff-Parkinson-White syndrome) were excluded. Patients with cardiogenic shock, prior coronary artery bypass grafting, and increased risk of bleeding were also not included. Further, expected inability to perform the first coronary angiography at 45 minutes was an exclusion criterion, as were chronic concomitant diseases, use of oral anticoagulant medication, and no informed consent. The start of the infusion of study medication was taken as the irrevocable entry of a patient into the study. Angiographic outcome and clinical complications were recorded.
Medication: Qualifying patients were treated either with 80-mg saruplase administered intravenously or a bolus dose of 20 mg and a 60- minute infusion of 60 mg, or 100 mg of alteplase administered as a bolus of 10 mg, and an infusion of 50 mg during the next hour, and a 40-mg infusion for the next 2 hours (the standard regimen at the time). Comedication consisted of heparin given as a 5,000-U bolus before the thrombolytic agent and another 5,000-U bolus before catheterization. After the first bolus, an intravenous drip of heparin was started and titrated according to activated partial thromboplastin time (1.5 to 2.5 X normal value), and continued until the second cardiac catheterization at 24 to 40 hours. In addition, all patients received 300 mg of acetylsalicylic acid administrated intravenously before study medication and continued once daily (80 to 120 mg orally).
Angiography: During the first catheterization, the first coronary angiogram had to be performed at 45 ± 7 minutes after the start of study medication, and repeated at 60 ± 7 minutes. If coronary flow was TIMI grade 0 to 2 at 60 minutes, the culprit artery was also visualized at 90 minutes. Thereafter, angioplasty could be performed in case of no or minimal flow (TIMI 0 to 1 flow). Patients with TIMI 3 flow at 60 minutes did not undergo a 90-minute angiography because of the small chance of reocclusion between 60 and 90 minutes.' At 24 to 40 hours a second
85 Chapter 6
catheterization was performed. All angiograms were centrally assessed by experienced cardiologists blinded to the therapy.
Statistical methods: The sample size of the study was calculated on the following assumptions: α = 0.05 (2-sided), and expected reocclusion rate of 5% for saruplase and 15% for alteplase (power 85%). To demonstrate a difference of >10%, 153 patients were needed. For an intent-to-treat analysis only patients with an open (TIMI flow grades 2 or 3) infarct- related artery, without an interventional procedure, at the end of the first catheterization entered analysis. Patients who did not undergo a second catheterization owing to technical reasons were excluded from analysis. Reocclusion was deemed to have occurred if (1) the vessel was documented as being closed at the second catheterization; (2) no second catheterization was performed owing to medical reasons, or an intervening intervention (coronary angioplasty or bypass) was performed; or (3) the second catheterization was not in the required time window. When the initial patency rate is of the order of 70% in both groups, 460 patients were needed to be randomized, leaving 306 patients for analysis of reocclusion. All data were collected and analyzed centrally by an independent statistic center. An intensive query system was used for missing or inconsistent data. Reocclusion and patency rates, and other binary variables were analyzed by a Fisher's exact test (2- sided).
RESULTS
Patient characteristics: Between December 1991 and February 1993, 473 patients were recruited from 17 centers in 8 countries: 236 patients were allocated to saruplase, and the other 237 patients to alteplase. The mean delay between onset of chest pain and start of medication was 170 minutes. This and other baseline data were nearly identical for both groups (Table 1).
86 Report of SESAM study group
Table 1. Baseline Data
Saruplase Alteplase 90 % CI (n = 236) (n = 237)
Mean age (yr ± SD) 57 + 9 57 + 10 -1.4-1.4 Men 192(81) 185(78) -9.4-2.8 Delay between pain onset 171 ±75 169 ±68 -12.9-8.9 and therapy (min ± SD) Infarct coronary artery Right 109(46) 106(45) Left anterior descending 93 (39) 95(40) Left circumflex 29(12) 29(12) Unclear 2 (0.8) 5(2.1) No catheterization 3(1.3) 2 (0.8)
Values are expressed as number (%) unless otherwise noted. CI = confidence interval.
Angiography:
Patency: The percentage of patients who underwent angiography at 45 minutes was 82.2%, at 60 minutes 94.7%, and at 24 to 40 hours 91.8%, indicating a high protocol compliance. Early patency rates (TIMI 2 or 3 flow) were slightly but not significantly higher in the saruplase treatment group (Table 2, Figure 1). With the inclusion of patients who already had TIMI 3 flow at 60 minutes but no repeat angiogram thereafter, and patients who had undergone angiography outside of the required time window, patency rates (at 90 minutes before additional intervention) were 79.9% and 81.4%, respectively (p = 0.72). During the first catheterization, TIMI 2 flow varied at the different time points, between 8% and 12% for both drugs (Figure 1). When considering the results after intervention, the overall patency rates at the end of the first catheterization were 94.0% and 92.3%, respectively (p = 0.58). At 24 to 40 hours, patency rates for all patients who underwent a catheterization were high and comparable between both groups, 94.0% and 93.6%, respectively.
87 Chapter 6
Table 2. Patency (Thrombolysis In Myocardial Infarction [TIMI] Trial 2 or 3 Flow) of Patients Who Had No Rescue Angioplasty
Saruplase Number (%) Alteplase Number (%)
45 min* 147/197 (75) 135/196(69) 60 min* 179/224 (80) 171/227(75) 90 minf 183/229(80) 188/231(81) 24 - 40 h* 168/170(99) 166/170(98)
•Patients with a medical reason for no catheterization were designated "closed " TNO angioplasty, patients outside time windows excluded. {Patients who were open (TIMI 2 or 3 flow) atfirst catheterizatio n without angioplasty. Note; All ρ values for comparison >0.2. patency rate (%)
100
80
60
40
D not done 20 Ώ TIMI grade 1 + 0 D TIMI grade 2 D TIMI grade 3 ЗД Saruplase Saruplase Saruplase Saruplase Alteplase Alteplase Alteplase Alteplase
FIGURE 1. Thrombolysis in Myocardial Infarction (TIMI) Trial flows are given at different time points for patients treated with saruplase and alteplase. SESAM = The Study in Europe with Saruplase and Alteplase in Myocardial infarction.
Reocclusion: Angiographically determined reocclusion rates at 24 to 40 hours were low in patients who did not undergo intervention: 2 patients (1.2%) receiving saruplase and 4 (2.4%) alteplase (p = 0.68). Furthermore, patients were considered to have had reocclusion if (re)angioplasty or coronary bypass surgery was performed (4 saruplase patients and 5
88 Report of SESAM study group alteplase patients) or because the patient died before a 24- to 40-hour catheterization procedure (3 saruplase patients). With use of an intent-to- treat analysis, including patients who did not undergo a 24- to 40-hour catheterization because of medical reasons (n = 20), or those who had undergone catheterization too early (n = 5), the reocclusion rates were 6.7% and 10.3%, respectively (p = 0.26). Outcome of rescue angioplasty at first catheterization: Angioplasty was performed in 57 patients at 90 minutes (Table 3). Rescue angioplasty was successful in 52 patients, resulting in a patency (TIMI 2 or 3 flow) of 91%. Reocclusion rates at 24 to 40 hours after successful angioplasty were somewhat higher in patients taking saruplase.
Table 3. Rescue Angioplasty
Saruplase Alteplase Number (%) Number (%)
TIMI 0-1 at 90 min 40/233 (17) 39/235 (17) TIMI 2 - 3 but unstable 5/233 (2) 1/235 (0.4) Angioplasty attempt 33/45(73) 24/40 (60) Successful angioplasty 30/33(91) 22/24 (92) Recatheterization at 24 - 40 h 27/30 (90) 20/22(91) Reocclusion 6/27 (22) 3/20(15)
Abbreviation as in Table 2.
Coronary angioplasty during hospital phase: Angioplasty was performed between the first and second catheterization because of recurrent ischemia, and during the second catheterization because of reocclusion or high-grade stenosis (Table 4). Angioplasty between the second catheterization and discharge was required for recurrent ischemia (spontaneous or during exercise testing), late reinfarction, or anatomic reasons.
89 Chaplor 6
Table 4. Number and Timing of Angioplasty During Hospital phase
Saruplase Alteplase (n = 236) (n = 237) 33(14) 24(10) Rescue angioplasty at first catheterization* Between catheterizations1 5(2) 7(3) Angioplasty at second 5(2) 18(8) catheterization* After second catheterization5 41(17) 32(14) Number of patients with at least 1 88 (37) 74(31) angioplasty •Angioplasty at first catheterization in case of Thrombolysis In Myocardial Infarction trial 0 or I flow + Between catheterizations for recurrent ischemia *At second catheterization for high-grade stenosis or reocclusion ^ After second catheterization for recurrent ischemia, remfarction, or anatomic reasons Values are expressed as number (%)
Clinical outcome: Complication rates were similar for saruplase and alteplase (Table 5).
Table 5. Clinical Outcome During Hospital Phase
Saruplase Alteplase (n = 236) (n = 237)
Death 11(4.7) 9 (3.8) Reinfarction 10(4.2) 10(4.2) Recurrent angina 41(17) 40(17) Coronary bypass surgery 9(3 8) 9(3 8) Angioplasty* 88 (37) 74(31) Severe bleeding 22 (9 3) 20 (8 4) Hemorrhagic stroke 2(0 8) 2 (0.8) Embolic stroke 2(0.8) 3(1.3) No major events 121 (51) 131 (55) "Some patients had >1 angioplasty Values are expressed as number (%).
90 Report of SESAM study group
Mortality and reinfarction rates were low. Patients who died were 5 years older than the study population. Most bleeding events were located at the catheter site: 13 of 22 patients and 14 of 20 patients, respectively. The rate of hemorrhagic stroke was the same in both groups. However, these patients were 11 years older than the total population.
Thrombolysis in myocardial infarction trial flow and clinical outcome: At the end of the first catheterization (including rescue angioplasty), 81% of the patients had TIMI 3 flow. The clinical outcome of the patients showed that patients with TIMI 0 to 1 flow had significantly higher mortality rates and lower reinfarction or recurrent angina than patients with TIMI 2 to 3 flow (Table 6). However, none of these 3 clinical parameters (alone or combined) was significantly different when patients with TIMI 2 and 3 flow were compared. Findings were independent of treatment allocation.
Table 6. Clinical Outcome and Thrombolysis In Myocardial Infarction Trial Flow at the Last View of the First Catheterization
TIMI 0 -1 TIMI 2 ТІМІЗ ρ Value
Number of patients 32(100) 55(100) 381 (100) 0.04* In-hospital death 4(13) 3(5.5) 13 (3.4) 0.44+ 0.63* Reinfarction* 0(0) 3 (5.5) 16(4.2) 0.72f 0.03* Recurrent angina* 1(3.1) 9(16) 70(18) 0.85f
»Difference between TIMI 0 to 1 and TIMI 2 to 3. difference between TIMI 2 and TIMI 3. *One saruplase-treated patient, who had recurrent angina and reinfarction, did not have afirst catheterization . Values are expressed as number (%). Abbreviation as in Table II.
Rescue angioplasty and clinical outcome: The group of patients in whom rescue angioplasty was successful had a mortality rate of 9.8%, a reinfarction rate of 3.4%, and a recurrent angina rate of 11.8%. Those with
91 Chapter 6 no rescue angioplasty had a mortality rate of 5.0%, reinfarction rate of 4.3%, and recurrent angina rate of 21.2%.
Discussion
Patency: Because one of the prime goals of thrombolytic therapy is to reopen vessels as soon as possible,6"9 it would appear that both saruplase and alteplase fulfill their role satisfactorily. The present study confirmed previous investigations,110 that saruplase results in fast reperfusion. At 45 minutes, 75% of the patients already had an open culprit artery. For the standard alteplase regimen, as used in this study, the usually quoted early patency rates (TIMI 2 or 3 flow) at 90 minutes vary between 64% and 76%." '2 Interestingly, the rates found in the present study are similar to those quoted for the front-loaded alteplase regimen of 81% to 91%.13"17 The Retaplase vs Alteplase Patency Investigation During myocardial infarction (RAPID) 1 and 2 studies18 showed slightly lower patency rates: 77% and 73%, respectively. However, patency rates for reteplase were significantly higher: 85% and 83%. The reason for the excellent patency rates of most of the more recent studies may be due to aggressive use of heparin. However, this is in contrast to the finding of Topol at al,19 that when heparin is given before thrombolysis the early acute patency rate changes very little although with continued heparinization the later patency (7 to 24 hours) is significantly better.20 Pretreatment with acetylsalicylic acid may also improve patency.
Reocclusion: Comparison of the reocclusion rates is valid because the patency rates of the last view of the first catheterization in patients not undergoing coronary angioplasty were virtually identical for saruplase- and alteplase-treated patients. Of patients undergoing a second cardiac catheterization and who were open without intervention at the first catheterization, only 2 patients (1.2%) treated with saruplase and 4 (2.4%) with alteplase had an angiographically documented reocclusion. For saruplase, this result again confirms the findings of a previous study,1 in which 1 of 108 patients had reocclusion as seen on angiography in the same time sequence. The outcome after alteplase treatment was clearly better than expected. During the SESAM study, 3 trials with similar design were published (2 with front-loaded alteplase) showing angiographically
92 Report of SESAM study group documented reocclusion rates of approximately 10% for alteplase.13"1 The Global Utilization of Streptokinase and TPA for Occluded arteries (GUSTO) trial17 showed 6% reocclusion rates 5 to 7 days after the onset of myocardial infarction in patients treated with front-loaded alteplase, but did not take into account patients not undergoing control angiography for medical reasons; the GUSTO trial rates agree with the 2.4% rate in this study. The adjunctive treatment was quite similar to this trial. Why the overall reocclusion rates in the SESAM study are low is unclear. It could be due to the adjunctive therapy of heparin and aspirin, or to better management of blood pressure (i.e., avoidance of hypotension).
Thrombolysis in Myocardial Infarction trial flow: Several studies have been published to indicate that patients with TIMI 2 flow at the end of the first cardiac catheterization have a poorer outcome than patients with TIMI 3 flow.21"26 In the present study, this finding could not be demonstrated because none of the usual clinical parameters (mortality, reinfarction, and recurrent angina) showed any difference between TIMI 2 and 3 patients (Table 4). However, there was an excess mortality in patients with TIMI 0 to 1 flow. The reason why the outcome of patients with TIMI 2 flow in this study is better than that reported in published studies is unclear. One explanation could be that the barrier between TIMI 1 and 2 flow is vaguely defined, and could be solved by frame counting. Another explanation may be the limited numbers of patients with TIMI 2 flow.
Rescue angioplasty: In this trial 65% of patients with TIMI 0 to 1 flow had rescue angioplasty. This percentage is comparable to previous trials with saruplase.2'6 In the Thrombolytic trial of Eminase in Acute Myocardial infarction 3 study, 43% of patients with TIMI 0 to 1 flow had such intervention.22 The beneficial effect of rescue angioplasty was previously demonstrated by Ellis et al.28 After successful rescue angioplasty the recurrence rate of angina (11.8% vs 21.2%) was lower than that in patients without angioplasty. The excess number of deaths (9.8% vs 5.0%) can be explained by the high rate of reocclusion after rescue angioplasty, and because rescue angioplasty is usually performed in patients with more extensive myocardial infarction.
Safety data: Safety data were similar for both saruplase and alteplase groups. The hospital mortality rates were low with both drugs, possibly
93 Chapter 6 because of the selection criteria (age <70 years, no cardiogenic shock) and the high and persistent patency rates. Incidence of recurrent angina and reinfarction rates during the hospital stay in this study were similar to other randomized trials with alteplase (2.6% to 4.0%).16,14 The stroke rates were comparable to those found in large randomized trials, bearing in mind the comparatively small size of this study. There is no indication that the stringent heparin regimen in this study compared with the GUSTO trial led to an increased risk of intracranial hemorrhage.17 Saruplase is not yet on the market. Its use will depend on cost/ benefit considerations.
Acknowledgment: Many people participated in the work presented in this article. We gratefully acknowledge the help of the departments of Cardiology of the following centers: С de Zwaan, MD, and coworkers of the Academic Hospital Maastricht, Maastricht, The Netherlands; coworkers of the Catharina Hospital, Eindhoven, The Netherlands; G. Pacouret, MD, and coworkers of Hôpital Trousseau, Tours, France; U. Hageleit, MD, and coworkers of the Marien Hospital, Wesel, Germany; coworkers of the Klinikum der Johannes Gutenberg Universität, Mainz, Germany; A. Betriu and coworkers of Hospital Clinico de Barcelona, Barcelona, Spain; coworkers of the Clinique Saint Joseph, Colmar, France; J. Zamorano, MD, and coworkers of Hospital Clinico de Madrid, Madrid, Spain; coworkers of the Hôpital de la Citadelle, Liège, Belgium; H. Bouchoucha, MD, and coworkers of the Centre Cardiologie du Nord, Paris, France; G.W. Tai, MD, and coworkers of the Stobhill General Hospital, Glasgow, United Kingdom; F.P. Jop, MD, and coworkers of the Klinikum der RWTH Aachen, Aachen, Germany; Α. Schuchert, MD, and coworkers of the Universtitätskrankenhaus Eppendorf, Hamburg, Germany; A. Caspi, MD, and coworkers of the Kaplan Hospital, Rehovot, Israel; T. Bonzel, MD, and coworkers of the Städtische Kliniken Fulda, Fulda, Germany; coworkers of the Reinier de Graef Hospital, Delft, The Netherlands; B. Eber, MD, and coworkers of the Medizinische Universitätsklinik, Graz, Austria; Prof. H.J. Trampisch. С. Schwarzweiler and coworkers of the Abteiling für Medizinische Informatik and Biomathematik, Bochum, Germany; monitors and other coworkers of Grüenthal GmbH, Aachen, Germany; and M. Muijtjens for her patience and typing of the manuscript.
94 Report of SESAM study group
APPENDIX
Participating centers (Department of Cardiology): University Hospital Maastricht, Maastricht, The Netherlands; Universitätsklinik Johannes Gutenberg, Mainz, Germany; Catharina Hospital, Eindhoven, The Netherlands; Centre Hospitalier Universitaire, Tours, France; Marien Hospital, Wesel, Germany; Hospital Clinico de Madrid, Madrid, Spain; Clinique Saint Joseph, Colmar, France; Hospital Clinico de Barcelona, Barcelona, Spain; Hôpital de la Citadelle, Liège, Belgium; Centre Cardiologie du Nord, Saint Denis/Paris, France; Stobhill General Hospital, Glasgow, United Kingdom; Klinikum der RWTH Aachen, Aachen, Germany; Universitätskrankenhaus Eppendorf, Hamburg, Germany; Kaplan Hospital, Rehovot, Israel; Städtische Kliniken, Fulda, Germany; Reinier de Graaf Hospital, Delft, The Netherlands; Medizinische Universitätsklinik, Graz, Austria; Abteilung fiir Medizinische Informatik und Biomathematik, Bochum, Germany; and Grünenthal GmbH, Aachen, Germany.
1. PRIMI Trial Study Group. Randomized double-blind trial of recombinant рго- urokinase against streptokinase in acute myocardial infarction. Lancet 1989;1:863-868. 2. The Belgian Saruplase Alteplase Trial Group. Effects of alteplase and saruplase on hemostatic variables: a single-blind, randomized trial in patients with acute myocardial infarction. Coron Artery Dis 1991;2:349-355. 3. Topoi EJ, Morris DC, Smalling RW, Schumacher RR, Taylor CR, Nishileuwer A, Liberman HA, Collen D, Tufte ME, Grossbarel EB, O'Neill WW. A multicenter, randomized, placebo-controlled trial of a new form of intravenous recombinant tissue-type plasminogen activator (activase) in acute myocardial infarction. J Am Coll Cardiol 1987;9:1205 - 1213. 4. Mueler HS, Rao AK, Forman SA. Thrombolysis In Myocardial Infarction (TIMI): comparative studies of coronary reperfusion and systemic fibrinogenolysis with two forms of recombinant tissue-type plasminogen activator. J Am Coll Cardiol 1987; 10:479 - 490. 5. Sherry S. Recombinant tissue plasminogen activator (rt-PA): is it the thrombolytic agent of choice in an evolving myocardial infarction? Am J Cardiol 1987;59:984-989. 6. Dalen JE, Gore JM, Braunwald E, Borer J, Goldberg RJ, Passamani ER, Forman S, Knatterud G. Six and twelve-month follow-up of the phase I
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Thrombolysis In Myocardial Infarction (TIMI) trial. Am J Cardiol 1988;62:179-185. 7. Vermeer F, Simoons ML, Bär FW, Tijssen JG, Domburg RT, Semiys PW, Verheugt FW, Res JC, de Zwaan С, van de Laarse A, Krauss XH, Lubsen J, Hugenholtz PG. Which patients benefit most from early thrombolytic therapy with intracoronary streptokinase? Circulation I986;74:1379 - 1389. 8. Bär FW, Frederiks J, de Swart H, van Ommen V, de Zwaan С, Vermeer F, Gorgels AP, Wellens HJJ. Comparison between acute PTCA after failed thrombolysis and immediate PTCA without thrombolysis in acute myocardial infarction. J Am Coll Cardiol 1994;405A:954 - 943. 9. Bär FW, Vermeer F, De Zwaan С, Ramentol M, Braat S, Simoons ML, Hermens WT, van der Laarsse A, Verheugt FW, Krauss XH, Wellens HJJ. Value of admission electrocardiogram in predicting outcome of thrombolytic therapy in acute myocardial infarction. Am J Cardiol 1987;59:6 - 13. 10. Tebbe U, Windeler J, Boesl I, Massberg I, Barth H, on behalve of the LIMITS study group. Thrombolysis with recombinant unglycosylated single chain urokinase-type plasminogen activator (saruplase) in acute myocardial infarction: influence of heparin on early patency rate (LIMITS study). J Am Coll Cardiol 1995;26:365 - 373. 11. Grines CL, Nissen SE, Booth DC, Gurley JC, Chelliah N, Wolf R, Blankenship J, Branco MC, Bennett К, DeMaria AN. A prospective, randomized trial comparing combination half-dose tissue-type plasminogen activator and streptokinase with full-dose tissue-type plasminogen activator. Circulation 1991;84:540-549. 12. Carney RJ, Murphy G A, Brandt TR, Daley PJ, Pickering E, White HJ, McDonough TJ, Virmilya SK, Teichman SL. Randomized angiographic trial of recombinant tissue-type plasminogen activator (alteplase) in myocardial infarction. J Am Coll Cardiol 1992;20:17 - 23. 13. Neuhaus KL, Tebbe U, Gottwik M, Weber M A J, Feuerer W, Niederer W, Haerer W, Praetorius F, Grosser KD, Huhmann W, Hoepp HW, Alber G, Sheikhzadeh A, Schneider B. Intravenous recombinant tissue plasminogen activator (rt-PA) and urokinase in acute myocardial infarction: results of the German Activator Urokinase Study (GAUS). J Am Coll Cardiol 1988; 12:581 -587. 14. Neuhaus KL, Von Essen R, Tebbe U, Vogt A, Roth M, Riess M, Niederer W, Forycki F, Wirtzfeld A, Maeurer W, Limbourg P, Merx W, Haerten K. Improved thrombolysis in acute myocardial infarction with front loaded administration of alteplase: results of the rt-PA-APSAC Patency Study (TAPS). J Am Coll Cardiol 1992;19:885 - 891. 15. Neuhaus KL, Feuerer W, Jeep-Tebbe S, Niederer W, Vogt A, Tebbe U. Improved thrombolysis with a modified dose regimen of recombinant tissue- type plasminogen activator. J Am Coll Cardiol 1989; 14:1566 - 1569.
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16. Ross AM, Lundergan С, Thompson M, Reiner J, Deychak Y, Rohrbeck S, Cooyne K, Walker P, Cho S, Greenhouse S. Lee К, Granger С, Wildermann Ν, Fink С, Harry S, Allison D, Draoui Y, Corstigan G, Fox D. The effects of tissue plasminogen activator, streptokinase, or both on coronary-artery patency, ventricular function, and survival after acute myocardial infarction. ./V EnglJMed 1993;329:1615 - 1622. 17. The GUSTO angiographic investigators. The effects of tissue plasminogen activator, streptokinase, or both on coronary artery patency, ventricular function, and survival after acute myocardial infarction. ./V Engl J Med 1993;329:1615-1622. 18. Weaver WD. Results of the RAPID 1 and RAPID 2 thrombolytic trials in acute myocardial infarction. Eur Heart J1996; 17(suppl E): 14 - 20. 19. Topol EJ, George BS, Kereiakes DJ, Stump DC, Candela RJ, Abbottsmit CW, Aronson L, Pickel A, Boswick JM, Lee KL, Ellis SG, Califf RM. A randomized controlled trial of intravenous tissue plasminogen activator and early intravenous heparin in acute myocardial infarction. Circulation 1989;79:281-286. 20. Hsia J, Kleiman Ν, Aguirre F, Chairman BR, Roberts R, Ross AM. Heparin- induced prolongation of partial thromboplastin time after thrombolysis: relation to coronary artery patency. J Am Coll Cardiol 1992;20:31 - 35. 21. Karagounis LA, Sorensen SG, Menlove RL, Moreno F, Anderson JL. Does thrombolysis in myocardial infarction (TIMI) perfusion grade 2 represent a mostly patent artery or a mostly occluded artery? Enzymatic and electrocardiographic evidence from the TEAM 2 study. J Am Coll Cardiol 1992;19:1 -10. 22. Anderson JL, Karagounis LA, Becker LC, Sorensen SG, Menloe RL. TIMI perfusion grade 3 but not grade 2 results in improved outcome after thrombolysis for myocardial infarction. Ventriculographic, enzymatic and electrocardiographic evidence from the TEAM 3 study. Circulation 1993;87:1829-1839. 23. Lincoff AM, Topol EJ. Illusion of reperfusion. Does anyone achieve optimal reperfusion during acute myocardial infarction. Circulation 1993;87:1792 - 1805. 24. Vogt A, von Essen R, Tebbe U, Feuerer W, Appel KF, Nauhaus KL. Impact of early perfusion status of the infarct-related artery after thrombolysis for acute myocardial infarction: retrospective analysis of four German multicentre studies. J Am Coll Cardiol 1993;21:1391 - 1395. 25. Ohman EM, Califf RM, Topol EJ, Candela R, Abbottsmith C, Ellis S, Sigmon KN, Kereiakes D, George B, Stack R. Consequences of reocclusion after successful reperfusion therapy in acute myocardial infarction. Circulation 1990;82:781-791.
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26 Anderson JL, Karagounis LA, Becker LC, Sorensen SG, Menlove RL TIMI perfusion grade 3 but not grade 2 results in improved outcome after thrombolysis for myocardial infarction Circulation 1993,87 1829- 1839 27 Bar FW, Meyer J, Uebis R, Lange S, Barth H, Groves R, Vermeer F The effect of taprostene in patients with acute myocardial infarction treated with thrombolytic therapy Results of the "START" study Eur Heart J 1993,14 1118-1126 28 Ellis SG, da Silva ER, Heyndrickx G, Talley JD, Cemigliaro C, Steg G, Spauldmg С Nobuyoshi M, Erbel R, Vassanelli C, Topoi EJ Randomized comparison of rescue angioplasty with conservative management of patients with early failure of thrombolysis for acute anterior myocardial infarction Circulation 1994,90 2280-2284
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Randomized, Double-Blind Study Comparing Saruplase With Streptokinase Therapy in Acute Myocardial Infarction: The COMPASS Equivalence Trial
Ulrich Tebbe, MD, Rolf Michels, MD, Jennifer Adgey, MD, Frcp, Jean Botami, MD, Avi Caspi, MD, Bernard Charbonnier, MD, Jürgen Windeler, MD, Hannes Barth, MD, Robert Groves, PhD, Gwyn R. Hopkins, BSc, MRCP, MFPM, William Fennell, MD, FRCPI, Amadeo Betriu, MD, Mikhail Ruda, MD, Johannes Mlczoch, MD.
For the comparison trial of saruplase and streptokinase (COMPASS) investigators
Lippe-Detmold, Bochum and Aachen, Germany, Eindhoven, The Netherlands, Belfast, Northern Ireland, United Kingdom, Liège, Belgium, Rehovot, Israel, Tours, France, Cork, Ireland, Barcelona, Spain, Moscow, Russian Federation, and Vienna, Austria
Abstract: Objectives. This study sought to demonstrate the equivalence of saruplase and streptokinase in terms of 30-day mortality Background. The use of thrombolytic agents m the treatment of acute myocardial infarction is well established and has been shown to substantially reduce post- myocardial infarction mortality Methods. Three thousand eighty-nine patients with symptoms compatible with those of acute myocardial infarction for <6 h entered the study at a total of 104 centers and were randomized to receive streptokinase (1 5-MU infusion over 60 mm) or saruplase (20-mg bolus and 60-mg infusion over 60 mm) In the saruplase group, a bolus of heparin (5,000 IU) was administered before saruplase, and a corresponding blinded double-dummy placebo bolus was administered before streptokinase, All patients received intravenous heparin infusions for >24 h starting 30 mm after the end of the thrombolytic infusions, the infusions were titrated to maintain an activated partial thromboplastin time at 1 5 to 2 5 times that of normal Results. Death of any cause up to 30 days after randomization occurred in 88 (5 7%) of 1,542 patients randomized to receive saruplase and 104 (6 7%) of 1,547 patients randomized to receive streptokinase (odds ratio 0 84, ρ < 0 01 for equivalence) Hemorrhagic strokes occurred more often in patients receiving saruplase (0 9% vs 0 3%), whereas thromboembolic strokes were more prevalent
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in the streptokmase-treated patients (0 5% vs. 1 0%) The rate of bleeding was similar in the two treatment groups (10,4% vs 10 9%) Hypotension and cardiogenic shock occurred less frequently in the saruplase group Reinfarction rates were similar Conclusions. Saruplase is a clinically safe and effective thrombolytic medication This profile ranks saruplase favorably among the currently available thrombolytic agents
The use of thrombolytic agents in the treatment of acute myocardial infarction is well established (1); the most widely used thrombolytic agent in Europe is streptokinase. Newer thrombolytic agents have been developed based on endogenous plasminogen activators, including tissue- type plasminogen activator (t-PA [e.g., alteplase]) and urokinase-type plasminogen activator (u-ΡΑ [e.g., saruplase]). Saruplase, a recombinant, unglycosylated, human, single-chain u-ΡΑ is a protein of known amino acid sequence that is produced through the use of genetically transformed Escherichia coli bacteria (2). Because saruplase has an amino acid structure that is identical to that of endogenous u-ΡΑ, there should be no risk of allergic reactions or antigenicity, as occurs with xenobiotic compounds like streptokinase. Saruplase has been shown to be more efficacious than streptokinase in restoring coronary artery patency (3). Although patency trials may demonstrate superiority in terms of clot lysis times and restoration of perfusion of the ischemic myocardium, because of their limited statistical power they cannot confirm diferences in survival rates. Conventional mortality trials require many tens of thousands of patients to demonstrate the superiority of one thrombolytic agent over another. In the present study, we took a novel approach to demonstrate the efficacy of a new agent by demonstrating that infarct survival is at least as good as that with a current standard. The results of a similar trial designed to show the equivalence of reteplase with streptokinase that began in August 1993 have heen recently reported (4).
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Abbreviations and Acronyms
APTT = activated partial thromboplastin time ASA = acetylsalicylic acid CABG = coronary artery bypass graft surgery COMPASS = Comparative Trial of Saruplase Versus Streptokinase GUSTO Global Utilization of Streptokinase and TPA [tissue-type Plasminogen activator] for Occluded Coronary Arteries INJECT International Joint Efficacy Comparison of Thrombolytics trial PTCA = percutaneous transluminal coronary angioplasty TIMI = Thrombolysis in Myocardial Infarction t-PA = tissue-type plasminogen activator u-PA urokinase-type plasminogen activator
Methods
Study organization. A total of 104 centers in 10 West European countries, Russia and Israel participated in the study (see Appendix). Ethics committee appraisal of the protocol was obtained for all centers involved from the hospital or regional or national institutional review board. International logistics coordination was performed by the sponsor (Grünenthal GmbH). An international steering committee met at regular intervals to review the course of the study. The study was conducted according to good clinical practice guidelines (5), and source data auditing was routinely performed.
Patients. Male or female patients >20 years old presenting to a study center within 6 h after the onset of acute myocardial infarction symptoms were screened for entry into the study. Eligible patients exhibited nitrate- resistant chest pain lasting for >30 min, with ST segment elevation >0.1 mV in two or more frontal plane leads or >0.2 mV in two or more precordial leads. Exclusion criteria included severe hypertension (systolic blood pressure >200 mm Hg or diastolic blood pressure >100 mm Hg on admission or hypertensive retinopathy grade HI or IV) or known intracranial, gastrointestinal, clotting, hepatic, pulmonary, renal, urogenital, vascular or other disorders that could increase the risk of
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bleeding after thrombolysis. Patients with major trauma or surgical procedures within 1 month were excluded, as were those with malignancy or known sensitivity to streptokinase or exposure to streptokinase within the past year or streptococcal infection within the past 3 months. In June 1993 (after -800 patients had been entered), the protocol was amended to allow patients >75 years old to enter the study. All patients gave witnessed or written informed consent.
Inclusion/exclusion criteria Informed consent 1 ASA (200-400 mg, po oriv) І Heparin placebo bolus ι V 5000 IU (to hepann)
Saruplase bolus ι ν placebo 20 mg/1 mm t = 0h (lo saruplase) I Saruplase Streptokinase 60 mg/ 60 mm 1 5x1CWlU/60min parallel ι ν + + infusions placebo placebo (to streptokinase) (to saruplase)
hepann ι ν infusion (a)PTT 1 5 to 2 5x normal ASApo >75mg/day I 30 day follow-up I 1 year follow-up
Figure 1. Study protocol for the administration of trial medication, iv (i.v.) = intravenous; po = oral.
Randomization and treatment strategy. The investigators used numbered, prerandomized medication kits in ascending order. The medication was supplied by Grünenthal GmbH, Aachen, Germany. Each kit contained either 1) heparin (5,000 IU for initial intravenous bolus), saruplase (20-mg vial for intravenous bolus and 60 mg for 60-min intravenous infusion) and streptokinase placebo, or 2) placebo to heparin (for initial intravenous bolus), streptokinase (1.5 MU for 60-min intravenous infusion) and saruplase placebo (Fig. 1 ). Additional therapy. Oral or intravenous acetylsalicylic acid (ASA) was administered as a loading dose before thrombolysis (200 to 400 mg), followed by a daily oral maintenance dose of >75 mg.
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Heparin was administered to all patients for >24 h, starting as an intravenous infusion 30 min after the end of the thrombolytic infusions. The starting dose was 15 IU/kg per h, with dose titration to achieve an activated partial thromboplastin time (aPTT) of 1.5 to 2.5 times the control value. The use of intravenous nitrates was recommended. All other medication administered in the hospital, including additional thrombolytic agents, was permitted if considered necessary and was documented. Invasive coronary procedures, including percutaneous transluminal coronary angioplasty (PTCA) and coronary artery bypass graft surgery (CABG), were permitted at the investigator's discretion. Clinical data, end points and follow-up. The primary end point was death from any cause at 30 days after the start of study treatment. Other prospectively defined end points up to 30 days were cerebral events, reinfarction and reintervention, such as PTCA, CABG or further thrombolytic treatment. In-hospital end point events included bleeding, hypotension and allergic reactions. Patients were followed up for 1 year. To determine possible antibody formation to the thrombolytic medication, blood samples were taken before treatment, at hospital discharge and at the 1-month follow-up visit. Analysis was performed at Grünenthal GmbH. Data management and quality assurance. Patient data were collected anonymously to retain confidentiality. All case report forms were checked at monitoring visits for accuracy, consistency and completeness on the basis of hospital files. Audits were performed at randomly chosen centers. Statistical analysis. The medical faculty of the University of Bochum, Germany, Institute for Information Technology and Biomathematics served as the study data and statistics center. The trial was designed as an equivalence trial to test a preformulated definition of the clinical equivalence of a saruplase regimen with that of the standard streptokinase regimen on the basis of 30-day mortality rates. Equivalence was defined as an odds ratio <1.5 for all-cause mortality at 30 days for all patients randomized. Using an alpha value of 0.05 (one-tailed), a power of 80% and a generalized Fisher exact test, it was calculated that 1,500 patients/treatment group would be necessary. Because the study was adequately powered to detect equivalence only by this a priori definition, no conclusions can be drawn as to the superiority of saruplase in this respect.
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Results
Patients. Between August 1992 and July 1994, a total of 3,089 patients were entered into the study at 104 hospitals in 12 countries: Austria (82 patients), Belgium (105 patients), United Kingdom (252 patients), France (127 patients), Germany (550 patients), Ireland (144 patients), Israel (332 patients), Italy (18 patients), The Netherlands (1,038 patients), Portugal (20 patients), Russian Federation (177 patients) and Spain (245 patients).
Table 1. Clinical Baseline Characteristics of Patients Receiving Saruplase or Streptokinase
Saruplase SK 90% CI of Ρ Value (n = 1,542) (n = 1,547) Difference
Age (yr) 59.3 (10.7) 60.4(10.8) -1.7 to-0.5 0.004 Men 80.1% 79.3% -1.6 to 3.2 Height (cm) 171.0(8.1) 170.8 (8.2) - 0.3 to 0.7 0.582 Weight (kg) 76.9(12.8) 77.1 (12.0) -1.0 to 0.6 Pulse rate (min"1) 74.4(17.0) 75.2(16.9) -1.8 to 0.2 0.167 SBP (mm Hg) 134.1 (24.0) 134.6(23.2) -1.9 to 0.9 DBP (mm Hg) 80.0(13.6) 80.1 (13.4) -0.9 to 0.7 Killip class I 85.2% 85.5% II 12.9% 13.0% III 1.1% 0.7% IV 0.8% 0.7% Time from Sx to 181 (83) 183(82) -6.9 to 2.9 study Tx (min)
Data presented are mean (SD) or percent of patients. CI = confidence interval; DBP = diastolic blood pressure; SBP = systemic blood pressure; SK = streptokinase; Sx = symptom; Tx = therapy,
The treatment groups were well matched at baseline (Tables 1 and 2). Only one baseline variable - age - differed (p < 0.05) between the groups. The diagnosis of myocardial infarction was confirmed in 96.7% of patients according to World Health Organization criteria of enzyme changes,
104 Report of COMPASS investigators symptoms and electrocardiography. A history of previous myocardial infarction was reported for 14.4% of patients (Table 2). Study medications. The loading dose of ASA was administered in 95.6% of patients before the start of the study therapy, and an additional 1.1% of patients had taken ASA a few hours before admission. The intravenous heparin (5,000 IU) or placebo bolus was administered to 99.1% of patients. Infusion of the randomly assigned double-blind thrombolytic agent was started in 99.4% of patients. Intravenous heparin infusions, targeted to an activated partial prothrombin time of 1.5 to 2.5 times normal levels, were started in 99.0% of patients and continued for a median time of 5 days in both treatment groups. Intravenous nitrate infusion was started before lysis in 79.6% of patients. The prevalence of early concomitant therapy was as follows (saruplase vs. streptokinase): 61.4% versus 62.6% for beta-adrenergic blocking agents; 27.8% versus 26.0% for calcium channel blocking agents; and 30.7% versus 31.9%» for angiotensin-converting enzyme inhibitors, with no marked difference in use between the groups.
Table 2. Comparability of Previous Medical History of Patients Receiving Saruplase or Streptokinase at Randomization
Saruplase SK 90% CI of Ρ Value (n = 1,542) (n = 1,547) Difference
Infarct site AL 45.4% 47.2% -5.4 to 1.8 IP 54.4% 52.7% -1.9 to 5.3 MI confirmed 96.9% 96.6% - 1.0 to 1.6 Prev Ml 14.2% 14.7% -3.0 to 2.1 Prev lysis Tx 3.6% 2.8% - 0.5 to 2.2 0.179 Diabetes 12.1% 10.6% -0.8 to 3.8 0.181 Hypertension 24.8% 25.0% -3.4 to 2.9 Angina 14.5% 13.5% -1.5 to 3.5 Current smoker 53.8% 49.6% 0.6 to 7.8 0.018
Data presented are percent of patients. AL = anterolateral; IP = inferoposterior; Ml = myocardial infarction; Prev = previous; other abbreviations as in Table 1.
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Primary end point. Death from any cause up to 30 days after randomization (primary study end point) occurred in 88 (5.7%) of 1,542 patients randomized to the saruplase group and in 104 (6.7%) of 1,547 patients randomized to the streptokinase group (odds ratio 0.84, ρ < 0.01 for equivalence) (Table 3, Fig. 2). The median time to death in the saruplase treatment group was 2.4 days, and in that for the streptokinase treatment group was 2.8 days. The main cause of death was cardiogenic shock. The 1-year mortality rate was 8.2% (126 of 1,528) for saruplase treatment and 9.6% (147 of 1,530) for streptokinase treatment.
Table 3. Causes of Death in Patients Receiving Saruplase or Streptokinase
Saruplase SK 90% CI of Ρ Value (n = 1,542) (n = 1,547) Difference
30-day mortality 88 (5.7%) 104(6.7%) -2.8 to 0.8 0.242 1-yr mortality 126 (8.2%) 147 (9.6%) -3.4 to 0.7 0.193 Main cause of death at 30 days Ventricular rupture/ 19(21.6%) 7 (6.7%) 4.0 to 25.8 0.003 tamponade Cardiogenic shock 29 (33.0%) 44 (42.3%) -24.1 to 5.4 0.183 Ventricular 18(20.5%) 28 (26.9%) -19.5 to 6.6 0.295 fibrillation/asystole Stroke 10(11.4%) 5 (4.8%) -2.3 to 15.4 0.092 Other 12(13.6%) 20(19.2%) -17.1 to 5.9 0.300
Data presented are number (%) of patients. Abbreviations as in Table 1.
Other clinical end points. Hypotension and cardiogenic shock were more common in the streptokinase-treated patients (Table 4). The incidence of arrhythmias and angina was similar in the two treatment groups. The rate of reinfarction was marginally higher in the saruplase treatment group and was 5.4% versus 4.5% at 30 days and 8.2% versus 7.1% at 1 year for saruplase versus streptokinase. PTCA was performed in 11.3% versus 10.3%) of patients and CABG in 3.4% versus 3.9%) (saruplase vs. streptokinase) during the first 30 days.
706 Repon of COMPASS investigators survival distribution function estimate 1,00 0.99 At 30 days saruplase 5 71% 0,98 streptokinase 6 72% 0,97 - difference (SA-SK) -1 01% 95% CI of difference [-2 78%, 0 75%] 0,96 0,95 0,94 0,93 0,92 0,91 - 0,90 - 0,89 - 0,88 τ τ т ι г 50 100 150 200 250 300 350 365 time after entry to trial (days)
Figure 2. One-year mortality rate Kaplan-Meier curves for patients randomized to receive saruplase (Sa [solid curve]) or streptokinase (Sk [dashed curve]) CI = confidence interval.
There was no difference in bleeding rate between the treatment groups. Severe bleeding was seldom seen, and the need for transfusion was rare (Table 5). The overall incidence of strokes (1.4% versus 1.4%) within the first 30 days was similar to that in other trials in which thrombolytic agents have been used (Table 6). Hemorrhagic strokes occurred more often in the saruplase group (0.9% versus 0.3%), whereas thromboembolic strokes were more prevalent in the streptokinase group (0.5% versus 1.0%). There was a slight tendency for more patients to die of stroke after treatment with saruplase (13 of 22 versus 9 of 21). The incidence of transient ischemic attacks was the same for both groups. Additional strokes were reported from day 30 to 1 year (1.1% versus 1.0%) and were mostly thromboembolic (0.7% versus 0.5%) or unspecified (0.3% versus 0.3%). Reactions that may have had an underlying allergic cause were reported in 1.6% of saruplase-treated patients and in 4.1% of streptokinase-treated patients (p < 0.001). Severe reactions were reported
107 Chapter 7 in two saruplase-treated patients (both >3 days after treatment and not thought by the investigator to be related to the trial medication) and 12 streptokinase-treated patients (all within the first day and in 11 thought by the investigator to be at least probably related to the trial medication). In >250 blood samples from saruplase-treated patients, no antibody was detected to saruplase or any possible contaminating E. coli protein. All samples (both at discharge and at 30 days) taken from the streptokinase- treated patients demonstrated antibody formation.
Table 4. Cardiac Events up to 30 Days in Patients Receiving Saruplase or Streptokinase
Saruplase SK 90% CI of Ρ Value (n = 1,542) (n = 1,547) Difference VF 99 (6.4%) 126(8.1%) -3.6 to 0.2 VT 276(17.9%) 275(17.8%) -2.6 to 2.9 0.065 Bradycardia 340 (22.0%) 306(19.8%) -0.7 to 5.2 Re infarction 83 (5.4%) 69 (4.5%) -0.7 to 2.5 0.121 Pericarditis 56(3.6%) 42 (2.7%) -0.4 to 2.2 0.236 Hypotension 484(31.4%) 590(38.1%) -10.2 to-3.3 0.146 Cardiogenic 51 (3.3%) 71 (4.6%) -2.7 to 0.2 0.001 shock Angina 426 (27.6%) 427 (27.6%) -3.2 to 3.2 0.067
Data presented are number (%) of patients. VF = ventricular fibrillation; VT = ventricular tachycardia; other abbreviations as in Table 1.
108 Report of COMPASS investigators
Table 5. In-Hospital Bleeding Events (excluding stroke) in Patients Receiving Saruplase or Streptokinase
Saruplase SK 90% CI of Ρ Value (n = 1,542) (n = 1,547) Difference
Mild 160(10.4%) 169(10.9%) -2.8 to 1.7 Moderate 106(6.9%) 109(7.0%) -2.0 to 1.7 Severe 32(2.1%) 38 (2.5%) -1.5 to 0.7 0.477 GI 14 9 Urogenital 1 2 Puncture site 12 20 Hb drop/anemia 5 7
Data presented are number or number (%) of patients. Gl = gastrointestinal; Hb hemoglobin; other abbreviations as in Table 1.
Table 6. Strokes and Transient Ischemic Attacks at 30 Days in Patients Receiving Saruplase or Streptokinase
Saruplase SK 90% CI of Ρ Value (n = 1,542) (n = 1,547) Difference
TIA 7 (0.5%) 8 (0.5%) -0.6 to 0.5 All strokes 22(1.4%) 21 (1.4%) -0.8 to 1.0 0.840 Hemorrhagic 14 (0.9%) 5 (0.3%) -0.0 to 1.2 0.038 Thromboembolic 8 (0.5%) 15(1.0%) -1.1 to 0.2 0.145 Unclassified 0 (0.0%) 1 (0.1%) -0.3 to 0.1 Outcome of stroke Death up to 30 13 (0.9%) 9 (0.6%) - 0.4 to 0.9 days Disabling 5 (0.3%) 6 (0.4%) - 0.5 to 0.4 No deficit 5 (0.3%) 7 (0.4%) - 0.6 to 0.4
Data presented are numher (%) of patients. Abbreviations as in Table 1.
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Discussion
Concomitant medication. The standard saruplase treatment regimen includes a previous heparin bolus (5,000 IU) and was shown in the Liquemin in Myocardial Infarction During Thrombolysis With Saruplase (LIMITS) trial (6) to be associated with enhanced thrombolytic efficacy at no marked cost to the safety of saruplase. Heparin placebo was given before streptokinase because this is not the standard procedure for streptokinase therapy (7). Therefore, the present study was a comparison of two treatment regimens. Heparin was administered to all patients after thrombolysis. This heparin regimen was chosen because at the time of the study, heparin was considered an important adjunctive therapy and was widely used, and although not of proven use with streptokinase, it appeared to be safe based on the Global Utilization of Streptokinase and TPA for Occluded Coronary arteries (GUSTO) study (8), albeit associated with a tendency for an increased incidence of intracranial hemorrhage. Primary end point, study size and statistics. Hospital mortality rates after myocardial infarction have been declining steadily over the past 20 years (9), and thrombolytic therapy has certainly been a contributing factor. Among patients with definite electrocardiographic ST segment elevation who presented early after symptom onset and were considered to be eligible for thrombolytic therapy, a mortality rate of 6% to 10% at 1 month may be expected after streptokinase therapy (4,8,10,11). To further lower this already low rate, any new thrombolytic agent must be greatly superior to streptokinase. In the GUSTO study (8), >40,000 patients were treated in four study arms to eventually show a 1% increase in survival benefit of t-PA over streptokinase. The tremendous consumption of resources in these trials led to reluctance to develop new thrombolytic drugs with a potentially more favorable safety/ efficacy relation. Innovative statistical designs to demonstrate clinically defined equivalence, rather than superiority, are gaining recognition by drug regulatory authorities as a way of preventing this stagnation of thrombolytic drug research and development. The present study was a test of clinical equivalence, defined as an odds ratio <1.5 for the 30-day mortality rate after treatment with saruplase or streptokinase. As in the International Joint Efficacy Comparison of Thrombolytics (INJECT) study (4), it was presumed during the planning of the study that there was a small mortality benefit over streptokinase. The collaborators of the INJECT study performed a meta-analysis of 13
110 Repon of COMPASS investigators placebo-controlled studies of thrombolysis showing a mean mortality difference of 2.7% at 35 days. The lower limit of the one-sided 95% confidence interval was 2.1%. In the COMPASS study, the mortality diference (saruplase minus streptokinase) was - 1%, with a 95% confidence interval of - 2.7% to 0.7%. Although the final study size was calculated on an odds ratio of 1.5 (mortality under streptokinase 7%, alpha = 5% and beta = 20%), the results of the study showed an odds ratio of 0.84. The upper limit of the one-sided 95% confidence interval was 1.09%, and that for a one-sided 97.5% confidence interval was 1.14%. Therefore, a placebo response of >2% can be excluded. In the end, both the present and the INJECT studies showed that the mortality rate for the new thrombolytic agent was at least equivalent to that for streptokinase. Patency and mortality. A major breakthrough in comparative thrombolysis trials was the confirmation of the open artery hypothesis by the angiographic arm of the GUSTO study, in which it was shown that early restoration of coronary artery patency (Thrombolysis in Myocardial Infarction [TIMI] grade 3 angiographic perfusion) is a powerful predictor of survival at 30 days after thrombolysis (12,13). On this basis, the trend seen in the present study to lower mortality in the saruplase group (5.7%) compared with the streptokinase group (6.7%) is reinforced by the results of a previous coronary artery patency trial comparing saruplase therapy with streptokinase (3). In the double-blind Prourokinase in Myocardial Infarction trial (PRIMI) trial of 401 patients, patency (TIMI grade 2 or 3 angiographic flow) was more rapidly restored with saruplase than with streptokinase. At 60 min after the start of therapy, the patency rate was 48.0% for patients in the streptokinase group versus 71.8%» for patients in the saruplase group (p < 0.001); by 90 min, the respective patency rates were 63.9% and 71.2%, respectively (p = 0.15). The high patency rates with low reocclusion rates have been confirmed in a study of saruplase versus alteplase (14). The 90-min patency rates were 80% for patients receiving saruplase and 81% for patients receiving alteplase, with reocclusion rates of 1.2% and 2.4%, respectively. It is very probable that the trend to lower mortality rates in the saruplase group in the COMPASS trial is indeed real. The primary study end point was achieved because the standard saruplase regimen was proved to be at least clinically equivalent to streptokinase (p < 0.001) in terms of 30-day survival rates. In comparing the reference (streptokinase) group outcomes with those from other randomized trials, a comparable 30-
111 Chapter 7
day mortality rate was seen in the GUSTO trial (8) (7.4% for the streptokinase group with intravenous heparin versus 6.7% in the COMPASS trial), but a slightly higher rate (9.5%) occurred in the INJECT trial (4). Strokes. From a clinical perspective, treatment with saruplase is at least equivalent and tends toward a 15% relative reduction in mortality compared with streptokinase therapy. Because this is all-cause mortality, it already includes a possible increase in fatal hemorrhagic strokes, which arguably may be related to an increased thrombolytic efficacy itself. Due to the increased rate of severe nonfatal hemorrhagic strokes possibly associated with the more efficacious thrombolytic regimens, it has been argued that a more revealing outcome variable is the combined end point of death or disabling stroke. Disabling stroke is defined as stroke that results in a patient who is no longer able of looking after himself or herself, which could have an economic impact. The number of disabling strokes was almost the same in both treatment groups, and there is therefore no change in the trend seen with the mortality data in favor of saruplase. Stroke rates overall were similar between the two treatment groups, but there is a distinct trend in the saruplase treatment group toward a lower incidence of thromboembolic strokes and a higher incidence of hemorrhagic strokes than with the streptokinase group. Speculative stroke classification was avoided; only when computer tomography or an autopsy was performed and the etiology was confirmed was a classification made. Others are termed "unclassified." Hemorrhagic stroke rates for streptokinase have been reported of 0.1% to 0.5% (8,10,11,15,16), and for thromboembolic strokes, the rates are 0.7% to 1.3%. Therefore, the results with streptokinase in this study are in broad agreement with the published data. Although the precise mechanism of action of saruplase is not known, ~30% of the dose administered in the regimen used in this study is converted to two-chain u-PA (17), which probably contributes significantly to lysis and side effects. Allergic-type reactions. A number of possible allergic reactions were reported in patients treated with saruplase, but no antibody directed toward saruplase was detected; these reactions could be attributed to other procedures or medications, such as ASA. All samples taken from patients administered streptokinase demonstrated antibody formation, and the incidence of both severe and other allergic-type reactions was much higher than that for the saruplase-treated group.
112 Report of COMPASS investigators
Conclusions. The present study demonstrated that the 30-day mortality rate for saruplase treatment is at least equivalent to that for streptokinase treatment, with an overall stroke rate and outcome after stroke that appear to be similar for the two treatments
Appendix
Participating Investigators and Institutions for the COMPASS Trial
Steering Committee Members and National Coordinators Prof U Tebbe, Detmold, Germany (Chairman), Prof A A J Adgey, Belfast, United Kingdom, Dr J Boland, Liege, Belgium, Dr A Caspi, Tel Aviv, Israel, Prof В Charbonnier, Tours, France, Dr W Fennell, Cork, Ireland, Dr A Betriu, Barcelona, Spam, Dr R Michels, MD, Eindhoven, The Netherlands, Prof M Y Ruda, Moscow, Russian Federation, Prof J Mlczoch, Vienna, Austria Safety Committee Prof D de Bono, Leicester, United Kingdom, Prof В Charbonnier, Tours, France, Prof H Schmutzler, Berlin, Germany Trial Management and Support Prof Dr J Windeler (Statician, Bochum/ Heidelberg, Germany), G R Hopkins, Dr R Groves (Coordinators, Grunenthal GmbH, Germany), Dr S Barnes, A Dreßen, Dr J Goldberg, D Weber, A Schuckelt, A Plum, С Tabbert, S Classen, M von Fisenne (Monitors, Grünenthal GmbH, Germany) Investigators: Austria Prof W Klein, Med Universitätsklinik, Graz, Prof Ρ Kühn, Krankenhaus der Barmherzigen Schwestern, Linz, Prof J Mlczoch, Krankenhaus Lamz, Vienna Belgium Prof L Bossaert, Academisch Ziekenhuis UIA, Antwerp, Dr R Ranquin, Middelheim Ziekenhuis, Antwerp Dr M Quinonez, Centre Hospitalier du Bois de l'Abbaye, Seraing, Dr С Degauque, L'Hôpital Civil deVerviers, Verviers France Dr M, Lang, Centre Hospitalier General de Blois, Blois, Dr G Doll, Hôpital François Rabelais, Chinon, Dr H Lardoux, Centre Hospitalier Gilles de Corbeil, Corbeil Essonnes, Prof С Thery, Hôpital Cardiologique, Lille, Dr G Laval, Hôpital du Hasenrain, Mulhouse, Prof J Hertault, Centre Hospitalier Regional et Universitaire, Nîmes, Dr Ρ Deutsch, Hôpital Broussais, Saint Malo, Prof R Haiat, Centre Hospitalier General, Samt Germain-en-Laye Germany Dr В Ducker, St Manen-Hospital, Ahaus, Dr W Lieb, Akademisches Lehrkrankenhaus, Alfeld, Dr С Bethge, Judisches Krankenhaus Berlin, Berlin, Prof W Jaedicke, Evang Krankenhaus, Castrop- Rauxel, Dr H Heuer, St Johannes Hospital, Dortmund, Dr W W Wucherpfennig, Krankenhaus St Martini, Duderstadt, Dr H Bock, Franz- Hospital, Duimen, Dr H Kronert, Kreiskrankenhaus, Eschwege, Dr W Himmel, Evang Krankenhaus, Bad Gandersheim Dr E Goede, Kreiskrankenhaus, Goslar,
113 Chapter 7
Prof W Teichmann, Klinik und Poliklinik der MLU, Halle/Saale, Dr R-D Beythien, St Sixtus-Hospital, Haltern, Dr W Sehnert, Evang Krankenhaus, Herne, Dr Ρ von Lowis of Menar, Evang Krankenhaus, Holzminden, Prof R Simon, Med Universitätsklinik, Kiel, Dr F Kersting, Kreiskrankenhaus Evang Stift St Martin, Kolblcnz, Prof К D Grosser Krankenanstalten, Krefeld, Dr L Engelmann, Zentrum fur Innere Medizin, Universität Leipzig, Prof W Jansen, Klinikum, Leverkusen, Dr J M Rustige, Klinik der Stadt, Ludwigshafen, Prof J Meyer, Klinikum der Johannes-Gutenberg-Universitat, Mainz, Dr V Kotter, Evang Krankenhaus, Mulheim, Prof M Luther, Kreis- krankenhaus Pasing, München, Prof A Schomig, Klinik Rechts der Isar, München, Prof Τ Kleine, Albert-Schweitzer-Krankenhaus, Northeim, Dr Ρ Lazarus, Klinikum, Schwerin, Prof Η R Ochs, Marienkrankenhaus, Soest, Dr J Ebel, Kreiskrankenhaus Charlottenstift, Stadtoldendorf, Dr A Schmidt, Karl-Olga-Krankenhaus, Stuttgart, Prof К Hagemann, Evang Krankenhaus Unna, Unna, Prof К Haerten, Manen-Hospital, Wesel Israel Dr Τ Rosenfeld, Haemek Hospital, Añila, Prof N Roguin, Government Hospital, Nahanya, Prof S Sclarovsky, Beihnson Medical Center, Petach-Tikvah, Dr A Caspi, Kaplan Hospital, Rehovot, Dr A Τ Marmor, Rebecca Zief Hospital, Safed Italy Prof G Specchia, MD, Ospedale San Matheo, Pavia, Prof С Cernigliaro, Ospedale Maggiore, Novara The Netherlands Prof F W A Verheugt, Dr G Veen, Vrije Universiteit Ziekenhuis, Amsterdam, Dr M Bijl, Wilhelmina Ziekenhuis, Assen, Dr J R M Peters, Maasziekenhuis, Boxmeer, Ρ H J M Dunselman, Ignatius Ziekenhuis, Breda, Dr A J Withagen, Renier de Graaf Ziekenhuis, Delft, Dr R Michels, Cathanna Ziekenhuis, Eindhoven, Dr L Rehk van Wely, Diaconessenhuis, Eindhoven, Dr W van Ekelen, St Annaziekenhuis, Geldrop, Dr L H Takens, Martini Ziekenhuis, Groningen, Dr Ρ Bendermacher, Elkerliek Ziekenhuis, Helmond, Dr J van der Pol, Bosch Medicentrum, Hertogenbosch, Dr W M Schenkel, Medisch Centrum Leeuwarden, Dr F Vermeer, Academisch Ziekenhuis, Maastricht, Dr J H Kingma, Stichting Antonius Ziekenhuis, Nieuwegein, Dr R Bergshoeff, Ziekenhuis Overvecht, Utrecht, Dr А С Bredero, Diakonessenhuis, Utrecht, Dr A H Bosma, St Joseph Ziekenhuis, Veldhoven, Dr H С Klomps, St Jans Gasthuis, Weert Ireland Dr W Fennell, Cork Regional Hospital, Cork, Prof M В Murphy, Mercy Hospital, Cork, Prof J H Horgan, Beaumont Private Clinic, Dublin, Dr В Maurer, St Vincent's Hospital, Dublin, Dr D D Sugrue, Mater Misencordiae Hospital, Dublin, Dr M J Walsh, St James's Hospital, Dublin Portugal Dr A Correla, Hospital de Sao Marcos de Braga, Braga, Prof R Seabra-Gomes, Hospital de Santa Cruz, Linda-A-Velha, Dr L Mourao, Hospital de Sao Francisco Xavier, Lisbon, Dr J Coucello, Hospital Distrital, Portimao Russian Federation Prof V Doschicin, Central Military Hospital of the Russian Ministry for Defense, Moscow, Prof A Ρ Golikov, Cardiological Clinic of Skhfosovsky Scientific Research Institute, Moscow, Prof А К Gruzdev, Central Clinic of the Medical Centre of the Government, Moscow, Prof V 1 Ivashkin,
114 Report of COMPASS investigators
Main Military Clinical Hospital, Moscow; Prof. A. J. Ivleva, Hospital Mo. 64, Moscow; Prof. G. A. Petrakov, Central Hospital of the Ministry of Internal Affairs, Moscow; Prof. M. Y. Ruda, Cardiology Research Centre, Moscow; Prof. V. I. Simonov, Central Hospital No. 7, Moscow. Spain: Prof. A. Betriu, Hospital Clinico, Barcelona; Dr. J. Figueras, Ciudad Sanitaria Vail D'Hebron, Barcelona; Dr. X. Sabate, Hospital Principes de Espana, Barcelona; Dr. R. Coma, Hospital Doce de Octubre, Madrid; Dr. F. J. Lacoma and Dr. A. Algora, Hospital Severo Ochoa de Lcganes, Madrid; Dr. J. L. Carpintero, Hospital Clinico, Malaga; Dr. M. Ruano, Hospital Universitari La Fe; Dr. J. Arzubiaga, Hospital Civil de Basurto, Bilbao; Dr. J. Froufe, Hospital Cruces de Bilbao, Vizcaya; Dra. E. Civeira, Hospital Clinico, Zaragoza. United Kingdom: Dr. R. D, Watson, Dudley Road Hospital, Birmingham; Prof. A. A. J. Adgey, Royal Victoria Hospital, Belfast; Dr. A. J. Moriarty, Craigavon Area Hospital, Craigavon; Dr. J. P. S. Varma, Erne Hospital, Enniskillen; Dr. H. M. Dunn, Altnagelvin Area Hospital, Londonderry.
115 Chapter 7
References
1. Fibrinolytic Therapy Trialists' (FTT) Collaborative Group. Indications for fibrinolytic therapy in suspected acute myocardial infarction: collaborative overview of early mortality and major morbidity results from all randomised trials of more than 1000 patients. Lancet 1994;343:311-22. 2. Flohé L. Recombinant human pro-urokinase (non-glycosylated). Drugs Future 1986;11:851-2. 3. PRIMI Trial Study Group. Randomised double blind trial of recombinant prourokinase against streptokinase in acute myocardial infarction. Lancet 1989;1:863-8. 4. International Joint Efficacy Comparison of Thrombolytics. Randomised, double-blind comparison of reteplase double-holus administration with streptokinase in acute myocardial infarction (INJECT): trial to investigate equivalence. Lancet 1995;346:329-36. 5. Good Clinical Practice for Trials on Medicinal Products in the EC. Brussels: Commission of the European Community, 1990:III/3976/88-EN. 6. Tcbbe U, Windeier J, Boesl I, et al., on behalf of the LIMITS Study Group. Thrombolysis with recombinant unglycosylated single-chain urokinase-type plasminogen activator (saruplase) in acute myocardial infarction: Influence of heparin on early patency rate (LIMITS study). J Am Coll Cardiol 1995;26:365-73. 7. Task Force on the Management of Acute Myocardial Infarction of the European Society of Cardiology. Acute myocardial infarction: pre-hospital and in-hospital management. Eur Heart J 1996;17:43-63. 8. The GUSTO Investigators. An international randomized trial comparing four thrombolytic strategies for acute myocardial infarction. N Engl J Med 1993;329:673-82. 9. Gheorghiade M, Ruzumna P, Borzak S, Havstad S, AH A, Goldstein S. Decline in the rate of hospital mortality from acute myocardial infarction: impact of changing management strategies. Am Heart J 1996; 131:250 -6. 10. ISIS-2 (Second International Study of Infarct Survival) Collaborative Group. Randomised trial of intravenous streptokinase, oral aspirin, both, or neither among 17,187 cases of suspected acute myocardial infarction: ISIS-2. Lancet 1988;2:349-60. 11. ISIS-3 (Third International Study of Infarct Survival) Collaborative Group. ISIS-3: a randomised trial of intravenous streptokinase versus tissue plasminogen activator versus anistreplase and of aspirin plus heparin versus aspirin alone among 41,299 cases of suspected acute myocardial infarction. Lancet 1992;339;753-70.
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12. Kleiman NS, White HD, Ohman EM, et al., for the GUSTO Investigators. Mortality within 24 hours of thrombolysis for myocardial infarction: the importance of early reperfusion. Circulation 1994;90:2658-65. 13. Simes RJ, Topol EJ, Holmes DR, et al., for the GUSTO-1 Investigators, link between the angiography substudy and mortality outcomes in a large randomized trial of myocardial reperfusion: importance of early and complete infarct artery reperfusion. Circulation 1995;91:1923 - S. 14. Bär FW, Meyer J, Vermeer F, et al., for the SESAM Study Group. Early patency and reocclusion: comparison of saruplase and alteplase in acute myocardial infarction: results of the randomized, multicentre SESAM study (Study in Europe with Saruplase and Alteplase in Myocardial Infarction). Am J Cardiol 1997;79:727-32. 15. Gruppo Italiano per lo Studio della Streptochinasi nell'Infarto Miocardico. GISSI-2: a factorial randomised trial of alteplase versus streptokinase and heparin versus no heparin among 12,490 patients with acute myocardial infarction. Lancet 1990;336:65-71. 16. ISAM Study Group. A prospective trial of intravenous streptokinase in acute myocardial infarction (ISAM). N Engl J Med 1986;314:1465 - 71. 17. Koster RW, Cohen AF, Hopkins GR, et al. Pharmacokinetics and pharma codynamics of saruplase, an unglycosylated single-chain urokinase-type plasminogen activator, in patients with acute myocardial infarction. Thromb Haemost 1994;71:740-4.
117 118 CHAPTER 8
Pharmacokinetics and hemostatic effects of saruplase in patients with acute myocardial infarction. Comparison of infusion, single bolus, and split-bolus administration.
H.R. Michels, MD', J.J.M.L. Hof/mann, Pft.D2, F.W.H.M. Bär, MD 3 'Department of Cardiology, Catharina Hospital, Eindhoven, the Netherlands.2 Department of Clinical Laboratories, Haemostasis Division, Catharina Hospital, Eindhoven, the Netherlands. 3Department of Cardiology, Academic Hospital Maastricht, the Netherlands.
Abstract. Background. Saruplase, or unglycosylated, single-chain urokinase-type plasminogen activator (scu-PA) selectively activates fibrin-bound plasminogen, and is subsequently converted to its two-chain derivative tcu-PA (urokinase) by plasmin. The efficacy of a 20 mg i.v. bolus followed by an infusion of 60 mg over lh (standard regimen) has been demonstrated in acute myocardial infarction (AMI). The Bolus Administration of Saruplase in Europe (BASE) study compared the efficacy of standard therapy, single bolus (80 mg) and split-bolus (2 χ 40 mg at 30 min interval) in AMI. Aim. In a sub-study of BASE the pharmacokinetics of total u-ΡΑ activity (amidolytic activity after plasmin treatment), high molecular weight (HMW) u- PA antigen and tcu-PA activity were compared in patients receiving standard therapy (n = 4), single bolus (n = 4) or split-bolus (n =5). Results. Total u-ΡΑ activity and HMW u-ΡΑ antigen were similar. The maximum concentration (СтШъ, mean ± S.D) of total u-ΡΑ activity was 2.2 ± 0.3 ßg/ml after standard therapy, 16.3 ± 3.9 ßg/ml after single bolus and 8.2 ± 1.6 ug/ml after split-bolus. The area under the concentration vs. time curve (A UC) values of total u-ΡΑ activity were 1.7 ±0.1 μg/ml*h (standard therapy), 4.0 ± 0.9 μg/ml*h (bolus) and 3.0 ± 0.7 ßg/ml*h (split-bolus). The dominant initial half-lives (t 'Δ a) were 7.1 ±1.1 min (standard), 8.8 ±0.8 min (bolus), and 5.1 ±2.1 min (split-bolus). Maximum plasma concentrations of of tcu-PA activity were observed at 5.2
±7 min (standard), 21 ± 10 min (bolus), and 42 ± 2 min (split-bolus). Cmax was lowest after standard therapy (0.6 ± 0.3 μg/ml), highest after bolus (4.2 ±2.2
119 Chapter 8
μζ/τηΐ) and approximately twice as high as standard therapy after split-bolus (1.3±0.8μ&πιΙ). After standard therapy the mean fibrinogen concentration decreased gradually from approximately 300 mg/dl to 70 mg/dl at 90 and 120 mm. After a single bolus fibrinogen concentration decreased below the limit of quantification within 30 min and remained so for at least 120 min. Directly after the second 40 mg of the split-bolus the fibrinogen levels demonstrated an accelerated and more pronounced decrease to approximately 65 mg/dl at 90 and 120 min. Conclusion. A single bolus results in very high early total u-ΡΑ activity which accelerates the appearance of tcu-PA activity andfibrinogen consumption. The pharmacokinetics and hemostatic effects of the split-bolus regimen are more comparable to those of standard therapy.
Keywords: saruplase, pharmacokinetics, bolus administration.
Abbreviated abstract.
Saruplase (scu-PA) is an effective thrombolytic agent when given as a bolus of 20 mg followed by an infusion of 60 mg/lh (standard regimen). We compared the pharmacokinetics of standard regimen (n = 4) to those of a single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, at 30 min interval, η = 5). A single bolus was found to result in very high early total u-ΡΑ activity and u-ΡΑ antigen concentrations which accelerate and enhance the generation of tcu-PA activity, leading to accelerated and enhanced systemic plasminogen activation. The effects of the split-bolus regimen on hemostasis were more similar to those of standard therapy.
Introduction.
Saruplase is the full-length unglycosylated single-chain urokinase-type plasminogen activator (scu-PA) obtained from genetically engineered Escherichia coli1. Scu-PA has intrinsic plasminogen activation potential which in vivo is focused on the plasminogen bound to the fibrin clot2,3 The plasmin formed locally splits scu-PA into two-chain urokinase-type plasminogen activator (tcu-PA), or urokinase.
120 Pharmacokinetics of saruplase
Experiments indicated that saruplase could be given as an intravenous bolus, followed by a 1 hour infusion4. Two dose-finding trials indicated that a 20 mg bolus followed by a 60 mg infusion is an effective dose regimen5'6. The efficacy of this regimen has been confirmed in studies that compared saruplase to streptokinase7,8, to urokinase9, and to recombinant tissue-type plasminogen activator (rt-PA)10'". A pharmacokinetic study with this regimen estimated an overall conversion of approximately 28% of saruplase into its active tcu-PA degradation product urokinase12. An i.v. bolus of urokinase of 2 million IU has been demonstrated to be safe and effective in achieving patency and in improving cardiac performance in AMI1314. Thrombolytic therapy has its maximal myocardial salvage and survival advantage when administered early after onset of symptoms. After one hour the efficacy profile declines rapidly and steeply and remains rather flat from 2 to 6 hours and beyond15"18. Bolus administration of a thrombolytic agent is simple and easy, and facilitates early prehospital, home or ambulance administration. Bolus administration of saruplase may combine the advantage of a high initial concentration of clot-specific scu-PA and very early conversion of scu-PA into its systemically active derivative tcu-PA. The multicenter double-blind, randomized Bolus Administration of Saruplase in Europe (BASE) pilot study in 8 European hospitals, compared the efficacy and safety of the i.v. administration of saruplase as a single bolus ( 80 mg and 60 mg), a split-bolus (2 χ 40 mg administered with 30 min interval), and the standard regimen (20 mg bolus followed by an infusion of 60 mg over lh). Comparable efficacy and safety profiles were observed in patients treated with the 80 mg single bolus and the standard regimen. The 60 mg single bolus was safe but did not meet with predefined minimal patency rates indicating insufficient effectiveness and was therefore abandoned prematurely. The highest patency rate and the highest complication rate were found with the split-bolus regimen (F. Bär accepted for publication, J Thrombosis and Thrombolysis). The pharmacokinetic profile of the standard regimen of saruplase has been studied previously12,19,20. Since the elimination capacities for saruplase protein could be limited, the pharmacokinetics of the bolus regimens could not easily be extrapolated from the available data. This applied especially to the appearance of the saruplase conversion product tcu-PA. The profile of hemostatic alterations by the standard regimen of saruplase lies
121 Chapter 8 between that of streptokinase and rt-PA710, and is quite similar to that of urokinase9'20,21. In this single centre sub-study of BASE, the pharmacokinetic behaviour of saruplase was assessed by measuring total u-ΡΑ activity (amidolytic activity after plasmin treatment) and high molecular weight (HMW) u-ΡΑ antigen. Plasma activity of tcu-PA was determined in order to obtain an estimate of saruplase conversion. Plasma concentrations of fibrinogen, plasminogen, and a2-antiplasmin were measured as indicators of systemic plasminogen activation.
Methods.
Patients. Included in the pharmacokinetic sub-study were patients who were included in the BASE study, and who had also consented to participate in the pharmacokinetic sub-study. Both protocols were approved by the Hospital Ethics Committee and by the Central Ethics and Advisory Board. The clinical part of the pharmacokinetic sub- study was performed in the Catharina hospital, Eindhoven, NL. The samples were analysed at the department of Human Pharmacology / Pharmacokinetics, Grünenthal GmbH, Aachen , Germany.
Investigatory study medication. Saruplase and placebo to saruplase were prepared by Grünenthal GmbH and were provided in vials containing 20 mg saruplase or placebo to saruplase, and 60 mg saruplase or placebo to saruplase. Patients were given either standard therapy (20 mg bolus + 60 mg over Ih), a single bolus (80 mg), or a split-bolus (2 χ 40 mg, at 30 min interval) using a randomized double- blind technique with 3 parallel treatment arms, where each regimen included placebo medications for the other arms (double dummy technique).
Blood sampling. Blood was sampled through an indwelling 19 gauge needle exclusively for this purpose in the arm opposite to that used for study medication administration. Blood samples for the analysis of the pharmacokinetic parameters were taken before study drug administration, and at 3, 5, 10, 15, 28, 33, 35, 40, 45, 58, 90, and 120 min after start of study drug administration. BASE being a double-
122 Pharmacokinetics ofsaruplase blind study, one sampling scheme had to fit all administration schemes. The pharmacokinetics of standard therapy have already been investigated13, so the sampling scheme was optimised to fit the single and split-bolus regimens. The resulting scheme had only two sampling times at the end of the infusion, and therefore the terminal phase after standard therapy could not be determined. Consequently the AUC values after standard therapy are underestimated. The samples were stabilised with citrate, aprotinin and benzamidine (CAB)23. The samples were shipped on dry ice and stored at -60°C to -80°C. Blood samples for the analysis of the hemostatic parameters were taken before study drug administration, and at 15, 28, 45, 58, 90 and 120 min after start of study drug administration. Pharmacokinetics: the saruplase-related analytes measured were high molecular weight urokinase-type plasminogen activator antigen (HMW u-ΡΑ antigen), total urokinase-type plasminogen activator activity (total u-ΡΑ activity) and the activity of two-chain u-ΡΑ (tcu-PA activity). HMW u-ΡΑ antigen comprises the entire molecule, containing the 'kringle' domain as well as the active site domain; hence HMW u-ΡΑ antigen includes the scu-PA as well as the tcu-PA forms of the molecule. HMW u-ΡΑ antigen was determined using a sandwich enzyme-linked immunosorbent assay (ELISA) using anti-LMW u-PA antibodies as capturing antibodies and anti-kringle antibodies as detection antibodies. The antibodies were obtained from rabbits. The second (detection) antibody was biotinylated. For signal generation streptavidin-peroxidase was used. Peroxidase activity was determined by ABTS/H2O2. The activity of tcu-PA was determined using an ¡mmuno activity assay23, without revious activation of the immuno- bound sample by plasmin. Total u-ΡΑ activity was measured with this same assay, but after activation of the immuno-bound sample with plasmin, so that the enzymatically inactive scu-PA is converted to its active tcu-PA form. The results of the determination of total u-PA activity and HMW u-PA antigen should be identical, unless the specific activity of u-PA is changed in vivo. The pharmacokinetic parameters were calculated with a two-compartmental model, using the TOPFIT program23, except after standard administration, where a one- compartmental model was used, because the terminal phase was beyond final sampling (120 min). Tcu-PA activity concentrations were evaluated by a model independent evaluation method developed by the departments of Biometry and Pharmacokinetics of Grünenthal GmbH.
123 Chapter 8
Hemostatic parameters. The hemostatic parameters studied were fibrinogen, plasminogen and ct2 -antiplasminogen. For the determination of fibrinogen the multifibrintest (Behringwerke AG, Germany) was used. Plasminogen was determined using the АСА Du Pont discrete clinical analyser, method 52 (plasminogen) (Du Pont de Nemours, Germany). For the determination of a2-antiplasmin the R Berichrom a2-antiplasmin diagnostic test was used (Behringwerke AG, Germany).
Statistical evaluation and presentation of results. Concentration values were checked for symmetrical distribution by comparing the arithmetic mean with the corresponding median values. Because of the small numbers of patients and incompatibility of administrations and sampling schemes, no biometrical calculations e.g. to evaluate differences in the administration schemes, were performed. All results are presented as means ± standard deviation (S.D.), with the exception of the presentation of the mean plasma concentrations of tcu-PA activity, where the standard error of the mean (S.E.M) was used as a measure of variation.
Abbreviations. A summary of abbreviations and of acronyms is given.
scu-PA single chain urokinase plasminogen activator (used here synonymously with saruplase) tcu-PA two-chain urokinase-type plasminogen activator, urokinase derived from scu-PA (saruplase) by breaking the peptide bond behind Lys 1S8 u-ΡΑ urokinase-type plasminogen activator, encompassing both tcu-PA and scu-PA IAA immunoactivity assay used for measuring the activities of tcu-PA and scu-PA in plasm ELISA enzyme-linked immunosorbent assay for measuring antigen concentrations cmax maximum concentration tmax time at which cmax is reached AUC area under the concentration vs time curve (from t=0) CI tot total plasma clearance Vc apparent volume of distribution of the central compartment Vss apparent volume of distribution at steady state VD, 0 apparent volume of distribution in the terminal disposition phase t'/2,kel overall elimination half-life t'/2,a half-life of the initial phase t'A, В half-life of the terminal disposition phase MRT mean residence time AUC a portion of the initial phase related to the total AUC AUC ß portion of the terminal disposition phase related to the total AUC
124 Pharmacokinetics of saruplase
Results.
Between December 1993 and December 1994, 192 patients in 8 European centres were included in BASE. Forty-three (22%) of these patients were included at the Catharina Hospital, Eindhoven, NL. Between February 1994 and December 1994 thirteen of these 43 patients were included in the pharmacokinetic sub-study. The demographic data of these patients are shown in table 1.
Table 1: Demographic data Standard = 20 mg bolus + 60 mg/1h, single bolus = 80 mg, split bolus = 2 χ 40 mg, 30 min interval
Sex Age Body weight Height Regimen (years) (kg) (cm) m 64 97 169 Standard m 53 86 178 Standard m 36 82 180 Standard m 67 113 167 Standard m 53 75 ne bolus m 70 74 167 bolus m 70 72 180 bolus f 68 65 170 bolus m 37 91 173 split bolus m 58 68 ne split bolus m 49 90 ne split bolus m 45 91 ne split bolus m 49 90 187 split bolus η e = not estimated
Pharmacokinetic parameters. The evaluation after standard therapy was performed using a one-compartment model, although previous studies have indicated that the decay of plasma concentrations is best described by a two-compartment model12'20. The pharmacokinetic parameters of total u-ΡΑ activity after standard therapy, single bolus and split-bolus are shown in table 2. In comparison to an earlier study with the last sampling at 240 min12, we found a lower AUC value and a higher total clearance after standard therapy (768 ± 37, vs 427 ± 113 ml/min). The AUC values obtained for protein equivalents of total u-ΡΑ activity were 1.7 ± 0.1 u/ml*h (standard therapy), 4.0 ± 0.9 u7ml*h (bolus) and 3.0 ± 0.7 uAnl*h
125 Chapter 8
(split-bolus). The dominant initial half-lives (t г α) were 7.1 ± 1.1 min (standard), 8.8 ± 0.8 min (bolus), and 5.1 ±2.1 min (split-bolus).
Table 2: Pharmacokinetic parameters of total u-ΡΑ activity, expressed as single bolus (80 mg), and split-bolus (2 χ 40 mg, 30 min interval), п.е. = not estimated.
Standard Bolus Split bolus Parameter 20 + 60 mg 80 mg 2 χ 40 mg
Cmax [Mg/ml] 2 2 ± 0 35 1631 ±388 8 18 ±1 55 AUC [\ig/mi h] 1 74 ± 0 09 4 02 * 0 93 2 96 ± 0 73 Cltot [ml/mm] 768 ± 37 346 * 84 475± 124 Vc[L] 7 91 ±1 38 4 59 ± 1 29 4 13 ±0 76 Vss (L] η e 5 28 ± 1 30 6 61 ±2 29 Vd, ß [L] ne 25 3 ± 9 3 152±58 tW, α [min] 7 1 ±11 88±08 5 1±2 1 t'/¡, D [min] 53 4 ±27 6 204i66 MRT [mm] 13 8 ± 2 9 17 1±14 148±27 AUC, α[%] ne 94 5 ± 4 2 68 8 ± 13 6 AUC, ß [%] ne 55±42 312*136
The pharmacokinetic parameters of tcu-PA activity after the three administration regimens are shown in table 3. Maximum plasma concentrations of tcu-PA activity were observed at 52 ± 7 min (standard), 21 ± 10 min (single bolus), and 42+2 min (split-bolus).
Table 3: Pharmacokinetic parameters of tcu-PA activity after standard (20 mg bolus + 60 mg/1h, η = 4), single bolus (80 mg, η = 4), and split- bolus (2 χ 40 mg, 30 interval, η = 5) of saruplase (mean ± SD).
Standard Bolus Split bolus Parameter 20 + 60 mg 80 mg 2 χ 40 mg
Cmax [ng/ml] 0 62 ± 0 32 4 ! 5 ± 2 24 1 31 ±084 tmax [min] 52 ±7 21 ± 10 42 ±2 AUC [>g/ml»h] 0 S3 ± 0 24 1 89 ± 0 80 0 76 ± 0 31 t'A [mm] 12 8± 1 4 21 1 ±97 26 4 ±21 0 MRT [min] 22 6 ± 1 1 26 1 ± 4 4 38 ±60
Maximum tcu-PA concentrations were highest after bolus (4.2 ± 2.2 ц^ті) and lowest after standard therapy (0.62 ± 0.3 μg/ml). Τ 'Λ α of
126 Pharmacokinetics of saruplase tcu-PA activity was 12.8 ± 1.4 min for standard therapy, 21.1 ± 9.7 min for single bolus, and 26.4 ±21.0 min for split-bolus administration. The ratio of the mean AUC values of tcu-PA activity and total u-PA activity was highest for single bolus (0.47) and comparable for standard therapy (0.30) and split-bolus (0.26). The pharmacokinetic parameters of HMW-u-PA antigen after the three administration schemes are shown in table 4.
Table 4: Pharmacokinetic parameters of HMW u-ΡΑ antigen after standard (20 mg + 60mg/1h, η = 4), single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, 30 min interval, η = 5) of saruplase (mean ± SD). Standard Bolus Split bolus Parameter 20 + 60 mg 80 mg 2 χ 40 mg
Cmax [μ§/ιη1] 1 81 ±0 20 13 65 ±3 02 6 96 ±1 88 AUC ^g/ml h] 1 82 ± 0 22 3 74 ±0 81 2 52 ±0 83 Cltot [ml/min] 739 ±88 370 ±81 582 ±207 Ve [LJ 100± 1 2 5 5 ± 1 4 52± 1 7 Vss [L] - 68±58 58± 12 Vd, ß [L] - 27 3 ± 12 7 16 7 ± 8 5 t'/2, α [min] 95±14 97±08 55±20 t'A, ß [mm] - 50 4 ±18 6 210±48 MRT [mm] 155±23 20 1 ± 1 8 12 8 ±4 8 AUC, α [%] - 92 4 ± 1 9 81 0±54 AUC, ß [%] - 7 6± 1 9 190±54
The results are similar to those of total u-ΡΑ activity. Figure 1 shows the rate and extent of conversion of scu-PA into tcu-PA (values of the plasma concentrations of tcu-PA activity expressed as protein equivalents ± the standard error of the mean, S.E.M.) No tcu-PA activity was detectable 3 min after the 20 mg bolus of standard therapy. Three min after the 80 mg single bolus the plasmaconcentration of tcu- PA activity had already reached 0.638 ± 0.379 μg/ml. No tcu-PA activity was detectable 3 min after the first 40 mg of the split-bolus. Plasma concentration of tcu-PA activity reached a maximum of 0.608 ± 0.169 μg/ml after 58 min with standard therapy, a maximum of 4.085 ± 1.117 μg/ml after 15 min with bolus administration and an initial peak of 0.364 ± 0.075 μ&Ίτι1 after 15 min, followed by a maximum of 0.631 ± 0.288 μg/ml after 35 min with split-bolus administration (table 5).
127 Chapter 8
Table 5: Plasma concentrations of tcu-PA activity (цд/ті) expressed as protein equivalents after standard (20 mg + 60 mg/1h, η = 4), single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, 30 min interval, η = 5) of saruplase (mean ± SEM) η.e. = not estimated.
Time Statutari Bolus Split Bolus (min) 20 mg+ 60 mg 80 mg 40 mg + 40 mg mean SEM mean SEM mean SEM 3 ne ne 0 638 0 379 η e ne 5 0018 0 004 1 128 0 494 0 108 0 054 10 0 125 0 026 2 955 1040 0 265 0 114 15 0 221 0 043 4 085 1 117 0 364 0 075 28 0 327 0 058 2 156 0 222 0216 0 031 33 0 322 0 050 1593 0 168 0 266 0 057 35 0 334 0 050 1334 0 101 0631 0 288 40 0 356 0 038 1060 0 119 1211 0 344 45 0 483 0 126 0 692 0071 1283 0 254 58 0 608 0 169 0 272 0 039 0 790 0 240 90 0 123 0 022 0 040 0 006 0 048 0014 120 0019 0 004 0016 0 002 0016 -
In summary, bolus administration caused the highest maximum concentration of tcu-PA activity in the shortest time, indicating an early and high plasma conversion rate from saruplase to tcu-PA.
• standard • bolus * split bolus
Figure 1: Plasma concentrations of tcu-PA activity (цд/ті) after standard (20 mg + 60 mg/1h, η = 4), single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, 30 min interval, η = 5) of saruplase.
128 Pharmacokinetics ofsaruplase
Hemostatic parameters. Fibrinogen (figure 2): after standard therapy the fibrinogen concentration decreased from approximately 300 mg/dl to 70 mg/dl at 90 and 120 min after start of treatment. The drop in fibrinogen concentration started slowly after the first 28 min and accelerated after 45 min, to reach its nadir at 90 min.
Fig- I. Fibrinogen levels (mg/dl) mg/dl
standard bolus split bolus 300 - ^^ • - - *• ·
250 - * · .
200 - ••-·*.
150 - • '* . 100 -
ш 50 - •
u I I I I I I 1 0 30 60 120 (mm) Figure 2: Plasma concentrations of fibrinogen (mg/dl) after standard (20 mg + 60 mg/1h, η = 4), single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, 30 min interval, η = 5) of saruplase
After single bolus administration fibrinogen concentration decreased very fast. After 15 min the fibrinogen values had already decreased below the limit of quantification in 2 out of 4 patients. After the first 40 mg of the split-bolus the fibrinogen levels were not essentially affected, but after the second 40 mg bolus a more rapid and pronounced decrease was seen. Alpha2-antiplasmin (figure 3): after standard therapy the pre dose concentration of a2-antiplasmin decreased from 83 ± 3% of the concentration of the control sample to 9 ± 3% at 90 min. Decrease started slowly between 15 and 28 min and accelerated then. Essentially the same pattern was seen after split-bolus. After single bolus administration the baseline concentration of 73 ± 12% decreased rapidly to 6 ± 4% after 15 min and remained low until the last sample at 120 min.
129 Chapter 8
Fig. 3. Alpha 2-antiplasmin (% controls) % control 90
standard
120 (mm)
Figure 3: Plasma concentrations of a2-antiplasmin (% control) after standard (20 mg + 60 mg/1h, η = 4), single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, 30 min interval, η = 5) of saruplase (mean ± SD).
Plasminogen (figure 4): after standard therapy the plasminogen concentration decreased from 93 ± 13% of control to approximately 23% of control at 90 to 120 min. Fig. 4. Plasminogen (% controls) % control
и и —•— standard 90 Л \v* - -•- - bolus 80 "• split bolus \ 70 \ 60 \ \\ 50 \ ^ \ 40 V * ^^~""*~«^j» * * 30
20
ι 1 ι I I ι 30 60 90 120 (mm) Figure 4: Plasmaconcentrations of plasminogen (% control) after standard (20 mg = 60 mg/1h, η = 4), single bolus (80 mg, η = 4), and split-bolus (2 χ 40 mg, 30 mm interval, η = 5)
130 Pharmacokinetics of samplase
A similar pattern was seen for split-bolus. After single bolus the plasminogen concentration decreased rapidly from 85 ± 6% of control to 25 ± 3% of control at 28 min and remained low to the end of the investigation at 120 min.
Discussion.
The pharmacokinetic sub-study had to be conducted under the limitations of the double-blind main study BASE. The sample scheme had to be identical irrespective of the administration scheme. The pharmacokinetics and pharmacodynamics of the standard regimen have already been described in detail12 and the sampling scheme in this study was optimised to fit the single and split-bolus regimens. In this study the number of samples was reduced to 13 ending at 120 min, instead of 16 ending at 240 min, as in earlier studies12'20. This scheme fitted the single bolus and split-bolus administration, but it was impossible to demonstrate the terminal phase after standard administration. Consequently a single compartment model had to be used for the pharmacokinetic evaluation after the standard regimen, although it is known that the decay of plasma concentrations of samplase fits a two-compartment model12. Therefore the AUC values after the standard regimen are underestimated. This becomes apparent from table 2, where the AUC after the standard regimen is half that expected from the UC's of the two other regimens. Consequently the total plasma clearance (Cltot), which equals Dose/AUC, is overestimated by at least a factor 2 (measured 768 ml/min, expected approximately 400 ml/min). The total clearance values obtained after single bolus and split-bolus administration are well in the range observed in earlier studies'2,20 after administration of 20 mg bolus followed by a 1 h infusion of 60 mg. The rate and amount of tcu-PA generated after administration of the single bolus is very high in comparison to standard therapy and to split-bolus administration (figure 1). High maximum concentrations are achieved almost instantly. The rate and amount of tcu-PA generated after administration of the split-bolus displays similarity to that obtained after standard administration until administration of the second 40 mg bolus, when a more rapid and pronounced increase of tcu-PA activity is seen,
131 Chapter 8 however to a lesser extent than after single bolus administration. The ratio of the mean AUC values of tcu-PA activity and total u-ΡΑ activity was similar for the standard regimen and for split-bolus administration but much higher after single bolus administration, indicating that more saruplase (47%) is converted into tcu-PA after single bolus administration. The decrease of hemostatic parameters was fastest and most pronounced after administration of the single bolus. The most moderate decrease was achieved with the standard regimen. After the first 40 mg of the split-bolus-regimen the fibrinogen values are not essentially affected, but directly after the second 40 mg a more rapid and pronounced decrease of the fibrinogen concentrations is seen. The decrease of the hemostatic parameters correlates with the increase of the tcu-PA activity in plasma. The single bolus regimen caused very high plasma concentrations of u-ΡΑ antigen and activity. Since concentration-dependent diffusion and conversion processes can be expected to proceed fastest in this group because of the high plasma concentrations, an accelerated plasmin generation and enhanced conversion of saruplase to tcu-PA appear plausible consequences. Obviously, the systemic plasmin generated by the first bolus of the split-bolus regimen could still be controlled by plasma inhibitors, but these appear to have been exhausted when the second bolus was given resulting in the typical alterations of plasminemia. With the single bolus regimen these changes were observed to be faster and more pronounced. Within 15 min a2-antiplasmin reached its lowest level (figure 3). It is difficult to understand why the decrease of plasminogen (figure 4) remains stable at about 23% of control throughout the period of investigation. Essentially all fibrinogen is consumed after 45 min and this should mean that (nearly) all plasminogen should be converted to plasmin by that time. This is also compatible with the results of α2- antiplasmin (figure 3). Perhaps the assay is not entirely specific and other substances than plasminogen interfered. Relationship of pharmacokinetics and hemostatic effects to clinical safety and efficacy. The relationship between the pharmacokinetic and hemostatic data and the clinical results from BASE is not entirely clear. The clinical results of the BASE pilot trial indicate that the efficacy and safety of the standard regimen and the single bolus are quite similar, while the split-bolus regimen has a positive influence on efficacy and a negative one on safety. Intuitively one would be inclined to expect this latter negative effect on safety from the single bolus
132 Pharmacokinetics of saruplase regimen, but apparently more hemostatic disturbance does not cause bleeding, while on the other hand the generation of less tcu-PA activity with less hemostatic changes is associated with an increased risk of bleeding when a split-bolus is administered. The mechanism is not readily explainable. From the experience with rt-PA and the mutants of wild type t-PA it has become clear that clot-specificity is associated with less hemostatic disturbance but with an increased risk of severe bleeding complications (i.e. intracranial hemorrhage), in comparison with streptokinase. Due to the very early and very high conversion of scu-PA into tcu-PA with single bolus administration, the single bolus may behave more like a systemic (non-specific) lytic agent. On the other hand one may speculate whether the second bolus of the split- bolus regimen intensifies the clot-specific properties. In any case, high systemic tcu-PA activity and profound hemostatic disturbance did not negatively influence the safety and efficacy of saruplase in this study. One might even speculate that the early high plasma concentration of thrombolytic activity might have contributed to the efficacy of the regimen due to an accelerated diffusion into the occluded vessel. As judged from clinical outcome the single bolus regimen was as safe and effective as the standard regimen, and was not associated with a clinically relevant plasminogen steal phenomenon which is believed to abrogate thrombolytic efficacy due to plasminogen depletion. This effect could have been less pronounced with the split-bolus regimen. At least plasma plasminogen was not exhausted by the first of the two boluses. However, the efficacy and safety of the single bolus regimen suggest that the rapid processes in the initial treatment phase of the single bolus regimen of saruplase are sufficient to achieve, and to maintain acceptable reperfusion, without causing more bleeding omplications. Therefore single bolus administration has been selected for further clinical evaluation.
Conclusion.
The BASE pilot study indicates, that the safety and efficacy of the standard regimen and the single bolus regimen are comparable. The pharmacokinetic sub-study of BASE demonstrates that the pharmacokinetics and hemostatic changes after a single bolus differ
133 Chapter 8 from those of the split-bolus regimen and the standard regimen. After the single bolus very high maximum concentrations of tcu-PA activity are seen very early indicating rapid and extensive conversion of saruplase into tcu-PA. The pharmacokinetics and hemostatic effects of the split-bolus regimen are more comparable to those of standard therapy.
Acknowledgements^ We thank Ingrid v.d. Kerkhof, Monique v.d. Broek and Guy v. Dael for their secretarial and graphical contribution to this manuscript. The study was sponsored by Grünenthal GmbH, the manufacturer of saruplase. We thank Gwyn Hopkins and Wolfgang Giinzler for their scientific contributions.
134 Pharmacokinetics of saruplase
References.
1. Holmes WE, Pennica D, Blaber M et al.. Cloning and expression of the gene for pro-urokinasein Escherichia coli. Biotechnology 1985; 3: 923- 929. 2. Hanbücken FW, Schneider J, Günzler WA et al. Selective fibrinolytic activity of recombinant human pro-urokinase (single-chain urokinase-type plasminogen activator) from bacteria. Drug Res 1987; 37: 993-997. 3. Gurewich V, Pannell R, Louie S et al. Effective and fibrin-specific clot lysis by a zymogen precursor form of urokinase (pro-urokinase). A study in vitro and in two animal species. J Clin Invest 1984; 73: 1731-1739. 4. Flameng W, Vanhaecke J, Stump D et al. Coronary thrombolysis by intravenous infusion of recombinant single-chain urokinase-type plasminogen activator or recombinant urokinase in baboons. Effect on regional blood flow, infarct size and hemostasis. J Am Coll Cardiol 1986; 8: 118-124. 5. Van de Werf F, Vanhaecke J, De Geest H et al. Coronary thrombolysis with recombinant single-chain urokinase-type plasminogen activator in patients with acute myocardial infarction. Circulation 1986; 74: 1066- 1070. 6. Diefenbach C, Erbel R, Pop Τ et al. Recombinant single-chain urokinase- type plasminogen activator during acute myocardial infarction. Am J Cardiol 1988;61:966-970. 7. PRIMI Trial study group. Randomized double-blind trial of recombinant pro-urokinase in acute myocardial infarction. Lancet 1989; 1: 863-868. 8. Tebbe U, Michels R, Adgey J et al. Randomized double-blind study comparing saruplase with streptokinase therapy in acute myocardial infarction: the COMPASS equivalence trial. J Am Coll Cardiol 1998; 31: 487-493. 9. Michels R, Hoffmann H, Windeler J et al. A double-blind multicentre comparison of saruplase and urokinase in the treatment of acute myocardial infarction. Report of the SUTAMI study group. J Thromb Thromboll995;2: 17-124. 10. The Belgian saruplase alteplase study group. Effects of alteplase and saruplase on hemostatic variables: a single-blind randomized trial in patients with acute myocardial infarction. Coron Artery Dis 1991; 2: 349- 355. 11. Bär F, Meyer J, Vermeer F et al. Early patency and reocclusion: comparison of saruplase and alteplase in acute myocardial infarction. Am J Cardiol 1997; 79: 727-732.
135 Chapter 8
urokinase-type plasminogen activator, in patients with acute myocardial infarction. Thromb Haemost 1994; 71: 740-744. 13. Mathey DG, Schofer J, Sheehan FH et al. Intravenous urokinase in acute myocardial infarction.Am J Cardiol 1985; 55: 878-882. 14. Penco M, Fedele F, Agati L et al. Effects of systemic treatment with urokinase (UK) on left ventricular function. Eur Heart J 1987; 8 (Suppl 2): 24. 15. Reimer KA, Lowe JE, Rasmussen MM, Jennings RB. The wave front phenomenon of ischemic cell death I. Myocardial infarct size vs duration of coronary occlusion in dogs. Circulation 1977; 56: 7786-794. 16. Rentrop P, Smith H, Painter L, Holt J. Changes in left ventricular ejection fraction after intracoronary thrombolytic therapy: Results of the Registry of the European Society of Cardiology. Circulation 1983; 68 (Suppl 1): 55-60. 17. Gruppo Italiano per lo Studio della Streptochinase nell'Infarto Miocardico (GISSI). Effectiveness of intravenous thrombolytic treatment in acute myocardial infarction. Lancet 1986; 1: 397-401. 18. ISIS-2 (Second International Study of Infarct Survival) Collaborative Group. Randomised trial of intravenous streptokinase, oral aspirin, both or neither among 17,187 cases of suspected myocardial infarction (ISIS- 2). Lancet 1988; 2: 349-360. 19. Griensven van JMT, Koster RW, Hopkins GR, et al. Effect of changes in liver blood flow on the pharmacokinetics of saruplase in patients with acute myocardial infarction. Thromb Haemost 1997; 78: 1015-1020. 20. Poeppelmeier J, Beier H, Carlsson J et al. Comparison of the pharmacokinetics and effects on the hemostatic system of saruplase and urokinase in patients with acute myocardial infarction. J Thromb Thrombol 1996;3:385-390. 21. Michels HR, Hoffmann JJML, Windeier J, Hopkins GR. Hemostatic changes after thrombolytic therapy with saruplase (unglycosylated single- chain urokinase-type plasminogen activator and urokinase (two-chain urokinase-type plasminogen activator). Blood Coagul Fibrinolysis 1996; 7:766-771. 22. Günzler WA, Beier H, Flohé L. Activity and antigen of saruplase and two- chain urokinase related plasminogen activator are stabilized by a combination of aprotonin and benzamidine in citrated plasma. Fibrinolysis 1990; 4 (Suppl 2): 145-147. 23. Heinzel G, Woloszak R, Thomann P. TOPFIT 2.0, Pharmacokinetic and Pharmacodynamic data analysis for the PC, 1993; Gustav Fisher , Stuttgart, Jena, New York.
736 CHAPTER 9
Fibrinolysis with recombinant single-chain urokinase- type plasminogen activator (saruplase) in acute myocardial infarction. Summary of the results of clinical trials.
H.R. Michels ', MD., F. W.H.M. Bär 2, Professor of Cardiology, F. W.A. Verheugt3, Professor of Cardiology
' Catharina Hospital Eindhoven, 2 Academie Hospital Maastricht, Academie Hospital Nijmegen
Summary.
Background. The ideal fibrinolytic agent should produce rapid, maximal and sustained coronary reperfusion avoid re-occlusion, bleeding complications and allergic reactions The administration should be easy, preferably as a single bolus Aim It is our aim to examine the results of the clinical trials with saruplase in the light of these criteria Saruplase is the full-length unglycosylated human recombinant single-chain urohnase-type plasminogen activator (scu-PA) obtained from genetically engineered Escherichia coli Results. Patency data from trials comparing saruplase with streptokinase or recombinant tissue-type plasminogen activator (rt-PA) show saruplase to be very fast-acting Although the late patency rates do not differ from those obtained with urokinase, the mortality data suggest an earlier onset of action than achieved with urokinase The 30-day mortality with saruplase and its net clinical benefit (mortality plus disabling stroke) are at least equivalent to that of streptokinase The haemorrhagic stroke rate associated with saruplase is slightly but not significantly higher than that seen with streptokinase, and comparable to that seen with rt-PA In comparison to streptokinase saruplase showed less bleeding complications, the overall rates of bleeding complications for saruplase and urokinase were identical, whereas the rates of bleeding complications of saruplase and rt-PA were quite comparable Saruplase has the advantage over streptokinase of being a physiological fibrinolytic agent and to date no anti-saruplase antibodies have been detected, whereas an immunological reaction to streptokinase is well established
137 Chapter 9
Being its parent molecule, saruplase and urokinase have similar qualities, but saruplase has intrinsic clot-specific action, and may act faster The efficacy and safety of saruplase when given as a single bolus or when given as a bolus (25%) followed by a one -hour infusion are quite comparable Conclusion. Saruplase is fast-acting, safe and effective, and without allergic response Saruplase compares favourably with streptokinase, urokinase and rt-PA Saruplase can be administered as a single bolus.
Key words: saruplase, fibrinolysis, acute myocardial infarction.
Introduction.
Rapid and sustained coronary artery reperfusion and the prevention of (sub)acute reocclusion, bleeding complications, and allergic reactions are the main objectives of coronary fibrinolysis in acute myocardial infarction (AMI). In addition the ideal fibrinolytic agent should also be easy to prepare and administer, and be cost-effective. Saruplase, the full-length unglycosylated human recombinant single-chain urokinase-type plasminogen activator (scu-PA), obtained from genetically engineered Escherichia coli \ is examined in the light of these criteria 2. Scu-PA is activated by plasmin to obtain full proteolytic activity3. It was shown to be fibrin-specific, i.e. to be able to lyse clots without causing much systemic fibrinogenolysis4. In contrast to t-PA, its fibrin-specificity is not the result of direct fibrin binding 4,5,6. Scu-PA has intrinsic plasminogen activation potential, which in vivo is focused on the plasminogen bound to the fibrin clot 6I°. The plasmin formed locally splits scu-PA into two-chain urokinase-type plasminogen activator (tcu-PA) or urokinase.
Clinical studies.
Dose-finding studies.
Two dose-finding reperfusion studies in AMI patients with proven occluded coronary arteries with saruplase have indicated that favourable reperfusion results were to be expected with a 20 mg bolus and a 60 mg infusion over 60 min""12(Tablel).
138 Summary and conclusions
Table 1. Saruplase: Dosefindings studies11,1 2
Study TIMI (min) TIMI Dose:
40 mg 70 mg 80 mg v.d. Werf 60 3 2/8 7/9 60 2-3 6/8 Diefenbach 60 2-3 3/12 7/12 90 2-3 4/12 10/12 11. Circulation 1986;74:1066-1070. 12. Am J Cardiol 1988;61:966-970.
In the first dose-finding trial n , 8 patients received a bolus of 10 mg followed by an infusion of 30 mg over 60 min, and 9 patients received a 10 mg bolus followed by an infusion of 60 mg over 60 min. Of the 8 patients receiving a total dose of 40 mg saruplase only 2 had rapid complete opacification (TIMI grade 3) flow at 60 min and 6 out of 8 had rapid or delayed but complete opacification (TIMI grade 2-3), whereas 7 out of 9 patients receiving a total dose of 70 mg showed rapid complete filling (TIMI grade 3 flow) at 60 min. In the second dose-finding trial'2, 12 patients received a bolus of 10 mg, followed by an infusion of 30 mg over 60 min, and 12 patients received a bolus of 20 mg followed by an infusion of 60 mg over 60 min. At 60 min 3 out of 12 in the 40 mg group and 7 out of 12 in the 80 mg group had TIMI grade 2-3 flow. At 90 min, 4 out of 12 patients in the 40 mg group had TIMI grade 2-3 flow, and 10 out of 12 patients receiving 80 mg had TIMI grade 2-3 flow.
Heparin-interaction studies.
Based on animal experience1314 the LIMITS (Liquemin in Myocardial Infarction during Thrombolysis with saruplase) trial15 evaluated the effect of a bolus dose of heparin on the efficacy and safety of saruplase in patients with an acute myocardial infarction. The trial was designed to evaluate the
139 Chapter 9 effect of adjunctive unfractionated heparin on patency, incidence of bleeding complications, and haemostasis (Table 2).
Table 2. Saruplase Interaction with heparin LIMITS15
Sarupl aseÌ + héparine Saruplase + placebo N = 56 N=62 TIMI grade flow 78.6 56.5 6-12 h (%) Bleeding (%): all 14.3 8.1 severe 1.8 1.6 Stroke (%) 0 0 Death (%) 5.4 14.5 15 JACC 1995,26 365-373
The study aimed to include 200 patients with an acute myocardial infarction who could be treated within 6 hours after onset of symptoms. Patients were randomized to a bolus of heparin, 5000 U, or placebo, prior to thrombolysis with saruplase 20 mg bolus followed by an infusion of 60 mg over 60 min. Starting 30 min after completion of the infusion of saruplase, an i.v. infusion of heparin was given for 5 days. After inclusion of 56 patients in the heparin pre-treatment group and 62 patients in the placebo group, the central ethics and advisory board recommended the termination of the study, due to the evident superiority of heparin pre-treatment. The addition of a bolus of heparin prior to saruplase increased the patency rate (TIMI grade 2 and 3 ) at 6-12 hours after the start of fibrinolytic therapy from 56.5% to 78.6%. The group which had received pre-thrombolysis heparin had fewer in-hospital deaths than the placebo group (5.4% vs 14.5%), however there were a few more bleeding complications (14.3% vs 8.1%). There were no cerebrovascular accidents or allergic reactions in either group. The results of this study led to the recommendation that an i.v. bolus of heparin should precede the administration of saruplase.
140 Summary and conclusions
Comparison of saruplase with other fibrinolytic agents.
Streptokinase. Saruplase was compared to streptokinase in the PRIMI (Randomised double-blind trial of recombinant Pro-urokinase against streptokinase in acute myocardial infarction) trial16 and in the COMPASS (Randomized, double-blind study comparing saruplase with streptokinase therapy in acute myocardial infarction) trial17. In the PRIMI trial 401 patients with a first acute myocardial infarction were treated within 4 hours of onset of symptoms with 80 mg saruplase i.V., given as a 20 mg bolus followed by 60 mg infusion over 60 min (198 patients) or 1.5 million I.U. streptokinase infused over 60 min (203 patients). The primary end-point of the study was the patency (TIMI grade flow 2-3) of the infarct-related coronary artery 90 after start of infusion. Patency at 60 min was a secondary end-point (Table 3).
Table 3. Saruplase: Comparison of patency (TIMI 2-3) with streptokinase. PRIMI16
Drug Patients Patency 60 min 90 min Saruplase 198 71.8%* 71.2%** Streptokinase 203 48.0% 63.9% 16. Lancet 1989;1:863-868. (*p<0.001) (**p<0.15)
Other secondary endpoints were the rate of reocclusion during the first 24 to 36 hours, and haemostatic changes. Patency rates at 60 min were 71.8% for saruplase and 48.0% for streptokinase (p < 0.001) and at 90 min 71.2% and 63.9% (p = 0.15). At 24-36 hours 4.9% of patients treated with saruplase and 4.3% of patients treated with streptokinase showed reocclusion, however of those patients with patent arteries at 90 min in whom no further intervention was done, angiographic reocclusion was 1% for the saruplase patients and 4% for the streptokinase patients. At the end of the fibrinolytic infusion (60 min) fibrinogen concentration had decreased to 0.44 (0.23-1.27) g/1 (median, 1st and 3rd quartile) in patients treated with saruplase and to 0.17 (0.06- 0.27) g/1 in patients treated with streptokinase. Concentrations of fibrin(ogen) degradation products rose to 96 (24-240) mg/1 after saruplase and to 240 (192-360 ) mg/1 after streptokinase. Bleeding complications were
141 Chapter 9
14.1% in saruplase patients compared to 24.6% in streptokinase patients (p <0.01). Haemorrhagic stroke occurred in 1% of saruplase patients and in 0.5% of streptokinase patients (NS). In-hospital mortality rates were 3.5% for the saruplase patients and 4.9% for the streptokinase patients.The PRIMI trial demonstrated earlier reperfusion, higher patency rate, less haemostatic changes, and less bleeding complications for saruplase in comparison to streptokinase. Patency trials like the PRIMI trial show superiority in terms of reperfusion but they lack the statistical power to confirm differences in mortality. Mortality trials require tens of thousands of patients to demonstrate superiority in terms of mortality. Equivalence trials need less patients to demonstrate the efficacy of a new agent by demonstrating equivalence in terms of mortality to the current standard. This novel approach recently came to attention. The INJECT (International Joint Efficacy Comparison of Thrombolysis) trial, started in 1993 was designed to investigate equivalence of reteplase double-bolus injection and streptokinase18. In 1995 the INJECT investigators were the first to publish the results of a thrombolysis equivalence trial. The COMPASS Equivalence Trial17 sought to demonstrate the equivalence of saruplase and streptokinase in terms of 30-day mortality. This trial defined equivalence as the upper limit of the 95% confidence interval of the odds ratio for all cause mortality at 30 days for all patients randomized, not to exceed 1.5. Between 1992 and 1994 this study enrolled 3,089 patients. 30-Day mortality of any cause was 5.7% for patients randomized to saruplase and 6.7% for patients randomized to streptokinase (odds ratio 0.84, ρ < 0.01 for equivalence). Although haemorrhagic strokes were more prevalent in the saruplase-treated patients (0.9% vs 0.5%), the overall stroke rates were identical (1.4%). Further the net clinical benefit which corresponds to the combined rates of overall mortality and permanent disability from stroke was also found to be in favour of saruplase (6.0% vs 7.1%). Bleeding complications were not different for the two treatment arms (mild: 10.4% vs 10.9%; severe 2.1% vs 2.5%). Hypotension and cardiogenic shock were more common in the patients treated with streptokinase. Reinfarction rates were similar. All samples from streptokinase-patients showed antibody formation, whereas no antibody directed against saruplase was detected.
Thus the COMPASS study demonstrated that the 30-day mortality rate for saruplase treatment is at least equivalent to that of streptokinase (Table 4). This also applies to the net clinical benefit of saruplase treatment. In over 250 samples from saruplase-treated patients, no antibody against saruplase
142 Summary and conclusions or Escherichia coli protein was detected, while all samples, at discharge and at 30 days from the streptokinase-treated patients demonstrated antibody formation.
Table 4. Saruplase: Mortality equivalence with streptokinase. COMPASS17
Saruplase Streptokinase Ρ - value N=1.542 N=1.547
30-day Mortality (%) 5.7 6.7 0.242
1-year Mortality (%) 2 9J, 0.193 17. JACC 1998;31:487-493.
Urokinase. The human urinary enzyme urokinase or tcu-PA is an effective but expensive fibrinolytic agent that has been registered for acute myocardial infarction in some, but not all, European countries. For these reasons urokinase has not gained widespread popularity in the Netherlands. Therefore some of the trials with urokinase are reported briefly. Early myocardial infarction trials, where urokinase was given at low dose over a long period of time had disappointing results 19"21. Later trials with an i.v. bolus of 2 million I.U. demonstrated the drug to be safe and effective in achieving patency and in improving cardiac performance 22,23. More recently an i.v. bolus of 1.5 million I.U. followed by an infusion of 1.5 million I.U. in 1 hour, resulted in a patency similar to that of t-PA (alteplase) and appeared equally save despite causing more haemostatic disturbance 24. Recanalization of chronically occluded aortocoronary saphenous vene bypass grafts with long term, low dose direct infusion of urokinase has proved to be successful.25. A combination of urokinase and rt-PA was associated with a lower rate of reocclusion, a lower rate of recurrent ischemia, and fewer adverse events than either urokinase or rt-PA alone 26'27. These same investigators compared the same combination with different dose regimens of rt-PA, and found a patency rate of 77% for the combination 28. In a trial that was designed to study the effect of urokinase before angioplasty on thrombus formation and ischaemia in patients with unstable angina 29'30, and in which two different intracoronary doses of urokinase were tested, more complications were found with urokinase than with placebo. Unlike urokinase, or tcu-PA, its parent molecule scu-PA can selectively activate fibrin-bound plasmin. The induced clot lysis is amplified by plasmin-
143 Chapter 9 triggered conversion of scu-PA to urokinase. After the results of the PRIMI study became available, it became interesting to compare saruplase and urokinase. In the SUTAMI trial (Double-blind Multicenter Comparison of the Efficacy and Safety of Saruplase and Urokinase in the Treatment of Acute myocardial Infarction)31'33 543 patients with acute myocardial infarction within 6 hours of onset of symptoms were randomized to saruplase (20 mg bolus plus 60 mg infusion) or urokinase (1.5 million I.U. bolus plus 1.5 million I.U. infusion). The primary endpoint of this study was the patency rate at 24 to 72 hours. With hindsight the assumption that late patency, like early patency, may differ between plasminogen activators, was wrong, however at the time of the conception of this trial the "catch up phenomenon" was not yet fully appreciated 3A. So not surprisingly the patency rates at 24-72 hours were comparable: 75.4% and 74.2%, respectively. It cannot be excluded, that an early patency difference was missed, because of the design of the study. There was a trend toward more haemorrhagic strokes in the saruplase group (1.1%) vs the urokinase (0%) group ( ρ = 0.25), but in-hospital mortality had the opposite trend: saruplase 4.4% and urokinase 8.1% ( ρ = 0.08) . The mortality rates for anterior infarction reflect the overall rates: 5.5% for saruplase and 11.0% for urokinase (Table.5).
Table 5. Saruplase: Comparison with urokinase. SUTAMI31 Saruplase Urokinase Ρ - value
TIMI 2-3 at 24-72 h (%) 75.4 74.2 0.77 ICH(%) 1.1 0 0.25 Mortality (%) 4.4 8.1 0.08
31.JThromb Thrombol 1995;2:117-124. (ICH = Intracranial haemorrhage)
Fibrinogen degradation products rose markedly after saruplase, from 0.43 mg/1 at baseline to 160 mg/1 at 2 hours and were still elevated to 0.67 mg/1 at 36-72 hours, whereas the increase after urokinase (from 0.45 mg/1 to 89.0 mg/1) was significantly (p < 0.001) smaller (Fig. 1). Other haemostatic changes were virtually identical. Thus, at the dosis used in the SUTAMI
144 Summary and conclusions study and in other clinical trials, the intrinsic clot selectivity of saruplase is completely masked. Fig. 1. Fibrinogen degredation products (median, mg/L)
1Í3
150-
125^
100 i
75"
50- variance analysis *·, Ν. p<0 001 ft./\ 25- "··
и ι 1 I 1 0 1 2 6-12 24-36 36-72 h
ь- saruplase 0 43 0 43 0 43 0 43 0 43 0 43 "О f- urokinase 0 45 0 45 0 45 0 45 0 45 0 45
Figure 1: Fibrinogen degradation products after saruplase and urokinase. theSUTAMl study.
A pharmacokinetic study 35 demonstrated the conversion of approximately 28% of saruplase into its active two-chain degradation product and the haemostatic effects observed in our study confirm early and substantial conversion of scu-PA into tcu-PA by plasmin. The generation of more fibrinogen degradation products should be considered a dose effect. The median concentrations of both thrombin-antithrombin III complex (TAT) and prothrombin fragments 1 +2 (Fi + 2) significantly increased 3- to 6-fold after the therapy, indicating extensive activation of the coagulation system. Following heparin administration, both parameters returned to normal in most patients. There was never any significant difference in these coagulation parameters between patients with and without reinfarction. Therefore, in contrast to other findings these parameters were no useful indicators of reinfaction in this study32. No antibody against saruplase was detected in the samples of 241 saruplase- treated patients, and no antibody against Escherichia coli protein was detected in 19 randomly selected samples from saruplase-treated patients.
145 Chapter 9
Alteplase. Saruplase was compared with alteplase in the Belgian Saruplase Alteplase Trial (SAT)36 and in the SESAM trial (Comparison of Saruplase and Alteplase in Acute myocardial infarction)37.The primary aim of the SAT trial was to study the effects of alteplase and saruplase on haemostatic variables. Secondary endpoints were patency of the infarct-related artery 24 to 72 hours after start of thrombolytic treatment and the safety of the two thrombolytic regimens. Fifty-two patients with acute myocardial infarction within 4 hours of onset of symptoms were randomized to saruplase 20 mg bolus i.v. followed by 60 mg i.v. over 1 hour, orto alteplase 10 mg bolus i.v. followed by an infusion of 50 mg i.v. over 1 hour, and then 40 mg in 2 hours. Heparin was started 30 min after the end of thrombolytic treatment and continued for 5 days during which no antiplatelet drugs were allowed. Alteplase caused significantly less haemostatic disturbances than saruplase. Fibrinogen, plasminogen and а.г - antiplasmin levels were consistently higher throughout the period of measurements from baseline to 24 hours, and alteplase generated less fibrinogen degradation products. However, in contrast to the assumption that the degree of haemostatic alteration correlates with bleeding, such bleeding complications were more frequent in the alteplase group. Haemorrhagic stroke occurred in 1 patient in the alteplase group. The SESAM trial 37 compared saruplase and alteplase in acute myocardial infarction with the primary aim to compare reocclusion after 24-40 hours. Other endpoints were patency rates at 45, 60 and 90 min, and safety data (Table 6). In this study 473 patients were randomized to saruplase (n = 236), 20 mg bolus i.v. plus 60 mg i.v. over 1 hour or to alteplase (n = 237), 10 mg bolus i.v. plus 50 mg i.v. infusion over the next hour, followed by 40 mg i.v. infusion for the next 2 hours. Heparin was given as a bolus 5000 U before fibrinolytic therapy followed by an infusion, that was continued for 24-40 hours. The angiographic reocclusion rate was remarkably low in the saruplase group, although not significantly different to that of the alteplase group (1.2% for the saruplase group and 2.4% for the alteplase group). Angiography showed very high patency rates (TIMI grade 2 and 3 flow) for saruplase and alteplase: 45 min, 74.6% vs 68.9% (p = 0.22); 60 min, 79.9% vs 75.3% (p = 0.26), 90 min, 79.9% vs 81.4% (p = 0.72). Haemorrhagic stroke occurred in 0.8% of patients in each group. Thus, in this study the safety and efficacy of saruplase and alteplase over 3 hours, are very similar. Although the outmoded 3-hour regimen of alteplase was used in this study,
146 Summary and conclusions patency rates of 75.3% at 60 min, and of 81.4% at 90 min in 237 patients in the alteplase arm of the SESAM study were comparable to those obtained with front-loaded t-PA in the three small studies that changed the infusion policy of alteplase to front-loaded administration38"40: in 74 patients Neuhaus 38 demonstrated a patency of 74 % at 60 min, and 90.5% at 90 min; in 143 patients the RAAMI trial39 demonstrated a patency of 62.0% at 60 min, and of 82.0% at 90 min, and in 209 patients the TAPS trial40 showed a patency of 74.0% at 60 min, and of 85.0% at 90 min.
Table 6. Saruplase: Comparison with alteplase. Sesam37
Saruplase Alteplase Ρ - value
Reocclusion 24-40 h(%) 1.2 2.4 P=0.68 TIMI 2-3 at 45 min (%) 74.6 68.9 P=0.22 TIMI 2-3 at 60 min (%) 79.9 75.3 P=0.26 TIMI 2-3 at 90 min (%) 79.9 81.4 P=0.72
37. Am J Cardiol 1997;79:727-732.
In conclusion, the standard therapy with saruplase compares favourably in terms of efficacy with other thrombolytic medications. Saruplase acts faster than streptokinase and its efficacy is at least comparable to that of urokinase and alteplase. Although standard therapy with saruplase causes less haemostatic changes than streptokinase, similar changes as urokinase, and more changes than alteplase (figure 2), its clinical safety profile is very similar to that of the other plasminogen activators. The rate of intracranial haemorhage seen with saruplase is higher than that seen with streptokinase, though not significantly so, and not different from that seen with t-PA16,17,37. Sofar no antibody against saruplase or against Escherichia coli protein has been detected and no allergic reaction has occurred in patients treated once or twice17'31.
Bolus administration. Bolus therapy facilitates pre-hospital administration of thrombolytic therapy. Before focusing on saruplase, the history of bolus therapy and the new development of mutants of t-PA with a prolonged hálf-life, designed for
147 Chapter 9 bolus therapy, are discussed. So far two fibrinolytic agents have been licensed for bolus administration, anistreplase for a single bolus and reteplase for double bolus administrations, 30 min apart. In anistreplase, or anisoylated plasminogen-streptokinase activator complex (APSAC), acylation with a p-anisoyl group reversibly inactivates the catalytic centre41. Deacylation and, thereby, activation occurs spontaneously at the fibrin surface and in the circulation. Anistreplase has a plasma half- life of 70 min42. In comparison to streptokinase, anistreplase causes more fibrinogen depletion and longer total fibrinolytic activity with less reocclusion43'44. Anistreplase has no survival benefit over streptokinase45. Although domiciliary thrombolysis with anistreplase demonstrated halving of mortality at one year46, anistreplase has not taken advantage of being the first fibrinolytic agent available for single bolus administration. It has all the shortcomings of streptokinase, and it is doubtful whether it can compete against the mutants of wild type t-PA and other physiological newcomers in the field, if these can be given as a single bolus.
Fig.2. Fibrinogen concentration (g/L) 4
0—r Ί I I I I I I Ι Γ 0 1 2 6-12 24-36 36-72 hours Figure 2: Fibrinogen concentrations after saruplase, urokinase, streptokinase, and alteplase.
Reteplase is a unglycosylated single-chain deletion mutant of t-PA expressed in Escherichia coli, with a lower fibrin-binding and a half-life of approximately 18 min47. Double bolus administration (10 U + 10 U, interval 30 min) of reteplase proved superior to t-PA in achieving 90-min TIMI grade
148 Summary and conclusions
3 flow and showed less ventricular dysfunction and cardiogenic shock ' The equivalence of reteplase and streptokinase in terms of mortality has been established18, however a study that compared reteplase and alteplase in 15,059 patients found no superiority in 30-day mortality 50. Its ease of administration renders reteplase an attractive alternative to alteplase. TNK-t- PA is a mutant of t-PA without deletions, but with amino acid substitutions at 3 sites, produced in Chinese hamster ovary (CHO) cells. TNK-t-PA has high fibrin specifity due to avid fibrin binding, a half-life that is prolonged to 18 min, and is resistant to plasminogen activator inhibitor-1 (PAI-1). In animal experiments it produced rapid and complete thrombolysis with a decreased risk of intracranial haemorrhage 51,52. Angiographic, pharmacokinetic and clinical dose-ranging studies have been completed 53,54. A single 40 mg bolus achieves similar 90 min TIMI grade 3 flow as front- loaded t-PA54. The preliminary results (American College of Cardiology, 48th Annual Scientific Session, 1999) of a mortality trial (ASSENT-2), that compares TNK-t-PA and front-loaded alteplase in 16.505 patients show no differences in 30 day mortality (6.16% vs 6.18%), or in intracranial haemorrhage (0.93% vs 0.94%). Lanetoplase or η-PA is a deletion and a point mutant of native t-PA produced in HO cells, with reduced fibrin binding and a half-life of 30-40 min55. A clinical dose-ranging trial, comparing weight-adjusted η-PA with front-loaded t-PA, has shown a better patency at 90 min, without haemorrhagic stroke in patients treated with 120 kU/kg η-PA56. Surprisingly, the preliminary results (American College of Cardiology, 48th Annual Scientific Sessions, 1999) of a large mortality trial (Intime-II), that compares η-PA with front-loaded t-PA in 15.078 patients, demonstrate significantly more intracranial haemorrhage with η-PA than with t-PA (1.13% vs 0.62%, ρ = 0.03). This study showed no difference in 30 day mortality (6.77% vs 6.60%) for η-PA and t-PA. In an early attempt to use a single bolus regimen in AMI, the efficacy of a bolus of 2 million I.U. of urokinase has been demonstrated22,23, but this has not been further developed. A single bolus t- PA did not result in satisfactory patency57, but two boluses 30 min apart did58 and a 100 mg double-bolus alteplase has demonstrated a TIMI grade 3 flow of 88% at 90 min59. A subsequent trial in 7,169 patients demonstrated no superiority of double bolus over front-loaded t-PA in terms of mortality and did not change the administration policy of t-PA60. The Bolus Administration of Saruplase in Europe (BASE) pilot study compared the efficacy in terms of persistent patency (an open vessel at 90 min and at 24 h) and safety of a 20 mg i.v. bolus followed by an infusion of
149 Chapter 9
60mg/h (standard regimen) with single boluses of 80 mg and 60 mg, and with a split-bolus of 40 mg with a 30 min interval. This pilot study showed comparable efficacy and safety profiles of the 80 mg bolus and and of standard therapy. The 60 mg bolus was safe but not sufficiently effective, and was therefore abandoned early. The highest patency rate, but also the highest mortality rate were found for the split-bolus regimen. The single bolus of 80 mg produced a patency rate comparable to that achieved by the 40 mg split-bolus, with a death rate of 6%, which was comparable to the standard regimen 61. The pharmacokinetics and haemostasiology were studied in a substudy of BASE (H.R. Michels, submitted for publication). The prematurely abandoned ineffective 60 mg bolus regimen was excluded from this substudy. The single bolus was found to result in very high early total u-ΡΑ activity in the circulation, which accelerates and enhances the generation of tcu-PA activity, leading to accelerated and enhanced fibrinogen consumption (Figure 3, 4).
Fig. 3. Plasma concentration of tcu-PA activity (pg/ml) с [pg/ml] D standard bolus split bolus 4 - • - »- + ..φ.
3 - •
• 2 - • 1 • w • • _î-->-4— -- *·* ·*· = -^*-.*--- *—---:¿:-_-.- ..„ о -1—1 '"""' _ · — 0 30 60 90 120 t (min) Figure 3: Tcu-PA activity after bolus 80 mg, split-bolus 2 χ 40 mg, and standard regimen.
The Bolus versus Infusion in RescupaseR (saruplase) Development (BIRD) study was designed to test the equivalence of efficacy and safety of the standard regimen and the 80 mg single bolus injection of saruplase62.
150 Summary and conclusions
Primary endpoint was 30 day-mortality. Secondary endpoints were stroke and severe bleeding. 2410 Patients were randomized to the standard regimen or 80 mg single bolus. Fig.4. Fibrinogen levels (mg/dl) mg/dl
ou - standard bolus split bolus 00 - • • · *• - • 50 -
'"•*··.. 00 - ' · - *.
so - • * . 00 -
50 - •
30 60 120 (mm) Figure 4: Fibrinogen consumption after bolus 80 mg, split-bolus 2 χ 40 mg, and standard regimen.
The death rates in both groups were comparable (6.0% and 5.9%). The rates of intracranial haemorrhage and severe bleeding were identical. The rate of reinfarction was slightly higher in the patients receiving the single bolus (Table 7).
Table 7. Saruplase: 30-Day mortality. Bolus vs Standard. BIRD55 Bolus 80 mg Standard:bolus 20mg, infusion 60 mg/l h
30-day mortality (%) 6.0 5.9 ICH (%) 0.8 0.7 Severe bleeding (%) 2.5 2.1 Reinfarction (%) 6.5 5Λ 55. Circulation 1998,98 (supplì): 1-505. (ICH = Intracranial haemorrhage)
151 Chapter 9
Conclusions. Haemostasiology of standard therapy .Treatment with saruplase given as an i.v. bolus of 20 mg followed by an i.v. infusion of 60 mg over 1 hour (standard therapy) causes less haemostatic changes than streptokinase treatment. The haemostatic changes following standard therapy of saruplase and urokinase treatment are very similar, but saruplase generates more degradation of fibrinogen. Standard therapy with saruplase causes more haemostatic disturbances than standard treatment with alteplase. However, haemostatic alterations do not seem to affect clinical safety. Haemostasiology of single bolus therapy. The administration of a single bolus of 80 mg saruplase results in very high early total u-ΡΑ activity and accelerated and enhanced generation of tcu-PA activity in comparison to standard therapy. Consequently the administration of a single bolus of 80 mg saruplase results in accelerated and enhanced plasminogen activation and fibrinogen degradation. There was no evidence of a plasminogen steal phenomenon affecting thrombolytic efficacy. Reperfusion and patency. Saruplase demonstrates earlier reperfusion and higher patency rates than streptokinase. The patency rates at 45, 60 and 90 min after saruplase treatment and after alteplase treatment are not significantly different. Reocclusion. The angiographic reocclusion rates after saruplase treatment were remarkably low, but not significantly different to that of the alteplase group in the SESAM trial. Mortality. The 30-day mortality rate for saruplase treatment is at least equivalent to that of streptokinase treatment. The 30-day mortality rates after a single bolus of 80 mg saruplase and a bolus of 20 mg followed by an infusion of 60 mg over 1 hour are equivalent. Intracranial haemorrhage. The rate of intracranial haemorrhage with saruplase is higher than than that seen with streptokinase, but not significantly so. The rate of intracranial haemorrhage with saruplase is not different from that seen with alteplase. The rates of intracranial haemorrhage and severe bleeding after a single bolus of 80 mg saruplase and after a 20 mg bolus followed by an infusion of 60 mg over 1 hour are comparable. Net clinical benefit. The net clinical benefit, as indicated by the overall mortality plus the disabling stroke rate, is at least equivalent to that of streptokinase despite a higher rate of intracranial Haemorrhage. Antibody formation and allergy. No antibody against saruplase or against Escherichia coli protein have been detected. No saruplase-related allergic reaction has occurred in patients treated once or twice.
152 Summary and conclusions
Acknowledgement. We thank Guy van Dael for his graphical assistence.
References.
1. Holmes W, Pennica D, Blaber M et al: Cloning and expression of the gene for pro-urokinase in Escherichia coli. Biotechnology 1985; 3: 923-929. 2. Michels HR. Saruplase in the context of other thrombolytic medications. Rev. Contemp. Pharmacother. 1998; 9: 403-409. 3. Gurewich V, Pannell R: Fibrin-specificity and efficacy of proteolysis induced by single-chain urokinase in plasma. Thromb. Haemost. 1983; 50: 321. 4. Gurewich V, Pannell R, Louie S , Kelley P, Suddith RL, Greenlee R. Effective and fibrin- specific clot lysis by a zymogen precursor form of urokinase (pro- urokinase). J. Clin. Invest. 1984; 73: 1731-1739. 5. Stump D, Thienpont M, Collen D. Urokinase-related proteins in human urine. J. Biol. Chem. 1986; 261: 1267-1273. 6. Lijnen H, Zamarron С, Blabber M, Winkler ME, Collen D: Activation of plasminogen by pro-urokinase, I. Mechanism. J. Biol. Chem. 1986; 261: 1253- 1258. 7. Collen D, Stassen J, Blaber M, Winkler M, Verstraete M . Biological and thrombolytic properties of proenzyme and active forms of human urokinase. III. Thrombolytic properties of natural and recombinant urokinase in rabbits with experimental jugular vene thrombosis. Throm. Haemost. 1984; 52: 27-30. 8. Pannell R, Gurewich V. Pro-urokinase - a study of its stability in plasma and of a mechanism for its selective fibrinolytic effect. Blood 1986; 67:1215-1223. 9. Hanbrücken F, Schneider J, Gunzler W, Friderichs E, Giertz H, Flohé L. Selective fibrinolytic activity of recombinant human pro-urokinase (single-chain urokinase-type plasminogen activator) from bacteria. Arzneimittelforschung 1987;37:993-997. 10. Gurewich V. The sequential, complementary and synergistic activation of fibrin- bound plasminogen by tissue plasminogen activator and pro-urokinase. Fibrinolysis 1989;3:59-66. 11. van de Werf F, Vanhaecke J, de Geest H, Verstraete D, Collen D. Coronary thrombolysis with recombinant single-chain urokinase-type plasminogen activator in patients with acute myocardial infarction. Circulation 1986; 74: 1066-1070. 12. Diefenbach C, Erbel R, Pop Τ et al: Recombinant single-chain urokinase-type plasminogen activator during acute myocardial infarction. Am J Cardiol 1988; 61:966-970. 13. Schneider J. Interactions of saruplase with acetylsalisylic acid, heparin, glyceryl nitrate, tranexamic acid, and aprotinin in a rabbit pulmonary thrombosis model. Arzneimittelforschung 1990; 40(11): 1180-1184.
153 Chapter 9
14. Schneider J. Heparin and the thrombin inhibitor argatroban enhance fibrinolysis by infused or bolus-injected saruplase (r-scu-PA) in rabbit femoral artery thrombosis. Thromb Res 1991; 64: 677-689. 15. Tebbe U, Windeier J, Bösl 1 et al: Thrombolysis with recombinant unglycosylated single-chain urokinase-type plasminogen activator (saruplase) in acute myocardial infarction: influence of heparin on early patency rate (LIMITS Study). J Am Coll Cardiol 1995; 26:365-373. 16. PRIMI Trial Study Group. Randomised double-blind trial of recombinant pro- urokinase against streptokinase in acute myocardial infarction. Lancet 1989; 1: 863-868. 17. Tebbe U, Michels R, Adgey J et al. Randomized double-blind study comparing saruplase with streptokinase therapy in acute myocardial infarction. J Am Coll Cardiol 1998;31:487-493. 18. INJECT: International Joint Efficacy Comparison of Thrombolysis. Randomised, double-blind comparison of reteplase double-bolus administration with streptokinase in acute myocardial infarction (INJECT): trial to investigate equivalence. Lancet 1995; 346: 329-336. 19. European Collaborative Study. Controlled trial of urokinase in myocardial infarction. Lancet 1975; 2: 624-626. 20. Gormsen J, Tidstrom B, Fedderson C, Ploug J. Biochemical evaluation of low dose urokinase in acute myocardial infarction. A double-blind study. Acta Med. Scandl973; 194:191-198. 21. Brochier M, Raynaud R, Planiol Τ et al. Le traitement par l'urokinase des infarctus du myocarde et syndromes de menace. Etude randomisée de 120 cas. Arch. Mal. Coeur 1975; 68:563-569. 22. Mathey DG, Schofer J, Sheehan FH, Becher H, Tilsner V, Dodge H. Intravenous urokinase in acute myocardial infarction. Am J Cardiol 1985; 5:878-82. 23. Penco M, Fedele F, Agati L et al. Effects of systemic treatment with urokinase UK) on left ventricular function. Eur Heart J 1987; 8 (suppl. 2): 24. 24. Neuhaus K-L, Tebbe U, Gottwick M et al. Intravenous recombinant tissue plasminogen activator (rt-PA) and urokinase in acute myocardial infarction. Result of the German Activator Urokinase Study (GAUS). J Am Coll Cardiol 1988; 12:581-587. 25. Hartmann JR, McKeever LS, O'Neill WW et al. Recanalization of chronically occluded aortocoronary saphenous vein bypass grafts with long-term low dose direct infusion of urokinase trial (ROBUST): a serial trial. J Am Coll Cardiol 60-66. 26. Califf RM, Topoi EJ, Stack RS et al. Evaluation of combination thrombolytic therapy and timing of cardiac catheterization in acute myocardial infarction. Results of Thrombolysis and Angioplasty in Myocardial infarction - phase 5 randomised trial. TAMI Study Group. Circulation 1991; 83: 1543-1546.
154 Summary and conclusions
27. Ward SR, Sutton JM, Pieper KS Schwaiger M, Caliif RM, Topoi EJ. Effects of thrombolytic regimen, early catheterization, and predischarge angiographic variables on six-week left ventricular function. Am J Cardiol 1997; 79:539-544. 28. Wall TC, Califf RM, George BS et al. Accelerated plasminogen activator dose regimens for coronary thrombolysis. Thrombolysis and Angioplasty in Myocardial Infarction - 7 (TAM1-7). J. Am Coll Cardiol 1992; 19: 482-489. 29. Ambrose JA, Almeida OD, Sharma SK el al. Adjunctive thrombolytic therapy during angioplasty for ischemic rest angina. Results of the TAUSA trial. TAUSA investigators. Thrombolysis and Angioplasty in Unstable Angina trial. Circulation 1994; 90: 69-77. 30. Mehran R, Ambrose JA, Bongu RM et al. Angioplasty of complex lesions in ischemic rest angina: results of the Thrombolysis and Angioplasty in Unstable Angina (TAUSA) trial. J Am Coll Cardiol 1995; 26: 961-966. 31. Michels R, Hoffmann H, Windeier J et al. A double-blind multicenterr comparison of the efficacy and safety of sarupkase and urokinase in the treatment of acute myocardial infarction: report of the SUT AMI study group. J. Thromb Thrombol 1995; 2: 117-124. 32. Hoffmann J, Michels HR, Windeler J, Günzler WA. Plasma markers of thrombin activity during coronary thrombolytic therapy with saruplase or urokinase: no prediction of infarction. Fibrinolysis 1993; 7: 330-334. 33. Michels HR, Hoffmann JJ, Windeier J, Hopkins GR. Hemostatic changes after thrombolytic therapy with saruplase (unglycosylated single-chain urokinase-type plasminogen activator) and urokinase (two-chain urokinase-type plasminogen activator). Blood Coag. and Fibrinolysis 1996; 7: 766-771. 34. Granger CB, Califf RM, Topoi EJ. Thrombolytic therapy for acute myocardial infarction. A review. Drugs 1992; 44: 293-325.. 35. Koster RW, Cohen AF, Hopkins GR, Beier H, Günzler WA, Wouw PAV. Pharmacokinetics and pharmacodynamics of saruplase, an unglycosylated single-chain urokinase-type plasminogen activator, in patients with acute myocardial infarction. Thromb. Haemostasis 1994; 71: 740-744.. 36. The Belgian Saruplase Alteplase Trial Group. Effects of alteplase and saruplase on hemostatic variables: a single-blind, randomized trial in patients with acute myocardial infarction. Coron. Artery Dis 1991; 2: 349-355. 37. Bär F, Meyer J, Vermeer F et al. Comparison of saruplase and alteplase in acute myocardial infarction. Am. J. Cardiol. 1997; 79: 727-732. 38. Neuhaus K-L, Feuerer W, Tebbe SJ, et al: Improved thrombolysis with a modified dose regimen of recombinant tissue-type plasminogen activator. J AM Coll Card 1989; 14: 1566-1569. 39. Carney R, Brandt TR, Daley Ρ et al: Increased efficacy of rt-PA by more rapid administration: the RAAMI trial. Circulation 1990; 82 (suppl III): III-538. 40. Neuhaus K-L, von Essen R, Tebbe U et al. Improved thrombolysis with front- loaded administration of alteplase: results of the rt-PA-ABSAC Patency Study (TAPS). J Am Coll Cardiol. 1992; 19: 885-91.
155 Chapter 9
41. Smith RAG, Dupe RJ, English PD, et al: Fibrinolysis with acyl-enzymes: a new approach to thrombolytic therapy. Nature 1981; 290: 505-508. 42. Staniforth DH, Smith RAG, Hibbs M: Streptokinase and anisoylated streptokinase plasminogen complex. Their action on haemostasis in human volunteers. Eur J Clin Pharmacol 1983; 24: 751-756. 43. Hoffmann JJML, Bonnier JJRM, de Swart JBRM: Systemic effects of thrombolytic drugs in acute myocardial infarction: comparison of intravenous APSAC (BRL 26921) and intracoronary streptokinase. Fibrinolysis 1987; 1: 225-230. 44. Bonnier JJRM, Visser RF, Klomps HC, Hoffmann JJML and the Dutch Invasive Reperfusion Study Group. Am J Cardiol 1988; 62: 25-30. 45. ISIS-3 (Third International Study of Infarct Survival) Collaborative Group: ISIS-3: A randomised comparison of streptokinase vs tissue plasminogen activator vs anistreplase and of aspirin plus heparin vs aspirin alone among 41,299 cases of suspected acute myocardial infarction. Lancet 1992; 339: 753- 770. 46. Rawles J, on behalf of the GREAT Group. Haqlving mortality at 1 year by domociliary thrombolysis in the Grampian Region Early Anistreplase Trial (GREAT). J Am Coll Cardiol 1994; 23: 1-5. 47. Noble S, McTavish D. Reteplae. A review of its pharmacological properties and clinical efficacy in the management of acute myocardial infarction. Drugs 1996; 4: 589-605. 48. Smalling RW, Bode C, Kalbfleisch J et al. More rapid, complete, and stable coronary thrombolysis with bolus administration of reteplase compared with alteplase infusion in acute myocardial infarction. Circulation 1995; 91: 2725- 2732. 49. Bode C, Smalling RW, Berg G et al. Randomized comparison of coronary thrombolysis achieved with double-bolus reteplase (recombinant plasminogen activator) and front-loaded, accelerated alteplase (recombinant tissue-type plasminogen activator) in patients with acute myocardial infarction. Circulation 1996;94:891-898. 50. The Global Use of Strategies to Open Occluded Coronary arteries (GUSTO III) investigators. A comparison of reteplase with alteplase for acute myocardial infarction. N Engl J Med 1997; 337:1118-1123. 51. Benedict CR, Refino CJ, Keyt BA, et al. New variant of human tissue plasminogen activator (TPA) with enhanced efficacy and lower incidence of bleeding compared with recombinant human TPA. Circulation 1995; 92: 3032- 3040. 52. Thomas GR, Thibodeaux H, Errett CJ et al. A long-half-life and fibrin-specific form of tissue plasminogen activator in rabbit models of embolic stroke and peripheral bleeding. Stroke 1994; 25: 2072-2079.
156 Summary and conclusions
53. Cannon CP, McCabe CH, Gibson CM et al. TNK-tisue plasminogen activator in acute myocardial infarction. Results of the Thrombolysis in myocardial infarction (TIMI) 10A dose-ranging trial. Circulation 1997; 95:351-356. 54. Cannon CP, Gibson CM, McCabe CH et al. TNK-tissue plasminogen activator compared with front-loaded alteplase in acute myocardial infarction: results of the TIMI 10B trail, thrombolysis in Myocardial infarction (TIMI) 10B Investigators. Circulation 1998; 98: 2805-2814. 55. Larsen GR, Timony GA, Horgan PG et al. Protein engineering at novel plasminogen activators with increased thrombolytic potency in rabbits relative to activase. J Biol Chem 1991; 266: 8156-8161. 56. den Heijer P, Vermeer F, Ambrosioni E et al. Evaluation of a weight-adjusted single- bolus plasminogen activator in patients with myocardial infarction. A double-blind, randomized angiographic trial of lanetoplase versus alteplase. Circulation 1998; 98: 2117-2125. 57. Tranchesi B, Chamone DF, Cobbaert C, vd Werf F, Vanhove Ρ, Verstraete M. Coronary recanalization rate after intravenous bolus of alteplase in acute myocardial infarction. Am J Cardiol 1991; 68: 161-165. 58. Gemmili JD, Hogg KJ, Maclnryre PD, Booth N, Rae AP, Dunn FG, Hillis WS. A pilot study of the efficacy and safety of bolus administration of alteplase in acute myocardial infarction. Br Heart J 1991; 66: 134-138. 59. Purvis JA, McNeill AJ, Siddiqui et al. Efficacy of 100 mg of double-bolus alteplase in achieving complete perfusion in the treatment of acute myocardial infarction. J.Am.Coll.Cardiol. 1994;23:6-10. 60. The Continuous Infusion versus Double-Bolus Administration of Alteplase (COBALT) Investigators. A comparison of continuous infusion of alteplase with double-bolus administration for acute myocardial infarction. N Engl J Med 1997; 337: 1124-30. 61. Bär FW, Meyer J, Vermeer F et al. Bolus administration of saruplase in Europe (BASE) a pilot study in patients with acute myocardial infarction. J Thromb Thrombolysis 1998; 6: 147-153. 62. Bär FW, Hopkins G, Dickhoet S. The Bolus versus Infusion in Rescuepase (saruplase) Development (BIRD) study in 2410 Patients with Acute Myocardial Infarction. Circulation 1998; 98: 1-505.
157 158 CHAPTER 10
Samenvatting en conclusies.
Saruplase (unglycosylated human recombinant single-chain urokinase-type plasminogen activator, scu-PA) wordt geproduceerd door genetisch gemanipuleerde Escherichia coli. Saruplase heeft een intrinsieke plasminogeen activerende werking, die gericht is op het aan fibrine gebonden plasminogeen. Het lokaal gevormde piasmine splitst saruplase in urokinase (two-chain urokinase-type plasminogen activator, tcu-PA).
Klinische studies
Dosering
Uit onderzoek naar een effectieve dosering bleek, dat er goede resultaten werden bereikt met een intraveneuze bolus van 20 mg gevolgd door een intraveneus infuuus van 60 mg in 1 uur (standaard behandeling).
Interactie met héparine
Uit dierproeven en uit onderzoek bij mensen bleek dat er een betere reperfiisie optrad indien de toediening van saruplase werd voorafgegaan door toediening van een bolus héparine van 5000 I.E.
Vergelijking met andere fibrinolytica
Streptokinase. Saruplase werd met streptokinase vergeleken in het PRIMI onderzoek. In dit onderzoek werden 401 patiënten dubbel-blind gerandomiseerd behandeld met saruplase (20 mg bolus + 60 mg infuus in 1 uur), of met 1,5 miljoen eenheden streptokinase in een infuus in 1 uur. Het primaire doel van deze studie was een vergelijking van het aantal open arteriën , 90 minuten na start van de behandeling. Er was een duidelijk
159 Chapter 10
maar niet significant verschil ten voordele van saruplase (71,2% vs 63,9%, ρ = 0,15), bovendien bleek het verschil 60 minuten na start van de behandeling wel significant (71,8& vs 48,0%, ρ < 0,001). Saruplase veroorzaakte een geringere verstoring van de hémostase, en minder bloedingen (14,1% vs 4,6%, ρ < 0,01). Het COMPASS onderzoek vergeleek de mortaliteit na een behandeling met saruplase of streptokinase na 30 dagen. Het betrof een equivalentie onderzoek, waarbij equivalentie werd gedefinieerd als de bovenste limiet van het 95% confidentie interval van een odds ratio voor dood door alle oorzaken, van minder dan 1,5. In dit onderzoek werden 3089 patiënten gerandomiseerd. De sterfte na 30 dagen bedroeg 5,7% voor saruplase en 6, 7% voor streptokinase (odds ratio 0,84, ρ <0,01). Er was geen verschil in bloedingscomplicaties, en hoewel meer patiënten in de saruplase groep een hersenbloeding doormaakten dan in de streptokinase groep, bereikte dit verschil geen statistische significantie (0,9% vs 0,5%), en werd het netto klinische effect hierdoor niet negatief beïnvloed. Er werden geen antilichamen tegen saruplase aangetoond terwijl alle patiënten die met streptokinase waren behandeld antilichamen vertoonden. Urokinase. In het SUTAMI onderzoek werden 543 patiënten gerandomiseerd naar een behandeling met saruplase of urokinase. Het primaire doel was de beoordeling van het aantal open bloedvaten na 24 tot 72 uur. Achteraf was de keuze van dit eindpunt onjuist, maar bij het ontwerpen van de studie was het bestaan van een laat inhaaleffect (catch up phenomenon), waardoor er na 24 uur geen verschillen meer worden gezien tussen de verschillende fi brino lytica, nog niet goed bekend. Het aantal open bloedvaten na 24-72 uur bleek gelijk (75,4% vs 75,2%). Ondanks het optreden van meer hersenbloedingen in de saruplase groep( 1,1% vs 0%, p=0,25), was de sterfte in de saruplase groep duidelijk, maar niet significant lager ( 4,4% vs 8,1%). De hemostatische veranderingen na beide middelen kwamen nauwkeurig overeen, met uitzondering van de fibrinogeen degradatie producten, die een significant grotere stijging vertoonden na saruplase, waarschijnlijk het gevolg van een conversie van ca. 28% scu-PA naar tcu-PA, hetgeen de gebruikte dosering urokinase beduidend overtreft. Alteplase. Saruplase werd met alteplase vergeleken in het SAT en in het SESAM onderzoek. Het SAT onderzoek, bij 52 patiënten, had als doel een vergelijking van de hemostatische parameters. Alteplase veroorzaakte significant minder verstoring van de hémostase dan saruplase, maar meer bloedingscomplicaties en de enige hersenbloeding trad op in de alteplase
160 Samenvatting en conclusies groep. In het SESAM onderzoek werden 473 patiënten gerandomiseerd met als primaire doel, de reocclusie na 24-40 uur te vergelijken. Ander doel was het aantal open bloedvaten 45, 60 en 90 minuten na start van de behandeling te vergelijken. De angiografische reocclusie toonde een gering maar niet significant verschil (saruplase 1,2%, alteplase 2,4%). Het aantal open bloedvaten na saruplase of alteplase vertoonde evenmin significante verschillen: 45 min, 74,6% vs 68,9%; 60 min, 79,9% vs 75,3%; 90 min 79,9% vs 81,4%. In beide groepen kreeg 0,8% een hersenbloeding.
Bolus, split-bolus of standaard behandeling.
Het BASE pilot onderzoek vergeleek de effectiviteit (open vaten na 90 min en 24 uur) en de veiligheid van standaard therapie met die van een bolus van 80 mg, een bolus van 60 mg en een split-bolus van 2 χ 40 mg met een interval van 30 minuten. De effectiviteit en veiligheid van de 80 mg bolus en standaard therapie waren vergelijkbaar. De split-bolus was zeer effectief maar geassocieerd met een hoge mortaliteit en de 60 mg bolus was niet effectief genoeg. In een substudie van de BASE studie vergeleken wij de farmacokinetiek en hemostatische veranderingen van de bolus van 80 mg, de split-bolus en de standaard behandeling. De bolus van 80 mg veroorzaakte een zeer vroege en zeer hoge tcu-PA activiteit en een snel en aanhoudend verlies van fibrinogeen. Het BIRD onderzoek bij 2410 patiënten vergeleek de veiligheid en effectiviteit van de bolus van 80 mg met die van de standaard behandeling. Het primaire eindpunt was de sterfte na 30 dagen. De sterfte na de bolus of na standaard behandeling was vergelijkbaar (6,0% vs 5,9%), in beide groepen traden evenveel hersenbloedingen op, er waren iets meer reinfarkten na de bolus.
Conclusies.
Standaard behandeling met saruplase verstoort de hémostase minder dan streptokinase, even veel als urokinase, en meer dan alteplase. Toediening van een bolus van 80 mg saruplase veroorzaakt een snelle defibrinogenatie, zonder dat dit aanleiding geeft tot meer bloedingen. Na saruplase gaan de bloedvaten sneller open dan na streptokinase, en
161 Chapter 10 ongeveer even snel als na alteplase. De sterfte na 30 dagen is tenminste vergelijkbaar met die streptokinase, en er zijn een verschillen tussen standaard behandeling en bolus toediening. Saruplase veroorzaakt meer hersenbloedingen dan streptokinase. Het aantal hersenbloedingen na saruplase of alteplase verschilt niet. Bolus toediening en standaard behandeling veroorzaken evenveel hersenbloedingen. Er zijn geen antilichamen tegen saruplase of tegen E. coli eiwit aangetoond, en er waren geen allergische reacties na een eerste of tweede behandeling met saruplase.
162 Dankbetuiging.
Vanaf het PRIMI onderzoek was de afdeling cardiologie van het Catharina Ziekenhuis betrokken bij het klinische onderzoek met saruplase. Het PRIMI onderzoek zelf is geen apart deel van dit proefschrift, omdat destijds terecht aan Fekry El Deep de eer van het co-auteurschap werd gegund. Hij verwerkte dit onderzoek in zijn fraai in leer gebonden proefschrift, dat hij te Cairo verdedigde. In dit proefschrift bombardeert hij mij tot professor, wellicht stimuleerde dit mijn klinisch-wetenschappelijke ambitie. De samenwerking met de afdeling Klinisch Onderzoek van de firma Grünenthal was een bijzondere ervaring; ik ben aan Hannes Barth, Gwyn Hopkins, Michael von Fisenne, Robert Groves, Wolfgang Günzler en alle overige medewerkers dank verschuldigd voor de correcte en plezierige samenwerking. Gwyn Hopkins schaafde mijn Engels bij. Voor het hoofdstuk over de farmacokinetiek maakte ik dankbaar gebruik van de expertise van Wolfgang Günzler. De directie van het Catharina ziekenhuis ben ik dankbaar voor hun positieve houding ten aanzien van patiëntgebonden klinisch onderzoek. De verpleegkundigen van de afdeling cardiologie ben ik dank verschuldigd voor hun loyale medewerking, in het bijzonder dank aan het hoofd van de CCU, Rob Storm van Leeuwen en de beide "research nurses" Harold Helmes en Peter Coppes, die steeds de soms ingewikkelde onderzoeksprotocollen feilloos vertaalden naar de gevraagde verpleegkundige inspanning. Vaak moesten de patiënten op ongelegen ogenblikken een hartcatherisatie ondergaan, het werken op de hartcatherisatiekamer (HCK) van het Catharina ziekenhuis is echter altijd, ook bij nacht en ontij, een hoogtepunt in het klinische bestaan. Geeft hun koffiekamer ten onrechte de indruk van een eenzijdige belangstelling, de "Hall of Fame" op weg naar de HCK weerspiegelt het veelzijdige talent van de ploeg van onze chef dr. Berry van Gelder. Dank aan het Klinisch Chemisch Laboratorium voor de goede samenwerking, Hans Hoffmann is op het gebied van hémostase even deskundig als bescheiden. Ad Loomans werkte onverstoorbaar door ondanks de lawine van cardiologisch onderzoek. Dank voor de plaatjes aan Guy van Dael, die het werk, dat zijn vader begon als hoofd van de Audiovisuele Dienst van het Catharina Ziekenhuis, schitterend heeft uitgebouwd.
163 Dank aan de assistenten cardiologie, die de patiënten vaak en correct includeerden. Onze eigen onderzoeksafdeling CATHREINE groeide mee met het onderzoek naar saruplase, dank aan Antoinette, Hanny, Ingrid, Pétrie, Linda, en Kristel voor de deskundige en toegewijde administratieve ondersteuning en voor de plezierige sfeer. Hooggeleerde Verheugt, beste Freek, dat ik uiteindelijk dan toch promoveer, dank ik aan jouw niet aflatende aansporingen. Het moment, dat jij de CCU van het thoraxcentrum van het Dijkzigt ziekenhuis Rotterdam binnenstapte was het begin van een hartelijke en vrolijke vrienschap. Ik prijs mij gelukkig met onze samenwerking in het bestuur van de Nederlandse Vereniging voor Cardiologie. Hooggeleerde Bär, beste Frits, het welslagen van het klinische onderzoek naar saruplase is grootdeels te danken aan jouw bezielende leiding. Nu dit onderzoek afgerond is, zie ik uit naar voortzetting van de vruchtbare samenwerking tussen onze beide klinieken. Op belangrijke momomenten zoekt men steun van oudere, wijzere maten. Mijn paranimfen Dorus Relik en Hans Bonnier gaan mij voor in anciënniteit. Hun tegengesteld voorkomen en karakter weerspiegelen de gefacetteerdheid, die soms onze maatschap even doet flonkeren als een goedgeslepen edelsteen. Piet Borsje en Mamdouh El Gamal mogen trots zijn op wat zij mede opbouwden. Aangevoerd door Jacques Kooolen en Nico Pij Is leiden thans Dorus, Hans, Frank Bracke, Kathinka Peels, Albert Meijer en Cees-Joost Botman assistenten op, die de leermeesters stimuleren en als het goed is zullen overtreffen. Terwijl ik schreef, werkten mijn maten, dank.
Velen werkten mee aan het onderzoek met saruplase. Het is een voorrecht dat ik, schakeltje in een ketting, radertje in een goed geoliede machine, van de inspanningen van anderen gebruik mag maken voor dit proefschrift. Dit proefschrift is opgedragen aan de belangrijkste vrouwen in mijn leven: mijn moeder, vrouw en dochters: verleden, heden, toekomst. Oorsprong, inspiratie en doel. Dat mijn vader, die mij andere dan materiële waarden en normen meegaf en zelf geen tijd nam voor een promotie, deze dag niet meemaakt, is jammer. Ik had meer mijn best moeten doen.
164 Curriculum vitae.
Rolf Michels was bom on 7 august 1945 in Paramaribo, Suriname, where he attended elementary school (Juliana school) and secondary school (Hendrik school). In 1960 a stricter discipline was needed and found at the Protestants Christelijk Jongens Internaat and the Christelijke H.B.S. in Culemborg, where he graduated in 1965 (H.B.S.-B). In 1965 he started the study of medicine at the Rijks Universiteit Groningen. In 1972 he returned to his motherland as a junior doctor, to follow internships in surgery (St. Vincentius Hospital Paramaribo, dr. F. Tjong Ayong), obstetrics and gynaecology (St. Vincentius Hospital, dr. P. Favery) and industrial medicine (Suralco, Moengo, dr. I Guicherit). May 30, 1973 he was certificated as medical doctor, and that same year he started as resident in the department of internal medicine at the St Elisabeth Hospital Amersfoort (dr. D. Bonte). In 1974 he started his training at the Thorax Center Rotterdam (Prof. P.G. Hugenholtz). November 1, 1978 he was certificated as cardiologist. From 1978 to 1980 he held the position of director of Coronary Care at the Thorax Center. In 1980 he accepted the invitation to join the cardiovascular group at the Catharina Hospital Eindhoven, where two years before Hans Bonnier, also from the Thorax Center Rotterdam, had given new impetus to the restart of cardiopulmonary surgery. From 1983 to 1986 he served as chairman of the Medical Staffai the Catharina Hospital. He was a member of the Board of the Netherlands Society of Cardiology (NVVC) from 1987 to 1993, where he served as president from 1990 to 1993. During this latter episode he served qualitate qua as member of the Board of the Netherlands Heart Foundation (NHS). He was one of the founders of the Netherlands Institute for Continuing Cardiovascular Education (Cardiovasculair Onderwijs Instituut, CVOI), where he served as chairman of the Board from 1993 to 1996. Currently he is chairman of the Quality Committee of the NVVC. He is member of the Board of the Cardiac Therapy Research Institute Eindhoven, CATHREINE. He is (co-)author of several publications on cardiogenic shock, intra-aortic balloon pumping, unstable angina, thrombolysis and percutaneous transluminal coronary angioplasty (PTCA). He is a fellow of the European Society of Cardiology. Rolf Michels is married with Renske Reijnders, of Zeist. They have four daughters, Michelle, Jeanne-Marie, Jacqueline and Danielle.
165 166 го го Ш m —ι Ν Ν N —ι ω О) СЛ ι-*- О СО ш
О 5 Э к о — —h о 3 3 Ъ i Φ о ß° с 5 S о со (D Q)
=3 II II II
3 5 φ с oo с Φ Q. co 3 13 СО О φ 3 ω TD
Fibrinolysis with recombinant single-chain
urokinase-type plasminogen activator
(saruplase) in acute myocardial infarction.
Rolf Michels Legend to the picture on the cover:
Primary structure of saruplase, one of the largest and most complex proteins ever produced in recombinant bacteria. The molecule is a single peptide chain of 411 amino acids which represents a molecular weight of 46 344 daltons. Circles symbolize single amino acids, which are assigned by single letter code. Bars between cysteines (C ) indicate disulphide bridges. Asteriks mark the amino acids representing the active side of the serine proteinase. Arrows indicate proteolytic cleavage sites. At Lys , conversion into active two-chain high molecular weight (HMW tcu-PA) derivative, at Lys l35, conversion into low molecular weight (LMW tcu- PA) derivative. Domains are marked by parentheses. EGF = epidermal growth factor-like domain.
(W.A.Günzïer. Rev ContempPharmacother 1998; 9: 355-362).
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