Safety of plasmin in the setting of concomitant aspirin and heparin administration in an animal model of bleeding

Abstract

Summary. Plasmin is a direct thrombolytic which has been shown to have a strikingly favorable bene®t to risk pro®le in comparison with plasminogen activators, notably tissue plasminogen activator (t-PA). As heparin is known to increase the risk of hemorrhage when co-administered with a plasminogen activator, we asked whether adjunct antithrombotic agents such as aspirin and heparin would affect the safety of plasmin. Three groups of rabbits were administered plasmin at a dose (4 mg kg À1 ) designed to induce signi®cant decreases in antiplasmin, ®brinogen and factor (F)VIII, to about 25, 40 and 40%, respectively, of baseline values, but not cause prolongation of the ear puncture bleeding time. In a blinded and randomized trial, the results show that an intravenous aspirin bolus plus heparin administered as a bolus followed by a maintenance continuous infusion did not signi®cantly prolong the bleeding time during plasmin infusion. These data indicate that in the rabbit, concomitant use of aspirin plus heparin does not affect the safety of a therapeutic dose of plasmin.Keywords: aspirin, heparin, plasmin, safety.Locally delivered thrombolytic therapy is effective for hastening the lysis of deep vein thromboses [1,2] and thrombosed catheters [3] and for improving the clinical outcome of peripheral arterial graft occlusion [4] or ischemic stroke [5]. All agents that are currently approved and used widely for clinical thrombolysis are plasminogen activators (PA) [6], but their use is uniformly associated with an increased risk of hemorrhagic complications, even when delivered regionally to a thrombus [7,8]. We recently reported studies of the direct-acting proteolytic enzyme plasmin in animal models of thrombolysis and bleeding, and show a dramatic superiority in the safety of plasmin over TPA [9,10], without sacri®ce of thrombolytic ef®cacy [11].The combined use of heparin or hirudin anticoagulant therapy with PA increases the risk of bleeding [12±15]. Thus, the rate of intracranial hemorrhage in patients with peripheral arterial occlusion treated with regional recombinant urokinase was 0.5% (1/210), compared with a rate of 4.8% (3/62) when heparin was co-administered with the PA [15]. Since our prior studies of plasmin were performed without concomitant anticoagulant administration, the experimental conditions may not have re¯ected actual clinical conditions in which heparin, aspirin and/or other antithrombotic therapies are utilized with PA. Thus, the study reported here tests whether the hemostatic safety of plasmin [9,10] is disrupted by the concomitant administration of aspirin or by aspirin plus heparin. MethodsHuman plasmin (BAY 57±9602, Lot TPM03 bulk) was puri®ed by SK activation of human blood plasma-derived plasminogen. The starting material for plasminogen puri®cation was the sodium caprylate precipitate from an intravenous immunoglobulin process [16]. Plasminogen was puri®ed by adsorption on lysine-Sepharose FF (Amersham Pharmacia, Uppsala, Sweden) and eluted with epsilon-aminocaproic acid. The resulti...

Direct fibrinolytic agents: biochemical attributes, preclinical foundation and clinical potential

“…Intravenous aspirin and heparin administered in addition to plasmin (4 mg kg )1 ) did not prolong the primary bleeding time [56]. These results translate to a significant margin of hemostatic safety for plasmin as opposed to t-PA [57].…”

“…However, prolongation of the bleeding time did not occur until complete depletion of fibrinogen and FVIII was induced at 8 mg kg )1 , which is three-fold to four-fold more than is needed for thrombolysis. Intravenous aspirin and heparin administered in addition to plasmin (4 mg kg )1 ) did not prolong the primary bleeding time [56]. These results translate to a significant margin of hemostatic safety for plasmin as opposed to t-PA [57].…”

“…A phase I/II dose selection and safety trial is underway in patients with recent (category I or IIa) lower extremity ischemia [71 (NCT 00418483)] caused by non-embolic thrombosis (> 10 cm in length) of an infrainguinal graft or native In vitro retracted clots: greater lytic activity than t-PA after catheter injection [39] Distal abdominal aorta thrombosis (rabbit): equivalent to t-PA with unimpeded blood flow in aorta, superior in thrombolysis and flow restoration with impeded flow (limited plasminogen supply) [16] Arteriovenous graft thrombosis (porcine): superior thrombolysis than with t-PA [38] Middle cerebral artery thrombo-occlusion: effective thrombolysis [54] Ear-puncture bleeding model (rabbit): hemostatic safety at suprathrombolytic dosages, as compared with bleeding caused by t-PA at any therapeutic dosage [16,55,57]. No effect with systemic heparin aspirin [56] Mini-plasmin Femoral artery thrombosis (canine): greater efficacy than t-PA [58] Middle cerebral artery ligation (mouse, hamster): reduced ischemic injury after systemic administration [59] Micro-plasmin Extracorporeal loop thrombus (rabbit): equivalent efficacy to plasmin, slightly less than with t-PA; bleeding with t-PA only [49] Cerebral ischemia (rabbit, mouse, rat): systemic (not local administration) shows less intracranial hemorrhage than caused by t-PA, decreased cerebral injury as good or better than with t-PA, but question of adverse outcomes with higher dosages [59][60][61][62][63][64] Vitreolysis (cat, rabbit, pig): vitreous detachment induced by local injection [65][66][67]…”

Cross-Species Pharmacologic Evaluation of Plasmin as a Direct-Acting Thrombolytic Agent: Ex Vivo Evaluation for Large Animal Model Development

“…Plasmin, derived from fractionated human plasma, has been shown to be efficacious and safe in preclinical animal models of thrombosis and recurrent bleeding models (1,4,13). In addition, plasmin, the active fibrinolytic enzyme responsible for fibrin clot degradation, would be a unique agent for regional thrombolysis in the treatment of thrombosis because it bypasses the need for plasminogen that all plasminogen activators have for thrombolytic activity.…”

The evolution of recombinant thrombolytics: Current status and future directions

“…140 Several studies conducted in animal models indicate the effectiveness and safety of plasmin administration for thrombolytic therapy. 141,142 In a phase I human trial, 24 mg plasmin (Talecris Biotherapeutics, Inc.) effectively induced thrombolysis of hemodialysis graft occlusions. 143 Several plasmin derivatives have also been created for application as direct thrombolytics.…”