Research

Experimental & Translational Stroke Medicine

, 4:10

Open Access This content is freely available online to anyone, anywhere at any time.

Safety evaluation of a recombinant plasmin derivative lacking kringles 2-5 and rt-PA in a rat model of transient ischemic stroke

  • R Christian CrumrineAffiliated withResearch and Pre-clinical Development, Grifols Therapeutics, Inc Email author 
  • , Victor J MarderAffiliated withDivision of Hematology/Medical Oncology, Department of Medicine, David Geffen School of Medicine at UCLA
  • , G McLeod TaylorAffiliated withResearch and Pre-clinical Development, Grifols Therapeutics, Inc
  • , Joseph C LaMannaAffiliated withDepartment of Physiology and Biophysics, Case Western Reserve University Email author 
  • , Constantinos P TsipisAffiliated withDepartment of Physiology and Biophysics, Case Western Reserve University
  • , Valery NovokhatnyAffiliated withResearch and Pre-clinical Development, Grifols Therapeutics, Inc Email author 
  • , Philip ScuderiAffiliated withResearch and Pre-clinical Development, Grifols Therapeutics, Inc Email author 
  • , Stephen R PettewayJrAffiliated withResearch and Pre-clinical Development, Grifols Therapeutics, Inc Email author 
  • , Vikram AroraAffiliated withResearch and Pre-clinical Development, Grifols Therapeutics, Inc Email author 

Abstract

Background

Tissue type plasminogen activator is the only approved thrombolytic agent for the treatment of ischemic stroke. However, it carries the disadvantage of a 10-fold increase in symptomatic and asymptomatic intracranial hemorrhage. A safer thrombolytic agent may improve patient prognosis and increase patient participation in thrombolytic treatment. A novel direct-acting thrombolytic agent, Δ(K2-K5) plasmin, promising an improved safety profile was examined for safety in the snare ligature model of stroke in the rat.

Methods

Male spontaneously hypertensive rats were subjected to 6 hours middle cerebral artery occlusion followed by 18 hours reflow. Beginning 1 minute before reflow, they were dosed with saline, vehicle, Δ(K2-K5) plasmin (0.15, 0.5, 1.5, and 5 mg/kg) or recombinant tissue-type plasminogen activator (10 and 30 mg/kg) by local intra-arterial infusion lasting 10 to 60 minutes. The rats were assessed for bleeding score, infarct volume, modified Bederson score and general behavioral score. In a parallel study, temporal progression of infarct volume was determined. In an in vitro study, whole blood clots from humans, canines and rats were exposed to Δ(K2-K5). Clot lysis was monitored by absorbance at 280 nm.

Results

The main focus of this study was intracranial hemorrhage safety. Δ(K2-K5) plasmin treatment at the highest dose caused no more intracranial hemorrhage than the lowest dose of recombinant tissue type plasminogen activator, but showed at least a 5-fold superior safety margin. Secondary results include: temporal infarct volume progression shows that the greatest expansion of infarct volume occurs within 2–3 hours of middle cerebral artery occlusion in the spontaneously hypertensive rat. A spike in infarct volume was observed at 6 hours ischemia with reflow. Δ(K2-K5) plasmin tended to reduce infarct volume and improve behavior compared to controls. In vitro data suggests that Δ(K2-K5) plasmin is equally effective at lysing clots from humans, canines and rats.

Conclusions

The superior intracranial hemorrhage safety profile of the direct-acting thrombolytic Δ(K2-K5) plasmin compared with recombinant tissue type plasminogen activator makes this agent a good candidate for clinical evaluation in the treatment of acute ischemic stroke.

Keywords

Ischemic stroke Δ(K2-K5) plasmin Intracranial hemorrhage Spontaneously hypertensive rat model Recombinant tissue-type plasminogen activator (rt-PA) Middle cerebral artery occlusion (MCAo)