Thrombomodulin-Dependent Protein C Activator Treatment Improves the Short-Term Outcome of Experimental Ischemic Stroke in Mice.

High dose human or mouse activated protein C (APC) treatment is neuroprotective in the transient intraluminal middle cerebral artery occlusion (MCAO) and reperfusion model of ischemic stroke, in mice. However, the therapeutic potential of systemic APC treatment in stroke might be limited because it can disable hemostasis. Low dose thrombomodulin-dependent protein C activator (PCA) enzyme treatment can generate antithrombotic APC on the intraluminal surface without profound systemic anticoagulation and antihemostatic effects in primates. We thus explored whether PCA treatment could produce short term outcome benefits in comparison to fibrinolysis with tPA in the mouse stroke model. Stroke was induced by MCAO between 0 and 60 min using an intraluminal filament that causes progressive and occlusive secondary thrombosis in the MCA region. Cortical perfusion as a measure of thrombotic occlusion was monitored from start until 75 min, i.e., following removal of the intraluminal filament. Cerebral infarct size (% brain volume, TTC stain), edema (% brain volume; calculated), and neurological damage scores (scale 0 – 5) were assessed in survivors at sacrifice next day. All animals (N=12 to 16 in each treatment group) were evaluated for macroscopic hemorrhage (intracerebral or other sites) on autopsy after death. Treatments with vehicle (saline bolus), recombinant PCA thrombin analog W215A/E217A (0.025 mg/kg IV bolus) or plasminogen activator (2.5 mg/kg Bolus, [tPA-B], or 10 mg/kg Infusion for 45 min, [tPA-I]) were started 15 min after initiation of MCAO. One day mortality rates were similar: 29% in the PCA, 38% in the tPA-B, and 23% in the tPA-I treatment groups vs. 23% in vehicle controls (P>0.05 each). Cerebral infarct size was reduced by 62%, 43%, and 73% in the PCA bolus, tPA-B, and tPA-I treated groups, respectively (p<0.05 for each). Macroscopic hemorrhage (intracranial and/or other sites) was found in 15% (2/13), 14% (2/14), 44% (7/16), and 31% (4/13) in the vehicle, PCA, tPA-B, and tPA-I-treated groups, respectively. One day neurological scores averaged 1.5 in PCA-treated (p<0.05), 3.2 in tPA-B (NS), and 2.2 in tPA-I treated (p<0.05) versus 3.9 in saline vehicle-treated animals. All enzyme treatments significantly reduced secondary (thrombotic) vascular occlusions, as evidenced by 62% (PCA), 66% (tPA-B), and 72% (tPA-I) improvement of early cortical reperfusion vs. vehicle-treated controls (p<0.05 each). The means and SEM are illustrated in the figure below. The apparent efficacy and safety of intermediate dose PCA in this model suggests that PCA treatment might have therapeutic utility in acute ischemic stroke, e.g. when reperfusion of the MCA is achieved by tPA treatment or other modalities.

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