SMTP-7, a novel small-molecule thrombolytic for ischemic stroke: a study in rodents and primates - PubMed (original) (raw)

SMTP-7, a novel small-molecule thrombolytic for ischemic stroke: a study in rodents and primates

Hironobu Sawada et al. J Cereb Blood Flow Metab. 2014 Feb.

Abstract

SMTP-7 (Stachybotrys microspora triprenyl phenol-7), a small molecule that promotes plasminogen activation through the modulation of plasminogen conformation, has excellent therapeutic activity against cerebral infarction in several rodent models. Detailed evaluations of SMTP-7 in a primate stroke model are needed for effective, safe drug development. Here we evaluated SMTP-7 in a monkey photochemical-induced thrombotic middle cerebral artery (MCA) occlusion model (n=6), in which MCA occlusion was followed by recanalization/reocclusion. SMTP-7 (10 mg/kg, intravenous infusion) significantly increased the postinfusion MCA recanalization rate (32.5-fold, P=0.043) and ameliorated the post-24-h neurologic deficit (by 29%, P=0.02), cerebral infarct (by 46%, P=0.033), and cerebral hemorrhage (by 51%, P=0.013) compared with the vehicle control animals. In normal monkeys, SMTP-7 did not affect general physiologic or hemostatic variables, including coagulation and platelet parameters. Investigations in rodent models of transient and permanent focal cerebral ischemia, as well as arterial thrombosis and bleeding tests, suggest a role for SMTP-7's regulated profibrinolytic action and neuroprotective properties in the monkey MCA occlusion model. In conclusion, SMTP-7 is effective in treating thrombotic stroke in monkeys. SMTP-7 is thus a promising candidate for the development of alternative therapy for ischemic stroke.

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Figures

Figure 1

Figure 1

Effects of SMTP-7 (Stachybotrys microspora triprenyl phenol-7) on blood flow and neurologic deficits in a monkey thrombotic middle cerebral artery (MCA) occlusion model. (A) Blood flow profile after photochemical induction of thrombotic MCA occlusion. The experimental design and patency status of the MCA blood flow in individual animals are shown. The complete stoppage of the blood flow was regarded as occlusion and is indicated as a filled bar, and reperfusion is represented by an open bar. (B) Time for initial occlusion. (C) Postinfusion recanalization time. The sum of the duration of recanalization during the 30-minute postinfusion period is shown. (D) Neurologic deficit scores. The total of the scores was 100, comprising consciousness (range 0 to 28), sensory system (range 0 to 22), motor system (range 0 to 32), and skeletal muscle coordination (range 0 to 18). _P_-values by unpaired Student's _t_-test are shown. There were six animals in each group.

Figure 2

Figure 2

Effects of SMTP-7 (Stachybotrys microspora triprenyl phenol-7) on cerebral infarction and hemorrhage in a monkey thrombotic middle cerebral artery (MCA) occlusion model. (A) Cerebral infarction. Images are representative sections stained with TTC (2,3,5-triphenyltetrazolium chloride). Infarct areas are marked. (B) The quantification of infarction volume corrected for edema. The degree of edema was as follows: control, 25.5±2.4% SMTP-7, 20.5±2.1%. The difference was significant (_P_=0.005 by unpaired Student's _t_-test). (C) Hemorrhagic area. Images are representative nonstained sections. Arrows indicate the hemorrhage area, which is marked by a dashed circle. (D) The quantification of the hemorrhagic area. _P_-values by unpaired Student's _t_-test are shown. There were six animals in each group.

Figure 3

Figure 3

Effects of SMTP-7 (Stachybotrys microspora triprenyl phenol-7) on neurologic deficits and cerebral infarct in transient and permanent focal cerebral ischemia models in rats. (A and B) Neurologic deficit scores and infarction sizes corrected for edema in a transient focal ischemia model. The total of the neurologic deficit scores was 15, comprising forelimb flexion (range 0 to 3), hindlimb flexion (range 0 to 3), forelimb motor function (range 0 to 3), postural reaction after lateral pressure (range 0 to 3), and the animal's posture (range 0 to 3). The insets in A and B show the experimental design and representative TTC (2,3,5-triphenyltetrazolium chloride)-stained brain sections from vehicle-treated and SMTP-7 (10 mg/kg)-treated animals, respectively. *P<0.05 compared with vehicle control by Kruskal–Wallis test (A) or Dunnett test (B). The degree of edema was as follows: control, 15.2±3.3% 1 mg/kg, 11.4±4.4% 3 mg/kg, 12.2±3.8% 10 mg/kg, 9.8±3.0%. The difference between the control group and the 10 mg/kg group was significant (_P_=0.02 by Tukey's test). There were 10 animals in each group. (C and D) Neurologic deficit scores and infarction size corrected for edema in the permanent focal ischemia model. The definition of the neurologic deficit score is the same as that described for panel A. The inset in C shows the experimental design. There was no significant difference between the groups in neurologic score. *P<0.05 by unpaired Student's _t_-test for infarction size corrected for edema. The degree of edema was 4.2±11.3% for the control group and 12.7±6.3% for the SMTP-7 group (_P_=0.076 by unpaired Student's _t_-test). There were 10 animals in each group.

Figure 4

Figure 4

Effects of SMTP-7 (Stachybotrys microspora triprenyl phenol-7) on thrombotic carotid artery occlusion, bleeding, and plasmin formation in vivo. (A) Initial occlusion time in a rat carotid artery occlusion model. **P<0.01 versus vehicle control by Mann–Whitney U_-test; #P<0.05 versus control by Kruskal–Wallis test. (B) Bleeding time in normal mice. *P<0.05 versus control; **P<0.01 versus control; ##P<0.01 versus 100 mg/kg SMTP-7 group; ††_P<0.01 versus 1 mg/kg tissue-type plasminogen activator (t-PA) group by Tukey–Kramer test. There were five to eight animals in each group. (C) Plasmin–_α_2-antiplasmin complex (Pm–AP) levels in normal mice. **P<0.01 versus control by Tukey test. Insets in panels A to C show the experimental designs. There were five animals in each group. (D) In vitro analysis of the formation of Pm–AP and t-PA–plasminogen activator inhibitor-1 (PAI-1) in the presence of SMTP-7. Mouse plasma supplemented with _α_2-antiplasmin or PAI-1 was incubated with plasmin or t-PA in the presence of the indicated concentrations of SMTP-7 to assess the effects of SMTP-7 on the formation of Pm–AP and t-PA–PAI-1 in vitro. Data are representative of duplicate experiments. Positions of Pm–AP and t-PA–PAI-1 standards are shown. The extended gel images are shown in Supplementary Figure 5.

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