Development of the Plasmin-independent Thrombolytic Agent for the Treatment of Acute Ischemic Stroke

项目概要 由血管内血凝块引起的血栓形成是这三种疾病中最常见的潜在病理 主要心血管疾病：缺血性心脏病、中风和静脉血栓栓塞。显着地 高发病率及其致命后果迫切需要寻找安全有效的溶栓药物 酶，在过去的几十年里仍然广泛存在。尽管付出了努力，目前 溶栓剂有显着的缺点，包括出血并发症。目前所有溶栓药物 参与纤溶酶原活化为纤溶酶。不幸的是，非特异性蛋白水解活性 纤溶酶降解参与止血和伤口愈合的必需蛋白质。因此，纤溶酶的临床应用 依赖溶栓治疗受到限制，需要在中风后 3 小时内开始治疗 避免致命的出血并发症。仍然迫切需要新的溶栓剂来改善 治疗无需担心出血和组织损伤并发症。 SNJ Pharma, Inc. 鉴定出出现的 HtrA1（高温要求 A1，命名为凝血酶） 对血管内凝块发挥强大的溶栓活性，同时允许伤口正常愈合。与所有不同 目前可用的溶栓剂中，HtrA1 蛋白酶不会将纤溶酶原激活为纤溶酶，纤溶酶是一种非特异性物质。 蛋白酶负责溶栓治疗中致命的出血并发症。 HtrA1 的高特异性 酶活性能够选择性降解病理血栓，同时保留止血凝块， 消除大出血的风险。体内小鼠模型，包括伤口切除和尾部出血测定， 证明用 HtrA1 治疗的动物与目前的治疗相比没有表现出伤口愈合损伤 溶栓剂。此外，给予HtrA1可完全溶解尾部血栓小鼠的血栓 静脉注射时，肺栓塞小鼠体内会出现血管内血栓，从而导致 抢救了血栓栓塞。在此，在确认 HtrA1 活性的特异性并保留有益的 止血凝块，我们的目的是评估 HtrA1 蛋白酶的治疗效果及其治疗血栓的潜力 使用体外血栓切除模型和体内光血栓 AIS 治疗急性缺血性中风 (AIS) 模型。我们使用第一种纤溶酶非依赖性溶栓酶成功完成该提案将 提出 HtrA1 蛋白酶作为 AIS 的革命性治疗候选物，且不存在局限性有限的缺点 没有出血和组织损伤并发症的治疗窗口。

Project Summary Thrombosis caused by intravascular blood clots is the most common underlying pathology of the three major cardiovascular disorders: ischemic heart disease, stroke, and venous thromboembolism. The significantly high incidence rate, as well as its fatal consequences, has called the search for safe and effective thrombolytic enzymes, which remained extensive throughout the past several decades. Despite the effort, current thrombolytics have significant drawbacks, including bleeding complications. All the current thrombolytic agents are involved in the activation of plasminogen to plasmin. Unfortunately, the non-specific proteolytic activity of plasmin degrades essential proteins involved in hemostasis and wound healing. Thus, the clinical use of plasmin- dependent thrombolytic therapy has been limited, requiring the initiation of treatment within 3 hours from stroke onset to avoid fatal hemorrhagic complications. A need for new thrombolytics remains urgent for improved treatment without concerns of hemorrhage and tissue injury complications. SNJ Pharma, Inc. identified HtrA1 (high-temperature requirement A1, named as thrombase) that appears to exert strong thrombolytic activity on intravascular clots while permitting normal wound healing. Unlike all currently available thrombolytics, HtrA1 protease does not activate plasminogen to plasmin, a non-specific protease responsible for fatal hemorrhagic complications from thrombolytic therapy. The high specificity of HtrA1 enzyme activity enables selective degradation of pathological thrombi while preserving hemostatic clots, removing the risk of massive bleeding. In vivo murine models, including wound excision and tail bleeding assays, demonstrated that animals treated with HtrA1 exhibited no damage in wound healing in contrast to current thrombolytics. In addition, HtrA1 administration completely dissolved blood thrombi in mice with tail thrombosis and intravascular thrombi in mice with pulmonary embolism when administrated intravenously, resulting in rescued thromboembolism. Here, upon confirming the specificity of HtrA1 activity and preservation of beneficial hemostatic clots, we aim to evaluate the therapeutic efficacies of HtrA1 protease and its potential for the treatment of acute ischemic stroke (AIS) using in vitro thrombectomy model and in vivo photothrombotic AIS model. Our successful completion of this proposal with the first plasmin-independent thrombolytic enzyme will present HtrA1 protease as a revolutionary therapeutic candidate for AIS without the drawback of a limited treatment window in the absence of hemorrhage and tissue injury complications.