THROMBOLYSIS BY RAPID DIRECT-ACTING FIBRINOLYTIC AGENTS

快速直接作用纤溶剂的溶栓作用

• 批准号: 3342469
• 负责人: FRANCIS S MARKLAND
• 金额: $ 14.95万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-01-01 至 1986-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3342469
• 关键词: blood coagulation tests; blood tests; cardiovascular disorder diagnosis; disease /disorder model; drug adverse effect; drug metabolism; fibrinogen; fibrinolysis; fibrinolytic agents; gel electrophoresis; gel filtration chromatography; histology; human therapy evaluation; human tissue; ion exchange chromatography; plasminogen activator; reptile poison; thromboembolism

Thromboembolic vascular disease comprises a significant fraction of all morbidity and mortality in the United States. Thrombolytic therapy, using systemically or selectively administered plasminogen activators, has gained increasing acceptance for the treatment of thromboembolic disease. However, further improvements in thrombolytic therapy are still required. Our search for a more effective and safer agent, led us to investigate snake venoms. In 1956, several investigators characterized fibrinolytic properties of various snake venoms and mentioned that purified venom fractions may be clinically useful as thrombolytic agents. Studies have, therefore, been initiated in our laboratories to purify and characterize fibrinolytic snake venom enzymes. These investigations employ a new method for purifying fibrinolytic enzymes and a new in vivo angiographic model for evaluating their therapeutic effectiveness. We have purified a fibrinolytic enzyme from southern copperhead venom by combination of gel filtration on Sephadex G-100 and ion exchange chromatography on carboxymethylcellulose and diethylaminoethylcellulose. The enzyme proved to be homogeneous by polyacrylamide gel electrophoresis in sodium dodecylsulfate. The enzyme does not appear to activate plasminogen, rather it has a direct lytic effect on fibrin. In vivo activity will be assessed against fresh thromboemboli introduced into rabbit kidneys. Rabbit renal arteries will be cateterized and following control arteriograms, standard volumes of clot (rabbit or human), will be injected into each kidney of fully heparinized rabbits. The test fibrinolytic enzyme will be infused into one renal artery only, while the contralateral renal artery serves as control. Thrombolysis will be assessed arteriographically at intervals up to 5 hours. Once significant in vivo thrombolytic activity of the test agent has been demonstrated in the rabbit, adverse effects will be assessed. Acute and chronic renal toxicity will be evaluated arteriographically, histologically, and functionally, following infusion into non-embolized kidneys. Cardiovascular effects will be evaluated by EKG and manometry following intrarenal, and then intracoronary, infusion. Possible alterations of coagulation parameters (thrombin time, fibrinogen concentration, clotting time) will be determined. Acute and chronic neurotoxicity will be tested after intracarotid infusion. In this way, the therapeutic ratio of several enzyme preparations will be compared. Those enzymes with the highest therapeutic ratios will be further studied to characterize their biochemical properties and in vitro activity on human fibrinogen, fibrin and blood coagulation proteins. If these investigations confirm our preliminary in vivo findings that purified snake venom enzymes are potent and safe fibrinolytic agents, these enzymes may find widespread application in the treatment of human thromboembolic disease.

血栓栓塞性血管疾病占所有 美国的发病率和死亡率。 溶栓治疗，使用 全身性或选择性给予纤溶酶原激活剂，已经获得了 增加了对血栓栓塞性疾病治疗的接受度。 然而，溶栓治疗仍需进一步改进。 为了寻找更有效更安全的药剂，我们进行了调查 蛇毒 1956年，几位研究人员将纤溶酶 各种蛇毒的性质，并提到，纯化的毒液， 级分在临床上可用作血栓溶解剂。 研究表明， 因此，在我们的实验室中开始纯化和表征 蛇毒纤溶酶 这些调查采用了一种新的方法 用于纯化纤维蛋白溶解酶和一种新的用于 评估其治疗效果。 我们已经净化了 凝胶结合法分离南方铜斑蛇毒纤溶酶 Sephadex G-100柱层析， 羧甲基纤维素和二乙基氨基乙基纤维素。 这种酶证明了 通过聚丙烯酰胺凝胶电泳在钠中均匀 十二烷基硫酸盐。 这种酶似乎不激活纤溶酶原， 它对纤维蛋白具有直接溶解作用。 将评估体内活性 防止新鲜血栓栓塞进入兔肾。 兔肾 将对动脉进行分类，并遵循对照动脉造影，标准 将一定体积的凝块（家兔或人）注射到 完全肝素化的兔子。 将输注供试纤溶酶 仅进入一个肾动脉，而对侧肾动脉用作 控制 将在以下时间间隔进行动脉造影评估溶栓 到5小时。 一旦体内溶栓活性显著， 代理已被证明在兔子，不良反应将 评估。 将评价急性和慢性肾毒性 灌注后的动脉造影、组织学和功能 注入未栓塞的肾脏 将通过以下方式评价心血管效应： 肾内和冠状动脉内输注后的EKG和测压。 凝血参数（凝血酶时间、纤维蛋白原）的可能变化 浓度、凝血时间）。 急性和慢性 将在颈动脉内输注后测试神经毒性。 如此则 比较几种酶制剂的疗效。 那些 具有最高治疗比率的酶将被进一步研究， 表征它们的生物化学性质和对人的体外活性 纤维蛋白原、纤维蛋白和凝血蛋白。 如果这些调查 证实了我们初步的体内研究结果，纯化的蛇毒酶 是有效和安全的纤维蛋白溶解剂，这些酶可能会发现广泛的 在治疗人血栓栓塞性疾病中的应用。