毕赤酵母表达去氨普酶及其突变体的溶栓活性研究

中国临床药理学与治疗学 ›› 2010, Vol. 15 ›› Issue (6): 645-650.

毕赤酵母表达去氨普酶及其突变体的溶栓活性研究

严守升, 宫新江, 郭永起, 李剑凤, 武国栋, 王庆民, 孙丽霞, 杨清敏, 王晶翼

齐鲁制药有限公司药物研究院,济南 250100,山东

收稿日期:2010-03-22 修回日期:2010-05-27 出版日期:2010-06-26 发布日期:2020-09-16
通讯作者: 王晶翼,男,博士,教授,主要从事创新药物研发工作。 Tel: 0531-83126988 E-mail: jingyi.wang@qilu-pharma.com
作者简介:严守升,男,博士,主要从事新药临床前药理研究工作。Tel: 0531-83126946 E-mail: shousheng.yan@qilu-pharma.com

Thrombolysis activities of desmoteplase and mutants expressed in yeast Pichia methanolica

YAN Shou-sheng, GONG Xin-jiang, GUO Yong-qi, LI Jian-feng, WU Guo-dong, WANG Qing-min, SUN Li-xia, YANG Qing-min, WANG Jing-yi

Research and Development Department, Qilu Pharmaceutical Co., Ltd., Jinan 250100,Shandong, China

Received:2010-03-22 Revised:2010-05-27 Online:2010-06-26 Published:2020-09-16

摘要/Abstract

摘要： 目的: 研究毕赤酵母表达野生型去氨普酶(DSPA^wt)及其突变体F195和QNRR的溶栓活性,为开发新型溶栓类药物奠定基础。方法: 以阿替普酶(rt-PA)和中国仓鼠卵巢细胞表达野生型DSPAα1为阳性对照,采用大鼠动-静脉旁路血栓模型,通过对血栓重量、出血时间、大鼠凝血指标凝血酶时间(TT)、凝血酶原时间(PT)、活化部分凝血酶原时间(APTT)和纤维蛋白降解产物D-二聚体的测定考察毕赤酵母表达DSPA^wt及其两种突变体的溶栓活性。结果: DSPA^wt及其突变体F195和QNRR在 0.7 mg/kg 和 1.5 mg/kg 剂量下均有一定的溶栓活性,其中 DSPA^wt溶栓作用较弱,有效动物数少,溶栓率较低,相同剂量突变体F195和QNRR溶栓作用显著增强,溶栓效果与DSPAα1接近。此外,不同剂量各样品均能在一定程度上延长大鼠出血时间,延长大鼠血浆TT、PT和APTT,提高血浆中D-二聚体含量。结论: 通过酵母表达系统有望获得具有高溶栓活性的突变型DSPA,从而开发成新一代溶栓类药物。

关键词: 去氨普酶, 溶栓活性, 动-静脉旁路血栓模型, 纤溶酶原激活剂

Abstract: AIM: To study the thrombolysis activities of wide type Desmodus rotundus Salivary Plasminogen Activivator (DSPAwt) and its mutations F195 and QNRR expressed in yeast Pichia methanolica. METHODS: The rat model of artery-vein bypass thrombosis was used to measure the thrombolysis activities of DSPA^wt, F195 and QNRR, in which indexes such as weight of thrombus, bleeding time (BT), thrombin time (TT), prothrombin time (PT), activated partial thromboplastin time (APTT) and D-dimer were investigated. DSPA was expressed in CHO cells (DSPAαl) and the launched thrombolytic drug rt-PA were used as positive controls. RESULTS: Positive controls as well as DSPA^wt, F195 and QNRR at doses of 0.7 mg/kg and 1.5 mg/kg individually showed thrombolysis potency, which decreased the weight of thrombus, delayed the levels of BT, TT, PT, APTT and elevated the level of D-dimer. F195, QNRR and DSPAαl had the similar higher activities, while DSPA^wt possessed limited thrombolysis effects. CONCLUSION: The results suggest a novel method to express DSPAs for further development as a novel type of drug for treatment of thrombotic diseases such as acute ischemic stoke.

Key words: Desmodus rotundus, Thrombolysis activity, Model of artery-vein bypass thrombosis, Plasminogen activivator

中图分类号:

R965.2

严守升, 宫新江, 郭永起, 李剑凤, 武国栋, 王庆民, 孙丽霞, 杨清敏, 王晶翼. 毕赤酵母表达去氨普酶及其突变体的溶栓活性研究[J]. 中国临床药理学与治疗学, 2010, 15(6): 645-650.

YAN Shou-sheng, GONG Xin-jiang, GUO Yong-qi, LI Jian-feng, WU Guo-dong, WANG Qing-min, SUN Li-xia, YANG Qing-min, WANG Jing-yi. Thrombolysis activities of desmoteplase and mutants expressed in yeast Pichia methanolica[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2010, 15(6): 645-650.

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