CN101668778B - 低出血抗凝血融合蛋白的制备及其应用 - Google Patents
低出血抗凝血融合蛋白的制备及其应用 Download PDFInfo
- Publication number
- CN101668778B CN101668778B CN2007800463406A CN200780046340A CN101668778B CN 101668778 B CN101668778 B CN 101668778B CN 2007800463406 A CN2007800463406 A CN 2007800463406A CN 200780046340 A CN200780046340 A CN 200780046340A CN 101668778 B CN101668778 B CN 101668778B
- Authority
- CN
- China
- Prior art keywords
- corecognition
- cracked
- oligopeptides
- thrombin
- hirudin
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
- C12N15/62—DNA sequences coding for fusion proteins
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P7/00—Drugs for disorders of the blood or the extracellular fluid
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P7/00—Drugs for disorders of the blood or the extracellular fluid
- A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P7/00—Drugs for disorders of the blood or the extracellular fluid
- A61P7/04—Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
- A61P9/10—Drugs for disorders of the cardiovascular system for treating ischaemic or atherosclerotic diseases, e.g. antianginal drugs, coronary vasodilators, drugs for myocardial infarction, retinopathy, cerebrovascula insufficiency, renal arteriosclerosis
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/46—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
- C07K14/47—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
- C07K14/4701—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals not used
- C07K14/4711—Alzheimer's disease; Amyloid plaque core protein
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/81—Protease inhibitors
- C07K14/8107—Endopeptidase (E.C. 3.4.21-99) inhibitors
- C07K14/811—Serine protease (E.C. 3.4.21) inhibitors
- C07K14/8121—Serpins
- C07K14/8128—Antithrombin III
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/81—Protease inhibitors
- C07K14/815—Protease inhibitors from leeches, e.g. hirudin, eglin
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/64—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue
- C12N9/6421—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue from mammals
- C12N9/6424—Serine endopeptidases (3.4.21)
- C12N9/6432—Coagulation factor Xa (3.4.21.6)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/64—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue
- C12N9/6421—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue from mammals
- C12N9/6424—Serine endopeptidases (3.4.21)
- C12N9/6443—Coagulation factor XIa (3.4.21.27)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y304/00—Hydrolases acting on peptide bonds, i.e. peptidases (3.4)
- C12Y304/21—Serine endopeptidases (3.4.21)
- C12Y304/21006—Coagulation factor Xa (3.4.21.6)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y304/00—Hydrolases acting on peptide bonds, i.e. peptidases (3.4)
- C12Y304/21—Serine endopeptidases (3.4.21)
- C12Y304/21027—Coagulation factor XIa (3.4.21.27)
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Genetics & Genomics (AREA)
- General Health & Medical Sciences (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Zoology (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Wood Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Biomedical Technology (AREA)
- General Engineering & Computer Science (AREA)
- Biophysics (AREA)
- Biotechnology (AREA)
- Gastroenterology & Hepatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Microbiology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Toxicology (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Hematology (AREA)
- Diabetes (AREA)
- Tropical Medicine & Parasitology (AREA)
- Neurology (AREA)
- Physics & Mathematics (AREA)
- Plant Pathology (AREA)
- Heart & Thoracic Surgery (AREA)
- Cardiology (AREA)
- Vascular Medicine (AREA)
- Urology & Nephrology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Peptides Or Proteins (AREA)
Abstract
提供了含有被凝血因子XIa和凝血因子Xa、或凝血酶和凝血因子Xa共同识别并裂解的寡肽的抗凝血融合蛋白。还提供了该抗凝血融合蛋白的制备方法和医药学用途。
Description
技术领域
本发明属于生物技术领域,涉及一类低出血抗凝血物质的结构和制备方法及其在防治血栓性疾病和血栓相关性疾病中的应用。具体而言,本发明涉及抗凝血物质与能被数个凝血因子(包括凝血酶、凝血因子Xa(FXa)、凝血因子XIa(FXIa)等)识别并裂解的氨基酸序列连接而成的新物质,以及这些新物质的制备和医药学用途。
背景技术
心脑血管疾病是近年来威胁人类健康和生命的头号杀手,血栓形成是许多心脑血管疾病的重要诱因,抗凝剂是防治血栓形成的重要药物。目前临床上广泛应用的抗凝剂主要是肝素,肝素的一个重要缺点是引发血小板减少症(thrombocytopenia)。现在又发展的低分子量肝素可以减轻肝素应用中的风险程度,但不能根本克服它的缺点。水蛭素是近年来欧洲已经上市的一个新型抗凝药物,它是凝血酶的直接抑制剂,但它对凝血酶的强抑制作用,可导致凝血相关参数,如出血时间、APTT、TT和PT等急剧升高,从而伴有全身或系统性出血风险。
现有抗凝药物存在的上述问题已经越来越多的受到人们的关注,作为一个理想的抗凝药物,在全身用药条件下,应当具有明确的抗凝抗栓效果,但又不引起出血副作用,使临床用药安全性提高。
为此,本发明思想的指导原则的关键是使抗凝剂的抗凝活性进行条件特异性释放,即在正常条件下,该类物质无抗凝活性,而当凝血系统被激活,有血栓形成可能或有血栓形成时,该类物质的抗凝活性才在该局部释放,营造不易形成血栓的微环境(即在局部出现抗凝活性),防止血栓形成或已有血栓的继续增长,甚至溶解已形成的微小栓子以达到防治血栓形成的功能。这就克服了肝素、水蛭素等抗凝剂用药引起系统性出血的风险。
例如水蛭素是由65~66个氨基酸组成的单链多肽。其氨基端可以与凝血酶催化活性位点结合,具有抗凝活性,羧基端与凝血酶的底物识别位点结合,对凝血酶有很强的特异性亲和作用。本文设计了一个封闭水蛭素氨基末端的措施,以达到暂时封闭水蛭素的抗凝活性的目的。当体内凝血系统被激活发生血栓时,利用血栓发生过程中引发的生物化学变化的特点,使氨基末端被封闭的水蛭素重新回复为水蛭素原形,在可能发生或已发生的血栓的局部发挥特异性抗凝作用,从而降低水蛭素引起系统性出血的危险,成为一类新型的、安全的、有效的抗凝剂。
在上述发明思想的指导下,本文对水蛭素进行了修饰。由于本实验室曾在水蛭素的氨基末端连接上可被凝血酶识别并裂解的寡肽(该水蛭素衍生物命名为TH)或凝血因子Xa识别并裂解的寡肽(该水蛭素衍生物命名为FH),以便一旦凝血系统被激活,则释放水蛭素的抗凝活性,起到抗凝和抗栓作用,且减少出血副作用。结果表明是有一定作用,但是效果还差。因此我们进而改变思路,考虑采取连接可被数个凝血因子或其它凝血因子识别并裂解的寡肽进行修饰,希望可以提高效果,达到具有实际应用的目的。当然这些寡肽在血栓发生时能否高效率地被相应的凝血因子识别并裂解,根据有关文献看来目前并无法推测。本实验室就试验制备了两个相关蛋白:①水蛭素的氨基末端连接上可被凝血酶和凝血因子Xa分别识别并裂解的寡肽,该水蛭素衍生物命名为GH;②水蛭素的氨基末端连接上可被凝血因子XIa和凝血因子Xa分别识别并裂解的寡肽,该水蛭素衍生物命名为EH。结果表明该两个水蛭素衍生物在体外和体内正常情况下不具有抗凝活性,一旦体内凝血系统被激活,则在以上凝血因子的共同作用下,可高效地局部释放水蛭素的抗凝活性,起到抗凝和抗栓作用。当然,在体内不存在血栓的部位,则无抗凝活性,故其出血副作用也显著降低。因此,它不同于水蛭素、肝素等抗凝剂,是一类安全、有效的抗凝抗栓剂。从而,此类具有低出血特征的抗凝剂对于防治血栓形成的应用具有重要意义。
发明目的
本发明的目的在于提供一类本身无抗凝活性,而当有血栓发生倾向或血栓发生时,可以在发生血栓局部释放其抗凝活性,从而防治血栓形成的物质。
发明内容
本发明现已发现,利用凝血因子XIa和凝血因子Xa分别识别的序列或凝血酶和凝血因子Xa分别识别的序列连接抗凝血物质可实现对此抗凝血物质抗凝活性的封闭,并在一定条件下裂解。衍生的抗凝血物质具有如下的特征:抗凝血物质如水蛭素的氨基末端被上述两个凝血因子分别识别的序列封闭后,在体外和血液系统非血栓部位没有抗凝活性,从而避免或减小了因抗凝血物质如水蛭素引起的系统出血的副作用;衍生的抗凝血物质只有在血栓发生时,在血栓部位特有的凝血因子的作用下才局部释放出游离的抗凝血物质,发挥防治血栓的作用,因而全身出血副作用明显降低。而且,由两个凝血因子分别识别的序列封闭抗凝血物质如水蛭素的氨基末端,在体内可以由两个凝血因子共同对其进行裂解,其效果大为优于由单个凝血因子的识别序列对抗凝血物质如水蛭素的氨基末端进行封闭。本发明基于上述特征的发现现已完成。
因此,本发明第一方面涉及含有被凝血因子XIa和凝血因子Xa共同识别并裂解的寡肽或被凝血酶和凝血因子Xa共同识别并裂解的寡肽的抗凝血物质。
本发明再一方面涉及含有被凝血因子XIa和凝血因子Xa共同识别并裂解的寡肽或凝血酶和凝血因子Xa共同识别并裂解的寡肽的抗凝血物质的制备方法,包括将凝血因子XIa和凝血因子Xa共同的识别序列或凝血酶和凝血因子Xa共同识别的序列所对应的碱基序列与抗凝血蛋白基因进行连接,然后这些基因构建至适合表达的载体,如pBV220,pPIC9,pPIC9K等,这些含有目的基因的重组载体在适宜的宿主中表达,例如在大肠杆菌,酵母或动物细胞体系中表达而得到新的抗凝血物质。
本发明再一方面涉及含有上述新的抗凝血物质及药用载体或赋形剂的药物组合物。
根据本发明,术语“抗凝物质”或“抗凝血物质”是指能够抗凝血的物质,如水蛭素、抗凝血酶III、蛇毒等,或它们的突变体。或其它具有抗凝作用的物质,优选水蛭素或其突变体。
根据本发明,术语“凝血因子XIa和凝血因子Xa共同识别的连接肽或凝血酶和凝血因子Xa共同识别的连接肽”或“凝血因子XIa和凝血因子Xa共同识别的寡肽或凝血酶和凝血因子Xa共同识别的寡肽”是指三肽序列EPR(GluProArg)或含有EPR的肽段,或五肽序列GVYAR(GlyValTyrAlaArg)或含有GVYAR的肽段。
根据本发明,本发明的含有被凝血因子XIa和凝血因子Xa共同识别并裂解的寡肽或凝血酶和凝血因子Xa共同识别并裂解的寡肽的抗凝血物质优选在水蛭素氨基末端连接上EPR或GVYAR。
根据本发明,本发明的含有被凝血因子XIa和凝血因子Xa共同识别并裂解的寡肽或凝血酶和凝血因子Xa共同识别并裂解的寡肽的抗凝血物质可在适宜的宿主系统中表达,优选在大肠杆菌或酵母中表达。
附图说明
图1表示EH和GH的核苷酸序列。
图2表示EH和GH的氨基酸序列。
下面的实施例用来进一步说明本发明,但并不意味着对发明的任何限制。
实施例EPR-HV2(EH)和GVYAR-HV2(GH)的制备及其防治血栓形成功能
一、EH蛋白和GH蛋白的制备
通过PCR将限制性酶切位点Xho I和凝血因子XIa和凝血因子Xa共同识别序列EPR或凝血酶和凝血因子Xa共同识别序列GVYAR对应的碱基序列引入水蛭素(HV2)基因的上游,在HV2基因下游引入EcoR I酶切位点,将该基因连接到相同酶切的pPIC9质粒上,得到重组质粒pPIC9-EH和pPIC9-GH。将pPIC9-EH和pPIC9-GH和pPIC9K用BamH I和Sal I双酶切后进行连接,得到pPIC9K-EH和pPIC9K-GH。将两个重组质粒电转化重组到酵母基因组,利用甲醇进行诱导表达。表达产物经分离纯化得到目的蛋白EH和GH。
二、EH和GH的生物活性
1.体外活性分析
GH、EH、FH和TH分别用凝血酶、FXa和FXIa进行裂解,裂解后采用纤维蛋白凝块法测定产物的抗凝活性。结果见表1。
表1 GH、EH、FH和TH被凝血酶、FXa和FXIa裂解前后抗凝活性分析
由表1可看出,GH可被凝血酶和FXa两种凝血因子分别识别并裂解,EH可以被FXa和FXIa两种凝血因子分别识别并裂解,而FH只被FXa识别并裂解,TH只被凝血酶识别并裂解。
2.GH和EH的抗栓作用和出血副作用分析
①GH和EH在大鼠颈总动脉血栓模型中的抗栓作用和出血副作用分析。结果见表2和表3。
表2 GH、FH、EH和TH对大鼠颈总动脉血栓形成时间的影响
#P<0.05,与NS组相比##P<0.01,与NS组相比
表3 EH、GH、FH和TH对大鼠颈总动脉血栓模型血浆TT的影响
NS-生理盐水
*P<0.05,与NS组相比;**P<0.01,与NS组相比
ΔP<0.01,HV组与其它各组相比
②GH、EH、TH和FH在小鼠割尾出血模型中的出血副作用分析。结果见表4。
表4 EH、GH、FH和TH对小鼠割尾出血时间的影响
*P<0.05,与NS组相比;ΔP<0.01,HV2组与其它各组相比;
#P<0.05,GH组与FH组和TH组相比
从表2可看出,EH、GH、FH和TH可延长大鼠颈总动脉血栓形成时间,且成剂量依赖关系,说明这些水蛭素衍生物都具有抗动脉血栓形成功能,而且彼此效果基本类似,但在出血副作用方面则大不相同,从表3和表4可看出,与HV2相比,GH、EH、FH和TH对出血指标的影响较小,说明其比HV2的安全性要高;而其中GH、EH比FH、TH对出血指标的影响更要小得多,特别是GH的出血时间更明显低于FH和TH组,说明GH、EH减低出血副作用方面要明显优于FH或TH。
Claims (5)
1.含有可被两种凝血因子共同识别并裂解的寡肽的抗凝血蛋白,其中,可被两种凝血因子共同识别并裂解的寡肽为能被凝血因子XIa和凝血因子Xa共同识别并裂解的氨基酸序列EPR,或为能被凝血酶和凝血因子Xa共同识别并裂解的氨基酸序列GVYAR,其中所述抗凝血蛋白是水蛭素,所述寡肽连接在水蛭素氨基末端。
2.权利要求1的含有可被两种凝血因子共同识别并裂解的寡肽的抗凝血蛋白,其是氨基酸序列如SEQ ID NO:3所示的抗凝血蛋白EH。
3.权利要求1的含有可被两种凝血因子共同识别并裂解的寡肽的抗凝血蛋白,其是氨基酸序列如SEQ ID NO:4所示的抗凝血蛋白GH。
4.制备权利要求1的含有可被两种凝血因子共同识别并裂解的寡肽的抗凝血蛋白的方法,其包括将编码EPR或GVYAR的碱基序列引入抗凝血蛋白基因的上游,然后这些基因通过适合的表达载体在大肠杆菌,酵母或动物细胞体系中表达。
5.包含权利要求1-3任一项的含有可被两种凝血因子共同识别并裂解的寡肽的抗凝血蛋白及药用载体或赋形剂的药物组合物。
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN2007800463406A CN101668778B (zh) | 2006-12-15 | 2007-12-11 | 低出血抗凝血融合蛋白的制备及其应用 |
Applications Claiming Priority (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN200610165756.7 | 2006-12-15 | ||
CN200610165756 | 2006-12-15 | ||
PCT/CN2007/003526 WO2008071081A1 (zh) | 2006-12-15 | 2007-12-11 | 低出血抗凝血融合蛋白的制备及其应用 |
CN2007800463406A CN101668778B (zh) | 2006-12-15 | 2007-12-11 | 低出血抗凝血融合蛋白的制备及其应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN101668778A CN101668778A (zh) | 2010-03-10 |
CN101668778B true CN101668778B (zh) | 2012-10-03 |
Family
ID=39511252
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN2007800463406A Active CN101668778B (zh) | 2006-12-15 | 2007-12-11 | 低出血抗凝血融合蛋白的制备及其应用 |
Country Status (6)
Country | Link |
---|---|
US (1) | US8101379B2 (zh) |
EP (1) | EP2103630B1 (zh) |
JP (1) | JP5345069B2 (zh) |
CN (1) | CN101668778B (zh) |
AT (1) | ATE533789T1 (zh) |
WO (1) | WO2008071081A1 (zh) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109206522A (zh) * | 2017-07-07 | 2019-01-15 | 北京三有利和泽生物科技有限公司 | 一种长效抗凝血融合蛋白及其应用 |
Families Citing this family (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
PT1737889E (pt) | 2004-10-19 | 2010-12-13 | Lonza Ag | Método para síntese de péptidos em fase sólida |
CN102180973B (zh) * | 2011-03-18 | 2012-08-29 | 重庆大学 | 靶向多功能防栓融合蛋白及其制备方法和应用 |
CN115572329B (zh) * | 2021-06-21 | 2024-02-06 | 王大勇 | 一组活性增强代谢较慢的菲牛蛭基因重组水蛭素及其制备方法 |
CN113956339B (zh) * | 2021-10-28 | 2023-02-24 | 中国药科大学 | 宽体金线蛭抗凝血因子XIa多肽及其应用 |
Family Cites Families (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5759542A (en) * | 1994-08-05 | 1998-06-02 | New England Deaconess Hospital Corporation | Compositions and methods for the delivery of drugs by platelets for the treatment of cardiovascular and other diseases |
AU713303B2 (en) * | 1995-06-12 | 1999-11-25 | Stichting Centraal Laboratorium Van De Bloedtransfusiedienst Van Het Nederlandse Rode Kruis | Factor IX binding peptides, derived from factor VIII and their use as inhibitors of blood coagulation |
US5910481A (en) * | 1995-11-13 | 1999-06-08 | Immuno Ag | Hybrid proteins with modified activity |
CN1480466A (zh) * | 2002-09-03 | 2004-03-10 | �й������ž�����ҽѧ��ѧԺ����ҽ | 一类溶栓抗凝双功能融合蛋白及应用 |
CN1896108B (zh) | 2005-06-01 | 2012-01-04 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | 特异性抗凝血物质的制备及其应用 |
-
2007
- 2007-12-11 WO PCT/CN2007/003526 patent/WO2008071081A1/zh active Application Filing
- 2007-12-11 CN CN2007800463406A patent/CN101668778B/zh active Active
- 2007-12-11 JP JP2009540580A patent/JP5345069B2/ja active Active
- 2007-12-11 US US12/519,309 patent/US8101379B2/en active Active
- 2007-12-11 AT AT07845883T patent/ATE533789T1/de active
- 2007-12-11 EP EP07845883A patent/EP2103630B1/en active Active
Non-Patent Citations (3)
Title |
---|
Richard J. Jenny,et al..A critical review of the methods for cleavage of fusion proteins with thrombin and factor Xa.《Protein Expression and Purification》.2003,第31卷第2页左栏第2段. * |
Susan M. Saporito-Irwin and William E. Van Nostrand.Coagulation Factor XIa Cleaves the RHDS Sequence and Abolishes the Cell Adhesive Properties of the Amyloid b-Protein.《THE JOURNAL OF BIOLOGICAL CHEMISTRY》.1995,第270卷(第44期),第26265-26269页,参见第26266页右栏第3段,26267页左栏第1段. * |
牛晋阳 等.新型水蛭素嵌合抗栓剂的构建表达与功能研究.《中国生物工程杂志》.2006,第26卷(第4期),摘要,第36页右栏第2行-第37页左栏第31行,第38页左栏最后1行-右栏第10行. * |
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN109206522A (zh) * | 2017-07-07 | 2019-01-15 | 北京三有利和泽生物科技有限公司 | 一种长效抗凝血融合蛋白及其应用 |
CN109206522B (zh) * | 2017-07-07 | 2021-11-09 | 北京三有利和泽生物科技有限公司 | 一种长效抗凝血融合蛋白及其应用 |
Also Published As
Publication number | Publication date |
---|---|
WO2008071081A8 (zh) | 2009-08-13 |
US20100113345A1 (en) | 2010-05-06 |
US8101379B2 (en) | 2012-01-24 |
EP2103630B1 (en) | 2011-11-16 |
EP2103630A1 (en) | 2009-09-23 |
EP2103630A4 (en) | 2010-07-28 |
WO2008071081A1 (zh) | 2008-06-19 |
CN101668778A (zh) | 2010-03-10 |
ATE533789T1 (de) | 2011-12-15 |
US20100234291A2 (en) | 2010-09-16 |
JP2010512349A (ja) | 2010-04-22 |
JP5345069B2 (ja) | 2013-11-20 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Stevens et al. | Protease–proteoglycan complexes of mouse and human mast cells and importance of their β‐tryptase–heparin complexes in inflammation and innate immunity | |
CN101668778B (zh) | 低出血抗凝血融合蛋白的制备及其应用 | |
CN101190945A (zh) | 特异性抗凝血物质的制备及其应用 | |
TW494104B (en) | Peptide having antithronbotic activity and process for producing the same | |
CN105848668A (zh) | 治疗或预防与出血或低凝血相关联的病状的疗法 | |
CA2108566A1 (en) | Method for inhibiting adhesion of white blood cells to endothelial cells | |
CN1301268C (zh) | 溶栓和抗凝双功能融合蛋白及应用 | |
CN1896108B (zh) | 特异性抗凝血物质的制备及其应用 | |
Caughey | Mast cell chymases and tryptases: phylogeny, family relations, and biogenesis | |
JP2019503341A (ja) | 植物細胞に生物学的に封入された、治療用タンパク質の、疾患の治療のための関心のある細胞型への標的化された送達 | |
Edgington et al. | The molecular biology of initiation of coagulation by tissue factor | |
CN113508126B (zh) | 新型肽及其用途 | |
CN102724995A (zh) | 富含具有延伸的α链的血纤蛋白原的血纤蛋白原制备物 | |
CN102443065A (zh) | 特异性抗凝血物质的制备及其应用 | |
TW200529870A (en) | Therapeutic use of factor XI | |
CA3089175A1 (en) | Extracorporeal device and matrix for removing fibrinolytic proteins from biological fluids, methods and uses thereof | |
CN100494366C (zh) | 血凝酶 | |
CN108676090B (zh) | 一种纤溶酶抑制剂及其应用 | |
KR950704488A (ko) | 원구류의 타액으로부터 제조된 트롬빈 억제제(Clotting Inhibitor Made From Protostomia Saliva) | |
CN103626850B (zh) | 具有细胞穿透功能的多肽及其在药物递送中的用途 | |
WO2005017139A1 (fr) | Thrombine provenant de venin de agkistrodon acutus utilisee comme medicament pour traiter une hemorragie | |
CN109295042A (zh) | 一种天然方格星虫纤溶酶的制备方法及其应用 | |
Mod | Engineered soluble protein C pathway receptors as novel pro-haemostatic agents | |
WO2011068853A2 (en) | TREATMENT OF IgE-MEDIATED DISEASE | |
FI109766B (fi) | Menetelmä eskaraasia sisältävän proteolyyttisen seoksen valmistamiseksi |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
C06 | Publication | ||
PB01 | Publication | ||
C10 | Entry into substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
C14 | Grant of patent or utility model | ||
GR01 | Patent grant | ||
TR01 | Transfer of patent right |
Effective date of registration: 20171124 Address after: No. 27 Taiping Road, Beijing, Haidian District Co-patentee after: BEIJING SANYOU LIHEZE BIOTECHNOLOGY CO., LTD. Patentee after: Emission and Radiation Medical Research Institute Address before: No. 27 Taiping Road, Beijing Co-patentee before: Beijing Sanly Science & Technology Development Co., Ltd. Patentee before: Emission and Radiation Medical Research Institute |
|
TR01 | Transfer of patent right |