PLASMINOGEN/ANGIOSTATIN CONVERTING ENZYME

纤溶酶原/血管抑制素转换酶

• 批准号: 2010213
• 负责人: GERALD A SOFF
• 金额: $ 20.52万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-04-01 至 2001-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2010213
• 关键词: angiogenesis inhibitors; antineoplastics; complementary DNA; enzyme activity; laboratory mouse; metastasis; neoplasm /cancer; neoplastic growth; plasma; plasminogen; protein metabolism; protein sequence; serine proteinases; tissue /cell culture

Angiostatin is a potent inhibitor of metastatic growth of some solid tumors. It is derived by proteolysis of plasminogen, and consists of the kringle 1-4 domain of plasminogen. Although the enzyme that converts plasminogen to angiostatin is expressed by some primary tumors, the identity of this enzyme, and which tumors express the enzyme remain unknown. We have demonstrated that the PC-3 and DU-145 human prostate cancer lines express the enzyme (or enzymes) necessary and sufficient for angiostatin generation, and this is tentatively designated as plasminogen- angiostatin converting enzyme (PACE). Additional data indicate that PACE activity is a serine protease. In preliminary experiments several purification tools have been identified and characterized which will allow for purification of PACE during this project. These include Reactive Red 120-Agarose (Dye-Ligand chromatography), Hi-Q-Sepharose (anion exchange chromatography) and soybean trypsin inhibitor-sepharose (affinity chromatography). The angiostatin generated in vitro by PC-3 cells is bioactive based on inhibition of endothelial cell proliferation and migration. Angiostatin antigen has been demonstrated in the plasma of athymic mice bearing PC-3 tumors. This proposal is to utilize the purification tools identified to complete the purification of the PACE enzyme or enzymes. If the enzyme is novel, the cDNA will be cloned. Monoclonal antibodies to PACE will be generated which will facilitate development of immunoassays and immunohistochemical staining for PACE. The kinetics of the interaction between PACE and plasminogen in vitro, as well as potential antiangiogenic and anti-metastatic effects of PACE administration in vivo will be analyzed.

血管抑素是一种有效的抑制肿瘤转移生长的药物， 肿瘤的 它是由纤溶酶原的蛋白水解而得，由 纤溶酶原Kringle 1-4结构域。 虽然这种酶 一些原发性肿瘤表达纤溶酶原转化为血管抑素， 这种酶的身份，以及哪些肿瘤表达这种酶仍然存在 未知 我们已经证明PC-3和DU-145人前列腺 癌细胞系表达的酶（或酶）是必需的，足以 血管生成抑制素，这是暂时指定为纤溶酶原- 血管抑素转化酶（PACE）。 数据显示，PACE 活性是丝氨酸蛋白酶。 在初步实验中， 已经鉴定并表征了纯化工具， 用于在该项目期间净化PACE。 其中包括活性红 120-琼脂糖（染料-配体色谱法），Hi-Q-琼脂糖（阴离子交换 层析）和大豆胰蛋白酶亲和层析-琼脂糖凝胶（亲和层析 色谱法）。 由PC-3细胞体外产生的血管抑素是 基于抑制内皮细胞增殖的生物活性， 迁移 血管抑素抗原已被证明在血浆中的 携带PC-3肿瘤的无胸腺小鼠。 这项建议是利用 为完成PACE的净化而确定的净化工具 酶或酶。 如果这种酶是新的，则将克隆cDNA。 将产生PACE的单克隆抗体， PACE的免疫测定和免疫组织化学染色的发展。 体外PACE与纤溶酶原相互作用的动力学， 以及PACE的潜在抗血管生成和抗转移作用 将分析体内施用。