TARGETING ANGIOGENESIS WITH TOXIN VEGF FUSION PROTEINS

描述：（申请人的描述） 该项目的总体目标是开发一种商业上可行的细菌 毒素-血管内皮生长因子融合蛋白 靶向血管生成部位的内皮细胞，主要在固体中 肿瘤和生长转移。 VEGF与内皮细胞相互作用 特异性KDR/flk-1受体，在内皮细胞中过表达， 相对于其在静止内皮上的表达的血管生成位点。 在该项目的第一阶段，我们构建，表达和纯化了两个 新的融合蛋白，SLT-VEGF/L和SLT-VEGF/S，分别含有， 志贺样毒素Ⅰ A亚单位及其功能活性片段 A亚单位的 出血性 结肠炎和溶血性尿毒综合征， 内皮细胞对炎症特别敏感。 SLT-VEGF/L和SLT-VEGF/S蛋白选择性抑制内皮细胞生长 过表达KDR/flk-1受体的细胞，IC-50为0.15 nM。 以低 纳摩尔浓度SLT-VEGF/L对内皮细胞具有细胞毒性 过表达KDR/flk-1受体。 然而，这些蛋白质并不影响 具有低数量KDR/flk-1受体的生长中的内皮细胞，静止 内皮细胞和生长中的非内皮细胞。 这些结果提供了一个 SLT-VEGF融合蛋白的II期体内试验的基本原理 血管生成的选择性抑制剂。 我们还将测试SLT-VEGF是否 蛋白质可以通过增强递送而与其它抗肿瘤药物协同作用 通过受损的肿瘤内皮或通过创造缺氧条件， 生物还原疗法 最后，我们将优化SLT-VEGF融合蛋白 以实现高水平的表达、活性和稳定性， 临床前和临床试验所必需的。 拟定商业应用： 该研究的潜在商业应用的简要总结：抑制血管生成但不影响正常内皮或正常细胞的VEGF-毒素融合蛋白将成为用于治疗血管生成依赖性病理的商业产品。

DESCRIPTION: (Applicant's Description) The overall goal of this project is to develop a commercially viable bacterial toxin-VEGF (vascular endothelial growth factor) fusion protein for selective targeting of endothelial cells at sites of angiogenesis, primarily in solid tumors and growing metastases. VEGF interacts with the endothelial cell specific KDR/flk-1 receptor that is overexpressed in endothelial cells at the sites of angiogenesis relative to its expression on quiescent endothelium. In Phase I of this project we have constructed, expressed and purified two novel fusion proteins, SLT-VEGF/L and SLT-VEGF/S which contain, respectively, The A subunit of Shiga-like toxin I (SLT) and a functionally active fragment of the A subunit. SLT causes hemorrhagic colitis and hemolytic uremic syndrome by damaging endothelial cells suggesting that endothelial cells are particularly sensitive to SLT . SLT-VEGF/L and SLT-VEGF/S proteins selectively inhibit growth of endothelial cells overexpressing KDR/flk-1 receptors with IC-50 of 0.15 nM. At low nanomolar concentrations SLT-VEGF/L is cytotoxic for endothelial cells overexpressing KDR/flk-l receptors. However, these proteins do not affect growing endothelial cells with low numbers of KDR/flk-l receptors, quiescent endothelial cells, and growing non-endothelial cells. These results provide a rationale for Phase II in vivo testing of SLT-VEGF fusion proteins as highly selective inhibitors of angiogenesis. We will also test whether SLT-VEGF proteins may synergize with other antitumor drugs by enhancing delivery through damaged tumor endothelium or by creating hypoxic conditions for bioreductive therapeutics. Finally, we will optimize SLT-VEGF fusion proteins to achieve high levels of expression, activity, and stability that are necessary for preclinical and clinical trials. PROPOSED COMMERCIAL APPLICATION: A brief summary of the potential commercial applications of the research: VEGF-toxin fusion proteins that inhibit angiogenesis but do not affect normal endothelium or normal cells will be commercial products for therapy of angioaenesis-dependent pathologies.