Long-Acting Endostatin for Treating Cancer

描述(由申请人提供)： 内皮抑素是一种20 kDa的非糖基化蛋白，是一种特异性的内皮细胞增殖抑制因子，是一种有效的抗血管生成药物。由于几个原因，内皮抑素的治疗效果一直难以评估。内皮抑素的可溶性制剂很快失去活性，并受到与储存、处理和纯化方法有关的技术困难的困扰。此外，由于其血清半衰期相对较短，需要大量的蛋白质才能在体内看到显著的生物活性。我们的研究目标是创造新的内皮抑素蛋白，既改善了物理特性，又提高了体外和体内活性。我们完成这一任务的方法包括通过位置特异性的共价加成聚乙二醇部分来修饰内皮抑素。通过利用内皮抑素的已知结构，我们已经能够将特定的半胱氨酸突变工程到蛋白质中，作为硫醇反应钉的附着位置。这一策略允许合理设计具有特定结构的全活性蛋白质类似物，并克服了使用标准赖氨酸反应性聚乙二醇化试剂修饰蛋白质时失去生物活性和产物异质性的问题。在这笔赠款的第二阶段，我们将在体外和体内评估一些这些聚乙二醇化的内皮抑素突变蛋白。根据我们的经验和文献报道，聚乙二醇化的人内皮抑素应该具有更高的稳定性、更大的效力和更低的剂量要求。这将允许更多的患者获得该药物，因为据估计，超过900万癌症患者可能从抗血管生成治疗中受益。

DESCRIPTION (provided by applicant): Endostatin is a 20 kDa non-glycosylated protein that is a specific inhibitor of endothelial proliferation and a potent anti-angiogenesis agent. The therapeutic effects of endostatin have been difficult to evaluate for several reasons. Soluble preparations of endostatin quickly lose activity and are plagued by technical difficulties related to storage, handling, and purification methods. In addition, because of its relatively short serum half-life, large quantities of protein are needed to see significant bioactivity in vivo. The goal of our research is to create novel endostatin proteins with both improved physical characteristics and enhanced in vitro and in vivo activities. Our approach towards accomplishing this task involves modifying endostatin by the site-specific covalent addition of a polyethylene glycol (PEG) moiety. By using the known structure of endostatin, we have been able to engineer into the protein, specific cysteine mutations to serve as attachment sites for thiol-reacfive PEGs. This strategy allows for the rational design of fully active protein analogues of defined structure and overcomes the problem of loss of bioactivity and product heterogeneity when proteins are modified using standard lysine-reactive PEG reagents. During the Phase II portion of this grant we will evaluate a number of these PEGylated endostatin muteins both in vitro and in vivo. Based upon our experience and literature reports, PEGylated human endostatin should have enhanced stability, greater potency and lower dose requirements. This will allow a larger number of patients access to the drug since it is estimated that more than 9 million cancer patients may benefit from anti-angiogenesis therapy.