Synthetic Heparan Sulfate: Probing Biosynthesis to Prepare Defined Drugs

说明(申请人提供)：硫酸乙酰肝素(HS)是一类多糖，包括著名的药物肝素，在细胞生长和发育中发挥重要作用。在哺乳动物中观察到的HS的许多结构变体被假设为在健康和疾病期间以不同的方式调节生长因子介导的信号和调节止血。我们的项目将探索HS结构变化是如何在生物合成/分解代谢过程中产生的，以及绘制路线以制备更具选择性和更强大的预期生物活性的更明确的分子。一个有利的假设是，这种模式 HS结构域(由N-磺酸(NS)和N-乙酰(NA)结构域组成)编码的信息调节HS与生长因子、细胞因子和凝血因子等蛋白质的相互作用。目前，人们很难理解HS的生物合成修饰和结构域的放置是如何控制的，以及哪些HS结构域结构在不同的生物系统中具有最高的活性。我们将应用我们最新开发的合成方法来回答该领域的关键问题。将生成具有大小定义和放置定义的NS和NA结构域的HS聚合物的小文库。然后，这些HS聚合物将被生物合成酶修饰，包括O-磺基转移酶、C5-异构体酶和/或分解代谢酶，包括Endo-6-Oendosulate ase和Heparanase。一种有重点的组合方法将用于生产特定的HS聚合物，以测试两种截然相反的生长因子信号模型，这是细胞增殖、胚胎发育和癌症中的关键事件。在这个项目中，我们的具体目标是：目标1：用化学酶法合成一个定义了NS和NA结构域的HS聚合物库。目的2.分析不同酶对HS多糖库的修饰模式。目的3：研究HS作为辅受体成纤维细胞的结构/功能关系 生长因子受体信号转导。

DESCRIPTION (provided by applicant): Heparan sulfate (HS), a class of polysaccharides that includes the well-known drug heparin, plays essential roles in cell growth and development. The many structural variants of HS observed in mammals have been hypothesized to differentially modulate growth factor-mediated signaling and regulate hemostasis during health and disease. Our project will explore how the HS structural variation is generated during biosynthesis/catabolism as well as map routes to prepare more defined molecules having greater selectivity and more potent desired bioactivities. A favored hypothesis is that the pattern of HS domains (comprised of both N-sulfo (NS) and N-acetyl (NA) domains) encodes information that modulates HS interaction with proteins like growth factors, cytokines, and clotting factors. Currently, it is very difficult to understand how HS biosynthetic modification an domain placement is controlled and which HS domain structures possess the highest activities in different biological systems. We will apply our newly developed synthetic methodology to answer key questions in the field. A small library of HS polymers having size-defined and placement-defined NS and NA domains will be generated. These HS polymers will then be modified by biosynthetic enzymes, including O-sulfotransferases, and C5-epimerase and/or catabolic enzymes including endo-6- Oendosulfatases and heparanase. A focused combinatorial approach will be used to produce defined HS polymers to test two opposing models of growth factor signaling, a key event in cell proliferation, embryonic development, and cancer. In this project our specific aims are: Aim 1: To chemoenzymatically synthesize a small library of HS polymers having defined NS and NA domains. Aim 2. To analyze the modification patterns generated through the action of various enzymes on the HS polysaccharide library. Aim 3: To characterize the structure/function relationship of HS activity as the co-receptor fibroblast growth factor receptor signaling.