Module Assembly and Evaluation of Artificial Glycosaminoglycans

磺化糖胺聚糖（GAGs）及其类似物戊聚糖聚硫酸酯（PPS）、葡聚糖硫酸酯（DS）和肝素可抑制朊病毒感染细胞中异常朊蛋白（PrP^<Sc>）的形成，延长瘙痒病感染动物的潜伏期。gag的磺化不完全受调控，可能的磺化位点是随机的。这一特性阻碍了对参与细胞表面各种生物事件的gag基本结构的阐明。为了解决GAGs在抑制PrP^<Sc>形成中的构效关系，我们采用基于碳水化合物模块概念的合成策略组装了一系列糖胺聚糖类似物。在合成的糖苷及其共聚物中，单体4-磺基- n -乙酰氨基葡萄糖胺（4SGN）和聚-4SGN和聚-6-磺基- n -乙酰氨基葡萄糖胺（6SGN）两种共聚物抑制PrP^<Sc>的形成，50%有效剂量低于20 μg/ml。连续治疗抑制效果更明显。它们降低了细胞朊蛋白的表达，但不影响细胞生长。结构比较表明，C-2上的n -乙酰基与C-4或C-6上的硫酸盐基重合可能参与抑制PrP^<Sc>的形成。然而，无论是单体的还是二聚体的6SGN，都没有表现出抑制作用，但聚6SGN表现出抑制作用，这表明多价构型在6SGN作用中的重要性。这些结果表明，人工硫酸糖苷及其聚合物不仅可用于分析GAGs的构效关系，而且可用于开发新的朊病毒疾病治疗化合物。

Sulfated glycosaminoglycans (GAGs) and their analogues such as pentosan polysulfate (PPS), dextran sulfate (DS), and heparin inhibit abnormal isoform of prion protein (PrP^<Sc>) formation in prion-infected cells and prolong the incubation time of scrapie-infected animals. Sulfation of GAGs is not completely regulated and possible sulfation sites are randomly sulfated. This property impedes an elucidation of fundamental structures of GAGs that are involved in diverse biological events on cell surfaces. To address the structure-activity relationship of GAGs in the inhibition of PrP^<Sc> formation, we assembled a series of glycosaminoglycans analogues by applying our synthetic strategy based on the concept of carbohydrate modules. Among the synthetic glycosides and their copolymers thus derived and examined, monomeric 4-sulfo-N-acetyl-glucosamine (4SGN), and two copolymers, poly-4SGN and poly-6-sulfo-N-acetyl-glucosamine (6SGN), inhibited PrP^<Sc> formation with 50% effective dose lower than 20 μg/ml. The inhibitory effect became more evident in consecutive treatment. They reduced the expression of cellular prion protein but did not affect cell growth. Structural comparison suggested that coincidence of N-acetyl group at C-2 with sulfate group at C-4 or C-6 might be involved in the inhibition of PrP^<Sc> formation. However, neither monomeric nor dimeric 6SGN, but poly-6SGN showed the inhibitory effect, suggesting the importance of polyvalent configuration in the effect of 6SGN. These results indicate that the artificially sulfated glycosides and their polymers are useful not only for the analysis of structure-activity relationship of GAGs but also for the improvement of new therapeutic compounds for prion diseases.