Modified multivalent poly-N-acetyllactosamine glycans as novel ligands of human galectin-3

近年来，碳水化合物在生物系统中的功能作用一直是一个热门的研究领域。碳水化合物以前被认为是保护细胞表面的惰性分子，现在被认为是细胞过程中重要的信息传递者。细胞表面的糖蛋白和糖脂可以作为蛋白质的配体，从而为细胞间的黏附提供锚，或为病毒和细菌的入侵提供受体。它们在调节分化过程中的作用是公认的。因此，糖共轭化合物和糖模拟物具有巨大的药用潜力。对于碳水化合物的生理应用，必须解决两个重要问题：i)多价性--由天然结构(多触角糖蛋白)中存在的相当弱的碳水化合物-蛋白质相互作用决定；ii)结构的稳定性和保护天然O-糖苷键不被生物体中的许多糖苷酶断裂。这一障碍可以通过制备人工多价新糖共轭化合物和通过引入非自然的碳水化合物连接导致类似糖的概念来克服。由于碳水化合物的高度复杂性和内在性质，复杂的多价或多线性糖簇的化学扩展实际上是不可能的，例如，存在大量非常相似的OH基团，这些基团只能通过复杂和费力的保护/去保护策略来区分。在合成复杂的碳水化合物结构方面，酶法已成为有机化学的一种可行的替代方法，特别是由于其选择性和简单性。在这里，与糖苷酶相比，糖基转移酶由于其绝对的立体选择性和高的区域选择性，为选择性修饰这类结构提供了理想的工具。由于这些酶可以修饰自然界中复杂的多天线结构，它们能够作用于含有LacNAc结构的多线性和多价糖簇。在Elling和Kren基团糖基转移酶的当前联合提案中，将应用糖合成酶和半乳糖氧化酶作为糖链修饰酶来创建连接到稳定连接物的修饰的聚-LacNAc文库。将使用两个小组的互补方法和材料。人Galectin-3是肿瘤进展和血管生成中重要的Galectin，其与人Galectin-3结合的选择性应通过糖基转移酶的级联反应中的特定糖链表位的合成来解决。这个携带聚-LacNAc寡聚体的小表位文库将用于下一步解决其多价呈现问题。这将通过选择性氧化聚-LacNAc低聚物的半乳糖部分和随后携带多聚-LacNAc低聚物的表位的化学结合来实现。通过这种方式，将获得一个修饰的非自然支化聚-LacNAc寡聚物库，作为Gal-3的潜在抑制剂。

The functional role of carbohydrates in biological systems has been an area of intense research in recent years. Carbohydrates, formerly considered as inert molecules protecting the cell surface, are now recognized as important information transmitters in cellular processes. Cell surface glycoproteins and glycolipids can serve as protein ligands, thereby providing an anchor for intercellular adhesion or a receptor for viral and bacterial invasion. Their role in regulation of differentiation processes is generally acknowledged. Glycoconjugates and glycomimetics have, therefore, a significant pharmaceutical potential. For physiological applications of carbohydrates two important issues have to be addressed: i) multivalency - dictated by the fact of rather weak carbohydrate-protein interactions occurring in natural structures (multiantennary glycoproteins); ii) stability of structure and protection from cleavage of natural O-glycosidic bonds by numerous glycosidases in the organism. This obstacle can be overcome by preparation of artificial multivalent neoglycoconjugates and by introduction of unnatural carbohydrate linkages leading to a glycomimetic concept. Chemical extension of complex multivalent or multi-linear glycoclusters is virtually impossible due to high complexity and intrinsic properties of carbohydrates, e.g., presence of a large number of rather similar OH groups that can only be distinguished by complex and labor-demanding protection/deprotection strategy. Enzymatic methods have already become a viable alternative to organic chemistry in the synthesis of complex carbohydrate structures, especially due to their selectivity and simplicity. Here, in contrast to glycosidases, glycosyltransferases offer an ideal tool for selective modification of such structures due to their absolute stereoselectivity and high regioselectivity. Since these enzymes modify complex multi-antennary structures in nature they are able to work on multi-linear and multivalent glycoclusters containing LacNAc structures. In the present joint proposal of the Elling and Kren group glycosyltransferases, glycosynthases and galactose oxidase as glycan modifying enzyme will be applied to create a library of modified poly-LacNAc conjugated to stable linkers. Complementary approaches and materials from both groups will be used. Selectivity for binding to human galectin-3, an important galectin in tumor progression and angiogenesis, shall be addressed by the synthesis of specific glycan epitopes in cascade reactions of glycosyltransferases. This small library of epitope carrying poly-LacNAc oligomers will be used for the next step addressing their multivalent presentation. This shall be accomplished by selective oxidation of galactose moieties of poly-LacNAc oligomers and subsequent chemical conjugation of epitope carrying poly-LacNAc oligomers. In this way a library of modified unnatural branched poly-LacNAc oligomers will be obtained as potential inhibitors of Gal-3.