| While this thesis contains two distinct parts, it is united in using chemical biology approaches in the field of glycoimmunology. In part one, I set out to identify inhibitors of sialyl (STs)- and fucosyltransferases (FUTs), enzymes that add sialic acid and fucose, respectively, to the tips of glycan chains. While gene ablation studies in mice have highlighted the pharmacological potential of inhibiting these enzymes for the treatment of immune-cell mediated diseases, biosynthetic inhibitors are lacking. Towards this end, a broadly applicable ST and FUT inhibitor-screening assay was developed and used to screen 16,000 small molecules against 5 STs and FUTs allowing for the identification of various classes of inhibitors. The next chapter describes a complimentary approach using fluorinated sialic acid and fucose analogs, which, upon feeding to cells, are readily taken up and metabolically converted into fluorinated donor substrate analogs intracellularly. These molecules act not only as well-established competitive inhibitors of sialyl- and fucosyltransferase, but also inhibit the de novo synthesis of the natural donor substrates, resulting in profound inhibition of a variety of sialylated and fucosylated glycans.;Part two of this thesis aims to exploit the restricted expression patterns and endocytic capacity of the Siglec family for cell-directed therapies. While Siglecs bind to sialic acid containing glycans with relatively low affinity and selectivity, limited work has suggested that appending unnatural substitutents to the C9 and C5 position of this "privileged scaffold" can give increases in affinity and selectivity for individual Siglecs. To systematically explore this, a library of ∼240 9- and 5-substituted sialosides was synthesized using solution-phase Cu(I)-accelerated azide-alkyne cycloaddition reactions. Subsequent microarray printing of the library followed by Siglec-screening led to the identification of ligand analogs which, when displayed on liposomal nanoparticles, allow for selective targeting of Siglec-9 and Siglec-10 expressing cells in peripheral human blood. This approach was then expanded by connecting the synthesis and screening procedures such that they both happened 'on-chip'. In so doing, the analog library size was dramatically increased (∼1200 members) with far less synthetic effort, allowing for the identification of a Siglec-7 specific sialic acid analog for cell targeting applications. |
