Fucosylation of Disaccharide Precursors of Sialyl LewisX Inhibit Selectin-mediated Cell Adhesion

Abstract

We showed previously that HL-60 and F9 mouse embryonal carcinoma cells will take up and deblock peracetylated Gal␤1-4GlcNAc␤-O-naphthalenemethanol (Gal␤1-4GlcNAc-NM) and use the disaccharide as a primer of oligosaccharide chains (Sarkar, A. K., Fritz, T. A., Taylor, W. H., and Esko, J. D. (1995) Proc. Natl. Acad. Sci. U. S. A. 92, 3323-3327). We now report that another disaccharide, acetylated GlcNAc␤1-3Gal-naphthalenemethanol (GlcNAc␤1-3Gal-NM), has even greater potency and that both compounds will inhibit sialyl Lewis X (sLe x )-dependent cell adhesion. When fed to U937 cells, acetylated forms of Gal␤1-4GlcNAc-NM and GlcNAc␤1-3Gal-NM primed oligosaccharides in a dosedependent manner. Analysis of compounds assembled on Gal␤1-4GlcNAc-NM showed only one product, namely Gal␤1-4(Fuc␣1-3)GlcNAc-NM. In contrast, GlcNAc␤1-3Gal-NM generated Gal␤1-4GlcNAc␤1-3Gal-NM, Gal␤1-4(Fuc␣1-3)GlcNAc␤1-3Gal-NM, NeuAc␣2-3Gal␤-1-4GlcNAc␤1-3Gal-NM, and NeuAc␣2-3Gal␤1-4(Fuc␣1-3)GlcNAc␤1-3Gal-NM. Both compounds decreased the incorporation of [ 3 H]fucose into cellular glycoconjugates, without affecting the incorporation of [ 3 H]mannosamine, a precursor of sialic acid residues. Moreover, the overall extent of sialylation was not affected based on the reactivity of cells to fluorescein isothiocyanateconjugated Maackia amurensis lectin. Priming inhibited expression of sLe x on cell surface glycoconjugates, which reduced E-selectin-dependent cell adhesion to tumor necrosis factor-␣-activated human umbilical vein endothelial cells. GlcNAc␤1-3Gal-NM and Gal␤1-4Glc-NAc-NM represent starting points for making enzymespecific, site-directed inhibitors of glycosyltransferases that could act in living cells.

Peracetylated N-Acetylmannosamine, a Synthetic Sugar Molecule, Efficiently Rescues Muscle Phenotype and Biochemical Defects in Mouse Model of Sialic Acid-deficient Myopathy

“…This distinct efficiency exhibited by Ac 4 ManNAc may be explained by two factors. First, O-acetylation of ManNAc provides hydrophobic modifications on the hydroxyl groups of the molecule because of the properties endowed by acetyl esters that facilitate passive diffusion of the compound into a cell (39). Second, O-acyl ManNAc analogs are believed to be sequestered in cellular membranes that serve as a "reservoir" for these compounds (35) as only a minor fraction is believed to be converted by cells to sialic acid.…”

Expression Patterns of α2,3-Sialyltransferases and α1,3-Fucosyltransferases Determine the Mode of Sialyl Lewis X Inhibition by Disaccharide Decoys

“…Cells take up the glycosides by passive diffusion, deacetylate them with endogenous carboxyesterases, and use them as substrates for glycosyltransferases, resulting in the generation of oligosaccharides containing Lewis type antigens or precursor structures (23)(24)(25). Both compounds are nontoxic up to 100 M based on trypan blue exclusion and normal growth rates.…”

“…Suitably conjugated disaccharides also act as primers, such as per-O-acetylated Gal␤1,4GlcNAc␤-O-naphthalenemethanol (AcGGn-NM), GlcNAc␤1,3Gal␤-O-naphthalenemethanol (AcGnG-NM), and Gal␤1,3GalNAc␣-O-naphthalenemethanol, since these compounds resemble intermediates en route to mature O-linked chains of glycoproteins. Disaccharide-based primers act at much lower concentrations than N-acetylgalac-tosaminides (10 -50 M versus 1-2 mM, respectively) (23)(24)(25) and have proven effective in blocking leukocyte adhesion to immobilized selectins in vitro (24). This finding encouraged us to examine their effect on human carcinoma cells, since positive results might suggest their use as antimetastatic agents.…”

Syntheses of Oligosaccharides Using Cell Function

“…Furthermore, those primers were also found to be inhibitors for endogenous glycan biosynthesis (6)(7)(8)(9)(10). The primers having bezyl and naphtyl group as aglycon tend to stay in cells because of their hydrophobicity.…”