Search Results (3)

The biological activity of therapeutic antibodies is highly influenced by their glycosylation profile. A valuable method for increasing the cytotoxic efficacy of antibodies, which are used, for example, in cancer treatment, is the reduction of core fucosylation, as this enhances the elimination of target cells through antibody-dependent cell-mediated cytotoxicity. Development of fucose analogues is currently the most promising strategy to reduce core fucosylation without cell line engineering. Since peracetylated sugars display enhanced cell permeability over the highly polar free hydroxy sugars, this work sought to compare the efficacy of peracetylated sugars to their unprotected forms. Two potent fucose analogues, 2-deoxy-2-fluorofucose and 5-alkynylfucose, and their acetylated forms were compared for their effects on fucosylation. 5-alkynylfucose proved to be more potent than 2-deoxy-2-fluorofucose at reducing core fucosylation but was associated with a significant higher incorporation of the alkynylated fucose analogue. Acetylation of the sugar yielded only slightly lower fucosylation levels suggesting that acetylation has a minor impact on cellular entry. Even though the efficacy of all tested components was confirmed, results presented in this study also show a significant incorporation of unnatural fucose analogues into the glycosylation pattern of the produced IgG, with unknown effect on safety and potency of the monoclonal antibody. Full article

Manipulations of cell surface glycosylation or glycan decoration of selected proteins hold immense potential for exploring structure-activity relations or increasing glycoprotein quality. Metabolic glycoengineering describes the strategy where exogenously supplied sugar analogues intercept biosynthetic pathways and are incorporated into glycoconjugates. Low membrane permeability, which so far limited the large-scale adaption of this technology, can be addressed by the introduction of acylated monosaccharides. In this work, we investigated tetra-O-acetylated, -propanoylated and -polyethylene glycol (PEG)ylated fucoses. Concentrations of up to 500 µM had no substantial effects on viability and recombinant glycoprotein production of human embryonic kidney (HEK)-293T cells. Analogues applied to an engineered Chinese hamster ovary (CHO) cell line with blocked fucose de novo synthesis revealed an increase in cell surface and recombinant antibody fucosylation as proved by lectin blotting, mass spectrometry and monosaccharide analysis. Significant fucose incorporation was achieved for tetra-O-acetylated and -propanoylated fucoses already at 20 µM. Sequential fucosylation of the recombinant glycoprotein, achieved by the application of increasing concentrations of PEGylated fucose up to 70 µM, correlated with a reduced antibody’s binding activity in a Fcγ receptor IIIa (FcγRIIIa) binding assay. Our results provide further insights to modulate fucosylation by exploiting the salvage pathway via metabolic glycoengineering. Full article

The novel glycosphingolipid, b-D-GalNAcp(1®4)[a-D-Fucp(1®3)]-b-D-GlcNAcp(1®)Cer (A), isolated from the marine sponge Aplysinella rhax has a unique structure, with D-fucose and N-acetyl-D-galactosamine moieties attached to a reducing-end N-acetyl-D-glucosamine through an a1®3 and b1®4 linkage, respectively. We synthesized glycolipid 1 and some non-natural di- and trisaccharide analogues 2-6 containing a D-fucose residue. Among these compounds, the natural type showed the most potent nitric oxide (NO) production inhibitory activity against LPS-induced J774.1 cells. Our results indicate that both the presence of a D-Fuca1-3GlcNAc-linkage and the ceramide aglycon portion are crucial for optimal NO inhibition. Full article