Development of glycosylated human interleukin-1alpha, neoglyco IL-1alpha, by coupling with D-galactose monosaccharide: synthesis and purification.
ABSTRACT In order to develop glycosylated cytokine, recombinant human IL-1alpha was chemically modified with galactose monosaccharide. Galactose with C9 spacer, 8-(hydrazinocarbonyl)octyl beta-D-galactopyranoside (3), was synthesized by glycosylation of C9 spacer, methyl 9-hydroxynonanoate, with acetobromogalactose, followed by deacetylation and hydrazidation. Total yield of 3 was 43.6% in three steps. Compound 3 was coupled to IL-1alpha by the acyl azide method. The glycosylated IL-1 was purified by anion-exchange chromatography, and galactose coupled to IL-1 was confirmed by R. communis lectin blotting. Based on the molecular weight, the average number of carbohydrate molecules introduced per molecule of IL-1alpha was estimated to be 9.1.
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ABSTRACT: In order to study the effect of glycosylation on its biological activities, and to develop TNFalpha with less deleterious effects, recombinant human TNFalpha was chemically coupled with N-acetylneuraminic acid (NeuAc). NeuAc with C9 spacer was coupled to TNFalpha by acyl azide method. Two glycosylated TNFalphas, designated L NeuAc-TNFalpha and H NeuAc-TNFalpha, were purified by anion-exchange chromatography. NeuAc coupling to TNFalpha was confirmed by lectin blotting. Average number of carbohydrate molecules introduced per molecule of L NeuAc-TNFalpha and H NeuAc-TNFalpha were estimated to be 1.0 and 1.5, respectively. We examined a variety of TNFalpha activities in vitro, including antiproliferative or cytotoxic activities to tumor cells, proliferative effect on fibroblast cells, stimulatory effects on IL-6 production by melanoma cells and NF-kappaB activation in hepatoma cells. L NeuAc-TNFalpha and H NeuAc-TNFalpha exhibited reduced activities about 1/3 and 1/10 as compared to native TNFalpha in all the activities performed in vitro.Cancer Immunology and Immunotherapy 05/2007; 56(4):545-53. · 3.64 Impact Factor