Synthesis of glycosylated human interleukin-1alpha, neoglyco IL-1alpha, coupled with N-acetylneuraminic acid

Abstract

In order to develop glycosylated cytokine, recombinant human IL-1alpha was chemically modified with N-acetylneuraminic acid (NANA). NANA with C9 spacer, 8-(hydrazinocarbonyl)octyl 5-acetamido-3, 5-dideoxy-D-glycero-alpha-D-galacto-2-nonulo-pyranosidonic acid potassium salt (6), was synthesized by glycosylation of C9 spacer, 8-[2-N-(benzyloxycarbonyl)hydrazinocarbonyl]octanol, with methylthio glycoside derivatives of NANA in the presence of molecular sieves 3A and methyl (methylthio)sulfonium trifrate in propionitrile, followed by separation of a and beta anomers with a column chromatography and deprotection. Compound 6 was coupled to IL-1alpha by the acyl azide method. The glycosylated IL-1 was purified by anion-exchange chromatography, and NANA coupled to IL-1 was confirmed by oxidation with NaIO4. Based on the molecular weight average number of carbohydrate molecules introduced per molecule of IL-1alpha was estimated to be 2.9.