Isolation of Positional Isomers of Mono-poly(ethylene glycol)ylated Interferon/#alpha#-2A and the Determination of Their Biochemical and Biological Characteristics

摘要

MonoPEGylated interferon #alpha#-2a (monoPEG-IFN) is produced by conjugating interferon #alpha#-2a to a chemically reactive form of polyethylene glycol-5000 in 1:1 ratio, via a urea linkage. Potentially, all 11 lysine residues and the N-terminal amino group in interferon #alpha#-2a are available for PEGylation. Thus monoPEG-IFN is a mixture of individual species, each having a unique site of PEGylation. A method involving sulfopropyl-high perfrmance liquid chromatography (SP-HPLC) which exploits the minute local charage differences between the individual monoPEG-IFN #alpha#-2a species (positional isomers) is used to separate them. Eleven positional isomers were isolated by this method, each having a specific lysine as their PEGylation site indicating all eleven lysines of interferon #alpha#-2a were PEGylated. However, N-terminal amino group was not derivatized. All isolated species exhibit antiviral and antiproliferative activities.