"Pegylation of Recombinant Human Interferon Alpha-2 and Characterization of Pegylated Products”

Abstract

Recombinant human interferon- alpha 2 (rhIFN-α2) is FDA approved for treatment of some viral and malignant diseases. Pegylated rhIFN-α2 (Pegasys and PEG-Intron) have highly improved pharmacokinetics, pharmacodynamics and therapeutic efficiency compared to native protein. In this work, we studied the pegylation of purified properly refolded rhIFN -α2b using linear (20kDa) PEG-NHS to prepare pegylated rhIFN-α2b with high stability and activity. The effect of different parameters like rhIFN-α2b final concentration, pH, rhIFN-α2b/PEG molar ratios and reaction time on the efficiency of pegylation (high percentage of monopegylated rhIFN-α2b) have been studied in small scale (100µl) pegylation reaction trials. Study of the percentages of different components of these reactions (mono, di, polypegylated rhIFN-α2b and unpegylated rhIFN-α2b) indicated that, 2h is optimum time to complete the reaction. The pegylation efficiency increased at pH 8 (57.9%) by reducing the protein concentration to 1mg/ml and reducing the rhIFN-α2b/PEG ratio to 1:2.