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Circumventing the heterogeneity and instability of human serum albumin-interferon-α2b fusion protein by altering its orientation

Paper ID Volume ID Publish Year Pages File Format Full-Text
25170 43558 2007 8 PDF Available
Title
Circumventing the heterogeneity and instability of human serum albumin-interferon-α2b fusion protein by altering its orientation
Abstract

Albuferon is a novel long-acting interferon resulted from the direct genetic fusion of human albumin and interferon-α2b (HSA-IFN-α2b). Albuferon, co-developed by Human Genome Sciences Inc. and Novartis, is currently in late stage development for the treatment of hepatitis C. It was unexpected that HSA-IFN-α2b secreted from Pichia pastoris migrated as doublets on non-reducing SDS-PAGE and was prone to form covalent aggregates in aqueous solution. The heterogeneity and instability of HSA-IFN-α2b lowered its recovery rate to about 10% and necessitated lyophilized formulation. Site-directed mutagenesis revealed that the heterogeneity and instability of HSA-IFN-α2b was caused by the incomplete disulfide bridge formation between Cys1 and Cys98 of IFN-α2b. To alleviate the structural perturbation of IFN-α2b by HSA, IFN-α2b-HSA fusion protein, in which IFN-α2b was located at the N-terminus, was created. IFN-α2b-HSA was shown to be homogeneous and stable at 37 °C for at least 10 days. The improved homogeneity and stability of IFN-α2b-HSA increased the recovery rate by 2.5-fold and made the development of stable solution formulation possible. In vitro antiviral assays showed that both fusion proteins retained the activity of IFN-α2b, and the EC50 of HSA-IFN-α2b, and IFN-α2b-HSA was calculated to be 120 ± 12.5, and 160 ± 1 1.3 ng/ml, respectively. The increased recovery rate and the possibility of solution formulation of IFN-α2b-HSA may compensate for its slightly decreased in vitro activity, and makes it to be a promising therapeutic agent that deserves further evaluation.

Keywords
Long-acting interferon; Human serum albumin fusion protein; Pichia pastoris
First Page Preview
Circumventing the heterogeneity and instability of human serum albumin-interferon-α2b fusion protein by altering its orientation
Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 131, Issue 3, 15 September 2007, Pages 245–252
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering