Oxidized and Aggregated Recombinant Human Interferon Beta is Immunogenic in Human Interferon Beta Transgenic Mice

Publication Type:

Journal Article

Source:

Pharm Res, Volume 28, Number 10, pp. 2393-402 (2011)

ISBN:

1573-904X (Electronic)07

DOI Name (links to online publication)

10.1007/s11095-011-0451-4

Abstract:

PURPOSE: To study the effect of oxidation on the structure of recombinant human interferon beta-1a (rhIFNbeta-1a) and its immunogenicity in wild-type and immune-tolerant transgenic mice. METHODS: Untreated rhIFNbeta-1a was degraded by metal-catalyzed oxidation, H(2)O(2)-mediated oxidation, and guanidine-mediated unfolding/refolding. Four rhIFNbeta-1a preparations with different levels of oxidation and aggregation were injected intraperitoneally in mice 15x during 3 weeks. Both binding and neutralizing antibodies were measured. RESULTS: All rhIFNbeta-1a preparations contained substantial amounts of aggregates. Metal-catalyzed oxidized rhIFNbeta-1a contained high levels of covalent aggregates as compared with untreated rhIFNbeta-1a. H(2)O(2)-treated rhIFNbeta-1a showed an increase in oligomer and unrecovered protein content by HP-SEC; RP-HPLC revealed protein oxidation. Guanidine-treated rhIFNbeta-1a mostly consisted of dimers and oligomers and some non-covalent aggregates smaller in size than those in untreated rhIFNbeta-1a. All degraded samples showed alterations in tertiary protein structure. Wild-type mice showed equally high antibody responses against all preparations. Transgenic mice were discriminative, showing elevated antibody responses against both metal-catalyzed oxidized and H(2)O(2)-treated rhIFNbeta-1a as compared to untreated and guanidine-treated rhIFNbeta-1a. CONCLUSIONS: Oxidation-mediated aggregation increased the immunogenicity of rhIFNbeta-1a in transgenic mice, whereas aggregated preparations devoid of measurable oxidation levels were hardly immunogenic.

09/02/2012