Antibody response to aggregated human interferon alpha2b in wild-type and transgenic immune tolerant mice depends on type and level of aggregation
✍ Scribed by Suzanne Hermeling; Huub Schellekens; Coen Maas; Martijn F.B.G. Gebbink; Daan J.A. Crommelin; Wim Jiskoot
- Publisher
- John Wiley and Sons
- Year
- 2006
- Tongue
- English
- Weight
- 301 KB
- Volume
- 95
- Category
- Article
- ISSN
- 0022-3549
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✦ Synopsis
The aim of this study was to determine the sensitivity of transgenic immune tolerant mice for the type and level of aggregation of recombinant human interferon alpha2b (rhIFNa2b). RhIFNa2b was aggregated by metal-catalyzed oxidation or by incubation at elevated temperature and various pHs. Native rhIFNa2b was mixed with oxidized rhIFNa2b at different ratios to obtain samples with different aggregation levels. The preparations were characterized by UV and fluorescence spectroscopy, gel permeation chromatography (GPC), dynamic light scattering (DLS), sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting, and ELISA. The immunogenicity was evaluated in wild-type mice and transgenic mice immune tolerant for hIFNa2. Sera were analyzed by ELISA for the presence of rhIFNa2bspecific antibodies. The oxidized and aged preparations widely differed regarding the level and nature of aggregates. All preparations containing aggregates increased the immune response in the wild-type mice as compared to native rhIFNa2b and were able to break the tolerance of the transgenic mice. The more native-like the conformation of the aggregated proteins, the more immunogenic the preparations were in the transgenic mice. The native-like aggregates prepared via metal catalysis induced a dose-dependent loss of tolerance in the transgenic mice. In conclusion, the transgenic mouse model can be used to screen rhIFNa2b formulations for low levels of immunogenic aggregates obtained under accelerated storage conditions.