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Structural characterization and immunogenicity in wildtype and immune tolerant mice of degraded recombinant human interferon alpha2b

2005-11Book part Open access
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dc.contributor.authorHermeling, Suzanne
dc.contributor.authorCaetano, Liliana Aranha
dc.contributor.authorDamen, J. Mirjam A.
dc.contributor.authorSlijper, Monique
dc.contributor.authorSchellekens, Huub
dc.contributor.authorCrommelin, Daan J. A.
dc.contributor.authorJiskoot, Wim
dc.date.accessioned2013-02-27T16:54:31Z
dc.date.available2013-02-27T16:54:31Z
dc.date.issued2005-11
dc.description.abstractPurpose - To study the influence of protein structure on the immunogenicity in wildtype and immune tolerant mice of well-characterized degradation products of recombinant human interferon alpha2b (rhIFNα2b). Methods - RhIFNα2b was degraded by metal catalyzed oxidation (M), crosslinking with glutaraldehyde (G), oxidation with hydrogen peroxide (H) and incubation in a boiling water bath (B). The products were characterized with UV absorption, circular dichroism and fluorescence spectroscopy, gel permeation chromatography, reversed-phase HPLC, SDS-PAGE, Western blotting and mass spectrometry. The immunogenicity of the products was evaluated in wildtype mice and in transgenic mice immune tolerant for hIFNα2. Serum antibodies were detected by ELISA or surface plasmon resonance. Results - M-rhIFNα2b contained covalently aggregated rhIFNα2b with three methionines partly oxidized to methionine sulfoxides. G-rhIFNα2b contained covalent aggregates and did not show changes in secondary structure. H-rhIFNα2b was only chemically changed with four partly oxidized methionines. B-rhIFNα2b was largely unfolded and heavily aggregated. Native (N) rhIFNα2b was immunogenic in the wildtype mice but not in the transgenic mice, showing that the latter were immune tolerant for rhIFNα2b. The antirhIFNα2b antibody levels in the wildtype mice depended on the degradation product: M-rhIFNα2b > H-rhIFNα2b ~ N-rhIFNα2b >> B-rhIFNα2b; G-rhIFNα2b did not induce anti-rhIFNα2b antibodies. In the transgenic mice, only M-rhIFNα2b could break the immune tolerance. Conclusions - RhIFNα2b immunogenicity is related to its structural integrity. Moreover, the immunogenicity of aggregated rhIFNα2b depends on the structure and orientation of the constituent protein molecules and/or on the aggregate size.por
dc.identifier.citationHermeling S, Caetano LA, Damen JM, Slijper M, Schellekens H, Crommelin DJ, Jiskoot W. Structural characterization and immunogenicity in wildtype and immune tolerant mice of degraded recombinant human interferon alpha2b. In Hermerling S. Structural aspects of the immunogenicity of therapeutic proteins: transgenic animals as predictors for breaking immune tolerance [Ph. D. Tesis]. Utrecht: Utrecht University; 2005. p. 95-118.por
dc.identifier.isbn90-393-4067-6
dc.identifier.urihttp://hdl.handle.net/10400.21/2307
dc.language.isoengpor
dc.peerreviewedyespor
dc.publisherUtrecht Universitypor
dc.relation.publisherversionhttp://igitur-archive.library.uu.nl/dissertations/2005-1129-200145/c6.pdfpor
dc.subjectImmunogenicitypor
dc.subjectDegraded recombinant human interferon alpha2bpor
dc.subjectBiopharmaceuticspor
dc.titleStructural characterization and immunogenicity in wildtype and immune tolerant mice of degraded recombinant human interferon alpha2bpor
dc.typebook part
dspace.entity.typePublication
oaire.citation.endPage118por
oaire.citation.startPage95por
rcaap.rightsopenAccesspor
rcaap.typebookPartpor

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