caa a22 4500 445313226 CHVBK 20180317142637.0 cr unu---uuuuu 170323e20111001xx s 000 0 eng 10.1007/s11095-011-0451-4 doi (NATIONALLICENCE)springer-10.1007/s11095-011-0451-4 Oxidized and Aggregated Recombinant Human Interferon Beta is Immunogenic in Human Interferon Beta Transgenic Mice [Elektronische Daten] [Miranda van Beers, Melody Sauerborn, Francesca Gilli, Vera Brinks, Huub Schellekens, Wim Jiskoot] ABSTRACT: Purpose: To study the effect of oxidation on the structure of recombinant human interferon beta-1a (rhIFNβ-1a) and its immunogenicity in wild-type and immune-tolerant transgenic mice. Methods: Untreated rhIFNβ-1a was degraded by metal-catalyzed oxidation, H2O2-mediated oxidation, and guanidine-mediated unfolding/refolding. Four rhIFNβ-1a preparations with different levels of oxidation and aggregation were injected intraperitoneally in mice 15× during 3weeks. Both binding and neutralizing antibodies were measured. Results: All rhIFNβ-1a preparations contained substantial amounts of aggregates. Metal-catalyzed oxidized rhIFNβ-1a contained high levels of covalent aggregates as compared with untreated rhIFNβ-1a. H2O2-treated rhIFNβ-1a showed an increase in oligomer and unrecovered protein content by HP-SEC; RP-HPLC revealed protein oxidation. Guanidine-treated rhIFNβ-1a mostly consisted of dimers and oligomers and some non-covalent aggregates smaller in size than those in untreated rhIFNβ-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 H2O2-treated rhIFNβ-1a as compared to untreated and guanidine-treated rhIFNβ-1a. Conclusions: Oxidation-mediated aggregation increased the immunogenicity of rhIFNβ-1a in transgenic mice, whereas aggregated preparations devoid of measurable oxidation levels were hardly immunogenic. The Author(s), 2011 antibodies nationallicence immunogenicity nationallicence oxidation nationallicence protein aggregates nationallicence recombinant human interferon beta nationallicence BABs : binding antibodies nationallicence hIFNβ : human interferon beta nationallicence i.p. : intraperitoneally nationallicence IgG : immunoglobulin G nationallicence MxA : myxovirus resistance protein A nationallicence NABs : neutralizing antibodies nationallicence rhIFNα-2b : recombinant human interferon alpha-2b nationallicence rhIFNβ : recombinant human interferon beta nationallicence rhIFNβ-1a : recombinant human interferon beta-1a nationallicence rhIFNβ-1b : recombinant human interferon beta-1b nationallicence SDS : sodium dodecyl sulfate nationallicence TRU/ml : ten-fold reduction units per ml nationallicence van Beers Miranda Division of Drug Delivery Technology, Leiden/Amsterdam Center for Drug Research (LACDR), Leiden University, Einsteinweg 55, 2333 CC, Leiden, The Netherlands aut Sauerborn Melody Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands aut Gilli Francesca Clinical Neurobiology Unit, Neuroscience Institute Cavalieri Ottolenghi (NICO), University Hospital San Luigi Gonzaga, Orbassano, TO, Italy aut Brinks Vera Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands aut Schellekens Huub Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands aut Jiskoot Wim Division of Drug Delivery Technology, Leiden/Amsterdam Center for Drug Research (LACDR), Leiden University, Einsteinweg 55, 2333 CC, Leiden, The Netherlands aut Pharmaceutical Research Springer US; http://www.springer-ny.com 28/10(2011-10-01), 2393-2402 0724-8741 28:10<2393 2011 28 11095 https://doi.org/10.1007/s11095-011-0451-4 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s11095-011-0451-4 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- van Beers Miranda Division of Drug Delivery Technology, Leiden/Amsterdam Center for Drug Research (LACDR), Leiden University, Einsteinweg 55, 2333 CC, Leiden, The Netherlands aut NATIONALLICENCE 700 1- Sauerborn Melody Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands aut NATIONALLICENCE 700 1- Gilli Francesca Clinical Neurobiology Unit, Neuroscience Institute Cavalieri Ottolenghi (NICO), University Hospital San Luigi Gonzaga, Orbassano, TO, Italy aut NATIONALLICENCE 700 1- Brinks Vera Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands aut NATIONALLICENCE 700 1- Schellekens Huub Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands aut NATIONALLICENCE 700 1- Jiskoot Wim Division of Drug Delivery Technology, Leiden/Amsterdam Center for Drug Research (LACDR), Leiden University, Einsteinweg 55, 2333 CC, Leiden, The Netherlands aut NATIONALLICENCE 773 0- Pharmaceutical Research Springer US; http://www.springer-ny.com 28/10(2011-10-01), 2393-2402 0724-8741 28:10<2393 2011 28 11095 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer