Investigation of the Immunogenicity of Different Types of Aggregates of a Murine Monoclonal Antibody in Mice

• Published in: Pharmaceutical research Vol. 32; no. 2; pp. 430 - 444
• Main Authors: Freitag, Angelika J., Shomali, Maliheh, Michalakis, Stylianos, Biel, Martin, Siedler, Michael, Kaymakcalan, Zehra, Carpenter, John F., Randolph, Theodore W., Winter, Gerhard, Engert, Julia
• Format: Journal Article
• Language: English
• Published: Boston Springer US 01.02.2015; Springer Nature B.V
• Subjects: Aggregates; Animals; Antibodies, Monoclonal - genetics; Antibodies, Monoclonal - immunology; Antibody Formation - drug effects; Antibody Formation - immunology; Biochemistry; Biomedical and Life Sciences; Biomedical Engineering and Bioengineering; Biomedicine; Female; Immunogenetic Phenomena - drug effects; Immunogenetic Phenomena - immunology; Immunoglobulin G - genetics; Immunoglobulin G - immunology; Immunology; Medical Law; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Monoclonal antibodies; Pharmaceutical sciences; Pharmacology/Toxicology; Pharmacy; Proteins; Research Paper; Rodents; Monoclonal antibody; Immunogenicity; Wild-type mice; Protein aggregates; Protein particles
• ISSN: 0724-8741; 1573-904X
• DOI: 10.1007/s11095-014-1472-6

Purpose The potential contribution of protein aggregates to the unwanted immunogenicity of protein pharmaceuticals is a major concern. In the present study a murine monoclonal antibody was utilized to study the immunogenicity of different types of aggregates in mice. Samples containing defined types of aggregates were prepared by processes such as stirring, agitation, exposure to ultraviolet (UV) light and exposure to elevated temperatures. Methods Aggregates were analyzed by size-exclusion chromatography, light obscuration, turbidimetry, infrared (IR) spectroscopy and UV spectroscopy. Samples were separated into fractions based on aggregate size by asymmetrical flow field-flow fractionation or by centrifugation. Samples containing different types and sizes of aggregates were subsequently administered to C57BL/6 J and BALB/c mice, and serum was analyzed for the presence of anti-IgG1, anti-IgG2a, anti-IgG2b and anti-IgG3 antibodies. In addition, the pharmacokinetic profile of the murine antibody was investigated. Results In this study, samples containing high numbers of different types of aggregates were administered in order to challenge the in vivo system. The magnitude of immune response depends on the nature of the aggregates. The most immunogenic aggregates were of relatively large and insoluble nature, with perturbed, non-native structures. Conclusion This study shows that not all protein drug aggregates are equally immunogenic.