Evaluation of capillary zone electrophoresis for charge heterogeneity testing of monoclonal antibodies

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2015-03, Vol.983-984, p.101-110
• Main Authors: Moritz, Bernd, Schnaible, Volker, Kiessig, Steffen, Heyne, Andrea, Wild, Markus, Finkler, Christof, Christians, Stefan, Mueller, Kerstin, Zhang, Li, Furuya, Kenji, Hassel, Marc, Hamm, Melissa, Rustandi, Richard, He, Yan, Solano, Oscar Salas, Whitmore, Colin, Park, Sung Ae, Hansen, Dietmar, Santos, Marcia, Lies, Mark
• Format: Article
• Language: English
• Subjects: Analysis of variance; Antibodies, Monoclonal - analysis; Antibodies, Monoclonal - chemistry; Antibody; Capillarity; Capillary zone electrophoresis; Charge; Chromatography; Chromatography, Ion Exchange; Electrophoresis, Capillary - methods; Heterogeneity; High throughput; Hydrogen-Ion Concentration; Intercompany study; Isoelectric Focusing; Pharmaceuticals; Reproducibility; Reproducibility of Results; Separation; Statistical analysis; Statistical methods
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2014.12.024

•CZE is a robust method for charge heterogeneity testing of biopharmaceuticals.•An intercompany study delivered precise results without prior training.•Implementation of CZE is very easy and allows high throughput applications.•CZE is suitable for use in a GMP environment of pharmaceutical industry. Within pharmaceutical industry charge heterogeneity testing of biopharmaceuticals has to be reproducible and fast. It should pass method validation according to ICH Q2. Classical approaches for the analysis of the charge heterogeneity of biopharmaceuticals are ion exchange chromatography (IEC) and isoelectric focusing (IEF). As an alternative approach, also capillary zone electrophoresis (CZE) was expected to allow reliable charge heterogeneity profiling by separation according to the analyte's net charge and hydrodynamic radius. Aim of this study was to assess if CZE possesses all of the required features. Therefore, beside lab internal validation of this method also an international cross company study was organized. It was shown that CZE is applicable across a broad pI range between 7.4 and 9.5. The coefficient of correlation was above 0.99 which demonstrated linearity. Precision by repeatability was around 1% (maximum relative standard deviation per level) and accuracy by recovery was around 100% (mean recovery per level). Accuracy was further verified by direct comparison of IEC, IEF and CZE, which in this case showed comparable %CPA results for all three methods. However, best resolution for the investigated MAb was obtained with CZE. In dependence on sample concentration the detection limit was between 1 and 3%. Within the intercompany study for CZE the same stressed and non-stressed samples were analyzed in each of the 11 participating labs. The finally obtained dataset contained more than 1000 separations which provided an extended dataset for further statistical evaluation. Among the different labs no significant differences between the peak profiles were observed. Mean driver for dropouts in quantitative evaluation was linked to the performance of some participating labs while the impact of the method performance was negligible. In comparison to a 50cm capillary there was a slightly better separation of impurities and drug substance related compounds with a 30cm capillary which demonstrates that an increased stability indicating potential can be combined with the increased separation velocity and high throughput capability of a shorter capillary. Sep