Global intercompany assessment of ICIEF platform comparability for the characterization of therapeutic proteins

Abstract

An international team spanning 19 sites across 18 biopharmaceutical and in vitro diagnostics companies in the United States, Europe, and China, along with one regulatory agency, was formed to compare the precision and robustness of imaged CIEF (ICIEF) for the charge heterogeneity analysis of the National Institute of Standards and Technology (NIST) mAb and a rhPD-L1-Fc fusion protein on the iCE3 and the Maurice instruments. This information has been requested to help companies better understand how these instruments compare and how to transition ICIEF methods from iCE3 to the Maurice instrument. The different laboratories performed ICIEF on the NIST mAb and rhPD-L1-Fc with both the iCE3 and Maurice using analytical methods specifically developed for each of the molecules. After processing the electropherograms, statistical evaluation of the data was performed to determine consistencies within and between laboratory and outlying information. The apparent isoelectric point (pI) data generated, based on two-point calibration, for the main isoform of the NIST mAb showed high precision between laboratories, with RSD values of less than 0.3% on both instruments. The SDs for the NIST mAb and the rhPD-L1-Fc charged variants percent peak area values for both instruments are less than 1.02% across different laboratories. These results validate the appropriate use of both the iCE3 and Maurice for ICIEF in the biopharmaceutical industry in support of process development and regulatory submissions of biotherapeutic molecules. Further, the data comparability between the iCE3 and Maurice illustrates that the Maurice platform is a next-generation replacement for the iCE3 that provides comparable data.

Charge heterogeneity of therapeutic monoclonal antibodies by different cIEF systems: views on the current situation

“…It has been demonstrated that these differences do not affect the separation efficiency, so that the different icIEF instruments currently available provide comparable results. 16 , 17 In both systems, pI values of the sample are determined after calibration with pI-markers, assuming a linear dependence throughout the pH gradient.…”

Strategies for capillary electrophoresis: Method development and validation for pharmaceutical and biological applications—Updated and completely revised edition

“…CIEF and iCIEF have matured into techniques that are routinely applied in analytical laboratories, especially in the biopharmaceutical industry. Exemplary applications include, but are not limited to, monoclonal antibodies [192,[209][210][211][212][213][214][215], bispecific antibodies [216][217][218], ADCs [219,220], PEGylated proteins [221,222], glycoproteins, in particular erythropoietin [213,223,224], and fusion proteins [213,225]. Because this subsection is part of the former "proteins" section, which was extended to all biopharmaceuticals, it should not go unmentioned that the range of applications has been expanded beyond proteins and includes oligosaccharides [226], LNPs and related mRNA vaccines [227][228][229][230], virus-like particles [231,232], poliovirus [230,233], adeno-associated virus [230,[234][235][236][237], papillomavirus [238], and bacteria and yeast cells [230].…”

“…For iCIEF, RSD values ≤0.8% for the apparent pI values and ≤11% (outliers removed) for the percentage area were reported [209]. A recent intercompany study using two iCIEF instruments reported RSDs values ≤0.28% for the apparent pI and RSDs values ≤6.25% for the percentage area of NISTmAb [215]. Kahle et al used the NISTmAb and infliximab in a comparative study and achieved RSD values of 0.05% for the apparent pI value and 1.3% for the percentage area of the NISTmAb main peak, whereas for the infliximab main peak, the reported RSD values were 0.04% and 2.4% for pI and percentage area, respectively.…”

“…For iCIEF, RSD values ≤0.8% for the apparent p I values and ≤11% (outliers removed) for the percentage area were reported [209]. A recent intercompany study using two iCIEF instruments reported RSDs values ≤0.28% for the apparent p I and RSDs values ≤6.25% for the percentage area of NISTmAb [215]. Kahle et al.…”

Fingerprinting trimeric SARS-CoV-2 RBD by capillary isoelectric focusing with whole-column imaging detection

“…The recombinant proteins will show heterogeneity in different expression systems and under different production process conditions, contributed by the post-translational modifications (PTMs) [ 16 , 17 ]. The isoelectric point (pI) is a powerful tool to determine the presence of protein isoforms [ 18 ]. Furthermore, it is noted that the isoelectric point is one of the prime keys for understanding various biochemical properties of protein sequences [ 19 , 20 ].…”