Bridging size and charge variants of a therapeutic monoclonal antibody by two-dimensional liquid chromatography
| Autor: | Tapan K. Das, Zhi Chen, Duohai Pan, Kaimeng Zhou, Gurusamy Balakrishnan, Ming Zeng, Sung-Joo Park |
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| Rok vydání: | 2019 |
| Předmět: |
Anions
medicine.drug_class Clinical Biochemistry Ion chromatography Size-exclusion chromatography Pharmaceutical Science Fractionation Chemical Fractionation Monoclonal antibody 01 natural sciences Analytical Chemistry Hydrophobic effect Cations Drug Discovery medicine Spectroscopy Chromatography Ion exchange biology 010405 organic chemistry Chemistry Elution 010401 analytical chemistry Antibodies Monoclonal 0104 chemical sciences biology.protein Antibody Chromatography Liquid |
| Zdroj: | Journal of pharmaceutical and biomedical analysis. 183 |
| ISSN: | 1873-264X |
| Popis: | Monoclonal antibodies are heterogeneous in nature and may contain numerous variants with differences in size, charge, and hydrophobicity, which may impact clinical efficacy, immunogenicity, and safety. Characterization of antibody variants is necessary to build structure-function correlation and establish a proper control strategy. Isolation and enrichment of variants by conventional chromatographic peak fractionation is labor-intensive and time-consuming. The instability of fractions during isolation and subsequent characterization may also be a concern. Hence, it is desirable to analyze antibody variants in an online and real-time manner. Here we demonstrate a 2D-LC methodology - multiple heart-cutting IEC-SEC- as an investigational tool to facilitate a charge variant characterization study. Both IEC modes - anion exchange (AEX) and cation exchange (CEX) chromatography are discussed. Using this approach, direct bridging of size and charge variants of an antibody molecule was achieved without offline peak fractionation. It was observed that antibody aggregates elute late on both the AEX and CEX columns, presumably due to secondary hydrophobic interactions. Additionally, we overcame the solvent mismatch issue and developed a 2D SEC-IEC method to confirm the bridging results. This is the first reported SEC-IEC 2D-LC application for the characterization of antibody size and charge variants. |
| Databáze: | OpenAIRE |
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