Method Validation for Determination of Charge Variants in Adalimumab by Ion-Exchange Chromatography
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Abstract
Background: Adalimumab, fully human recombinant immunoglobulin G1, is a highly specific monoclonal antibody that binds to Tumor necrosis factor-alpha (TNF-α) and prevents its interaction with receptors. Currently, all versions of adalimumab both originator and biosimilar, are widely used to treat rheumatoid arthritis and other inflammatory diseases. During its commercial manufacturing, adalimumab is produced by biotechnological techniques that may change the charge variants on biomolecules. These modifications may impact on biological activity, immunogenicity, and stability. Consequently, the quality control of these drugs is significant in order to ensure that patients receive quality drugs according to the international standard. Characterization of protein charge variants using ion-exchange chromatography, including ionic strength condition or pH gradient mode, is useful for qualitative and quantitative analysis.
Objectives: This study aimed to validate a method for the determination of charge variants in adalimumab by ion-exchange chromatography according to the International Conference on Harmonization (ICH) guidelines and to be used as a standard protocol in laboratory quality control of marketed adalimumab prior to release for supply in the country.
Methods: Validation for the determination of charge isoforms in adalimumab was performed by a pH-gradient cation-exchange chromatography with testing parameters including specificity, precision, accuracy, linearity, limit of quantitation, limit of detection, and robustness.
Results: The results indicated that the method was specific for adalimumab determination with linearity over the concentration range of 1.00 to 10.00 mg/ml. The percentage of relative standard deviations of repeatability and intermediate precision were less than 5%. The percentage of recovery in the accuracy range was 90% to 110%, while the limits of quantitation and detection were 2.50 and 0.70 mg/ml, respectively. The method demonstrates robustness when changing the shelf life of the mobile phase.
Conclusions: The results of the study suggested that this method had specificity, precision, accuracy, linearity, and robustness. Hence, it is suitable for a standard method in quality control of adalimumab products for registration purposes in Thailand.
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