Monoclonal antibodies (mAbs) are made from highly uniform immune cells, which are clones of a single parent cell. mAbs have monovalent affinity because they bind to the same epitope (the site where the antibody recognizes the antigen). Almost any substance can produce mAbs with binding specificity, and then they can be used to detect or purify the substance.
In 1975, Kohler and Milstein created the in vitro hybridoma technology and obtained murine mAbs, opening a new era of monoclonal antibody technology. With the development of molecular biology, antibody library, and transgenic technologies, mAbs have evolved several stages: chimeric, humanized, and whole human monoclonal antibodies. mAbs have been widely used in the treatment of diseases because of their specificity, homogeneity, and mass production ability. Therapeutic monoclonal antibodies work through a variety of mechanisms, such as blocking the function of the target molecule, inducing cell apoptosis (cells that express the target molecule), or regulating signaling pathways.
Monoclonal antibody drugs are one of the fastest growing areas in biopharmaceutical R&D, with a compound annual growth rate of more than 40%. Due to the high price and high profit of monoclonal antibody drugs, the development of monoclonal antibody drugs is still an important driving force for the development of the biopharmaceutical industry. At least 570 therapeutic mAbs are undergoing clinical trials worldwide. As of December 2019, the US FDA has approved 79 therapeutic monoclonal antibodies. Between 2008 and 2017, 48 mAbs were approved. As of the end of 2017, 61 mAbs are available for clinical use. According to the official, 18 new antibodies were approved between 2018 and 2019. In the past 25 years, mAbs have become the main treatment for many diseases and can be used to treat tumors, autoimmune diseases, infectious diseases, and transplant rejection. With the advancement of technologies, mAbs discovery and development are faster and more efficient.
Every monoclonal antibody drug needs to undergo extensive and detailed characterization studies to obtain the approval required for clinical trials and finally reaching the market. The European Medicines Agency (EMA) issued a regulatory document on mAb characterization in July 2009, stating that researchers should study the characterization of monoclonal antibodies in detail. The monoclonal antibody characterization should include the determination of the physical and chemical and structural properties, purity, impurities, and other aspects of the mAb according to the International Conference on Harmonization (ICH) guide Q6B(3).
ICH guide Q6B provides a set of internationally recognized principles for the characterization of biotechnology products to support the market application of monoclonal antibodies. The document suggests that all monoclonal antibody drugs should undergo the following analyses:
| Service categories | Specific analysis items |
|---|---|
| Primary structure analysis | Complete molecular weight |
| Desugared molecular weight | |
| Peptide coverage | |
| N/C terminal amino acid sequence analysis | |
| Peptide mapping | |
| Advanced structural analysis | Disulfide bond localization |
| Nuclear magnetic resonance | |
| Thermal stability | |
| Circular dichroism | |
| Protein modification analysis | Oxidation/Deamidation |
| N-terminal analysis | |
| C-terminal analysis | |
| Phosphorylation, acetylation, glycosylation | |
| Glycosylation site | |
| N-glycan profile analysis | |
| Impurity analysis | Purity |
| Charge heterogeneity | |
| Host cell protein residue | |
| Protein A residue | |
| Residual amount of foreign DNA |
