Global characterization of modifications to the charge isomers of IgG antibody

Xinling Cui; Wei Mi; Zhishang Hu; Xiaoyu Li; Bo Meng; Xinyuan Zhao; Xiaohong Qian; Tao Zhu; Wantao Ying

doi:10.1016/j.jpha.2020.11.006

Volume 12 Issue 1

Feb. 2022

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Article Navigation > Journal of Pharmaceutical Analysis > 2022 > 12(1): 156-163

Xinling Cui, Wei Mi, Zhishang Hu, Xiaoyu Li, Bo Meng, Xinyuan Zhao, Xiaohong Qian, Tao Zhu, Wantao Ying. Global characterization of modifications to the charge isomers of IgG antibody[J]. Journal of Pharmaceutical Analysis, 2022, 12(1): 156-163. doi: 10.1016/j.jpha.2020.11.006

Citation:

Xinling Cui, Wei Mi, Zhishang Hu, Xiaoyu Li, Bo Meng, Xinyuan Zhao, Xiaohong Qian, Tao Zhu, Wantao Ying. Global characterization of modifications to the charge isomers of IgG antibody[J]. Journal of Pharmaceutical Analysis, 2022, 12(1): 156-163. doi: 10.1016/j.jpha.2020.11.006

Citation:

Xinling Cui, Wei Mi, Zhishang Hu, Xiaoyu Li, Bo Meng, Xinyuan Zhao, Xiaohong Qian, Tao Zhu, Wantao Ying. Global characterization of modifications to the charge isomers of IgG antibody[J]. Journal of Pharmaceutical Analysis, 2022, 12(1): 156-163. doi: 10.1016/j.jpha.2020.11.006

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Global characterization of modifications to the charge isomers of IgG antibody

doi: 10.1016/j.jpha.2020.11.006

a. State Key Laboratory of Proteomics, Beijing Proteome Research Center, National Center for Protein Sciences (Beijing), Beijing Institute of Lifeomics, Beijing, China;
b. National Institute of Metrology, Center for Advanced Measurement Science, Beijing, China;
c. National Institute of Metrology, Chemical Metrology & Analytical Science Division, Beijing, China;
d. CanSino Bio Corporation Tianjin Key Laboratory of Recombinant Bacterial Recombination and Bacterial Vaccines, Tianjin, China

Funds:

We are grateful for the financial support from the National Key Program for Basic Research of China (Grant Nos.: 2018YFC0910302 and 2017YFF0205400), the National Natural Science Foundation of China (Grant No.: 81530021), and Innovation Foundation of Medicine (Grant Nos.: BWS14J052 and 16CXZ027).

Received Date: Mar. 12, 2020
Accepted Date: Nov. 17, 2020
Rev Recd Date: Nov. 08, 2020
Publish Date: Nov. 25, 2020

Abstract

Abstract

Posttranslational modifications of antibody products affect their stability, charge distribution, and drug activity and are thus a critical quality attribute. The comprehensive mapping of antibody modifications and different charge isomers (CIs) is of utmost importance, but is challenging. We intended to quantitatively characterize the posttranslational modification status of CIs of antibody drugs and explore the impact of posttranslational modifications on charge heterogeneity. The CIs of antibodies were fractionated by strong cation exchange chromatography and verified by capillary isoelectric focusing-whole column imaging detection, followed by stepwise structural characterization at three levels. First, the differences between CIs were explored at the intact protein level using a top-down mass spectrometry approach; this showed differences in glycoforms and deamidation status. Second, at the peptide level, common modifications of oxidation, deamidation, and glycosylation were identified. Peptide mapping showed nonuniform deamidation and glycoform distribution among CIs. In total, 10 N-glycoforms were detected by peptide mapping. Finally, an in-depth analysis of glycan variants of CIs was performed through the detection of enriched glycopeptides. Qualitative and quantitative analyses demonstrated the dynamics of 24 N-glycoforms. The results revealed that sialic acid modification is a critical factor accounting for charge heterogeneity, which is otherwise missed in peptide mapping and intact molecular weight analyses. This study demonstrated the importance of the comprehensive analyses of antibody CIs and provides a reference method for the quality control of biopharmaceutical analysis.
- Antibody,
- Charge isomers,
- Mass spectrometry,
- Posttranslational modification,
- Glycopeptide

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References(37)

References

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