LC-MS analysis combined with principal component analysis and soft independent modelling by class analogy for a better detection of changes in N-glycosylation profiles of therapeutic glycoproteins

详细信息    查看全文

• 作者：Ana Planinc ; Bieke Dejaegher ; Yvan Vander Heyden&#8230
• 关键词：Therapeutic glycoproteins ; N ; Glycans ; Principal component analysis (PCA) ; Soft independent modelling by class analogy (SIMCA) ; High ; resolution mass spectrometry (HRMS) ; Hydrophilic interaction liquid chromatography (HILIC)
• 刊名：Analytical and Bioanalytical Chemistry
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：409
• 期：2
• 页码：477-485
• 全文大小：
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Analytical Chemistry; Biochemistry, general; Laboratory Medicine; Characterization and Evaluation of Materials; Food Science; Monitoring/Environmental Analysis;
• 出版者：Springer Berlin Heidelberg
• ISSN：1618-2650
• 卷排序：409

文摘

Therapeutic proteins are among the top selling drugs in the pharmaceutical industry. More than 60 % of the approved therapeutic proteins are glycosylated. Nowadays, it is well accepted that changes in glycosylation may affect the safety and the efficacy of the therapeutic proteins. For this reason, it is important to characterize both the protein and the glycan structures. In this study, analytical and data processing methods were developed ensuring an easier characterization of glycoprofiles. N-glycans were (i) enzymatically released using peptide-N-glycosidase F (PNGase F), (ii) reduced, and (iii) analyzed by hydrophilic interaction liquid chromatography coupled to a high-resolution mass spectrometer (HILIC-HRMS). Glycosylation changes were analyzed in human plasma immunoglobulin G samples which had previously been artificially modified by adding other glycoproteins (such as ribonuclease B and fetuin) or by digesting with enzyme (neuraminidase). Principal component analysis (PCA) and classification through soft independent modelling by class analogy (SIMCA) were used to detect minor glycosylation changes. Using HILIC-MS-PCA/SIMCA approach, it was possible to detect small changes in N-glycosylation, which had not been detected directly from the extracted-ion chromatograms, which is current technique to detect N-glycosylation changes in batch-to-batch analysis. The HILIC-MS-PCA/SIMCA approach is highly sensitive approach due to the sensitivity of MS and appropriate data processing approaches. It could help in assessing the changes in glycosylation, controlling batch-to-batch consistency, and establishing acceptance limits according to the glycosylation changes, ensuring safety and efficacy.