Investigating the Degradation Behaviors of a Therapeutic Monoclonal Antibody Associated with pH and Buffer Species

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• 作者：Songyan Zheng ; Difei Qiu ; Monica Adams ; Jinjiang Li ; Rao V. Mantri…
• 关键词：KEY WORDSaggregation ; conformational stability ; fragmentation ; monoclonal antibody (mAb) ; protein stability ; thermal stability
• 刊名：AAPS PharmSciTech
• 出版年：2017
• 出版时间：January 2017
• 年：2017
• 卷：18
• 期：1
• 页码：42-48
• 全文大小：783KB
• 刊物主题：Pharmacology/Toxicology; Biotechnology; Biochemistry, general; Pharmacy;
• 出版者：Springer US
• ISSN：1530-9932
• 卷排序：18

文摘

This study aimed in understanding the degradation behaviors of an IgG 1 subtype therapeutic monoclonal antibody A (mAb-A) associated with pH and buffer species. The information obtained in this study can augment conventional, stability-based screening paradigms by providing the direction necessary for efficient experimental design. Differential scanning calorimetry (DSC) was used for studying conformational stability. Dynamic light scattering (DLS) was utilized to generate B<sub>22sub>*, a modified second virial coefficient for the character of protein-protein interaction. Size-exclusion chromatography (SEC) and hydrophobic interaction chromatography (HIC) were employed to separate degradation products. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used for determining the molecular size and liquid chromatography mass spectrometry (LC-MS) were used for identifying the sequence of the separated fragments. The results showed that both pH and buffer species played the roles in controlling the degradation behaviors of mAb-A, but the pH was more significant. In particular, pH 4.5 induced additional thermal transition peaks occurring at a low temperature compared with pH 6.5. A continual temperature-stress study illustrated that the additional thermal transition peaks related to the least stable structure and a greater fragmentation. Although mAb-A showed the comparable conformational structures and an identical amount of aggregates at time zero between the different types of buffer species at pH 6.5, the aggregation formation rate showed a buffer species-dependent discrepancy over a temperature-stress period. It was found that the levels of aggregations associated with the magnitudes of protein-protein interaction forces.