Hybrid Mass Spectrometry Methods Reveal Lot-to-Lot Differences and Delineate the Effects of Glycosylation on the Tertiary Structure of Herceptin

摘要: To quantify the measurable variations in structure of a biopharmaceutical product we systematically evaluate three lots Herceptin®, two mAb standards and an intact Fc-hinge fragment. Each is examined states; glycan intact, truncated (following endoS2 treatment) fully deglycosylated. Despite equivalence at protein level, each lot Herceptin® gives distinctive signature different mass spectrometry approaches. Ion mobility (IM-MS) shows that API, attached N-glycans reduce conformational spread by 10.5–25%. Hydrogen/deuterium exchange (HDX-MS) data support this, with lower global deuterium uptake solution when comparing to deglycosylated protein. HDX-MS activated IM-MS map influence glycans on reveal allosteric effects which extend far beyond Fc domains into Fab region. Taken together, these findings supplied interactive sets establish acceptance criteria application for MS based characterisation biosimilars novel therapeutic mAbs.