Intact mass analysis of a therapeutic monoclonal antibody biosimilar
28th Mar 2022
Intact glycan ratio monitoring of Trastuzumab using a SCIEX X500B Mass Spectrometer
Biologics represent a growing class of pharmaceuticals being developed to target a variety of disease and illness vectors. Monoclonal antibodies (mAbs) represent the main class of these with twelve more being approved in 2020 by the FDA for cancers, viruses, and other targets. In contrast to traditional small molecule drugs, mAbs are much more complex with a large degree of heterogeneity. Some of these features such as glycan profile and accurate mass are defined as critical quality attributes (CQAs). Identifying and confirming these CQAs can play a critical role in development or production of mAbs.
Here we outline an analysis strategy to confirm the intact molecular weight and glycan assignment for a common mAb scaffold targeting the HER2 receptor. All data was collected on a SCIEX X500B Mass Spectrometer with an EXION HPLC. The chromatography was performed with a bioZen Intact XB-C8 Column (3.6 µm, 50 x 2.1 mm).
Figure 1: RAW MS of trastuzumab biosimilar. Charge states are observed between +38 and +68. See more here.
Table 1: Observed trastuzumab glycoforms masses. Reconstructed masses match previously published data. See more here.
Mass reconstruction was performed using BioPharmaView Flex software from SCIEX to match the intact masses of the protein and glycoforms. Masses were compared to expected previously published work with highly comparably masses and ratios (Agilent 5991-8445EN).
Figure 2: Reconstructed spectra of trastuzumab biosimilar. Various glycoforms have been assigned based upon accurate mass in line with previously published data. See more here.
Figure 2 shows the reconstructed mass spectrum and the assigned glycoforms. Labelled glycoforms all are present without C-terminal lysines. Intermediate peaks are unlabelled but are forms with a single lysine loss. See more here.
