Epitope Mapping for a Preclinical Bevacizumab (Avastin)Biosimilar on an Extended Construct of Vascular Endothelial GrowthFactor A Using Millisecond Hydrogen–Deuterium Exchange MassSpectrometry

The success of Bevacizumab (Avastin), a monoclonal antibody (mAb) anti-cancer drug targeting Vascular Endothelial Growth Factor A (VEGF-A), has motivated the development of biosimilars. Establishing target epitope similarity using epitope mapping is a critical step in pre-clinical mAb biosimilar development. Here we use Time Resolved ElectroSpray Ionization Hydrogen Deuterium Exchange Mass Spectrometry (TRESI-HDX MS) to rapidly compare the epitopes of commercial Avastin and a biosimilar in pre-clinical development (ApoBev) on an extended construct of VEGF-A. The Avastin and ApoBev epitopes determined in our experiments agree with each other and with the known epitope derived from the Avastin Fab domain / truncated VEGF co-crystal structure. However, subtly different allosteric effects observed exclusively at short (ms) HDX labeling times may reflect a slightly different binding mode for ApoBev.