Investigation of the Charge Variant Profile of Non-cleavable Conjugated Antibodies

This research aimed to analyze the charge variant profiles of monoclonal antibodies (mAbs) conjugated to maytansine derivatives or tomaymycin compounds via a non-cleavable linker, utilizing the imaging capillary isoelectric focusing (icIEF) technique. Initially, the charge variant profiles for three mAbs were assessed, revealing both major and minor variants among the samples. Specifically, mAB1 and mAB2 displayed two distinct charge variants with isoelectric points (pI) of 9.00 and 8.95, respectively. In contrast, mAB3 showed a predominant charge variant with a pI of 8.50, along with two minor variants (pI values of 8.30 and 8.60). The conjugation of mAB1 involved a maytansine derivative using non-cleavable linkers, while mAB2 and mAB3 were linked to tomaymycin molecules. The resulting non-cleavable antibody conjugates exhibited greater heterogeneity and acidity than their unconjugated forms. The observed pI ranges were 7.4 to 8.9 for mAB1 conjugates (∆pI: 1.4), 8.2 to 8.9 for mAB2 (∆pI: 0.7), and 7.4 to 8.4 for mAB3 (∆pI: 1). The icIEF method proved effective for tracking the charge profiles of antibody-drug conjugates (ADCs), showing excellent repeatability for both intra-day and inter-day measurements for unconjugated mAbs and ADCs.