Pharmaceutica Analytica Acta

Multiplex analysis of mAb aggregates and charge variants by size exclusion-cation exchange chromatography coupled to mass spectrometry

23^rd Global summit on Analytical and Bioanalytical Techniques

September 12-13, 2022 | Webinar

Giulia Lambiase

BioPharmaceuticals Development, R&D, AstraZeneca, Cambridge, UK

Scientific Tracks Abstracts: Pharm Anal Acta

Abstract:

Monoclonal antibodies (mAbs) are extremely complex due to the presence of structural modifications resulting from enzymatic and chemical reactions such as glycosylation, glycation, deamidation, isomerisation, oxidation, aggregation and fragmentation. Size and charge variants analysis are carried out from the early stages of drug development throughout product lifetime to investigate product degradation pathways and optimize process conditions. However, conventional analytical workstreams for size and charge variant characterization are both time and sample demanding, requiring the application of multiple analytical methods. This study presents the development of a novel 2D-LC/MS approach combining both aggregate and charge variant profiling of a mAb candidate in a single method. Aggregate quantification was performed in the first dimension (1D) by size exclusion chromatography SEC, followed by online fraction transfer of the monomer peak to the second dimension (2D) by a heart-cutting for charge variant analysis by cation exchange chromatography (CEX). Aiming to maximise the information obtained from minimal sample and time required for analysis, a salt-based separation with UV detection was developed for supporting the processing of a large number of samples to facilitate high-throughput process development (HTPD). In addition, a mass spectrometry (MS) compatible SEC-CEX separation was developed enabling online charge variant peak identification. This study presented the ability to multiplex mAb size and charge variants analysis by coupling SEC with CEX in a 2D-LC set-up. To date, this is the first 2D SEC-CEX-UV(-MS) application for intact mAb analysis.

Biography :

Giulia Lambiase holds an MSc in Pharmacy from the University of Bologna and a PhD in Chemical and Biological Engineering from the University of Sheffield in partnership with AstraZeneca. Giulia has over 6 years experience in advanced analytical methodologies and chemometric tools applied to protein drug analysis with major focus on chromatography and mass spectrometry methods, multivariate analysis and statistical experimental design. She is now a Senior Scientist in the Analytical Sciences team at AstraZeneca. Outcome from her PhD work led to three publications in reputed journals.