Know Your Variants and Impurities: Mass Spectrometry in Bioprocess Development
Presented by Christopher Yu, Genentech, a Member of the Roche Group
Mass spectrometry has increasingly become an indispensable tool in bioprocess development. Recent advances in instrument capability provide the required sensitivity, resolution, speed and dynamic range for detection of low level product related variants, modifications, and process related impurities. A common approach is peptide mapping with reversed phase HPLC or UHPLC, with online LC-MS/MS analysis using data dependent acquisition. This combination of two different modes of high resolution separation provides the foundation for routine detection of species present at levels below 1%.
Three specific areas of mass spectrometry application in bioprocess development are included in this presentation. The first is sequence variant analysis in support of cell line selection and cell culture process development. Low level, previously unknown amino acid substitutions due to either mutation or mistranslation are identified with the use of Mascot error tolerant search. Recent studies on analytical approaches towards improved detection coverage and the use of appropriate assay control will be presented. The second area is the characterization of product color due to oxidation and advanced glycation end products modifications on lysine and cysteine residues. These variants are highly heterogeneous in nature and are present at very low levels, both representing significant challenges in analytical technologies. Cell culture media and process conditions that yield reduced color modification have been identified. We describe in the third area recent advances in the use of mass spectrometry to complement the conventional immune assay approaches for host cell protein (HCP) analysis. Analytical methodology is presented for both the identification and quantification of HCP to support the optimization of purification process.
In summary, we demonstrate that mass spectrometry is a critical analytical tool for fast and efficient bioprocess development of safe, consistent, and high quality products.