on 11-22-201306:35 AM - edited on 10-15-202107:45 AM by AnalyteGuru
Samonig M, Huber C, Scheffler K. Application Note 591 Monoclonal antibodies (mAbs) are one of the fastest growing classes of pharmaceutical products. They play a major role in the treatment of a variety of conditions such as cancer, infectious diseases, allergies, inflammation, and auto-immune diseases. Because mAbs can exhibit significant heterogeneity, extensive analytical characterization is required to obtain approval for a new mAb as a therapeutic product. Mass spectrometry has become an essential tool in the characterization of mAbs, providing molecular weight determinations of intact proteins as well as separated light and heavy chains, elucidation of glycosylation and glycan structures, confirmation of correct amino acid sequences, and identification of impurities such as host cell proteins (HCP) inherent to the production process.
Rituximab, which is known under the trade names Rituxan® (Biogen Idec/Genentech) in the United States and MabThera® (Roche) in Europe, is a recombinantly
produced, monoclonal chimeric antibody against the protein CD20. It was one of the first new generation drugs in cancer immune therapy. Rituximab was approved by the U.S. Food and Drug Administration in 1997 and by the European Commission in 1998 for cancer therapy of malignant lymphomas. The variable domain of the antibody targets the cell surface molecule CD20, that can be found in some non-Hodgkin lymphomas. In this application note, the capabilities and performance of the Q Exactive benchtop Orbitrap mass spectrometer in analyzing the intact and reduced forms of rituximab are demonstrated as well as sequence confirmation analyses using a combined top-down and bottom-up approach.
Furthermore, the sensitivity of two chromatographic setups using monolithic columns coupled online to the mass spectrometer is evaluated. The data obtained demonstrate superior resolution and mass accuracy of the Q Exactive mass spectrometer and present it as a high confidence screening tool for accelerated and accurate biopharmaceutical product development and characterization.
Division of Chemistry and Bioanalytics, University of Salzburg, Salzburg, Austria; Thermo Fisher Scientific, Dreieich, Germany