RT journal article
T1 N-glycosylation profile analysis of Trastuzumab biosimilar candidates by Normal Phase Liquid Chromatography and MALDI-TOF MS approaches
A1 Sanchez-De Melo, Ivan
A1 Grassi, Paola
A1 Ochoa, Francisco
A1 Bolívar Pérez, Jorge
A1 García Cózar, Francisco José
A1 Durán Ruiz, María del Carmen
A2 Bioquímica y Biología MolecularMicrobiología, Medicina Preventiva, Salud Pública
K1 Monoclonal antibodies
K1 Biosimilars
K1 N-glycosylation
K1 MALDI-TOF MS
K1 Normal Phase Liquid
K1 Chromatography
K1 2AB fluorescent labeling, HILIC
AB The pharmaceutical market has entered an era in which the production of new therapeutics is being often replaced by “biosimilars”, copies of already commercialized products waiting for the patents to expire in order to be distributed in a more competitive and affordable manners. Due to its relevance, the ErbB2-targeted monoclonal antibody Trastuzumab (Herceptin) used as breast cancer therapy is one of the main targets in the production of biosimilars. A major challenge is to produce antibodies with the same or the closest N-glycosylation pattern seen in the commercialized drug. Several factors, such as growing conditions or cell types employed, can determine the final composition and structure of the glycans, significantly affecting the properties of the generated antibodies. Therefore, an appropriate characterization is essential. In the present study, we describe two different but complementary strategies to characterize the N-glycosylation of two biosimilar candidates of Trastuzumab. In the first case, N-glycans are fluorescently labeled and separated by Normal Phase HPLC. Different sugars will elute at different times and can be identified using specific oligosaccharide standards. In the second approach, released glycans are permethylated and analyzed by MALDI-TOF MS, being able to determine the structure because of the differential sugar masses.Biological significanceThe characterization of the N-glycosylation sites of therapeutic recombinant monoclonalantibodies (mAbs) is usually one of the most critical and time consuming steps in thedeveloping process of biosimilars or any other glycosylated drug. Herein we describe twodifferent but complementary approaches to characterize mAbs glycosylation patterns, theuse of glycan fluorescence labeling coupled to HPLC and MALDI-TOF MS profile analysis.This article is part of a Special Issue entitled: HUPO 2014.
PB Elsevier
YR 2015
FD 2015-04
LK http://hdl.handle.net/10498/20379
UL http://hdl.handle.net/10498/20379
LA eng
DS Repositorio Institucional de la Universidad de Cádiz
RD 21-sep-2026