title: Comparability study of Rituximab originator and follow-on biopharmaceutical.
abstract: Immunglobolin G (IgG)-based biopharmaceuticals are emerging on the pharmaceuticals market due to their high target selectivity in different diseases. In parallel, a growing interest by other companies to produce similar or highly similar follow-on biologics exits, once the patent of blockbuster biotherapeutics is about to expire. In correlation to their complex structure, an analytical challenge is facing the approval of these biosimilars. Health authorities (e.g. FDA and EMA) have issued several guidelines to define critical quality attributes during manufacturing process changes. In the current study, physicochemical characterization using state-of-the-art analytics was applied to analyse intact mass, post-translational modifications (PTMs) and higher order structure of Rituximab and one of its biosimilars. Intact mass analysis, middle-up approach as well as subunit analysis revealed similar glycoforms but additional lysine variants in the biosimilar. The N-glycosylation site was confirmed for both, the originator and the biosimilar. PTMs and higher order structure were confirmed to be similar. A special focus was given to N-glycosylation due to its potential to monitor the batch-to-batch consistency and alteration during the production bioprocess. Comparison of the N-glycosylation profiles obtained from three batches of the biosimilar and the reference product showed quantitative variations, although the N-glycans were qualitatively similar. Furthermore, a head-to-head comparability of functional properties was performed to investigate the impact of glycosylation alteration and PTMs on potency within the biosimilar batches and between originator and follow-on biodrug. The data affirm that the difference is still in the acceptable range for biosimilarity.
authors.affiliation: Institute of Pharmacy, Department of Biology, Chemistry, Pharmacy, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany; Labor für Biochemie, Department of Life Sciences & Technology, Beuth Hochschule für Technik Berlin, Seestraße 64, 13347 Berlin, Germany.
authors.forename: Othman
authors.initials: O
authors.lastname: Montacir
authors.affiliation: Institute of Pharmacy, Department of Biology, Chemistry, Pharmacy, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany.
authors.forename: Houda
authors.initials: H
authors.lastname: Montacir
authors.affiliation: Institut für Chemie und Biochemie, Department of Biology, Chemistry, Pharmacy, Freie Universität Berlin, Thielallee 63, 14195 Berlin, Germany.
authors.forename: Murat
authors.initials: M
authors.lastname: Eravci
authors.affiliation: Großgerätezentrum BioSupraMol, Department of Biology, Chemistry, Pharmacy, Freie Universität Berlin, Takustr. 3, 14195 Berlin, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Springer
authors.affiliation: Labor für Biochemie, Department of Life Sciences & Technology, Beuth Hochschule für Technik Berlin, Seestraße 64, 13347 Berlin, Germany.
authors.forename: Stephan
authors.initials: S
authors.lastname: Hinderlich
authors.affiliation: AryoGen Pharmed, Cross Tajbakhsh Street, 24th Kilometer Makhsous, Tehran, Iran.
authors.forename: Amirhossein
authors.initials: A
authors.lastname: Saadati
authors.affiliation: Institute of Pharmacy, Department of Biology, Chemistry, Pharmacy, Freie Universität Berlin, Königin-Luise-Str. 2+4, 14195 Berlin, Germany. Electronic address: maria.parr@fu-berlin.de.
authors.forename: Maria Kristina
authors.initials: MK
authors.lastname: Parr
mesh_terms: D059451:Biosimilar Pharmaceuticals; D006031:Glycosylation; D011134:Polysaccharides; D011499:Protein Processing, Post-Translational; D000069283:Rituximab / Q000494:pharmacology*
chemical_list: D059451:Biosimilar Pharmaceuticals; D011134:Polysaccharides; D000069283:Rituximab
keywords: Biopharmaceutical; Functional properties; Glycosylation; Mass spectrometry; Physicochemical characterization