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Physicochemical and functional comparability between the proposed biosimilar rituximab GP2013 and originator rituximab.

ABSTRACT: Regulatory approval for a biosimilar product is provided on the basis of its comparability to an originator product. A thorough physicochemical and functional comparability exercise is a key element in demonstrating biosimilarity. Here we report the characterization of a proposed biosimilar rituximab (GP2013) and originator rituximab.To compare GP2013 with originator rituximab using an extensive array of routine analytical and extended characterization methods.Primary and higher order protein structures were analyzed using a variety of methods that included high-performance liquid chromatography electrospray ionization mass spectrometry (HPLC-ESI-MS), peptide mapping with UV and MS detection, circular dichroism (CD), Fourier transform infrared (FTIR) spectroscopy, hydrogen deuterium exchange (HDX) MS, 1D (1)H nuclear magnetic resonance (NMR) spectroscopy, X-ray crystallography and differential scanning calorimetry (DSC). Charge and amino acid modifications were assessed using cation exchange chromatography (CEX) and peptide mapping using reversed-phase (RP) HPLC. Boronate affinity chromatography was used to determine the relative amount of glycation. Glycans were identified and quantified after 2-aminobenzamide (2-AB) labeling and separation using normal phase HPLC with fluorescence and MS detection, respectively. Glycan site occupancy was determined using reducing capillary electrophoresis with sodium dodecyl sulfate (CE-SDS). Size heterogeneity was determined using reducing and non-reducing CE-SDS, size exclusion chromatography (SEC) and asymmetric flow field flow fractionation (AF4). Biological characterization included a series of bioassays (in vitro target binding, antibody-dependent cell-mediated cytotoxicity [ADCC], complement-dependent cytotoxicity [CDC] and apoptosis) and surface plasmon resonance (SPR) Fc receptor binding assays.Intact mass analysis of GP2013 and the heavy and light chains using RP HPLC-ESI-MS revealed the expected molecular mass of rituximab. The amino acid sequence was shown to be identical between GP2013 and the originator rituximab. Further sequence confirmation using RP-HPLC-UV/MS peptide mapping showed non-distinguishable chromatograms for Lys-C digested GP2013 and originator rituximab. The higher order structure of GP2013 was shown to be indistinguishable from originator rituximab using a large panel of redundant and orthogonal methods. GP2013 and originator rituximab were comparable with regard to charge variants, specific amino acid modifications and the glycan pattern. GP2013 was also shown to have similar purity, aggregate and particle levels when compared with the originator. Functionally, and by using a comprehensive set of bioassays and binding assays covering a broad range of rituximab's functional activities, GP2013 could not be distinguished from originator rituximab.GP2013 was shown to be physicochemically highly similar to originator rituximab at the level of primary and higher order structure, post-translational modifications and size variants. An extensive functional characterization package indicated that GP2013 has the same biological properties as originator rituximab.

SUBMITTER: Visser J

PROVIDER: S-EPMC3775154 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature

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Physicochemical and functional comparability between the proposed biosimilar rituximab GP2013 and originator rituximab.

Visser Jan J Feuerstein Isabel I Stangler Thomas T Schmiederer Timo T Fritsch Cornelius C Schiestl Martin M

BioDrugs : clinical immunotherapeutics, biopharmaceuticals and gene therapy 20131001 5

<h4>Background</h4>Regulatory approval for a biosimilar product is provided on the basis of its comparability to an originator product. A thorough physicochemical and functional comparability exercise is a key element in demonstrating biosimilarity. Here we report the characterization of a proposed biosimilar rituximab (GP2013) and originator rituximab.<h4>Objective</h4>To compare GP2013 with originator rituximab using an extensive array of routine analytical and extended characterization method ...[more]

PMID: 23649935

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Project description:Development of bio-therapeutics has exhibited exponential growth in China over the past decade. However, no biosimilar drug has been approved in China (CN) due to the lack of a national biosimilar regulatory guidance. HLX01, a rituximab biosimilar developed in China under European Medicines Agency biosimilar guidelines and requirements, was the first such drug submitted for regulatory review in China, and it is expected to receive approval there as a biosimilar product. To demonstrate the analytical similarities of HLX01, CN-rituximab (sourced in China but manufactured in Europe) and EU-rituximab (sourced and manufactured in Europe), an extensive 3-way physicochemical and functional similarity assessment using a series of orthogonal and state-of-the-art techniques was conducted, following the similarity requirement guidelines recently published by China's Center for Drug Evaluation. The results of the similarity study showed an identical protein amino acid sequence and highly similar primary structures between HLX01 and the reference product (RP) MabThera®, along with high similarities in higher order structures, potency, integrity, purity and impurity profiles, biological and immunological binding functions, as well as degradation behaviors under stress conditions. In addition, HLX01 presented slightly lower aggregates and better photostability compared with the RP. Despite slight changes in relative abundance of glycan moieties and heavy chain C-terminal lysine modification, no differences in biological activities and immunological properties were observed between the RP and HLX01. In conclusion, HLX01 is highly similar to CN- and EU-sourced RP in terms of physicochemical properties and biological activities, suggesting similar product quality, efﬁcacy, and safety. The regulatory requirements interpreted and applied towards the HLX01 marketing application sets a precedent for analytical similarity assessment of biosimilar products in China.

Dataset Information

Physicochemical and functional comparability between the proposed biosimilar rituximab GP2013 and originator rituximab.

Publications

Physicochemical and functional comparability between the proposed biosimilar rituximab GP2013 and originator rituximab.

Similar Datasets

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