Engineering Chinese Hamster Ovary cell recombinant protein glycosylation
department or school: Applied Science, Faculty of
department or school: Chemical and Biological Engineering, Department of
data provider: DSpace
degree campus: UBCV
creator: Gupta, Shivani
issued: 2023-01-02T23:51:40Z
issued: 2023
related degree: Doctor of Philosophy - PhD
degree grantor: University of British Columbia
description: Since 2015 more than 39 biosimilars have been approved by the FDA, with over 20 launched, providing alternatives for widely used protein therapeutics. A major challenge in biosimilar development is the genetic differences in the host cell lines used to manufacture these biologics. Many biologics approved before 2011 were expressed in murine cell lines, whereas Chinese Hamster Ovary (CHO) cells have become the preferred hosts for production. Differences between glycosylation have been identified in biologics produced using murine CHO cell line. In the case of monoclonal antibodies (mAbs), differences in glycans can significantly impact critical antibody effector function, binding activity, efficacy, and in vivo half-life. To more closely match the reference biologic (RB) glycosylation, a mAb expressing CHO cell line was genetically engineered to express mAb with murine-like glycans. This was done by selectively co-expressing two key murine enzymes, α1,3-galactosyltransferase (GGTA1) and cytidine monophosphate-N-Acetylneuraminic acid hydroxylase (CMAH). These CHO cells were then analyzed using a spectrum of analytical methods to demonstrate biosimilarity, including high-resolution mass spectrometry and biochemical and cell-based assays. In fed-batch cultures, two CHO cell clones were identified that maintained stable production while matching the glycosylation profile and function of the reference product. This study demonstrates the feasibility of engineering CHO cells to express mAbs with murine glycans, to reduce the residual uncertainty regarding biosimilarity, resulting in a higher probability of regulatory approval and potentially reduced costs and time in development. In parallel, a high-throughput CHO-specific assay was designed and evaluated to quantify N-glycosylation-related enzyme mRNA expression levels. This assay is referred to as the N-glycan enzyme mRNA expression profiler. Two different mAbs produced by two CHO cell hosts were investigated to assess t…
aggregated CHO: https://circle.library.ubc.ca/rest/handle/2429/83532
note: ENGINEERING CHINESE HAMSTER OVARY CELL RECOMBINANT PROTEIN GLYCOSYLATION by Shivani Gupta M.S. Northwestern University, 2010 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in The Faculty of Graduate and Postdoctoral Studies (Chemical and Biological Engineering) THE UNIVERSITY OF BRITISH COLUMBIA (Vancouver) December 2022 © Shivani Gupta, 2022 ii The following individuals certify that they have read, and recommend to the Faculty of Graduate and Postdoctoral Studies for acceptance, the dissertation entitled: Engineering Chinese Hamster Ovary Cell Recombinant Protein Glycosylation submitted by Shivani Gupta in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Chemical and Biological Engineering Examining Committee: James Piret, Professor, Chemical and Biological Engineering, UBC Supervisor Chetan Goudar, Vice President, Attribute Sciences, Amgen Inc. Supervisory Committee Member Stephen Withers, Professor, Department of Chemistry, UBC University Examiner Vikramaditya G. Yadav, Associate Professor, Chemical and Biological Engineering, UBC University Examiner Additional Supervisory Committee Members: Charles Haynes, Professor, Chemical and Biological Engineering, UBC Supervisory Committee Member iii Abstract Since 2015 more than 39 biosimilars have been approved by the FDA, with over 20 launched, providing alternatives for widely used protein therapeutics. A major challenge in biosimilar development is the genetic differences in the host cell lines used to manufacture these biologics. Many biologics approved before 2011 were expressed in murine cell lines, whereas Chinese Hamster Ovary (CHO) cells have become the preferred hosts for production. Differences between glycosylation have been identified in biologics produced using murine CHO cell line. In the case of monoclonal antibodies (mAbs), differences in glycans can significantly impact critical antibody effector function, binding act…
has type: Thesis/Dissertation
date issued: 2023-05
is shown at: 10.14288/1.0422924
language: eng
degree discipline: Chemical and Biological Engineering
provider: Vancouver : University of British Columbia Library
publisher: University of British Columbia
rights: Attribution-NonCommercial-NoDerivatives 4.0 International
rights URI: http://creativecommons.org/licenses/by-nc-nd/4.0/
scholar level: Graduate
contributor: Piret, James
type: Text
identifier URI: http://hdl.handle.net/2429/83532