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Efficient clinical-grade γ-retroviral vector purification by high-speed centrifugation for CAR T cell manufacturing

Mekkaoui, Leila; Tejerizo, Jose G; Abreu, Sara; Rubat, Lydie; Nikoniuk, Aleksandra; Macmorland, William; Horlock, Claire; ... Pule, Martin; + view all (2023) Efficient clinical-grade γ-retroviral vector purification by high-speed centrifugation for CAR T cell manufacturing. Molecular Therapy - Methods and Clinical Development , 28 pp. 116-128. 10.1016/j.omtm.2022.12.006. Green open access

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Abstract

γ-Retroviral vectors (γ-RV) are powerful tools for gene therapy applications. Current clinical vectors are produced from stable producer cell lines which require minimal further downstream processing, while purification schemes for γ-RV produced by transient transfection have not been thoroughly investigated. We aimed to develop a method to purify transiently produced γ-RV for early clinical studies. Here, we report a simple one-step purification method by high-speed centrifugation for γ-RV produced by transient transfection for clinical application. High-speed centrifugation enabled the concentration of viral titers in the range of 107-108 TU/mL with >80% overall recovery. Analysis of research-grade concentrated vector revealed sufficient reduction in product- and process-related impurities. Furthermore, product characterization of clinical-grade γ-RV by BioReliance demonstrated two-logs lower impurities per transducing unit compared with regulatory authority-approved stable producer cell line vector for clinical application. In terms of CAR T cell manufacturing, clinical-grade γ-RV produced by transient transfection and purified by high-speed centrifugation was similar to γ-RV produced from a clinical-grade stable producer cell line. This method will be of value for studies using γ-RV to bridge vector supply between early- and late-stage clinical trials.

Type: Article
Title: Efficient clinical-grade γ-retroviral vector purification by high-speed centrifugation for CAR T cell manufacturing
Location: United States
Open access status: An open access version is available from UCL Discovery
DOI: 10.1016/j.omtm.2022.12.006
Publisher version: https://doi.org/10.1016/j.omtm.2022.12.006
Language: English
Additional information: © 2022 The Authors. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
Keywords: CAR T cells, downstream processing, high-speed centrifugation, manufacturing, γ-retroviral vectors
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Cancer Institute
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Cancer Institute > Research Department of Haematology
URI: https://discovery.ucl.ac.uk/id/eprint/10163093
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