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Lock, Dominik; Monjezi, Razieh; Brandes, Caroline; Bates, Stephan; Lennartz, Simon; Teppert, Karin; Gehrke, Leon; Karasakalidou-Seidt, Rafailla; Lukic, Teodora; Schmeer, Marco; Schleef, Martin; Werchau, Niels; Eyrich, Matthias; Assenmacher, Mario; Kaiser, Andrew; Prommersberger, Sabrina; Schaser, Thomas; Hudecek, Michael

Automated, scaled, transposon-based production of CAR T cells

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  • Media type: E-Article
  • Title: Automated, scaled, transposon-based production of CAR T cells
  • Contributor: Lock, Dominik; Monjezi, Razieh; Brandes, Caroline; Bates, Stephan; Lennartz, Simon; Teppert, Karin; Gehrke, Leon; Karasakalidou-Seidt, Rafailla; Lukic, Teodora; Schmeer, Marco; Schleef, Martin; Werchau, Niels; Eyrich, Matthias; Assenmacher, Mario; Kaiser, Andrew; Prommersberger, Sabrina; Schaser, Thomas; Hudecek, Michael
  • imprint: BMJ, 2022
  • Published in: Journal for ImmunoTherapy of Cancer
  • Language: English
  • DOI: 10.1136/jitc-2022-005189
  • ISSN: 2051-1426
  • Keywords: Cancer Research ; Pharmacology ; Oncology ; Molecular Medicine ; Immunology ; Immunology and Allergy
  • Origination:
  • Footnote:
  • Description: <jats:sec><jats:title>Background</jats:title><jats:p>There is an increasing demand for chimeric antigen receptor (CAR) T cell products from patients and care givers. Here, we established an automated manufacturing process for CAR T cells on the CliniMACS Prodigy platform that is scaled to provide therapeutic doses and achieves gene-transfer with virus-free Sleeping Beauty (SB) transposition.</jats:p></jats:sec><jats:sec><jats:title>Methods</jats:title><jats:p>We used an advanced CliniMACS Prodigy that is connected to an electroporator unit and performed a series of small-scale development and large-scale confirmation runs with primary human T cells. Transposition was accomplished with minicircle (MC) DNA-encoded SB100X transposase and pT2 transposon encoding a CD19 CAR.</jats:p></jats:sec><jats:sec><jats:title>Results</jats:title><jats:p>We defined a bi-pulse electroporation shock with bi-directional and unidirectional electric field, respectively, that permitted efficient MC insertion and maintained a high frequency of viable T cells. In three large scale runs, 2E8 T cells were enriched from leukapheresis product, activated, gene-engineered and expanded to yield up to 3.5E9 total T cells/1.4E9 CAR-modified T cells within 12 days (CAR-modified T cells: 28.8%±12.3%). The resulting cell product contained highly pure T cells (97.3±1.6%) with balanced CD4/CD8 ratio and a high frequency of T cells with central memory phenotype (87.5%±10.4%). The transposon copy number was 7.0, 9.4 and 6.8 in runs #1–3, respectively, and gene analyses showed a balanced expression of activation/exhaustion markers. The CD19 CAR T cell product conferred potent anti-lymphoma reactivity in pre-clinical models. Notably, the operator hands-on-time was substantially reduced compared with conventional non-automated CAR T cell manufacturing campaigns.</jats:p></jats:sec><jats:sec><jats:title>Conclusions</jats:title><jats:p>We report on the first automated transposon-based manufacturing process for CAR T cells that is ready for formal validation and use in clinical manufacturing campaigns. This process and platform have the potential to facilitate access of patients to CAR T cell therapy and to accelerate scaled, multiplexed manufacturing both in the academic and industry setting.</jats:p></jats:sec>
  • Access State: Open Access