Lock D, Monjezi R, Brandes C, Bates S, Lennartz S, Teppert K, Gehrke L, Karasakalidou-Seidt R, Lukic T, Schmeer M, Schleef M, Werchau N, Eyrich M, Assenmacher M, Kaiser A, Prommersberger S, Schaser T, Hudecek M (2022)
Publication Type: Journal article
Publication year: 2022
Book Volume: 10
Article Number: e005189
Journal Issue: 9
Background 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. Methods 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. Results 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. Conclusions 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.
APA:
Lock, D., Monjezi, R., Brandes, C., Bates, S., Lennartz, S., Teppert, K.,... Hudecek, M. (2022). Automated, scaled, transposon-based production of CAR T cells. Journal for ImmunoTherapy of Cancer, 10(9). https://doi.org/10.1136/jitc-2022-005189
MLA:
Lock, Dominik, et al. "Automated, scaled, transposon-based production of CAR T cells." Journal for ImmunoTherapy of Cancer 10.9 (2022).
BibTeX: Download