Establishing the scalable manufacture of primary human T-cells in an automated stirred-tank bioreactor

Costariol, Elena, Rotondi, Marco, Amini, Arman, Hewitt, Christopher, Nienow, Alvin, Heathman, Thomas R.J., Micheletti, Martina and Rafiq, Qasim (2019). Establishing the scalable manufacture of primary human T-cells in an automated stirred-tank bioreactor. Biotechnology and Bioengineering ,


Advanced cell and gene therapies such as chimeric antigen receptor T-cell immunotherapies (CAR-T), present a novel therapeutic modality for the treatment of acute and chronic conditions including acute lymphoblastic leukaemia and non-Hodgkin lymphoma. However, the development of such immunotherapies requires the manufacture of large numbers of T-cells which remains a major translational and commercial bottleneck due to the manual, small-scale and often static culturing systems used for their production. Such systems are used because there is an unsubstantiated concern that primary T-cells are shear sensitive, or prefer static conditions, and therefore do not grow as effectively in more scalable, agitated systems, such as stirred-tank bioreactors, as compared to T-flasks and culture bags. In this study, we demonstrate, that not only can T-cells be cultivated in an automated stirred-tank bioreactor system (ambr® 250), but that their growth is consistently and significantly better than that in T-flask static culture, with equivalent cell quality. Moreover, we demonstrate that at progressively higher agitation rates over the range studied here, and thereby higher specific power inputs (P/M W kg-1), the higher the final viable T-cell density; i. e., a cell density of 4.65 ± 0.24 x 106 viable cells ml-1 obtained at the highest P/M of 74 x 10-4 W kg-1 in comparison to 0.91 ± 0.07 x 106 viable cells ml-1 at the lowest P/M of 3.1 x 10-4 W kg-1. We posit that this improvement is due to the inability at the lower agitation rates to effectively suspend the Dynabeads®, which are required to activate the T-cells; and that contact between them is improved at the higher agitation rates. Importantly, from the data obtained, there is no indication that T-cells prefer being grown under static conditions or are sensitive to fluid dynamic stresses within a stirred-tank bioreactor system at the agitation speeds investigated. Indeed, the opposite has proven to be the case, whereby the cells grow better under higher agitation speeds whilst maintaining their quality. This study is the first demonstration of primary T-cell ex vivo manufacture activated by Dynabeads® in an automated stirred-tank bioreactor system such as the ambr® 250 and the findings have the potential to be applied to multiple other cell candidates for advanced therapy applications.

Publication DOI:
Divisions: Life & Health Sciences
Additional Information: This is the peer reviewed version of the following article: Costariol, E. , Rotondi, M. , Amini, A. , Hewitt, C. J., Nienow, A. W., Heathman, T. R., Micheletti, M. and Rafiq, Q. A. (2019), Establishing the scalable manufacture of primary human T‐cells in an automated stirred‐tank bioreactor. Biotechnol. Bioeng. Accepted Author Manuscript, which has been published in final form at  This article may be used for non-commercial purposes in accordance With Wiley Terms and Conditions for self-archiving.
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Related URLs: https://onlinel ... .1002/bit.27088 (Publisher URL)
Published Online Date: 2019-06-11
Authors: Costariol, Elena
Rotondi, Marco
Amini, Arman
Hewitt, Christopher ( 0000-0001-6988-6071)
Nienow, Alvin
Heathman, Thomas R.J.
Micheletti, Martina
Rafiq, Qasim



Version: Accepted Version

Access Restriction: Restricted to Repository staff only until 11 June 2020.

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