Pierre Springuel, Tiffany Hood, Fern Slingsby, Timo Schmidberger, Nicola Bevan, Noushin Dianat, Julia Hengst, Qasim A Rafiq
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引用次数: 0
Abstract
The ex vivo expansion of autologous chimeric antigen receptor (CAR) T cells to reach a therapeutic dose significantly prolongs manufacturing time and increases overall costs. The common use of animal- or human-derived serum in T cell expansion culture media further contributes to process variability, costs and introduces additional safety concerns. To address these challenges, this study focused on intensifying CAR-T cell expansion using perfusion processes in xeno-free (XF) and serum-free (SF) culture medium. The impacts of alternative tangential flow (ATF) perfusion rates, perfusion start times and donor variability were evaluated using a Design of Experiments (DOE) approach in the Ambr® 250 High-Throughput Perfusion stirred-tank bioreactor. This allowed the identification of optimal combinations of perfusion parameters on a per-donor basis, enabling 4.5-fold improvements in final cell yields and over 50% reductions in the expansion time required to reach a representative CAR-T dose compared to a fed-batch process. Subsequent process development then established an adaptive perfusion strategy enabling 130 ± 9.7-fold expansions to achieve final cell densities of 33.5 ± 3 × 106 cells/mL while reducing medium requirements by 11% without compromising CAR-T cell quality attributes compared to static well-plate cultures. Harvested cells predominantly expressed naïve and central memory markers, low levels of exhaustion markers, and maintained cytotoxicity and cytokine release in vitro. This study demonstrates the potential of optimising and adapting perfusion strategies in XF/SF-culture medium to enhance CAR-T cell yields, shorten expansion times and reduce medium consumption while addressing patient variability in clinical manufacturing. Key considerations for future implementation and improvement of adaptive perfusion feeds for clinical CAR-T manufacturing are also discussed.
期刊介绍:
The translation of new discoveries in medicine to clinical routine has never been easy. During the second half of the last century, thanks to the progress in chemistry, biochemistry and pharmacology, we have seen the development and the application of a large number of drugs and devices aimed at the treatment of symptoms, blocking unwanted pathways and, in the case of infectious diseases, fighting the micro-organisms responsible. However, we are facing, today, a dramatic change in the therapeutic approach to pathologies and diseases. Indeed, the challenge of the present and the next decade is to fully restore the physiological status of the diseased organism and to completely regenerate tissue and organs when they are so seriously affected that treatments cannot be limited to the repression of symptoms or to the repair of damage. This is being made possible thanks to the major developments made in basic cell and molecular biology, including stem cell science, growth factor delivery, gene isolation and transfection, the advances in bioengineering and nanotechnology, including development of new biomaterials, biofabrication technologies and use of bioreactors, and the big improvements in diagnostic tools and imaging of cells, tissues and organs.
In today`s world, an enhancement of communication between multidisciplinary experts, together with the promotion of joint projects and close collaborations among scientists, engineers, industry people, regulatory agencies and physicians are absolute requirements for the success of any attempt to develop and clinically apply a new biological therapy or an innovative device involving the collective use of biomaterials, cells and/or bioactive molecules. “Frontiers in Bioengineering and Biotechnology” aspires to be a forum for all people involved in the process by bridging the gap too often existing between a discovery in the basic sciences and its clinical application.