Homogeneous shear distribution improves NK-92 cell cytotoxicity in a clinically relevant 2 L membrane-stirred bioreactor

Abstract

The immortalized NK-92 cell line is widely used to study natural killer (NK) cellbiology and develop immunotherapies. NK-92 cells exhibit strong cytotoxicactivity against tumor and virus-infected cells and are often used as afunctional surrogate for primary NK cells. Beyond research applications, NK-92cells are currently being evaluated in clinical trials as an allogeneic, off-the-shelfcell therapy. NK cells are typically cultured in static systems, which limitsscalability. For clinical and commercial applications, large-scale cell expansionrequires scalable platforms, such as bioreactors. However, conventional bubbleaerated bioreactors generate shear stress and foam, which can impairproliferation during prolonged culture and compromise cell quality. To addressthese limitations, a membrane-based stirring and aeration system was comparedto a conventional pitched-blade impeller with microsparger aeration in 2 L stirredtank bioreactors. NK-92 cells were expanded from static pre-cultures into shakeflasks and subsequently inoculated into each bioreactor with continuous feedingto maintain ideal nutrient supply. Both systems supported comparable growth,viability, and metabolic profiles. Cells showed comparable growth, viability andmetabolism profile in both systems. However, cells expanded in the membranebased system exhibited markedly higher cytotoxicity and cytotoxic capacity.Computational fluid dynamic simulations of both systems suggest that thisobservation is likely attributable to the more homogeneous shear distributionin the membrane-stirred setup compared to the pitched-blade configuration.Overall, this work presents a novel cultivation method to produce highly cytotoxicNK-92 cells in a well scalable stirred-tank bioreactor platform for allogeneic offthe-shelf cell therapy.

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