Cryobiological aspects of upscaling cryopreservation for encapsulated liver cell therapies.

For the efficient delivery of a cell therapy a treatment must be provided rapidly, at clinical scale, contain a sufficient active cellular component (biomass), and adhere to a multitude of regulatory requirements. Cryopreservation permits many of these demands to be met more readily. Here we present the cryopreservation and recovery of large volume (2.5L) alginate encapsulated liver cell spheroids (AELS), suitable for use with a novel bioartificial liver device (HepatiCan™) for the treatment of those suffering from acute liver failure (ALF), in regulatory approved cryobags and a cryopreservation process optimised for large volumes. By first assessing the thermal profiles of large scale cryobags with a thermal mimic, the feasibility of cryopreserving a full patient dose simultaneously (3x cryobags containing 833 ml biomass each) was investigated, allowing for small and subsequently large-scale testing of cellular functional recoveries. Work presented here demonstrates that optimised reproducible cooling and warming profiles could be achieved with these large volumes, leading to high biomass recoveries at full clinical scale. The recovered AELS also had high regeneration potential, achieving full pre-freeze viable cell densities within 3 days, indicating that the cell therapy could be delivered rapidly to patients with ALF. This study has presented the feasibility for rapid delivery of large volume cell therapies, whilst further research into improved speed of post-thaw recovery is warranted.