An independent publication in Nature: Scientific Reports cites using fresh leukopaks sourced from HemaCare in their T cell cryopreservation study. The authors, who work at the Cell and Gene Therapy Catapult in London, are studying the impact of cooling and thawing rates on cryopreserved human peripheral blood-derived T cells. In order for the emerging cell and gene therapy industry to effectively ship and store cell-based therapies to global markets, it will be necessary to cryopreserve cell therapy starting materials, as well as the final therapeutic products. To retain peak cellular function, and therefore therapeutic efficacy upon patient administration, it is necessary to understand how to optimize the cryopreservation process.
Catapult scientists realized that to ensure a cell therapy product with reproducible efficacy, they needed to have a thorough understanding of the actual mechanics of cooling and thawing as it applies to their target cells. CAR-T cell therapies have recently earned the FDA’s seal of approval based on the exceptionally positive outcomes observed with these therapies in cancer treatment. Since then, there has been a significant upswing in the number of CAR-T based therapeutics in clinical development. For this reason, the researchers have been interested in safeguarding the efficacy of T cell therapies at the point of the clinical delivery, by optimizing cryopreservation techniques.
The authors focused on exploring how the interaction between cooling and warming rates during freeze-thaw affects cell function and viability. Simultaneously, they planned to carry out a cryomicroscopy study which would allow them to observe and measure ice crystal formation. They elected to use standard cryopreservation vials, and a standard DMSO-based cryoprotectant (CryoStor 10®) for the bulk of their study. The authors obtained fresh HemaCare leukopaks from healthy donors, from which they isolated and expanded human T cells, using standard immunomagnetic separation and cell culture techniques. It’s critical to have consistently high cell viability counts in the initial leukapheresis sample to carry out such studies.
Using a specialized cryomicroscopy technique, the authors were able to directly correlate ice crystal formation during freezing, and any recrystallization during thawing, to loss of cell viability and proliferative capacity. They then analyzed the effect of various cooling and warming rates on post thaw viability and proliferation. The results were complicated. As expected, slow cooling rates (lower than 10 °C min−1) combined with rapid warming rates gave the best outcome. As the rate of cooling increases, there is a potential of ice recrystallization upon warming, leading to a loss in cell viability. This risk drops significantly, however, at slower cooling rates. Very high viability, total viable cell numbers and cell proliferation were observed following very slow rates of cooling (0.1 °C min−1), even in the face of slower rates of warming. Immune function and apoptosis rates were not examined for the purposes of this study.
This research provides valuable insight into the effects of changes in ice crystallization on both cryopreserved cell therapy starting materials and cryopreserved cell therapies. At HemaCare, we strongly support research that helps define optimal conditions for the clinical delivery of cell therapy products.
- Baboo J., et al. The Impact of Varying Cooling and Thawing Rates on the Quality of Cryopreserved Human Peripheral Blood T Cells. Nature: Scientific Reports. 9: 3417. Mar 2019.