How to Improve Cryopreservation of Leukopaks and Apheresis Products

Apheresis products/leukopaks are heterogeneous cell mixtures. Many of these cell types are of clinical interest. For example, apheresis products contain populations of T-Cells (CD3+CD4+ and CD3+CD8+), B-cells, and natural killer cells. These cell types can be isolated from the heterogeneous cell population, genetically modified, and/or expanded in culture for a cell therapy product. Typically, all apheresis products/leukopaks are cryopreserved using the same cryopreservation protocol (e.g. cooling rate and composition of freezing solution). The different cell types present in a leukopak have different biological characteristics and not surprisingly, different freezing responses. A recent study (Pi et al., 2020) demonstrated that different cell subpopulations had different freezing responses when subjected to the same freezing conditions (Fig 1). CD3+CD4+ cells survived more readily than CD3+CD8+ cells, and NK cells also had a poor outcome.  

So, what do we do to improve the preservation of heterogeneous cell populations? Current methods of preservation result in depletion of the CD3+CD8+ relative to the CD3+CD4+. The Evia Bio approach is to use a computational algorithm to improve post-thaw recovery of mixed cell populations when high levels of post-thaw recovery are desired for multiple cell types. Figure 2 demonstrates an optimization strategy used to improve post-thaw recovery of both CD3+CD4+ and CD3+CD8+ subpopulations present in leukopaks. We developed an approach that reduced cell losses in the CD3+CD8+ subpopulation while still maintaining a high recovery of the CD3+CD4+ fraction of cells. This novel approach can be used to improve the preservation of other heterogeneous cell populations with different freezing responses.

References:

Pi, C.-H., Hornberger, K., Dosa, P., & Hubel, A. (2020). Understanding the freezing responses of T cells and other subsets of human peripheral blood mononuclear cells using DSMO-free cryoprotectants. Cytotherapy, 22(5), 291–300. https://doi.org/10.1016/j.jcyt.2020.01.013