Cryopreservation has no effect on function of natural killer cells differentiated in vitro from umbilical cord blood CD34 + cells

Background aims Natural killer (NK) cells offer the potential for a powerful cellular immunotherapy because they can target malignant cells without being direct effectors of graft-versus-host disease. We have previously shown that high numbers of functional NK cells can be differentiated in vitro from umbilical cord blood (CB) CD34+ cells. To develop a readily available, off-the-shelf cellular product, it is essential that NK cells differentiated in vitro can be frozen and thawed while maintaining the same phenotype and functions. Methods We evaluated the phenotype and function of fresh and frozen NK cells differentiated in vitro. We also assessed whether the concentration of NK cells at the time of freezing had an impact on cell viability. Results We found that cell concentration of NK cells at the time of freezing did not have an impact on their viability and on cell recovery post-thaw. Moreover, freezing of differentiated NK cells in vitro did not affect their phenotype, cytotoxicity and degranulation capacity toward K562 cells, cytokine production and proliferation. Conclusions We are therefore able to generate large numbers of functional NK cells from CB CD34+ cells that maintain the same phenotype and function post-cryopreservation, which will allow for multiple infusions of a highly cytotoxic NK cell product.