Natural Killer Cells Unruffled by Cryopreservation – If Done Right

A breast cancer cell – one of the targets of natural killer cells. Image Source: Wikimedia. Released by The National Cancer Institute.

Natural killer cells are desirable for tumor-targeted cell therapies, but difficult to obtain in sufficient quantities. Deviating from previous results, scientists now show that those cells can be cryopreserved without impacting their function – using BioCision’s CoolCell® freezing containers.

Natural killer (NK) cells are part of our body’s innate immune system – our first line of defense against viral attacks or tumors. In James Bond-like fashion, these cells strike and kill, with high precision, intruders that have built themselves a home within unsuspecting host cells. NK cells are not easily fooled – they can discern “self” from “missing self” by the lower levels of a cell surface component called major histocompatibility complex I- and then attack only those cells that show “missing self”. Such cells are either stressed by the presence of pathogens such as viruses or may have turned cancerous.

Because of their ability to seek out and destroy tumor cells, natural killer cells have gained much attention for use in allogeneic cell immunotherapy approaches. For therapy, NK cells can be obtained from donors through filtering of peripheral blood or from umbilical cord blood. Either way, the cells have to be expanded to reach clinically relevant numbers. Consequently, cryopreservation of NK cells becomes a desirable goal as this would allow repeated treatments with the same cellular product. Sadly, previous studies had shown loss of natural killer cell function immediately after thawing (we recently reported on the challenges during cryopreservation of natural killer cells). British scientists now decided to re-examine the issue and report their encouraging findings in a recent paper. [1]

Rather than expanding harvested NK cells, the researchers chose a different tack. They decided to induce CD34+ stem cells from umbilical cord blood in vitro to develop into natural killer cells, using a previously established protocol. [2] After obtaining the CD34+-derived NK cells, the scientists investigated the effects of cryopreservation on cell phenotype and function. For the crucial cryopreservation step, they utilized BioCision’s CoolCell® freezing containers, to reliably provide the best possible slow-freezing conditions that enable maximal cell viability after thawing.

Lo and behold, they were unable to find any effects of cryopreservation on functionality of the natural killer cells. This was not for lack of trying. The researchers investigated all relevant parameters. They measured cytotoxicity against K562 erythroleukemic cells, expression of eleven key receptors with regulatory activities on NK cell functions, and cell proliferation in response to IL-2. Nothing. The cryopreserved natural killer cells also released antimicrobial cytotoxic and other molecules from granules to the same extent as fresh cells, when stimulated. They also generated the same amount of IFN-γ.

Finally, the authors concluded that “[o]ur data clearly show that cryopreservation does not affect functionality or phenotype of NK cells differentiated in vitro.” So, what was different from the previous attempts? The authors speculate that the concentration of the freezing medium components such as serum and DMSO as well as the source of the natural killer cells may have played a role.

Bottom line: after cryopreservation, those NK cells still looked to kill. We should have known. James Bond, after all, tends to live twice and die another day.


[1] Domogala, A. et al. Cryopreservation has no effect on function of natural killer cells differentiated in vitro from umbilical cord blood CD34+ cells. Cytotherapy 2016 Apr 15. [Epub ahead of print], PMID 27090754.

[2] Grzywacz, B. et al. Coordinated acquisition of inhibitory and activating receptors and functional properties by developing human natural killer cells. Blood 108 (12):3824–33. 2006.