title: Highly efficient IL-21 and feeder cell-drivenex vivoexpansion of human NK cells with therapeutic activity in a xenograft mouse model of melanoma.
abstract: Natural killer (NK) cells are promising antitumor effector cells, but the generation of sufficient NK cell numbers for adoptive immunotherapy remains challenging. Therefore, we developed a method for highly efficientex vivoexpansion of human NK cells.Ex vivoexpansion of NK cells in medium containing IL-2 and irradiated clinical-grade feeder cells (EBV-LCL) induced a 22-fold NK cell expansion after one week that was significantly increased to 53-fold by IL-21. Repeated stimulation with irradiated EBV-LCL and IL-2 and addition of IL-21 at the initiation of the culture allowed sustained NK cell proliferation with 1011-fold NK cell expansion after 6 weeks. Compared to naive NK cells, expanded NK cells upregulated TRAIL, NKG2D, and DNAM-1, had superior cytotoxicity against tumor cell linesin vitroand produced more IFNγ and TNF-α upon PMA/Iono stimulation. Most importantly, adoptive transfer of NK cells expanded using feeder cells, IL-2 and IL-21 led to significant inhibition of tumor growth in a melanoma xenograft mouse model, which was greater than with NK cells activated with IL-2 alone. Intriguingly, adoptively transferred NK cells maintained their enhanced production of IFNγ and TNF-α uponex vivorestimulation, although they rapidly lost their capacity to degranulate and mediate tumor cytotoxicity after thein vivotransfer. In conclusion, we developed a protocol forex vivoNK cell expansion that results in outstanding cell yields. The expanded NK cells possess potent antitumor activityin vitroandin vivoand could be utilized at high numbers for adoptive immunotherapy in the clinic.
authors.affiliation: Innate Immunity Group, German Cancer Research Center, Heidelberg, Germany; Miltenyi Biotec GmbH, Bergisch Gladbach, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Granzin
authors.affiliation: Innate Immunity Group, German Cancer Research Center , Heidelberg, Germany.
authors.forename: Ana
authors.initials: A
authors.lastname: Stojanovic
authors.affiliation: Innate Immunity Group, German Cancer Research Center , Heidelberg, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Miller
authors.affiliation: Section of Transplantation Immunotherapy, Hematology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health , Bethesda, MD, USA.
authors.forename: Richard
authors.initials: R
authors.lastname: Childs
authors.affiliation: Miltenyi Biotec GmbH, Bergisch Gladbach , Germany.
authors.forename: Volker
authors.initials: V
authors.lastname: Huppert
authors.affiliation: Innate Immunity Group, German Cancer Research Center , Heidelberg, Germany.
authors.forename: Adelheid
authors.initials: A
authors.lastname: Cerwenka
keywords: Cancer immunotherapy; EBV-LCL; Interleukin-21; NK cell expansion; Natural killer cells; melanoma; xenograft mouse model