Growth and Activation of Natural Killer Cells Ex Vivo from Children with Neuroblastoma for Adoptive Cell Therapy
Menée in vitro, cette étude évalue la faisabilité d'une méthode pour faire croître ex vivo des cellules NK activées à partir de cellules mononucléées du sang périphérique d'enfants atteints d'un neuroblastome à haut risque
Résumé en anglais
Purpose: Adoptive transfer of natural killer (NK) cells combined with tumor-specific monoclonal antibodies (mAbs) has therapeutic potential for malignancies. We determined if large numbers of activated NK (aNK) cells can be grown ex vivo from peripheral blood mononuclear cells (PBMC) of children with high-risk neuroblastoma using artificial antigen-presenting cells (aAPC).
Experimental Design: Irradiated K562-derived Clone 9.mbIL21 aAPC were co-cultured with PBMC, and propagated NK cells were characterized with flow cytometry, cytotoxicity assays, Luminex® multi-cytokine assays, and a NOD/SCID mouse model of disseminated neuroblastoma.
Results: Co-culturing patient PBMC with aAPC for 14 days induced 2,363±443-fold expansion of CD56+CD3-CD14- NK cells with 83±4% purity (n=10). Results were similar with PBMC from normal donors (n=5). Expression of DNAM-1, NKG2D, FcγRIII/CD16 and CD56 increased 6±3, 10±2, 21±20, and 18±3-fold respectively on day 14 compared to day 0, demonstrating activation of NK cells. In vitro, aNK cells were highly cytotoxic against neuroblastoma cell lines, and killing was enhanced with GD2-specific monoclonal antibody ch14.18. When mediating cytotoxicity with ch14.18, release of TNFα, GM-CSF, IFNγ, sCD40L, CCL2/MCP-1, CXCL9/MIG, and CXCL11/I-TAC by aNK cells increased 4-, 5- 6-, 15-, 265-, 917- and 363-fold (151 to 9,121 pg/mL), respectively, compared to aNK cells alone. Survival of NOD/SCID mice bearing disseminated neuroblastoma improved when treated with thawed and immediately intravenously infused cryopreserved aNK cells compared to un-treated mice and was further improved when ch14.18 was added.
Conclusion: Propagation of large numbers of aNK cells that maintain potent anti-neuroblastoma activities when cryopreserved supports clinical testing of adoptive cell therapy with ch14.18.
