Dendritic cells (DCs) are the most potent antigen-presenting cells, and have thus been used in clinical cancer vaccines. However, the effects of DC vaccines are still limited, leading researchers to explore novel ways to make them effective. In this study, we investigated whether human monocyte-derived DCs generated via the addition of interleukin 15 (IL-15) had a higher capacity to induce antigen-specific T cells compared to conventional DCs. We isolated CD14+ monocytes from peripheral blood from multiple myeloma (MM) patients, and induced immature DCs with granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-4 in the presence or absence of IL-15 for 4-6 days. Then we generated mature DCs (mDCs) with lipopolysaccharide for another 2 days [IL-15 mDCs (6 days), IL-15 mDCs (8 days), and conventional mDCs (8 days)]. IL-15 mDCs (6 days) showed higher expression of MHC I and II, CD40, CD86, and CCR7, and the secretion of IFN-γ was significantly higher compared to conventional mDCs. IL-15 mDCs (6 days) showed superior polarization of naïve T cells toward Th1 cells and a higher proportion of activated T cells, cytokine-induced killer (CIK) cells, and natural killer (NK) cells for inducing strong cytotoxicity against myeloma cells, and lower proportion of regulatory T cells compared to conventional mDCs. These data imply that novel multipotent mDCs generated by the addition of IL-15, which can be cultivated in 6 days, resulted in outstanding activation of T cells, CIK cells and NK cells, and may facilitate cellular immunotherapy for cancer patients.

The development of new treatment agents in recent decades has significantly improved the survival of patients with multiple myeloma (MM). Nonetheless, MM remains an incurable disease; therefore, novel combination therapies are required. Natural killer (NK) cells are one of the safest immunotherapeutic options. In this study, we found that the anti-myeloma activity of expanded NK cells (eNKs) was improved by daratumumab, lenalidomide, and dexamethasone (DRd) in an MM xenograft mouse model. NK cells expanded from peripheral blood mononuclear cells collected from MM patients were highly cytotoxic against DRd pretreated tumor cells in vitro. To mimic the clinical protocol, a human MM xenograft model was developed using human RPMI8226-RFP-FLuc cells in NOD/SCID IL-2Rγnull (NSG) mice. MM bearing mice were randomly divided into six groups: no treatment, eNK, Rd, Rd + eNKs, DRd, and DRd + eNKs. DRd significantly enhanced the cytotoxicity of eNKs by upregulating NK cell activation ligands and effector function. DRd in combination with eNKs significantly reduced the serum M-protein level and prolonged mouse survival. In addition, DRd significantly increased the persistence of eNK and homing to MM sites. These results show that the anti-myeloma activity of ex vivo-expanded and activated NK cells is augmented by the immunomodulatory effect of DRd in MM-bearing mice, suggesting the therapeutic potential of this combination for MM patients.

BACKGROUND: Natural killer (NK) cell-based immunotherapy is a promising treatment approach for multiple myeloma (MM), but obtaining a sufficient number of activated NK cells remains challenging. Here, we report an improved method to generate ex vivo expanded NK (eNK) cells from MM patients based on genetic engineering of K562 cells to express OX40 ligand and membrane-bound (mb) IL-18 and IL-21. METHODS: K562-OX40L-mbIL-18/-21 cells were generated by transducing K562-OX40L cells with a lentiviral vector encoding mbIL-18 and mbIL-21, and these were used as feeder cells to expand NK cells from peripheral blood mononuclear cells of healthy donors (HDs) and MM patients in the presence of IL-2/IL-15. Purity, expansion rate, receptor expression, and functions of eNK cells were determined over four weeks of culture. RESULTS: NK cell expansion was enhanced by short exposure of soluble IL-18 and IL-21 with K562-OX40L cells. Co-culture of NK cells with K562-OX40L-mbIL-18/-21 cells resulted in remarkable expansion of NK cells from HDs (9,860-fold) and MM patients (4,929-fold) over the 28-day culture period. Moreover, eNK cells showed increased expression of major activation markers and enhanced cytotoxicity towards target K562, U266, and RPMI8226 cells. CONCLUSIONS: Our data suggest that genetically engineered K562 cells expressing OX40L, mbIL-18, and mbIL-21 improve the expansion of NK cells, increase activation signals, and enhance their cytolytic activity towards MM cells.