NK cells represent a first line of defense and play a role in immunological regulation of cancers. Their frequency and/or function may be associated with disease prognosis of various cancers. NK cells are progressively dysregulated in multiple myeloma (MM) patients. To restore and facilitate the antitumor NK cell effect, NK cells are adequately needed to be stimulated. But, an optimal strategy for manipulating NK cells has not been determined. In the current study, we have assessed the frequency and expression of two costimulatory molecules, NKG2D and DNAM-1 on NK cells as well as NK cell ligands on myeloma cells from peripheral blood mononuclear cells (PBMCs) and/or bone marrow mononuclear cells (BMMNCs) of 34 MM patients. We also assessed the proliferation of NK cells as a result of adjunctive effect of NKT cells that had been stimulated with CD1d and α-GalCer complex. Results demonstrated that NKT and NK cells in PBMCs were correlated with those in BMMNCs. Also, activated NKT cells enhanced the proliferation of NK cells and expression of NKG2D and DNAM-1 on NK cells. In addition, we demonstrated that NK cell-mediated antitumor effects driven by NKT cell activation were depended on these co-stimulatory molecules in response to the NKG2D and DNAM-1 ligands on myeloma cells. Thus, we demonstrated that NK cell response following NKT cell activation was improved beyond immune suppressive status in patients. This NK cell adjunctive therapy by activating NKT cell-targeted approach should be nominated as a potential new target for treatment of MM.