Nature Products Enhance NKG2D Ligands Expression of CD123⁺CD34⁺CD38⁻ Leukemia Stem Cells for Stimulating Cytotoxicity of NKG2D⁺ Cells to Themselves

CD123⁺CD34⁺CD38⁻ leukemia cells regarded as leukemia stem cells, not only refractory to chemotherapeutics but also resistant to immune response such as cytotoxicity of prime natural killer cells, usually lead leukemia to relapse. It is essential to eliminate leukemia stem cells at utmost for cure. Here we would observe whether nature products could enhance CD123⁺CD34⁺CD38⁻ leukemia cells to express NKG2D ligands for arousing cytotoxicity of NKG2D⁺ cells to thoroughly eliminate leukemia stem cells. First, CD123⁺CD34⁺CD38⁻ leukemia cells, isolated from human acute leukemia cell line KG1a, whose MLL gene was broken apart and rearranged, of which over 60% were in G₀ phase of cell cycle, which could form about 80 colonies per 1000 cells in methylcellulose after 21 days. What’s more, they were worse than CD123⁺CD34⁺CD38⁺ leukemia cells on resistance to cytotoxicity of natural killer cells(Killed ratio <19%). CD123⁺CD34⁺CD38⁻ leukemia cells from KG1a had characteristic of defined leukemia stem cells. Then, NKG2D⁺ cells from human peripheral blood monocytes were stimulated by cytokines and CpG, whose cytotoxicity against K562 cells reached (96.5±1.23)% analyzed by LDH releasing assay, suggesting NKG2D⁺ cells had better cytotoxicity than prime natural killer cells (69.31±8.45)%. However, cytotoxicity of NKG2D⁺ cells against CD123⁺CD34⁺CD38⁻ leukemia stem cells was only(49.17±3.90)%, still higher than of prime natural killer cells (19±2.63)%. Therefore, whether to arouse cytotoxicity of NKG2D⁺ cells to deplete leukemia stem cells become essential. Finally, three kinds of nature products would be selected, including resveratrol (RV, a polyphenol found in berries), matrine and arsenic trioxide (ATO), which could decrease proliferation and survival of CD123⁺CD34⁺CD38⁻ leukemia cells and whose 50% effective concentration (EC50) were respectively 56μM, 500μg/ml and 4.0μM. Importantly, EC50 of resveratrol could up regulate NKG2D ligands ULBP1 to (53.28±8.60)%, ULBP2 to (36.54±11.45)% and ULBP3 to (65.08±14.44)% on the surface of CD123⁺CD34⁺CD38⁻ leukemia cells, and stimulate cytolytic ability of NKG2D⁺ cells, so that cytotoxicity of NKG2D⁺ cells against them reached from (49.17±3.90)% to (92.33±2.63)%. Similarly to matrine and arsenic trioxide, they could each up regulate ULBP2 from to 9.71±2.37% and ULBP1 to 24.38±3.65%, resulting in cytotoxicity of NKG2D⁺ cells against them reaching to 73.20±3.41% and 55.70±0.53%. In conclusions, cytotoxicity of NKG2D⁺ cells was better than of prime natural killer cells against not only K562, but also CD123⁺CD34⁺CD38⁻ leukemia cells, which had characteristic of leukemia stem cells, even to resist killing of prime natural killer cells. Nature products, resveratrol, matrine and arsenic trioxide, could enhance NKG2D ligands expression of CD123+CD34+CD38− leukemia stem cells for stimulating cytotoxicity of NKG2D+ cells to leukemia stem cells. This finding could provide a new strategy for eliminate leukemia stem cells.