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Rapamycin enhances dimethyl sulfoxide-mediated growth arrest in human myelogenous leukemia cells

Introduction: The pharmacological inhibitors of phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway have been proposed in the treatment of leukemia based on their antiproliferative effects, but their possible role in differentiation therapy is less explored. Dimethyl sulfoxide (DMSO) is a potent inducer of granulocytic differentiation of acute myeloid leukemia (AML) lines. The aim of the present study was to test for the possible synergistic effects of rapamycin and DMSO on growth arrest and differentiation of AML. Materials and Methods: Myeloblastic (AML-M2) HL- 60, promyelocytic (AML-M3) NB4, monocytic (AML-M5) U937, immature (AML-M6) KG-1, and erythro- megakaryocytic K562 cells were maintained in exponential growth and differentiated in the presence of DMSO (0.6 or 1.25%). The number of viable cells was quantified using a hemocytometer. The expression of differentiation markers and the cell cycle distribution of propidium iodide- labeled cells were determined by FACS analyses. Cell morphology was evaluated on May-Grunwald- Giemsa-stained cytospin preparations. The level of phosphorylated (Thr389) and total p70 S6 Kinase (p70 S6K) in cell lysates was determined by Western blot analysis. Results: Western blot analysis demonstrated that rapamycin (20 nM) completely reduced the level of Thr389-phosphorylated p70 S6K. When applied for 96 h, rapamycin (20nM) alone exerted minimal antiproliferative effects, DMSO inhibited proliferation in a dose-dependent manner, and the combination of rapamycin and DMSO reduced the number of viable cells significantly more than either agent alone. FACS analysis of propidium iodide-labeled cells revealed a synergistic effect of rapamycin and DMSO on cell cycle arrest in G0/G1 phase. In NB4 cells, rapamycin had no statistically significant effects on DMSO-mediated increase in the expression of CD11b, but increased apoptosis. Conclusion: Rapamycin potentiates growth- inhibitory effects of DMSO in the established leukemia lines. These results suggest that mTOR inhibitors may have beneficial effects in combination with differentiation agents in therapy of AML.

rapamycin; DMSO; acute myeloid leukemia

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