stilbenes and Leukemia--Erythroblastic--Acute

Multidrug resistance (MDR) is a major obstacle in the clinical therapy of hematological malignancies. P-glycoprotein (P-gp) overexpression results in reduction of intracellular drug concentration with a consequence that the cytotoxicity of anti-tumor drugs is decreased, which leads to MDR in K562/ADR cells. In this study, we found that resveratrol enhanced the anti-proliferative activity of bestatin in K562/ADR cells. Co-treatment with resveratrol, IC50 values of bestatin in K562/ADR cells significantly decreased and activation of caspase-3 and caspase-8 increased, which indicated that resveratrol potentiated bestatin-induced apoptosis. Resveratrol increased the intracellular concentration of bestatin through inhibiting P-gp function and downregulating P-gp expression at mRNA and protein levels, which increased anti-proliferative activity of bestatin in K562/ADR cells. Resveratrol decreased the phosphorylation of Akt and mTOR but did not affect the phosphorylations of JNK or ERK1/2. These results demonstrated that resveratrol could increase the anti-proliferative activity of bestatin through downregulating P-gp expression via suppressing the PI3K/Akt/mTOR signaling pathway.

Topics: Antineoplastic Agents; Apoptosis; ATP Binding Cassette Transporter, Subfamily B, Member 1; Blotting, Western; Caspases; Cell Proliferation; Cell Survival; Down-Regulation; Doxorubicin; Drug Resistance, Neoplasm; Drug Synergism; Humans; K562 Cells; Leucine; Leukemia, Erythroblastic, Acute; Phosphatidylinositol 3-Kinases; Phosphorylation; Proto-Oncogene Proteins c-akt; Resveratrol; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Stilbenes; TOR Serine-Threonine Kinases

CLL cells are characterized by high levels of proteins that are post-translationally modified by O-linked β-N-acetylglucosamine (O-GlcNAc) moieties, but it is not clear whether O-GlcNAc is a relevant therapeutic target. The neutraceutical resveratrol is cytotoxic to chronic lymphocytic leukemia cells in vitro. In this study, we found that resveratrol has therapeutic activity as a single agent in vivo in both human chronic lymphocytic leukemia patients and mice with erythroleukemia. Blood and splenic O-GlcNAc levels reflected the changes in tumor burden. Resveratrol directly lowered O-GlcNAc levels in leukemia cells through proteasomal activation, but increasing O-GlcNAc levels in vitro did not prevent cell death. These findings suggest that resveratrol has potential as a novel treatment for some forms of chronic and acute leukemia, and the measurement of O-GlcNAc levels could be a surrogate marker for therapeutic responses.

Topics: 3T3 Cells; Acetylglucosamine; Animals; Drug Antagonism; Humans; Interferons; Leukemia, Erythroblastic, Acute; Leukemia, Lymphocytic, Chronic, B-Cell; Mice; Proteins; Resveratrol; Stilbenes

Using structure-based virtual screening, we previously identified a novel stilbenoid inhibitor of Jak2 tyrosine kinase named G6. Here, we hypothesized that G6 suppresses Jak2-V617F-mediated human pathological cell growth in vitro and in vivo. We found that G6 inhibited proliferation of the Jak2-V617F expressing human erythroleukemia (HEL) cell line by promoting marked cell cycle arrest and inducing apoptosis. The G6-dependent increase in apoptosis levels was concomitant with increased caspase 3/7 activity and cleavage of PARP. G6 also selectively inhibited phosphorylation of STAT5, a downstream signaling target of Jak2. Using a mouse model of Jak2-V617F-mediated hyperplasia, we found that G6 significantly decreased the percentage of blast cells in the peripheral blood, reduced splenomegaly, and corrected a pathologically low myeloid to erythroid ratio in the bone marrow by eliminating HEL cell engraftment in this tissue. In addition, drug efficacy correlated with the presence of G6 in the plasma, marrow, and spleen. Collectively, these data demonstrate that the stilbenoid compound, G6, suppresses Jak2-V617F-mediated aberrant cell growth. As such, G6 may be a potential therapeutic lead candidate against Jak2-mediated, human disease.

Topics: Amino Acid Substitution; Animals; Apoptosis; Cell Cycle; Cell Line, Tumor; Disease Models, Animal; Humans; Janus Kinase 2; Leukemia, Erythroblastic, Acute; Mice; Mice, Mutant Strains; Mutation, Missense; Protein Kinase Inhibitors; Signal Transduction; STAT5 Transcription Factor; Stilbenes

Hyperkinetic Jak2 tyrosine kinase signaling has been implicated in several human diseases including leukemia, lymphoma, myeloma, and the myeloproliferative neoplasms. Using structure-based virtual screening, we previously identified a novel Jak2 inhibitor named G6. We showed that G6 specifically inhibits Jak2 kinase activity and suppresses Jak2-mediated cellular proliferation. To elucidate the molecular and biochemical mechanisms by which G6 inhibits Jak2-mediated cellular proliferation, we treated Jak2-V617F expressing human erythroleukemia (HEL) cells for 12 h with either vehicle control or 25 μM of the drug and compared protein expression profiles using two-dimensional gel electrophoresis. One differentially expressed protein identified by electrospray mass spectroscopy was the intermediate filament protein, vimentin. It was present in DMSO treated cells but absent in G6 treated cells. HEL cells treated with G6 showed both time- and dose-dependent cleavage of vimentin as well as a marked reorganization of vimentin intermediate filaments within intact cells. In a mouse model of Jak2-V617F mediated human erythroleukemia, G6 also decreased the levels of vimentin protein, in vivo. The G6-induced cleavage of vimentin was found to be Jak2-dependent and calpain-mediated. Furthermore, we found that intracellular calcium mobilization is essential and sufficient for the cleavage of vimentin. Finally, we show that the cleavage of vimentin intermediate filaments, per se, is sufficient to reduce HEL cell viability. Collectively, these results suggest that G6-induced inhibition of Jak2-mediated pathogenic cell growth is concomitant with the disruption of intracellular vimentin filaments. As such, this work describes a novel pathway for the targeting of Jak2-mediated pathological cell growth.

Topics: Animals; Calpain; Cell Death; Cell Line, Tumor; Humans; Janus Kinase 2; Leukemia, Erythroblastic, Acute; Mice; Mice, Inbred NOD; Mice, SCID; Protein Kinase Inhibitors; Spectrometry, Mass, Electrospray Ionization; Stilbenes; Vimentin

Resveratrol, a natural dietary polyphenol, has been postulated to be implicated in the cardioprotective effect of red wine and the low incidence of breast and prostate cancers among vegetarians and Orientals respectively. This compound inhibits ribonucleotide reductase as does hydroxyurea, the first therapeutic agent used in the treatment of sickle cell disease. Using the human erythroleukaemic K562 cell line as an in vitro model, we show here that 50 micromol/l of resveratrol induced a higher haemoglobin production (sevenfold) in K562 cells than 500 micromol/l of hydroxyurea (3.5-fold). This erythroid differentiation was linked to a dose- and time-dependent inhibition of cell proliferation associated with an equivalent increased expression of p21 mRNA, but with a higher increased level of p21 protein (sixfold) for cells treated with resveratrol than for those treated with hydroxyurea (1.5-fold). We also show that 50 micromol/l of resveratrol and 25 micromol/l of hydroxyurea induced variable but similar enhancements of fetal haemoglobin synthesis in cultured erythroid progenitors for the majority of the sickle cell patients studied. These inductions were linked to, but not correlated with, a variable decrease in erythroid burst-forming unit clone number. Taken together, these results show that resveratrol merits further investigations in sickle cell disease therapy.

Topics: Anemia, Sickle Cell; Antioxidants; Blotting, Western; Cell Differentiation; Cell Division; Cell Line; Cells, Cultured; Dose-Response Relationship, Drug; Enzyme Inhibitors; Erythroid Precursor Cells; Fetal Hemoglobin; Gene Expression; Granulocyte-Macrophage Colony-Stimulating Factor; Hemoglobins; Humans; Hydroxyurea; Leukemia, Erythroblastic, Acute; Models, Biological; Proto-Oncogene Proteins p21(ras); Resveratrol; Reverse Transcriptase Polymerase Chain Reaction; Ribonucleotide Reductases; Stem Cells; Stilbenes; Time Factors