azd-6244 and Leukemia--Myeloid

Molecular aberrations of the Ras/Raf/mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK)/ERK and/or Murine double minute (MDM2)/p53 signaling pathways have been reported in 80% and 50% of primary acute myeloid leukemia (AML) samples and confer poor outcome. In this study, antileukemic effects of combined MEK inhibition by AZD6244 and nongenotoxic p53 activation by MDM2 antagonist Nutlin-3a were investigated. Simultaneous blockade of MEK and MDM2 signaling by AZD6244 and Nutlin-3a triggered synergistic proapoptotic responses in AML cell lines [combination index (CI) = 0.06 +/- 0.03 and 0.43 +/- 0.03 in OCI/AML3 and MOLM13 cells, respectively] and in primary AML cells (CI = 0.52 +/- 0.01). Mechanistically, the combination upregulated levels of BH3-only proteins Puma and Bim, in part via transcriptional upregulation of the FOXO3a transcription factor. Suppression of Puma and Bim by short interfering RNA rescued OCI/AML3 cells from AZD/Nutlin-induced apoptosis. These results strongly indicate the therapeutic potential of combined MEK/MDM2 blockade in AML and implicate Puma and Bim as major regulators of AML cell survival.

Topics: Acute Disease; Apoptosis; Apoptosis Regulatory Proteins; Bcl-2-Like Protein 11; Benzimidazoles; Drug Synergism; Extracellular Signal-Regulated MAP Kinases; Forkhead Box Protein O3; Forkhead Transcription Factors; HL-60 Cells; Humans; Imidazoles; Leukemia, Myeloid; MAP Kinase Signaling System; Membrane Proteins; Myeloid Cell Leukemia Sequence 1 Protein; Phosphorylation; Piperazines; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Proto-Oncogene Proteins c-mdm2; U937 Cells

We have recently reported that the mitogen-activated protein kinase/ERK kinase (MEK) inhibitor AZD6244 (ARRY-142886) strikingly potentiated the effects of histone deacetylase inhibitor to induce growth arrest and apoptosis of acute myelogeneous leukemia (AML) cells in association with enhanced upregulation of p21(waf1). This study examined the effects of the MEK inhibitor on the action of DNA methyltransferase inhibitor 5-Aza-2'-deoxycytidine (5-AzadC), another epigenetic agent in AML cells. AZD6244 significantly potentiated the ability of 5-AzadC to induce growth arrest and apoptosis of NB4, and freshly isolated AML cells. In parallel, 5-AzadC induced expression of p21(waf1) in AML cells, which was potently enhanced in the presence of AZD6244. Further studies explored the molecular mechanisms by which 5-AzadC induced expression of p21(waf1) and found that a low dose of 5-AzadC (1 microM) induced acetylation of histone H3 on the p21(waf1) gene promoter; however, higher dose of this compound (3 or 5 microM) potently induced DNA damage as assessed by expression of gammaH2AX, in NB4 cells. These effects were strikingly enhanced by concomitant blockade of MEK signaling. Furthermore, knock-down of p21(waf1) by the siRNA rescued NB4 cells from 5-AzadC-mediated growth inhibition. Taken together, combination of 5-AzadC and the MEK inhibitor may be useful for treatment of individuals with a subset of AML.

Topics: Acetylation; Aged; Apoptosis; Azacitidine; Benzimidazoles; Blotting, Western; Cell Proliferation; Chromatin Immunoprecipitation; Cyclin-Dependent Kinase Inhibitor p21; Decitabine; DNA Modification Methylases; Drug Synergism; Enzyme Inhibitors; Flow Cytometry; Gene Expression Regulation; Histone Deacetylase Inhibitors; Histone Deacetylases; Histones; Humans; Leukemia, Myeloid; MAP Kinase Kinase 1; Middle Aged; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Small Interfering; Signal Transduction; Transfection; Tumor Cells, Cultured