anisomycin and quinoline

Mixed lineage kinase 7 (MLK7) is a mitogen-activated protein kinase kinase kinase (MAPKKK) that activates the pro-apoptotic signaling pathways p38 and JNK. A library of potential kinase inhibitors was screened, and a series of dihydropyrrolopyrazole quinolines was identified as highly potent inhibitors of MLK7 in vitro catalytic activity. Of this series, an aryl-substituted dihydropyrrolopyrazole quinoline (DHP-2) demonstrated an IC50 of 70 nM for inhibition of pJNK formation in COS-7 cell MLK7/JNK co-transfection assays. In stimulated cells, DHP-2 at 200 nM or MLK7 small interfering RNA completely blocked anisomycin and UV induced but had no effect on interleukin-1beta or tumor necrosis factor-alpha-induced p38 and JNK activation. Additionally, the compound blocked anisomycin and UV-induced apoptosis in COS-7 cells. Heart tissue homogenates from MLK7 transgenic mice treated with DHP-2 at 30 mg/kg had reduced JNK and p38 activation with no apparent effect on ERK activation, demonstrating that this compound can be used to block MLK7-driven MAPK pathway activation in vivo. Taken together, these data demonstrate that MLK7 is the MAPKKK required for modulation of the stress-activated MAPKs downstream of anisomycin and UV stimulation and that DHP-2 can be used to block MLK7 pathway activation in cells as well as in vivo.

Topics: Animals; Anisomycin; Apoptosis; Blotting, Western; Catalysis; COS Cells; Cytokines; DNA Fragmentation; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Enzyme Activation; Enzyme Inhibitors; Glutathione Transferase; Humans; Inhibitory Concentration 50; Interleukin-1; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 4; MAP Kinase Kinase Kinases; Mice; Mitogen-Activated Protein Kinase Kinases; Models, Chemical; Muscle Proteins; Myocardium; Nucleic Acid Synthesis Inhibitors; p38 Mitogen-Activated Protein Kinases; Plasmids; Protein Serine-Threonine Kinases; Pyrazoles; Quinolines; RNA, Small Interfering; Signal Transduction; Time Factors; Transfection; Transgenes; Tumor Necrosis Factor-alpha; Ultraviolet Rays