title: Trimidox induces apoptosis via cytochrome c release in NALM-6 human B cell leukaemia cells.
abstract: Trimidox (3,4,5-trihydroxybenzamidoxime) has been shown to reduce the activity of ribonucleotide reductase accompanied by growth inhibition and the differentiation of mammalian cells. Here we examine the induction of apoptosis by trimidox in several human leukaemia cell lines, focusing on the release of cytochrome c and the activation of caspase proteases in the human B cell line NALM-6. Induction of apoptosis by trimidox (300 microM) was detected in NALM-6, HL-60 (premyelocytic leukaemia cells), MOLT-4 (an acute lymphoblastic leukaemia cells), Jurkat (a T-cell leukaemia cells), U937 (expressing many monocyte-like characteristics), and K562 (erythroleukaemia). NALM-6 was most affected by trimidox among leukaemia cells; therefore, we employed NALM-6 cells in the subsequent experiments. The cells showed a time-dependent increase in DNA damage after trimidox (250 microM) treatment. A significant increase in the amount of cytochrome c release was detected after treatment with trimidox. Bcl-2 and Bax protein expressions were not changed by trimidox. Caspase-3 and -9 were activated by incubation with trimidox, whereas caspase-8 was not. Furthermore, trimidox-induced apoptosis was prevented by a broad-spectrum caspase inhibitor, a caspase-3, and a caspase-9 inhibitor, but not by a caspase-8 inhibitor. Inhibition of c-Jun NH2-terminal kinase (JNK) by SP600125 appreciably protected cells from trimidox-induced apoptosis, but no effect inhibition of p38 mitogen-activated protein kinase (MAPK) by SB203580. In contrast, extracellular signal-regulated kinase (ERK) inhibitors U0126 and PD98059 strongly potentiated the apoptotic effect of trimidox. This report shows that the induction of apoptosis by trimidox occurs through a cytochrome c-dependent pathway, which sequentially activates caspase-3 and caspase-9.
authors.affiliation: Department of Pharmacology and Toxicology, Cancer Research Institute, Tohoku Pharmaceutical University, Komatsushima, Sendai, Japan. syu-kan@tohoku-pharm.ac.jp
authors.forename: Syu-Ichi
authors.initials: S
authors.lastname: Kanno
authors.forename: Kohji
authors.initials: K
authors.lastname: Uwai
authors.forename: Ayako
authors.initials: A
authors.lastname: Tomizawa
authors.forename: Takako
authors.initials: T
authors.lastname: Hiura
authors.forename: Yuu
authors.initials: Y
authors.lastname: Osanai
authors.forename: Mayuko
authors.initials: M
authors.lastname: Ujibe
authors.forename: Mitsuhiro
authors.initials: M
authors.lastname: Takeshita
authors.forename: Takaharu
authors.initials: T
authors.lastname: Ohtake
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Kimura
authors.forename: Masaaki
authors.initials: M
authors.lastname: Ishikawa
mesh_terms: D000873:Anthracenes / Q000494:pharmacology; D017209:Apoptosis; D001550:Benzamidines / Q000633:toxicity*; D002070:Butadienes / Q000494:pharmacology; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism*; D045744:Cell Line, Tumor; D045304:Cytochromes c / Q000378:metabolism*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000633:toxicity*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D015448:Leukemia, B-Cell; D009570:Nitriles / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology; D012264:Ribonucleotide Reductases / Q000037:antagonists & inhibitors; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors
chemical_list: C074761:3,4,5-trihydroxybenzamidoxime; D000873:Anthracenes; D001550:Benzamidines; D002070:Butadienes; D004791:Enzyme Inhibitors; D009570:Nitriles; D009842:Oligopeptides; C113580:U 0126; C110772:benzoylcarbonyl-aspartyl-glutamyl-valyl-aspartyl-fluoromethyl ketone; C403754:benzyloxycarbonyl-leucyl-glutamyl-histidyl-aspartic acid fluoromethyl ketone; C432165:pyrazolanthrone; D045304:Cytochromes c; D012264:Ribonucleotide Reductases; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases