title: The novel triterpenoid 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO) potently enhances apoptosis induced by tumor necrosis factor in human leukemia cells.
abstract: Tumor necrosis factor (TNF) is a potent activator of the nuclear factor-kappaB (NF-kappaB) pathway that leads to up-regulation of anti-apoptotic proteins. Hence, TNF induces apoptosis in the presence of inhibitors of protein or RNA synthesis. We report that a novel triterpenoid, 2-cyano-3,12-dioxooleana-1,9,-dien-28-oic acid (CDDO) inhibits NF-kappaB-mediated gene expression at a step after translocation of activated NF-kappaB to the nucleus. This effect appears specific for the NF-kappaB pathway as CDDO does not inhibit gene expression induced by the phorbol ester 12-0-tetradecanoylphorbol-13-acetate (TPA). CDDO in combination with TNF caused a dramatic increase in apoptosis in ML-1 leukemia cells that was associated with activation of caspase-8, cleavage of Bid, translocation of Bax, cytochrome c release, and caspase-3 activation. Experiments with caspase inhibitors demonstrated that caspase-8 was an initiator of this pathway. TNF also induced a transient activation of c-Jun N-terminal kinase (JNK), which upon addition of CDDO was converted to a sustained activation. The activation of JNK was also dependent on caspase-8. Sustained activation of JNK is frequently pro-apoptotic, yet inhibition of JNK did not prevent Bax translocation or cytochrome c release, demonstrating its lack of involvement in CDDO/TNF-induced apoptosis. Apoptosis was acutely induced by CDDO/TNF in every leukemia cell line tested including those that overexpress Bcl-x(L), suggesting that the mitochondrial pathway is not required for apoptosis by this combination. These results suggest that the apoptotic potency of the CDDO/TNF combination occurs through selective inhibition of NF-kappaB-dependent anti-apoptotic proteins, bypassing potential mitochondrial resistance mechanisms, and thus may provide a basis for the development of novel approaches to the treatment of leukemia.
authors.affiliation: Department of Pharmacology and Toxicology, Dartmouth Medical School, Hanover, New Hampshire 03755, USA.
authors.forename: Terrance A
authors.initials: TA
authors.lastname: Stadheim
authors.forename: Nanjoo
authors.initials: N
authors.lastname: Suh
authors.forename: Neema
authors.initials: N
authors.lastname: Ganju
authors.forename: Michael B
authors.initials: MB
authors.lastname: Sporn
authors.forename: Alan
authors.initials: A
authors.lastname: Eastman
mesh_terms: D017209:Apoptosis; D002273:Carcinogens; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D003574:Cytochrome c Group / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D006801:Humans; D020672:I-kappa B Proteins; D015151:Immunoblotting; D048031:JNK Mitogen-Activated Protein Kinases; D007938:Leukemia / Q000473:pathology*; D048670:MAP Kinase Kinase 4; D008928:Mitochondria / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism; D009569:Nitric Oxide / Q000037:antagonists & inhibitors; D009828:Oleanolic Acid / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000494:pharmacology*; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate; D013997:Time Factors; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein
chemical_list: C116817:2-cyano-3,12-dioxoolean-1,9-dien-28-oic acid; C494826:BAX protein, human; C494811:BCL2L1 protein, human; D002273:Carcinogens; D002843:Chromatin; D003574:Cytochrome c Group; D004268:DNA-Binding Proteins; D020672:I-kappa B Proteins; D016328:NF-kappa B; C000606204:NFKBIA protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D014409:Tumor Necrosis Factor-alpha; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D000072000:NF-KappaB Inhibitor alpha; D009569:Nitric Oxide; D009828:Oleanolic Acid; D048031:JNK Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D013755:Tetradecanoylphorbol Acetate