Geldanamycin decreases Raf-1 and Akt levels and induces apoptosis in neuroblastomas.
Neuroblastomas are the most common extracranial solid tumors of childhood. These tumors are associated with an overall poor prognosis, particularly for advanced stage disease. The benzoquinone ansamycin antibiotic, geldanamycin (GA), exhibits potent antitumor activity in certain cancer cell lines by destabilizing important signal transduction proteins (e.g., Raf-1 and Akt). The purpose of our study was to determine whether GA can alter the expression of Raf-1 and Akt, which have been shown to be critical for neuronal cell survival, and induce apoptosis of neuroblastoma cells. Human neuroblastoma cells (SH-SY5Y, SK-N-SH and LAN-1) were treated with GA for a variable period of time. Cell viability was assessed with MTT assays. Apoptosis was assessed with DNA fragmentation ELISA, TUNEL-flow cytometric assay, Western blot and caspase activities. We found that GA decreases cell viability and induces apoptosis in the SH-SY5Y human neuroblastoma cell line. These effects were mediated through activation of caspase-9 and -3, mitochondrial release of cytochrome c and subsequent PARP cleavage. GA-induced apoptosis was associated with a reduction in the level and activity of Raf-1 and Akt. The importance of these proteins was further demonstrated by induction of apoptosis in SH-SY5Y cells by a combination of U0126 (MEK1/2 inhibitor) and LY294002 (an inhibitor of PI3K). Similar to SH-SY5Y cells, other human neuroblastoma cells (SK-N-SH and LAN-1) were sensitive to the effects of GA-induced apoptosis. Taken together, our findings suggest that GA may be a novel therapeutic agent, which may be effective in the treatment of neuroblastomas.
Kim
Sunghoon
S
Department of Surgery, The University of Texas Medical Branch, Galveston 77555, USA.
Kang
Junghee
J
Hu
Wanqin
W
Evers
B Mark
BM
Chung
Dai H
DH
D017209:Apoptosis / Q000187:drug effects*; D016227:Benzoquinones; D015153:Blotting, Western; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D003574:Cytochrome c Group / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D047029:Lactams, Macrocyclic; D020935:MAP Kinase Signaling System; D008928:Mitochondria / Q000502:physiology*; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D011233:Precipitin Tests; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D011809:Quinones / Q000494:pharmacology*; D015398:Signal Transduction / Q000502:physiology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
D016227:Benzoquinones; D061945:Caspase Inhibitors; D003574:Cytochrome c Group; D004791:Enzyme Inhibitors; D047029:Lactams, Macrocyclic; D011518:Proto-Oncogene Proteins; D011809:Quinones; D011065:Poly(ADP-ribose) Polymerases; D011505:Protein-Tyrosine Kinases; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D019908:Proto-Oncogene Proteins c-raf; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; C001277:geldanamycin