title: X-linked inhibitor of apoptosis (XIAP) blocks Apo2 ligand/tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis of prostate cancer cells in the presence of mitochondrial activation: sensitization by overexpression of second mitochondria-derived activator of caspase/direct IAP-binding protein with low pl (Smac/DIABLO).
abstract: The resistance to Apo2 ligand (Apo2L)/tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated apoptosis could be overcome by treatment with subtoxic concentrations of actinomycin D (Act D) in prostate tumor cells. Furthermore, the sensitization to Apo2L/TRAIL-mediated apoptosis by Act D positively correlated with selective down-regulation of X-linked inhibitor of apoptosis (XIAP). In this study, we examined whether second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pl (Smac/DIABLO), a known inhibitor of apoptosis (IAP)-neutralizing protein, sensitizes resistant prostate tumor cells to Apo2L/TRAIL-mediated apoptosis. The prostate tumor cell line CL-1 was treated with Apo2L/TRAIL, Act D, or a combination of the two. The apoptosis-mediated signaling pathway was examined by Western blotting and flow cytometry. Furthermore, CL-1 cells transfected with the anti-IAP inhibitor Smac/DIABLO were examined for sensitivity to Apo2L/TRAIL. Whereas Apo2L/TRAIL induced the release of cytochrome c and endogenous Smac/DIABLO in the CL-1 tumor cells, the cytosolic levels of both molecules were not sufficient to induce apoptosis. Transient transfectants with a Smac/DIABLO cDNA encoding a neutralizing inhibitor of IAPs were sensitized to Apo2L/TRAIL-mediated apoptosis. The sensitization to Apo2L/TRAIL by Smac/DIABLO overexpression was a result of synergistic activation of caspases-3, -9, and -8. Treatment of the Smac/DIABLO transient transfectant with Apo2L/TRAIL enhanced the release of Smac/DIABLO from mitochondria and led to reduction of IAP family proteins (XIAP, c-IAP1, and c-IAP2). These results show that Smac/DIABLO can sensitize CL-1 tumor cells to Apo2L/TRAIL-mediated apoptosis. Thus, up-regulation of Smac/DIABLO and sensitization to Apo2L/TRAIL-mediated apoptosis are of potential clinical application in the immunotherapy of drug-/Apo2L/TRAIL-resistant tumors.
authors.affiliation: Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles School of Medicine, Jonsson Comprehensive Cancer Center, University of California, Los Angeles, California 90095, USA.
authors.forename: Chuen-Pei
authors.initials: CP
authors.lastname: Ng
authors.forename: Benjamin
authors.initials: B
authors.lastname: Bonavida
mesh_terms: D000911:Antibodies, Monoclonal / Q000378:metabolism; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins / Q000378:metabolism*; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002455:Cell Division; D003574:Cytochrome c Group / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D053938:DNA Fragmentation; D018076:DNA, Complementary / Q000378:metabolism; D003609:Dactinomycin / Q000494:pharmacology; D004789:Enzyme Activation; D005434:Flow Cytometry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D007167:Immunotherapy; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D008297:Male; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism*; D008928:Mitochondria / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000378:metabolism; D011506:Proteins / Q000494:pharmacology / Q000502:physiology*; D015398:Signal Transduction; D053221:TNF-Related Apoptosis-Inducing Ligand; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry* / Q000378:metabolism*; D015854:Up-Regulation; D051636:X-Linked Inhibitor of Apoptosis Protein
chemical_list: D000911:Antibodies, Monoclonal; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D003574:Cytochrome c Group; C412403:DIABLO protein, human; D018076:DNA, Complementary; D047908:Intracellular Signaling Peptides and Proteins; D008024:Ligands; D008562:Membrane Glycoproteins; D024101:Mitochondrial Proteins; D011506:Proteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; D003609:Dactinomycin; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases