Intracellular regulation of tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis in human multiple myeloma cells.
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL, Apo2 ligand) effectively kills multiple myeloma (MM) cells in vitro irrespective of refractoriness to dexamethasone and chemotherapy. Because clinical trials with this anticancer agent are expected shortly, we investigated the signaling pathway of TRAIL-induced apoptosis in MM. We detected rapid cleavage of caspases-8, -9, -3, and -6, as well as the caspase substrates poly(ADP-ribose) polymerase (PARP) and DNA fragmentation factor-45 (DFF45), but not caspase-10, upon TRAIL treatment in sensitive MM cells, pointing to caspase-8 as the apical caspase of TRAIL signaling in MM cells. These phenomena were not observed or were significantly delayed in TRAIL-resistant MM cells, suggesting that resistance may arise from inhibition at the level of caspase-8 activation. Higher levels of expression for various apoptosis inhibitors, including FLICE-inhibitory protein (FLIP), and lower procaspase-8 levels were present in TRAIL-resistant cells and sensitivity was restored by the protein synthesis inhibitor cycloheximide (CHX) and the protein kinase C (PKC) inhibitor bisindolylmaleimide (BIM), which both lowered FLIP and cellular inhibitor of apoptosis protein-2 (cIAP-2) protein levels. Forced expression of procaspase-8 or FLIP antisense oligonucleotides also sensitized TRAIL-resistant cells to TRAIL. Moreover, the cell permeable nuclear factor (NF)-kappaB inhibitor SN50, which sensitizes TRAIL-resistant cells to TRAIL, also inhibited cIAP2 protein expression. Finally, CHX, BIM, and SN50 facilitated the cleavage and activation of procaspase-8 in TRAIL-resistant cells, confirming that inhibition of TRAIL-induced apoptosis occurs at this level and that these agents sensitize MM cells by relieving this block. Our data set a framework for the clinical use of approaches that sensitize MM cells to TRAIL by agents that inhibit FLIP and cIAP-2 expression or augment caspase-8 activity.
Mitsiades
Nicholas
N
Department of Adult Oncology, Dana Farber Cancer Institute, Harvard Medical School, 44 Binney Street, Mayer Bldg., Boston, MA 02115, USA.
Mitsiades
Constantine S
CS
Poulaki
Vassiliki
V
Anderson
Kenneth C
KC
Treon
Steven P
SP
D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D051017:Apoptosis Regulatory Proteins; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; D002352:Carrier Proteins / Q000378:metabolism / Q000502:physiology; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism / Q000502:physiology; D019008:Drug Resistance, Neoplasm / Q000502:physiology; D004789:Enzyme Activation / Q000502:physiology; D004792:Enzyme Precursors / Q000378:metabolism / Q000502:physiology; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins / Q000378:metabolism / Q000494:pharmacology* / Q000502:physiology; D009101:Multiple Myeloma / Q000188:drug therapy / Q000201:enzymology / Q000473:pathology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D053221:TNF-Related Apoptosis-Inducing Ligand; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism / Q000494:pharmacology* / Q000502:physiology; D014764:Viral Proteins / Q000378:metabolism / Q000502:physiology
D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D053446:CASP8 and FADD-Like Apoptosis Regulating Protein; C506386:CFLAR protein, human; D002352:Carrier Proteins; D004792:Enzyme Precursors; D051034:Inhibitor of Apoptosis Proteins; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; D014764:Viral Proteins; C086430:inhibitor of apoptosis, Nucleopolyhedrovirus; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases