Expression of Smac/DIABLO in ovarian carcinoma cells induces apoptosis via a caspase-9-mediated pathway.
We have constructed Ad CMV-Smac, a recombinant adenovirus encoding Smac/DIABLO, the recently described second mitochondrial activator of caspases. Transfection of ovarian carcinoma cells with Ad CMV-Smac at multiplicities of infection of 3-60 pfu/cell leads to increasing apoptosis in a dose-dependent manner. Western blot analysis confirms that Smac-induced apoptosis proceeds via a pathway mediated primarily by caspase-9 that can be inhibited by zLEHD-fmk and overexpression of the X-linked inhibitor of apoptosis protein (XIAP). In contrast, there is no cleavage of either caspase-8 or caspase-12. Ad CMV-Smac appears to induce apoptosis independently of cytochrome c release from mitochondria and is not inhibited by overexpression of Bcl-2. Ad CMV-Smac can combine with other proapoptotic factors, such as cisplatin, paclitaxel, and procaspase-3, to produce greater levels of apoptosis in transfected cells.
Cancer Research UK Molecular Oncology Unit, Imperial College School of Medicine, Hammersmith Hospital, W12 ONN, London, UK. iain.mcneish@cancer.org.uk
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McNeish
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Bell
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McKay
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Tenev
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Marani
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Lemoine
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics*; D051017:Apoptosis Regulatory Proteins; D002277:Carcinoma / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000235:genetics / Q000378:metabolism*; D003574:Cytochrome c Group / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism*; D010051:Ovarian Neoplasms / Q000188:drug therapy / Q000201:enzymology* / Q000235:genetics; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D014407:Tumor Cells, Cultured; D051636:X-Linked Inhibitor of Apoptosis Protein
D000970:Antineoplastic Agents; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D003574:Cytochrome c Group; C412403:DIABLO protein, human; D004791:Enzyme Inhibitors; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D024101:Mitochondrial Proteins; D011506:Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D011993:Recombinant Fusion Proteins; D051636:X-Linked Inhibitor of Apoptosis Protein; C496866:XIAP protein, human; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases