Abstract
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.
MeSH terms
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Antineoplastic Agents / pharmacology
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Apoptosis / drug effects
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Apoptosis / genetics*
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Apoptosis Regulatory Proteins
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Carcinoma / drug therapy
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Carcinoma / enzymology*
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Carcinoma / genetics
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Carrier Proteins / genetics
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Carrier Proteins / metabolism*
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Caspase 3
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Caspase 9
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Caspases / genetics
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Caspases / metabolism*
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Cytochrome c Group / metabolism
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Enzyme Inhibitors / pharmacology
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Enzyme Precursors / genetics
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Enzyme Precursors / metabolism
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Female
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Gene Expression Regulation, Neoplastic / genetics*
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Genetic Vectors
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Humans
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Intracellular Signaling Peptides and Proteins
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Mitochondrial Proteins / genetics
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Mitochondrial Proteins / metabolism*
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Ovarian Neoplasms / drug therapy
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Ovarian Neoplasms / enzymology*
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Ovarian Neoplasms / genetics
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Proteins / genetics
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Proteins / metabolism
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Proto-Oncogene Proteins c-bcl-2 / genetics
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Proto-Oncogene Proteins c-bcl-2 / metabolism
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Recombinant Fusion Proteins
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Tumor Cells, Cultured
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X-Linked Inhibitor of Apoptosis Protein
Substances
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Antineoplastic Agents
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Apoptosis Regulatory Proteins
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Carrier Proteins
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Cytochrome c Group
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DIABLO protein, human
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Enzyme Inhibitors
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Enzyme Precursors
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Intracellular Signaling Peptides and Proteins
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Mitochondrial Proteins
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Proteins
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Proto-Oncogene Proteins c-bcl-2
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Recombinant Fusion Proteins
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X-Linked Inhibitor of Apoptosis Protein
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XIAP protein, human
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CASP3 protein, human
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CASP9 protein, human
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Caspase 3
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Caspase 9
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Caspases