Abstract
Caspase-8 is frequently deleted or silenced in neuroblastoma and other solid tumor such as medulloblastoma and small cell lung carcinoma. Caspase-8 expression can be re-established in neuroblastoma cell lines by treatment with demethylating agents or with IFN-gamma. Here we show that four different retinoic acid (RA) derivatives also increase caspase-8 protein expression in neuroblastoma, medulloblastoma and small cell lung carcinoma cell lines. This increase in protein expression is mirrored by an increase in RNA expression in NB cells. However, the promoter region of the caspase-8 gene was not responsible for the induction of caspase-8 expression. Rather, we identified another intronic region containing a CREB binding site that was required for maximal induction of caspase-8 via RA. DNA-protein interaction assays revealed increased phospho-CREB binding to this response element in RA-treated NB cells. Furthermore, mutations of the CREB binding site completely blocked caspase-8 induction in the luciferase reporter system assay and transfection of dominant-negative form of CREB repressed the up-regulation of caspase-8 by RA. Importantly, RA-released cells maintained caspase-8 expression for at least 2-5 days and were more sensitive to doxorubicin and TNFalpha. Thus, RA treatment in conjunction with TNFalpha and/or subsets of cytotoxic agents may have therapeutic benefits.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Antibiotics, Antineoplastic / pharmacology
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Antineoplastic Agents / pharmacology*
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Apoptosis / drug effects*
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Apoptosis / physiology
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Blotting, Western
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Carcinoma, Small Cell / drug therapy
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Carcinoma, Small Cell / genetics
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Carcinoma, Small Cell / pathology
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Caspase 8 / genetics*
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Caspase 8 / metabolism
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Cell Proliferation / drug effects
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Chromatin Immunoprecipitation
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Cyclic AMP Response Element-Binding Protein / genetics
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Cyclic AMP Response Element-Binding Protein / metabolism*
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DNA Methylation
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Doxorubicin / pharmacology
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Electrophoretic Mobility Shift Assay
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Humans
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Introns / genetics
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Luciferases / metabolism
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Lung Neoplasms / drug therapy
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Lung Neoplasms / genetics
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Lung Neoplasms / pathology
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Medulloblastoma / drug therapy
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Medulloblastoma / genetics
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Medulloblastoma / pathology
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Mutation / genetics
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Neuroblastoma / drug therapy
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Neuroblastoma / genetics
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Neuroblastoma / pathology*
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Phosphorylation / drug effects
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Promoter Regions, Genetic
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RNA, Messenger / genetics
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RNA, Messenger / metabolism
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Response Elements / genetics
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Signal Transduction
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Transcription, Genetic*
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Transcriptional Activation
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Tretinoin / pharmacology*
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Tumor Cells, Cultured
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Tumor Necrosis Factor-alpha / pharmacology
Substances
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Antibiotics, Antineoplastic
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Antineoplastic Agents
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Cyclic AMP Response Element-Binding Protein
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RNA, Messenger
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Tumor Necrosis Factor-alpha
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Tretinoin
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Doxorubicin
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Luciferases
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Caspase 8