E2F-1 transcriptional activity is a critical determinant of Mdm2 antagonist-induced apoptosis in human tumor cell lines.
Nutlin-3 is a selective inhibitor of the p53-Mdm2 interaction, and inhibits growth in most tumor cells with wild-type p53. However, it only induces apoptosis in subsets of tumor cells. We report that the apoptotic response induced by Nutlin-3 correlates with its antitumor effects in xenograft models in athymic mice. We have investigated signals that sensitize cells to undergo apoptosis induced by Nutlin-3. We demonstrate that adenovirus E1A increases Nutlin-3-induced apoptosis through pRb inhibition in mouse embryonic fibroblast cells in a p53-dependent manner. Consistent with this, pRb depletion by siRNA transfection with Nutlin-3 synergistically increases apoptosis in HCT116 human colon cancer cells, which are insensitive to induction of apoptosis by Nutlin-3 alone. As pRb is a key negative regulator of E2F, we asked whether E2F transcriptional activity determines the apoptotic response of cancer cells to Nutlin-3. We demonstrate that transcriptional activity of E2F correlates with the apoptotic response to Nutlin-3 in various tumor cells and depletion of E2F-1 suppresses Nutlin-3-induced apoptosis in cells possessing high transcriptional activity of E2F, including retinoblastoma cells harboring mutated Rb with wild-type p53. Furthermore, we report that expression of the p53 homologue p73, a target of E2F-1, is markedly increased by Nutlin-3 in Rb-mutated tumor cells harboring wild-type p53. Depletion of p73 by siRNA transfection suppresses Nutlin-3-induced apoptosis in these cells. Taken together, our results demonstrate that E2F-1 transcriptional activity is a critical determinant of Mdm2 antagonist-induced apoptosis and p73 is important for E2F-1-mediated apoptosis induced by Nutlin-3, especially in tumor cells with mutated Rb. Furthermore, our results suggest that tumor cells, including Rb mutated cells, which harbor wild-type p53 and high E2F transcriptional activity, could be a good target for Mdm2 antagonist therapy.
Helen Diller Family Comprehensive Cancer Center and Cancer Research Institute, University of California, San Francisco, CA 94115, USA.
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Kitagawa
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Aonuma
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Lee
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Fukutake
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McCormick
D000818:Animals; D017209:Apoptosis / Q000502:physiology; D018797:Cell Cycle Proteins; D049109:Cell Proliferation; D002478:Cells, Cultured; D050687:E2F1 Transcription Factor / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D016193:G1 Phase / Q000502:physiology; D006801:Humans; D007093:Imidazoles / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D016688:Mice, Inbred NOD; D008819:Mice, Nude; D009154:Mutation / Q000235:genetics; D009369:Neoplasms / Q000378:metabolism / Q000473:pathology*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010879:Piperazines / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000494:pharmacology; D016192:Resting Phase, Cell Cycle / Q000502:physiology; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism; D013237:Stereoisomerism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism
D018797:Cell Cycle Proteins; D050687:E2F1 Transcription Factor; D007093:Imidazoles; C482775:MDM4 protein, human; D009687:Nuclear Proteins; D010879:Piperazines; D011518:Proto-Oncogene Proteins; D034741:RNA, Small Interfering; D016160:Retinoblastoma Protein; D016159:Tumor Suppressor Protein p53; C482205:nutlin 3; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2