Stabilisation of p53 enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-κB activation

• Published in: British journal of cancer 2011-09, Vol.105 (7), p.1012-1022
• Main Authors: Pan, D, Pan, L-Z, Hill, R, Marcato, P, Shmulevitz, M, Vassilev, L T, Lee, P W K
• Format: Article
• Language: English
• Subjects: 631/250/516/1909; 631/80/82/23; 692/699/67/1059; 692/699/67/581; Annexin V; Apoptosis; Bax protein; Biological and medical sciences; Biomedical and Life Sciences; Biomedicine; Blotting, Western; Cancer; Cancer Research; Cell Cycle; Cell Line, Tumor; Cell Nucleus; Cell Proliferation; Confocal microscopy; Cytotoxicity; Drug Resistance; Epidemiology; Humans; Imidazoles - pharmacology; Immunoenzyme Techniques; Luciferases - metabolism; Medical sciences; Molecular Diagnostics; Molecular Medicine; Mutation; Mutation - genetics; NF- Kappa B protein; NF-kappa B - genetics; NF-kappa B - metabolism; Oncology; Oncolysis; p53 protein; Piperazines - pharmacology; Polymerase chain reaction; Protein Transport; Proto-Oncogene Proteins c-mdm2 - antagonists & inhibitors; Proto-Oncogene Proteins c-mdm2 - genetics; Proto-Oncogene Proteins c-mdm2 - metabolism; Reoviridae - physiology; Reovirus; Replication; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; Titration; Translocation; Tumor cell lines; Tumor Suppressor Protein p53 - chemistry; Tumor Suppressor Protein p53 - genetics; Tumor Suppressor Protein p53 - metabolism; Tumors; Western blotting
• ISSN: 0007-0920; 1532-1827
• DOI: 10.1038/bjc.2011.325

Background: Naturally oncolytic reovirus preferentially kills cancer cells, making it a promising cancer therapeutic. Mutations in tumour suppressor p53 are prevalent in cancers, yet the role of p53 in reovirus oncolysis is relatively unexplored. Methods: Human cancer cell lines were exposed to Nutlin-3a, reovirus or a combination of the two and cells were processed for reovirus titration, western blot, real-time PCR and apoptosis assay using Annexin V and 7-AAD staining. Confocal microscopy was used to determine translocation of the NF- κ B p65 subunit. Results: We show that despite similar reovirus replication in p53 +/+ and p53 −/− cells, stabilisation of p53 by Nutlin-3a significantly enhanced reovirus-induced apoptosis and hence virus release and dissemination while having no direct effect on virus replication. Enhanced apoptosis by Nutlin-3a was not observed in p53 −/− or p53 knockdown cells; however, increased expression of Bax and p21 are required. Moreover, elevated NF- κ B activation in reovirus-infected cells following Nutlin-3a treatment was necessary for enhanced reovirus-induced apoptosis, as synergistic cytotoxicity was overcome by specific NF- κ B inhibitors. Conclusion: Nutlin-3a treatment enhances reovirus-induced apoptosis and virus spread through p53-dependent NF- κ B activation, and combination of reovirus and Nutlin-3a might represent an improved therapy against cancers harbouring wild-type p53.