title: Stabilisation of p53 enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-κB activation.
abstract: 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-κB 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-κB activation in reovirus-infected cells following Nutlin-3a treatment was necessary for enhanced reovirus-induced apoptosis, as synergistic cytotoxicity was overcome by specific NF-κB inhibitors.
  
  
  CONCLUSIONS
  Nutlin-3a treatment enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-κB activation, and combination of reovirus and Nutlin-3a might represent an improved therapy against cancers harbouring wild-type p53.
authors.affiliation: Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 4R2.
authors.forename: D
authors.initials: D
authors.lastname: Pan
authors.forename: L-Z
authors.initials: LZ
authors.lastname: Pan
authors.forename: R
authors.initials: R
authors.lastname: Hill
authors.forename: P
authors.initials: P
authors.lastname: Marcato
authors.forename: M
authors.initials: M
authors.lastname: Shmulevitz
authors.forename: L T
authors.initials: LT
authors.lastname: Vassilev
authors.forename: P W K
authors.initials: PW
authors.lastname: Lee
mesh_terms: D017209:Apoptosis; D015153:Blotting, Western; D002453:Cell Cycle; D045744:Cell Line, Tumor; D002467:Cell Nucleus; D049109:Cell Proliferation; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D007124:Immunoenzyme Techniques; D008156:Luciferases / Q000378:metabolism; D009154:Mutation / Q000235:genetics*; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D010879:Piperazines / Q000494:pharmacology; D021381:Protein Transport; D051736:Proto-Oncogene Proteins c-mdm2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D012087:Reoviridae / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D007093:Imidazoles; D016328:NF-kappa B; D010879:Piperazines; D012333:RNA, Messenger; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; C482205:nutlin 3; D008156:Luciferases; D051736:Proto-Oncogene Proteins c-mdm2