title: Loss of p16 expression has prognostic significance in human nasopharyngeal carcinoma.
abstract: OBJECTIVE
  p16 is an important inhibitor of cell cycle progression; absence of p16 can thus result in increased cellular proliferation. In nasopharyngeal carcinoma (NPC), absence of p16 has been reported in association with presence of the EBV and pRb. We therefore examined p16, pRb, and EBV-encoded RNA (EBER) expression in biopsy specimens from 84 patients with newly diagnosed NPC, who were treated primarily with curative radiation therapy. Our objective was to determine whether there was a correlation between these parameters and clinical outcome in NPC.
  
  
  METHODS
  Sections were cut from archival formalin-fixed, paraffin-embedded tumor blocks from NPC patients. p16 and pRb expression were determined using polyclonal and monoclonal antibodies, respectively. The presence of EBV was determined by in situ hybridization for EBER. The percentage of positively staining tumor nuclei was scored for p16 or pRb immunoreactivity. Relative intensity and proportion of cells with EBER signals were also documented.
  
  
  RESULTS
  p16 expression was reduced (</=5% positive immunoexpression) in 59 of 84 (70%) NPC samples; in contrast, pRb was observed in all (100%) tumors. EBER signals were detected in 67 of 83 (81%) NPC specimens. There was a weak correlation between EBER presence and loss of p16 (P = 0.1). Using a Cox regression model controlling for known prognostic parameters, such as age, weight loss, and tumor stage, complete absence of p16 expression (0%, i.e., no immunostaining identified throughout the specimen) was associated with an inferior overall survival rate (P = 0.022). In addition, EBER-positive NPC was strongly associated with improved overall survival (P = 0.005) as reported previously (Shi et al., Cancer, 94: 1997, 2002).
  
  
  CONCLUSIONS
  These results provide the first evidence suggesting that inactivation of p16 appears to be a significant predictor for poor overall survival in NPC. Given that reduced p16 expression is observed in the majority of patients with NPC, this indicates that therapeutic strategies targeting the p16 pathway may be a biologically rational approach for NPC. The favorable prognostic value of EBER suggests that future clinical trials with NPC should consider stratifying for EBER status.
authors.affiliation: Department of Otolaryngology, Princess Margaret Hospital/Ontario Cancer Institute, Toronto M5G 2M9, Canada.
authors.forename: Antti A
authors.initials: AA
authors.lastname: Mäkitie
authors.forename: Christina
authors.initials: C
authors.lastname: MacMillan
authors.forename: James
authors.initials: J
authors.lastname: Ho
authors.forename: Wei
authors.initials: W
authors.lastname: Shi
authors.forename: Andrew
authors.initials: A
authors.lastname: Lee
authors.forename: Brian
authors.initials: B
authors.lastname: O'Sullivan
authors.forename: David
authors.initials: D
authors.lastname: Payne
authors.forename: Melania
authors.initials: M
authors.lastname: Pintilie
authors.forename: Bernard
authors.initials: B
authors.lastname: Cummings
authors.forename: John
authors.initials: J
authors.lastname: Waldron
authors.forename: Padraig
authors.initials: P
authors.lastname: Warde
authors.forename: Jonathan
authors.initials: J
authors.lastname: Irish
authors.forename: Dale
authors.initials: D
authors.lastname: Brown
authors.forename: Ralph
authors.initials: R
authors.lastname: Gilbert
authors.forename: Patrick
authors.initials: P
authors.lastname: Gullane
authors.forename: Fei-Fei
authors.initials: FF
authors.lastname: Liu
authors.forename: Suzanne
authors.initials: S
authors.lastname: Kamel-Reid
mesh_terms: D000328:Adult; D000368:Aged; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000032:analysis*; D018572:Disease-Free Survival; D005260:Female; D004854:Herpesvirus 4, Human / Q000235:genetics; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D009303:Nasopharyngeal Neoplasms / Q000737:chemistry* / Q000401:mortality / Q000821:virology; D011379:Prognosis; D012367:RNA, Viral / Q000032:analysis; D016160:Retinoblastoma Protein / Q000032:analysis
chemical_list: D019941:Cyclin-Dependent Kinase Inhibitor p16; D012367:RNA, Viral; D016160:Retinoblastoma Protein