title: Quantitative Epstein-Barr virus DNA analysis and detection of gene promoter hypermethylation in nasopharyngeal (NP) brushing samples from patients with NP carcinoma.
abstract: OBJECTIVE
  Nasopharyngeal carcinoma (NPC) is highly prevalent in southern China and characterized by a strong association with EBV. We aimed to detect EBV DNA and cancer-related gene promoter hypermethylation in nasopharyngeal (NP) brushing samples and provide a novel noninvasive approach for NPC detection.
  
  
  METHODS
  Twenty-eight NPC cases and 26 noncancerous subjects were prospectively recruited. NP brushing samples were subjected to quantitative real-time PCR analysis of EBV DNA and methylation-specific PCR analysis of the DAP-kinase, RASSF1A, and p16 genes.
  
  
  RESULTS
  EBV DNA quantity in NP brushing samples from NPC patients (median, 8.94 copies/actin) was significantly higher than that of controls (median, 0 copies/actin; P < 0.0001). Twenty-seven of 28 NPC patients had detectable EBV DNA in NP brushes, whereas 25 of 26 controls had undetectable or very low levels of EBV DNA. Elevated EBV DNA level in brushing samples as a tumor marker had a sensitivity of 96.4% and a specificity of 96.2% for NPC detection. Moreover, T(1) disease had a significantly lower EBV DNA level as compared with locally more advanced disease (P = 0.037). In brushing samples of NPC patients, the frequencies of DAP-kinase, RASSF1A, and p16 promoter hypermethylation were 50.0%, 39.3%, and 46.4%, respectively. Seventy-eight percent of cases showed methylation of at least one gene. No aberrant hypermethylation was detected in control samples.
  
  
  CONCLUSIONS
  Our study demonstrated the feasibility of detecting multiple molecular tumor markers in NP brushing samples with a high sensitivity and specificity for NPC detection. It offers a powerful yet noninvasive approach for the diagnosis of NPC in high-risk populations.
authors.affiliation: Department of Anatomical and Cellular Pathology, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong Special Administrative Region, People's Republic of China.
authors.forename: Joanna H M
authors.initials: JH
authors.lastname: Tong
authors.forename: Raymond K Y
authors.initials: RK
authors.lastname: Tsang
authors.forename: Kwok-Wai
authors.initials: KW
authors.lastname: Lo
authors.forename: John K S
authors.initials: JK
authors.lastname: Woo
authors.forename: Joseph
authors.initials: J
authors.lastname: Kwong
authors.forename: Michael W Y
authors.initials: MW
authors.lastname: Chan
authors.forename: Alexander R
authors.initials: AR
authors.lastname: Chang
authors.forename: Charles A
authors.initials: CA
authors.lastname: van Hasselt
authors.forename: Dolly P
authors.initials: DP
authors.lastname: Huang
authors.forename: Ka-Fai
authors.initials: KF
authors.lastname: To
mesh_terms: D000328:Adult; D000368:Aged; D002277:Carcinoma / Q000097:blood / Q000821:virology*; D002681:China; D019175:DNA Methylation; D004279:DNA, Viral; D005260:Female; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D007070:Immunoglobulin A / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009303:Nasopharyngeal Neoplasms / Q000097:blood / Q000821:virology*; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity
chemical_list: D004279:DNA, Viral; D007070:Immunoglobulin A