A comparison study of different PCR assays in measuring circulating plasma epstein-barr virus DNA levels in patients with nasopharyngeal carcinoma.
OBJECTIVE
To compare the performance of three PCR assays in measuring circulating Epstein-Barr virus (EBV). DNA levels in nasopharyngeal carcinoma patients and to confirm its prognostic significance.


METHODS
Plasma from 58 newly diagnosed nasopharyngeal carcinoma patients were collected before, during, and every 3 to 6 months after radiotherapy. EBV DNA levels were determined by real-time quantitative PCR using primer/probe sets for polymerase-1 (Pol-1), latent membrane protein 2 (Lmp2), and BamHI-W. Pretreatment levels from the three assays were correlated with each other and serial measurements from the Pol-1 assay were correlated with clinical variables.


RESULTS
Pol-1 was more accurate than BamHI-W in predicting EBV DNA concentrations in cell lines. Of the three assays, BamHI-W yielded the highest concentrations followed by Pol-1 in plasmas (n = 23). The correlation coefficient was 0.99 (P < 0.0001) for Pol-1 and Lmp2, 0.66 (P < 0.0001) for Pol-1 and BamHI-W, and 0.55 (P < 0.0001) for BamHI-W and Lmp2. Elevated pretreatment DNA levels as detected by Pol-1 were correlated with advanced nodal stage (P = 0.04) and overall stage (P = 0.028). There was no correlation between pretreatment EBV DNA levels and freedom-from-relapse or overall survival; however, there was a significant correlation between posttreatment levels and these variables. The 2-year freedom-from-relapse and overall survival rates were 92% and 94% for patients with undetectable, and 37% and 55% for those with detectable, posttreatment levels (P < 0.0001 and P < 0.002).


CONCLUSIONS
The three PCR assays yielded similar results in detecting EBV DNA in plasmas. The Pol-1-detected posttreatment EBV DNA level was the strongest predictor for treatment outcomes.
Le
Quynh-Thu
QT
Department of Radiation Oncology, Stanford University, Stanford, California 94305-5407, USA. qle@stanford.edu
Jones
Carol D
CD
Yau
Tsz-Kok
TK
Shirazi
Haider A
HA
Wong
Priscilla H
PH
Thomas
Ella N
EN
Patterson
Bruce K
BK
Lee
Anne W M
AW
Zehnder
James L
JL
D000293:Adolescent; D000328:Adult; D000368:Aged; D004256:DNA Polymerase I / Q000235:genetics; D004279:DNA, Viral / Q000097:blood*; D015245:Deoxyribonuclease BamHI / Q000235:genetics; D020031:Epstein-Barr Virus Infections / Q000097:blood / Q000473:pathology / Q000821:virology*; D005260:Female; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D008297:Male; D008875:Middle Aged; D009303:Nasopharyngeal Neoplasms / Q000097:blood / Q000473:pathology / Q000821:virology*; D009364:Neoplasm Recurrence, Local; D016133:Polymerase Chain Reaction / Q000379:methods*; D013997:Time Factors; D014763:Viral Matrix Proteins / Q000235:genetics
D004279:DNA, Viral; C037662:EBV-associated membrane antigen, Epstein-Barr virus; D014763:Viral Matrix Proteins; D004256:DNA Polymerase I; D015245:Deoxyribonuclease BamHI