Cathryn E. Tune, Per-Gunnar Liavaag, Jeremy L. Freeman, Michiel W. M. van den Brekel, Thomas Shpitzer, Jeroen D. F. Kerrebijn, David Payne, Jonathan C. Irish, Raymond Ng, Roy K. Cheung, Hans-Michael Dosch

Affiliations of authors: C. E. Tune, P.-G. Liavaag, R. K. Cheung, H.-M. Dosch, Division of Immunology and Cancer Research, Hospital for Sick Children, Toronto, ON, Canada; J. L. Freeman, M. W. M. van den Brekel, T. Shpitzer, J. D. F. Kerrebijn, Department of Otolaryngology, Mount Sinai Hospital, Toronto; D. Payne, Division of Radiation Oncology, Princess Margaret Hospital, Toronto; J. C. Irish, Department of Otolaryngology, Toronto Hospital-General Division; R. Ng, Centenary Health Center, Scarborough, ON.

BACKGROUND: Nasopharyngeal carcinoma (NPC) is an important tumor in many countries. Ethnic and regional factors strongly influence disease risk. NPC is usually diagnosed late in disease development, and 10-year survival rates are as low as 10%. Epstein-Barr virus (EBV), a possibly causative agent, is present in all cells of essentially all undifferentiated NPCs. We wished to determine the following: 1) whether an ambulatory nasopharyngeal brush biopsy could provide sufficient tumor cell DNA for the detection of EBV and 2) whether the detection of EBV in this locale reflects the presence of tumor cells or simply EBV carrier status. METHODS: We collected nasopharyngeal tissue via ambulatory brush biopsies from 21 patients with newly diagnosed NPC and from 157 subjects with other otolaryngologic complaints. The majority of study subjects were from high-risk populations. Sample DNA was analyzed for the presence of EBV genomic sequences by use of the polymerase chain reaction (PCR). RESULTS: Ninety-six percent of samples yielded sufficient DNA for PCR amplification. Nineteen of 21 patients with NPC brushed positive for EBV DNA, while all but two (1.3%) of 149 informative control subjects were negative for EBV (two-sided P<.0001). One of the EBV-positive control subjects had an EBV-positive inverted sinonasal papilloma; the other EBV-positive control subject exhibited no overt clinical disease. CONCLUSION: Demonstration of EBV DNA in nasopharyngeal brush biopsy specimens detects NPC with a sensitivity of at least 90% (95% confidence interval = 89.63%-91.32%) and a specificity of approximately 99% (95% confidence interval = 98.64%-98.68%). This technique merits further testing as a possible ambulatory screening strategy in high-risk populations.

Nasopharyngeal carcinoma (NPC) is a relatively rare tumor in Caucasians, but it occurs at high frequency in Pacific and Mediterranean rim countries, among the Inuit, and in coastal regions of Africa. NPC has strong genetic and environmental roots (1); e.g., 1%-8% of Southern Chinese are at risk of developing NPC (2). In Chinese immigrants to North America, NPC risk declines rapidly, but it remains 40-fold to 50-fold higher than that of resident Caucasians (3). More than 90% of NPC patients in North America are of Asian, Mediterranean, or African descent. In a worldwide perspective, NPC is a major cause of morbidity and mortality.

With often minimal, nonspecific local symptoms (4), more than two thirds of patients are diagnosed only late in the disease process by endoscopy and/or biopsies of metastatic lymph nodes (5). Primary radiotherapy of NPC usually induces remission, but relapse rates are high and are associated with increasing resistance to therapy (6). Important sites of treatment failure are the nasopharynx and distant metastases; 10-year survival rates are as low as 10% (5). Earlier detection would almost certainly improve overall survival (7).

Epstein-Barr virus (EBV) is a large herpesvirus carried by nearly all human adults; homologous viruses are carried by nonhuman primates (8). EBV is usually latent, infrequently producing infectious progeny (9). Cells maintain from few to more than 50 virus copies as nonintegrated, double-stranded DNA episomes (10). B lymphocytes constitute the likely sole EBV reservoir, while persistent infection of epithelial cells is characteristic of malignant tumors such as NPC (11).

EBV is closely and perhaps causally linked to NPC; essentially all tumors and preinvasive lesions of the nasopharynx (12) carry clonal EBV genomes, and viral infection precedes malignant transformation (13-15). Accordingly, detection of EBV genomic DNA in nasopharyngeal biopsy specimens may predict the presence of NPC (16). However, the practical utility of diagnostic EBV detection is uncertain, since most human adults carry the virus. The high prevalence of EBV-specific serologies in carriers reduces its utility for prediction of NPC risk: Population screening through serology and endoscopy has not been satisfactory (17,18). We thus sought to test in a clinical setting an ambulatory brush biopsy procedure with polymerase chain reaction (PCR)-based EBV detection for identification of subjects who are likely to harbor NPC.

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