Correspondence

Epstein–Barr Virus and Nasopharyngeal Cancer

N Engl J Med 1996; 334:122-123January 11, 1996

Article

To the Editor:

The thought-provoking results of Pathmanathan et al. (Sept. 14 issue)1 concerning the role of Epstein–Barr virus (EBV) in nasopharyngeal cancer are of great interest and may have therapeutic implications, especially in areas of the world where the disease is endemic. There is, however, one point I would like clarified.

Do the investigators hold that the only evolutionary path for nasopharyngeal cancer is through infection of an epithelial stem cell followed by transformation, clonal expansion, and development into metastatic cancer? From their study, this would seem to be the case, since the episomal EBV DNA in each case contained a homogeneous terminal and there was no evidence of the expression of viral products that normally signify a productive infection.

In a previous paper, Raab-Traub and Flynn showed that invasive nasopharyngeal cancers also contain unique clonal populations of EBV episomal DNA.2 In situ hybridization was not performed, so it could not be categorically stated that all the tumor cells contained clonal EBV DNA. Moreover, the investigators themselves cited a study reporting that EBV DNA was sporadically distributed throughout a nasopharyngeal cancer in situ.3 These data appear to leave open the possibility that, though a clonal infection with EBV does occur, the involved cells may already have undergone malignant transformation.

It would be helpful if the investigators would comment on the possibility that not all invasive nasopharyngeal cancers develop from an EBV-infected stem cell, but that infection occurs only after carcinoma has developed. Although the EBV genome may not be required for transformation, it is possible that the interaction of the viral and host genomes results in a unique phenotype.

A.I. Meisler, M.D.
Veterans Affairs Medical Center, Pittsburgh, PA 15240

3 References

1
Pathmanathan R, Prasad U, Sadler R, Flynn K, Raab-Traub N. Clonal proliferations of cells infected with Epstein-Barr virus in preinvasive lesions related to nasopharyngeal carcinoma. N Engl J Med 1995;333:693-698
Full Text | Web of Science | Medline

2
Raab-Traub N, Flynn K. The structure of the termini of the Epstein-Barr virus as a marker of clonal cellular proliferation. Cell 1986;47:883-889
CrossRef | Web of Science | Medline

3
Yeung WM, Zong YS, Chiu CT, et al. Epstein-Barr virus carriage by nasopharyngeal carcinoma in situ. Int J Cancer 1993;53:746-750
CrossRef | Web of Science | Medline

To the Editor:

Pathmanathan et al. concluded that only latent, and not lytic, EBV infection is characteristic of preinvasive lesions that may lead to the development of nasopharyngeal cancer. However, there is increasing evidence that the EBV lytic cycle is reactivated in nasopharyngeal cancer and probably also in its preinvasive lesions. For example, the clinical onset of nasopharyngeal cancer is preceded by the appearance of high titers of antibodies to EBV replicative antigens such as EA and VCA. Moreover, most patients with nasopharyngeal cancer have antibodies against the EBV transactivator protein ZEBRA, which induces the switch from the latent to the productive cycle of EBV.1 In contrast to the findings of Pathmanathan et al., Martel-Renoir et al.2 recently showed that specimens of nasopharyngeal cancer expressed the Zebra protein in all cases. Most specimens also expressed the lytic (truncated) form of latent membrane protein 1 (LMP-1), but only 50 percent were positive for the latent form of LMP-1.2 Other lytic genes (BMLF1 and BRLF1) were expressed in a few specimens. These findings suggest that an abortive lytic cycle occurs in nasopharyngeal cancer.2

To identify LMP-1 in neoplastic cells, Pathmanathan et al. used the monoclonal antibodies CS1 through CS4, which are raised against the carboxyl terminal of LMP-1 and which would react not only with the latent but also with the lytic form of LMP-1. Therefore, it seems to be impossible to distinguish between these two different forms of LMP-1 in the histologic sections shown in Figure 1 and Figure 2 of the article by Pathmanathan et al.

Since the intensity of immunostaining for LMP-1 increases near the luminal surface (Figure 2B and Figure 2D of the article) and since the neoplastic specimens showed “some squamous maturation of proliferating cells in the topmost layers” (Figure 1A and Figure 1C), one might suggest that the expression of (lytic?) LMP-1 is dependent on the differentiation status of the epithelial cell, as was recently shown for the BZLF1 promoter of EBV in benign squamous epithelial cells in humans.3

Further studies are needed to show whether the expression of either the latent or the lytic form of LMP-1 can really play a crucial (oncogenic) part in the process of transformation of a previously benign epithelial cell into its malignant counterpart.

Volker Schuster, M.D.
Tanja Pukrop, Ph.D.
University of Würzburg, 97080 Würzburg, Germany

3 References

1
Joab I, Nicolas JC, Schwaab G. Detection of anti-Epstein-Barr-virus transactivator (ZEBRA) antibodies in sera from patients with nasopharyngeal carcinoma. Int J Cancer 1991;48:647-649
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2
Martel-Renoir D, Grunewald V, Touitou R, Schwaab G, Joab I. Qualitative analysis of the expression of Epstein-Barr virus lytic genes in nasopharyngeal carcinoma biopsies. J Gen Virol 1995;76:1401-1408
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3
Karimi L, Crawford DH, Speck S, Nicholson LJ. Identification of an epithelial cell differentiation responsive region within the BZLF1 promoter of the Epstein-Barr virus. J Gen Virol 1995;76:759-765
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Author/Editor Response

Dr. Raab-Traub replies:

To the Editor: In response to Dr. Meisler: the question of whether nasopharyngeal cancer begins to develop before EBV infection or whether EBV subsequently infects the developing tumor was the point of the study. We found that early preinvasive lesions were not only extremely rare but also uniformly infected with EBV that was clonal. Although our original study that identified clonal EBV in nasopharyngeal cancer did not use in situ hybridization, subsequent work has shown that samples of nasopharyngeal cancer consistently contain EBV, with markers of EBV infection in all cells.1,2 The only other study of EBV in dysplasia detected EBV DNA in a subgroup of the cells.3 We believe that the difference may be due to differences in sensitivity between methods that detect EBV-encoded RNA or LMP-1 and in situ DNA hybridization, and that EBV infection occurs before the development of the carcinoma.

In response to Drs. Schuster and Pukrop: abortive or permissive infection with EBV replication does occur in a subgroup of the tumor cells. We have previously shown that transcription associated with viral replication is detectable in some nasopharyngeal cancers, and our analysis of the EBV termini also identified linear forms in some nasopharyngeal cancers.1,4 These findings suggested that viral replication may occur in some cells and might be the source of the antigenic stimulus that promotes the high titers of antibodies to replicative antigens. Indeed, specific replicative transcripts have been detected with expression of the Z antigen in a few cells. Drs. Schuster and Pukrop apparently believe that we may have detected the lytic form of LMP-1 in the tumors. However, since the lytic form of LMP-1 is expressed after Z is produced, it is unlikely that such a late gene product would be detected in all cells, but Z only in rare cells.

Furthermore, the termini assay only detected linear forms at low levels in a single sample of preinvasive neoplasia, and all cells expressed the EBV-encoded RNAs, which are not expressed in hairy leukoplakia, the only known example of a permissive infection in vivo.5 Since rising titers of antibodies to EBV replicative antigens are an excellent prognostic indicator, viral reactivation and replication are important steps in the development of the tumor. However, because herpesvirus replication eventually results in the death of the host cell, it is difficult to envision how a tumor would develop from a permissively infected cell. The fact that Z is detected in rare cells whereas EBV-encoded RNA and LMP-1 are detected in all cells clearly indicates that nasopharyngeal cancer represents a latent, transforming infection.

Nancy Raab-Traub, Ph.D.
Lineberger Comprehensive Cancer Center, Chapel Hill, NC 27599-7295

5 References

1
Raab-Traub N, Flynn K. The structure of the termini of the Epstein-Barr virus as a marker of clonal cellular proliferation. Cell 1986;47:883-889
CrossRef | Web of Science | Medline

2
Wu TC, Mann RB, Epstein JI, et al. Abundant expression of EBER1 small nuclear RNA in nasopharyngeal carcinoma: a morphologically distinctive target for detection of Epstein-Barr virus in formalin-fixed paraffin-embedded carcinoma specimens. Am J Pathol 1991;138:1461-1469
Web of Science | Medline

3
Yeung WM, Zong YS, Chiu CT, et al. Epstein-Barr virus carriage by nasopharyngeal carcinoma in situ. Int J Cancer 1993;53:746-750
CrossRef | Web of Science | Medline

4
Raab-Traub N, Hood R, Yang CS, Henry B II, Pagano JS. Epstein-Barr virus transcription in nasopharyngeal carcinoma. J Virol 1983;48:580-590
Web of Science | Medline

5
Gilligan K, Rajadurai P, Resnick L, Raab-Traub N. Epstein-Barr virus small nuclear RNAs are not expressed in permissively infected cells in AIDS-associated leukoplakia. Proc Natl Acad Sci U S A 1990;87:8790-8794
CrossRef | Web of Science | Medline

Citing Articles (1)

Citing Articles

1
Jian-Hong Lu, Yun-Lian Tang, Hai-Bo Yu, Jian-Hua Zhou, Chun-Yan Fu, Xi Zeng, Zheng-Yuan Yu, Hong-Ling Yin, Ming-Hua Wu, Jun-Yi Zhang, Xiao-Ling Li, Gui-Yuan Li. (2010) Epstein–Barr virus facilitates the malignant potential of immortalized epithelial cells: from latent genome to viral production and maintenance. Laboratory Investigation 90:2, 196-209
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