No evidence for the presence of Epstein-Barr virus in squamous cell carcinoma of the mobile tongue.

Torben Wilms,Lena Norberg Spaak,Robin Fåhraeus,Luigi Califano,Christos Loizou,Katarina Olofsson,Giuseppe Colella,Nicola Sgaramella,Karin Nylander,Pretty S Philip,Renato Franco,Gulfaraz Khan,Asma Hassani,Philip J Coates,Joseph S Pagano

doi:10.1371/journal.pone.0184201

Abstract

Squamous cell carcinoma of the head and neck (SCCHN) comprises a large group of cancers in the oral cavity and nasopharyngeal area that typically arise in older males in association with alcohol/tobacco usage. Within the oral cavity, the mobile tongue is the most common site for tumour development. The incidence of tongue squamous cell carcinoma (TSCC) is increasing in younger people, which has been suggested to associate with a viral aetiology. Two common human oncogenic viruses, human papilloma virus (HPV) and Epstein-Barr virus (EBV) are known causes of certain types of SCCHN, namely the oropharynx and nasopharynx, respectively. EBV infects most adults worldwide through oral transmission and establishes a latent infection, with sporadic productive viral replication and release of virus in the oral cavity throughout life. In view of the prevalence of EBV in the oral cavity and recent data indicating that it infects tongue epithelial cells and establishes latency, we examined 98 cases of primary squamous cell carcinoma of the mobile tongue and 15 cases of tonsillar squamous cell carcinoma for the presence of EBV-encoded RNAs (EBERs), EBV DNA and an EBV-encoded protein, EBNA-1. A commercially available in situ hybridisation kit targeting EBER transcripts (EBER-ISH) showed a positive signal in the cytoplasm and/or nuclei of tumour cells in 43% of TSCCs. However, application of control probes and RNase A digestion using in-house developed EBER-ISH showed identical EBER staining patterns, indicating non-specific signals. PCR analysis of the BamH1 W repeat sequences did not identify EBV genomes in tumour samples. Immunohistochemistry for EBNA-1 was also negative. These data exclude EBV as a potential player in TSCC in both old and young patients and highlight the importance of appropriate controls for EBER-ISH in investigating EBV in human diseases.

Highlights

Squamous cell carcinoma of the head and neck (SCCHN) comprises a large group of tumours in the head and neck region and occurs mainly in older people who have a history of alcohol and tobacco use
We have previously used PCR for all human papilloma virus (HPV) types and in situ hybridisation for HPV16, the most common high risk HPV seen in association with SCCHN
As HPV is not the only oncogenic virus present in the oral cavity and implicated in the development of tongue squamous cell carcinomas (SCC) (TSCC), we investigated whether there is evidence for a potential role of Epstein-Barr virus (EBV) in this tumour type

Summary

Introduction

Squamous cell carcinoma of the head and neck (SCCHN) comprises a large group of tumours in the head and neck region and occurs mainly in older people who have a history of alcohol and tobacco use. We have recently shown that TSCCs are not associated with HPV [7], leaving EBV as a potentially causative oncogenic virus in this tumour type. During the natural life cycle of EBV, persistent latent infection occurs in B-lymphocytes and epithelial cells in the nasopharynx are thought to be the site for viral replication and shedding into the saliva [8, 9]. This general picture may not be entirely accurate, with recent evidence that EBV may establish latency in basal epithelial cells in the oral cavity (reviewed in [10])

Methods

Results

Conclusion