Reactive Nitrogen Species-dependent DNA Damage in EBV-associated Nasopharyngeal Carcinoma: the Relation to STAT3 Activation and EGFR Expression

Nasopharyngeal carcinoma (NPC) is strongly associated with Epstein-Barr virus (EBV) infection. Recently, reactive nitrogen and oxygen species are considered to participate in inflammation-related carcinogenesis through DNA damage. In our study, we obtained biopsy and surgical specimens of nasopharyngeal tissues from NPC patients in southern China, and performed double immunofluorescent staining to examine the formation of 8-nitroguanine, a nitrative DNA lesion and 8-oxo-7,8-dihydro-2'-deoxyguanosine, an oxidative DNA lesion, in these specimens. Strong DNA lesions were observed in cancer cells and inflammatory cells in stroma of NPC patients. Intensive immunoreactivity of iNOS was detected in the cytoplasm of 8-nitroguanine-positive cancer cells. DNA lesions and iNOS expression were also observed in epithelial cells of EBV-positive patients with chronic nasopharyngitis, although their intensities were significantly weaker than those in NPC patients. In EBV-negative subjects, no or little DNA lesions and iNOS expression were observed. EGFR and phosphorylated STAT3 were strongly expressed in cancer cells of NPC patients, but NF-kappaB was not expressed, suggesting that STAT3-dependent mechanism is important for NPC carcinogenesis. IL-6 was expressed mainly in inflammatory cells of nasopharyngeal tissues of EBV-infected patients. EBV-encoded RNAs (EBERs) and latent membrane protein 1 (LMP1) were detected in cancer cells from all EBV-infected patients. In vitro cell system, nuclear accumulation of EGFR was observed in LMP1-expressing cells, and IL-6 induced phosphorylated STAT3 and iNOS. These data suggest that nuclear accumulation of EGFR and STAT3 activation by IL-6 play the key role in iNOS expression and resultant DNA damage, leading to EBV-mediated NPC.