Epstein-Barr Virus in HIV-Associated Conjunctival Squamous Cell Carcinoma and Transcriptional Reprogramming of Conjunctival Epithelial Cells

The involvement of Epstein-Barr virus (EBV) in conjunctival squamous cell carcinoma (cSCC) remains incompletely defined. This study integrated analyses of primary tumors with an experimental model of the conjunctival epithelium. Five Nigerian tumors from patients with HIV and three Japanese tumors from patients without HIV were examined by EBV-encoded RNA in situ hybridization (EBER-ISH), EBV DNA PCR, and virome analysis of RNA sequencing data. EBER localized to epithelial tumor cells in four Nigerian specimens and none of the Japanese specimens. EBV DNA and RNA were each detected in three Nigerian tumors, with every EBER-positive tumor supported by at least one additional assay. Human telomerase reverse transcriptase-immortalized conjunctival epithelial cells infected with Akata-GFP-EBV expressed latency-associated and lytic viral genes. Comparison of three infected and three control cultures identified 96 differentially expressed host genes and 23 enriched Hallmark gene sets at a false discovery rate below 0.05. RT-qPCR confirmed increased HES1, CXCL8, CXCL2, and LOX and decreased FGFBP1, CSTA, SAPCD2, and TMEM40 expression. Prominent responses involved inflammatory signaling, epithelial remodeling, metabolism, and cell-cycle regulation. These findings identify tumor-cell-associated EBV in a subset of HIV-associated cSCC and establish an epithelial model for investigating infection-associated host responses.