Integrative bioinformatics prioritises exosome-associated Epstein- Barr virus microRNA targets in gastric cancer

Abstract Background Epstein-Barr virus (EBV)-associated gastric cancer (EBVaGC) is characterised by substantial immune-cell infiltration and frequent amplification or overexpression of programmed death (PD)-ligand 1 and PD-ligand 2. Nevertheless, the molecular mechanisms that may contribute to immune regulation in this cancer subtype remain incompletely understood. EBV-positive SNU719 gastric cancer cells produce BamHI-A rightward transcript (BART) microRNAs (miRNAs), some of which are packaged into exosomes while also remaining detectable in the parental cells. This study aimed to identify EBV BART miRNAs relatively enriched in SNU719-derived exosomes and to investigate their candidate interactions with human transcripts involved in innate immune signalling. Results Publicly available datasets were integrated through a CLASH-guided multilayer computational framework. Parental-cell and exosomal miRNA profiles were compared to identify exosome-enriched EBV BART miRNAs. SNU719-specific CLASH data were then used to identify reproducible viral miRNA interactions with human transcripts in the toll-like receptor (TLR)/nuclear factor kappa-light-chain-enhancer of activated B cells/interferon regulatory factor pathway. Network topology and perturbation analyses ranked the most influential miRNA candidates, while SNU719 RNA-seq data confirmed expression of most prioritised human target transcripts. This integrated analysis identified 11 reproducible miRNA-target interactions and prioritised BART17-5p, BART8-3p, and BART10-3p as the strongest candidate exosome-associated EBV BART miRNAs. The Cancer Genome Atlas-Stomach Adenocarcinoma analysis did not reveal significant EBV-status-dependent differences in bulk TLR-related pathway scores. Conclusions The study provides a reusable computational framework for prioritizing exosome-associated viral miRNA-human target interactions from public multi-omic datasets. BART17-5p, BART8-3p, and BART10-3p emerged as experimentally testable candidates linked to innate immune signalling in EBVaGC.