Abstract P52: Towards Profiling DNA Modifications of Virus and Human Host DNA in Epstein-Barr Virus (EBV)-positive Nasopharyngeal Carcinoma (NPC) with Nanopore Ultra-Long Reads Sequencing

Abstract Nasopharyngeal Carcinoma (NPC) is a highly metastatic malignancy, often diagnosed at advanced stages with poor prognosis and survival rates. It is strongly associated with Epstein-Barr virus (EBV) exposure and Chinese/Asian ethnic backgrounds, leading to approximately 300 new cases annually in Singapore. EBV plays a critical role in NPC development, with both its latent and lytic phases contributing to tumorigenesis. The virus precisely controls its methylation pattern to regulate latent gene expression and lytic reactivation. In contrast, disruptions in DNA modification patterns in humans are frequently observed in tumorigenesis, suggesting transcriptional dysregulation. Currently, few studies have explored limited loci in human and EBV epigenomes, with most only profiling population-level average methylation status in NPC. These studies cannot distinguish between 5-hydroxymethylcytosine (5hmC) and 5-methylcytosine (5mC), which have opposing regulatory roles. Moreover, existing methods fail to access repetitive regions, which comprise significant portions of both the EBV and human genomes. To address these gaps, we generated whole-genome, single-molecule, single-base resolution profiles of 5mC and 5hmC modifications in EBV and human DNA from different NPC tumor samples and cell lines using direct Nanopore ultralong-read sequencing. This approach offers unique insights into the complexities of EBV and human DNA modifications during NPC development. We discovered that distinct EBV strains cause contrasting 5mC changes in both human and EBV epigenomes, with some inducing hypermethylation and others hypomethylation. While hypomethylation was previously linked to APOBEC activity, our findings suggest that EBV strain-specific sequences play a significant role. Notably, in EBV epigenome, 5mC differences are concentrated in repetitive regions, whereas 5hmC profiles are consistent across strains, with persistent 5hmC modifications at specific loci. Our study provides valuable epigenome knowledge to the field of NPC research. Further data and detailed mechanistic studies are crucial to deepening our understanding of disease onset, progression, and advancing early detection and therapeutic strategies. Citation Format: Li Zhe, Yao Fei, Sia Yee Yen, Melvin Chua Lee Kiang, Joshua Tay, Liu Jianjun. Towards Profiling DNA Modifications of Virus and Human Host DNA in Epstein-Barr Virus (EBV) -positive Nasopharyngeal Carcinoma (NPC) with Nanopore Ultra-Long Reads Sequencing abstract. In: Proceedings of Frontiers in Cancer Science 2024; 2024 Nov 13-15; Singapore. Philadelphia (PA): AACR; Cancer Res 2025;85 (15Suppl): Abstract nr P52.