EBV-associated histone modifications regulate DNA damage repair pathway and associate with cisplatin resistance in nasopharyngeal epithelial malignancy

Introduction: Epstein-Barr virus (EBV) infection is associated with various cancers. Some studies have confirmed EBV can induce hypermethylation to suppress the expression of several key cancer-related genes in gastric cancer. Previously, we showed that nasopharyngeal carcinoma (NPC), an EBV-associated malignancy, is characterized by low mutation rate, but harbors extensive hypermethylation compared with other cancer types. Also, the de novo methylated regions in NPC extensively overlap with the histone bivalent marks (H3K4me3 and H3K27me3) derived from human embryonic stem cells . This suggests an important role of EBV in regulating histone modifications in NPC development. Hence, further studies focusing on the EBV-associated histone modifications are necessary. Aim: This study aims to elucidate the role of EBV in regulating a transcription-activation histone mark (H3K4me3) and a suppressive mark (H3K27me3) in nasopharyngeal epithelial (NPE) cells. Methodologies: Two pairs of EBV+/- NPE cell lines were used for chromatin immunoprecipitation sequencing (ChIP-Seq) to identify the genes regulated by EBV-associated histone modifications. ChIP-QPCR and RT-QPCR were utilized to validate the ChIP-Seq results. The candidate genes and pathways identified from the ChIP-Seq results were validated by in vitro and in vivo functional studies. Results: A panel of 16 DNA damage repair family members were identified with significant reduction of H3K4me3 in the EBV+ NPE cells. One of the candidate genes, MLH1, was validated with down-regulation in the EBV+ NPC cell lines and NPC specimens, and the expression level was demonstrated to associate with cisplatin resistance. Also, the base excision repair (BER) pathway was significantly enriched with reduction of H3K4me3 after EBV infection. Down-regulation of the 7 members from BER was validated in the EBV+ NPE cell lines and NPC specimens. The defective DNA damage repair in the EBV+ NPE cell lines was further validated by the comet assay. Furthermore, after EBV infection, gain of H3K27me3 in MLH1 and the 7 members from BER pathway was identified by ChIP-QPCR. Interestingly, our results from bisulfite sequencing reveals that the promoter regions of the candidate genes are hypomethylated in both EBV +/- NPE cells. Conclusion: EBV modulates histone bivalent marks independent of the promoter DNA methylation status to regulate DNA damage repair in the host cells. Acknowledgements: This work was supported by the Research Grants Council of the Hong Kong Special Administrative Region, People’s Republic of China: Grant number AoE/M-06/08 to MLL and Seed Funding Programme for Basic Research of HKU 201308159003 to AKLC. Citation Format: Merrin Man Long Leong, Arthur Kwok Leung Cheung, Wei Dai, Sai Wah Tsao, Maria Li Lung. EBV-associated histone modifications regulate DNA damage repair pathway and associate with cisplatin resistance in nasopharyngeal epithelial malignancy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5192.