Abstract Liver cancer, primarily hepatocellular carcinoma (HCC), is the fifth most common cancers and the third leading cause of cancer mortality worldwide. In recent years, deregulated epigenetic mechanisms were found important in a wide range of human carcinogenesis, including HCC. The epigenetic changes elicited by various environmental factors and lifestyles (such as hepatitis B virus (HBV) and hepatitis C virus (HCV) infection, chronic alcohol intake) were suspected to promote HCC development. However, the underlying mechanism and specific gene targets are still unknown. Our previous study suggested that deregulation of epigenetic regulators was a common event in human HCC. Among 90 epigenetic modifiers analyzed in human HCC, Euchromatic histone-lysine N-methyltransferase 2 (EHMT2/G9a) was the most significantly up-regulated one. G9a is a SET domain containing histone methyltransferase specific for histone H3 lysine 9 mono- and di-methylation. Previous work had revealed that G9a is important for embryonic development and adipocyte differentiation. However, little is known about functional roles of G9a in human HCC. We hypothesize that deregulation of G9a may led to aberrant epigenetic changes and contribute to initiation and progression of human HCC. In this study, we found that G9a mRNA expression in primary HCC was 3 folder than their corresponding non-tumor liver and 10 fold higher than normal livers. The up-regulation of G9a was detected in 63% (24/38) of primary HCC samples and was significantly associated with the presence of venous invasion (P = 0.006). The frequent G9a up-regulation in HCC was attributed to gene amplification and microRNA deregulation. We showed that 53% of HCC samples have 3 or more copies of G9a gene. In addition, we identified that miR-1 was a post-transcriptional regulator of G9a. Ectopic expression of miR-1 suppressed G9a 3′-untranslated-region-coupled luciferase activity. Functionally, knockdown of G9a significantly suppressed HCC cell proliferation, migration and colony formation. We also demonstrated that UNC0638, as a G9a specific inhibitor, could effectively inhibit HCC cell growth and cause cell morphology changes. By utilizing nude mice model, we showed that depletion of G9a drastically inhibited in vivo tumorigenicity. Our finding suggested that up-regulation of G9a contributed to the liver carcinogenesis and could be a therapeutic target for HCC treatment. Citation Format: Lai Wei, Felice Ho-Ching Tsang, Jeffery Chi-Fai Lau, Sandy Leung-Kuen Au, Joyce Man-Fong Lee, Carmen Chak-Lui Wong, Irene Oi-Lin Ng, Chun-Ming Wong. Deregulation of G9a and its functional roles in liver cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5147. doi:10.1158/1538-7445.AM2014-5147