Regulation of HK2 expression through alterations in CpG methylation of the HK2 promoter during progression of hepatocellular carcinoma

// Hyun Gyu Lee 1 , Hyemi Kim 1, 2 , Taekwon Son 3 , Youngtae Jeong 4 , Seung Up Kim 5 , Seung Myung Dong 6 , Young Nyun Park 2, 7 , Jong Doo Lee 8 , Jae Myun Lee 1, 2 , Jeon Han Park 1 1 Department of Microbiology and Immunology, Institute for Immunology and Immunological Diseases, Yonsei University College of Medicine, Seoul, Republic of Korea 2 Brain Korea 21 PLUS Project for Medical Sciences, Yonsei University College of Medicine, Seoul, Republic of Korea 3 Research Institute of Pharmaceutical Sciences, College of Pharmacy, Seoul National University, Seoul, Republic of Korea 4 Stanford Cancer Institute and Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, California, USA 5 Department of Internal Medicine, Institute of Gastroenterology, Yonsei University College of Medicine, Seoul, Republic of Korea 6 Research Institute, National Cancer Center, Goyang, Gyeonggi-do, Republic of Korea 7 Department of Pathology, Yonsei University College of Medicine, Seoul, Republic of Korea 8 Department of Nuclear Medicine, International St. Mary's Hospital, Catholic Kwandong University College of Medicine, Incheon, Republic of Korea Correspondence to: Jeon Han Park, email: jhpark5277@yuhs.ac Keywords: hexokinase 2, HumanMethylation450 BeadChip, HK2-CIMP, hypoxia response element, HIF-1α Received: March 22, 2016 Accepted: May 23, 2016 Published: May 30, 2016 ABSTRACT Hexokinase 2 (HK2) is a rate-determining enzyme in aerobic glycolysis, a process upregulated in tumor cells. HK2 expression is controlled by various transcription factors and epigenetic alterations and is heterogeneous in hepatocellular carcinomas (HCCs), though the cause of this heterogeneity is not known. DNA methylation in the HK2 promoter CpG island ( HK2 -CGI) and its surrounding regions (shore and shelf) has not previously been evaluated, but may provide clues about the regulation of HK2 expression. Here, we compared HK2 promoter methylation in HCCs and adjacent non-cancerous liver tissues using a HumanMethylation450 BeadChip array. We found that, while the HK2 -CGI N-shore was hypomethylated, thereby enhancing HK2 expression, the HK2 -CGI was itself hypermethylated in some HCCs. This hypermethylation suppressed HK2 expression by inhibiting interactions between HIF-1α and a hypoxia response element (HRE) located at –234/–230. HCCs that were HK2 negative and had distinct promoter CGI methylation were denoted as having a HK2 -CGI methylation phenotype ( HK2 -CIMP), which was associated with poor clinical outcome. These findings indicate that HK2 -CGI N-shore hypomethylation and HK2 -CGI hypermethylation affect HK2 expression by influencing the interaction between HIF 1α and HRE. HK2 -CGI hypermethylation induces HK2 -CIMP and could represent a prognostic biomarker for HCC.