Abstract 4949: Epigenetic silence of ASPP1 and ASPP2 genes promotes tumor growth in HBV-positive hepatocellular carcinoma

Abstract

Abstract The Ankyrin-repeat-, SH3-domain- and proline-rich-region containing protein (ASPP) family of proteins regulates apoptosis through interaction with p53 and its family members. This study evaluates the epigenetic regulation of ASPP1 and ASPP2 in HBV-positive hepatocellular carcinoma (HCC) and explores the effects of down-regulation of ASPP1 and ASPP2 on the development of HCC. The expression of ASPP1 and ASPP2 was diminished in HCC cells by epigenetic silence owing to hypermethylation of ASPP1 and ASPP2 promoters as detected by methylation-specific PCR (MS-PCR) and 5-Aza-2′-Deoxycytidine treatment. Analyses of 51 paired HCC and surrounding non-tumor tissues revealed that methylation of ASPP1 and ASPP2 was associated with the decreased expression of ASPP1 and ASPP2 in tumor tissues (38.1% vs 6.7% in ASPP1, P = 0.018, and 50% vs 15% in ASPP2, P = 0.012). The correlations of the expression and the methylation of ASPP1 and ASPP2 with p53 gene status were further analyzed. HCCs harboring wild type p53 gene were more frequent to have decreased ASPP2 expression (P = 0.028). Whereas no statistic significance was found between down-regulation of ASPP1 and p53 gene status (P = 0.704). Methylation of ASPP1 and/or ASPP2 in the surrounding non-tumor tissues was closely related with the tumor size (P = 0.031) and the tumor stage (P = 0.010). This result indicates that methylation of ASPP1 and ASPP2 may participate in the early stage of neoplasia. Moreover, ASPP2 became methylated upon HBV X protein (HBx) expression. The expression of HBx enhanced the recruitment of DNMT1 and DNMT3A on ASPP2 promoter to initiate DNA methylation, and subsequent increased the binding of methyl-CpG binding proteins on its promoter to suppress ASPP2 expression. The suppressive effects on tumor growth by ASPP1 and ASPP2 were examined with RNA interference mediated gene silence. Down-regulation of ASPP1 and ASPP2 promoted the growth of HCC cells in soft agar and in nude mice, and decreased the sensitivity of HCC cells to apoptotic stimuli. In conclusion, ASPP1 and ASPP2 genes are frequently down-regulated by DNA methylation in HBV-positive HCC, which may play important roles in the development of HCC. These findings provide a new insight into the molecular mechanisms leading to hepatocarcinogenesis and may have potent therapeutic applications. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4949.