Promoter Methylation status of HIN-1 associated with outcomes of ovarian clear cell adenocarcinoma

Background: This study is to analyze promoter methylation of various tumor suppressor genes in different types of ovarian carcinoma and to identify potential therapeutic targets of ovarian clear cell adenocarcinoma (OCCA). Materials and Methods: The promoter methylation statuses of 40 genes in primary ovarian carcinomas including 47 clear- and 63 non-clear-cell type tissues, 6 OCCA cell lines, 29 benign ovarian endometriotic cysts, and 31 normal controls were analyzed by methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA). The MS-MLPA results were correlated with clinicopathological features and outcomes of 47 OCCA patients. Functions of the target genes were further explored by Western Blot Analysis, apoptosis assay, and caspase-3/7 activity analysis. Results: Frequencies of methylated RASSF1A, CDH13, CACNA1A, HIN-1, and sFRP5 genes in OCCA tissues were significantly higher than those in non-OCCA cancerous tissues and benign endometriotic cysts. The expected 5-year PFS and OS for patients with methylated promoters of HIN-1 and CACNA1A genes were significantly worse than those for patients without methylated HIN-1 (28% vs. 54%, p = 0.047 for PFS; 30% vs. 62%, p = 0.002 for OS, respectively) and CACNA1A (30% vs. 63%, p=0.01 for PFS; 40% vs. 75%, p=0.02 for OS). When the HIN-1 gene was over-expressed in ES2 cells, a significant reduction in cell growth and induction of apoptosis, and increasing paclitaxel sensitivity by reducing phosphorylation of Akt were observed. Conclusions: Methylation of HIN-1 and CACNA1A promoters are two novel epigenetic biomarkers associated with poor outcomes in OCCA patients. Ectopic expression of the HIN-1 gene increased paclitaxel sensitivity through Akt pathway.