Abstract 1154: Detection of aberrant methylation of the RASSF1A gene as a biomarker for hepatocellular carcinoma.

Abstract

Abstract Hepatocellular carcinoma (HCC) is an aggressive malignancy that is responsible for between 250,000 and 800,000 deaths annually worldwide. HCC has a poor prognosis with a 5-year survival rate of less than 15% mainly due to a lack of early detection. The current biomarker for HCC is the alpha-fetoprotein (AFP), which has a low sensitivity of 40-60%. There is a necessity for non-invasive, convenient, and sensitive methods to detect the cancer in its curative stages to improve prognosis. Previous studies from our laboratory showed that the location of the CpG site methylation studied could impact the marker performance in distinguishing HCC from other liver diseases. The aberrant methylation of the tumor-suppressor RASSF1A gene has been extensively studied as a marker for a variety of cancers, including liver cancer. However, some studies found that the gene was methylated even in non-cancerous liver tissues. Thus, in order to explore the possibility of detecting aberrant methylation of the tumor-suppressor RASSF1A gene as a biomarker for HCC screening, we analyzed the methylation profiles of 57 CpG sites in three regions of the promoter - P1, P2, and E1 - in normal, hepatitic, cirrhotic, adjacent non-HCC, and HCC tissue samples through bisulfite DNA sequencing. We discovered that the P1 region was most specifically methylated to HCC. We then confirmed the findings by developing a methylation specific PCR and analyzing a larger sample size. The data were analyzed and the P1 test was found to have an area under the curve (AUROC) of 0.90, whereas the E1 and P2 tests had AUROC of 0.84 and 0.72, respectively. At a cutoff of 10 copies for our analysis, the P1 MSP gave a sensitivity of 80.8% and specificity of 85.7%. Furthermore, we found that out of 62 AFP-negative samples, mRASSF1A detected 54 (87.1%). By combining AFP and mRASSF1A, 107 of 115 (93.0%) of HCC samples were detected as cancerous. Subsequently, we developed a PCR assay to detect the methylated RASSF1A gene in the urine of patients. This assay was then used to test the urine DNA isolated from patients with hepatitic, cirrhotic, or HCC after which the data were statistically evaluated. The potential of a urine DNA test for HCC screening is discussed. Citation Format: Surbhi Jain, Lijia Xie, Batbold Boldbaatar, Ying-Hsiu Su, Wei Song. Detection of aberrant methylation of the RASSF1A gene as a biomarker for hepatocellular carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1154. doi:10.1158/1538-7445.AM2013-1154