Frequent concomitant epigenetic silencing of SOX1 and secreted frizzled-related proteins (SFRPs) in human hepatocellular carcinoma.

Background And Aim: Except for genetic mutations, epigenetic changes are also involved in the development of human cancers. Recently, we have identified SOX1, SRY (sex determining region Y)-box 1, is hypermethylated in cervical cancer and ovarian cancer. Therefore, we investigated whether promoter hypermethylation of SOX1 is common in hepatocellular carcinoma (HCC).

Methods: We used methylation-specific polymerase chain reaction (MS-PCR) and bisulfite sequencing to analyze the methyaltion level of the SOX1 promoter in seven HCC cell lines, 54 clinical HCCs, 42 cirrhotic livers, 21 livers with chronic hepatitis, and 15 control livers. Then, we employed quantitative MS-PCR (QMSP) to validate in an independent set of samples (60 paired HCCs and 30 control livers). Finally, we used luciferase reporter and colony formation assay to check the effect of SOX1 in HCC.

Results: Promoter methylation of SOX1 was significantly frequent in HCC cell lines and clinical HCCs, cirrhotic livers, but not in control livers (P < 0.0001). There is a significant correlation between downregulation of SOX1 expression and promoter methylation. QMSP results confirmed that promoter hypermethylation of SOX1 is significantly more frequent in HCCs than control livers (P < 0.0001). The frequency of SOX1 methylation in patients with secreted frizzled-related proteins (SFRPs) methylation is significantly higher than in patients without SFRPs methylation (P < 0.0001). Furthermore, ectopic expression of SOX1 could suppress T-cell factor-dependent transcriptional activity and colony formation number in HCCs.

Conclusions: Concomitant epigenetic silencing of SOX1 and SFRPs through promoter hypermethylation is frequent in HCCs, and this might contribute to abnormal activation of canonical Wnt signal pathway.