Transcriptional silencing of secreted frizzled related protein 1 (SFRP1) by promoter hypermethylation in non-small-cell lung cancer

Abstract

Secreted frizzled related protein 1 (SFRP 1) is an antagonist of the transmembrane frizzled receptor, a component of the Wnt signaling pathway, and has been suggested to be a candidate tumor suppressor in several human malignancies. Since SFRP 1 is located at chromosome 8 p 11, where lung cancers also exhibit frequent allelic loss, we hypothesized that the inactivation of SFRP 1 is also involved in lung carcinogenesis. To substantiate this, we performed mutational analysis of SFRP 1 for 29 non-small-cell lung cancer (NSCLC) and 25 small-cell lung cancer (SCLC) cell lines, and expression analysis for the same cell lines. Although somatic mutations were not detected in the coding sequence, downregulation of SFRP 1 was observed in 14 (48%) NSCLC and nine (36%) SCLC cell lines. We analysed epigenetic alteration of the SFRP 1 promoter region and detected hypermethylation in 15 (52%) of 29 NSCLC cell lines, two (8%) of 25 SCLC cell lines, and 44 (55%) of 80 primary lung tumors. By comparing the methylation status with SFRP 1 expression, we found a significant correlation between them. We also performed loss of heterozygosity (LOH) analysis and found that 15 (38%) of 40 informative surgical specimens had LOH in the SFRP 1 gene locus. Furthermore, we performed colony formation assay of two NSCLC cell lines (NCI-H 460 and NCI-H 2009) and found the reduction of colony formation with SFRP 1 transfection. In addition, we also detected that SFRP 1 inhibits the transcriptional activity of beta-catenin, which is thought to be a downstream molecule of SFRP 1, with luciferase reporter assay. Our current studies demonstrated that the SFRP 1 gene is frequently downregulated by promoter hypermethylation and suppresses tumor growth activity of lung cancer cells, which suggests that SFRP 1 is a candidate tumor suppressor gene for lung cancer.