Diagnostic role of Wnt pathway gene promoter methylation in non small cell lung cancer

// Shunlin Liu 1, * , Xiaoying Chen 2, * , Ruhua Chen 3, * , Jinzhi Wang 4, * , Guoliang Zhu 5 , Jianzhong Jiang 6 , Hongwei Wang 7 , Shiwei Duan 2 , Jianan Huang 1 1 Department of Respiratory Medicine, The First Affiliated Hospital of Soochow University, Suzhou, Jiangsu 215006, China 2 Medical Genetics Center, School of Medicine, Ningbo University, Ningbo, Zhejiang 315211, China 3 Department of Respiratory Medicine, Affiliated Yixing Hospital of Jiangsu University, Yixing, Jiangsu 214200, China 4 Department of Cell Biology, School of Medicine, Soochow University, Suzhou, Jiangsu 215007, China 5 Department of Pathology, Huzhou First People's Hospital, Huzhou, Zhejiang 313000, China 6 Department of Geriatrics, Affiliated Yixing Hospital of Jiangsu University, Yixing, Jiangsu 214200, China 7 Realgen Biotechnology Co., Ltd. Zhangjiang High Technology Park, Shanghai 201203, China * Co-first authors Correspondence to: Jianan Huang, email: huang_jian_an@163.com Shiwei Duan, email: duanshiwei@nbu.edu.cn Keywords: non-small cell lung cancer, quantitative methylation-specific PCR, DNA methylation, diagnosis, Wnt pathway Received: November 22, 2016      Accepted: March 21, 2017      Published: March 31, 2017 ABSTRACT Wnt signal pathway genes are known to be involved with cancer development. Here we tested the hypothesis whether DNA methylation of genes part of the Wnt signaling pathway could help the diagnosis of non-small cell lung cancer (NSCLC). The methylation levels of SFRP1, SFRP2, WIF1 and PRKCB in 111 NSCLC patients were evaluated by quantitative methylation-specific PCR (qMSP). Promoter methylation levels of four candidate genes were significantly higher in tumor tissues compared with the adjacent tissues. SFRP1, SFRP2 and PRKCB genes were all shown to be good predictors of NSCLC risk ( SFRP1 : AUC = 0.711; SFRP2 : AUC = 0.631; PRKCB : AUC = 0.650). The combined analysis showed that the methylation status of the four genes had a sensitivity of 70.3% and a specificity of 73.9% in the prediction of NSCLC risk for study cohort. A higher diagnostic value with an AUC of 0.945 (95% CI: 0.923–0.967, sensitivity: 90.6%, specificity: 93.0%) was found in TCGA cohort. In addition, SFRP1 and SFRP2 hypermethylation events were specific to male patients. Further TCGA data mining analysis suggested that SFRP1 _cg15839448, SFRP2 _cg05774801, and WIF1 _cg21383810 were inversely associated with the host gene expression. Moreover, GEO database analysis showed that 5'-Aza-deoxycytidine was able to upregulate gene expression in several lung cancer cell lines. Subsequent dual-luciferase reporter assay showed a crucial regulatory function of PRKCB promoter. In summary, our study showed that a panel of Wnt signal pathway genes ( SFRP1, SFRP2, WIF1 and PRKCB ) had the potential as methylation biomarkers in the diagnosis of NSCLC.