Epigenetic silencing of NKD2, a major component of Wnt signaling, promotes breast cancer growth

// Yan Dong 1, 2 , Baoping Cao 1, 3 , Meiying Zhang 1, 3 , Weidong Han 2 , James G. Herman 4 , Francois Fuks 5 , Yali Zhao 6 , Mingzhou Guo 1 1 Department of Gastroenterology & Hepatology, Chinese PLA General Hospital, Beijing 100853, China 2 Department of Molecular Biology, Institute of Basic Medicine, School of Life Sciences, Chinese PLA General Hospital, Beijing 100853, China 3 Medical College of NanKai University, Tianjin 300071, China 4 The Hillman Cancer Center, University of Pittsburgh Cancer Institute, Pittsburgh, PA 15213, U.S.A 5 Laboratory of Cancer Epigenetics, Free University of Brussels (U.L.B.), 808 route de Lennik, Brussels 1070, Belgium 6 Central Laboratory, The Affiliated Hainan Hospital of Chinese PLA General Hospital, Hai Tang Wan, Sanya 572013, China Correspondence to: Mingzhou Guo, e-mail: mzguo1@gmail.com Yali Zhao, e-mail: zhaoyl301@163.com Keywords: NKD2, DNA methylation, Wnt signaling, epigenetics, breast cancer Received: February 27, 2015 Accepted: June 08, 2015 Published: June 19, 2015 ABSTRACT Naked cuticle homolog 2 (NKD2) has been reported to antagonize Wnt signaling in zebrafish, mouse and mammals. The aim of this study is to investigate the epigenetic changes and mechanisms of NKD2 in human breast cancer development. Six breast cancer cell lines (MCF-7, ZR75–1, MDA-MB-468, MDA-MB-231, T47D and BT474) and 68 cases of primary human breast cancer were studied using methylation specific PCR, immunohistochemistry, western blot, flow cytometry techniques and a xenograft mouse model. The expression of NKD1 and NKD2 was regulated by promoter region methylation in breast cancer cells. No NKD1 methylation was found in primary human breast cancer. NKD2 was methylated in 51.4% (35/68) of human primary breast cancer samples. NKD2 methylation was significantly associated with reduction of NKD2 expression, and tumor stage ( p < 0.05). NKD2 suppressed breast cancer cell proliferation both in vitro and in vivo . NKD2 induced G1/S arrest and inhibited Wnt signaling in breast cancer cells. In conclusion, NKD2 is frequently methylated in human breast cancer, and the expression of NKD2 is regulated by promoter region methylation. NKD2 suppresses breast cancer proliferation by inhibiting Wnt signaling.