Contents lists available at ScienceDirect Phytomedicine journal homepage: www.elsevier.com/locate/phymed Original Article Crucifera sulforaphane (SFN) inhibits the growth of nasopharyngeal carcinoma through DNA methyltransferase 1 (DNMT1)/Wnt inhibitory factor 1 (WIF1) axis Luo Chen a , Lai Sheung Chan a , Hong Lok Lung a , Timothy Tak Chun Yip b,c , Roger Kai Cheong Ngan b,c , Jonathan Woon Chung Wong a , Kwok Wai Lo d , Wai Tong Ng e , Anne Wing Mui Lee f , George Sai Wah Tsao g , Maria Li Lung f , Nai Ki Mak a, ⁎ a Department of Biology, Hong Kong Baptist University, Kowloon, Hong Kong, China b Department of Clinical Oncology, Queen Elizabeth Hospital Hong Kong, Kowloon, Hong Kong, China c Center for Nasopharyngeal Carcinoma Research, University of Hong Kong, Pokfulam, Hong Kong, China d Department of Anatomical and Cellular Pathology and State Key Laboratory of Translational Oncology, The Chinese University of Hong Kong, Shatin, NT, Hong Kong, China e Clinical Oncology, Pamela Youde Nethersole Eastern Hospital, Chai Wan, Hong Kong, China f Department of Clinical Oncology, Center for Nasopharyngeal Carcinoma Research, University of Hong Kong, Pokfulam, Hong Kong, China g Department of Anatomy, Center for Nasopharyngeal Carcinoma Research, University of Hong Kong, Pokfulam, Hong Kong, China ARTICLE INFO Keywords: Sulforaphane Nasopharyngeal carcinoma DNMT1 WIF1 ABSTRACT Background: Sulforaphane (SFN), a natural compound present in cruciferous vegetable, has been shown to possess anti-cancer activities. Cancer stem cell (CSC) in bulk tumor is generally considered as treatment resistant cell and involved in cancer recurrence. The effects of SFN on nasopharyngeal carcinoma (NPC) CSCs have not yet been explored. Purpose: The present study aims to examine the anti-tumor activities of SFN on NPC cells with CSC-like prop- erties and the underlying mechanisms. Methods: NPC cells growing in monolayer culture, CSCs-enriched NPC tumor spheres, and also the NPC nude mice xenograft were used to study the anti-tumor activities of SFN on NPC. The population of cells expressing CSC-associated markers was evaluated using flow cytometry and aldehyde dehydrogenase (ALDH) activity assay. The effect of DNA methyltransferase 1 (DNMT1) on the growth of NPC cells was analyzed by using small in- terfering RNA (siRNA)-mediated silencing method. Results: SFN was found to inhibit the formation of CSC-enriched NPC tumor spheres and reduce the population of cells with CSC-associated properties (SRY (Sex determining Region Y)-box 2 (SOX2) and ALDH). In the functional study, SFN was found to restore the expression of Wnt inhibitory factor 1 (WIF1) and the effect was accompanied with the downregulation of DNMT1. The functional activities of WIF1 and DNMT1 were confirmed using exogenously added recombinant WIF1 and siRNA knockdown of DNMT1. Moreover, SFN was found to inhibit the in vivo growth of C666-1 cells and enhance the anti-tumor effects of cisplatin. Conclusion: Taken together, we demonstrated that SFN could suppress the growth of NPC cells via the DNMT1/ WIF1 axis. Introduction Nasopharyngeal carcinoma (NPC) is closely associated with latent infection of Epstein-Barr virus (EBV) and nearly all NPC tumors contain EBV (Hildesheim et al., 2002). C666-1 is not only the cell line con- sistently harboring EBV, but also known to be resistant to cisplatin treatment (Cheung et al., 1999; Lun et al., 2012). These features make it a suitable choice for studying NPC. Patients with early stage NPC can be https://doi.org/10.1016/j.phymed.2019.153058 Received 10 December 2018; Received in revised form 26 July 2019; Accepted 29 July 2019 Abbreviations: ALDH, Aldehyde Dehydrogenase; CSC, Cancer Stem Cell; DNMT1, DNA Methyltransferase 1; EBV, Epstein-Barr Virus; EGF, Epidermal Growth Factor; FGF, Fibroblast Growth Factor; IGF, Insulin-like Growth Factor; NPC, Nasopharyngeal Carcinoma; SFN, Sulforaphane; SOX2, SRY (sex determining region Y)-box 2; WIF1, Wnt Inhibitory Factor 1 ⁎ Corresponding author. E-mail address: nkmak@hkbu.edu.hk (N.K. Mak). Phytomedicine 63 (2019) 153058 0944-7113/ © 2019 Published by Elsevier GmbH. T