Cancer Biology and Signal Transduction Inhibition of SIRT1 Signaling Sensitizes the Antitumor Activity of Silybin against Human Lung Adenocarcinoma Cells In Vitro and In Vivo

Although silybin, a natural flavonolignan, has been shown to exhibit potent antitumor activities against various types of cancers, including lung cancer, the molecular mechanisms behind these activities remain unclear. Silent information regulator 1 (SIRT1) is a conserved NADþ-dependent deacetylase that has been implicated in the modulation of transcriptional silencing and cell survival. Furthermore, it plays a key role in carcinogenesis through the deacetylation of important regulatory proteins, including p53. In this study, we investigated the antitumor activity of silybin towards human lung adenocarcinoma cells in vitro and in vivo and explored the role of the SIRT1 signaling pathway in this process. Silybin treatment resulted in a doseand timedependent decrease in lung adenocarcinomaA549 cell viability. In addition, silybin exhibited strong antitumor activity illustrated by reductions in tumor cell adhesion, migratory capability, and glutathione levels and by increased apoptotic indices and reactive oxygen species levels. Silybin treatment also downregulated SIRT1 and upregulated p53 acetylation. SIRT1 siRNA (in vitro) or cambinol (a known SIRT1 inhibitor used for in vivo studies) further enhanced the antitumor activity of silybin. In summary, silybin is a potent inhibitor of lung adenocarcinoma cell growth that interferes with SIRT1 signaling, and this inhibition is a novel mechanism of silybin action thatmaybeused for therapeutic intervention in lungadenocarcinoma treatment.MolCancerTher; 13(7); 1–13. 2014 AACR. Introduction Lung cancer is the leading cause of cancer burden worldwide with more than 3 million cases and 1 million deaths occurring annually (1). Non–small cell lung cancer (NSCLC) subtypes (adenocarcinoma, squamous cell carcinoma, and large cell carcinoma) account for 80% to 85% of all lung cancers. The majority of patients with NSCLC are diagnosed during advanced stages and exhibit inoperable local or distant metastases (2). Despite extensive research, the overall 5-year survival rate of patients with lung cancer is only 8% to 14%, and this rate has improved only marginally over the past 30 years (1, 3). These alarming statistics suggest that additional strategies are needed to control this disease. An essential requirement for any successful longterm cancer chemoprevention strategy is that the chemopreventive agent exhibits little or no toxicity (4). An example of such an agent is silybin (the chemical structure of which is shown in Fig. 1A), a flavanone present in milk thistle (Silybum marianum L.) that is used as a hepatoprotective agent and has been marketed as a dietary supplement (5). Silybin exhibits strong chemopreventive and anticancer properties towards various models of colon (6), liver (7), kidney (8), brain (9), prostate, and breast cancer (10), among others. Moreover, no median lethal dose (lethal dose, 50%; LD50) for silybin has been reported in laboratory animals, and it has also been considered exceptionally safe due to its extremely low toxicity in both animals and humans during acute or chronic administration; these findings emphasize the importance of utilizing this effective biologic agent in cancer chemoprevention studies (1, 11). With respect to lung cancer, silybin has been shown to cause significant growth inhibition and apoptosis in both small cell lung cancers and NSCLCs (1, 3, 11). However, the mechanisms responsible for the antitumor effects of silybin in lung adenocarcinoma have not yet been fully elucidated. Authors' Affiliations: Department of Cardiovascular Surgery, Xijing Hospital; Department of Thoracic Surgery, Tangdu Hospital; Department of Prosthodontics, School of Stomatology, The Fourth Military Medical University, Xi'an; and Department of Clinical Laboratory, The First Affiliated Hospital of General Hospital of PLA, Beijing, China Note: Supplementary data for this article are available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/). Z. Liang, Y. Yang, and Haibin Wang contributed equally to this work. Corresponding Authors:Wensheng Chen, Department of Cardiovascular Surgery, Xijing Hospital, The Fourth Military Medical University, 127 ChangleWestRoad,Xi'an 710032,China. Phone: 86-29-8477-5314; Fax: 86-298321-0092; E-mail: [email protected]; and Weixun Duan, [email protected] doi: 10.1158/1535-7163.MCT-13-0942 2014 American Association for Cancer Research. Molecular Cancer Therapeutics www.aacrjournals.org OF1 on June 20, 2017. © 2014 American Association for Cancer Research. mct.aacrjournals.org Downloaded from Published OnlineFirst May 5, 2014; DOI: 10.1158/1535-7163.MCT-13-0942