Mutation Research 707 (2011) 42–52 Contents lists available at ScienceDirect Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis journal homepage: www.elsevier.com/locate/molmut Community address: www.elsevier.com/locate/mutres Antroquinonol inhibits NSCLC proliferation by altering PI3K/mTOR proteins and miRNA expression profiles V. Bharath Kumar a , Ta-Chun Yuan a , Je-Wen Liou b , Chih-Jen Yang c , Ping-Jyun Sung d , Ching-Feng Weng a,∗ a Department of Life Science and Institute of Biotechnology, National Dong Hwa University, Hualien, Taiwan b Department of Biochemistry, School of Medicine, Tzu-Chi University, Hualien, Taiwan c Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan d Graduate Institute of Marine Biotechnology, Department of Life Science, National Dong Hwa University, Pingtung, Taiwan article info Article history: Received 25 September 2010 Received in revised form 11 December 2010 Accepted 19 December 2010 Available online 24 December 2010 Keywords: Antroquinonol PI3K/mTOR miRNA cdc2 proteins PARP Caspase cleavage abstract Antroquinonol a derivative of Antrodia camphorata has been reported to have antitumor effects against various cancer cells. However, the effect of antroquinonol on cell signalling and survival pathways in non- small cell lung cancer (NSCLC) cells has not been fully demarcated. Here we report that antroquinonol treatment significantly reduced the proliferation of three NSCLC cells. Treatment of A549 cells with antro- quinonol increased cell shrinkage, apoptotic vacuoles, pore formation, TUNEL positive cells and increased Sub-G1 cell population with respect to time and dose dependent manner. Antroquinonol treatment not only increased the Sub-G1 accumulation but also reduced the protein levels of cdc2 without altering the expression of cyclin B1, cdc25C, pcdc2, and pcdc25C. Antroquinonol induced apoptosis was associated with disrupted mitochondrial membrane potential and activation of Caspase 3 and PARP cleavage in A549 cells. Moreover, antroquinonol treatment down regulated the expression of Bcl2 proteins, which was correlated with the decreased PI3K and mTOR protein levels without altering pro apoptotic and anti apoptotic proteins. Results from the microarray analysis demonstrated that antroquinonol altered the expression level of miRNAs compared with untreated control in A549 cells. The data collectively suggested the antiproliferative effect of antroquinonol on NSCLC A549 cells, which provides useful infor- mation for understanding the anticancer mechanism influenced by antroquinonol and is the first report to suggest that antroquinonol may be a promising chemotherapeutic agent for lung cancer. © 2010 Elsevier B.V. All rights reserved. 1. Introduction Lung cancer remains the leading cause of cancer-related death in the worldwide. Among them, 80% of lung cancers are non- small cell lung cancer (NSCLC). Adenocarcinoma is a subgroup of NSCLC and some other tumour histotype shows resistance to radi- ation, chemotherapy and platinum-based doublet chemotherapy while other tumour subtypes display very low response rate for the chemotherapy [1–5]. One of every three cancer-related deaths accounts for lung cancer with an overall survival of 5-years, having less than 15% survival probability [6,7]. Similarly in two thirds of cases the cancer might have start spreading during the time of diag- nosis, due to limited therapeutic options [8,9]. Till now numerous new cytotoxic agents have been introduced in NSCLC treatment, only small improvements have been observed in the survival of patients with advanced or metastatic lung cancer [10]. Thus it ∗ Corresponding author. Tel.: +886 3 8633637; fax: +886 3 8630255. E-mail address: cfweng@mail.ndhu.edu.tw (C.-F. Weng). is important to consider a new bioactive compound from natu- ral products, such as phytochemicals that can selectively inhibit tumour cells growth without affecting normal cells [11,12]. Phytochemicals represent a new class of compounds with anti-carcinogenic properties that are gaining attention in the treat- ment of human cancers, thus offering promised new options for the development of effective chemopreventive and chemothera- peutic strategies, particularly those that can be administered as dietary supplements [13]. Antrodia camphorata (A. camphorata) is a new basidiomycete in the Polyporaceae (Aphyllophorales), caus- ing brown heart rot in Cinnamomum kanehirai hay (Lauraceae) in Taiwan has been identified as distinct species of the genus Antrodia [14,15]. A. camphorate is used for the treatment of diar- rhoea, abdominal pain, hypertension, itching of the skin, and liver cancer [16]. Previous studies have shown that growth inhibitory effects of A. camphorate in prostate cancer, bladder carcinoma, and breast cancer cells [17–19]. Furthermore, ethanol extracts of A. camphorata mycelia (SACE and Fraction-6) can induce apopto- sis of NSCLC cells by down regulating the synthesis of galectin-1, RhoGDI-, human calpain small (regulatory) subunit and eIF5A 0027-5107/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.mrfmmm.2010.12.009