Anti-Tumor Activity of a Novel Compound-CDF Is Mediated by Regulating miR-21, miR-200, and PTEN in Pancreatic Cancer Bin Bao1., Shadan Ali2., Dejuan Kong1, Sanila H. Sarkar1, Zhiwei Wang1, Sanjeev Banerjee1, Amro Aboukameel2, Subhash Padhye3, Philip A. Philip2, Fazlul H. Sarkar1* 1 Department of Pathology, Wayne State University, Detroit, Michigan, United States of America, 2 Division of Hematology/Oncology Karmanos Cancer Institute, Wayne State University, Detroit, Michigan, United States of America, 3 Dr. D. Y. Patil University, Pimpri, Pune, India Abstract Background: The existence of cancer stem cells (CSCs) or cancer stem-like cells in a tumor mass is believed to be responsible for tumor recurrence because of their intrinsic and extrinsic drug-resistance characteristics. Therefore, targeted killing of CSCs would be a newer strategy for the prevention of tumor recurrence and/or treatment by overcoming drug- resistance. We have developed a novel synthetic compound-CDF, which showed greater bioavailability in animal tissues such as pancreas, and also induced cell growth inhibition and apoptosis, which was mediated by inactivation of NF-kB, COX-2, and VEGF in pancreatic cancer (PC) cells. Methodology/Principal Findings: In the current study we showed, for the first time, that CDF could significantly inhibit the sphere-forming ability (pancreatospheres) of PC cells consistent with increased disintegration of pancreatospheres, which was associated with attenuation of CSC markers (CD44 and EpCAM), especially in gemcitabine-resistant (MIAPaCa-2) PC cells containing high proportion of CSCs consistent with increased miR-21 and decreased miR-200. In a xenograft mouse model of human PC, CDF treatment significantly inhibited tumor growth, which was associated with decreased NF-kB DNA binding activity, COX-2, and miR-21 expression, and increased PTEN and miR-200 expression in tumor remnants. Conclusions/Significance: These results strongly suggest that the anti-tumor activity of CDF is associated with inhibition of CSC function via down-regulation of CSC-associated signaling pathways. Therefore, CDF could be useful for the prevention of tumor recurrence and/or treatment of PC with better treatment outcome in the future. Citation: Bao B, Ali S, Kong D, Sarkar SH, Wang Z, et al. (2011) Anti-Tumor Activity of a Novel Compound-CDF Is Mediated by Regulating miR-21, miR-200, and PTEN in Pancreatic Cancer. PLoS ONE 6(3): e17850. doi:10.1371/journal.pone.0017850 Editor: Lin Zhang, University of Pennsylvania, United States of America Received December 13, 2010; Accepted February 10, 2011; Published March 9, 2011 Copyright: 2011 Bao et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Funding: National Cancer Institute, NIH grants 5R01CA131151, 3R01CA131151-02S1, and 5R01CA132794 (F.H. Sarkar). We thank Puschelberg and Guido foundations for their generous financial contribution. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. * E-mail: fsarkar@med.wayne.edu . These authors contributed equally to this work. Introduction Pancreatic cancer (PC) is one of the most lethal malignant diseases with worst prognosis, which is ranked as the fourth leading cause of cancer-related deaths in the United States [1]. Over the past two decades, numerous efforts have been made in improving treatment and survival PC patients but the outcome has been disappointing. This disappointing outcome is due to many factors among which de novo resistance (intrinsic) and acquired (extrinsic) resistance to conventional therapeutics (chemotherapy and radiation therapy) including gemcitabine alone or in-combination with other cytotoxic or targeted agents. Emerging evidence suggest that the resistance could in fact be due to the enriched existence of tumor initiating cells, also classified as cancer stem-like cells (CSC) in a tumor mass [2–6]. The CSCs have the capacity of self-renewal and the potential to regenerate into all types of differentiated cells giving rise to heterogeneous tumor cell populations in a tumor mass, which contributes to tumor aggressiveness [2–6]. Thus, the failure to eliminate these special cells is considered to be one of the underlying causes of poor treatment outcome with conventional therapeutics, suggesting that newer and novel therapeutic strategies must be developed for the targeted killing of drug resistant CSCs in order to eradicate the risk of tumor recurrence for improving the survival of patients diagnosed with PC. In search of novel yet non-toxic agents, attention has been focused on natural agents for several years. One such agent is curcumin (diferuloylmethane), which is derived from the plant Curcuma longa (Linn) grown in tropical Southeast Asia [7–9]. Curcumin has been shown to inhibit the growth of a variety of tumor cells; however, the poor bioavailability of curcumin limits its application in the clinic. Recently, we have developed a novel synthetic analogue of curcumin, 3,4-difluoro-benzo-curcumin [we named it as Difluorinated-Curcumin or in short CDF [10,11]], which showed greater bioavailability in pancreatic tissues, and also PLoS ONE | www.plosone.org 1 March 2011 | Volume 6 | Issue 3 | e17850