Toxicology 264 (2009) 192–197 Contents lists available at ScienceDirect Toxicology journal homepage: www.elsevier.com/locate/toxicol Antiproliferative effect of flavomannin-6,6 ′ -dimethylether from Tricholoma equestre on Caco-2 cells Gisela Pachón-Pe ˜ na a , Fernando J. Reyes-Zurita b , Gerard Deffieux c , Amaia Azqueta d , Adela López de Cerain d , Josep J. Centelles a , Edmond E. Creppy e,∗ , Marta Cascante a,∗∗ a Department of Biochemistry and Molecular Biology, Institute of Biomedicine of the University of Barcelona (IBUB) and Unit Associated with CSIC, Diagonal 645, 08028- Barcelona, Spain b Department of Biochemistry and Molecular Biology I, Faculty of Sciences, University of Granada, Campus Fuentenueva s/n, 18071 Granada, Spain c Laboratoire de Mycologie et Biotechnologie végétale, Université de Bordeaux 2, Victor Ségalen, Faculté des Sciences Pharmaceutiques, 146, rue Léo-Saignat, 33076 Bordeaux, France d Departamento de Ciencias de la Alimentación, Fisiología y Toxicología, Facultad de Farmacia, Universidad de Navarra, Irunlarrea s/n, 31008 Pamplona, Spain e Laboratoire de Toxicologie et d’Hygiène Appliquée, Université de Bordeaux 2, Victor Ségalen, Faculté des Sciences Pharmaceutiques, 146, rue Léo-Saignat, 33076 Bordeaux, France article info Article history: Received 8 June 2009 Received in revised form 5 August 2009 Accepted 11 August 2009 Available online 15 August 2009 Keywords: Tricholoma equestre (L.)P.Kumm Flavomannin-6,6 ′ -dimethylether Caco-2 Cell growth inhibition Cell cycle p27 Apoptosis DNA fragmentation abstract Several studies have been performed reporting antitumoral activity of different mushroom extracts. The current study reports the antiproliferative activity of flavomannin-6,6 ′ -dimethylether obtained from a very common edible mushroom: Tricholoma equestre (L.)P.Kumm, and the characterization of its effects at molecular level. Concentrations causing 50% and 80% growth inhibition on human adenocarcinoma col- orectal Caco-2 cells were determined (in g/mL: IC 50 = 96 ± 3 after 24 h and 78 ± 7 after 48 h, IC 80 = 112 ± 4 after 24 h and 90 ± 3 after 48 h) by using MTT method. It was demonstrated that flavomannin-6,6 ′ - dimethylether induced an arrest in G0/G1 phase of the cell cycle by flow cytometry analysis and an increase of p27 protein level by Western blot. Furthermore, this compound did not induce apoptosis by flow cytometry or DNA fragmentation by gel electrophoresis. Thus, it could be a promising agent due to its cytostatic effect against Caco-2 tumoral cells, and the absence of a genotoxic effect. © 2009 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Cancer occurs as a result of stepwise accumulation of genetic changes, which follow the failure of the homeostatic mechanisms that govern normal cell proliferation (Hann et al., 2002). An alter- ation of multi-genetic and epigenetic molecular events is required for cancer development, thus indicating a strong strategy to pre- vent cancer using some compounds, which are able to arrest or reverse carcinogenic changes before the appearance of the malig- nant disease (Singh and Agarwal, 2006). A great quantity of new techniques facilitates the fast and simultaneous screening of var- ious novel compounds for their antitumor activities at molecular and biochemical levels. Although sometimes the random screen- ∗ Corresponding author. ∗∗ Corresponding author. Tel.: +34 93 4021593; fax: +34 93 4021559. E-mail addresses: giselapachon@yahoo.com (G. Pachón-Pe ˜ na), ferjes@ugr.es (F.J. Reyes-Zurita), Gerard.Deffieux@phyto.u-bordeaux2.fr (G. Deffieux), o.a.azqueta@medisin.uio.no (A. Azqueta), acerain@unav.es (A.L. de Cerain), josepcentelles@ub.edu (J.J. Centelles), edmond.creppy@tox.u-bordeaux2.fr (E.E. Creppy), martacascante@ub.edu (M. Cascante). ing of various substances seems pointless and time-consuming, many anticancer agents from plants and marine organisms have been discovered recently (Singh and Agarwal, 2006; Singh et al., 2008; Molinski et al., 2009). Cell cycle progression is an important regulated biological event in normal cells, which becomes almost universally aberrant or deregulated in transformed and neoplastic cells. In this regard, targeting deregulated cell cycle progression and its modulation by various natural and synthetic agents are gaining widespread attention to control the unchecked growth and prolif- eration in cancer cells (Singh and Agarwal, 2006). The expanded knowledge of molecular basis of tumorigenesis and metastasis, together with the inherently vast structural diversity of natural compounds found in mushrooms, provided unique opportunities for discovering new drugs that rationally target the abnormal molecular and biochemical signals leading to cancer (Konno, 2007). There is a significant interest in the use of mushroom and/or mushroom extracts as a dietary supplement based on theories that they enhance immune function and promote health. There have been a number of attempts to determine antitumor activities of mushrooms. Such studies are important because many com- pounds from mushrooms do potentially have significant biological activity (Konno, 2007; Zaidman et al., 2005; Sullivan et al., 2006). 0300-483X/$ – see front matter © 2009 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.tox.2009.08.009