Jacaranone Induces Apoptosis in Melanoma Cells via ROS-Mediated Downregulation of Akt and p38 MAPK Activation and Displays Antitumor Activity In Vivo Mariana H. Massaoka1, Alisson L. Matsuo 1 , Carlos R. Figueiredo 1 , Camyla F. Farias1, Nata´ lia Girola1, Denise C. Arruda1, Jorge A. B. Scutti1, Paulete Romoff2, Oriana A. Favero2, Marcelo J. P. Ferreira2, Joa˜ o H. G. Lago3, Luiz R. Travassos1* 1 Unidade de Oncologia Experimental, Universidade Federal de Sa˜o Paulo, Sa˜o Paulo, Sa˜o Paulo, Brazil, 2 Centro de Cieˆncias e Humanidades e Centro de Cieˆncias Biol o´gicas e da Sa u´de, Universidade Presbiteriana Mackenzie, Sa˜o Paulo, Sa˜o Paulo, Brazil, 3 Instituto de Cieˆncias Ambientais, Qu´ımicas e Farmaceˆuticas, Universidade Federal de Sa˜o Paulo, Diadema, Sa˜o Paulo, Brazil Abstract Background: Malignant melanoma is a deadly type of metastatic skin cancer with increased incidence over the past 30 years. Despite the advanced knowledge on the biology, immunobiology and molecular genetics of melanoma, the alternatives of treatment are limited with poor prognosis. On clinical trials, natural products and among them redox-active quinones have been tested in the attempt to control the growth of cancer cells. Recently, we isolated jacaranone from Pentacalia desiderabilis, a benzoquinone derivative that showed a broad antitumor activity and protective anti-melanoma effect in a syngeneic model. The purified substance is active at micromolar concentrations, is not hemolytic, and is not toxic in na¨ıve mice. Methodology/Principal Findings: The jacaranone antitumor activity was shown against several human cancer cell lines in vitro. Moreover, the induction of apoptosis in murine melanoma cells and jacaranone antitumor activity in vivo, in a melanoma experimental model, were also shown. Jacaranone renders antiproliferative and proapoptotic responses in tumor cells, by acting on Akt and p38 MAPK signaling pathways through generation of reactive oxygen species (ROS). The free radical scavenger N-acetyl-cysteine (NAC) was able to completely suppress cell death induced by jacaranone as it blocked Akt downregulation, p38 MAPK activation as well as upregulation of proapoptotic Bax. Notably, treatment of melanoma growing subcutaneously in mice with jacaranone significantly extended the mean survival times in a dose- dependent manner. Conclusions/Significance: The results provide evidence for the mechanisms of action of jacaranone and emphasize the potential use of this quinone for the treatment of melanoma. Citation: Massaoka MH, Matsuo AL, Figueiredo CR, Farias CF, Girola N, et al. (2012) Jacaranone Induces Apoptosis in Melanoma Cells via ROS-Mediated Downregulation of Akt and p38 MAPK Activation and Displays Antitumor Activity In Vivo. PLoS ONE 7(6): e38698. doi:10.1371/journal.pone.0038698 Editor: Salvatore V. Pizzo, Duke University Medical Center, United States of America Received January 23, 2012; Accepted May 9, 2012; Published June 6, 2012 Copyright: 2012 Massaoka 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: The State of Sa˜o Paulo Research Support Foundation (FAPESP), Brazil, supported this work through grant 2010/51423-0 as well as the Brazilian National Research Council (CNPq). 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: luiztravassos@gmail.com Introduction Among the conventional antitumor cytotoxic chemotherapies, many compounds are derived from natural products [1,2], and commonly used agents to treat human cancers belong to the quinone class of organic compounds [3]. Antitumor quinones (e.g. dauno- rubicin and doxorubicin) can undergo reversible enzymatic re- duction and oxidation, and form oxygen reactive species (ROS). The role of ROS as secondary messengers in intracellular signaling cascades has been investigated extensively in the last two decades [4]. There is evidence that low levels of ROS can induce cell proliferation and genetic instability, which may contribute to the oncogenic phenotype of cancer cells. At high concentration ROS can promote cellular senescence and apoptosis, and therefore may function as antitumorigenic species [5]. Indeed, cancer cells frequently exhibit abnormal redox status associated with increased basal production of ROS, and thus cannot tolerate higher levels of free radicals [6]. Therefore, the use of compounds that interfere in redox regulation is a promising strategy to selectively target tumors [4]. Recently, we isolated jacaranone from leaves of Pentacalia desiderabilis [7]. Although potential antiproliferative protective effects have been described for extracts and purified jacaranone from Senecio leucanthemifolius Poiret [8], the mechanism of action and the antitumor activity of this quinone has not been previously investigated. In the present work, we show that jacaranone induces biochemical and morphological alterations of apoptosis mediated by generation of reactive oxygen species (ROS). Investigation of the signaling pathways affected by jacaranone showed down- regulation of Akt, activation of p38 MAPK and upregulation of PLoS ONE | www.plosone.org 1 June 2012 | Volume 7 | Issue 6 | e38698