An Integrated Drosophila Model System Reveals Unique Properties for F14512, a Novel Polyamine-Containing Anticancer Drug That Targets Topoisomerase II Sonia Chelouah1,2¤a, Caroline Monod-Wissler1,2¤b, Christian Bailly3, Jean-Marc Barret3, Nicolas Guilbaud3, Ste´ phane Vispe´ 3*, Emmanuel Ka¨ s1,2* 1 Universite´ de Toulouse, UPS, Universite´ Paul Sabatier, Laboratoire de Biologie Mole´culaire Eucaryote; Toulouse; France, 2 CNRS, Centre National de la Recherche Scientifique, UMR5099, Laboratoire de Biologie Mole´culaire Eucaryote, Toulouse, France, 3 Centre de Recherche en Oncologie Expe´rimentale, Institut de Recherche Pierre Fabre, Toulouse, France Abstract F14512 is a novel anti-tumor molecule based on an epipodophyllotoxin core coupled to a cancer-cell vectoring spermine moiety. This polyamine linkage is assumed to ensure the preferential uptake of F14512 by cancer cells, strong interaction with DNA and potent inhibition of topoisomerase II (Topo II). The antitumor activity of F14512 in human tumor models is significantly higher than that of other epipodophyllotoxins in spite of a lower induction of DNA breakage. Hence, the demonstrated superiority of F14512 over other Topo II poisons might not result solely from its preferential uptake by cancer cells, but could also be due to unique effects on Topo II interactions with DNA. To further dissect the mechanism of action of F14512, we used Drosophila melanogaster mutants whose genetic background leads to an easily scored phenotype that is sensitive to changes in Topo II activity and/or localization. F14512 has antiproliferative properties in Drosophila cells and stabilizes ternary Topo II/DNA cleavable complexes at unique sites located in moderately repeated sequences, suggesting that the drug specifically targets a select and limited subset of genomic sequences. Feeding F14512 to developing mutant Drosophila larvae led to the recovery of flies expressing a striking phenotype, "Eye wide shut," where one eye is replaced by a first thoracic segment. Other recovered F14512-induced gain- and loss-of-function phenotypes similarly correspond to precise genetic dysfunctions. These complex in vivo results obtained in a whole developing organism can be reconciled with known genetic anomalies and constitute a remarkable instance of specific alterations of gene expression by ingestion of a drug. "Drosophila-based anticancer pharmacology" hence reveals unique properties for F14512, demonstrating the usefulness of an assay system that provides a low-cost, rapid and effective complement to mammalian models and permits the elucidation of fundamental mechanisms of action of candidate drugs of therapeutic interest in humans. Citation: Chelouah S, Monod-Wissler C, Bailly C, Barret J-M, Guilbaud N, et al. (2011) An Integrated Drosophila Model System Reveals Unique Properties for F14512, a Novel Polyamine-Containing Anticancer Drug That Targets Topoisomerase II. PLoS ONE 6(8): e23597. doi:10.1371/journal.pone.0023597 Editor: Claudine Mayer, Institut Pasteur, France Received April 4, 2011; Accepted July 21, 2011; Published August 10, 2011 Copyright: 2011 Chelouah 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: EK was funded by the Centre National de la Recherche Scientifique (CNRS, http://www.cnrs.fr/), the Association pour la Recherche sur le Cancer (ARC, http://www.arc-cancer.net/, grant No. 4897), the Ligue Nationale Contre le Cancer (LNCC, http://www.ligue-cancer.net/) and the Agence Nationale de la Recherche (ANR, http://www.agence-nationale-recherche.fr/, grant DNAMOTORS No. ANR-06-BLAN-0419 CSD 8). These funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Commercial funding to EK was provided in part by the Institut de Recherche Pierre Fabre, which played a role in study design, part of data collection and analysis, decision to publish and contributed to preparation of the manuscript. Competing Interests: The authors have read the journal’s policy and have the following conflicts: Christian Bailly, Jean-Marc Barret, Nicolas Guilbaud and Ste´phane Vispe´ are fulltime employees of Institut de Recherche Pierre Fabre (IRPF), which developed F14512, a patented drug, and conducts its ongoing phase I clinical trials; Research in Emmanuel Ka¨s’s lab was funded in part by IRPF, of which EK is neither an employee nor a consultant and is not involved in patents, products in development or marketed products. This does not alter the authors’ adherence to all the PLoS ONE policies on sharing data and materials. * E-mail: kas@ibcg.biotoul.fr (EK); stephane.vispe@pierre-fabre.com (SV) ¤a Current address: Institut de Cance´rologie Gustave Roussy, Signalisation, Noyaux et Innovations en Cance´rologie, UMR 8126 CNRS, Villejuif, France ¤b Current address: Universite´ de Toulouse 3, Centre de Biologie du De´veloppement UMR5547 CNRS, Toulouse, France Introduction Topoisomerase II (Topo II) is an essential enzyme that handles topological reactions in all living cells and is required for different aspects of DNA metabolism, from the resolution of constraints resulting from packaging of DNA into chromatin, to its transcription and replication and, most importantly, the conden- sation [1] and segregation of sister chromatids in the course of cell division (reviewed in [2,3,4]). As such, it constitutes a target of choice for anticancer drugs ([5,6,7] and references therein). Because the basic Topo II-mediated reaction is central to such key aspects of cellular activities, drugs that target its catalytic cycle and, most commonly, poison it, have been extensively used in anti- cancer therapies [8]. These include etoposide (VP16) and teniposide (VM26) still being widely used in human [6,9]. However, widespread applications have been limited by their high cytotoxicity to normal cells as well as the induction of drug resistance [10,11]. In proliferating cells, Topo II is an abundant enzyme, with approximately 2–36105 molecules/nucleus [1] preferentially lo- calized at sites found every 50–80 kbp [12]. Treatment of cells with Topo II poisons such as etoposide or teniposide can result in PLoS ONE | www.plosone.org 1 August 2011 | Volume 6 | Issue 8 | e23597