Therapeutic Discovery Inhibition of Dynamin by Dynole 34-2 Induces Cell Death following Cytokinesis Failure in Cancer Cells Megan Chircop 1 , Swetha Perera 1 , Anna Mariana 1 , Hui Lau 4 , Maggie P.C. Ma 1 , Jayne Gilbert 2 , Nigel C. Jones 5 , Christopher P. Gordon 3 , Kelly A. Young 3 , Andrew Morokoff 4 , Jennette Sakoff 2 , Terence J. O’Brien 5 , Adam McCluskey 3 , and Phillip J. Robinson 1 Abstract Inhibitors of mitotic proteins such as Aurora kinase and polo-like kinase have shown promise in preclinical or early clinical development for cancer treatment. We have reported that the MiTMAB class of dynamin small molecule inhibitors are new antimitotic agents with a novel mechanism of action, blocking cytokinesis. Here, we examined 5 of the most potent of a new series of dynamin GTPase inhibitors called dynoles. They all induced cytokinesis failure at the point of abscission, consistent with inhibition of dynamin while not affecting other cell cycle stages. All 5 dynoles inhibited cell proliferation (MTT and colony formation assays) in 11 cancer cell lines. The most potent GTPase inhibitor, dynole 34-2, also induced apoptosis, as revealed by cell blebbing, DNA fragmentation, and PARP cleavage. Cell death was induced specifically following cytokinesis failure, suggesting that dynole 34-2 selectively targets dividing cells. Dividing HeLa cells were more sensitive to the antiproliferative properties of all 5 dynoles compared with nondividing cells, and nontumorigenic fibroblasts were less sensitive to cell death induced by dynole 34-2. Thus, the dynoles are a second class of dynamin GTPase inhibitors, with dynole 34-2 as the lead compound, that are novel antimitotic compounds acting specifically at the abscission stage. Mol Cancer Ther; 10(9); 1553–62. Ó2011 AACR. Introduction Several small molecule inhibitors targeting mitotic proteins have shown promise in preclinical or early clinical development as cancer treatments (1). These mostly act at early mitotic stages, primarily disrupting metaphase–anaphase transition (and spindle assembly checkpoint; SAC) by targeting cyclin-dependent kinase, Aurora kinase, polo-like kinase (Plk), or kinesin spindle protein (1, 2). These new mitotic inhibitors prevent pro- liferation of most tumor cells in vitro and reduce tumor volume in vivo by inhibiting growth and/or triggering cell death following an aberrant mitotic event leading to aneuploidy (1, 2). Such compounds are expected to have a more favorable therapeutic window than currently used chemotherapeutic agents, for example, paclitaxel (1), as they would spare nondividing, nondifferentiating cells. The endocytic protein, dynamin II (dynII), may also be a novel mitotic target for development of selective anti- cancer drugs, because dynII exclusively functions during the abscission stage of cytokinesis and is not required for progression through any other cell cycle phase (3). Dyna- min inhibitors tested to date exclusively block cytokinesis (3), a cell cycle stage not yet targeted for chemotherapeu- tic intervention. We recently reported the anticancer properties of dyna- min inhibitors within the long chain amines and ammo- nium salts series (MiTMAB; refs. 3–5). dynII is best known for its role in membrane trafficking processes, specifically in clathrin-mediated endocytosis (6–8). The MiTMABs inhibit dynamin-dependent receptor- mediated endocytosis (RME; refs. 5, 9). Consistent with a role for dynamin in cytokinesis (3, 6, 7, 10–14), the MiTMABs induce cytokinesis failure in synchronized HeLa cells, specifically blocking abscission (3). Unlike the Aurora kinase and Plk inhibitors, dynamin inhibitors tested so far do not affect progression through any other mitosis stage. Like Aurora kinases, Plk or kinesin spindle protein inhibitors (1), MiTMABs have antiproliferative and cytotoxicity properties that seem to be selective for cancer cells (3). Several other small molecule inhibitors of dynamin have been developed by our group and others, including Authors' Affiliations: 1 Children's Medical Research Institute, The Uni- versity of Sydney, 214 Hawkesbury Road, Westmead; 2 Department of Medical Oncology, Newcastle Mater Misericordiae Hospital, Edith Street, Waratah; 3 Chemistry, School of Environmental & Life Sciences, The University of Newcastle, Callaghan, New South Wales; and Departments of 4 Surgery and 5 Medicine, Royal Melbourne Hospital, University of Melbourne, Parkville, Victoria, Australia Note: Supplementary material for this article is available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/). Corresponding Author: Megan Chircop, Children's Medical Research Institute, The University of Sydney, Locked Bag 23, Wentworthville, NSW 2145, Australia. Phone: 61-2-9687-2800; Fax: 61-2-9687-2120; E-mail: mchircop@cmri.org.au doi: 10.1158/1535-7163.MCT-11-0067 Ó2011 American Association for Cancer Research. Molecular Cancer Therapeutics www.aacrjournals.org 1553 on October 4, 2021. © 2011 American Association for Cancer Research. mct.aacrjournals.org Downloaded from Published OnlineFirst July 12, 2011; DOI: 10.1158/1535-7163.MCT-11-0067