PRECLINICAL REPORTSCamptothecin and Fas receptor agonists synergistically induce medulloblastoma cell death: ROS-dependent mechanismsLi, Yanga; Rory Goodwin, Courtneya b; Sang, Yingyinga; Rosen, Eliot M.e; Laterra, Johna b c d; Xia, Shulia d Author Information aHugo W. Moser Research Institute at Kennedy Krieger Departments of bNeuroscience cOncology dNeurology, School of Medicine, Johns Hopkins University, Baltimore, Maryland eDepartment of Oncology, Lombardi Cancer Center, Washington, DC, USA Correspondence to Dr Shuli Xia, Kennedy Krieger Research Institute, 707 N. Broadway, Room 400K, Baltimore, MD 21205, USA Tel: +1 443 923 2683; fax: +1 443 923 2695; e-mail: [email protected] Received 3 April 2009 Revised form accepted 20 June 2009 Anti-Cancer Drugs 20(9):p 770-778, October 2009. | DOI: 10.1097/CAD.0b013e32832fe472 Buy Metrics Abstract Medulloblastoma, a common malignant pediatric brain tumor, is highly resistant to death receptor-mediated apoptosis despite death receptor expression by tumor cells. Developing new strategies to overcome this resistance to death receptor activation could positively impact therapeutic outcomes. We explored the modulation of death receptor-induced medulloblastoma cell death by the topoisomerase I inhibitor camptothecin (CPT). CPT significantly increased the human medulloblastoma DAOY cell death response to agonistic anti-Fas antibody (CH-11). Cell death after CPT, CH-11, and CPT+CH-11 treatment was 9, 7, and 33%, respectively. Isobologram analysis showed that CH-11 and CPT act synergistically to induce cell death in DAOY cells. A similar pattern of synergism between CPT and CH-11 was found in ONS-76 medulloblastoma cells. Synergistic cell death was found to be predominantly apoptotic involving both extrinsic and intrinsic pathways as evidenced by annexin V staining, cleavage of caspases (3, 8, and 9), Bid and PARP, and cytoprotection by caspase inhibitors. Flow cytometric analyses showed that expression of cell surface Fas or Fas ligand did not change with drug treatment. Western blot analyses showed that the combination of CH-11+CPT induced a significant decrease in XIAP levels. Furthermore, reactive oxygen species, especially O2−, were elevated after CPT treatment, and even more so by the CH-11+CPT treatment. The antioxidants glutathione and N-acetyl-cysteine prevented cell death induced by CPT+CH-11. Moreover, the mitochondrial respiratory chain complex I inhibitor rotenone potentiated CH-11-induced apoptosis in DAOY cells. Taken together, these findings show that CPT synergizes with Fas activation to induce medulloblastoma apoptosis through a mechanism involving reactive oxygen species and oxidative stress pathways. © 2009 Lippincott Williams & Wilkins, Inc.