Arch Pharm Res Vol 32, No 11, 1545-1554, 2009 DOI 10.1007/s12272-009-2106-0 1545 Snake Venom Toxin Inhibits Cell Growth through Induction of Apoptosis in Neuroblastoma Cells Mi Hee Park 1 , Dong Ju Son 1 , Dong Hoon Kwak 1 , Ho Sueb Song 2 , Ki-Wan Oh 1 , Hwan-Soo Yoo 1 , Yong Moon Lee 1 , Min Jong Song 3 , and Jin Tae Hong 1 1 College of Pharmacy and CBITRC, Chungbuk National University, Cheongju 361-763, Korea, 2 College of Oriental Medi- cine, Kyungwon University, San 65 Bokjeong-dong, Sujeong-gu, Seongnam, Gyeonggii 461-701, Korea, and 3 Obstetrics and Gynecology, SEOUL ST.MARY'S HOSPITAL, The Catholic University, Korea (Received May 18, 2009/Revised September 6, 2009/Accepted September 6, 2009) Snake venom toxin from Vipera lebetina turanica can induce apoptosis in many cancer cell lines, but there is no study about the apoptotic effect of snake venom toxin on human neuro- blastoma cells. In this study, we investigated the apoptotic effect of snake venom toxin in human neuroblastoma SK-N-MC and SK-N-SH cells. Our result showed that cell detachment and apoptotic cell death were increased by snake venom toxin (1.25-10 µg/mL), but normal neuronal cells were not affected. Consistent with the induction of apoptosis, the level of reac- tive oxygen species (ROS) was increased, but mitochondrial membrane potential (MMP) was disrupted by treatment with snake venom toxin. However, the glutathione prevented snake venom toxin-induced cell growth inhibition. Snake venom toxin also increased the expression of pro-apoptotic protein Bax, but down-regulated anti-apoptotic protein Bcl-2. Therefore, these results showed that snake venom toxin from Vipera lebetina turanica causes apoptotic cell death of neuroblastoma cells through ROS dependent MMP disruption, and suggested that snake venom toxin may be applicable as an anti-cancer agent for neuroblastoma. Key words: Snake venom toxin, Apoptosis, Neuroblastoma cells INTRODUCTION Neuroblastoma, the most common extracranial solid tumor of infancy, accounts for 15% of deaths from all childhood cancers (Torkin et al., 2005). The prognosis is both age- and stage-dependent, with metastases frequently occurring in older children (Ginkel et al., 2007). For these high-risk neuroblastoma patients, the prospect of long-term survival is poor despite of the intensive multimodal therapy. As survival rates for stage IV disease are still < 50%, it is important to develop more effective modes of therapy (Combaret et al., 2008). Cytotoxic agents currently used in the treatment of neuroblastoma mediate cell death by activating key elements of the apoptotic signaling pathways (Choi et al., 2008; Mühlethaler-Mottet et al., 2008; Pagnan et al., 2009). Apoptotic or programmed cell death is a different form of cell death regulated by internal genetic pro- grams and characterized by specific morphological and biochemical properties. Several biochemical markers have been identified in the induction of apoptotic cell death including increase of reactive oxygen species (ROS), decrease of mitochondrial membrane potential (MMP) and changes in the level of Bcl-2 and Bax proteins in mitochondria (Duan et al., 2009; Itahana et al., 2008; Martínez-Pastor et al., 2009; Saleem et al., 2008). In the intrinsic apoptotic cell death pathway, ROS are potent inducers of oxidative damage and have been proposed as critical regulators of apoptotic cell death. ROS can induce opening of the permeability transition pore through oxidation-dependent mechani- sms (Armstrong et al., 2002; Miller et al., 2003). Loss of membrane potential can cause apoptotic cell death by the release of cytochrome c from mitochondria, and a decrease in the Bcl-2-Bax ratio associated with mitochondria (Cervia et al., 2007; Pasquier et al., Correspondence to: Jin Tae Hong, College of Pharmacy and CBITRC, Chungbuk National University, Cheongju 361-763, Korea Tel: 82-43-261-2813, Fax: 82-43-268-2732 E-mail: jinthong@cbucc.chungbuk.ac.kr