592 Research Article Does the lipid membrane composition of arsonoliposomes affect their anticancer activity? A cell culture study Paraskevi Zagana 1 , Maria Haikou 1 , Eleftheria Giannopoulou 2 , Panayiotis V. Ioannou 3 and Sophia G. Antimisiaris 1, 4 1 Laboratory of Pharmaceutical Technology, Department of Pharmacy, University of Patras, Patras, Greece 2 Laboratory of Pharmacology, School of Medicine, University of Patras, Patras, Greece 3 Department of Chemistry, University of Patras, Patras, Greece 4 Institute of Chemical Engineering and High-Temperature Processes-FORTH, Patras, Greece Sonicated arsonoliposomes were prepared using arsonolipid with palmitic acid acyl chain (C16), mixed with phosphatidylcholine (PC)-based or 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)- based, and cholesterol (Chol) with C16/DSPC/Chol 8:12:10 molar ratio. PEG-lipid (1,2-distearoyl-sn- glycero-3-phosphoethanolamine conjugated to polyethylenoglycol 2000) containing vesicles (PEGy- lated-arsonoliposomes; PC-based and DSPC-based) were also prepared. The cytotoxicity of these arsonoliposomes towards different cancer cells (human promyelocytic leukaemia NB4, Prostatic can- cer PC3, human breast adenocarcinoma MDA-MB-468, human T-lymphocyte (MT-4) and also towards human umbilical vein endothelial cells (HUVECs) was evaluated by calculating the arsonoli- posome-induced growth inhibition of the cells by the MTTassay. IC-50 values were interpolated from cell number/arsonoliposome concentration curves. The results reveal that all types of arsonolipo- somes evaluated significantly inhibit the growth of most of the cancer cells studied (PC3, NB4, MT4) with the exception of the MDA-MB-468 breast cancer cells which were minimally affected by arso- noliposomes; in some cases even less than HUVEC. Nevertheless, for the same cell type the differ- ences between the different types of arsonoliposomes were significant but not proportional to their stability, indicating that the formation of arsonoliposomes with very stable membranes is not a prob- lem for their anticancer activity. Thereby it is concluded that arsonoliposome composition should be adjusted in accordance to their in vivo kinetics and the desired, for each specific application, biodistri- bution of As and/or encapsulated drug. Keywords: Anticancer / Arsenic / Arsonolipid / Arsonoliposome / Lipid composition / Received: April 25, 2007; revised: June 16, 2007; accepted: June 18, 2007 1 Introduction Liposomes containing a type of Arsonolipids (Ars) (Fig. 1), referred to as , arsonoliposomes’ have been prepared and characterized in our laboratories previously [1]. As sum- marized before [2], promising results were obtained with some of the arsonoliposome types prepared, for which a dif- ferential toxicity towards cancer and normal cells was dem- onstrated [3, 4] as well as in vitro antiparasitic activity [5]. However, it has been found that the membrane integrity of arsonoliposomes [6, 7] and their in vivo distribution (of arsenic) [8, 9] are influenced by the lipid composition of the arsonoliposome membrane, and this has been pinpointed as the reason – or at least one of the reasons – for the different antiparasitic activities demonstrated (in vitro and in vivo), by different arsonoliposome types (with different lipid com- positions) [10]. The glutathione induced reduction of As(V) (of Ars) to As(III) which has higher toxicity, and the fact that gluta- thione concentration is much higher in tumour cells as com- pared to normal cells [11], is one of the mechanisms pro- posed to explain the demonstrated differential toxicity of Correspondence: Dr. Sophia G. Antimisiaris, Department of Phar- macy, University of Patras, University Campus, Rio 26500, Patras, Greece E-mail: santimis@upatras.gr Fax: +30-2610-996-302 Abbreviations: DSPC, 1,2-distearoyl-sn-glycero-3-phosphocholine; FCS, fetal calf serum; HUVEC, human umbilical vein endothelial cell; PC, phosphatidylcholine i 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.mnf-journal.com DOI 10.1002/mnfr.200700474 Mol. Nutr. Food Res. 2009, 53, 592 – 599