Cytotoxicity of an Anti-cancer Lysophospholipid through Selective Modification of Lipid Raft Composition

Abstract

Edelfosine is a prototypical member of the alkylphosphocholine class of antitumor drugs. Saccharomyces cerevisiae was used to screen for genes that modulate edelfosine cytotoxicity and identified sterol and sphingolipid pathways as relevant regulators. Edelfosine addition to yeast resulted in the selective partitioning of the essential plasma membrane protein Pma1p out of lipid rafts. Microscopic analysis revealed that Pma1p moved from the plasma membrane to intracellular punctate regions and finally localized to the vacuole. Consistent with altered sterol and sphingolipid synthesis resulting in increased edelfosine sensitivity, mislocalization of Pma1p was preceded by the movement of sterols out of the plasma membrane. Cells with enfeebled endocytosis and vacuolar protease activities prevented edelfosine-mediated (i) mobilization of sterols, (ii) loss of Pma1p from lipid rafts, and (iii) cell death. The activities of proteins and signaling processes are meaningfully altered by changes in lipid raft biophysical properties. This study points to a novel mode of action for an anti-cancer drug through modification of plasma membrane lipid composition resulting in the displacement of an essential protein from lipid rafts.The synthetic lipid edelfosine (also known as 1-O-octadecyl-2-Omethyl-rac-glycero-3-phosphocholine or ET-18-OCH3) is a prototypical member of the alkylphosphocholine class of cancer chemotherapeutic drugs. Alkylphosphocholines are effective drugs as they contain ether-linked fatty acids, as opposed to ester-linked fatty acids prevalent in endogenous phospholipids, and thus are much more resistant to cellular degradation by phospholipases (1-3). Because of their similarity in structure to phosphatidylcholine (PC), 5 the main experimentations to determine a mechanism of action for edelfosine and other alkylphosphocholines have focused on PC metabolism. This course of action was supported by observations that only alkylphospholipids with choline head groups, but not head groups found on other phospholipids such as ethanolamine or serine, were effective antitumor agents (1). Edelfosine and other choline-containing alkylphospholipids were found to inhibit PC synthesis and this correlated with inhibition of cell growth in various cancer cell lines (4 -7).Further metabolic labeling demonstrated that alkylphosphocholine drugs can inhibit the synthesis of PC-derived sphingomyelin, and this correlated with increased ceramide mass. Inhibition of de novo ceramide synthesis by the addition of the ceramide synthase inhibitor fumonisin B1 decreased ceramide levels and this was associated with increased resistance to alkylphosphocholines (8). As ceramide is a lipid second messenger whose accumulation can result in cytostasis or apoptosis (9, 10), an increase in cellular ceramide-mediated signaling was an alternate hypothesis proposed for alkylphosphocholine-mediated cell death (8).To gain further insight into the mechanism of action of alkylphosphocholine-mediated cytotoxicity we performed a genetic screen in Saccha...

Lipid-dependent surface transport of the proton pumping ATPase: A model to study plasma membrane biogenesis in yeast

“…Indicating that phosphorylation and lipid raft association may play important roles in maintaining protein stability at the plasma membrane [19,20]. This view is consistent with the fact that disruption of membrane microdomains at the plasma membrane by incubation with the anti-cancer drug edelfosine, an alkylglycerophosphocholine, disrupts raft association of plasma membrane localized Pma1 and induces its vacuolar turnover [21].…”

Antitumor alkyl-lysophospholipid analog edelfosine induces apoptosis in pancreatic cancer by targeting endoplasmic reticulum

“…We have previously shown that edelfosine increases cytosolic [Ca 2 þ ] in HeLa cells (Nieto-Miguel et al, 2007). On these grounds, and taking into account the interaction of edelfosine with cell membranes and lipids (Gajate et al, , 2009a(Gajate et al, , 2009bZaremberg et al, 2005;Mollinedo and Gajate, 2006;Torrecillas et al, 2006;Busto et al, 2007Busto et al, , 2008Ausili et al, 2008;, it might be envisaged that the increase in cytosolic [Ca 2 þ ] could be due to an increased permeability of the plasma membrane, resulting in Ca 2 þ entry from the extracellular medium, which it then leads to the herein reported increase in [Ca 2 þ ] ER , well above the steady state levels in control cells. The effect on [Ca 2 þ ] ER may seem puzzling, because ER stress has been rather associated with ER-stored Ca 2 þ release.…”

Pivotal role of dihydrofolate reductase knockdown in the anticancer activity of 2-hydroxyoleic acid

“…Such modifications can be used for therapeutic purposes, an approach termed ''membrane-lipid therapy'' (7,24). Indeed, lipid molecules such as edelfosine and mitelfosine are potent antineoplastic drugs that target the membrane (25,26), and they induce apoptosis in cancer cells through mechanisms that alter membrane lipid raft organization (26). By contrast with oleic acid, ␣-hydroxy derivatives of fatty acids (e.g., HOA) are less prone to be used by cells, and they are therefore likely to display better pharmacological properties (27).…”