C6-ceramide

Ferrying of molecules into cells via entry through caveolae may represent a way to traffic specifically cytotoxic molecules to specific action sites. For example, elevating the intracellular level of the sphingolipid ceramide is known to exert antimitogenic and proapoptotic effects. While ceramide is cell-permeable and displays antiapoptotic properties in vitro, systemic in vivo use of ceramide is hampered by its hydrophobicity. Using a C6-ceramide formulation in pegylated liposomes was shown to elicit a sixfold reduction in solid phase tumors, when compared to unloaded liposomes in a mouse model of breast adenocarcinoma [68],... 

H]-ethanolamine into PE was inhibited by C6-ceramide in a dose and time-dependent manner (b) delayed disappearance of label from CDP-choline and CDP-ethanolamine in pulse-chase experiments indicated impaired conversion of these CDP-metaboUtes to PC and PE by CPT and EPT, respectively (c) the activities of CPT and EPT are decreased upon C6-ceramide treatment (see Eigure 2 adapted from Bladergroen et al. 1999b). In contrast to BHK cells the activity of CT was not affected significantly in rat-2 fibroblasts by short-chain ceramides. 

Figure 2. Effect of Ca-ceramide on phosphatidylcholine and phosphatidylethanolamine synthesis in rat-2 fibroblasts. Cells were treated for 2 h in the absence (open bars) or presence (hatched bars) of 25pM C6-ceramide and the following parameters were determined i) the incorporation of [ H]choline and [ H]ethanolamine into phosphatidylcholine (PC) and phosphatidylethanolamine (PE), respectively (panel A) and in CDP-choline and CDP-ethanolamine, respectively (panel B) and ii) the in vitro activity of choline- and ethanolaminephosphotransferase (CPT and EPT) (panel C).

In vitro and in vivo studies have shown that short-chain cer-amides induce growth arrest (154). A fluorescent analog of C6-ceramide partitioned into caveolin-enriched microdomains of rat aorta vascular smooth muscle cells and led to growth arrest via activation of PKC-zeta, which is recruited to lipid microdomains and subsequently reduces the activity of Akt (155). 

Other laboratories have also shown that enzymes of the CDP-choline pathway for the production of PC in the cells, including CPT, show reduced activity when cells are incubated with cell-permeable C2/C6 ceramides (Bladergroen et al., 1999 Ramos et al., 2002), and it has been predicted that this inhibition may be due to the competitive inhibition by ceramides owing to the similarity in the stmcture to one of the substrates for CPT, DAG. 

FIGURE 13 Blood and tumor concentrations of bioactive ceramide-lipid C6 in tumorbearing mice were maintained over a 48-h period (a) 10- and 40-mg/kg doses of liposomal-C6 followed first-order kinetics with blood concentration exceeding in vitro IC50 sustaibed at 48h (b) at these doses steady-state concentration of C6 in tumor tissue achieved at 60min. The 40-mg/kg dose maintained a concentration above the desired IC50 up to 48 h. (Reprinted from ref. 431 with permission of American Association for Cancer Research.)... 

Drug-incorporated STPP liposomes are prepared by adding 10 mole% ceramide C6 to the lipid composition mentioned in step 1 in Subheading 3.1 earlier. 

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