Liposomes Containing Cationic Dimethyl Dioctadecyl Ammonium Bromide: Formulation, Quality Control, and Lipofection Efficiency

Abstract

This article describes a novel, simple, and relatively inexpensive method to prepare cationic liposomes using an ethanol injection/pressure extrusion method. The study also demonstrated that binding erythrosine dye to cationic liposomes results in a shift of the absorption maximum of the dye from 528 nm to 549 nm at pH 4.25, allowing quantification and visualization of these vesicles. In addition, a relatively simple Ficoll-based gradient centrifugation method for separation of lipoplexes from unbound molecules is presented. Laboratory-formulated dimethyl dioctadecyl ammonium bromide (DDAB) containing liposomes were just as efficient in complexing nucleic acids as commercially available types, and binding increased as the positive to neutral lipid ratio was increased. Transfection efficiency of the DDAB-containing liposomes increased as the ratio of cationic to neutral lipid was increased from 1:1 to 4:1 with either PtdChol or DOPE as the neutral lipid. A concomitant increase in cytotoxicity of CSU-SA1 cancer cells was noted as the ratio of positive to neutral lipid of the liposomes was increased. Nevertheless, our present study showed that the 2:1 liposome is a good choice since it delivers functional plasmids at a comparable rate to commercial liposome formulations, has similar toxicities to the less harmful commercial liposomes, and is at least 1000-fold more economical to prepare inhouse, a major factor to be considered in preclinical and clinical studies with these carriers.

In Vitro Antifungal Activities of Inhibitors of Phospholipases from the Fungal Pathogen Cryptococcus neoformans

“…Three of the compounds tested are nontoxic to humans and have already been approved for use in other situations: compound O as a component in liposomes for drug delivery (12), compound D as an anticholinergic (26), and compound A as a bactericidal agent for the treatment of gingivitis (9). Compound P is known as an ion-pairing reagent (30).…”

Cellular VPS4 Is Required for Efficient Entry and Egress of Budded Virions of Autographa californica Multiple Nucleopolyhedrovirus

“…Unless specified otherwise, Sf9 and High 5 cells were seeded at a density of 1 ϫ 10 6 cells per well in six-well plates for transfections. Transfection of plasmid DNAs was performed using a standard CaPO 4 precipitation procedure (8,9), and bacmid transfections were performed with a laboratory-formulated dimethyl dioctadecyl ammonium bromide (DDAB)/dioleoyl phosphatidylethanolamine (DOPE) liposome method (19). For viral infections, the virus was incubated on cells for 1 h, and then cells were washed once in TNMFH.…”

Cytotoxicity issues pertinent to lipoplex-mediated gene therapy in-vivo

“…Lipoplex-mediated toxicity is closely associated with the charge ratio between the cationic lipid species in the formulation and the nucleic acid, as well as the dose of lipoplexes administered (Dass et al 2002). Higher charge ratios are generally more toxic to a variety of cell types, including cancer cell lines.…”

“…Lipid-related eOE ects include the cationic lipid component making up the vesicle (Dass 1998), cationic to neutral lipid ratio (Dass et al 2002), and the type of neutral lipid in the vesicle (Boussif et al 2001). Formulation modi® cation-related eOE ects include docking of lipoplexes on to larger solid particles such as polystyrene divinylbenzene microspheres , inclusion of fusogenic viral coat proteins (Namoto et al 1998), or the presence of other non-lipidic biomolecules in the precomplexed liposomes (Chen et al 2000).…”