DOTAP/DOPE ratio and cell type determine transfection efficiency with DOTAP-liposomes.

The effects of lipid compositions on their physicochemical properties and transfection efficiencies were investigated. Four liposome formulations with different 1,2-dioleoyl-3-trimethylammoniumpropane (DOTAP) to dioleoylphosphatidylethanolamine (DOPE) weight ratios were investigated, that is, weight ratios 1:0 (T1P0), 3:1 (T3P1), 1:1 (T1P1), and 1:3 (T1P3). Mean sizes of liposomes were influenced by their lipid composition and the preparation concentration at the time of sonication. Zeta potentials of liposomes were inversely correlated with their liposome sizes. However, neither liposome sizes nor zeta potentials were correlated with transfection efficiency. The optimum composition of liposomes was cell-line dependent (T1P0 and T3P1 for Huh7 and AGS, T3P1 and T1P1 for COS7, and T1P1 and T1P3 for A549). The shape of lipoplexes was changed from lamellar to inverted hexagonal structure according to the increased ratio of DOPE, but there was no definite advantage of specific structure in transfection efficiency throughout all used cell lines. However, cellular internalization was consistently faster in T1P0, T3P1, T1P1 compared to T1P3 in all cell lines, suggesting the importance of endosomal escape. Our findings show that the transfection efficiency of DOTAP liposomes is mainly influenced by lipid composition and cell type, and not by size or zeta potential.

Efficient delivery of plasmid DNA using cholesterol-based cationic lipids containing polyamines and ether linkages.

Cationic liposomes are broadly used as non-viral vectors to deliver genetic materials that can be used to treat various diseases including cancer. To circumvent problems associated with cationic liposome-mediated delivery systems such as low transfection efficiency and serum-induced inhibition, cholesterol-based cationic lipids have been synthesized that resist the effects of serum. The introduction of an ether-type linkage and extension of the aminopropyl head group on the cholesterol backbone increased the transfection efficiency and DNA binding affinity compared to a carbamoyl-type linkage and a mono aminopropyl head group, respectively. Under optimal conditions, each liposome formulation showed higher transfection efficiency in AGS and Huh-7 cells than commercially available cationic liposomes, particularly in the presence of serum. The following molecular structures were found to have a positive effect on transfection properties: (i) extended aminopropyl head groups for a strong binding affinity to plasmid DNA; (ii) an ether linkage that favors electrostatic binding to plasmid DNA; and (iii) a cholesterol backbone for serum resistance.

Transfection property of a new cholesterol-based cationic lipid containing tri-2-hydroxyethylamine as gene delivery vehicle.

A novel cholesterol-based cationic lipid containing a tri-2- hydroxyethylamine head group and ether linker (Chol- THEA) was synthesized and examined as a potent gene delivery vehicle. In the preparation of cationic liposome, the addition of DOPE as helper lipid significantly increased the transfection efficiency. To find the optimum transfection efficiency, we screened various weight ratios of DOPE and liposome/DNA (N/P). The best transfection efficiency was found at the Chol-THEA:DOPE weight ratio of 1:1 and N/P weight ratio of 10~15. Most of the plasmid DNA was retarded by this liposome at the optimum N/P weight ratio of 10. The transfection efficiency of Chol-THEA liposome was compared with DOTAP, Lipofectamine, and DMRIE-C using the luciferase assay and GFP expression. Chol-THEA liposome with low toxicity had better or similar potency of gene delivery compared with commercial liposomes in COS-7, Huh-7, and MCF-7 cells. Therefore, Chol-THEA could be a useful non-viral vector for gene delivery.

The synthesis of cholesterol-based cationic lipids with trimethylamine head and the effect of spacer structures on transfection efficiency.

Five cholesterol-based cationic lipids were newly synthesized based on cholest-5-en-3ß-oxyethane-N,N,N-trimethylammonium bromide (Chol-ETA) structure where the cholesterol backbone is linked to cationic head via various lengths of ether-linked carbon spacer. The transfection efficiency of these compounds was increased in order of three (Chol-PRO)