Delivery and Pathway in MCF7 Cells of DNA Vectorized by Cationic Liposomes Derived from Cholesterol

Abstract

We have investigated the delivery and the pathway in tumoral MCF7 cells of DNA carried by liposomes prepared from (trimethyl aminoethane carbamoyl cholesterol iodide (TMAE-Chol), a cholesterol-based cationic lipid with a quaternary ammonium on the polar head. The structure of DNA-liposome complexes depends on the length of DNA and on the lipid-DNA charge ratio X. Spherical beads constitute fine structures of the observed complexes even when they appear as aggregates. For oligonucleotide transfer, dissociation from liposomes after transfection, penetration of the oligonucleotides into nuclei, and a long resident time were observed. For plasmid transfer, a correlation between the variation in the transfection level and the ultrastructure of complexes was demonstrated. The results showed a cellular route of lipid/plasmid complexes from the beginning by endocytosis, entrapped into endosomes, released by the latter until entry in the perinuclear area, and then penetration of plasmids inside the nuclei resulting in the observed expression of the beta-galactosidase gene.

Biotin-directed assembly of targeted modular lipoplexes and their transfection of human hepatoma cells in vitro

“…However, these large liposomes have been shown previously to form complexes with DNA that present as clusters (100-200 nm) composed of small unilamellar vesicles in the 30-50 nm size range (Singh et al, 2006). These observations are consistent with the ultrastructure reported for lipoplexes formed between liposomes containing the monocationic cholesteryl cytofectin trimethylaminoethane carbamoyl cholesterol (TMAEC-chol) and the plasmid pCMV-β (Cao et al, 2000).…”

Cholesteryl Cytofectins with Primary Amino Head Groups Transfect Transformed Human Epithelial Cell Lines Efficiently

“…Similar clusters have been described by others (Percot et al 2004;Cao et al 2000). The shape adopted by lipoplexes is influenced by the size of plasmid DNA, the type of liposome, and the conditions and mechanisms adopted for lipoplex formation (Sun et al 2004).…”

“…It is understood that primary amino groups on cytofectins MS10 and MS11 are protonated at pH 7.5, as the pK b values for such groups are about 3.5. This, in large measure, explains the apparent excess positive charge observed in such complexes when full DNA retardation is observed in band shift assays (Cao et al 2000;Kisoon et al 2002).…”

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

“…Lipoplexes have been shown to enter cells via clathrininvolved endocytosis, and to become entrapped in endosomes, being released from these vesicular structures and gaining entry into the perinuclear area before ® nally being taken up into the nucleus (Cao et al 2000). Friend et al (1996) describe vesicular and reticular intranuclear membranes probably resulting from fusion of lipoplexes with the nuclear envelope.…”

“…Liposomes formulated without neutral lipid(s) have inferior rates of transfection (Lasic & Pearlman 1996). That lipoplexes may be stored for as long as a year in sterile water (Cao et al 2000) and may be administered in-vivo via the vascular route (Dass 1998) highlights the versatile usefulness of these vehicles. However, shelf-life is highly dependent on the chemical constituents in the formulation as some formulations tend to form macroscopic aggregates over time (Das & Niven 2001).…”