1-2-dioleoyloxy-3-(trimethylammonium)propane and 1-2-distearoylphosphatidylethanolamine

Acanthamoeba keratitis is an ophthalmic disease with no specific treatment that specially affects contact lens users. The silencing of serine phosphatase (SP) and glycogen phosphorylase (GP) proteins produced by Acanthamoeba has been shown to significantly reduce the cytopathic effect, although no vehicle was proposed yet to deliver the siRNA sequences to the trophozoites. In this study, PEGylated cationic liposomes were proposed and optimized using Box-Behnken design. The influence of DOTAP:DOPE ratio, DSPE-PEG concentration, and siRNA/DOTAP charge ratio were evaluated over both biological response and physicochemical properties of liposomes. The ratio of DOTAP:DOPE had an effect in the trophozoite activity whereas the charge ratio influenced both size and protease activity. The predicted values were very close to the observed values, yielding a formulation with good activity and toxicity profile, which was used in the following experiments. A murine model of ocular keratitis was treated with siGP + siSP-loaded liposomes, as well as their respective controls, and combined treatment of liposomes and chlorhexidine. After 15 days of eight daily administrations, the liposomal complex combined with chlorhexidine was the only treatment able to reverse the more severe lesions associated with keratitis. There was 60% complete regression in corneal damage, with histological sections demonstrating the presence of an integral epithelium, without lymphocytic infiltrate. The set of results demonstrate the efficacy of a combined therapy based on siRNA with classical drugs for a better prognosis of keratitis caused by Acanthamoeba.

Topics: Acanthamoeba; Acanthamoeba Keratitis; Animals; Chlorhexidine; Cornea; Disease Models, Animal; Drug Administration Schedule; Drug Compounding; Drug Delivery Systems; Drug Therapy, Combination; Factor Analysis, Statistical; Fatty Acids, Monounsaturated; Gene Expression Regulation; Glycogen Phosphorylase; Humans; Liposomes; Phosphatidylethanolamines; Phosphoric Monoester Hydrolases; Polyethylene Glycols; Protozoan Proteins; Quaternary Ammonium Compounds; Rats; Rats, Wistar; RNA, Small Interfering; Trophozoites

Drug-fortified cationic liposomes of 6‑methoxy‑2‑naphthylacetic acid (6‑MNA) were prepared and characterized by various techniques. The residence time of drug-fortified liposomes in joint cavity was evaluated by intra-articular (IA) administration of the radio-labeled (

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Arthritis, Experimental; Cartilage, Articular; Cell Survival; Delayed-Action Preparations; Fatty Acids, Monounsaturated; Liposomes; Male; Mice; Naphthaleneacetic Acids; NIH 3T3 Cells; Phosphatidylcholines; Phosphatidylethanolamines; Quaternary Ammonium Compounds; Rats, Sprague-Dawley

When exposed to cancer cells, cytotoxic drugs such as doxorubicin (DOX) can lead to the induction of heat shock protein 90 (Hsp90), a molecular chaperone associated with a number of cancer-related client proteins, and result in cell survival. Co-administration of DOX with tanespimycin (TNP), an Hsp90 inhibitor, can sensitize the cancer cells to the cytotoxic effects of DOX. The effect of such a combination has been found to depend on the schedule of administration. Sequential administration of DOX and TNP has been linked to highly synergistic combination effects. Therefore, we aimed to develop folate-receptor targeted hybrid lipid-core nanocapsules comprising a hybrid lipid core lodging TNP and a polymeric corona lodging DOX (F-DTN). These nanocarriers were capable of delivering DOX and TNP sequentially, which was well demonstrated by an in vitro release study. The in vitro release profiles displayed pH-dependent and sustained release features. F-DTN exhibited excellent morphological characteristics with highly monodispersed particles. In vitro tests with F-DTN in MCF-7 cell line demonstrated exceptional cytotoxicity, with high cellular uptake and apoptosis. These findings were appreciably more assertive than tests with free individual drugs (DOX, TNP), free drug combination (DOX/TNP), or non-folate receptor-targeted hybrid lipid-core nanocapsules (DTN). In vivo pharmacokinetic study revealed noticeable enhancement of bioavailability and plasma circulation time of the drugs when encapsulated in the carrier system. Therefore, hybrid lipid-core nanocapsules have the potential to be utilized for application in folate receptor-targeted combination chemotherapy.

Topics: A549 Cells; Animals; Antineoplastic Agents; Apoptosis; Benzoquinones; Biomarkers; Dimethyl Sulfoxide; Doxorubicin; Drug Delivery Systems; Fatty Acids, Monounsaturated; Folate Receptor 1; Folic Acid; Gene Expression; Humans; Injections, Intravenous; Lactams, Macrocyclic; Male; MCF-7 Cells; Micelles; Nanocapsules; Neoplasm Proteins; Phosphatidylethanolamines; Quaternary Ammonium Compounds; Rats; Rats, Sprague-Dawley

Nonviral gene delivery vectors hold great promise for gene therapy due to the safety concerns with viral vectors. However, the application of nonviral vectors is hindered by their low transfection efficiency. Herein, in order to tackle this challenge, we developed a nonviral vector integrating lipids, sleeping beauty transposon system and 8-mer stem cell targeting peptides for safe and efficient gene delivery to hard-to-transfect mesenchymal stem cells (MSCs). The 8-mer MSC-targeting peptides, when synthetically reiterated in three folds and chemically presented on the surface, significantly promoted the resultant lipid-based nanoparticles (LBNs) to deliver VEGF gene into MSCs with a high transfection efficiency (∼52%) and long-lasting gene expression (for longer than 170 h) when compared to nonreiterated peptides. However, the reiterated stem cell targeting peptides do not enable the highly efficient gene transfer to other control cells. This work suggests that the surface presentation of the reiterated stem cell-targeting peptides on the nonviral vectors is a promising method for improving the efficiency of cell-specific nonviral gene transfection in stem cells.

Topics: Amino Acid Sequence; Animals; Cell Survival; Endocytosis; Fatty Acids, Monounsaturated; Gene Expression; Gene Transfer Techniques; Liposomes; Mesenchymal Stem Cells; Molecular Sequence Data; Nanoparticles; Particle Size; Peptides; Phosphatidylethanolamines; Quaternary Ammonium Compounds; Rats; Transgenes; Transposases; Vascular Endothelial Growth Factor A

Targeted gene delivery offers immense potential for clinical applications. Liposomes decorated with targeting ligands have been extensively used for both in vitro and in vivo gene delivery. Lipoplexes with high cholesterol content that result in cholesterol domain formation within the complexes have been shown to exhibit enhanced transfection in vitro and resistance to serum-induced aggregation. In the present study, folate was employed as a targeting ligand that was conjugated with either cholesterol or a diacyl lipid (DSPE), and these conjugates were incorporated into lipoplexes formulated with DOTAP/cholesterol (wt/wt: 31/69) that are known to possess cholesterol nanodomains. Cellular uptake and transfection of these lipoplexes in the presence of 50% serum were examined when the ligand was located within or excluded from the cholesterol nanodomain. Lipoplexes with folate-cholesterol exhibited a 50-fold increase in transfection compared to those with folate-DSPE, while the cellular uptake level is only 40% of that with folate-DSPE. These results indicate that the presence of the ligand within the cholesterol domain promotes more productive transfection in cultured cells, and intracellular trafficking of the lipoplexes after entry into cells plays a crucial role in gene delivery.

Topics: Calorimetry, Differential Scanning; Cell Line, Tumor; Cholesterol; Fatty Acids, Monounsaturated; Flow Cytometry; Folic Acid; Gene Transfer Techniques; Humans; Liposomes; Phosphatidylethanolamines; Quaternary Ammonium Compounds; Scattering, Radiation

Several reports have shown a fast and efficient translocation of TAT-modified lipoplexes and particles into the cell cytoplasm. However, neither the uptake mechanism nor the biological effect of TAT-modified lipoplexes has been studied in detail. In this report we show that the increase in gene transfer of TAT-modified lipoplexes depends on the amount of cationic lipid in the lipoplexes and on the way TAT was coupled to the lipoplexes. We demonstrate that the cellular uptake of both TAT-modified and unmodified lipoplexes is very fast and, in contrast to previous publications, temperature-dependent. Additionally, after internalization TAT-modified as well as unmodified lipoplexes end up in lysosomal vesicles, indicating the involvement of clathrin-mediated endocytosis. Furthermore, chlorpromazine, a specific inhibitor of clathrin-dependent endocytosis, strongly inhibits the cellular uptake and biological activity of both the TAT-modified and unmodified lipoplexes. We also found that the uptake and biological activity of these lipoplexes are diminished when cholesterol in the cell membrane was bound by filipin, an inhibitor of the lipid-raft mediated pathway. Considering these data, we conclude that TAT-modified and unmodified lipoplexes are mainly internalized via a cholesterol-dependent clathrin-mediated pathway.

Topics: Animals; Chlorocebus aethiops; Cholesterol; Clathrin; COS Cells; Endocytosis; Fatty Acids, Monounsaturated; Filipin; Fluorescein-5-isothiocyanate; Fluorescent Dyes; Gene Products, tat; Gene Transfer Techniques; Genetic Therapy; Liposomes; Lysosomes; Membrane Microdomains; Models, Molecular; Molecular Structure; Phosphatidylethanolamines; Polyethylene Glycols; Quaternary Ammonium Compounds; Temperature; Time Factors

Topics: Animals; Cell Line; Choline; Coronary Restenosis; DNA; Fatty Acids, Monounsaturated; Gene Targeting; Genes, Reporter; Green Fluorescent Proteins; Liposomes; Luminescent Proteins; Muscle, Smooth, Vascular; Phosphatidylethanolamines; Polyethylene Glycols; Polyethyleneimine; Polylysine; Protamines; Quaternary Ammonium Compounds; Rats; Transfection