Gemini Amphiphile-Based Lipoplexes for Efficient Gene Delivery: Synthesis, Formulation Development, Characterization, Gene Transfection, and Biodistribution Studies

Abstract

Some quaternary gemini amphiphiles (GAs) were synthesized as nonviral gene delivery carriers. The critical miceller concentration values of these amphiphiles are indicative of their superior surface-active properties. All of the synthesized GAs, alone or along with lipids like cholesterol and/or dioleoylphosphatidyl ethanolamine (DOPE), were formulated as liposomes. Formulations of GAs with DOPE showed average particle diameters of 326–400 nm with positive ζ-potential (30.1–46.4 mV). The lipoplexes of theses formulations showed complete pDNA retention at the base at a N/P ratio higher than 1.0 in gel retardation study. The GAs were effective in condensing pDNA into a ψ-phase, as indicated by circular dichroism study, and provided complete protection of the pDNA against the enzyme DNase at a N/P ratio more than 1. In vitro cell line studies showed that GA liposomal formulations caused β-gal expression and offered a higher transfection efficiency than that of liposomes prepared with the help of N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium methyl-sulfate (DOTAP)/DOPE and dicyclocarbodiimide (DCC)/DOPE but comparable to those of Lipofectamine 2000 in A549 and HeLa cell lines. Modulation of head group polarity significantly affected the transfection efficacy of GAs. The cell viabilities of almost all of the formulations were comparable to those of the standards (DCC/DOPE and DOTAP/DOPE liposomes). Incorporation of cholesterol [GA/DOPE/cholesterol in the ratio of 1:1:1] further improved the serum compatibility of the formulations and improved the transfection efficacy when evaluated in A549 and HeLa cell lines. Fluorescence-assisted cell sorting studies showed comparable number of transfected cells to Lipofectamine 2000 in the HeLa cell line. Intracellular trafficking studies using confocal microscopy indicated transfection of the HeLa cells with the reporter gene within 30 min of lipoplex treatment. γ-Scintigraphy using 99mTc-labeled lipoplexes showed higher concentrations of the lipoplexes in vital tissues like liver, spleen, lungs, and kidneys.

Barriers and Strategies of Cationic Liposomes for Cancer Gene Therapy

“…Fusogenic lipid, DOPE, is a neutral helper lipid that is commonly added to cationic liposomal formulation for promoting membrane fusion-mediated endosomal escape. 95 , 150 , 151 , 152 , 153 , 154 Meanwhile, as mentioned above, lipoplexes could also enter into the cells via membrane fusion-mediated internalization, which favors the phosphoethanolamine (PE) lipid-enriched lipoplexes. 95 However, the amount of DOPE added to the liposome is controversial and most likely related to the structure and σ M of the cationic lipids.…”

Transgene expression in mice of the Opa1 mitochondrial transmembrane protein through bicontinuous cubic lipoplexes containing gemini imidazolium surfactants

“…Intravenous administration of pH-sensitive sugar-based gemini cationic lipids trough the tail vein leaded to a transfection efficiency that was sevenfold higher than the commercially available lipofection agents in lung and heart after 24 h of treatment [ 65 ]. However, the intravenous administration through the tail vein of quaternary gemini cationic lipids-based lipoplexes resulted in their accumulation mainly in liver and spleen for 24 h that precedes a first accumulation in lungs within the first minutes after injection [ 16 ]. Again, the rapid redistribution of lipoplexes from lungs to vital organs for drug elimination such as liver, spleen or kidneys is observed upon an intravenous administration.…”

“…Although lipoplexes have been shown as efficient DNA carriers in living cells [ 7 , 14 ] clinical studies using lipoplexes are still scarce. It is believed that lipoplexes, similarly to liposomes, can be rapidly recognized and removed from the circulation by the reticuloendothelial system (RES) [ 15 ] when administrated in animal models, thus sinking any significant localization in targeted cells [ 16 ]. Since three decades ago, it is known that polyethyleneglycol (PEG)-coated liposomes improve their stability and increase their half-lives in circulation [ 17 ].…”

Multicompartmental Lipopolyplex as Vehicle for Antigens and Genes Delivery in Vaccine Formulations

“…Some of these problems can be circumvented by employing non-viral vehicles such as cationic liposomes or polymers, which form complexes with DNA to favor its transport to the target. These vehicles have been developed and evolved to improve their efficacy of transport and delivery [ 24 , 25 ] and can be combined to synergize their capacities, forming lipopolyplexes. The multicompartmental lipopolyplexes permit the incorporation of antigens with diverse chemical nature.…”