Novel N,N '-diacyl-1,3-diaminopropyl-2-carbamoyl bivalent cationic lipids for gene delivery--synthesis, in vitro transfection activity, and physicochemical characterization.
Novel N,N'-diacyl-1,3-diaminopropyl-2-carbamoyl bivalent cationic lipids were synthesized and their physicochemical properties in lamellar assemblies with and without plasmid DNA were evaluated to elucidate the structural requirements of these double-chained pH-sensitive surfactants for potent non-viral gene delivery and expression. The highest in vitro transfection efficacies were induced at +/-4:1 by the dimyristoyl, dipalmitoyl and dioleoyl derivatives 1,3lb2, 1,3lb3 and 1,3lb5, respectively, without inclusion of helper lipids. Transfection activities were reduced in the presence of either 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine alone or in combination with cholesterol for all derivatives except 1,3lb5, which maintained reporter gene expression levels at +/-4:1 and yielded increased lipofection activity at a lower charge ratio of +/-2:1. Ethidium bromide displacement indicated efficient plasmid DNA binding and compaction by the transfection-competent analogs. Dynamic light-scattering and electrophoretic mobility studies revealed lipoplexes of the active lipids with large particle sizes (mean diameter>or=500 nm) and zeta potentials with positive values (low ionic strength) or below neutrality (high ionic strength). Langmuir film balance studies showed high in-plane elasticity of these derivatives in isolation. In agreement with the monolayer experiments, fluorescence polarization studies verified the fluid nature of the highly transfection-efficient amphiphiles, with gel-to-liquid crystalline phase transitions below physiological temperature. The active compounds also interacted with endosome-mimicking vesicles to a greater extent than the poorly active derivative 1,3lb4, as revealed by fluorescence resonance energy transfer experiments. Taken together, the results suggest that well-hydrated and highly elastic cationic lipids with increased acyl chain fluidity and minimal cytotoxicity elicit high transfection activity.
Spelios
Michael
M
Division of Pharmaceutical Sciences, Arnold & Marie Schwartz College of Pharmacy & Health Sciences, Long Island University, Brooklyn, NY 11201, USA.
Savva
Michalakis
M
D002412:Cations / Q000737:chemistry; D004123:Dimethylamines / Q000138:chemical synthesis / Q000737:chemistry; D005227:Fatty Acids / Q000138:chemical synthesis / Q000737:chemistry*; D007700:Kinetics; D010316:Particle Size; D010957:Plasmids / Q000737:chemistry; D011412:Propanolamines / Q000737:chemistry; D013501:Surface-Active Agents / Q000737:chemistry; D013696:Temperature; D014162:Transfection
D002412:Cations; D004123:Dimethylamines; D005227:Fatty Acids; D011412:Propanolamines; D013501:Surface-Active Agents