title: Low-pH-sensitive poly(ethylene glycol) (PEG)-stabilized plasmid nanolipoparticles: effects of PEG chain length, lipid composition and assembly conditions on gene delivery.
abstract: BACKGROUND
  We have studied the effects of the poly(ethylene glycol) (PEG) chain length and acyl chain composition on the pH-sensitivity of acid-labile PEG-diorthoester (POD) lipids. The optimal conditions are described for preparation of DNA plasmid encapsulated POD nanolipoparticles (NLPs) which mediate high gene delivery activity in vitro with moderate cytotoxicity.
  
  
  RESULTS
  A series of POD lipids with various PEG chain lengths (750, 2000, and 5000 Da) or acyl chains (distearoyl 18:0 or dioleoyl 18:1) were incorporated into DNA containing NLPs or model liposomes as a stealth and bioresponsive component. We investigated the collapse kinetics of the POD-stabilized liposomes when the PEG chain length was changed. We optimized a detergent dialysis method to encapsulate plasmid DNA into NLPs prepared from a mixture of the various POD lipids, cationic lipid and phosphatidylethanolamine lipid. A critical concentration (28 mM) of n-octyl-beta-D-glucopyranoside (OG) enabled high encapsulation of DNA plasmid into 100 nm particles with a neutral surface charge. The POD NLPs are stable at pH 8.5 but rapidly collapse (approximately 10 min) into aggregates at pH 5.0. In the detergent solution there is a metastable DNA-lipid intermediate that evolves into a stable NLP if the detergent is removed shortly after adding DNA to the lipid-detergent mixture. The rank order of transfection activity from NLPs containing PEG-lipid was POD 750 > POD 5000 = POD 2000 > non-pH-sensitive PEG-lipid. The particle size stability was in the reverse order. Binding of the NLPs to cells reached a maximum level by 12 hours. The POD NLPs had slightly less transfection activity but considerably lower cytotoxicity than the PEI-DNA polyplex.
  
  
  CONCLUSIONS
  Of the PEG-orthoester lipids tested, POD 2000 is the better choice for the preparation of sterically stabilized NLPs with a small particle diameter, good stability, low cytotoxicity, and satisfactory transfection activity.
authors.affiliation: Department of Biopharmaceutical Sciences and Pharmaceutical Chemistry, University of California at San Francisco, San Francisco, CA 94143-0446, USA.
authors.forename: Weijun
authors.initials: W
authors.lastname: Li
authors.forename: Zhaohua
authors.initials: Z
authors.lastname: Huang
authors.forename: J Andrew
authors.initials: JA
authors.lastname: MacKay
authors.forename: Stefan
authors.initials: S
authors.lastname: Grube
authors.forename: Francis C
authors.initials: FC
authors.lastname: Szoka
mesh_terms: D000818:Animals; D002460:Cell Line; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D004305:Dose-Response Relationship, Drug; D005347:Fibroblasts / Q000187:drug effects; D018014:Gene Transfer Techniques; D005909:Glioblastoma / Q000188:drug therapy; D006801:Humans; D006863:Hydrogen-Ion Concentration; D020128:Inhibitory Concentration 50; D008055:Lipids / Q000737:chemistry; D008081:Liposomes / Q000737:chemistry; D008546:Melanoma, Experimental / Q000188:drug therapy; D051379:Mice; D049329:Nanostructures; D010957:Plasmids; D011092:Polyethylene Glycols / Q000138:chemical synthesis / Q000737:chemistry* / Q000633:toxicity; D013997:Time Factors
chemical_list: D008055:Lipids; D008081:Liposomes; D011092:Polyethylene Glycols