Document Detail


Formation of plasmid-based transfection complexes with an acid-labile cationic lipid: characterization of in vitro and in vivo gene transfer.
MedLine Citation:
PMID:  12403065     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
PURPOSE: This study tests the hypothesis that gene transfer efficiency may be improved through the use of transiently stable transfection complexes that degrade within endosomal compartments and promote plasmid escape into the cytosol. METHOD: An acid labile cationic lipid, O-(2R-1,2-di-O-(1'Z, 9'Z-octadecadienyl)-glycerol)-3-N-(bis-2-aminoethyl)-carbamate (BCAT), was designed, synthesized, and tested for enhanced gene transfer activity relative to non-labile controls. RESULTS: The O-alkenyl chains of BCAT were completely hydrolyzed after 4 h incubation in pH 4.5 buffer at 25 degrees C. Addition of BCAT to plasmid DNA in 40% ethanol followed by ethanol evaporation yielded transfection complexes that transfected several cell types in the presence of fetal calf serum and without the need of a helper lipid. Transfection complexes prepared from BCAT displayed higher luciferase expression than the corresponding DCAT complexes (an acid-insensitive derivative of BCAT) for all cell types tested. Uptake studies showed that this increase was not due to a difference in the amount of DNA being delivered. FAGS analysis for GFP expression showed that BCAT transfection complexes yielded 1.6 more transfected cells and 20% higher log mean fluorescence than DCAT transfection complexes. In vivo gene transfer was demonstrated in subcutaneous tumor-bearing mice by systemic administration of a 60 microg plasmid dose. Expression was observed in the lungs and in the tumor, with the highest activity being observed in the lungs. CONCLUSIONS: Our results show that increased transfection can be obtained by coupling the cationic headgroup to the hydrophobic amphiphilic tails via acid-labile bonds. Acid-catalyzed release of the alkyl chains should facilitate dissociation of the cationic lipid headgroup from the plasmid, thus accelerating one of the rate-limiting steps in cationic lipid mediated transfection.
Authors:
Jeremy A Boomer; David H Thompson; Sean M Sullivan
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.    
Journal Detail:
Title:  Pharmaceutical research     Volume:  19     ISSN:  0724-8741     ISO Abbreviation:  Pharm. Res.     Publication Date:  2002 Sep 
Date Detail:
Created Date:  2002-10-29     Completed Date:  2003-04-09     Revised Date:  2007-11-14    
Medline Journal Info:
Nlm Unique ID:  8406521     Medline TA:  Pharm Res     Country:  United States    
Other Details:
Languages:  eng     Pagination:  1292-301     Citation Subset:  IM    
Affiliation:
Department of Chemistry, Purdue University, West Lafayette, Indiana 47907-1393, USA.
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MeSH Terms
Descriptor/Qualifier:
3T3 Cells
Acids
Animals
Cations / chemistry,  pharmacology
Cell Line
Chemistry, Pharmaceutical
Gene Transfer Techniques
Humans
Lipids / chemistry,  pharmacology
Mice
Mice, Inbred C3H
Plasmids / chemistry*,  genetics,  pharmacology*
Transfection / methods*
Grant Support
ID/Acronym/Agency:
GM55266/GM/NIGMS NIH HHS
Chemical
Reg. No./Substance:
0/Acids; 0/Cations; 0/Lipids

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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