Polymer-based gene delivery with low cytotoxicity by a unique balance of side-chain termini.
Protein expression after delivery of plasmid DNA to the cell nucleus depends on the processes of transcription and translation. Cytotoxic gene-delivery systems may compromise these processes and limit protein expression. This situation is perhaps most prevalent in current nonviral polycationic gene-delivery systems in which the polycationic nature of the delivery system can lead to cytotoxicity. To approach the problem of creating nontoxic but effective gene-delivery systems, we hypothesized that by optimizing the balance between polymer cationic density with endosomal escape moieties, effective gene transfer with low cytotoxicity could be created. As a model system, we synthesized a series of polymers whose side-chain termini varied with respect to the balance of cationic centers and endosomal escape moieties. Specifically, by polymer-analogous amidation we conjugated imidazole groups to the epsilon-amines of polylysine in varying mole ratios (73.5 mol % imidazole, 82.5 mol % imidazole, and 86.5 mol % imidazole). The primary epsilon-amine terminus of polylysine served as a model for the cationic centers, whereas the imidazole groups served as a model for the endosomal escape moieties. These polymers condensed plasmid DNA into nanostructures <150 nm and possessed little cytotoxicity in vitro. Transfection efficiency, as measured by luciferase protein expression, increased with increasing imidazole content of the polymers in a nonlinear relationship. The polymer with the highest imidazole content (86.5 mol %) mediated the highest protein expression, with levels equal to those mediated by polyethylenimine, but with little to no cytotoxicity.
Putnam
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D
Department of Chemical Engineering, E25-342, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
Gentry
C A
CA
Pack
D W
DW
Langer
R
R
D000818:Animals; D001672:Biocompatible Materials; D002460:Cell Line; D002470:Cell Survival; D003587:Cytomegalovirus; D004337:Drug Carriers; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D007093:Imidazoles; D007202:Indicators and Reagents; D008156:Luciferases / Q000032:analysis / Q000235:genetics*; D008264:Macrophages; D051379:Mice; D010957:Plasmids; D011107:Polylysine; D013329:Structure-Activity Relationship; D014162:Transfection / Q000379:methods*
D001672:Biocompatible Materials; D004337:Drug Carriers; D007093:Imidazoles; D007202:Indicators and Reagents; D011107:Polylysine; D008156:Luciferases