Novel serum-tolerant lipoplexes target the folate receptor efficiently.
Gene transfer using non-viral vectors is a promising approach for the safe delivery of nucleic acid therapeutics. In this study, we investigate a lipid-based system for targeted gene delivery to malignant cells overexpressing the folate receptor (FR). Cationic liposomes were formulated with and without the targeting ligand folate conjugated to distearoylphosphatidyl ethanolamine polyethylene glycol 2000 (DSPE-PEG2000), the novel cytofectin 3β[N(N(1),N(1)-dimethlaminopropylsuccinamidoethane)-carbamoyl]cholesterol (SGO4), which contains a 13atom, 15Å spacer element, and the helper lipid, dioleoylphosphatidylethanolamine (DOPE). Physicochemical parameters of the liposomes and lipoplexes were obtained by zeta sizing, zeta potential measurement and cryo-TEM. DNA-binding and protection capabilities of liposomes were confirmed by gel retardation assays, EtBr intercalation and nuclease protection assays. The complexes were assessed in an in vitro system for their effect on cell viability using the MTT assay, and gene transfection activity using the luciferase assay in three cell lines; HEK293 (FR-negative), HeLa (FR(+)-positive), KB (FR(++)-positive). Low cytotoxicities were observed in all cell lines, while transgene activity promoted by folate-tagged lipoplexes in FR-positive lines was tenfold greater than that by untargeted constructs and cell entry by folate complexes was demonstrably by FR mediation. These liposome formulations have the design capacity for in vivo application and may therefore be promising candidates for further development.
Non-Viral Gene Delivery Laboratory, Discipline of Biochemistry, Westville Campus, University of KwaZulu-Natal, P Bag X54001, Durban 4000, South Africa.
Sridevi
S
Gorle
Non-Viral Gene Delivery Laboratory, Discipline of Biochemistry, Westville Campus, University of KwaZulu-Natal, P Bag X54001, Durban 4000, South Africa. Electronic address: ariattim@ukzn.ac.za.
Mario
M
Ariatti
Non-Viral Gene Delivery Laboratory, Discipline of Biochemistry, Westville Campus, University of KwaZulu-Natal, P Bag X54001, Durban 4000, South Africa.
Moganavelli
M
Singh
D002470:Cell Survival / Q000187:drug effects; D002784:Cholesterol / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000737:chemistry*; D004247:DNA / Q000008:administration & dosage / Q000737:chemistry*; D005492:Folic Acid / Q000008:administration & dosage / Q000737:chemistry*; D058972:Folic Acid Transporters / Q000378:metabolism; D018014:Gene Transfer Techniques; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D007624:KB Cells; D008081:Liposomes; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000008:administration & dosage / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000008:administration & dosage / Q000737:chemistry*; D044967:Serum
C519184:1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-methoxy-poly(ethylene glycol 2000); D058972:Folic Acid Transporters; D008081:Liposomes; D010714:Phosphatidylethanolamines; D011092:Polyethylene Glycols; C020888:1,2-dielaidoylphosphatidylethanolamine; D004247:DNA; D005492:Folic Acid; D002784:Cholesterol; D008156:Luciferases
Folate receptor; Gene therapy; Lipid nanoparticles; Liposome; Non-viral vectors; Transfection