Asymmetrical N4,N9-diacyl spermines: SAR studies of nonviral lipopolyamine vectors for efficient siRNA delivery with silencing of EGFP reporter gene.
Our aim is to study the effects of varying the two acyl moieties in synthesized N(4),N(9)-diacyl spermines on siRNA formulations and their delivery efficiency in cell lines. Six novel asymmetrical lipopolyamines, [N(4)-cholesteryloxy-3-carbonyl-N(9)-oleoyl-, N(4)-decanoyl-N(9)-oleoyl-, N(4)-decanoyl-N(9)-stearoyl-, N(4)-lithocholoyl-N(9)-oleoyl-, N(4)-myristoleoyl-N(9)-myristoyl-, and N(4)-oleoyl-N(9)-stearoyl]-1,12-diamino-4,9-diazadodecane, were assessed for their abilities to bind to siRNA, studied using a RiboGreen intercalation assay, and to form nanoparticles. Their siRNA delivery efficiencies were quantified in FEK4 primary skin cells and in an immortalized cancer cell line (HtTA) using a fluorescein-tagged siRNA, and compared with formulations of N(4),N(9)-dioleoyl-1,12-diamino-4,9-diazadodecane and of a leading transfecting agent, TransIT-TKO. Transfection was measured in terms of siRNA delivery and silencing of EGFP reporter gene in HeLa cells. By incorporating two different acyl moieties, changing their length and oxidation level in a controlled manner, we show efficient fluorescein-tagged siRNA formulation, delivery, and knock-down of EGFP reporter gene. N(4)-Oleoyl-N(9)-stearoyl spermine and N(4)-myristoleoyl-N(9)-myristoyl spermine are effective siRNA delivery vectors typically resulting in 89% cell delivery and gene silencing to 34% in the presence of serum, comparable with the results obtained with TransIT-TKO; adding a second lipid chain is better than incorporating a steroid moiety.
Department of Pharmacy and Pharmacology, University of Bath , Bath BA2 7AY, U.K.
Ian S
IS
Blagbrough
Abdelkader A
AA
Metwally
Hassan M
HM
Ghonaim
D002460:Cell Line; D045744:Cell Line, Tumor; D002626:Chemistry, Pharmaceutical / Q000379:methods; D020868:Gene Silencing / Q000187:drug effects; D018014:Gene Transfer Techniques; D017930:Genes, Reporter / Q000235:genetics*; D018773:Genes, erbB-1 / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D053758:Nanoparticles / Q000008:administration & dosage; D010316:Particle Size; D011073:Polyamines / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics*; D013096:Spermine / Q000494:pharmacology*; D014162:Transfection
D011073:Polyamines; D034741:RNA, Small Interfering; D013096:Spermine