Delivery of siRNA mediated by histidine-containing reducible polycations.
Histidine containing reducible polycations based on CH(6)K(3)H(6)C monomers (His6 RPCs), are highly effective DNA transfection agents combining pH buffering endosomal escape mechanisms with rapid unpackaging following reduction in the cytoplasm. We examined their ability to mediate siRNA uptake into cells focusing on hepatocyte delivery. Co-delivery of EGFP siRNA with pEGFP plasmid DNA reduced reporter gene expression by 85%. However while DNA transfection efficiency increased with polymer size, with 162 k His6 RPCs proving the most effective, delivery of siRNA alone to EGFP stably expressing cells was only possible using 36-80 k polymers. Analysis of particle sizes showed that 80 k polymers formed more compact siRNA complexes than 162 k polymers. The reducible nature of the polymer was necessary for siRNA activity, since siRNA combined with non-reducible polylysine showed little activity. Incorporation of a targeting peptide from the Plasmodium falciparum circumsporozoite (CS) protein onto His6 RPCs, significantly improved transfection of plasmid DNA and siRNA activity in hepatocytes, but not in most non-liver cells tested. siRNA targeted to the hepatitis B virus surface antigen delivered by CS-His6 RPC, mediated falls in both mRNA and protein expression, suggesting that this delivery system could be developed for potential therapies for viral hepatitis.
Stevenson
Mark
M
Department of Clinical Pharmacology, University of Oxford, Old Road Campus, Oxford OX3 7DQ, UK. mark.stevenson@clinpharm.ox.ac.uk
Ramos-Perez
Victor
V
Singh
Surjeet
S
Soliman
Mahmoud
M
Preece
Jon A
JA
Briggs
Simon S
SS
Read
Martin L
ML
Seymour
Leonard W
LW
D000818:Animals; D019556:COS Cells; D045744:Cell Line, Tumor; D002522:Chlorocebus aethiops; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000008:administration & dosage* / Q000235:genetics; D004337:Drug Carriers / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics; D006514:Hepatitis B Surface Antigens / Q000235:genetics; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006639:Histidine / Q000737:chemistry*; D006801:Humans; D008156:Luciferases / Q000235:genetics; D051379:Mice; D008970:Molecular Weight; D010963:Plasmodium falciparum; D011073:Polyamines / Q000737:chemistry*; D000071228:Polyelectrolytes; D015800:Protozoan Proteins / Q000737:chemistry; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D004247:DNA; D004337:Drug Carriers; D049452:Green Fluorescent Proteins; D006514:Hepatitis B Surface Antigens; D006639:Histidine; D008156:Luciferases; D011073:Polyamines; D000071228:Polyelectrolytes; D015800:Protozoan Proteins; D034741:RNA, Small Interfering; C041595:circumsporozoite protein, Protozoan