Optimization of poly(amido amine)s as vectors for siRNA delivery.
By Michael addition polymerization of N,N'-cystaminebisacrylamide (CBA) with variable ratios of 4-amino-1-butanol (ABOL) and ethylene diamine (EDA) or triethylenetetramine (TETA), poly(amido amine) copolymers could be obtained with tunable charge densities. The copolymers were optimized to serve as nonviral vectors in RNA interference (RNAi) to form stable, nanosized polyplexes with siRNA with maximum transfection efficacy. It was observed that at least 20-30% EDA or TETA amino units in the copolymers is necessary to encapsulate siRNA into small and stable polyplexes (< 200 nm). Incorporation of higher amounts of EDA or TETA in the copolymers did not further improve polyplex formation and stability, but the increased cationic charge in these copolymers resulted in increased cytotoxicity and hemolytic activity. Copolymers with 20% EDA showed excellent gene silencing properties in vitro (70% luciferase knockdown in H1299 cells) with negligible cytotoxicity.
Department of Biomedical Chemistry, Faculty of Science and Technology, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, PO Box 217, 7500 AE Enschede, The Netherlands.
L J
LJ
van der Aa
P
P
Vader
G
G
Storm
R M
RM
Schiffelers
J F J
JF
Engbersen
D000178:Acrylamides / Q000737:chemistry; D000605:Amino Alcohols / Q000737:chemistry; D001682:Biological Availability; D002021:Buffers; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004229:Dithiothreitol / Q000737:chemistry; D004587:Electrophoresis, Agar Gel; D005029:Ethylenediamines / Q000737:chemistry; D015870:Gene Expression / Q000235:genetics; D018014:Gene Transfer Techniques; D005978:Glutathione / Q000737:chemistry; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D049409:Luciferases, Firefly / Q000235:genetics / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008970:Molecular Weight; D009690:Nucleic Acid Conformation; D009757:Nylons / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D010316:Particle Size; D010957:Plasmids / Q000008:administration & dosage / Q000737:chemistry; D011073:Polyamines / Q000138:chemical synthesis / Q000737:chemistry* / Q000494:pharmacology; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000737:chemistry / Q000493:pharmacokinetics; D055672:Static Electricity; D014162:Transfection; D014266:Trientine / Q000737:chemistry
D000178:Acrylamides; D000605:Amino Alcohols; D002021:Buffers; D005029:Ethylenediamines; D009757:Nylons; D011073:Polyamines; D034741:RNA, Small Interfering; C089584:4-aminobutanol; D049452:Green Fluorescent Proteins; C031234:ethylenediamine; D049409:Luciferases, Firefly; D005978:Glutathione; D014266:Trientine; D004229:Dithiothreitol