Bioresponsive hyperbranched polymers for siRNA and miRNA delivery.
This work presents the novel use of reducible hyperbranched (rHB) polymers for delivery of RNA interference (RNAi) therapeutics. Cationic poly(amido amine) hyperbranched polymers that contain different contents of reducible disulfide to nonreducible linkages (0%, 17%, 25%, and 50%) were used to form interpolyelectrolyte polyplexes with siRNA and precursor miRNA (pre-miRNA). Atomic force microscopy (AFM) revealed rHB complexes of ∼100 nm in size, which exhibited redox-activated disassembly in the presence of dithiothreitol (DTT). The complexes were avidly internalized and showed no cellular toxicity in an endogenous enhanced green fluorescence protein (EGFP) expressing H1299 human lung cancer cell line. The highest specific EGFP gene silencing (∼75%) was achieved with rHB (17%)/siRNA complexes at a weight-to-weight (w/w) ratio of 40 that correlated with the ability for this polymer to successfully transfect pre-miRNA. Evaluation of temporal silencing levels over 72 h revealed incremental knockdown that reached a maximum at 72 h for the rHB (50%) complexes, in contrast to maximum knockdown at 24 h that remained relatively consistent, thereafter, for the rHB (17%), rHB (25%), and non-rHB complexes. The role of particle disassembly for intracellular targeting and modulation of gene silencing addressed in this work are important considerations in the development of this and other next-generation delivery systems.
Rahbek
Ulrik L
UL
Interdisciplinary Nanoscience Center (iNANO), University of Aarhus, Aarhus C, Denmark.
Nielsen
Anne F
AF
Dong
Mingdong
M
You
Yezi
Y
Chauchereau
Anne
A
Oupicky
David
D
Besenbacher
Flemming
F
Kjems
Jørgen
J
Howard
Kenneth A
KA
D045744:Cell Line, Tumor; D004573:Electrolytes / Q000737:chemistry; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D020868:Gene Silencing; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics; D035683:MicroRNAs / Q000008:administration & dosage*; D018625:Microscopy, Atomic Force; D010084:Oxidation-Reduction; D011073:Polyamines / Q000737:chemistry; D011108:Polymers / Q000737:chemistry*; D034741:RNA, Small Interfering / Q000008:administration & dosage*; D013997:Time Factors; D014162:Transfection
D004573:Electrolytes; D035683:MicroRNAs; D011073:Polyamines; D011108:Polymers; D034741:RNA, Small Interfering; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins