Dataset Information

Modulating Polymer-siRNA Binding Does Not Promote Polyplex-Mediated Silencing.

ABSTRACT: The development of delivery vehicles for small interfering RNAs (siRNAs) remains a bottleneck to widespread clinical use. Cationic polymers represent an important class of potential delivery vehicles. In this study, we used alkyne-azide click chemistry to synthesize a variety of cationic poly(propargyl glycolide) backbone polymers to bind and deliver siRNAs. We demonstrated control over the binding interactions of these polymers and siRNAs by varying binding strength by more than three orders of magnitude. Binding strength was found to meet or exceed that of commercially available transfection agents. Our polymers effectively delivered siRNAs with no detectable cytotoxicity. Despite accumulation of siRNAs at levels comparable with commercial reagents, we did not observe silencing of the targeted protein. The implications of our results for future siRNA delivery vehicle design are discussed.

SUBMITTER: Splichal RC

PROVIDER: S-EPMC8215420 | biostudies-literature | 2021 Jun

REPOSITORIES: biostudies-literature

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Modulating Polymer-siRNA Binding Does Not Promote Polyplex-Mediated Silencing.

Splichal R Chauncey RC Gredell Joseph A JA Vogel Erin B EB Malefyt Amanda A Comiskey Georgina G Smith Milton R MR Chan Christina C Walton S Patrick SP

Nucleic acid therapeutics 20200729 3

The development of delivery vehicles for small interfering RNAs (siRNAs) remains a bottleneck to widespread clinical use. Cationic polymers represent an important class of potential delivery vehicles. In this study, we used alkyne-azide click chemistry to synthesize a variety of cationic poly(propargyl glycolide) backbone polymers to bind and deliver siRNAs. We demonstrated control over the binding interactions of these polymers and siRNAs by varying binding strength by more than three orders of ...[more]

PMID: 32749923

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Project description:When nanoparticles interact with their environment, the nature of that interaction is governed largely by the properties of its outermost surface layer. Here, we exploit the exceptional properties of a common disaccharide, trehalose, which is well-known for its unique biological stabilization effects. To this end, we have developed a synthetic procedure that readily affords a polymer of this disaccharide, poly(methacrylamidotrehalose) or "poly(trehalose)" and diblock copolycations containing this polymer with 51 repeat units chain extended with aminoethylmethacrylamide (AEMA) at three degrees of polymerization (n = 34, 65, and 84). Two series of experiments were conducted to study these diblock copolymers in detail and to compare their properties to two control polymers [PEG-P(AEMA) and P(AEMA)]. First, we demonstrate that the poly(trehalose) coating ensures colloidal stability of polyplexes containing siRNA in the presence of high salt concentrations and serum proteins. Poly(trehalose) retains the ability of trehalose to lower the phase transition energy associated with water freezing and can protect siRNA polyplexes during freeze-drying, allowing complete nanoparticle resuspension without loss of biological function. Second, we show that siRNA polyplexes coated with poly(trehalose) have exceptional cellular internalization into glioblastoma cells that proceeds with zero-order kinetics. Moreover, the amount of siRNA delivered by poly(trehalose) block copolycations can be controlled by the siRNA concentration in cell culture media. Using confocal microscopy we show that trehalose-coated polyplexes undergo active trafficking in cytoplasm upon internalization and significant siRNA-induced target gene down-regulation was achieved with an IC50 of 19 nM. These findings coupled with a negligible cytotoxicity suggests that poly(trehalose) has the potential to serve as an important component of therapeutic nanoparticle formulations of nucleic acids and has great promise to be extended as a new coating for other nanobased technologies and macromolecules, in particular, those related to nanomedicine applications.

Project description:Formation of stable, long-circulating siRNA polyplexes is a significant challenge in translation of intravenously-delivered, polymeric RNAi cancer therapies. Here, we report that siRNA hydrophobization through conjugation to palmitic acid (siPA) improves stability, in vivo pharmacokinetics, and tumor gene silencing of PEGylated nanopolyplexes (siPA-NPs) with balanced cationic and hydrophobic content in the core relative to the analogous polyplexes formed with unmodified siRNA, si-NPs. Hydrophobized siPA loaded into the NPs at a lower charge ratio (N(+):P(-)) relative to unmodified siRNA, and siPA-NPs had superior resistance to siRNA cargo unpackaging in comparison to si-NPs upon exposure to the competing polyanion heparin and serum. In vitro, siPA-NPs increased uptake in MDA-MB-231 breast cancer cells (100% positive cells vs. 60% positive cells) but exhibited equivalent silencing of the model gene luciferase relative to si-NPs. In vivo in a murine model, the circulation half-life of intravenously-injected siPA-NPs was double that of si-NPs, resulting in a >2-fold increase in siRNA biodistribution to orthotopic MDA-MB-231 mammary tumors. The increased circulation half-life of siPA-NPs was dependent upon the hydrophobic interactions of the siRNA and the NP core component and not just siRNA hydrophobization, as siPA did not contribute to improved circulation time relative to unmodified siRNA when delivered using polyplexes with a fully cationic core. Intravenous delivery of siPA-NPs also achieved significant silencing of the model gene luciferase in vivo (∼40% at 24 h after one treatment and ∼60% at 48 h after two treatments) in the murine MDA-MB-231 tumor model, while si-NPs only produced a significant silencing effect after two treatments. These data suggest that stabilization of PEGylated siRNA polyplexes through a combination of hydrophobic and electrostatic interactions between siRNA cargo and the polymeric carrier improves in vivo pharmacokinetics and tumor gene silencing relative to conventional formulations that are stabilized solely by electrostatic interactions.

Dataset Information

Modulating Polymer-siRNA Binding Does Not Promote Polyplex-Mediated Silencing.

Publications

Modulating Polymer-siRNA Binding Does Not Promote Polyplex-Mediated Silencing.

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