Targeting the Blind Spot of Polycationic Nanocarrier-Based siRNA Delivery

Abstract

Polycationic nanocarriers attract increasing attention to the field of siRNA delivery. We investigated the self-assembly of siRNA vs pDNA with polycations, which are broadly used for nonviral gene and siRNA delivery. Although polyethyleneimine (PEI) was routinely adopted as siRNA carrier based on its efficacy in delivering pDNA, it has not been investigated yet why PEI efficiently delivers pDNA to cells but is controversially discussed in terms of efficacy for siRNA delivery. We are the first to investigate the self-assembly of PEI/siRNA vs PEI/pDNA and the steps of complexation and aggregation through different levels of hierarchy on the atomic and molecular scale with the novel synergistic use of molecular modeling, molecular dynamics simulation, isothermal titration calorimetry, and other characterization techniques. We are also the fist to elucidate atomic interactions, size, shape, stoichiometry, and association dynamics for polyplexes containing siRNA vs pDNA. Our investigation highlights differences in the hierarchical mechanism of formation of related polycation–siRNA and polycation–pDNA complexes. The results of fluorescence quenching assays indicated a biphasic behavior of siRNA binding with polycations where molecular reorganization of the siRNA within the polycations occurred at lower N/P ratios (nitrogen/phosphorus). Our results, for the first time, emphasize a biphasic behavior in siRNA complexation and the importance of low N/P ratios, which allow for excellent siRNA delivery efficiency. Our investigation highlights the formulation of siRNA complexes from a thermodynamic point of view and opens new perspectives to advance the rational design of new siRNA delivery systems.

Mixing-sequence-dependent nucleic acid complexation and gene transfer efficiency by polyethylenimine

“…1B and 2B). 29 This can be explained by the fact that a significant conformational change is required for plasmid DNA to bind to cationic polymers, while small siRNA's binding to cationic polymers do not require such a change. However, a sum of electrostatically attractive forces exerted on one plasmid DNA molecule is greater than those on some siRNA molecules, generating higher molecular shielding upon complete complexation ( Fig.…”

Systematic coarse-grained modeling of complexation between small interfering RNA and polycations

“…3(b)). The absence of siRNA bending (as compared to complexation with hyperbranched PEI 11 ) is easily understood from the very flexible nature of linear PEI backbone, which enables it to adapt its conformations during the binding process. As the PEG grafts tend to be located away from the siRNA, their presence does not have a strong effect on the siRNA shape.…”

“…[8][9][10] Protective carriers are needed to deliver siRNA to the target site to avoid degradation by nucleases, to facilitate cellular uptake, and so on. PEI appears to be a promising carrier, as it offers pH-buffering properties, 6 high affinity to siRNA, 11 high genedelivery efficiency, 12,13 and the advantage of being easily modified with other functional groups. 14 In spite of this great potential, there exist several challenges that prevent the clinical use of such gene-delivery systems.…”

“…11 Sun et al performed AA simulations involving multiple siRNA molecules to study the effect of lipid substitution on PEI-mediated siRNA complexation.…”

Impact of molecular weight and degree of conjugation on the thermodynamics of DNA complexation and stability of polyethylenimine-graft-poly(ethylene glycol) copolymers

“…Recently, isothermal titration calorimetry (ITC) has been used to determine the thermodynamics of the DNA or siRNA interactions of PEI, poly-L-lysine, poly(2-(dimethylamino) ethyl methacrylate), poly(2-(diethylamino)ethyl methacrylate), poly (glycoamidoamine) and chitosan [43][44][45][46][47][48][49][50][51][52][53][54]. The difficulty with application of ITC is that not only binding heat is detected.…”