Polyelectrolyte Complexation of Oligonucleotides by Charged Hydrophobic—Neutral Hydrophilic Block Copolymers

Abstract

Polyelectrolyte complex micelles (PCMs, core-shell nanoparticles formed by complexation of a polyelectrolyte with a polyelectrolyte-hydrophilic neutral block copolymer) offer a solution to the critical problem of delivering therapeutic nucleic acids, Despite this, few systematic studies have been conducted on how parameters such as polycation charge density, hydrophobicity, and choice of charged group influence PCM properties, despite evidence that these strongly influence the complexation behavior of polyelectrolyte homopolymers. In this article, we report a comparison of oligonucleotide PCMs and polyelectrolyte complexes formed by poly(lysine) and poly((vinylbenzyl) trimethylammonium) (PVBTMA), a styrenic polycation with comparatively higher charge density, increased hydrophobicity, and a permanent positive charge. All of these differences have been individually suggested to provide increased complex stability, but we find that PVBTMA in fact complexes oligonucleotides more weakly than does poly(lysine), as measured by stability versus added salt. Using small angle X-ray scattering and electron microscopy, we find that PCMs formed from both cationic blocks exhibit very similar structure-property relationships, with PCM radius determined by the cationic block size and shape controlled by the hybridization state of the oligonucleotides. These observations narrow the design space for optimizing therapeutic PCMs and provide new insights into the rich polymer physics of polyelectrolyte self-assembly.

Assembly and Characterization of Polyelectrolyte Complex Micelles

“…This allows for control of PCM size by choice of block polymer. The data shown here is for pLys-PEG with 88 nt/bp DNA and has been previously reported 26 . , scattering is dominated by the individual polymers inside the PCM core, fit here by a power law (green) with low q cutoff.…”

“…Recent work by our group and others in the field has begun to address this problem by developing structure-property, and in some cases structure-property-function relationships for PCMs formed from nucleic acids and various cationic-neutral polymers 7,[25][26][27] . Two consistent themes that have emerged from these studies are the importance of developing well-controlled, repeatable protocols for PCM assembly and the benefit of using multiple techniques to characterize the resulting nanoparticles.…”

“…The polyelectrolytes are mixed in high salt conditions where electrostatic attractions are screened, then the salt concentration is slowly lowered to allow the polyelectrolytes to settle into their most energetically favorable configurations, analogous to the slow cooling process of thermal annealing. Using this protocol, we are regularly able to achieve exceptionally low polydispersity and high repeatability for oligonucleotide PCMs 7,26 . Finally, we describe how four separate measurement techniques can be used to characterize PCMs over a very wide range of length scales, from external morphology to internal structure: DLS, multi-angle light scattering (MALS), small angle X-ray scattering (SAXS), and transmission electron microscopy (TEM).…”

“…generated in a high throughput manner 28 . The choice of charged group strongly affects the stability of ion pairing and shape of micelles 26 , and PCM size has been shown to increase with the length of the charged block 5,7,26 (Figure 2), thus allowing PCM properties to be tuned for the requirements of a desired application. For linear diblocks, we have found that the charged block should have at least 10 charges and be strongly charged at the desired pH.…”

“…When using oligonucleotides as a polyanion, the hybridization state and chemical structure help determine the propensity for self-assembly and the characteristics of the resulting PCM 5,7,26 . Optimizing these, within the requirements for biological efficacy (if using PCMs for delivery) will increase the likelihood of forming the desired structures.…”