Polycation Architecture and Assembly Direct Successful Gene Delivery: Micelleplexes Outperform Polyplexes via Optimal DNA Packaging

Abstract

Cellular delivery of biomacromolecules is vital to medical research and therapeutic development. Cationic polymers are promising and affordable candidate vehicles for these precious payloads. However, the impact of polycation architecture and solution assembly on the biological mechanisms and efficacy of these vehicles has not been clearly defined. In this study, four polymers containing the same cationic poly­(2-(dimethylamino)­ethyl methacrylate) (D) block but placed in different architectures have been synthesized, characterized, and compared for cargo binding and biological performance. The D homopolymer and its diblock copolymer poly­(ethylene glycol)-block-poly­(2-(dimethyl­amino) ethyl methacrylate) (OD) readily encapsulate pDNA to form polyplexes. Two amphiphilic block polymer variants, poly­(2-(dimethylamino)­ethyl methacrylate)-block-poly­(n-butyl methacrylate) (DB) and poly­(ethylene glycol)-block-poly­(2-(dimethyl­amino)­ethyl methacrylate)-block-poly­(n-butyl methacrylate) (ODB), self-assemble into micelles, which template pDNA winding around the cationic corona to form micelleplexes. Micelleplexes were found to have superior delivery efficiency compared to polyplexes and detailed physicochemical and biological characterizations were performed to pinpoint the mechanisms by testing hypotheses related to cellular internalization, intracellular trafficking, and pDNA unpackaging. For the first time, we find that the higher concentration of amines housed in micelleplexes stimulates both cellular internalization and potential endosomal escape, and the physical motif of pDNA winding into micelleplexes, reminiscent of DNA compaction by histones in chromatin, preserves the pDNA secondary structure in its native B form. This likely allows greater payload accessibility for protein expression with micelleplexes compared to polyplexes, which tightly condense pDNA and significantly distort its helicity. This work provides important guidance for the design of successful biomolecular delivery systems via optimizing the physicochemical properties.

On Complex Coacervate Core Micelles: Structure-Function Perspectives

“…Whereas 70% of HEK293T cells were successfully transfected, when subjected to micelleplexes with pDNA encoding for green fluorescence protein (GFP), only 40% and 10% of HEK293T were transfected when instead jetPEI and conventional polyplexes were utilized. This dramatic improvement in transfection efficiency is presumably due to the preservation of the native conformation of pDNA within micelleplexes, which are essentially aggregates of pDNA and positively charged micelles [ 87 ]. The pDNA configuration in micelleplexes appears analogous to the DNA compaction by histones.…”

pH‐Sensitive Polycations for siRNA Delivery: Effect of Asymmetric Structures of Tertiary Amine Groups

“…The reason for this phenomenon was that the form of micelleplexes formed by polycationic micelles/gene could cause higher gene silencing efficiency than PICs. [ 16 ] To verify the phenomenon, Circular dichroism (CD) was carried out. As shown in Figure S2, Supporting Information, the PEI showed greater influence on the siRNA secondary structure and a red shift in the signal was observed.…”

“…[ 9,10 ] Non‐viral vehicles, as the viral vehicles succedaneum, have the advantages of low immunogenicity, low toxicity, and low cost. [ 11–13 ] Among the non‐viral vehicles, the polycations have become the outshining tactics for gene delivery due to their controlled preparation and flexible modification, [ 14 ] such as poly(2‐dimethylaminoethyl methacrylate) (PDMAEMA), [ 15,16 ] polyethylenimine (PEI), [ 17 ] poly(L‐lysine) (PLL), [ 18,19 ] and polyamidoamine dendrimer (PAMAM), [ 20 ] which could condense gene through charge interaction to form nanosized polyion complex (PIC). Such kind of vectors with abundant amine had presented excellent siRNA binding capacity and delivery efficiency, but the additional charge caused low biocompatibility and high biotoxicity.…”

Understanding the Interaction of Polyelectrolyte Architectures with Proteins and Biosystems

“…Here, cationic polyelectrolytes are used to compact DNA and RNA by formation of so‐called polyplexes [20] . The micelles and aggregates formed by this interaction may then form more complex supramolecular structures [174] . Polyethyleneimine (PEI) has been the cationic polyelectrolyte of choice [175, 176] .…”