Polymer–Peptide Delivery Platforms: Effect of Oligopeptide Orientation on Polymer-Based DNA Delivery

Abstract

The success of non-viral transfection using polymers hinges on efficient nuclear uptake of nucleic acid cargo and overcoming intra- and extracellular barriers. By incorporating PKKKRKV heptapeptide pendent groups as nuclear localization signals (NLS) on a polymer backbone, we demonstrate protein expression levels higher than obtained from JetPEI™ and Lipofectamine™ 2000, the latter being notorious for coupling high transfection efficiency with cytotoxicity. The orientation of the NLS peptide grafts markedly affected transfection performance. Polymers with the sequence attached to the backbone from the valine residue achieved higher nuclear translocation relative to those having the NLS groups attached in the opposite orientation. The differences in nuclear localization and DNA complexation strength between the two orientations correlated with a striking difference in protein expression, both in cell culture and in vivo. Polyplexes formed from these comb polymer structures exhibited transfection efficiencies superior to those of Lipofectamine 2000 but with greatly reduced toxicity. Moreover, these novel polymers enabled high reporter gene expression in mice when administered by intramuscular ultrasound-mediated delivery, demonstrating their therapeutic promise in vivo.

Recent progress in the science of complex coacervation

“…Both architecture and stiffness effects have been widely explored in the context of charged complexes, such as between DNA and branched polyelectrolytes or dendrimers, as a way to tune the interactions in polyplexes. 133,144,185,186 This understanding has been extended Figure 7c. 34 For both simulation and theory, salt preferentially partitions to the supernatant phase.…”

“…35,[43][44][45]55,75,77,136 Additionally, solid-phase synthesis enables precise control over chemical sequence, 70,[137][138][139][140][141][142][143] and can be combined with methods for controlled polymerization to allow for the preparation of well-controlled comb polymer architectures. 46,144 Controlled polymerization also has allowed for the synthesis of random copolymers to facilitate the introduction of multiple functionalities. 64 Lastly, the expanding breadth of synthetic approaches for preparing charged polymers has allowed for the design of elegant experiments to test specific molecular features, such as hydrophobicity 53 or polarity.…”

Polymer‐mediated gene therapy: Recent advances and merging of delivery techniques

“…This approach indicates that integrin targeting, coupled with optimized vehicle designs with free polymer exclusion, enhances the delivery of therapeutic genes while minimizing potential toxic effects. intramuscular gene delivery (with or without ultrasound mediation) in mice by NLS-and rNLS-based polyplexes (Parelkar et al, 2014) A redox-sensitive, detachable PEG gene delivery system, also described by Kataoka, utilized a P-[Asp(DET)] polymer that promoted significantly higher in vivo gene expression of human TNF-α and antitumor effects in a pancreatic cancer mouse model following administration into the peritoneal cavity of mice (Kumagai et al, 2012;Takae et al, 2008). While this delivery system promised elimination of pancreatic tumors, it lacked targeting modalities to ensure accumulation at target tumors and minimization of any cytotoxic effects.…”

Functional droplets that recognize, collect, and transport debris on surfaces

“…This key component of the extracellular protein fibronectin is now widely implemented in synthetic biomaterials for promoting cell adhesion or improving cell uptake ( 8 ). For drug delivery vehicles, integration of nuclear localization sequences into the delivery system offers the potential to realize nature’s specificity in comparatively simple synthetic polymers, gels, and the like ( 9 , 10 ). More generally, once the fundamental characteristics of any of a variety of biological functions are identified, methods can be designed for reducing biological complexity to scalable materials, minimizing the extent of functionality required to achieve a desired outcome.…”