Peptide-Functionalized Poly(ethylene glycol) Star Polymers: DNA Delivery Vehicles with Multivalent Molecular Architecture

Abstract

Exploring the development of nonviral nucleic acid delivery vectors with progressive, specific, and novel designs in molecular architecture is a fundamental way to investigate how aspects of chemical and physical structure impact the transfection process. In this study, macromolecules comprised of a four-arm star poly(ethylene glycol) and termini modified with one of five different heparin binding peptides have been investigated for their ability to bind, compact, and deliver DNA to mammalian cells in vitro. These new delivery vectors combine a PEG-derived stabilizing moiety with peptides that exhibit unique cell-surface binding ability in a molecular architecture that permits multivalent presentation of the cationic peptides. Five peptide sequences of varying heparin binding affinity were studied; each was found to sufficiently bind heparin for biological application. Additionally, the macromolecules were able to bind and compact DNA into particles of proper size for endocytosis. In biological studies, the PEG-star peptides displayed a range of toxicity and transfection efficiency dependent on the peptide identity. The vectors equipped with peptides of highest heparin binding affinity were found to bind DNA tightly, increase levels of cellular internalization, and display the most promising transfection qualities. Our results suggest heparin binding peptides with specific sequences hold more potential than nonspecific cationic polymers to optimize transfection efficiency while maintaining cell viability. Furthermore, the built-in multivalency of these macromolecules may allow simultaneous binding of both DNA at the core of the polyplex and heparan sulfate on the surface of the cell. This scheme may facilitate a bridging transport mechanism, tethering DNA to the surface of the cell and subsequently ushering therapeutic nucleic acids into the cell. This multivalent star shape is therefore a promising architectural feature that may be exploited in the design of future polycationic gene delivery vectors.

Applications of Orthogonal “Click” Chemistries in the Synthesis of Functional Soft Materials

“…The efficiency of this chemistry has also been exploited in non-protein systems, as exemplified by the group of Kiick, who conjugated maleimido-functionalized low molecular weight heparin to star PEGs in order to create non-covalently crosslinked hydrogels, 428 as well as in the preparation of gene delivery vectors based on vinyl sulfone-terminated four-arm PEG conjugated with cysteine-functionalized heparin binding peptides and poly(ethylene imine). 429 Random copolymers of 2-(dimethylamino) ethyl methacrylate (DMAEMA) and aminoethyl methacrylate (AEMA) with the amino groups modified with N -succinimidyl 3-(2-pyridyldithio)propionate (SPDP) have also been conjugated to both transferrin (Tf) or the F(ab') fragment of mAb323/A3 by the formation of reversible disulfide linkages with coupling efficiencies > 90%; similarly high coupling efficiencies 430 have been obtained also for malemido-based strategies. SPDP groups have also been utilized by Maynard in combination with ATRP to prepare poly(hydroxyl ethyl methyl acrylate) (poly(HEMA)) and for conjugation to BSA via formation of reversible disulfide linkages.…”

Z-RAFT star polymerization of styrene: Comprehensive characterization using size-exclusion chromatography

“…In addition, star polymers are becoming increasingly important for life sciences, where they e.g. find applications in the field of drug release [6], serve as unimolecular polymeric micelles [7,8], and are used as nucleic acid delivery vectors [9]. A lot of effort was put into the investigation of properties of stars [10,11] as well as into the development of new methods for their synthesis [12].…”

PEGylation of Melittin: Structural Characterization and Hemostatic Effects

“…There are fewer studies that optimized the PEGylation of peptides for biological studies and characterized the impact of individual modifications through each arm [48]. Our study used a multi-arm PEG to increase the drug payload; thus allowing for the optimization of MLT interactions with biological systems [27].…”

“…PEG has also been utilized to produce nano-objects and tissue regenerative matrices in combination with helical peptides [24] and biological molecules [25]. Bioconjugation with multiple-arm PEG or 'multi-PEGylation' has been mainly used for making hydrogels [26] and gene delivery [27,28]. New PEGs with multiple functionalities at the end [29] and side groups [30] have been studied as a high drug-loading carrier, overcoming the limited payload of linear PEGs.…”