Abstract
The administration of gene-editing tools has been proposed as a promising therapeutic approach for correcting mutations that cause diseases. Gene-editing tools, composed of relatively large plasmid DNA constructs that often need to be co-delivered with a guiding protein, are unable to spontaneously penetrate mammalian cells. Although viral vectors facilitate DNA delivery, they are restricted by the size of the plasmid to carry. In this work, we describe a strategy for the stable encapsulation of the gene-editing tool piggyBac transposon into Poly (β-amino ester) nanoparticles (NPs). We propose a non-covalent and a covalent strategy for stabilization of the nanoformulation to slow down release kinetics and enhance intracellular delivery. We found that the formulation prepared by covalently crosslinking Poly (β-amino ester) NPs are capable to translocate into the cytoplasm and nuclei of human glioblastoma (U87MG) cells within 1 h of co-culturing, without the need of a targeting moiety. Once internalized, the nanoformulation dissociates, delivering the plasmid presumably as a response to the intracellular acidic pH. Transfection efficiency is confirmed by green fluorescence protein (GFP) expression in U87MG cells. Covalently stabilized Poly (β-amino ester) NPs are able to transfect ~55% of cells causing non-cytotoxic effects. The strategy described in this work may serve for the efficient non-viral delivery of other gene-editing tools.
Highlights
In recent years gene-editing therapies have emerged as a promising avenue for the treatment of various diseases ranging from genetic disorders [1] and infections [2,3,4] to cancer [5,6,7]
We study the fabrication of nanoparticles (NPs) composed of the PBAE copolymer poly–block–poly(1,4-butanediol)–diacrylate-β,hydroxyamylamine
PEG-PDHA NPs display a relatively small and narrow size distribution, and the PEG chains are at high density forming a thick shell layer on the NPs surface (Figure 1), we found that PBCAG releases very fast from these NPs, hindering gene delivery
Summary
In recent years gene-editing therapies have emerged as a promising avenue for the treatment of various diseases ranging from genetic disorders [1] and infections [2,3,4] to cancer [5,6,7]. For cancer treatment, gene-editing therapy works by inhibiting or editing the expression of the gene of interest. The most common delivery systems utilized in gene therapy are viral vectors, which use the virus capsid as a means to introduce genetic materials. Viral vectors are very effective in gene delivery as they are built upon the natural ability of viruses to infect cells while being modified to be used in therapy [19,20,21].
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