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. 2024 Oct 15;14(44):32637-32647.
doi: 10.1039/d4ra05978f. eCollection 2024 Oct 9.

Furan-rich, biobased transfection agents as potential oligomeric candidates for intracellular plasmid DNA delivery

Ashique Al Hoque  1   2 , Prakash Kannaboina  3 , Yeabstega Abraham  4 , Masfique Mehedi  4 , Mukund P Sibi  3 , Mohiuddin Quadir  1
Affiliations

Furan-rich, biobased transfection agents as potential oligomeric candidates for intracellular plasmid DNA delivery

Ashique Al Hoque et al. RSC Adv. .

Abstract

Biobased, DNA delivery vectors have been synthesized with a core motif composed of 2,5-bishydroxymethylfuran (BHMF) readily available from an important biomass feedstock 5-hydroxymethyl furfural (HMF). To generate the product, BHMF was first converted to 2,5-furan bishydroxymethyl diacrylate (2,5-FDA), which was later conjugated with different types of secondary amines. Rich in tertiary nitrogen, these oligomeric FDA-amino esters demonstrated stable electrostatic interactions with negatively charged plasmid DNA in an aqueous environment. We evaluated synthetic routes toward these plasmid DNA-binding amino esters (pFASTs), identified their nanoscale features, and attempted to establish their structure-property relationship in the context of the DNA delivery. Our preliminary studies show that the pFASTs formed stable complexes with the plasmid DNA. Dynamic light scattering indicated that the DNA polyplexes of pFASTs have hydrodynamic diameters within the size range of 100-150 nm with a surface charge (ζ-potential) ranging from -10 to +33 mV, depending on pFAST type. These oligomeric amino esters rich in furan motif were also found to successfully transfect the GFP-expressing plasmid DNA intracellularly. Collectively, this study establishes a new route to produce DNA transfection agents from sustainable resources that can be used for transferring genetic materials for humans, veterinary, and agrochemical purposes.

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Conflict of interest statement

Authors declare no conflict of interest.

Figures

Scheme 1
Scheme 1. Synthetic pathway to pFAST-1, 2 and 3via Michael addition of secondary amines to 2,5-FDA. These products were purified by repeated precipitation from diethyl ether.
Fig. 1
Fig. 1. (A) PicoGreen assay indicating the extent of DNA complexation with pFAST systems where PEG-PLK was used as a control (B) DNA binding assay through agarose gel retardation. (B1) DNA–pFASTs at 1 : 10 ratio where wells 1–4 represent free DNA, and DNA complexed with pFAST-1, pFAST-2, and pFAST-3 respectively, (B2) DNA–pFASTs at 1 : 50 ratio, and (B3) DNA : pFASTs at 1 : 100 ratio where wells 1–4 represents free DNA, pFAST-1, pFAST-2, and pFAST-3, respectively.
Fig. 2
Fig. 2. (A) Particle size distribution of pBR322 DNA/pFAST polyplexes prepared with pFAST-1–3 at a DNA : pFAST ratio of 1 : 50. (B) Effect of complexation ratio of gWiz-GFP DNA/pFAST on the hydrodynamic diameters of polyplexes as a function of pFAST type.
Fig. 3
Fig. 3. Transmission electron microscopy of DNA/pFAST polyplexes, (A) pFAST-1, (B) pFAST-2 and (C) pFAST-3.
Fig. 4
Fig. 4. Cytotoxicity profiles of pFAST 1–3 compared to PEI and PEG-PLK in two different cell lines: (A) HFF2 human fibroblasts, and (B) KRAS-mutant MIA PaCa-2 cells.
Fig. 5
Fig. 5. Plasmid DNA delivery in human lung airway epithelial A549 cells using nanoparticle formulations. gWiz-GFP plasmid was delivered to A549 cells using two nanoparticle formulations: (A1) (shown in the top panel) gWiz-GFP DNA/pFAST-1 polyplexes at 1 : 10 DNA/pFAST ratio and (B1) (shown in the bottom papne) gWiz-GFP DNA/pFAST-2 polyplexes at 1 : 10 DNA/pFAST ratio. At 48 hours post-incubation, GFP expression (successful intracellular delivery) in the A549 cells was observed under a Leica DMi8 epifluorescence microscope. For convenience, a 3D-rendering in the bottom panel shows the relative position of GFP within the cytosol.
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