Targeting Cancer Cells Overexpressing Folate Receptors with New Terpolymer-Based Nanocapsules: Toward a Novel Targeted DNA Delivery System for Cancer Therapy (original) (raw)
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Intracellular Delivery of Nanometric DNA Particles via the Folate Receptor
Bioconjugate Chemistry, 2002
The size of condensed DNA particles is a key determinant for both diffusion to target cells in vivo and intracellular trafficking. The smallest complexes are obtained when each DNA molecule collapses individually. This was achieved using a designed cationic thiol-detergent, tetradecyl-cysteinyl-ornithine (C 14 COrn). The resulting particles were subsequently stabilized by air-induced dimerization of the detergent into a disulfide lipid on the DNA template. Particles are anionic (zeta potential ) -45 mV), and their size (30 nm) corresponds to the volume of a single plasmid DNA molecule. The electrophoretic mobility of the condensed DNA, though quasi-neutralized, was found higher than that of the extended DNA. Moreover, the dimerized (C 14 COrn) 2 lipid was found to be an efficient transfection reagent for various cell lines. In an attempt to achieve extended circulation times and to target tumors by systemic delivery, we have coated the particles with PEG-folate residues. Plasmid DNA was condensed into monomolecular particles as described above and coated by simple mixing with DPPE-PEG-folate. Physicochemical measurements showed particles coated with 2% of DPPE-PEG 3400folate remain monomolecular and are stable in the cell-culture medium. Caveolae-mediated cell entry was demonstrated by ligand-dependence, by competition with excess folic acid as well as by confocal microscopy.
Biomaterials, 2010
Targeted gene delivery systems have attracted great attention due to their potential in directing the therapeutic genes to the target cells. However, due to their low efficiency, most of the successful applications of polymeric vectors have been focused on genes which can achieve robust expression. Minicircle DNA (mcDNA) is a powerful candidate in terms of improving gene expression and prolonging the lifespan of gene expression. In this study, we have combined folate/poly(ethylene glycol) modified polyethylenimine and mcDNA as a new tumor gene delivery system. We found that folate-labeled polyplexes were homogenous, with a size ranging from 60 to 85 nm. mcDNA increased folate-labeled vector based gene expression 2e8 fold in folate receptor-positive cells. Results of folic acid competition assay indicated that mcDNA mediated by folate-labeled vector were internalized into cells through receptormediated endocytosis. The investigation of the endocytosis pathway of the polyplexes showed that a large portion of them escaped from endo/lysosome and the polyplexes were associated before being separated in the nucleus. Furthermore, in vivo optical imaging and luciferase assays demonstrated that systemic delivery of the folate-labeled polyplexes resulted in preferential accumulation of transgenes in folate receptor-positive tumors, and mcDNA mediated approach achieved 2.3 fold higher gene expressions in tumors than conventional plasmid. Cytotoxicity assays showed that PEG-shielding of the polyplexes reduced the toxicity of PEI.
Long-circulating DNA lipid nanocapsules as new vector for passive tumor targeting
Biomaterials, 2010
Systemic gene delivery systems are needed for therapeutic application to organs that are inaccessible by percutaneous injection. Currently, the main objective is the development of a stable and non-toxic vector that can encapsulate and deliver foreign genetic material to target cells. To this end, DNA, complexed with cationic lipids i.e DOTAP/DOPE, was encapsulated into lipid nanocapsules (LNCs) leading to the formation of stable nanocarriers (DNA LNCs) with a size inferior to 130nm. Amphiphilic and flexible poly (ethylene glycol) (PEG) polymer coatings [PEG lipid derivative (DSPE-mPEG 2000 ) or F108 poloxamer] at different concentrations were selected to make DNA LNCs stealthy. Some of these coated lipid nanocapsules were able to inhibit complement activation and were not phagocytised in vitro by macrophagic THP-1 cells whereas uncoated DNA LNCs accumulated in the vacuolar compartment of THP-1 cells. These results correlated with a significant increase of in vivo circulation time in mice especially for DSPE-mPEG 2000 10mM and an early half-life time (t 1/2 of distribution) 5-fold greater than for non-coated DNA LNCs (7.1h vs 1.4h). Finally, a tumor accumulation assessed by in vivo fluorescence imaging system was evidenced for these coated LNCs as a passive targeting without causing any hepatic damage.
Efficient Nonviral Gene Therapy Using Folate-Targeted Chitosan-DNA Nanoparticles In Vitro
ISRN Pharmaceutics, 2012
Nonviral cationic polymers like chitosan can be combined with DNA to protect it from degradation. The chitosan is a biocompatible, biodegradable, nontoxic, and cheap polycationic polymer with low immunogenicity. The objective of this study was to synthesize and then assess different chitosan-DNA nanoparticles and to select the best ones for selectivein vitrotransfection in human epidermoid carcinoma (KB) cell lines. It revealed that different combinations of molecular weight, the presence or absence of folic acid ligand, and different plasmid DNA sizes can lead to nanoparticles with various diameters and diverse transfection efficiencies. The intracellular trafficking, nuclear uptake, and localization are also studied by confocal microscopy, which confirmed that DNA was delivered to cell nuclei to be expressed.
Modular Construction of Multifunctional Bioresponsive Cell-Targeted Nanoparticles for Gene Delivery
Bioconjugate Chemistry, 2011
Multifunctional and modular block copolymers prepared from biocompatible monomers and linked by a bioreducible disulfide linkage have been prepared using a combination of ring-opening and atomtransfer radical polymerizations (ATRP). The presence of terminal functionality via ATRP allowed cell-targeting folic acid groups to be attached in a controllable manner, while the block copolymer architecture enabled well-defined nanoparticles to be prepared by a water-oil-water double emulsion procedure to encapsulate DNA with high efficiency. Gene delivery assays in a Calu-3 cell line indicated specific folatereceptor-mediated uptake of the nanoparticles, and triggered release of the DNA payload via cleavage of the disulfide link resulted in enhanced transgene expression compared to nonbioreducible analogues. These materials offer a promising and generic means to deliver a wide variety of therapeutic payloads to cells in a selective and tunable way.
Biomaterials, 2007
Poor water solubility and low transfection efficiency of chitosan are major drawbacks for its use as a gene delivery carrier. PEGylation can increase its solubility, and folate conjugation may improve gene transfection efficiency due to promoted uptake of folate receptorbearing tumor cells. The aim of this study was to synthesize and characterize folate-poly(ethylene glycol)-grafted chitosan (FA-PEG-Chi) for targeted plasmid DNA delivery to tumor cells. Gel electrophoresis study showed strong DNA binding ability of modified chitosan. The pH 50 values, defined as the pH when the transmittance of a polymer solution at 600 nm has reached 50% of the original value, suggested that the water solubility of PEGylated chitosan had improved significantly. Regression analysis of pH 50 value as a function of substitution degree of PEG yielded an almost linear correlation for PEG-Chi and FA-PEG-Chi. The solubility of PEGylated chitosan decreased slightly by further conjugation of folic acid due to the relatively more hydrophobic nature of folic acid when compared to PEG. In addition, the chitosan-based DNA complexes did not induce remarkable cytotoxicity against HEK 293 cells. FA-PEG-Chi can be a promising gene carrier due to its solubility in physiological pH, efficiency in condensing DNA, low cytotoxicity and targeting ability. r
Colloids and Surfaces B: Biointerfaces, 2015
Folic acid-conjugated d-␣-tocopheryl polyethylene glycol 1000 succinate (TPGS-FOL) decorated methotrexate (MTX)-conjugated nanoparticles were developed for targeted delivery of MTX to folate receptor-expressed tumor cells. The synthesis of TPGS-FOL followed 3-step process. Firstly, the terminal hydroxyl group of TPGS was converted to sulfonyl chloride using mesyl chloride in comparison with nosyl and tosyl chlorides. The highest conversion efficiency and yield were obtained by mesyl chloride due to the formation of higher reactive intermediate in a presence of triethylamine. Secondly, the substitution of sulfonyl group by sodium azide produced considerably high yield with conversion efficiency of over 90%. Lastly, the coupling reaction of azido-substituted TPGS and propargyl folamide by click reaction resulted in 96% conjugation efficiency without polymer degradation. To fabricate the folate receptortargeted nanoparticles, 10 and 20%mol MTX-conjugated PEGylated poly(-caprolactone) nanoparticles were decorated with TPGS-FOL. The size and size distribution of MTX-conjugated nanoparticles relatively increased with %MTX. The MTX release from the nanoparticles was accelerated in acidic medium with an increase of %MTX but retarded in physiological pH medium. The decoration of TPGS-FOL onto the nanoparticles slightly enlarged the size and size distribution of the nanoparticles; however, it did not affect the surface charge. The cytotoxicity and cellular uptake of MCF-7 cells demonstrated that 10% MTX-conjugated nanoparticles and FOL-decorated nanoparticles possessed higher toxicity and uptake efficiency than 20% MTX-conjugated nanoparticles and undecorated nanoparticles, respectively. The results indicated that FOL-10% MTX-conjugated nanoparticles exhibited potential targeted delivery of MTX to folate receptor-expressed cancer cells.
Tumor transfection after systemic injection of DNA lipid nanocapsules
Biomaterials, 2011
With the goal of generating an efficient vector for systemic gene delivery, a new kind of nanocarrier consisting of lipid nanocapsules encapsulating DOTAP/DOPE lipoplexes (DNA LNCs) was pegylated by the post-insertion of amphiphilic and flexible polymers. The aim of this surface modification was to create a long-circulating vector, able to circulate in the blood stream and efficient in transfecting tumoral cells after passive targeting by enhanced permeability and retention effect (EPR effect). PEG conformation, electrostatic features, and hydrophylicity are known to be important factors able to influence the pharmacokinetic behaviour of vectors. In this context, the surface structure characteristics of the newly pegylated DNA LNCs were studied by measuring electrophoretic mobility as a function of ionic strength in order to establish a correlation between surface properties and in vivo performance of the vector. Finally, thanks to this PEGylation, gene expression was measured up to 84-fold higher in tumor compared to other tested organs after intravenous injection. The present results indicate that PEGylated DNA LNCs are promising carriers for an efficient cancer gene therapy.