Gene delivery system of pDNA using the blood glycoprotein fetuin

Fetuin is a biocompatible plasma protein and strongly enhances phagocytosis of bacteria, DNA and apoptotic cells by peripheral blood cells such as monocytes, macrophages and dendritic cells. We developed a novel gene delivery system: ternary complexes constructed with pDNA, polyethylenimine (PEI) and fetuin. Without covalent binding, fetuin was able to coat pDNA–PEI complexes, and stable anionic nanoparticles formed at a weight ratio greater than 30. Optimised pDNA–PEI–fetuin complexes significantly decreased the cytotoxicity of pDNA–PEI complexes in the melanoma cell line B16F10. Furthermore, the pDNA–PEI–fetuin complexes had higher transgene efficiency compared to that of commercial lipofectin previously reported in B16F10 cells despite an anionic surface. The pDNA–PEI–fetuin complexes did not agglutinate with erythrocytes. The pDNA–PEI–fetuin complexes had high gene expression in the spleen after intravenous administration in mice. Thus, the pDNA–PEI–fetuin complexes were a useful in vivo gene delivery system with tropism for the spleen.     

Secure and effective gene delivery system of plasmid DNA coated by polynucleotide

Polynucleotides are anionic macromolecules which are expected to transfer into the targeted cells through specific uptake mechanisms. So, we developed polynucleotides coating complexes of plasmid DNA (pDNA) and polyethylenimine (PEI) for a secure and efficient gene delivery system and evaluated their usefulness. Polyadenylic acid (polyA), polyuridylic acid (polyU), polycytidylic acid (polyC), and polyguanylic acid (polyG) were examined as the coating materials. pDNA/PEI/polyA, pDNA/PEI/polyU, and pDNA/PEI/polyC complexes formed nanoparticles with a negative surface charge although pDNA/PEI/polyG was aggregated. The pDNA/PEI/polyC complex showed high transgene efficiency in B16-F10 cells although there was little efficiency in pDNA/PEI/polyA and pDNA/PEI/polyU complexes. An inhibition study strongly indicated the specific uptake mechanism of pDNA/PEI/polyC complex. Polynucleotide coating complexes had lower cytotoxicity than pDNA/PEI complex. The pDNA/PEI/polyC complex showed high gene expression selectively in the spleen after intravenous injection into mice. The pDNA/PEI/polyC complex showed no agglutination with erythrocytes and no acute toxicity although these were observed in pDNA/PEI complex. Thus, we developed polynucleotide coating complexes as novel vectors for clinical gene therapy, and the pDNA/PEI/polyC complex as a useful candidate for a gene delivery system.     

Cyclic RGD-poly(ethylene glycol)-polyethyleneimine is more suitable for glioblastoma targeting gene transfer in vivo

Arginine-glycine-aspartic acid (RGD) is a widely chosen ligand to improve the specific gene targeting transfection efficiency of polyethyleneimine (PEI) in vivo. However, the optimal RGD conjugating mode, RGD-poly(ethylene glycol)-PEI (RGD-PEG-PEI) or RGD-PEI-methoxyl poly(ethylene glycol) (RGD-PEI-mPEG) still remains controversial. In this study, RGD-PEG-PEI and RGD-PEI-mPEG were synthesized and compared with respects to their glioblastoma cell-binding capability and tumor-targeting ability of their complexes with plasmid DNA. These results demonstrated that RGD-PEG-PEI/plasmid enhanced green fluorescent protein (pEGFP)-N2 complexes had higher binding affinities with U87 cells than RGD-PEI-mPEG/pEGFP-N2 complexes. The gene transfection was also performed on U87 cells in vitro and in vivo. In vitro, both of the RGD-modified PEI derivatives enhanced the gene transfection efficiency to some extent. However, all of the complexes (with or without RGD modification) had high transfection efficiency. The biodistribution of RGD-PEG-PEI/pEGFP-N2 complexes in mice bearing subcutaneous glioblastomas were significantly greater than that of RGD-PEI-mPEG/pEGFP-N2 complexes, suggesting a more efficient gene transfection in vivo. In the RGD-PEG-PEI, the use of a PEG spacer was particularly important. These results indicated that RGD-PEG-PEI was more suitable for targeted gene transfer in vivo.     

Nanoparticles electrostatically coated with folic acid for effective gene therapy

We developed a novel vector, electrostatically coated poly(ethylenimine) (PEI)/pDNA complexes with folic acid (FA). Without covalent binding, the FA molecules could coat the PEI/pDNA complexes, and stable anionic nanoparticles were formed at a charge ratio greater than 60. The addition of FA markedly decreased the cytotoxicity of the cationic PEI/pDNA complexes to the melanoma cell line, B16-F10 cells, which regularly expressed FA-specific receptor (FR). Furthermore, the anionic FA60/PEI/pDNA complexes showed high transgene efficiency via the FR-mediated pathway in B16-F10 cells. The FA60/PEI/pDNA complexes did not show agglutination with erythrocytes. After the intravenous injection of FA60/PEI/pDNA complexes into mice, a higher transgene efficiency than PEI/pDNA complexes was observed in the liver, kidney, spleen, and lung with FR. The gene expressions of FA60/PEI/pDNA complexes were significantly inhibited by preadministration of FA. Thus, the FA60/PEI/pDNA complexes were useful for effective gene therapy.     

Glioma and microenvironment dual targeted nanocarrier for improved antiglioblastoma efficacy

Drug delivery systems based on nanoparticles (nano-DDS) have aroused attentions for the treatment of glioblastoma (GBM), the most malignant brain cancer with a dismal prognosis. However, there are still numerous unmet challenges for traditional nano-DDS, such as the poor nanoparticle penetration, short retention in the GBM parenchyma and low glioma targeting ability. Herein, we used Pep-1 and CREKA peptides to construct a novel multifunctional GBM targeting nano-DDS (PC-NP). Pep-1 was used to overcome the blood–brain tumor barrier (BBTB) and home to glioma cells via interleukin-13 receptor-α2-mediated endocytosis, and CREKA was used to bind to fibrin–fibronectin complexes abundantly expressed in tumor microenvironment for enhanced retention in the GBM. Biological studies showed that the cellular uptake of PC-NP by U87MG cells was significantly enhanced compared with the non-targeting NP. Furthermore, CREKA modification increased the binding capacity of PC-NP to fibrin–fibronectin complexes as confirmed by the competition experiment. In accordance with the increased cellular uptake, PC-NP remarkably increased the cytotoxicity of its payload paclitaxel (PTX) against U87MG cells with an IC₅₀ of 0.176?μg/mL. In vivo fluorescence imaging and antiglioma efficacy evaluation further confirmed that PC-NP accumulated effectively and penetrated deeply into GBM tissue. PC-NP-PTX exhibited a median survival time as long as 61?days in intracranial GBM-bearing mice. In conclusion, our findings indicated PC-NP as a promising nano-DDS for GBM targeting delivery of anticancer drugs.     

Intracellular uptake and release of poly(ethyleneimine)-co-poly(methyl methacrylate) nanoparticle/pDNA complexes for gene delivery

Our previous studies demonstrated that cationic nanoparticles composed of well-defined poly(methyl methacrylate) (PMMA) cores surrounded by a hairly poly(ethyleneimine) (PEI) shells have comparative advantages over the PEI system for gene delivery. In this study, we focused on the intracellular uptake and release of PEI-PMMA nanoparticle/pDNA complexes. The behavior of the nanoparticle/pDNA complexes in recipient cells was monitored by using confocal laser scanning microscopy. We found that the nanoparticle/pDNA complexes were internalized very effectively by endocytosis. In the recipient cells the nanoparticles were found localized in the cytoplasm. At the same time, the pDNA carried by the nanoparticles successfully detached from the nanoparticles and localized in the nucleus of the HeLa cells.     

Anti-cancer effect of R3V6 peptide-mediated delivery of an anti-microRNA-21 antisense-oligodeoxynucleotide in a glioblastoma animal model

MicroRNA-21 (miR-21) expression in glioblastoma inhibits the expression of pro-apoptotic genes, thereby promoting tumor growth. A previous study showed that the amphiphilic R3V6 peptide is an efficient carrier of the anti-miR-21 antisense oligodeoxynucleotide (antisense-ODN) into cells in vitro. In the current study, in vivo delivery of antisense-ODN using the R3V6 peptide was evaluated in a glioblastoma animal model. In vitro transfection showed that the R3V6 peptide delivered antisense-ODN more efficiently than polyethylenimine (25?kDa, PEI25k) in C6 glioblastoma cells. For in vivo evaluation, antisense-ODN/R3V6 complex was injected intratumorally into a C6 glioblastoma xenograft animal model. Tumor growth was suppressed by the injection of the antisense-ODN/R3V6 complex, compared with the antisense-ODN/PEI25k and scrambled-antisense-ODN (scr-antisense-ODN)/R3V6 complexes. Real-time RT-PCR showed that miR-21 levels were reduced most efficiently by the antisense-ODNR3V6 complex in tumors. Due to inhibition of miR-21, expression of the programed cell death 4 (PDCD4) gene was promoted in tumors, resulting in the induction of apoptosis of tumor cells. These results suggest that delivery of antisense-ODN using R3V6 peptides may be useful for the development of antisense-ODN therapy for glioblastoma.     

Poly(l-lactide-co-glycolide) nanospheres conjugated with a nuclear localization signal for delivery of plasmid DNA

Polymeric nanospheres fabricated from biodegradable poly(lactide-co-glycolide) (PLGA) have been extensively investigated for applications in gene delivery. In this study, we show that the covalent conjugation of a nuclear localization signal (NLS, SV40 peptide) on PLGA nanospheres enhances the gene transfection efficiency. NLS conjugated PLGA copolymer was prepared by using a coupling reaction between maleimide-terminated PLGA copolymer and NLS in the presence of Imject maleimide conjugation buffer. PLGA nanospheres encapsulating plasmid (pDNA) were prepared by using a double emulsion-solvent evaporation method. The kinetics of in vitro release of pDNA from PLGA nanospheres was determined with UV in phosphate buffered saline (PBS). Gene transfection efficiency in human dermal fibroblasts was tested in vitro using nanospheres encapsulating the luciferase gene. The conjugation of the NLS peptide to the PLGA nanospheres could improve the nuclear localization and/or cellular uptake of PLGA nanosphere/pDNA constructs and thereby improve the transfection efficiency of a PLGA nanosphere gene delivery system. The pDNA was released from PLGA nanospheres over nine days. NLS conjugation enhanced the gene transfection efficiency in vitro by 1.2 ～ 3.2-fold over 13 days. PLGA/pDNA nanospheres appeared to be superior to PEI/pDNA complexes for the long-term expression of pDNA. Furthermore, the level of the sustained gene expression of the PLGA nanospheres was enhanced by the conjugation of NLS to the PLGA nanospheres. This study showed that the NLS conjugation enhanced the gene transfection efficiency of the PLGA nanosphere gene delivery system in vitro and that the enhanced gene expression was sustained for at least 13 days.     

Glioma homing peptide-modified PEG-PCL nanoparticles for enhanced anti-glioma therapy

Background: Peptide-mediated drug delivery system (DDS) has been increasingly used to promote on-demand treatment efficacy of cancers. Herein, LTLRWVGLMS (LS10) peptide is selected as the functional ligand for specific glioma-targeting drug delivery. LS10 peptide selectively binds to NG2 proteins that are widely overexpressed in the glioma cells and restricted in normal tissue. LS10 peptide-decorated DDS is expected to hold vast promises in glioma therapy and decrease unwanted side effects.

Methods: LS10 peptide was conjugated on the surface of poly(ethylene glycol)-poly(?-caprolactone) (PEG-PCL) nanoparticles via 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride/N-hydroxysulfosuccinimide coupling reaction. Using U87MG cells as the glioma cell model, cellular uptake, internalization mechanism, cellular cytotoxicity and apoptosis were investigated. 1,1′-Dioctadecyl-3,3,3′,3′-tetramethyl indotricarbocyanine iodide were used as fluorescence probes to investigated in vivo glioma targeting capability of LS10-NP. The glioma therapeutic efficacy of paclitaxel-loaded LS10-NP was studied on glioma-bearing nude mice.

Results: The LS10-NP with size of 119?nm enhanced cellular uptake on U87MG cells, increased cytotoxicity of the loaded paclitaxel (PTX), and improved penetration in 3D U87MG glioma spheres. In vivo biodistribution experiments showed that LS10-NP exhibited the enhanced drug localization at glioma site, which resulted in prolonged survival time of glioma-bearing mice.

Conclusion: Our results indicated that LS10 peptide-modified nanoparticulate DDS could significantly improve the anti-glioma efficacy.     

Enhancement of gene transfection into human dendritic cells using cationic PLGA nanospheres with a synthesized nuclear localization signal

Effective delivery of DNA encoding antigen into the dendritic cells (DCs), which are non-dividing cells, is very important for the development of DNA vaccines. In a previous study, we developed the PLGA nanospheres that contained a cationic nanomaterial and showed high transfection efficiency in COS7 cells, which divide. In the present study, to produce an effective vector for the DNA vaccines, the gene expression and intracellular trafficking of pDNA complexed with PLGA/PEI nanospheres, in combination with an NF-κB analog as a nuclear localization signal (NLS) and electroporation were evaluated in human monocyte-derived DCs (hMoDCs). Cellular uptake of pDNA both in COS7 cells and hMoDCs was enhanced using the PLGA/PEI nanospheres. On the other hand, the PLGA/PEI nanospheres significantly promoted the transfection in COS7 cells, but had almost no effect on transfection in hMoDCs. The intranuclear transport of pDNA by PLGA/PEI nanospheres in COS7 cells was significantly higher than that in hMoDCs. These results indicate that pDNA complexed with PLGA/PEI nanospheres cannot enter into the nuclei of non-dividing cells. However, PLGA/PEI nanospheres combinated with NLS and electroporation (experimental permeation enhancer) greatly elevated the transfection efficiency by improvement of not only intracellular uptake but also intranuclear transport of pDNA in the hMoDCs. Thus, this delivery system using nanospheres combined with synthesized NLS might be applicable to DC-based gene vaccines when much non-invasive application such as needle-free injector should be required.     

A Potential Targeting Gene Vector Based on Biotinylated Polyethyleneimine/Avidin Bioconjugates

Purpose  To improve the gene delivery efficiency and safety of non-viral vector in liver cells, avidin, which exhibited good biocompatibility and remarkable accumulation in liver, was bioconjugated with biotinylated polyethylenimine to obtain a novel gene vector. Materials and methods  Biotinylated polyethyleneimine/avidin bioconjugate (ABP) was synthesized through grafting biotin to high molecular weight branched polyethylenimine (PEI, 25 kDa) and then bioconjugating with avidin by the biotin-avidin interaction. Physiochemical characteristics of ABP/pDNA complexes were analyzed, and in vitro cytotoxicity and transfection of ABP were also evaluated in HepG2, Hela and 293 T cells by using 25 kDa PEI as the control. Results  It was found that ABP was able to condense pDNA efficiently at N/P ratio of 4. The particle sizes of ABP/pDNA complexes were less than 220 nm, and the average surface charges were around 27 mV at the N/P ratio ranging from 2 to 60. Among three different cell lines, ABP and its DNA complexes demonstrated much lower cytotoxicity and higher transfection efficacy in HepG2 cells as compared with 25 kDa PEI. Conclusion  ABP presented higher transfection efficacy and safety in HepG2 cells due to the biocompatibility of avidin and the specific interactions between avidin and HepG2 cells.     

Delivery of anti-microRNA-21 antisense-oligodeoxynucleotide using amphiphilic peptides for glioblastoma gene therapy

Inhibition of microRNA-21 (miR-21) has been shown to promote apoptosis of cancer cells and to reduce tumor size in glioblastoma. However, efficient carriers for antisense-oligodeoxynucleotide (antisense-ODN) against miR-21 have not yet been developed. In this study, the R3V6 peptide (R3V6) was evaluated as a carrier of antisense-ODN. In a gel retardation assay, R3V6 formed a complex with an antisense-ODN. The serum stability assay showed that R3V6 protected it from nucleases more efficiently than polyethylenimine (PEI; 25?kDa, PEI25k). A Renilla luciferase gene with a 3′-untranslated region (3′-UTR) recognizable by miR-21 (psiCHECK2-miR-21-UTR) was constructed for the antisense-ODN assay. psiCHECK2-miR-21-UTR expressed less Renilla luciferase in the cells with a higher level of miR-21 due to the effect of miR-21. In an in vitro transfection assay, the R3V6 peptide delivered anti-miR-21 antisense-ODN into cells more efficiently than PEI (25?kDa, PEI25k) and lipofectamine. As a result, antisense-ODN/R3V6 complex inhibited miR-21 and increased Renilla luciferase expression more efficiently than antisense-ODN/PEI25k or antisense-ODN/Lipofectamine complexes in both C6 and A172 glioblastoma cells. Furthermore, the antisense-ODN/R3V6 complexes reduced the level of miR-21 and induced apoptosis of glioblastoma cells. These results suggest that the R3V6 peptide may be a useful carrier of antisense-ODN for glioblastoma gene therapy.     

T7 peptide-functionalized nanoparticles utilizing RNA interference for glioma dual targeting

Among all the malignant brain tumors, glioma is the deadliest and most common form with poor prognosis. Gene therapy is regarded as a promising way to halt the progress of the disease or even cure the tumor and RNA interference (RNAi) stands out. However, the existence of the blood–brain barrier (BBB) and blood tumor barrier (BTB) limits the delivery of these therapeutic genes. In this work, the delivery system targeting to the transferrin (Tf) receptor highly expressed on both BBB and glioma was successfully synthesized and would not compete with endogenous Tf. U87 cells stably express luciferase were employed here to simulate tumor and the RNAi experiments in vitro and in vivo validated that the gene silencing activity was 2.17-fold higher with the targeting ligand modification. The dual-targeting gene delivery system exhibits a series of advantages, such as high efficiency, low toxicity, stability and high transaction efficiency, which may provide new opportunities in RNAi therapeutics and nanomedicine of brain tumors.     

Galactomannan-PEI based non-viral vectors for targeted delivery of plasmid to macrophages and hepatocytes

Intracellular nature and diversified locations of infectious and parasitic diseases such as leishmaniasis, trypanosomiasis, tuberculosis and hepatitis B and C pose a significant global burden and challenge to the scientists working in the area of drug discovery and drug delivery. The macrophages and hepatocytes are considered as potential target sites as they together play an important role in various infectious diseases. The present study scrutinizes the applicability of a natural biopolymer-based chemical vectors, capable of targeting both macrophages and hepatocytes, that can form a complex with plasmid and administer it into cells to produce a desired protein. The investigations were made to develop a novel series of gene carriers by conjugating depolymerized galactomannan (guar gum), a biocompatible polysaccharide with low molecular weight branched PEI (LMWP). A series of conjugates were developed and characterized using physicochemical techniques. All the GP/pDNA complexes showed significantly higher transfection efficiency with GP-3/pDNA, one of the best formulations, showed ∼2.0–7.7-folds higher transfection efficacy when compared with the standard transfection reagents. Further, GP-3/pDNA displayed significantly higher target specific transfection efficiency under both in vitro and in vivo conditions. The data demonstrate the potential of GP vectors to deliver nucleic acids simultaneously to macrophages and hepatocytes in gene delivery applications.     

Insights into the mechanism of magnetofection using MNPs-PEI/pDNA/free PEI magnetofectins

Magnetofection is an efficient new physical gene transfection technology. Despite its effective gene delivery capability, till now relatively little work has been conducted on the mechanism of magnetofection, especially the intracellular fates of the components of magnetofectins and their effects on magnetofection. In this study, we investigated the mechanism of magnetofection using magnetofectins that were prepared via electrostatic self-assembly of the three components: polyethyleneimine (PEI)-coated magnetic nanoparticles (MNPs-PEI), plasmid DNA (pDNA) and PEI in the free form (free PEI). TEM observation and agarose gel electrophoresis assays have indicated MNPs play the role of driving magnetofectins to the cell surface without entering into the nucleus. Confocal microscopic tracking of fluorescence-labeled PEI has shown that the free PEI (green) can be found in the nucleus but almost all of the MNPs-PEI (red) are confined in the cytoplasm in COS-7 cells 30 min post-transfection or in SPC-A1 cells 90 min post-transfection, implying that the pDNA/PEI complex must separate from MNPs-PEI before entering into the nucleus. In addition, reporter gene assays showed the magnetofectins, in which the free PEI was absent, failed to transfect SPC-A1 or COS-7 cell lines; and there was an optimal ratio of the constituents of magnectofectins to achieve optimal transfection efficiency by balancing stable complex formation and facile release of PEI/pDNA from the complex. In summary, our findings further the knowledge of magnetofection and can be helpful for the design and preparation of gene delivery vehicles for effective magnetofection.     

Physical stability and in-vitro gene expression efficiency of nebulised lipid-peptide-DNA complexes

The lower respiratory tract provides a number of disease targets for gene therapy. Nebulisation is the most practical system for the aerosolisation of non-viral gene delivery systems. The aerosolisation process represents a significant challenge to the maintenance of the physical stability and biological activity of the gene vector. In this study we investigate the role of a condensing polycationic peptide on the stability and efficiency of nebulised lipid-DNA complexes. Complexes prepared from the cationic lipid 1, 2-dioleoyl-3-trimethylammonium propane (DOTAP) and plasmid DNA (pDNA) at mass (w/w) ratios of 12:1, 6:1 and 3:1, and complexes prepared from DOTAP, the polycationic peptide, protamine, and pDNA (LPD) at 3:2:1 w/w ratio were nebulised using a Pari LC Plus jet nebuliser. Samples from the nebuliser reservoir (pre- and post-nebulisation) and from the aerosol mist were collected and investigated for changes, including: particle diameter, retention of in-vitro transfection activity and the relative concentration and nature of the complexed pDNA remaining after the nebulisation procedure. The process of jet nebulisation adversely affected the physical stability of lipid:pDNA complexes with only those formulated at 12:1 w/w DOTAP:pDNA able to maintain their pre-nebulisation particle size distribution (145+/-3 nm pre-nebulisation vs. 142+/-2 nm aerosol mist) and preserve significant pDNA integrity in the reservoir (35% of pre-nebulisation pDNA band intensity). The LPD complexes were smaller (102+/-1 nm pre-nebulisation vs. 113+/-2 nm aerosol mist) with considerably greater retention of pDNA integrity in the reservoir (90% of pre-nebulisation pDNA band intensity). In contrast the concentration of pDNA in the aerosol mist for both the 12:1 w/w DOTAP:pDNA and LPD complexes were significantly reduced (10 and 12% of pre-nebulised values, respectively). Despite reduced pDNA concentration the transfection (% cells transfected) mediated by aerosol mist for the nebulised complexes was comparatively efficient (LPD aerosol mist 26 vs. 40% for pre-nebulised complex; the respective values for 12: 1 w/w DOTAP:pDNA were 12 vs. 28%). The physical stability and biological activity of nebulised lipid:pDNA complexes can be improved by inclusion of a condensing polycationic peptide such as protamine. The incorporation of the peptide precludes the use of potentially toxic excesses of lipid and charge and may act as a platform for the covalent attachment of peptide signals mediating sub-cellular targetting.     

Poly(L-lactic acid)/polyethylenimine nanoparticles as plasmid DNA carriers

Non-viral vectors such as liposomes, polycations, and nanoparticles have been used as gene delivery systems. In this study, we prepared and characterized biodegradable poly(L-lactic acid) (PLA)/polyethylenimine (PEI) nanoparticles as gene carriers. pCMV/β-gal and pEGFP-C1 were utilized as model plasmid DNAs (pDNA). Nanoparticles were prepared using a double emulsion-solvent evaporation technique, and their pDNA binding capacity was assessed by agarose gel electrophoresis. Transfection was studied in HEK 293 and HeLa cell lines, and the transfection efficiencies were determined by β-galactosidase assay or flow cytometry. Three kinds of PLA/PEI systems were studied by varying the molecular weight of PEI. The PLA/PEI 25K system had a higher transfection efficiency than the PLA/PEI 0.8K or PLA/PEI 750K systems. The transfection efficiency was found to be dependent on the ratio of PLA/PEI nanoparticles to pDNA with an optimum ratio of 60:1 (w/w). The cytotoxicity was dependent on the quantity of PLA/PEI nanoparticles used, but it was comparable to that of commercial Lipofectin™. These results demonstrate the potential of PLA/PEI nanoparticles as gene carriers.     

Physicochemical characterization of poly(L-lactic acid) and poly(D,L-lactide-co-glycolide) nanoparticles with polyethylenimine as gene delivery carrier

Polymer nanoparticles have been used as non-viral gene delivery systems and drug delivery systems. In this study, biodegradable poly(L-lactic acid) (PLA)/polyethylenimine (PEI) and poly(D,L-lactide-co-glycolide) (PLGA)/PEI nanoparticles were prepared and characterized as gene delivery systems. The PLA/PEI and PLGA/PEI nanoparticles, which were prepared by a diafiltration method, had spherical shapes and smooth surface characteristics. The size of nanoparticles was controlled by the amount of PEI, which acted as a hydrophilic moiety, which effectively reduced the interfacial energy between the particle surface and the aqueous media. The nanoparticles showed an excellent dispersive stability under storage in a phosphate-buffered saline solution for 12 days. The positive zeta-potentials for the nanoparticles decreased and changed to negative values with increasing plasmid DNA (pDNA) content. Agarose gel electrophoresis showed that the complex formation between the nanoparticles and the pDNA coincided with the zeta-potential results. The results of in vitro transfection and cell viability on HEK 293 cells indicated that the nanoparticles could be used as gene delivery carriers.     

Transferrin receptor targeting segment T7 containing peptide gene delivery vectors for efficient transfection of brain tumor cells

Successful gene therapy for brain tumors are often limited by two important factors, the existence of blood brain barrier (BBB) and inefficient transfection of brain tumor cells. In this study, we designed a series of peptide-based gene delivery vectors decorated with T7 segment for binding the transferrin (Tf) receptors which were highly expressed on brain tumor cells, and evaluated their ability of gene delivery. The physicochemical properties of peptide vectors or peptide/DNA complexes were studied as well. The in vitro transfection efficiency was investigated in normal and glioma cell lines. Among these complexes, PT-02/DNA complexes showed the highest transfection efficiency in glioma cells and low cytotoxicity in normal cell lines, and it could transport DNA across the BBB model in vitro. Furthermore, PT-02/DNA could deliver pIRES2-EGFP into the brain site of zebrafish in vivo. The designed peptide vectors offered a promising way for glioma gene therapy.