Jialiang Xia

Hydrophobic polyphenylalanine-grafted hyperbranched polyethylenimine and its in vitro gene transfection.

A series of amphiphilic multi-armed PPn copolymers were prepared by ROP of Phe-NCA with PEI-25k as a macroinitiator. The particle size of the PPn/DNA complexes was about 100 nm and the zeta potentials were below 20 mV. An MTT assay demonstrated that all the PPn copolymers had lower cytotoxicity compared to PEI-25k. In vitro gene transfection studies were also conducted in HeLa, 293 and CT 26 cells. The optimal quantity of hydrophobic phenylalanine segments in PP80 led to higher transfection efficiency in various cell lines based on this study. The results indicate that PP80 was the best candidate for gene delivery among these PPn copolymers.

PEI Conjugated Gold Nanoparticles: Efficient Gene Carriers with Visible Fluorescence

Low molecular weight polyethyleneimines were conjugated onto gold nanoparticles to form Au-PEI conjugates. Au-PEI is the first reported gene carrier which has both high transfection efficiency and strong fluorescence. Au-PEI showed higher transfection efficiency and it can be used for bioimaging because it can be detected by confocal laser scanning microscopy and in vivo bioimaging system.

A highly efficient siRNA carrier of PBLG modified hyperbranched PEI

In search for effective non-viral gene vectors for the delivery of siRNA, a copolymer was designed and synthesized by grafting hydrophobic poly(gamma-benzyl L-glutamate) segment (PBLG) to hyperbranched polyethylenimine (PEI-PBLG). PEI-PBLG could efficiently deliver siRNA to cells to silence the targeted gene. Markedly, PEI-PBLG caused lower cytotoxicity in comparison to unmodified PEI. The siRNA complexed with PEI-PBLG showed a remarkable knockdown (75.23% relative to untreated cells, without changing the medium after 6 h of incubation) of the targeted luciferase gene in stable expressing luciferase CT26 cells while the Lipofectamine2000 and unmodified PEI could only achieve knockdown rates of 57.92% and 15.31%, respectively. The siRNA complexed with PEI-PBLG also demonstrated that it had greater gene silencing ability than unmodified PEI and Lipofectamin2000 in both 4T1 cells stably transfected with the luciferase gene and HeLa cells transiently transfected with the luciferase gene. The internalization efficiency of carrier/Alexa647-labeled siRNA was quantified using flow cytometry. PEI-PBLG/Alexa647-labeled siRNA showed internalization efficiency of 52.67% while PEI and Lipofectamine2000 demonstrated 27.23% and 37.91%, respectively. Confocal laser scanning microscopy (CLSM) assay also indicated that PEI-PBLG induced higher cell uptake efficiency than other commercial reagents. PEI-PBLG was shown to be a promising siRNA carrier with potential application in cancer therapy.

Charge-conversional zwitterionic copolymer as pH-sensitive shielding system for effective tumor treatment

UNLABELLED A novel pH-responsive gene delivery system for tumor acidity-targeted pDNA delivery is prepared by introducing a rapid charge-conversional zwitterionic copolymer to the positive surface of PEI/pDNA complexes through electrostatic interaction. The shielding system (OEAL) consists of oligoethylenimine (OEI), poly(l-aspartate) (PBLA), and poly(l-lysine) (PLL). The charge-conversional behavior of the OEAL/PEI/DNA ternary complex is evaluated by zeta potential assay. The surface charges of the complexes can change from negative to positive in the pH range of 7.4-6.8. Under a simulative in vivo environment, OEAL/PEI/DNA exhibits promotion of cellular uptake by tumor cells and enhanced gene transfection efficiency because of its good charge-conversional properties. Antitumor experiments further show that the pH-responsive charge-conversional system can mediate a therapeutic gene that can induce tumor apoptosis (pKH3-rev-casp-3) to achieve effective tumor inhibition. Accordingly, OEAL can be regarded as a promising tumor microenvironment-sensitive gene delivery shielding system for antitumor therapy. STATEMENT OF SIGNIFICANCE This manuscript focused on the novel pH-responsive gene delivery system for tumor acidity-targeted pDNA delivery. The novel system is prepared by introducing a rapid charge-conversional zwitterionic copolymer, consisting of oligoethylenimine, poly(l-aspartate) and poly(l-lysine), to the positive surface of PEI/pDNA complexes. The surface charges of the complexes can change from negative to positive from pH 7.4 to 6.8. OEAL/PEI/DNA shows promoting cellular uptake by tumor cells and enhanced gene transfection efficiency. The antitumor experiments further show that the pH responsive charge conversional system can mediate pKH3-rev-casp-3 to achieve effective tumor inhibition. Accordingly, OEAL can be regarded as a promising tumor microenvironment sensitive gene delivery shielding system for antitumor therapy.

Polyglutamic acid based polyanionic shielding system for polycationic gene carriers

For the purpose of increasing the in vivo stability of polycation gene carriers, we prepared a kind of pH-sensitive poly(ethylene glycol)-poly (γ-benzyl-L-glutamate-co-glutamic acid) (PEG-PGA(65), 65 denotes the molar ratio of glutamic acid in poly(γ-benzyl-L-glutamate-co-glutamic acid)). PEG-PGA(65) showed low cytotoxicity and could shield the positive charge of DNA/PEI (1:1) polyplexes efficiently. The transfection was enhanced due to the partially charge shielding in HeLa cell line at pH of 7.4. There was almost no transfection efficiency when the surface charge of the ternary particles turned to negative at pH of 7.4. However, the transfection efficiency recovered a lot by culturing at pH of 6.0 at the beginning of transfection. Confocal microscopic observation and flow cytometry results showed DNA/PEI polyplexes should be efficiently released and endocytosized at pH 6.0, because of the pH triggered deshielding action of PEG-PGA(65). Due to the good biocompatibility and suitable pH triggered shielding/deshielding property, PEG-PGA(65) could be a potential shielding system for polycationic gene carriers used in vivo.

Oligoethylenimines Grafted to PEGylated Poly(β-amino ester)s for Gene Delivery

Novel polymers composed of net-like PEGylated poly(β-amino ester) (N-P-1, M(w) = 6900 or N-P-2, M(w) = 21,400) and oligoethylenimine (OEI) (OEI423 or OEI600) were synthesized and evaluated as gene carriers. The molecular weights of these polymers were well-controlled by the concentration of the cross-linking reaction. The synthesized polymers showed high biodegradability, less cytotoxicity, and efficient DNA retard ability. The N-P-1-OEI600/DNA complex showed much slower aggregation in the presence of 10 and 20% serum solutions. In vitro transfection assays, N-P-2-OEI423, N-P-1-OEI600, and N-P-2-OEI600 showed enhanced transfection efficiency compared with the PEI25K control in the presence or in the absence of serum in different cell lines. In particular, in Cos-7 cells, the transfection efficiency of N-P-1-OEI600 was 20.9 times higher than that of PEI25K in the presence of serum. The polymer/DNA complex stability, lower cytotoxicity, and higher transfection efficiency in the presence of serum revealed that N-P-1-OEI600 could be a potential nonviral gene carrier for In Vivo application.

SYNTHESIS AND CHARACTERIZATION OF GENIPIN CROSS-LINKED OLIGOETHYLENIMINE FOR GENE DELIVERY: SYNTHESIS AND CHARACTERIZATION OF GENIPIN CROSS-LINKED OLIGOETHYLENIMINE FOR GENE DELIVERY

A novel fluorescent cationic polymer, genipin cross-linked branched oligoethylenimine (1800 Da, OEI1800), GP, was successfully synthesized in 70% ethanol solution and evaluated as a non-viral gene carrier. The copolymers were characterized by nuclear magnetic resonance spectroscopy ((1)H-NMR) and gel permeation chromatography (GPC). The complexes of copolymers/DNA based on GP were measured by dynamic light scattering and Zeta potential instrument. The results showed that the synthesized copolymers could efficiently condense DNA into nanosized particles (120 similar to 150 nm) with positive surface charges (+ 20 similar to 25 mV). The DNA-binding ability of the copolymers was further examined by gel retardation assy. When the mass ratio of polymer and DNA was 0.25, the plasmids were fully retarded in 1% agarose gel by gel retardation electrophoresis. Moreover, the in vitro cytotoxicity and transfection efficiency of the GP copolymers were respectively tested by the MTT and luciferase assay,using OEI1800 and PEI25k as the control in the HEK293 cells lines. The cell toxicity and gene transfection evaluations showed that the copolymers exhibited lower cytotoxicity and higher gene transfection efficiency than PEI25k in the HEK293 cell lines. Finally, the carriers could be used as the fluorescent labeling materials. Therefore, the copolymers may have potential biomedical application as non-viral gene carriers.

pH-Triggered Sheddable Shielding System for Polycationic Gene Carriers

For improving the therapeutic efficiency of tumors and decreasing undesirable side effects, ternary complexes were developed by coating pH-sensitive PEG-b-PLL-g-succinylsulfathiazole (hereafter abbreviated as PPSD) with DNA/PEI polyplexes via electrostatic interaction. PPSD can efficiently shield the surface charge of DNA/PEI. The gene transfection efficiency of ternary complexes was lower than that of DNA/PEI at pH 7.4; however, it recovered to the same level as that of DNA/PEI at pH 6.0, attributed to the pH-triggered release of DNA/PEI from ternary complexes. Cell uptake results also exhibited the same trend as transfection at different pH values. The suitable ability for pH-triggered shielding/deshielding estimated that PPSD demonstrates potential as a shielding system for use in in vivo gene delivery.

Synthesis of oligoethylenimine grafted net-poly(amino ester) and their application in gene delivery

Summary Novel net-polymers were prepared by grafting oligoethylenimine (OEI) on net-poly(amino ester) (N-P-g-OEI) and applied as gene carriers. N-P-g-OEI can effectively condense DNA to form small-sized complex particles (