Polyethylenimine (PEI) is a class of cationic polymers proven to be effective for gene delivery. However, PEI is nondegradable and the molecular weight of PEI affects the cytotoxicity and gene transfer activity. Aiming to prepare a biodegradable gene vector with high transfection efficiency and low cytotoxicity, we conjugated low molecular weight (LMW) PEIs to the biodegradable backbone polyglutamic acids derivative (PEG-b-PBLG) by aminolysis to form PEIs combined PEG-b-PLG-g-PEIs (GGI). Two copolymers, GGI 30 and GGI 40, were synthesized. The chemistry of GGI was characterized using IR, 1H NMR and 13C NMR, GPC, and CD, respectively. The degradation behaviors of copolymer GGI in papain solution were investigated. GGIs showed good DNA condensation ability and high protection of DNA from nuclease degradation. The zeta potential of the GGI/pDNA polyplexes was approximately 15 mV, and the particle size was in the range 102-138 nm at N/P ratios between 10 and 30. The particle size and the morphology of the polyplex was further confirmed by transmission electron microscope (TEM). In cytotoxicity assay, GGIs were significantly less toxic than PEI 25k. The degradation product of GGI exhibited negligible effects on cells even at high copolymer concentration. The results of GFP flow cytometry and fluorescence imaging showed that the trasnfection efficiencies of GGIs were all markedly higher than PEI 25k in Hela, HepG2, Bel 7402, and 293 cell lines. Importantly, the presence of serum had a lower inhibitive effect on the transfection activity of GGI in comparison to PEI 25k and Lipofectamine 2000. Therefore, PEG-b-PLG-g-PEI copolymers may be attractive cationic polymers for nonviral gene therapy.

We have developed a nanocarrier drug-delivery system based on micelles formed by a new class of well-defined linear PEGylated two-arm oligomer of cholic acids in aqueous solution. By varying the length of the linear PEG chains and the configuration of cholic acid oligomer, one can easily finetune the physicochemical properties of the amphiphilic polymers and the resulting micelles. These include particle size, critical micelle concentration, and drug loading capacity. High level of hydrophobic anticancer drugs such as PTX, etoposide and SN-38 can be readily loaded into such nanocarriers. The loading capacity of the nanocarrier for PTX (PTX) is extremely high (12.0 mg/ mL), which is equivalent to 37.5% (w/w) of the total mass of the micelle. PTX-loaded nanocarriers are much more stable than Abraxane® (PTX/human serum albumin nanoaggregate) when stored in bovine serum albumin solution or dog plasma. PTX release profile from the micelles is burst-free and sustained over a period of seven days. The anti-tumor activity of PTX-loaded nanocarriers against ovarian cancer cell line in vitro, with continuous drug exposure, is similar to Taxol® (formulation of PTX dissolved in Cremophor EL and ethanol) or Abraxane®. Targeted drug delivery to tumor site with these novel micelles was demonstrated by near infrared fluorescence (NIRF) imaging in nude mice bearing ovarian cancer xenograft. Furthermore, PTX-loaded nanocarriers demonstrated superior anti-tumor efficacy compared to Taxol® at equivalent PTX dose in ovarian cancer xenograft model.

An ideal gene carrier is required both in safety and efficiency for transfection. Polyethylenimine (PEI), a well-studied cationic polymer, has been proved with high transfection efficiency, but is reported as toxicity in many cell lines. In this study, PEI was coupled with polyethylene glycol (PEG) to reduce its cytotoxicity. PEG-PEI copolymers were synthesized with isoporon diisocyanate (IPDI) in two steps. A set of PEG-PEI with different PEG molecular weights (MWs) and amounts of PEG were synthesized. The molecular structure of the resulting copolymers was evaluated by nuclear magnetic resonance spectroscopy ((1)H NMR), infrared spectroscopy (IR), and gel permeation chromatography (GPC), all of which had successfully verified formation of the copolymers. The particle size and zeta potential of polymer/DNA complexes were measured, and their cytotoxicity and transfection efficiency in Hela cells were evaluated. We found that the copolymer block structure significantly influenced not only the physicochemical properties of complexes, but also their cytotoxicity and transfection efficiency. PEG (5 kDa) significantly reduced the diameter of the spherical complexes. The zeta potential of complexes was reduced with increasing amount of PEG grafting. Cytotoxicity was dependent not on PEG MW but on the amount of PEG grafting. Copolymer PEG-PEI (2-25-1) with 1.89 PEG (2 kDa) was proved to be more efficient for in vitro gene transfer. In conclusion, PEG MW and the degree of PEGylation were found to significantly influence the biological activity of PEG-PEI/DNA complexes. These results provide new sights into the studies using block copolymer as gene delivery systems.

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Abstract. In the present study, we developed novel insulin-loaded hyaluronic acid (HA) nanoparticles for insulin delivery. The insulin-loaded HA nanoparticles were prepared by reverse-emulsion-freeze-drying method. This method led to a homogenous population of small HA nanoparticles with average size of 182.2 nm and achieved high insulin entrapment efficiencies (approximately 95%). The pH-sensitive HA nanoparticles as an oral delivery carrier showed advantages in protecting insulin against the strongly acidic environment of the stomach, and not destroying the junction integrity of epithelial cells which promise long-term safety for chronic insulin treatment. The results of transport experiments suggested that insulinloaded HA nanoparticles were transported across Caco-2 cell monolayers mainly via transcellular pathway and their apparent permeability coefficient from apical to basolateral had more than twofold increase compared with insulin solution. The efflux ratio of P app (B to A) to P app (A to B) less than 1 demonstrated that HA nanoparticle-mediated transport of insulin across Caco-2 cell monolayers underwent active transport. The results of permeability through the rat small intestine confirmed that HA nanoparticles significantly enhanced insulin transport through the duodenum and ileum. Diabetic rats treated with oral insulin-loaded HA nanoparticles also showed stronger hypoglycemic effects than insulin solution. Therefore, these HA nanoparticles could be a promising candidate for oral insulin delivery.