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To develop a carrier for targeted drug delivery and triggered drug release in a reducing-tumor environment, reduction-sensitive hyaluronic acid-g-stearic acid (HCS) micelles were synthesized using a coupling agent. The HCS 40% was shown to have a more compact particle size than HCS 20% and the particle size of doxorubicin (DOX)-loaded HCS (HCSD) was increased relative to that of HCS micelles. The behavior of DOX release from HCSD showed that DOX was rapidly released in GSH (10 mM) solution. The site-specific targeting effect of HCSD nanoparticles was investigated by cellular uptake and competition assay at HCT116 and CT26 cell lines. An in vivo study of HCSD revealed that tumor suppression and site-specific targeted delivery of HCSD nanoparticles in HCT116-xenografted tumors were more superb than in the CT26-xenografted tumor. These results suggest that HCSD nanoparticles can be expected to have high therapeutic efficacy because they enable targeted drug delivery and rapid drug release., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Branched polyethylenimine (bPEI) was well known as high transfection agent, which has many amine group. However, utilization of bPEI was limited due to high toxicity. To solve these problems, bPEI was introduced to low molecular weight water-soluble chitosan (LMWSC) with coupling agent. In addition, hyaluronic acid (HA), one of natural anion polymer, was introduced to binary complex of pDNA/bPEI-grafted LMWSC (LMPEI) to target the specific cancer cell and impart the serum resistant. Ternary complexes of pDNA/LMPEI/HA were prepared by electrostatic charge interaction and their binding affinity and DNase protection assay were conducted by gel retardation assay. Particle size of ternary complexes showed that had each 482 ± 245.4 (pDNA/LMPEI2%/HA, 1:16:1, w/w/w) and 410 ± 78.5 nm (pDNA/LMPEI4%/HA, 1:16:2, w/w/w). Moreover, to demonstrate serum-resistant effect of ternary complexes, particle size of them was measured according to incubated time (0-10 h) under serum condition. Transfection assay of ternary complexes showed that their transfection efficiency in CD44-receptor overexpressed HCT116 cell was higher than CD44-receptor negative CT26 cell. Additionally, intracellular uptake of ternary complexes with propidium iodide (PI)-labeled pDNA was observed to confirm targeting effect and cellular internalization by fluorescence microscopy. These results suggest that ternary complexes are superb gene carrier with excellent serum-resistant and high gene transfection., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Transferrin/PEG/O-carboxymethyl chitosan/fatty acid/paclitaxel (TPOCFP) micelles were tested for suitability as a drug carrier characterized by low cytotoxicity, sustained release, high cellular uptake, and site-specific targeted delivery of hydrophobic drugs. Characterization, drug content, encapsulation efficiency, and in vitro drug release were investigated. When the feeding amount of paclitaxel (PTX) was increased, the drug content increased, but loading efficiency decreased. TPOCFP micelles had a spherical shape, with a particle size of approximately 140-649 nm. In vitro cell cytotoxicity and hemolysis assays were conducted to confirm the safety of the micelles. Anticancer activity and confocal laser scanning microscopy (CLSM) were used to confirm the targeting efficiency of target ligand-modified TPOCFP micelles. Anticancer activity and CLSM results clearly demonstrated that transferrin-modified TPOCFP micelles were quickly taken up by the cell. The endocytic pathway of TPOCFP micelles was analyzed by flow cytometry, revealing transfection via receptor-mediated endocytosis. These results suggest that PTX-encapsulated TPOCFP micelles may be used as an effective cancer-targeting drug delivery system for chemotherapy., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The aim of this study is to prepare delivery vehicles of paclitaxel using low molecular weight water-soluble chitosan (LMWSC) and evaluate them as an anticancer drug delivery system. LMWSC was modified with methoxy polyethylene glycol (LMWSC-MPEG, ChitoPEG), and then it was conjugated with cholesterol (LMWSC-MPEG-Chol). Core-shell type LMWSC-MPEG-Chol nanoparticles (LMWSC-NPs) were prepared by the dialysis method, and the core-shell structure was confirmed by 1H NMR analysis. To this polymer, paclitaxel was encapsulated and core-shell type nanoparticles were prepared. The release tests indicated that release of paclitaxel from the core-shell type nanoparticles and its transport across the dialysis membrane was slower than dialysis of free paclitaxel. In a cytotoxicity study using CT26 cell, the paclitaxel-encapsulated core-shell type nanoparticles (LMWSC-NPs) showed a toxicity against tumor cells similar to paclitaxel itself. The results of a tumor inhibition test with CT26 implanted upon mouse tumor models in vivo indicated that the application of a dose of 10 mg/kg of LMWSC-NPT showed a superior survival rate, and a slower tumor growth than when paclitaxel alone was administered, although the tumor growth and survival rate were not significantly changed at a dose of 2 mg/kg. The LMWSC-NPT dose above 10mg/kg showed a superior antitumor activity., (Copyright © 2010 Elsevier B.V. All rights reserved.)

This study is aimed to develop an efficient oral vaccine carrier which specifically targets the follicle-associated epithelium region of Peyer's patch (PP). M cell-homing peptide was selected by the phase display technique and its targeting efficiency was validated using chitosan nanoparticles (CNs) conjugated with the discovered peptide. A phage clone encoding CKSTHPLSC (CKS9) peptide sequence was selected by analysis of comparative superiority in transcytosis efficacy across the M cell layer in vitro and in vivo among the candidates. CKS9 was chemically conjugated to water-soluble chitosan (WSC) and the CKS9-immobilized chitosan nanoparticles (CKS9-CNs) were prepared by ionic gelation of CKS9-WSC with tripolyphosphate, yielding spherical nanoparticles around 226.2 +/- 41.9 nm. The targeting ability of CKS9-CNs to the M cell and to the PP regions of rat small intestine was investigated by in vitro transcytosis assay and closed ileal loop assay, respectively, and was visualized by fluorescence-microscopy analysis. CKS9-CNs were transported more effectively across the M cell model and accumulated more specifically into PP regions in comparison with CNs, indicating that CKS9 peptide prominently enhanced the targeting and transcytosis ability of CNs to PP regions. These results suggest that the CKS9-CNs could be used as a new carrier for oral vaccine delivery., (Copyright 2010 Elsevier Ltd. All rights reserved.)

The aim of this research is to develop a novel branched polyethylenimine (PEI)-like polycation as a potential gene carrier with high gene transfection efficiency and low toxicity. In particular, alpha,beta-poly(l-aspartate-graft-PEI) (Asp-g-PEI), a pseudo-branched PEI, was synthesized by the ring-opening reaction of poly(l-succinimide) (PSI) with low molecular weight branched PEI (LMW PEI, MW=600 and 1200). Good plasmid condensation and protection ability of Asp-g-PEI were confirmed by agarose gel electrophoresis assay. Asp-g-PEI/DNA complexes showed high positive zeta potential, narrow size distribution, good dispersity and a compact spherical shape with size below 250nm when the N/P ratio was above 5, suggesting that they can be endocytosed. Cytotoxicity of Asp-g-PEI/DNA complexes was rather lower than that of PEI25K/DNA complexes, especially at high N/P ratio. The most efficient gene transfection of Asp-g-PEI/DNA complexes was similar or a little higher than that of PEI25K in 293T, HeLa and HepG2 cell lines, while almost 4 and 6 times higher than that of parent PEI1200 and PEI600, respectively, in HeLa cell line; as the molecular weight of parent PEI in Asp-g-PEI was increased from 600 to 1200, the transfection efficiency showed a tendency to decrease. The mechanism of Asp-g-PEI-mediated gene transfection was attributed to the "proton sponge effect" due to PEI in the copolymer.

The aim of this study was to develop a novel polymeric magnetic nanoprobe as an MRI contrast agent to target hepatocytes, as well as to evaluate the targeting ability of the nanoprobe with MRI in vivo. Superparamagnetic iron oxide nanocrystals (SPIONs) were synthesized by a thermal decomposition and seed growth method. An 1-ethyl-3-(3-(dimethylamino)-propyl) carbodiimide (EDC)-mediated reaction coupled water-soluble chitosan (WSC) to linoleic acid (LA). Twelve-nanometer-sized SPIONs were incorporated into the core of self-assembled WSC-LA nanoparticles. The morphology and size distribution of the SPION-loaded WSC-LA nanoparticles (SCLNs) were determined by transmittance electron microscopy (TEM) and dynamic light scattering (DLS), respectively. The encapsulation of SPIONs in the WSC-LA nanoparticles reduced the cytotoxicity of bare iron particles and enhanced their dispersion ability in water. The clustering of SPIONs into WSC-LA nanoparticles showed ultrasensitive magnetic behavior. After in vivo intravascular SCLN injection, MRI revealed relative signal enhancement in the liver. The localization of SCLN in hepatocytes was confirmed by Prussian blue staining and TEM analysis. We have successfully developed an ultrasensitive SCLN that effectively targets hepatocytes. The SCLN can be used as a contrast agent to aid in the diagnosis of hepatic diseases.

The multiblock copolymer composed of poly(gamma-benzyl L-glutamate) (PBLG) and poly(ethylene oxide) (PEO) was synthesized to prepare polymeric micelles as an anticancer drug carrier. Adriamycin (ADR) used as an anticancer drug was incorporated into the polymeric micelles prepared by the multiblock copolymer. The higher the drug feeding ratio, the higher the drug loading contents and the lower the drug loading efficiency. The increased drug feeding ratio resulted in increased particle sizes. At all of the formulations, particle sizes were less than 150 nm. The particles were observed as spherical shapes. ADR release from ADR-loaded polymeric micelles in vitro was decreased with an increased drug loading contents. In in vitro antitumor activity test using CT 26 tumor cells, polymeric micelles showed almost similar cytotoxicity when compared to ADR itself while polymeric micelles themselves did not affect cytotoxicity. In in vivo antitumor activity test using mice tumor xenograft model, the polymeric micelles showed improved survivability of mice with minimized weight changes and excellent tumor growth suppression efficacy. Polymeric micelles of the multiblock copolymer suggested to be a good candidate for anticancer drug delivery carrier.

Chitosan and chitosan derivatives have been proposed as alternative and biocompatible cationic polymers for non-viral gene delivery. However, the low transfection efficiency and low specificity of chitosan is an aspect of this approach that must be addressed prior to any clinical applications. In the present study a chitosan derivative, galactosylated poly(ethylene glycol)-chitosan-graft-polyethylenimine (Gal-PEG-CHI-g-PEI), was investigated as a potential hepatocyte-targeting gene carrier. The composition of Gal-PEG-CHI-g-PEI was characterized using (1)H nuclear magnetic resonance ((1)H NMR), and the particle size and zeta potential of Gal-PEG-CHI-g-PEI/DNA complexes were measured using dynamic light scattering (DLS). The Gal-PEG-CHI-g-PEI exhibited lower cytotoxicity compared to PEI 25K as a control. Likewise, Gal-PEG-CHI-g-PEI/DNA complexes showed good hepatocyte specificity. Furthermore, Gal-PEG-CHI-g-PEI/DNA complexes transfected liver cells more effectively than PEI 25K in vivo after intravenous (i.v.) administration. Together, these results suggest that Gal-PEG-CHI-g-PEI, which has improved transfection efficiency and hepatocyte specificity both in vitro and in vivo, may be useful for gene therapy.

The aim of this study was to prepare ciprofloxacin HCl (CIP)-encapsulated poly(dl-lactide-co-glycolide) (PLGA) copolymer nanoparticles and its antibacterial potential was evaluated with pathogenic bacteria, Escherichia coli (E. coli), in vitro and in vivo. CIP-encapsulated nanoparticles of PLGA were prepared by multiple emulsion solvent evaporation method. PLGA nanoparticles showed spherical shapes with particle sizes around 100-300 nm. Loading efficiency was lower than 50% (w/w) because of water-solubility properties of CIP. At drug release study, CIP showed initial burst effect for 12 h and then continuously released for 2 weeks. At in vitro antibacterial activity test, CIP-encapsulated nanoparticles showed relatively lower antibacterial activity compared to free CIP due to the sustained release characteristics of nanoparticles. However, CIP-encapsulated PLGA nanoparticles (doses: 25 mg CIP/kg of mice) effectively inhibited the growth of bacteria due to the sustained release characteristics of nanoparticles, while free CIP was less effective on the inhibition of bacterial growth. These results indicated that CIP-encapsulated PLGA nanoparticles have superior effectiveness to inhibit the growth of bacteria in vivo.

Chitosan has been investigated as a non-viral vector because it has several advantages such as biocompatibility, biodegradability and low toxicity with high cationic potential. However, the low specificity and low transfection efficiency of chitosan need to be solved prior to clinical application. In this paper, we focused on the galactose or mannose ligand modification of chitosan for enhancement of cell specificity and transfection efficiency via receptor-mediated endocytosis in vitro and in vivo.

To develop chitosan based efficient gene carriers, highly purified chitosan oligosaccharides (COSs) were chemically modified with deoxycholic acid (DOCA). Owing to the amphiphilic characters, the DOCA-conjugated COSs (COSDs) formed self-aggregated nanoparticles in aqueous milieu. The physicochemical characterization revealed that the particle size of the nanoparticles was in the range of 200 approximately 240 nm and the critical aggregation concentration (cacs) was 0.012 approximately 0.046 g/L, depending on the degree of substitution (DS). As efficient gene carriers, the COSD nanoparticles showed superior gene condensation and protection of condensed gene from endonuclease attack than unmodified COSs. Furthermore, COSDs showed great potential for gene carrier with the high level of gene transfection efficiencies, even in the presence of serum. Considered with the negligible cytotoxic effects, DOCA-modified chitosan oligosaccharides can be considered as potential candidates for efficient non-viral gene carriers.

Glucosylated polyethylenimine (GPEI) was synthesized as a tumor-targeting gene carrier through facilitative glucose metabolism by tumor glucose transporter. Particle sizes of GPEI/DNA complex increased in proportion to glucose content of GPEI, whereas surface charge of the complex was not dependent on glucosylation, partially due to inefficient shielding of the short hydrophilic group introduced. GPEI with higher glucosylation (36 mol-%) had no cytotoxic effect on cells even at polymer concentrations higher than 200 microg/mL. Compared to unglucosylated PEI, glucosylation induced less than one-order decrease of transfection efficiency. Transfection of GPEI/DNA complex into tumor cells possibly occurred through specific interaction between glucose-related cell receptors and glucose moiety of GPEI. Gamma imaging technique revealed GPEI/DNA complex was distributed in liver, spleen, and tumors.

Water-soluble chitosan (WSC) was used to increase the stability of chitosan in water and decrease the cytotoxicity induced by acetic acid. Lactobionic acid (LA) bearing galactose group was coupled with WSC for hepatocytes specificity. The composition of galactose in galactosylated chitosan (GC) was determined by NMR spectroscopy. The GC was complexed with plasmid DNA in various GC/DNA (N/P) charge ratios and the resulting complex was characterized by dynamic light scattering, gel retardation, and turbidity to determine the particle sizes, complex formation, and complex stability, respectively. Cytotoxicity and transfection efficiency of GC were also studied in cultured HepG2 human hepatoblastoma cell line and HeLa human cervix epithelial carcinoma cells. The complete GC/DNA complex was formed at the charge ratio of 5 and the GC/DNA complex to DNase I resistance was obtained. Particle sizes decreased with increasing charge ratio of GC to DNA and had a minimum value around 120 nm at the charge ratio of 5. And no significant difference in particle sizes from the charge ratio of 5-20 was found. The suspension of GC/DNA complexes exhibited no significant change in turbidity at the charge ratios of 10, indicating the complete shielding of DNA charge. Cytotoxicity study showed that GC prepared by the water-soluble chitosan had no cytotoxic effects on cells. And transfection efficiency into HepG2, which has asialoglycoprotein receptors (ASGP-R), was higher than that into HeLa without ASGP-R.