• Asian Pacific Journal of Cancer Prevention
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• v.15 no.17
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• pp.7113-7118
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• 2014

Background: HSP90 may be overexpressed in cancer cells which are greatly dependent on Hsp90 function. Geldanamycin derivative 17 allylamino-17-demethoxygeldanamycin (17-AAG) inhibits the function and expression of HSP90. 17-AAG has poor water-solubility which is a potential problem for clinical practice. In this study for improving the stability and solubility of molecules in drug delivery systems we used a ${\beta}$-cyclodextrin-17AAG complex. Materials and Methods: To assess cytotoxic effects of ${\beta}$-cyclodextrin-17AAG complexes and free 17AAG, colorimetric cell viability (MTT) assays were performed. Cells were treated with equal concentrations of ${\beta}$-cyclodextrin- 17AAG complex and free 17AAG and Hsp90 gene expression levels in the two groups was compared by real-time PCR. Results: MTT assay confirmed that ${\beta}$-cyclodextrin- 17AAG complex enhanced 17AAG cytotoxicity and drug delivery in T47D breast cancer cells. The level of Hsp90 gene expression in cells treated with ${\beta}$-cyclodextrin- 17AAG complex was lower than that of cells treated with free 17AAG (P=0.001). Conclusions: The results demonstrated that ${\beta}$-cyclodextrin- 17AAG complexes are more effective than free 17AAG in down-regulating HSP90 expression due to enhanced ${\beta}$-cyclodextrin-17AAG uptake by cells. Therefore, ${\beta}$-cyclodextrin could be superior carrier for this kind of hydrophobic agent.

• Polymer(Korea)
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• v.38 no.1
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• pp.85-92
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• 2014

To develop water-soluble chitosan as an effient gene delivery carrier, chitosan oligosaccharides (COSs) with various molecular weights (MW) were studied for gene transfection agents. MWs of COSs fractionated by ultrafiltration techniques were identified as narrow MW distributions with the average MW ranging from 1 to 10 kDa through gel permeation chromatography (GPC) measurement depending on the applied ultrafiltration membranes. Their structural characterizations were analyzed by FTIR spectrophotometer and $^1H$ NMR. The degree of deacetylation was determined by UV spectroscopy showing the degree of deacetylation above 90%. The relative cell viabilities were maintained over 100% (10 mg/mL), independent of the MW of the fractionated COSs. The fractionated COSs of 10 mg/mL concentration with narrow MW distributions showed non-cytotoxicity in Caco-2 cells.

• Journal of Microbiology and Biotechnology
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• v.16 no.9
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• pp.1369-1376
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• 2006

Self-organized nanogels were prepared from pullulan/biotin conjugates (PU/Bio) for the development of an effective anticancer drug delivery system. The degree of biotin substitution was 11, 19, and 24 biotin groups per 100 anhydroglucose units of pullulan. The physicochemical properties of the nanogels (PU/Bio1, 2 and 3) in aqueous media were characterized by dynamic light scattering, transmission electron microscopy, and fluorescence spectroscopy. The mean diameter of all the samples was less than 300 nm with a unimodal size distribution. The critical aggregation concentrations (CACs) of the nanoparticles in distilled water were $2.8{\times}10^{-2},\;1.6{\times}10^{-2}$, and $0.7{\times}10^{-2}mg/ml$ for the PU/Bio1, 2, and 3, respectively. The aggregation behavior of the nanogels indicated that biotin can perform as a hydrophobic moiety. To observe the specific interaction with a hepatic carcinoma cell line (HepG2), the conjugates were labeled with rhodamine B isothiocyanate (RITC) and their intensities measured using a fluorescence microplate reader. The HepG2 cells treated with the fluorescence-labeled PU/Bio nanoparticles were strongly luminated compared with the control (pullulan). Confocal laser microscopy also confirmed internalization of the PU/Bio nanogels into the cancer cells. Such results demonstrated that the biotin in the conjugate acted as both a hydrophobic moiety for self-assembly and a tumor-targeting moiety for specific interaction with tumor cells. Consequently, PU/Bio nanogels would appear to be a useful drug carrier for the treatment of liver cancer.

• Journal of Microbiology and Biotechnology
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• v.28 no.3
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• pp.367-374
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• 2018

RNA interference provides an effective tool for developing antitumor therapies. Cell-penetrating peptides (CPPs) are delivery vectors widely used to efficiently transport small-interfering RNA (siRNA) to intracellular targets. In this study, we investigated the efficacy of the cancer-specific CPP carrier BR2 to specifically transport siRNA to cancer-target cells. Our results showed that BR2 formed a complex with anti-vascular endothelial growth factor siRNA (siVEGF) that exhibited the appropriate size and surface charge for in vivo treatment. Additionally, the BR2-VEGF siRNA complex exhibited significant serum stability and high levels of gene-silencing effects in vitro. Moreover, the transfection efficiency of the complex into a cancer cell line was higher than that observed in non-cancer cell lines, resulting in downregulated intracellular VEGF levels in HeLa cells and comprehensively improved antitumor efficacy in the absence of significant toxicity. These results indicated that BR2 has significant potential for the safe, efficient, and specific delivery of siRNA for diverse applications.

• Bulletin of the Korean Chemical Society
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• v.32 no.2
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• pp.651-655
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• 2011

We synthesized a novel cationic dendrimer consisting of a poly(amidoamine) dendrimer (PAMAM, generation 4) backbone with both L-arginine (Arg) at the termini and 6-aminohexanoic acid (Ahx) between the original core polymer and the peripheral Arg units. The sequential chemical modification of PAMAM G4 with Ahx and Arg resulted in higher transfection efficiency with much less cytotoxicity. PAMAM G4-Ahx-Arg formed stable polyplexes at weight ratios of 8:1 or higher (polymer: plasmid DNA), and the mean polyplex diameter was $180{\pm}20nm$. PAMAM G4-Ahx-Arg showed much higher transfection ability than PAMAM G4 or PAMAM G4-Ahx. Furthermore, PAMAM G4-Ahx-Arg was much less cytotoxic than PEI25KD and PAMAM G4-Arg. In addition to Arg grafting of the PAMAM dendrimer, which endows a higher transfection capability, the addition of Ahx spacer increased dendrimer hydrophobicity, introduced flexibility into the conjugated amino acids, and reduced cytotoxicity. Overall, it appears that the concomitant modification of PAMAM with Ahx and Arg could lead to new PAMAM conjugates with better performances.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.11
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• pp.6949-6953
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• 2013

Background: Nowadays, the encapsulation of cytotoxic chemotherapeutic agents is attracting interest as a method for drug delivery. We hypothesized that the efficiency of helenalin might be maximized by encapsulation in ${\beta}$-cyclodextrin nanoparticles. Helenalin, with a hydrophobic structure obtained from flowers of Arnica chamissonis and Arnica Montana, has anti-cancer and anti-inflammatory activity but low water solubility and bioavailability. ${\beta}$-Cyclodextrin (${\beta}$-CD) is a cyclic oligosaccharide comprising seven D-glucopyranoside units, linked through 1,4-glycosidic bonds. Materials and Methods: To test our hypothesis, we prepared ${\beta}$-cyclodextrin-helenalin complexes to determine their inhibitory effects on telomerase gene expression by real-time polymerase chain reaction (q-PCR) and cytotoxic effects by colorimetric cell viability (MTT) assay. Results: MTT assay showed that not only ${\beta}$-cyclodextrin has no cytotoxic effect on its own but also it demonstrated that ${\beta}$-cyclodextrin-helenalin complexes inhibited the growth of the T47D breast cancer cell line in a time and dose-dependent manner. Our q-PCR results showed that the expression of telomerase gene was effectively reduced as the concentration of ${\beta}$-cyclodextrin-helenalin complexes increased. Conclusions: ${\beta}$-Cyclodextrin-helenalin complexes exerted cytotoxic effects on T47D cells through down-regulation of telomerase expression and by enhancing Helenalin uptake by cells. Therefore, ${\beta}$-cyclodextrin could be superior carrier for this kind of hydrophobic agent.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.10
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• pp.4203-4209
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• 2015

Exosomes, membranous nanovesicles, naturally carry bio-macromolecules or miRNA and play impoetant roles in tumor pathogenesis. Here, we showed that macrophages cell-derived exosomes can function as vehicles to deliver exogenous miR-21 inhibitor into BGC-823 gastric cancer cells. Exosomes loaded with miR-21inhibitor significantly increased miR-21 levels in BGC-823, but miR-21inhibitor loaded in exosomes exerted an opposite effect. miRNA transfected with exosomes had less cellular toxicity to host cells compared to conventional transfection methods. The miR-21inhibitor loaded exosomes promoted the migration ability and reduced apoptosis of BGC-823 gastric cancer cells. These observations indicate that miR-21 acts as a tumor promoter by targeting the PDCD4 gene and preventing apoptosis of gastric cancer cells through inhibition of PDCD4 expression. Furthermore, exosome -mediated miR-21 inhibitor delivery resulted in functionally more efficient inhibition and less cellular toxicity compared to conventional transfection methods. Similar approaches could be useful in modification of target biomolecules in vitro and in vivo. These findings contribute to our understanding of the functions of miR-21 and exosomes as a carrier for therapy of gastric cancer.

• Journal of Veterinary Science
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• v.25 no.1
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• pp.4.1-4.14
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• 2024

Background: Lawsonia intracellularis is the causative agent of proliferative enteropathy and is associated with several outbreaks, causing substantial economic loss to the porcine industry. Objectives: In this study, we focused on demonstrating the protective effect in the mouse model through the immunological bases of two vaccine strains against porcine proliferative enteritis. Methods: We used live-attenuated Salmonella Typhimurium (ST) secreting two selected immunogenic LI antigens (Lawsonia autotransporter A epitopes and flagellin [FliC]-peptidoglycan-associated lipoprotein-FliC) as the vaccine carrier. The constructs were cloned into a Salmonella expression vector (pJHL65) and transformed into the ST strain (JOL912). The expression of immunogenic proteins within Salmonella was evaluated via immunoblotting. Results: Immunizing BALB/c mice orally and subcutaneously induced high levels of LI-specific systemic immunoglobulin G and mucosal secretory immunoglobulin A. In immunized mice, there was significant upregulation of interferon-γ and interleukin-4 cytokine mRNA and an increase in the subpopulations of cluster of differentiation (CD) 4⁺ and CD 8⁺ T lymphocytes upon splenocytes re-stimulation with LI antigens. We observed significant protection in C57BL/6 mice against challenge with 10^6.9 times the median tissue culture infectious dose of LI or 2 × 10⁹ colony-forming units of the virulent ST strain. Immunizing mice with either individual vaccine strains or co-mixture inhibited bacterial proliferation, with a marked reduction in the percentage of mice shedding Lawsonia in their feces. Conclusions: Salmonella-mediated LI gene delivery induces robust humoral and cellular immune reactions, leading to significant protection against LI and salmonellosis.

• The Korean Journal of Nuclear Medicine
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• v.38 no.3
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• pp.253-258
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• 2004

Purpose: Chitosan has been studied as a non-viral gene delivery vector, drug delivery carrier, metal chelator, food additive, and radiopharmaceutical, among other things. Recently, galactose-graft chitosan was studied as a non-viral gene and drug delivery vector to target hepatocytes. The aim of this study was to investigate the usefulness of nuclear imaging for in vivo evaluation of targeting the hepatocyte by galactose grafting. Methods and Materials: Galactosyl methylated chitosan (GMC) was produced by methylation to lactobionic acid coupled chitosan. Cytotoxicity of $^{99m}Tc$-GMC was determined by MTT assay. Rabbits were injected via their auricular vein with $^{99m}Tc$-GMC and $^{99m}Tc$-methylated chitosan (MC), the latter of which does not contain a galactose group, and images were acquired with a gamma camera equipped with a parallel hole collimator. The composition of the galactose group in galactosylated chitosan (GC), as well as the tri-, di-, or mono-methylation of GMC, was confirmed by NMR spectroscopy. Results: The results of MTT assay indicated that $^{99m}Tc$-GMC was non-toxic. $^{99m}Tc$-GMC specifically accumulated in the liver within 10 minutes of injection and maintained high hepatic uptake. In contrast, $^{99m}Tc$-MC showed faint liver uptake. $^{99m}Tc$-GMC scintigraphy of rabbits showed that the galactose ligand principally targeted the liver while the chitosan functionalities led to excretion through the urinary system. Conclusion: Bioconjugation with a specific ligand endows some degree of targetability to an administered molecule or drug, as in the case of galactose for hepatocyte in vivo, and evaluating said targetabililty is a clear example of the great benefit proffered by nuclear imaging.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.17
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• pp.7749-7754
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• 2015

Background: Currently, cationic liposome has become the commonly used vehicles for gene transfection. Furthermore, one of the most significant steps in microRNAs expression studies is transferring microRNAs into cell cultures successfully. In this study we aim to approach the feasibility of transfection of cervical cancer cell lines mediated by liposome and to obtain the optimized transfection condition for cervical cancer cell lines. Materials and Methods: $Lipofectamine^{TM}2000$ as the carrier, miR-101 mimic was transfected into Hela cells and Siha cells. Using green fluorescent protein as reporter gene, to set different groups according to cell seeding density, the amount of miRNA, miRNA and the proportion of Liposomes, Whether to add serum into medium to study their impact on the liposomal transfection efficiency. Finally, MTT assay was used to analyze the relative minimal cell toxicity of liposome reagents. Results: The seeding density of Hela cell line and Siha are $1.5{\times}10^4$ (per well of 24 well plates), miRNA amount is 1ul of both, the ratio of miRNA and liposome is 1:0.5 of Hela cell line; 1:0.7 of Siha cell line respectively, after 24 hours we can get the highest transfection efficiency. Compared with serum medium, only Siha cells cultured with serum-free medium obtained higher transfection efficiency before transfection (P<0.01). MTT assay showed that according to the above conditions which has the lowest cytotoxicity. Conclusions: The method of Liposome to transfected is a suitable way and it can be an efficient reagent for miRNA delivery for Hela cells and Siha cells in vitro. It may serve as a reference for the further research or application.