Chung Kil Song

Epigallocatechin-3-Gallate Enhances CD8+ T Cell–Mediated Antitumor Immunity Induced by DNA Vaccination

Immunotherapy and chemotherapy are generally effective against small tumors in animal models of cancer. However, these treatment regimens are generally ineffective against large, bulky tumors. We have found that a multimodality treatment regimen using DNA vaccination in combination with chemotherapeutic agent epigallocatechin-3-gallate (EGCG), a compound found in green tea, is effective in inhibiting large tumor growth. EGCG was found to induce tumor cellular apoptosis in a dose-dependent manner. The combination of EGCG and DNA vaccination led to an enhanced tumor-specific T-cell immune response and enhanced antitumor effects, resulting in a higher cure rate than either immunotherapy or EGCG alone. In addition, combined DNA vaccination and oral EGCG treatment provided long-term antitumor protection in cured mice. Cured animals rejected a challenge of E7-expressing tumors, such as TC-1 and B16E7, but not a challenge of B16 7 weeks after the combined treatment, showing antigen-specific immune responses. These results suggest that multimodality treatment strategies, such as combining immunotherapy with a tumor-killing cancer drug, may be a more effective anticancer strategy than single-modality treatments.

A novel vesicular carrier, transethosome, for enhanced skin delivery of voriconazole: Characterization and in vitro/in vivo evaluation

This study describes a novel carrier, transethosome, for enhanced skin delivery of voriconazole. Transethosomes (TELs) are composed of phospholipid, ethanol, water and edge activator (surfactants) or permeation enhancer (oleic acid). Characterization of the TELs was based on results from recovery, particle size, transmission electron microscopy (TEM), zeta potential and elasticity studies. In addition, skin permeation profile was obtained using static vertical diffusion Franz cells and hairless mouse skin treated with TELs containing 0.3% (w/w) voriconazole, and compared with those of ethosomes (ELs), deformable liposomes (DLs), conventional liposomes (CLs) and control (polyethylene glycol, PG) solutions. The recovery of the studied vesicles was above 90% in all vesicles, as all of them contained ethanol (7-30%). There was no significant difference in the particles size of all vesicles. The TEM study revealed that the TELs were in irregular spherical shape, implying higher fluidity due to perturbed lipid bilayer compared to that of other vesicles which were of spherical shape. The zeta potential of vesicles containing sodium taurocholate or oleic acid showed higher negative value compared to other vesicles. The elasticities of ELs and TELs were much higher than that of CLs and DLs. Moreover, TELs dramatically enhanced the skin permeation of voriconazole compared to the control and other vesicles (p<0.05). Moreover, the TELs enhanced both in vitro and in vivo skin deposition of voriconazole in the dermis/epidermis region compared to DLs, CLs and control. Therefore, based on the current study, the novel carrier TELs could serve as an effective dermal delivery for voriconazole.

Disaccharide-modified liposomes and their in vitro intracellular uptake

Sterically stabilized liposomes (SSL) were known to be accumulated passively in cancer due to the effect of enhanced permeability and retention (EPR). However, drug delivery via SSL to cancer seemed to show an insufficient improvement of chemotherapeutic efficacy. Herein, carbohydrate-binding proteins (lectins) of cell surface, which express on the plasmic membrane of many malignant cells, can be a good model of surface-modified liposomes. In this study, we investigated the in vitro characteristics of liposomes of which the surface was modified with a disaccharide molecule, sucrose or maltose. The disaccharide-modified lipids such as sucrose-modified lipid and maltose-modified lipid, in which the disaccharide was conjugated to the one end of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-(polyethylene glycol)-2000 (DSPE-PEG2000), was synthesized. The disaccharide-modified liposomes were prepared by thin film-hydration method and then doxorubicin (DOX), an anticancer drug, was loaded to the prepared liposomes by the remote loading method with ammonium ion gradient. Flow cytometry and confocal microscopy analyses showed that the disaccharide-modified liposomes enhanced the intracellular uptake of liposomes into various cancer cell lines via lectin-mediated endocytosis. The disaccharide-modified liposomes in which DOX was loaded inside of liposomes exhibited higher cytotoxicity against various cancer cells than DOX-loaded SSL did. These results suggest that disaccharide-modified liposomes may be promising cancer targeting carriers which can enhance intracellular uptake and cytotoxicity of the drug-loaded liposomes via lectin-mediated endocytosis.

Microemulsion-based Hydrogel Formulation of Itraconazole for Topical Delivery

The present study was aimed at preparing microemulsion-based hydrogel (MBH) for the skin delivery of itraconazole. Microemulsion prepared with Transcutol as a surfactant, benzyl alcohol as an oil and the mixture of ethanol and phasphatidyl choline (3:2) as a cosurfactant were characterized by solubility, phase diagram, particle size. MBHs were prepared using 0.7 % of xanthan gum (F1-1) or carbopol 940 (F1-2) as gelling agents and characterized by viscosity studies. The in vitro permeation data obtained by using the Franz diffusion cells and hairless mouse skin showed that the optimized microemulsion (F1) consisting of itraconazole (1% w/w), benzyl alcohol (10% w/w), Transcutol (10% w/w) and the mixture of ethanol and phospahtidylcholine (3:2) (10% w/w) and water (49% w/w) showed significant difference in the flux (~1 µg/ cm 2 /h) with their corresponding MBHs (0.25-0.64 µg/cm 2 /h). However, the in vitro skin drug content showed no significant difference between F1 and F1-1, while F1-2 showed significantly low skin drug content. The effect of the amount of drug loading (0.02, 1 and 1.5% w/w) on the optimized MBH (F1-2) showed that the permeation and skin drug content increased with higher drug loading (1.5%). The in vivo study of the optimized MBH (F1-2 with1.5% w/w drug loading) showed that this formulation could be used as a potential topical formulation for itraconazole. Key wordsItraconazole, Benzyl alcohol, Microemulsion-based gel, Topical skin delivery

Enhanced in vitro cellular uptake of P-gp substrate by poloxamer-modified liposomes (PMLs) in MDR cancer cells

Poloxamer-modified liposomes (PMLs) were prepared using poloxamers (P85 and F68) by the thin-film hydration method for overcoming the multidrug resistance and thereby enhancing the intracellular uptake of specific substrates of P-gp, rhodamine 123 (R123). The prepared liposomes, plain liposomes (PLs) and PMLs, were characterized by particle size, zeta potential and drug entrapment efficiency, and assessed by in vitro cellular uptake using KB and KBV20C (P-gp over-expression cell line) cells. The transmission electron microscopy study revealed the spherical shape of the prepared liposomes. No significant difference was observed between the PMLs and liposome without poloxamer (PLs) in the particle size (∼160 nm) and zeta potential (∼−5 mV). The in vitro cellular uptake study showed that P85-modified liposomes (PML-P85) significantly increased the internalization of R123 in MDR tumour cells. Our results showed that PML-P85 could be an effective carrier for anticancer drugs in MDR cancer therapy.

Solid Dispersions as a Drug Delivery System

ABSTRACT − Solid dispersion, defined as the dispersion of one or more active ingredient in a carrier or matrix at solidstate, is an efficient strategy for improving dissolution of poorly water-soluble drugs for enhancement of their bioavailabilit y.Compared to other conventional formulations such as tablets or capsules, solid dispersion which can be prepared by variousmethods has many advantages. However, despite numerous studies which have been carried out, limitations for com-mercializing these products remain to be solved. For example, during the manufacturing process or storage, amorphous formof solid dispersion can be converted into crystalline form. That is, the dissolution rate of solid dispersion would continuouslydecrease during storage, resulting in a product of no value. To resolve these problems, studies have been conducted on theeffects of excipients. In fact, modification of the solid dispersions to overcome these disadvantages has progressed from thefirst generation to the recent third generation products. In this review, an overview on solid dispersions in general will begiven with emphasis on the various manufacturing processes which include the use of polymers and on the stabilizationstrategies which include methods to prevent crystallization.Key words

In vitro and in vivo evaluation of N,N,N-trimethylphytosphingosine-iodide (TMP) in liposomes for the treatment of angiogenesis and metastasis

Phytosphingosine and methyl derivatives are important mediators on cellular processes, and are associated with cell growth and death. The antitumor activity of N,N,N-trimethylphytosphingosine-iodide (TMP) as a novel potent inhibitor of angiogenesis and metastasis was evaluated in B16F10 murine melanoma cells. The results indicated that TMP itself effectively inhibited in vitro cell migration, tube formation, and the expression of angiogenic factors as well as in vivo lung metastasis. However, TMP slightly suppressed in vivo experimental tumor metastasis in its free form and induced side effects including hemolysis and local side effects. Therefore, in an attempt to reduce the toxicity and the undesirable side effects of TMP, a liposomal formulation was prepared and tested for its effectiveness. TMP liposomes retained the effectiveness of TMP in vitro while side effects were reduced, and both in vivo experimental and spontaneous tumor metastasis were significantly suppressed. These results support the conclusion that TMP effectively inhibits in vitro angiogenesis as well as in vivo metastasis, and a liposomal formulation is more efficient delivery system for TMP treatment than solution.

Poloxamer-based solid dispersions for oral delivery of docetaxel: Differential effects of F68 and P85 on oral docetaxel bioavailability.

Development of an oral docetaxel formulation has been hindered mainly due to its poor solubility and oral bioavailability. The aim of this study was to develop poloxamer F68/P85-based solid dispersions (SDs) for the oral delivery of docetaxel and investigate their in vivo pharmacokinetic impacts on the systemic absorption of docetaxel given orally, in comparison with a SD based on F68 alone. The F68 and/or P85-based docetaxel SDs were prepared with varying the contents of poloxamers and then evaluated in terms of morphology, crystallinity, solubility, dissolution, permeation across rat intestinal segments, and oral pharmacokinetics in rats. As a result, the SDs successfully changed the crystalline properties of docetaxel and enhanced the drug solubility and dissolution. The SD prepared with F68 alone significantly enhanced the dissolution but not intestinal permeation of docetaxel, leading to only limited enhancement of oral bioavailability (1.39-fold increase). Notably, however, the F68/P85-based SD significantly enhanced both the dissolution and intestinal permeation of docetaxel, achieving a markedly improved oral bioavailability (2.97-fold increase). Therefore, the present results suggest that the intestinal permeation factor should be taken into account when designing SD formulations for the oral delivery of BCS class IV drugs including docetaxel, and that P85 could serve as a potential formulation excipient for enhancing the intestinal permeation of docetaxel.

AAPE proliposomes for topical atopic dermatitis treatment

Abstract Context: Anti-inflammatory effect of advanced adipose stem cell derived protein extract (AAPE) could be improved by minimising protein degradation. Objective: To develop a proliposomal formulation of AAPE for the treatment of topical atopic dermatitis. Materials and methods: Proliposomal powder was manufactured by evaporating a solution of soy phosphatidyl choline, AAPE and Poloxamer 407 in ethanol under vacuum on sorbitol powder. Characterisation of proliposomes (zeta potential, diameter, stability and flowability) as well as in vivo efficacy in a dermatitis mouse model was investigated. Results and discussion: Reconstitution of the proliposomal powder formed liposomes of 589 ± 3.6 nm diameter with zeta potential of −51.33 ± 0.36 mV. Protein stability was maintained up to 90 days at 25 °C as proliposomes. In vivo studies on atopic dermatitis mouse model showed a significant reduction in IgE levels after topical AAPE proliposome treatment. Conclusion: AAPE proliposomes maintained protein stability and showed promising results for atopic dermatitis treatment.

Abstract 3228: Preparation and in vivo evaluation of a phytosphingosine derivative, N,N,N-Trimethylphytosphingosine (TMP), in liposomes for the treatment of tumor metastasis

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Purpose: The aim of this study is to evaluate the cytotoxicity and anti-metastatic effect of a phytosphingosine derivative, N,N,N-trimethylphytosphingosine.iodide (TMP-I), and to compare it with that of TMP-I liposome. Background: Sphingolipids are known for ortho-signaling and anti-adhesion features as well as intracellular signal messenger in cell differentiation, proliferation and apoptosis. Sphingosine and sphinganine, structural analogs of sphingolipids were recognized as negative modulator of transmembrane signaling through protein kinase C (PKC) that plays an important role in the cancer treatment. Among them, phytosphingosine has been known for antitumor and anti-invasive effect through phytosphingosine-induced apoptosis mechanism. Recently it has been reported that N-monomethylphytosphingosine and N,N-dimethylphytosphingosine induced apoptosis via mitochondria-involved apoptosis such as caspase-8, caspase-3 and caspase-9 and this prompt us for this study. Methods: TMP-I solution was evaluated for cytotoxicity in various cancer cell lines and hemolysis in rat blood cells at various concentrations. The prepared TMP-I liposomes (DPPC:Chol:TMP-I at 5:5:1 mole ratio) were characterized by particle size and studied for in vitro cytotoxicity, cell migration and in vivo angiogenesis in rats. Furthermore, to evaluate the anti-metastatic effect of TMP-I liposomes, C57BL6 mice (n=5) were inoculated with B16F10 cells in tail vein for direct metastasis and footpad for spontaneous metastasis studies. Results: The prepared TMP-I liposomes were with a mean size of 150 nm. TMP-I solution showed significantly higher cytotoxicity and hemolysis (≥100 μg/ml) compared to that of TMP-I liposomes in several cancer cell lines. TMP-I liposomes showed significantly low cell migration and angiogenesis compared to control formulations. The in vivo study in mice showed that the numbers of lung nodules in the mice treated with TMP-I liposomes were significantly lower compared to that of control formulations. Conclusion: Our results suggested that the TMP-I liposome could be used as a potential carrier for the treatment of tumor metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3228. doi:10.1158/1538-7445.AM2011-3228