Sungpil Cho

Preparation and characterization of glycol chitin as a new thermogelling polymer for biomedical applications.

In this study, a new thermo-sensitive polymer, glycol chitin, was synthesized by controlled N-acetylation of glycol chitosan and evaluated as a thermogelling system. The physico-chemical properties of glycol chitins with different degrees of acetylation (DA) were investigated in terms of degradation, cytotoxicity, rheological properties, and in vitro and in vivo gel formation. Aqueous solutions of glycol chitins were flowable freely at room temperature but quickly became a durable gel at body temperature. Thermo-reversible sol-gel transition properties were observed with fast gelation kinetics. Glycol chitins with higher DA showed faster degradation in the presence of lysozyme. They exhibited no significant biological toxicity against human cell lines. An anti-cancer drug, doxorubicin, could be incorporated into the hydrogel by a simple mixing process and released in a sustained pattern over 13 days. Our findings suggest that glycol chitins could be useful as a new thermogelling biomaterial for drug delivery and injectable tissue engineering.

Surface plasmon resonance studies of the direct interaction between a drug/intestinal brush border membrane

AbstractPurpose.We describe here a new method to estimate the oral drug permeability from the small intestine using surface plasmon resonance (SPR) technology. The interaction between drugs and brush border membrane (BBM) surfaces immobilized on biosensor chip were evaluated by measuring the SPR response signal. Methods. BBM vesicles, isolated from Sprague-Dawley (SD) rats, were immobilized onto the L1 chip composed of dextran derivatives with modified lipophilic residues. A SPR (BIAcore 3000) was used with L1 chip, and it was carried out in a running buffer, HEPES-buffered saline containing 0.1% DMSO. Fourteen drugs for the SPR experiments were flowed over the BBM immobilized L1 chip, and the response levels according to the BBM surfaces were evaluated directly in a continuous flow system. Results. The immobilized BBM surface on L1 chip was very stable, and it was regenerated by injecting a new BBM vesicle solution. It was evident that drug binding events, using BBM surfaces, directly provides information that predicts the Fa value in human for transcellularly absorbed drugs. The throughput to assay each drug at a single concentration is 100 drugs for 24 h. Conclusions. The interaction between drugs and small intestinal surfaces was successfully assayed using SPR technology, and this SPR analysis exhibited advantages: lack of labeling requirements, the real-time acquirement of various results, and the repeated use for various drugs.

Preparation and Characterization of Reconstructed Small Intestinal Brush Border Membranes for Surface Plasmon Resonance Analysis

AbstractPurpose. To prepare the surface generated by small intestinal brush border membrane vesicles (BBMVs) for the surface plasmon resonance (SPR) analysis, which allows the real-time measurement of binding events occurring on the intestinal membrane. Methods. BBMVs were isolated from Sprague-Dawley rats, suspended in HEPES-buffered saline, and flowed over the surface of a SPR sensor chip composed of dextran derivatives modified with lipophilic residues. The surface coverage was determined from binding of bovine serum albumin to BBMV-immobilized sensor chip. The performance of BBMVs immobilized was evaluated by their interaction with otilonium bromide and bile salts. Results. The stable BBMV surface was achieved when BBMV suspension was flowed over the sensor chip for 8 h at a rate of 2 μl/min. The flow of otilonium bromide resulted in an increased SPR signal because of its binding to calcium channel, which is known to be distributed over the gastrointestinal tact. When bile salts were flowed over ileal and duodenal BBMV surfaces, respectively, a slightly higher SPR signal was observed in the ileal BBMV surface, indicating the specific interaction of bile salts with bile acid transporters. Conclusions. BBMV surfaces may be useful for the estimation of binding events on the intestinal membrane by SPR analysis, especially for the drugs that are orally administrated.

Design and strategies for bile acid mediated therapy and imaging

Bioinspired materials have received substantial attention across biomedical, biological, and drug delivery research because of their high biocompatibility and lower toxicity compared with synthetic materials. Bile acids, well-established biomimetic biomolecules, have been reported with respect to their potential applications as carriers of drugs or imaging contrast agents and, most importantly, as oral absorption enhancers. This review introduced the potential mechanisms involved in the oral absorption of bile acids and their derivatives and further focused on the intelligent applications of bile acids or modified bile acids that respond to biological cues as potential oral absorption enhancers for peptides and macromolecular drugs. Our investigations via the modifications of bile acids with various linkers have demonstrated their effects on the degree of oral absorption. Furthermore, we summarized the reports regarding the development of bile acid formulations for the oral delivery of optical imaging contrast agents for GI tract imaging, as well as anticancer drug delivery. Our opinions regarding the utilization of bile acids for biological and biomedical applications provide clear and concise guidance to investigators with respect to the merits and demerits of bile acid use and the selection of appropriate bile acids based on the requirements for improved biomedical applications.

Mucoadhesive hybrid gel improves intraperitoneal platinum delivery.

A leading cause of death and suffering in patients with abdominal or pelvic malignancies is progression of peritoneal surface disease. Changes in the use of chemotherapy have shown significant survival benefits for intraperitoneal or combined intraperitoneal and intravenous treatment following optimal surgical cytoreduction. However, broader clinical use of intraperitoneal therapy has not reached its full potential due to limited efficacy, accessibility and nonspecific toxicity. To overcome these problems, we developed a mucoadhesive hybrid gel (HG) for a local, intraperitoneal drug delivery. In vivo studies confirmed reliable adherence and residence of the gel to the peritoneal sidewall for at least 72 h exhibiting no signs of tissue toxicity. Functionally active CDDP was released from HG within 2h and was equal to free CDDP in vitro. Moreover, intraperitoneal application of HG-CDDP significantly enhanced CDDP accumulation in the genomic DNA of peritoneal tissues compared to the same CDDP dose administered intravenously. These findings indicate the potential application of this hybrid gel as a mucoadhesive drug carrier amendable to use for intraperitoneal drug delivery and possible expansion for use on other mucosal surfaces of the female reproductive tract.

Endometrial cancers harboring mutated fibroblast growth factor receptor 2 protein are successfully treated with a new small tyrosine kinase inhibitor in an orthotopic mouse model

Objectives AL3818 (anlotinib) is a receptor tyrosine kinase inhibitor targeting vascular endothelial growth factor receptors (VEGFR1, VEGFR2/KDR, and VEGFR3), stem cell factor receptor (C-kit), platelet-derived growth factor (PDGFβ), and fibroblast growth factor receptors (FGFR1, FGFR2, and FGFR3). This study evaluates the efficacy of AL3818 studying tumor regression in an orthotopic murine endometrial cancer model. Methods We tested the cytotoxicity of AL3818 on a panel of 7 human endometrial cancer cell lines expressing either wild-type or mutant FGFR2 and also assessed the in vivo antitumor efficacy in a murine, orthotopic AN3CA endometrial cancer model. AL3818 was administered daily per os either alone or in combination with carboplatin and paclitaxel, which represent the current standard of adjuvant care for endometrial cancer. Results AL3818 significantly reduces AN3CA cell number in vitro, characterized by high expression of a mutated FGFR2 protein. Daily oral administration of AL3818 (5 mg/kg) resulted in a complete response in 55% of animals treated and in a reduced tumor volume, as well as decreased tumor weights of AN3CA tumors by 94% and 96%, respectively, following a 29-day treatment cycle. Whereas carboplatin and paclitaxel failed to alter tumor growth, the combination with AL3818 did not seem to exhibit a superior effect when compared with AL3818 treatment alone. Conclusions AL3818 shows superior efficacy for the treatment of endometrial cancer irresponsive to conventional carboplatin and paclitaxel combination and warrants further investigation.

Combinatorial gene construct and non-viral delivery for anti-obesity in diet-induced obese mice

The combinatorial peptidergic therapy of islet amyloid polypeptide (IAPP) and leptin (LEP) analogues was once an optimistic option in treating obese animals and patients. However, the need for frequent administrations and its negative side effects prevent it from being a viable choice. Here, we developed a combinatorial gene therapy of IAPP and LEP, where two genes are inserted into a single plasmid with self-cleaving furin and 2A sites to treat diet-induced obese (DIO) mice. The developed plasmid DNA (pDNA) individually produced both IAPP and LEP peptides in vitro and in vivo. The pDNA was delivered with a non-viral polymeric carrier, and its once-a-week administrations demonstrated a synergistic loss of body weight and significant reductions of fat mass, blood glucose, and lipid levels in DIO mice. The results suggest that the combinatorial gene therapy would have higher potential than the peptidergic approach for future translation due to its improved practicability.

Oral siRNA Delivery to Treat Colorectal Liver Metastases

Convenient multiple dosing makes oral administration an ideal route for delivery of therapeutic siRNA. However, hostile GI environments and nonspecific biological trafficking prevent achieving appropriate bioavailability of siRNA. Here, an orally administered AuNP-siRNA-glycol chitosan-taurocholic acid nanoparticle (AR-GT NPs) was developed to selectively deliver Akt2 siRNA and treat colorectal liver metastases (CLM). AR-GT NPs are dual padlocked nonviral vectors in which the initially formed AuNP-siRNA (AR) conjugates are further encompassed by bifunctional glycol chitosan-taurocholic acid (GT) conjugates. Covering the surface of AR with GT protected the Akt2 siRNA from GI degradation, facilitated active transport through enterocytes, and enhanced selective accumulation in CLM. Our studies in CLM animal models resulted in the reduction in Akt2 production, followed by initiation of apoptosis in cancer cells after oral administration of Akt2 siRNA-loaded AR-GT. This therapeutic siRNA delivery system may be a promising approach in treating liver-associated diseases.

Abstract 3244: Treatment of endometrial cancer cells with a new small tyrosine kinase inhibitor targeting mutated fibroblast growth factor receptor-2

Endometrial cancer (EC) is the most common uterine malignancy in industrialized countries and is the most frequent gynecologic cancer in the United States with an estimated 60,050 new cases in 2016. Traditionally observed in peri-, postmenopausal women, the incidence rate of EC has steadily increased among younger women of reproductive age over the past two decades partially owing to a more sedentary lifestyle and rampant obesity epidemic. Surgery in combination with either radio- and or chemotherapy is commonly curative for the early stage of the disease. However, patients with recurrent or metastatic disease, who only respond poorly to cytotoxic chemotherapy are in desperate need of therapeutic alternatives as are patients of reproductive age. Thus, innovative, safe and more effective therapies are mandated. Several studies have demonstrated the increased expression of various tyrosine kinase receptors involved in the development of EC. We hypothesized that a targeted therapeutic approach using a pan-tyrosine kinase inhibitor would prevent the proliferation and progression of EC. Anlotinib (AL3818) is a multi-targeted receptor tyrosine kinase inhibitor targeting vascular endothelial growth factor receptors 1 to 3 (VEGFR1-3), stem cell factor receptor (C-kit), platelet-derived growth factor (PDGFβ), and fibroblast growth factor receptors 1 to 3 (FGFR1-3). We tested a panel of seven human endometrial cancer cell lines (AN3Ca, Ishikawa, HEC1A, HEC1B, KLE, MFE280 and MFE296) to assess the cytotoxicity of anlotinib in vitro. A lower IC50 value was observed in AN3Ca cells (25 nM), a cell line characterized by a high level of expression of an FGFR2 mutant protein, while cell lines expressing FGFR2 wild-type are less sensitive to anlotinib (HEC1A: 33 μM, or HEC1B:36 μM). Somatic mutations of FGFR2 have been observed in 12% of EC, indicating the potential for this drug in a subset of EC patients. In summary, these results suggest improved efficacy of anlotinib for the treatment of ECs expressing an increased level of FGFR2 mutant proteins. The safety and efficacy of anlotinib are currently being evaluated in an orthotopic AN3Ca mouse model of EC. Experimental groups include a tumor bearing control not subjected to any treatment, anlotinib alone, carboplatin/paclitaxel, a combination of anlotinib and carboplatin/paclitaxel. Primary study endpoints include tumor growth retardation, the delayed onset of local metastases as well as reduced toxicity as secondary endpoint. Citation Format: Sebastien Taurin, Chieh-Hsiang Yang, Maria Reyes, Sungpil Cho, Elke A. Jarboe, Theresa L. Werner, Demetrius M. Coombs, Paul Chen, Margit M. Janat-Amsbury. Treatment of endometrial cancer cells with a new small tyrosine kinase inhibitor targeting mutated fibroblast growth factor receptor-2 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3244. doi:10.1158/1538-7445.AM2017-3244

Enhanced thermogenic program by non-viral delivery of combinatory browning genes to treat diet-induced obesity in mice.

Thermogenic program (also known as browning) is a promising and attractive anti-obesity approach. Islet amyloid polypeptide (IAPP) and irisin have emerged as potential browning hormones that hold high potential to treat obesity. Here, we have constructed a dual browning gene system containing both IAPP and irisin (derived from fibronectin type III domain containing 5; FNDC5) combined with 2A and furin self-cleavage sites. Intraperitoneal administration of the construct complexed with a linear polyethylenimine into diet-induced obese mice demonstrated the elevation of anti-obesogenic effects characterized as the decreased body weight, adiposity, and levels of glucose and insulin. In addition, the construct delivery increased energy expenditure and the expression of core molecular determinants associated with browning. The additional advantages of the dual browning gene construct delivery compared to both single gene construct delivery and dual peptide delivery can be emphasized on efficacy and practicability. Hence, we have concluded that dual browning gene delivery makes it therapeutically attractive for diet-induced obesity treatment.