Fibroblast growth factor receptor 4 increases epidermal growth factor receptor (EGFR) signaling by inducing amphiregulin expression and attenuates response to EGFR inhibitors in colon cancer.

Fibroblast growth factor receptor 4 (FGFR4) is known to induce cancer cell proliferation, invasion, and antiapoptosis through activation of RAS/RAF/ERK and PI3K/AKT pathways, which are also known as major molecular bases of colon cancer carcinogenesis related with epidermal growth factor receptor (EGFR) signaling. However, the interaction between FGFR4 and EGFR signaling in regard to colon cancer progression is unclear. Here, we investigated a potential cross-talk between FGFR4 and EGFR, and the effect of anti-EGFR therapy in colon cancer treatment. To explore the biological roles of FGFR4 in cancer progression, RNA sequencing was carried out using FGFR4 transfected colon cell lines. Gene ontology data showed the upregulation of genes related to EGFR signaling, and we identified that FGFR4 overexpression secretes EGFR ligands such as amphiregulin (AREG) with consequent activation of EGFR and ErbB3. This result was also shown in in vivo study and the cooperative interaction between EGFR and FGFR4 promoted tumor growth. In addition, FGFR4 overexpression reduced cetuximab-induced cytotoxicity and the combination of FGFR4 inhibitor (BLU9931) and cetuximab showed profound antitumor effect compared to cetuximab alone. Clinically, we found the positive correlation between FGFR4 and AREG expression in tumor tissue, but not in normal tissue, from colon cancer patients and these expressions were significantly correlated with poor overall survival in patients treated with cetuximab. Therefore, our results provide the novel mechanism of FGFR4 in connection with EGFR activation and the combination of FGFR4 inhibitor and cetuximab could be a promising therapeutic option to achieve the optimal response to anti-EGFR therapy in colon cancer.

Cryomilling-induced solid dispersion of poor glass forming/poorly water-soluble mefenamic acid with polyvinylpyrrolidone K12.

The effect of mechanical impact on the polymorphic transformation of mefenamic acid (MFA) and the formation of a solid dispersion of mefenamic acid, a poor glass forming/poorly-water soluble compound, with polyvinylpyrrolidone (PVP) K12 was investigated. The implication of solid dispersion formation on solubility enhancement of MFA, prepared by cryomilling, was investigated. Solid state characterization was conducted using powder X-ray diffraction (PXRD) and Fourier-transform infrared (FTIR) spectroscopy combined with crystal structure analysis. Apparent solubility of the mixtures in pH 7.4 buffer was measured. A calculation to compare the powder patterns and FTIR spectra of solid dispersions with the corresponding physical mixtures was conducted. Solid state characterization showed that (1) MFA I transformed to MFA II when pure MFA I was cryogenically milled (CM); and (2) MFA forms a solid dispersion when MFA was cryogenically milled with PVP K12. FTIR spectral analysis showed that hydrogen bonding facilitated by mechanical impact played a major role in forming solid dispersions. The apparent solubility of MFA was significantly improved by making a solid dispersion with PVP K12 via cryomilling. This study highlights the importance of cryomilling with a good hydrogen bond forming excipient as a technique to prepare solid dispersion, especially when a compound shows a poor glass forming ability and therefore, is not easy to form amorphous forms by conventional method.

Effects of seaweed Laminaria japonica extracts on skin moisturizing activity in vivo.

Twelve species of edible seaweed from the coast of Korea were screened for skin moisturizing activity. We placed the lead of a Corneometer on an approximately 6-cm2 test area of the forearm and measured both untreated skin (control) and skin treated with test moisturizing creams either containing or not containing 5% water:propylene glycol (50:50) extracts of seaweeds. Over the 8-h observation period, the strongest activity of the Laminaria japonica extracts occurred at the 2-h period. For the 10% extract, hydration with the L. japonica extract increased by 14.44% compared with a placebo. Transepidermal water loss (TEWL) was also measured using a test cream with 10% L. japonica extract. For up to 8 h after applying the creams, TEWL was decreased to 4.01 g/cm2, which was approximately 20% of that seen with the control. We suggest that the L. japonica extract hydrates skin via the humectants and hydrocolloids that it contains. To confirm the safety of L. japonica extracts, we performed a patch test on human skin. The results suggested that at moderate doses humans can safely use the extracts. For commercial applications, we evaluated the physicochemical characteristics of the test cream products, including Hunter L, a, and b values; pH; refractive index; and coefficient of viscosity. L. japonica extract did not affect overall formulations of the test cream product in any of the tested aspects. These results suggest that L. japonica extract is a promising ingredient in moisturizing formulations.

Low-dose thiazide diuretics in children with idiopathic renal hypercalciuria.

AIM: To evaluate the therapeutic effect of hydrochlorothiazide in idiopathic renal hypercalciuria. METHODS: We retrospectively analysed the data of 28 children (6.0±4.1 years, M:F=19:9) diagnosed as having idiopathic renal hypercalciuria from the years 1991 to 2008. The dose of hydrochlorothiazide was initially 0.5 mg/kg/day and gradually increased to achieve the appropriate hypocalciuric effect (urinary calcium/creatinine<0.2 mg/mg) in some unresponsive patients. RESULTS: Twenty-two patients (79%) had gross haematuria, 6 (21%) microscopic haematuria, 2 left flank pain, 6 (21%) urolithiasis and 9 (32%) urinary tract infection at the diagnosis of hypercalciuria. The low doses (0.5 mg/kg/day) of hydrochlorothiazide reduced urinary calcium excretion in 25 patients (89%) and 3 (11%) required the increased doses (1-2 mg/kg/day). Haematuria and urolithiasis gradually resolved in accordance with the improvement of hypercalciuria. Nineteen patients (68%) maintaining hypocalciuria during hydrochlorothiazide therapy were discontinued after 12.5±5.3 months of treatment. Eleven of the 19 patients maintained normocalciuria, while 8 showed increased urinary calcium excretion at 2.9±2.3 months after treatment was stopped, requiring thiazide retreatment. CONCLUSION: Our results suggest that low dose (0.5 mg/kg/day) of hydrochlorothiazide may be safe and effective in controlling renal hypercalciuria in children.

Lgals3bp suppresses colon inflammation and tumorigenesis through the downregulation of TAK1-NF-κB signaling.

Galectin 3-binding protein (LGALS3BP, also known as 90K) is a multifunctional glycoprotein involved in immunity and cancer. However, its precise role in colon inflammation and tumorigenesis remains unclear. Here, we showed that Lgals3bp-/- mice were highly susceptible to colitis and colon tumorigenesis, accompanied by the induction of inflammatory responses. In acute colitis, NF-κB was highly activated in the colon of Lgals3bp-/- mice, leading to the excessive production of pro-inflammatory cytokines, such as IL-6, TNFα, and IL-1ß. Mechanistically, Lgals3bp suppressed NF-κB through the downregulation of TAK1 in colon epithelial cells. There was no significant difference in the pro-inflammatory cytokine levels between wild-type and Lgals3bp-/- mice in a chronic inflammatory state, during colon tumorigenesis. Instead, Lgals3bp-/- mice showed elevated levels of GM-CSF, compared to those in WT mice. We also found that GM-CSF promoted the accumulation of myeloid-derived suppressor cells and ultimately increased colon tumorigenesis in Lgals3bp-/- mice. Taken together, Lgals3bp plays a critical role in the suppression of colitis and colon tumorigenesis through the downregulation of the TAK1-NF-κB-cytokine axis. These findings suggest that LGALS3BP is a novel immunotherapeutic target for colon inflammation and tumorigenesis.

IL-12 Enhances Immune Response by Modulation of Myeloid Derived Suppressor Cells in Tumor Microenvironment.

Myeloid derived suppressor cells (MDSCs) are a heterogenous population of immature cells that play a critical role in tumor associated immune suppression. In tumor conditions, the population of MDSCs increases. The main feature of these cells is their ability to suppress the T cell response in antigen specific or nonspecific manners depending on the condition of T cell activation. IL-12 can modulate MDSC in preliminary reports, so we investigated how IL-12 can affect MDSC in a tumor microenvironment. After implanting tumor based cells on syngeneic host, 4T-1/BALB/c or EL4/C57BL6 mice, MDSCs (Gr1+CD11b+) were isolated from splenocytes. Isolated MDSCs were treated with GM-CSF with or without IL-12 and analyzed based on their phenotypes and functions. Treatment of MDSC with IL-12 increased co-stimulatory molecules of CD80, CD86, OX-40L, enhancing the DC phenotype (CD11c) and maturation markers such as p-NF-κB and p-GSK3ß. In addition to a change of surface markers, T-cell suppressive function of MDSC after IL-12 treatment was significantly improved compared with the control MDSC. In addition, PD-L1+F4/80+ macrophages, which show aninhibitory effect in phagocytosis, were decreased after IL-12 treatment. The changes of cell surface expression of CD80, CD86, MHC class II were also shown in vivo. Our results showed that the IL-12 can modulate MDSC into APC and recover the macrophage function. These results suggested that IL-12 plays a role in improving the tumor immune microenvironment through MDSC modulation.

Bavachin and isobavachalcone, acyl-coenzyme A: cholesterol acyltransferase inhibitors from Psoralea corylifolia.

Acyl-coenzyme A: cholesterol acyltransferase (ACAT) catalyzes cholesterol esterification and plays important roles in intestinal absorption of cholesterol, hepatic production of lipoproteins and accumulation of cholesteryl ester within macrophages and smooth muscle cells. Ethanol extract of Psoralea corylifolia showed a significant inhibition of ACAT enzyme. Via bioactivity-guided fractionation of the ethanol extract of Psoralea corylifolia, two prenylated flavonoids were isolated. Their structures were determined as bavachin (1) and isobavachalcone (2) by spectroscopic analysis ((1)H-, (13)C-NMR, 2DNMR, and ESI-MS). The IC(50) values were 86.0 (1) and 48.0 (2) microM in the ACAT assay system using rat liver microsome. Compound 2 also decreased cholesteryl ester formations in HepG2 cells. In addition, this compound showed a noncompetitive type of inhibition of ACAT.

Penicillium griseofulvum F1959, high-production strain of pyripyropene a, specific inhibitor of acyl-CoA: cholesterol acyltransferase 2.

Acyl-coenzyme A: cholesterol acyltransferase (ACAT) catalyzes cholesterol esterification and plays an important role in the intestinal absorption of cholesterol, hepatic production of lipoproteins, and accumulation of cholesteryl ester within cells. During the course of screening to find ACAT inhibitors from microbial sources, the present authors isolated pyripyropene A from Penicillium griseofulvum F1959. Pyripyropene A, an ACAT2-specific inhibitor, has already been produced from Aspergillus fumigatus. Yet, Aspergillus fumigatus is a pathogen and only produces a limited amount of pyripyropene A, making the isolation of pyripyropene A troublesome. In contrast, Penicillium griseofulvum F1959 was found to produce approximately 28 times more pyripyropene A than Aspergillus fumigatus, plus this report also describes the ideal conditions for the production of pyripyropene A by Penicillium griseofulvum F1959 and its subsequent purification.

Anti-inflammatory effects of ß-sitosterol-ß-D-glucoside from Trachelospermum jasminoides (Apocynaceae) in lipopolysaccharide-stimulated RAW 264.7 murine macrophages.

Trachelospermum jasminoides (Apocynaceae) has pharmacological effects that include anti-inflammatory, anti-bacterial and anti-viral activities, which have been observed from various studies. Of these pharmacological effects, the anti-inflammatory capacity of compounds from T. jasminoides is not yet known exactly. In this study, we investigated the compound that can be used for the suppression of lipopolysacchaide (LPS) stimulated inflammatory responses in macrophages among the five isolated compounds. ß-sitosterol-ß-D-glucoside (1) was found to reduce nitric oxide (NO) production from LPS-induced RAW 264.7 cells the most. In addition, compound 1 strongly inhibited the interleukin 6 (IL-6) activities of stimulated macrophages. Treatment of RAW 264.7 cells with compound 1 reduced secretion of inflammatory elements including tumour necrosis factor - alpha (TNF-α) and interleukin 1 beta (IL-1ß). Thus, compound 1 may be a useful candidate for the development of new drugs to treat endotoxemia and inflammation accompanied by the overproduction of NO.

Renal stone associated with the ketogenic diet in a 5-year old girl with intractable epilepsy.

In this paper, we report on a 5-year-old girl who developed a renal stone while following the ketogenic diet to treat refractory seizure disorder. Three months after initiating the ketogenic diet, she developed severe abdominal pain and vomiting. The spot urine calcium-to-creatinine (Ca/Cr) ratio and 24-hour urine evaluation showed hypercalciuria. Computed tomography (CT) imaging revealed a stone in the right ureteropelvic junction, resulting in hydronephrosis of the right kidney. The renal stone disappeared 5 days after conservative treatment; the patient's microscopic hematuria resolved concurrently. In light of this case report, we recommend regularly monitoring the urine Ca/Cr ratio with ultrasonography for further development of renal stones in patients following the ketogenic diet. If these patients exhibit evidence of symptomatic hypercalciuria or cyristalluria, liberalization of fluid restriction and urine alkalization using oral potassium citrate should be considered.

Inhibitory activity of diacylglycerol acyltransferase by glabrol isolated from the roots of licorice.

Acyl-coenzyme A: diacylglycerol acyltransferase (DGAT, EC 2.3.1.20) catalyzes triglyceride synthesis in the glycerol phosphate pathway. It has relations with the excess supply and accumulation of triglycerides. Therefore, DGAT inhibitors may act as a potential therapy for obesity and type 2 diabetes. Five flavonoids were isolated from the ethanol extracts of licorice roots, using an in vitro DGAT inhibitory assay. One isoprenyl flavonoid showed most potential inhibition of DGAT on five flavonoids (1-5). On the basis of spectral evidences, the compound was identified as glabrol (5). Compound 5 inhibited rat liver microsomal DGAT activity with an IC50 value of 8.0 microM, but the IC50 value for four flavonoids (1-4) was more than 100 microM. In addition, glabrol showed a noncompetitive type of inhibition against DGAT. These data suggest that potential therapy for the treatment in obesity and type 2 diabetes patients by licorice roots might be related with its DGAT inhibitory effect.