Currently, only a few medicines have been approved for use in the clinical treatment of chronic pain, but they are not fully satisfying due to their side effects. From the view that radical treatment, rather than simply treating symptoms, is more important in addressing life-long chronic pain, we have been investigating translational research for a mechanism-based medicine to treat pain. Through the characterization of various types of peripheral and central neuropathic pain in mice, we discovered that lysophosphatidic acid (LPA) plays roles in definitive mechanisms of the development and maintenance of neuropathic pain. We found LPA₁ receptor- and LPA₃ receptor-mediated amplification of LPA production could be a key mechanism underlying the initiation and maintenance of this pain. We have developed stress-induced fibromyalgia models, and have revealed that LPA₁ receptor-signaling also plays key roles in the mechanism. Throughout these studies, we found that LPA plays a key role in pain memory, and that LPA₁ receptor- and LPA₃ receptor-antagonists could reverse the established pain, and thereby cure the disease source of pain.

To examine the effect of tanshinone IIA on Angiotensin II (Ang II)-induced proliferation and autophagy in vascular smooth muscle cells (VSMCs) and the related mechanism. VSMCs were treated with Ang II with or without tanshinone IIA (1, 5 and 10 µg/mL), and the proliferation, apoptosis in cells with different treatment were examined by methylthiazolyl tetrazolium (MTT) and flow cytometry methods. Moreover, the expression of autophagy related proteins and mitogen-activated protein kinase (MAPK) signaling molecules were examined by RT-quantitative (q)PCR and Western blot methods. Ang II induced significantly increase in the proliferation and autophagy of VSMCs, and the MAPK signaling was activated. Tanshinone IIA can attenuate Ang II-induced effects via down-regulating the MAPK signaling pathway. Tanshinone IIA can inhibit Ang II-induced proliferation and autophagy of VSMCs via regulating the MAPK signaling pathway.

Autophagy plays key roles in the development of acute pancreatitis (AP) and the regulation of impaired autophagy has therapeutic potential. The objective of the present study was to investigate whether pharmacological inhibition of autophagy could ameliorate AP in mice and examine the underlying mechanisms. In current study, by imaging-based high-throughput screening, a novel spautin-1 derivative spautin-A41 was identified as a potent autophagy inhibitor. Mice treated with spautin-A41 were resistant to the cerulein-induced elevation of serum pancreatic enzyme activities and pancreatic apoptosis. Mechanistically, spautin-A41 effectively reduced the expression levels of Class III phosphatidylinositol 3 (PI3) kinase complexes and subsequently ameliorated pancreatitis by inhibiting the formation of autophagosome. Therefore, pharmacological inhibition of autophagy by spautin-A41 may serve as new target for treating or lessening the severity of AP.

Investigation of the occurrence time of adverse drug reactions helps to prevent the development and aggravation of adverse reactions, but the expression time of ganciclovir-induced adverse events has not been elucidated. In this study, using databases of spontaneous adverse event reports, the Japanese Adverse Drug Event Report database (JADER) and the U.S. Food and Drug Administration (FDA)’s Adverse Event Reporting System (FAERS), the incidence of adverse reactions due to ganciclovir and their expression time were analyzed. As a result of calculation of the reporting odds ratio (ROR) and 95% confidence interval for individual main adverse reactions of ganciclovir (cytopenia, leukopenia, thrombocytopenia, liver damage, and acute renal failure), a signal was detected for all adverse reactions in both databases, except for liver damage in JADER. Furthermore, the Weibull distribution was performed for the analysis of onset time of each ganciclovir-induced adverse event. The results of Weibull parameter α and β values of each adverse event in both JADER and FAERS suggested that most adverse events occurred within 30 d and classified into the early failure type, except that thrombocytopenia and acute renal failure in JADER classified into the random failure type. Based on these findings, it concluded that the paying attention to signs of each ganciclovir-induced adverse event is required from the early phase after ganciclovir administration. However, in FAERS, development after a long-term course also accounted for 11%, suggesting that long-term periodic monitoring of adverse reactions would be also required.

Tyrosine kinase inhibitors (TKIs) are used as the first choice for chronic myeloid leukemia (CML) pharmacotherapeutics. Some patients taking these drugs showed good therapeutic reactivity despite the disappearance of drugs from blood. We investigated whether these drugs have sustained effects even after their disappearance and whether their effects depend on their amounts of intracellular accumulation. Cell proliferation after exposure of K562 cells or Multidrug resistance-1 (MDR-1)-transfected K562 cells was determined by a cell counting kit-8 assay. The intracellular accumulation amount of the drug showing a sustained cytostatic effect was measured by ultra high performance liquid chromatography mass spectrometry. Cell viability decreased in a culture time-dependent manner after washing out nilotinib and dasatinib. The sustained cytostatic effect of dasatinib, but not that of nilotinib, correlated with the intracellular accumulation level. In contrast, imatinib showed continuous a cytostatic effect after drug washout for long-term exposure but not after drug washout for short-term exposure. These results suggest that a good response in patients with a low serum concentration of imatinib, nilotinib or dasatinib may be due to the cytostatic effect of that drug continues even after its disappearance in plasma.

Spirogyra neglecta (SN), commonly named “Tao” in Thai, is a genus of filamentous green macroalgae. SN contains polyphenols such as isoquercetin, catechin, hydroquinone and kaempferol. These constituents exhibit beneficial effects including anti-oxidant, anti-gastric ulcer, anti-hyperglycaemia and anti-hyperlipidaemia in both in vitro and in vivo models. Whether SN extract (SNE) has an anti-inflammatory effect in vivo remains unclear. This study examined the effect of SNE on renal function and renal organic transport in lipopolysaccharide (LPS)-induced renal inflammation in rats. Rats were randomised and divided into normal saline (NS), NS supplemented with 1000 mg/kg body weight (BW) of SNE (NS + SNE), intraperitoneally injected with 12 mg/kg BW of LPS and LPS treated with 1000 mg/kg BW of SNE (LPS + SNE). Biochemical parameters in serum and urine, lipid peroxidation concentration, kidney function and renal organic anion and cation transports were determined. LPS-injected rats developed renal injury and inflammation by increasing urine microalbumin, total malondialdehyde (MDA) and inflammatory cytokines, tumor necrosis factor (TNF)-α and interleukin (IL)-1β protein expression, respectively. In addition, uptake of renal organic anion, [³H]-oestrone sulphate (ES), was reduced in LPS-injected rats together with increased expression of organic anion transporter 3 (Oat3). However, the renal injury and inflammation, as well as impaired Oat3 function and protein expression, were restored in LPS + SNE rats. Accordingly, SNE could be developed as nutraceutical product to prevent inflammation-induced nephrotoxicity.

Hazardous drugs (HD), which need to be handled with care, may be administered through a feeding tube using the simple suspension method. However, instrument contamination during HD administration with the simple suspension method remains unclear. Therefore, to minimize such contamination during the simple suspension method using an injector, we propose the following exposure countermeasures method: (1) Wear two layers of gloves. (2) Prepare injectors for administration and flushing. (3) Use caps. (4) Replace outer gloves after the removal of tablets from the press-through package (PTP) sheet. (5) Handle drugs on a tray. (6) Inject while wrapping the connection site between the injector for administration and the tube with gauze. (7) Wrap the connection site between the injector and tube with gauze. (8) Do not point the injector downward. To establish whether these countermeasures method are effective, 16 ward nurses who routinely administer drugs via a feeding tube were enrolled as subjects. By visual evaluation, we compared differences in instrument contamination between a suspension using a medicine cup and administration via a feeding tube (the conventional method) and the exposure countermeasures method. Exposure with the countermeasures method under our instruction was markedly lower than that with the conventional method. Furthermore, after implementing the exposure countermeasures method, most nurses noted that caution and awareness of exposure countermeasures increased. Thus, to minimize exposure, we recommend the implementation of the exposure countermeasures method and increasing knowledge and awareness of measures against exposure.

Colorectal cancer (CRC) is one of the most common malignant tumors and the third leading cause of cancer-related deaths in the world. It was reported that sophocarpine could attenuate the progression of CRC in mice. However, the mechanisms by which sophocarpine regulate the proliferation and migration in CRC remain unclear. Thus, this study aimed to investigate anti-tumor mechanisms of sophocarpine in CRC cells. CCK-8 assay, wound healing assay and transwell migration were used to detect cell proliferation and migration, respectively. In addition, Western blotting and enzyme-linked immunosorbent assay (ELISA) were used to further detect protein expressions and cytokines in vitro. The results revealed that sophocarpine significantly inhibited proliferation in HCT116 and SW620 cells, respectively. Meanwhile, sophocarpine inhibited CRC cells migration via downregulation of the levels of N-cadherin, matrix metalloproteinase (MMP)-9, phosphorylated extracellular signal-regulated kinase (p-ERK), p-mitogen-activated protein kinase kinase (MEK), vascular endothelial growth factor (VEGF)-A, VEGF-C and VEGF-D. Moreover, overexpression of MEK reversed the anti-migration effects of sophocarpine on CRC cells via upregulation of VEGF-A/C/D. Our findings indicated that sophocarpine could inhibit CRC cells migration via downregulation of MEK/ERK/VEGF pathway. Thus, sophocarpine may act as a potential agent for the treatment of CRC.

Uridine 5′-diphospho-glucuronosyltransferase (UGT), a metabolic enzyme of irinotecan active metabolite, has two genetic polymorphisms (UGT1A1*6 and UGT1A1*28). In UGT1A1 homozygous or heterozygous patients, metabolism is delayed and the risk of developing adverse effects is increased, and therefore, dose reduction of irinotecan is considered. However, the specific dose reduction rate of irinotecan for heterozygous patients is uncertain. We studied the necessity of irinotecan dose reduction and its optimal dose in UGT1A1 heterozygous patients with lung cancer. Patients with lung cancer treated with irinotecan in the Tokushima University Hospital or Tokushima Municipal Hospital were included in this study. The dose of irinotecan was evaluated based on the relative dose intensity (RDI). The time to treatment failure (TTF) was defined as the period until treatment change, death, or progressive disease based on response evaluation criteria of solid tumors. We targeted 31 patients treated with irinotecan: 12 wild types (WT), 14 heterozygotes, and 1 complex heterozygote and 4 homozygotes. There was no significant difference in the TTF, but the mean RDI during the entire treatment period was significantly different in the wild type (79%), heterozygous (62%), and complex heterozygous and homozygous groups (46%). In addition, the proportion of patients who completed treatment without dose reduction in the WT group tended to be higher than that in the other groups. For lung cancer patients with UGT1A1 heterozygote types who start irinotecan therapy, reducing the initial dose by approximately 20% might be a safer chemotherapy without decreasing the therapeutic effect.

As alveolar macrophages are attractive targets for the treatment of tuberculosis, effective methods for delivery to alveolar macrophages are under development. We investigated a pulmonary formulation for the efficient delivery of high water-soluble drugs at high concentration targeting alveolar macrophages. In this study, a surfactant-coated high water-soluble drug complex (SDC, a hydrophobic dried emulsion), which can preferably target alveolar macrophages and be expected to deliver drug at a high concentration, was prepared in the first process. OCT313, a high water-soluble sugar derivative with anti-tuberculosis activity was used. Then, a unique two-solution, mixing-type nozzle was used to prepare the SDC nanoparticles in mannitol (MAN) microparticles (SDC/MAN microparticles) because it was difficult to disperse the SDC nanoparticles in aqueous solution. The single micron size of OCT313–SDC/MAN microparticles contained OCT313–SDC nanoparticles (mean particle size of OCT313–SDC nanoparticles, 277.9 nm; drug contents, 1.31 ± 0.041 wt%). We found that the treatment of SDC/MAN microparticles exhibited significantly higher drug accumulation in macrophage cells (Raw264.7 cells, 7.5-fold, at 4 h after treatment) in vitro and in alveolar macrophages in rats (9.1-fold, at 4 h after treatment) in vivo than that of drug alone. These results suggest that the SDC/MAN microparticle formulation prepared by spray drying through a two-solution mixing-type nozzle provides efficient delivery of a water-soluble drug targeting alveolar macrophages and may be useful for tuberculosis treatment.

The aim of the study was to investigate the changes in the reactive oxygen species (ROS), Sirt1, p53 and acetylated p53 in brain tissue of newborn rats exposed to hyperoxia to clarify the role of Sirt1 signaling pathway in brain injury. Neonate rats were randomly divided into normoxic group and hyperoxic group. Rats in the normoxic group were exposed to room air while the rats in the hyperoxic group were put in a hyperoxic chamber (80 ± 5% oxygen) for 1 to 14 d. Data, including weight growth, the water content of brain tissue, hematoxyline and eosin (H&E) and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (Tunel) stain, ROS expression, the relative expression of Sirt1 mRNA and p53 mRNA, and the protein relative expression of Sirt1, p53 and acetylated p53 were analyzed at 1, 7 and 14 d after exposure. A reduced body weight and increased water content were observed in the brain tissue of hyperoxic group compared to normoxic group. HE staining and Tunel staining of brain tissue suggested that cell damaged after hyperoxic exposure. RT-PCR and Western blot results showed that the expression of Sirt1 in the hyperoxic group was lower than that in the normoxic group while the expression of p53 was higher than that in the normoxic group. In addition, Western blot data indicated acetylated p53 expression was higher in the hyperoxic group. Hyperoxic exposure can lead to brain injury in newborn Sprague-Dawley (SD) rats. These events might be regulated by the Sirt1 pathway, which downregulated the deacetylation of p53.

Vitamin D has an immune-modulating effect, related to the pathophysiology of asthma and chronic obstructive pulmonary disease (COPD). However, few studies have focused on the difference between patients with asthma and COPD in the association of circulating vitamin D levels with clinical outcomes. We sought to investigate the associations of circulating vitamin D levels with health-related QOL (HR-QOL), severity, and exacerbations. Subjects included 152 asthma patients and 50 COPD patients. We measured plasma concentrations of 25-hydroxyvitamin D₃ [25(OH)D₃]. HR-QOL was assessed using the EuroQoL 5-Dimension (EQ-5D) and the 12-item Short Form Health Survey (SF-12) scales. Exacerbations were recorded during a one-year follow-up. Associations between plasma 25 (OH)D₃ concentrations and outcome variables were evaluated using linear regression. Plasma concentrations of 25(OH)D₃ were positively associated with the EQ-5D index value and the SF-12 physical component score in patients with asthma; however, such associations were not observed in patients with COPD. A significant association between severity and plasma concentrations of 25(OH)D₃ was found only in patients with COPD. The hazard ratios (95% confidence interval) of plasma 25(OH)D₃ concentrations (per 1 ng/mL decrease) for time to first exacerbation was 1.38 (1.10–1.75; p = 0.006) and 0.95 (0.87–1.03; p = 0.179) in patients with COPD and asthma, respectively. Lower concentrations of plasma 25(OH)D₃ contributed to lower HR-QOL in patients with asthma, and were associated with severity and risk of future exacerbations in patients with COPD.

Anandamide (AEA) played potent neuroprotective activities via cannabinoid type 1 (CB1) and 2 (CB2) receptor. N-Linoleyltyrosine (NITyr), as an AEA analogue, was synthesized in our laboratory and evaluated the neuroprotective effects and mechanisms for the first time. NITyr was synthesized via substitution reaction. The neuroprotective effects of NITyr were evaluated in a gerbil model of transient cerebral ischemia. Each gerbil was subjected to open field test (OFT), Rotard rod test (RRT), Morris water maze (MWM) successively and executed after animal behaviors. Part of the brain was stained with hematoxylin and eosin (HE) and Nissl staining, and the rest for biochemical analysis. NITyr could not increase spontaneous locomotor activity and ameliorate the anxiety behavior in the OFT but could improve the motor coordination in the RRT and the spatial memory impairment in the MWM. Immunohistochemically, NITyr could attenuate the ischemia-induced neural loss in the hippocampus. The Enzyme-linked immunosorbent assay (ELISA) suggested that NITyr ameliorated the inflammation and oxidative stress. Consistently, NITyr could up-regulate the expressions of p-phosphadylinositol 3-kinase (PI3K) and p-Akt but not PI3K and Akt in the hippocampus. In addition to oxidative stress, CB2 receptor antagonist AM630 but not CB1 receptor antagonist AM251 could reverse the above phenomena. However, CB1 receptor antagonist AM251 could reverse oxidative stress. Accordingly, NITyr could up-regulate the expressions of CB2 but not CB1. NITyr could improve the motor coordination, learning and memory impairments, neural loss in the hippocampus and the inflammation of the mice via CB2 receptor involvement of PI3K/Akt signaling pathway.

Intracerebroventricular (icv) injection of transient receptor potential vanilloid 4 (TRPV4) agonists 4α-phorbol-12, 13-didecanoate (4α-PDD) and GSK101690A increased urinary excretion under the physiological condition. TRPV4 antagonists ruthenium red and HC-067047 significantly blocked increased urinary volume after intragastric administration of water and 4α-PDD-induced diuresis. Administration of the TRPV4 agonists did not significantly change the plasma concentration of vasopressin or atrial natriuretic factor. Pretreatment with indomethacin inhibited the diuresis induced by 4α-PDD. Moreover, icv injection of prostaglandin (PG) F_2α produced diuretic effects. These findings indicate that central TRPV4 regulates urine excretion, which contributes to systemic water homeostasis in vivo. The underlying mechanisms are suggested to involve PG synthesis, but not release of vasopressin or atrial natriuretic factor.

Since grape extract (GE) contains oligomeric proanthocyanidins and numerous polyphenols, dietary GE supplements may exert protective effects against various diseases. The present study investigated the pharmacological effects of GE derived from Chardonnay in vitro and in vivo. GE (100 µg/mL) completely inhibited tumor necrosis factor-α-induced endothelin-1, monocyte chemoattractant protein-1, interleukin-1β, and intercellular adhesion molecule-1 gene expression in cultured endothelial cells. GE also strongly stimulated the phosphatidylinositol 3-kinase (PI3K)/Akt/endothelial nitric oxide synthase (eNOS) pathway. In the in vivo study, the effects of GE on ischemic acute kidney injury (AKI) were examined using male C57bl/6J wild-type and eNOS^−/− mice. Right nephrectomized mice were exposed to 45 min of ischemia in the left kidney and this was followed by reperfusion. Although renal functional parameters in AKI mice significantly increased 48 h after reperfusion, the administration of GE (0.1 and 1 mg/kg, intravenous (i.v.)) 5 min before ischemia dose-dependently improved post-ischemic renal dysfunction in wild-type mice. Renal histopathological studies on AKI mice revealed tubular necrosis, proteinaceous casts in tubuli, and medullary congestion. The administration of GE ameliorated this damage in wild-type mice, but not in eNOS^−/− mice. Furthermore, GE significantly restored decreases in the renal nitric oxide metabolite content due to ischemia in wild-type mice, but not in eNOS^−/− mice. Thus, eNOS is closely involved in the renoprotective effects of GE, strongly suggesting that GE supplements are useful as a prophylactic treatment for the development of ischemic AKI.

Onset of severe hyponatremia following cisplatin (CDDP) administration has been previously reported. However, the risk factors associated with hyponatremia still remain unclear. We conducted a retrospective, single-center, case-control study to identify independent risk factors of severe hyponatremia in patients with various types of cancers. Adult patients who received intravenous CDDP administration between January 2012 and December 2017 met the inclusion criteria. The investigators recorded patients’ demographics and clinical information retrospectively, and assessed the lowest serum sodium level within 21 d of the first CDDP administration. Risk factors for grade ≥3 hyponatremia were examined via a logistic regression analysis. Among a total of 472 patients, fifty patients (10.6%) developed grade ≥3 hyponatremia. Univariate analysis revealed that age (≥65 years), presence of small cell lung or esophageal cancer, and lower sodium concentrations in the serum (<138 mEq/L) were significantly associated with grade 3 and 4 hyponatremia. Additionally, multivariable logistic regression analysis showed that the presence of small cell lung cancer (adjusted odds ratio, 3.26; 95% confidence interval (CI), 1.07–10.00) and lower sodium concentrations in the serum (<138 mEq/L) (adjusted odds ratio, 6.18; 95%CI, 3.21–11.90) were independent risk factors of grade 3 and 4 hyponatremia. Thus, serum sodium concentrations in patients with these risk factors should be closely monitored after CDDP administration.

Pharmaceutical applications of three dimensional (3D) printing technology are increasing following the approval of 3D-printed tablets by the U.S. Food and Drug Administration. Semi-solid extrusion-type 3D printers are used to 3D print hydrogel- and paste-based materials. We previously developed tablet formulations for semi-solid extrusion-type 3D bioprinters. In the present study, we extended our study to the preparation of muco-adhesive oral film formulations to 3D bioprint mouth ulcer pharmaceuticals. We focused on hydroxypropyl methylcellulose (HPMC)-based catechin (model drug)-loaded hydrogel formulations and found that the viscosity of a hydrogel formulation is dependent on the HPMC concentration, and that viscosity is important for facile 3D printing. HPMC-based films were prepared using two different drying methods (air drying and freeze drying). The films exhibited different drug dissolution profiles, and increasing the amount of HPMC in the film delayed drug dissolution. The fabrication of HPMC-based catechin-loaded films with different shapes provides a model of individualized, on-demand pharmaceuticals. Our results support the flexible application of 3D bioprinters (semi-solid extrusion-type 3D printers) for preparing film formulations.

Tumor necrosis factor-α (TNF-α) converting enzyme/a disintegrin and metalloproteinase domain-containing protein 17 (TACE/ADAM17) is a key sheddase that releases TNF-α from its inactive precursor and is thought as a new drug target to inhibit TNF-α production. In the present study, pharmacological effects of a novel TACE selective inhibitor, JTP-96193, on type 2 diabetes and diabetic peripheral neuropathy (DPN) as its major complication was examined. Enzyme inhibitory activity of JTP-96193 on TACE and other ADAMs was measured in in vitro. High fat-induced obese mice and type 2 diabetic KK-A^y mice were used to evaluate the effect of JTP-96193 on insulin resistance. Finally, streptozotocin (STZ)-induced diabetic mice were treated with JTP-96193 to evaluate the sciatic motor nerve conduction velocities (MNCV). JTP-96193 selectively inhibited human TACE activity with IC₅₀ value of 5.4 nM and showed more than 1800-fold selectivity against other matrix metalloproteinases. In mouse models of obesity and diabetes, JTP-96193 reduced the TNF-α release from the fat tissue and prevented development of diabetes and improved insulin resistance, respectively. Furthermore, JTP-96193 prevented delay of sciatic MNCV without any effects on blood glucose or insulin levels in STZ-induced diabetic mice. TACE inhibitor is effective on insulin resistance and DPN independent from glucose-lowering effect. These pharmacological properties of JTP-96193 may be helpful to treat type 2 diabetes accompanied by its microvascular complications.

4,4-Diisothiocyanatostilbene disulfonic acid (DIDS), an antagonist of anion channel including voltage-dependent anion channel (VDAC), acts as both neurotoxicant and neuroprotectant, resulting in the controversy. VDAC contributes to neuronal apoptosis and is a candidate target protein of 15-deoxy-Δ^12,14-prostaglandin J₂ (15d-PGJ₂). Caspase-3 is activated during neuronal apoptosis caused by 15d-PGJ₂. In the present study, we ascertained whether DIDS was neuroprotective or neurotoxic in the primary culture of rat cortical neurons. Neuronal cell viabilities were primarily evaluated by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H tetrazolium bromide (MTT) reduction assay. Plasma membrane integrity and apoptosis were detected by the staining of propidium iodide (PI) and Hoechst33342, respectively. Alternatively, apoptosis was also measured by caspase-3 assay kit. DIDS did not prevent neurons from undergoing the 15d-PGJ₂-induced apoptosis. In contrast, DIDS caused neuronal cell death in a concentration-dependent manner by itself, confirming its neurotoxicity. The sublethal application of DIDS did not decrease MTT-reducing activity, increase caspase-3 activity, condense chromatin, allow PI to enter neuron and degenerate neuronal morphology significantly. Interestingly, DIDS enhanced the 15d-PGJ₂-induced neuronal apoptosis markedly under the sublethal condition. To our knowledge, this is the first report of synergistic effects of DIDS on the neurotoxicity of 15d-PGJ₂.

Growth of cancer cells is more highly dependent on various types of amino acids than that of normal cells, and thus prevention of amino acid requirement has been recognized as strategies for cancer therapies. In this study, we found that deprivation of cysteine (Cys) in culturing media prevented the growth of various types of human cancer cell lines. Cys is easily converted to cystine (Cys–Cys) in media and uptaken into cells by cystine/glutamate transporter (xCT). The incorporated Cys–Cys is decomposed into Cys, and used for synthesis of glutathione that suppresses reactive oxygen species-induced cell damage. Therefore, we examined whether a selective xCT inhibitor erastin prevented the growth of human cancer cell lines. As a result, erastin significantly prevented the proliferation of various types of human cancer cells. Among them, MDA-MB-231 breast cancer cells were identified as the most erastin-sensitive cells. To investigate the ability of erastin to prevent growth of tumor in mice, MDA-MB-231 breast cancer cells were implanted into BALB/c nude female mice kept under standardized light/dark cycle conditions. The growth of tumor implanted in mice was significantly suppressed by administration of erastin during the light phase, whereas its administration during the dark phase failed to suppress the tumor growth. The dosing time-dependency of erastin-induced cystine/cysteine deprivation was closely related to that of its anti-tumor effects. Our present findings suggest that the anti-tumor efficacy of erastin in tumor-bearing mice is improved by optimizing the dosing schedule.

Elevated intraocular pressure (IOP) is the major cause of glaucoma, which is the second leading cause of blindness. However, current glaucoma treatments cannot completely regulate IOP and progression of glaucoma. Our group recently found that autotaxin (ATX) activity in human aqueous humor (AH) was positively correlated with increased IOP in various subtypes of glaucoma. To develop new IOP-lowering treatments, we generated a novel ATX inhibitor as an ophthalmic drug by high-throughput screening, followed by inhibitor optimization. Administration of the optimized ATX inhibitor (Aiprenon) reduced IOP in laser-treated mice exhibiting elevated IOP and higher level of ATX activity in AH and normal mice in vivo. The stimulation of ATX induced outflow resistance in the trabecular pathway; however, administration of Aiprenon recovered the outflow resistance in vitro. The in vitro experiments implied that the IOP-lowering effect of Aiprenon could be correlated with the altered cellular behavior of trabecular meshwork (TM) and Schlemm’s canal endothelial (SC) cells. Overall, our findings showed that ATX had major impact in regulating IOP as a target molecule, and potent ATX inhibitors such as Aiprenon could be a promising therapeutic approach for lowering IOP.

Skin exposure to UV rays causes the production of reactive oxygen species (ROS), and it is a major risk factor for various skin disorders and diseases. In particular, exposure to UV-A is a major cause of photoaging. We have previously isolated 2ʹ,3ʹ-dihydroxy-4ʹ,6ʹ-dimethoxychalcone (DDC) from green perilla leaves as an activator of the nuclear factor erythroid 2-related factor-2 (Nrf2)–antioxidant response element (ARE) and demonstrated the protective effects of DDC both in vitro and in vivo in PC12 cells and Parkinson’s disease models, respectively. In this study, we used HaCaT cells to examine the effects of DDC on ROS production and cell damage induced by UV-A. Our results indicated that UV-A irradiation in HaCaT cells increased ROS production in an energy-dependent manner. In addition, cell viability decreased in an energy-dependent manner 24 h after UV-A irradiation. However, treatment with DDC 24 h prior to UV-A irradiation significantly suppressed UV-A radiation-induced ROS production. In addition, DDC showed cytoprotective effects when used 24 h before and after UV-A irradiation. Treatment with DDC for 24 h also increased the expression levels of heme oxygenase-1 (HO-1) in a concentration-dependent manner. Pretreatment with the HO-1 inhibitor followed by DDC treatment before UV-A irradiation for 24 h reduced ROS production and the cytoprotective effect. These results suggest that DDC increases the expression levels of HO-1 and protects HaCaT cells through the suppression of UV radiation-induced ROS production.

Amyloid β protein (Aβ) causes neurotoxicity and cognitive impairment in Alzheimer’s disease (AD). Oxidative stress is closely related to the pathogenesis of AD. We have previously reported that 2′,3′-dihydroxy-4′,6′-dimethoxychalcone (DDC), a component of green perilla, enhances cellular resistance to oxidative damage through the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2)–antioxidant response element (ARE) pathway. Here, we investigated the effects of DDC on cortical neuronal death induced by Aβ. When Aβ and DDC had been preincubated for 3 h, the aggregation of Aβ was significantly suppressed. In this condition, we found that DDC provided a neuroprotective action on Aβ-induced cytotoxicity. Treatment with DDC for 24 h increased the expression of heme oxygenase-1 (HO-1), and this was controlled by the activation of the Nrf2–ARE pathway. However, DDC did not affect Aβ-induced neuronal death under any of these conditions. These results suggest that DDC prevents the aggregation of Aβ and inhibits neuronal death induced by Aβ, and although it activates the Nrf2–ARE pathway, this mechanism is less involved its neuroprotective effect.

GGsTop is a highly potent and specific, and irreversible γ-glutamyl transpeptidase (GGT) inhibitor without any influence on glutamine amidotransferases. The aim of the present study was to investigate the involvement of GGT in ischemia/reperfusion-induced cardiac dysfunction by assessing the effects of a treatment with GGsTop. Using a Langendorff apparatus, excised rat hearts underwent 40 min of global ischemia without irrigation and then 30 min of reperfusion. GGT activity was markedly increased in cardiac tissues exposed to ischemia, and was inhibited by the treatment with GGsTop. Exacerbation of cardiac functional parameters caused by ischemia and reperfusion, namely the reduction of left ventricular (LV) developed pressure and the maximum and negative minimum values of the first derivative of LV pressure, and the increment in LV end-diastolic pressure was significantly attenuated by GGsTop treatment. The treatment with GGsTop suppressed excessive norepinephrine release in the coronary perfusate, a marker for myocardial dysfunction, after ischemia/reperfusion. In addition, oxidative stress indicators in myocardium, including superoxide and malondialdehyde, after ischemia/reperfusion were significantly low in the presence of GGsTop. These observations demonstrate that enhanced GGT activity contributes to cardiac damage after myocardial ischemia/reperfusion, possibly via increased oxidative stress and subsequent norepinephrine overflow. GGT inhibitors have potential as a therapeutic strategy to prevent myocardial ischemia/reperfusion injury in vivo.

Novel bivalent twin-drug type hydantoin derivatives were evaluated in vitro using a human brain glioma cell line (U251) and a human carcinoma cell line (KB3-1). Among the 5-substituted hydantoin derivatives (1a–b and 2a–d) examined in this study, bivalent symmetrical 5-substituted hydantoin derivative 1b showed the highest anti-proliferative activity towards both U251 and KB3-1 cells. The values of anti-proliferative activity (IC₅₀) of this hydantoin derivative against the two cell lines (U251 and KB3-1) were 0.46 and 5.21 µM, respectively. The anti-proliferative activity of all of the compounds except for compounds 2a and 2d against U251 cells was higher than that of cisplatin. Bivalent symmetrical compound 1b had a biphenylmethane linker in the molecule. All of the tested bivalent hydantoin derivatives showed higher activity against U251 cells than against KB3-1 cells. For twin-drug type hydantoin derivatives 2a–d, which have a linear methylene linker in the molecules, it was found that methylene linker length in these molecules have an effect on the anti-proliferative activity against U251 and KB3-1 cells.