Evaluation of the repeated-dose liver and gastrointestinal tract micronucleus assays with 22 chemicals using young adult rats: summary of the collaborative study by the Collaborative Study Group for the Micronucleus Test (CSGMT)/The Japanese Environmental Mutagen Society (JEMS) - Mammalian Mutagenicity Study Group (MMS)
PubMed | Toray Industries Inc, Nissan Chemical Industries Ltd., Yakult Honsha Co., LSI Corporation and 16 more.
Type: | Journal: Mutation research. Genetic toxicology and environmental mutagenesis | Year: 2015
The repeated-dose liver micronucleus (RDLMN) assay using young adult rats has the potential to detect hepatocarcinogens. We conducted a collaborative study to assess the performance of this assay and to evaluate the possibility of integrating it into general toxicological studies. Twenty-four testing laboratories belonging to the Mammalian Mutagenicity Study Group, a subgroup of the Japanese Environmental Mutagen Society, participated in this trial. Twenty-two model chemicals, including some hepatocarcinogens, were tested in 14- and/or 28-day RDLMN assays. As a result, 14 out of the 16 hepatocarcinogens were positive, including 9 genotoxic hepatocarcinogens, which were reported negative in the bone marrow/peripheral blood micronucleus (MN) assay by a single treatment. These outcomes show the high sensitivity of the RDLMN assay to hepatocarcinogens. Regarding the specificity, 4 out of the 6 non-liver targeted genotoxic carcinogens gave negative responses. This shows the high organ specificity of the RDLMN assay. In addition to the RDLMN assay, we simultaneously conducted gastrointestinal tract MN assays using 6 of the above carcinogens as an optional trial of the collaborative study. The MN assay using the glandular stomach, which is the first contact site of the test chemical when administered by oral gavage, was able to detect chromosomal aberrations with 3 test chemicals including a stomach-targeted carcinogen. The treatment regime was the 14- and/or 28-day repeated-dose, and the regime is sufficiently promising to incorporate these methods into repeated-dose toxicological studies. The outcomes of our collaborative study indicated that the new techniques to detect chromosomal aberrations in vivo in several tissues worked successfully.
Ohsumi K.,Astellas Pharma Inc. |
Masaki T.,Astellas Research Technologies Co. |
Takase S.,Astellas Pharma Inc. |
Watanabe M.,Astellas Pharma Inc. |
Fujie A.,Astellas Pharma Inc.
Journal of Antibiotics | Year: 2014
A novel antifungal agent, AS2077715, was isolated from the fermentation broth of a fungal strain (339855) identified as a new Capnodium species based on morphological characteristics and large-subunit ribosomal DNA sequencing. AS2077715 was isolated as a white powder via solvent extraction, HP-20 and ODS-B column chromatography and crystallization, and was determined to have the molecular formula C 25 H 41 NO 7. AS2077715 has a structure related to that of funiculosin, an inhibitor of mitochondrial cytochrome bc 1 complex (complex III), and showed antifungal activity against Trichophyton species.
Vulgatibacter incomptus gen. nov., sp. nov. and Labilithrix luteola gen. nov., sp. nov., two myxobacteria isolated from soil in Yakushima Island, and the description of Vulgatibacteraceae fam. nov., Labilitrichaceae fam. nov. and Anaeromyxobacteraceae fam. nov
Yamamoto E.,Astellas Research Technologies Co. |
Muramatsu H.,Astellas Research Technologies Co. |
Nagai K.,Astellas Research Technologies Co.
International journal of systematic and evolutionary microbiology | Year: 2014
Two myxobacterial strains (designated B00001(T) and B00002(T)) were isolated from forest soil samples collected from Yakushima Island, Kagoshima, Japan. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strains B00001(T) and B00002(T) respectively formed independent branches within the suborders Cystobacterineae and Sorangiineae and were most closely related to Cystobacter armeniaca DSM 14710(T) (90.4% similarity) and Byssovorax cruenta DSM 14553(T) (91.3%). Neither strain showed typical features of myxobacteria such as bacteriolytic action or fruiting body formation, but both had high DNA G+C contents (66.3-68.3 mol%). Swarming motility was observed in strain B00002(T) only. Cells of both strains were vegetative, chemoheterotrophic, mesophilic, strictly aerobic, Gram-negative, motile rods, and both strains exhibited esterase lipase (C8), leucine arylamidase, naphthol-AS-BI-phosphohydrolase and β-galactosidase activities. Strain B00001(T) contained MK-7 as the predominant respiratory quinone and the major fatty acid was iso-C15:0. In contrast, strain B00002(T) contained MK-8 as the major cellular quinone and the major fatty acids were C16 : 1ω5c and iso-C17 : 0. Based on the phenotypic and genotypic data presented, strains B00001(T) and B00002(T) represent novel genera and species, for which we propose the names Vulgatibacter incomptus gen. nov., sp. nov. and Labilithrix luteola gen. nov., sp. nov., respectively. The type strains of Vulgatibacter incomptus and Labilithrix luteola are B00001(T) ( = NBRC 109945(T) = DSM 27710(T)) and B00002(T) ( = NBRC 109946(T) = DSM 27648(T)), respectively. The new genera are assigned to the new families Vulgatibacteraceae fam. nov. and Labilitrichaceae fam. nov., respectively. In addition, Anaeromyxobacteraceae fam. nov., is proposed to accommodate the genus Anaeromyxobacter, which is related to the genus Vulgatibacter. © 2014 IUMS.
Nagashima T.,Astellas Pharma Inc. |
Shigematsu N.,Astellas Research Technologies Co. |
Maruki R.,Astellas Research Technologies Co. |
Urano Y.,Astellas Pharma Inc. |
And 4 more authors.
Molecular Pharmacology | Year: 2010
Excessive hepatic glucose production through the gluconeogenesis pathway is partially responsible for the elevated glucose levels observed in patients with type 2 diabetes mellitus (T2DM). The forkhead transcription factor forkhead box O1 (Foxo1) plays a crucial role in mediating the effect of insulin on hepatic gluconeogenesis. Here, using a db/db mouse model, we demonstrate the effectiveness of Foxo1 inhibitor, an orally active small-molecule compound, as a therapeutic drug for treating T2DM. Using mass spectrometric affinity screening, we discovered a series of compounds that bind to Foxo1, identifying among them the compound, 5-amino-7-(cyclohexylamino)-1-ethyl-6-fluoro-4-oxo-1,4- dihydroquinoline-3-carboxylic acid (AS1842856), which potently inhibits human Foxo1 transactivation and reduces glucose production through the inhibition of glucose-6 phosphatase and phosphoenolpyruvate carboxykinase mRNA levels in a rat hepatic cell line. Oral administration of AS1842856 to diabetic db/db mice led to a drastic decrease in fasting plasma glucose level via the inhibition of hepatic gluconeogenic genes, whereas administration to normal mice had no effect on the fasting plasma glucose level. Treatment with AS1842856 also suppressed an increase in plasma glucose level caused by pyruvate injection in both normal and db/db mice. Taken together, these findings indicate that the Foxo1 inhibitor represents a new class of drugs for use in treating T2DM. Copyright © 2010 The American Society for Pharmacology and Experimental Therapeutics.
PubMed | Astellas Research Technologies Co. and Astellas Pharma Inc.
Type: Journal Article | Journal: The Journal of antibiotics | Year: 2015
FR901459, a product of the fungus Stachybotrys chartarum No. 19392, is a derivative of cyclosporin A (CsA) and a powerful immunosuppressant that binds cyclophilin. Recently, it was reported that CsA was effective against hepatitis C virus (HCV). However, FR901459 lacks active moieties, which are essential for synthesizing more potent and safer derivatives of this anti-HCV agent. Here we identified an actinomycete strain (designated 7887) that was capable of efficient bioconversion of FR901459. Structural elucidation of the isolated bioconversion products (1-7) revealed that compounds 1-4 were mono-hydroxylated at the position of 1-MeBmt or 9-MeLeu, whereas compounds 5-7 were bis-hydroxylated at both positions. The results of morphological and chemical characterization, as well as phylogenetic analysis of 16S ribosomal DNA (rDNA), suggested that strain 7887 belonged to the genus Lentzea. Comparison of the FR901459 conversion activity of strain 7887 with several other Lentzea strains revealed that although all examined strains metabolized FR901459, strain 7887 had a characteristic profile with respect to bioconversion products. Taken together, these findings suggest that strain 7887 can be used to derivative FR901459 to produce a chemical template for further chemical modifications that may provide more effective and safer anti-HCV drugs.
PubMed | Astellas Research Technologies Co.
Type: Evaluation Studies | Journal: Assay and drug development technologies | Year: 2015
Transport assays using P-gp-expressing cell lines are commonly used to identify P-gp substrates and inhibitors in drug discovery. The P-gp cell-based assay is performed manually in 12- or 24-well plates and requires improvement for high-throughput screening. In this study, we established an efficient semiautomated 96-well transport assay using LLC-PK1 cells transfected with human P-gp. The protocol was optimized with a microplate washer for exchanging media and buffer to enhance throughput. P-gp substrates and inhibitors, and the paracellular marker Dextran Texas Red were used to validate the 96-well transport assay. Cell monolayer integrity after washing by a microplate washer was confirmed by measuring paracellular permeability of Dextran Texas Red. Permeability and net flux ratio of the P-gp substrates and the inhibitory potency of the P-gp inhibitors were comparable in 24- and 96-well plates. The regression value of net flux ratio of P-gp substrates was high between the two formats (r=0.99). The optimized 96-well transport assay using the microplate washer was found to be an efficient high-throughput screening tool that provided the same quality data as the 24-well plate for the identification of P-gp substrates and inhibitors in drug discovery.
PubMed | Astellas Research Technologies Co.
Type: Journal Article | Journal: Experimental animals | Year: 2015
To confirm our hypothesis that the sex and age of cynomolgus monkeys influences the effect of training, we employed a new training technique designed to increase the animals affinity for animal care personnel. During 151 days of training, monkeys aged 2 to 10 years accepted each 3 raisins/3 times/day, and communicated with animal care personnel (5 times/day). Behavior was scored using integers between -1 and 5. Before training, 35 of the 61 monkeys refused raisins offered directly by animal care personnel (Score -1, 0 and 1). After training, 28 of these 35 monkeys (80%) accepted raisins offered directly by animal care personnel (>Score 2). The mean score of monkeys increased from 1.2 0.1 to 4.3 0.2. The minimum training period required for monkeys to reach Score 2 was longer for females than for males. After 151 days, 6 of the 31 females and 1 of the 30 males still refused raisins offered directly by animal care personnel. Beneficial effects of training were obtained in both young and adult monkeys. These results indicate that our new training technique markedly improves the affinity of monkeys for animal care personnel, and that these effects tend to vary by sex but not age. In addition, abnormal behavior and symptoms of monkeys were improved by this training.
An automated method by which effects of compounds on locomotor activity and spontaneous neuropathic pain-specific movements can be simultaneously evaluated in rats with chronic-constriction nerve injury
PubMed | Astellas Research Technologies Co.
Type: | Journal: European journal of pharmaceutical sciences : official journal of the European Federation for Pharmaceutical Sciences | Year: 2016
Neuropathic pain patients are characterized by evoked pain (hyperalgesia and allodynia) and spontaneous pain, the latter of which is the predominant symptom. In animal models of neuropathic pain, effects of test compounds on spontaneous pain-related behaviors have been evaluated by direct visual observation. In addition, by performing another locomotor activity experiment in normal animals, it is also indispensable to examine whether test compounds cause motor impairment to avoid overestimation of their analgesic activities. In the present study, we developed spontaneous pain-specific and automated evaluation method by improving a previous method for measuring movements of the injured hind limb in unilateral chronic constriction nerve injury (CCI) rats. Rats with unilateral CCI were implanted with strong and weak magnets in each hind limb, respectively. Limb movements were automatically detected as spiked waveforms in electromagnetic field analyzing system. Movements in each limb were analyzed separately according to differences in their respective wave amplitudes, and aberrant movements of injured limb were specifically detected on basis of the asymmetry between injured and uninjured limb movements. Consequently, the incidence ratio of spontaneous pain in injured limb, which was not affected by individual locomotive activities, was able to be obtained as a new evaluation index. The incidence ratio of spontaneous pain revealed substantial difference between CCI and sham rats with only a small variation in the value. Further, according to the frequency of movement of the uninjured limb, it could be determined simultaneously whether a test compound causes sedative effect or not, resulting in eliminating the need for another experiment in normal animals.
Yoshida S.,Astellas Pharma Inc. |
Kasai M.,Astellas Pharma Inc. |
Kimura T.,Astellas Research Technologies Co. |
Akiba T.,Astellas Pharma Inc. |
And 2 more authors.
Organic Process Research and Development | Year: 2012
Process research and development of a practical and scalable synthetic route toward compound (S)-1 and compound (R)-1, which are potent selective dual antagonists for 5-HT2B and 5-HT7 receptors, respectively, is described. The medicinal chemistry route and second generation route were also unattractive for large-scale use for a variety of reasons. The new synthetic method does not require any purification by column chromatography for all steps and highly exothermic reactions. Additionally, we developed an efficient method of optical resolution in which each carboxylic acid isomer was separated with chiral amine in high yield and high enantiopurity. This highly efficient and scalable process was successfully demonstrated in the large scale synthesis of compound (S)-1 and compound (R)-1 in high enantiopurity. © 2012 American Chemical Society.
PubMed | Astellas Research Technologies Co.
Type: Journal Article | Journal: The Journal of veterinary medical science | Year: 2016
To determine the effect of long-distance (approximately 600 km) road transportation on the blood biochemistry of laboratory animals, we investigated the changes in serum biochemical parameters in healthy cynomolgus monkeys and beagle dogs transported by truck from Osaka to Tsukuba, Japan. The concentrations of serum cortisol, total bilirubin and aspartate aminotransferase in monkeys increased during transportation. Serum cortisol and total bilirubin levels in dogs also increased during transportation, but serum triglyceride decreased. Serum parameter values in truck-transported monkeys and dogs returned to baseline levels within two weeks following arrival. Taken together, these results suggest that a two-week acclimation period is the minimum duration required for adaptation following road transportation.