Synthesis and biological evaluation of loxoprofen derivatives

Non-steroidal anti-inflammatory drugs (NSAIDs) achieve their anti-inflammatory actions through an inhibitory effect on cyclooxygenase (COX). Two COX subtypes, COX-1 and COX-2, are responsible for the majority of COX activity at the gastrointestinal mucosa and in tissues with inflammation, respectively. We previously suggested that both gastric mucosal cell death due to the membrane permeabilization activity of NSAIDs and COX-inhibition at the gastric mucosa are involved in NSAID-induced gastric lesions. We have also reported that loxoprofen has the lowest membrane permeabilization activity among the NSAIDs we tested. In this study, we synthesized a series of loxoprofen derivatives and examined their membrane permeabilization activities and inhibitory effects on COX-1 and COX-2. Among these derivatives, 2-{4'-hydroxy-5-[(2-oxocyclopentyl)methyl]biphenyl-2-yl}propanoate 31 has a specificity for COX-2 over COX-1. Compared to loxoprofen, oral administration of 31 to rats produced fewer gastric lesions but showed an equivalent anti-inflammatory effect. These results suggest that 31 is likely to be a therapeutically beneficial and safer NSAID.     

Discovery of orteronel (TAK-700), a naphthylmethylimidazole derivative, as a highly selective 17,20-lyase inhibitor with potential utility in the treatment of prostate cancer

A novel naphthylmethylimidazole derivative 1 and its related compounds were identified as 17,20-lyase inhibitors. Based on the structure-activity relationship around the naphthalene scaffold and the results of a docking study of 1a in the homology model of 17,20-lyase, the 6,7-dihydro-5H-pyrrolo[1,2-c]imidazole derivative (+)-3c was synthesized and identified as a potent and highly selective 17,20-lyase inhibitor. Biological evaluation of (+)-3c at a dose of 1mg/kg in a male monkey model revealed marked reductions in both serum testosterone and dehydroepiandrosterone concentrations. Therefore, (+)-3c (termed orteronel [TAK-700]) was selected as a candidate for clinical evaluation and is currently in phase III clinical trials for the treatment of castration-resistant prostate cancer.     

Design and synthesis of novel allosteric MEK inhibitor CH4987655 as an orally available anticancer agent

The MAP kinase pathway is one of the most important pathways involved in cell proliferation and differentiation, and its components are promising targets for antitumor drugs. Design and synthesis of a novel MEK inhibitor, based on the 3D-structural information of the target enzyme, and then multidimensional optimization including metabolic stability, physicochemical properties and safety profiles were effectively performed and led to the identification of a clinical candidate for an orally available potent MEK inhibitor, CH4987655, possessing a unique 3-oxo-oxazinane ring structure at the 5-position of the benzamide core structure. CH4987655 exhibits slow dissociation from the MEK enzyme, remarkable in vivo antitumor efficacy both in mono- and combination therapy, desirable metabolic stability, and insignificant MEK inhibition in mouse brain, implying few CNS-related side effects in human. An excellent PK profile and clear target inhibition in PBMC were demonstrated in a healthy volunteer clinical study.     

Resilient Plant–Bird Interactions in a Volcanic Island Ecosystem: Pollination of Japanese Camellia Mediated by the Japanese White-Eye

Observations of interspecies interactions during volcanic activity provide important opportunities to study how organisms respond to environmental devastation. Japanese camellia (Camellia japonica L.) and its main avian pollinator, the Japanese White-eye (Zosterops japonica), offer an excellent example of such an interaction as key members of the biotic community on Miyake-jima, which erupted in 2000 and continues to emit volcanic gases. Both species exhibit higher resistance to volcanic damage than other species. We examined the effects of volcanic activity on this plant–pollinator system by estimating pollen flow and the genetic diversity of the next generation. The results showed that despite a decrease in Camellia flowers, the partitioning of allelic richness among mother-tree pollen pools and seeds decreased while the migration rate of pollen from outside the study plot and the pollen donor diversity within a fruit increased as the index of volcanic damage increased. In areas with low food (flower) density due to volcanic damage, Z. japonica ranged over larger areas to satisfy its energy needs rather than moving to areas with higher food density. Consequently, the genetic diversity of the seeds (the next plant generation) increased with the index of volcanic damage. The results were consistent with previously published data on the movement of Z. japonica based on radio tracking and the genetic diversity of Camellia pollen adhering to pollinators. Overall, our results indicated that compensation mechanisms ensured better pollination after volcanic disturbance.     

An Immunohistochemical Analysis of Tissue Thrombin Expression in the Human Atria

Objective

Thrombin, the final coagulation product of the coagulation cascade, has been demonstrated to have many physiological effects, including pro-fibrotic actions via protease-activated receptor (PAR)-1. Recent investigations have demonstrated that activation of the cardiac local coagulation system was associated with atrial fibrillation. However, the distribution of thrombin in the heart, especially difference between the atria and the ventricle, remains to be clarified. We herein investigated the expression of thrombin and other related proteins, as well as tissue fibrosis, in the human left atria and left ventricle.

Methods

We examined the expression of thrombin and other related molecules in the autopsied hearts of patients with and without atrial fibrillation. An immunohistochemical analysis was performed in the left atria and the left ventricle.

Results

The thrombin was immunohistologically detected in both the left atria and the left ventricles. Other than in the myocardium, the expression of thrombin was observed in the endocardium and the subendocardium of the left atrium. Thrombin was more highly expressed in the left atrium compared to the left ventricle, which was concomitant with more tissue fibrosis and inflammation, as detected by CD68 expression, in the left atrium. We also confirmed the expression of prothrombin in the left atrium. The expression of PAR-1 was observed in the endocardium, subendocardium and myocardium in the left atrium. In patients with atrial fibrillation, strong thrombin expression was observed in the left atrium.

Conclusions

The strong expression levels of thrombin, prothrombin and PAR-1 were demonstrated in the atrial tissues of human autopsied hearts.     

Domestication Process of the Goat Revealed by an Analysis of the Nearly Complete Mitochondrial Protein-Encoding Genes

Goats (Capra hircus) are one of the oldest domesticated species, and they are kept all over the world as an essential resource for meat, milk, and fiber. Although recent archeological and molecular biological studies suggested that they originated in West Asia, their domestication processes such as the timing of population expansion and the dynamics of their selection pressures are little known. With the aim of addressing these issues, the nearly complete mitochondrial protein-encoding genes were determined from East, Southeast, and South Asian populations. Our coalescent time estimations suggest that the timing of their major population expansions was in the Late Pleistocene and significantly predates the beginning of their domestication in the Neolithic era (≈10,000 years ago). The ω (ratio of non-synonymous rate/synonymous substitution rate) for each lineage was also estimated. We found that the ω of the globally distributed haplogroup A which is inherited by more than 90% of goats examined, turned out to be extremely low, suggesting that they are under severe selection pressure probably due to their large population size. Conversely, the ω of the Asian-specific haplogroup B inherited by about 5% of goats was relatively high. Although recent molecular studies suggest that domestication of animals may tend to relax selective constraints, the opposite pattern observed in our goat mitochondrial genome data indicates the process of domestication is more complex than may be presently appreciated and cannot be explained only by a simple relaxation model.     

A Critical Role of Fatty Acid Binding Protein 4 and 5 (FABP4/5) in the Systemic Response to Fasting

During prolonged fasting, fatty acid (FA) released from adipose tissue is a major energy source for peripheral tissues, including the heart, skeletal muscle and liver. We recently showed that FA binding protein 4 (FABP4) and FABP5, which are abundantly expressed in adipocytes and macrophages, are prominently expressed in capillary endothelial cells in the heart and skeletal muscle. In addition, mice deficient for both FABP4 and FABP5 (FABP4/5 DKO mice) exhibited defective uptake of FA with compensatory up-regulation of glucose consumption in these tissues during fasting. Here we showed that deletion of FABP4/5 resulted in a marked perturbation of metabolism in response to prolonged fasting, including hyperketotic hypoglycemia and hepatic steatosis. Blood glucose levels were reduced, whereas the levels of non-esterified FA (NEFA) and ketone bodies were markedly increased during fasting. In addition, the uptake of the ¹²⁵I-BMIPP FA analogue in the DKO livers was markedly increased after fasting. Consistent with an increased influx of NEFA into the liver, DKO mice showed marked hepatic steatosis after a 48-hr fast. Although gluconeogenesis was observed shortly after fasting, the substrates for gluconeogenesis were reduced during prolonged fasting, resulting in insufficient gluconeogenesis and enhanced hypoglycemia. These metabolic responses to prolonged fasting in DKO mice were readily reversed by re-feeding. Taken together, these data strongly suggested that a maladaptive response to fasting in DKO mice occurred as a result of an increased influx of NEFA into the liver and pronounced hypoglycemia. Together with our previous study, the metabolic consequence found in the present study is likely to be attributed to an impairment of FA uptake in the heart and skeletal muscle. Thus, our data provided evidence that peripheral uptake of FA via capillary endothelial FABP4/5 is crucial for systemic metabolism and may establish FABP4/5 as potentially novel targets for the modulation of energy homeostasis.     

Estimation of breeding population size using DNA‐based pedigree reconstruction in brown bears

Robust estimates of demographic parameters are critical for effective wildlife conservation and management but are difficult to obtain for elusive species. We estimated the breeding and adult population sizes, as well as the minimum population size, in a high‐density brown bear population on the Shiretoko Peninsula, in Hokkaido, Japan, using DNA‐based pedigree reconstruction. A total of 1288 individuals, collected in and around the Shiretoko Peninsula between 1998 and 2020, were genotyped at 21 microsatellite loci. Among them, 499 individuals were identified by intensive genetic sampling conducted in two consecutive years (2019 and 2020) mainly by noninvasive methods (e.g., hair and fecal DNA). Among them, both parents were assigned for 330 bears, and either maternity or paternity was assigned to 47 and 76 individuals, respectively. The subsequent pedigree reconstruction indicated a range of breeding and adult (≥4 years old) population sizes: 128–173 for female breeders and 66–91 male breeders, and 155–200 for female adults and 84–109 male adults. The minimum population size was estimated to be 449 (252 females and 197 males) in 2019. Long‐term continuous genetic sampling prior to a short‐term intensive survey would enable parentage to be identified in a population with a high probability, thus enabling reliable estimates of breeding population size for elusive species.