Long-term dynamics of the network structures in seed dispersal associated with fluctuations in bird migration and fruit abundance patterns.

In temperate zones, seed-dispersal networks by migratory birds are formed on long time scale. In mid-October from 2005 to 2016, to explore the dynamics of the network structures, we examined interannual variability of fruit abundance, bird migration, and seed-dispersal networks in central Japan. For 12 years, the fruit abundance exhibited a remarkable fluctuation across years, with the number of fruiting plants and matured fruits fluctuating repeatedly every other year, leading to the periodic fluctuations. The abundance of migratory birds was also fluctuated. According to the abundance of fruits and migratory birds, the 12 years was classified into three types: frugivores and fruits were abundant, frugivores were abundant but fruits were scarce, and frugivores were scarce. The seed-dispersal networks were investigated by collecting faeces and vomits of migrants. Of the 6652 samples collected from 15 bird species, 1671 (25.1%) included seeds from 60 plant species. Main dispersers were composed of Turdus pallidus, T. obscurus, and Zosterops japonicus. The network structures were almost nested for 12 years. Specifically, the nested structure was developed in years when fruit abundance was low. GLM analyses showed the abundance of migrants, particularly T. pallidus and T. obscurus, had strong positive effects on nested structure. It may be caused by the fact the two Turdus species were more frequently functioning as generalist dispersers when fruit abundance was lower. Our study suggested fruit abundance and foraging behaviour of frugivores determine the network structures of seed dispersal on long time scale.

Characteristics of the joint position sense in children with developmental dyslexia.

[Purpose] Developmental dyslexia is a disorder in which reading and writing of characters is difficult. The present study investigated age-dependent joint position sense of the forearm and wrist and whether children with developmental dyslexia have less joint position sense than typically developing children. [Participants and Methods] The participants were comprised of 84 typically developing elementary school students, 12 university students, and 2 children with developmental dyslexia. Joint position sense was evaluated using the reproduction method based on four tasks. The participants were divided into three age groups. The children with developmental dyslexia were compared with the typically developing children in the same age group. [Results] Significant negative correlations were found between the reproduction error of the typically developing children and that of the university students in most tasks. The children with developmental dyslexia showed increased reproduction error relative to the reproduction error of the typically developing children in the same age group in 4 of the 8 tasks. [Conclusion] The accuracy of the joint position sense improved with development. However, the joint position sense of the children with developmental dyslexia was lower than that of the typically developing children in the same age group. The difficulty in writing experienced by children with developmental dyslexia may be related to joint position sensing impairment due to impaired joint position sense.

Perfect chemomechanical coupling of FoF1-ATP synthase.

FoF1-ATP synthase (FoF1) couples H+ flow in Fo domain and ATP synthesis/hydrolysis in F1 domain through rotation of the central rotor shaft, and the H+/ATP ratio is crucial to understand the coupling mechanism and energy yield in cells. Although H+/ATP ratio of the perfectly coupling enzyme can be predicted from the copy number of catalytic ß subunits and that of H+ binding c subunits as c/ß, the actual H+/ATP ratio can vary depending on coupling efficiency. Here, we report actual H+/ATP ratio of thermophilic Bacillus FoF1, whose c/ß is 10/3. Proteoliposomes reconstituted with the FoF1 were energized with ΔpH and Δψ by the acid-base transition and by valinomycin-mediated diffusion potential of K+ under various [ATP]/([ADP]⋅[Pi]) conditions, and the initial rate of ATP synthesis/hydrolysis was measured. Analyses of thermodynamically equilibrated states, where net ATP synthesis/hydrolysis is zero, show linear correlation between the chemical potential of ATP synthesis/hydrolysis and the proton motive force, giving the slope of the linear function, that is, H+/ATP ratio, 3.3 ± 0.1. This value agrees well with the c/ß ratio. Thus, chemomechanical coupling between Fo and F1 is perfect.

Successful Treatment of Bilocular Walled-off Necrosis with Transmural Naso-cyst Continuous Irrigation: A Case Report.

Recently, transmural naso-cyst continuous irrigation (TNCCI) has been reported as an effective and safe treatment for walled-off necrosis (WON). We herein report a case of bilocular WON that was successfully treated with TNCCI. The patient was a 60-year-old man. The patient underwent endoscopic ultrasound-guided cyst drainage of the main cavity and subcavity using a single transluminal gateway transcystic multiple drainage technique, which was ineffective. Subsequently, a lumen-apposing metal stent (LAMS) was placed in the main cavity and TNCCI was successfully performed in the subcavity. TNCCI with LAMS was effective in treating bilocular WON.

Design and synthesis of an androgen receptor pure antagonist (CH5137291) for the treatment of castration-resistant prostate cancer.

A series of 5,5-dimethylthiohydantoin derivatives were synthesized and evaluated for androgen receptor pure antagonistic activities for the treatment of castration-resistant prostate cancer. Since CH4933468, which we reported previously, had a problem with agonist metabolites, novel thiohydantoin derivatives were identified by applying two strategies. One was the replacement of the alkylsulfonamide moiety by a phenylsulfonamide to avoid the production of agonist metabolites. The other was the replacement of the phenyl ring with a pyridine ring to improve in vivo potency and reduce hERG affinity. Pharmacological assays indicated that CH5137291 (17b) was a potent AR pure antagonist which did not produce the agonist metabolite. Moreover, CH5137291 completely inhibited in vivo tumor growth of LNCaP-BC2, a castration-resistant prostate cancer model.

Outcome of colorectal cancer in Diamond-Blackfan syndrome with a ribosomal protein S19 mutation.

Diamond-Blackfan anemia is an autosomal dominant syndrome, characterized by anemia and a predisposition for malignancies. Ribosomal proteins are responsible for this syndrome, and the incidence of colorectal cancer in patients with this syndrome is higher than the general population. This patient's Diamond-Blackfan anemia was caused by a novel ribosomal protein S19 gene mutation, and he received chemotherapy for colorectal cancer caused by it. In his cancer, ribosomal proteins S19 and TP53 were overexpressed. He received 5FU and cetuximab; however, his anemia made chemotherapy difficult, and he did not survive long. Patients with Diamond-Blackfan anemia should be screened earlier and more often for colorectal cancer than usual.

Characterization of multinuclear hepatocytes induced in rats by mitemcinal (GM-611), an erythromycin derivative.

Mitemcinal is an erythromycin derivative with motilin agonistic action, developed as a gastrointestinal motor-activating agent. The characteristics of mitemcinal-induced multinuclear hepatocytes (MNHs, hepatocytes with three or more nuclei per cell) from detailed morphological observations together with the results of a study on the mechanisms of MNH formation by combining cytocentrifuge preparations with 5-bromo-2'-deoxyuridine cumulative labeling are reported. MNHs were observed only in rats in the high-dose groups of the subchronic study, with a higher incidence in females and reversibility after twenty-eight days of drug withdrawal, but not observed in dogs. In the chronic study, the incidence increased relative to the dose. Histopathologically, MNHs were preferentially observed in the centrilobular zone, without nuclear atypia or mitotic figures. In the cell kinetic study, the labeling pattern of MNHs included all-positive, all-negative, and mixed labeling patterns of nuclei. The all-negative pattern indicated that the cells were formed by fusion of nondividing cells. The current results indicate that the cell kinetic approach effectively demonstrated the mechanism of mitemcinal-induced MNHs as fusion of hepatocytes and that drug-induced disturbance of mitosis is not involved in the multinucleation of MNHs by mitemcinal.

Monophasic action potential in anaesthetized guinea pigs as a biomarker for prediction of liability for drug-induced delayed ventricular repolarization.

INTRODUCTION: Drug-induced QT interval prolongation has been one of the critical issues for developing new chemical entities and pharmaceutical companies need to evaluate the risk early in the development stage. At such stage, guinea pigs are appropriate due to their small size requiring only small amounts of test drugs. The purpose of this study was to determine the utility of guinea pig monophasic action potential (MAP) using 12 reference drugs in order to clarify prediction of the QT interval prolonging risk. METHODS: Male guinea pigs were anaesthetized with pentobarbital (40 mg/kg, i.p.). Parameters analyzed were epicardial MAP duration (MAP(90)) at sinus rhythm (MAP(90(sinus))) and MAP(90) during atrial pacing (MAP(90(pacing))). Test drugs were administered to animals intravenously and cumulatively. RESULTS: Vehicle control did not affect the parameters tested. All 8 QT-prolonging drugs prolonged MAP(90(sinus)) and MAP(90(pacing)) dose-dependently, whereas all 4 non-QT-prolonging drugs showed no or very slight prolongations of these MAP(90) parameters. Rank order potency of MAP(90(pacing)) prolongations by the QT-prolonging drugs tended to correspond to clinical plasma concentrations associated with QT interval prolongations or Torsades de Pointes but showed less of a link with hERG inhibition activities. CONCLUSION: The present study demonstrates that the MAP model using anaesthetized guinea pigs could predict the liability of drugs for QT interval prolongation with high accuracy. QT assessment using the combination of the hERG assay with high sensitivity and the current in vivo assay would be desirable for early risk assessment within drug development.

Preclinical electrophysiology assays of mitemcinal (GM-611), a novel prokinetic agent derived from erythromycin.

Mitemcinal (GM-611) is a novel erythromycin-derived prokinetic agent that acts as an agonist at the motilin receptor. Erythromycin has shown QT prolongation and torsades de pointes (TdP) in humans and cisapride, a second class of prokinetic agents typified by the 5-HT(4) receptor agonist, has been terminated due to TdP. In this study an extended series of safety pharmacology protocols and evaluations have been undertaken to assess the potential risk of mitemcinal on QT prolongation or proarrhythmic effects. Mitemcinal and its metabolites, GM-577 and GM-625, inhibited the human ether-a-go-go-related gene (HERG) tail current in a concentration-dependent manner with IC(50) values of 20.2, 41.7, and 55.0 microM, respectively. Administration of 10 mg/kg mitemcinal in anesthetized guinea pigs resulted in a slight prolongation of the monophasic action potential (MAP) duration during atrial pacing at the plasma concentration of mitemcinal 1.1 microM, with low maximum increases in MAPD(70) (6.6%) and MAPD(90) (4.6%) relative to vehicle. A 10-min infusion of 20 mg/kg of mitemcinal in a proarrhythmic rabbit model did not evoke TdP even when QT and corrected QT (QTc) intervals were significantly prolonged. In this study, the Cmax plasma-free concentration of mitemcinal indicates that the prolongation was more than 400-fold that of the therapeutic dose. Our findings of a wide safety margin and the absence of TdP within this margin suggest that mitemcinal may provide sufficient safety in clinical use.

Hemodynamic and electrophysiological effects of mitemcinal (GM-611), a novel prokinetic agent derived from erythromycin in a halothane-anesthetized canine model.

Mitemcinal (GM-611) is a novel erythromycin-derived prokinetic agent that acts as an agonist at the motilin receptor. We investigated the QT-prolonging effects of mitemcinal using a halothane-anesthetized canine model. Intravenous administration of mitemcinal at doses of more than 8.3 mg/kg per 10 min significantly prolonged the QT interval corrected by Fridericia's corrections. Mitemcinal exhibited a bradycardiac effect and produced significantly greater prolongation in monophasic action potential duration (MAP(90)) at sinus rhythm compared with MAP(90) at pacing and showed reverse use-dependent prolongation of repolarization, suggesting that the negative chronotropic effect of mitemcinal potentiates the prolongation of the repolarization period. A technique using MAP/pacing electrodes allowed measurements of both MAP(90) and effective refractory period (ERP) simultaneously at the same ventricular site. Although mitemcinal slightly prolonged the MAP(90(CL400)) and ERP in comparison with the control group at the dose of 25 mg/kg per 10 min, the terminal repolarization period, the difference between MAP(90(CL400)) and ERP, did not increase suggesting the absence of a proarrhythmic effect even with a 7000-fold for the therapeutic blood concentration as free level. The electrophysiological results from mitemcinal in this study indicate that the risk of serious arrhythmia such as torsades de pointes, a major clinical concern related to QT interval prolongation, might be low.

QT-RR relationships and suitable QT correction formulas for halothane-anesthetized dogs.

Several QT correction (QTc) formulas have been used for assessing the QT liability of drugs. However, they are known to under- and over-correct the QT interval and tend to be specific to species and experimental conditions. The purpose of this study was to determine a suitable formula for halothane-anesthetized dogs highly sensitive to drug-induced QT interval prolongation. Twenty dogs were anesthetized with 1.5% halothane and the relationship between the QT and RR intervals were obtained by changing the heart rate under atrial pacing conditions. The QT interval was corrected for the RR interval by applying 4 published formulas (Bazett, Fridericia, Van de Water, and Matsunaga); Fridericia's formula (QTcF = QT/RR(0.33)) showed the least slope and lowest R(2) value for the linear regression of QTc intervals against RR intervals, indicating that it dissociated changes in heart rate most effectively. An optimized formula (QTcX = QT/RR(0.3879)) is defined by analysis of covariance and represents a correction algorithm superior to Fridericia's formula. For both Fridericia's and the optimized formula, QT-prolonging drugs (d,l-sotalol, astemizole) showed QTc interval prolongation. A non-QT-prolonging drug (d,l-propranolol) failed to prolong the QTc interval. In addition, drug-induced changes in QTcF and QTcX intervals were highly correlated with those of the QT interval paced at a cycle length of 500 msec. These findings suggest that Fridericia's and the optimized formula, although the optimized is a little bit better, are suitable for correcting the QT interval in halothane-anesthetized dogs and help to evaluate the potential QT prolongation of drugs with high accuracy.

Retinal toxicity induced by small-molecule Hsp90 inhibitors in beagle dogs.

Heat shock protein 90 (Hsp90) is a constitutively expressed molecular chaperone and plays an important role in the folding of client proteins with key regulatory roles in growth, survival, differentiation and metastasis. Because inhibition of Hsp90 degrades multiple oncogenic client proteins, it is considered to be an attractive anticancer therapy, and clinical trials of several Hsp90 inhibitors have been carried out. In the present study, two structurally distinct Hsp90 inhibitors, CH5164840 and CH5449302, were orally administered to beagle dogs to evaluate systemic toxicity. CH5164840 induced symptoms that suggest visual disorder, and ophthalmological observation and electroretinography (ERG) revealed loss of pupillary light reflex and abnormal waveforms, respectively. Histopathological examination showed changes in the photoreceptor cell layer and the outer nuclear layer of retina. On the other hand, while there were no clinical symptoms related to visual disorder, animals treated with CH5449302 showed similar abnormalities of ERG responses and histopathological changes in the photoreceptor cell layer and the outer nuclear layer of retina. The visual symptoms and abnormalities of ERG responses were noted at an earlier stage or lower dose than other toxicities in both compounds. Considering that two structurally distinct Hsp90 inhibitors induced a retinal toxicity in dogs after repeated administration, and that visual disorders were also reported in some clinical trials of Hsp90 inhibitors, it would seem highly likely that Hsp90 inhibition induces retinal toxicity. Also, our study indicated that a detailed ocular examination to evaluate the safety of Hsp90 inhibitors would be useful in both preclinical and clinical studies.

Proteomic identification of the macrophage-capping protein as a protein contributing to the malignant features of hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most deadly cancers worldwide. We performed a proteomic study to understand the molecular mechanisms underlying metastasis in HCC. Among the 3491 protein spots observed by two-dimensional difference gel electrophoresis (2D-DIGE), we found that 197 and 88 protein spots had statistically significant differences in intensity between tumor and non-tumor tissues and between the tumors with and without vascular invasion, respectively. Mass spectrometry was used to identify the proteins corresponding to those protein spots. We found that compared to tumor tissues without vascular invasion, those with vascular invasion showed markedly upregulated expression of the macrophage-capping protein (CapG). The association of increased CapG expression with vascular invasion in the tumor tissues was confirmed by western blotting. CapG expression levels were equal for non-tumor tissues and tumor tissues without venous invasion, as assessed by 2D-DIGE and western blotting. Silencing of CapG reduced tumor invasion without affecting the proliferation of the HCC cells. These observations suggested that CapG is involved in the process of metastasis by promoting the invasiveness of tumor cells. It may therefore be worth investigating the clinical usefulness of CapG as a biomarker in risk-stratification therapy and as a therapeutic target in HCC.

Synthesis of an insulated molecular wire by click polymerization.

We developed a new method for the synthesis of an organic-soluble insulated molecular wire (IMW) with permethylated cyclodextrin (PMCD); this method involves click polymerization of linked [2]rotaxane containing azide and alkynyl groups at both ends of a π-conjugated guest.

Accurate detection of drug-induced delayed ventricular repolarization with a suitable correction formula in Langendorff guinea pig heart.

The aims of this study were to determine a suitable method to correct the ventricular repolarization period against the RR interval in isolated perfused Langendorff guinea pig heart and to clarify the reliability of this model using several drugs. QT and RR intervals from an electrocardiogram and the epicardial monophasic action potential duration (MAP(90)) were measured. Two drugs clinically known to be QT-prolonging (E-4031, moxifloxacin) and two known to be non-QT-prolonging (verapamil, zatebradine) were used for the study. To determine a method of correcting the ventricular repolarization period against RR interval, heart rates were slowed with 0.3 µM zatebradine, a specific bradycardiac agent, and then accelerated with atrial pacing to obtain a wide range of MAP(90)/RR relationships. An exponential rate-correction model elicited the most appropriate algorithm for the relationship among the four models tested. Based on linear regression analysis, the exponential showed superior dissociation of corrected MAP(90)s against RR intervals than generic Bazett's and Fridericia's formulae. E-4031 and moxifloxacin prolonged the corrected QT (QTc) intervals and MAP(90) under atrial pacing at a cycle length of 0.25 sec (MAP(90(pacing))) dose-dependently; verapamil and zatebradine failed to prolong them, indicating that the reliability of this model was excellent. MAP(90(pacing)) prolongation by moxifloxacin, the positive compound in the clinical "Thorough QT/QTc Study", was seen at around QTc-prolonging concentrations in clinic, suggesting that the sensitivity would be appropriate for QT evaluation. We therefore concluded that the isolated guinea pig heart model is sufficiently sensitive and useful for assessing the potential QT prolongation of drugs.

Application of probabilistic analysis for precisely correcting the QT interval for heart rate in telemetered common marmosets.

INTRODUCTION: QT intervals are strongly influenced by preceeding heart rate history and are also characterized by rate-independent variability, leading to difficulty in precise rate-correction of the raw QT interval. The present study elucidates a novel analytical method that effectively addresses this problematic phenomenon in telemetered common marmosets. METHODS: ECGs were collected from telemetered common marmosets (male and female) and analyzed by computerized algorithms. Descriptive statistics were calculated from the mean of QT intervals for 5-ms increments of RR. The QT interval was corrected for the RR interval by applying Bazett's, Fridericia's, and individual probabilistic QT rate-correction formulae. RESULTS: The linear regression of log-transformed QT and RR intervals derived from a probabilistic approach yielded a well-correlated QT-RR fit. Assessed as the slope of the QTc-RR interval, application of individual probabilistic QT rate-corrections resulted in the most effective dissociation of the effects of rate from the raw QT interval, compared to generic rate-correction formulae. Using individual corrections, the QTc was stable while the interquartile range (IQR) of the QTc distribution was stable, spanning 5-10 ms for each subject over all physiological RR intervals. Heart rate variability distributions were centered about unity during both photoperiods and sinus arrhythmia was far less pronounced compared with measurements in dogs. DISCUSSION: Probabilistic QT rate-correction eliminated the confounding effects of heart rate and provided a stable QTc baseline. These results indicate that application of this method of analysis in telemetered common marmosets results in a high degree of sensitivity for the consistent detection of small (5-10 ms) changes in the QTc interval.

Sensitivity of common marmosets to detect drug-induced QT interval prolongation: moxifloxacin case study.

INTRODUCTION: Moxifloxacin is the most widely used positive reference agent in clinical cardiac repolarization studies, but it has not been characterized in common marmosets which are uniquely suited to studies in early-stage development due to their small size and minimal test article requirements. The purpose of this study was to evaluate the sensitivity of the common marmoset to detect moxifloxacin-associated QT interval prolongation. METHODS: Eight telemetered common marmosets were monitored for 24 h following oral administration of moxifloxacin by gavage at 0, 10, 30, and 100 mg/kg using a Latin square design. Concurrently, a pharmacokinetic evaluation in 8 non-telemetered animals was conducted. A rate-corrected QT (QTc) interval was derived using an individual probabilistic QT rate-correction. QTc (placebo-adjusted QTc change from the individual baseline) was calculated and the relationship between pharmacokinetics (PK) and pharmacodynamics (PD) was analyzed. RESULTS: A slight, but not significant, increase in QTc was detected with 10 mg/kg of moxifloxacin. Moxifloxacin at 30 and 100 mg/kg elicited dose-dependent increases in QTc of 14.0+/-3.6 and 35.0+/-6.2 ms, respectively, with associated total moxifloxacin C(max) values of 6.5+/-0.5 and 16.5+/-1.6 microg/mL, respectively. From the PK/PD relationship, the plasma concentration which would attain QTc of 5 to 10 ms was estimated to be 1.67-3.73 microg/mL. The results were consistent with typical clinical trial results (QTc of 6.6-14.8 ms at 2.5-3.5 microg/mL). CONCLUSIONS: The present study demonstrates that the common marmoset is highly sensitive to moxifloxacin-associated changes in cardiac repolarization, assessed as QTc. As such, this species is suitable for precise and reliable detection of small, but significant, drug-associated increases in QTc interval. Thus, the common marmoset should be regarded as a validated animal model for the detection of QT risk in early-stage drug development and represents an important addition to the current in vivo armamentarium.

Cardiovascular safety profile of MA-2029, a novel motilin receptor antagonist.

The aim of this study was to assess the cardiovascular effect of MA-2029, a selective motilin receptor antagonist highly expected for the treatment of irritable bowel syndrome (IBS). MA-2029 inhibited the human ether-a-go-go-related gene (hERG) current at 100 microg/ml, but shortened action potential duration (APD) in isolated guinea pig papillary muscles at 10 and 100 microg/ml and the corrected QT (QTc) interval after oral administration of 30 and 300 mg/kg in conscious telemetered dogs. The discrepancy was probably caused by blockade of the Ca(2+) channel because MA-2029 inhibited the Ca(2+) current in isolated guinea pig myocytes. MA-2029 at 100 microg/ml also decreased the maximum rising velocity and action potential amplitude in the action potential study, indicating that MA-2029 has Na(+) channel blocking potential. In the cardiovascular study, MA-2029 at 30 mg/kg induced slight cardiovascular changes such as hypotension, QTc shortening, and PR prolongation possibly caused by Ca(2+) channel blockade. The plasma concentration at 4 hr after 30 mg/kg administration was 2.10 microg/ml, 200-fold higher than the effective concentration of MA-2029 as a motilin receptor antagonist. These results suggest that MA-2029 has sufficient cardiovascular safety although it inhibits multiple ion channels at supra-effective concentrations. On the other hand, cisapride, an effective IBS drug, showed clear hERG inhibition and APD prolongation at 100 ng/ml. Cisapride exhibited a narrow safety margin because it caused QT prolongation potential even at the therapeutic concentration. In conclusion, MA-2029 is a novel drug highly expected for the treatment of IBS with lower cardiovascular risk than cisapride.

Assessment of the carcinogenic potential of mitemcinal (GM-611): increased incidence of malignant lymphoma in a rat carcinogenicity study.

Mitemcinal is an erythromycin derivative, which acts as an agonist of the motilin receptor. For assessment of the carcinogenicity of mitemcinal, we conducted a short-term carcinogenicity study in p53 (+/-) C57BL/6 mice and a 104-week carcinogenicity study in CD(SD)IGS rats. There was no evidence of a carcinogenic potential in mouse when administered for 26 consecutive weeks at levels up to 250 mg/kg/day. In the rat study, an increased incidence of lymphoma was noted in 5/60 males and 8/60 females of the high dose group (60 mg/kg/day) compared to 1/60 and 0/60 in control males and females, respectively, with statistical significance in females. Rat lymphomas include different immunomorphologic types (T- or B-cell lineage). Immunohistochemical analysis revealed that lymphomas from mitemcinal-treated rats and spontaneous cases were of T-cell lineage. The overall weight of evidence suggests that the incidence of spontaneous lymphoma was enhanced in the rat study. They also indicate that the increased incidence of lymphomas was based on a non-genotoxic effect with a threshold dose-response and that the tumorigenesis was based on the strain or species specificity of background factors. The high dose in the rat study is approximately 1600-fold higher (AUC) than that of the clinical dose, a sufficient margin of safety for the clinical dose. We conclude that the risk of carcinogenesis due to mitemcinal in humans can be considered to be minimal and is to represent an acceptable risk for the continued administration of mitemcinal to humans.