Hirobumi Suzuki

ITZ-1, a Client-Selective Hsp90 Inhibitor, Efficiently Induces Heat Shock Factor 1 Activation

ITZ-1 is a chondroprotective agent that inhibits interleukin-1beta-induced matrix metalloproteinase-13 (MMP-13) production and suppresses nitric oxide-induced chondrocyte death. Here we describe its mechanisms of action. Heat shock protein 90 (Hsp90) was identified as a specific ITZ-1-binding protein. Almost all known Hsp90 inhibitors have been reported to bind to the Hsp90 N-terminal ATP-binding site and to simultaneously induce degradation and activation of its multiple client proteins. However, within the Hsp90 client proteins, ITZ-1 strongly induces heat shock factor-1 (HSF1) activation and causes mild Raf-1 degradation, but scarcely induces degradation of a broad range of Hsp90 client proteins by binding to the Hsp90 C terminus. These results may explain ITZ-1s inhibition of MMP-13 production, its cytoprotective effect, and its lower cytotoxicity. These results suggest that ITZ-1 is a client-selective Hsp90 inhibitor.

TAK-063, a PDE10A Inhibitor with Balanced Activation of Direct and Indirect Pathways, Provides Potent Antipsychotic-Like Effects in Multiple Paradigms

Phosphodiesterase 10A (PDE10A) inhibitors are expected to be novel drugs for schizophrenia through activation of both direct and indirect pathway medium spiny neurons. However, excess activation of the direct pathway by a dopamine D1 receptor agonist SKF82958 canceled antipsychotic-like effects of a dopamine D2 receptor antagonist haloperidol in methamphetamine (METH)-induced hyperactivity in rats. Thus, balanced activation of these pathways may be critical for PDE10A inhibitors. Current antipsychotics and the novel PDE10A inhibitor TAK-063, but not the selective PDE10A inhibitor MP-10, produced dose-dependent antipsychotic-like effects in METH-induced hyperactivity and prepulse inhibition in rodents. TAK-063 and MP-10 activated the indirect pathway to a similar extent; however, MP-10 caused greater activation of the direct pathway than did TAK-063. Interestingly, the off-rate of TAK-063 from PDE10A in rat brain sections was faster than that of MP-10, and a slower off-rate PDE10A inhibitor with TAK-063-like chemical structure showed an MP-10-like pharmacological profile. In general, faster off-rate enzyme inhibitors are more sensitive than slower off-rate inhibitors to binding inhibition by enzyme substrates. As expected, TAK-063 was more sensitive than MP-10 to binding inhibition by cyclic nucleotides. Moreover, an immunohistochemistry study suggested that cyclic adenosine monophosphate levels in the direct pathway were higher than those in the indirect pathway. These data can explain why TAK-063 showed partial activation of the direct pathway compared with MP-10. The findings presented here suggest that TAK-063’s antipsychotic-like efficacy may be attributable to its unique pharmacological properties, resulting in balanced activation of the direct and indirect striatal pathways.

BTZO-15, an ARE-Activator, Ameliorates DSS- and TNBS-Induced Colitis in Rats

Inflammatory bowel disease (IBD) is a group of chronic inflammatory disorders that are primarily represented by ulcerative colitis and Crohns disease. The etiology of IBD is not well understood; however, oxidative stress is considered a potential etiological and/or triggering factor for IBD. We have recently reported the identification of BTZO-1, an activator of antioxidant response element (ARE)-mediated gene expression, which protects cardiomyocytes from oxidative stress-induced insults. Here we describe the potential of BTZO-15, an active BTZO-1 derivative for ARE-activation with a favorable ADME-Tox profile, for the treatment of IBD. BTZO-15 induced expression of heme oxygenase-1 (HO-1), an ARE-regulated cytoprotective protein, and inhibited NO-induced cell death in IEC-18 cells. Large intestine shortening, rectum weight gain, diarrhea, intestinal bleeding, and an increase in rectal myeloperoxidase (MPO) activity were observed in a dextran sulfate sodium (DSS)-induced colitis rat model. Oral administration of BTZO-15 induced HO-1 expression in the rectum and attenuated DSS-induced changes. Furthermore BTZO-15 reduced the ulcerated area and rectal MPO activity in 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced colitis rats without affecting rectal TNF-α levels. These results suggest that BTZO-15 is a promising compound for a novel IBD therapeutic drug with ARE activation properties.

FTBMT, a Novel and Selective GPR52 Agonist, Demonstrates Antipsychotic-Like and Procognitive Effects in Rodents, Revealing a Potential Therapeutic Agent for Schizophrenia

GPR52 is a Gs-coupled G protein–coupled receptor that is predominantly expressed in the striatum and nucleus accumbens (NAc) and was recently proposed as a potential therapeutic target for schizophrenia. In the current study, we investigated the in vitro and in vivo pharmacologic activities of a novel GPR52 agonist, 4-(3-(3-fluoro-5-(trifluoromethyl)benzyl)-5-methyl-1H-1,2,4-triazol-1-yl)-2-methylbenzamide (FTBMT). FTBMT functioned as a selective GPR52 agonist in vitro and in vivo, as demonstrated by the activation of Camp signaling in striatal neurons. FTBMT inhibited MK-801–induced hyperactivity, an animal model for acute psychosis, without causing catalepsy in mice. The c-fos expression also revealed that FTBMT preferentially induced neuronal activation in the shell of the Nac compared with the striatum, thereby supporting its antipsychotic-like activity with less catalepsy. Furthermore, FTBMT improved recognition memory in a novel object-recognition test and attenuated MK-801–induced working memory deficits in a radial arm maze test in rats. These recognitive effects were supported by the results of FTBMT-induced c-fos expression in the brain regions related to cognition, including the medial prefrontal cortex, entorhinal cortex, and hippocampus. Taken together, these findings suggest that FTBMT shows antipsychotic and recognitive properties without causing catalepsy in rodents. Given its unique pharmacologic profile, which differs from that of current antipsychotics, FTBMT may provide a new therapeutic option for the treatment of positive and cognitive symptoms of schizophrenia.

Combined treatment with a selective PDE10A inhibitor TAK‐063 and either haloperidol or olanzapine at subeffective doses produces potent antipsychotic‐like effects without affecting plasma prolactin levels and cataleptic responses in rodents

Activation of indirect pathway medium spiny neurons (MSNs) via promotion of cAMP production is the principal mechanism of action of current antipsychotics with dopamine D2 receptor antagonism. TAK‐063 [1‐[2‐fluoro‐4‐(1H‐pyrazol‐1‐yl)phenyl]‐5‐methoxy‐3‐(1‐phenyl‐1H‐pyrazol‐5‐yl)pyridazin‐4(1H)‐one] is a novel phosphodiesterase 10A inhibitor that activates both direct and indirect pathway MSNs through increasing both cAMP and cGMP levels by inhibition of their degradation. The activation of indirect pathway MSNs through the distinct mechanism of action of these drugs raises the possibility of augmented pharmacological effects by combination therapy. In this study, we evaluated the potential of combination therapy with TAK‐063 and current antipsychotics, such as haloperidol or olanzapine after oral administration. Combined treatment with TAK‐063 and either haloperidol or olanzapine produced a significant increase in phosphorylation of glutamate receptor subunit 1 in the rat striatum. An electrophysiological study using rat corticostriatal slices showed that TAK‐063 enhanced N‐methyl‐D‐aspartic acid receptor‐mediated synaptic responses in both direct and indirect pathway MSNs to a similar extent. Further evaluation using pathway‐specific markers revealed that coadministration of TAK‐063 with haloperidol or olanzapine additively activated the indirect pathway, but not the direct pathway. Combined treatment with TAK‐063 and either haloperidol or olanzapine at subeffective doses produced significant effects on methamphetamine‐ or MK‐801‐induced hyperactivity in rats and MK‐801‐induced deficits in prepulse inhibition in mice. TAK‐063 at 0.1 mg/kg did not affect plasma prolactin levels and cataleptic response from antipsychotics in rats. Thus, TAK‐063 may produce augmented antipsychotic‐like activities in combination with antipsychotics without effects on plasma prolactin levels and cataleptic responses in rodents.

Etiology and long-term rebleeding of endoscopic ulcerative lesions in the small bowel in patients with obscure gastrointestinal bleeding: A multicenter cohort study: Management of small bowel ulcerations

Among patients with obscure gastrointestinal bleeding (OGIB), endoscopic ulcerative lesions in the small bowel have diverse etiologies and often cause rebleeding. Certain characteristics of patients or ulcerations may be reasonable indications for diagnostic balloon‐assisted endoscopy (BAE) to assess etiology and may be risks of rebleeding; however, these characteristics are unclear. We aimed to elucidate appropriate indications for diagnostic BAE and predictors of long‐term rebleeding in patients with small bowel ulcerative lesions.

Colonoscopy reduces colorectal cancer mortality: A multicenter, long-term, colonoscopy-based cohort study

Background There are limited colonoscopy-based cohort data concerning the effectiveness of colonoscopy in reducing colorectal cancer deaths. The aim of this study was to clarify whether colonoscopy reduces colorectal cancer mortality. Methods A cohort of 18,816 patients who underwent colonoscopy without a diagnosis of colorectal cancer between 2001 and 2010 at high colonoscopy procedure volume centers was selected. Patient characteristics and colonoscopy findings were assessed. The main endpoint was colorectal cancer death (all, right-sided, and left-sided cancers), and data were censored at the time of the final visit or the final colonoscopy. The standardized all colorectal, colon, and rectal cancer mortality rates were estimated with reference to those of the general Japanese population. Additional outcome was all- cause death and the standardized all-cause mortality rate was also estimated. Results The total observed person-year mortality for colorectal cancer was 67,119. Of these, 4, 3, and 1 patients died from colorectal, colon, and rectal cancers, respectively; these values were significantly lower than the number of expected deaths in the general population, estimated to be 53.1, 34.0, and 19.1, respectively. The standardized mortalities for all colorectal, colon, and rectal cancers were 0.08 (95% confidence interval (CI), 0.02–0.17), 0.09 (95% CI, 0.02–0.22), and 0.05 (95% CI, 0.0002–0.21), respectively. There were 586 all-cause deaths (3.11%) during the observation period. The standardized all-cause mortality ratios were 0.22 (95% CI, 0.206–0.23). Conclusions The colorectal cancer mortality of patients who received colonoscopy without colorectal cancer diagnosis decreased significantly compared with that of individuals in the general population. These results were compatible even in patients with right-sided colon cancer.

BTZO-2, an antioxidant response element-activator, provides protection against lethal endotoxic shock in mice.

We recently reported a unique antioxidant response element (ARE)-activator, BTZO-1, which induced expression of cytoprotective proteins such as heme oxygenase-1 (HO-1) and suppressed oxidative stress-induced cardiomyocyte apoptosis via binding to macrophage migration inhibitory factor (MIF). HO-1 induction and apoptosis inhibition have been reported to improve the outcomes following experimental sepsis by protecting the organs. Therefore, we investigated the potential of BTZO-2, an active BTZO-1 derivative, as a drug for sepsis. BTZO-2 significantly protected mice from the endotoxic shock induced by 5mg/kg lipopolysaccharide (LPS); survival rates increased from 42% to 100%. In contrast, BTZO-2 did not provide significant protection to mice from the shock induced by 10 μg/kg LPS together with d-galactosamine (d-GalN, hepatocyte-specific transcription inhibitor) (LPS/d-GalN). Hepatic HO-1 protein was up-regulated by BTZO-2 in mice injected with 5mg/kg LPS, but not in those injected with 10 μg/kg LPS/d-GalN. Interestingly, BTZO-2 showed little or no effect on LPS-induced up-regulation of plasma cytokine levels in mice. Thus, the organ protection mediated by HO-1 may have a pivotal role in the pharmacological effect of BTZO-2. These results suggest that BTZO-2 is a promising compound for a novel drug for sepsis.

S223. COMBINED TREATMENT WITH A SELECTIVE PDE10A INHIBITOR TAK-063 AND ANTIPSYCHOTICS AT SUBEFFECTIVE DOSES PRODUCES POTENT ANTIPSYCHOTIC-LIKE EFFECTS WITHOUT EXACERBATING SIDE EFFECTS PROFILE IN RODENTS

Abstract Background Activation of indirect pathway medium spiny neurons (MSNs) via promotion of cAMP production is the principal mechanism of action (MOA) of current antipsychotics with dopamine D2 receptor antagonism. Phosphodiesterase 10A (PDE10A) inhibitors activate both direct and indirect pathway MSNs by increasing both cAMP and cGMP levels by inhibiting their degradation, which might be expected to promote activation of intracellular signaling similar to that of D2 antagonists in the indirect pathway MSNs. Thus, the activation of the indirect MSN pathway through the distinct MOA of these compounds raises the possibility of augmented pharmacologic effects with combined treatment. In this study, we compared gene-regulation patterns in the indirect pathway MSNs induced by the PDE10A inhibitors T-773 and T-609, and the D2 antagonist haloperidol, using a cell-type-specific comprehensive gene expression analysis in Drd2-bacTRAP (translating ribosome affinity purification) mice. The pharmacologic effects of combined treatment with another PDE10A inhibitor, TAK-063, and clinically used antipsychotics, haloperidol (HAL) and olanzapine (OLA), were evaluated in multiple rodent models. Methods Male ICR mice, Drd2-bacTRAP mice, and Sprague-Dawley rats were used. The indirect pathway MSN-specific gene expression changes by T-773, T-609, and HAL were investigated using RNA sequencing of striatal samples of Drd2-bacTRAP mice. The activation of MSNs in rats was evaluated by measuring glutamate receptor subunit 1 phosphorylation (pGluR1) levels. An in vitro electrophysiological study on the corticostrial pathway in rats was conducted in a slice preparation. The activation of each MSN pathway was assessed by inducing genes as pathway-specific markers: enkephalin for the indirect pathway and substance P for the direct pathway. Suppression of MK-801- or methamphetamine (METH)-induced hyperactivity was assessed by measuring locomotor activity for 2 hours after administration of these stimulants to rats. Improvement of prepulse inhibition (PPI) was investigated in a MK-801-induced PPI deficit mouse model. Results Translational profiling in Drd2-bacTRAP mice treated with the PDE10A selective inhibitors, T-773 and T-609, and with HAL suggested regulatory of a largely overlapping signaling pathway by these compounds in the indirect pathway MSNs: 87% of the genes regulated by HAL were also regulated by both T-773 and T-609. Combined treatment with TAK-063 and either HAL or OLA produced an augmented effect on pGluR1 in the rat striatum. An electrophysiological study in rat brain slices indicated that TAK-063 enhanced synaptic responses to a similar extent in both direct and indirect pathway MSNs. Additional evaluation using MSN pathway-specific markers revealed that coadministration of TAK-063 with HAL or OLA additively activated the indirect pathway, but not the direct pathway. Combined treatment with TAK-063 (0.1 mg/kg p.o.) and either HAL (0.3 mg/kg p.o.) or OLA (3 mg/kg p.o.) at subeffective doses produced augmented effects on METH- or MK-801–induced hyperactivity in rats and MK-801–induced PPI deficits in mice. TAK-063 at 0.1 mg/kg did not affect plasma prolactin levels and cataleptic response induced by HAL or OLA in rats. Discussion PDE10A inhibitors and HAL showed similar patterns of gene regulation in indirect pathway MSNs in mice. Combined treatment with TAK-063 and either HAL or OLA at subeffective doses produced significant antipsychotic-like effects but no augmentation of the plasma prolactin level and cataleptic response. Although further preclinical and clinical studies will be needed, TAK-063 may provide a novel mechanism as a PDE10A inhibitor for use as combination therapy in schizophrenia.

Genetic deletion of GPR52 enhances the locomotor-stimulating effect of an adenosine A2A receptor antagonist in mice: A potential role of GPR52 in the function of striatopallidal neurons

G protein-coupled receptor 52 (GPR52) is largely co-expressed with dopamine D2 receptor (DRD2) in the striatum and nucleus accumbens, and this expression pattern is similar to that of adenosine A2A receptor (ADORA2A). GPR52 has been proposed as a therapeutic target for positive symptoms of schizophrenia, based on observations from pharmacological and transgenic mouse studies. However, the physiological role of GPR52 in dopaminergic functions in the basal ganglia remains unclear. Here, we used GPR52 knockout (KO) mice to examine the role of GPR52 in dopamine receptor-mediated and ADORA2A-mediated locomotor activity and dopamine receptor signaling. High expression of GPR52 protein in the striatum, nucleus accumbens, and lateral globus pallidus of wild type (WT) littermates was confirmed by immunohistochemical analysis. GPR52 KO and WT mice exhibited almost identical locomotor responses to the dopamine releaser methamphetamine and the N-methyl-d-aspartate antagonist MK-801. In contrast, the locomotor response to the ADORA2A antagonist istradefylline was significantly augmented in GPR52 KO mice compared to WT mice. Gene expression analysis revealed that striatal expression of DRD2, but not of dopamine D1 receptor and ADORA2A, was significantly decreased in GPR52 KO mice. Moreover, a significant reduction in the mRNA expression of enkephalin, a marker of the activity of striatopallidal neurons, was observed in the striatum of GPR52 KO mice, suggesting that GPR52 deletion could enhance DRD2 signaling. Taken together, these results imply the physiological relevance of GPR52 in modulating the function of striatopallidal neurons, possibly by interaction of GPR52 with ADORA2A and DRD2.