Discovery of a novel 9-position modified second-generation anti-HCV candidate via bioconversion and semi-synthesis of FR901459.

Evidence that hepatitis C virus (HCV) utilizes cellular cyclophilin proteins in the virus replication cycle has increased attention on cyclophilin inhibitors as attractive therapeutic targets in the treatment of HCV. Previous reports have described a number of non-immunosuppressive cyclophilin inhibitors, most of which require many synthetic steps for their preparation. Sasamura et al. have previously reported the isolation of bioconversion derivative 4. This analog is a convenient starting point for optimization due to the presence of the readily modifiable primary hydroxyl group and because it shows moderate anti-HCV activity and decreased immunosuppressive activity. We have also established an efficient C-alkylation reaction at the 3-position. Through a detailed structure-activity relationship study, we discovered a new type of clinical candidate 14 which requires a short synthetic process and has potent anti-HCV activity and reduced immunosuppressive activity, as well as improved aqueous solubility and pharmacokinetics.

Population doublings of murine CD4(+) memory T cells during continuous antigen stimulation in vivo.

We investigated the expansion rate of CD4(+) memory T cells using a newly developed in vivo system. Neonatal thymectomy abrogates the subsequent production of T cells and induces autoimmune gastritis (AIG) by the activation of CD4(+) T cells; this disease was transferred into athymic nude mice through the inoculation of splenic CD4(+) memory T cells. The transferred CD4(+) T cells increased logarithmically in number during the first 2months in the spleen of the recipients. The serial transfer of these splenocytes at two-month intervals revealed that the numbers of the AIG-transferable generations were inversely correlated with the age of the first AIG donors. The duration of the AIG-promoting capacity of CD4(+) T cells under continuous antigenic stimulation in vivo was approximately equivalent-one and a half years. These results indicate that there exists an intrinsic population doubling limit in memory CD4(+) T cells similar to that of self-renewing naïve ones.

Synthesis of neoglycosphingolipid from methoxyamino-functionalized ceramide.

An efficient approach for the synthesis of a methoxyamino-functionalized ceramide was established from phytosphingosine using specific Nß→Nα acyl migration of the octadecanoyl group during the removal of Nα-Fmoc protective group. One step glycoblotting reaction of the ceramide mimic with lactose afforded a neoglycosphingolipid showing potent inhibitory activity against recombinant endoglycoceramidase II from Rhodococcus sp.

Crossmodal correspondences between visual features and tastes in preschoolers: an exploratory study.

Introduction: Adults possess a natural inclination to associate sensory cues derived from distinct modalities, such as the pairing of sweet with pink. However, studies exploring crossmodal correspondences in children, particularly in the sensory pairing of visual features and tastes, are scant, leaving unanswered questions regarding the developmental trajectory of crossmodal correspondences. The present study investigates whether Japanese preschool children demonstrate specific biases in shape-color, shape-taste, and color-taste associations. Methods: In a series of in-person experiments, 92 children between 3 to 6 years of age completed matching tasks utilizing paper stimuli. Results: Children exhibit crossmodal correspondences in shape-color (circle-red and asymmetrical star-yellow), shape-taste (triangle-salty and circle-sweet), and color-taste (yellow-sour, black-bitter, and pink-sweet) associations. Moreover, children's choices are not influenced by their individual preferences. Discussion: The crossmodal correspondences observed in this study have been observed in previous research on adults from the same (Japanese) culture, although adults showed more crossmodal correspondences than the children in this study (e.g., pink-circle, triangle-sour, and green-bitter). Thus, while some crossmodal correspondences emerge during childhood, others may require additional time to develop, thereby highlighting the importance of understanding the cognitive mechanisms underlying crossmodal correspondences from an ontogenic perspective.

Suppression of lactate production in fed-batch culture of some lactic acid bacteria with sucrose as the carbon source.

We report a method for suppression of lactate production by lactic acid bacteria (LAB) in culture. LAB produce lactate to regenerate NAD+ that is consumed during glycolysis. Glucose suppresses NAD+ regeneration pathways other than lactate dehydrogenase and non-glycolytic ATP production pathways. Therefore, the carbon source was changed to sucrose, and fed-batch culture was performed to limit the glycolytic flux and thus suppress lactate production. As a result, lactate productivity (i.e., the amount of lactate produced per amount of grown cell) in the sucrose/fed-batch culture was decreased compared to that in glucose/batch culture, in all five LAB strains examined. The productivity level decreased to 24% and 46% in Lactobacillus reuteri JCM 1112 and Lactococcus lactis JCM 7638, respectively. Metabolic flux analysis of Lactobacillus reuteri JCM 1112 revealed increased contributions of the mannitol production pathway to NAD+ regeneration and the arginine deiminase pathway to ATP production in the sucrose/fed-batch culture.

Synthesis and Characterization of N,N-Dimethylformamide-Protected Palladium Nanoparticles and Their Use in the Suzuki-Miyaura Cross-Coupling Reaction.

Herein, the synthesis of new N,N-dimethylformamide (DMF)-protected palladium nanoparticles (Pd NPs-OAc) employing Pd (OAc)2 (= Pd(OCOCH3)2) as the NP precursor is reported. Pd NPs-OAc were comprehensively characterized by transmission electron microscopy, FT-IR, NMR, and X-ray photoelectron spectroscopy to determine the Pd NP size distribution and the coordination state of DMF. Pd NPs-OAc were compared with Pd NPs-Cl, using PdCl2 as the NP precursor. The Suzuki-Miyaura cross-coupling reaction proceeded efficiently in the presence of Pd NPs-OAc and a high catalytic activity was observed with a turnover number of up to 1.5 × 105. Furthermore, the Pd NP-OAc catalysts could be recycled at least five times.

Discovery of quinazolinone and quinoxaline derivatives as potent and selective poly(ADP-ribose) polymerase-1/2 inhibitors.

Two classes of quinazolinone derivatives and quinoxaline derivatives were identified as potent and selective poly(ADP-ribose) polymerase-1 and 2 (PARP-1) and (PARP-2) inhibitors, respectively. In PARP enzyme assays using recombinant PARP-1 and PARP-2, quinazolinone derivatives displayed relatively high selectivity for PARP-1 and quinoxaline derivatives showed superior selectivity for PARP-2. SBDD analysis via a combination of X-ray structural study and homology modeling suggested distinct interactions of inhibitors with PARP-1 and PARP-2. These findings provide a new structural framework for the design of selective inhibitors for PARP-1 and PARP-2.

Rational approaches to discovery of orally active and brain-penetrable quinazolinone inhibitors of poly(ADP-ribose)polymerase.

A novel class of quinazolinone derivatives as potent poly(ADP-ribose)polymerase-1 (PARP-1) inhibitors has been discovered. Key to success was application of a rational discovery strategy involving structure-based design, combinatorial chemistry, and classical SAR for improvement of potency and bioavailability. The new inhibitors were shown to bind to the nicotinamide-ribose binding site (NI site) and the adenosine-ribose binding site (AD site) of NAD+.

Rational design of conformationally restricted quinazolinone inhibitors of poly(ADP-ribose)polymerase.

A successful design of conformationally restricted novel quinazolinone derivatives linked via a cyclopentene moiety as potent poly(ADP-ribose)polymerase-1 (PARP-1) inhibitors has been developed. One selected member of the new series, 8-chloro-2-[(3S)-3-(4-phenylpiperidin-1-yl)cyclopent-1-en-1-yl]quinazolin-4(3H)-one (S-16d), was found to be highly potent with IC(50)=8.7 nM and good brain penetration.

Discovery of potent and selective PARP-1 and PARP-2 inhibitors: SBDD analysis via a combination of X-ray structural study and homology modeling.

We disclose herein our efforts aimed at discovery of selective PARP-1 and PARP-2 inhibitors. We have recently discovered several novel classes of quinazolinones, quinazolidinones, and quinoxalines as potent PARP-1 inhibitors, which may represent attractive therapeutic candidates. In PARP enzyme assays using recombinant PARP-1 and PARP-2, the quinazolinone derivatives displayed relatively high selectivity for PARP-1 and quinoxaline derivatives showed superior selectivity for PARP-2, and the quinazolidinone derivatives did not have selectivity for PARP-1/2. Structure-based drug design analysis via a combination of X-ray structural study utilizing the complexes of inhibitors and human PARP-1 catalytic domain, and homology modeling using murine PARP-2 suggested distinct interactions of inhibitors with PARP-1 and PARP-2. These findings provide a new structural framework for the design of selective inhibitors for PARP-1 and PARP-2.

4-Phenyl-1,2,3,6-tetrahydropyridine, an excellent fragment to improve the potency of PARP-1 inhibitors.

We have shown that a 4-phenyl-1,2,3,6-tetrahydropyridine fragment plays an important role in improving inhibitory potency against poly(ADP-ribose) polymerase-1 (PARP-1). Various benzamide analogues linked with this fragment via alkyl spacers have been prepared and evaluated. As a result, some of them have been found to be highly potent PARP-1 inhibitors.

Neuroprotective effects of a novel poly(ADP-ribose) polymerase-1 inhibitor, 2-[3-[4-(4-chlorophenyl)-1-piperazinyl] propyl]-4(3H)-quinazolinone (FR255595), in an in vitro model of cell death and in mouse 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine model of Parkinson's disease.

The massive activation of poly(ADP-ribose) polymerase-1 (PARP-1) by DNA-damaging stimuli, such as exposure to reactive oxygen species (ROS), can lead to cell injury via severe, irreversible depletion of the NAD and ATP pool, and PARP-1 inhibitors have been expected to rescue neurons from degeneration in a number of disease models. We have recently identified 2-[3-[4-(4-chlorophenyl)-1-piperazinyl] propyl]-4(3H)-quinazolinone (FR255595) as a novel and potent PARP-1 inhibitor through structure-based drug design and high-throughput screening. This compound potently inhibited PARP activity with an IC(50) value of 11 nM and was orally active and highly brain penetrable. Here, we show that prevention of PARP activation by FR255595 protects against both ROS-induced cells injury in vitro and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced nigrostriatal dopaminergic damage in an in vivo Parkinson's disease (PD) model. In cell death models in vitro, exposure of hydrogen peroxide induced cell death with PARP overactivation in PC12 cells and SH-SY5Y cells, and pre- and post-treatment with FR255595 (10(-9)-10(-5) M) significantly reduced PARP activation and cell death. In mouse MPTP model, MPTP (20 mg/kg i.p.) intoxication lead to PARP activation and cell damage in the nigrostriatal dopaminergic pathway, which was significantly ameliorated by oral administration of FR255595 (10-32 mg/kg), both in the substantia nigra and in the striatum via marked reduction of PARP activation, even with delayed treatment. These findings clearly indicate that the novel PARP-1 inhibitor FR255595 exerts neuroprotective effect through its potent PARP-1 inhibitory actions in PD model, suggesting that the drug could be an attractive candidate for several neurodegenerative disorders, including PD.

A new poly(ADP-ribose) polymerase inhibitor, FR261529 [2-(4-chlorophenyl)-5-quinoxalinecarboxamide], ameliorates methamphetamine-induced dopaminergic neurotoxicity in mice.

Methamphetamine (METH) administration in mice, results in a chronic dopamine (DA) depletion associated with nerve terminal damage, with DA oxidation and generation of reactive oxygen species (ROS) primarily mediating this neurotoxicity. The oxidative stress induced by METH putatively activates nuclear enzyme poly(ADP-ribose) polymerase (PARP), with excessive PARP activation eventually leading to cell death. In this study, we show that prevention of PARP activation by treatment with FR261529 [2-(4-chlorophenyl)-5-quinoxalinecarboxamide], the compound that was recently identified as a novel PARP inhibitor (IC50 for PARP-1 = 33 nM, IC50 for PARP-2 = 7 nM), protects against both ROS-induced cells injury in vitro and METH-induced dopaminergic neuronal damage in an in vivo Parkinson's disease (PD) model. In PC12 cells, exposure of hydrogen peroxide or METH markedly induced PARP activation, and treatment with FR261529 (1 microM) significantly reduced PARP activation and attenuated cell death. In the mouse METH model, METH (15 mg/kg x 2 i.p., 2 h apart) intoxication accelerated DA metabolism and oxidation in the striatum, with subsequent cell damage in nigrostriatal dopaminergic neurons after 4 days. Oral administration of FR261529 (10 or 32 mg/kg) attenuated the damage of dopaminergic neurons via marked reduction of PARP activity and not via changes in dopamine metabolism or body temperature. These findings indicate that the neuroprotective effects of a novel PARP inhibitor, FR261529, were accompanied by inhibition of METH-induced PARP activation, suggesting that METH induces nigrostriatal dopaminergic neurodegeneration involving PARP activation and also orally active and brain-penetrable PARP inhibitor FR261529 could be a novel attractive therapeutic candidate for neurodegenerative disorders such as PD.

A novel and potent poly(ADP-ribose) polymerase-1 inhibitor, FR247304 (5-chloro-2-[3-(4-phenyl-3,6-dihydro-1(2H)-pyridinyl)propyl]-4(3H)-quinazolinone), attenuates neuronal damage in in vitro and in vivo models of cerebral ischemia.

The activation of poly(ADP-ribose) polymerase-1 (PARP-1) after exposure to nitric oxide or oxygen-free radicals can lead to cell injury via severe, irreversible depletion of NAD. Genetic deletion or pharmacological inhibition of PARP-1 attenuates brain injury after focal ischemia and neurotoxicity in several neurodegenerative models in animals. FR247304 (5-chloro-2-[3-(4-phenyl-3,6-dihydro-1(2H)-pyridinyl)propyl]-4(3H)-quinazolinone) is a novel PARP-1 inhibitor that has recently been identified through structure-based drug design. In an enzyme kinetic analysis, FR247304 exhibits potent and competitive inhibition of PARP-1 activity, with a K(i) value of 35 nM. Here, we show that prevention of PARP activation by FR247304 treatment protects against both reactive oxygen species-induced PC12 cell injury in vitro and ischemic brain injury in vivo. In cell death model, treatment with FR247304 (10(-8)-10(-5) M) significantly reduced NAD depletion by PARP-1 inhibition and attenuated cell death after hydrogen peroxide (100 microM) exposure. After 90 min of middle cerebral artery occlusion in rats, poly(ADP-ribosy)lation and NAD depletion were markedly increased in the cortex and striatum from 1 h after reperfusion. The increased poly(ADP-ribose) immunoreactivity and NAD depletion were attenuated by FR247304 (32 mg/kg i.p.) treatment, and FR247304 significantly decreased ischemic brain damage measured at 24 h after reperfusion. Whereas other PARP inhibitors such as 3-aminobenzamide and PJ34 [N-(6-oxo-5,6-dihydro-phenanthridin-2-yl)-N,N-dimethylactamide] showed similar neuroprotective actions, they were less potent in in vitro assays and less efficacious in an in vivo model compared with FR247304. These results indicate that the novel PARP-1 inhibitor FR247304 exerts its neuroprotective efficacy in in vitro and in vivo experimental models of cerebral ischemia via potent PARP-1 inhibition and also suggest that FR247304 or its derivatives could be attractive therapeutic candidates for stroke and neurodegenerative disease.