Identification of novel lysine demethylase 5-selective inhibitors by inhibitor-based fragment merging strategy.

Histone lysine demethylases (KDMs) have drawn much attention as targets of therapeutic agents. KDM5 proteins, which are Fe(II)/α-ketoglutarate-dependent demethylases, are associated with oncogenesis and drug resistance in cancer cells, and KDM5-selective inhibitors are expected to be anticancer drugs. However, few cell-active KDM5 inhibitors have been reported and there is an obvious need to discover more. In this study, we pursued the identification of highly potent and cell-active KDM5-selective inhibitors. Based on the reported KDM5 inhibitors, we designed several compounds by strategically merging two fragments for competitive inhibition with α-ketoglutarate and for KDM5-selective inhibition. Among them, compounds 10 and 13, which have a 3-cyano pyrazolo[1,5-a]pyrimidin-7-one scaffold, exhibited strong KDM5-inhibitory activity and significant KDM5 selectivity. In cellular assays using human lung cancer cell line A549, 10 and 13 increased the levels of trimethylated lysine 4 on histone H3, which is a specific substrate of KDM5s, and induced growth inhibition of A549 cells. These results should provide a basis for the development of cell-active KDM5 inhibitors to highlight the validity of our inhibitor-based fragment merging strategy.

Identification of novel plasminogen activator inhibitor-1 inhibitors with improved oral bioavailability: Structure optimization of N-acylanthranilic acid derivatives.

Novel plasminogen activator inhibitor-1 (PAI-1) inhibitors with highly improved oral bioavailability were discovered by structure-activity relationship studies on N-acyl-5-chloroanthranilic acid derivatives. Because lipophilic N-acyl groups seemed to be important for the anthranilic acid derivatives to strongly inhibit PAI-1, synthesis of compounds in which 5-chloroanthranilic acid was bound to a variety of highly lipophilic moieties with appropriate linkers was investigated. As the result it appeared that some of the derivatives possessing aryl- or heteroaryl-substituted phenyl groups in the acyl chain had potent in vitro PAI-1 inhibitory activity. Oral absorbability of typical compounds was also evaluated in rats, and compounds 40, 55, 60 and 76 which have diverse chemical structure with each other were selected for further pharmacological evaluation.

Design and synthesis of novel and highly-active pan-histone deacetylase (pan-HDAC) inhibitors.

Histone deacetylase (HDAC) inhibitions are known to elicit anticancer effects. We designed and synthesized several HDAC inhibitors. Among these compounds, compound 40 exhibited a more than 10-fold stronger inhibitory activity compared with that of suberoylanilide hydroxamic acid (SAHA) against each human HDAC isozyme in vitro (IC50 values of 40: HDAC1, 0.0038µM; HDAC2, 0.0082µM; HDAC3, 0.015µM; HDAC8, 0.0060µM; HDAC4, 0.058µM; HDAC9, 0.0052µM; HDAC6, 0.058µM). The dose of the administered HDAC inhibitors that contain hydroxamic acid as the zinc-binding group may be reduced by 40. Because the carbostyril subunit is a time-tested structural component of drugs and biologically active compounds, 40 most likely exhibits good absorption, distribution, metabolism, excretion, and toxicity (ADMET). Thus, compound 40 is expected to be a promising therapeutic agent or chemical tool for the investigation of life process.

Structure-activity relationships of new N-acylanthranilic acid derivatives as plasminogen activator inhibitor-1 inhibitors.

Novel anthranilic acid derivatives having substituted N-acyl side chains were designed and synthesized for evaluation as plasminogen activator inhibitor-1 (PAI-1) inhibitors. Compounds with a 4-diphenylmethyl-1-piperazinyl moiety on the acyl side chains in general exhibited potent in vitro PAI-1 inhibitory activity and good pharmacokinetic profiles after oral administration in rats. Compound 16f (TM5275) was identified as a promising candidate for further pharmacological evaluation.

A novel inhibitor of plasminogen activator inhibitor-1 provides antithrombotic benefits devoid of bleeding effect in nonhuman primates.

Inhibition of plasminogen activator inhibitor (PAI)-1 is useful to treat several disorders including thrombosis. An inhibitor of PAI-1 (TM5275) was newly identified by an extensive study of structure-activity relationship based on a lead compound (TM5007) which was obtained through virtual screening by docking simulations. Its antithrombotic efficacy and adverse effects were tested in vivo in rats and nonhuman primates (cynomolgus monkey). TM5275, administered orally in rats (1 to 10 mg/kg), has an antithrombotic effect equivalent to that of ticlopidine (500 mg/kg) in an arterial venous shunt thrombosis model and to that of clopidogrel (3 mg/kg) in a ferric chloride-treated carotid artery thrombosis model. TM5275 does not modify activated partial thromboplastin time and prothrombin time or platelet activity and does not prolong bleeding time. Combined with tissue plasminogen activator, TM5275 improves the latter's therapeutic efficacy and reduces its adverse effect. Administered to a monkey model of photochemical induced arterial thrombosis, TM5275 (10 mg/kg) has the same antithrombotic effect as clopidogrel (10 mg/kg), without enhanced bleeding. This study documents the antithrombotic benefits of a novel, more powerful, PAI-1 inhibitor in rats and, for the first time, in nonhuman primates. These effects are obtained without adverse effect on bleeding time.

A ribonucleoside with pyrimido[4,5-d]pyrimidine-2,4,5,7-(1H,3H,6H,8H)-tetraone as a nucleobase, which universally binds to natural nucleosides.

A ribonucleoside with pyrimido[4,5-d]pyrimidine-2,4,5,7-(1H,3H,6H,8H)-tetraone, which is expected to serve as a universal base, was synthesized and binding affinity of this artificial nucleoside to natural nucleosides was investigated by UV analysis. As the result, it was revealed that the artificial derivative universally forms a base pair with four kinds of natural deoxyribonucleosides.

Palladium(II)-Catalyzed Asymmetric 1,3-Dipolar Cycloaddition of Nitrones to 3-Alkenoyl-1,3-oxazolidin-2-ones.

Chiral phosphinepalladium(II)-catalyzed asymmetric 1,3-dipolar cycloaddition of nitrones to α,ß-unsaturated carboxylic acid derivatives has been investigated. In the presence of a catalytic amount of [Pd(NCMe)2{(S)-tolbinap}](BF4)2 [TolBINAP = 2,2'-bis(di-p-tolylphosphino)-1,1'-binaphthyl], the reaction of 3-alkenoyl-1,3-oxazolidin-2-ones as dipolarophiles and N-substituted N-benzylidenenitrones has been successfully performed to give isoxazolidine derivatives in high yields with high enantioselectivities. For example, 3-((2,5-dimethyl-3-phenylisoxazolidin-4-yl)carbonyl)-1,3-oxazolidin-2-one was obtained from the reaction of N-benzylidenemethylamine N-oxide and 3-crotonoyl-1,3-oxazolidin-2-one in 89% yield with 60% endo selectivity and 91% ee of the endo isomer. The cycloaddition of N-benzylidenebenzylamine N-oxide and 3-crotonoyl-1,3-oxazolidin-2-one afforded 3-((2-benzyl-5-methyl-3-phenylisoxazolidin-4-yl)carbonyl)-1,3-oxazolidin-2-one in 94% yield with 93% endo selectivity and 89% ee of the endo isomer. Remarkably, the endo/exo selectivity of the products depended on the N-substituent group of the nitrones. These selectivities were explained using molecular modeling.