Antitumor activity of a novel oral signal transducer and activator of transcription 3 inhibitor YHO-1701.

The signal transducer and activator of transcription 3 (STAT3) signaling pathway is a key mediator of cancer cell proliferation, survival and invasion. Aberrant STAT3 has been demonstrated in various malignant cancers. YHO-1701 is a novel quinolinecarboxamide derivative generated from STX-0119. Here, we examined the effect of YHO-1701 on STAT3 and evaluated antitumor activity of YHO-1701 as a single agent and in combination. YHO-1701 inhibited STAT3-SH2 binding to phospho-Tyr peptide selectively and more potently than STX-0119 in biochemical assays. Molecular docking studies with STAT3 suggested more stable interaction of YHO-1701 with the SH2 domain. YHO-1701 exhibited approximately 10-fold stronger activity than STX-0119 in abrogating the STAT3 signaling pathway of human oral cancer cell line SAS. YHO-1701 also blocked multi-step events by inhibiting STAT3 dimerization and suppressed STAT3 promoter activity. As expected, YHO-1701 exerted strong antiproliferative activity against human cancer cell lines addicted to STAT3 signaling. Orally administered YHO-1701 showed statistically significant antitumor effects with long exposure to high levels of YHO-1701 at tumor sites in SAS xenograft models. Moreover, combination regimen with sorafenib led to significantly stronger antitumor activity. In addition, the suppression level of survivin (a downstream target) was superior for the combination as compared with monotherapy groups within tumor tissues. Thus, YHO-1701 had a favorable specificity for STAT3 and pharmacokinetics after oral treatment; it also contributed to the enhanced antitumor activity of sorafenib. The evidence presented here provides justification using for this approach in future clinical settings.

Combretastatin A4 disodium phosphate-induced myocardial injury.

Histopathological and electrocardiographic features of myocardial lesions induced by combretastatin A4 disodium phosphate (CA4DP) were evaluated, and the relation between myocardial lesions and vascular changes and the direct toxic effect of CA4DP on cardiomyocytes were discussed. We induced myocardial lesions by administration of CA4DP to rats and evaluated myocardial damage by histopathologic examination and electrocardiography. We evaluated blood pressure (BP) of CA4DP-treated rats and effects of CA4DP on cellular impedance-based contractility of human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs). The results revealed multifocal myocardial necrosis with a predilection for the interventricular septum and subendocardial regions of the apex of the left ventricular wall, injury of capillaries, morphological change of the ST junction, and QT interval prolongation. The histopathological profile of myocardial lesions suggested that CA4DP induced a lack of myocardial blood flow. CA4DP increased the diastolic BP and showed direct effects on hiPS-CMs. These results suggest that CA4DP induces dysfunction of small arteries and capillaries and has direct toxicity in cardiomyocytes. Therefore, it is thought that CA4DP induced capillary and myocardial injury due to collapse of the microcirculation in the myocardium. Moreover, the direct toxic effect of CA4DP on cardiomyocytes induced myocardial lesions in a coordinated manner.

AutoGPA: An Automated 3D-QSAR Method Based on Pharmacophore Alignment and Grid Potential Analysis.

3D-QSAR approach has been widely applied and proven to be useful in the case where no reliable crystal structure of the complex between a biologically active molecule and the receptor is available. At the same time, however, it also has highlighted the sensitivity of this approach. The main requirement of the traditional 3D-QSAR method is that molecules should be correctly overlaid in what is assumed to be the bioactive conformation. Identifying an active conformation of a flexible molecule is technically difficult. It has been a bottleneck in the application of the 3D-QSAR method. We have developed a 3D-QSAR software named AutoGPA especially based on an automatic pharmacophore alignment method in order to overcome this problem which has discouraged general medicinal chemists from applying the 3D-QSAR methods to their "real-world" problems. Applications of AutoGPA to three inhibitor-receptor systems have demonstrated that without any prior information about the three-dimensional structure of the bioactive conformations AutoGPA can automatically generate reliable 3D-QSAR models. In this paper, the concept of AutoGPA and the application results will be described.

Novel acrylonitrile derivatives, YHO-13177 and YHO-13351, reverse BCRP/ABCG2-mediated drug resistance in vitro and in vivo.

Breast cancer resistance protein (BCRP/ABCG2) confers resistance to anticancer drugs such as 7-ethyl-10-hydroxycamptothecin (SN-38, an active metabolite of irinotecan), mitoxantrone, and topotecan. In this study, we examined the reversing effects of YHO-13177, a novel acrylonitrile derivative, and its water-soluble diethylaminoacetate prodrug YHO-13351 on the BCRP-mediated drug resistance. YHO-13177 potentiated the cytotoxicity of SN-38, mitoxantrone, and topotecan in both BCRP-transduced human colon cancer HCT116 (HCT116/BCRP) cells and SN-38-resistant human lung cancer A549 (A549/SN4) cells that express BCRP, but had little effect in the parental cells. In addition, YHO-13177 potentiated the cytotoxicity of SN-38 in human lung cancer NCI-H460 and NCI-H23, myeloma RPMI-8226, and pancreatic cancer AsPC-1 cells that intrinsically expressed BCRP. In contrast, it had no effect on P-glycoprotein-mediated paclitaxel resistance in MDR1-transduced human leukemia K562 cells and multidrug resistance-related protein 1-mediated doxorubicin resistance in MRP1-transfected human epidermoid cancer KB-3-1 cells. YHO-13177 increased the intracellular accumulation of Hoechst 33342, a substrate of BCRP, at 30 minutes and partially suppressed the expression of BCRP protein at more than 24 hours after its treatment in both HCT116/BCRP and A549/SN4 cells. In mice, YHO-13351 was rapidly converted into YHO-13177 after its oral or intravenous administration. Coadministration of irinotecan with YHO-13351 significantly increased the survival time of mice inoculated with BCRP-transduced murine leukemia P388 cells and suppressed the tumor growth in an HCT116/BCRP xenograft model, whereas irinotecan alone had little effect in these tumor models. These findings suggest that YHO-13351, a prodrug of YHO-13177, could be clinically useful for reversing BCRP-mediated drug resistance in cancer chemotherapy.

Streptomycin alleviates irinotecan-induced delayed-onset diarrhea in rats by a mechanism other than inhibition of ß-glucuronidase activity in intestinal lumen.

Irinotecan hydrochloride (CPT-11) is a useful drug for cancer chemotherapy but sometimes induces severe diarrhea clinically. CPT-11 is mainly activated to SN-38 by carboxylesterase (CES) and then detoxified to SN-38 glucuronide (SN-38G) by UDP-glucuronosyltransferase (UGT) in the liver. SN-38G is excreted via bile and de-conjugated to SN-38 by ß-glucuronidase (ß-GLU) in the intestinal content. In order to clarify the alleviative effect of antibiotics on CPT-11-induced diarrhea, we examined whether penicillin G and streptomycin (SM) alleviate CPT-11-induced delayed-onset diarrhea using three diarrheal models, i.e., Wistar rats with repeated dosing of CPT-11 (60 mg/kg/day i.v. for 4 consecutive days) and Wistar and Gunn rats with a single dosing of CPT-11 (200 and 20 mg/kg i.v., respectively). Gunn rats have an inherited deficiency of UGT1A and cannot conjugate SN-38 to SN-38G. Therefore, onset of CPT-11-induced diarrhea in Gunn rats is not affected by ß-GLU activity. SM alleviated diarrhea in all three diarrheal models. The alleviation of diarrhea by SM in Gunn rats indicated that the effect of SM occurred by a mechanism other than the inhibition of ß-GLU activity. SM decreased CPT-11 and/or SN-38 concentrations in intestinal tissues and alleviated epithelial damage from the ileum to colon. SM did not inhibit ß-GLU activity in the cecal content. SM also inhibited the intestinal absorption of CPT-11 and decreased CES activity and increased UGT activity in the intestinal epithelium. These findings indicated that SM decreased the exposure of CPT-11 and SN-38 to the intestinal epithelium by inhibiting the absorption of CPT-11 from the intestinal lumen and the change of CES and UGT activities in the intestinal epithelium and alleviated delayed-onset diarrhea.

Characteristic amino acid distribution around segments unique to allergens.

Epitopes are located at the surface of allergens with which antibodies specifically bind. On the assumption that fragments unique to allergens have common, characteristic amino acid sequences, we compared the amino acid sequences of allergens with those of non-allergens. Segments around fragments unique to allergens showed wavelet-like distributions for several amino acids. Charged residues, alanine and glycine had positive peaks at the centre of the unique segments with small valleys on both sides, while aromatic residues, proline and cysteine showed the inverse distribution. Furthermore, the wavelet-like distribution of amino acids could be represented by a universal distribution function together with an index characterizing the intensity of the wavelet. Using the universal distribution function and the novel index of amino acids, we developed a simple method for extracting segments and fragments that are unique to allergens. The significance of the universal distribution function and the novel index is also discussed, by comparing the plot of the allergen-unique fragments index and dynamic fluctuation in the three dimensional structure of birch pollen allergen as both a single molecule and a complex with the corresponding antibody.

SOSUI-GramN: high performance prediction for sub-cellular localization of proteins in gram-negative bacteria.

A predictive software system, SOSUI-GramN, was developed for assessing the subcellular localization of proteins in Gram-negative bacteria. The system does not require the sequence homology data of any known sequences; instead, it uses only physicochemical parameters of the N- and C-terminal signal sequences, and the total sequence. The precision of the prediction system for subcellular localization to extracellular, outer membrane, periplasm, inner membrane and cytoplasmic medium was 92.3%, 89.4%, 86.4%, 97.5% and 93.5%, respectively, with corresponding recall rates of 70.3%, 87.5%, 76.0%, 97.5% and 88.4%, respectively. The overall performance for precision and recall obtained using this method was 92.9% and 86.7%, respectively. The comparison of performance of SOSUI-GramN with that of other methods showed the performance of prediction for extracellular proteins, as well as inner and outer membrane proteins, was either superior or equivalent to that obtained with other systems. SOSUI-GramN particularly improved the accuracy for predictions of extracellular proteins which is an area of weakness common to the other methods.

Potent, exceptionally selective, orally bioavailable inhibitors of TNF-alpha Converting Enzyme (TACE): novel 2-substituted-1H-benzo[d]imidazol-1-yl)methyl)benzamide P1' substituents.

Novel ((2-substituted-1H-benzo[d]imidazol-1-yl)methyl)benzamides were found to be excellent P1' substituents in conjunction with unique constrained beta-amino hydroxamic acid scaffolds for the discovery of potent selective inhibitors of TNF-alpha Converting Enzyme (TACE). Optimized examples proved potent for TACE, exceptionally selective over a wide panel of MMP and ADAM proteases, potent in the suppression of LPS-induced TNF-alpha in human whole blood and orally bioavailable.

Alpha,Beta-cyclic-beta-benzamido hydroxamic acids: Novel oxaspiro[4.4]nonane templates for the discovery of potent, selective, orally bioavailable inhibitors of tumor necrosis factor-alpha converting enzyme (TACE).

Two novel oxaspiro[4.4]nonane beta-benzamido hydroxamic scaffolds have been synthesized in enantio- and diasteriomerically pure form. These templates proved to be exceptional platforms that have led to the discovery of potent inhibitors of TACE that are active in a cellular assay measuring suppression of LPS-induced TNF-alpha. Furthermore, these inhibitors are selective against related MMPs, demonstrate permeability in a Caco-2 assay, and display good oral bioavailability.

Alpha,beta-cyclic-beta-benzamido hydroxamic acids: novel templates for the design, synthesis, and evaluation of selective inhibitors of TNF-alpha converting enzyme (TACE).

Selective inhibitors of TNF-alpha Converting Enzyme (TACE) based on (1R,2S)-cyclopentyl, (3S,4S)-pyrrolidinyl, and (3R,4S)-tetrahydrofuranyl beta-benzamido hydroxamic acids have been synthesized and evaluated. This study has led to the discovery of novel inhibitors whose profiles include activity against TACE in an enzyme assay, potency in the suppression of LPS-stimulated TNF-alpha in human whole blood, selectivity against a panel of MMPs and oral bioavailability.

Synthesis and structure-activity relationship of a novel, non-hydroxamate series of TNF-alpha converting enzyme inhibitors.

A novel series of TNF-alpha converting enzyme (TACE) inhibitors which are non-hydroxamate have been discovered. These compounds use a triazolethione moiety as the zinc binding ligand and exhibit IC50 values from 1.5 to 100 nM in a porcine TACE assay. They also have excellent selectivities over other MMPs.

Discovery of low nanomolar non-hydroxamate inhibitors of tumor necrosis factor-alpha converting enzyme (TACE).

Using a pyrimidine-2,4,6-trione motif as a zinc-binding group, a series of selective inhibitors of tumor necrosis factor-alpha converting enzyme (TACE) was discovered. Optimization of initial lead 1 resulted in a potent inhibitor (51), with an IC(50) of 2 nM in a porcine TACE assay. To the best of our knowledge, compound 51 and related analogues represent first examples of non-hydroxamate-based inhibitors of TACE with single digit nanomolar potency.

Intramolecular palladium-catalyzed allylic alkylation: enantio- and diastereoselective synthesis of [2.2.2] bicycles.

Pd-catalyzed asymmetric allylic alkylation provides both enantio- and diastereoselectivity in formation of bicyclo [2.2.2] octan-2,3-diones and quinuclidin-2-ones, the latter potential precursors to quinine alkaloids. [reaction: see text]