Kurarinone induced p53-independent G0/G1 cell cycle arrest by degradation of K-RAS via WDR76 in human colorectal cancer cells.

Kurarinone (KR), a naturally occurring flavonoid in Sophora flavescens Aiton and a traditional herbal medicine, reportedly has anti-cancer activity against various cancer types both in vitro and in vivo. However, the cellular mechanism of KR remains unknown. Therefore, we aimed to elucidate the mechanism of cell cycle arrest induced by KR in human colorectal cancer cells. KR not only reduced cell proliferation but also induced G0/G1 arrest of colorectal cancer cell lines. The results of western blotting analysis showed that KR reduced the protein levels of cyclin D1/D3 and CDK4/6 by downregulating signaling proteins such as K-RAS, c-MYC, and p-extracellular signal-regulated kinase. Additionally, KR arrested the cell cycle in the G0/G1 phase in a p53-independent manner, and decreased the protein level of K-RAS by proteasomal degradation dependent on WDR76, an E3 ubiquitin ligase. From these results, we propose that KR could be a potent anti-cancer agent, acting through the degradation of K-RAS dependent on WDR76, regardless of the p53 status.

Production of Bioactive 3'-Hydroxystilbene Compounds Using the Flavin-Dependent Monooxygenase Sam5.

The flavin-dependent monooxygenase Sam5 was previously reported to be a bifunctional hydroxylase with a coumarte 3-hydroxylase and a resveratrol 3'-hydroxylase activity. In this article, we showed the Sam5 enzyme has 3'-hydroxylation activities for methylated resveratrol (pinostilbene and pterostilbene), hydroxylated resveratrol (oxyresveratrol) and glycosylated resveratrol (piceid) as substrates. However, the use of piceid, a glycone type stilbene, as a substrate for bioconversion experiments with the Sam5 enzyme expressed in, Escherichia coli does not convert to the hydroxylated compound astringin, but it has converted in vitro enzyme reactions. Finally, we report a novel catalytic activity of Sam5 monooxygenase for the synthesis of piceatannol derivatives, 3'-hydroxylated stilbene compounds. Development of this bioproduction method for the hydroxylation of stilbenes is challenging because of the difficulty in expressing P450-type hydroxylase in E. coli and regionspecific chemical synthesis.

Four new neolignans isolated from Eleutherococcus senticosus and their protein tyrosine phosphatase 1B inhibitory activity (PTP1B).

Four new compounds, erythro-7'E-4-hydroxy-3,3'-dimethoxy-8,5'-oxyneoligna-7'-ene-7,9-diol-9'-al (1), (7S,8S)-4-hydroxy-3,1',3'-trimethoxy-4',7-epoxy-8,5'-neolign-9-ol (5), (7S,8S,7'E)-5-hydroxy-3,3'-dimethoxy-4',7-epoxy-8,5'-neolign-7'-ene-9,9'-diol (6) and (7S,8S,7'E)-5-hydroxy-3,3',9'-trimethoxy-4'-7-epoxy-8,5'-neolign-7'-ene-9-ol (7). Along with four known compounds (2-4, 8) were isolated from the EtOAc-soluble extract of Eleutherococcus senticosus. Their structures were elucidated on the basis of spectroscopic and physicochemical analyses. All the compounds were evaluated for in vitro inhibitory activity against PTP1B, VHR and PP1. Among them, compounds 1-4 and 6-8 were found to exhibit selective inhibitory activity on PTP1B with IC50 values ranging from 17.2±1.6 to 32.7±1.2µM.

New neo-lignan from Acanthopanax senticosus with protein tyrosine phosphatase 1B inhibitory activity.

New neo-lignan, (7S, 8R)-3-hydroxyl-4-methoxyl-balanophonin (1), together with seven known compounds (2-8) were isolated from the EtOAc-soluble extract of Acanthopanax senticosus. The structure of the new neo-lignan was elucidated with spectroscopic and physico-chemical analyses. All the isolates were evaluated for in vitro inhibitory activity against PTP1B, VHR and PP1. Among them, the new compound (1) was found to exhibit selective inhibitory activity on PTP1B with IC50 value 15.2 ± 1.4 µM.

Structures and biological activities of azaphilones produced by Penicillium sp. KCB11A109 from a ginseng field.

Twelve metabolites, including five highly oxygenated azaphilones, geumsanols A-E, along with seven known analogues were isolated from Penicillium sp. KCB11A109, a fungus derived from a ginseng field. Their structures were assigned by spectroscopic means (NMR and MS), and stereochemistries were determined by extensive spectroscopic analyses ((1)H-(1)H coupling constants, NOESY, and HETLOC) and chemical derivatizations (modified Mosher's method and acetonide formation). The isolates were evaluated for their anticancer, antimicrobial, antimalarial activities, and phenotypic effects in zebrafish development. Of these compounds possessing no pyranoquinone core, only geumsanol E exhibited cytotoxic activities and toxic effects on zebrafish embryos, suggesting that a double bond at C-11 and C-12 is important for biological activity.

Eupafolin suppresses prostate cancer by targeting phosphatidylinositol 3-kinase-mediated Akt signaling.

Phosphatase and tensin homolog (PTEN) loss or mutation consistently activates the phosphatidylinositol 3-kinase (PI3-K)/Akt signaling pathway, which contributes to the progression and invasiveness of prostate cancer. Furthermore, the PTEN/PI3-K/Akt and Ras/MAPK pathways cooperate to promote the epithelial-mesenchymal transition (EMT) and metastasis initiated from prostate stem/progenitor cells. For these reasons, the PTEN/PI3-K/Akt pathway is considered as an attractive target for both chemoprevention and chemotherapy. Herein we report that eupafolin, a natural compound found in common sage, inhibited proliferation of prostate cancer cells. Protein content analysis indicated that phosphorylation of Akt and its downstream kinases was inhibited by eupafolin treatment. Pull-down assay and in vitro kinase assay results indicated that eupafolin could bind with PI3-K and attenuate its kinase activity. Eupafolin also exhibited tumor suppressive effects in vivo in an athymic nude mouse model. Overall, these results suggested that eupafolin exerts antitumor effects by targeting PI3-K.

Hypo-pigmenting effect of sesquiterpenes from Inula britannica in B16 melanoma cells.

During the course of screens to identify anti-melanogenic agents from natural resources, we found that the methanol extract of the dried flower of Inula britannica L. inhibited melanin synthesis in cultured melanoma cells stimulated with 3-isobutyl-1-methylxanthine (IBMX). A bioassay-guided isolation of the chloroform fraction of the I. britannica using an in vitro melanogenesis inhibition assay led to the isolation of sesquiterpenes, 1-O-acetylbritannilactone (1), britannilactone (2) and neobritannilactone B (3). Compounds 1 and 2 significantly reduced melanin production in a dose-dependent manner with IC50 values of 13.3 and 15.5 µM, respectively, whereas compound 3 was found to be cytotoxic. Compound 1 also inhibited the tyrosinase activity only in cell based-systems. Western blot analysis indicated that compound 1 inhibited melanogenesis by activating extracellular signal-regulated kinase (ERK) and Akt signaling and also inhibiting cAMP related binding protein, which regulates its downstream pathway, including tyrosinase, tyrosinase related protein-1 and TRP-2. These results demonstrated that compound 1, a major sesquiterpene from the flowers of I. britannica, exhibited anti-melanogenic activity by suppression of tyrosinase expression via ERK and Akt signaling. Taken together, our results suggest that these compounds may act as potent natural skin-lightening agents.

Protein tyrosine phosphatase 1B inhibitory activity of 24-norursane triterpenes isolated from Weigela subsessilis.

During the screening effort to discover new types of protein tyrosine phosphatase 1B (PTP1B) inhibitors, it was found that a MeOH extract of the leaves and stems of Weigela subsessilis (Caprifoliaceae) inhibited the enzyme activity. By means of an in vitro bioassay-guided fractionation on the MeOH extract, two 24-norursane triterpenes, ilekudinol A (1) and ilekudinol B (2), were isolated as active metabolites. Compounds 1 and 2 inhibited PTP1B with IC(50) values of 29.1 ± 2.8 and 5.3 ± 0.5 µM, respectively. Kinetic studies suggest that both 1 and 2 are non-competitive inhibitors of PTP1B. The findings indicate that the free carboxyl group at C-28 in this type of triterpenes plays a critical role in the inhibition of PTP1B.

Evaluation of human neutrophil elastase inhibitory effect of iridoid glycosides from Hedyotis diffusa.

Five iridoid glycosides were isolated from the MeOH extract of Hedyotis diffusa, and their structures were elucidated as E-6-O-p-methoxycinnamoyl scandoside methyl ester (1), Z-6-O-p-methoxycinnamoyl scandoside methyl ester (2), E-6-O-p-feruloyl scandoside methyl ester (3), E-6-O-p-coumaroyl scandoside methyl ester (4), and Z-6-O-p-coumaroyl scandoside methyl ester (5) by interpretation of their spectroscopic data. All the isolated compounds were evaluated for human neutrophil elastase inhibitory effect, and compound 1 showed potent activity with an IC(50) value of 18.0muM. The molecular docking simulation suggested a structural model for the inhibition of human neutrophil elastase by compound 1.

Protein tyrosine phosphatase 1B inhibitors isolated from Morus bombycis.

Bioassay-guided fractionation of the chloroform-soluble fraction of Morus bombycis, using an in vitro PTP1B inhibitory assay led to the identification of three 2-arylbenzofurans, albafuran A (1), mulberrofuran W (2) and mulberrofuran D (6), along with three chalcone-derived Diels-Alder products, kuwanon J (3), kuwanon R (4), and kuwanon V (5). Compounds 1-6 showed remarkable inhibitory activity against PTP1B with IC(50) values ranging from 2.7 to 13.8 microM. Inhibition kinetics were analyzed by Lineweaver-Burk plots, which suggested that compounds 1-6 inhibited PTP1B in a mixed-type manner. The present results indicate that the respective lipophilic and hydroxyl groups of 2-arylbenzofurans and chalcone-derived Diels-Alder products play an important role in inhibition of PTP1B.

PTP1B inhibitory effects of tridepside and related metabolites isolated from the Antarctic lichen Umbilicaria antarctica.

The selective inhibition of PTP1B has been widely recognized as a potential drug target for the treatment of type 2 diabetes and obesity. In the course of screening for PTP1B inhibitory natural products, the MeOH extract of the dried sample of the Antarctic lichen Umbilicaria antarctica was found to exhibit significant inhibitory effect, and the bioassay-guided fractionation and purification afforded three related lichen metabolites 1-3. Compounds 1-3 were identified as gyrophoric acid (1), lecanoric acid (2), and methyl orsellinate (3) mainly by analysis of NMR and MS data. These compounds inhibited PTP1B activity with 50% inhibitory concentration values of 3.6 +/- 0.04 microM, 31 +/- 2.7 microM, and 277 +/- 8.6 microM, respectively. Furthermore, the kinetic analysis of PTP1B inhibition by compound 1 suggested that the compound inhibited PTP1B activity in a non-competitive manner.

Inhibition of protein tyrosine phosphatase 1B by lupeol and lupenone isolated from Sorbus commixta.

Protein tyrosine phosphatase 1B (PTP1B) appears to be an attractive target for the development of new drugs for type 2 diabetes and obesity. In our preliminary test, a MeOH extract of the stem barks of Sorbus commixta Hedl. (Rosaceae) showed strong PTP1B inhibitory activity. Bioassay-guided fractionation of the MeOH extract resulted in the isolation of two lupane-type triterpenes, lupenone (1) and lupeol (2). Compounds 1 and 2 inhibited PTP1B with IC(50) values of 13.7 +/- 2.1 and 5.6 +/- 0.9 microM, respectively. Kinetic studies revealed that both the compounds 1 and 2 are non-competitive inhibitors of PTP1B that decrease V(max) values with no effect on K(m) values.

Protein tyrosine phosphatase 1B inhibitory effects of depsidone and pseudodepsidone metabolites from the Antarctic lichen Stereocaulon alpinum.

Seven phenolic lichen metabolites (1-7) have been isolated from a methanol extract of the Antarctic lichen Stereocaulon alpinum by various chromatographic methods. The structures of these compounds were determined mainly by analysis of NMR spectroscopic data. A depsidone-type compound, lobaric acid (1) and two pseudodepsidone-type compounds, 2 and 3, exhibited potent inhibitory activity against protein tyrosine phosphatase 1B (PTP1B) with IC(50) values of 0.87microM, 6.86microM, and 2.48microM, respectively. Kinetic analyses of PTP1B inhibition by compounds 1 and 2 suggested that these compounds inhibited PTP1B activity in a non-competitive manner.

Protein tyrosine phosphatase 1B inhibitory by dammaranes from Vietnamese Giao-Co-Lam tea.

ETHNOPHARMACOLOGICAL RELEVANCE: Gynostemma pentaphyllum (Thunb.) tea was used in Vietnamese folk medicine as anti-diabetic agent. AIM OF THE STUDY: This study was aimed to investigate the inhibitory activities of fractions and constituents isolated from Gynostemma pentaphyllum on protein tyrosine phosphatase 1B (PTP1B) since it has been proposed as a treatment therapy for type 2 diabetes and obesity. MATERIALS AND METHODS: The 70% EtOH extract, CHCl3 fraction, EtOAc fraction, BuOH fraction, and seven isolated dammarane triterpenes were evaluated for their inhibitory activity in protein phosphatase enzymes (PTP1B and VHR). RESULTS: CHCl3-soluble fraction showed a dose-dependent inhibitory activity of the PTP1B enzyme with the IC50 value of 30.5 microg/mL. Among seven tested compounds, compounds 6 showed the most potent PTP1B inhibitory activity with IC50 value of 5.3+/-0.4 microM compared to a range 15.7-28.5 microM for the other six compounds. The inhibition mode of 6 was competitive toward p-NPP with a K(i) value of 2.8 microM. CONCLUSION: These study results suggested that the PTP1B inhibitory activity of these dammaranes may enable this plant to play an important role in the treatment of diabetes.

Genistein-derivatives from Tetracera scandens stimulate glucose-uptake in L6 myotubes.

An EtOAc-soluble partition of the MeOH extract of a branch of Tetracera scandens (Dilleniaceae family) was subjected to a glucose-uptake assay, which led to the isolation and identification of five isoflavones of previously known structure namely, genistein (1), its derivatives 3',5'-diprenylgenistein (2), 6,8-diprenylgenistein (3), derrone (4) and alpinumisoflavone (5). Of these, compounds 2--5 exhibited significant glucose-uptake activity in basal and insulin-stimulated L6 myotubes. The findings from adenosine monophosphate-activated kinase (AMPK) activation and glucose transport protein4 (GLUT4) and GLUT1 over-expression revealed certain characteristics of compounds 2--5. These compounds inhibited protein tyrosine phosphatase 1B (PTP1B) activities with IC50 values ranging from 20.63 +/- 0.17 to 37.52 +/- 0.31 microM. No muscle cell toxicity was reported with compounds 3--5, while compounds 1 and 2 reduced muscle cell viability with IC50 values of 34.27 +/- 0.35 and 18.69 +/- 0.19 microM, respectively. It was concluded that T. scandens and its constituents exerted highly desirable activities on type 2 diabetes mellitus treatment since they significantly stimulated the uptake of glucose, AMPK phosphorylation, GLUT4 and GLUT1 mRNA expressions and PTP1B inhibition in L6 myotubes.

Prenylisoflavonoids from Erythrina senegalensis as novel HIV-1 protease inhibitors.

Eight compounds were isolated from the CH (2)Cl (2) extracts of ERYTHRINA SENEGALENSIS to assess HIV-1 protease (PR) activity inhibition. The prenylated isoflavone structures, identified by spectroscopic analysis, were 8-prenylluteone ( 1), auriculatin ( 2), erysenegalensein O ( 3), erysenegalensein D ( 4), erysenegalensein N ( 5), derrone ( 6), alpinumisoflavone ( 7), and 6,8-diprenylgenistein ( 8). The constituents showed dose-dependent inhibitory activities on HIV-1 PR with IC (50) values from 0.5 to 30.0 muM. Compounds 1 - 5 possessing two hydroxy groups in the 2' and 4' positions of the B ring, potently inhibited HIV-1 PR activity. In addition, 6,8-diprenylgenistein ( 8) with two prenyl groups in the 6 and 8 positions of the A ring and one hydroxy group in the 4' position of B-ring was the most potent HIV-1 PR inhibitor.

Isoprenylated flavonoids from the stem bark of Erythrina abyssinica.

Three new prenylated flavanones, abyssinoflavanones V, VI, and VII (1-3), together with eight known flavanones (4-11) and two chalcones (12-13), were isolated from the stem bark of Erythrina abyssinica. Their structures were elucidated on the basis of spectroscopic and physicochemical analyses. All the isolates, with the exception of 1 and 8, strongly inhibited PTP1B activity in an in vitro assay with IC50 values ranging from 14.2 +/- 1.7 to 26.7 +/- 1.2 microM.

Protein tyrosine phosphatase-1B inhibitory activity of isoprenylated flavonoids isolated from Erythrina mildbraedii.

Inhibition of protein tyrosine phosphatase-1B (PTP1B) has been proposed as a therapy for treatment of type-2 diabetes and obesity. Bioassay-guided fractionation of an EtOAc-soluble extract of the root bark of Erythrina mildbraedii, using an in vitro PTP1B inhibitory assay, resulted in the isolation of three new isoprenylated flavonoids, abyssinone-IV-4'-O-methyl ether (2), 7-hydroxy-4'-methoxy-3'-(3-hydroxy-3-methyl-trans-but-1-enyl)-5'-(3-methylbut-2-enyl)flavanone (3), and abyssinone-VI-4-O-methyl ether (6), along with six known flavonoids, abyssinone-V-4'-O-methyl ether (1), abyssinone-V (4), abyssinone-IV (5), sigmoidin E (7), 4'-hydroxy-5,7-dimethoxyisoflavone (8), and alpinumisoflavone (9). Compounds 1 and 2, 4-7, and 9 inhibited PTP1B activity, with IC50 values ranging from 14.8 +/- 1.1 to 39.7 +/- 2.5 microM. On the basis of the data obtained, flavanones and chalcones with isoprenyl groups may be considered as a new class of PTP1B inhibitors.

PTP1B inhibitory activity of kaurane diterpenes isolated from Siegesbeckia glabrescens.

Protein tyrosine phosphatase 1B (PTP1B) is considered as a therapeutic target for the treatment of diabetes and obesity. In our preliminary screening study, a MeOH extract of the aerial part of Siegesbeckia glabrescens was found to inhibit PTP1B activity at 30 microg/mL. Bioassay-guided fractionation led to the isolation of two active diterpenes, ent-16betaH, 17-isobutyryloxy-kauran-19-oic acid (1) and ent-16betaH, 17-acetoxy-18-isobutyryloxy-kauran-19-oic acid (2), along with ent- 16betaH, 17-hydroxykauran-19-oic acid (3). Compounds 1 and 2 inhibited the PTP1B activity with IC50 values of 8.7 +/- 0.9 and 30.6 +/- 2.1 microM, respectively. Kinetic studies suggest that both 1 and 2 are non-competitive inhibitors of PTP1B. However, compound 3 substituted with a hydroxyl group at C-17 in kaurane-type showed no inhibitory effects towards PTP1B.

Inhibition of protein tyrosine phosphatase 1B by prenylated isoflavonoids isolated from the stem bark of Erythrina addisoniae.

It has been suggested that protein tyrosine phosphatase 1B (PTP1B) inhibitors might be a therapeutic target for the treatment of type 2 diabetes and obesity. A bioassay-guided phytochemical study of the EtOAc extract of the stem bark of Erythrina addisoniae (Leguminosae) resulted in the identification of a new PTP1B inhibitory compound, 5,2',4'-trihydroxy-6-(gamma,gamma-dimethylallyl)-2''',2'''-dimethyldihydropyrano[5''',6''']isoflavanone ( 6), along with five known prenylated isoflavonoids, orientanol E ( 1), senegalensin ( 2), warangalone ( 3), warangalone 4'-methyl ether ( 4) and 2,3-dihydroauriculatin ( 5). Compounds 1, 5 and 6 inhibited PTP1B with IC (50) values ranging from 2.6 +/- 0.5 to 10.1 +/- 0.3 microM. Our results indicate that hydroxylation at both 2'- and 4'-positions in the B-ring and cyclization between a hydroxy group at C-7 and one of the prenyl groups at C-6 or C-8 in the A-ring may be important for activity. Thus, compounds 5 and 6 could be a new class of natural PTP1B inhibitors.