Physodic acid sensitizes LNCaP prostate cancer cells to TRAIL-induced apoptosis.

In spite of the extensive research for developing new therapies, prostate cancer is still one of the major human diseases with poor prognosis and high mortality. Therefore, with the aim of identifying novel agents with antigrowth and pro-apoptotic activity on prostate cancer cells, in the present study, we evaluated the effect of lichen secondary metabolite physodic acid on cell growth in human prostate cancer cells. In addition, we tested the apoptotic activity of physodic acid on TRAIL-resistant LNCaP cells in combination with TRAIL. The cell viability was measured using MTT assay. LDH release, a marker of membrane breakdown, was also measured. For the detection of apoptosis, the evaluation of DNA fragmentation and caspase-3 activity assay were employed. The expression of proteins was detected by Western blot analysis. It was observed that physodic acid showed a dose-response relationship in the range of 12.5-50 µM concentrations in LNCaP and DU-145 cells, activating an apoptotic process. In addition, physodic acid sensitizes LNCaP cells to TRAIL-induced apoptosis. The combination of physodic acid with other anti-prostate cancer therapies could be considered a promising strategy that warrants further investigations.

Potential Effect of Pseudevernia furfuracea (L.) Zopf Extract and Metabolite Physodic Acid on Tumour Microenvironment Modulation in MCF-10A Cells.

Lichens comprise a number of unique secondary metabolites with remarkable biological activities and have become an interesting research topic for cancer therapy. However, only a few of these metabolites have been assessed for their effectiveness against various in vitro models. Therefore, the aim of the present study was to assess the effect of extract Pseudevernia furfuracea (L.) Zopf (PSE) and its metabolite physodic acid (Phy) on tumour microenvironment (TME) modulation, focusing on epithelial-mesenchymal transition (EMT), cancer-associated fibroblasts (CAFs) transformation and angiogenesis. Here, we demonstrate, by using flow cytometry, Western blot and immunofluorescence microscopy, that tested compounds inhibited the EMT process in MCF-10A breast cells through decreasing the level of different mesenchymal markers in a time- and dose-dependent manner. By the same mechanisms, PSE and Phy suppressed the function of Transforming growth factor beta (TGF-ß)-stimulated fibroblasts. Moreover, PSE and Phy resulted in a decreasing level of the TGF-ß canonical pathway Smad2/3, which is essential for tumour growth. Furthermore, PSE and Phy inhibited angiogenesis ex ovo in a quail embryo chorioallantoic model, which indicates their potential anti-angiogenic activity. These results also provided the first evidence of the modulation of TME by these substances.

Synthesis and Target Identification of Benzoxepane Derivatives as Potential Anti-Neuroinflammatory Agents for Ischemic Stroke.

Benzoxepane derivatives were designed and synthesized, and one hit compound emerged as being effective in vitro with low toxicity. In vivo, this hit compound ameliorated both sickness behavior through anti-inflammation in LPS-induced neuroinflammatory mice model and cerebral ischemic injury through anti-neuroinflammation in rats subjected to transient middle cerebral artery occlusion. Target fishing for the hit compound using photoaffinity probes led to identification of PKM2 as the target protein responsible for anti-inflammatory effect of the hit compound. Furthermore, the hit exhibited an anti-neuroinflammatory effect in vitro and in vivo by inhibiting PKM2-mediated glycolysis and NLRP3 activation, indicating PKM2 as a novel target for neuroinflammation and its related brain disorders. This hit compound has a better safety profile compared to shikonin, a reported PKM2 inhibitor, identifying it as a lead compound in targeting PKM2 for the treatment of inflammation-related diseases.

Expanding antibiotic chemical space around the nidulin pharmacophore.

Reinvestigating antibiotic scaffolds that were identified during the Golden Age of antibiotic discovery, but have long since been "forgotten", has proven to be an effective strategy for delivering next-generation antibiotics capable of combatting multidrug-resistant superbugs. In this study, we have revisited the trichloro-substituted depsidone, nidulin, as a selective and unexploited antibiotic lead produced by the fungus Aspergillus unguis. Manipulation of halide ion concentration proved to be a powerful tool for modulating secondary metabolite production and triggering quiescent pathways in A. unguis. Supplementation of the culture media with chloride resulted in a shift in co-metabolite profile to dichlorounguinols and nornidulin at the expense of the non-chlorinated parent, unguinol. Surprisingly, only marginal enhancement of nidulin was observed, suggesting O-methylation may be rate-limiting. Similarly, supplementation of the media with bromide led to the production of the corresponding bromo-analogues, but also resulted in a novel family of depsides, the unguidepsides. Unexpectedly, depletion of chloride from the media halted the biosynthesis of the non-chlorinated parent compound, unguinol, and redirected biosynthesis to a novel family of ring-opened analogues, the unguinolic acids. Supplementation of the media with a range of unnatural salicylic acids failed to yield the corresponding nidulin analogues, suggesting the compounds may be biosynthesised by a single polyketide synthase. In total, 12 new and 11 previously reported nidulin analogues were isolated, characterised and assayed for in vitro activity against a panel of bacteria, fungi and mammalian cells, providing a comprehensive structure-activity profile for the nidulin scaffold.

Effects of two lichen acids isolated from Pseudevernia furfuracea (L.) Zopf in cultured human lymphocytes.

The present study aims at assessing the efficacies of olivetoric acid (OA) and physodic acid (PA) isolated from Pseudevernia furfuracea (L.) Zopf (Parmeliaceae) in human lymphocytes (HLs) in vitro. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assays were performed to establish cytotoxicity in HLs. Besides, oxidative stress and genotoxicity were monitored by estimating the changes of total oxidative stress (TOS) and 8-hydroxy-2'-deoxyguanosine (8-OH-dG) levels, respectively, in HLs. At the same time, OA- and PA-induced total antioxidant capacity (TAC) levels in HLs were determined. Although especially low concentrations of OA (IC50=109.94 mg/L) and PA (IC50=665.49 mg/L) did not show cytotoxic effect at high levels in HLs, it was revealed that cytotoxicity was significantly (p<0.05) associated with oxidative stress and genotoxicity via correlation analysis. While TOS level in HLs did not statistically (p>0.05) increase in the presence of all treatments (0.5-100 mg/L) of PA, TAC level was increased by PA applications in certain concentrations (0.5-10 mg/L). Overall, the obtained data indicate that OA and especially PA as lichen compounds that do not cause oxidative stress can be a new resource of therapeutics as recognized in the present study with their high antioxidant features.

Lichen-derived caperatic acid and physodic acid inhibit Wnt signaling in colorectal cancer cells.

Lichens are a source of secondary metabolites which possess important biological activities, including antioxidant, antibacterial, anti-inflammatory, and cytotoxic effects. The anticancer activity of lichens was shown in many types of tumors, including colorectal cancers (CRC). Several studies revealed that the application of lichen extracts diminished the proliferation of CRC cells and induced apoptosis. Colon carcinogenesis is associated with aberrations in Wnt signaling. Elevated transcriptional activity of ß-catenin induces cell survival, proliferation, and migration. Thus, the inhibition of Wnt signaling is a promising therapeutic strategy in colorectal cancer. The aim of this study was the evaluation of the effects of lichen-derived depsides (atranorin, lecanoric acid, squamatic acid) and depsidones (physodic acid, salazinic acid) and a poly-carboxylic fatty acid-caperatic acid, on Wnt signaling in HCT116 and DLD-1 colorectal cancer cell lines. HCT116 cells were more sensitive to the modulatory effects of the compounds. PKF118-310, which was used as a reference ß-catenin inhibitor, dose-dependently reduced the expression of the classical ß-catenin target gene-Axin2 in both cell lines. Lecanoric acid slightly reduced Axin2 expression in HCT116 cells while caperatic acid tended to reduce Axin2 expression in both cell lines. Physodic acid much more potently decreased Axin2 expression in HCT116 cells than in DLD-1 cells. Physodic acid and caperatic acid also diminished the expression of survivin and MMP7 in a cell line and time-dependent manner. None of the compounds affected the nuclear translocation of ß-catenin. This is the first report showing the ability of caperatic acid and physodic acid to modulate ß-catenin-dependent transcription.

Tubulin-binding dibenz[c,e]oxepines: Part 2. Structural variation and biological evaluation as tumour vasculature disrupting agents.

5,7-Dihydro-3,9,10,11-tetramethoxybenz[c,e]oxepin-4-ol 1, prepared from a dibenzyl ether precursor via Pd-catalysed intramolecular direct arylation, possesses broad-spectrum in vitro cytotoxicity towards various tumour cell lines, and induces vascular shutdown, necrosis and growth delay in tumour xenografts in mice at sub-toxic doses. The biological properties of 1 and related compounds can be attributed to their ability to inhibit microtubule assembly at the micromolar level, by binding reversibly to the same site of the tubulin αß-heterodimer as colchicine 2 and the allocolchinol, N-acetylcolchinol 4.

Evaluation of antioxidant and cytotoxic effects of olivetoric and physodic acid in cultured human amnion fibroblasts.

It is known that lichens are utilized for the treatment of many diseases including ulcer, diabetes, and cancer for many years. Secondary metabolites in the structure of the lichens provide various activity properties for them. In the present study, cytotoxic and oxidative effects of main constituents of Pseudevernia furfuracea (L.) Zopf (Parmeliaceae), olivetoric acid (OA), and physodic acid (PA) were investigated on cultured human amnion fibroblasts (HAFs). OA and PA were isolated from P. furfuracea using column chromatography and their structures were determined by proton nuclear magnetic resonance and carbon-13 nuclear magnetic resonance. HAFs were incubated during 48 h in the presence of OA and PA, at different concentrations from 6.25 mg/L to 200 mg/L. OA showed higher cytotoxicity than PA. In fact, median inhibitory concentration values of OA and PA were 571.27 and 3373.69 mg/L, respectively. The lower concentrations (<50 mg/L) of OA and PA did not cause oxidative stress and genotoxicity; furthermore, they supported anti-oxidative capacity of HAFs. Therefore, all these data suggested that both tested metabolites, especially PA might be developed as natural health medicine to protect human body against oxidative stress and genotoxicity. As far as we know, this is the first report on the cytotoxic and anti-oxidative activities of OA and PA on HAFs.

Lichen-derived compounds show potential for central nervous system therapeutics.

BACKGROUND: Natural products from lichens are widely investigated for their biological properties, yet their potential as central nervous system (CNS) therapeutic agents is less explored. PURPOSE: The present study investigated the neuroactive properties of selected lichen compounds (atranorin, perlatolic acid, physodic acid and usnic acid), for their neurotrophic, neurogenic and acetylcholine esterase (AChE) activities. METHODS: Neurotrophic activity (neurite outgrowth) was determined using murine neuroblastoma Neuro2A cells. A MTT assay was performed to assess the cytotoxicity of compounds at optimum neurotrophic activity. Neuro2A cells treated with neurotrophic lichen compounds were used for RT-PCR to evaluate the induction of genes that code for the neurotrophic markers BDNF and NGF. Immunoblotting was used to assess acetyl H3 and H4 levels, the epigenetic markers associated with neurotrophic and/or neurogenic activity. The neurogenic property of the compounds was determined using murine hippocampal primary cultures. AChE inhibition activity was performed using a modified Ellman's esterase method. RESULTS: Lichen compounds atranorin, perlatolic acid, physodic acid and (+)-usnic acid showed neurotrophic activity in a preliminary cell-based screening based on Neuro2A neurite outgrowth. Except for usnic acid, no cytotoxic effects were observed for the two depsides (atranorin and perlatolic acid) and the alkyl depsidone (physodic acid). Perlatolic acid appears to be promising, as it also exhibited AChE inhibition activity and potent proneurogenic activity. The neurotrophic lichen compounds (atranorin, perlatolic acid, physodic acid) modulated the gene expression of BDNF and NGF. In addition, perlatolic acid showed increased protein levels of acetyl H3 and H4 in Neuro2A cells. CONCLUSION: These lichen depsides and depsidones showed neuroactive properties in vitro (Neuro2A cells) and ex vivo (primary neural stem or progenitor cells), suggesting their potential to treat CNS disorders.

Depsidones from Lichens as Natural Product Inhibitors of M-Phase Phosphoprotein 1, a Human Kinesin Required for Cytokinesis.

M-Phase Phosphoprotein 1 (MPP1), a microtubule plus end directed kinesin, is required for the completion of cytokinesis. Previous studies have shown that MPP1 is upregulated in various types of bladder cancer. This article describes inhibitor screening leading to the identification of a new class of natural product inhibitors of MPP1. Two compounds with structural similarity, norlobaridone (1) and physodic acid (2), were found to inhibit MPP1. Physodic acid is not competitive with ATP, indicating the presence of an allosteric inhibitor-binding pocket. Initial drug-like property screening indicates that physodic acid is more soluble than norlobaridone and has more favorable lipophilicity. However, both suffer from high clearance in human microsomal stability assays mediated by the lability of the lactone ring as well as hydroxylation of the alkyl chains as shown by metabolite identification studies. In cell-based assays physodic acid is a weak inhibitor with EC50 values of about 30 µM in a range of tumor cell lines. The two depsidones identified and characterized here could be used for future improvement of their activity against MPP1 and will be useful chemical probes for studying this unique molecular motor in more depth.

Cytotoxic activity of physodic acid and acetone extract from Hypogymnia physodes against breast cancer cell lines.

CONTEXT: Lichens produce specific secondary metabolites with different biological activity. OBJECTIVE: This study investigated the cytotoxic effects of physodic acid, in addition to the total phenolic content and cytotoxic and antioxidant activity of acetone extract from Hypogymnia physodes (L.) Nyl. (Parmeliaceae). MATERIALS AND METHODS: Cytotoxicity of physodic acid (0.1-100 µM) was assessed in MDA-MB-231, MCF-7 and T-47D breast cancer cell lines and a nontumorigenic MCF-10A cell line using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, neutral red uptake and crystal violet assays during 72 h of incubation. An MTT assay was also used to assess the cytotoxic effects of the acetone extract (0.1-100 µg/mL) in the MDA-MB-231, MCF-7, T-47D breast cancer cell lines after 72 h. The total phenolic content of the acetone extract, expressed as the gallic acid equivalent, was investigated using Folin-Ciocalteu reagent. The antioxidant activity of the extract was assessed by 2,2-diphenyl-1-picrylhydrazyl and ferric-reducing antioxidant power assays. RESULTS: The cytotoxic activity of physodic acid appeared to be strong in the tumorigenic cell lines (IC50 46.0-93.9 µM). The compound was inactive against the nontumorigenic MCF-10A cell line (IC50 >100 µM). The acetone extract showed cytotoxicity in the breast cancer cell lines (IC50 46.2-110.4 µg/mL). The acetone extract was characterized by a high content of polyphenols, and it had significant antioxidant activity. DISCUSSION AND CONCLUSION: Physodic acid and acetone extract from H. physodes displayed cytotoxic effects in the breast cancer cell lines. Furthermore, acetone extract from H. physodes possessed significant antioxidant properties.

In vitro antitumor activities of the lichen compounds olivetoric, physodic and psoromic acid in rat neuron and glioblastoma cells.

Context Since methods utilised in the treatment of glioblastoma multiforme (GBM) are inadequate and have too many side effects, usage of herbal products in the treatment process comes into prominence. Lichens are symbiotic organisms used for medicinal purposes for many years. There are various anticancer treatments about components of two lichen species used in the present study. Objective Antitumor potential of three lichen secondary metabolites including olivetoric acid (OLA) and physodic acid (PHA) isolated from Pseudevernia furfuracea (L.) Zopf (Parmeliaceae) and psoromic acid (PSA) isolated from Rhizoplaca melanophthalma (DC.) Leuckert (Lecanoraceae) were investigated on human U87MG-GBM cell lines and primary rat cerebral cortex (PRCC) cells for the first time. Materials and methods PRCC cells used as healthy brain cells were obtained from Sprague-Dawley rats. The treatments were carried out on the cells cultured for 48 h. Cytotoxic effects of different concentrations (2.5, 5, 10, 20 and 40 mg/L) of metabolites on the cells were determined via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase (LDH) analyses. Total antioxidant capacity (TAC) and total oxidant status (TOS) parameters were used for assessing oxidative alterations. Oxidative DNA damage potentials of metabolites were investigated via evaluating 8-hydroxy-2'-deoxyguanosine (8-OH-dG) levels. Results Median inhibitory concentration (IC50) values of OLA, PHA and PSA were 125.71, 698.19 and 79.40 mg/L for PRCC cells and 17.55, 410.72 and 56.22 mg/L for U87MG cells, respectively. It was revealed that cytotoxic effects of these metabolites showed positive correlation with concentration, LDH activity and oxidative DNA damage. Discussion and conclusion The present findings obtained in this study revealed that primarily OLA and then PSA had high potential for use in the treatment of GBM.

Crystal structure-based discovery of a novel synthesized PARP1 inhibitor (OL-1) with apoptosis-inducing mechanisms in triple-negative breast cancer.

Poly (ADP-ribose) polymerase-1 (PARP1) is a highly conserved enzyme focused on the self-repair of cellular DNA damage. Until now, numbers of PARP inhibitors have been reported and used for breast cancer therapy in recent years, especially in TNBC. However, developing a new type PARP inhibitor with distinctive skeleton is alternatively promising strategy for TNBC therapy. In this study, based on co-crystallization studies and pharmacophore-docking-based virtual screening, we discovered a series of dihydrodibenzo[b,e]-oxepin compounds as PARP1 inhibitors. Lead optimization result in the identification of compound OL-1 (2-(11-(3-(dimethylamino)propylidene)-6,11- dihydrodibenzo[b,e]oxepin )-2-yl)acetohydrazide), which has a novel chemical scaffold and unique binding interaction with PARP1 protein. OL-1 demonstrated excellent potency (inhibiting PARP1 enzyme activity with IC50 = 0.079 µM), as well as inhibiting PARP-modulated PARylation and cell proliferation in MDA-MB-436 cells (BRAC1 mutation). In addition, OL-1 also inhibited cell migration that closely related to cancer metastasis and displayed remarkable anti-tumor efficacy in MDA-MB-436 xenograft model without apparent toxicities. These findings highlight a new small-molecule PAPR1 inhibitor (OL-1) that has the potential to impact future TNBC therapy.

Olopatadine inhibits exocytosis in rat peritoneal mast cells by counteracting membrane surface deformation.

BACKGROUND/AIMS: Besides its anti-allergic properties as a histamine receptor antagonist, olopatadine stabilizes mast cells by inhibiting the release of chemokines. Since olopatadine bears amphiphilic features and is preferentially partitioned into the lipid bilayers of the plasma membrane, it would induce some morphological changes in mast cells and thus affect the process of exocytosis. METHODS: Employing the standard patch-clamp whole-cell recording technique, we examined the effects of olopatadine and other anti-allergic drugs on the membrane capacitance (Cm) in rat peritoneal mast cells during exocytosis. Using confocal imaging of a water-soluble fluorescent dye, lucifer yellow, we also examined their effects on the deformation of the plasma membrane. RESULTS: Low concentrations of olopatadine (1 or 10 µM) did not significantly affect the GTP-γ-S-induced increase in the Cm. However, 100 µM and 1 mM olopatadine almost totally suppressed the increase in the Cm. Additionally, these doses completely washed out the trapping of the dye on the cell surface, indicating that olopatadine counteracted the membrane surface deformation induced by exocytosis. As shown by electron microscopy, olopatadine generated inward membrane bending in mast cells. CONCLUSION: This study provides electrophysiological evidence for the first time that olopatadine dose-dependently inhibits the process of exocytosis in rat peritoneal mast cells. Such mast cell stabilizing properties of olopatadine may be attributed to its counteracting effects on the plasma membrane deformation in degranulating mast cells.

Azelastine desensitization of transient receptor potential vanilloid 1: a potential mechanism explaining its therapeutic effect in nonallergic rhinitis.

BACKGROUND: Capsaicin, a prototypic transient receptor potential vanilloid 1 (TRPV1) agonist, has been shown to be more clinically effective in the treatment of nonallergic rhinitis (NAR) compared with other rhinitis subtypes. Azelastine has also been found to be clinically effective in the treatment of NAR but its mechanism(s) of action is still poorly elucidated. This study was designed to determine, using in vitro cell lines, whether topical therapies including azelastine have activity on TRPV1 ion channels similar to capsaicin. METHODS: The effects of capsaicin (1 µM), azelastine (30 µM), bepotastine (10 µM), olopatadine (10 µM), and fluticasone (200 µM) on TRPV1 channels using mice neuronal cells (Cath.a), as surrogates for submucosal sensory neurons, and human nasal epithelial cells (hNEC) were determined and compared. For azelastine, bepotastine, and capsaicin, which elicited an agonist effect on TRPV1, live cell [Ca(2+)] signaling in Cath.a cells and hNECs expressing TRPV1 were performed in the absence and presence of capsazepine at 10 µM (a TRPV1 antagonist) or using wild-type mouse embryonic fibroblasts (wtMEFs) that express TRPV1 ion channels and TRPV1 homozygous null mutant (TRPV1-/-) knockout MEF cells as controls to establish TRPV1 channel selectivity. As azelastine has previously been found clinically effective in NAR, additional experiments were performed to determine its ability to desensitize TRPV1 ion channels and its effect on regulating intracellular calcium homeostasis. RESULTS: Cath.a cells treated with azelastine, bepotastine, or capsaicin showed a significant increase in TRPV1-dependant (Ca(2+)) specific cytosolic fluorescence. Continuous treatment with azelastine or capsaicin resulted in desensitization of TRPV1 channels. In hNECs, azelastine stimulation resulted in Ca(2+) shifts from the cytosol to mitochondria and overexpression of hematopoietic cell-specific Lyn substrate 1-associated protein X1, which may thus be effective in cytosolic Ca(2+) homeostasis. CONCLUSION: Azelastine, similar to capsaicin, exhibits direct activity on TRPV1 ion channels that may represent a novel mechanistic pathway explaining its clinical efficacy in NAR.

Evaluation of the antihistamine effects of olopatadine and levocetirizine during a 24-h period: a double-blind, randomized, cross-over, placebo-controlled comparison in skin responses induced by histamine iontophoresis.

The antihistamine effects of olopatadine and levocetirizine, in standard-dose application described in their information (5 mg twice a day for olopatadine; 5 mg once daily for levocetirizine), were examined from 11.5 to 24 h after application. The test was designed in a double-blind, randomized, cross-over, placebo-controlled study of 12 healthy volunteers on histamine-induced flare and wheal response using an iontophoresis technique. The suppressive effect of olopatadine on the wheals induced by a 0.1-mA histamine iontophoresis lasted for 24 h after dosing. Both drugs inhibited flare induced by histamine iontophoresis almost completely until 24 h after the first administration. Suppression of the 0.2-mA-induced wheal response by levocetirizine, taken once daily, decreased with time, although 0.1-mA-induced flare was almost completely suppressed by the drug. Olopatadine completely suppressed even the wheal response induced by a 0.2-mA histamine iontophoresis. Compared with the placebo, the two drugs significantly suppressed the subjective itching assessed by visual analog scale at all intervals. There were no significant differences in subjective drowsiness and objective cognitive function between drug- and placebo-treated subjects. These results demonstrate that olopatadine seems to be more potent than levocetirizine when administrated in a standard dose. In conclusion, mild to moderate urticaria could be controlled by standard application as described in their information. On the other hand, severe urticaria could be managed by a standard application of olopatadine, but levocetirizine may need an additional dose to control severe urticaria.

Antiallergic effects of pigments isolated from green sea urchin (Strongylocentrotus droebachiensis) shells.

This study was undertaken to evaluate possible antiallergic effects of an extract of pigments from green sea urchin (Strongylocentrotus droebachiensis) shells. Effects were studied on animal models - guinea pig ileum contraction, rabbit eyes allergic conjunctivitis, and rabbit local skin irritation. The extract significantly reduced, in a dose-dependent manner, the histamine-induced contractions of the isolated guinea pig ileum with ID50 =1.2 µg/mL (in equivalents of spinochrome B), had an inhibitory effect on the model of ocular allergic inflammation surpassing the reference drug olopatadine, and did not show any irritating effect in rabbits. The extract predominantly contained polyhydroxy-1,4-naphthoquinone which would be responsible for the pharmacological activity. The active compounds of the extract were evaluated in silico with molecular docking. Molecular docking into H1R receptor structures obtained from molecular dynamic simulations showed that all spinochrome derivatives bind to the receptor active site, but spinochrome monomers fit better to it. The results of the present study suggest possibilities for the development of new agents for treating allergic diseases on the base of pigments from sea urchins shells.

Metabolically stable dibenzo[b,e]oxepin-11(6H)-ones as highly selective p38 MAP kinase inhibitors: optimizing anti-cytokine activity in human whole blood.

Five series of metabolically stable disubstituted dibenzo[b,e]oxepin-11(6H)-ones were synthesized and tested in a p38α enzyme assay for their inhibition of tumor necrosis factor-α (TNF-α) release in human whole blood. Compared to the monosubstituted dibenzo[b,e]oxepin-11(6H)-one derivatives, it has been shown that the additional introduction of hydrophilic residues at position 9 leads to a substantial improvement of the inhibitory potency and metabolic stability. Using protein X-ray crystallography, the binding mode of the disubstituted dibenzoxepinones and the induction of a glyince flip in the hinge region were confirmed. The most potent compound of this series, 32e, shows an outstanding biological activity on isolated p38α, with an IC50 value of 1.6 nM, extraordinary selectivity (by a factor >1000, Kinase WholePanelProfiler), and low ATP competitiveness. The ability to inhibit the release of TNF-α from human whole blood was optimized down to an IC50 value of 125 nM. With the promising dibenzoxepinone inhibitor 3i, a pharmacokinetic study in mice was conducted.

Effect of Hypogymnia physodes extracts and their depsidones on micronucleus distribution in human lymphocytes.

Three lichen depsidones, physodalic acid (1), physodic acid (2), and 3-hydroxy physodic acid (3), were isolated from Hypogymnia physodes diethyl ether extract using column chromatography, and their structures determined by comparing their UV, 1H and 13C NMR spectroscopic and MS data with those given in the literature, as well as with data computed by CHEM draw ultra 11 software. The contents of 1, 2 and 3 were determined in the methanol (ME), acetone (AE), and diethyl ether (EE) extracts using reversed-phase high performance liquid chromatography with photodiode array detection. The extracts, isolated depsidones 1-3 and EE fraction F23 (consisting of 90% 2 and 3, in the ratio 5.5: 1) were evaluated for their in vitro effects on chromosome aberrations in peripheral human lymphocytes using the cytochalasin-B blocked micronucleus (CBMN) assay in doses of 1 microg/mL and 2 microg/mL of final culture solution. The frequency of MN was scored in binucleated cells, and nuclear proliferation index was calculated. It was found that 1, 2, 3, F23, and EE at 1.0 microg/mL exerted a beneficial effect on lymphocyte cells giving a significant decrease of the frequency of MN in comparison with the positive control Amifostin WR-2721.Among the tested depsidones at a concentration of 1 microg/mL, 3 exhibited the most prominent effect decreasing the frequency of MN by 30.3%, followed by 2 (28.2%) and 1 (22.0%). The extracts were less effective than the isolated depsidones.