Identification of Non-Macrocyclic Small Molecule Inhibitors against the NS3/4A Serine Protease of Hepatitis C Virus through in Silico Screening

Drug discovery and design for inhibition of the Hepatitis C Virus (HCV) NS3/4A serine protease is a major challenge. The broad, shallow, and generally featureless nature of the active site makes it a difficult target for "hit" selection especially using standard docking programs. There are several macrocyclic NS3/4A protease inhibitors that have been approved or are in clinical trials to treat chronic HCV (alone or as combination therapy), but most of the current therapies for HCV infection have untoward side effects, indicating a continuing medical need for the discovery of novel therapeutics with improved efficacy. In this study, we designed and implemented a two-tiered and progressive docking regime that successfully identified five non-macrocyclic small molecules that show inhibitory activity in the low micromolar range. Of these, four compounds show varying inhibition against HCV subgenotypes 1b, 1a, 2a, and 4d. The top inhibitor (3) has an IC(50) value of 15 μM against both subgenotypes 1b and 2a of the NS3/4A protease enzyme. Another inhibitor, 1, inhibits all four subgenotypes with moderate activity, showing highest activity for genotype 2a (24 μM). The five inhibitors presented in this study could be valuable candidates for future hit to lead optimization. Additionally, enzyme-inhibitor interaction models presented herein provide key information regarding structural differences between the active sites of the NS3/4A protease of the HCV subgenotype 1a and 1b that might explain the variable inhibitory activity between subgenotypes of the small molecule inhibitors identified here.     

Medicinal Flowers. Part 29. Acylated Oleanane‐Type Triterpene Bisdesmosides: Perennisaponins G,H, I,J, K,L, and M with Pancreatic Lipase Inhibitory Activity from the Flowers of Bellis perennis

The MeOH extract from the flowers of Bellis perennis was found to show pancreatic‐lipase inhibitory activity (IC₅₀ 455 μg/ml). From the extract, seven new triterpene saponins named perennisaponins G ( 1 ; IC₅₀ 163 μM ), H ( 2 ; 137 μM ), I ( 3 ; 147 μM ), J ( 4 ; 148 μM ), K ( 5 ; 223 μM ), L ( 6 ; 81.4 μM ), and M ( 7 ; 195 μM ) were isolated as pancreatic lipase inhibitors. The stereostructures of 1 – 7 were elucidated on the basis of chemical and spectroscopic evidence.     

Prussian Blue Modified Carbon Ionic Liquid Electrode: Electrochemical Characterization and Its Application for Hydrogen Peroxide and Glucose Measurements

Prussian blue modified carbon ionic liquid electrodes (PB‐CILEs) were fabricated using chemical and electrochemical procedures. Chemically fabricated PB‐CILE exhibited an excellent sensitivity (0.0866 μA μM^?1), low detection limit (0.01 μM) and two linear ranges (0.01–1 and 1–600 μM) toward hydrogen peroxide. Then, glucose oxidase (GOx) was immobilized on the surface of PB‐CILE to fabricate glucose biosensor using three different procedures involving cross linking with glutaraldehyde (GLU) and bovine serum albumin (BSA), entrapment into the Nafion matrix and covering with a sol‐gel layer. Glucose biosensor fabricated using cross linking procedure showed the best sensitivity (0.0019 μA μM^?1) and operational stability for glucose.     

Antioxidant inhibitors potentiate the cytotoxicity of photodynamic therapy

Photodynamic therapy (PDT) is an increasingly popular anticancer treatment that uses photosensitizer, light and tissue oxygen to generate cytotoxic reactive oxygen species (ROS) within illuminated cells. Acting to counteract ROS-mediated damage are various cellular antioxidant pathways. In this study, we combined PDT with specific antioxidant inhibitors to potentiate PDT cytotoxicity in MCF-7 cancer cells. We used disulphonated aluminium phthalocyanine photosensitizer plus various combinations of the antioxidant inhibitors: diethyl-dithiocarbamate (DDC, a Cu/Zn-SOD inhibitor), 2-methoxyestradiol (2-ME, a Mn-SOD inhibitor), l-buthionine sulfoximine (BSO, a glutathione synthesis inhibitor) and 3-amino-1,2,4-triazole (3-AT, a catalase inhibitor). BSO, singly or in combination with other antioxidant inhibitors, significantly potentiated PDT cytotoxicity, corresponding with increased ROS levels and apoptosis. The greatest potentiation of cell death over PDT alone was seen when cells were preincubated for 24 h with 300 μM BSO plus 10 mM 3-AT (1.62-fold potentiation) or 300 μM BSO plus 1 μM 2-ME (1.52-fold), or with a combination of all four inhibitors (300 μM BSO, 10 mM 3-AT, 1 μM 2-ME and 10 μM DDC: 1.4-fold). As many of these inhibitors have already been clinically tested, this work facilitates future in vivo studies.     

Platinum(IV) coordination compounds containing 5-methyl-1,2,4-triazolo[1,5-a]pyrimidin-7(4H)-one as nonleaving ligand. Molecular and cytotoxicity in vitro characterization

Novel platinum(IV) coordination compounds with 5-methyl-1,2,4-triazolo[1,5-a]pyrimidin-7(4H)-one (HmtpO): cis-trans-[PtCl(2)(OH)(2)(NH(3))(HmtpO)] (1), cis-trans-[PtCl(5)(HmtpO)][(CH(3))(2)NH(2)] (2) have been prepared and structurally characterized by spectroscopic methods ((1)H, IR and X-ray crystallography (2)). The X-ray results indicate that the local geometry around the platinum(IV) centre approximates a typical octahedral arrangement with nitrogen atom N3 of the HmtpO and three chloride atoms in equatorial positions. The remaining two axial positions are occupied by two chlorides. The preliminary assessment of antitumor properties of (1) was performed as an in vitro antiproliferative activity against HL-60 human acute promyelocytic leukemia and HCV29T bladder cancer. The cis-trans-[PtCl(2)(OH)(2)(NH(3))(HmtpO)] (1) exhibits higher cytotoxic activity against HL-60 (IC(50)=6.4 μM) than cisplatin.     

Spectroelectrochemical investigation of intramolecular and interfacial electron-transfer rates reveals differences between nitrite reductase at rest and during turnover

A combined fluorescence and electrochemical method is described that is used to simultaneously monitor the type-1 copper oxidation state and the nitrite turnover rate of a nitrite reductase (NiR) from Alcaligenes faecalis S-6. The catalytic activity of NiR is measured electrochemically by exploiting a direct electron transfer to fluorescently labeled enzyme molecules immobilized on modified gold electrodes, whereas the redox state of the type-1 copper site is determined from fluorescence intensity changes caused by Fo?rster resonance energy transfer (FRET) between a fluorophore attached to NiR and its type-1 copper site. The homotrimeric structure of the enzyme is reflected in heterogeneous interfacial electron-transfer kinetics with two monomers having a 25-fold slower kinetics than the third monomer. The intramolecular electron-transfer rate between the type-1 and type-2 copper site changes at high nitrite concentration (≥520 μM), resulting in an inhibition effect at low pH and a catalytic gain in enzyme activity at high pH. We propose that the intramolecular rate is significantly reduced in turnover conditions compared to the enzyme at rest, with an exception at low pH/nitrite conditions. This effect is attributed to slower reduction rate of type-2 copper center due to a rate-limiting protonation step of residues in the enzyme's active site, gating the intramolecular electron transfer.     

A search for BACE inhibitors reveals new biosynthetically related pyrrolidones, furanones and pyrroles from a southern Australian marine sponge, Ianthella sp

Fractionation of a southern Australian marine sponge, Ianthella sp., yielded sixteen metabolites including a new class of pyrrolidone, ianthellidones A-F (1-6), a new class of furanone, ianthellidones G-H (7-8), new and known lamellarins, lamellarins O1 (9), O2 (10), O (11) and Q (12), plus the known 4-hydroxybenzaldehyde (13), 4-hydroxybenzoic acid (14), 4-methoxybenzoic acid (15) and ethyl 4-hydroxybenzoate (16). Structures for all Ianthella metabolites were determined by detailed spectroscopic analysis, supported by a plausible biosynthetic relationship. The ianthellidones were non-cytotoxic towards two human colon cancer cell lines (SW620 and SW620 Ad300), as well as Gram +ve and Gram -ve bacteria, and a fungus. Ianthellidone F (6) and lamellarins O2 (10) and O (11) displayed modest BACE inhibitory properties (IC(50) > 10 μM), while lamellarin O1 (9) was more potent (IC(50) < 10 μM). Lamellarin O (11) exhibited modest cytotoxicity towards SW620 and SW620 Ad300 cell lines (IC(50) > 22 μM), was an inhibitor of the multi-drug resistance efflux pump P-glycoprotein, and displayed selective growth inhibitory activity against the Gram +ve bacterium Bacillus subtilis (ATCC 6633) (IC(50) 2.5 μM).     

Dendroaspis natriuretic peptide regulates the cardiac L-type Ca2+ channel activity by the phosphorylation of α1c proteins

Dendroaspis natriuretic peptide (DNP), a new member of the natriuretic peptide family, is structurally similar to atrial, brain, and C-type natriuretic peptides. However, the effects of DNP on the cardiac function are poorly defined. In the present study, we examined the effect of DNP on the cardiac L-type Ca(2+) channels in rabbit ventricular myocytes. DNP inhibited the L-type Ca(2+) current (I(Ca,L)) in a concentration dependent manner with a IC(50) of 25.5 nM, which was blocked by an inhibitor of protein kinase G (PKG), KT5823 (1 μM). DNP did not affect the voltage dependence of activation and inactivation of I(Ca,L). The α(1c) subunit of cardiac L-type Ca(2+) channel proteins was phosphorylated by the treatment of DNP (1 μM), which was completely blocked by KT5823 (1 μM). Finally, DNP also caused the shortening of action potential duration in rabbit ventricular tissue by 22.3 ± 4.2% of the control (n = 6), which was completely blocked by KT5823 (1 μM). These results clearly indicate that DNP inhibits the L-type Ca(2+) channel activity by phosphorylating the Ca(2+) channel protein via PKG activation.     

Titanium silicalite-1 zeolite microparticles for enzymeless H2O2 detection

In this communication, we demonstrate for the first time that titanium silicalite-1 zeolite microparticles (TSZMs) can effectively catalyze the reduction of H(2)O(2), leading to an enzymeless H(2)O(2) sensor with a linear detection range from 100 μM to 40 mM (r = 0.994) and a detection limit of 0.5 μM at a signal-to-noise ratio of 3.     

Pseuduvarines A and B, two new cytotoxic dioxoaporphine alkaloids from Pseuduvaria rugosa

Pseuduvarines A (1) and B (2), two new dioxoaporphine alkaloids with an amino moiety, were isolated from the stem bark of Pseuduvaria rugosa and their structures were elucidated by combination of 2D-NMR spectroscopic analysis. Pseuduvarines A (1) and B (2) showed cytotoxicity against MCF7, HepG2, and HL-60 (1: IC??, 0.9, 21.7, and >50.0 μM, respectively, 2: IC?? >50.0, 15.7, and 12.4 μM, respectively).     

Fullerene Black Modified Screen Printed Electrodes for the Quantification of Acetaminophen and Guanine

Fullerene Black (FB) and Extracted Fullerene Black (EFB) were used in modified screen‐printed electrodes producing electrochemical transducers (FB‐SPEs and EFB‐SPEs). A complete electrochemical study was performed and the best results are obtained working with FB‐SPEs, especially in terms of: 1. improved electron‐transfer kinetic mechanisms and 2. sensitivity and selectivity toward Acetaminophen (Ac) and Guanine (G). These latter represent two important electro‐active targets to quantify in medicine field application, because: Ac is a preferred alternative (as analgesic‐antipyretic agent) to aspirin, particularly for patients who cannot tolerate aspirin; the oxidation signal of G is useful for the fabrication of emerging analytical tools, such as DNA chipsand user‐friendly diagnostic devices. Ac and G are quantify by using FB‐SPEs electrochemical devices, with an extended linearity (1–300 μM for Ac; 0.1–300 μM for G), an excellent sensitivity (2.82 μA μM⁻¹ cm⁻² in the case of Ac; and 0.183 μA μM⁻¹ cm⁻² in the case of G), a low detection limit (0.01 μM for Ac; 0.005 μM for G), a very good reproducibility (both: intra‐; inter‐electrodes reproducibility RSD % ranging from 0.3–0.5 for Ac; and 0.50–0.85 for G) and a very fast response time (6 s for Ac; 5 s in the case of G). In addition, high selectivity is obtained at FB‐SPEs, meaning that the FB‐SPEs electrochemical transducers are suitable to simultaneously quantify Ac and G in real samples, having several different (highly concentrated) interference.     

Synthesis,cytotoxic, and carbonic anhydrase inhibitory effects of new 2-(3-(4-methoxyphenyl)-5-(aryl)-4,5-dihydro-1H-pyrazol-1-yl)benzo[d]thiazole derivatives

2-(3-[4-Methoxyphenyl]-5-aryl-4,5-dihydro-1H-pyrazol-1-yl)benzo[d]thiazoles ( 1b-7b ) were synthesized for the first time except 1b , and spectral methods such as ¹H NMR, ¹³C NMR and HRMS were utilized to illuminate the chemical structures of the synthesized compounds. Phenyl ( 1b ), 2-methoxyphenyl ( 2b ), 4-methoxyphenyl ( 3b ), 4-methoxy-3-hydroxyphenyl ( 4b ), 2,5-dimethoxyphenyl ( 5b ), 3,4,5-trimethoxyphenyl ( 6b ), or thiophene-2-yl ( 7b ) was used as a aryl part. The inhibitory effects of the compounds were evaluated toward human carbonic anhydrase I and II enzymes (hCA I and hCA II). In vitro cytotoxic effects of the compounds against human oral squamous carcinomas and human normal oral cells were carried out via MTT. The compounds ( 1b-7b ) had Ki values of 36.87 ± 11.62-66.24 ± 2.99 μM (hCA I) and 22.66 ± 1.41-89.95 ± 6.25 μM (hCA II). Compounds 1b (Ki = 36.87 ± 11.62 μM) toward hCA I, 6b (Ki = 22.66 ± 1.41 μM) toward hCA II had significant enzyme inhibitory potency. Compound 6b had the highest tumor selectivity (TS = 29.3) and potency selectivity expression (PSE = 272.3) values. Therefore, compounds 1b and 6b with CAs inhibition effect and compound 6b with the cytotoxicity may be forwarded to further studies as potent compounds.     

Spectrophotometric flow-injection determination of urea in body fluids by using an immobilized urease reactor

A flow system involving a packed-bed enzyme reactor (volume 180 μl) with urease immobilized covalently on poly(glycidyl methacrylate)-coated porous glass is used for determining urea in blood serum and urine. Enzymatically produced ammonia is converted to an indophenolate dye (by oxidative coupling with hypochlorite and sodium salicylate), which is detected spectrophotometrically at 700 nm. The calibration graph is rectilinear for 25–500 μM urea when injecting samples (75 μl) diluted 1:50 for serum or 1:1000 for urine at a frequency of 60 h^?1; the relative standard deviation is 1.1% for ten injections of 300 μM urea. The immobilized urease is stabilized by the addition of disodium EDTA, sodium azide and 2-mercaptoethanol to a 0.2 M phosphate buffer (pH 6.9) used as the carrier stream, which serves also as a preservative for longterm storage of the urease reactor packing at 4°C.     

Nitrile-Containing Fischerindoles from the Cultured Cyanobacterium Fischerella sp

Chemical investigation of the cultured cyanobacterium Fischerella sp. (SAG strain number 46.79) led to the isolation of four nitrile-containing indole alkaloids, namely 12-epi-fischerindole I nitrile (1), deschloro 12-epi-fischerindole I nitrile (2), 12-epi-fischerindole W nitrile (3), and deschloro 12-epi-fischerindole W nitrile (4) along with a known metabolite hapalosin. The structures were determined by detailed spectroscopic analyses on the basis of 1D and 2D NMR and HRESIMS data. All isolates were evaluated for cytotoxicity against human cancer cells and for 20S proteasome inhibition. Deschloro 12-epi-fischerindole I nitrile (2) was found to be weakly cytotoxic against HT-29 cells with an ED(50) value of 23 μM. Hapalosin showed weak cytotoxicity against HT-29 and MCF-7 cells with ED(50) values of 22 and 27 μM, respectively, as well as moderate 20S proteasome inhibition with an IC(50) value of 12 μM. Compounds 1-4 all contain a nitrile moiety instead of the isonitrile found in all fischerindoles reported to date. Compounds 3 and 4 also display a new carbon skeleton, in which a six-membered ring replaces the five-membered ring normally found in fischerindole-type alkaloids.     

Electrochemical Investigation of Ruxolitinib: Sensitive Voltammetric Assay in Drug Product and Human Serum by Using Different Solid Electrodes

Ruxolitinib (RUX), a compound of the pyrrolopyrimidines class with activity as a tyrosine kinase inhibitory drug, is used to treat myelofibrosis. This study is reported for the detailed electrochemical behavior of RUX. The effects of supporting electrolyte, pH, and scanning rate on the peak potentials and currents of RUX were investigated by BDDE and GCE using different voltammetric techniques. Under optimum experimental conditions, calibration curves for RUX were obtained as 4 μM–80 μM and 1 μM–80 μM with a limit of detection (LOD) of 0.517 μM and 0.192 μM by the GCE and BDDE, respectively using DPV.     

Voltammetric Sensing of Caffeine in Food Sample Using Cu-MOF and Graphene

Cu-MOF/graphene composite were obtained by solvothermal and electrochemical methods. The interaction and formation of Cu-MOF/graphene (GE) composite was systematically studied with the support of various characterizations including X-ray diffraction (XRD), field emission scanning electron microscopy (FE-SEM) and cyclic voltammogram (CV). The Cu-MOF/graphene composite coated glassy carbon (GC) electrode exhibited good catalytic performance towards the electro-oxidation of caffeine in neutral condition. The modified electrode displayed good linear range (5 μM to 450 μM), sensitivity (0.710 μA μM⁻¹ cm⁻²), detection limit (1.38 μM), selectivity, high stability and reproducibility. Finally, the fabricated electrode extended into successful detection of caffeine in tea and coffee samples with good recoveries.     

Selective and Sensitive Determination of Paracetamol and Levodopa with Using Electropolymerized 3,5-Diamino-1,2,4-triazole Film on Glassy Carbon Electrode

For determination of levodopa (L-DOPA) and paracetamol (PAR), selective, effective and sensitive electrochemical sensor based on 3,5-diamino-1,2,4-triazole (35DT) on glassy carbon (GC) electrode was reported. The DPV method was used to obtain the oxidation peak current of L-DOPA which increased linearly with its concentration at the range of 1–99 μM and 99–480 μM. The linear relationship between the concentration of PAR and its peak current was obtained in the range of 0.3–55 μM and 55–475 μM. The limit of detection values for PAR and L-DOPA were found to be 0.1 and 0.25 μM, respectively. Satisfactory results were obtained in pharmaceutical samples.     

Synthesis and evaluation of in vitro bioactivity for polysubstituted N-arylpyrazole derivatives

New polysubstituted N-arylpyrazole derivatives were synthesized from N1-arylsydnone with acetylene and boronic acid, including 2-thiophenyl, 3-thiophenyl, 2-benzo[b]thiophenyl, or dibenzothiophenyl-4-boronic acid, via 1,3-dipolar cycloaddition and Suzuki coupling reaction. Based on the growth inhibitory activity results against lung carcinoma (NCI-H226), nasopharyngeal (NPC-TW01), and T-cell leukemia (Jurkat) cancer cells, compounds 5d and 7d with dibenzothiophenyl bioisostere possessed the significant inhibitory activity for NPC-TW01 (32 μM and 16 μM) and NCI-H226 (16 μM and 8.9 μM), respectively.     

Directed synthesis of μ-1,3,5 bridged dicyanamides by thermal decomposition of μ-1,5 bridged precursor compounds

Reaction of transition-metal dicyanamides with pyridazine leads to the formation of the ligand-rich 1 : 2 (1 : 2 = ratio between metal salt and organic co-ligand) compounds [M(dca)(2)(pydz)(2)](n) (dca = dicyanamide, pydz = pyridazine) with M = Mn (1-Mn), Fe (1-Fe), Co (1-Co), Ni (1-Ni). In their crystal structures linear polymeric M-(dca)(2)-M chains are found, in which the M(ii) cations are μ-1,5 bridged by the dca anions. The pydz ligands are terminally N-bonded to the cations, which are octahedrally coordinated by two pydz ligands and four dca anions. On heating these precursor compounds, 1-Mn, 1-Fe and 1-Co transform quantitatively into new ligand-deficient 1 : 1 intermediate compounds of composition [M(dca)(2)(pydz)](n) with M = Mn (2-Mn), Fe (2-Fe) and Co (2-Co). Investigations by IR spectroscopy, and single crystal X-ray structure analysis, show that the intermediates form a more condensed layered structure in which half of the pristine μ-1,5 bridged dca anions become μ-1,3,5 bridging. This structural transformation is accompanied by a pronounced change of their magnetic properties: whereas the ligand-rich 1 : 2 compounds show only Curie-Weiss paramagnetism, the ligand-deficient 1 : 1 intermediates show either antiferro- or ferromagnetic ordering at lower temperatures mediated by the three-atom pathway of the μ-1,3,5 bridging dca anions.     

Synthesis and cytotoxic activity of some novel N-pyridinyl-2-(6-phenylimidazo[2,1-b]thiazol-3-yl)acetamide derivatives

A series of novel compounds bearing imidazo[2,1-b]thiazole scaffolds were designed and synthesized based on the optimization of the virtual screening hit compound N-(6-morpholinopyridin-3-yl)-2-(6-phenylimidazo[2,1-b]thiazol-3-yl)acetamide (5a), and tested for their cytotoxicity against human cancer cell lines, including HepG2 and MDA-MB-231. The results indicated that the compound 2-(6-(4-chlorophenyl)imidazo[2,1-b]thiazol-3-yl)-N-(6-(4-(4-methoxybenzyl)piperazin-1-yl)pyridin-3-yl)acetamide (5l), with slightly higher inhibition on VEGFR2 than 5a (5.72% and 3.76% inhibitory rate at 20 μM, respectively), was a potential inhibitor against MDA-MB-231 (IC(50) = 1.4 μM) compared with sorafenib (IC(50) = 5.2 μM), and showed more selectivity against MDA-MB-231 than HepG2 cell line (IC(50) = 22.6 μM).