HTS identifies novel and specific uncompetitive inhibitors of the two-component NS2B-NS3 proteinase of West Nile virus.


:West Nile virus (WNV), a member of the Flavividae family, is a mosquito-borne, emerging pathogen. In addition to WNV, the family includes dengue, yellow fever, and Japanese encephalitis viruses, which affect millions of individuals worldwide. Because countermeasures are currently unavailable, flaviviral therapy is urgently required. The flaviviral two-component nonstructural NS2B-NS3 proteinase (protease [pro]) is essential for viral life cycle and, consequently, is a promising drug target. We report here the results of the miniaturization of an NS2B-NS3pro activity assay, followed by high-throughput screening of the National Institutes of Health's 65,000 compound library and identification of novel, uncompetitive inhibitors of WNV NS2B-NS3pro that appear to interfere with the productive interactions of the NS2B cofactor with the NS3pro domain. We anticipate that following structure optimization, the identified probes could form the foundation for the design of novel and specific therapeutics for WNV infection. We also provide the structural basis for additional species-selective allosteric inhibitors of flaviviruses.


Assay Drug Dev Technol


Johnston PA,Phillips J,Shun TY,Shinde S,Lazo JS,Huryn DM,Myers MC,Ratnikov BI,Smith JW,Su Y,Dahl R,Cosford ND,Shiryaev SA,Strongin AY




Has Abstract


2007-12-01 00:00:00












  • Transgenic mice targeting the heart unveil G protein-coupled receptor kinases as therapeutic targets.

    abstract::GRKs critically regulate betaAR signaling via receptor phosphorylation and the triggering of desensitization. In the heart, betaARs control the chronotropic, lusitropic, and inotropic responses to the catecholamine neurotransmitters, norepinephrine and epinephrine. Signaling through cardiac betaARs is significantly im...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章,评审


    authors: Iaccarino G,Koch WJ

    更新日期:2003-04-01 00:00:00

  • High-content assay for measuring intracellular growth of Leishmania in human macrophages.

    abstract::Leishmania species are sandfly-transmitted protozoan parasites that cause a spectrum of diseases, ranging from localized skin lesions to fatal visceral disease, in more than 12 million people worldwide. These parasites primarily target macrophages in their mammalian hosts and proliferate as non-motile amastigotes in t...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Dagley MJ,Saunders EC,Simpson KJ,McConville MJ

    更新日期:2015-09-01 00:00:00

  • Multi-photon excitation of intrinsic protein fluorescence and its application to pharmaceutical drug screening.

    abstract::The majority of proteins contain intrinsic fluorophores as natural sensors of molecular structures, dynamics, and interactions. The intrinsic protein fluorescence signal allows for the label-free and, hence, undisturbed and rapid study of protein-ligand interactions. Ultraviolet-based drug screening is hampered by the...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Buehler C,Dreessen J,Mueller K,So PT,Schilb A,Hassiepen U,Stoeckli KA,Auer M

    更新日期:2005-04-01 00:00:00

  • A Flexible Multiplatform Bioanalytical Strategy for Measurement of Total Circulating Shed Target Receptors: Application to Soluble B Cell Maturation Antigen Levels in the Presence of a Bispecific Antibody Drug.

    abstract::B cell maturation antigen (BCMA) is a membrane-bound receptor that is overexpressed on multiple myeloma cells and can be targeted with biotherapeutics. Soluble shed forms of membrane-associated receptors in circulation can act as a drug sink, especially when it is present in high molar ratio compared to drug concentra...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Stauffer A,Ray C,Hall M

    更新日期:2021-01-01 00:00:00

  • High-Throughput Screening of Na(V)1.7 Modulators Using a Giga-Seal Automated Patch Clamp Instrument.

    abstract::Voltage-gated sodium (Na(V)) channels have an essential role in the initiation and propagation of action potentials in excitable cells, such as neurons. Of these channels, Na(V)1.7 has been indicated as a key channel for pain sensation. While extensive efforts have gone into discovering novel Na(V)1.7 modulating compo...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Chambers C,Witton I,Adams C,Marrington L,Kammonen J

    更新日期:2016-03-01 00:00:00

  • Miniaturized, ultra-high throughput screening of tyrosine kinases using homogeneous, competitive fluorescence immunoassays.

    abstract::Assay miniaturization and the implementation of high-density 1,536-microwell screening increase the speed and efficiency of screening and lead discovery. To serve this need, a platform of miniaturizable assay technologies has been assembled for specific biological targets. This platform will enable initiation and comp...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Beasley JR,McCoy PM,Walker TL,Dunn DA

    更新日期:2004-04-01 00:00:00

  • Assay development and multivariate scoring for high-content discovery of chemoprotectants of endoplasmic-reticulum-stress-mediated amylin-induced cytotoxicity in pancreatic beta cells.

    abstract::The underlying pathogenesis of type-II diabetes mellitus is in the dysfunction and selective loss of pancreatic islet β-cells, which ultimately leads to underproduction of endogenous insulin. Amylin, a 37-amino-acid human hormone that is cosecreted with insulin, helps regulate gastric emptying and maintain blood gluco...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Law CJ,Ashcroft HA,Zheng W,Sexton JZ

    更新日期:2014-09-01 00:00:00

  • Validating a firefly luciferase-based high-throughput screening assay for antimalarial drug discovery.

    abstract::The emergence and spread of multidrug-resistant Plasmodium falciparum and recent detection of potential artemisinin-resistant strains in Southeast Asia highlight the importance of developing novel antimalarial therapies. Using a previously generated stable transgenic P. falciparum line with high-level firefly lucifera...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Che P,Cui L,Kutsch O,Cui L,Li Q

    更新日期:2012-02-01 00:00:00

  • Development and validation of a cell-based assay for the nuclear receptor retinoid-related orphan receptor gamma.

    abstract::The nuclear receptor retinoid-related orphan receptor gamma (RORγ) has become an attractive target for drug discovery due to its important role in the development and differentiation of Th17 cells, a subset of T cells that produce interleukin-17 and are involved in the pathogenesis of human inflammatory and autoimmune...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Wilkinson J,Hallis T,Hermanson S,Bi K

    更新日期:2011-04-01 00:00:00

  • Melt Dispersion Adsorbed onto Porous Carriers: An Effective Method to Enhance the Dissolution and Flow Properties of Raloxifene Hydrochloride.

    abstract::The objective of the present investigation is to enhance the dissolution and flow properties of raloxifene hydrochloride (RXH), a biopharmaceutical classification system class II drug. Melt dispersion of RXH with polyethylene glycol (PEG) 6000 was prepared by the fusion method. The melt dispersion was then adsorbed on...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Mahajan AN,Surti N,Patel P,Patel A,Shah D,Patel V

    更新日期:2020-08-01 00:00:00

  • WGA-coated yttrium oxide beads enable an imaging-based adenosine 2a receptor binding scintillation proximity assay suitable for high throughput screening.

    abstract::Adenosine receptors belong to the superfamily of G protein-coupled receptors and are involved in a variety of physiologic functions. Traditionally, binding assays to detect adenosine 2a (A2a) antagonists and agonists have used filtration methods that are cumbersome to run and not amenable to HTS. We developed scintill...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Bryant R,McGuinness D,Turek-Etienne T,Guyer D,Yu L,Howells L,Caravano J,Zhai Y,Lachowicz J

    更新日期:2004-06-01 00:00:00

  • Agonists of the orphan human G2A receptor identified from inducible G2A expression and beta-lactamase reporter screen.

    abstract::The G protein-coupled receptor (GPCR) G2A (for G2 accumulation) was identified as a stress-inducible antiproliferative cell cycle regulator. Targeted G2A gene deletion in mice resulted in systemic lupus erythematosus-like and atherosclerotic lesion phenotypes. These findings suggested that G2A may be a therapeutic tar...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Bercher M,Hanson B,van Staden C,Wu K,Ng GY,Lee PH

    更新日期:2009-04-01 00:00:00

  • In Vitro Evaluation of Small Molecule Delivery into Articular Cartilage: Effect of Synovial Clearance and Compressive Load.

    abstract::Intra-articular injection of drug depots is considered as a therapeutic strategy for the treatment of osteoarthritis. In this study, we designed an in vitro assay in a previously described bioreactor system to evaluate the uptake of a small molecule drug mimic as a function of drug clearance by the synovium and compre...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Spitters TWGM,Stamatialis D,Petit A,Leeuw MGW,Karperien M

    更新日期:2019-05-01 00:00:00

  • Steroid hormone receptors and drug discovery: therapeutic opportunities and assay designs.

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    journal_title:Assay and drug development technologies

    pub_type: 杂志文章,评审


    authors: Bai C,Schmidt A,Freedman LP

    更新日期:2003-12-01 00:00:00

  • Digital movie analysis for quantification of beating frequencies, chronotropic effects, and beating areas in cardiomyocyte cultures.

    abstract::Software, named Cardio Analyser, was developed for digital movie analysis of beating frequencies, drug-induced chronotropic effects, and quantification of beating areas of contracting cardiomyocyte cultures. A major novelty of the software is the introduction of automated noise filtering and automated movie analysis o...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Stummann TC,Wronski M,Sobanski T,Kumpfmueller B,Hareng L,Bremer S,Whelan MP

    更新日期:2008-06-01 00:00:00

  • Reconfiguring the AR-TIF2 Protein-Protein Interaction HCS Assay in Prostate Cancer Cells and Characterizing the Hits from a LOPAC Screen.

    abstract::The continued activation of androgen receptor (AR) transcription and elevated expression of AR and transcriptional intermediary factor 2 (TIF2) coactivator observed in prostate cancer (CaP) recurrence and the development of castration-resistant CaP (CRPC) support a screening strategy for small-molecule inhibitors of A...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Fancher AT,Hua Y,Camarco DP,Close DA,Strock CJ,Johnston PA

    更新日期:2016-10-01 00:00:00

  • Expression and purification of recombinant human apolipoprotein A-II in Pichia pastoris.

    abstract::Apolipoprotein A-II (ApoA-II) is the second most abundant protein constituent of high-density lipoprotein (HDL). The physiologic role of ApoA-II is poorly defined. ApoA-II may inhibit lecithin:cholesterol acyltransferase and cholesteryl-ester-transfer protein activities, but may increase the hepatic lipase activity. A...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Su M,Qi Y,Wang M,Chang W,Peng S,Xu T,Wang D

    更新日期:2013-10-01 00:00:00

  • Development of a protein biochip to identify 6 monoclonal antibodies against subtypes of recombinant human interferons.

    abstract::Recombinant human interferons (rhIFNs) are broadly used as effective therapeutic agents with antiviral, antitumor, and immune-modulating properties. Advances in protein biochip technology have benefited the medical community greatly, making true parallelism, miniaturization, and high throughput possible. In this study...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Xu Z,Du W,Zhang P,Wang X,Ma X,Shi L,Song L

    更新日期:2010-04-01 00:00:00

  • A beta-lactamase reporter assay for monitoring the activation of the smoothened pathway.

    abstract::The seven-transmembrane protein Smoothened (Smo) mediates the cellular response to the Hedgehog protein signal and is involved in cell growth and differentiation during embryonic development. Stimulation of the Smo pathway is directly implicated in tissue maintenance and repair, but overactivation of Smo could lead to...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Osak B,Fitzgerald LR

    更新日期:2007-12-01 00:00:00

  • Analysis of purified tubulin in high concentration of glutamate for application in high throughput screening for microtubule-stabilizing agents.

    abstract::High-throughput screening (HTS) is a powerful approach for the discovery of potential and effective drugs from a number of candidates. The dynamics of tubulin assembly and disassembly in vitro are an important target for the screening of anti-tumor agents. However, previously described methods are not amenable for HTS...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Dong H,Li YZ,Hu W

    更新日期:2004-12-01 00:00:00

  • Development and validation of HTS flux assay for endogenously expressed chloride channels in a CHO-K1 cell line.

    abstract::An atomic absorption spectroscopy-based detection system was employed to develop a new non-radioactive flux assay for chloride (Cl-) channels in a high throughput format. Cl- flux is assayed by measuring the extent to which Cl- precipitates an excess amount of silver ions (Ag+). A linear correlation was observed betwe...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Gill S,Gill R,Xie Y,Wicks D,Liang D

    更新日期:2006-02-01 00:00:00

  • High-throughput flow cytometry to detect selective inhibitors of ABCB1, ABCC1, and ABCG2 transporters.

    abstract::Up-regulation of pump (transporter) expression and selection of resistant cancer cells result in cancer multidrug resistance to diverse substrates of these transporters. While more than 48 members of the ATP binding cassette (ABC) transporter superfamily have been identified, up to now only three human ABC transporter...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Ivnitski-Steele I,Larson RS,Lovato DM,Khawaja HM,Winter SS,Oprea TI,Sklar LA,Edwards BS

    更新日期:2008-04-01 00:00:00

  • A flexible multiwell format for immunofluorescence screening microscopy of small-molecule inhibitors.

    abstract::Large-scale screens in mammalian cells demand for flexible high-throughput screening platforms that allow to analyze cellular traits on a genome-wide level or to identify small-molecule inhibitors (SMIs) from complex compound libraries. In this study we developed and tested high-density cell arrays made out of polydim...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Scholz AK,Klebl BM,Morkel M,Lehrach H,Dahl A,Lange BM

    更新日期:2010-10-01 00:00:00

  • Drug Discovery Approaches Utilizing Three-Dimensional Cell Culture.

    abstract::Historically, two-dimensional (2D) cell culture has been the preferred method of producing disease models in vitro. Recently, there has been a move away from 2D culture in favor of generating three-dimensional (3D) multicellular structures, which are thought to be more representative of the in vivo environment. This t...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章,评审


    authors: Ryan SL,Baird AM,Vaz G,Urquhart AJ,Senge M,Richard DJ,O'Byrne KJ,Davies AM

    更新日期:2016-01-01 00:00:00

  • Dynamic mass redistribution as a means to measure and differentiate signaling via opioid and cannabinoid receptors.

    abstract::Classically, G protein-coupled receptor activation by a ligand has been viewed as producing a defined response such as activation of a G protein, activation or inhibition of adenylyl cyclase, or stimulation of phospholipase C and/or alteration in calcium flux. Newer concepts of ligand-directed signaling recognize that...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Codd EE,Mabus JR,Murray BS,Zhang SP,Flores CM

    更新日期:2011-08-01 00:00:00

  • Examining Ligand-Based Stabilization of Proteins in Cells with MEK1 Kinase Inhibitors.

    abstract::In this study, we describe the evaluation of a cell-based protein stability assay using β-galactosidase fragment complementation technology performed in two independent laboratories. The assay is based on the ability of certain ligands to bind to a protein leading to a ligand-protein complex that has a different stabi...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Auld DS,Davis CA,Jimenez M,Knight S,Orme JP

    更新日期:2015-06-01 00:00:00

  • Use of multiple assay endpoints to investigate the effects of incubation time, dose of toxin, and plating density in cell-based cytotoxicity assays.

    abstract::Here we show the results of comparing cell viability, cytotoxicity, and apoptosis assays for measuring the time- and dose-dependent toxic effects of tamoxifen on HepG2 cells. The quantitation of adenosine 5'-triphosphate (ATP), 5-(3-carboxymethoxyphenyl)-2-(4,5- dimethylthiazolyl)-3-(4-sulfophenyl) tetrazolium, inner ...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Riss TL,Moravec RA

    更新日期:2004-02-01 00:00:00

  • Combination of Resveratrol and Quercetin Causes Cell Growth Inhibition, DNA Damage, Cell Cycle Arrest, and Apoptosis in Oral Cancer Cells.

    abstract:: Resveratrol and quercetin alone are well reported to have anticancer potential, but their combination studies are very inadequate. We have examined their combination in Cal-33 and SCC-15 oral cancer cells (OCCs) and noncancerous HEK-293 cells. Combination of 10 μM concentration of each resveratrol...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Singh V,Singh R,Kujur PK,Singh RP

    更新日期:2020-07-01 00:00:00

  • Characterization of an in vitro differentiation assay for pancreatic-like cell development from murine embryonic stem cells: detailed gene expression analysis.

    abstract::Embryonic stem (ES) cell technology may serve as a platform for the discovery of drugs to treat diseases such as diabetes. However, because of difficulties in establishing reliable ES cell differentiation methods and in creating cost-effective plating conditions for the high-throughput format, screening for molecules ...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Chen C,Chai J,Singh L,Kuo CY,Jin L,Feng T,Marzano S,Galeni S,Zhang N,Iacovino M,Qin L,Hara M,Stein R,Bromberg JS,Kyba M,Ku HT

    更新日期:2011-08-01 00:00:00

  • CypHer 5: a generic approach for measuring the activation and trafficking of G protein-coupled receptors in live cells.

    abstract::GPCRs are one of the most popular classes of therapeutic drug targets. It is therefore important to design specific assay formats to readily identify ligands at these receptors. CypHer 5 technology utilizes the general ability of GPCRs to be internalized into the endosomal pathway of a cell in response to agonist liga...

    journal_title:Assay and drug development technologies

    pub_type: 杂志文章


    authors: Adie EJ,Francis MJ,Davies J,Smith L,Marenghi A,Hather C,Hadingham K,Michael NP,Milligan G,Game S

    更新日期:2003-04-01 00:00:00