A Perspective on Implementing a Quantitative Systems Pharmacology Platform for Drug Discovery and the Advancement of Personalized Medicine
Drug candidates exhibiting well-defined pharmacokinetic and pharmacodynamic profiles that are otherwise safe often fail to demonstrate proof-of-concept in phase II and III trials. Innovation in drug discovery and development has been identified as a critical need for improving the efficiency of drug discovery, especially through collaborations between academia, government agencies, and industry. To address the innovation challenge, we describe a comprehensive, unbiased, integrated, and iterative quantitative systems pharmacology (QSP)–driven drug discovery and development strategy and platform that we have implemente...
Source: Journal of Biomolecular Screening - June 20, 2016 Category: Molecular Biology Authors: Stern, A. M., Schurdak, M. E., Bahar, I., Berg, J. M., Taylor, D. L. Tags: Perspective Article Source Type: research
Characterization of Early Cortical Neural Network Development in Multiwell Microelectrode Array Plates
We examined neural network ontogeny using microelectrode array (MEA) recordings made in multiwell MEA (mwMEA) plates over the first 12 days in vitro (DIV). In primary cortical cultures, action potential spiking activity developed rapidly between DIV 5 and 12. Spiking was sporadic and unorganized at early DIV, and became progressively more organized with time, with bursting parameters, synchrony, and network bursting increasing between DIV 5 and 12. We selected 12 features to describe network activity; principal components analysis using these features demonstrated segregation of data by age at both the well and plate level...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Cotterill, E., Hall, D., Wallace, K., Mundy, W. R., Eglen, S. J., Shafer, T. J. Tags: Original Research Source Type: research
Characterization of Differentiated SH-SY5Y as Neuronal Screening Model Reveals Increased Oxidative Vulnerability
In this study, we present an optimized neuronal differentiation protocol for SH-SY5Y that requires only two work steps and 6 days. After differentiation, the cells present increased levels of ATP and plasma membrane activity but reduced expression of energetic stress response genes. Differentiation results in reduced mitochondrial membrane potential and decreased robustness toward perturbations with 6-hydroxydopamine. We are convinced that the presented differentiation method will leverage genetic and chemical high-throughput screening projects targeting pathways that are involved in the selective vulnerability of neurons ...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Forster, J. I., Köglsberger, S., Trefois, C., Boyd, O., Baumuratov, A. S., Buck, L., Balling, R., Antony, P. M. A. Tags: Original Research Source Type: research
Development of an HTRF Assay for the Detection and Characterization of Inhibitors of Catechol-O-Methyltransferase
Catechol-O-methyltransferase (COMT) plays an important role in the deactivation of catecholamine neurotransmitters and hormones. Inhibitors of COMT, such as tolcapone and entacapone, are used clinically in the treatment of Parkinson’s disease. Discovery of novel inhibitors has been hampered by a lack of suitable assays for high-throughput screening (HTS). Although assays using esculetin have been developed, these are affected by fluorescence, a common property of catechol-type compounds. We have therefore evaluated a new homogenous time-resolved fluorescence (HTRF)–based assay from CisBio (Codolet, France), whi...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Kimos, M., Burton, M., Urbain, D., Caudron, D., Martini, M., Famelart, M., Gillard, M., Barrow, J., Wood, M. Tags: Original Research Source Type: research
Kinetic Analysis of Membrane Potential Dye Response to NaV1.7 Channel Activation Identifies Antagonists with Pharmacological Selectivity against NaV1.5
The NaV1.7 voltage-gated sodium channel is a highly valued target for the treatment of neuropathic pain due to its expression in pain-sensing neurons and human genetic mutations in the gene encoding NaV1.7, resulting in either loss-of-function (e.g., congenital analgesia) or gain-of-function (e.g., paroxysmal extreme pain disorder) pain phenotypes. We exploited existing technologies in a novel manner to identify selective antagonists of NaV1.7. A full-deck high-throughput screen was developed for both NaV1.7 and cardiac NaV1.5 channels using a cell-based membrane potential dye FLIPR assay. In assay development, known local...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Finley, M., Cassaday, J., Kreamer, T., Li, X., Solly, K., ODonnell, G., Clements, M., Converso, A., Cook, S., Daley, C., Kraus, R., Lai, M.-T., Layton, M., Lemaire, W., Staas, D., Wang, J. Tags: Original Research Source Type: research
An Integrated Approach for Screening and Identification of Positive Allosteric Modulators of N-Methyl-D-Aspartate Receptors
We report a novel screening approach for identification and characterization of NMDAR-PAMs. The approach combines high-throughput fluorescence imaging with automated electrophysiological recording of glutamate-evoked responses in HEK-293 cells expressing NR1/NR2A NMDAR subunits. Initial high-throughput screening (HTS) of a chemical library containing >810,000 compounds using a calcium flux assay in 1536-well plate format identified a total of 864 NMDAR-PAMs. Concentration response determination in both calcium flux and automated electrophysiological assays found several novel chemical series with EC50 values between 0.4...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Jambrina, E., Cerne, R., Smith, E., Scampavia, L., Cuadrado, M., Findlay, J., Krambis, M. J., Wakulchik, M., Chase, P., Brunavs, M., Burris, K. D., Gallagher, P., Spicer, T. P., Ursu, D. Tags: Original Research Source Type: research
At-Line Cellular Screening Methodology for Bioactives in Mixtures Targeting the {alpha}7-Nicotinic Acetylcholine Receptor
This study describes screening methodology for identifying bioactive compounds in mixtures acting on the α7-nAChR. The methodology developed combines liquid chromatography (LC) coupled via a split with both an at-line calcium (Ca2+)-flux assay and high-resolution mass spectrometry (MS). This allows evaluation of α7-nAChR responses after LC separation, while parallel MS enables compound identification. The methodology was optimized for analysis of agonists and positive allosteric modulators, and was successfully applied to screening of the hallucinogen mushroom Psilocybe Mckennaii. The crude mushroom extract was...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Otvos, R. A., Mladic, M., Arias-Alpizar, G., Niessen, W. M. A., Somsen, G. W., Smit, A. B., Kool, J. Tags: Original Research Source Type: research
Discovery of Novel Inhibitors of the Tautomerase Activity of Macrophage Migration Inhibitory Factor (MIF)
Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine associated with multiple diseases, including neurodegenerative disorders. With the ultimate goal of providing novel chemotypes as starting points for development of disease-modifying therapeutics for neurodegeneration, we endeavored to screen the GSK compound collection for MIF inhibitors using a miniaturized, activity-based kinetic assay. The assay monitors the increase in absorbance at 320 nm resulting from keto-to-enol tautomerization of 4-hydroxyphenylpyruvate, a reaction catalyzed by MIF. We ran a full-diversity screen evaluating the inhibitory...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Zapatero, M. C., Perez, P., Vazquez, M. J., Colmenarejo, G., de los Frailes, M., Ramon, F. Tags: Original Research Source Type: research
Discovery of Enhancers of the Secretion of Leukemia Inhibitory Factor for the Treatment of Multiple Sclerosis
We describe here the development of a phenotypic assay and screening of 1.7 million compounds to identify LIF enhancers using U87 MG cells. Five chemically tractable series of compounds and a few singletons were selected for further progression. Some of them were also active in a different LIF-expressing cell line and in primary rat astrocytes. Although further studies would be required to deconvolute the targets involved in LIF induction and to confirm activity of hits in more disease-relevant assays, our results have demonstrated the potential of the phenotypic approach to identify specific and chemically tractable small...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Vela, L., Caballero, I., Fang, L., Liu, Q., Ramon, F., Diez, E., de los Frailes, M. Tags: Original Research Source Type: research
A KNIME-Based Analysis of the Zebrafish Photomotor Response Clusters the Phenotypes of 14 Classes of Neuroactive Molecules
In this study, we developed an automated analysis workflow using the KNIME Analytics Platform and made it freely accessible. This workflow allows us to simultaneously calculate a behavioral fingerprint for all analyzed compounds and to further process the data. Furthermore, to further characterize the potential of PMR for mechanism prediction, we performed PMR analysis of 767 neuroactive compounds covering 14 different receptor classes using the KNIME workflow. We observed a true positive rate of 25% and a false negative rate of 75% in our screening conditions. Among the true positives, all receptor classes were represente...
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Copmans, D., Meinl, T., Dietz, C., van Leeuwen, M., Ortmann, J., Berthold, M. R., de Witte, P. A. M. Tags: Original Research Source Type: research
JBS Special Issue: Innovative Screening Methodologies to Identify New Compounds for the Treatment of Central Nervous System Disorders
(Source: Journal of Biomolecular Screening)
Source: Journal of Biomolecular Screening - May 19, 2016 Category: Molecular Biology Authors: Burris, K. D., Dworetzky, S. I. Tags: Editorial Source Type: research
An Automated Microscale Thermophoresis Screening Approach for Fragment-Based Lead Discovery
Fragment-based lead discovery has proved to be an effective alternative to high-throughput screenings in identifying chemical matter that can be developed into robust lead compounds. The search for optimal combinations of biophysical techniques that can correctly and efficiently identify and quantify binding can be challenging due to the physicochemical properties of fragments. In order to minimize the time and costs of screening, optimal combinations of biophysical techniques with maximal information content, sensitivity, and robustness are needed. Here we describe an approach utilizing automated microscale thermophoresis...
Source: Journal of Biomolecular Screening - March 18, 2016 Category: Molecular Biology Authors: Linke, P., Amaning, K., Maschberger, M., Vallee, F., Steier, V., Baaske, P., Duhr, S., Breitsprecher, D., Rak, A. Tags: Technical Notes Source Type: research
A High-Throughput Assay for the Detection of {alpha}-Dystroglycan N-Terminus in Human Uterine Fluid to Determine Uterine Receptivity
In this study, we first validated three monoclonal antibodies raised against α-DG-N in our system, and then established a sandwich ELISA suitable for the detection of α-DG-N in human uterine fluid. This ELISA detected significantly higher concentrations of α-DG-N in uterine fluid of women in the receptive phase. We believe this newly established α-DG-N ELISA may provide an important tool in the development of noninvasive strategies to detect uterine receptivity in women. (Source: Journal of Biomolecular Screening)
Source: Journal of Biomolecular Screening - March 18, 2016 Category: Molecular Biology Authors: Heng, S., Vollenhoven, B., Rombauts, L. J., Nie, G. Tags: Technical Notes Source Type: research
Phenotypic Screening of Drug Library in Actively Differentiating Mouse Embryonic Stem Cells
Phenotypic screening enables the discovery of new drug leads with novel targets. ES cells differentiate into different lineages by successively making use of different subsets of the genome’s possible macromolecular interactions. If a compound effectively targets just one of these interactions, it derails the developmental pathway to produce a phenotypical change. The OTRADI microsource spectrum library of 2000 approved drug components, natural products, and bioactive components was screened for compounds that can induce phenotypic changes in ES cell cultures at 10 µM after 3 days. Twenty-one compounds that ind...
Source: Journal of Biomolecular Screening - March 18, 2016 Category: Molecular Biology Authors: Billard, B., Chang, C.-N., Singh, A. J., Gross, M. K., Allen, R., Kioussi, C. Tags: Technical Notes Source Type: research
An Efficient and Economical Assay to Screen for Triclosan Binding to FabI
Triclosan is an effective inhibitor for enoyl acyl carrier protein reductase (ENR) in fatty acid biosynthesis. Triclosan-resistant mutants of ENR have emerged. Thus, it is important to detect these triclosan-resistant mutations in ENR. Generally, enzyme activity assays on the mutants are used to determine the effect of triclosan on ENR activity. Since the substrates are linked to acyl carrier protein (ACP), the assays are challenging due to the need to prepare the ACP and link it to the substrates. Non-ACP-linked (coenzyme A [CoA]-linked) substrates can be used in some ENR, but not in all. Consequently, screening for tricl...
Source: Journal of Biomolecular Screening - March 18, 2016 Category: Molecular Biology Authors: Demissie, R. D., Kabre, P., Tuntland, M. L., Fung, L. W.- M. Tags: Original Research Source Type: research
