scholarly journals High-throughput screening for amyloid-β binding natural small-molecules based on the combinational use of biolayer interferometry and UHPLC−DAD-Q/TOF-MS/MS

2021 ◽  
Author(s):  
Minsong Guo ◽  
Fengdan Zhu ◽  
Wenqiao Qiu ◽  
Gan Qiao ◽  
Betty Yuen-Kwan Law ◽  
...  
Keyword(s):  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Amyloid Β ◽  
Biolayer Interferometry ◽  
Tof Ms

scholarly journals High Throughput Screening at the Membrane Interface Reveals New Inhibitors of Amyloid-β

2019 ◽  
Author(s):  
Sarah J. Cox ◽  
Brian Lam ◽  
Ajay Prasad ◽  
Hannah A. Marietta ◽  
Nicholas V. Stander ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Cell Membrane ◽  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Amyloid Β ◽  
Amyloid Aggregation ◽  
Membrane Interface

Amyloid-β aggregation at the cell-membrane of neruonal cells is implicated as a source of toxicity for Alzheimer’s disease. Small molecules have been studied for their ability to supress amyloid aggregation and toxicity, but the presence of membranes negate their activity. Here, we have identified 5 small molecules that are active at the membrane interface.

scholarly journals A High Throughput Screening Assay for Anti-Mycobacterial Small Molecules Based on Adenylate Kinase Release as a Reporter of Cell Lysis

PLoS ONE ◽  
2015 ◽  
Vol 10 (6) ◽  
pp. e0129234 ◽  
Author(s):  
Lauren Forbes ◽  
Katherine Ebsworth-Mojica ◽  
Louis DiDone ◽  
Shao-Gang Li ◽  
Joel S. Freundlich ◽  
...  
Keyword(s):  
Small Molecules ◽  
High Throughput ◽  
Adenylate Kinase ◽  
High Throughput Screening ◽  
Cell Lysis ◽  
Screening Assay ◽  
High Throughput Screening Assay ◽  
Adenylate Kinase Release

In pursuit of the ideal antifungal agent for Candida infections: high-throughput screening of small molecules

2014 ◽  
Vol 19 (11) ◽  
pp. 1721-1730 ◽  
Author(s):  
Sarah S.W. Wong ◽  
Lakshman P. Samaranayake ◽  
Chaminda J. Seneviratne
Keyword(s):  
Small Molecules ◽  
High Throughput ◽  
Antifungal Agent ◽  
High Throughput Screening ◽  
Candida Infections ◽  
The Ideal

scholarly journals A Fluorescence-based Lymphocyte Assay Suitable for High-throughput Screening of Small Molecules

10.3791/55199 ◽  
2017 ◽  
Author(s):  
Ahmed Fouda ◽  
Mahasti Tahsini ◽  
Fatemeh Khodayarian ◽  
Fatimah Al-nafisah ◽  
Moutih Rafei
Keyword(s):  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening

scholarly journals Development of a Simple Assay Protocol for High-Throughput Screening of Mycobacterium tuberculosis Glutamine Synthetase for the Identification of Novel Inhibitors

2005 ◽  
Vol 10 (7) ◽  
pp. 725-729 ◽  
Author(s):  
Upasana Singh ◽  
Vinita Panchanadikar ◽  
Dhiman Sarkar
Keyword(s):  
Escherichia Coli ◽  
Mycobacterium Tuberculosis ◽  
Glutamine Synthetase ◽  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Coli Strain ◽  
Compound Library ◽  
Essential Enzyme ◽  
Assay Protocol

Mycobacterium tuberculosis glutamine synthetase (GS) is an essential enzyme involved in the pathogenicity of the organism. The screening of a compound library using a robust high-throughput screening (HTS) assay is currently thought to be the most efficient way of getting lead molecules, which are potent inhibitors for this enzyme. The authors have purified the enzyme to a >90% level from the recombinant Escherichia coli strain YMC21E, and it was used for partial characterization as well as standardization experiments. The results indicated that the Kmof the enzyme for L-glutamine and hydroxylamine were 60 mM and 8.3 mM, respectively. The Km for ADP, arsenate, and Mn2+ were 2 [.proportional]M, 5 [.proportional]M, and 25 [.proportional]M, respectively. When the components were adjusted according to their Km values, the activity remained constant for at least 3 h at both 25° C and 37° C. The Z′ factor determined in microplate format indicated robustness of the assay. When the signal/noise ratios were determined for different assay volumes, it was observed that the 200-[.proportional]l volume was found to be optimum. The DMSO tolerance of the enzyme was checked up to 10%, with minimal inhibition. The IC50 value determined for L-methionine S-sulfoximine on the enzyme activity was 3 mM. Approximately 18,000 small molecules could be screened per day using this protocol by a Beckman Coulter HTS setup.

scholarly journals High-Throughput Screening for Drugs That Inhibit Papain-Like Protease in SARS-CoV-2

2020 ◽  
Vol 25 (10) ◽  
pp. 1152-1161
Author(s):  
Emery Smith ◽  
Meredith E. Davis-Gardner ◽  
Ruben D. Garcia-Ordonez ◽  
Tu-Trinh Nguyen ◽  
Mitchell Hull ◽  
...  
Keyword(s):  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Clinical Stage ◽  
Severe Pneumonia ◽  
Selective Inhibition ◽  
Peptide Cleavage ◽  
Global Pandemic ◽  
Drug Registration ◽  
Medical Countermeasures

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in late 2019 has triggered an ongoing global pandemic whereby infection may result in a lethal severe pneumonia-like disease designated as coronavirus disease 2019 (COVID-19). To date, millions of confirmed cases and hundreds of thousands of deaths have been reported worldwide, and there are currently no medical countermeasures available to prevent or treat the disease. The purported development of a vaccine could require at least 1–4 years, while the typical timeline from hit finding to drug registration of an antiviral is >10 years. Thus, repositioning of known drugs can significantly accelerate the development and deployment of therapies for COVID-19. To identify therapeutics that can be repurposed as SARS-CoV-2 antivirals, we developed and initiated a high-throughput cell-based screen that incorporates the essential viral papain-like protease (PLpro) and its peptide cleavage site into a luciferase complementation assay to evaluate the efficacy of known drugs encompassing approximately 15,000 clinical-stage or US Food and Drug Administration (FDA)-approved small molecules. Confirmed inhibitors were also tested to determine their cytotoxic properties. Here, we report the identification of four clinically relevant drugs that exhibit selective inhibition of the SARS-CoV-2 viral PLpro.

scholarly journals A Novel High-Throughput Screening Platform Identifies Itaconate Derivatives from Marine Penicillium antarcticum as Inhibitors of Mesenchymal Stem Cell Differentiation

Marine Drugs ◽  
2020 ◽  
Vol 18 (4) ◽  
pp. 192
Author(s):  
Pietro Marchese ◽  
Nipun Mahajan ◽  
Enda O’Connell ◽  
Howard Fearnhead ◽  
Maria Tuohy ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Chondrogenic Differentiation ◽  
Stem Cell Differentiation ◽  
Ectopic Calcification ◽  
Tissue Calcification ◽  
Screening Platform

Worldwide diffused diseases such as osteoarthritis, atherosclerosis or chronic kidney disease are associated with a tissue calcification process which may involve unexpected local stem cell differentiation. Current pharmacological treatments for such musculoskeletal conditions are weakly effective, sometimes extremely expensive and often absent. The potential to develop new therapies is represented by the discovery of small molecules modulating resident progenitor cell differentiation to prevent aberrant tissue calcification. The marine environment is a rich reserve of compounds with pharmaceutical potential and many novel molecules are isolated from macro and microorganisms annually. The potential of small molecules synthetized by marine filamentous fungi to influence the osteogenic and chondrogenic differentiation of human mesenchymal stem/stromal cells (hMSCs) was investigated using a novel, high-throughput automated screening platform. Metabolites synthetized by the marine-derived fungus Penicillium antarcticum were evaluated on the platform. Itaconic acid derivatives were identified as inhibitors of calcium elaboration into the matrix of osteogenically differentiated hMSCs and also inhibited hMSC chondrogenic differentiation, highlighting their capacity to impair ectopic calcification. Bioactive small molecule discovery is critical to address ectopic tissue calcification and the use of biologically relevant assays to identify naturally occurring metabolites from marine sources represents a strategy that can contribute to this effort.

scholarly journals An Ultra-High Throughput Cell-Based Screen for Wee1 Degradation Inhibitors

2010 ◽  
Vol 15 (8) ◽  
pp. 907-917 ◽  
Author(s):  
Franck Madoux ◽  
Scott Simanski ◽  
Peter Chase ◽  
Jitendra K. Mishra ◽  
William R. Roush ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Fusion Protein ◽  
Small Molecules ◽  
High Throughput ◽  
High Throughput Screening ◽  
Cell Cycle Progression ◽  
Ubiquitin Proteasome Pathway ◽  
Subsequent Degradation ◽  
Ubiquitin Proteasome ◽  
Proteasome Pathway

The tyrosine kinase Wee1 is part of a key cellular sensing mechanism that signals completion of DNA replication, ensuring proper timing of entry into mitosis. Wee1 acts as an inhibitor of mitotic entry by phosphorylating cyclin-dependent kinase CDK1. Wee1 activity is mainly regulated at the protein level through its phosphorylation and subsequent degradation by the ubiquitin proteasome pathway. To facilitate identification of small molecules preventing Wee1 degradation, a homogeneous cell-based assay was developed using HeLa cells transiently transfected with a Wee1-luciferase fusion protein. To ensure ultra-high-throughput screening (uHTS) compatibility, the assay was scaled to a 1536-well plate format and cells were transfected in bulk and cryopreserved. This miniaturized homogeneous assay demonstrated robust performance, with a calculated Z′ factor of 0.65 ± 0.05. The assay was screened against a publicly available library of ~218,000 compounds to identify Wee1 stabilizers. Nonselective, cytotoxic, and promiscuous compounds were rapidly triaged through the use of a similarly formatted counterscreen that measured stabilization of an N-cyclin B-luciferase fusion protein, as well as execution of viability assessment in the parental HeLa cell line. This screening campaign led to the discovery of 4 unrelated cell-permeable small molecules that showed selective Wee1-luciferase stabilization with micromolar potency. One of these compounds, SID4243143 (ML 118), was shown to inhibit cell cycle progression, underscoring the importance of Wee1 degradation to the cell cycle. Results suggest that this uHTS approach is suitable for identifying selective chemical probes that prevent Wee1 degradation and generally applicable to discovering inhibitors of the ubiquitin proteasome pathway.

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