scholarly journals High-Content Screening: A New Approach to Easing Key Bottlenecks in the Drug Discovery Process

1997 ◽  
Vol 2 (4) ◽  
pp. 249-259 ◽  
Author(s):  
Kenneth A. Giuliano ◽  
Robbin L. DeBiasio ◽  
R. Terry Dunlay ◽  
Albert Gough ◽  
Joanne M. Volosky ◽  
...  
Keyword(s):  
Drug Discovery ◽  
High Throughput Screening ◽  
Nuclear Dna ◽  
Spatial Information ◽  
Fluorescent Protein ◽  
Human Tumor ◽  
Nuclear Dna Content ◽  
High Content Screening ◽  
Drug Induced ◽  
Cell Functions

Recent improvements in target discovery and high throughput screening (HTS) have increased the pressure at key points along the drug discovery pipeline. High-content screening (HCS) was developed to ease bottlenecks that have formed at target validation and lead optimization points in the pipeline. HCS defines the role of targets in cell functions by combining fluorescence-based reagents with the ArrayScan™ System to automatically extract temporal and spatial information about target activities within cells. The ArrayScan System is a tabletop instrument that includes optics for subcellular resolution of fluorescence signals from many cells in a field within a well of a microtiter plate. One demonstrated application is a high-content screen designed to measure the drug-induced transport of a green fluorescent protein-human glucocorticoid receptor chimeric protein from the cytoplasm to the nucleus of human tumor cells. A high-content screen is also described for the multiparametric measurement of apoptosis. This single screen provides measurements of nuclear size and shape changes, nuclear DNA content, mitochondrial potential, and actin-cytoskeletal rearrangements during drug-induced programmed cell death. The next generation HCS system is a miniaturized screening platform, the CellChip™ System, that will increase the throughput of HCS, while integrating HCS with HTS on the same platform.

scholarly journals Development and pilot screen of novel high content assay for down regulators of expression of heterogenous nuclear ribonuclear protein H2

2020 ◽  
Author(s):  
Juan Diez ◽  
Sumitha Rajendrarao ◽  
Shadi A. Baajour ◽  
Praathibha Sripadhan ◽  
Timothy P. Spicer ◽  
...  
Keyword(s):  
Metastatic Melanoma ◽  
High Throughput Screening ◽  
Large Scale ◽  
Fluorescent Protein ◽  
High Content Screening ◽  
Screening Assay ◽  
Down Regulation ◽  
Metastatic Melanoma Cell ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Nuclear Ribonucleoprotein

ABSTRACTDespite recent advances in melanoma drug discovery, the average overall survival of patients with late stage metastatic melanoma is approximately 3 years, suggesting a need for new approaches and melanoma therapeutic targets. Previously we identified heterogeneous nuclear ribonucleoprotein H2 as a potential target of anti-melanoma compound 2155-14 (Palrasu et al, Cell Physiol Biochem 2019;53:656-86). In the present study, we endeavored to develop an assay to enable a high throughput screening campaign to identify drug-like molecules acting via down regulation of heterogeneous nuclear ribonucleoprotein H that can be used for melanoma therapy and research.ResultsWe established a cell-based platform using metastatic melanoma cell line WM266-4 expressing hnRNPH2 conjugated with green fluorescent protein to enable assay development and screening. High Content Screening assay was developed and validated in 384 well plate format, followed by miniaturization to 1,536 well plate format. All plate-based QC parameters were acceptable: %CV = 6.7±0.3, S/B = 21±2.1, Z’ = 0.75±0.04. Pilot screen of FDA-approved drug library (n=1,400 compounds) demonstrated hit rate of 0.5%. Two compounds demonstrated pharmacological response and were authenticated by western blot analysis.ConclusionsWe developed a highly robust HTS-amenable high content screening assay capable of monitoring down regulation of hnRNPH2. This assay is thus capable of identifying authentic down regulators of hnRNPH1 and 2 in a large compound collection and, therefore, is amenable to a large-scale screening effort.

A novel multi-parametric high content screening assay in ciPTEC-OAT1 to predict drug-induced nephrotoxicity during drug discovery

2018 ◽  
Vol 92 (10) ◽  
pp. 3175-3190 ◽  
Author(s):  
Anna-Karin Sjögren ◽  
Katarina Breitholtz ◽  
Ernst Ahlberg ◽  
Lucas Milton ◽  
Malin Forsgard ◽  
...  
Keyword(s):  
Drug Discovery ◽  
High Content Screening ◽  
Screening Assay ◽  
Drug Induced

scholarly journals Flow cytometry may allow microscope-independent detection of holocentric chromosomes in plants

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
František Zedek ◽  
Pavel Veselý ◽  
Lucie Horová ◽  
Petr Bureš
Keyword(s):  
Flow Cytometry ◽  
Dna Content ◽  
High Throughput Screening ◽  
Nuclear Dna ◽  
Nuclear Dna Content ◽  
Holocentric Chromosomes ◽  
Induce Cell Cycle Arrest ◽  
Unequal Distribution ◽  
Plant Phylogeny ◽  
Radiation Induced

Abstract Two chromosomal structures, known as monocentric and holocentric chromosomes, have evolved in eukaryotes. Acentric fragments of monocentric chromosomes are unequally distributed to daughter cells and/or lost, while holocentric fragments are inherited normally. In monocentric species, unequal distribution should generate chimeras of cells with different nuclear DNA content. We investigated whether such differences in monocentric species are detectable by flow cytometry (FCM) as (i) a decreased nuclear DNA content and (ii) an increased coefficient of variance (CV) of the G1 peak after gamma radiation-induced fragmentation. We compared 13 monocentric and 9 holocentric plant species. Unexpectedly, monocentrics and holocentrics did not differ with respect to parameters (i) and (ii) in their response to gamma irradiation. However, we found that the proportion of G2 nuclei was highly elevated in monocentrics after irradiation, while holocentrics were negligibly affected. Therefore, we hypothesize that DNA-damaging agents induce cell cycle arrest leading to endopolyploidy only in monocentric and not (or to much lesser extent) in holocentric plants. While current microscope-dependent methods for holocentrism detection are unreliable for small and numerous chromosomes, which are common in holocentrics, FCM can use somatic nuclei. Thus, FCM may be a rapid and reliable method of high-throughput screening for holocentric candidates across plant phylogeny.

scholarly journals Development and Pilot Screen of Novel High Content Assay for Down Regulators of Expression of Heterogenous Nuclear Ribonuclear Protein H2

2021 ◽  
Vol 55 (3) ◽  
pp. 265-276
Keyword(s):  
Metastatic Melanoma ◽  
High Throughput Screening ◽  
Large Scale ◽  
Fluorescent Protein ◽  
High Content Screening ◽  
Screening Assay ◽  
Down Regulation ◽  
Metastatic Melanoma Cell ◽  
Heterogeneous Nuclear Ribonucleoprotein ◽  
Nuclear Ribonucleoprotein

BACKGROUND/AIMS: Despite recent advances in melanoma drug discovery, the average overall survival of patients with late-stage metastatic melanoma is approximately 3 years, suggesting a need for new approaches and melanoma therapeutic targets. Previously we identified heterogeneous nuclear ribonucleoprotein H2 as a potential target of anti-melanoma compound 2155-14 (Palrasu et al., Cell Physiol Biochem 2019;53:656-686). In the present study, we endeavored to develop an assay to enable a high throughput screening campaign to identify drug-like molecules acting via down regulation of heterogeneous nuclear ribonucleoprotein H2 that can be used for melanoma therapy and research. METHODS: We established a cell-based platform using metastatic melanoma cell line WM266-4 expressing hnRNPH2 conjugated with green fluorescent protein to enable assay development and screening. High Content Screening assay was developed and validated in 384 well plate format, followed by miniaturization to 1,536 well plate format. RESULTS: All plate-based QC parameters were acceptable: %CV = 6.7±0.3, S/B = 21±2.1, Z' = 0.75±0.04. Pilot screen of FDA-approved drug library (n=1,400 compounds) demonstrated hit rate of 0.5%. Two compounds demonstrated pharmacological response and were authenticated by western blot analysis. CONCLUSION: We developed a highly robust HTS-amenable high content screening assay capable of monitoring down regulation of hnRNPH2. This assay is thus capable of identifying authentic down regulators of hnRNPH1 and 2 in a large compound collection and, therefore, is amenable to a large-scale screening effort.

Upscaling and Automation of Electrophysiology: Toward High Throughput Screening in Ion Channel Drug Discovery

2003 ◽  
Vol 9 (1) ◽  
pp. 49-58
Author(s):  
Margit Asmild ◽  
Nicholas Oswald ◽  
Karen M. Krzywkowski ◽  
Søren Friis ◽  
Rasmus B. Jacobsen ◽  
...  
Keyword(s):  
Drug Discovery ◽  
Ion Channel ◽  
High Throughput ◽  
High Throughput Screening
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