High-Throughput Screening: today’s biochemical and cell-based approaches

Blay, Vincent; Tolani, Bhairavi; Ho, Sunita P.; Arkin, Michelle R.

doi:10.1016/j.drudis.2020.07.024

Cited by 176 publications

(171 citation statements)

References 135 publications

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“…Therefore, it has become standard procedure to co-transfect a second constitutively active reporter plasmid to allow correction for such effects. Stable reporter lines have recently gained popularity, for example, in high-throughput screening procedures [ 7 ]. The generation of a stable reporter line requires a selectable marker to remove cells that did not incorporate the reporter.…”

Section: Introductionmentioning

confidence: 99%

Current Practice in Bicistronic IRES Reporter Use: A Systematic Review

Akker

Zacchini

Housmans

et al. 2021

IJMS

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Bicistronic reporter assays have been instrumental for transgene expression, understanding of internal ribosomal entry site (IRES) translation, and identification of novel cap-independent translational elements (CITE). We observed a large methodological variability in the use of bicistronic reporter assays and data presentation or normalization procedures. Therefore, we systematically searched the literature for bicistronic IRES reporter studies and analyzed methodological details, data visualization, and normalization procedures. Two hundred fifty-seven publications were identified using our search strategy (published 1994–2020). Experimental studies on eukaryotic adherent cell systems and the cell-free translation assay were included for further analysis. We evaluated the following methodological details for 176 full text articles: the bicistronic reporter design, the cell line or type, transfection methods, and time point of analyses post-transfection. For the cell-free translation assay, we focused on methods of in vitro transcription, type of translation lysate, and incubation times and assay temperature. Data can be presented in multiple ways: raw data from individual cistrons, a ratio of the two, or fold changes thereof. In addition, many different control experiments have been suggested when studying IRES-mediated translation. In addition, many different normalization and control experiments have been suggested when studying IRES-mediated translation. Therefore, we also categorized and summarized their use. Our unbiased analyses provide a representative overview of bicistronic IRES reporter use. We identified parameters that were reported inconsistently or incompletely, which could hamper data reproduction and interpretation. On the basis of our analyses, we encourage adhering to a number of practices that should improve transparency of bicistronic reporter data presentation and improve methodological descriptions to facilitate data replication.

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Section: Introductionmentioning

confidence: 99%

Current Practice in Bicistronic IRES Reporter Use: A Systematic Review

Akker

Zacchini

Housmans

et al. 2021

IJMS

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“…Public-domain cocrystal structures frequently provide useful precompetitive information concerning binding of tool compounds to potential drug discovery targets ( 56 ). Even more powerful are the many cocrystal structure studies carried out within biopharmaceutical companies that directly assess in 3D how small-molecule hits coming from biochemical or cell-based assays ( 57 ) or biophysical measurements ( 58 ) bind to would-be drug targets. In silico virtual screening exercises carried out computationally with millions of small molecules can also provide useful information regarding potential starting points for medicinal chemistry.…”

Section: Utility Of Pdb Structures For Small-molecule Drug Discovery and Developmentmentioning

confidence: 99%

Impact of structural biologists and the Protein Data Bank on small-molecule drug discovery and development

Burley

2021

Journal of Biological Chemistry

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The Protein Data Bank (PDB) is an international core data resource central to fundamental biology, biomedicine, bioenergy, and biotechnology/bioengineering. Now celebrating its 50th anniversary, the PDB houses >175,000 experimentally determined atomic structures of proteins, nucleic acids, and their complexes with one another and small molecules and drugs. The importance of three-dimensional (3D) biostructure information for research and education obtains from the intimate link between molecular form and function evident throughout biology. Among the most prolific consumers of PDB data are biomedical researchers, who rely on the open access resource as the authoritative source of well-validated, expertly curated biostructures. This review recounts how the PDB grew from just seven protein structures to contain more than 49,000 structures of human proteins that have proven critical for understanding their roles in human health and disease. It then describes how these structures are used in academe and industry to validate drug targets, assess target druggability, characterize how tool compounds and other small-molecules bind to drug targets, guide medicinal chemistry optimization of binding affinity and selectivity, and overcome challenges during preclinical drug development. Three case studies drawn from oncology exemplify how structural biologists and open access to PDB structures impacted recent regulatory approvals of antineoplastic drugs.

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“…In this regard, recent advances in artificial intelligence (AI) and neural network learning as well as in silico library screening with molecular docking hold the promise for increased screening efficiency. Several readouts may be used for HTS, including fluorescence labeling methods for specific molecular targets, reporter-gene luminescent assays for transcriptional activity, mass spectrometry analysis for proteomic changes, and cell-based phenotypic screens [ 67 ]. Another strategy to decrease the time from initial screening to drug development is using drug repurposing.…”

Section: Tools For Drug Discoverymentioning

confidence: 99%

Emerging hiPSC Models for Drug Discovery in Neurodegenerative Diseases

Trudler

Ghatak

Lipton

2021

IJMS

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Neurodegenerative diseases affect millions of people worldwide and are characterized by the chronic and progressive deterioration of neural function. Neurodegenerative diseases, such as Alzheimer’s disease (AD), Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS), and Huntington’s disease (HD), represent a huge social and economic burden due to increasing prevalence in our aging society, severity of symptoms, and lack of effective disease-modifying therapies. This lack of effective treatments is partly due to a lack of reliable models. Modeling neurodegenerative diseases is difficult because of poor access to human samples (restricted in general to postmortem tissue) and limited knowledge of disease mechanisms in a human context. Animal models play an instrumental role in understanding these diseases but fail to comprehensively represent the full extent of disease due to critical differences between humans and other mammals. The advent of human-induced pluripotent stem cell (hiPSC) technology presents an advantageous system that complements animal models of neurodegenerative diseases. Coupled with advances in gene-editing technologies, hiPSC-derived neural cells from patients and healthy donors now allow disease modeling using human samples that can be used for drug discovery.

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High-Throughput Screening: today’s biochemical and cell-based approaches

Cited by 176 publications

References 135 publications

Current Practice in Bicistronic IRES Reporter Use: A Systematic Review

Current Practice in Bicistronic IRES Reporter Use: A Systematic Review

Impact of structural biologists and the Protein Data Bank on small-molecule drug discovery and development

Emerging hiPSC Models for Drug Discovery in Neurodegenerative Diseases

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