scholarly journals Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of nsp14/nsp10 exoribonuclease

2021 ◽  
Vol 478 (13) ◽  
pp. 2445-2464
Author(s):  
Berta Canal ◽  
Allison W. McClure ◽  
Joseph F. Curran ◽  
Mary Wu ◽  
Rachel Ulferts ◽  
...  
Keyword(s):  
Antiviral Treatment ◽  
Genome Replication ◽  
Economic Costs ◽  
Cell Culture Model ◽  
Antiviral Compounds ◽  
Biochemical Assays ◽  
Culture Model ◽  
Viral Genome Replication ◽  
Great Pressure

SARS-CoV-2 is a coronavirus that emerged in 2019 and rapidly spread across the world causing a deadly pandemic with tremendous social and economic costs. Healthcare systems worldwide are under great pressure, and there is an urgent need for effective antiviral treatments. The only currently approved antiviral treatment for COVID-19 is remdesivir, an inhibitor of viral genome replication. SARS-CoV-2 proliferation relies on the enzymatic activities of the non-structural proteins (nsp), which makes them interesting targets for the development of new antiviral treatments. With the aim to identify novel SARS-CoV-2 antivirals, we have purified the exoribonuclease/methyltransferase (nsp14) and its cofactor (nsp10) and developed biochemical assays compatible with high-throughput approaches to screen for exoribonuclease inhibitors. We have screened a library of over 5000 commercial compounds and identified patulin and aurintricarboxylic acid (ATA) as inhibitors of nsp14 exoribonuclease in vitro. We found that patulin and ATA inhibit replication of SARS-CoV-2 in a VERO E6 cell-culture model. These two new antiviral compounds will be valuable tools for further coronavirus research as well as potentially contributing to new therapeutic opportunities for COVID-19.

scholarly journals Identifying SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of Nsp14/nsp10 Exoribonuclease

2021 ◽  
Author(s):  
Clovis Basier ◽  
Souradeep Basu ◽  
Rupert Beale ◽  
Agustina P Bertolin ◽  
Berta Canal ◽  
...  
Keyword(s):  
Antiviral Treatment ◽  
Genome Replication ◽  
Economic Costs ◽  
Cell Culture Model ◽  
Antiviral Compounds ◽  
Biochemical Assays ◽  
Culture Model ◽  
Viral Genome Replication ◽  
Great Pressure

SARS-CoV-2 is a coronavirus that emerged in 2019 and rapidly spread across the world causing a deadly pandemic with tremendous social and economic costs. Healthcare systems worldwide are under great pressure, and there is urgent need for effective antiviral treatments. The only currently approved antiviral treatment for COVID-19 is remdesivir, an inhibitor of viral genome replication. SARS-CoV-2 proliferation relies on the enzymatic activities of the non-structural proteins (nsp), which makes them interesting targets for the development of new antiviral treatments. With the aim to identify novel SARS-CoV-2 antivirals, we have purified the exoribonuclease/methyltransferase (nsp14) and its cofactor (nsp10) and developed biochemical assays compatible with high-throughput approaches to screen for exoribonuclease inhibitors. We have screened a library of over 5000 commercial compounds and identified patulin and aurintricarboxylic acid (ATA) as inhibitors of nsp14 exoribonuclease in vitro. We found that patulin and ATA inhibit replication of SARS-CoV-2 in a VERO E6 cell-culture model. These two new antiviral compounds will be valuable tools for further coronavirus research as well as potentially contributing to new therapeutic opportunities for COVID-19.

scholarly journals Identifying SARS-CoV-2 Antiviral Compounds by Screening for Small Molecule Inhibitors of Nsp15 Endoribonuclease

2021 ◽  
Author(s):  
Rupert Beale ◽  
Agustina P Bertolin ◽  
Berta Canal ◽  
Tom D Deegan ◽  
John FX Diffley ◽  
...  
Keyword(s):  
Immune Evasion ◽  
Antiviral Drug ◽  
Genome Replication ◽  
Chemical Library ◽  
Antiviral Compounds ◽  
Biochemical Assays ◽  
Viral Genome Replication ◽  
Affect Disease Severity ◽  
Endoribonuclease Activity

SARS-CoV-2 is responsible for COVID-19, a human disease that has caused over 2 million deaths, stretched health systems to near-breaking point and endangered the economies of countries and families around the world. Antiviral treatments to combat COVID-19 are currently lacking. Remdesivir, the only antiviral drug approved for the treatment of COVID-19, can affect disease severity, but better treatments are needed. SARS-CoV-2 encodes 16 non-structural proteins (nsp) that possess different enzymatic activities with important roles in viral genome replication, transcription and host immune evasion. One key aspect of host immune evasion is performed by the uridine-directed endoribonuclease activity of nsp15. Here we describe the expression and purification of nsp15 recombinant protein. We have developed biochemical assays to follow its activity, and we have found evidence for allosteric behaviour. We screened a custom chemical library of over 5000 compounds to identify nsp15 endoribonuclease inhibitors, and we identified and validated NSC95397 as an inhibitor of nsp15 endoribonuclease in vitro. Although NSC95397 did not inhibit SARS-CoV-2 growth in VERO E6 cells, further studies will be required to determine the effect of nsp15 inhibition on host immune evasion.

scholarly journals Identifying SARS-CoV-2 antiviral compounds by screening for small molecule inhibitors of nsp15 endoribonuclease

2021 ◽  
Vol 478 (13) ◽  
pp. 2465-2479 ◽  
Author(s):  
Berta Canal ◽  
Ryo Fujisawa ◽  
Allison W. McClure ◽  
Tom D. Deegan ◽  
Mary Wu ◽  
...  
Keyword(s):  
Immune Evasion ◽  
Antiviral Drug ◽  
Genome Replication ◽  
Chemical Library ◽  
Antiviral Compounds ◽  
Biochemical Assays ◽  
Viral Genome Replication ◽  
Affect Disease Severity ◽  
Endoribonuclease Activity

SARS-CoV-2 is responsible for COVID-19, a human disease that has caused over 2 million deaths, stretched health systems to near-breaking point and endangered economies of countries and families around the world. Antiviral treatments to combat COVID-19 are currently lacking. Remdesivir, the only antiviral drug approved for the treatment of COVID-19, can affect disease severity, but better treatments are needed. SARS-CoV-2 encodes 16 non-structural proteins (nsp) that possess different enzymatic activities with important roles in viral genome replication, transcription and host immune evasion. One key aspect of host immune evasion is performed by the uridine-directed endoribonuclease activity of nsp15. Here we describe the expression and purification of nsp15 recombinant protein. We have developed biochemical assays to follow its activity, and we have found evidence for allosteric behaviour. We screened a custom chemical library of over 5000 compounds to identify nsp15 endoribonuclease inhibitors, and we identified and validated NSC95397 as an inhibitor of nsp15 endoribonuclease in vitro. Although NSC95397 did not inhibit SARS-CoV-2 growth in VERO E6 cells, further studies will be required to determine the effect of nsp15 inhibition on host immune evasion.

1120: Citrate and Vitamin E Blunt the SWL-Induced Free Radical Surge in an In-Vitro MDCK Cell Culture Model

2004 ◽  
Vol 171 (4S) ◽  
pp. 295-295
Author(s):  
Fernando C. Delvecchio ◽  
Ricardo M. Brizuela ◽  
Karen J. Byer ◽  
W. Patrick Springhart ◽  
Saeed R. Khan ◽  
...  
Keyword(s):  
Cell Culture ◽  
Free Radical ◽  
Vitamin E ◽  
Mdck Cell ◽  
Cell Culture Model ◽  
Culture Model

scholarly journals An In Vitro Cell Culture Model for Pyoverdine-Mediated Virulence

Pathogens ◽  
2020 ◽  
Vol 10 (1) ◽  
pp. 9
Author(s):  
Donghoon Kang ◽  
Natalia V. Kirienko
Keyword(s):  
Cell Culture ◽  
Virulence Factors ◽  
Model Organisms ◽  
Cell Culture Model ◽  
Chronic Infections ◽  
In Vitro Cell Culture ◽  
Mitochondrial Homeostasis ◽  
Culture Model ◽  
Wide Range

Pseudomonas aeruginosa is a multidrug-resistant, opportunistic pathogen that utilizes a wide-range of virulence factors to cause acute, life-threatening infections in immunocompromised patients, especially those in intensive care units. It also causes debilitating chronic infections that shorten lives and worsen the quality of life for cystic fibrosis patients. One of the key virulence factors in P. aeruginosa is the siderophore pyoverdine, which provides the pathogen with iron during infection, regulates the production of secreted toxins, and disrupts host iron and mitochondrial homeostasis. These roles have been characterized in model organisms such as Caenorhabditis elegans and mice. However, an intermediary system, using cell culture to investigate the activity of this siderophore has been absent. In this report, we describe such a system, using murine macrophages treated with pyoverdine. We demonstrate that pyoverdine-rich filtrates from P. aeruginosa exhibit substantial cytotoxicity, and that the inhibition of pyoverdine production (genetic or chemical) is sufficient to mitigate virulence. Furthermore, consistent with previous observations made in C. elegans, pyoverdine translocates into cells and disrupts host mitochondrial homeostasis. Most importantly, we observe a strong correlation between pyoverdine production and virulence in P. aeruginosa clinical isolates, confirming pyoverdine’s value as a promising target for therapeutic intervention. This in vitro cell culture model will allow rapid validation of pyoverdine antivirulents in a simple but physiologically relevant manner.

scholarly journals Development of an in vitro cell culture model to study milk to plasma ratios of therapeutic drugs

2013 ◽  
Vol 45 (4) ◽  
pp. 325 ◽  
Author(s):  
Anurupa Maitra ◽  
Shahnaz Patel ◽  
VijayR Bhate ◽  
VilliS Toddywalla ◽  
MaithiliA Athavale
Keyword(s):  
Cell Culture ◽  
Cell Culture Model ◽  
In Vitro Cell Culture ◽  
Therapeutic Drugs ◽  
Culture Model

scholarly journals Effects of Ethanol and Acetaldehyde on Tight Junction Integrity: In Vitro Study in a Three Dimensional Intestinal Epithelial Cell Culture Model

PLoS ONE ◽  
2012 ◽  
Vol 7 (4) ◽  
pp. e35008 ◽  
Author(s):  
Elhaseen Elamin ◽  
Daisy Jonkers ◽  
Kati Juuti-Uusitalo ◽  
Sven van IJzendoorn ◽  
Freddy Troost ◽  
...  
Keyword(s):  
Cell Culture ◽  
Epithelial Cell ◽  
Intestinal Epithelial Cell ◽  
Three Dimensional ◽  
In Vitro Study ◽  
Intestinal Epithelial ◽  
Cell Culture Model ◽  
Epithelial Cell Culture ◽  
Culture Model

scholarly journals Human-specific dual regulations of FXR-activation for reduction of fatty liver using in vitro cell culture model

2019 ◽  
Vol 64 (2) ◽  
pp. 112-123 ◽  
Author(s):  
Teruo Miyazaki ◽  
Akira Honda ◽  
Tadashi Ikegami ◽  
Takashi Iida ◽  
Yasushi Matsuzaki
Keyword(s):  
Cell Culture ◽  
Fatty Liver ◽  
Cell Culture Model ◽  
Culture Model ◽  
Human Specific

scholarly journals 2-Deoxy-D-Glucose Attenuates Isoflurane-Induced Cytotoxicity in an In Vitro Cell Culture Model of H4 Human Neuroglioma Cells

2011 ◽  
Vol 113 (6) ◽  
pp. 1468-1475 ◽  
Author(s):  
Jun Zhang ◽  
Yuanlin Dong ◽  
Zhipeng Xu ◽  
Yiying Zhang ◽  
Chuxiong Pan ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Culture Model ◽  
In Vitro Cell Culture ◽  
Culture Model
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