TY - JOUR
T1 - SARS-CoV-2 Nucleocapsid Protein TR-FRET Assay Amenable to High Throughput Screening
AU - Gorshkov, Kirill
AU - Morales Vasquez, Desarey
AU - Chiem, Kevin
AU - Ye, Chengjin
AU - Nguyen Tran, Bruce
AU - Carlos De La Torre, Juan
AU - Moran, Thomas
AU - Chen, Catherine Z.
AU - Martinez-Sobrido, Luis
AU - Zheng, Wei
N1 - Funding Information:
We thank Dr. Charles Chiu, M.D./Ph.D., Director, UCSF-Abbott Viral Diagnostics and Discovery Center at UCSF School of Medicine for the isolate SARS-CoV-2/human/USA/CA-UCSF-0001C/2020. This research was supported in part by the Intramural Research Program of the National Center for Advancing Translational Sciences, NIH. We thank the development group at Columbia Biosciences for their assistance in reagent development.
Publisher Copyright:
©
PY - 2022/1/14
Y1 - 2022/1/14
N2 - Drug development for specific antiviral agents against coronavirus disease 2019 (COVID-19) is still an unmet medical need as the pandemic continues to spread globally. Although huge efforts for drug repurposing and compound screens have been put forth, only a few compounds are in late-stage clinical trials. New approaches and assays are needed to accelerate COVID-19 drug discovery and development. Here, we report a time-resolved fluorescence resonance energy transfer-based assay that detects the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (NP) produced in infected cells. It uses two specific anti-NP monoclonal antibodies conjugated to donor and acceptor fluorophores that produce a robust ratiometric signal for high throughput screening of large compound collections. Using this assay, we measured a half maximal inhibitory concentration (IC50) for remdesivir of 9.3 μM against infection with SARS-CoV-2 USA/WA1/2020 (WA-1). The assay also detected SARS-CoV-2 South African (Beta, β), Brazilian/Japanese P.1 (Gamma, γ), and Californian (Epsilon, ϵ) variants of concern (VoC). Therefore, this homogeneous SARS-CoV-2 NP detection assay can be used for accelerating lead compound discovery for drug development and for evaluating drug efficacy against emerging SARS-CoV-2 VoC.
AB - Drug development for specific antiviral agents against coronavirus disease 2019 (COVID-19) is still an unmet medical need as the pandemic continues to spread globally. Although huge efforts for drug repurposing and compound screens have been put forth, only a few compounds are in late-stage clinical trials. New approaches and assays are needed to accelerate COVID-19 drug discovery and development. Here, we report a time-resolved fluorescence resonance energy transfer-based assay that detects the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid protein (NP) produced in infected cells. It uses two specific anti-NP monoclonal antibodies conjugated to donor and acceptor fluorophores that produce a robust ratiometric signal for high throughput screening of large compound collections. Using this assay, we measured a half maximal inhibitory concentration (IC50) for remdesivir of 9.3 μM against infection with SARS-CoV-2 USA/WA1/2020 (WA-1). The assay also detected SARS-CoV-2 South African (Beta, β), Brazilian/Japanese P.1 (Gamma, γ), and Californian (Epsilon, ϵ) variants of concern (VoC). Therefore, this homogeneous SARS-CoV-2 NP detection assay can be used for accelerating lead compound discovery for drug development and for evaluating drug efficacy against emerging SARS-CoV-2 VoC.
KW - SARS-CoV-2
KW - TR-FRET
KW - assay development
KW - nucleocapsid phosphoprotein
KW - variants of concern
UR - http://www.scopus.com/inward/record.url?scp=85122734695&partnerID=8YFLogxK
U2 - 10.1021/acsptsci.1c00182
DO - 10.1021/acsptsci.1c00182
M3 - Article
AN - SCOPUS:85122734695
SN - 2575-9108
VL - 5
SP - 8
EP - 19
JO - ACS Pharmacology and Translational Science
JF - ACS Pharmacology and Translational Science
IS - 1
ER -