Development of CRISPR as an Antiviral Strategy to Combat SARS-CoV-2 and Influenza

Timothy R Abbott; Girija Dhamdhere; Yanxia Liu; Xueqiu Lin; Laine Goudy; Leiping Zeng; Augustine Chemparathy; Stephen Chmura; Nicholas S Heaton; Robert Debs; Tara Pande; Drew Endy; Marie F La Russa; David B Lewis; Lei S Qi

doi:10.1016/j.cell.2020.04.020

Development of CRISPR as an Antiviral Strategy to Combat SARS-CoV-2 and Influenza

Cell. 2020 May 14;181(4):865-876.e12. doi: 10.1016/j.cell.2020.04.020. Epub 2020 Apr 29.

Authors

Affiliations

¹ Department of Bioengineering, Stanford University, Stanford, CA 94305, USA.
² Department of Pediatrics, Stanford University, Stanford, CA 94305, USA.
³ Department of Management Science and Engineering, Stanford University, Stanford, CA 94305, USA.
⁴ DNARx, San Francisco, CA 94107, USA.
⁵ Department of Molecular Genetics and Microbiology, Duke University School of Medicine, Durham, NC 27710, USA.
⁶ Los Altos High School, Los Altos, CA 94022, USA.
⁷ Department of Bioengineering, Stanford University, Stanford, CA 94305, USA. Electronic address: mlarussa@stanford.edu.
⁸ Department of Pediatrics, Stanford University, Stanford, CA 94305, USA. Electronic address: dblewis@stanford.edu.
⁹ Department of Bioengineering, Stanford University, Stanford, CA 94305, USA; Department of Chemical and Systems Biology, Stanford University, Stanford, CA 94305, USA; ChEM-H, Stanford University, Stanford, CA 94305, USA. Electronic address: stanley.qi@stanford.edu.

Abstract

The coronavirus disease 2019 (COVID-19) pandemic, caused by the SARS-CoV-2 virus, has highlighted the need for antiviral approaches that can target emerging viruses with no effective vaccines or pharmaceuticals. Here, we demonstrate a CRISPR-Cas13-based strategy, PAC-MAN (prophylactic antiviral CRISPR in human cells), for viral inhibition that can effectively degrade RNA from SARS-CoV-2 sequences and live influenza A virus (IAV) in human lung epithelial cells. We designed and screened CRISPR RNAs (crRNAs) targeting conserved viral regions and identified functional crRNAs targeting SARS-CoV-2. This approach effectively reduced H1N1 IAV load in respiratory epithelial cells. Our bioinformatic analysis showed that a group of only six crRNAs can target more than 90% of all coronaviruses. With the development of a safe and effective system for respiratory tract delivery, PAC-MAN has the potential to become an important pan-coronavirus inhibition strategy.

Keywords: 2019-nCoV; COVID-19; CRISPR; Cas13; IAV; RdRP; SARS-CoV-2; antiviral; influenza; nucleocapsid.

MeSH terms

A549 Cells
Antibiotic Prophylaxis / methods
Antiviral Agents / pharmacology*
Base Sequence
Betacoronavirus / drug effects*
Betacoronavirus / genetics
Betacoronavirus / growth & development
COVID-19
CRISPR-Cas Systems*
Clustered Regularly Interspaced Short Palindromic Repeats
Computer Simulation
Conserved Sequence
Coronavirus / drug effects
Coronavirus / genetics
Coronavirus / growth & development
Coronavirus Infections / drug therapy
Coronavirus Nucleocapsid Proteins
Coronavirus RNA-Dependent RNA Polymerase
Epithelial Cells / virology
Humans
Influenza A Virus, H1N1 Subtype / drug effects*
Influenza A Virus, H1N1 Subtype / genetics
Influenza A Virus, H1N1 Subtype / growth & development
Lung / pathology
Lung / virology
Nucleocapsid Proteins / genetics
Pandemics
Phosphoproteins
Phylogeny
Pneumonia, Viral / drug therapy
RNA, Viral / antagonists & inhibitors*
RNA-Dependent RNA Polymerase / genetics
SARS-CoV-2
Viral Nonstructural Proteins / genetics

Substances

Antiviral Agents
Coronavirus Nucleocapsid Proteins
Nucleocapsid Proteins
Phosphoproteins
RNA, Viral
Viral Nonstructural Proteins
nucleocapsid phosphoprotein, SARS-CoV-2
Coronavirus RNA-Dependent RNA Polymerase
NSP12 protein, SARS-CoV-2
RNA-Dependent RNA Polymerase