CRISPR-Cas13d effectively targets SARS-CoV-2 variants, including Delta and Omicron, and inhibits viral infection

Zongzhi Liu; Xiang Gao; Chuanwen Kan; Lingyu Li; Yuan Zhang; Yibo Gao; Shengyuan Zhang; Liangji Zhou; Hui Zhao; Mingkun Li; Zheng Zhang; Yingli Sun

doi:10.1002/mco2.208

CRISPR-Cas13d effectively targets SARS-CoV-2 variants, including Delta and Omicron, and inhibits viral infection

MedComm (2020). 2023 Jan 31;4(1):e208. doi: 10.1002/mco2.208. eCollection 2023 Feb.

Authors

Zongzhi Liu^{1

2

3}, Xiang Gao⁴, Chuanwen Kan^{2

5}, Lingyu Li^{1

2

3}, Yuan Zhang⁶, Yibo Gao⁷, Shengyuan Zhang⁴, Liangji Zhou⁶, Hui Zhao^{6

8

9}, Mingkun Li^{2

10}, Zheng Zhang⁴, Yingli Sun^{1

2

3}

Affiliations

¹ Central Laboratory National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital and Shenzhen Hospital Chinese Academy of Medical Sciences and Peking Union Medical College Shenzhen China.
² University of Chinese Academy of Sciences Beijing China.
³ CAS Key Laboratory of Genome Sciences and Information Beijing Institute of Genomics Chinese Academy of Sciences/China National Center for Bioinformation Beijing China.
⁴ Institute of Hepatology National Clinical Research Center for Infectious Disease School of Medicine Shenzhen Third People's Hospital The Second Affiliated Hospital Southern University of Science and Technology Shenzhen Guangdong China.
⁵ Beijing Institute of Genomics Chinese Academy of Sciences, China National Center for Bioinformation Beijing China.
⁶ Key Laboratory for Regenerative Medicine Ministry of Education School of Biomedical Sciences Faculty of Medicine The Chinese University of Hong Kong Hong Kong.
⁷ National Cancer Center/National Clinical Research Center for Cancer/Cancer Hospital Chinese Academy of Medical Sciences and Peking Union Medical College Beijing China.
⁸ Kunming Institute of Zoology, The Chinese University of Hong Kong (KIZ-CUHK) Joint Laboratory of Bioresources and Molecular Research of Common Diseases The Chinese University of Hong Kong Hong Kong.
⁹ Hong Kong Branch of CAS Center for Excellence in Animal Evolution and Genetics The Chinese University of Hong Kong Hong Kong.
¹⁰ Key Laboratory of Genomic and Precision Medicine Beijing Institute of Genomics Chinese Academy of Sciences China National Center for Bioinformation Beijing China.

Abstract

The recent pandemic of variants of concern (VOC) of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) highlights the need for innovative anti-SARS-CoV-2 approaches in addition to vaccines and antiviral therapeutics. Here, we demonstrate that a CRISPR-Cas13-based strategy against SARS-CoV-2 can effectively degrade viral RNA. First, we conducted a cytological infection experiment, screened CRISPR-associated RNAs (crRNAs) targeting conserved regions of viruses, and used an in vitro system to validate functional crRNAs. Reprogrammed Cas13d effectors targeting NSP13, NSP14, and nucleocapsid transcripts achieved >99% silencing efficiency in human cells which are infected with coronavirus 2, including the emerging variants in the last 2 years, B.1, B.1.1.7 (Alpha), D614G B.1.351 (Beta), and B.1.617 (Delta). Furthermore, we conducted bioinformatics data analysis. We collected the sequence information of COVID-19 and its variants from China, and phylogenetic analysis revealed that these crRNA oligos could target almost 100% of the SARS-CoV family, including the emerging new variant, Omicron. The reprogrammed Cas13d exhibited high specificity, efficiency, and rapid deployment properties; therefore, it is promising for antiviral drug development. This system could possibly be used to protect against unexpected SARS-CoV-2 variants carrying multiple mutations.

Keywords: CRISPR‐Cas system; RNAi therapy; SARS‐CoV‐2; gene editing.