AAV-CRISPR-Cas13 eliminates human enterovirus and prevents death of infected mice
Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved..
BACKGROUND: RNA viruses account for many human diseases and pandemic events but are often not targetable by traditional therapeutics modalities. Here, we demonstrate that adeno-associated virus (AAV) -delivered CRISPR-Cas13 directly targets and eliminates the positive-strand EV-A71 RNA virus in cells and infected mice.
METHODS: We developed a Cas13gRNAtor bioinformatics pipeline to design CRISPR guide RNAs (gRNAs) that cleave conserved viral sequences across the virus phylogeny and developed an AAV-CRISPR-Cas13 therapeutics using in vitro viral plaque assay and in vivo EV-A71 lethally-infected mouse model.
FINDINGS: We show that treatment with a pool of AAV-CRISPR-Cas13-gRNAs designed using the bioinformatics pipeline effectively blocks viral replication and reduces viral titers in cells by >99.99%. We further demonstrate that AAV-CRISPR-Cas13-gRNAs prophylactically and therapeutically inhibited viral replication in infected mouse tissues and prevented death in a lethally challenged EV-A71-infected mouse model.
INTERPRETATION: Our results show that the bioinformatics pipeline designs efficient CRISPR-Cas13 gRNAs for direct viral RNA targeting to reduce viral loads. Additionally, this new antiviral AAV-CRISPR-Cas13 modality represents an effective direct-acting prophylactic and therapeutic agent against lethal RNA viral infections.
FUNDING: Agency for Science, Technology and Research (A∗STAR) Assured Research Budget, A∗STAR Central Research Fund UIBR SC18/21-1089UI, A∗STAR Industrial Alignment Fund Pre-Positioning (IAF-PP) grant H17/01/a0/012, MOE Tier 2 2017 (MOE2017-T2-1-078; MOE-T2EP30221-0005), and NUHSRO/2020/050/RO5+5/NUHS-COVID/4.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2023 |
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| Erschienen: |
2023 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:93 |
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| Enthalten in: |
EBioMedicine - 93(2023) vom: 30. Juli, Seite 104682 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Keng, Choong Tat [VerfasserIn] |
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| Links: |
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| Themen: |
Adeno-associated viral vectors |
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| Anmerkungen: |
Date Completed 17.07.2023 Date Revised 26.07.2023 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1016/j.ebiom.2023.104682 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| 520 | |a Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved. | ||
| 520 | |a BACKGROUND: RNA viruses account for many human diseases and pandemic events but are often not targetable by traditional therapeutics modalities. Here, we demonstrate that adeno-associated virus (AAV) -delivered CRISPR-Cas13 directly targets and eliminates the positive-strand EV-A71 RNA virus in cells and infected mice | ||
| 520 | |a METHODS: We developed a Cas13gRNAtor bioinformatics pipeline to design CRISPR guide RNAs (gRNAs) that cleave conserved viral sequences across the virus phylogeny and developed an AAV-CRISPR-Cas13 therapeutics using in vitro viral plaque assay and in vivo EV-A71 lethally-infected mouse model | ||
| 520 | |a FINDINGS: We show that treatment with a pool of AAV-CRISPR-Cas13-gRNAs designed using the bioinformatics pipeline effectively blocks viral replication and reduces viral titers in cells by >99.99%. We further demonstrate that AAV-CRISPR-Cas13-gRNAs prophylactically and therapeutically inhibited viral replication in infected mouse tissues and prevented death in a lethally challenged EV-A71-infected mouse model | ||
| 520 | |a INTERPRETATION: Our results show that the bioinformatics pipeline designs efficient CRISPR-Cas13 gRNAs for direct viral RNA targeting to reduce viral loads. Additionally, this new antiviral AAV-CRISPR-Cas13 modality represents an effective direct-acting prophylactic and therapeutic agent against lethal RNA viral infections | ||
| 520 | |a FUNDING: Agency for Science, Technology and Research (A∗STAR) Assured Research Budget, A∗STAR Central Research Fund UIBR SC18/21-1089UI, A∗STAR Industrial Alignment Fund Pre-Positioning (IAF-PP) grant H17/01/a0/012, MOE Tier 2 2017 (MOE2017-T2-1-078; MOE-T2EP30221-0005), and NUHSRO/2020/050/RO5+5/NUHS-COVID/4 | ||
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| 700 | 1 | |a Chia, Bing Shao |e verfasserin |4 aut | |
| 700 | 1 | |a Vasandani, Suraj Rajan |e verfasserin |4 aut | |
| 700 | 1 | |a Lim, Daryl Shern |e verfasserin |4 aut | |
| 700 | 1 | |a Guo, Ke |e verfasserin |4 aut | |
| 700 | 1 | |a Wong, Yi Hao |e verfasserin |4 aut | |
| 700 | 1 | |a Mok, Chee Keng |e verfasserin |4 aut | |
| 700 | 1 | |a Chu, Justin Jang Hann |e verfasserin |4 aut | |
| 700 | 1 | |a Chew, Wei Leong |e verfasserin |4 aut | |
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