Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were intranasally infected with 10 3 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 4 hours post infection, 15 μg SLR14 or vehicle were intravenously administered. Weight loss and survival were monitored daily up to 14 DPI. Death was recorded when mice were found dead in the cage, moribund, or at 80% of original body weight. In a separate cohort, lung tissues were collected for virological and immunological analysis 5 DPI. b,c , Weight loss ( b ) and survival ( c ) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI. d,e , Measurement of genomic viral RNA in the lung 5 DPI by reverse-transcription quantitative PCR (RT-qPCR) against SARS-CoV-2 N gene using CDCN1 ( d ) or CDCN2 ( e ) primer-probe sets. f , Measurement of infectious virus titer in the lung 5 DPI by plaque assay. Limit of detection (LOD): 10 2 PFU/mL. g–i , Measurement of expression of interferon stimulated genes (ISG) Cxcl9 ( g ), Isg15 ( h ) and Usp18 ( i ) in the lung 5 DPI by RT-qPCR. j,k , Frequency of CD11b + CD64 + macrophages of CD45 + cells ( j ) and mean fluorescence intensity of MHCII on Ly6C high monocytes ( k ) in the lung 5 DPI by flow cytometry. Mean ± s.e.m.; Statistical significance was calculated by log-rank Mantel–Cox test ( c ) or one-way ANOVA followed by Tukey correction ( d–k ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Data are representative of two independent experiments.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Plaque Assay, Expressing, Fluorescence, Flow Cytometry

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were intranasally infected with 10 3 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 2 hours before infection, 15 μg SLR14 or vehicle were intravenously administered 7 DPI. 24 hours before SLR14 injection, half of SLR14-treated mice was additionally given 2 mg anti-IFNAR antibodies. Weight loss and survival were monitored daily up to 14 DPI. Death was recorded when mice were found dead in the cage, moribund, or at 80% of original body weight. In a separate cohort, lung tissues were collected for virological analysis 3, 6, and 8 DPI. B–d , Weight loss ( b,c ) and survival ( d ) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI. E–g , Measurement of genomic viral RNA in the lung parenchyma 3, 6, and 8 DPI by RT-qPCR using the CDCN2 primer-probe set. H–j , Measurement of genomic viral RNA in the trachea 3, 6, and 8 DPI by RT-qPCR using the CDCN2 primer-probe set. Mean ± s.e.m.; Statistical significance was calculated by log-rank Mantel–Cox test ( d ) or one-way ANOVA followed by Tukey correction ( e–j ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Data are representative of two independent experiments.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Injection, Quantitative RT-PCR

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were intranasally infected with 10 3 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 2 hours before infection, 15 μg SLR14 or vehicle were intravenously administered. 24 hours before SLR14 injection, half of SLR14-treated mice was additionally given 2 mg anti-IFNAR antibodies. Nasal washes were collected for virological analysis 3, 6, and 8 DPI. b–d , Measurement of genomic viral RNA in the nasal wash 3, 6, and 8 DPI by RT-qPCR using the CDCN2 primer-probe set. Mean ± s.e.m.; Statistical significance was calculated by one-way ANOVA followed by Tukey correction ( b–d ); *P ≤ 0.05, **P ≤ 0.01,***P ≤ 0.001, ****P ≤ 0.0001.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Injection, Quantitative RT-PCR

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: Ifnar −/− mice were intratracheally administered with 10 11 genome copies of AAV9- hACE2 and let rest for 2 weeks before intranasal infection with 10 6 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 15 μg SLR14 or vehicle were intravenously administered at 4 hours after infection. Lung tissues were collected for virological analysis 4 DPI. b , Measurement of genomic viral RNA 4 DPI by RT-qPCR using the CDCN2 primer-probe set. Mean ± s.e.m.; Statistical significance was calculated by one-way ANOVA followed by Tukey correction ( b ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Quantitative RT-PCR

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were i.v. injected with 15 μg AF647- conjugated SLR14 or vehicle. Lung tissues were collected for SLR14 uptake analysis by flow cytometry 4 hours post injection. Lung tissues from vehicle-injected controls were also collected as negative controls. b , Frequency of indicated immune and non-immune cell types among SLR14 + cells versus total lung cells. c , Frequency of indicated macrophage populations among SLR14 + cells or total lung cells. d , Distribution index (frequency of a given cell type in the SLR14 + compartment/frequency of all cells) of indicated immune and non-immune cell types. e , Distribution index of indicated macrophage populations.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Injection, Flow Cytometry

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were intranasally infected with 10 3 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 15 μg SLR14 were intravenously administered at 16 hours before, 2 hours before, 4 hours after, 24 hours after, 48 hours after, or 72 hours after infection. Weight loss and survival were monitored daily up to 14 DPI. Death was recorded when mice were found dead in the cage, moribund, or at 80% of original body weight. B–d , Weight loss ( b,c ) and survival ( d ) of prophylactically SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI. E–g , Weight loss ( e,f ) and survival ( g ) of therapeutically SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI. Mean ± s.e.m.; Statistical significance was calculated by log-rank Mantel–Cox test ( d,g ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Data are pooled from of three independent experiments.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: Rag −/− mice were intratracheally administered with 10 11 genome copies of AAV9- hACE2 and let rest for 2 weeks before intranasal infection with 10 6 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 200 μL convalescent sera or PBS were intravenously administered 7 DPI. Lung tissues were collected for virological analysis 14 DPI. b , Measurement of genomic viral RNA in the lung 14 DPI by RT-qPCR. c , Measurement of infectious virus titer in the lung 14 DPI by plaque assay. Limit of detection (LOD) for plaque assay: 10 2 PFU/mL. Mean ± s.e.m.; Statistical significance was calculated by one-way ANOVA followed by Tukey correction ( b , c ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Data are pooled from two independent experiments.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Quantitative RT-PCR, Plaque Assay

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: Rag −/− mice were intratracheally administered with 10 11 genome copies of AAV9- hACE2 and let rest for 2 weeks before intranasal infection with 10 6 PFU SARS-CoV-2 (2019n-CoV/USA_WA1/2020). 15 μg SLR14 or vehicle were intravenously administered 7 DPI. 24 hours before SLR14 injection, half of SLR14-treated mice was additionally given 2 mg anti-IFNAR antibodies. Lung tissues were collected for virological analysis 14 DPI. b , Measurement of genomic viral RNA in the lung 14 DPI by RT-qPCR. c , Measurement of infectious virus in the lung 14 DPI by plaque assay. Limit of detection (LOD): 10 2 PFU/mL. Mean ± s.e.m.; Statistical significance was calculated by one-way ANOVA followed by Tukey correction ( b,c ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Data are pooled from two independent experiments.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Injection, Quantitative RT-PCR, Plaque Assay

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were intranasally infected with 10 4 B.1.526, B.1.351, or B.1.1.7 variants. 15 μg SLR14 or vehicle were intravenously administered at 4 hours after infection. Weight loss and survival were monitored daily up to 14 DPI. Death was recorded when mice were found dead in the cage, moribund, or at 80% of original body weight. b–d , Weight loss (b,c) and survival (d) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following P.1 infection. e–g, Weight loss (e,f) and survival (g) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following B.1.526 infection. h–j, Weight loss (h,i) and survival (j) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following B.1.351 infection. k–m, Weight loss (k,l) and survival (m) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following B.1.1.7 infection. Mean ± s.e.m.; Statistical significance was calculated by log-rank Mantel–Cox test (d,g,j,m); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001. Data are pooled from two independent experiments.

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection

Journal: bioRxiv

Article Title: A stem-loop RNA RIG-I agonist confers prophylactic and therapeutic protection against acute and chronic SARS-CoV-2 infection in mice

doi: 10.1101/2021.06.16.448754

Figure Lengend Snippet: a , Experimental scheme: K18-hACE2 mice were intranasally infected with 10 4 , 3.3 × 10 3 , or 10 3 PFU B.1.1.7 variant. 15 μg SLR14 or vehicle were intravenously administered at 4 hours after infection. Weight loss and survival were monitored daily up to 14 DPI. Death was recorded when mice were found dead in the cage, moribund, or at 80% of original body weight. B–d , Weight loss ( b,c ) and survival ( d ) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following infection with 10 4 PFU B.1.1.7. e–g , Weight loss ( e,f ) and survival ( g ) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following infection with 3.3 × 10 3 PFU B.1.1.7. h-j , Weight loss ( h,i ) and survival (j) of SLR14- and vehicle-treated K18-hACE2 mice from 1 to 14 DPI following infection with 103 PFU B.1.1.7. Mean ± s.e.m.; Statistical significance was calculated by log-rank Mantel–Cox test ( d,g,j ); *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001

Article Snippet: AAV9 vector encoding hACE2 was purchased from Vector Biolabs (AAV9-CMV-hACE2).

Techniques: Infection, Variant Assay