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. 2021 Jan:63:103153.
doi: 10.1016/j.ebiom.2020.103153. Epub 2020 Dec 3.

Prophylactic intranasal administration of a TLR2/6 agonist reduces upper respiratory tract viral shedding in a SARS-CoV-2 challenge ferret model

Pamela C Proud  1 , Daphne Tsitoura  2 , Robert J Watson  1 , Brendon Y Chua  3 , Marilyn J Aram  1 , Kevin R Bewley  1 , Breeze E Cavell  1 , Rebecca Cobb  1 , Stuart Dowall  1 , Susan A Fotheringham  1 , Catherine M K Ho  1 , Vanessa Lucas  1 , Didier Ngabo  1 , Emma Rayner  1 , Kathryn A Ryan  1 , Gillian S Slack  1 , Stephen Thomas  1 , Nadina I Wand  1 , Paul Yeates  1 , Christophe Demaison  2 , Weiguang Zeng  3 , Ian Holmes  2 , David C Jackson  3 , Nathan W Bartlett  4 , Francesca Mercuri  5 , Miles W Carroll  6
Affiliations

Prophylactic intranasal administration of a TLR2/6 agonist reduces upper respiratory tract viral shedding in a SARS-CoV-2 challenge ferret model

Pamela C Proud et al. EBioMedicine. 2021 Jan.

Abstract

Background: The novel human coronavirus SARS-CoV-2 is a major ongoing global threat with huge economic burden. Like all respiratory viruses, SARS-CoV-2 initiates infection in the upper respiratory tract (URT). Infected individuals are often asymptomatic, yet highly infectious and readily transmit virus. A therapy that restricts initial replication in the URT has the potential to prevent progression of severe lower respiratory tract disease as well as limiting person-to-person transmission.

Methods: SARS-CoV-2 Victoria/01/2020 was passaged in Vero/hSLAM cells and virus titre determined by plaque assay. Challenge virus was delivered by intranasal instillation to female ferrets at 5.0 × 106 pfu/ml. Treatment groups received intranasal INNA-051, developed by Ena Respiratory. SARS-CoV-2 RNA was detected using the 2019-nCoV CDC RUO Kit and QuantStudio™ 7 Flex Real-Time PCR System. Histopathological analysis was performed using cut tissues stained with haematoxylin and eosin (H&E).

Findings: We show that prophylactic intra-nasal administration of the TLR2/6 agonist INNA-051 in a SARS-CoV-2 ferret infection model effectively reduces levels of viral RNA in the nose and throat. After 5 days post-exposure to SARS-CoV-2, INNA-051 significantly reduced virus in throat swabs (p=<0.0001) by up to a 24 fold (96% reduction) and in nasal wash (p=0.0107) up to a 15 fold (93% reduction) in comparison to untreated animals.

Interpretation: The results of our study support clinical development of a therapy based on prophylactic TLR2/6 innate immune activation in the URT, to reduce SARS-CoV-2 transmission and provide protection against COVID-19.

Funding: This work was funded by Ena Respiratory, Melbourne, Australia.

Keywords: COVID-19; Ferret; INNA-051; SARS-CoV-2; TLR-2; Viral shedding.

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Conflict of interest statement

Declaration of Competing Interests Authors report grants from Ena Respiratory, during the conduct of the study. W. Zeng and D.C. Jackson reports grants from Ena Therapeutics, during the conduct of the study. D. Tsitoura, C. Demaison, F. Mercuri, I. Holmes and N.W. Bartlett reports personal fees and other from Ena Therapeutics, outside the submitted work. D.C. Jackson, W. Zeng and B.Y. Chua reports other from Ena Therapeutics, outside the submitted work. Dr. Holmes reports personal fees from Ena Therapeutics, outside the submitted work. In addition, D. Tsitoura, C. Demaison and F. Mercuri have a patent AU 2020901709 pending to Ena Therapeutics. D.C Jackson, W. Zeng and C. Demaison have a patent PCT/AU2011/001225 issued to Ena Therapeutics. D.C Jackson, W. Zeng, I. Holmes and C. Demaison have a patent PCT/AU2020/050660 pending to Ena Therapeutics.

Figures

Fig 1
Fig. 1
Clinical observations. (a) Schematic of experimental design. Ferrets received INNA-051 and PBS treatments 4 days and 1 day prior to challenge with 5.0 × 106 pfu/ml SARS-CoV-2. Nasal wash and throat swabs were collected at days 1, 3, 5, 7, 10 & 12 post challenge (p.c.) for all treatment groups and control group. Scheduled culls were performed for 6/24 ferrets on day 3 p.c. (group 1 n=2, group 2 n=2, group 4 n=2) and 18/24 ferrets on days 12-14 p.c. (b) Temperatures were measured twice daily (approximately 8 hours apart), using implanted temperature/ID chips. Mean temperatures +/- standard error of the mean (SEM) are displayed. Temperature dip post SARS-CoV-2 challenge (*) was attributed to sedation. Group 1 animals were first to come out of sedation and therefore recover temperatures. (b) Weight was recorded daily and percentage change from the ferret weight prior to treatment plotted. Mean percentage weight change +/- SEM are displayed. Weight dips on days 4 and 8 p.c. (*) attributed to 2/4 ferrets in group 2 losing weight due to thirst after upturning water bowl overnight. Ferrets in group 3 (2/6 animals) lost weight for same reason on day 8 p.c.
Fig 2
Fig. 2
Viral RNA shedding following SARS-CoV-2 challenge. Nasal wash and throat swabs were collected for all treatment groups and vehicle control group at days 1, 3 (n=6 for groups 1-4), 5, 7, 10 & 12 p.c. (n=4 groups 1,2&4, n=6 group 3). Lung tissue was collected at necropsy on scheduled cull day 3 (n=6) and end cull days 12-14 (n=18). Viral RNA was quantified by RT-qPCR. (a) Nasal wash (b) Throat swab (c) Lung tissue. Geometric mean +/- standard deviation are displayed on the graphs. Dashed horizontal lines denote the lower limit of quantification (LLOQ) and lower limit of detection (LLOD). Day 7 nasal wash for group 4 had viral RNA quantified for 3/4 ferrets; no sample was available for processing. Statistical significance (95% CI of differences) in comparison to the control group using two-way ANOVA Dunnett's multiple comparisons test are displayed above the error bars (*). Fig a) day 3 group 3 (p=0.0155), day 5 group 1 (p=0.0244), group 2 (p=0.0107) and group 3 (p=0.0071), day 7 group 2 (p=0.0054). Fig b) day 1 group 1 (p=0.0129), day 3 group 3 (p=0.0345), day 5 group 1 (p=0.0002), group 2 (p=<0.0001) and group 3 (p=0.0039), day 7 group 1 (p=0.0014), group 2 (p=<0.0001) and group 3 (p=0.0002).
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References

    1. Cui J, Li F, Shi Z-L. Origin and evolution of pathogenic coronaviruses. Nat Rev Microbiol. 2019;17(3):181–192. - PMC - PubMed
    1. Li X, Zai J, Zhao Q, Nie Q, Li Y, Foley BT. Evolutionary history, potential intermediate animal host, and cross-species analyses of SARS-CoV-2. J Med Virol. 2020;92(6) - PMC - PubMed
    1. Zhong NS, Zheng BJ, Li YM, Poon LLM, Xie ZH, Chan KH. Epidemiology and cause of severe acute respiratory syndrome (SARS) in Guangdong, People's Republic of China, in February, 2003. Lancet North Am Ed. 2003;362(9393):1353–1358. - PMC - PubMed
    1. Zaki AM, van Boheemen S, Bestebroer TM, Osterhaus ADME, Fouchier RAM. Isolation of a novel coronavirus from a man with pneumonia in Saudi Arabia. N Engl J Med. 2012;367(19):1814–1820. - PubMed
    1. Lu R, Zhao X, Li J, Niu P, Yang B, Wu H. Genomic characterisation and epidemiology of 2019 novel coronavirus: implications for virus origins and receptor binding. Lancet North Am Ed. 2020;395(10224):565–574. - PMC - PubMed

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