Induction of interferon-stimulated genes correlates with reduced growth of influenza A virus in the lungs after RIG-I agonist treatment of ferrets

Intracellular RIG-I receptors represent key innate sensors of RNA virus infection and RIG-I activation results in the induction of hundreds of host effector genes, including interferon-stimulated genes (ISGs). Synthetic RNA agonists targeting RIG-I have shown promise as antivirals against a broad spectrum of viruses, including influenza A virus (IAV), both in vitro and in mouse models of infection. Herein, we demonstrate that treatment of a ferret airway epithelial (FRL) cell line with RIG-I agonist rapidly and potently induced expression of a broad range of ISGs and resulted in potent inhibition of growth of different IAV strains. In ferrets, a single intravenous injection of RIG-I agonist was associated with upregulated ISG expression in peripheral blood mononuclear cells (PBMC) and lung tissue, but not in nasal tissues. In a ferret model of viral contact transmission, a single treatment of recipient animals 24 hr prior to co-housing with IAV-infected donors did not reduce virus transmission and shedding but did result in reduced virus titres in lungs six days after treatment. A single treatment of the IAV-infected donor animals also resulted in reduced virus titres in the lungs two days later. Thus, a single intravenous treatment with RIG-I agonist prior to infection or to ferrets with an established IAV infection can reduce virus growth in the lungs. These findings suggest that RIG-I agonists could be developed further as effective antiviral treatment to limit the impact of severe lower airway disease during IAV infections. FRL cells were transfected with 3pRNA, ctrl RNA or treated with recombinant ferret IFN-a and RNA was isolated at indicated time points, prepared with a QuantSeq 3´-mRNA Library Prep Kit (Lexogen) and sequenced on a NovaSeq 6000 (Illumina). The RNA-seq data was processed with nf-core RNA-seq v1.4.2 pipeline using STAR for alignment and featureCounts for gene quantification. The library strandedness parameter was set to forward and the reference was set to domestic ferret genome (GenBank: AEYP00000000.1).