Small molecule RIG-I agonist serves to adjuvant broad, multifaceted influenza virus vaccine immunity

We identified a small molecule compound, KIN1148, that directly binds RIG-I to drive IRF3 and NF B activation and expression of innate immune genes, cytokines and chemokines. KIN1148 activates RIG-I in an RNA- and ATP-independent manner and does not induce a canonical antiviral interferon (IFN) gene program traditionally associated with RIG-I activation. When administered in combination with a vaccine against influenza A virus (IAV), KIN1148 induces both neutralizing antibody and broadly cross-protective IAV-specific T cell responses compared to vaccination alone, which induces poor responses. In this study, we demonstrate that KIN1148 directly engages RIG-I to activate IRF3- and NFB-dependent innate immune responses, making it the first small molecule RIG-I agonist to be identified. Biochemical studies show that KIN1148 binds to RIG-I to drive RIG-I self-oligomerization and downstream signaling activation in an RNA- and ATP-independent manner. We further find that transcriptional programs induced by KIN1148 treatment exhibit shared and unique signatures to that induced by other methods of RIG-I activation, including Sendai virus (SeV) infection and PAMP RNA transfection. KIN1148 adjuvants a split virus (SV) vaccine at suboptimal dose to protect mice from lethal challenge with a recombinant highly pathogenic avian H5N1 influenza virus, A/Vietnam/1203/2004. Differentiated THP-1 cells were treated with small molecule compounds (KIN1000 and KIN1148 at 0.625, 2.5 or 10 micromoles/liter) or 25 IU/mL IFN Beta diluted in cRPMI supplemented with 0.5% (v/v) DMSO, or treated with cRPMI supplemented with 0.5% (v/v) DMSO alone (DMSO). Control cells infected with 25 HAU/mL SeV were maintained in cRPMI supplemented with 0.5% (v/v) DMSO after removal of the virus inoculum.