Toll-like Receptors (TLR) 7 and 8 are sensors of the innate immune system that trigger inflammatory cytokine and Type I IFN production in response RNA degradation products. Our laboratory has recently uncovered a novel mechanism for TLR7/8 regulation, whereby naturally-occurring 2′-O-methyl (2′-OMe)-modified RNA fragments, as short as three bases, potently antagonise TLR7 and TLR8 to prevent autoimmunity. Starting from these natural ligands, we undertook a medicinal chemistry approach to optimise both the potency and selectivity of 3-base 2′-OMe oligonucleotides (oligos) to inhibit TLR7 and TLR8. Using a TLR7-driven animal model of skin inflammation, we demonstrated that topical application of a modified murine TLR7-inhibitory oligo greatly ameliorates skin inflammation while reducing pro-inflammatory gene expression in vivo. Additional studies show that co-packaging a TLR7 inhibitory oligo with mRNA within lipid nanoparticles (LNPs) effectively reduced mRNA-driven reactogenicity, without compromising mRNA expression or the physical properties of the LNPs.
Excitingly, lead optimisation studies of a human-specific 3-base oligo identified that a single base modification allows for dual inhibition of human TLR7/8. In vitro studies in human blood confirmed these oligos inhibit TLR7/8 activation by potent small molecule agonists, like resiquimod. As such, we are currently pursuing the development of a dual human TLR7/8 antagonistic 3-base oligonucleotide (SOF-SKN™) for the treatment of inflammatory skin diseases.
Collectively, our work establishes that select short oligonucleotides can be rationally designed to effectively co-opt a natural immune checkpoint in inflammation, with a wide range of potential therapeutic applications in chronic and auto-inflammatory disorders, and for the modulation of innate immune responses to RNA therapeutics, including mRNA vaccines.