Discovery of Highly Potent C7 Methyl Acetate Substituted Imidazoquinoline-Based TLR7 and TLR8 Agonists

A series of highly substituted imidazoquinolines were synthesized and evaluated for Toll-like receptor (TLR) 7 and 8 activity and potency in stimulating pro-inflammatory cytokine production. The approach applies structure activity relationship data to evaluate the potentiating effects of ring substitutions to the C7-position. Our results show a clear correlation between the inclusion of a C7 methyl acetate group and TLR7 and 8 activity of substituted imidazoquinoline analogs. The most active compounds identified also contain C2 butyl and N1 aminobenzyl substitutions with several analogs displaying single digit nanomolar TLR7 agonist activity. The immunostimulatory effects of the compounds were further evaluated in vitro using human peripheral blood mononuclear cells (PBMCs) and mouse bone marrow-derived dendritic cells (BMDCs). The most potent compounds induce significant production of TNF-α, IL-1β, and IFN-γ by PBMC and expression of MHC II, CD40, and CD80 on BMDCs. A structural model of the TLR7-agonist complex is presented that shows the C7 acetate forms stabilizing H-bonding contacts with several charged amino acid sidechains near the surface of the receptor. The collective results provide strong evidence that correlates the enhanced potency of the C7 acetate analogs to recognition elements at this new site, providing insight to the design of TLR7 and TLR8 ligands.