Structure−Activity Relationships Towards the Identification of High-Potency Selective Human Toll-Like Receptor-7 Agonist

Toll-like receptors (TLRs) act as the “sentinel” of the immune system to link innate immune responses with adaptive immunity. TLR7 agonists are highly immunostimulatory and can be exploited as powerful vaccine adjuvants. A structure-activity relationship study was conducted on the TLR7-active imidazoquinoline (IMDQ) scaffold, starting with 1-benzyl-2-butyl-1H-imidazo[4,5-c]quinolin-4-amine (BBIQ) as a lead structure. A systematic exploration of electron withdrawing as well as electron donating substituents at the para-position of benzyl group at N-1 position of IMDQ scaffold led to the identification of a highly active para-hydroxymethyl IMDQ analogue with an EC50 value of 0.23 µM for human TLR7 with marginal activity for human TLR8, thereby indicating it as a TLR7-specific agonist that was 37-fold more potent than imiquimod. Its bio-steric para-aminomethyl analogue was a dual TLR7 and TLR8 agonist. Molecular modelling was performed which revealed the TLR8 activity of the IMDQ scaffold to be associated with the presence of amino functionality in the benzyl group. TLR7-biased activity was driven by the forming of multiple H-bonds with TLR7 which not formed when the IMDQ scaffold compounds were docked with TLR8. Finally, the role of the IMDQ scaffold agonists as vaccine adjuvants was tested with a Covid-19 vaccine in mice, which showed that TLR7 activity even in the absence of TLR8 activity was sufficient for potentiation of anti-spike protein antibody production, suggesting that TLR7 specific agonists may make suitable vaccine adjuvants.