Small-molecule inhibition of TLR8 through stabilization of its resting state

Abstract

Endosomal Toll-like receptors (TLR3/7/8/9) are highly analogous sensors for various viral or bacterial RNA/DNA molecular patterns. Nonetheless, few small-molecules can selectively modulate these TLRs. In this manuscript, we identified the first human TLR8-specific small-molecule antagonists via a novel inhibition mechanism. Crystal structures of two distinct TLR8-ligand complexes validated a unique binding site on the protein-protein interface of the TLR8 homodimer. Upon binding to this new site, the small-molecule ligands stabilize the preformed TLR8 dimer in its resting state, preventing activation. As a proof of concept of their therapeutic potential, we have demonstrated that these drug-like inhibitors are able to suppress TLR8-mediated proinflammatory signaling in various cell lines, human primary cells, and patient specimens. These results not only suggest a novel strategy for TLR inhibitor design, but also shed critical mechanistic insight into these clinically important immune receptors.

Sensing of HIV-1 by TLR8 activates human T cells and reverses latency

“…2b). To further assess the specificity of the TLR8 response, we utilized TLR8 inhibitors that stabilize the TLR8 dimer in its inactive state 45,46 . CD4+ T cells were pre-treated with the structurally optimized TLR8 inhibitors CU-CPT9a and CU-CPT9b, or the negative control compound CU-CPT6, for 2 h prior to TCR activation and stimulation with CL264, CL75, pU/ pLA or the TLR2 ligand FSL-1 for 24 h. TLR8-induced IL-6 was significantly reduced by inhibitors CU-CPT9a and CU-CPT9b ( Fig.…”

Agonist and antagonist ligands of toll-like receptors 7 and 8: Ingenious tools for therapeutic purposes

“…TLR antagonists are generally modified agonists that bind TLRs but fail to induce the signal transduction [69,283]. Recently highlighting the existence of antagonist pockets on TLRs 7/8 will also allow specific structure-based design using molecular modeling tools to produce new potent structures [9,284]. They represent novel therapies to treat or prevent diseases cited herein [285].…”

“…Recently, Zhang et al identified, for the first time, human TLR8 small antagonist molecules with a novel inhibition mechanism [304]. SAR studies on the pyrazolo[1,5-a]pyrimidine series led to identification of the compound: CU-CPT8m (64), which proved to be the most active with an IC50 of 67 nM and a strong ability to inhibit the production of pro-inflammatory cytokines (Fig.…”

RNA is taking its Toll: Impact of RNA‐specific Toll‐like receptors on health and disease

“…Understanding the receptor‐ligand interactions and molecular pathways is of great importance in order to restrain or enhance these immune reactions. This knowledge recently led to the development of new TLR8 antagonists, the CU‐CPT 9 derivates, which stabilize the receptor's resting state to inhibit its activation . The antagonists possess IC50 values in a nM‐pM range and exhibit high specificity with the ability to discriminate between TLR7 and TLR8 which is superior compared to previous inhibitors .…”