Toll-like Receptor (TLR) plays a key role in the innate immune system. Specifically, TLRs are one-way transmembrane receptors that recognize structurally conserved molecules derived from microorganisms. Besides, TLR signals by recruiting specific adaptor molecules, leading to the transcription factor NF-κB and IRF activation. TLRs are located on the cell surface or intracellular compartments, and they recognize different or overlapping PAMPs. Each TLR consists of an extracellular domain and a cytoplasmic Toll/IL-1 receptor (TIR) domain that initiates downstream signal transduction.

Moreover, TLR7 and TLR8 are members closely related to the phylogeny and structure of the TLR family. Both TLR7 and TLR8 have been identified as natural receptors for single-stranded RNA. Furthermore, they are effective activators of innate immune response after virus infection. The small molecule agonists of TLR7 and tlr8 have aroused great interest in cancer research because of their profound antitumor activity. Here, we will introduce a potent TLR7/8 agonist, CL097 (HY-128799).

First of all, CL097, a potent TLR7/8 agonist, induces pro-inflammatory cytokines in macrophages. Importantly, CL097 induces NADPH oxidase priming, resulting in an increase of the fMLF-stimulated ROS production.

In the second place, CL097 induces activation of NF-κB at 0.1 μM in TLR7 transfected HEK293 cells and at 4 μM in TLR8-transfected HEK293 cells. Particularly, CL097 induces hyperactivation of the NADPH oxidase by stimulating the phosphorylation of p47phox on selective sites in human neutrophils. It suggests that p38 MAPK, ERK1/2, protein kinase C, and Pin1 control this process. Obviously, CL097 induces the phosphorylation of p47phox on specific sites and enhances fMLF-induced p47phox phosphorylation.

Last but not the least, CL097 induces efficient diabetogenic Cytotoxic T lymphocyte (CTL) function in NOD mice. Besides, CL097 with 5 mg/kg by s.c. alone causes a modest specific lysis of the target peptide.

All in all, CL097, a potent TLR7/8 agonist, induces pro-inflammatory cytokines.