CL097 hydrochloride is a potent TLR7 and TLR8 agonist. CL097 hydrochloride induces pro-inflammatory cytokines in macrophages and NADPH oxidase priming, thereby increasing the fMLF-stimulated ROS production [2].

CL097 effectively activates NF-κB at concentrations of 0.1 μM in TLR7-transfected HEK293 cells and 4 μM in TLR8-transfected HEK293 cells [1]. It also triggers a hyperactivation of NADPH oxidase in human neutrophils, primarily by promoting phosphorylation of p47phox at specific sites. This activation suggests the involvement of p38 MAPK, ERK1/2, protein kinase C, and Pin1 in regulating the process. Additionally, CL097 not only induces phosphorylation of p47phox on these specific sites but also amplifies fMLF-induced phosphorylation of p47phox [2]. Western blot analysis on neutrophils, with concentrations ranging from 0 to 10 μg/mL and a pretreatment duration of 30 minutes, demonstrated a concentration-dependent increase in p47phox phosphorylation [2].

Stimulation with CL097 and a CD40 agonist enhances diabetogenic cytotoxic T lymphocyte (CTL) functionality in NOD mice. When administered subcutaneously (s.c.) at a dosage of 5 mg/kg, CL097 alone elicits a specific target peptide lysis rate of approximately 25%. However, the concurrent administration of CL097 (5 mg/kg, s.c.) and a CD40 agonist (10 mg/kg, intraperitoneally (i.p.)) significantly enhances the specific lysis, doubling the efficacy seen with CL097 alone against IGRP-peptide-coated targets. This study utilized female 8.3 NOD mice aged 5-6 weeks, highlighting a potentiated immune response through combined treatment.