E6446 (20 mg/kg, pO.) almost entirely inhibits the production of IL-6 induced by CpG1668, and in a dose-dependent manner, diminishes the development of ANA (anti-nuclear antibodies) in mice at levels of 20 and 60 mg/kg^[1].

E6446 (20, 60 mg/kg, pO.) inhibits TLR9 signaling in mice in a dose-dependent fashion. At doses of 60, 120 mg/kg, orally, it prevents TLR and LPS-induced septic shock hyperresponsiveness in rodent malaria, reduces TLR responsiveness in acute malaria, and inhibits activation of both TLR7 and TLR9^[2].

E6446 is a powerful and orally active inhibitor of TLR7 and TLR9. It effectively inhibits DNA activation of HEK:TLR9 cells with an IC50 of 10 nM, yet its efficacy is significantly reduced in suppressing LPS endotoxin activation of HEK:TLR4 cells or R848 activation of HEK:TLR7 cells. E6446 strongly reduces IL-6 production triggered by CpG2216 but does not affect IL-6 production induced by the TLR3 ligand poly inosine-cytosine. The inhibitory effect of E6446 on TLR7 varies with the ligand; it is a strong inhibitor of IL-6 production induced by RNA but less effective against IL-6 induction by the small molecule imidazoquinoline ligand R-848. E6446 inhibits the TLR9-DNA interaction in vitro, with an IC50 ranging from 1 to 10 µM^[1].

E6446 (0.01-0.03 μM) specifically blocks TLR9 activation by CpG ODN 2006, and inhibits TLR7/8 activation by the imidazoquinoline compound R848 at concentrations of 2-8 μM. It diminishes TLR4 activation by 50% at 30 μM, exhibiting IC50 values of 0.01 μM in HEK-TLR9 cells activated with oligo 2006 and 0.23 μM in human PBMCs activated with oligo 2216, respectively^[2].