IDO1/TDO-IN-4 displays potent inhibition of both IDO1 (IC50 = 3.53 μM) and TDO (IC50 = 1.15 μM) and has an acceptable safety profile and pharmacokinetic properties. IDO1/TDO-IN-4 forms a hydrogen bond with IDO1, and π π stacking interaction with TDO. IDO1/TDO-IN-4 can be used in the research of depression and inflammation-induced depression [1].

IDO1/TDO-IN-4 (compound 28, 0-2 µM, 1 h) effectively suppresses the activation of LPS-induced BV2 microglial cells, as indicated by morphological changes, reducing pro-inflammatory agent production, and enhancing IL-10 expression. It also diminishes IDO1 expression, blocking excessive tryptophan breakdown via the kynurenine pathway. RT-PCR analysis shows that, within these concentrations and incubation time, IDO1/TDO-IN-4 reduces COX2, iNOS, TNF-α, and IL-1β synthesis while elevating IL-10 levels in 100 ng/mL LPS-stimulated BV2 microglial cells.

IDO1/TDO-IN-4, also known as compound 28, administered at 20 mg/kg intraperitoneally on days 1, 2, and 3, effectively mitigates LPS-induced neuroinflammation and depressive-like behavior in mice, demonstrating significant reduction in microglial activation and inflammatory markers (TNF-α, IL-1β, and iNOS) within the hippocampus, and suppressing the LPS-induced upregulation of IDO1. In pharmacokinetic assessments in male C57BL/6J mice, IDO1/TDO-IN-4, regardless of being administered intraperitoneally or intravenously, showed high exposure and volume of distribution at steady state, with notable pharmacokinetic parameters including a half-life (T 1/2) of 2.31 hours (i.v.) and 0.77 hours (i.p.), maximum concentration (C max) of 5543.99 ng/mL (i.v.) and 3878 ng/mL (i.p.), and a bioavailability (F %) of 52.55.