Anti-neuroinflammatory polyprenylated acylphloroglucinols from Hypericum patulum: LC-MS/MS-guided isolation, semi-synthesis, and TLR4 inhibition.

Alzheimer's disease (AD) is a serious disease that poses a major threat to human health. Up till now, there is no effective drug on the market that could completely cure AD. In this study, 45 polycyclic polyprenylated acylphloroglucinol (PPAP) analogs, including 13 new ones, were isolated from Hypericum patulum by LC-MS/MS guided isolation with molecular networking analysis. Architecturally, all isolates consist of 12 cage-like PPAPs, 24 type-A PPAPs, four seco-PPAPs, three spiroType PPAPs, and two adamantane-type PPAPs. Semi-synthesis was applied for the first time to assign the relative and absolute configuration of those PPAPs (1-8) with an oxidized prenyl group. Those compounds were identified by various spectral data, quantum chemical calculations, and single crystal X-ray diffraction. Fourteen compounds exhibited better NO inhibitory effect with IC50 values of 1.97 ± 0.44-13.97 ± 1.45 μM than that of the positive control, minocycline (IC50: 14.09 ± 1.14) in BV-2 cells. The structure activity relationship was also discussed. Further mechanistic study for a high content compound (4) implied that this compound could suppress the expression of TLR4, prohibit TLR4/NF-κB/JNK signaling pathway, and reduce iNOS and COX-2 expression. Compound 4 could directly bind to TLR4 via cellular thermal shift assay (CETSA), surface plasmon resonance assay (SPR), and molecular docking. What's more, compound 4 exhibited remarkable anti-AD effects without toxicity in zebrafish model. Thus, compound 4 is a potential anti-AD lead compound by targeting TLR4.