Discovery of anti-neuroinflammatory agents from 1,4,5,6-tetrahydrobenzo[2,3]oxepino[4,5-d]pyrimidin-2-amine derivatives by regulating microglia polarization.

Neuroinflammation mediated by microglia activation leads to various neurodegenerative and neurological disorders. In order to develop more and better options for this disorders, a series of 3,4-dihydrobenzo[b]oxepin-5(2H)-one derivatives (BZPs, 6-19) and novel 1,4,5,6-tetrahydrobenzo[2,3]oxepino[4,5-d]pyrimidin-2-amine derivatives (BPMs, 20-33) were synthesized and screened the anti-neuroinflamamtion effects. 3,5-bis-trifluoromethylphenyl-substituted BPM 29 showed more potent anti-neuroinflammatory activity and no toxicity to BV2 microglia cells in vitro. 29 significantly reduced the number of M1 phenotype of microglia cells, but significantly increased the number of M2 phenotype of microglia cells in lipopolysaccharide (LPS)-induced BV2 microglia cells. 29 significantly reduced the secretion of inflammatory cytokines (IL-18, IL-1ß, TNF-α), but increased the secretion of anti-inflammatory cytokines (IL-10) from LPS-induced BV2 microglia cells. Also, 29 inhibited the NOD-like receptor NLRP3 inflammasome formation, and down-regulated the expression of M2 isoform of pyruvate kinase in LPS-induced BV2 microglia cells. In vivo, 29 reduced the neuroinflammation in cuprizone-induced inflammatory and demyelinating mice by reducing the expression of inducible nitric-oxide synthase, but increased the expression of CD206. Taken together, 29 might be a prospective anti-neuroinflammatory compound for neuroinflammatory and demyelinating disease by alleviating microglia activation.

Discovery of anti-hepatoma agents from 1,4,5,6,7,8-hexahydropyrido[4,3-d]pyrimidine by inhibiting PI3K/AKT/NF-κB pathway activation.

In order to obtain new anti-hepatoma drugs with low toxicity, some 1,4,5,6,7,8-hexahydropyrido[4,3-d]pyrimidines (PPMs, 4a-t) were synthesized in this study. Many of them showed significant anti-hepatoma effects against HCC cells and low toxicity toward HHL-5 cells. Combined with their anti-hepatoma activity and toxicity, 4-CF3-substituted 4k was selected as an effective lead compound. Preliminary mechanistic studies revealed that 4k could up-regulate the expression levels of Bax and caspase-3 proteins, down-regulate the expression levels of Bcl-2 protein, promote significant apoptosis of HepG2, and block cells in G2-M phase to prevent cells from completing mitosis. Also, 4k could significantly inhibit the activation of PI3K/AKT/NF-κB pathway by blocking the phosphorylation of PI3K, AKT, NF-κB/p65 and IFN-γ-induced nuclear transport. Docking analysis showed that 4k could reasonably bind to the active sites of Bcl-2, NF-κB/p65, PI3K and AKT. This result suggested that 4k could be used as a new type of NF-κB inhibitor, which provides a scientific basis for further research into the treatment of hepatoma.

Trifluoromethyl-substituted 3,5-bis(arylidene)-4-piperidones as potential anti-hepatoma and anti-inflammation agents by inhibiting NF-кB activation.

Some methoxy-, hydroxyl-, pyridyl-, or fluoro-substituted 3,5-bis(arylidene)-4-piperidones (BAPs) could reduce inflammation and promote hepatoma cell apoptosis by inhibiting activation of NF-κB, especially after introduction of trifluoromethyl. Herein, a series of trifluoromethyl-substituted BAPs (4-30) were synthesised and the biological activities were evaluated. We successfully found the most potential 16, which contains three trifluoromethyl substituents and exhibits the best anti-tumour and anti-inflammatory activities. Preliminary mechanism research revealed that 16 could promote HepG2 cell apoptosis in a dose-dependent manner by down-regulating the expression of Bcl-2 and up-regulating the expression of Bax, C-caspase-3. Meanwhile, 16 inhibited activation of NF-κB by directly inhibiting the phosphorylation of p65 and IκBα induced by LPS, together with indirectly inhibiting MAPK pathway, thereby exhibiting both anti-hepatoma and anti-inflammatory activities. Molecular docking confirmed that 16 could bind to the active sites of Bcl-2, p65, and p38 reasonably. The above results suggested that 16 has enormous potential to be developed as a multifunctional agent for the clinical treatment of liver cancers and inflammatory diseases.

Discovery of novel NF-кB inhibitor based on scaffold hopping: 1,4,5,6,7,8-hexahydropyrido[4,3-d]pyrimidine.

NF-κB is a key signaling pathway molecule linking hepatoma and chronic inflammation. Inhibition of NF-κB activation can alleviate inflammation, and promote hepatoma cell apoptosis. In this study, a series of fluoro-substituted 1,4,5,6,7,8-hexahydropyrido[4,3-d]pyrimidines (PPMs, 31-57) were synthesized from 3,5-bis(arylidene)-4-piperidones (BAPs, 4-30) based on scaffold hopping. We successfully discovered the most potent 43 substituted by electron-withdrawing substitutes (3-F and 4-CF3) exhibited less toxicity and higher anti-inflammatory activity. Preliminary mechanistic studies revealed that 43 induced dose-dependent cell apoptosis at cell and protein level, while inhibited NF-κB activation by suppressing LPS-induced phosphorylation levels of p65, IκBα and Akt, and by indirectly suppressing MAPK signaling, and by inhibiting the nuclear translocation of NF-κB induced by TNF-α or LPS. Docking analysis verified simulated 43 could reasonably bind to the active site of Bcl-2, p65 and p38 proteins. This compound, as a novel NF-κB inhibitor, also demonstrated both anti-inflammatory and anti-hepatoma activities, warranting its further development as a potential multifunctional agent for the clinical treatment of liver cancers and inflammatory diseases.