[Inhibitory Effect of Cinobufotalin on Macrophage Inflammatory Factor Storm and Its Mechanism].

OBJECTIVE: To investigate the inflammatory effects of Cinobufotalin on monocytes in resting state and macrophages in activated state and its molecular mechanism. METHODS: THP-1 cells were stimulated with Phorbol 12-myristate 13-acetate to induce differentiation into macrophages. Lipopolysaccharides was added to activate macrophages in order to establish macrophage activation model. Cinobufotalin was added to the inflammatory cell model for 24 h as a treatment. CCK-8 was used to detect cell proliferation, Annexin V /PI double staining flow cytometry was used to detect cell apoptosis, flow cytometry was used to detect macrophage activation, and cytometric bead array was used to detect cytokines. Transcriptome sequencing was used to explore the gene expression profile regulated by Cinobufotalin. Changes in the significantly regulated molecules were verified by real-time quantitative polymerase chain reaction and Western blot. RESULTS: 1∶25 concentration of Cinobufotalin significantly inhibited the proliferation of resting monocytes(P<0.01), and induced apoptosis(P<0.01), especially the activated macrophages(P<0.001, P<0.001). Cinobufotalin significantly inhibited the activation of macrophages, and significantly down-regulated the inflammatory cytokines(IL-6, TNF-α, IL-1ß, IL-8) released by activated macrophages(P＜0.001). Its mechanism was achieved by inhibiting TLR4/MYD88/P-IκBa signaling pathway. CONCLUSION: Cinobufotalin can inhibit the inflammatory factors produced by the over-activation of macrophages through TLR4/MYD88/P-IκBa pathway, which is expected to be applied to the treatment and research of diseases related to the over-release of inflammatory factors.

Facile Synthesis and Cytotoxicity of Phenazine-Chromene Hybrid Molecules Derived from Phenazine Natural Product.

AIM AND OBJECTIVE: Small molecule targeted drugs can effectively reduce the toxicity and side effects of drugs, and improve the efficacy of drugs by their specific antitumor activity. Hence, the development of small molecular targeted drugs for cancer has important significance. This study was undertaken to design and synthesize novel phenazine-chromene hybrid molecules in order to optimize the structure and improve the efficacy of this kind of hybrids. MATERIALS AND METHODS: O-diaminobenzene was used as starting material to synthesize twentyfour heterocyclic compounds designed as hybrid molecules of phenazine and 4H-chromene pharmacophores by facile methods. The structures of the compound were confirmed by 1H NMR, 13C NMR and HRMS. Furthermore, the synthesized compounds were evaluated for in vitro activity against four human cancer cell lines and two non-cancer cell lines by MTT test. RESULTS: Some compounds showed strong cytotoxic activities against HepG2 and A549 cancer lines (IC50 = 5-10 µM). Comparing 2i with 2l, the introduction of hydrophilic groups on the phenazine core could not improve the antiproliferative activity significantly. Except 2d and 3c, compounds owning chlorine substituent on the 4H-chromene pharmacophore seemingly contribute to enhance the compounds' antiproliferative activity. Specially, compound 3c showed highest cytotoxicity against A549 cells with IC50 values of 3.3±0.4 µM. Furthermore, all compounds showed low or no cytotoxicity against HUVEC and L02 non-cancer cells in vitro. CONCLUSION: Compound 3c may be used as potential lead molecule against A549 cancer cells.