Identification of natural compounds targeting Annexin A2 with an anti-cancer effect

Annexin A2, a multifunctional tumor associated protein, promotes nuclear factor-kappa B (NF-κB) activation by interacting with NF-κB p50 subunit and facilitating its nuclear translocation. Here we demonstrated that two ginsenosides Rg5 (G-Rg5) and Rk1 (G-Rk1), with similar structure, directly bound to Annexin A2 by molecular docking and cellular thermal shift assay. Both Rg5 and Rk1 inhibited the interaction between Annexin A2 and NF-κB p50 subunit, their translocation to nuclear and NF-κB activation. Inhibition of NF-κB by these two ginsenosides decreased the expression of inhibitor of apoptosis proteins (IAPs), leading to caspase activation and apoptosis. Over expression of K302A Annexin A2, a mutant version of Annexin A2, which fails to interact with G-Rg5 and G-Rk1, effectively reduced the NF-κB inhibitory effect and apoptosis induced by G-Rg5 and G-Rk1. In addition, the knockdown of Annexin A2 largely enhanced NF-κB activation and apoptosis induced by the two molecules, indicating that the effects of G-Rg5 and G-Rk1 on NF-κB were mainly mediated by Annexin A2. Taken together, this study for the first time demonstrated that G-Rg5 and G-Rk1 inhibit tumor cell growth by targeting Annexin A2 and NF-κB pathway, and G-Rg5 and G-Rk1 might be promising natural compounds for targeted cancer therapy.

Ginsenoside Rk1 suppresses pro-inflammatory responses in lipopolysaccharide-stimulated RAW264.7 cells by inhibiting the Jak2/Stat3 pathway / 中国天然药物

The saponin ginsenoside Rk1 is a major compound isolated from ginseng. Ginsenoside Rk1 has been reported to have anti-inflammatory and anti-tumor properties and to be involved in the regulation of metabolism. However, the effect and mechanism of anti-inflammatory action of ginsenoside Rk1 has not been fully clarified. We investigated whether ginsenoside Rk1 could suppress the inflammatory response in lipopolysaccharide-stimulated RAW264.7 macrophages and to explore its mechanism of the action. RAW264.7 cells were treated with LPS (1 μg·mL) in the absence or the presence of Ginsenoside Rk1 (10, 20, and 40 μmol·L). Then the inflammatory factors were tested with Griess reagents, ELISA, and RT-PCR. The proteins were analyzed by Western blotting. Ginsenoside Rk1 inhibited lipopolysaccharide-induced expression of nitric oxide (NO), interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α, and monocyte chemotactic protein (MCP)-1. Ginsenoside Rk1 inhibited the lipopolysaccharide-stimulated phosphorylation of NF-κB and janus kinase (Jak)2 and signal transducer and activator of transcription (Stat)3 at Ser727 and Tyr705. These data suggested that ginsenoside Rk1 could inhibit expression of inflammatory mediators and suppress inflammation further by blocking activation of NF-κB and the Jak2/Stat3 pathway in LPS-stimulated RAW264.7 cells.

Simultaneous Determination of Four Kinds of Rare Saponins in Black Ginseng by HPLC / 中国药学杂志

OBJECTIVE: To establish an HPLC method for determining four kinds of rare saponins, ie, 20(S)-Rg3, 20(R)-Rg3, Rk1, and Rg5 in black ginseng. METHODS: COSMOSIL C18-PAQ (4.6 mm×250 mm, 5 μm) column was used and temperature was maintained at 30℃. Gradient elution was conducted using mobile phase consisting of acetonitrile and water at a flow rate of 1.0 mL·min-1. The detection wavelength was set at 203 nm. The injected sample volume was 10 μL. RESULTS: Good resolution was achieved for the four rare saponins in the ranges of 0.051-0.256 (r=0.999 9), 0.009-0.280(r=0.999 7), 0.051-0.303(r=0.999 9) and 0.093-0.279 mg·mL-1(r=0.999 9) for 20(S)-Rg3, 20(R)-Rg3, Rk1, and Rg5, respectively. The corresponding average recovery rates were 103.9%, 99.2%, 97.0%, and 100.6%, and the standard deviations were 0.71%, 0.73%, 1.97%, and 0.57%, respectively. CONCLUSION: The method is accurate, simple, reliable and reproducible for the determination of saponins in black ginseng. The determination result can be used as a reference for the rational medication, quality control, and further study of black ginseng.

Identification of saponins from Panax notoginseng in metabolites of rats / 中国中药杂志

UPLC-QTOF-MS/MS was used to identify metabolites in rat blood, urine and feces after the administration of n-butanol extract derived from steamed notoginseng. The metabolic process of saponins came from steamed notoginseng was analyzed. The metabolites were processed by PeakView software, and identified according to the structural characteristics of prototype compounds and the accurate qualitative and quantitative changes of common metabolic pathways. Four saponins metabolites were identified based on MS/MS information of metabolites, namely ginsenoside Rh₄, Rk₃, Rk₁, Rg₅,and their 15 metabolites were verified. The metabolic pathways of the four ginsenosides in n-butanol extract included glucuronidation, desugar, sulfation, dehydromethylation, and branch loss. The metabolites of main active saponin components derived from steamed Panax notoginseng were analyzed from the perspective of qualitative analysis. And the material basis for the efficacy of steamed notoginseng was further clarified.

A new triterpene natural product from stems and leaves of Panax ginseng / 中草药

Objective: To study the chemical constituents of saponins in the stems and leaves of Panax ginseng. Methods: The chemical constituents were isolated and purified by various chromatographic methods, and their structures were identified by NMR and MS data analysis. Results: Nine compounds were isolated and identified as 3β,6α,12β,25-tetrahydroxy-dammar-E-20(22)-ene-6-O-α-L-rhamnopyranosyl-(1→2)-β-D-glucopyranoside (1), sanchinoside B1 (2), 3β,6α,12β-dammar-E-20(22)-ene-3,6,12,25-tetraol (3), ginsenoside Rk3 (4), ginsenoside Rh4 (5), notoginsenoside T2 (6), 3β,6α,12β-dammar-20(21),24-diene-3,6,12-triol (7), ginsenoside Rk1 (8), and ginsenoside Rg5 (9). Conclusion: Compound 1 is a new natural product and the other eight compounds are all isolated from the stems and leaves of P. ginseng for the first time.

The effect of ginsenoside Rk1 in junctional protein of severe preeclamptic placenta / 대한산부인과학회지

OBJECTIVE: To investigate the differential expression of junctional proteins in the normal and preeclamptic human placenta and the effect of ginsenoside Rk1 in junctional proteins. METHODS: Placental tissues from 10 women with severe preeclampsia and 5 normal women were collected at the time of their cesarean section. Five of 10 preeclamptic women were complicated with intrauterine growth restriction (IUGR). Immunohistochemistry and Western blotting was employed to localize junctional proteins (zo-1, occludin and plakoglobin) positive cells. The placental explant culture was performed to investigate if Rk1 can attenuate the expression of junctional proteins (zo-1, occluding and plakoglobin) induced by deferoxamine-induced hypoxia. Rk1 was treated at the day 3 and Western blot analysis was performed for protein quantification. RESULTS: There was no different expression of zo-1 and plakoglobin among all the study groups. Occludin showed negative at the endothelial cells of the terminal villi in both normal and preeclampsia groups. At the endothelial cells of the stem villi, occludin was detected in both normal and severe preeclamptic placenta with normal fetal growth. However, severe preeclampsia with IUGR were decreased expression of occludin at the endothelial cells of the stem villi. When we administered Rk1 to the placenta treated with DFO, expression of occludin was not different. CONCLUSION: The placental expression of zo-1 and plakoglobin were not different among the study groups, while that of occludin was significantly decreased at the endothelium of stem villi in severe preeclampsia with IUGR. Rk-1 showed no effect on the placental junctional proteins. These results suggest that occludin may play a role in pathophysiology of fetal growth restriction in utero.