Cinobufotalin

Tumour Biol. 2015 Feb 28.

Ceramide production mediates cinobufotalin-induced growth inhibition and apoptosis in cultured hepatocellular carcinoma cells.[Pubmed: 25724183]

Hepatocellular carcinoma (HCC) is a highly aggressive and lethal neoplasm with poor prognosis. The aim of this study is to investigate the anticancer activity of Cinobufotalin, a bufadienolide isolated from toad venom, in cultured HCC cells, and to study the underlying mechanisms.
METHODS AND RESULTS:
We found that Cinobufotalin (at nmol/L) significantly inhibited HCC cell growth and survival while inducing considerable cell apoptosis. Further, Cinobufotalin inhibited sphingosine kinase 1 (SphK1) activity and induced pro-apoptotic ceramide production. Ceramide synthase-1 small hairpin RNA (shRNA)-depletion inhibited Cinobufotalin-induced ceramide production and HCC cell apoptosis. On the other hand, the glucosylceramide synthase (GCS) inhibitor 1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP) facilitated Cinobufotalin-induced ceramide production and cell apoptosis. SphK1 inhibitor II (SKI-II), similar to Cinobufotalin, increased cellular ceramide level and promoted HCC cell apoptosis. Finally, we observed that Cinobufotalin inactivated Akt-S6K1 signaling in HepG2 cells, which was again inhibited by ceramide synthase-1 shRNA-depletion.
CONCLUSIONS:
In conclusion, the results of this study suggest that Cinobufotalin induces growth inhibition and apoptosis in cultured HCC cells through ceramide production. Cinobufotalin may be investigated as a novel anti-HCC agent.

Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2014 Oct;22(5):1306-10.

[Reversal effect of cinobufacini on multidrug resistance of Raji/ADR cells and its mechanisms].[Pubmed: 25338578]

The aim of this study was to explore the reversing effect of cinobufacini on multidrug resistance of Raji/ADR cells and its mechanisms.
METHODS AND RESULTS:
The growth inhibitory rate, half inhibitory concentration (IC50), reversing multiples to adriamycin- resistance were detected by MTT, and the curve of growth inhibitory rate was drawn; the MDR-1 and MRP-1 gene transcription was determined by RT-PCR; the expressions of P-gp and MRP-1 proteins were assayed by Western blot and flow cytometry. The results showed that the inhibitory rates of cinobufacini on Raji and Raji/ADR cells at 72 h were 75.6% and 69.3% respectively, the IC50 were 3.9 mmol/L and 4.6 mmol/L without significant difference (P > 0.05). The reversing multiples to adriamycin-resistance were 255.7 multiples, the transcription of mdr-1 and mrp-1 genes and the expression of P-gp and MRP-1 proteins significantly decreased (P < 0.05) in Raji/ADR cells after the treatment with Cinobufotalin.
CONCLUSIONS:
It is concluded that Cinobufotalin can reverse the adriamycin-resistance in Raji/ADR cells and the expression of P-gp and MRP-1 proteins were down-regulated through the transcriptional pathway. The Cinobufotalin is an effective reversal agent for the multidrug resistance of tumors.

Life Sci. 1998;63(9):781-8.

Neutralization of cardiac toxins oleandrin, oleandrigenin, bufalin, and cinobufotalin by digibind: monitoring the effect by measuring free digitoxin concentrations.[Pubmed: 9740315]

Oleandrin plant poisoning is common in children and the plant extract is used in Chinese medicines. The toxicity is due to oleandrin and the deglycosylated metabolite oleandrigenin. Bufalin and Cinobufotalin (toad cardiac toxins) are also widely used in Chinese medicines like Chan SU, and Lu-Shen -WU. Severe toxicity from bufalin after consumption of toad soup has been reported.
METHODS AND RESULTS:
Taking advantage of structural similarities of these toxins with digitoxin, we demonstrated that these compounds can be rapidly detected in blood by the fluorescence polarization immunoassay for digitoxin. The cross reactivities of these compounds with digoxin assay were much lower. For example, when a drug free serum was supplemented with 10 microg/ml of oleandrin, we observed 127.7 ng/ml of digitoxin equivalent but only 2.4 ng/ml of digoxin equivalent concentration. Digibind neutralized all cardiac toxins studied as evidenced by significant fall of free concentrations. When aliquots of serum pool containing 50.0 microg/ml of oleandrin were supplemented with 0, 10.0, 25.0, 50.0, 100, and 200 microg/ml of digibind, the mean free concentrations were 30.6, 23.3, 16.0, 10.7, 7.8 and 5.5 microg/ml respectively. Similarly, with 50.0 microg/ml of oleandrigenin (total concentration: 36.2 ng/ml), the free concentration was 14.5 ng/ml digitoxin equivalent in the absence of digibind and 5.4 ng/ml in the presence of 200 microg/ml of digibind. In another specimen containing 500 ng/ml bufalin (total concentration: 156.9 ng/ml), the free concentration was 8.6 ng/ml in the absence of digibind and none detected in the presence of 100.0 microg/ml digibind.
CONCLUSIONS:
Because such neutralization may also occur in vivo, digibind may be useful in treating patients exposed to these toxins.

Pharmacology & Clinics of Chinese Materia Medica, 2007, 23(4):54-6.

Effect of cinobufotalin on the dendritic cells derived from peripheral blood of patients with chronic hepatitis B[Reference: WebLink]

To study the effect of Cinobufotalin on the morphous,phenotype and function of dendritic cells derived from peripheral blood of patients with chronic hepatitis B(CHB).
METHODS AND RESULTS:
All investigated people were divided into three group: normal control group, CHB group and CHB+Cinobufotalin (CHB+Cino) group. Mononuclear cells from the peripheral blood of patients with CHB and healthy adults were isolated and were cultured in the medium contained GM-CSF(100ng/ml) and IL-4(500u/ml) to induce DCs. After cultured 48 h, Cinobufotalin were added into the culture media of DCs of CHB+Cino grou. On culture day 7, the morphous of DCs was observed and taken pictures; the expression of CD40, HLA-DR, CD80 and CD86 molecules on DCs were detected by flow cytometer; the DCs' capacities of stimulating allogeneic lymphocyte proliferation was assayed with cell count kit 8(CCK8); the concentration of IL-12 in the supernatants of DCs' cultures was detected by ELISA.Compared with healthy controls, the DCs derived from peripheral blood of the patients of CHB group displayed immature cell morphous and had lower expression of surface molecules and lower capacities of stimulating allogeneic lymphocyte proliferation and secreting IL-12. But compared with the CHB group, the DCs incubated with Cinobufotalin( CHB+Cino group) displayed mature cell appearance like normal people's DCs and had higher expression of surface molecules including CD40, CD80 and CD86, and the abilities of stimulating allogeneic lymphocyte proliferation and secreting IL-12 of the DCs of CHB+Cino group were obviously higher.
CONCLUSIONS:
Cinobufotalin can promote the DCs derived from peripheral blood of patients with chronic hepatitis B to mature and effectively enhance its(the DCs') capabilities, therefore the treatment of Cinobufotalin may potentiate the antiviral immunity of the patients with CHB.