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Research Area
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Description
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Prostate cancer,Breast cancer |
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Biological Activity
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Description
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Zibotentan (ZD4054) is a specific Endothelin A (ETA) antagonist with IC50 of 21 nM. |
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Targets
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Endothelin A (ETA) |
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IC50 |
21 nM [1] |
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In Vitro
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As Zibotentan specifically inhibits ETA-mediated antiapoptotic effects, but not ETB-mediated proapoptotic effects in human and rat smooth muscle cells, Zibotentan binds to endothelin A receptor (ETA) with high affinity with Ki of 13 nM, and has no affinity for endothelin B receptor (ETB) with IC50 of >10 μM. [1] Zibotentan treatment at 1 μM inhibits ET-1 induced mitogenic activity in ovarian carcinoma cell lines HEY and OVCA 433 secreting ET-1 and expressing ETA and ETB mRNA. [2] ZD4054 (1 μM) inhibits ET-1 induced EGFR transactivation in HEY and OVCA 433 cells. Zibotentan (1 μM) reverts ET-1 mediated epithelial-mesenchymal transition (EMT), by enhancing E-cadherin expression and promoter activity, and inhibiting vascular endothelial growth factor (VEGF) secretion and invasiveness in HEY and OVCA 433 cells. [3] Zibotentan also potently inhibits the basal and ET-1 induced cell proliferation in SKOV-3 and A-2780 cells, associated with the inhibition of AKT and p42/44MAPK phosphorylation, and with increased apoptosis through the inhibition of bcl-2 and activation of caspase-3 and poly(ADP-ribose) polymerase proteins. [4] |
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In Vivo
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Administration of Zibotentan at 10 mg/kg/day for 21 days potently inhibits the growth of HEY ovarian carcinoma xenografts in mice by 69% with no associated toxicity, which is in association with the blocking of cell proliferation evaluated by 37% inhibition of the Ki-67 expression, and the 62% inhibition of tumor-induced vascularization. Consistently, Zibotentan treatment significantly inhibits the expression of matrix metalloproteinase-2 (MMP-2) and VEGF, as well as the activation of p42/44 MAPK and EGFR, and potently enhances the expression of E-cadherin. [3] |
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Clinical Trials
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A Phase III study of Zibotentan in patients with hormone resistant prostate cancer (HRPC) and bone metastases has been completed. |
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Features
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Combination Therapy
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Description
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Zibotentan (1 μM) in combination with Gefitinib (1 μM) results in a greater inhibition of ET-1 induced EGFR, MAPK, and AKT phosphorylation in HEY and OVCA 433 cells, as well as a significant decrease in cell proliferation (65%), invasion (52%), and VEGF production (50%), accompanied by a 2-fold increase in apoptosis. Zibotentan administered i.p. at 10 mg/kg/day together with oral administration of Gefitinib at 125 mg/kg/day for 21 days significantly induce tumor growth inhibition by 82% of HEY xenografts, more potently compared with each drug treatment alone with ~69% tumor growth inhibition, and the growth delay in established tumors persists for up to 4 weeks after the termination of the combined treatment. The combination of Zibotentan and Gefitinib induces more significant inhibition of tumor neovascularization (78%) compared with either Zibotentan (62%) or Gefitinib (45%) treatment alone, and markedly decreased expression of Ki-67 by 46% versus 37% or 30%, which is associated with more potent inhibition of the expression/activation of MMP-2, VEGF, MAPK and EGFR, and the enhancement of E-cadherin expression, compare with single-agent treatment. [3] The combination of Zibotentan (10 mg/kg/day) and Paclitaxel (three 20 mg/kg i.v. doses every 4 days) produces additive antitumor effects in HEY ovarian cancer xenografts, with 40% of mice remaining tumor-free. [4] |
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Protocol
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Kinase Assay
[1]
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| Receptor-binding assays |
The inhibition by Zibotentan (varying concentrations) of 125iodine-ET-1 binding to cloned human ETA is assessed using standard radioligand-binding techniques. Human recombinant ETA is expressed in mouse erythroleukaemic cells, and cell membranes prepared for competitive binding studies using 125iodine-ET-1 as the radioligand. Incubations are carried out in triplicate in the presence of Zibotentan, 100 pM to 100 μM in half-log increments, and inhibition of ET-1 binding is expressed as the geometric mean pIC50 value (concentration to inhibit 50% of binding) with a 95% confidence interval (CI). The affinity of Zibotentan for cloned human ETA is also assessed using the equation of Cheng and Prusoff to determine the equilibrium dissociation constant (Ki) in a further receptor-binding screen utilizing a greater number of concentration-response curves determined in three separate studies. |
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Cell Assay
[3]
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| Cell Lines |
HEY and OVCA 433 |
| Concentrations |
Dissolved in DMSO, final concentrations 1 μM |
| Incubation Time |
48 hours |
| Methods |
Cells are serum starved by incubation for 24 hours in serum-free DMEM before exposed to Zibotentan for 48 hours. After the treatment, cells are lysed and the supernatant is recovered and assayed for histone-associated DNA fragments, at 405 nm by the use of a microplate reader. For detection of early apoptotic events, floating and adherent cells are collected. Cells are double stained with FITC-conjugated Annexin V and propidium iodide using the Vybrant Apoptosis Kit and are immediately analyzed by cytofluorometric analysis. |
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Animal Study
[3]
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| Animal Models |
Female athymic (nu+/nu+) mice bearing established HEY human ovarian carcinoma xenografts |
| Formulation |
Dissolved in DMSO, and diluted in PBS |
| Doses |
10 mg/kg/day |
| Administration |
Treated i.p. |
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References |
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[1] Morris CD, et al. Br J Cancer, 2005, 92(12), 2148-2152.
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[2] Rosanò L, et al. Exp Biol Med (Maywood), 2006, 231(6), 1132-1135.
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[3] Rosanò L, et al. Cancer Res, 2007, 67(13), 6351-6359.
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[4] Rosanò L, et al. Mol Cancer Ther, 2007, 6(7), 2003-2011.
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