Background
Parbendazole is an inhibitor of microtubule assembly (EC50 = 3 ?M).1 It reduces viability of HeLa cells (EC50 = 0.53 ?M) and induces genotoxic stress in a reporter assay using HEK293 cells.2 Parbendazole reduces, but does not eradicate, worm burden in goats with naturally acquired or experimental nematode infections when administered at a dose of 15 mg/kg.3 It is teratogenic to lamb embryos when administered to pregnant ewes at a dose of 60 mg/kg during the third week of pregnancy.
1.Havercroft, J.C., Quinlan, R.A., and Gull, K.Binding of parbendazole to tubulin and its influence on microtubules in tissue-culture cells as revealed by immunofluorescence microscopyJ. Cell Sci.49195-204(1981)
2.Lo, Y.-C., Senese, S., France, B., et al.Computational cell cycle profiling of cancer cells for prioritizing FDA-approved drugs with repurposing potentialSci. Rep.7(1)(2017)
3.Charles, T.P., Pompeu, J., and Miranda, D.B.Efficacy of three broad-spectrum anthelmintics against gastrointestinal nematode infections of goatsVet. Parasitol.34(1-2)71-75(1989)
Protocol
Kinase experiment: | Pure tubulin is obtained from sheep brain by 2 cycles of assembly and disassembly in vitro. Immediately prior to use the protein is centrifuged at 130000 g for 30 min to remove any aggregates. It is used at a protein concentration of 0-2 mg/mL in 0.025 M Pipes buffer, 0-5 mM EGTA, 0-25 mM Mg2SOsup>4, 0.1 mM GTP. Drug binding is determined by equilibrium dialysis using concentrations of parbendazole between 0.1 μM and 4 μM, and 2% (v/v) DMF (dimethyl formamide) as a carrier. Equilibrium is achieved by constant stirring for 2 h at 26°C, bovine serum albumin being used as a standard. 200 μL aliquots are counted in PCS in a 25-200B liquid scintillation counter[2]. |
Cell experiment: | Vero cells, an established cell line derived from monkey kidney are seeded in DMEM supplemented with 10% (v/v) foetal calf serum onto glass coverslips in multiwell dishes. They are allowed to settle, and spread for 2-5 h in a humid atmosphere at 37°C. After this time the medium is changed to DMEM containing 2, 10 or 20 μM parbendazole and 1% (v/v) DMSO controls contained 1 % (v/v) DMSO or had no additions[2]. |
参考文献: [1]. Lo YC, et al. Computational Cell Cycle Profiling of Cancer Cells for Prioritizing FDA-Approved Drugs with Repurposing Potential. Sci Rep. 2017 Sep 12;7(1):11261.
[2]. Havercroft JC, et al. Binding of parbendazole to tubulin and its influence on microtubules in tissue-culture cells as revealed by immunofluorescence microscopy. J Cell Sci. 1981 Jun;49:195-204.
[3]. Foster KE, et al. A mutant beta-tubulin confers resistance to the action of benzimidazole-carbamate microtubule inhibitors both in vivo and in vitro. Eur J Biochem. 1987 Mar 16;163(3):449-55. |
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