HTS assays for tubulin and tubulin associated proteins
With microtubules playing a central and vital role in cell division as well as vesicle and cargo transport in cells, targeting microtubule dynamics is a well-established area of drug discovery. Cytoskeleton, Inc. are the world leaders at providing the scientific community with tubulin proteins and tubulin related assays.
The optical density based tubulin polymerization assay (Cat. # BK004P) is well established in the pharmaceutical sector and is highly referenced in scientific literature. It is supported by patented technology for lyophilized tubulin and each customer obtains a free license to use these preparations for screening and drug development purposes. The BK006P tubulin polymerization assay uses highly purified tubulin, which is a good secondary screen for determining IC50s and tubulin specificity. The fluorescence tubulin polymerization assay (Cat. # BK011P) is less established in the field but is a good alternative for HTS because of the price savings over the absorbance based assay.
For more selective anti-cancer drug development a very exciting secondary screen can be achieved using Cancer Cell Line Tubulin. Currently, HeLa and MCF-7 cell line tubulins can be used in the fluorescence polymerization assay (Cat. # H001 and H005, respectively). It has been shown that anti-cancer drugs have different affinities for these tubulins compared to bovine brain (neuronal) tubulin. This indicates that a drug development program aimed at increasing the affinity for cancer cell line tubulin over neuronal tubulin may lead to greater targeting efficiency toward cancer cells.
We also provide HTS assays for other types of tubulins, such as fungal tubulins, plant tubulins or bacterial tubulin analogs. If you are interested in assaying a specific tubulin not listed on our website, please contact our technical support to discuss how we can provide you with the specific tubulin type you want.
Cytoskeleton's Tubulin and MAPs products have been cited hundreds of times over the past 18 years. A select few are described here, for more citations on individual products please use the "Citations" tab on each individual product page.
Tubulin polymerization HTS assay using >99% pure tubulin, OD based - Porcine (Cat. # BK006P)
O'Boyle NM, Carr M, Greene LM, Bergin O, Nathwani SM, McCabe T, Lloyd DG, Zisterer DM, Meegan MJ. (2010). Synthesis and Evaluation of Azetidinone Analogues of Combretastatin A-4 as Tubulin Targeting Agents.. J Med Chem.
Jacob Kushkuley, Walter K. H. Chan, Sangmook Lee, Joel Eyer, Jean-Francois Leterrier, Franck Letournel and Thomas B. Shea (2009). Neurofilament cross-bridging competes with kinesin-dependent association of neurofilaments with microtubules. J Cell Science 122 ,3579-86.
Chen, Z., Merta, P. J., Lin, N. H., Tahir, S. K., Kovar, P., Sham, H. L. and Zhang, H. (2005). A-432411, a novel indolinone compound that disrupts spindle pole formation and inhibits human cancer cell growth. Mol. Cancer Ther. 4, 562-568.
Huang, Y. T., Huang, D. M., Guh, J. H., Chen, I. L., Tzeng, C. C. and Teng, C. M. (2005). CIL-102 interacts with microtubule polymerization and causes mitotic arrest following apoptosis in the human prostate cancer PC-3 cell line. J. Biol. Chem. 280, 2771-2779.
Rouzier, R., Rajan, R., Wagner, P., Hess, K. R., Gold, D. L., Stec, J., Ayers, M., Ross, J. S., Zhang, P., Buchholz, T. A. et al. (2005). Microtubule-associated protein tau: A marker of paclitaxel sensitivity in breast cancer. Proc. Natl. Acad. Sci. U. S. A. 102, 8315-8320.
Jiang, J. D., Denner, L., Ling, Y. H., Li, J. N., Davis, A., Wang, Y., Li, Y., Roboz, J., Wang, L. G., Perez-Soler, R. et al. (2002). Double blockade of cell cycle at G1-S transition and M phase by 3-iodoacetamido benzoyl ethyl ester, a new type of tubulin ligand. Cancer Res. 62, 6080-6088.
Mooberry, S. L., Tien, G., Hernandez, A. H., Plubrukarn, A. and Davidson, B. S. (1999). Laulimalide and isolaulimalide, new paclitaxel-like microtubule-stabilizing agents. Cancer Res. 59, 653-660.
Tubulin ligand competition assay (Cat. # CDS15)
Davis, P. D., Dougherty, G. J., Blakey, D. C., Galbraith, S. M., Tozer, G. M., Holder, A. L., Naylor, M. A., Nolan, J., Stratford, M. R., Chaplin, D. J. et al. (2002). ZD6126: a novel vascular-targeting agent that causes selective destruction of tumor vasculature. Cancer Res. 62, 7247-7253.
Question 1: What is the most economical way to screen for anti-microtubule compounds?
Answer 1: Based on Cytoskeleton’s many years of experience in working with cytoskeletal proteins such as tubulin and our recent innovation of the tubulin polymerization assay, we recommend the fluorescence-based tubulin polymerization assay kit (Cat. # BK011P) for characterizing and screening anti-microtubule compounds. This kit provides all of the necessary reagents and buffers to perform up to 96 reactions in the included 96 well plate. The fluorescence-based assay is very economical because it requires 1/3 the amount of tubulin (100 ug) and 1/2 the reaction volume (50 ul) of traditional absorbance-based polymerization assays. Besides being able to do more reactions with the same amount of 99% pure tubulin, the fluorescence assay also offers twice the sensitivity of the absorbance-based assay with a lower coefficient of variation (11% vs 13%).
Question 2: How can we selectively target a particular cancer cell line through tubulin-targeted screening?
Answer 2: Cytoskeleton is pleased to offer specialized tubulins from different cancer cell lines for improved drug targeting. We currently offer highly purified tubulin isolated from the HeLa and MCF-7 cancer cell lines (Cat. # H001 and H005, respectively). We can also isolate tubulin from other cancer cell lines or tissue samples on a custom services basis.
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