Determination of IC50 at RSK2
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Compounds were assessed for their ability to inhibit the phosphorylation of a substrate peptide by RSK2 as described previously (Boyer et al., 2011). In brief, human RSK2 protein (Invitrogen, Carlsbad, CA) was used to measure kinase activity utilizing Kinase GloPlus (Promega, Madison, WI) that uses a luciferin-luciferase based detection reagent to quantify residual ATP. The relative light unit signal was measured on an LJL Analyst (Molecular Devices, Sunnyvale, CA) in luminescence mode using 384 aperture; relative light unit signals were converted to percentage of control; the IC50 was fitted to a standard four-parameter logistic equation.
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Applications
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BIX 02565 (1, 3, and 10 mg/kg) elicited decreases in both mean arterial pressure and heart rate in anesthetized rats. Moreover, BIX 02565 (30, 100, and 300 mg/kg) also induced concentration-dependent decreases in mean arterial pressure (MAP) after each daily oral dose in telemetry-instrumented conscious rats.
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References:
1. Fryer, R. M., Muthukumarana, A., Chen, R. R., Smith, J. D., Mazurek, S. N., Harrington, K. E., Dinallo, R. M., Burke, J., DiCapua, F. M., Guo, X., Kirrane, T. M., Snow, R. J., Zhang, Y., Soleymanzadeh, F., Madwed, J. B., Kashem, M. A., Kugler, S. Z., O'Neill, M. M., Harrison, P. C., Reinhart, G. A. and Boyer, S. J. (2012) Mitigation of off-target adrenergic binding and effects on cardiovascular function in the discovery of novel ribosomal S6 kinase 2 inhibitors. J Pharmacol Exp Ther. 340, 492-500
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