Abstract
The advancement of a kinase inhibitor throughout drug discovery and development is predicated upon its selectivity towards the target of interest. Thus, profiling the compound against a broad panel of kinases is important for providing a better understanding of its activity and for obviating any off-target activities that can result in undesirable consequences. To assess the selectivity and potency of an inhibitor against multiple kinases, it is desirable to use a universal assay that can monitor the activity of all classes of kinases regardless of the nature of their substrates. The luminescent ADP-Glo kinase assay is a universal platform that measures kinase activity by quantifying the amount of the common kinase reaction product ADP. Here we present a method using standardized kinase profiling systems for inhibitor profiling studies based on ADP detection by luminescence. The kinase profiling systems are sets of kinases organized by family, presented in multi-tube strips containing eight enzymes, each with corresponding substrate strips, and standardized for optimal kinase activity. We show that using the kinase profiling strips we could quickly and easily generate multiple selectivity profiles using small or large kinase panels, and identify compound promiscuity within the kinome.
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References
Hunter T (2000) Signaling-2000 and beyond. Cell 100:113–127
Manning G, Whyte DB, Martinez R et al (2002) The protein kinase complement of the human genome. Science 298:1912–1934
Cohen P (2002) Protein kinases—the major drug targets of the twenty-first century? Nat Rev Drug Discov 1:309–315
Widakowich C, de Castro G Jr, de Azambuja E et al (2007) Review: side effects of approved molecular targeted therapies in solid cancers. Oncologist 12:1443–1455
Castoldi RE, Pennella G, Saturno GS et al (2007) Assessing and managing toxicities induced by kinase inhibitors. Curr Opin Drug Discov Devel 10:53–57
Olaharski AJ, Gonzaludo N, Bitter H et al (2009) Identification of a Kinase Profile that Predicts Chromosome Damage Induced by Small Molecule Kinase Inhibitors. PLoS Comput Biol 5, e1000446. doi:10.1371/journal.pcbi.1000446
Bain J, Plater L, Elliott M et al (2007) The selectivity of protein kinase inhibitors: a further update. Biochem J 408:297–315
Jia Y, Quinn CM, Kwak S, Talanian RV (2008) Current in vitro kinase assay technologies: The quest for a universal format. Curr Drug Discov Technol 5:59–69
Zegzouti H, Zdanovskaia M, Hsiao K, Goueli SA (2009) ADP-Glo: A Bioluminescent and homogeneous ADP monitoring assay for kinases. Assay Drug Dev Technol 7:560–572
Krishnamurty R, Maly DJ (2007) Chemical genomic and proteomic methods for determining kinase inhibitor selectivity. Comb Chem High Throughput Screen 10:652–666
Miduturu CV, Deng X, Kwiatkowski N et al (2011) High-throughput kinase profiling: a more efficient approach toward the discovery of new kinase inhibitors. Chem Biol 18:868–879
Anastassiadis T, Deacon SW, Devarajan K et al (2011) Comprehensive assay of kinase catalytic activity reveals features of kinase inhibitor selectivity. Nat Biotechnol 29:1039–1045
Fabian MA, Biggs WH 3rd, Treiber DK et al (2005) A small molecule-kinase interaction map for clinical kinase inhibitors. Nat Biotechnol 23:329–336
Karaman MW, Herrgard S, Treiber DK et al (2008) A quantitative analysis of kinase inhibitor selectivity. Nat Biotechnol 26:127–132
Card M, Caldwell C, Min H et al (2009) High-Throughput Biochemical Kinase Selectivity Assays: Panel Development and Screening Applications. J Biomol Screen 14:31–42
Larson B, Banks P, Zegzouti H, Goueli SA (2009) A simple and robust automated kinase profiling platform using luminescent ADP accumulation technology. Assay Drug Dev Technol 7:573–584
Zegzouti H, Alves J, Worzella T et al (2011) Screening and profiling kinase inhibitors with a luminescent ADP detection platform. Promega Corporation Web site. http://www.promega.com/resources/pubhub/screening-and-profiling-kinase-inhibitors-with-a-luminescent-adp-detection-platform. Accessed 21 April 2015
Tanega C, Shen M, Mott BT et al (2009) Comparison of bioluminescent kinase assays using substrate depletion and product formation. Assay Drug Dev Technol 7:606–614
Davis MI, Sasaki AT, Shen M et al (2013) A Homogeneous, High-Throughput Assay for Phosphatidylinositol 5-Phosphate 4-Kinase with a Novel, Rapid Substrate Preparation. PLoS One 8, e54127. doi:10.1371/journal.pone.0054127
Li H, Totoritis RD, Lor LA et al (2009) Evaluation of an antibody-free ADP detection assay: ADP-Glo. Assay Drug Dev Technol 7:598–605
Zegzouti H, Hennek J, Alves J, Goueli S (2015) Kinase strips for routine creation of inhibitor selectivity profiles: a novel approach to targeted profiling. SLAS Conference 2015. http://www.promega.com/resources/scientific_posters/posters/kinase-strips-forcreationinhibitorselectivityprofilesnovel-approachtargetedprofilingposter/. Accessed 30 April 2015.
Blasina A, Hallin J, Chen E et al (2008) Breaching the DNA damage checkpoint via PF-00477736, a novel small-molecule inhibitor of checkpoint kinase 1. Mol Cancer Ther 7:2394–2404
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Zegzouti, H., Hennek, J., Goueli, S.A. (2016). Using Bioluminescent Kinase Profiling Strips to Identify Kinase Inhibitor Selectivity and Promiscuity. In: Zegzouti, H., Goueli, S. (eds) Kinase Screening and Profiling. Methods in Molecular Biology, vol 1360. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3073-9_5
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DOI: https://doi.org/10.1007/978-1-4939-3073-9_5
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3072-2
Online ISBN: 978-1-4939-3073-9
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