title: Bioluminescent kinase strips: A novel approach to targeted and flexible kinase inhibitor profiling.
abstract: In addition to target efficacy, drug safety is a major requirement during the drug discovery process and is influenced by target specificity. Therefore, it is imperative that every new drug candidate be profiled against various liability panels that include protein kinases. Here, an effective methodology to streamline kinase inhibitor profiling is described. An accessible standardized profiling system for 112 protein kinases covering all branches of the kinome was developed. This approach consists of creating different sets of kinases and their corresponding substrates in multi-tube strips. The kinase stocks are pre-standardized for optimal kinase activity and used for inhibitor profiling using a bioluminescent ADP detection assay. We show that these strips can routinely generate inhibitor selectivity profiles for small or broad kinase family panels. Lipid kinases were also assembled in strip format and profiled together with protein kinases. We identified two specific PI3K inhibitors that have off-target effects on CK2 that were not reported before and would have been missed if compounds were not profiled against lipid and protein kinases simultaneously. To validate the accuracy of the data generated by this method, we confirmed that the inhibition potencies observed are consistent with published values produced by more complex technologies such as radioactivity assays.
authors.affiliation: R&D Department, Promega Corporation, Madison, WI 53711, USA.
authors.forename: J
authors.initials: J
authors.lastname: Hennek
authors.affiliation: R&D Department, Promega Corporation, Madison, WI 53711, USA.
authors.forename: J
authors.initials: J
authors.lastname: Alves
authors.affiliation: SignalChem Pharmaceuticals, Richmond, British Columbia V6V 2J2, Canada.
authors.forename: E
authors.initials: E
authors.lastname: Yao
authors.affiliation: R&D Department, Promega Corporation, Madison, WI 53711, USA; Department of Pathology and Laboratory Medicine, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA.
authors.forename: S A
authors.initials: SA
authors.lastname: Goueli
authors.affiliation: R&D Department, Promega Corporation, Madison, WI 53711, USA. Electronic address: hicham.zegzouti@promega.com.
authors.forename: H
authors.initials: H
authors.lastname: Zegzouti
mesh_terms: D000244:Adenosine Diphosphate / Q000032:analysis / Q000378:metabolism; D057075:Enzyme Assays; D020128:Inhibitory Concentration 50; D008156:Luciferases / Q000737:chemistry / Q000378:metabolism; D008163:Luminescent Measurements; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D000244:Adenosine Diphosphate; D008156:Luciferases; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases
keywords: ADP detection; Bioluminescence; Kinase assay; Kinase inhibitor; Kinase profiling; Selectivity profiles