Design, Synthese und Optimierung von CK1δ Inhibitoren

Protein Kinase CK1δ is an important determinant in development and progression of severe pathogenic disorders such as Alzheimer's disease, amyotrophic lateral sclerosis, familial advanced sleep phase syndrome, and cancer. Consequently, potent and selevctive small molecule kinase inhibitors have been pursued with increasing interest over the past decade. However, the existence of further five evolutionary conserved human CK1 isoforms that possess similar, different, or even opposite physiological and pathophysiological implications renders the design of suitable candidates enormously comlicated. Especially highly related CK1 isoforms δ and ε remain difficult to discriminate and have therefore often been reported as redundant, rather by means of missing evidence. The present study reports on in silico design, synthesis, and biological evaluation of novel and optimized 4,5-diaryl-imidazoles as potent dual-specific ATP-competitive inhibitors of CK1 isoforms δ and ε as well as p38α mitogen-activated protein kinase. In fact, most promising candidate is among the most effective CK1 inhibitors published to date with an IC50 value in the low single-digit nanomolar range and good selectivity in a panel of 321 protein kinases. Consequently, it holds the potential to serve as an important biological tool for pharmacological evaluation of CK1-dependent activity such as in stem cell applications.Die Protein Kinase CK1δ ist einer der bestimmenden Faktoren in Entwicklung und Progression schwerwiegender pathogener Störungen wie der Alzheimer-Erkrankung, der amyotrophen Lateralsklerose, des familiären vorverlagerten Schlafphasen Syndroms und Krebs. Es ist daher kaum verwunderlich, dass die Bestrebungen hinsichtlich potenter und selektiver niedermolekularer CK1δ Inhibitoren innerhalb der vergangenen Dekade kontinuierlich zugenommen haben. Allerdings ist die Entwicklung derartiger Hemmstoffe durch die Existenz weiterer fünf evolutionär konservierter humaner CK1 Isoformen mit zum Teil ähnlicher, unterschiedlicher, oder sogar gegensätzlicher physiologischer und pathophysiologischer Bedeutung erheblich erschwert. Insbesondere die hochkonservierten Isoformen δ und ε sind bis heute durch kleine Moleküle nicht erfolgreich unterscheidbar und wurden daher in verschiedenen Zusammenhängen als redundant klassifiziert, ohne dass jedoch ausreichend Evidenz für diese Annahme geliefert werden konnte. Die vorliegende Arbeit beschreibt das in silico basierte Design, die Synthese und die biologische Charakterisierung neuer und optimierter 4,5-Diaryl-imidazole als potente und dual-spezifische ATP-kompetitive Inhibitoren der CK1 Isoformen δ und ε sowie der p38α Mitogen-aktivierten Protein Kinase. Tatsächlich handelt es sich bei der vielversprechendsten Verbindung um einen der potentesten CK1 Hemmstoffe die bis heute publiziert werden konnten. Der Inhibitor weist eine halbmaximale Hemmkonzentration im niedrigen einstelligen nanomolaren Bereich auf, zeigt gute Selektivität über 321 Protein Kinasen und ist biologisch aktiv in verschiedenen humanen Tumorzelllinien. Die Verbindung besitzt somit das Potential als biologisches Tool in der fort-schreitenden Untersuchung der pharmakologischen Bedeutung der CK1 entscheidend Beitrag zu leisten, gerade im Hinblick auf die zunehmende Anwendung Stammzell-basierter Testsysteme

A CK1 FRET biosensor reveals that DDX3X is an essential activator of CK1ε

Casein kinase 1 (CK1) plays central roles in various signal transduction pathways and performs many cellular activities. For many years CK1 was thought to act independently of modulatory subunits and in a constitutive manner. Recently, DEAD box RNA helicases, in particular DEAD box RNA helicase 3 X-linked (DDX3X), were found to stimulate CK1 activity in vitro In order to observe CK1 activity in living cells and to study its interaction with DDX3X, we developed a CK1-specific FRET biosensor. This tool revealed that DDX3X is indeed required for full CK1 activity in living cells. Two counteracting mechanisms control the activity of these enzymes. Phosphorylation by CK1 impairs the ATPase activity of DDX3X and RNA destabilizes the DDX3X-CK1 complex. We identified possible sites of interaction between DDX3X and CK1. While mutations identified in the DDX3X genes of human medulloblastoma patients can enhance CK1 activity in living cells, the mechanism of CK1 activation by DDX3X points to a possible therapeutic approach in CK1-related diseases such as those caused by tumors driven by aberrant Wnt/beta-catenin and Sonic hedgehog (SHH) activation. Indeed, CK1 peptides can reduce CK1 activity

Optimized 4,5-Diarylimidazoles as Potent/Selective Inhibitors of Protein Kinase CK1 delta and Their Structural Relation to p38 alpha MAPK

The involvement of protein kinase CK1 delta in the pathogenesis of severe disorders such as Alzheimer's disease, amyotrophic lateral sclerosis, familial advanced sleep phase syndrome, and cancer has dramatically increased interest in the development of effective small molecule inhibitors for both therapeutic application and basic research. Unfortunately, the design of CK1 isoform-specific compounds has proved to be highly complicated due to the existence of six evolutionarily conserved human CK1 members that possess similar, different, or even opposite physiological and pathophysiological implications. Consequently, only few potent and selective CK1 delta inhibitors have been reported so far and structurally divergent approaches are urgently needed in order to establish SAR that might enable complete discrimination of CK1 isoforms and related p38 alpha MAPK. In this study we report on design and characterization of optimized 4,5-diarylimidazoles as highly effective ATP-competitive inhibitors of CK1 delta with compounds 11b (IC50 CK1 delta = 4 nM, IC50 CK1 epsilon = 25 nM), 12a (IC50 CK1 delta = 19 nM, IC50 CK1 epsilon = 227 nM), and 16b (IC50 CK1 delta = 8 nM, IC50 CK1 epsilon = 81 nM) being among the most potent CK1 delta-targeting agents published to date. Inhibitor compound 11b, displaying potential as a pharmacological tool, has further been profiled over a panel of 321 protein kinases exhibiting high selectivity. Cellular efficacy has been evaluated in human pancreatic cancer cell lines Colo357 (EC50 = 3.5 mu M) and Panc89 (EC50 = 1.5 mu M). SAR is substantiated by X-ray crystallographic analysis of 16b in CK1 delta and 11b in p38 alpha

Optimized 4,5-diarylimidazoles as potent/selective inhibitors of protein kinase CK1δ and their structural relation to p38α MAPK

The involvement of protein kinase CK1δ in the pathogenesis of severe disorders such as Alzheimer’s disease, amyotrophic lateral sclerosis, familial advanced sleep phase syndrome, and cancer has dramatically increased interest in the development of effective small molecule inhibitors for both therapeutic application and basic research. Unfortunately, the design of CK1 isoform-specific compounds has proved to be highly complicated due to the existence of six evolutionarily conserved human CK1 members that possess similar, different, or even opposite physiological and pathophysiological implications. Consequently, only few potent and selective CK1δ inhibitors have been reported so far and structurally divergent approaches are urgently needed in order to establish SAR that might enable complete discrimination of CK1 isoforms and related p38α MAPK. In this study we report on design and characterization of optimized 4,5-diarylimidazoles as highly effective ATP-competitive inhibitors of CK1δ with compounds 11b (IC50 CK1δ = 4 nM, IC50 CK1ε = 25 nM), 12a (IC50 CK1δ = 19 nM, IC50 CK1ε = 227 nM), and 16b (IC50 CK1δ = 8 nM, IC50 CK1ε = 81 nM) being among the most potent CK1δ-targeting agents published to date. Inhibitor compound 11b, displaying potential as a pharmacological tool, has further been profiled over a panel of 321 protein kinases exhibiting high selectivity. Cellular efficacy has been evaluated in human pancreatic cancer cell lines Colo357 (EC50 = 3.5 µM) and Panc89 (EC50 = 1.5 µM). SAR is substantiated by X-ray crystallographic analysis of 16b in CK1δ and 11b in p38α