How Selective Are Pharmacological Inhibitors of Cell-Cycle-Regulating Cyclin-Dependent Kinases?

Cyclin-dependent kinases (CDKs) are an important and emerging class of drug targets for which many small-molecule inhibitors have been developed. However, there is often insufficient data available on the selectivity of CDK inhibitors (CDKi) to attribute the effects on the presumed target CDK to these inhibitors. Here, we highlight discrepancies between the kinase selectivity of CDKi and the phenotype exhibited; we evaluated 31 CDKi (claimed to target CDK1-4) for activity toward CDKs 1, 2, 4, 5, 7, 9 and for effects on the cell cycle. Our results suggest that most CDKi should be reclassified as pan-selective and should not be used as a tool. In addition, some compounds did not even inhibit CDKs as their primary cellular targets; for example, NU6140 showed potent inhibition of Aurora kinases. We also established an online database of commercially available CDKi for critical evaluation of their utility as molecular probes. Our results should help researchers select the most relevant chemical tools for their specific applications.

[1]  Sarah A. Boswell,et al.  Multi-Omics Profiling Establishes the Polypharmacology of FDA Approved CSK4/6 Inhibitors and Its Impact on Drug Response , 2018 .

[2]  W. Tap,et al.  CDK4/6 Inhibitors: The Mechanism of Action May Not Be as Simple as Once Thought. , 2018, Cancer cell.

[3]  V. Kryštof,et al.  Selective inhibition reveals cyclin-dependent kinase 2 as another kinase that phosphorylates the androgen receptor at serine 81. , 2018, Biochimica et biophysica acta. Molecular cell research.

[4]  M. Hochberg,et al.  Spatial competition constrains resistance to targeted cancer therapy , 2017, Nature Communications.

[5]  Heiner Koch,et al.  The target landscape of clinical kinase drugs , 2017, Science.

[6]  Esther S. Kim,et al.  Abemaciclib: First Global Approval , 2017, Drugs.

[7]  Gang Wu,et al.  Cyclin-dependent kinase 5 controls vasculogenic mimicry formation in non-small cell lung cancer via the FAK-AKT signaling pathway. , 2017, Biochemical and biophysical research communications.

[8]  Daniel Svozil,et al.  Probes &Drugs portal: an interactive, open data resource for chemical biology , 2017, Nature Methods.

[9]  J. Hutcheson,et al.  Biological specificity of CDK4/6 inhibitors: dose response relationship, in vivo signaling, and composite response signature , 2017, Oncotarget.

[10]  Jürgen Bajorath,et al.  Mapping of inhibitors and activity data to the human kinome and exploring promiscuity from a ligand and target perspective , 2017, Chemical biology & drug design.

[11]  Kuen-Feng Chen,et al.  Palbociclib induces activation of AMPK and inhibits hepatocellular carcinoma in a CDK4/6‐independent manner , 2017, Molecular oncology.

[12]  Jian Du,et al.  Preclinical characterization of abemaciclib in hormone receptor positive breast cancer , 2017, Oncotarget.

[13]  Yahiya Y. Syed.  Ribociclib: First Global Approval , 2017, Drugs.

[14]  Lenka Munoz,et al.  Non-kinase targets of protein kinase inhibitors , 2017, Nature Reviews Drug Discovery.

[15]  P. Sicinski,et al.  Cell cycle proteins as promising targets in cancer therapy , 2017, Nature Reviews Cancer.

[16]  M. Noble,et al.  Cyclin-Dependent Kinase (CDK) Inhibitors: Structure–Activity Relationships and Insights into the CDK-2 Selectivity of 6-Substituted 2-Arylaminopurines , 2016, Journal of medicinal chemistry.

[17]  Jia Wang,et al.  Cyclin-Dependent Kinase 2 Promotes Tumor Proliferation and Induces Radio Resistance in Glioblastoma , 2016, Translational oncology.

[18]  M. Kaur,et al.  Design strategies, structure activity relationship and mechanistic insights for purines as kinase inhibitors. , 2016, European journal of medicinal chemistry.

[19]  M. Strnad,et al.  5-Substituted 3-isopropyl-7-[4-(2-pyridyl)benzyl]amino-1(2)H-pyrazolo[4,3-d]pyrimidines with anti-proliferative activity as potent and selective inhibitors of cyclin-dependent kinases. , 2016, European journal of medicinal chemistry.

[20]  K. Berka,et al.  Synthesis, biological evaluation and molecular modeling of a novel series of 7-azaindole based tri-heterocyclic compounds as potent CDK2/Cyclin E inhibitors. , 2016, European journal of medicinal chemistry.

[21]  U. Rix,et al.  Chemoproteomics Reveals Novel Protein and Lipid Kinase Targets of Clinical CDK4/6 Inhibitors in Lung Cancer. , 2015, ACS chemical biology.

[22]  M. Noble,et al.  Identification and Characterization of an Irreversible Inhibitor of CDK2 , 2015, Chemistry & biology.

[23]  John P. Overington,et al.  The promise and peril of chemical probes. , 2015, Nature chemical biology.

[24]  E. Reddy,et al.  Dual Inhibition of CDK4/Rb and PI3K/AKT/mTOR Pathways by ON123300 Induces Synthetic Lethality in Mantle Cell Lymphomas , 2015, Leukemia.

[25]  V. Kryštof,et al.  Novel arylazopyrazole inhibitors of cyclin-dependent kinases. , 2015, Bioorganic & medicinal chemistry.

[26]  B. Kuster,et al.  Optimized chemical proteomics assay for kinase inhibitor profiling. , 2015, Journal of proteome research.

[27]  Eugenia G. Giannopoulou,et al.  CDK/CK1 inhibitors roscovitine and CR8 downregulate amplified MYCN in neuroblastoma cells , 2014, Oncogene.

[28]  David Shum,et al.  Small-Molecule Inhibitors of SETD8 with Cellular Activity , 2014, ACS chemical biology.

[29]  P. Nordlund,et al.  The cellular thermal shift assay for evaluating drug target interactions in cells , 2014, Nature Protocols.

[30]  Julian Blagg,et al.  Chemical biology approaches to target validation in cancer. , 2014, Current opinion in pharmacology.

[31]  M. Malumbres.  Cyclin-dependent kinases , 2014, Genome Biology.

[32]  P. Iversen,et al.  Preclinical characterization of the CDK4/6 inhibitor LY2835219: in-vivo cell cycle-dependent/independent anti-tumor activities alone/in combination with gemcitabine , 2014, Investigational New Drugs.

[33]  A. Kozubík,et al.  The role of high cell density in the promotion of neuroendocrine transdifferentiation of prostate cancer cells , 2014, Molecular Cancer.

[34]  Y. Chai,et al.  CDK2-dependent phosphorylation of Suv39H1 is involved in control of heterochromatin replication during cell cycle progression , 2014, Nucleic acids research.

[35]  M. Noble,et al.  An Inhibitor’s-Eye View of the ATP-Binding Site of CDKs in Different Regulatory States , 2014, ACS chemical biology.

[36]  L. Wodicka,et al.  Dual kinase-bromodomain inhibitors for rationally designed polypharmacology , 2014, Nature chemical biology.

[37]  Sung Hyun Lee,et al.  A specific inhibitor of CDK1, RO-3306, reversibly arrests meiosis during in vitro maturation of porcine oocytes. , 2014, Animal reproduction science.

[38]  R. Sutherland,et al.  Targeting cyclin-dependent kinase 1 (CDK1) but not CDK4/6 or CDK2 is selectively lethal to MYC-dependent human breast cancer cells , 2014, BMC Cancer.

[39]  E. Reddy,et al.  Discovery of 8-Cyclopentyl-2-[4-(4-methyl-piperazin-1-yl)-phenylamino]-7-oxo-7,8-dihydro-pyrido[2,3-d]pyrimidine-6-carbonitrile (7x) as a Potent Inhibitor of Cyclin-Dependent Kinase 4 (CDK4) and AMPK-Related Kinase 5 (ARK5) , 2014, Journal of medicinal chemistry.

[40]  N. Manfrini,et al.  Regulation of the DNA damage response by cyclin-dependent kinases. , 2013, Journal of molecular biology.

[41]  Jun Lu,et al.  CDK5 is essential for TGF-β1-induced epithelial-mesenchymal transition and breast cancer progression , 2013, Scientific Reports.

[42]  S. Olesen,et al.  Cyclin-dependent kinase inhibitor dinaciclib interacts with the acetyl-lysine recognition site of bromodomains. , 2013, ACS chemical biology.

[43]  P. Sicinski,et al.  The kinase-independent, second life of CDK6 in transcription. , 2013, Cancer cell.

[44]  P. Kaldis,et al.  Cdks, cyclins and CKIs: roles beyond cell cycle regulation , 2013, Development.

[45]  Karel Berka,et al.  A novel series of highly potent 2,6,9-trisubstituted purine cyclin-dependent kinase inhibitors. , 2013, Journal of medicinal chemistry.

[46]  P. Nordlund,et al.  Monitoring Drug Target Engagement in Cells and Tissues Using the Cellular Thermal Shift Assay , 2013, Science.

[47]  M. Strnad,et al.  Synthesis and in vitro biological evaluation of 2,6,9-trisubstituted purines targeting multiple cyclin-dependent kinases. , 2013, European journal of medicinal chemistry.

[48]  F. Hoh,et al.  An integrated chemical biology approach provides insight into Cdk2 functional redundancy and inhibitor sensitivity. , 2012, Chemistry & biology.

[49]  Joost C. M. Uitdehaag,et al.  A guide to picking the most selective kinase inhibitor tool compounds for pharmacological validation of drug targets , 2012, British journal of pharmacology.

[50]  Mindy I. Davis,et al.  Comprehensive analysis of kinase inhibitor selectivity , 2011, Nature Biotechnology.

[51]  Theonie Anastassiadis,et al.  Comprehensive assay of kinase catalytic activity reveals features of kinase inhibitor selectivity , 2011, Nature biotechnology.

[52]  Jernej Ule,et al.  The Cyclin K/Cdk12 complex maintains genomic stability via regulation of expression of DNA damage response genes. , 2011, Genes & development.

[53]  N. Curtin,et al.  Preclinical in vitro and in vivo evaluation of the potent and specific cyclin-dependent kinase 2 inhibitor NU6102 and a water soluble prodrug NU6301. , 2011, European journal of cancer.

[54]  P. Butler,et al.  Cyclin-Dependent Kinase 5 Promotes Pancreatic β-Cell Survival via Fak-Akt Signaling Pathways , 2011, Diabetes.

[55]  Ian Collins,et al.  Probing the Probes: Fitness Factors For Small Molecule Tools , 2010, Chemistry & biology.

[56]  S. Gollin,et al.  Deficiency in Myosin Light Chain Phosphorylation Causes Cytokinesis Failure and Multipolarity in Cancer Cells , 2010, Oncogene.

[57]  Stephen V Frye,et al.  The art of the chemical probe. , 2010, Nature chemical biology.

[58]  Xiao-min Luo,et al.  B5, a novel pyrrole-substituted indolinone, exerts potent antitumor efficacy through G2/M cell cycle arrest , 2010, Investigational new drugs.

[59]  N. Curtin,et al.  Pre-clinical evaluation of cyclin-dependent kinase 2 and 1 inhibition in anti-estrogen-sensitive and resistant breast cancer cells , 2009, British Journal of Cancer.

[60]  A. Isacchi,et al.  Identification of N,1,4,4-tetramethyl-8-{[4-(4-methylpiperazin-1-yl)phenyl]amino}-4,5-dihydro-1H-pyrazolo[4,3-h]quinazoline-3-carboxamide (PHA-848125), a potent, orally available cyclin dependent kinase inhibitor. , 2009, Journal of medicinal chemistry.

[61]  M. Andreeff,et al.  Cyclin‐dependent kinase 1 inhibitor RO‐3306 enhances p53‐mediated Bax activation and mitochondrial apoptosis in AML , 2009, Cancer science.

[62]  S. Rabindran,et al.  Discovery of 4-(benzylaminomethylene)isoquinoline-1,3-(2H,4H)-diones and 4-[(pyridylmethyl)aminomethylene]isoquinoline-1,3-(2H,4H)-diones as potent and selective inhibitors of the cyclin-dependent kinase 4. , 2009, Journal of medicinal chemistry.

[63]  P. Greengard,et al.  Roscovitine-derived, dual-specificity inhibitors of cyclin-dependent kinases and casein kinases 1. , 2008, Journal of medicinal chemistry.

[64]  L Meijer,et al.  CR8, a potent and selective, roscovitine-derived inhibitor of cyclin-dependent kinases , 2008, Oncogene.

[65]  L. Krasinska,et al.  Selective chemical inhibition as a tool to study Cdk1 and Cdk2 functions in the cell cycle , 2008, Cell cycle.

[66]  S. Knapp,et al.  A systematic interaction map of validated kinase inhibitors with Ser/Thr kinases , 2007, Proceedings of the National Academy of Sciences.

[67]  S. Knapp,et al.  Crystal Structures of the p21-Activated Kinases PAK4, PAK5, and PAK6 Reveal Catalytic Domain Plasticity of Active Group II PAKs , 2007, Structure.

[68]  G. Bubley,et al.  Androgen receptor phosphorylation and stabilization in prostate cancer by cyclin-dependent kinase 1 , 2006, Proceedings of the National Academy of Sciences.

[69]  Hongmao Sun,et al.  Selective small-molecule inhibitor reveals critical mitotic functions of human CDK1. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[70]  S. Emanuel,et al.  1-Acyl-1H-[1,2,4]triazole-3,5-diamine analogues as novel and potent anticancer cyclin-dependent kinase inhibitors: synthesis and evaluation of biological activities. , 2005, Journal of medicinal chemistry.

[71]  L. Wodicka,et al.  A small molecule–kinase interaction map for clinical kinase inhibitors , 2005, Nature Biotechnology.

[72]  L. Meijer,et al.  Novel CDK inhibition profiles of structurally varied 1-aza-9-oxafluorenes. , 2005, Bioorganic & medicinal chemistry letters.

[73]  Christopher B. Cooper,et al.  Discovery and SAR of 2-aminothiazole inhibitors of cyclin-dependent kinase 5/p25 as a potential treatment for Alzheimer's disease. , 2004, Bioorganic & medicinal chemistry letters.

[74]  Yuzhu Chen,et al.  N2-substituted O6-cyclohexylmethylguanine derivatives: potent inhibitors of cyclin-dependent kinases 1 and 2. , 2004, Journal of medicinal chemistry.

[75]  A. Osnowski,et al.  2-Anilino-4-(thiazol-5-yl)pyrimidine CDK inhibitors: synthesis, SAR analysis, X-ray crystallography, and biological activity. , 2004, Journal of medicinal chemistry.

[76]  P. Greengard,et al.  Structural basis for the synthesis of indirubins as potent and selective inhibitors of glycogen synthase kinase-3 and cyclin-dependent kinases. , 2004, Journal of medicinal chemistry.

[77]  P. Greengard,et al.  GSK-3-selective inhibitors derived from Tyrian purple indirubins. , 2003, Chemistry & biology.

[78]  Terence Hui,et al.  SU9516: biochemical analysis of cdk inhibition and crystal structure in complex with cdk2. , 2003, Biochemical and biophysical research communications.

[79]  S. A. Watkins,et al.  Synthesis, structure-activity relationship, and biological studies of indolocarbazoles as potent cyclin D1-CDK4 inhibitors. , 2003, Journal of medicinal chemistry.

[80]  L. Meijer,et al.  Evaluation of the first cytostatically active 1-aza-9-oxafluorenes as novel selective CDK1 inhibitors with P-glycoprotein modulating properties. , 2003, Journal of medicinal chemistry.

[81]  D. Fabbro,et al.  Selective in vivo and in vitro effects of a small molecule inhibitor of cyclin-dependent kinase 4. , 2001, Journal of the National Cancer Institute.

[82]  B G Benson,et al.  Prevention of chemotherapy-induced alopecia in rats by CDK inhibitors. , 2001, Science.

[83]  A H Calvert,et al.  Identification of novel purine and pyrimidine cyclin-dependent kinase inhibitors with distinct molecular interactions and tumor cell growth inhibition profiles. , 2000, Journal of medicinal chemistry.

[84]  A Joly,et al.  CVT-313, a Specific and Potent Inhibitor of CDK2 That Prevents Neointimal Proliferation* , 1997, The Journal of Biological Chemistry.

[85]  Julian Blagg,et al.  A public-private partnership to unlock the untargeted kinome. , 2013, Nature chemical biology.

[86]  Mindy I. Davis,et al.  A quantitative analysis of kinase inhibitor selectivity , 2008, Nature Biotechnology.

[87]  L. Di,et al.  5.20 – Chemical Stability , 2007 .

[88]  Doriano Fabbro,et al.  Chemical and biological profile of dual Cdk1 and Cdk2 inhibitors. , 2003, Current medicinal chemistry. Anti-cancer agents.