The specificities of protein kinase inhibitors: an update

We have previously examined the specificities of 28 commercially available compounds, reported to be relatively selective inhibitors of particular serine/threonine-specific protein kinases [Davies, Reddy, Caivano and Cohen (2000) Biochem. J. 351, 95—105]. In the present study, we have extended this analysis to a further 14 compounds. Of these, indirubin-3′-monoxime, SP 600125, KT 5823 and ML-9 were found to inhibit a number of protein kinases and conclusions drawn from their use in cell-based assays are likely to be erroneous. Kenpaullone, Alsterpaullone, Purvalanol, Roscovitine, pyrazolopyrimidine 1 (PP1), PP2 and ML-7 were more specific, but still inhibited two or more protein kinases with similar potency. Our results suggest that the combined use of Roscovitine and Kenpaullone may be useful for identifying substrates and physiological roles of cyclin-dependent protein kinases, whereas the combined use of Kenpaullone and LiCl may be useful for identifying substrates and physiological roles of glycogen synthase kinase 3. The combined use of SU 6656 and either PP1 or PP2 may be useful for identifying substrates of Src family members. Epigallocatechin 3-gallate, one of the main polyphenolic constituents of tea, inhibited two of the 28 protein kinases in the panel, dual-specificity, tyrosine-phosphorylated and regulated kinase 1A (DYRK1A; IC50 = 0.33μM) and p38-regulated/activated kinase (PRAK; IC50 = 1.0μM).

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Targeting Protein Kinase Inhibitors with Traditional Chinese Medicine

Current Drug Targets ◽

10.2174/1389450120666190802125959 ◽

2019 ◽

Vol 20 (15) ◽

pp. 1505-1516

Author(s):

Yangyang Zhang ◽

Minghua Liu ◽

Jun Wang ◽

Jianlin Huang ◽

Mingyue Guo ◽

...

Keyword(s):

Chinese Medicine ◽

Protein Kinase ◽

Traditional Chinese Medicine ◽

Protein Kinases ◽

Anticancer Agents ◽

Kinase Inhibitors ◽

Specific Protein ◽

Protein Kinase Inhibitors ◽

Bioactive Substances ◽

Phosphoinositide 3 Kinase

Protein kinases play critical roles in the control of cell growth, proliferation, migration, and angiogenesis, through their catalytic activity. Over the past years, numerous protein kinase inhibitors have been identified and are being successfully used clinically. Traditional Chinese medicine (TCM) represents a large class of bioactive substances, and some of them display anticancer activity via inhibiting protein kinases signal pathway. Some of the TCM have been used to treat tumors clinically in China for many years. The p38mitogen-activated protein kinase (MAPK), phosphoinositide 3-kinase, serine/threonine-specific protein kinases (PI3K/AKT/mTOR), and extracellular signal-regulated kinases (ERK) pathways are considered important signals in cancer cell development. In the present article, the recent progress of TCM that exhibited significant inhibitory activity towards a range of protein kinases is discussed. The clinical efficacy of TCM with inhibitory effects on protein kinases in treating a tumor is also presented. The article also discussed the prospects and problems in the development of anticancer agents with TCM.

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Specificity and mechanism of action of some commonly used protein kinase inhibitors

Biochemical Journal ◽

10.1042/bj3510095 ◽

2000 ◽

Vol 351 (1) ◽

pp. 95-105 ◽

Cited By ~ 1822

Author(s):

Stephen P. DAVIES ◽

Helen REDDY ◽

Matilde CAIVANO ◽

Philip COHEN

Keyword(s):

Protein Kinase ◽

Protein Kinases ◽

Kinase Inhibitors ◽

Specific Protein ◽

Protein Kinase Inhibitors ◽

Mitogen Activated Protein Kinase ◽

Phosphatidylinositol 3 Kinase ◽

Selective Inhibitors ◽

Mitogen Activated Protein ◽

Additional Protein

The specificities of 28 commercially available compounds reported to be relatively selective inhibitors of particular serine/threonine-specific protein kinases have been examined against a large panel of protein kinases. The compounds KT 5720, Rottlerin and quercetin were found to inhibit many protein kinases, sometimes much more potently than their presumed targets, and conclusions drawn from their use in cell-based experiments are likely to be erroneous. Ro 318220 and related bisindoylmaleimides, as well as H89, HA1077 and Y 27632, were more selective inhibitors, but still inhibited two or more protein kinases with similar potency. LY 294002 was found to inhibit casein kinase-2 with similar potency to phosphoinositide (phosphatidylinositol) 3-kinase. The compounds with the most impressive selectivity profiles were KN62, PD 98059, U0126, PD 184352, rapamycin, wortmannin, SB 203580 and SB 202190. U0126 and PD 184352, like PD 98059, were found to block the mitogen-activated protein kinase (MAPK) cascade in cell-based assays by preventing the activation of MAPK kinase (MKK1), and not by inhibiting MKK1 activity directly. Apart from rapamycin and PD 184352, even the most selective inhibitors affected at least one additional protein kinase. Our results demonstrate that the specificities of protein kinase inhibitors cannot be assessed simply by studying their effect on kinases that are closely related in primary structure. We propose guidelines for the use of protein kinase inhibitors in cell-based assays.

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Rapid redistribution of F-actin induced by NGF and cyclicamp in PC12D cells was inhibited by specific protein kinase inhibitors

Neuroscience Research Supplements ◽

10.1016/0921-8696(91)90779-m ◽

1991 ◽

Vol 16 ◽

pp. 48

Author(s):

Misao Iwanaga ◽

Mamoru Sano

Keyword(s):

Protein Kinase ◽

Kinase Inhibitors ◽

Specific Protein ◽

Protein Kinase Inhibitors ◽

Specific Protein Kinase

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Multiple mechanisms for the phosphorylation of C-terminal regulatory sites in rabbit muscle glycogen synthase expressed in COS cells

Biochemical Journal ◽

10.1042/bj3130045 ◽

1996 ◽

Vol 313 (1) ◽

pp. 45-50 ◽

Cited By ~ 21

Author(s):

Alexander V. SKURAT ◽

Peter J. ROACH

Keyword(s):

Protein Kinase ◽

Protein Kinases ◽

Glycogen Synthase ◽

Glycogen Synthase Kinase ◽

Glycogen Synthase Kinase 3 ◽

Rabbit Muscle ◽

Cos Cells ◽

Additional Mutation ◽

Regulatory Sites

Glycogen synthase can be inactivated by sequential phosphorylation at the C-terminal residues Ser652 (site 4), Ser648 (site 3c), Ser644 (site 3b) and Ser640 (site 3a) catalysed by glycogen synthase kinase-3. In vitro, glycogen synthase kinase-3 action requires that glycogen synthase has first been phosphorylated at Ser656 (site 5) by casein kinase II. Recently we demonstrated that inactivation is linked only to phosphorylation at site 3a and site 3b, and that, in COS cells, modification of these sites can occur by alternative mechanisms independent of any C-terminal phosphorylations [Skurat and Roach (1995) J. Biol. Chem. 270, 12491-12497]. To address these mechanisms multiple Ser → Ala mutations were introduced in glycogen synthase such that only site 3a or site 3b remained intact. Additional mutation of Arg637 → Gln eliminated phosphorylation of site 3a, indicating that Arg637 may be important for recognition of site 3a by its corresponding protein kinase(s). Similarly, additional mutation of Pro645 → Ala eliminated phosphorylation of site 3b, indicating a possible involvement of ‘proline-directed’ protein kinase(s). Mutation of Arg637 alone did not activate glycogen synthase as expected from the loss of phosphorylation at site 3a. Rather, mutation of both Arg637 and the Ser → Ala substitution at site 3b was required for substantial activation. The results suggest that sites 3a and 3b can be phosphorylated independently of one another by distinct protein kinases. However, phosphorylation of site 3b can potentiate phosphorylation of site 3a, by an enzyme such as glycogen synthase kinase-3.

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Receptor-mediated endocytosis of urokinase-type plasminogen activator is regulated by cAMP-dependent protein kinase

Journal of Cell Science ◽

10.1242/jcs.110.12.1395 ◽

1997 ◽

Vol 110 (12) ◽

pp. 1395-1402 ◽

Cited By ~ 1

Author(s):

L. Goretzki ◽

B.M. Mueller

Keyword(s):

Protein Kinase ◽

Plasminogen Activator ◽

Kinase Inhibitors ◽

Melanoma Cells ◽

Protein Kinase Inhibitors ◽

Dependent Protein Kinase ◽

Camp Dependent Protein Kinase ◽

Type Plasminogen Activator ◽

Urokinase Type Plasminogen Activator ◽

Dependent Protein

Internalization of the urokinase-type plasminogen activator (uPA) requires two receptors, the uPA receptor (uPAR) and the low density lipoprotein receptor-related protein (LRP)/alpha2-macroglobulin (alpha2M) receptor. Here, we address whether protein kinases are involved in the internalization of uPA by human melanoma cells. Initially, we found that the internalization of uPA was significantly inhibited by the serine/threonine protein kinase inhibitors staurosporine, K-252a and H-89, but not by the tyrosine kinase inhibitors, genistein and lavendustin A. Internalization of uPA was also inhibited by a pseudosubstrate peptide for cAMP-dependent protein kinase (PKA), but not by a pseudosubstrate peptide for protein kinase C. We confirmed a requirement for PKA-activity and implicated a specific isoform by using an antisense oligonucleotide against the regulatory subunit RI alpha of PKA which suppresses PKA-I activity. Exposure of cells to this oligonucleotide led to a specific, dose-dependent decrease in RI alpha protein and to a significant inhibition in the rate of uPA internalization. We further demonstrate that treatment of melanoma cells with either H-89 or PKA RI alpha antisense oligonucleotides also resulted in a decreased internalization of two other ligands of LRP, activated alpha2M and lactoferrin, indicating that PKA activity is associated with LRP. Finally, we demonstrate that PKA activity is also required for the internalization of transferrin, but not for the internalization of the epidermal growth factor or adenovirus 2, suggesting that in melanoma cells, PKA activity is not generally required for clathrin-mediated endocytosis, but is rather associated with specific internalization receptors.

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