A Cdk9 inhibitor
LDC000067 (LDC067) is a novel specific inhibitor of CDK9 with IC50 value of 44 ± 10 nM [1].
Cyclin-dependent kinase 9 (CDK9) is a cyclin-dependent kinase. CDK9 and cyclin T form the positive transcription elongation factor b (P-TEFb) complex for RNA polymerase II and functions by phosphorylating the C-terminal domain of the largest subunit of RNA polymerase II [1].
LDC000067 (LDC067) is a novel and highly specific CDK9 inhibitor. LDC000067 exhibited selectivity for CDK9 over other CDKs in the range of 55-fold (vs. CDK2) to over 230-fold (vs. CDK6 and CDK7). LDC067 also inhibited transcription in a dose-dependent and ATP-competitive manner. In whole cells, LDC000067 induced the tumor suppressor protein p53 activation and apoptosis. LDC000067 also selectively reduced short-lived mRNAs, including those that encode regulators of apoptosis and proliferation such as MYC and MCL1 [1].
Reference:
[1].? Albert TK, Rigault C, Eickhoff J, et al. Characterization of molecular and cellular functions of the cyclin-dependent kinase CDK9 using a novel specific inhibitor. Br J Pharmacol, 2014, 171(1): 55-68.
| Kinase experiment [1]: | |
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Inhibitory activities |
Kinase inhibition by LDC000067 was measured in a radio metric assay, which directly measured kinase catalytic activity towards a specific substrate. Briefly, 10 μM LDC000067 or DMSO as solvent control, were added to base reaction buffer (10 mM MgCl2, 1 mM EDTA, 20 mM HEPES pH 7.5, 2 mM DTT, 0.02 mg·mL-1 BSA, 0.1 mM Na3VO4, 0.02% Brij35, 1% DMSO), containing cofactors and substrates, which were required by the individual kinase. 10 μCi of [γ-33P]-APP (10mCi·mL-1, 3000Ci·mmol-1, Perkin Elmer) was added to the reaction mixture. And such kinase reaction incubated for 120min at room temperature. Reactions were found on P81 ion exchange paper, and filters generally washed in 0.75% phosphoric acid before radiometric quantification. Each protein kinase was measured in duplicate, and its catalytic activity expressed as residual kinase activity, which shows the percentage of average substrate phophorylation in contrast with the solvent control reaction. |
| Cell experiment [1]: | |
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Cell lines |
HEK293T cells, THP1 cells |
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Preparation method |
The solubility of this compound in DMSO is >10 mM. General tips for obtaining a higher concentration: Please warm the tube at 37℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months. |
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Reaction Conditions |
Competitive kinase binding displacement assay: 90 min.DNA microarray analysis assay: 90 min. |
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Applications |
LDC000067 inhibits CDK9 with an IC50 value of 44(10 nM, and its selectivity for CDK9 over other CDKs is in the range of 55-fold to over 230-fold, especially higher selectivity in an ATP-competitive kinase binding assay. Besides, effects of LDC000067 in whole cells contain induction of the tumour suppressor protein p53 and apoptosis. Moreover, gene expression profiling of cells treated with LDC000067 demonstrate selective reduction of short-lived mRNAs, which encode regulators of proliferation and apoptosis, such as MCL1 and MYC. |
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参考文献: [1]. T K Albert1, C Rigault1, J Eickhoff, K Baumgart, C Antrecht1, B Klebl, G Mittler and M Meisterernst. Characterization of molecular and cellular functions of the cyclin-dependent kinase CDK9 using a novel specific inhibitor. British Journal of Pharmacology. 2014, 171: 55–68. | |
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