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Open AccessArticle

Characterizing CDK8/19 Inhibitors through a NFκB-Dependent Cell-Based Assay

1
Department of Drug Discovery and Biomedical Sciences, University of South Carolina, Columbia, SC 29208, USA
2
Senex Biotechnology, Inc., Columbia, SC 29208, USA
*
Authors to whom correspondence should be addressed.
Cells 2019, 8(10), 1208; https://doi.org/10.3390/cells8101208
Received: 9 September 2019 / Revised: 2 October 2019 / Accepted: 4 October 2019 / Published: 6 October 2019
(This article belongs to the Special Issue The Role of Mediator Kinase in Cancer )
Cell-based assays for CDK8/19 inhibition are not easily defined, since there are no known cellular functions unique to these kinases. To solve this problem, we generated derivatives of 293 cells with CRISPR knockout of one or both of CDK8 and CDK19. Double knockout (dKO) of CDK8 and CDK19 together (but not individually) decreased the induction of transcription by NFκB (a CDK8/19-potentiated transcription factor) and abrogated the effect of CDK8/19 inhibitors on such induction. We generated wild type (WT) and dKO cell lines expressing luciferase from an NFκB-dependent promoter. Inhibitors selective for CDK8/19 over other CDKs decreased TNFα-induced luciferase expression in WT cells by ~80% with no effect on luciferase induction in dKO cells. In contrast, non-selective CDK inhibitors flavopiridol and dinaciclib and a CDK7/12/13 inhibitor THZ1 (but not CDK4/6 inhibitor palbociclib) suppressed luciferase induction in both WT and dKO cells, indicating a distinct role for other CDKs in the NFκB pathway. We used this assay to characterize a series of thienopyridines with in vitro bone anabolic activity, one of which was identified as a selective CDK8/19 inhibitor. Thienopyridines inhibited luciferase induction in the WT but not dKO cells and their IC50 values in the WT reporter assay showed near-perfect correlation (R2 = 0.98) with their reported activities in a bone anabolic activity assay, confirming that the latter function is mediated by CDK8/19 and validating our assay as a robust and quantitative method for CDK8/19 inhibition. View Full-Text
Keywords: CDK8; CDK19; CDK inhibitors; NFκB; thienopyridines; cell-based assays CDK8; CDK19; CDK inhibitors; NFκB; thienopyridines; cell-based assays
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MDPI and ACS Style

Li, J.; Ji, H.; Porter, D.C.; Broude, E.V.; Roninson, I.B.; Chen, M. Characterizing CDK8/19 Inhibitors through a NFκB-Dependent Cell-Based Assay. Cells 2019, 8, 1208. https://doi.org/10.3390/cells8101208

AMA Style

Li J, Ji H, Porter DC, Broude EV, Roninson IB, Chen M. Characterizing CDK8/19 Inhibitors through a NFκB-Dependent Cell-Based Assay. Cells. 2019; 8(10):1208. https://doi.org/10.3390/cells8101208

Chicago/Turabian Style

Li, Jing; Ji, Hao; Porter, Donald C.; Broude, Eugenia V.; Roninson, Igor B.; Chen, Mengqian. 2019. "Characterizing CDK8/19 Inhibitors through a NFκB-Dependent Cell-Based Assay" Cells 8, no. 10: 1208. https://doi.org/10.3390/cells8101208

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