There is unmet need for chemical tools to explore the role of the Mediator complex in human pathologies ranging from cancer to cardiovascular disease. Here we determine that CCT251545, a small molecule WNT-pathway inhibitor discovered through cell-based screening, is a potent and selective chemical probe for the human Mediator complex-associated protein kinases CDK8 and CDK19 with >100-fold selectivity over 291 other kinases. X-ray crystallography demonstrates a Type 1 binding mode involving insertion of the CDK8 C-terminus into the ligand binding site. InReprints and permissions information is available online at http://www.nature.com/reprints/index.html.Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use
The
Mediator complex-associated cyclin-dependent kinase CDK8 has
been implicated in human disease, particularly in colorectal cancer
where it has been reported as a putative oncogene. Here we report
the discovery of 109 (CCT251921), a potent, selective,
and orally bioavailable inhibitor of CDK8 with equipotent affinity
for CDK19. We describe a structure-based design approach leading to
the discovery of a 3,4,5-trisubstituted-2-aminopyridine series and
present the application of physicochemical property analyses to successfully
reduce in vivo metabolic clearance, minimize transporter-mediated
biliary elimination while maintaining acceptable aqueous solubility.
Compound 109 affords the optimal compromise of in vitro
biochemical, pharmacokinetic, and physicochemical properties and is
suitable for progression to animal models of cancer.
Mediator-associated kinases CDK8/19 are context-dependent drivers or suppressors of tumorigenesis. Their inhibition is predicted to have pleiotropic effects, but it is unclear whether this will impact on the clinical utility of CDK8/19 inhibitors. We discovered two series of potent chemical probes with high selectivity for CDK8/19. Despite pharmacodynamic evidence for robust on-target activity, the compounds exhibited modest, though significant, efficacy against human tumor lines and patient-derived xenografts. Altered gene expression was consistent with CDK8/19 inhibition, including profiles associated with super-enhancers, immune and inflammatory responses and stem cell function. In a mouse model expressing oncogenic beta-catenin, treatment shifted cells within hyperplastic intestinal crypts from a stem cell to a transit amplifying phenotype. In two species, neither probe was tolerated at therapeutically-relevant exposures. The complex nature of the toxicity observed with two structurally-differentiated chemical series is consistent with on-target effects posing significant challenges to the clinical development of CDK8/19 inhibitors.DOI:
http://dx.doi.org/10.7554/eLife.20722.001
WNT
signaling is frequently deregulated in malignancy, particularly
in colon cancer, and plays a key role in the generation and maintenance
of cancer stem cells. We report the discovery and optimization of
a 3,4,5-trisubstituted pyridine 9 using a high-throughput
cell-based reporter assay of WNT pathway activity. We demonstrate
a twisted conformation about the pyridine–piperidine bond of 9 by small-molecule X-ray crystallography. Medicinal chemistry
optimization to maintain this twisted conformation, cognisant of physicochemical
properties likely to maintain good cell permeability, led to 74 (CCT251545), a potent small-molecule inhibitor of WNT signaling
with good oral pharmacokinetics. We demonstrate inhibition of WNT
pathway activity in a solid human tumor xenograft model with evidence
for tumor growth inhibition following oral dosing. This work provides
a successful example of hypothesis-driven medicinal chemistry optimization
from a singleton hit against a cell-based pathway assay without knowledge
of the biochemical target.
The mediator complex-associated cyclin dependent kinase CDK8 regulates β-catenin-dependent transcription following activation of WNT signaling. Multiple lines of evidence suggest CDK8 may act as an oncogene in the development of colorectal cancer. Here we describe the successful optimization of an imidazo-thiadiazole series of CDK8 inhibitors that was identified in a high-throughput screening campaign and further progressed by structure-based design. In several optimization cycles, we improved the microsomal stability, potency, and kinase selectivity. The initial imidazo-thiadiazole scaffold was replaced by a 3-methyl-1H-pyrazolo[3,4-b]-pyridine which resulted in compound 25 (MSC2530818) that displayed excellent kinase selectivity, biochemical and cellular potency, microsomal stability, and is orally bioavailable. Furthermore, we demonstrated modulation of phospho-STAT1, a pharmacodynamic biomarker of CDK8 activity, and tumor growth inhibition in an APC mutant SW620 human colorectal carcinoma xenograft model after oral administration. Compound 25 demonstrated suitable potency and selectivity to progress into preclinical in vivo efficacy and safety studies.
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