CCT251545

CCT251545 was optimized from a phenotypic screening hit for inhibitors of WNT pathway activity. Impressively, this optimization was done without the benefit of structure-based design. CCT251545 has potent cell-based activity and good oral pharmokinetics (PK). The cellular targets of CCT251545 were subsequently determined to be CDK8/19 using SILAC-based affinity capture and rationalized by X-ray crystallography. The affinity-capture experiment suggests very high specificity for CDK8/19. However, direct kinase selectivity profiling experiments did not include the positive controls of CDK8 or CDK19. Therefore, a specificity score cannot be determined nor can it be shown that the compound was active in the assay. Efforts to connect CDK8/19 binding to cellular activity (e.g., WNT reporter gene-assay inhibition) would benefit from the use of a dominant drug resistant allele of CDK8/19. Such an allele could be expressed in cells to circumvent CDK8/19 inhibitory activity of CCT251545 and provide strong evidence that modulation of CDK8/19 by CCT251545 accounts for the observed cellular activity. Regardless, CCT251545 is a potent inhibitor of CDK8/19, has good PK properties and is expected to be highly specific for CDK8/19.

CCT251545 is a high-affinity ligand of CDK8 and CDK19 with biochemical IC50s for these targets of 7 and 6 nM, respectively. The compound exhibits high selectivity compared with other targets, as demonstrated by proteomics data (SILAC), and suggested by testing in an expanded kinase panel. CCT251545 exhibits potent target engagement in cells (reduction of phospho-STAT1-Ser727; IC50=9 nM) at concentrations consistent with those that reduced WNT-reporter activity in the same cell lines. In vivo pharmacokinetic properties were investigated in mice, rats, and dogs and found adequate for in vivo testing (moderate clearance and F >50% at a low oral dose of 0.5 mg/kg). CCT25145 inhibits the growth of MMTV-WNT-1 breast cancers transplanted from genetically engineered mice as well as APC-mutant SW620 human colorectal cancer xenografts, consistent with in vivo pharmacodynamic data (reduction of p-STATSer727) and at exposure levels that exceed cellular GI50's. Taken together, CCT251545 is a well suited and selective probe for CDK8/19 with proven cellular and in vivo target engagement and efficacy.