Identification and analysis of novel CDK8 inhibitors

Chronic inflammation is associated with an assortment of ailments. Current anti-inflammatory treatments are non-specific and their effectiveness may vary based on the patient and illness. Identifying treatments targeting additional inflammatory signaling mediators may lead to novel therapeutics. Previous studies have shown cyclin-dependent kinase 8 (CDK8) to play an important role in mediating inflammatory signaling. Importantly, CDK8 is druggable and tailoring a small molecule for increased potency and selectivity may lead to novel therapeutics. This thesis sought to identify novel CDK8 inhibitors. Potential inhibitors were selected for analysis from a structure-based virtual screening (SBVS) protocol. To improve the screening hit rate, compounds were ranked by pharmacological scores. From the screening protocol in this study, 13 potential inhibitors were selected for evaluation. Enzymatic assays confirmed a CDK8 inhibitor, E966-0578. Further testing of the compound revealed favorable inhibitory concentration (IC50 1,684.4 nM). A total of 22 analogs were sourced to analyze the structural-activity relationship (SAR) to identify chemical components that can lead to CDK8 inhibitory activity. Analog F059-1017 was identified to exhibit even greater CDK8 potency (IC50 558.1 nM) and was selected for further analysis. Cellular assays revealed F059-1017 to have low cytotoxicity in vitro. Cells treated with F059-1017 not only displayed reduced CDK8 activity but also reduced phosphorylation and expression of downstream inflammatory signaling mediators. Small molecules are known to be associated with off-target activity, which could lead to unwanted side effects. Selectivity of F059-1017 was assessed through kinase profiling against the whole kinome. This showed F059-1017 to be a CDK8 selective inhibitor. Further analysis of substructures obtained from compounds with associated bioactivity against CMGC kinome family and CDK8 showed F059-1017 to contain CDK8 specific substructures. Taken together, these findings report F059-1017 to be a novel and selective CDK8 inhibitor and could serve as a lead compound for further optimization against inflammation.