Discovery of LL-K8-22: A Selective, Durable, and Small-Molecule Degrader of the CDK8-Cyclin C Complex

Abstract

The CDK8−cyclin C complex is an important anti-tumor target, but unlike CDK8, cyclin C remains undruggable. Modulators regulating cyclin C activity directly are still under development. Here, a series of hydrophobic tagging-based degraders of the CDK8−cyclin C complex were designed, synthesized, and evaluated to identify the first dual degrader, LL-K8-22, which induced selective and synchronous degradation of CDK8 and cyclin C. Proteomic and immunoblot studies exhibited that LL-K8-22 significantly degraded CDK8 without reducing CDK19 and did not degrade other cyclin proteins except cyclin C. Moreover, LL-K8-22 showed enhanced anti-proliferative effects over its parental molecule, BI-1347, with potency increased by 5-fold in MDA-MB-468 cells. LL-K8-22 exhibited more pronounced effects on CDK8−cyclin C downstream signaling than BI-1347, suppressing STAT1 phosphorylation more persistently. RNA-sequencing analysis revealed that LL-K8-22 inhibited E2F-and MYC-driven carcinogenic transcriptional programs. Overall, LL-K8-22 is the first-in-class degrader of cyclin C and would be useful for studying the unknown functions of cyclin C.

Small-Molecule Hydrophobic Tagging: A Promising Strategy of Druglike Technology for Targeted Protein Degradation

“…90 Very recently, on the basis of the similar hydrophobic tagging strategy, Luo et al designed CDK8-cyclin C degraders and successfully degraded the undruggable target cyclin C and enhanced antitumor potency (Table 1). 95 Overall, these results indicated that menthoxyacetyl may be a useful pharmacological tool for investigating the unrecognized functions of complex proteins.…”

Uncovering PROTAC Sensitivity and Efficacy by Multidimensional Proteome Profiling: A Case for STAT3

“…The DIA method eliminates the need for sample labeling, simplifies the workflow, and avoids additional expenses associated with the purchase or synthesis of labeling reagents. We pioneered to apply DIA-MS to PROTAC research, achieving quicker and more flexible processing of multiple samples than that with isobaric labeling quantitative analysis in customized experimental designs. − …”

“…In previous studies, researchers used MS-based proteomics to determine the efficacy of PROTAC degraders with a single condition further with a series of treatment time points or doses. They mainly focused on the degradation of target proteins and differentially expressed proteins (DEPs) proteome-wide. − ,, The limited proteomic data analysis is insufficient to understand the PROTAC efficacy and sensitivity. In this study, we applied DIA-based MS to multidimensional proteomic profiling of PROTAC, named “DIA-MPP,” for in-depth and comprehensive proteome profiling with multiple conditions.…”

Discovery of HyT‐Based Degraders of CDK9‐Cyclin T1 Complex

“…The HyT technique has proven to be highly efficient in its ability to specifically dismantle multiple targets, including the AR, [47] Her3, [48] Tau, [49] EZH2, [50] ALK, [51] CDK9, [52] CDK8, [53] and PARP [54] . Adamantane is the most common hydrophobic tag, [55] while Boc 3 Arg is another effective option that can also trigger proteolysis.…”

“…The proposed mechanism is that hydrophobic moiety induced protein of interest (POI) loses its original folded state and then recruits endogenous chaperone, [45] or hydrophobic moiety recognized by a chaperone, thereby mediating proteasomal degradation of the POI. [46] The HyT technique has proven to be highly efficient in its ability to specifically dismantle multiple targets, including the AR, [47] Her3, [48] Tau, [49] EZH2, [50] ALK, [51] CDK9, [52] CDK8, [53] and PARP. [54] Adamantane is the most common hydrophobic tag, [55] while Boc 3 Arg is another effective option that can also trigger proteolysis.…”