Targeted Degradation of CDK9 Potently Disrupts the MYC Transcriptional Network.

Cyclin-dependent kinase 9 (CDK9) coordinates signaling events that regulate RNA polymerase II (Pol II) pause-release states. It is an important co-factor for transcription factors, such as MYC, that drive aberrant cell proliferation when their expression is deregulated. CDK9 modulation offers an approach for attenuating dysregulation in such transcriptional programs. As a result, numerous drug development campaigns to inhibit CDK9 kinase activity have been pursued. More recently, targeted degradation has emerged as an attractive approach. However, comprehensive evaluation of degradation versus inhibition is still critically needed to assess the biological contexts in which degradation might offer superior therapeutic benefits. We validated that CDK9 inhibition triggers a compensatory mechanism that dampens its effect on MYC expression and found that this feedback mechanism was absent when the kinase is degraded. Importantly, CDK9 degradation is more effective than its inhibition for disrupting MYC transcriptional regulatory circuitry likely through the abrogation of both enzymatic and scaffolding functions of CDK9. Highlights: - KI-CDK9d-32 is a highly potent and selective CDK9 degrader. - KI-CDK9d-32 leads to rapid downregulation of MYC protein and mRNA transcripts levels. - KI-CDK9d-32 represses canonical MYC pathways and leads to a destabilization of nucleolar homeostasis. - Multidrug resistance ABCB1 gene emerged as the strongest resistance marker for the CDK9 PROTAC degrader.

Exploration of bromodomain ligand-linker conjugation sites for efficient CBP/p300 heterobifunctional degrader activity.

Synthesis of proteolysis targeting chimeras (PROTACs) involves conjugation of an E3 ligase binding ligand to a ligand targeting a protein of interest via a rigid or flexible chemical linker. The choice of linker conjugation site on these ligands can be informed by structural analysis of ligand-target binding modes, the feasibility of synthetic procedures to access specific sites, and computational modeling of predicted ternary complex formations. Small molecules that target bromodomains - epigenetic readers of lysine acetylation - typically offer several potential options for linker conjugation sites. Here we describe how varying the linker attachment site (exit vector) on a CBP/p300 bromodomain ligand along with linker length affects PROTAC degradation activity and ternary complex formation. Using kinetic live cell assays of endogenous CBP and p300 protein abundance and bead-based proximity assays for ternary complexes, we describe the structure-activity relationships of a diverse library of CBP/p300 degraders (dCBPs).