Craig R. Stumpf

Supplementary Table S1 shows CRISPR screen hits

Supplementary file containing Figures S1-3 and Tables S1-4

Supplementary Figure S3 shows megakaryocyte colony formation data

Supplementary Table S4 shows information for the antibodies used in these studies

Supplemental Figure S2 shows PLX-3618 binding data to both bromodomains of BRD4

Supplemental Figure S1 shows BRD4 degradation kinetics in the HiBiT reporter cell line

Supplemental Table S3 shows the protein expression plasmids used in these studies

Supplementary Table S2 show anti-proliferative activity in AML cell lines (n = 2)

Targeted protein degradation (TPD) using the ubiquitin proteasome system (UPS) is a rapidly growing drug discovery modality to eliminate pathogenic proteins. Strategies for TPD have focused on heterobifunctional degraders that often suffer from poor drug-like properties, and molecular glues that rely on serendipitous discovery. Monovalent “direct” degraders represent an alternative approach, in which small molecules bind to a target protein and induce degradation of that protein through the recruitment of an E3 ligase complex. Using an ultra-high throughput cell-based screening platform, degraders of the bromodomain extraterminal protein BRD4 were identified and optimized to yield a lead compound, PLX-3618. In this paper, we demonstrate that PLX-3618 elicited UPS-mediated selective degradation of BRD4, resulting in potent antitumor activity in vitro and in vivo. Characterization of the degradation mechanism identified DCAF11 as the E3 ligase required for PLX-3618-mediated degradation of BRD4. Protein–protein interaction studies verified a BRD4:PLX-3618:DCAF11 ternary complex, and mutational studies provided further insights into the DCAF11-mediated degradation mechanism. Collectively, these results demonstrate the discovery and characterization of a novel small molecule that selectively degrades BRD4 through the recruitment of the E3 substrate receptor, DCAF11, and promotes potent antitumor activity in vivo.

Supplementary Table S1 shows CRISPR screen hits