Chemical probes for epigenetic proteins are essential tools for dissecting the molecular mechanisms for gene regulation and therapeutic development. The bromodomain and extra-terminal (BET) proteins are master transcriptional regulators. Despite promising therapeutic targets, selective small molecule inhibitors for a single bromodomain remain an unmet goal due to their high sequence similarity. Here, we address this challenge via a structure-activity relationship study using 1,4,5-trisubstituted imidazoles against the BRD4 N-terminal bromodomain (D1). Leading compounds 26 and 30 have 15 and 18 nM affinity against BRD4 D1 and over 500-fold selectivity against BRD2 D1 and BRD4 D2 via ITC. Broader BET selectivity was confirmed by fluorescence anisotropy, thermal shift, and CETSA. Despite BRD4 engagement, BRD4 D1 inhibition was unable to reduce c-Myc expression at low concentration in multiple myeloma cells. Conversely, for inflammation, IL-8 and chemokine downregulation were observed. These results provide new design rules for selective inhibitors of an individual BET bromodomain.
Bibliographical noteFunding Information:
This work was funded by Office of Discovery and Translation Grant at UMN, UL1 TR002494 (W.C.K.P), Center for Regenerative Medicine (V.S.), and the NIH MIRA awards, R35 GM140837-01 (W.C.K.P.) and R35 GM118047 (H.A.) A.D. was supported by the UMN Doctoral Dissertation Fellowship and a NIH chemistry–biology interface training grant (T32-GM008700/T32-GM132029-01). H.Z. was supported by UMN IEM Engineering in Medicine Doctoral Fellowship 2020. D.H. acknowledges funding from the Masonic Cancer Center at the University of Minnesota with resources from Minnesota Masonic Charities. R.L. is supported in part by St. Jude Cancer Center Support grant CA21765 and by the American Lebanese Syrian Associated Charities (ALSAC). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institute of Health or other funding agencies. X-ray diffraction data were collected at the Northeastern Collaborative Access Team beamlines, which are supported by NIGMS P30 GM124165. The Pilatus 6 M detector on 24-ID-C beamline is funded by a NIH-ORIP HEI grant (S10 RR029205).
© 2022 American Chemical Society.
- Binding Sites
- Cell Cycle Proteins/antagonists & inhibitors
- Cell Line, Tumor
- Drug Design
- Molecular Structure
- Protein Binding
- Protein Domains
- Proto-Oncogene Proteins c-myc/metabolism
- Structure-Activity Relationship
- Transcription Factors/antagonists & inhibitors
PubMed: MeSH publication types
- Research Support, Non-U.S. Gov't
- Journal Article
- Research Support, N.I.H., Extramural