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Selective inhibition of BET bromodomains

Abstract

Epigenetic proteins are intently pursued targets in ligand discovery. So far, successful efforts have been limited to chromatin modifying enzymes, or so-called epigenetic ‘writers’ and ‘erasers’. Potent inhibitors of histone binding modules have not yet been described. Here we report a cell-permeable small molecule (JQ1) that binds competitively to acetyl-lysine recognition motifs, or bromodomains. High potency and specificity towards a subset of human bromodomains is explained by co-crystal structures with bromodomain and extra-terminal (BET) family member BRD4, revealing excellent shape complementarity with the acetyl-lysine binding cavity. Recurrent translocation of BRD4 is observed in a genetically-defined, incurable subtype of human squamous carcinoma. Competitive binding by JQ1 displaces the BRD4 fusion oncoprotein from chromatin, prompting squamous differentiation and specific antiproliferative effects in BRD4-dependent cell lines and patient-derived xenograft models. These data establish proof-of-concept for targeting protein–protein interactions of epigenetic ‘readers’, and provide a versatile chemical scaffold for the development of chemical probes more broadly throughout the bromodomain family.

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Figure 1: Structure and selectivity of JQ1.
Figure 2: Characterization of BET complexes with (+)-JQ1.
Figure 3: Binding site comparison between N- and C-terminal bromodomains in complex with (+)-JQ1.
Figure 4: JQ1 binds BRD4 competitively with chromatin resulting in differentiation and growth arrest of NMC cells.
Figure 5: JQ1 promotes differentiation, tumour regression and improved survival in murine models of NMC.

Accession codes

Primary accessions

Protein Data Bank

Data deposits

Atomic coordinates and structure factors for the reported crystal structures have been deposited with the Protein Data Bank under accession codes 2OSS, 3MXF and 3ONI.

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Acknowledgements

We are grateful to U. Oppermann, S. Müller, S. Sallan, C. Lathan, P. Rahl, R. Young, K. Lee and K. Shaw for discussions and sharing unpublished information; K. Agu, S. Johnston and L. Li for analytical chemistry support; J. Daley for flow cytometry support; T. Bowman, T. Caron, C. Marvin and S. Rodig for immunohistochemistry; and A. Bass for sharing cell lines. The Structural Genomics Consortium is a registered charity (number 1097737) that receives funds from the Canadian Institutes for Health Research, the Canadian Foundation for Innovation, Genome Canada through the Ontario Genomics Institute, GlaxoSmithKline, Karolinska Institutet, the Knut and Alice Wallenberg Foundation, the Ontario Innovation Trust, the Ontario Ministry for Research and Innovation, Merck & Co., Inc., the Novartis Research Foundation, the Swedish Agency for Innovation Systems, the Swedish Foundation for Strategic Research and the Wellcome Trust. This research was supported by a Graduate Fellowship from the Chemistry-Biochemistry-Biology Interface Program at the University of Notre Dame, NIGMS T32-075762 (to Y.S.), the DF/HCC (to C.A.F. and J.E.B.), the National Institutes of Health, the Burroughs Wellcome Fund, and the Leukemia & Lymphoma Society (to J.E.B.).

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Contributions

P.F., J.Q., S.K. and J.E.B. designed the study, analysed data and wrote the manuscript. P.F. and S.P. performed and analysed biophysical studies. J.Q. and J.E.B. designed JQ1 and established the synthetic routes. Y.S. and O.W. completed docking and molecular dynamics studies. O.F. performed and analysed DSF. S.M. and N.L.T. contributed biochemical assays. M.R.M., M.P. and T.D.H. performed and analysed alpha-screen assays. W.B.S., M.J.C. and J.E.B. performed in vitro NMC studies and immunohistochemistry. E.M.M. performed flow cytometry studies. E.M.M. and N.W. performed proliferation studies. T.T.H., M.J.C., C.A.F. and J.E.B. completed FRAP studies. M.R.M. and B.S. performed expression analysis. Y.W., A.L.C. and A.L.K. completed in vivo efficacy studies. T.K. and I.F. expressed and purified proteins. S.K. and J.E.B. supervised the research.

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Correspondence to Stefan Knapp or James E. Bradner.

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The authors declare no competing financial interests.

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This file contains Supplementary Figures 1-17 with legends, Supplementary Methods, Supplementary Procedures, Supplementary Tables 1-5 and additional references. (PDF 3176 kb)

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Filippakopoulos, P., Qi, J., Picaud, S. et al. Selective inhibition of BET bromodomains. Nature 468, 1067–1073 (2010). https://doi.org/10.1038/nature09504

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