Triple-negative breast cancer (TNBC) is a heterogeneous and clinically aggressive disease for which there is no targeted therapy1,2,3. BET bromodomain inhibitors, which have shown efficacy in several models of cancer4,5,6, have not been evaluated in TNBC. These inhibitors displace BET bromodomain proteins such as BRD4 from chromatin by competing with their acetyl-lysine recognition modules, leading to inhibition of oncogenic transcriptional programs7,8,9. Here we report the preferential sensitivity of TNBCs to BET bromodomain inhibition in vitro and in vivo, establishing a rationale for clinical investigation and further motivation to understand mechanisms of resistance. In paired cell lines selected for acquired resistance to BET inhibition from previously sensitive TNBCs, we failed to identify gatekeeper mutations, new driver events or drug pump activation. BET-resistant TNBC cells remain dependent on wild-type BRD4, which supports transcription and cell proliferation in a bromodomain-independent manner. Proteomic studies of resistant TNBC identify strong association with MED1 and hyper-phosphorylation of BRD4 attributable to decreased activity of PP2A, identified here as a principal BRD4 serine phosphatase. Together, these studies provide a rationale for BET inhibition in TNBC and present mechanism-based combination strategies to anticipate clinical drug resistance.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.


Primary accessions

Gene Expression Omnibus

Data deposits

RNA-seq, ChIP-seq, and Chem-seq data have been deposited in the NCBI GEO database with the accession number GSE63584.


  1. 1.

    et al. Outcomes by tumor subtype and treatment pattern in women with small, node-negative breast cancer: a multi-institutional study. J. Clin. Oncol. 32, 2142–2150 (2014)

  2. 2.

    et al. Identification of human triple-negative breast cancer subtypes and preclinical models for selection of targeted therapies. J. Clin. Invest. 121, 2750–2767 (2011)

  3. 3.

    et al. Dissecting the heterogeneity of triple-negative breast cancer. J. Clin. Oncol. 30, 1879–1887 (2012)

  4. 4.

    et al. Targeting MYCN in neuroblastoma by BET bromodomain inhibition. Cancer Discov. 3, 308–323 (2013)

  5. 5.

    et al. BET bromodomain inhibition as a therapeutic strategy to target c-Myc. Cell 146, 904–917 (2011)

  6. 6.

    et al. Selective inhibition of BET bromodomains. Nature 468, 1067–1073 (2010)

  7. 7.

    & BET domain co-regulators in obesity, inflammation and cancer. Nature Rev. Cancer 12, 465–477 (2012)

  8. 8.

    et al. Discovery and characterization of super-enhancer-associated dependencies in diffuse large B cell lymphoma. Cancer Cell 24, 777–790 (2013)

  9. 9.

    et al. Super-enhancers in the control of cell identity and disease. Cell 155, 934–947 (2013)

  10. 10.

    et al. Comprehensive genomic analysis identifies novel subtypes and targets of triple-negative breast cancer. Clin. Cancer Res. 21, 1688–1698 (2015)

  11. 11.

    et al. Genome-wide localization of small molecules. Nature Biotechnol. 32, 92–96 (2014)

  12. 12.

    et al. Selective inhibition of tumor oncogenes by disruption of super-enhancers. Cell 153, 320–334 (2013)

  13. 13.

    et al. Master transcription factors and mediator establish super-enhancers at key cell identity genes. Cell 153, 307–319 (2013)

  14. 14.

    et al. The OCT4 pseudogene POU5F1B is amplified and promotes an aggressive phenotype in gastric cancer. Oncogene 34, 199–208 (2015)

  15. 15.

    HIF-1 mediates metabolic responses to intratumoral hypoxia and oncogenic mutations. J. Clin. Invest. 123, 3664–3671 (2013)

  16. 16.

    et al. Functional synergies yet distinct modulators affected by genetic alterations in common human cancers. Cancer Res. 71, 3471–3481 (2011)

  17. 17.

    et al. The JAK2/STAT3 signaling pathway is required for growth of CD44+CD24 stem cell-like breast cancer cells in human tumors. J. Clin. Invest. 121, 2723–2735 (2011)

  18. 18.

    & BH3 profiling in whole cells by fluorimeter or FACS. Methods 61, 156–164 (2013)

  19. 19.

    et al. Drug-induced death signaling strategy rapidly predicts cancer response to chemotherapy. Cell 160, 977–989 (2015)

  20. 20.

    et al. Endogenous purification reveals GREB1 as a key estrogen receptor regulatory factor. Cell Reports 3, 342–349 (2013)

  21. 21.

    , , , & Phospho switch triggers Brd4 chromatin binding and activator recruitment for gene-specific targeting. Mol. Cell 49, 843–857 (2013)

  22. 22.

    et al. PP2A inhibition determines poor outcome and doxorubicin resistance in early breast cancer and its activation shows promising therapeutic effects. Oncotarget 6, 4299–4314 (2015)

  23. 23.

    & Multiple pathways regulated by the tumor suppressor PP2A in transformation. Trends Mol. Med. 14, 152–160 (2008)

  24. 24.

    , & Protein phosphatase 2A regulatory subunits and cancer. Biochim. Biophys. Acta 1795, 1–15 (2009)

  25. 25.

    et al. Phenothiazines induce PP2A-mediated apoptosis in T cell acute lymphoblastic leukemia. J. Clin. Invest. 124, 644–655 (2014)

  26. 26.

    et al. Transcriptional plasticity promotes primary and acquired resistance to BET inhibition. Nature 525, 543–547 (2015)

  27. 27.

    et al. BET inhibitor resistance emerges from leukaemia stem cells. Nature 525, 538–542 (2015)

  28. 28.

    & Quantitative analysis of dose-effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv. Enzyme Regul. 22, 27–55 (1984)

  29. 29.

    et al. Molecular profiling of human mammary gland links breast cancer risk to a p27+ cell population with progenitor characteristics. Cell Stem Cell 13, 117–130 (2013)

  30. 30.

    et al. The CRAPome: a contaminant repository for affinity purification-mass spectrometry data. Nature Methods 10, 730–736 (2013)

  31. 31.

    , & TopHat: discovering splice junctions with RNA-Seq. Bioinformatics 25, 1105–1111 (2009)

  32. 32.

    et al. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nature Biotechnol. 28, 511–515 (2010)

  33. 33.

    , , & Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 10, R25 (2009)

  34. 34.

    et al. Model-based analysis of ChIP-Seq (MACS). Genome Biol. 9, R137 (2008)

  35. 35.

    et al. Transcriptional amplification in tumor cells with elevated c-Myc. Cell 151, 56–67 (2012)

  36. 36.

    et al. NF-κB directs dynamic super enhancer formation in inflammation and atherogenesis. Mol. Cell 56, 219–231 (2014)

Download references


We thank D. Silver and members of the Polyak and Bradner laboratories for their critical reading of this manuscript and useful discussions. We thank G. Brown for help with creating the word cloud figures. This work was supported by the NIH DF/HCC SPORE in Breast Cancer CA168504 (K.P., E.P.W., I.E.K., D.D., W.T.B., and J.E.B.), CA080111 (K.P. and M.B.), and CA103867 (C.M.C.), Susan G. Komen Foundation (S.S.), CPRIT RP110471 and RP140367 (C.M.C), Welch Foundation (C.M.C.), US Department of Defense CDMRP BC122003 (S.X.L.) and CA120184 (C.Y.L.), Princess Margaret Cancer Foundation (H.H.H.), Canada Foundation for Innovation and Ontario Research Fund CFI32372 (H.H.H.), NSERC discovery grant RGPIN-2015-04658 (H.H.H.), and the Harvard Ludwig Center for Cancer Research (J.E.B., M.B. and K.P.).

Author information

Author notes

    • Shaokun Shu
    • , Charles Y. Lin
    • , Housheng Hansen He
    •  & Robert M. Witwicki

    These authors contributed equally to this work.


  1. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts 02215, USA

    • Shaokun Shu
    • , Charles Y. Lin
    • , Housheng Hansen He
    • , Robert M. Witwicki
    • , Doris P. Tabassum
    • , Justin M. Roberts
    • , Michalina Janiszewska
    • , Sung Jin Huh
    • , Jeremy Ryan
    • , Ernest Doherty
    • , Daniel G. Stover
    • , Muhammad B. Ekram
    • , Guillermo Peluffo
    • , Jonathan Brown
    • , Ian E. Krop
    • , Deborah Dillon
    • , Michael McKeown
    • , Christopher Ott
    • , Jun Qi
    • , Min Ni
    • , Eric P. Winer
    • , Antony Letai
    • , Henry W. Long
    • , Myles Brown
    • , James E. Bradner
    •  & Kornelia Polyak
  2. Department of Medicine, Brigham and Women’s Hospital, and Department of Medicine, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Shaokun Shu
    • , Charles Y. Lin
    • , Housheng Hansen He
    • , Robert M. Witwicki
    • , Michalina Janiszewska
    • , Sung Jin Huh
    • , Jeremy Ryan
    • , Daniel G. Stover
    • , Muhammad B. Ekram
    • , Guillermo Peluffo
    • , Jonathan Brown
    • , Ian E. Krop
    • , Michael McKeown
    • , Christopher Ott
    • , Jun Qi
    • , Min Ni
    • , Eric P. Winer
    • , Antony Letai
    • , William T. Barry
    • , Myles Brown
    • , James E. Bradner
    •  & Kornelia Polyak
  3. Department of Biostatistics and Computational Biology, Dana-Farber Cancer Institute, and Department of Biostatistics, Harvard School of Public Health, Boston, Massachusetts 02115, USA

    • Housheng Hansen He
    • , Hao Guo
    • , William T. Barry
    • , X. Shirley Liu
    •  & Clifford A. Meyer
  4. Princess Margaret Cancer Center/University Health Network, Toronto, Ontario M5G1L7, Canada

    • Housheng Hansen He
    •  & Yi Liang
  5. Department of Medical Biophysics, University of Toronto, Toronto, Ontario M5G2M9, Canada

    • Housheng Hansen He
  6. Harvard University, Cambridge, Massachusetts 02138, USA

    • Ernest Doherty
  7. Cancer Research UK, Cambridge Institute, University of Cambridge, Cambridge CB2 0RE, UK

    • Hisham Mohammed
    • , Clive D’Santos
    •  & Jason S. Carroll
  8. Department of Pathology, Brigham and Women’s Hospital, and Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA

    • Deborah Dillon
  9. Center for Functional Cancer Epigenetics, Dana-Farber Cancer Institute, Boston, Massachusetts 02215, USA

    • Prakash K. Rao
    • , Melissa Duarte
    • , Henry W. Long
    • , Myles Brown
    • , X. Shirley Liu
    •  & Kornelia Polyak
  10. Simmons Comprehensive Cancer Center, Departments of Biochemistry and Pharmacology, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA

    • Shwu-Yuan Wu
    •  & Cheng-Ming Chiang
  11. Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA

    • Lars Anders
    •  & Richard A. Young
  12. Broad Institute, Cambridge, Massachusetts 02142, USA

    • X. Shirley Liu
    • , James E. Bradner
    •  & Kornelia Polyak


  1. Search for Shaokun Shu in:

  2. Search for Charles Y. Lin in:

  3. Search for Housheng Hansen He in:

  4. Search for Robert M. Witwicki in:

  5. Search for Doris P. Tabassum in:

  6. Search for Justin M. Roberts in:

  7. Search for Michalina Janiszewska in:

  8. Search for Sung Jin Huh in:

  9. Search for Yi Liang in:

  10. Search for Jeremy Ryan in:

  11. Search for Ernest Doherty in:

  12. Search for Hisham Mohammed in:

  13. Search for Hao Guo in:

  14. Search for Daniel G. Stover in:

  15. Search for Muhammad B. Ekram in:

  16. Search for Guillermo Peluffo in:

  17. Search for Jonathan Brown in:

  18. Search for Clive D’Santos in:

  19. Search for Ian E. Krop in:

  20. Search for Deborah Dillon in:

  21. Search for Michael McKeown in:

  22. Search for Christopher Ott in:

  23. Search for Jun Qi in:

  24. Search for Min Ni in:

  25. Search for Prakash K. Rao in:

  26. Search for Melissa Duarte in:

  27. Search for Shwu-Yuan Wu in:

  28. Search for Cheng-Ming Chiang in:

  29. Search for Lars Anders in:

  30. Search for Richard A. Young in:

  31. Search for Eric P. Winer in:

  32. Search for Antony Letai in:

  33. Search for William T. Barry in:

  34. Search for Jason S. Carroll in:

  35. Search for Henry W. Long in:

  36. Search for Myles Brown in:

  37. Search for X. Shirley Liu in:

  38. Search for Clifford A. Meyer in:

  39. Search for James E. Bradner in:

  40. Search for Kornelia Polyak in:


S.S. performed cell culture, xenograft, ChIP-seq, and RNA-seq experiments, and data analyses. C.Y.L. and C.A.M. performed genomic data analyses. H.H.H. helped with ChIP-seq and RNA-seq experiments and data analyses. R.M.W. performed cell culture, ChIP-seq experiments and data analyses. J.M.R. performed synergy studies. D.P.T. helped with immunofluorescence staining. M.J. and S.J.H. helped with confocal microscopy and image quantification. Y.L. helped with BRD4 ChIP-seq. M.B.E. and G.P. helped with cell cycle studies. E.D. helped with generating and testing BRD4 mutants. J.B. and L.A. performed Chem-seq. H.M., C.D. and J.S.C. conducted proteomic experiments and data analyses. C.O. and M.M. performed drug sensitivity screens. J.Q. synthesized BBI compounds. M.N. generated shRNA constructs. D.D., I.E.K. and E.P.W. generated the TMA and linked to clinical data. H.G., D.G.S. and W.T.B. performed TMA and statistical analyses. J.R. and A.L. performed BH3 profiling and data analyses. C.-M.C. and S.-Y.W. provided phospho-BRD4 antibody. P.K.R. and M.D. generated RNA-seq libraries. K.P. supervised with help from J.E.B., X.S.L., M.B., R.A.Y. and H.L. All authors helped to design the study and write the manuscript.

Competing interests

J.E.B. and R.A.Y. are founders of Syros Pharmaceuticals, J.E.B. is the founder of Tensha Therapeutics. K.P. receives research support from and is a consultant for Novartis Oncology.

Corresponding authors

Correspondence to James E. Bradner or Kornelia Polyak.

Extended data

Supplementary information

PDF files

  1. 1.

    Supplementary Figure 1

    This file contains uncropped gels.

Zip files

  1. 1.

    Supplementary Tables

    This zipped file contains Supplementary Tables 1-10 and a Supplementary Table guide.

Excel files

  1. 1.

    Supplementary Data

    This file contains Source data for the graphs.

About this article

Publication history






Further reading


By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Newsletter Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing