BRD4 associates with p53 in DNMT3A‐mutated leukemia cells and is implicated in apoptosis by the bromodomain inhibitor JQ1

Abstract

The bromodomain and extra terminal (BET) family protein bromodomain containing protein 4 (BRD4) is an epigenetic regulator recently identified as a therapeutic target for several hematological cancers, notably mixed lineage leukemia-fusion acute myeloid leukemia (MLL-AML). Here, we show that the BRD4 bromodomain inhibitor JQ1 is highly active against the p53-wild-type Ontario Cancer Institute (OCI)-AML3 cell line which carries mutations in nucleophosmin (NPM1) and DNA methyltransferase 3 (DNMT3A) genes commonly associated with poor prognostic disease. We find that JQ1 causes caspase 3/7-mediated apoptosis and DNA damage response in these cells. In combination studies, we show that histone deacetylase (HDAC) inhibitors, the HDM2 inhibitor Nutlin-3, and the anthracycline daunorubicin all enhance the apoptotic response of JQ1. These compounds all induce activation of p53 suggesting that JQ1 might sensitize AML cells to p53-mediated cell death. In further experiments, we show that BRD4 associates with acetylated p53 but that this association is not inhibited by JQ1 indicating that the protein–protein interaction does not involve bromodomain binding of acetylated lysines. Instead, we propose that JQ1 acts to prevent BRD4-mediated recruitment of p53 to chromatin targets following its activation in OCI-AML3 cells resulting in cell cycle arrest and apoptosis in a c-MYC-independent manner. Our data suggest that BET bromodomain inhibition might enhance current chemotherapy strategies in AML, notably in poor-risk DNMT3A/NPM1-mutated disease.

BRD4 localization to lineage-specific enhancers is associated with a distinct transcription factor repertoire

“…In turn, RNA Polymerase II (RNAPII) can be phosphorylated at Ser-2 residues within the carboxy-terminal heptapeptide repeat domain (CTD), which serves as a hallmark for transcriptional elongation to recruit various proteins involved in transcription-associated processes. In addition to binding to acetylated histone proteins, BRD4 has recently also been shown to directly interact with several transcription factors (TFs) including TWIST, p53, C/EBPα and C/EBPβ, ERG and NFκB in both acetylation-dependent and -independent manners (4–8). Moreover, proteomic analyses revealed the ability of the ET domain of BRD4 to interact with other chromatin-associated proteins like NSD3 and JMJD6 (9,10).…”

5. Chemische Open-Access-Sonden für epigenetische Zielstrukturen

“…Durch Interaktion mit dem Androgenrezeptor oder dem Tumorsuppressor 53 reguliert BRD4 die Androgen-oder p53-abhän-gigen Gene. Entsprechend zeigten BET-Inhibitoren vielversprechende Effekte bei Prostatakrebs oder in p53-abhängigen Leukämiemodellen (Asangani et al, 2014;Stewart et al, 2013 (Picaud et al, 2013) sie erzeugt. Präklinische Experimente bei Leukämie zeigen jedoch schon, dass sich auch Resistenzen gegen BET-Behandlung bilden können.…”

Epigenetic Therapy in Acute Myeloid Leukemia: Current and Future Directions

“…The bromodomain and extra terminal (BET) family of proteins include a number of Bromodomain-containing “reader” proteins (BRD [BRD2, BRD3, BRD4, and BRDT]) which use their acetyl-lysine recognition motifs (bromodomains) to read the post-translational acetylated motifs of histones and influence transcription of target genes 93, 94 . Dysregulation of BET adaptors has been shown to contribute to leukemogenesis across a variety of AML subtypes driven by different mutations 95 .…”

“…Inhibition of BRD4 using small hairpin inhibitory RNAs (shRNAs) or small molecule inhibitors (e.g. JQ1 and I-BET151) induces robust in vivo and in vitro anti-leukemic effects via down-regulation of oncogenes, suggesting this approach could be a potentially effective therapeutic strategy in some subtypes of AML 94, 95, 97–99 . Preclinical studies of the selective small molecule BRD4 inhibitors JQ1 and I-BET151 showed terminal myeloid differentiation and elimination of leukemia stem cells in MLL-rearranged AML, synergistic activity with HDAC inhibitors and anthracyclines to enhance p53-mediated apoptosis in DNMT3A / NPM1 -mutated leukemic cell lines, and activity against NPM1-mutated AML and JAK2V617F-driven neoplasms 94, 95, 98, 99 .…”