Combined treatment with CBP and BET inhibitors reverses inadvertent activation of detrimental super enhancer programs in DIPG cells

Maria Wiese,Angel M Carcaboso,Esther Hulleman,Christof M Kramm,Steven A Johnsen,Gunther Nussbaumer,Klaudia Kubiak,Gerrit H Gielen,Feda H Hamdan,Christopher Diederichs

doi:10.1038/s41419-020-02800-7

Abstract

Diffuse intrinsic pontine gliomas (DIPG) are the most aggressive brain tumors in children with 5-year survival rates of only 2%. About 85% of all DIPG are characterized by a lysine-to-methionine substitution in histone 3, which leads to global H3K27 hypomethylation accompanied by H3K27 hyperacetylation. Hyperacetylation in DIPG favors the action of the Bromodomain and Extra-Terminal (BET) protein BRD4, and leads to the reprogramming of the enhancer landscape contributing to the activation of DIPG super enhancer-driven oncogenes. The activity of the acetyltransferase CREB-binding protein (CBP) is enhanced by BRD4 and associated with acetylation of nucleosomes at super enhancers (SE). In addition, CBP contributes to transcriptional activation through its function as a scaffold and protein bridge. Monotherapy with either a CBP (ICG-001) or BET inhibitor (JQ1) led to the reduction of tumor-related characteristics. Interestingly, combined treatment induced strong cytotoxic effects in H3.3K27M-mutated DIPG cell lines. RNA sequencing and chromatin immunoprecipitation revealed that these effects were caused by the inactivation of DIPG SE-controlled tumor-related genes. However, single treatment with ICG-001 or JQ1, respectively, led to activation of a subgroup of detrimental super enhancers. Combinatorial treatment reversed the inadvertent activation of these super enhancers and rescued the effect of ICG-001 and JQ1 single treatment on enhancer-driven oncogenes in H3K27M-mutated DIPG, but not in H3 wild-type pedHGG cells. In conclusion, combinatorial treatment with CBP and BET inhibitors is highly efficient in H3K27M-mutant DIPG due to reversal of inadvertent activation of detrimental SE programs in comparison with monotherapy.

Highlights

Diffuse intrinsic pontine gliomas (DIPG) are pediatric high-grade gliomas accounting for approximately 10–15% of pediatric central nervous system tumors
To evaluate if H3K27M mutation leads to changes in cell growth in vitro, we investigated the potential of H3 wild-type (H3WT) and H3K27M-mut pediatric high-grade gliomas (pedHGG) and DIPG cell lines to form gliomaspheres
In accordance with previous reports demonstrating that H3K27M mutation confers a more stem cell-like phenotype2, we observed a stronger potential of the H3.3K27M-mutant VUMCDIPG-A and HSJD-DIPG-007 cell lines to form gliomaspheres under stem cell conditions

Summary

Introduction

Diffuse intrinsic pontine gliomas (DIPG) are pediatric high-grade gliomas (pedHGG) accounting for approximately 10–15% of pediatric central nervous system tumors. Acetylation at lysine 27 of histone 3 (H3K27ac) is mediated by Official journal of the Cell Death Differentiation Association. Wiese et al Cell Death and Disease (2020)11:673 the KAT3 family of acetyltransferases (HATs) including CREB-binding protein (CBP). In addition to its function as an acetyltransferase, CBP serves as a protein bridge that connects other transcription factors to the transcription machinery, and acts as scaffold protein during the formation of multicomponent transcriptional regulatory complexes. CBP is necessary for the recruitment of diverse transcription factors, including the Bromodomain and Extra-Terminal (BET) protein BRD4, to genomic regions displaying an enrichment of acetylated nucleosomes, thereby promoting transcription. BRD4 and CBP are promising pharmaceutical targets for treatment of DIPG and pedHGG5,6

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