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Updated: Aug 1, 2025
A Protocol for Rapid Post-mortem Cell Culture of Diffuse Intrinsic Pontine Glioma DIPG
Published on: March 7, 2017
Targeting SWI/SNF ATPases in H3.3K27M diffuse intrinsic pontine gliomas
Mateus Mota1,2, Stefan R Sweha1,2, Matt Pun1,2,3,4
1Laboratory of Brain Tumor Metabolism and Epigenetics, Department of Pathology, University of Michigan, Ann Arbor, MI 48109.
Abstract:
Diffuse midline gliomas (DMGs) including diffuse intrinsic pontine gliomas (DIPGs) bearing lysine-to-methionine mutations in histone H3 at lysine 27 (H3K27M) are lethal childhood brain cancers. These tumors harbor a global reduction in the transcriptional repressive mark H3K27me3 accompanied by an increase in the transcriptional activation mark H3K27ac. We postulated that H3K27M mutations, in addition to altering H3K27 modifications, reprogram the master chromatin remodeling switch/sucrose nonfermentable (SWI/SNF) complex. The SWI/SNF complex can exist in two main forms termed BAF and PBAF that play central roles in neurodevelopment and cancer. Moreover, BAF antagonizes PRC2, the main enzyme catalyzing H3K27me3. We demonstrate that H3K27M gliomas show increased protein levels of the SWI/SNF complex ATPase subunits SMARCA4 and SMARCA2, and the PBAF component PBRM1. Additionally, knockdown of mutant H3K27M lowered SMARCA4 protein levels. The proteolysis targeting chimera (PROTAC) AU-15330 that simultaneously targets SMARCA4, SMARCA2, and PBRM1 for degradation exhibits cytotoxicity in H3.3K27M but not H3 wild-type cells. AU-15330 lowered chromatin accessibility measured by ATAC-Seq at nonpromoter regions and reduced global H3K27ac levels. Integrated analysis of gene expression, proteomics, and chromatin accessibility in AU-15330-treated cells demonstrated reduction in the levels of FOXO1, a key member of the forkhead family of transcription factors. Moreover, genetic or pharmacologic targeting of FOXO1 resulted in cell death in H3K27M cells. Overall, our results suggest that H3K27M up-regulates SMARCA4 levels and combined targeting of SWI/SNF ATPases in H3.3K27M can serve as a potent therapeutic strategy for these deadly childhood brain tumors.
Insights
Histone H3K27M mutations in diffuse midline gliomas reprogram the SWI/SNF complex. Targeting SWI/SNF ATPases, like SMARCA4, shows promise for treating these lethal childhood brain cancers.
Area of Science:
- Oncology
- Epigenetics
- Molecular Biology
Background:
- Diffuse midline gliomas (DMGs) with H3K27M mutations are aggressive pediatric brain cancers.
- These tumors exhibit altered histone modifications, specifically reduced H3K27me3 and increased H3K27ac.
- The SWI/SNF chromatin remodeling complex is implicated in cancer and neurodevelopment.
Purpose of the Study:
- To investigate the role of H3K27M mutations in reprogramming the SWI/SNF complex.
- To explore the therapeutic potential of targeting SWI/SNF components in H3K27M gliomas.
Main Methods:
- Analysis of SWI/SNF subunit protein levels in H3K27M gliomas.
- Utilizing a PROTAC (AU-15330) to degrade SWI/SNF ATPase subunits (SMARCA4, SMARCA2) and PBRM1.
- Assessing cytotoxicity, chromatin accessibility (ATAC-Seq), and gene expression changes upon PROTAC treatment.
- Investigating the role of FOXO1 in H3K27M cells.
Main Results:
- H3K27M gliomas show elevated levels of SMARCA4, SMARCA2, and PBRM1.
- PROTAC AU-15330 demonstrated cytotoxicity specifically in H3.3K27M cells.
- AU-15330 reduced chromatin accessibility and global H3K27ac levels.
- Targeting FOXO1 also induced cell death in H3K27M cells.
Conclusions:
- H3K27M mutations up-regulate SMARCA4, impacting SWI/SNF complex activity.
- Combined targeting of SWI/SNF ATPases presents a potential therapeutic strategy for H3K27M-driven pediatric brain tumors.
- FOXO1 is identified as a relevant downstream target in these tumors.
