Oral Presentation 37th Lorne Cancer Conference 2025

Chromatin remodelling with combined FACT and BET inhibition disrupts oncogenic transcription in Diffuse Midline Glioma (113828)

Holly Holliday 1 2 , Aaminah Khan 1 2 , Anahid Ehteda 1 2 , Samuel E Ross 3 , Nisitha Jayatilleke 1 , Anjana Gopalakrishnan 1 , Hieu Nguyen 1 , Xinyi Guo 1 , Jie Liu 1 , Eyden Wang 1 , Alvin Lee 1 , Yolanda Colino Sanguino 1 2 , Daisy Kavanagh 4 , Robert J Weatheritt 4 , Fatima Valdes Mora 1 2 , Marcel E Dinger 3 , Chelsea Mayoh 1 2 , Benjamin S Rayner 1 2 , Maria Tsoli 1 2 , David Ziegler 1 2 5
  1. Children's Cancer Institute, Kensington, NSW, Australia
  2. School of Clinical Medicine, University of New South Wales, Sydney, NSW, Australia
  3. School of Life and Environmental Sciences, University of Sydney, Sydney, NSW, Australia
  4. Garvan Institute, Sydney, NSW, Australia
  5. Kid's Cancer Centre, Sydney Children's Hospital, Randwick, NSW, Australia

Aberrant epigenetic regulation is a hallmark of Diffuse Midline Glioma (DMG), an incurable paediatric brain tumour. The H3K27M driver histone mutation leads to transcriptional dysregulation, indicating that targeting the epigenome and transcription may be key therapeutic strategies against this highly aggressive cancer. One such target is the Facilitates Chromatin Transcription (FACT) histone chaperone. Using Cleavage Under Targets and Release Using Nuclease (CUT&RUN) we found FACT to be enriched at developmental gene promoters, coinciding with regions of open chromatin and binding motifs of core DMG regulatory transcription factors. Furthermore, FACT interacted and co-localised with the Bromodomain and Extra-Terminal Domain (BET) protein BRD4 at proximal promoters, suggesting functional cooperation between FACT and BRD4 in DMG. In vitro, a combinatorial therapeutic approach using the FACT inhibitor CBL0137, coupled with BET inhibition revealed potent and synergistic cytotoxicity across a range of DMG cultures, with H3K27M-mutant cells demonstrating heightened sensitivity. These results were recapitulated in vivo, significantly extending survival in three independent orthotopic PDX models of DMG. Using ATAC-seq and nascent RNA-seq, we observed that treatment with the FACT inhibitor CBL0137 led to reduced chromatin accessibility at transcription start sites and when combined with the BET inhibitor JQ1, this effect was markedly enhanced, resulting in a widespread transcriptional shutdown. This combination repressed general transcription including key oncogenic drivers such as MYC, PDGFRA, and MDM4. These findings were further validated at the protein level, confirming the downregulation of these oncogenes. Additionally, we noted aberrant splicing and upregulation of the antigen presentation machinery, suggesting enhanced immune visibility in response to treatment. These findings are being further investigated using single cell RNA-seq and spatial transcriptomics in an immunocompetent mouse model of DMG, to explore the therapeutic potential of heightened immune visibility alongside tumour cytotoxicity. Together, our data highlight the promise of targeting both FACT and BRD4 in DMG, offering a dual strategy to suppress DMG growth while potentially augmenting anti-tumour immunity.