Combined inhibition of FACT and BET disrupts transcription to suppress tumor growth in mouse models of diffuse midline glioma

Aberrant epigenetic regulation is a hallmark of diffuse midline glioma (DMG), an incurable pediatric brain tumor. 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. We found FACT to be enriched at developmental gene promoters, coinciding with open chromatin and binding motifs of core DMG regulatory transcription factors. Furthermore, FACT co-occurred with the bromodomain and extraterminal domain (BET) protein BRD4 at promoters and enhancers, 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. These results were recapitulated in vivo, extending survival in three independent orthotopic patient-derived xenograft models of DMG. Mechanistically, we show that CBL0137 treatment decreased chromatin accessibility and combined with BET inhibition to cause broad transcriptional collapse; silencing of several key oncogenes including <i>MYC</i>, <i>PDGFRA</i>, <i>MDM4</i>, and <i>SOX2</i>; and alterations to the splicing landscape. This combination also elicited immune-related effects, including activation of the interferon response and antigen presentation mechanisms in DMG cells and induction of an activated state in macrophages and T cells, as demonstrated in an immunocompetent setting with spatial transcriptomics. Together, our data highlight the therapeutic promise of simultaneously targeting FACT and BET proteins in DMG, offering a dual tumor-intrinsic and immune-mediated strategy for combating this devastating pediatric brain tumor.