Abstract Paediatric high-grade gliomas (HGGs) including diffuse midline gliomas (DMGs) account for the highest mortality in CNS tumours with 90% of patients surviving less than two years. The identification of hallmark H3K27/G34 mutations and oncogenic targets such as PDGFRA and PI3K has not yet led to improved survival. This therapeutic stagnation may be impacted by the overexpression of BCL-2 family anti-apoptotic proteins, which confer resistance to cell death. While the anti-apoptotic protein MCL-1 is a known driver of resistance in multiple cancers 1,2, its specific role in HGG/DMG remains unexplored. This study evaluates MCL-1 inhibitors in HGG/DMG and identifies potent combinatorial strategies to overcome resistance. Baseline protein and gene expression analyses, using Western blot and RT-qPCR, confirmed MCL-1 upregulation in a panel of DMG/HGG cultures. Selective MCL-1 inhibitors (S63845, MIK665, BRD-810) and the BCL-2 inhibitor venetoclax were screened in cytotoxicity assays, with MIK665 showing the greatest efficacy against both HGG and DMG cells. Crucially, these inhibitors exhibited no toxicity against healthy astrocytes and rat cardiomyoblasts at clinically relevant doses. In vivo, MCL1 inhibition (S63845) significantly extended the survival of two genetically distinct DMG models and one HGG orthotopic model. Notably, pharmacokinetic analysis detected S63845 levels in brain tissue, indicating that the drug crossed the blood-brain barrier, while histopathology showed reduced Ki67-positive proliferative tumour cells. Mechanistically, MIK665 treatment of DMG cells induced apoptosis, evidenced by cleaved-PARP accumulation and Annexin V positivity, while restoring H3K27 histone trimethylation, suggesting a potential novel role in epigenetic reprogramming. Given these findings, we evaluated MIK665 in combination with the FACT inhibitor, CBL-037 (currently in clinical evaluation, NCT04870944)3,4. This combination showed strong synergistic effects, particularly in DMG cells without any toxicity in healthy control cells. In conclusion, we demonstrate for the first time that MCL-1 inhibition achieves potent therapeutic efficacy against DMG/HGG. The discovery that MCL-1 inhibition restores H3K27me3 reveals a novel mechanistic link between apoptotic evasion and epigenetic dysregulation. The robust synergy with epigenetic inhibitor CBL-0137 provides a compelling rationale for further preclinical development in DMG and HGG tumours. 1. Adjumain, S. et al. Multidimensional, integrative profiling identifies BCL2L1 methylation as a predictor of MCL1 dependency in pediatric malignancies. JCI Insight 10, e184601 (2025). 2. Beroukhim, R. et al. The landscape of somatic copy-number alteration across human cancers. Nature 463, 899 (2010). 3. Ehteda, A. et al. Dual targeting of the epigenome via FACT complex and histone deacetylase is a potent treatment strategy for DIPG. Cell Rep 35, 108994 (2021). 4. Holliday, Holly, et al. “Combined inhibition of FACT and BET disrupts transcription to suppress tumor growth in mouse models of diffuse midline glioma.” Science Translational Medicine 18.832 (2026): eadr1557.
Lawrence et al. (Tue,) studied this question.