Epco-16. disrupting dot1l epigenetic activity reprograms glioblastoma stem cells towards a dopaminergic neuronal-like state

Abstract Glioblastoma is the most common and aggressive primary adult brain tumor, remaining incurable despite current therapeutic strategies. Through a genome-wide essentiality screen, we previously identified disruptor of telomeric silencing-1-like (DOT1L) as one of the most important epigenetic regulators in glioblastoma stem cells (GSCs). Chemogenic inhibition of DOT1L epigenetic activity was found to increase the expression of neuronal markers while decreasing the expression of cancer stem cell markers at the transcriptional and protein level, suggesting GSC differentiation towards a neuronal lineage. Thus, we sought to investigate the mechanism of DOT1L in GSCs and whether disruption of DOT1L epigenetic activity promotes functional integration of GSCs into neural circuits. Here, we profiled the overall transcriptional and chromatin accessibility landscape of GSCs following DOT1L inhibition. Interestingly, we identified that DOT1L inhibition results in increased chromatin accessibility at key gene loci involved in neural differentiation, synaptic transmission, in addition to dopamine synthesis, packaging, and release, which correlated with upregulated gene expression. Furthermore, these gene loci exhibited alterations of the DOT1L epigenetic mark, H3K79me2. Transcriptional motif analysis of GSCs following DOT1L inhibition revealed dysregulated accessibility of transcription factor motifs known to be involved in regulating cellular response to neuronal signals. Cell viability assessment of GSCs treated with a DOT1L inhibitor and a panel of neurotransmitters further confirmed that DOT1L inhibition sensitizes GSCs to neuronal signals. These data suggest that disrupting the epigenetic activity of DOT1L reprograms the epigenetic landscape of GSCs to promote a functional, dopaminergic, neuronal-like state that exhibits enhanced responsiveness to neuronal signals. Future experiments aim to investigate the impact of disrupted DOT1L epigenetic activity on GSC-neuronal bidirectional communication and the mechanisms by which this influences the progression and aggressiveness of glioblastoma.