Project description:The prevailing model of therapeutic resistance in cancer posits that malignant cells acquire genetic mutations in the context of therapeutic pressure that negate the efficacy of the drug. This mutation centric view of therapeutic resistance has recently been challenged by emerging evidence that shows a paucity of genetic diversity in malignant cells that evade therapeutic challenge emphasizing the role of epigenetic evolution in drug resistance. We have previously established that therapeutic resistance emerges in the absence of genetic adaptation from leukemia stem cells and whilst the clinical relevance of these findings has been established, what remains unanswered is whether transcriptional programs that confer resistance to therapies are pre-existing or acquired. It is also unclear how stable this adaptive transcriptional response is and whether it can be reversed. Here, we utilize a unique leukaemia model to highlight the key principles of epigenetic resistance. Using single cell RNA-seq with indexed flow sorting, we demonstrate that a form of Lamarckian evolution, namely transcriptional plasticity, drives epigenetic resistance. With a CRISPR-Cas9 screen we identify reciprocal regulators of the enhancer modification H3K4me1/2, LSD1 and MLL4, as important modulators of the resistant cell state. Knockdown or catalytic inhibition of LSD1 causes reversion to a drug sensitive state that is dependent on the lineage determining pioneer transcription factor Pu.1. Mechanistically, we establish that inhibition of LSD1 results in newly formed enhancers marked by H3K4me1/2, H3K27ac and Pu.1 binding, which facilitates the recruitment of transcriptional co-activators and activation of a differentiation gene expression program. We show that this enhancer-remodeling does not result in a simple linear reversion to the original chromatin state of drug naive cells, rather it reinstates the cells dependence on transcriptional co-activators including Med1 and Brd4 due to the formation of new enhancers with enhancer-promoter loops to canonical Brd4 target genes. Together these findings provide new insights into the molecular mechanisms that facilitate epigenetic resistance to cancer therapies and highlight new opportunities for therapeutic intervention to overcome transcriptional plasticity.

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Targeting enhancer switching overcomes non-genetic drug resistance in acute myeloid leukaemia

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