Dual Targeting of FLT3 and LSD1 Disrupts the MYC Super Enhancer Complex in Acute Myeloid Leukemia.

Clinical responses to kinase inhibitor therapy in acute myeloid leukemia (AML) are limited by the inevitable development of resistance. A major contributor to resistance is early epigenetic adaptation, leading to persistence of a small number of leukemia cells. Here we show that inhibition of the epigenetic regulator lysine-specific deme-thylase 1 (LSD1) augments the response to inhibitors of the FLT3 kinase in AML. We demonstrate that combined FLT3 and LSD1 inhibition results in synergistic cell death of FLT3-mutant AML cells via proliferative arrest and apoptosis. The drug combination synergistically activates a pro-differentiative epigenetic and transcriptional program while simultaneously suppressing the activity of MYC target genes. High resolution multi-modal epigenetic analyses revealed that combined FLT3 and LSD1 resulted in the suppression of MYC-bound promoters and the activation of PU.1-bound enhanc-ers. Forced expression of MYC partially abrogated the drug effect, and regulon en-richment analysis in primary AML samples nominated STAT5 as a putative regulator of MYC gene expression. STAT5 is highly bound to the MYC blood super-enhancer and inhibition of FLT3 results in a loss of STAT5 binding and a loss of super enhancer acti-vation. Furthermore, knockdown of STAT5 augments LSD1-inhibitor induced cell death. LSD1 inhibition also directly represses MYC target genes which show specific accumulation of the repressive LSD1 substrate H3K9me1. We validated these findings in 67 primary AML samples including 19 FLT3-ITD positive AML samples, with the vast majority demonstrating improved responses with the drug combination. High MYC regulon activity was a predictor of response to the drug combination and RNA-seq on drug treated AML samples revealed suppression of MYC target genes. Finally, single cell ATAC seq on primary AML blasts treated ex-vivo with combined FLT3 and LSD1 inhibition results in a shift from MYC super enhancer-high to a MYC super enhancer-low cell state. Collectively, these studies provide preclinical rationale for the investiga-tion of dual FLT3 and LSD1 inhibition in clinical trial.