Identification of epigenetic modifiers essential for growth and survival of AML1/ETO-positive leukemia
Aberrant gene expression patterns in acute myeloid leukemia (AML) with balanced chromosomal translocations are often associated with direct or indirect dysregulation of epigenetic modifiers. Previously, we have shown that the AML1/ETO (RUNX1/MTG8) fusion protein, encoded by the translocation (8;21)(q22;q22), leads to the epigenetic repression of its target genes. We aimed in this work to identify critical epigenetic modifiers, on which AML1/ETO-positive AML cells depend on for proliferation and survival using shRNA library screens and global transcriptomics approaches. Using shRNA library screens, we identified 41 commonly depleted genes in two AML1/ETO-positive cell lines Kasumi-1 and SKNO-1. Among them, several epigenetic regulators as DMAP1, SMYD1, SMYD2, BRD4, SETD8, SETD1A and HDAC8 were depleted. We validated genetically and pharmacologically DNMT1 and ATR using several AML1/ETO-positive and negative cell lines. We also showed in vivo differentiation of myeloblasts after treatment with the DNMT1-inhibitor decitabine in a patient with an AML1/ETO-positive AML. Bioinformatic analysis of global transcriptomics after AML1/ETO induction in 9/14/18-U937 cells identified 973 differentially expressed genes (DEGs) (648 up- and 325 downregulated). Pathway analysis revealed several interferon response pathways and metabolic pathways such as oxidative phosphorylation and hypoxia enriched after AML1/ETO induction. Three genes (PARP2, PRKCD and SMARCA4) were both downregulated after AML1/ETO induction, and also detected in the survival shRNA screens of both cell lines, decreasing survival and proliferation. In conclusion, using unbiased shRNA library screens and global transcriptomics, we have identified several driver epigenetic regulators for proliferation in AML1/ETO-positive AML. Some of these epigenetic regulators, such as DNMT1 and ATR, are susceptible to pharmacological inhibition by small molecules showing promising preclinical and clinical efficacy Overall design: We compare differential regulated genes after AML1/ETO indution in our generated 9/14/18-U937 cells after Ponasterone A compared to 9/14/18-U937 treated with vehicle (Ethanol), to 9/10/7-U937 LacZ clone treated with Ponasterone A and Ethanol as controls