Identification of MYC-Driven Vulnerabilities in Acute Myeloid Leukemia
Authors: Nathan Monica , Mahmoud Ahmed , Arthur Nancy BJ , Gruszczynska Agata , Sykes Stephen , Ferraro Francesca 
Published: December 2024

We1 and others2 have shown that MYC is a critical oncogene implicated in the pathogenesis of acute myeloid leukemia (AML). Gain-of-function mutations in Mac box I generate transplantable myeloid leukemia in mice with phenotypic characteristics similar to the overexpression of wild-type Myc. In this study, we aimed to identify a set of biologically distinct, non-redundant, direct MYC transcriptional targets critical for AML maintenance, referred to as the MYC-Leukemogenic Signature (MYC-LS).

To identify direct transcriptional targets essential for leukemogenesis, we performed an integrated analysis of the chromatin binding and gene expression profiles of pre-leukemic stem cells expressing oncogenic levels of wild-type or mutated MYC protein. We compared these profiles to those of normal murine hematopoietic progenitor cells. We validated these targets in fully transformed murine myeloid leukemia cells using both bulk and single-cell RNA sequencing. We then analyzed MYC-LS gene expression in human AML samples from the TCGA dataset and evaluated their functional relevance using DepMap Public 22Q4 Chronos Scores. Finally, we performed knockdown studies of selected targets in independent murine cell lines.

We identified 858 differentially expressed direct MYC targets (ANOVA, log FC &gt; ±2, FDR &lt; 0.05), with 472 upregulated and 386 downregulated in pre-leukemic hematopoietic progenitor cells compared to normal hematopoietic progenitor cells. GO-Elite analysis identified distinct clusters of dysregulated genes. Cluster 1 genes, upregulated in pre-leukemia cells, were enriched for nucleolar genes, pseudouridinylation genes, and nucleocytoplasmic transport genes. Cluster 2 genes, downregulated in pre-leukemia cells, were enriched for cell communication and immune genes. Of these genes, 135 were direct MYC transcriptional targets, comprising the MYC-Leukemogenic Signature (MYC-LS), including 133 protein-coding genes, one long non-coding RNA, and one microRNA. These targets showed significant changes in expression in MYC-overexpressing HSPCs (116 upregulated, 19 downregulated). Subcellular localization analysis revealed predominant nuclear localization for 77 out of 135 MYC-LS genes, including components of the H/ACA snoRNP complex and pseudouridine synthases pathways.

We examined the MYC-LS in human AML samples from the TCGA dataset and analyzed their functional relevance using DepMap Public 22Q4 Chronos Scores. In human AML samples, 107 out of 135 MYC-LS genes significantly correlated with MYC mRNA levels (Pearson correlation coefficients 0.24-0.73, Bonferroni p-value &lt; 0.05). Hierarchical clustering indicated significant dysregulation of 131 out of 135 MYC-LS genes in AML cells compared to normal human myeloid counterparts (FDR &lt; 0.05). DepMap analysis showed that 85 MYC-LS genes were essential for at least one myeloid cell line (median essentiality score -0.5, p &lt; 0.01), with 5 of the top 10 dependencies being genes in the nucleotide synthesis pathway. CTP synthase 1 (CTPS1), a gene involved in cellular response to cytarabine, a chemotherapy drug with the highest efficacy against AML cells, was identified as the top AML-specific dependency, validating the relevance of the MYC-LS genes as potential therapeutic targets. shRNA-mediated inhibition of selected MYC-LS targets significantly inhibited the growth of murine AML1 cells (murine leukemia driven by Dnmt3a and Flt3-ITD) and Mll-Af9 leukemias.

Our study describes the MYC-Leukemogenic Signature, a set of direct MYC target genes dysregulated in both murine and human AML cells. These genes are primarily involved in nucleotide synthesis, RNA metabolism, and inflammatory responses, and they are critical for leukemia cell survival and correlate with MYC expression. Importantly, our findings suggest that MYC mutations activate potentially targetable pathways in AML.

1 -Arthur NB, et al. Clin Cancer Res. 2024 Jun 7. doi: 10.1158/1078-0432.CCR-24-0926.

2- Freie B, et al. doi: 10.1101/gad.351292.123