3186 – TRANSCRIPTOMIC ANALYSIS OF SYNTHETIC HUMAN CORD BLOOD LEUKEMIAS REVEALS DEVELOPMENT STAGE-SPECIFIC DEPENDENCE ON N-MYC

Abstract

Descriptive sequencing of human leukemia samples has revealed the diversity and combinatorial complexity of oncogenic effector genes which can occur within all or just a subset of tumor cells. With the aim of deconvoluting the individual contributions of T-cell acute lymphoblastic leukemia (T-ALL) oncogenes, we employed our recently developed synthetic model of human T-ALL (Kusakabe et al, 2019) to perform a systematic analysis of transcriptional signatures associated with each of the four oncogenes used in our model. We identified synthetic leukemias to cluster based on two gene signatures which resembled distinct stages in T-cell development and had marked variation in MYCN expression, with its highest expression found on the more "immature" cluster. All synthetic leukemias carried activated NOTCH1, which is best known for upregulating MYC, but not MYCN. By analyzing publicly available RNA sequencing datasets, we found MYCN to be highly expressed in early T-cell progenitors (ETPs) and T-ALLs of more immature phenotype. To assess oncogene "addiction" to MYCN, cell lines and primary synthetic leukemias were transduced with shRNAs or sgRNAs to knock-down (KD) or knock-out (KO) MYCN, respectively. By both approaches, bulk cell growth and/or clonogenic activity were reduced in KD/KO cells as compared to controls, supporting their functional dependence on MYCN. MYCN overexpression resulted in increased clonogenic activity and differentiation delay of NOTCH1-transduced cells in vitro but was still insufficient to generate leukemia in vivo. We are currently exploring other genes with similar expression patterns to MYCN to identify upstream/downstream mediators of MYCN dependence in this context. Our results emphasize stage-specific oncogene dependencies and identify alternate therapeutic targets in T-ALLs with high MYCN expression.