MODELING and Targeting ERG-INDUCED Acute MYELOID LEUKEMIA (AML).
Authors: Smooha Gil , Birger Yehudit , Goldberg Liat , Jacob-Hirsch Jasmine , Amariglio Ninette , Rechavi Gideon , Levanon Ditsa , Groner Yoram , Taragan Anat , Kloog Yoel , Wen Jeremy Q , Crispino John , Radmacher Michael D. , Marcucci Guido , Bloomfield Clara D. , Izraeli Shai 
Published: October 2019

Abstract
Abstract 475
ERG is an oncogene located on the long arm of human chromosome 21 that encodes an ETS transcription factor that has been reported to be involved in normal and aberrant megakaryopoiesis (Salek-Ardakani et al Cancer Res. 2009;69:4665; Stankiewicz et al Blood 2009;113:3337). High ERG expression has also been reported associated with poor prognosis of cytogenetically normal AML (Marcucci et al J Clin Oncol. 2007;25:3337). Thus, deciphering the molecular targets and oncogenic pathways activated by overexpressed ERG protein is likely to lead to more effective therapy for ERG-related myeloid leukemias. Towards these goals we have combined in-vitro and in-vivo approaches. We have created transgenic mice expressing the ERG3 hematopoietic isoform under the VAV promoter. All these mice die from invasive acute megakaryocytic or undifferentiated myeloid leukemia by the age of 5 months. The leukemias are transplantable, and primary growth factor dependent leukemic cell lines, suitable for pharmacological and molecular studies, have been established. To identify ERG target genes and proteins, we have analyzed gene expression in two “mirror-image” cellular systems – shRNA mediated knockdown of ERG in Meg01 megakaryocytic leukemia cells and overexpression of ERG in K562 erythroleukemia cells. Gene Set Enrichment Analysis (GSEA) demonstrated that the “ERG gene-associated expression signature” in these cell lines is enriched with genes that were also identified in primary human AML characterized by ERG overexpression. By chromatin immunoprecipitation, we then identified specific, direct ERG targets and confirmed their expression in samples from ERG transgenic leukemias and primary human “ERG-overexpressing” AMLs. Surprisingly, we observed that the lymphoid kinase Bruton agammaglobulinemia tyrosine kinase (BTK) is an ERG direct target that is upregulated in both ERG overexpressing mouse and human leukemias. Preliminary experiments have demonstrated activity of a BTK inhibitor on ERG transgenic and ERG overexpressing human AML cells. ERG overexpression also induced marked activation of RAS signaling as supported by elevated levels of RAS-GTP and its downstream targets and significant growth inhibitory activity caused by a novel RAS inhibitor, farnesylthiosalicylic acid (Salirasib), that disrupts the spatiotemporal localization of active Ras (Rotblat et al Methods Enzymol. 2008;439:467) was observed. Dramatic growth inhibitory activity has also been induced by two novel compounds that induce megakaryocytic polyploidization, suggesting a potential role for “differentiation” therapy of ERG-related leukemias. Thus we have created a mouse model for ERG-related AML and characterized several genes and biochemical pathways that are susceptible to pharmacological inhibition. Our studies are not only likely to decipher the mechanisms by which ERG overexpression contributes to leukemogenesis, but also provide a platform for preclinical studies of novel therapeutics of these poor prognosis leukemias.

Disclosures:
No relevant conflicts of interest to declare.