3030 – THE EPIGENETIC REGULATOR LANDSCAPE OF STEMNESS NETWORKS IN ACUTE MYELOID LEUKEMIA

Several leukemia-associated oncoproteins activate transcriptional circuits resembling a stem-like state in acute myeloid leukemia (AML). This activation of “stemness” genes such as the clustered homeobox (HOX) genes, TALE domain proteins MEIS1, PBX1/3, and the Polycomb group gene BMI1 is achieved by enlisting the activity of specialized components of the epigenetic machinery. Leveraging a MEIS1 CRISPR knock-in fluorescent reporter and a pooled domain-focused high-density CRISPR screen, we comprehensively mapped epigenetic regulators critical for perpetuating these stemness networks in AML. Through this approach, we identified and independently validated members of eight distinct chromatin-modifying complexes as candidate regulators of the HOX/MEIS gene expression program. Single-cell and bulk RNA-seq studies revealed that some of the novel HOX regulators that we identified were required for the expression of a large proportion of the leukemia-associated transcriptome. Of particular interest to us was the TUDOR domain chromatin reader CCDC101. CRISPR-ko of CCDC101 significantly reduced the proliferation of cells with distinct HOX-activating mutations. ChIP-seq studies showed that CCDC101 occupied stem-cell associated gene loci in a TUDOR-domain dependent manner and CCDC101 deletion selectively attenuated their transcription. Further, CCDC101 knockout impaired blast colony formation and induced differentiation in the MLL-AF9 and CALM-AF10 mouse AML models but did not affect normal hematopoietic colony formation. CCDC101 deletion delayed disease latency in two distinct human AML cell line models in vivo and resulted in striking antiproliferative effects in AML patient cells. Our study revealed novel attractive nodes for therapeutic targeting of leukemia stem cells in AML and provides a framework to identify vulnerabilities against recalcitrant oncogenic networks.