CRISPR Screens Identify Candidate Therapeutic Targets in Leukemia

Abstract: Acute myeloid leukemia (AML) is a complex hematological malignancy marked by proliferation of immature myeloid cells with a dismal 5-year survival. A major challenge is the persistence of leukemia stem cells (LSCs) after standard treatment, leading to relapse. This thesis employs in vivo CRISPR/Cas9 screening to investigate critical AML and LSC molecular mechanisms, interrogating the dependancies on cell surface receptors for use as potential therapies. In our initial study (Article I), we identify the chemokine receptor CXCR4 as a crucial dependency of AML cell growth and survival. Using a murine model of AML driven by MLL::AF9, we find that CXCR4 loss triggers oxidative stress and differentiation in vivo, with CXCL12 ligand signaling being non-essential for leukemia development.Expanding our study to interrogate nearly one thousand cell surface receptors, we identify three additional AML dependencies. Among these, GLUT1, a primary cellular glucose transporter, emerges as a key regulator of energy metabolism, driving MLL::AF9 LSC survival (Article II). Inhibition of GLUT1 suppresses cellular bioenergetics, prompting autophagy as a metabolic adaptation. Notably, dual inhibition of GLUT1 and oxidative phosphorylation effectively eliminates human AML cells, especially for the RUNX1-mutated AML subtype.Furthermore, our research also reveals iron metabolism as another critical AML dependency (Article III). We found that disrupting iron uptake through genetic knockdown of Tfrc, encoding the transferrin receptor (TFR1), suppresses leukemia development in a p53-dependent manner, leading to transcriptional repression of antioxidant defense and mitochondrial respiration pathways. Patient-derived AML cells were selectively targeted upon iron chelation treatment. Additionally, our work uncovers the role of H2K1 in evading NK cell-mediated immune surveillance in vivo through disruption of NK cell maturation and activation (Article IV). Consistent with this finding, H2k1 disruption alone suffices to reverse this immune evasion, restoring NK cell-mediated anti-leukemic effects.In conclusion, this thesis highlights the value of CRISPR/Cas9 screens in identifying physiologically relevant AML dependencies. It offers insights into targetable LSC vulnerabilities, emphasizing the potential of metabolic targeting and combined treatments for improved AML therapies.