Acute myeloid leukemia (AML) is a disease characterized by overproduction of abnormally differentiated, hyper-proliferative myeloid cells known as blasts in bone-marrow and blood. Our laboratory has previously demonstrated that loss of epigenetic repression by the polycomb repressive complex 2 (PRC2), which is mediated by complex member metal response element binding transcription factor 2 (MTF2), drives chemo-resistance resulting in refractory AML. In this study, to identify MTF2-specific synthetic lethal interactions, a genome-scale CRISPR Knock-out (GeCKO) synthetic lethal screen was performed in matched MTF2-deficient and rescued THP-1 cells both in the absence and presence of the induction chemotherapeutic cytarabine. Following careful analysis of screening data using specialized software, 104 highly significant MTF2-specific synthetic lethal interactions as well as 15 cytarabine-specific synthetic lethal interactions were identified. Reduced stringency upon analysis helped to identify an additional seven MTF2-specific synthetic lethal interactions that could be targeted with commercially available small-molecule inhibitors. Among eight small molecule inhibitors, two DNA Polymerase A/Ribonucleotide Reductase Catalytic Subunit M1 (POLA/RRM1) dual inhibitors (clofarabine and fludarabine) were shown to induce toxicity with specificity for MTF2-deficient THP-1 cells at low concentrations only in the absence of cytarabine.
In the future, further testing of the therapeutic potential of clofarabine and fludarabine in treating MTF2-deficient AML will be conducted in patient derived bone-marrow aspirates which better represent the true clonal and hierarchical nature of this life-threatening malignancy. Furthermore, lentiviral delivery of short-hairpin RNAs (shRNAs) targeting highly significant, non-enzymatic MTF2 and cytarabine-specific synthetic lethal interactions will be performed in both THP-1 cells as well as in patient derived bone-marrow aspirates. Eventually, in vitro validated targets will be validated under in vivo conditions using a patient derived xenograft (PDX) preclinical animal model of AML using immunocompromised NOD scid gamma (NSG) mice.
| Identifer | oai:union.ndltd.org:uottawa.ca/oai:ruor.uottawa.ca:10393/39886 |
| Date | 28 November 2019 |
| Creators | Cafariello, Christopher |
| Contributors | Stanford, William |
| Publisher | Université d'Ottawa / University of Ottawa |
| Source Sets | Université d’Ottawa |
| Language | English |
| Detected Language | English |
| Type | Thesis |
| Format | application/pdf |
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