The Nemours Foundation

DLK1 As a Novel Target for Down Syndrome Myeloid Leukemia - Project Summary Down Syndrome (DS), With Triplication of Chromosome 21, Is Recognized as One of the Most Important Leukemia-predisposing Syndromes. 1-2% of DS Children Develop Myeloid Leukemia (DS-ML) Before Age 5, Which Is Preceded by a Pre-leukemic Phase Termed Transient Abnormal Myelopoiesis (tam). 30% of Tam Cases Progressing to DS-ML. Aside From Low Dose Chemotherapy, There Are No Treatment Options for Tam and No Preventative Measures to Stall DS-ML Onset. Nearly 10-15% of Children With DS-ML Are Either Refractory to Treatment or Suffer Early Relapse. These Children With Refractory Disease Face a Dismal Outcome With 3-YEAR Event-free Survival Less Than 21%. Moreover, Treatment-related Toxicity and Morbidity Is a Major Cause of Death in DS-ML Patients. Therefore, Novel Therapeutic Options Are Needed for This Rare Disease. Both Tam and DS- ML Are Characterized by the Pathognomonic Mutation in the Gene Encoding Essential Hematopoietic Transcription Factor GATA1, Resulting in N-terminally Truncated Mutant GATA1 Protein (GATA1S). Trisomy 21 and GATA1S Are Sufficient to Induce Tam, While Additional Co-operating Mutations in Genes Such as STAG2 Are Required for DS-ML Leukemogenesis. the Individual and Synergistic Contribution of These Genetic Events Towards DS-ML Refractoriness and Remodeling the Bone Marrow Microenvironment Remain Poorly Defined. Using CRISPR/CAS9 Mediated Gene Targeting for Stepwise Introduction of GATA1 or GATA1 and STAG2 Mutations in Ipscs With Trisomy 21, We Modelled Tam and DS-ML Respectively. We Generated Patient-derived Xenograft (PDX) Models Representing Refractory DS-ML. Our Transcriptome Analysis to Identify Novel DS-ML Specific Cell Surface Proteins Revealed Delta-like Non-canonical Notch Ligand 1 (DLK1) as One of the Top Targets. DLK1 Transcript Was Undetectable in Normal Bone Marrow Specimens, While It Was More Abundant in DS-ML Compared to Tam. Enhanced Expression of DLK1 Was Also Detected in Our Ipsc-generated DS-ML Models, But Not in Tam Models. Our Preliminary Data Shows That DLK1 Knockout in DS-ML Cell Line Suppressed Proliferation and Delayed Engraftment. Furthermore, Targeting DLK1 via an Immuno-conjugate Significantly Reduced Leukemic Burden and Prolonged Survival in Refractory DS-ML PDX Lines. These Results Indicate That DLK1 May Promote DS-ML Leukemogenesis and Can Be Used as a Therapeutic Target. Consistent With the Effect of DLK1 on the Inhibition of Interferon Signaling via Suppression of NOTCH1 Activity, We Identified That Downregulation of DLK1 Resulted in Increased Interferon Signaling. Interestingly, We Demonstrated That GATA1S Suppressed Interferon Signaling via Downregulation of the Rig-i Pathway in Ipsc-derived Tam and DS-ML Models. Exogenous Treatment of DS-ML Cells With Interferon-a Resurrected Interferon Signaling, Confirming That the Pathway Is Intact. Thus, Suppression of Interferon Signaling via GATA1S And/or DLK1 Is a DS-ML Dependency That Can Be Exploited Therapeutically. Upon Study Completion, We Will Identify Novel Treatment Options for Tam/ds-ml, and a Newly Characterized Target That May Also Be Exploited in Non-ds Leukemia.