Thursday, November 21, 2024
11/21/2024
Project Summary: RUNX1 is the DNA-binding subunit of the core binding factor (CBF) complex and a master-regulator transcription factor, which is involved in normal and malignant hematopoiesis. Somatic, heterozygous RUNX1 mutations commonly occur in Myelodysplastic Syndrome (MDS) (10%), as well as in secondary (s) or de novo AML (~10%). Germline mutations in RUNX1 cause the highly penetrant (~40%) autosomal dominant, Familial Platelet Disorder (FPD), which can evolve into myeloid malignancy (FPD-MM). Majority of mutant (mt) RUNX1 behave mostly as loss of function mutations, conferring relative therapy-resistance and poorer survival in patients with AML. Consequently, there is a strong unmet need to develop novel therapies for AML expressing somatic or germline mtRUNX1. Our preliminary studies have demonstrated for the first time that shRNA- mediated knockdown of RUNX1 (mutant and wild-type) or disruption of its binding to CBFβ induces greater lethality in AML progenitor cells (HPCs) expressing mtRUNX1 compared to wild-type (wt) RUNX1. We also found that the +24kb enhancer (eR1) within the intragenic super-enhancer (SE) of RUNX1 regulates its transcription in AML cells. The chromatin reader BET (Bromodomain Extra-terminal) protein (BETP) BRD4 promotes transcription of RUNX1 and its targets. BRD4 degradation or eviction from chromatin, or gene-editing of the +24kb RUNX1 eR1, induces lethality in AML cells. By determining and utilizing the mRNA signature from RUNX1-depleted (by shRNA) AML cells, we queried, through LINCS1000-CMap (Connectivity Mapping) analysis, for expression mimickers (EMs). We identified novel EMs that repress RUNX1 and its targets and induce significantly more apoptosis of AML cells expressing mtRUNX1 versus wtRUNX1. Therefore, the hypothesis motivating our studies is that knocking down of levels of RUNX1 and its targets will induce lethality not only in AML blasts expressing somatic mtRUNX1 but also in FPD/MM HPCs expressing germline mtRUNX1. The specific aims of studies proposed are: AIM 1: To determine impact on active enhancers, transcriptome and pre-clinical in vitro and in vivo efficacy of BETP antagonist along with its co-repression of RUNX1, BCL2 and CDK6, alone or in combination with BCL2 or CDK6 inhibitor, in AML blasts and patient- derived xenograft (PDX) models expressing somatic mutant RUNX1. Additionally, we will evaluate pre-clinical efficacy of co-targeting CRISPR-Cas9 screen-discovered top ‘druggable’ dependencies along with BETP antagonist against AML blasts expressing somatic mtRUNX1. AIM 2: To elucidate pre-clinical in vitro and in vivo efficacy of the EMs homoharringtonine (omacetaxine) or fedratinib alone and in combination with BETP antagonists against patient-derived AML blasts and PDX models expressing somatic mtRUNX1. AIM 3: To determine pre-clinical in vitro and in vivo efficacy of selected EMs that repress RUNX1 and its targets against patient-derived HPCs from FPD-MM expressing germline mtRUNX1 and other somatic co-mutations versus HPCs from RUNX1-FPD expressing only germline mtRUNX1.
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