Novel C/EBPalpha replacement therapy by microRNA mimics in acute myeloid leukemia

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Acute myeloid leukemia (AML) is a malignant disease of the hematopoietic system, and the outcome of patients suffering from AML is still very poor. The transcription factor C/EBP Alpha plays an important role in the normal differentiation of myeloid progenitor cells and subsequent cell death. An inactivation of C/EBP Alpha via different mechanisms is found in over 50% of AML cases, e.g. by inactivating mutations of the CEBPA-gene in 10% of cases. This leads to a block in differentiation and an accumulation of immature blast cells. Thus, it would be advantageous to re-activate C/EBP Alpha function in AML therapeutically, thus forcing the AML blasts to differentiate normally and die. MicroRNAs, a class of small non-coding RNAs, were identified as important regulators of normal hematopoiesis and AML development. We have previously shown that C/EBPα induces microRNAs miR-223, miR-34a, and miR-30c which are downregulated in C/EBP Alpha mutated AML. Within this proposed project we will follow the innovative idea to restore wild-type C/EBP Alpha function by replacing downstream microRNAs and thus mimicking “C/EBP Alpha-ness”, instead of "Fixing" C/EBP Alpha itself. We plan to identify further C/EBP Alpha-induced microRNAs using a global next-generation sequencing screen in CEBPA knock-out (KO) mice, and, additionally, AML patient samples with C/EBP Alpha-mutations. We will combine the previously discovered microRNAs miR-223, miR-34a and miR-30c with further microRNAs downregulated in CEBPA KO mice from our preliminary data into a cocktail we have called "Alpha MicroRNA Group 2016" (AMG-16). Utilizing PEI nanoparticles as a highly efficient and non-toxic delivery system we will test our cocktail in various in vitro AML cell lines and AML patient sample models. In addition, in vivo mouse models, such as C/EBP alpha KO mice and NSG mice with AML patient derived xenografts (PDX) cells will further bridge the gap between research and clinic. Subsequently, the microRNAs will be functionally investigated and their biological role analyzed. The specific aims are: 1. Identification and charaterization of C/EBPα-induced microRNAs in vitro and in vivo; 2. Development of a C/EBP alpha replacement therapy by microRNA mimics in AML model systems in vitro; 3. Development of a C/EBP alpha replacement therapy by microRNA mimics in AML model systems in vivo. We hypothesize that multiple mimics will show an additive therapeutic effect in comparison to single mimic therapy. Taken together, we will establish a novel C/EBP alpha replacement therapy by microRNA mimics in AML. Thereby, we attempt to establish the basis for a new therapeutic strategy to clinically enhance AML treatment success.