Project Summary: The past several decades have seen little progress in the development of new treatment options for AML patients. This is due in large part to our limited understanding of how many recurrent genetic alterations truly impact disease progression. Thus, there is a great need to identify novel genetic alterations that drive AML, establish a mechanistic understanding of how these changes influence disease progression, and develop therapeutic approaches for patients with these alterations. To this end, we recently identified that expression of the DNA and RNA binding protein hnRNP K is reduced in AML patients harboring a 9q21.32 deletion and this haploinsufficiency directly inhibits its transcriptional activation of p21 and C/EBPα (In press, Cancer Cell). In contrast to reduced expression, we also discovered that HNRNPK is amplified and overexpressed in over 30% of AML patients without this deletion, resulting in poor clinical responses and outcomes. Furthermore, multivariate analyses revealed that hnRNP K overexpression drives c-Myc expression and also cooperates with the most common alteration in AML (mutant NPM1) to exacerbate leukemogenesis. Given our current lack of understanding as to how hnRNP K overexpression impacts AML progression, this application will interrogate hnRNP K oncogenic functions and determine its cooperative relationship with mutant NPM1 in AML. Mechanistically, hnRNP K is also thought to activate expression of oncogenes like c-Myc, through its transcriptionally and translationally functions. Thus, exploring the role of hnRNP K in mediating c-Myc expression in AML is critical for understanding as to how hnRNP K overexpression drives disease progression, as this may represent an uncharacterized mechanism to drive c-Myc expression in the absence of MYC amplifications or translocations in AML. It is our hypothesis that hnRNP K is an uncharacterized oncogene that stimulates c-Myc expression and cooperates with mutant NPM1 to drive leukemogenesis. The objectives of this proposal are to understand the hnRNP K-mediated mechanisms that drive leukemogenesis, determine the synergism between hnRNP K and mutant NPM1, and identify therapeutic approaches to exploit these alterations. Using genetically defined primary patient samples and novel transgenic animal models, we will evaluate 1). how hnRNP K overexpression impacts leukemogenesis and myeloid differentiation, 2). how hnRNP K directly regulates the c- Myc pathway and therapeutic responses, 3). how hnRNP K overexpression synergizes with mutant NPM1 in driving leukemic progression 4). the mechanisms by which hnRNP K directly regulates unique transcriptional and translational programs governing leukemogenesis. These studies will fundamentally advance our understanding of hnRNP K oncogenic functions and determine whether hnRNP K can be used to risk stratify patients and serve as a marker for targeted therapy.