Project Summary Acute Myelogenous Leukemia (AML) is an aggressive, genetically heterogeneous cancer—affecting both children and adults—that arises from abnormal myeloid cells in the bone marrow (BM). While many patients achieve remission following induction chemotherapy, the 5-year survival remains a dismal 25% due to relapse with increased resistance to cancer drugs. AML remodels the BM microenvironment, in part by releasing membrane-bound extracellular vesicles (EVs) that dysregulate recipient cells. We have found that EVs transfer endoplasmic reticulum-stress to stromal cells, leading to cell type-specific phenotypic changes that alter the cellular composition of the BM. Interestingly, these EVs were also found to contain bone morphorgenic proteins (BMP)—potent growth factors implicated in drug resistance and cancer progression. Currently little is known about how EV-mediated transmission of ER-stress and BMPs contribute to forming a chemo-protective niche. Multiple studies have shown that ER-stress can be transmitted between cells in the tumor microenvironment, contributing to drug resistance in solid tumor models. In the context of AML, we have found that EVs alone are sufficient to transmit ER-stress and activate the Unfolded Protein Response (UPR) pathway in recipient stromal cells. In vivo, AML cells exhibit marked UPR due to physiologic stress in the tumor microenvironment. We have found that the upregulation of the UPR pathway coincides with the increased expression and packaging of multiple BMP types onto EVs. Since both UPR and BMPs have been previously implicated in promoting cancer survival and drug resistance, we propose to study the role of EVs in transferring adaptive change to recipient BM cells to form a chemo-protective environment. Additionally, due to the mounting evidence that AML-EVs remodel the BM microenvironment, blocking EV biogenesis has become an obvious therapeutic target for AML. We hypothesize that EV-mediated transmission of Unfolded Protein Response and bone morphogenic proteins promotes adaptive changes contributing to drug resistance in AML, which can be ameliorated by inhibiting the release of EVs. To test this hypothesis, In AIM 1, we will identify the contribution of EV-mediated transmission of ER-stress in promoting chemo-protective changes in both recipient stromal and AML cells, and determine if inhibiting exosome release can prevent these effects. In Aim 2 we will determine how EV-associated BMPs dysregulate stromal cells, and examine the effect of blocking EV-release and BMP-receptor signaling on AML progression. Our long-term goal is to answer long standing questions about how AML cells evade chemotherapy and develop new therapeutic strategies to reduce drug resistance in patients with AML.