Leukemia is the most common form of childhood cancer. Children with acute myeloid leukemia (AML) have less than 50% overall survival despite aggressive chemotherapy and bone marrow transplantation. Therefore, it is critical to understand the molecular pathogenesis of AML. We demonstrated that CREB is overexpressed in bone marrow cells from patients with AML but not in normal bone marrow or bone marrow from patients without active leukemia. Furthermore, CREB overexpression was associated with an increased risk of relapse and decreased event-free survival in patients with AML. Our preliminary results suggest that AML is a heterogeneous disease that is not well understood. We hypothesize that there is an uncoupling of differentiation and CREB expression in myeloid leukemia cells. We propose to study the role of CREB in normal and malignant myeloid cells to identify novel mechanisms of leukemogenesis and improve our understanding of the molecular pathways regulating myeloid cell proliferation and differentiation. In Specific Aim 1, we will characterize CREB expression and activation in primary normal myeloid cells and myeloid leukemia cells. Experiments are proposed to determine the expression of CREB in normal mouse embryos at different stages of hematopoietic development. We will also examine CREB expression in normal myeloid progenitor cells at different stages of myeloid differentiation. Finally, we will examine whether CREB is activated in primary leukemia cells. In Specific Aim 2, we will further characterize the biological phenotype of CREB overexpression and down regulation in myeloid leukemia cell lines and primary normal myeloid cells. Our preliminary results demonstrated that CREB overexpression leads to increased proliferation and survival of myeloid leukemia cells. CREB down regulation using RNA interference (RNAi) suppresses the growth and survival of leukemia cells. To study signaling pathways upstream of CREB, we will overexpress activated kinases and use RNAi technology to inhibit expression of kinases. In Specific Aim 3, we will characterize the phenotype of transgenic mice in which CREB overexpression is targeted to myeloid cells. Defects in hematopoiesis and development of leukemia will be determined in both CREB transgenic mice and a mouse bone marrow transplant model. These studies will define the role of CREB in both normal and malignant myelopoiesis.