Hematopoietic as well as leukemic stem cells (HSC/LSC) are dependent on interactions with extracellular matrix, soluble factors and cellular components of the bone marrow (BM) microenvironment for their survival. These interactions are hypothesized to promote the quiescence of the stem cell and their retention in the niche and influence the decisions regarding symmetric versus asymmetric self-renewal divisions. The signals that contribute to these decisions are just now being identified and likely involve the classical molecular pathways shown to be important in stem cells, including WNT/catenin, Notch, KitL/Kit and SDF/CXCR4 signaling. Each of these signals impacts directly or indirectly on the Rho family of small GTPase proteins, including the Rac family. These proteins play an essential role in the retention of HSC in the niche and in the survival, proliferation and differentiation decisions that are critical for sustaining hematopoiesis and maintaining the integrity of the stem cell compartment. We have recently demonstrated that activated Rac is increased in a novel model of human leukemia. In this model, the MLL-AF9 (MA9) leukemia oncogene is used to transform primary human CD34+ cells and promotes a rapid acute myeloid leukemia (AML) in immunodeficient mice. Suppression of Rac-GTP formation using the Rac inhibitor NSC23766, or shRNA knockdown of Rac expression, could effectively promote apoptosis and suppress proliferation under conditions where little or no inhibitory effect could be detected on normal human CD34+ cord blood cells. The inhibition of Rac and the induction of apoptosis correlate with the loss of Bcl protein expression, especially of Bcl-xL, which is an identified downstream target of Rac2 signaling. We propose to use our newly developed leukemia model to study the role of Rac signaling in MLL leukemia cells using in vitro assays measuring well-characterized phenotypes associated with Rac activity, including migration, survival, adhesion and proliferation (Aim1). We will extend these results in vivo using a xenograft system and a pharmacologic as well as RNAi approach to inhibit Rac activity as well as Bcl-xL function (Aim 2). This combinatorial approach should enable us to determine the importance of the Rac signaling pathway in leukemia and begin to dissect the specific mechanisms involved in the Rac addiction in MA9 AML.
We have recently shown that human blood stem cells can be induced to make leukemia upon introduction
|Mizukawa, Benjamin; Wei, Junping; Shrestha, Mahesh et al. (2011) Inhibition of Rac GTPase signaling and downstream prosurvival Bcl-2 proteins as combination targeted therapy in MLL-AF9 leukemia. Blood 118:5235-45|
|Wunderlich, M; Chou, F-S; Link, K A et al. (2010) AML xenograft efficiency is significantly improved in NOD/SCID-IL2RG mice constitutively expressing human SCF, GM-CSF and IL-3. Leukemia 24:1785-8|