| Acute myeloid leukemia (AML) consists of a heterogeneous population of malignant cells that appears to be hierarchically structured, just as the normal hematopoietic system. This heterogeneous population of AML cells contains only a limited number of primitive cells that are characterized by the immunophenotypic markers CD34+/CD38- and long-term engraftment potential in NOD-SCID mice. The leukemic stem cells (LSCs) have properties in common with normal HSCs, especially the capacity of stem cell self-renewal, but differ with regard to the differentiation program, which is often impaired. Apparently, the transforming events in HSCs or Multipotent Progenitor Cells (MPPs) trigger a genetic program that deregulates the differentiation program at several levels and enhances the self-renewal potential of immature stem and/or progenitor cells. At the molecular level this is reflected by the sustained activation of a number of proteins instead of the normal pattern of transient phosphorylation and de-phosphorylation. Recent studies in our lab have shown that the sustained activation of the transcription factor STAT5 in cord blood stem cells triggers the self-renewal potential of HSCs and strongly influences the interaction of HSCs with stromal cells, but limits differentiation along the myeloid lineage. This transcription factor is of interest in AML since a high number of AML cases demonstrate sustained STAT5 tyrosine phosphorylation due to mutations in cytokine receptors (Flt3, c-Kit), several chromosomal translocation products such as Tel/Jak2 and Tel/Abl, or due to autocrine produced growth factors. Apparently, this transcription factor has great impact on the genetic program that controls relevant properties of HSCs. The aim of this research line is to elucidate mechanisms by which several cellular defects in AML that converge at the level of STAT5 activation can control self-renewal and hampered myelopoiesis of the developing leukemic clone. We are currently optimizing culture conditions for AML cells and are trying to define alternative ways to isolate, study, and manipulate the leukemic hematopoietic stem cell. Ongoing studies include the downmodulation of STAT5 expression in the leukemic stem cell using RNAi to determine whether this restores myelopoiesis and alleviates self-renewal of the leukemic stem cell. Leukemic transformation has often been described as a multi-step process, in which an initial mutation results in enhanced proliferation, which in conjunction with mutations in the differentiation and apoptotic program, synergize in the leukemic event. Although our preliminary data indicate that a persistent activation of STAT5 in human CB CD34+ cells results in enhanced self-renewal on MS5 stromal cells, it is currently unclear whether this persistent activation of STAT5 is also sufficient to induce a leukemia-like phenotype in NOD-SCID mice. It is plausible that, as is the case for e.g. AML1-ETO, one single hit is not sufficient to induce a full leukemic phenotype in vivo. We are currently pursuing to build a multi-step leukemic transformation model in which constitutively activated STAT5A(1*6) together with N-Ras, NF-kB mutants, AML1-ETO or FLT3 ITDs are introduced into human stem and progenitor cells to deepen our insights into the development of leukemia. |
