Acute myeloid leukemia (AML) is a clonal neoplastic disease that originates from transformed cells which have progressively acquired critical genetic changes that disrupt key growth-regulatory pathways. Despite the established use and optimization of regimens applying polychemotherapy and the development of new agents that are effective at reducing the tumor burden, relapse continues to be the most common cause of death in AML. Recent experimental evidence proposes a pathogenetic model of AML in which at least two major molecular events are required for the development of malignant cells. One causes a differentiation arrest and a second confers proliferative properties to the cells, thereby leading to the formation of a pool of leukemia/cancer stem cells (LSC) that give rise to a hierarchy of functionally heterogeneous bulk tumor cells with more or less limited self-renewal capacity.
As a consequence, future treatments should not only aim at reducing the bulk tumor (blast) population but must be directed against those LSC if one aims at a cure of the disease. Similar to normal hematopoietic stem cells (HSC), LSCs in AML are quiescent and have unlimited self-renewal and clonogenic capacity. As they are quiescent, LSC commonly do not respond well to cell cycle-directed cytotoxic agents and thus contribute to treatment failure. However, elimination of the LSC compartment within the leukemia clone will be essential for cure of leukemia. Thus, defining the characteristics of LSC is critical for both, to understand the genesis of leukemia and to develop strategies by which these cells can be eradicated. However, the molecular mechanisms by which genetic and epigenetic events lead to the formation of LSC are largely unknown.
In the past, research efforts have been focusing on the bulk tumor cells, but this approach is hampered by at least two conceptual challenges: first, the bulk tumor cells appear at the final stage of disease and are hence likely to show many secondary alterations besides the primary oncogenic events that lead to cancer stem cell formation. Second, an adequate cellular control population is not available. This is particularly true for acute leukemias, in which a developmental block is a hallmark of the disease and results in a bulk tumor population of immature cells that cannot be easily compared to normal blood or bone marrow (BM) cells. In order to identify LSC pathways, novel experimental approaches other than the examination of bulk tumor cells need to be established.
Transcriptional analysis is a potentially powerful method to reveal deregulated transcriptional networks and their effect on the function of normal and malignant HSCs and progenitors, particularly as genes encoding transcription factors, which govern these networks, are frequently mutated, rearranged, or otherwise disrupted in human AML (for review see Tenen DG, Nat Rev Cancer 2003). The transcription factor PU.1 can serve as a good example as PU.1 is indispensable for myelomonocytic differentiation in normal hematopoiesis, and reduced function of PU.1 has been linked with the development of AML in mice and humans (McKercher et al., EMBO J 1996; Vangala et al., Blood 2003; Rosenbauer et al., Nat Genet 2004; Mueller et al., Blood 2006; Steidl et al., Nat Genet 2006; Steidl et al., J Clin Invest 2007). Targeted disruption of a distal enhancer, which reduces PU.1 expression levels by 80%, leads to the development of AML in mice. The course of disease includes a preleukemic phase followed by a leukemic phase with high numbers of malignant immature cells in the blood and marrow and frequent genetic aberrations including complex karyotypes. However, the molecular mechanisms underlying the formation of LSC and the transcriptional targets that mediate the tumor suppressor function of PU.1 in stem cells are still only poorly understood.
Findings from our own group and others demonstrate a critical role of PU.1 and JUNB in the genesis and function of LSC in mice and humans, and that PU.1 and JUNB are already deregulated in the early stem cell compartment. Our recent work has demonstrated proof of principle of analysis of disturbed transcription in specific stem and progenitor populations, and that this type of analysis is capable of defining targets that are essential for LSC function (Steidl et al., Nat Genet 2006; Steidl et al., J Clin Invest 2007).
The goal of our research is to better understand the molecular genetic events that progressively occur in distinct stem and progenitor compartments and how they lead to the formation and maintenance of LSC in myeloid malignancies including AML and MDS. The identification of these mechanisms is of major importance as the involved pathways will provide targets as well as markers for LSC-directed therapeutic approaches.
To address these questions we are using PU.1 knockdown-induced AML as a newly established model system as well as primary samples from patients with AML and MDS.