Granulocyte-macrophage progenitors as candidate leukemic stem cells in blast-crisis CML

Background: The progression of chronic myelogenous leukemia (CML) to blast crisis is supported by self-renewing leukemic stem cells. In normal mouse hematopoietic stem cells, the process of self-renewal involves the (beta)-catenin-signaling pathway. We investigated whether leukemic stem cells in CML also use the (beta)-catenin pathway for self-renewal. Methods: We used fluorescence-activated cell sorting to isolate hematopoietic stem cells, common myeloid progenitors, granulocyte-macrophage progenitors, and megakaryocyte-erythroid progenitors from marrow during several phases of CML and from normal marrow. BCR-ABL, (beta)-catenin, and LEF-1 transcripts were compared by means of a quantitative reverse-transcriptase-polymerase-chain-reaction assay in normal and CML hematopoietic stem cells and granulocyte-macrophage progenitors. Confocal fluorescence microscopy and a lymphoid enhancer factor/T-cell factor reporter assay were used to detect nuclear (beta)-catenin in these cells. In vitro replating assays were used to identify self-renewing cells as candidate leukemic stem cells, and the dependence of self-renewal on (beta)-catenin activation was tested by lentiviral transduction of hematopoietic progenitors with axin, an inhibitor of the (beta)-catenin pathway. Results: The granulocyte-macrophage progenitor pool from patients with CML in blast crisis and imatinib-resistant CML was expanded, expressed BCR-ABL, and had elevated levels of nuclear (beta)-catenin as compared with the levels in progenitors from normal marrow. Unlike normal granulocyte-macrophage progenitors, CML granulocyte-macrophage progenitors formed self-renewing, replatable myeloid colonies, and in vitro self-renewal capacity was reduced by enforced expression of axin. Conclusions: Activation of (beta)-catenin in CML granulocyte-macrophage progenitors appears to enhance the self-renewal activity and leukemic potential of these cells.