Detection, isolation, and stimulation of quiescent primitive leukemic progenitor cells from patients with acute myeloid leukemia (AML)

Although many acute myeioid leukemia (AML) colony-forming cells (CFCs) and long-term culture-initiating c ells (LTC-ICs) directly isolated from patients are actively cycling, quiescent progenitors are present in most samples. In the current study, H-3-thymidine (H-3-Tdr) suicide assays demonstrated that most NOD/ SCID mouse leukemia-initiating cells (NOD/SL-ICs) are quiescent in 6 of 7 AML samples. AML cells in G(0), G(1), and S/G(2)(+)M were isolated from 4 of these samples using Hoechst 33342/pyroninY staining and cell sorting. The progenitor content of each subpopulation was consistent with the H-3-Tdr suicide results, with, NOD/ SL-ICs found almost exclusively among G(0) cells While the cycling status of AML CFCs and LTC-ICs was more heterogeneous. Interestingly, after 72 hours in serum-free culture With or without Steel factor (SF), Fit-3 ligand (FL), and interleukin-3 (IL-3), most G(0) AML cells entered active cell cycle (percentage of AML cells remaining in G(0) at 72 hours, 1.2% to 37%, and 0% to 7.6%, in cultures without and with growth factors [GFs], respectively) while G(0) cells from normal lineage-depleted bone marrow remained quiescent in the absence of GF. All 4 AML samples showed evidence of autocrine production of 2 or more of SF, FL, IL-3, and granulocyte-macrophage colony-stimulating factor (GM-CSF); in addition, 3 of 4 sa moles. contained an internal tandem duplication of the FLT3 gene. In summary, quiescent leukemic cells, including NOD/SL-ICs, are present in most AML patients. Their spontaneous entry into active cell cycle in short-term culture might be explained by the deregulated GF signaling present in many AMLs. (C) 2003 by The American Society of Hematology.