Ex vivo expansion of enriched peripheral blood CD34+ progenitor cells by stem cell factor, interleukin-1 beta (IL-1 beta), IL-6, IL-3, interferon-gamma, and erythropoietin

Abstract

To provide sufficient numbers of peripheral blood progenitor cells (PBPCs) for repetitive use after high-dose chemotherapy, we investigated the ability of hematopoietic growth factor combinations to expand the number of clonogenic PBPCs ex vivo. Chemotherapy plus granulocyte colony-stimulating factor (G-CSF) mobilized CD34+ cells from 18 patients with metastatic solid tumors or refractory lymphomas were cultured for up to 28 days in a liquid culture system. The effects of interleukin-1 beta (IL-1), IL-3, IL-6, granulocyte-macrophage-CSF (GM-CSF), G-CSF, macrophage-CSF (M-CSF), stem cell factor (SCF), erythropoietin (EPO), leukemia inhibitory factor (LIF), and interferon-gamma, as well as 36 combinations of these factors were tested. A combination of five hematopoietic growth factors, including SCF, EPO, IL-1, IL-3, and IL-6, was identified as the optimal combination of growth factors for both the expansion of total nucleated cells as well as the expansion of clonogenic progenitor cells. Proliferation peaked at days 12 to 14, with a median 190-fold increase (range, 46- to 930-fold) of total clonogenic progenitor cells. Expanded progenitor cells generated myeloid (colony-forming unit-granulocyte-macrophage), erythroid (burst-forming unit-erythroid), as well as multilineage (colony-forming unit-granulocyte, erythrocyte, monocyte, megakaryocyte) colony-forming units. The number of multilineage colonies increased 250-fold (range, 33- to 589-fold) as compared with pre-expansion values. Moreover, the absolute number of early hematopoietic progenitor cells (CD34+/HLA-DR-; CD34+/CD38-), as well as the number of 4-HC-resistant progenitors within expanded cells increased significantly. Interferon-gamma was shown to synergize with the 5-factor combination, whereas the addition of GM-CSF significantly decreased the number of total clonogenic progenitor cells. Large-scale expansion of PB CD34+ cells (starting cell number, 1.5 x 10(6) CD34+ cells) in autologous plasma supplemented with the same 5-factor combination resulted in an equivalent expansion of progenitor cells as compared with the microculture system. In summary, our data indicate that chemotherapy plus G-CSF-mobilized PBPCs from cancer patients can be effectively expanded ex vivo. Moreover, our data suggest the feasibility of large-scale expansion of PBPCs, starting from small numbers of PB CD34+ cells. The number of cells expanded ex vivo might be sufficient for repetitive use after high-dose chemotherapy and might be candidate cells for therapeutic gene transfer.