Ex vivo expansion of megakaryocyte progenitor cells from normal bone marrow and peripheral blood and from patients with haematological malignancies

A number of haematological and non-haematological malignancies can be successfully treated using high-dose chemotherapy +/- irradiation followed by haematopoietic progenitor cell transplantation. Post transplant, thrombocytopenia and neutropenia always occur and patients require platelet transfusions. It may be possible to reduce the period of thrombocytopenia by re-infusion of ex vivo expanded megakaryocyte progenitors (MP), derived from the progenitor cell graft. We have investigated the expansion of MP from CD34(+) enriched cells from normal bone marrow (NBM) and peripheral blood (PB) and remission BM or PB samples from patients with haematological malignancies. CD34(+) cells were cultured in serum-free medium supplemented with thrombopoietin (TPO), interleukin 1 (IL-1), IL-6 and stem cell factor (SCF) for 7 d, then cell proliferation was assessed by flow cytometry using lineage-specific markers. It was possible to significantly expand the number of MP cells from all sources. There were no major differences in yields of MP from normal BM or PB, or BM from multiple myeloma and non-Hodgkin's lymphoma patients. However, expansion of MP in acute myeloid leukaemia samples was lower than all other samples and the number of megakaryocyte colony-forming units was reduced. Several cytokine combinations were evaluated to optimize UP expansion from NBM. Equivalent yields of MP were obtained using TPO and one of IL-1, IL-3, granulocyte-macrophagc colony-stimulating factor or SCF, suggesting that large cytokine combinations are not necessary for this procedure. It should be possible to scale up the culture conditions described to produce effective MP doses for clinical transplantation.