EX-Vivo expansion of neutrophil precursors and megakaryocytes from cd34+ cells using novel hematopoietic factors receptor agonists in serum free media

Two major complications of high dose chemotherapy are neutropenia and thrdmbocytopenia. Transfusion of ex vivo expanded blood progenitors could reduce the period of cytopenia by providing mature functional cells within days of infusion. We investigated the ability of threejiovel multi-functional hematopoietic growth factor receptor agonists: Synthokine dimer (Syn), Myelopoietin (MPO) and Promegapoietin (PMP) to generate megakaryocyte (MKs)and neutrophil precursors (NP). CD34+ cells purified using the Isolex(r) 300 from G-CSF mobilized apheresis products were cultured in X-VIVO 10(tm) + 1% Buminate containing Syn, MPO, PMP alone or in combination. Starting CFU content was 16.8E04±2.3. Cultures were evaluated at day 12 for cell numbers, phenotype and colony forming cells. Results are mean+/-SD (n=6). Factors Fold Total CFU % CD15+ %CD41a+ Increase (E04) cells (NP) cells (MK) Syn 77±26 8.1±1.8 46.4±2 4 7.4±3 3 MPO 109+44 109±47 670+13 5 5±1 8 PMP 128±61 154±78 55.3±1 6 9 Ol4 6 Syn+MPO 216±122 149+58 65.1±8.7 4 2±2 1 Syn+PMP 274+13.3 242±96 43.7±7 1 10.3±5.2 MPO+PMP 297±19.5 17.7+30 64.5±6.5 6.4+15 Syn+MPO+PMP 37.0+15.9 30.1+5.2 59.3t5 9 7.0+1.8 All three factors alone or in combination supported CD34+ cell proliferation and differentiation. Addition of Syn to MPO or PMP resulted in a significant increase in the total number of NP and MKs, respectively. MPO/PMP or PMP/MPO/Syn generated the largest number of NP and MKs. Syn/MPO/PMP resulted in a significantly higher number of CFU compared to the other combinations tested (p <.05). These data show that a serum free culture system using three novel growth factor receptor agonists can generate large numbers of NP and Mks.