Cytokine-mobilized peripheral blood progenitor cells for allogeneic reconstitution of miniature swine.

Abstract

Because of the relative ease of acquisition, increased yield, and improved engraftment characteristics, mobilized peripheral blood progenitor (stem) cells (PBSCs) have recently become the preferred source for hematopoietic stem cell transplantation. In our laboratory, procurement of a megadose of PBSCs is necessary for on-going studies evaluating non-myelosuppressive transplant regimens for the induction of mixed chimerism and allograft tolerance. To exploit hematopoietic growth factor synergy, we have sought to combine growth factors with proven utility to improve PBSC mobilization and maximize our PBSC procurement through an automated collection procedure. Mobilization characteristics of PBSCs were determined in 2-5-month-old miniature swine. Animals received either swine recombinant stem cell factor (pSCF, 100 microg/kg) and swine recombinant interleukin 3 (pIL-3, 100 microg/kg), administered intramuscularly for 8 days, or pSCF, pIL-3, and human recombinant granulocyte-colony stimulating factor (hG-CSF), at 10 microg/kg. Leukapheresis was performed beginning on day 5 of cytokine treatment and continued daily for 3 days. Collection of PBSCs from cytokine-mobilized animals via an automated leukapheresis procedure demonstrated a 10-fold increase in the number of total nucleated cells (TNC) (20-30 x 10(10) TNC) compared to bone marrow harvesting (2-3 x 10(10) total TNC). A more rapid rise in white blood cells (WBCs) was seen after administration of all three cytokines compared to pSCF and pIL-3 alone. An increase in colony-forming unit granulocyte-macrophage frequency measured daily from peripheral blood during cytokine treatment, was seen with the addition of hG-CSF to pSCF/pIL-3 correlating well with the rise in WBCs. Similarly, the addition of hG-CSF demonstrated a notable increase in the median progenitor cell yield from the 3-day leukapheresis procedure. Cytokine-mobilized PBSCs were capable of hematopoietic reconstitution. PBSCs mobilized with pSCF/pIL-3 were infused into an SLA-matched recipient conditioned with cyclophosphamide (50 mg/kg) and total body irradiation 1150 cGy. Neutrophil and platelet engraftment occurred on days 5 and 7, respectively, with minimal evidence of graft-versus-host disease. Complete donor chimerism has been demonstrated 331 days after transplant. Our preliminary results show that in this well-defined miniature swine model, recombinant swine cytokine combinations (pSCF, pIL-3 with or without hG-CSF) successfully mobilize a high yield of progenitor cells for allogeneic transplantation. Furthermore, these cytokine-mobilized PBSCs demonstrate the potential to reconstitute hematopoiesis and provide long-term engraftment in miniature swine.