Recent clinical data suggest that peripheral blood mononuclear cells mobilized with recombinant granulocyte stimulating factor (G-PBMC) may be a better product than aspirated marrow for achieving rapid hematopoietic reconstitution following myeloablative therapy. Currently, there is no satisfactory explanation for the rapid engraftment kinetics seen with G- PBMC. Although G- PBMC may contain three- to four-fold more CD34 cells than an average marrow harvest, studies have shown no correlation between CD34 cell number and rate of engraftment after reaching a threshold of 5 x 106 CD34 cells/kg. Therefore, an increase in CD34 cell number is unlikely to account for the difference in reconstitution. An alternative hypothesis is that the CD34 cells in G-PBMC are qualitatively different from those in marrow. To test this hypothesis, functionally distinct subpopulations of CD34 cells, enriched for long- term culture-initiating cells (LTC-IC) and colony-forming units (CFU), will be isolated from G-PBMC. The surface phenotype, proliferative potential, cytokine responsiveness, and novel gene expression of these cells will be compared to that of comparable populations isolated from marrow and cord blood. Clinical data also support the hypothesis that other cells, in addition to CD34 cells, may be qualitatively different. For example, the benefits of rapid engraftment seen with G-PBMC in the allogeneic setting have not been diminished by an increase in the incidence or severity of acute graft-versus- host disease (aGBHD). This is of particular interest since graft-versus-host reactions are attributed to donor T cells, and the G-PBMC product has, on average, one log more T cells than aspirated marrow. These observations would suggest that cell populations such as T cells and graft facilitating accessory cells may be quantitatively or qualitatively different in G-PBMC compared to marrow. To test this hypothesis, populations of accessory cells will be isolated from G-PBMC, and their effect on stromal cell function and alloreactivities will be analyzed in vitro and compared to data obtained with comparable populations isolated from marrow. The studies proposed in this application should help identify the functional components of human G- PBMC that may be required to achieve rapid stable engraftment in an allogeneic host. This information is imperative to the success of the overall goal, which is to optimize the use of G-PBMC and broaden its application to include the treatment of nonmalignant diseases like sickle cell anemia and other inborn errors of metabolism.
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