DONOR-TO-DONOR VARIABILITY IN THE EXPANSION POTENTIAL OF HUMAN BONE-MARROW CELLS IS REDUCED BY ACCESSORY CELLS BUT NOT BY SOLUBLE GROWTH-FACTORS

Clinical trials assessing the utility of cultured hematopoietic cells for the support of patients receiving high-dose chemotherapy are begin ning. Although many reports have described these cultures, little is k nown about the donor-to-donor variability that might be expected lo oc cur in widespread use. Therefore, this study was undertaken to assess variables which might predict and reduce the donor-to-donor variabilit y ill cell expansion potential. CD34-enriched cell cultures, plated to contain 3000 CD34(+)lin(-) cells per well, exhibited a wide range of cell output (0.02 to 5.07 x 10(6)) with a high coefficient of variatio n (C-v = 0.69, n = 52). The range in CFU-GM output was even greater (1 2 to 9455, C-v = 0.90). Addition of preformed stroma had a significant positive effect, and resulted in narrower ranges of eel (0.19 to 8.27 x 10(6), C-v = 0.41) and CFU-GM (218 to 17586, C-v = 0.54) output. A wide range of stromal-dependence was exhibited by CD34-enriched cells from different donors, with stroma augmenting cell output by 1.2- to 1 4-fold (mean 3.5), and CFU-GM output by 1.7- to 24-fold (mean 6.5). In contrast, changes in the soluble growth factor combination affected c ells from different donors in a similar fashion, thereby altering the mean level of performance without reducing donor-to-donor variability. Experiments were next performed to assess the relative contribution o f CD34(+)lin(-) cells and stromal cells tit culture variability by cul turing CD34(+)lin(-) cells from three donors on preformed stroma from three donors in parallel. Variability in culture output was attributed to the CD34(+)lin(-) cell donor, whereas stroma from different autolo gous or allogeneic donors gave similar performance. Therefore, both ex pansion potential and stromal-dependency were inherent characteristics of CD34(+)lin(-) cells from different donors. donor characteristics ( i.e., sex, age, weight, and height) and flow cytometric assays (i.e., CD34(+)lin(-) cell purity, and CD38(-), Thy-1(+), and c-kit(+) subsets thereof) were not well correlated with expansion potential. In contra st, many of the different biological characteristics (i.e., inoculum C FU-GM, cell and CFU-GM output, and stromal-dependency) were strongly c orrelated with each other. Mononuclear cell (MNC) cultures, which prov ide an accessory cell environment (including endogenous struma) in whi ch CD34(+)lin(-) cells grow, were compared with CD34-enriched cell cul tures. MNC cultures (containing 3000 CD34(+)lin(-) cells) were found t o give the greatest and most consistent cell (2.51 to 5.20 x 10(6), C- v = 0.17) and CFU-GM (2618 to 14,745, C-v = 0.46) output. These result s have significant implications for the design of clinical trials of c ultured hematopoietic cells, as well as for the understanding of diver sity in human stem cell behavior. Furthermore, the results demonstrate the importance of a large sample size in scientific studies of primar y human hematopoietic cell behavior.