Optimizing preparation of normal dendritic cells and bcr-abl(+) mature dendritic cells derived from immunomagnetically purified CD14(+) cells

The goal of this work was to optimize dendritic cell (DC) preparations obta ined from patients suffering from chronic myeloid leukemia (CML) and compar e them with DC prepared from normal CD14(+) mononuclear cells (MNC). We stu died normal DC and bcr-abl(+) leukemic DC (CML-DC) yields, expression of me mbrane molecules, differentiation status, and ability to stimulate T cells. We isolated DC precursors from PBMC by CD14-specific immunoadsorption and cultured them for 7 days in GM-CSF and IL-4, followed by a 3-day incubation to fully differentiate the cells. We evaluated cultures of CML-DC using RP MI 1640 medium supplemented with FBS and X-VIVO 15 medium containing human AB serum. In contrast to cells matured in RPMI 1640, virtually all cells in cubated in X-VIVO 15 expressed CD83, a marker of mature DC. CML-DC and norm al DC were indistinguishable in expression of CD83, resulting in the highes t percentage reported so far. The yields of normal DC and CML-DC from CD14( +) cells were indistinguishable. The percentage of bcr-abl(+) cells in PBMC varied among patients between 65% and 97% and the final CML-DC preparation s were >98% bcr-abl(+) the highest purity of bcr-abl(+) cells to date. Norm al DC and CML-DC were equally effective in stimulating proliferation of all ogeneic and autologous T cells. These techniques provide highly enriched, m ature, functional CML-DC.