Efficient presentation of tumor idiotype to autologous T cells by CD83(+) dendritic cells derived from highly purified circulating CD14(+) monocytes in multiple myeloma patients

Objectives. To generate mature and fully functional CD83(+) dendritic cells derived from circulating CD14(+) cells highly purified from the leukaphere sis products of multiple myeloma patients, Materials and Methods. CD14(+) monocytes mere selected by high-gradient mag netic separation and differentiated to immature dendritic cells with granul ocyte-macrophage colony-stimulating factor and interleukin-4 For 6-7 days a nd then induced to terminal maturation by the addition of tumor necrosis fa ctor-alpha. or stimulation with CD40 ligand, Dendritic cells were character ized by immunophenotyping, evaluation of soluble antigens uptake, cytokine secretion, capacity of stimulating allogeneic T cells, and ability of prese nting nominal antigens, including tumor idiotype, to autologous T lymphocyt es. Results, Phenotypic analysis showed that 90% +/- 6% of cells recovered afte r granulocyte-macrophage colony-stimulating factor and interleukin-4 stimul ation expressed all surface markers typical of immature dendritic cells and demonstrated a high capacity of uptaking soluble antigens as shown by the FITC-dextran assay, Subsequent exposure to maturation stimuli induced the d ownregulation of CD1a and upregulation of CD83, HLA-DR, costimulatory molec ules and induced the secretion of large amounts of interleukin-12 Mature CD 83(+) cells showed a diminished ability of antigen uptake whereas they prov ed to be potent stimulators of allogeneic T cells in a mixed lymphocyte rea ction, Monocyte-derived dendritic cells, pulsed before the addition of matu ration stimuli, were capable of presenting soluble proteins such as keyhole limpet hemocyanin and tetanus toxoid to autologous T cells for primary and secondary Immune response, respectively. Conversely, pulsing of mature (CD 83(+)) dendritic cells was less efficient for the induction of T-cell proli feration. More importantly, CD14(+) cells-derived dendritic cells stimulate d autologous T-cell proliferation in response to a tumor antigen such as th e patient-specific idiotype, Moreover, idiotype-pulsed dendritic cells indu ced the secretion of interleukin-2 and gamma-interferon by purified CD4(+) cells, T-cell activation was better achieved when Fab immunoglobulin fragme nts were used as compared with the whole protein. When dendritic cells deri ved from CD14(+) cells from healthy volunteers were analyzed, we did not fi nd any difference with samples from myeloma patients as for cell yield, phe notypic profile, and functional characteristics. Conclusion. These studies demonstrate that mobilized purified CD14(+) cells represent the optimal source for the production of a homogeneous cell popu lation of mature CD83(+) dendritic cells suitable for clinical trials in mu ltiple myeloma, (C) 2000 International Society for Experimental Hematology. Published by Elsevier Science Inc. Published by Elsevier Science Inc.