An optimised biphasic culture system for the generation of functional dendritic cells from patients with acute lymphoblastic leukaemia at presentation and in clinical remission

We have tested the hypothesis that functional dendritic cells (DC) may be g enerated from patients with acute lymphoblastic leukaemia (ALL). We evaluat ed the production of DC from blast cells taken at presentation from nine ch ildren with ALL. Blast cells were expanded in serum-free medium supplemente d with Flt3L, G-CSF, GM-CSF, IL-3, IL-6 and SCF for 7 days and subsequently stimulated with Flt3L, GM-CSF and TGF-beta for a further 14 days, with the addition of TNF-alpha for the final 48 h of culture. Cultured cells had th e morphological appearance of DC and expressed the DC-associated antigens C D1A (range 2-87%) and CD83 (15-44%). Expression of the co-stimulatory molec ules CD80 and CD86 was increased and the majority of these cells retained t heir expression of CD34 (73 +/- 4%) and HLA-DR (79 +/- 5%). Seven of the ni ne ALL had a leukaemia-specific abnormality and DC generated from five of t hese seven cases were derived from the leukaemic clone. Leukaemic DC derive d from four HLA-A*02-positive ALL pulsed with CMV-associated peptides could induce significant proliferation of peptide-specific CD8(+) T cells. This specificity was verified using tetrameric complexes of HLA class I/antigeni c peptide. DC could also be generated from cells taken at times of complete remission of ALL and from normal controls using these culture conditions. These findings show that functional DC can be generated both from ALL blast s and from patients in remission; these might be utilised in future for imm unotherapeutic strategies in the treatment of ALL.