Dendritic Cell-Based Immunotherapeutic Strategies in Adult Acute Lymphoid Leukemia Patients Treated with Conventional Therapy or with Imatinib.

In adult patients with acute lymphoid leukemia (ALL) conventional therapies can cure only about 20% of patients. Despite the high proportion of complete remissions, the prognosis remains poor because the disease is not eradicate and patients eventually relapse. It is thus important to develop new therapeutic strategies. Dendritic cell (DC)-based vaccines have been tested in different neoplastic conditions, including patients with hematological malignancies, and the possibility of eliciting specific responses has been documented. In this study, we evaluated in adult ALL patients in complete remission and off-therapy the possibility of generating DC (ALL-DC) and the ability of ALL-DC to stimulate allogeneic and autologous anti-leukemic T lymphocytes. In all 15 B-lineage and 2 T-lineage ALL cases studied it has been possible to generate adequate numbers of DC, i.e. 1–1.5 x 10⁶ from 20 ml of peripheral blood. After gradient separation and adhesion, monocytes were incubated with granulocyte/macrophage colony-stimulating-factor (GM-CSF) and interleukin-4 (IL-4) for 5 days, and tumor necrosis factor (TNF)-α for 48 hours. The expression of the DC markers CD1a, CD80, CD86, CD83 and CD40, evaluated by flow cytometry on ALL-DC, was comparable with that observed on DC derived from 10 normal donors. ALL-DC were capable of loading apoptotic autologous and allogenic leukemic bodies generated in vitro after incubation with Imatinib (for Bcr/Abl+ ALL) or Fludarabine. Successful loading has been demonstrated by flow cytometry utilizing the PKH67 fluorescent dye incubated with the leukemic cells before drug treatment. In all patients, the T-lymphoid population showed a proliferative capacity and an IFNγ and TNFα intracytoplasmic production, after PHA and PMA stimulation, comparable to that generated in lymphocytes from normal donors. Furthermore, in 6/7 B-lineage ALL patients tested DC-pulsed with apoptotic leukemic cells were capable of stimulating the proliferation of both allogenic and autologous lymphocytes. We also studied 4 further patients affected by B-lineage ALL who carried the molecular alteration Bcr/Abl. These patients were treated daily with Imatinib and at the time of the study were in complete morphologic remission. T lymphocytes, repeatedly evaluated, showed a normal proliferative response to PHA stimulation. The generation of DC has proven numerically and phenotypically comparable to that of normal donors, but in 2/3 cases analyzed DC showed a reduced ability to load apoptotic leukemic cells.

In conclusion, the results of this study indicate that DC can be effectively generated in vitro from adult ALL patients at the time of complete remission. These DC pulsed with leukemic apoptotic bodies are capable of stimulating an anti-leukemic T-lymphocyte response, also under autologous conditions. Our data suggest that in ALL patients in complete remission of their disease, an immunotherapeutic strategy utilizing DC could have a role in the control of the leukemia. In Bcr/Abl+ patients treated with Imatinib, further studies are necessary to fully define the competence of the DC compartment.