Vaccination of B-CLL Patients with Autologous Dendritic Cells Results in Immunological and Clinical Responses.

Abstract

B-cell chronic lymphocytic leukemia (B-CLL) is a very attractive disease to be targeted by immunotherapy due to its rather low progression and due to the availability of cancer cells in the peripheral blood. Dendritic cells (DCs) are the most professional antigen presenting cells, but are deficient in B-CLL. Therefore administration of ex vivo generated DCs might improve the anti-leukemic T cell response in patients with B-CLL. Our group reported the first clinical trial with allogeneic DCs stimulated with leukemic cells lysates in B-CLL patients. To answer the question whether autologous DCs might cause a similar immunotherapeutical effect we conducted the present study assessing the safety and feasibility as well as the clinical and immunological responses to autologous monocyte-derived DC immunotherapy in B-CLL patients. Precursor cells for DC generation were obtained using two methods: by plastic adherence of PBMC from leukapheresis or CD14 positive separation through magnetic beads of PBMC from 150mL of peripheral blood. Twelve patients at clinical stage 0 and 2 according to Rai were vaccinated intradermally up to eight times with a mean number of 7.4×106 DCs pulsed with B-CLL cell lysates. Specific CD8+ T cell responses to RHAMM and fibromodulin - recently described as tumor specific antigens (TAA) in B-CLL were evaluated by multi-parametric tetramer-based flow cytometry and ELISPOT assays. Furthermore, the percentage of CD4+CD25hiFoxP3+ T regulatory cells in the peripheral blood over the time of vaccination was assessed. We observed a decrease of peripheral blood leukocytes and B CD19+/CD5+ leukemic cells in five patients, three patients showed a stable disease and four patients progressed despite DC vaccination. The best clinical response was observed in the patients treated with DCs generated from adherent fraction of PBMC obtained by leukapheresis. No severe toxicities were noted. After the end of therapy the in vitro proliferation of autologous T cells stimulated by DC pulsed with lysates, as well as by lysates alone increased significantly comparing to the values before treatment (1391±2170, p<0.001; 3280±4842, p<0.001). A significant increase of specific cytotoxic CD8+ T lymphocytes against leukemia-associated antigens (RHAMM/CD168 and fibromodulin), was detected after DC vaccination. In patients with a clinical response, an increase of interleukin 12 (IL-12) serum levels (12±9 pg/ml vs. 6±7 pg/ml; p=0.02,) was noted. In patients with the most favorable clinical response we observed a substantial decrease of CD4+/CD25+ cells (14.3±11.3% vs. 28.6±12.5%) and an increase of CD8+/CD25+ cells percentage after DC therapy (49.3±55.7% vs. 8.9±1.9%), as well as a decrease of the frequency of CD4+CD25hiFOXP3+ T regulatory cells. Taken together, the study demonstrated that vaccination with autologous DC in CLL patients is feasible and safe. Immunological and to some extend hematological responses could be noted, justifying further investigation on this immunotherapeutical approach.