Generation of a Dendritic Cell‐based Vaccine in Chronic Lymphocytic Leukaemia Using CliniMACS Platform for Large‐scale Production

Abstract

We previously demonstrated that dendritic cells (DC) that have endocytosed apoptotic bodies of autologous leukemic cells (Apo‐DC) can boost antileukemic T‐cell responses. In this study, we report a description of the production procedure and product specification of the Apo‐DC vaccine preparations for clinical use. Enriched populations of CD14+ monocytic precursors and CD19+ leukaemic cells were obtained using CliniMACS technology from a single leukapheresis product. Apoptotic bodies were obtained by irradiating (5 Gy) CD19+ selected B cells. DC were generated ex vivo by culturing monocytes with granulocyte macrophage colony‐stimulating factor and interleukin‐4. Following coculture with apoptotic bodies, DCs were matured with tumour necrosis factor‐α. The mean percentage of CD14+ cells in the peripheral blood as well as in the leukapheresis product of the patients (n = 10) was approximately 2% (range, 0.8–3.3). Immunomagnetic selection using the CD14 reagent yielded a CD14+ population that was 91 ± 2.2% (mean ± SEM) pure. Immunomagnetic selection of CD19 expressing cells yielded a population that was 100 ± 0.03% pure. Cell viability immediately after selection was 97% and 98% after 7 days of culture. The Apo‐DC cellular vaccine product showed a mature phenotype, with a high rate of endocytosis (84%) of apoptotic leukemic B‐cells. In conclusion, despite significant variability in the circulating monocyte frequency of the chronic lymphocytic leukaemia patients, our method permitted the production of a DC vaccine with high reproducibility and conforming with recommended quality standards.

Vaccination with dendritic cells loaded with tumor apoptotic bodies (Apo-DC) in patients with chronic lymphocytic leukemia: effects of various adjuvants and definition of immune response criteria

“…Details of the cellular vaccine production from the first ten patients (cohorts I and II) have been reported in detail elsewhere [8]. Briefly, CD14?…”

“…The vaccine was produced at a certified GMP laboratory from a leukapheresis product in adherence to a previously validated protocol [8]. Briefly, CD14 ?…”

Complex evaluation of human monocyte‐derived dendritic cells for cancer immunotherapy

“…Common strategies used at present for DC maturation include the following: ( i ) combination of tumour-necrosis factor (TNF)-α, IL-1β, IL-6 and prostaglandin E2 (PGE2) or its modifications [13–16]; ( ii ) sole use of TNF-α [17–19]; ( iii ) cocktails containing TNF-α, IL-1β, interferons (IFN) and Toll-like receptor (TLR) ligands [20–22]; ( iv ) cocktails containing TLR ligands and IFN-γ [23–25] and ( v ) cocktails containing CD40 ligand [26,27]. Newly designed maturation cocktails are usually compared only with the combination of TNF-α, IL-1β, IL-6 and PGE2, despite current knowledge that this combination is not optimal for inducing strong anti-tumour immune reaction [21,28,29].…”

A phase I clinical trial combining dendritic cell vaccination with adoptive T cell transfer in patients with stage IV melanoma

“…The standard operating procedure (SOP) used for DC generation has, with minor modifications, already been applied in a number of clinical trials [23,24]. This 7-day protocol resulted in high numbers of mature antigen-loaded DC in all cases.…”