A full scale comparative study of methods for generation of functional Dendritic cells for use as cancer vaccines

The Elutra Cell Selection System was used to isolate monocytes after collection of leukapheresis product. The enriched monocytes were cultured in gas permeable Teflon bags with IL-4 and GM-CSF for 24 hours (Fast DC) or 5 days (Standard DC) to obtain immature DCs. The cells were then transfected with mRNA from the leukemia cell line Jurkat E6 by electroporation and incubated for additional 24 h or 2 days in the presence of pro-inflammatory cytokines (TNFα, IL-1β, IL-6 and PGE2) to obtain mature DCs.Mature Fast DC and Standard DC displayed comparable levels of many markers expressed on DC, including HLA-DR, CD83, CD86, CD208 and CCR7. However, compared to Standard DC, mature Fast DC was CD14high CD209low. Fast DC and Standard DC transfected with Jurkat E6-cell mRNA were equally able to elicit T cell specifically recognizing transfected DCs in vitro. IFNγ-secreting T cells were observed in both the CD4+ and CD8+ subsets.Our results indicate that mature Fast DC are functional antigen presenting cells (APCs) capable of inducing primary T-cell responses, and suggest that these cells may be valuable for generation of anti-tumor vaccines.Dendritic cells (DCs) are potent antigen-presenting cells (APCs) involved in the induction of T-cell-mediated immune responses and have appeared as important candidates for cellular-based therapies [1]. Immature DCs are found throughout the body where they capture and process antigens (Ag). When activated by danger signals the DCs start differentiation towards a mature penotype and migration to the T cell dependent areas of secondary lymphoid organs. During this process, they lose the capacity for Ag-capturing and upregulate major histocompatibility complex (MHC)- and costimulatory molecules for stimulation of naive T cells [2-4].In vitro, DCs can be differentiated from various cellular sources, including CD34+ progenitor cells from bone marrow (BM) and cord blood (CB), and monocytes obtained from peripheral blood mononuclear cells (PBMCs)