A full scale comparative study of methods for generation of functional Dendritic cells for use as cancer vaccines.
BACKGROUND
Dendritic cells (DCs) are professional antigen-presenting cells with the ability to induce primary T-cell responses and are commonly produced by culturing monocytes in the presence of IL-4 and GM-CSF for 5-7 days (Standard DC). Recently, Dauer and co-workers presented a modified protocol for differentiation of human monocytes into mature DCs within 48 hours (Fast DC). Here we report a functional comparison of the two strategies for generation of DCs from human monocytes with adaptions for large-scale clinical use.


METHODS
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 (TNFalpha, IL-1beta, IL-6 and PGE2) to obtain mature DCs.


RESULTS
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. IFNgamma-secreting T cells were observed in both the CD4+ and CD8+ subsets.


CONCLUSIONS
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.
Jarnjak-Jankovic
Silvija
S
Department of Pediatric Research, The National Hospital, Oslo, Norway. silvija.jankovic@rr-research.no
Hammerstad
Hege
H
Saebøe-Larssen
Stein
S
Kvalheim
Gunnar
G
Gaudernack
Gustav
G
D000938:Antigen-Presenting Cells / Q000276:immunology; D001781:Blood Component Removal; D019496:Cancer Vaccines / Q000276:immunology / Q000494:pharmacology*; D018929:Cell Culture Techniques / Q000379:methods; D045744:Cell Line, Tumor; D003713:Dendritic Cells / Q000166:cytology / Q000276:immunology*; D006801:Humans; D018189:Immunomagnetic Separation; D016130:Immunophenotyping; D008297:Male; D009000:Monocytes / Q000166:cytology*; D009369:Neoplasms / Q000517:prevention & control*; D012333:RNA, Messenger; D012680:Sensitivity and Specificity; D013601:T-Lymphocytes / Q000276:immunology; D014162:Transfection
D019496:Cancer Vaccines; D012333:RNA, Messenger