Short-term cultured, interleukin-15 differentiated dendritic cells have potent immunostimulatory properties.
BACKGROUND
Optimization of the current dendritic cell (DC) culture protocol in order to promote the therapeutic efficacy of DC-based immunotherapy is warranted. Alternative differentiation of monocyte-derived DCs using granulocyte macrophage colony-stimulating factor (GM-CSF) and interleukin (IL)-15 has been propagated as an attractive strategy in that regard. The applicability of these so-called IL-15 DCs has not yet been firmly established. We therefore developed a novel pre-clinical approach for the generation of IL-15 DCs with potent immunostimulatory properties.


METHODS
Human CD14+ monocytes were differentiated with GM-CSF and IL-15 into immature DCs. Monocyte-derived DCs, conventionally differentiated in the presence of GM-CSF and IL-4, served as control. Subsequent maturation of IL-15 DCs was induced using two clinical grade maturation protocols: (i) a classic combination of pro-inflammatory cytokines (tumor necrosis factor-alpha, IL-1beta, IL-6, prostaglandin E2) and (ii) a Toll-like receptor (TLR)7/8 agonist-based cocktail (R-848, interferon-gamma, TNF-alpha and prostaglandin E2). In addition, both short-term (2-3 days) and long-term (6-7 days) DC culture protocols were compared. The different DC populations were characterized with respect to their phenotypic profile, migratory properties, cytokine production and T cell stimulation capacity.


RESULTS
The use of a TLR7/8 agonist-based cocktail resulted in a more optimal maturation of IL-15 DCs, as reflected by the higher phenotypic expression of CD83 and costimulatory molecules (CD70, CD80, CD86). The functional superiority of TLR7/8-activated IL-15 DCs over conventionally matured IL-15 DCs was evidenced by their (i) higher migratory potential, (ii) advantageous cytokine secretion profile (interferon-gamma, IL-12p70) and (iii) superior capacity to stimulate autologous, antigen-specific T cell responses after passive peptide pulsing. Aside from a less pronounced production of bioactive IL-12p70, short-term versus long-term culture of TLR7/8-activated IL-15 DCs resulted in a migratory profile and T cell stimulation capacity that was in favour of short-term DC culture. In addition, we demonstrate that mRNA electroporation serves as an efficient antigen loading strategy of IL-15 DCs.


CONCLUSIONS
Here we show that short-term cultured and TLR7/8-activated IL-15 DCs fulfill all pre-clinical prerequisites of immunostimulatory DCs. The results of the present study might pave the way for the implementation of IL-15 DCs in immunotherapy protocols.
Anguille
Sébastien
S
University of Antwerp - Faculty of Medicine, Vaccine & Infectious Disease Institute (Vaxinfectio), Laboratory of Experimental Hematology, Universiteitsplein 1, B-2610 Wilrijk (Antwerp), Belgium. sebastien.anguille@uza.be
Smits
Evelien L J M
EL
Cools
Nathalie
N
Goossens
Herman
H
Berneman
Zwi N
ZN
Van Tendeloo
Vigor F I
VF
D000276:Adjuvants, Immunologic / Q000378:metabolism*; D000956:Antigens, Viral / Q000276:immunology; D018414:CD8-Positive T-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000821:virology; D002454:Cell Differentiation / Q000187:drug effects*; D002465:Cell Movement / Q000187:drug effects; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000166:cytology* / Q000187:drug effects / Q000276:immunology*; D018274:Electroporation; D000939:Epitopes; D006801:Humans; D018664:Interleukin-12 / Q000378:metabolism; D019409:Interleukin-15 / Q000494:pharmacology*; D010587:Phagocytosis / Q000187:drug effects; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011971:Receptors, Immunologic / Q000378:metabolism; D013997:Time Factors; D051199:Toll-Like Receptor 7 / Q000378:metabolism; D051200:Toll-Like Receptor 8 / Q000378:metabolism; D014162:Transfection
D000276:Adjuvants, Immunologic; D000956:Antigens, Viral; D000939:Epitopes; D019409:Interleukin-15; D012333:RNA, Messenger; D011971:Receptors, Immunologic; D051199:Toll-Like Receptor 7; D051200:Toll-Like Receptor 8; D018664:Interleukin-12