Abstract
Dendritic cell (DC)-based antitumor vaccines have proven to be a safe approach, but often fail to generate robust results between trials. Translation to the clinic has been hindered in part by the lack of standard operation procedures for vaccines production, namely the definition of optimal culture conditions during ex-vivo DC differentiation. Here we sought to compare the ability of three clinical grade serum-free media, DendriMACS, AIM-V, and X-VIVO 15, alongside with fetal bovine serum-supplemented Roswell Park Memorial Institute Medium (RPMI), to support the differentiation of monocyte-derived DCs (Mo-DCs). Under these different culture conditions, phenotype, cell metabolomic profiles, response to maturation stimuli, cytokines production, allogenic T cell stimulatory capacity, as well as priming of antigen-specific CD8+ T cells and activation of autologous natural killer (NK) cells were analyzed. Immature Mo-DCs differentiated in AIM-V or X-VIVO 15 presented lower levels of CD1c, CD1a, and higher expression of CD11c, when compared to cells obtained with DendriMACS. Upon stimulation, only AIM-V or X-VIVO 15 DCs acquired a full mature phenotype, which supports their enhanced capacity to polarize T helper cell type 1 subset, to prime antigen-specific CD8+ T cells and to activate NK cells. CD8+ T cells and NK cells resulting from co-culture with AIM-V or X-VIVO 15 DCs also showed superior cytolytic activity. 1H nuclear magnetic resonance-based metabolomic analysis revealed that superior DC immunostimulatory capacities correlate with an enhanced catabolism of amino acids and glucose. Overall, our data highlight the impact of critically defining the culture medium used in the production of DCs for clinical application in cancer immunotherapy. Moreover, the manipulation of metabolic state during differentiation could be envisaged as a strategy to enhance desired cell characteristics.
Highlights
Dendritic cells (DCs) comprise several subsets of highly specialized professional antigen-presenting cells with superior capacity to acquire, process and present antigens to naïve T-lymphocytes (T-cells) [1]
Monocytes were cultured with granulocytemacrophage colony-stimulating factor (GM-CSF) and IL-4 in three different serumfree media (SFM), AIM-V, X-VIVO-15, and DendriMACS, or with fetal bovine serum (FBS)-supplemented Roswell Park Memorial Institute Medium (RPMI)
After 6 days of differentiation, immature dendritic cell (iDC) were analyzed for their morphological appearance and expression levels of CD14, CD16, CD1a, CD1c, and CD11c
Summary
Dendritic cells (DCs) comprise several subsets of highly specialized professional antigen-presenting cells with superior capacity to acquire, process and present antigens to naïve T-lymphocytes (T-cells) [1] Considering their capacity to elicit strong antitumor immune responses, DCs have been extensively used in immunotherapeutic strategies to fight cancer [2, 3]. There are four main approaches exploring DCs in oncologic treatments: 1) non-targeted protein and nucleic acidbased vaccines captured by DCs in vivo; 2) direct targeting of antigens to DCs in vivo; 3) vaccines composed of ex vivo produced DCs, matured and loaded with tumor antigens; 4) biomaterial based platforms to recruit and program endogenous DCs [4] Among these approaches, ex vivo generated DCs are used in nearly 97% of the registered clinical trials, leukapheresisisolated CD14+ monocytic precursors being the primary source to produce monocyte-derived DCs (Mo-DCs) [5].
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