IFN-γ Enhances TH1 Polarisation of Monocyte-derived Dendritic Cells

Using serum-free conditions, human monocytederived dendritic cells (MoDCs) tend to mature insufficiently in a TH1-polarizing direction under approved and standardized clinical conditions. However, for the initiation of an efficient tumour antigen-specific cytotoxic T-cell response, the induction of a distinct TH1 response is favourable. Therefore, to improve TH1 polarisation, the influence of interferon-gamma (IFN-γ) on the maturation of MoDCs was investigated with clinical-grade cytokines or lipopolysaccharide (LPS) in serum-free medium focusing on the viability, phenotypic characteristics, cytokine profile and restimulating capacities. As in previous research, we confirmed that in respect of viability and phenotypic characteristics, cytokine cocktails consisting of tumour necrosis factor-alpha (TNF-α), interleukin (IL)-1β, IL-6 and prostaglandin (PG) E2, mature MoDCs most efficiently. However, these cytokine-matured MoDCs secreted relatively high levels of IL-10 and only low levels of IL-12p70. Remarkably, if IFN-γ was added, significantly lower levels of IL-10 concomitant with higher levels of IL-12p70 could be detected. Pretreatment with IFN-γ did not improve the phenotypic characteristics nor the TH1 polarisation of MoDCs. Nevertheless, MoDCs matured with clinical-grade cytokines and IFN-γ could be re-stimulated most effectively with IFN-γ. In conclusion, our work demonstrates that addition of INF-γ to clinical-grade cytokine cocktails readily matures MoDCs and enhances their TH1 polarisation efficiently under serum-free conditions. Dendritic cells (DCs) are most potent antigen-presenting cells playing a pivotal role in the induction of the immune response (1, 2). Fully matured DCs are effective activators of naive T-cells and are therefore regarded as important initiators of primary T-cell immune responses. In recent years, several methods have been established to generate DCs in vitro from monocytes isolated from the peripheral blood of volunteers or cancer patients, the so-called monocyte-derived DCs (MoDCs) (3, 4). Loaded with tumour antigen, MoDCs are increasingly applied as vaccines for cancer patients to induce a tumour antigen-specific cytotoxic T-cell (CTL) response (5-7). Nevertheless, an efficient induction of tumour antigen-specific antitumoural CTL response requires both breaking antigen-specific tolerance, as well as changing an immune-suppressive tumour-associated environment (8). As shown in preclinical studies, only fully matured MoDCs characterized by high expression levels of costimulatory molecules (8-10), high migratory activity (11), as well as the induction of a distinct TH1-response (12-15) were able to induce an efficient tumour antigen-specific CTL response in vivo. Unfortunately, 1467 *Both authors contributed equally to this work. Abbrevations: CCR, chemokine receptor; CD, cluster of differentiation; CTL, cytotoxic T lymphocytes; DCs, dendritic cells; moDCs, monocyte-derived dendritic cells; DC1, TH1 polarised dendritic cells; ELISPOT, enzyme-linked immunosorbent spot; FITC: fluorescein-5-isothiocyanat; GM-CSF, granulocytemacrophage colony stimulating factor; HLA, human leucocyte antigen; INF-γ, interferon-gamma; IL, interleukin; LPS, lipopolysaccharide; MHC, major histocompatibility complex; PBMC, peripheral blood mononuclear cells; PBS, phosphatebuffered saline; PE, phycoerythrin; PGE2, prostaglandin E2; THresponse, T-helper cell response; TLR, toll-like receptors; TNF, tumour necrosis factor. Correspondence to: Dr. Bernd Hildenbrand, Tumor Biology Center, Department of Clinical Research, Breisacherstr. 117, D-79106 Freiburg, Germany. Tel: +497612062922, Fax: +497612061861, email: [email protected]