Increased Tumor-Specific CD8⁺ T Cell Induction by Dendritic Cells Matured with a Clinical Grade TLR-Agonist in Combination with IFN-γ

Abstract

The limited response rate of cancer patients treated with dendritic cell (DC)-based vaccines indicates that vast improvements remain necessary. In many murine tumour models it has been demonstrated that the use of innate triggers (e.g. TLR triggers) in the maturation of DC results in higher efficacy. However, as few of these innate triggers are generated clinical grade, there remains a great necessity to fill the gap between fundamental mouse studies and a clinical trial in humans. In the present study we used a TLR2/4-agonist (FMKp which is available clinical grade) in combination with IFN-y (FIcocktail) in the maturation of elutriated monocyte-derived DC and compared it with the most used DC in current clinical trials (TNF -a/PGE-2, i.e, TP-cocktail). In addition to the assessment of CD4+ T cell polarizing capacity, we compared the quantity and intrinsic quality of induced CD8+ T cells of 2 different DC maturation protocols with all cells from the same donor. Besides differences in the cytokine profile, which could be coupled to increased Thl and Th17 polarization, we demonstrate in this study that FMKp/IFN-y matured DC are twice as effective in inducing cytotoxic T cells against known tumor antigens. Both DCs induced phenotypically equivalent effector memory CD8+ T cells that did not show a significant difference in their intrinsic capacity to kill tumor cells. These findings point to the therapeutic applicability of FI-DC as superior inducers of functional antigen-specific T cells. Their increased chemokine secretion is suggestive of a mechanism by which these DC may compensate for the limited migration observed for all ex vivo cultured DC when applied in patients.Dendritic cells (DC) are very efficient initiators of potent immune responses in vivo. These cells are therefore attractive tools for cellular immunotherapy in the treatment of cancer and viral infections that evade the regular immune surveillance (1-3). As a consequence, a range of methods to generate large amounts of DC ex vivo has become available. They can be generated from a variety of sources, but DC differentiation from monocytes with a growth factor cocktail containing IL-4 and GM-CSF is the current gold standard (4-5).Translating these culture methods into good manufacturing practice guidelines has allowed the introduction of DC immunotherapy into the

Klebsiella pneumoniae‐triggered DC recruit human NK cells in a CCR5‐dependent manner leading to increased CCL19‐responsiveness and activation of NK cells

“…For the TNF-a/PGE2 and IFN-g/FMKpmatured DC, 6 h maturation irreversibly triggered maturation as evidenced by the upregulation of MHC class II and costimulatory molecules (Supporting Information Fig. 1), which was comparable to the expression of these markers detected after 48 h maturation [22]. Filtered, cell-free, DC supernatant was harvested after 18 h of culture and transferred to the lower compartment of a Transwell s .…”

Inflammation restraining effects of prostaglandin E2 on natural killer–dendritic cell (NK-DC) interaction are imprinted during DC maturation

“…To this end, monocyte-derived DCs were matured with FMKp and IFN-␥ as described previously. 35 DCs were matured for 6 hours with IFN-␥/FMKp and different concentrations of PGE2. After washing, to remove all maturation factors, DCs were incubated for another 18 hours in medium only (total maturation of 24 hours).…”

“…35 In addition, highly purified monocyte fractions obtained by elutriation of leukapheresis products were used (purity, Ͼ 95%). Written informed consent was obtained from all leukapheresis donors in accordance with the Declaration of Helsinki.…”

“…22,35,39 Based on these observations, PGE2 is often incorporated in DC maturation cocktails used in clinical vaccination studies. Our data on the inhibitory effect of PGE2 on NK-DC interaction in combination with previous reports on enhanced T H 17 and T H 2 induction by PGE2-matured DCs and the decreased ability to induce antigenspecific CTLs 35 argue against the use of PGE2-matured DCs in DC-based vaccines, because it does not contribute to in vivo DC migration and has detrimental effects on immune effector mechanisms. In addition, Muthuswamy et al 48 showed that increased CCR7 expression of PGE2-matured DCs, compared with TLRmatured DCs, correlates with their lack of CCL19 production.…”

Cholesterol: An Inflammatory Compound

“…High levels of plasminogen activator inhibitor-1 result in reduced fibrinolytic capability, thereby contributing to increased risk for thromboembolic events (67)(68)(69)(70)(71).…”