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Prostaglandin E2 has been shown to enhance the maturation, migration, and antigen-presenting capacity of DCs. It is therefore included in many maturation cocktails for the generation of monocyte-derived DCs. Paradoxically, PGE2 is also an important tumor-derived immunosuppressive factor and has inhibitory effects on DC differentiation and function. To further investigate these seemingly contradictory results we studied whether the DC:T cell ratio has an impact on the outcome of the interaction between PGE2-treated DCs and T cells. Surprisingly, at high DC:T cell ratios T cell proliferation was inhibited while at low ratios PGE2-treated DCs displayed enhanced T cell-stimulatory properties. The inhibitory function of PGE2-treated DCs depended primarily on the PGE2-induced induction of indoleamine 2,3-dioxygenase competence. In summary, we show that PGE2-treated DCs can have either an immunogenic or tolerogenic function depending on the DC:T cell ratio. This finding could explain the conflicting results regarding the influence of PGE2 on DC function., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Immune tolerance is a central mechanism counteracting tumor-specific immunity and preventing effective anticancer immunotherapy. Induction of tolerance requires a specific environment in which tolerogenic dendritic cells (DCs) play an essential role deviating the immune response away from effective immunity. It was recently shown that maturation of DCs in the presence of PGE2 results in upregulation of indoleamine 2,3-dioxygenase (IDO) providing a potential mechanism for the development of DC-mediated Tcell tolerance. Here, we extend these findings, demonstrating a concomitant induction of IDO and secretion of soluble CD25 after DC maturation in the presence of PGE2. While maturation of DCs induced IDO expression on transcriptional level, only integration of PGE2 signaling led to up-regulation of functional IDO protein as well as significant expression of cell-surface and soluble CD25 protein. As a consequence, T-cell proliferation and cytokine production were significantly inhibited, which was mediated mainly by IDO-induced tryptophan depletion. Of importance, we demonstrate that different carcinoma entities associated with elevated levels of PGE2 coexpress CD25 and IDO in peritumoral dendritic cells, suggesting that PGE2 might influence IDO expression in human DCs in the tumor environment. We therefore suggest PGE2 to be a mediator of early events during induction of immune tolerance in cancer.

Despite the lack of tumor control, infiltration of immune cells has been demonstrated for several malignancies including non-Hodgkin's lymphoma. Since dendritic cells play a pivotal role in the initiation and control of the immune response, the frequency and phenotype of recently described sub-types of dendritic cells in non-Hodgkin's lymphoma were characterized. Myeloid and plasmacytoid dendritic cells were analysed in 55 non-Hodgkin's lymphoma and 33 reactive lymph nodes by flow cytometry and immunohistochemistry. Overall frequency of dendritic cells in reactive lymph nodes was higher than in non-Hodgkin's lymphoma while the pDC/mDCs ratio was comparable. The low frequency of dendritic cells in infiltrated lymph nodes was confirmed by immunohistochemistry; however, no significant difference in the distribution within lymphoid and tumor tissue was detected. For further characterization of the dendritic cells in non-Hodgkin's lymphoma, the expressions of adhesion molecules, costimulatory molecules, chemokine receptors and activation markers were assessed. Interestingly, a significantly decreased expression of CD62L and CCR7, receptors necessary for homing to lymph nodes, was identified in dendritic cells in non-Hodgkin's lymphoma, potentially explaining the lack of these cells. Taken together, dendritic cells are phenotypically altered and reduced in number in NHL, potentially contributing to the loss of tumor control in these patients.

Despite the still poorly understood complexity of tumor-host immune interactions, the use of cellular vaccines (particularly dendritic cells) has made it possible to reliably generate tumor antigen-specific T cells, both in animal models and in humans. These encouraging pre-clinical results have led to a translation of these immunotherapeutic strategies into clinical trials. With numerous trials still underway, their general outcome has so far been disappointing, and the discrepancy between pre-clinical data and clinical response rates is striking. Thus, either the pre-clinical models have not been representative of the human situation or the translation into human clinical trials is still sub-optimal. Here we suggest new avenues of clinical research to further improve cellular cancer immunotherapy.

MUC1 is a glycoprotein overexpressed in breast cancer and other adenocarcinomas, and is known to elicit cellular and humoral immunity directed against unglycosylated peptide epitopes in the repeat domain. Based on immunological evidence that O-linked glycans on repeat peptides remain intact during processing by dendritic cells (DC), we used MUC1 as a model to address the question which role O-linked glycans play in this process. We were able to identify the sites of proteolysis in MUC1 repeats and the enzyme(s) involved, and elucidated the site-specific effects of O-glycosylation on MUC1 processing by human and mouse DC. Peptides generated by the cellular processing machinery from native mucin or (glyco)peptides suggest specific cleavage at Gly13-Ser14, His20-Gly1 and Thr3-Ser4 peptide bonds in the tandem repeat GVTSAPDTRPAPGSTAPPAH resulting in the initial formation of STA27 or GVT20 and SAP17 as the final product with intact O-glycosylation. Human cathepsin L and the corresponding mouse enzyme in low-density endosomes were identified in vitro to catalyze this site-specific MUC1 proteolysis. O-Glycosylation controls the processing by preventing proteolysis of the Thr3-Ser4 peptide bond if either amino acid is glycosylated, and is responsible for the inertness of tumor-associated MUC1 glycoforms to effective DC processing by masking this cleavage site.

Abstract: Prostaglandin E2 has been shown to enhance the maturation, migration, and antigen-presenting capacity of DCs. It is therefore included in many maturation cocktails for the generation of monocyte-derived DCs. Paradoxically, PGE2 is also an important tumor-derived immunosuppressive factor and has inhibitory effects on DC differentiation and function. To further investigate these seemingly contradictory results we studied whether the DC:T cell ratio has an impact on the outcome of the interaction between PGE2-treated DCs and T cells. Surprisingly, at high DC:T cell ratios T cell proliferation was inhibited while at low ratios PGE2-treated DCs displayed enhanced T cell-stimulatory properties. The inhibitory function of PGE2-treated DCs depended primarily on the PGE2-induced induction of indoleamine 2,3-dioxygenase competence. In summary, we show that PGE2-treated DCs can have either an immunogenic or tolerogenic function depending on the DC:T cell ratio. This finding could explain the conflicting results regarding the influence of PGE2 on DC function. [Copyright &y& Elsevier]