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Mass spectrometric analysis identified the peptide recognized by a cytotoxic T lymphocyte (CTL) specific for the chemically induced BALB/c Meth A sarcoma as derived from a 17beta-hydroxysteroid dehydrogenase type 12 (Hsd17b12) pseudogene present in the BALB/c genome, but only expressed in Meth A sarcoma. The sequence of the peptide is TYDKIKTGL and corresponds to Hsd17b12(114-122) with threonine instead of isoleucine at codon 114 and is designated Hsd17b12(114T). Immunization of mice with an Hsd17b12(114T) peptide-pulsed dendritic cell-based vaccine or a non-viral plasmid construct expressing the Hsd17b12(114T) peptide protected the mice from lethal Meth A tumor challenge in tumor rejection assays. A Hsd17b12(114-122) peptide-pulsed vaccine was ineffective in inducing resistance in mice to Meth A sarcoma. These results confirm the immunogenicity of the identified tumor peptide, as well as demonstrate the efficacies of these vaccine vehicles. These findings suggest that the role of the human homolog of Hsd17b12, HSD17B12, as a potential human tumor antigen be explored.

There are differences in the clinical course of chronic viral hepatitis C between adults and children, but it is generally accepted that the disease has cell-mediated immune background. The aim of this study was to evaluate PBMC subsets in children with chronic hepatitis C before treatment in order to find some predictive factors, useful for patients management. Several PBMC subsets, in particular lymphoid and dendritic cell (DC) ones, were tested by flow cytometry in HCV(+) paediatric patients (n = 46) and in control children matched in terms of age and sex (n = 20). Data were subjected to extensive statistics. It was found that cells with cytotoxic potential such as CD8(+)CD28(-) T cells, NK and NKT cells as well as lineage(-)HLA-DR(+) DC were increased in per cent values, while CD4(+) T cells and CD4:CD8 ratio were decreased in hepatitis C group. In HCV(+) patients, CD4(+) T cells were inversely correlated with alanine aminotransferase (ALT) levels and with viraemia. DC subset of myeloid origin (CD11c(+)) assessed both in per cent values and as mean fluorescence intensity (MFI) of HLA-DR expression was shown to be downregulated in hepatitis patients, in spite of increased numbers. To conclude, PBMC subsets, and in particular DC, are affected by HCV chronic infection in children, reflected by the correlation with clinical parameters, such as ALT and viraemia.

The discovery of dendritic cells (DC) as professional antigen presenting cells has opened up new possibilities for their use in the development of cancer vaccines. Here we show that unpulsed BM-derived CD11c+CD8alpha-DC administered s.c. to mice enhanced their resistance to subsequent lethal tumor challenges. The tumor resistance-inducing activity of unpulsed DC was dependent on their maturation status, involved CD4+ and CD8+ T cell activities, and was abrogated by pulsing with an irrelevant class I MHC-restricted peptide. Although the BALB/c Meth A tumor system was employed extensively to demonstrate the tumor resistance inducing activity of unpulsed DC, the immunogenicity of this vaccine was evident in other inbred strains of mice against a variety of syngeneic tumors. The broad anti-tumor responses induced by unpulsed DC appears to be due to their capacity to present "self" tumor-associated antigens, such as the H2-Kd-restricted, wild type sequence p53(232-240) peptide. These findings highlight the potential broad applicability of unpulsed DC as a tumor vaccine.

Increased levels of interleukin (IL)-10 have been described as a negative prognostic indicator for survival in patients with various types of cancer. IL-10 exerts tolerogenic and immunosuppressive effects on dendritic cells, which are crucial for the induction of an antitumor immune response. Blood dendritic cell antigen (BDCA)-2 and BDCA-4 are specifically expressed by CD123(bright) CD11c- plasmacytoid dendritic cells; whereas BDCA-1 and BDCA-3 define 2 distinct subsets of CD11c+ myeloid dendritic cells. In this study, the T-helper cell (Th)1/Th2 cytokine serum profile of 65 hepatocellular carcinoma patients was assessed. We found that serum levels of IL-10 were substantially increased in hepatocellular carcinoma patients as compared with controls. Peripheral blood mononuclear cells from healthy volunteers were exposed to recombinant human (rh)IL-10 in vitro to additionally characterize its impact on distinct blood dendritic cell subsets. A dramatic decrease of all myeloid dendritic cell (MDC) and plasmacytoid dendritic cell (PDC) subsets was detectable after 24 hours of continuous rhIL-10 exposure. Moreover, the expression of HLA-DR, CD80 and CD86, was significantly reduced on rhIL-10-treated dendritic cell subsets. Direct ex vivo flow cytometric analysis of various dendritic cell subpopulations in peripheral blood from hepatocellular carcinoma patients revealed an immature phenotype and a substantial reduction of circulating dendritic cells that was associated with increased IL-10 concentrations in serum and with tumor progression. These findings confirm a predominantly immunosuppressive role of IL-10 for circulating dendritic cells in patients with hepatocellular carcinoma and, thus, may indicate novel aspects of tumor immune evasion.

Dendritic cells (DCs) induce and regulate T-cell-mediated immune responses. Circulating precursor (p)DC1 and pDC2 from patients with chronic hepatitis B virus (HBV) infection were quantified by flow cytometry. To assess their function, DC1 were cultured from patients and compared to those of healthy volunteers. HBV patients exhibited a significant decrease in the proportion of freshly isolated pDC1 to pDC2. DC1 propagated from patients showed lower expression of costimulatory molecules and impaired allostimulatory capacity in comparison to controls. After exposure to proinflammatory cytokines, expression of costimulatory molecules, secretion of interleukin-12 (IL-12) and allostimulatory properties increased, but capacity for T-cell stimulation and IL-12 production remained inferior to that of control DCs. HBV-DNA was amplified by polymerase chain reaction in DC1 cultured from all patients. Viral particles were visible in DC1 by electron microscopy. These results suggest that intracellular presence of HBV impairs DC1 functional maturation and subsequent deficits in T-lymphocyte activation may contribute to viral persistence.

It has been suggested that dendritic cells (DCs) are capable of ingesting apoptotic tumor cells (ATCs) and presenting tumor-associated antigens to immune cells. We evaluated the potential of human DCs, which have ingested ATCs, to serve as a source of antigenic epitopes for presentation to T cells specific for PCI-13, a squamous cell carcinoma of the head and neck cell line. Immature DCs (DCimm) generated in the presence of interleukin 4 and granulocyte machrophage colony-stimulating factor from peripheral blood monocytes of HLA-A2+ healthy donors were incubated in the presence of ATCs. Uptake of ATCs by DCs was monitored by flow cytometry and confocal microscopy after 2-18 h of coincubation. When DCs were matured (DCmat) in the presence of proinflammatory cytokines, their capacity to uptake ATCs was reduced. Responses of PCI-13-specific CD8+ T cells to unmodified PCI-13 cells and to DCimm or DCmat +/- ATCs or +/- tumor lysates were tested in gamma-IFN enzyme-linked immunospot and cytotoxicity assays. Unmodified tumor cells were found to be the best stimulators of antitumor activity of the established T-cell line, and ATCs alone were minimally stimulatory. However, DCs that ingested ATCs were able to present tumor antigens to CTLs, and DCimm and DCmat were almost equally stimulatory. When DCs plus various tumor-derived preparations were used as antigen-presenting cells with autologous HLA-A2+ T cells obtained from normal donors, DCs that had ingested ATCs were more effective in generating CD8+ CTLs than tumor cells alone or DCs pulsed with tumor lysates. The results indicate that human DCs fed with ATCs and then matured effectively generated T cell-mediated antitumor responses in vitro.