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In this paper we demonstrate that patients treated with chemotherapy and/or hematopoietic stem cell transplantation (HSCT) have highly significant reduced serum ascorbic acid (AA) levels compared to healthy controls. We recently observed in in vitro experiments that growth of both T and NK cells from hematopoietic stem cells is positively influenced by AA. It might be of clinical relevance to study the function and recovery of immune cells after intensive treatment, its correlation to AA serum levels and the possible effect of AA supplementation.

Background Aims: Natural killer (NK) cell-based immunotherapy is a promising treatment for a variety of malignancies. However, generating sufficient cell numbers for therapy remains a challenge. To achieve this, optimization of protocols is required., Methods: Mature NK cells were expanded from peripheral blood mononuclear cells PBMCs in the presence of anti-CD3 monoclonal antibody and interleukin-2. Additionally, NK-cell progenitors were generated from CD34(+) hematopoietic stem cells or different T/NK-cell progenitor populations. Generated NK cells were extensively phenotyped, and functionality was determined by means of cytotoxicity assay., Results: Addition of ascorbic acid (AA) resulted in more proliferation of NK cells without influencing NK-cell functionality. In more detail, PBMC-derived NK cells expanded 2362-fold (median, range: 90-31,351) in the presence of AA and were capable of killing tumor cells under normoxia and hypoxia. Moreover, hematopoietic stem cell-derived progenitors appeared to mature faster in the presence of AA, which was also observed in the NK-cell differentiation from early T/NK-cell progenitors., Conclusions: Mature NK cells proliferate faster in the presence of phospho-L-AA, resulting in higher cell numbers with accurate functional capacity, which is required for adoptive immunotherapy., (Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.)

Among prostaglandins (PGs), PGE2 is abundantly expressed in various malignancies and is probably one of many factors promoting tumor growth by inhibiting tumor immune surveillance. In the current study, we report on a novel mechanism by which PGE2 inhibits in vitro natural killer-dendritic cell (NK-DC) crosstalk and thereby innate and adaptive immune responses via its effect on NK-DC crosstalk. The presence of PGE2 during IFN-γ/membrane fraction of Klebsiella pneumoniae DC maturation inhibits the production of chemokines (CCL5, CCL19, and CXCL10) and cytokines (IL-12 and IL-18), which is cAMP-dependent and imprinted during DC maturation. As a consequence, these DCs fail to attract NK cells and show a decreased capacity to trigger NK cell IFN-γ production, which in turn leads to reduced T-helper 1 polarization. In addition, the presence of PGE2 during DC maturation impairs DC-mediated augmentation of NK-cell cytotoxicity. Opposed to their inhibitory effects on peripheral blood-derived NK cells, PGE2 matured DCs induce IL-22 secretion of inflammation constraining NKp44(+) NK cells present in mucosa-associated lymphoid tissue. The inhibition of NK-DC interaction is a novel regulatory property of PGE2 that is of possible relevance in dampening immune responses in vivo.

Mucin-1 (MUC1) is a heavily O-glycosylated, transmembrane protein that is expressed on the apical surface of most secretory epithelia. In malignantly transformed epithelia, MUC1 has lost its apical distribution, is underglycosylated and is secreted into the circulation. Due to the underglycosylation of MUC1, cancer-specific MUC1-Tn/STn antigens, which are highly immunogenic, become exposed. We aimed at developing a system that allows detection of antibodies directed to the native form of MUC1 and the underglycosylated MUC1-Tn epitopes. To this end, we made use of the Chinese Hamster Ovary (CHO) ldlD cell line stably transfected with MUC1. This cell line has a glycosylation defect, which can be reversed by addition of different monosaccharides to the cell culture and enables the production of cells expressing the MUC1-Tn glycoforms. After validation with glycospecific antibodies, the CHO-ldlD MUC1 system was used to detect serum MUC1 and MUC1-Tn antibodies. Using this system, we could confirm the presence of MUC1-Tn antibodies in the serum of a patient vaccinated with a truncated MUC1 peptide. This indicates that the CHO-ldlD MUC1 system represents a flow cytometry-based technique to detect antibodies binding to the underglycosylated MUC1 protein. This cellular system is complementary to the previously published methods to detect MUC1 serum antibodies, since the antibodies to the native protein are evaluated and therefore it can be effectively used for MUC1 antibody monitoring in vaccination studies as well as for functional assays., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Haploidentical hematopoietic stem cell transplantation (haplo-HSCT) is a treatment option for patients with hematopoietic malignancies that is hampered by treatment-related morbidity and mortality, in part the result of opportunistic infections, a direct consequence of delayed T-cell recovery. Thymic output can be improved by facilitation of thymic immigration, known to require precommitment of CD34(+) cells. We demonstrate that Delta-like ligand-mediated predifferentiation of mobilized CD34(+) cells in vitro results in a population of thymocyte-like cells arrested at a T/natural killer (NK)-cell progenitor stage. On intrahepatic transfer to Rag2(-/-)gamma(c)(-/-) mice, these cells selectively home to the thymus and differentiate toward surface T-cell receptor-alphabeta(+) mature T cells considerably faster than animals transplanted with noncultured CD34(+) cells. This finding creates the opportunity to develop an early T-cell reconstitution therapy to combine with HSCT.

Background: In melanoma patients vaccinated with monocyte-derived melanoma peptide-pulsed dendritic cells (DC), the delayed-type hypersensitivity (DTH) reactions have been examined as a surrogate marker to determine if acquired immunity is induced by DC vaccination. To date, however, only limited information has been reported as for histopathological analyses of DTH., Objective: To evaluate tumor-specific immunomonitoring histopathologically after DC vaccination in melanoma patients., Methods: Seven patients previously vaccinated with monocyte-derived melanoma peptide-pulsed DCs were challenged with recall antigenic peptide injection in the skin of the forearm. Using immunohistochemical techniques, the presence of immune cells and the expression of CD4, CD8, interleukin (IL)-2, IL-4, IL-10, Foxp3, CD1a, CD1d, and interferon (IFN)-gamma was investigated at the site of injection where a DTH reaction developed., Results: Strong DTH reactions from infiltrated erythema to bullae formation were detected in all 7 cases. Biopsies taken from the DTH site revealed heavy infiltration of mononuclear cells and eosinophils in the dermis and subcutaneous tissue. Cells staining positively for CD4, CD8, IL-2, IL-4, Foxp3, CD1d, and IFN-gamma were increased at the site 48h after antigen injection in all cases. Cells positive for IL-10 were never found in any patient. Regulatory T cells appeared 6h after injection and reached their maximum at day 7., Conclusions: The significant induction of CD8(+)T cells as well as both Th1 and Th2-type cells at the site of DTH suggests that effective antigen presentation leading to anti-tumor immune responses has taken place. Inhibitory mechanisms may also develop as the disappearance of the DTH response could be related to an increase in Foxp3+ cells.