Regulatory T cells, dendritic cells and neutrophils in patients with renal cell carcinoma.

We evaluated dendritic cells (DC), regulatory T lymphocytes (Treg) and neutrophils in 37 patients with newly diagnosed renal cell carcinoma (RCC) in the tumor and peripheral blood (PB) and correlated these parameters with tumor staging (early-T1, 2, late-T3, 4 and metastatic disease). The number of myeloid and plasmacytoid DC in blood of RCC patients was higher than in healthy controls. The percentage of myeloid dendritic cells (mDC) from CD45+ cells in tumors was higher in comparison with peripheral blood irrespective of disease stage. Higher percentage of these cells expressed a maturation marker in the periphery in the early stage (CD83 expressing cells). The number of plasmacytoid dendritic cells (pDC) in PB was similar in both early and late stage groups, but the early group displayed a significantly higher percentage of pDC in tumor cell suspension. Neutrophil counts in the peripheral blood of RCC patients were higher than in healthy controls, but the counts in both tumor stage groups were similar. The proportion of neutrophils from CD45+ cells was higher in late stage tumors. Higher percentage of Treg from CD4+ cells was detected in renal carcinoma tissue in comparison to PB with no difference between stages of the disease. Our results reflect the complex interplay between various cells of the immune system and the tumor microenvironment. Activation of dendritic cell subpopulations at early stages of the disease course is counterbalanced by the early appearance of T regulatory cells both in the periphery and tumor tissue. Later stages are characterized by the accumulation of neutrophils in the tumor. Appropriate timing of anticancer strategies, especially immunotherapy, should take these dynamics of the immune response in RCC patients into account.

Poly I: C-activated dendritic cells that were generated in CellGro for use in cancer immunotherapy trials.

BACKGROUND: For clinical applications, dendritic cells (DCs) need to be generated using GMP-approved reagents. In this study, we tested the characteristics of DCs generated in two clinical grade culture media and activated by three maturation stimuli, Poly I: C, LPS and the mixture of proinflammatory cytokines in order to identify the optimal combination of culture media and activation stimulus for the clinical use. METHOD: We tested DCs generation using two GMP-certified culture media, CellGro and RPMI+5% human AB serum and evaluated DCs morphology, viability and capapability to mature. We tested three maturation stimuli, PolyI:C, LPS and the mixture of proinflammatory cytokines consisting of IL-1, IL-6, TNF and prostaglandin E2. We evaluated the capacity of activated DCs to induce antigen-specific T cells and regulatory T lymphocytes. RESULTS: Cell culture in CellGro resulted in a higher yield of immature DCs resulting from increased number of adherent monocytes. DCs that were generated in CellGro and activated using Poly I:C were the most efficient in expanding antigen-specific T cells compared to the DCs that were generated in other media and activated using LPS or the cocktail of proinflammatory cytokines. A comparison of all tested combinations revealed that DCs that were generated in CellGro and activated using Poly I:C induced low numbers of regulatory T cells. CONCLUSION: In this study, we identified monocyte-derived DCs that were generated in CellGro and activated using Poly I:C as the most potent clinical-grade DCs for the induction of antigen-specific T cells.

Intensive physical activity increases peripheral blood dendritic cells.

We analyzed the frequency and absolute numbers of circulating myeloid and plasmacytoid DCs in peripheral blood and evaluated their maturation status to test the hypothesis that significant physical stress to the body might induce measurable changes in DCs subsets, phenotype and function, which would complete existing knowledge about the response of the cellular immune system to an acute exercise in top sportsmen. We evaluated the heart rate and draw blood samples before and after the physical load in 18 profesional ice-hockey players. We observed an increase in leukocytes numbers with a predominant increase in the population of DCs and lymphocytes after exercise. Both myeloid and plasmacytoid DCs increased significantly. We found a correlation between the increase of peripheral blood DCs and serum epinephrine and norepinephrine levels. Increase in peripheral blood DCs also correlates with the extent of heart rate elevation during exercise.

Paricalcitol (19-nor-1,25-dihydroxyvitamin D2) and calcitriol (1,25-dihydroxyvitamin D3) exert potent immunomodulatory effects on dendritic cells and inhibit induction of antigen-specific T cells.

Paricalcitol (19-nor-1,25/OH(2)/D(2)), a second generation vitamin D receptor (VDR) activator, is a synthetic analogue of vitamin D3. In contrast to calcitriol, paricalcitol has a reduced effect on intestinal calcium resorption thus avoiding undesirable hypercalcemia. Information about immunomodulatory activity of paricalcitol is scarce. In this study we show that, in all investigated aspects, paricalcitol retains significant immunomodulatory activity, comparable to calcitriol. Both VDR agonists impaired differentiation of immature dendritic cells (DCs) from monocytes. The presence of VDR agonists during DC differentiation abolished their capacity to be activated and, despite potent Toll-like receptor mediated stimulation, VDR agonist-treated DCs remained in the immature state. In accordance with these findings, VDR-treated DCs produced no bioactive IL-12 and had a significantly decreased capacity to induce antigen-specific T cells while the capacity to induce functional Tregs remained unchanged when compared to control DCs. As DCs and T cells play an important role in the pathogenesis of atherosclerosis, in end-stage renal disease patients, paricalcitol should be a VDR agonist of choice for the reduction of the risk of atherosclerosis due to its immunomodulatory effect proven in this study and known limited hypercalcemic effect. The immunomodulatory potency of paricalcitol makes it a drug of interest in the therapy of chronic immune-mediated inflammatory diseases.

FOCUS on FOCIS: combined chemo-immunotherapy for the treatment of hormone-refractory metastatic prostate cancer.

Immunotherapy has emerged as another treatment modality in cancer. The goal of immunotherapy in advanced cancer patients does not have to be the complete eradication of tumor cells but rather the restoration of a dynamic balance between tumor cells and the immune response. Appropriate combination of tumor mass reduction (by surgery and/or chemotherapy) and neutralization of tumor-induced immunosuppression might set the right conditions for the induction of anti-tumor immune response by active immunotherapy. We review experimental basis and key concepts of combined chemo-immunotherapy and document its principles in the case report of patient with hormone refractory metastatic prostate cancer with sinister prognosis. More than four hundred prostate cancer patients have been treated with DC-based immunotherapy and tumor-specific immune responses have been reported in two-thirds of them. In half of these patients, DC immunotherapy resulted in transient clinical responses. Tregs, among other factors, potently inhibit tumor-specific T cells. Prostate cancer patients have elevated numbers of circulating and tumor infiltrating Tregs and there is evidence that Tregs increase tumor growth in vivo. Because of the high frequency of circulating Tregs in our patients, we first administered metronomic cyclophosphamide. After obtaining IRB approval, we started regular vaccinations with dendritic cells (DCs) loaded with killed prostate cancer cells. In accordance with the principles of combined immunotherapy, we continued palliative chemotherapy with docetaxel to reduce the tumor cell burden. DC-based vaccination induced prostate cancer cell-specific immune response. Combined chemo-immunotherapy consisting of alternate courses of chemotherapy and vaccination with mature DCs pulsed with LNCap prostate cancer cell line led to the marked improvement in the clinical and laboratory presentation and to the decrease of PSA levels by more than 90%.

Kinetics of dendritic cells reconstitution and costimulatory molecules expression after myeloablative allogeneic haematopoetic stem cell transplantation: implications for the development of acute graft-versus host disease.

Allogeneic hematopoetic stem cell transplantation (HSCT) represents a unique opportunity to monitor the kinetics of reconstitution of dendritic cells (DCs) and their dynamics in distinct pathologies. We analyzed DCs reconstitution after myeloablative HSCT. We separately analyzed patients with acute GVHD. DCs were monitored from the earliest phase of hematopoetic reconstitution until day +365. Both myeloid DCs and plasmacytoid DCs appeared at earliest stages after engraftment and relative numbers within white blood cells compartment peaked between days 19-25 after HSCT. Their proportion then gradually declined and absolute numbers of both DC subsets remained lower than in controls during the whole follow-up. Patients with acute GVHD had significantly lower numbers of circulating DCs. Decrease in DC counts preceded onset of clinical symptoms by at least 24 h and was independent of corticosteroids administration. This study reveals quantification of plasmacytoid and myeloid DCs as a potential biomarker for the prediction of acute GVHD development.