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1.
Cell Rep ; 30(4): 1027-1038.e4, 2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-31995747

RESUMO

Plasmacytoid dendritic cells (pDCs) and type 2 conventional dendritic cells (cDC2s) are currently under evaluation for use in cancer vaccines. Although both DC subsets can activate adaptive and innate lymphocytes, their capacity to recruit such cells is rarely considered. Here, we show that pDCs and cDC2s display a striking difference in chemokine secretion, which correlates with the recruitment of distinct types of immune effector cells. Activated pDCs express high levels of CXCR3 ligands and attract more CD8+ T cells, CD56+ T cells, and γδ T cells in vitro, compared to cDC2s. Skin from melanoma patients shows an influx of immune effector cells following intradermal vaccination with pDCs or cDC2s, with pDCs inducing the strongest influx of lymphocytes known to possess cytolytic activity. These findings suggest that combining both DC subsets could unite the preferred chemoattractive properties of pDCs with the superior T cell priming properties of cDC2s to ultimately enhance vaccine efficacy.

2.
J Immunol Res ; 2019: 7458238, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31309123

RESUMO

The STAT signaling pathway is important in dendritic cell (DC) development and function. Tumor cells can induce STAT signaling, thereby inhibiting DC maturation and immunostimulatory functions, leading to hampered efficacy of DC-based immunotherapies. Platinum-based chemotherapeutics can inhibit STAT signaling, thereby making them an interesting tool to improve DC development and function. In this study, we provide a comprehensive overview of STAT expression and phosphorylation during DC differentiation and maturation and investigate the effects of platinum drugs on STAT signaling during these processes. Monocytes were differentiated into monocyte-derived DCs (moDCs) with IL-4 and GM-CSF and matured with cytokines or TLR ligands. STAT expression and phosphorylation were analyzed by western blotting, and moDC viability and phenotype were analyzed by flow cytometry. Platinum drugs were added at day 3 of differentiation or at the start of maturation to investigate regulation of the STAT signaling pathway. All STAT proteins were expressed during moDC differentiation and STAT1, STAT5, and STAT6 were phosphorylated. No significant changes occurred in the expression and phosphorylation state of the STAT proteins during differentiation. After maturation with TLR ligands, the expression of STAT1 increased, but other STAT proteins were not affected. Phosphorylation of STAT1 and STAT3 increased during maturation, where TLR ligands induced significantly higher levels of phosphorylation than cytokines. Platinum drugs cisplatin and oxaliplatin significantly inhibited phosphorylation of STAT6 during differentiation and maturation. Treatment did not affect the phenotype or viability of the cells. As STAT6 is an important regulator of DC function, these findings suggest a role for platinum-based chemotherapeutics to enhance DC function via inhibition of STAT signaling, thereby potentially enhancing efficacy of DC-based immunotherapies.


Assuntos
Antineoplásicos/farmacologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/metabolismo , Platina/farmacologia , Fatores de Transcrição STAT/metabolismo , Biomarcadores , Diferenciação Celular , Células Dendríticas/citologia , Células Dendríticas/imunologia , Expressão Gênica , Humanos , Imunofenotipagem , Fosforilação , Fatores de Transcrição STAT/genética , Transdução de Sinais
3.
Proc Natl Acad Sci U S A ; 116(17): 8463-8470, 2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-30962374

RESUMO

There are adaptive T-cell and antibody autoimmune responses to myelin-derived peptides in multiple sclerosis (MS) and to aquaporin-4 (AQP4) in neuromyelitis optica spectrum disorders (NMOSDs). Strategies aimed at antigen-specific tolerance to these autoantigens are thus indicated for these diseases. One approach involves induction of tolerance with engineered dendritic cells (tolDCs) loaded with specific antigens. We conducted an in-human phase 1b clinical trial testing increasing concentrations of autologous tolDCs loaded with peptides from various myelin proteins and from AQP4. We tested this approach in 12 patients, 8 with MS and 4 with NMOSD. The primary end point was the safety and tolerability, while secondary end points were clinical outcomes (relapses and disability), imaging (MRI and optical coherence tomography), and immunological responses. Therapy with tolDCs was well tolerated, without serious adverse events and with no therapy-related reactions. Patients remained stable clinically in terms of relapse, disability, and in various measurements using imaging. We observed a significant increase in the production of IL-10 levels in PBMCs stimulated with the peptides as well as an increase in the frequency of a regulatory T cell, known as Tr1, by week 12 of follow-up. In this phase 1b trial, we concluded that the i.v. administration of peptide-loaded dendritic cells is safe and feasible. Elicitation of specific IL-10 production by peptide-specific T cells in MS and NMOSD patients indicates that a key element in antigen specific tolerance is activated with this approach. The results warrant further clinical testing in larger trials.

4.
Cancer Immunol Immunother ; 67(9): 1425-1436, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30019146

RESUMO

There has recently been a paradigm shift in the field of dendritic cell (DC)-based immunotherapy, where several clinical studies have confirmed the feasibility and advantageousness of using directly isolated human blood-derived DCs over in vitro differentiated subsets. There are two major DC subsets found in blood; plasmacytoid DCs (pDCs) and myeloid DCs (mDCs), and both have been tested clinically. CD1c+ mDCs are highly efficient antigen-presenting cells that have the ability to secrete IL-12p70, while pDCs are professional IFN-α-secreting cells that are shown to induce innate immune responses in melanoma patients. Hence, combining mDCs and pDCs poses as an attractive, multi-functional vaccine approach. However, type I IFNs have been reported to inhibit IL-12p70 production and mDC-induced T-cell activation. In this study, we investigate the effect of IFN-α on mDC maturation and function. We demonstrate that both recombinant IFN-α and activated pDCs strongly enhance mDC maturation and increase IL-12p70 production. Co-cultured mDCs and pDCs additionally have beneficial effect on NK and NKT-cell activation and also enhances IFN-γ production by allogeneic T cells. In contrast, the presence of type I IFNs reduces the proliferative T-cell response. The mere presence of a small fraction of activated pDCs is sufficient for these effects and the required ratio between the subsets is non-stringent. Taken together, these results support the usage of mDCs and pDCs combined into one immunotherapeutic vaccine with broad immunostimulatory features.


Assuntos
Células Dendríticas/imunologia , Interferon Tipo I/farmacologia , Interleucina-12/biossíntese , Células Mieloides/imunologia , Antígenos CD1/imunologia , Antígenos CD1/farmacologia , Técnicas de Cocultura , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Glicoproteínas/imunologia , Glicoproteínas/farmacologia , Humanos , Imunidade Inata , Interferon Tipo I/imunologia , Interferon alfa-2 , Interferon-alfa/imunologia , Interferon-alfa/farmacologia , Interferon gama/biossíntese , Interferon gama/imunologia , Interleucina-12/imunologia , Interleucina-12/farmacologia , Ativação Linfocitária , Células Mieloides/citologia , Células Mieloides/efeitos dos fármacos , Quinolinas/farmacologia , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/farmacologia , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia
5.
Front Immunol ; 9: 1169, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29904379

RESUMO

The identification of activated T-lymphocytes restricted to myelin-derived immunogenic peptides in multiple sclerosis (MS) and aquaporin-4 water channel in neuromyelitis optica (NMO) in the blood of patients opened the possibility for developing highly selective and disease-specific therapeutic approaches. Antigen presenting cells and in particular dendritic cells (DCs) represent a strategy to inhibit pro-inflammatory T helper cells. DCs are located in peripheral and lymphoid tissues and are essential for homeostasis of T cell-dependent immune responses. The expression of a particular set of receptors involved in pathogen recognition confers to DCs the property to initiate immune responses. However, in the absence of danger signals different DC subsets have been revealed to induce active tolerance by inducing regulatory T cells, inhibiting pro-inflammatory T helper cells responses or both. Interestingly, several protocols to generate clinical-grade tolerogenic DC (Tol-DC) in vitro have been described, offering the possibility to restore the homeostasis to central nervous system-related antigens. In this review, we discuss about different DC subsets and their role in tolerance induction, the different protocols to generate Tol-DCs and preclinical studies in animal models as well as describe recent characterization of Tol-DCs for clinical application in autoimmune diseases and in particular in MS and NMO patients. In addition, we discuss the clinical trials ongoing based on Tol-DCs to treat different autoimmune diseases.


Assuntos
Células Dendríticas/imunologia , Imunoterapia/métodos , Esclerose Múltipla/terapia , Neuromielite Óptica/terapia , Linfócitos T Auxiliares-Indutores/imunologia , Alarminas/metabolismo , Animais , Apresentação do Antígeno , Aquaporina 4/imunologia , Autoantígenos/imunologia , Células Dendríticas/transplante , Humanos , Tolerância Imunológica , Imunoterapia/tendências , Ativação Linfocitária , Bainha de Mielina/imunologia , Especificidade do Receptor de Antígeno de Linfócitos T
7.
Front Oncol ; 8: 127, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29755954

RESUMO

Background and objective: Diffuse intrinsic pontine glioma (DIPG) is a lethal brainstem tumor in children. Dendritic cells (DCs) have T-cell stimulatory capacity and, therefore, potential antitumor activity for disease control. DCs vaccines have been shown to reactivate tumor-specific T cells in both clinical and preclinical settings. We designed a phase Ib immunotherapy (IT) clinical trial with the use of autologous dendritic cells (ADCs) pulsed with an allogeneic tumors cell-lines lysate in patients with newly diagnosed DIPG after irradiation (radiation therapy). Methods: Nine patients with newly diagnosed DIPG met enrollment criteria. Autologous dendritic cell vaccines (ADCV) were prepared from monocytes obtained by leukapheresis. Five ADCV doses were administered intradermally during induction phase. In the absence of tumor progression, patients received three boosts of tumor lysate every 3 months during the maintenance phase. Results: Vaccine fabrication was feasible in all patients included in the study. Non-specific KLH (9/9 patients) and specific (8/9 patients) antitumor response was identified by immunologic studies in peripheral blood mononuclear cells (PBMC). Immunological responses were also confirmed in the T lymphocytes isolated from the cerebrospinal fluid (CSF) of two patients. Vaccine administration resulted safe in all patients treated with this schema. Conclusion: These preliminary results demonstrate that ADCV preparation is feasible, safe, and generate a DIPG-specific immune response detected in PBMC and CSF. This strategy shows a promising backbone for future schemas of combination IT.

8.
J Leukoc Biol ; 102(3): 881-895, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28630103

RESUMO

Dendritic cells (DCs) are APCs essential in regulating the immune response. PGE2, produced during inflammation, has a pivotal role in the maturation of DCs and, therefore, is vital for the immune response. The large variety of biologic functions governed by PGE2 is mediated by its signaling through 4 distinct E-type prostanoid (EP) receptors. Immunogenic DCs express EP2 and EP4, which mediate the PGE2 signaling. However, the expression and function of EP receptors in human tolerogenic DCs (tol-DCs), which present an inhibitory phenotype, have not yet, to our knowledge, been assessed. To clarify the role of EP receptors in tol-DCs, we examined the expression of different EP receptors and their effect using selective agonists in human cells. We find that EP2 and EP3 expression are up-regulated in in vitro-generated tol-DCs compared with mature DCs (mDCs). Activation of EP2-EP4 has a direct effect on the surface expression of costimulatory molecules and maturation receptors, such as CD80, CD83, and CD86 or MHCII and CCR7 in tol-DCs, the latter being exclusively modulated by PGE2-EP4 signaling. Importantly, we find that EP2 and EP3 receptors are involved in tolerance induction through IL-10 production by tol-DCs. These results are in sharp contrast with the inflammatory role of EP4 Moreover, we show that DCs generated in the presence of agonists for EP receptors, induce naive T cell differentiation toward polarized Th1/Th17 cells. Given the differential effects of EP receptors, our results suggest that EP receptor agonist/antagonists might become relevant novel drug templates to modulate immune response.


Assuntos
Células Dendríticas/imunologia , Dinoprostona/farmacologia , Tolerância Imunológica/efeitos dos fármacos , Imunossupressores/farmacologia , Receptores de Prostaglandina E Subtipo EP2/imunologia , Receptores de Prostaglandina E Subtipo EP3/imunologia , Antígenos CD/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Interleucina-10/imunologia , Receptores CCR7/imunologia , Células Th1/imunologia , Células Th17/imunologia
9.
Int J Pharm ; 511(2): 785-93, 2016 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-27477102

RESUMO

The design of innovative strategies to selectively target cells, such antigen-presenting cells and dendritic cells, in vivo to induce immune tolerance is gaining interest and relevance for the treatment of immune-mediated diseases. A novel loaded-nanosystem strategy to generate tolerogenic dendritic cells (tol-DCs) was evaluated. Hence budesonide (BDS) was encapsulated in multiwalled polyurethane-polyurea nanoparticles (PUUa NPs-BDS) based on self-stratified polymers by hydrophobic interactions at the oil-water interface. DCs treated with encapsulated BDS presented a prominent downregulation of costimulatory molecules (CD80, CD83 and MHCII) and upregulation of inhibitory receptors. Moreover, DCs treated with these PUUa NPs-BDS also secreted large amounts of IL-10, a crucial anti-inflammatory cytokine to induce tolerance, and inhibited T lymphocyte activation in a specific manner compared to those cells generated with free BDS. These results demonstrate that PUUa NPs-BDS are a highly specific and efficient system through which to induce DCs with a tolerogenic profile. Given the capacity of PUUa NPs-BDS, this delivery system has a clear advantage for translation to in vivo studies.


Assuntos
Budesonida/farmacologia , Células Dendríticas/imunologia , Tolerância Imunológica/efeitos dos fármacos , Nanopartículas/química , Polímeros/química , Poliuretanos/química , Antígenos CD/metabolismo , Antígeno B7-1/metabolismo , Budesonida/química , Proliferação de Células/efeitos dos fármacos , Citocinas/metabolismo , Humanos , Imunoglobulinas/metabolismo , Interleucina-10/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Glicoproteínas de Membrana/metabolismo , Nanopartículas/ultraestrutura , Proteínas Nucleares/metabolismo , Tamanho da Partícula , Linfócitos T/efeitos dos fármacos , Transativadores/metabolismo
10.
J Leukoc Biol ; 97(4): 751-60, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25624460

RESUMO

The aim of this study was to test the hypothesis whether MERTK, which is up-regulated in human DCs treated with immunosuppressive agents, is directly involved in modulating T cell activation. MERTK is a member of the TAM family and contributes to regulating innate immune response to ACs by inhibiting DC activation in animal models. However, whether MERTK interacts directly with T cells has not been addressed. Here, we show that MERTK is highly expressed on dex-induced human tol-DCs and participates in their tolerogenic effect. Neutralization of MERTK in allogenic MLR, as well as autologous DC-T cell cultures, leads to increased T cell proliferation and IFN-γ production. Additionally, we identify a previously unrecognized noncell-autonomous regulatory function of MERTK expressed on DCs. Mer-Fc protein, used to mimic MERTK on DCs, suppresses naïve and antigen-specific memory T cell activation. This mechanism is mediated by the neutralization of the MERTK ligand PROS1. We find that MERTK and PROS1 are expressed in human T cells upon TCR activation and drive an autocrine proproliferative mechanism. Collectively, these results suggest that MERTK on DCs controls T cell activation and expansion through the competition for PROS1 interaction with MERTK in the T cells. In conclusion, this report identified MERTK as a potent suppressor of T cell response.


Assuntos
Linfócitos T CD4-Positivos/enzimologia , Células Dendríticas/enzimologia , Tolerância Imunológica/fisiologia , Ativação Linfocitária/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Receptores Proteína Tirosina Quinases/fisiologia , Antígenos de Bactérias/imunologia , Comunicação Autócrina , Proteínas Sanguíneas/fisiologia , Linfócitos T CD4-Positivos/citologia , Diferenciação Celular , Divisão Celular , Células Cultivadas , Técnicas de Cocultura , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Dexametasona/farmacologia , Indução Enzimática/efeitos dos fármacos , Humanos , Memória Imunológica , Testes de Liberação de Interferon-gama , Teste de Cultura Mista de Linfócitos , Monócitos/citologia , Especificidade do Receptor de Antígeno de Linfócitos T , Regulação para Cima/efeitos dos fármacos , c-Mer Tirosina Quinase
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