Assuntos
Imunomodulação/efeitos da radiação , Neoplasias/radioterapia , Radiocirurgia/métodos , Animais , Apresentação de Antígeno/efeitos da radiação , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Morte Celular Imunogênica , Imunoterapia/métodos , Camundongos , Neoplasias/imunologia , Hipofracionamento da Dose de Radiação , Fatores de Tempo , Microambiente Tumoral/imunologia , Microambiente Tumoral/efeitos da radiaçãoRESUMO
PURPOSE: Patients with central nervous system (CNS) tumors are typically treated with radiotherapy, but this is not curative and results in the upregulation of phosphorylated STAT3 (p-STAT3), which drives invasion, angiogenesis, and immune suppression. Therefore, we investigated the combined effect of an inhibitor of STAT3 and whole-brain radiotherapy (WBRT) in a murine model of glioma. EXPERIMENTAL DESIGN: C57BL/6 mice underwent intracerebral implantation of GL261 glioma cells, WBRT, and treatment with WP1066, a blood-brain barrier-penetrant inhibitor of the STAT3 pathway, or the two in combination. The role of the immune system was evaluated using tumor rechallenge strategies, immune-incompetent backgrounds, immunofluorescence, immune phenotyping of tumor-infiltrating immune cells (via flow cytometry), and NanoString gene expression analysis of 770 immune-related genes from immune cells, including those directly isolated from the tumor microenvironment. RESULTS: The combination of WP1066 and WBRT resulted in long-term survivors and enhanced median survival time relative to monotherapy in the GL261 glioma model (combination vs. control P < 0.0001). Immunologic memory appeared to be induced, because mice were protected during subsequent tumor rechallenge. The therapeutic effect of the combination was completely lost in immune-incompetent animals. NanoString analysis and immunofluorescence revealed immunologic reprograming in the CNS tumor microenvironment specifically affecting dendritic cell antigen presentation and T-cell effector functions. CONCLUSIONS: This study indicates that the combination of STAT3 inhibition and WBRT enhances the therapeutic effect against gliomas in the CNS by inducing dendritic cell and T-cell interactions in the CNS tumor.
Assuntos
Neoplasias Encefálicas/terapia , Comunicação Celular/imunologia , Quimiorradioterapia/métodos , Glioma/terapia , Fator de Transcrição STAT3/antagonistas & inibidores , Animais , Apresentação de Antígeno/efeitos dos fármacos , Apresentação de Antígeno/efeitos da radiação , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/patologia , Comunicação Celular/efeitos dos fármacos , Comunicação Celular/efeitos da radiação , Linhagem Celular Tumoral/ultraestrutura , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/efeitos da radiação , Modelos Animais de Doenças , Glioma/imunologia , Glioma/patologia , Humanos , Memória Imunológica/efeitos dos fármacos , Camundongos , Piridinas/administração & dosagem , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/efeitos da radiação , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Microambiente Tumoral/efeitos da radiação , Tirfostinas/administração & dosagemRESUMO
The development of immune checkpoint inhibitors (ICIs) is revolutionizing the way we think about cancer treatment. Even so, for most types of cancer, only a minority of patients currently benefit from ICI therapies. Intrinsic and acquired resistance to ICIs has focused research towards new combination therapy approaches that seek to increase response rates, the depth of remission and the durability of benefit. In this Review, we describe how radiotherapy, through its immunomodulating effects, represents a promising combination partner with ICIs. We describe how recent research on DNA damage response (DDR) inhibitors in combination with radiotherapy may be used to augment this approach. Radiotherapy can kill cancer cells while simultaneously triggering the release of pro-inflammatory mediators and increasing tumour-infiltrating immune cells - phenomena often described colloquially as turning immunologically 'cold' tumours 'hot'. Here, we focus on new developments illustrating the key role of tumour cell-autonomous signalling after radiotherapy. Radiotherapy-induced tumour cell micronuclei activate cytosolic nucleic acid sensor pathways, such as cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING), and propagation of the resulting inflammatory signals remodels the immune contexture of the tumour microenvironment. In parallel, radiation can impact immunosurveillance by modulating neoantigen expression. Finally, we highlight how tumour cell-autonomous mechanisms might be exploited by combining DDR inhibitors, ICIs and radiotherapy.
Assuntos
Neoplasias/etiologia , Neoplasias/patologia , Microambiente Tumoral , Animais , Apresentação de Antígeno/imunologia , Apresentação de Antígeno/efeitos da radiação , Biomarcadores Tumorais , Caspases/metabolismo , Reparo do DNA , Suscetibilidade a Doenças , Exossomos/metabolismo , Humanos , Proteínas de Membrana/metabolismo , Terapia de Alvo Molecular , Neoplasias/radioterapia , Nucleotidiltransferases/metabolismo , Processamento de Proteína Pós-Traducional , Radioterapia/efeitos adversos , Radioterapia/métodos , Transdução de Sinais , Microambiente Tumoral/imunologia , Microambiente Tumoral/efeitos da radiaçãoRESUMO
Alterations in gut microbiota impact the pathophysiology of several diseases, including cancer. Radiotherapy (RT), an established curative and palliative cancer treatment, exerts potent immune modulatory effects, inducing tumor-associated antigen (TAA) cross-priming with antitumor CD8+ T cell elicitation and abscopal effects. We tested whether the gut microbiota modulates antitumor immune response following RT distal to the gut. Vancomycin, an antibiotic that acts mainly on gram-positive bacteria and is restricted to the gut, potentiated the RT-induced antitumor immune response and tumor growth inhibition. This synergy was dependent on TAA cross presentation to cytolytic CD8+ T cells and on IFN-γ. Notably, butyrate, a metabolite produced by the vancomycin-depleted gut bacteria, abrogated the vancomycin effect. In conclusion, depletion of vancomycin-sensitive bacteria enhances the antitumor activity of RT, which has important clinical ramifications.
Assuntos
Apresentação de Antígeno/efeitos da radiação , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Microbioma Gastrointestinal , Neoplasias Experimentais , Animais , Apresentação de Antígeno/genética , Antígenos de Neoplasias/genética , Butiratos/imunologia , Linfócitos T CD8-Positivos/patologia , Células Dendríticas/patologia , Feminino , Microbioma Gastrointestinal/imunologia , Microbioma Gastrointestinal/efeitos da radiação , Camundongos , Camundongos Knockout , Neoplasias Experimentais/genética , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/patologia , Neoplasias Experimentais/radioterapiaRESUMO
The expression of antigens that are recognized by self-reactive T cells is essential for immune-mediated tumor rejection by immune checkpoint blockade (ICB) therapy. Growing evidence suggests that mutation-associated neoantigens drive ICB responses in tumors with high mutational burden. In most patients, only a few of the mutations in the cancer exome that are predicted to be immunogenic are recognized by T cells. One factor that limits this recognition is the level of expression of the mutated gene product in cancer cells. Substantial preclinical data show that radiation can convert the irradiated tumor into a site for priming of tumor-specific T cells, that is, an in situ vaccine, and can induce responses in otherwise ICB-resistant tumors. Critical for radiation-elicited T-cell activation is the induction of viral mimicry, which is mediated by the accumulation of cytosolic DNA in the irradiated cells, with consequent activation of the cyclic GMP-AMP synthase (cGAS)/stimulator of interferon (IFN) genes (STING) pathway and downstream production of type I IFN and other pro-inflammatory cytokines. Recent data suggest that radiation can also enhance cancer cell antigenicity by upregulating the expression of a large number of genes that are involved in the response to DNA damage and cellular stress, thus potentially exposing immunogenic mutations to the immune system. Here, we discuss how the principles of antigen presentation favor the presentation of peptides that are derived from newly synthesized proteins in irradiated cells. These concepts support a model that incorporates the presence of immunogenic mutations in genes that are upregulated by radiation to predict which patients might benefit from treatment with combinations of radiotherapy and ICB.
Assuntos
Apresentação de Antígeno/efeitos da radiação , Antígenos de Neoplasias/genética , Neoplasias/imunologia , Radioterapia/efeitos adversos , Animais , Antígenos de Neoplasias/imunologia , Humanos , Mutação , Neoplasias/genética , Neoplasias/radioterapiaRESUMO
Ultraviolet radiation (UVR) can have a beneficial biologic impact on skin, but it is also the most significant environmental risk factor for skin cancer development. Photocarcinogenesis comprises a complex interplay between the carcinogenic UVR, skin, and the immune system. UVB is absorbed by the superficial skin layers and is mainly responsible for direct DNA damage, which, if unrepaired, can lead to mutations in key cancer genes. UVA is less carcinogenic, penetrates deeper in the dermis, and mainly causes indirect oxidative damage to cellular DNA, proteins, and lipids, via photosensitized reactions. UVR not only induces mutagenesis, altering proliferation and differentiation of skin cells, but also has several immunosuppressive effects that compromise tumor immunosurveillance by impairing antigen presentation, inducing suppressive cells, and modulating the cytokine environment. This review focuses upon molecular and cellular effects of UVR, regarding its role in skin cancer development.
Assuntos
Carcinogênese/efeitos da radiação , Tolerância Imunológica/efeitos da radiação , Mutagênese/efeitos da radiação , Neoplasias Induzidas por Radiação , Neoplasias Cutâneas/etiologia , Terapia Ultravioleta/efeitos adversos , Apresentação de Antígeno/efeitos da radiação , Citocinas/efeitos da radiação , Dano ao DNA/efeitos da radiação , Humanos , Estresse Oxidativo/efeitos da radiação , Pele/efeitos da radiação , Dermatopatias/radioterapia , Linfócitos T Reguladores/efeitos da radiaçãoRESUMO
Radiation therapy (RT) is used for local tumor control through direct killing of tumor cells. Radiation-induced cell death can trigger tumor antigen-specific immune responses, but these are often noncurative. Radiation has been demonstrated to induce immunogenic modulation (IM) in various tumor types by altering the biology of surviving cells to render them more susceptible to T cell-mediated killing. Little is known about the mechanism(s) underlying IM elicited by sub-lethal radiation dosing. We have examined the molecular and immunogenic consequences of radiation exposure in breast, lung, and prostate human carcinoma cells. Radiation induced secretion of ATP and HMGB1 in both dying and surviving tumor cells. In vitro and in vivo tumor irradiation induced significant upregulation of multiple components of the antigen-processing machinery and calreticulin cell-surface expression. Augmented CTL lysis specific for several tumor-associated antigens was largely dictated by the presence of calreticulin on the surface of tumor cells and constituted an adaptive response to endoplasmic reticulum stress, mediated by activation of the unfolded protein response. This study provides evidence that radiation induces a continuum of immunogenic alterations in tumor biology, from immunogenic modulation to immunogenic cell death. We also expand the concept of immunogenic modulation, where surviving tumor cells recovering from radiation-induced endoplasmic reticulum stress become more sensitive to CTL killing. These observations offer a rationale for the combined use of radiation with immunotherapy, including for patients failing RT alone.
Assuntos
Apresentação de Antígeno/efeitos da radiação , Calreticulina/farmacologia , Neoplasias/imunologia , Neoplasias/radioterapia , Linfócitos T Citotóxicos/efeitos da radiação , Sequência de Aminoácidos , Apresentação de Antígeno/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/imunologia , Neoplasias da Mama/radioterapia , Linhagem Celular Tumoral , Relação Dose-Resposta à Radiação , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Estresse do Retículo Endoplasmático/imunologia , Estresse do Retículo Endoplasmático/efeitos da radiação , Feminino , Humanos , Imunogenética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/radioterapia , Masculino , Dados de Sequência Molecular , Neoplasias/tratamento farmacológico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/imunologia , Neoplasias da Próstata/radioterapia , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologiaRESUMO
Tumour chemotherapy with drugs is typically associated with severe systemic and local side effects for which reason immunotherapy represents a safer alternative. However, vaccination often fails to generate the required cytotoxic CD8 T-cell responses due to insufficient access of antigens to the cytosol and the MHC class I pathway of antigen presentation. One important issue of tumour research is therefore to develop strategies that allow cytosolic targeting or endosomal escape of tumour antigens. The objective of the current study was to test whether endocytosed antigen could be delivered to MHC class I by means of photochemical internalisation (PCI). Briefly, the antigen and the photosensitiser Amphinex were loaded in vitro onto bone-marrow-derived murine dendritic cells (DCs). After light activation, which is supposed to cause disruption of OVA- and Amphinex-containing endosomes, the DCs were cultured with OVA-specific CD8 T cells or used for immunisation of mice. PCI facilitated CD8 T-cell responses as measured by IFN-γ secretion in vitro and CD8 T-cell proliferation in vivo. In conclusion, the current proof-of-concept study is the first to describe PCI-mediated immunisation and the results revealed the feasibility of this novel technology in autologous vaccination for stimulation of CD8 T-cell responses.
Assuntos
Antígenos/administração & dosagem , Citosol/metabolismo , Genes MHC Classe I , Imunidade Celular/efeitos dos fármacos , Imunização/métodos , Fármacos Fotossensibilizantes/farmacologia , Linfócitos T/imunologia , Animais , Apresentação de Antígeno/efeitos dos fármacos , Apresentação de Antígeno/efeitos da radiação , Antígenos/metabolismo , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/imunologia , Células da Medula Óssea/efeitos da radiação , Células Cultivadas , Citosol/imunologia , Células Dendríticas/citologia , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/efeitos da radiação , Endocitose/efeitos dos fármacos , Endocitose/efeitos da radiação , Endossomos/química , Endossomos/metabolismo , Endossomos/efeitos da radiação , Estudos de Viabilidade , Feminino , Imunidade Celular/efeitos da radiação , Testes de Liberação de Interferon-gama , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fármacos Fotossensibilizantes/efeitos da radiação , Organismos Livres de Patógenos Específicos , Linfócitos T/metabolismoRESUMO
Dendritic cells (DC) can achieve cross-presentation of naturally-occurring tumor-associated antigens after phagocytosis and processing of dying tumor cells. They have been used in different clinical settings to vaccinate cancer patients. We have previously used gamma-irradiated MART-1 expressing melanoma cells as a source of antigens to vaccinate melanoma patients by injecting irradiated cells with BCG and GM-CSF or to load immature DC and use them as a vaccine. Other clinical trials have used IFN-gamma activated macrophage killer cells (MAK) to treat cancer patients. However, the clinical use of MAK has been based on their direct tumoricidal activity rather than on their ability to act as antigen-presenting cells to stimulate an adaptive antitumor response. Thus, in the present work, we compared the fate of MART-1 after phagocytosis of gamma-irradiated cells by clinical grade DC or MAK as well as the ability of these cells to cross present MART-1 to CD8(+) T cells. Using a high affinity antibody against MART-1, 2A9, which specifically stains melanoma tumors, melanoma cell lines and normal melanocytes, the expression level of MART-1 in melanoma cell lines could be related to their ability to stimulate IFN-gamma production by a MART-1 specific HLA-A*0201-restricted CD8(+) T cell clone. Confocal microscopy with Alexa Fluor®(647)-labelled 2A9 also showed that MART-1 could be detected in tumor cells attached and/or fused to phagocytes and even inside these cells as early as 1 h and up to 24 h or 48 h after initiation of co-cultures between gamma-irradiated melanoma cells and MAK or DC, respectively. Interestingly, MART-1 was cross-presented to MART-1 specific T cells by both MAK and DC co-cultured with melanoma gamma-irradiated cells for different time-points. Thus, naturally occurring MART-1 melanoma antigen can be taken-up from dying melanoma cells into DC or MAK and both cell types can induce specific CD8(+) T cell cross-presentation thereafter.
Assuntos
Apresentação de Antígeno/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas/imunologia , Raios gama , Antígeno MART-1/imunologia , Macrófagos/imunologia , Melanoma/imunologia , Fagocitose/imunologia , Apresentação de Antígeno/efeitos da radiação , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , Células Dendríticas/metabolismo , Regulação Neoplásica da Expressão Gênica/imunologia , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Antígeno HLA-A2/imunologia , Antígeno HLA-A2/metabolismo , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Antígeno MART-1/biossíntese , Macrófagos/metabolismo , Melanoma/metabolismo , Microscopia Confocal , Fagocitose/efeitos da radiaçãoRESUMO
Dendritic cells (DC) are endowed with the ability to cross-present antigens from other cell types to cognate T cells. DC are poised to meet polymorphonuclear leukocytes (PMNs) as a result of being co-attracted by interleukin-8 (IL-8), for instance as produced by tumor cells or infected tissue. Human monocyte-derived and mouse bone marrow-derived DC can readily internalize viable or UV-irradiated PMNs. Such internalization was abrogated at 4°C and partly inhibited by anti-CD18 mAb. In mice, DC which had internalized PMNs containing electroporated ovalbumin (OVA) protein, were able to cross-present the antigen to CD8 (OT-1) and CD4 (OT-2) TCR-transgenic T cells. Moreover, in humans, tumor cell debris is internalized by PMNs and the tumor-cell material can be subsequently taken up from the immunomagnetically re-isolated PMNs by DC. Importantly, if human neutrophils had endocytosed bacteria, they were able to trigger the maturation program of the DC. Moreover, when mouse PMNs with E. coli in their interior are co-injected in the foot pad with DC, many DC loaded with fluorescent material from the PMNs reach draining lymph nodes. Using CT26 (H-2(d)) mouse tumor cells, it was observed that if tumor cells are intracellularly loaded with OVA protein and UV-irradiated, they become phagocytic prey of H-2(d) PMNs. If such PMNs, that cannot present antigens to OT-1 T cells, are immunomagnetically re-isolated and phagocytosed by H-2(b) DC, such DC productively cross-present OVA antigen determinants to OT-1 T cells. Cross-presentation to adoptively transferred OT-1 lymphocytes at draining lymph nodes also take place when OVA-loaded PMNs (H-2(d)) are coinjected in the footpad of mice with autologous DC (H-2(b)). In summary, our results indicate that antigens phagocytosed by short-lived PMNs can be in turn internalized and productively cross-presented by DC.
Assuntos
Apresentação de Antígeno , Apoptose/imunologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Neutrófilos/citologia , Neutrófilos/imunologia , Animais , Apresentação de Antígeno/efeitos da radiação , Apoptose/efeitos da radiação , Transporte Biológico/efeitos da radiação , Medula Óssea/imunologia , Linhagem Celular Tumoral , Sobrevivência Celular/imunologia , Sobrevivência Celular/efeitos da radiação , Apresentação Cruzada/imunologia , Apresentação Cruzada/efeitos da radiação , Células Dendríticas/microbiologia , Células Dendríticas/efeitos da radiação , Escherichia coli/fisiologia , Humanos , Interleucina-8/metabolismo , Linfonodos/imunologia , Linfonodos/microbiologia , Camundongos , Neutrófilos/microbiologia , Neutrófilos/efeitos da radiação , Ovalbumina/metabolismo , Fagocitose/imunologia , Fagocitose/efeitos da radiação , Linfócitos T/imunologia , Raios UltravioletaRESUMO
Photodynamic therapy, unlikely to elicit drug-resistance, deserves attention as a strategy to counter this outstanding problem common to the chemotherapy of all diseases. Previously, we have broadened the applicability of this modality to photodynamic vaccination by exploiting the unusual properties of the trypanosomatid protozoa, Leishmania, i.e., their innate ability of homing to the phagolysosomes of the antigen-presenting cells and their selective photolysis therein, using transgenic mutants endogenously inducible for porphyrin accumulation. Here, we extended the utility of this host-parasite model for in vitro photodynamic therapy and vaccination by exploring exogenously supplied photosensitizers. Seventeen novel phthalocyanines (Pcs) were screened in vitro for their photolytic activity against cultured Leishmania. Pcs rendered cationic and soluble (csPcs) for cellular uptake were phototoxic to both parasite and host cells, i.e., macrophages and dendritic cells. The csPcs that targeted to mitochondria were more photolytic than those restricted to the endocytic compartments. Treatment of infected cells with endocytic csPcs resulted in their accumulation in Leishmania-containing phagolysosomes, indicative of reaching their target for photodynamic therapy, although their parasite versus host specificity is limited to a narrow range of csPc concentrations. In contrast, Leishmania pre-loaded with csPc were selectively photolyzed intracellularly, leaving host cells viable. Pre-illumination of such csPc-loaded Leishmania did not hinder their infectivity, but ensured their intracellular lysis. Ovalbumin (OVA) so delivered by photo-inactivated OVA transfectants to mouse macrophages and dendritic cells were co-presented with MHC Class I molecules by these antigen presenting cells to activate OVA epitope-specific CD8+T cells. The in vitro evidence presented here demonstrates for the first time not only the potential of endocytic csPcs for effective photodynamic therapy against Leishmania but also their utility in photo-inactivation of Leishmania to produce a safe carrier to express and deliver a defined antigen with enhanced cell-mediated immunity.
Assuntos
Descoberta de Drogas , Indóis/metabolismo , Espaço Intracelular/metabolismo , Leishmania/fisiologia , Leishmania/parasitologia , Fotoquimioterapia/métodos , Animais , Apresentação de Antígeno/efeitos dos fármacos , Apresentação de Antígeno/efeitos da radiação , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/parasitologia , Linfócitos T CD8-Positivos/efeitos da radiação , Linhagem Celular , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/parasitologia , Células Dendríticas/efeitos da radiação , Endocitose/efeitos dos fármacos , Endocitose/efeitos da radiação , Antígenos HLA/imunologia , Interações Hospedeiro-Parasita , Indóis/química , Indóis/farmacologia , Indóis/uso terapêutico , Espaço Intracelular/efeitos dos fármacos , Espaço Intracelular/efeitos da radiação , Isoindóis , Leishmania/efeitos dos fármacos , Luz , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/parasitologia , Macrófagos/efeitos da radiação , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Ovalbumina/imunologia , Fagossomos/efeitos dos fármacos , Fagossomos/metabolismo , Fagossomos/parasitologia , Fagossomos/efeitos da radiação , Fotólise/efeitos dos fármacos , Fotólise/efeitos da radiação , Solubilidade , Especificidade por SubstratoRESUMO
Extracorporeal photochemotherapy (ECP) is widely used to treat cutaneous T-cell lymphoma, graft-versus-host disease, and allografted organ rejection. Its clinical and experimental efficacy in cancer immunotherapy and autoreactive disorders suggests a novel mechanism. This study reveals that ECP induces a high percentage of processed monocytes to enter the antigen-presenting dendritic cell (DC) differentiation pathway, within a single day, without added cytokines, as determined by enhanced expression of relevant genes. The resulting DCs are capable of processing and presentation of exogenous and endogenous antigen and are largely maturationally synchronized, as assessed by the level of expression of costimulatory surface molecules. Principal component analysis of the ECP-induced monocyte transcriptome reveals that activation or suppression of more than 1100 genes produces a reproducible distinctive molecular signature, common to ECP-processed monocytes from normal subjects, and those from patients. Because ECP induces normal monocytes to enter the DC differentiation pathway, this phenomenon is independent of disease state. The efficiency with which ECP stimulates new functional DCs supports the possibility that these cells participate prominently in the clinical successes of the treatment. Appropriately modified by future advances, ECP may potentially offer a general source of therapeutic DCs.
Assuntos
Diferenciação Celular , Células Dendríticas/citologia , Expressão Gênica , Fotoferese , Apresentação de Antígeno/efeitos dos fármacos , Apresentação de Antígeno/fisiologia , Apresentação de Antígeno/efeitos da radiação , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos da radiação , Separação Celular , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/efeitos da radiação , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/efeitos da radiação , Doença Enxerto-Hospedeiro/imunologia , Humanos , Imunofenotipagem , Hibridização In Situ , Linfoma Cutâneo de Células T/imunologia , Monócitos/citologia , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Radiation therapy affects the immune system. In addition to killing radiosensitive immune cells, it can induce functional changes in those cells that survive. Our recent studies showed that the exposure of dendritic cells (DCs) to radiation in vitro influences their ability to present tumour antigen in vivo. Here we show that local radiation therapy of B16 melanoma tumours inhibits the development of systemic immunity to the melanoma antigen MART-1. This inhibition could not be overcome by intratumoral injection of DCs expressing human MART-1 after radiation therapy, suggesting that a form of immune suppression might have developed. On the other hand, injection of MART-expressing DCs prior to tumour irradiation was able to prevent inhibition from developing. These results suggest that local radiation therapy may block the generation of immunity under some circumstances and that strategies may be required to prevent this and allow radiation-induced cell death to translate fully into the development of systemic immunity.
Assuntos
Apresentação de Antígeno/efeitos da radiação , Antígenos de Neoplasias/imunologia , Células Dendríticas/efeitos da radiação , Melanoma Experimental/radioterapia , Proteínas de Neoplasias/imunologia , Neoplasias Cutâneas/radioterapia , Evasão Tumoral/efeitos dos fármacos , Animais , Células Dendríticas/imunologia , Antígeno MART-1 , Melanoma Experimental/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Doses de Radiação , Neoplasias Cutâneas/imunologiaRESUMO
Until now, the anti-tumor efficacy of synthetic oligodeoxynucleotides containing CpG motifs (CpG ODNs) has been reported in a number of preventive and therapeutic tumor models. Predominately class B CpG ODNs were used, relatively little has been reported regarding the class C CpG ODNs. The present study was, therefore, aimed at assessing the ability of CpG ODNs class C applied as a single agent and in combination with radiotherapy to induce the anti-tumor immunity in an experimental tumor model in mice (subcutaneous [s.c.] B16F1). Class C CpG ODNs applied three times as a single agent efficiently delayed the growth of s.c. B16F1 tumors. The combined therapy (CpG ODNs and tumor irradiation) remarkably enhanced the anti-tumor effect. The peritumoral (p.t.) application of CpG ODNs in combination with irradiation increased the number of dendritic cells (DCs) at the tumor site and improved the antigen loading and maturation of DCs. In conclusion, the combined therapy with CpG ODNs and irradiation creates a unique in situ DCs vaccine that could be easily applicable without prior knowledge of tumor antigens.
Assuntos
Vacinas Anticâncer , Processos de Crescimento Celular , Células Dendríticas/metabolismo , Melanoma Experimental/terapia , Oligodesoxirribonucleotídeos/administração & dosagem , Radioterapia , Animais , Apresentação de Antígeno/efeitos dos fármacos , Apresentação de Antígeno/efeitos da radiação , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Antígeno B7-2/biossíntese , Antígeno B7-2/genética , Antígeno B7-2/imunologia , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/efeitos da radiação , Processos de Crescimento Celular/efeitos dos fármacos , Processos de Crescimento Celular/efeitos da radiação , Terapia Combinada , Células Dendríticas/efeitos dos fármacos , Células Dendríticas/imunologia , Células Dendríticas/patologia , Células Dendríticas/efeitos da radiação , Feminino , Humanos , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Transplante de NeoplasiasRESUMO
BACKGROUND: Prostate hyperthermia and photodynamic therapy can be delivered by a variety of procedures which result in a wide range of temperatures and light energy and cause different kinds of cell death. METHODS: We have addressed the immunogenic effect of heating and UVC irradiation on the prostate cancer (PCa) cell line LNCaP, by studying the release of Danger Associated Molecule Pattern (DAMP) molecules HSP70 and HMGB1 and the dendritic cell (DC) antigen-presenting efficiency. RESULTS: Intracellular upmodulation and extracellular release of HSP70 were inversely correlated. Mild temperatures (43-47 degrees C) induced an early increase of intracellular HSP70, whereas the highest temperature (56 degrees C) induced its extrusion from the cell. Likewise, UVC caused an immediate migration of HSP70 into the cell medium in the absence of any intracellular modulation. 56 degrees C and UVC also induced a robust release of HMGB1. The release of DAMP molecules was closely associated with post-apoptotic membrane damage, as shown by double Annexin V/propidium iodide staining, whereas beta-tubulin, a structural component of cell membranes, was specifically induced by 56 degrees C heating. Tumor uptake strongly impaired the cytokine-driven maturation of DCs and 56 degrees C heating led to a significant recovery of CD83 and CCR7 DC maturation markers, but did not influence the antigen cross-presentation activity. On the contrary, UVC-treated LNCaP had negligible effects on DC maturation, but increased the cross-priming of tumor specific CTL. CONCLUSIONS: These data may be of use in the design of effective non-surgical PCa ablations that combine tumor destruction with long lasting immunity.
Assuntos
Células Dendríticas/imunologia , Proteína HMGB1/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Hipertermia Induzida , Neoplasias da Próstata/imunologia , Raios Ultravioleta , Apresentação de Antígeno/imunologia , Apresentação de Antígeno/efeitos da radiação , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Células Dendríticas/metabolismo , Células Dendríticas/efeitos da radiação , Raios gama , Proteína HMGB1/isolamento & purificação , Humanos , Masculino , Necrose/imunologia , Necrose/metabolismo , Necrose/radioterapia , Neoplasias da Próstata/radioterapia , Neoplasias da Próstata/terapia , Linfócitos T Citotóxicos/citologia , Linfócitos T Citotóxicos/imunologia , Tubulina (Proteína)/biossínteseRESUMO
Cutaneous immunity can be controlled by environmental factors such as ultraviolet (UV) irradiation. UV irradiation affects keratinocytes, antigen presenting cells, such as epidermal Langerhans cells (LC), and T lymphocytes. LC are specialized in antigen presentation. Upon encountering exogenous antigens they migrate to skin draining lymph nodes where they present skin-acquired antigens to naive T cells resulting in effector T cell differentiation. T cell effector functions depend on the activation state of LC, which can be influenced by UV irradiation. After completion T cell mediated cutaneous immune responses need to be downregulated. In this context, CD4(+)CD25(+) regulatory T cells have been shown to play an important role in the suppression of cellular immune responses via inhibition of T cell proliferation. Naturally occurring regulatory T cells develop in the thymus and on the molecular level members of the B7- and TNF-superfamilies are critically involved in the peripheral maintenance of CD4(+)CD25(+) T cells. Substantial evidence exists that peripheral regulatory T cells are responsive to environmental stimuli including UV irradiation. UV-induced regulatory T cells are expanded by UV-exposed cutaneous LC and recently, epidermal expression of vitamin D3 or RANKL (CD254) has been shown to connect the environment to the immune system via expansion of CD4(+)CD25(+) regulatory T cells.
Assuntos
Colecalciferol/metabolismo , Imunidade/efeitos da radiação , Ligante RANK/metabolismo , Receptores do Fator de Necrose Tumoral/metabolismo , Pele/imunologia , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/efeitos da radiação , Animais , Apresentação de Antígeno/efeitos da radiação , Antígenos CD4 , Diferenciação Celular/efeitos da radiação , Colecalciferol/imunologia , Exposição Ambiental , Humanos , Tolerância Imunológica/efeitos da radiação , Subunidade alfa de Receptor de Interleucina-2 , Ligante RANK/genética , Ligante RANK/imunologia , Receptores do Fator de Necrose Tumoral/genética , Receptores do Fator de Necrose Tumoral/imunologia , Transdução de Sinais/imunologia , Transdução de Sinais/efeitos da radiação , Pele/patologia , Pele/efeitos da radiação , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/patologia , Raios UltravioletaRESUMO
The immune response to cancer is critically determined by the way in which tumor cells die. As necrotic, stress-associated death can be associated with activation of antitumor immunity, whole tumor cell antigen loading strategies for dendritic cell (DC)-based vaccination have commonly used freeze-thaw "necrotic" lysates as an immunogenic source of tumor-associated antigens. In this study, the effect of such lysates on the ability of DCs to mature in response to well-established maturation stimuli was examined, and methods to enhance lysate-induced DC activation explored. Freeze-thaw lysates were prepared from murine tumor cell lines and their effects on bone marrow-derived DC maturation and function examined. Unmodified freeze-thaw tumor cell lysates inhibited the toll-like receptor-induced maturation and function of bone marrow-derived DCs, preventing up-regulation of CD40, CD86, and major histocompatibility complex class II, and reducing secretion of inflammatory cytokines [interleukin (IL)-12 p70, tumor necrosis factor-alpha, and IL-6]. Although IL-10 secretion was increased by lysate-pulsed DCs, this was not responsible for the observed suppression of IL-12. Although activation of the nuclear factor-kappaB pathway remained intact, the kinase activity of phosphorylated p38 mitogen-activated protein kinase was inhibited in lysate-pulsed DCs. Lysate-induced DC suppression was partially reversed in vitro by induction of tumor cell stress before lysis, and only DCs loaded with stressed lysates afforded protection against tumor challenge in vivo. These data suggest that ex vivo freeze-thaw of tumor cells does not effectively mimic in vivo immunogenic necrosis, and advocates careful characterization and optimization of tumor cell-derived vaccine sources for cancer immunotherapy.
Assuntos
Apresentação de Antígeno , Células Dendríticas/imunologia , Imunoterapia , Neoplasias/terapia , Animais , Apresentação de Antígeno/imunologia , Apresentação de Antígeno/efeitos da radiação , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/efeitos da radiação , Diferenciação Celular/imunologia , Diferenciação Celular/efeitos da radiação , Citocinas/imunologia , Citocinas/metabolismo , Células Dendríticas/patologia , Feminino , Congelamento , Regulação Neoplásica da Expressão Gênica/imunologia , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Temperatura Alta , Ativação Linfocitária/imunologia , Ativação Linfocitária/efeitos da radiação , Melanoma Experimental , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Necrose/imunologia , Neoplasias/patologia , Radiação Ionizante , Transdução de Sinais/imunologia , Transdução de Sinais/efeitos da radiação , Células Th1/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
We have previously reported that local tumor irradiation, without inducing cell death, can augment the therapeutic efficacy of intratumoral (IT) dendritic cell (DC) vaccination. This study examined potential mechanisms underlying radiation enhancement of IT DC therapy in this setting. Even though ionizing radiation did not mediate tumor cell killing, bone marrow-derived DCs acquired in vitro tumor antigens from irradiated D5 murine melanoma cells more efficiently than from untreated cells. This radiation-enhanced loading of DCs did not induce DC maturation, but was associated with improved cross-priming of T cells both in vitro and in vivo. Furthermore, in vivo pulsing of DCs with irradiated versus untreated tumor cells resulted in superior presentation of tumor antigens to T cells. In addition, tumor irradiation facilitated homing of IT administered DCs to the draining lymph node, possibly by down-regulating CCL21 expression within the tumor mass. Studies of the tumor microenvironment in irradiated versus untreated tumors did not reveal significant inflammatory changes. Moreover, radiation did not promote accumulation of CD4 or CD8 effector T cells within solid tumors. Our results indicate that, without inducing cytotoxicity, tumor irradiation can enhance the ability of DCs to capture tumor antigens, migrate to the draining lymph node, and present processed antigens to T cells. These findings may prove useful in designing future strategies for human cancer immunotherapy.
Assuntos
Vacinas Anticâncer/efeitos da radiação , Movimento Celular/efeitos da radiação , Células Dendríticas/imunologia , Células Dendríticas/efeitos da radiação , Animais , Apresentação de Antígeno/efeitos da radiação , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Vacinas Anticâncer/uso terapêutico , Quimiocina CCL21/imunologia , Quimiocina CCL21/metabolismo , Apresentação Cruzada/efeitos da radiação , Células Dendríticas/metabolismo , Fluoresceínas , Humanos , Imunoterapia Adotiva , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Melanoma Experimental/radioterapia , Camundongos , Camundongos Transgênicos , Radiação Ionizante , Succinimidas , Linfócitos T/citologia , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
We review the field of photoimmunology with emphasis on immunosuppression induced by ultraviolet B radiation. Recent studies have focused on UVB-induced alterations in epidermal Langerhans cell function, resulting in a shift from Th1 to Th2 phenotype and the activation of regulatory T cells as the source of IL-10 that is central to this form of immunosuppression.