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1.
Nat Immunol ; 24(9): 1487-1498, 2023 09.
Artículo en Inglés | MEDLINE | ID: mdl-37474653

RESUMEN

Malaria is caused by Plasmodium species transmitted by Anopheles mosquitoes. Following a mosquito bite, Plasmodium sporozoites migrate from skin to liver, where extensive replication occurs, emerging later as merozoites that can infect red blood cells and cause symptoms of disease. As liver tissue-resident memory T cells (Trm cells) have recently been shown to control liver-stage infections, we embarked on a messenger RNA (mRNA)-based vaccine strategy to induce liver Trm cells to prevent malaria. Although a standard mRNA vaccine was unable to generate liver Trm or protect against challenge with Plasmodium berghei sporozoites in mice, addition of an agonist that recruits T cell help from type I natural killer T cells under mRNA-vaccination conditions resulted in significant generation of liver Trm cells and effective protection. Moreover, whereas previous exposure of mice to blood-stage infection impaired traditional vaccines based on attenuated sporozoites, mRNA vaccination was unaffected, underlining the potential for such a rational mRNA-based strategy in malaria-endemic regions.


Asunto(s)
Vacunas contra la Malaria , Malaria , Animales , Ratones , Células T de Memoria , Malaria/prevención & control , Hígado , Plasmodium berghei/genética , Linfocitos T CD8-positivos
2.
Cancer Immunol Immunother ; 72(7): 2267-2282, 2023 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-36881133

RESUMEN

AIM: We have previously reported that polyfunctional T cell responses can be induced to the cancer testis antigen NY-ESO-1 in melanoma patients injected with mature autologous monocyte-derived dendritic cells (DCs) loaded with long NY-ESO-1-derived peptides together with α-galactosylceramide (α-GalCer), an agonist for type 1 Natural Killer T (NKT) cells. OBJECTIVE: To assess whether inclusion of α-GalCer in autologous NY-ESO-1 long peptide-pulsed DC vaccines (DCV + α-GalCer) improves T cell responses when compared to peptide-pulsed DC vaccines without α-GalCer (DCV). DESIGN, SETTING AND PARTICIPANTS: Single-centre blinded randomised controlled trial in patients ≥ 18 years old with histologically confirmed, fully resected stage II-IV malignant cutaneous melanoma, conducted between July 2015 and June 2018 at the Wellington Blood and Cancer Centre of the Capital and Coast District Health Board. INTERVENTIONS: Stage I. Patients were randomised to two cycles of DCV or DCV + α-GalCer (intravenous dose of 10 × 106 cells, interval of 28 days). Stage II. Patients assigned to DCV + α-GalCer were randomised to two further cycles of DCV + α-GalCer or observation, while patients initially assigned to DCV crossed over to two cycles of DCV + α-GalCer. OUTCOME MEASURES: Primary: Area under the curve (AUC) of mean NY-ESO-1-specific T cell count detected by ex vivo IFN-γ ELISpot in pre- and post-treatment blood samples, compared between treatment arms at Stage I. Secondary: Proportion of responders in each arm at Stage I; NKT cell count in each arm at Stage I; serum cytokine levels at Stage I; adverse events Stage I; T cell count for DCV + α-GalCer versus observation at Stage II, T cell count before versus after cross-over. RESULTS: Thirty-eight patients gave written informed consent; 5 were excluded before randomisation due to progressive disease or incomplete leukapheresis, 17 were assigned to DCV, and 16 to DCV + α-GalCer. The vaccines were well tolerated and associated with increases in mean total T cell count, predominantly CD4+ T cells, but the difference between the treatment arms was not statistically significant (difference - 6.85, 95% confidence interval, - 21.65 to 7.92; P = 0.36). No significant improvements in T cell response were associated with DCV + α-GalCer with increased dosing, or in the cross-over. However, the NKT cell response to α-GalCer-loaded vaccines was limited compared to previous studies, with mean circulating NKT cell levels not significantly increased in the DCV + α-GalCer arm and no significant differences in cytokine response between the treatment arms. CONCLUSIONS: A high population coverage of NY-ESO-1-specific T cell responses was achieved with a good safety profile, but we failed to demonstrate that loading with α-GalCer provided an additional advantage to the T cell response with this cellular vaccine design. CLINICAL TRIAL REGISTRATION: ACTRN12612001101875. Funded by the Health Research Council of New Zealand.


Asunto(s)
Melanoma , Neoplasias Cutáneas , Masculino , Humanos , Adolescente , Neoplasias Cutáneas/terapia , Neoplasias Cutáneas/metabolismo , Péptidos/metabolismo , Anticuerpos/metabolismo , Citocinas/metabolismo , Células Dendríticas , Antígenos de Neoplasias , Melanoma Cutáneo Maligno
3.
Bioconjug Chem ; 2023 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-37022946

RESUMEN

Synthetic vaccines that induce T cell responses to peptide epitopes are a promising immunotherapy for both communicable and noncommunicable diseases. Stimulating strong and sustained T cell responses requires antigen delivery to appropriately activated antigen presenting cells (APCs). One way this can be accomplished is by chemically conjugating immunogenic peptide epitopes with α-galactosylceramide (α-GalCer), a glycolipid that acts as an immune adjuvant by inducing stimulatory interactions between APCs and type I natural killer T (NKT) cells. Here we investigate whether increasing the ratio of antigen:adjuvant improves antigen-specific T cell responses. A series of conjugate vaccines was prepared in which one, two, four, or eight copies of an immunogenic peptide were covalently attached to a modified form of α-GalCer via the poly(ethoxyethylglycinamide) dendron scaffold. Initial attempts to synthesize these multivalent conjugate vaccines involved attaching the bicyclo[6.1.0]non-4-yne (BCN) group to the adjuvant-dendron structure followed by strain-promoted azide-alkyne cycloaddition of the peptide. Although this approach was successful for preparing vaccines with either one or two peptide copies, the synthesis of vaccines requiring attachment of four or eight BCN groups suffered from low yields due to cyclooctyne degradation. Instead, conjugate vaccines containing up to eight peptide copies were readily achieved through oxime ligation with adjuvant-dendron constructs decorated with the 8-oxo-nonanoyl group. When evaluating T cell responses to vaccination in mice, we confirmed a significant advantage to conjugation over admixes of peptide and α-GalCer, regardless of the peptide to adjuvant ratio, but there was no advantage to increasing the number of peptides attached. However, it was notable that the higher ratio conjugate vaccines required lower levels of NKT cell activation to be effective, which could be a safety advantage for future vaccine candidates.

4.
Allergy ; 76(10): 3155-3170, 2021 10.
Artículo en Inglés | MEDLINE | ID: mdl-34185885

RESUMEN

BACKGROUND: Mucosal-associated invariant T (MAIT) cells are unconventional T cells which recognize microbial metabolites presented by the major histocompatibility complex class I-related molecule MR1. Although MAIT cells have been shown to reside in human and murine skin, their contribution to atopic dermatitis (AD), an inflammatory skin disease associated with barrier dysfunction and microbial translocation, has not yet been determined. METHODS: Genetic deletion of MR1 and topical treatment with inhibitory MR1 ligands, which result in the absence and functional inhibition of MAIT cells, respectively, were used to investigate the role of MR1-dependent immune surveillance in a MC903-driven murine model of AD. RESULTS: The absence or inhibition of MR1 arrested AD disease progression through the blockade of both eosinophil activation and recruitment of IL-4- and IL-13-producing cells. In addition, the therapeutic efficacy of phototherapy against MC903-driven AD could be increased with prior application of folate, which photodegrades into the inhibitory MR1 ligand 6-formylpterin. CONCLUSION: We identified MAIT cells as sentinels and mediators of cutaneous type 2 immunity. Their pathogenic activity can be inhibited by topical application or endogenous generation, via phototherapy, of inhibitory MR1 ligands.


Asunto(s)
Dermatitis Atópica , Antígenos de Histocompatibilidad Clase I , Antígenos de Histocompatibilidad Menor , Células T Invariantes Asociadas a Mucosa , Terapia Ultravioleta , Animales , Dermatitis Atópica/terapia , Modelos Animales de Enfermedad , Ratones
5.
Cytometry A ; 97(8): 824-831, 2020 08.
Artículo en Inglés | MEDLINE | ID: mdl-32293794

RESUMEN

The arrival of mass cytometry (MC) and, more recently, spectral flow cytometry (SFC) has revolutionized the study of cellular, functional and phenotypic diversity, significantly increasing the number of characteristics measurable at the single-cell level. As a consequence, new computational techniques such as dimensionality reduction and/or clustering algorithms are necessary to analyze, clean, visualize, and interpret these high-dimensional data sets. In this small comparison study, we investigated splenocytes from the same sample by either MC or SFC and compared both high-dimensional data sets using expert gating, t-distributed stochastic neighbor embedding (t-SNE), uniform manifold approximation and projection (UMAP) analysis and FlowSOM. When we downsampled each data set to their equivalent cell numbers and parameters, our analysis yielded highly comparable results. Differences between the data sets only became apparent when the maximum number of parameters in each data set were assessed, due to differences in the number of recorded events or the maximum number of assessed parameters. Overall, our small comparison study suggests that mass cytometry and spectral flow cytometry both yield comparable results when analyzed manually or by high-dimensional clustering or dimensionality reduction algorithms such as t-SNE, UMAP, or FlowSOM. However, large scale studies combined with an in-depth technical analysis will be needed to assess differences between these technologies in more detail. © 2020 International Society for Advancement of Cytometry.


Asunto(s)
Algoritmos , Análisis de Datos , Análisis por Conglomerados , Citometría de Flujo
6.
Org Biomol Chem ; 17(5): 1225-1237, 2019 01 31.
Artículo en Inglés | MEDLINE | ID: mdl-30656346

RESUMEN

Activated NKT cells can stimulate antigen-presenting cells leading to enhanced peptide antigen-specific immunity. However, administration of potent NKT cell agonists like α-galactosylceramide (α-GalCer) can be associated with release of high levels of cytokines, and in some situations, hepatotoxicity. Here we show that it is possible to provoke sufficient NKT cell activity to stimulate strong antigen-specific T cell responses without these unwanted effects. This was achieved by chemically conjugating antigenic peptides to α-galactosylphytosphingosine (α-GalPhs), an NKT cell agonist with very weak activity based on structural characterisation and biological assays. Conjugation improved delivery to antigen-presenting cells in vivo, while use of a cathepsin-sensitive linker to release the α-GalPhs and peptide within the same cell promoted strong T cell activation and therapeutic anti-tumour responses in mice. The conjugates activated human NKT cells and enhanced human T cell responses to a viral peptide in vitro. Accordingly, we have demonstrated a means to safely exploit the immunostimulatory properties of NKT cells to enhance T cell activation for virus- and tumour-specific immunity.


Asunto(s)
Células Presentadoras de Antígenos/inmunología , Vacunas contra el Cáncer/administración & dosificación , Células T Asesinas Naturales/efectos de los fármacos , Células T Asesinas Naturales/inmunología , Neoplasias Experimentales/inmunología , Péptidos/administración & dosificación , Adyuvantes Inmunológicos , Animales , Antígenos CD1d/química , Vacunas contra el Cáncer/inmunología , Enfermedad Hepática Inducida por Sustancias y Drogas/prevención & control , Epítopos/química , Glucolípidos/química , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Neoplasias Experimentales/tratamiento farmacológico , Péptidos/química , Péptidos/inmunología
7.
J Immunol ; 199(8): 2631-2638, 2017 10 15.
Artículo en Inglés | MEDLINE | ID: mdl-28877992

RESUMEN

Mucosal-associated invariant T (MAIT) cells are innate T cells that recognize intermediates of the vitamin B2 biosynthetic pathway presented by the monomorphic MR1 molecule. It remains unclear whether, in addition to their cytolytic activity that is important in antimicrobial defense, MAIT cells have immune-modulatory functions that could enhance dendritic cell (DC) maturation. In this study, we investigated the molecular mechanisms dictating the interactions between human MAIT cells and DCs and demonstrate that human MAIT cells mature monocyte-derived and primary DCs in an MR1- and CD40L-dependent manner. Furthermore, we show that MAIT cell-derived signals synergize with microbial stimuli to induce secretion of bioactive IL-12 by DCs. Activation of human MAIT cells in whole blood leads to MR1- and cytokine-dependent NK cell transactivation. Our results underscore an important property of MAIT cells, which can be of translational relevance to rapidly orchestrate adaptive immunity through DC maturation.


Asunto(s)
Células Dendríticas/inmunología , Activación de Linfocitos , Células T Asesinas Naturales/inmunología , Ligando de CD40/metabolismo , Comunicación Celular , Diferenciación Celular , Células Cultivadas , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Inmunidad Mucosa , Interleucina-12/metabolismo , Antígenos de Histocompatibilidad Menor/metabolismo , Monocitos/inmunología , Receptor Cross-Talk , Riboflavina/inmunología , Riboflavina/metabolismo , Transducción de Señal
8.
Cancer Immunol Immunother ; 67(2): 285-298, 2018 02.
Artículo en Inglés | MEDLINE | ID: mdl-29094183

RESUMEN

Vaccines that elicit targeted tumor antigen-specific T-cell responses have the potential to be used as adjuvant therapy in patients with high risk of relapse. However, the responses induced by vaccines in cancer patients have generally been disappointing. To improve vaccine function, we investigated the possibility of exploiting the immunostimulatory capacity of type 1 Natural killer T (NKT) cells, a cell type enriched in lymphoid tissues that can trigger improved antigen-presenting function in dendritic cells (DCs). In this phase I dose escalation study, we treated eight patients with high-risk surgically resected stage II-IV melanoma with intravenous autologous monocyte-derived DCs loaded with the NKT cell agonist α-GalCer and peptides derived from the cancer testis antigen NY-ESO-1. Two synthetic long peptides spanning defined immunogenic regions of the NY-ESO-1 sequence were used. This therapy proved to be safe and immunologically effective, inducing increases in circulating NY-ESO-1-specific T cells that could be detected directly ex vivo in seven out of eight patients. These responses were achieved using as few as 5 × 105 peptide-loaded cells per dose. Analysis after in vitro restimulation showed increases in polyfunctional CD4+ and CD8+ T cells that were capable of manufacturing two or more cytokines simultaneously. Evidence of NKT cell proliferation and/or NKT cell-associated cytokine secretion was seen in most patients. In light of these strong responses, the concept of including NKT cell agonists in vaccine design requires further investigation.


Asunto(s)
Antígenos de Neoplasias/genética , Células Dendríticas/inmunología , Galactosilceramidas/inmunología , Melanoma/inmunología , Proteínas de la Membrana/genética , Antígenos de Neoplasias/metabolismo , Humanos , Proteínas de la Membrana/metabolismo
9.
J Immunol ; 195(3): 821-31, 2015 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-26078270

RESUMEN

The function of dendritic cells (DCs) can be modulated through multiple signals, including recognition of pathogen-associated molecular patterns, as well as signals provided by rapidly activated leukocytes in the local environment, such as innate-like T cells. In this article, we addressed the possibility that the roles of different murine DC subsets in cross-priming CD8(+) T cells can change with the nature and timing of activatory stimuli. We show that CD8α(+) DCs play a critical role in cross-priming CD8(+) T cell responses to circulating proteins that enter the spleen in close temporal association with ligands for TLRs and/or compounds that activate NKT cells. However, if NKT cells are activated first, then CD8α(-) DCs become conditioned to respond more vigorously to TLR ligation, and if triggered directly, these cells can also contribute to priming of CD8(+) T cell responses. In fact, the initial activation of NKT cells can condition multiple DC subsets to respond more effectively to TLR ligation, with plasmacytoid DCs making more IFN-α and both CD8α(+) and CD8α(-) DCs manufacturing more IL-12. These results suggest that different DC subsets can contribute to T cell priming if provided appropriately phased activatory stimuli, an observation that could be factored into the design of more effective vaccines.


Asunto(s)
Linfocitos T CD8-positivos/inmunología , Reactividad Cruzada/inmunología , Células Dendríticas/inmunología , Activación de Linfocitos/inmunología , Células T Asesinas Naturales/inmunología , Animales , Presentación de Antígeno/inmunología , Antígenos de Superficie/genética , Interferón-alfa/biosíntesis , Interferón-alfa/inmunología , Interleucina-12/biosíntesis , Lectinas Tipo C/genética , Lectinas de Unión a Manosa/genética , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Bazo/inmunología , Receptores Toll-Like/inmunología
10.
BMC Pulm Med ; 17(1): 169, 2017 Dec 04.
Artículo en Inglés | MEDLINE | ID: mdl-29202821

RESUMEN

BACKGROUND: Asthma inflammatory phenotypes are often defined by relative cell counts of airway eosinophils/neutrophils. However, the importance of neutrophilia remains unclear, as does the effect of ICS treatment on asthma phenotypes and airway neutrophil function. The purpose of this study was to assess asthma phenotype prevalence/characteristics in a community setting, and, in a nested preliminary study, determine how treatment changes affect phenotype stability and inflammation, with particular focus on airway neutrophils. METHODS: Fifty adult asthmatics and 39 non-asthmatics were assessed using questionnaires, skin prick tests, spirometry, exhaled nitric oxide (FENO) measurement, and sputum induction. Twenty-one asthmatics underwent further assessment following treatment optimisation (n = 11) or sub-optimisation (n = 10). RESULTS: Forty percent (20/50) had eosinophilic asthma (EA) and 8% had neutrophilic asthma. EA was associated with increased FENO, bronchodilator reversibility (BDR) and reduced lung function (p < 0.05). Following optimisation/sub-optimisation, the EA/NEA (non-eosinophilic asthma) phenotype changed in 11/21 (52%) asthmatics. In particular, fewer subjects had EA post treatment optimisation, but this was not statistically significant. However, a significant (p < 0.05) reduction in FENO, ACQ7 score, and BDR was observed after treatment optimisation, as well as an increase in FEV1-% predicted (p < 0.05). It was also associated with reduced eosinophils (p < 0.05) and enhanced neutrophil phagocytosis (p < 0.05) in EA only, and enhanced neutrophil oxidative burst in both EA and NEA (p < 0.05). CONCLUSIONS: In this community based population, non-eosinophilic asthma was common, less severe than EA, and at baseline most asthmatics showed no evidence of inflammation. In the nested change in treatment study, treatment optimisation was associated with reduced sputum eosinophils, improved symptoms and lung function, and enhanced neutrophil function, but a significant reduction in EA could not be demonstrated. TRIAL REGISTRATION: The nested change in treatment component of this study is registered at the Australia and New Zealand Clinical Trial Registry ( www.ANZCTR.org.au ) ACTRN12617001356358 . Registration date 27/09/2017. Retrospectively registered.


Asunto(s)
Corticoesteroides/uso terapéutico , Asma/tratamiento farmacológico , Broncodilatadores/uso terapéutico , Eosinofilia/tratamiento farmacológico , Neutrófilos/fisiología , Fenotipo , Administración por Inhalación , Adolescente , Corticoesteroides/administración & dosificación , Adulto , Anciano , Asma/fisiopatología , Pruebas Respiratorias , Femenino , Volumen Espiratorio Forzado , Humanos , Inflamación/fisiopatología , Masculino , Persona de Mediana Edad , Óxido Nítrico/análisis , Ápice del Flujo Espiratorio , Fagocitosis , Estallido Respiratorio , Esputo/citología , Encuestas y Cuestionarios , Adulto Joven
11.
Magn Reson Chem ; 55(5): 498-507, 2017 May.
Artículo en Inglés | MEDLINE | ID: mdl-27487091

RESUMEN

Complex materials are ubiquitous in science, engineering and nature. One important parameter for characterising their morphology is the degree of anisotropy. Magnetic resonance imaging offers non-invasive methods for quantitative measurements of the materials anisotropy, most commonly via diffusion tensor imaging and the subsequent extraction of the spatially resolved fractional anisotropy (FA) value. Here, we propose an alternative way of determining the FA as a sample average for cases where spatially resolved methods are not needed or not applicable. It is based on a particular diffusion-diffusion correlation spectroscopy protocol, allowing for the extraction of the mean (i.e. sample averaged) FA value. We demonstrate that mean FA values obtained from three anisotropic biological tissues are consistent with those extracted using diffusion tensor imaging. Moreover, we show that differences of mean FA values in healthy and tumour-bearing mouse brains allow to distinguish these tissue types. We anticipate that the proposed method will be beneficial in the wider context of medical and material science. Copyright © 2016 John Wiley & Sons, Ltd.


Asunto(s)
Espectroscopía de Resonancia Magnética/métodos , Animales , Anisotropía , Encéfalo/diagnóstico por imagen , Neoplasias Encefálicas/diagnóstico por imagen , Línea Celular Tumoral , Simulación por Computador , Daucus carota , Difusión , Imagen de Difusión Tensora , Humanos , Ratones
12.
Blood ; 124(19): 2953-63, 2014 Nov 06.
Artículo en Inglés | MEDLINE | ID: mdl-25237205

RESUMEN

Acute leukemias with adverse prognostic features carry a high relapse rate without allogeneic stem cell transplantation (allo-SCT). Allo-SCT has a high morbidity and is precluded for many patients because of advanced age or comorbidities. Postremission therapies with reduced toxicities are urgently needed. The murine acute leukemia model C1498 was used to study the efficacy of an intravenously administered vaccine consisting of irradiated leukemia cells loaded with the natural killer T (NKT)-cell agonist α-galactosylceramide (α-GalCer). Prophylactically, the vaccine was highly effective at preventing leukemia development through the downstream activities of activated NKT cells, which were dependent on splenic langerin(+)CD8α(+) dendritic cells and which led to stimulation of antileukemia CD4(+) and CD8(+) T cells. However, hosts with established leukemia received no protective benefit from the vaccine, despite inducing NKT-cell activation. Established leukemia was associated with increases in regulatory T cells and myeloid-derived suppressor cells, and the leukemic cells themselves were highly suppressive in vitro. Although this suppressive environment impaired both effector arms of the immune response, CD4(+) T-cell responses were more severely affected. When cytarabine chemotherapy was administered prior to vaccination, all animals in remission posttherapy were protected against rechallenge with viable leukemia cells.


Asunto(s)
Vacunas contra el Cáncer/farmacología , Citarabina/farmacología , Galactosilceramidas/inmunología , Células Asesinas Naturales/trasplante , Leucemia Mieloide/tratamiento farmacológico , Leucemia Mieloide/prevención & control , Enfermedad Aguda , Animales , Antimetabolitos Antineoplásicos/farmacología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Terapia Combinada , Células Dendríticas/inmunología , Proteínas Fluorescentes Verdes/genética , Células Asesinas Naturales/efectos de la radiación , Leucemia Mieloide/inmunología , Ratones Endogámicos C57BL , Ratones Transgénicos , Pronóstico , Prevención Secundaria/métodos , Trasplante Autólogo
13.
Nat Chem Biol ; 10(11): 943-9, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25282504

RESUMEN

Epitope-based peptide vaccines encompass minimal immunogenic regions of protein antigens to allow stimulation of precisely targeted adaptive immune responses. However, because efficacy is largely determined by the functional status of antigen-presenting cells (APCs) that acquire and present peptides to cells of the adaptive immune system, adjuvant compounds are needed to enhance immunogenicity. We present here a vaccine consisting of an allergen-derived peptide conjugated to a prodrug of the natural killer-like T (NKT) cell agonist α-galactosylceramide, which is highly effective in reducing inflammation in a mouse model of allergic airway inflammation. Unlike other peptide-adjuvant conjugates that directly activate APCs through pattern recognition pathways, this vaccine encourages third-party interactions with NKT cells to enhance APC function. Therapeutic efficacy was correlated with marked increases in the number and functional activity of allergen-specific cytotoxic T lymphocytes (CTLs), leading to suppression of immune infiltration into the lungs after allergen challenge in sensitized hosts.


Asunto(s)
Adyuvantes Inmunológicos , Hipersensibilidad/inmunología , Profármacos/química , Linfocitos T Citotóxicos/inmunología , Vacunas/inmunología , Alérgenos/administración & dosificación , Alérgenos/química , Alérgenos/inmunología , Animales , Antiinflamatorios/metabolismo , Antiinflamatorios/farmacología , Antiinflamatorios/uso terapéutico , Células Presentadoras de Antígenos/efectos de los fármacos , Células Presentadoras de Antígenos/inmunología , Modelos Animales de Enfermedad , Femenino , Galactosilceramidas/metabolismo , Galactosilceramidas/farmacología , Galactosilceramidas/uso terapéutico , Hipersensibilidad/tratamiento farmacológico , Inmunoglobulina E/sangre , Inflamación/tratamiento farmacológico , Masculino , Ratones , Ratones Endogámicos C57BL , Conformación Molecular , Células T Asesinas Naturales/citología , Células T Asesinas Naturales/efectos de los fármacos , Células T Asesinas Naturales/inmunología , Péptidos/administración & dosificación , Péptidos/química , Péptidos/inmunología , Profármacos/metabolismo , Linfocitos T Citotóxicos/efectos de los fármacos , Vacunas/administración & dosificación , Vacunas/síntesis química , Vacunas/química
14.
Int J Cancer ; 136(11): 2566-78, 2015 Jun 01.
Artículo en Inglés | MEDLINE | ID: mdl-25363661

RESUMEN

Glioblastoma multiforme (GBM) is a highly malignant brain tumor with an extremely short time to relapse following standard treatment. Since recurrent GBM is often resistant to subsequent radiotherapy and chemotherapy, immunotherapy has been proposed as an alternative treatment option. Although it is well established that GBM induces immune suppression, it is currently unclear what impact prior conventional therapy has on the ability of GBM cells to modulate the immune environment. In this study, we investigated the interaction between immune cells and glioma cells that had been exposed to chemotherapy or irradiation in vitro. We demonstrate that treated glioma cells are more immunosuppressive than untreated cells and form tumors at a faster rate in vivo in an animal model. Cultured supernatant from in vitro-treated primary human GBM cells were also shown to increase suppression, which was independent of accessory suppressor cells or T regulatory cell generation, and could act directly on CD4(+) and CD8(+) T cell proliferation. While a number of key immunosuppressive cytokines were overexpressed in the treated cells, including IL-10, IL-6 and GM-CSF, suppression could be alleviated in a number of treated GBM lines by inhibition of prostaglandin E2. These results reveal for the first time that conventional therapies can alter immunosuppressive pathways in GBM tumor cells, a finding with important implications for the combination of immunotherapy with standard treatment.


Asunto(s)
Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/patología , Citocinas/metabolismo , Glioblastoma/inmunología , Glioblastoma/patología , Animales , Neoplasias Encefálicas/terapia , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Línea Celular Tumoral , Proliferación Celular , Medios de Cultivo Condicionados/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de la radiación , Glioblastoma/terapia , Humanos , Terapia de Inmunosupresión , Ratones , Ratones Endogámicos C57BL , Trasplante de Neoplasias
15.
J Neurooncol ; 121(2): 319-29, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25366363

RESUMEN

There is no standard treatment for recurrent glioblastoma multiforme (GBM). Retreatment with temozolomide (TMZ) is one treatment option. We reasoned this could be more effective if combined with a vaccine that preferentially targeted TMZ-resistant cells. To test the feasibility and safety of such an approach, a phase 1 trial was conducted in which patients with GBM tumors relapsing after standard chemoradiotherapy were retreated with TMZ in combination with a vaccine consisting of monocyte-derived dendritic cells (DC) pulsed with autologous tumor cells that had previously been exposed to TMZ in vivo in the course of primary treatment. Of 14 participants, nine patients completed the initial phase of priming vaccinations and two cycles of TMZ, one proved to have radionecrosis, one rapidly progressed, and in three the yield of DC vaccine was insufficient to proceed with treatment. Other than expected toxicities related to TMZ, there were no adverse events attributable to the combined treatment. Two patients had objective radiological responses. Six month progression-free survival was 22 %, similar to retreatment with TMZ alone. Anti-tumor immune responses were assessed in peripheral blood mononuclear cells using interferon-γ ELISpot, with two patients meeting criteria for a vaccine-induced immune response, one of whom remained disease-free for nearly three years. Another patient with an anti-tumor immune response at baseline that was sustained post-vaccination experienced a 12-month period of progression-free survival. In summary, the combined treatment was safe and well-tolerated but feasibility in the recurrent setting was marginal. Evidence of immune responses in a few patients broadly correlated with better clinical outcome.


Asunto(s)
Antineoplásicos Alquilantes/uso terapéutico , Neoplasias Encefálicas/terapia , Vacunas contra el Cáncer/uso terapéutico , Dacarbazina/análogos & derivados , Glioblastoma/terapia , Recurrencia Local de Neoplasia/terapia , Adulto , Anciano , Encéfalo/patología , Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/patología , Vacunas contra el Cáncer/efectos adversos , Terapia Combinada/efectos adversos , Dacarbazina/efectos adversos , Dacarbazina/uso terapéutico , Células Dendríticas/inmunología , Supervivencia sin Enfermedad , Estudios de Factibilidad , Femenino , Glioblastoma/inmunología , Glioblastoma/patología , Humanos , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/inmunología , Recurrencia Local de Neoplasia/patología , Retratamiento , Temozolomida , Resultado del Tratamiento
16.
Crit Rev Oncog ; 29(1): 31-43, 2024.
Artículo en Inglés | MEDLINE | ID: mdl-38421712

RESUMEN

The capacity of type I natural killer T (NKT) cells to provide stimulatory signals to antigen-presenting cells has prompted preclinical research into the use of agonists as immune adjuvants, with much of this work focussed on stimulating T cell responses to cancer. In attempting to evaluate this approach in the clinic, our recent dendritic-cell based study failed to show an advantage to adding an agonist to the vaccine. Here we present potential limitations of the study, and suggest why other simpler strategies may be more effective. These include strategies to target antigen-presenting cells in the host, either through promoting efficient transfer from injected cell lines, facilitating uptake of antigen and agonist as injected conjugates, or encapsulating the components into injected nanovectors. While the vaccine landscape has changed with the rapid uptake of mRNA vaccines, we suggest that there is still a role for recruiting NKT cells in altering T cell differentiation programmes, notably the induction of resident memory T cells.


Asunto(s)
Células T Asesinas Naturales , Vacunas , Humanos , Vacunación , Diferenciación Celular
17.
ACS Chem Biol ; 19(6): 1366-1375, 2024 Jun 21.
Artículo en Inglés | MEDLINE | ID: mdl-38829263

RESUMEN

Eliciting an antihapten antibody response to vaccination typically requires the use of constructs where multiple copies of the hapten are covalently attached to a larger carrier molecule. The carrier is required to elicit T cell help via presentation of peptide epitopes on major histocompatibility complex (MHC) class II molecules; as such, attachment to full-sized proteins, alone or in a complex, is generally used to account for the significant MHC diversity in humans. While such carrier-based vaccines have proven extremely successful, particularly in protecting against bacterial diseases, they can be challenging to manufacture, and repeated use can be compromised by pre-existing immunity against the carrier. One approach to reducing these complications is to recruit help from type I natural killer T (NKT) cells, which exhibit limited diversity in their antigen receptors and respond to glycolipid antigens presented by the highly conserved presenting molecule CD1d. Synthetic vaccines for universal use can, therefore, be prepared by conjugating haptens to an NKT cell agonist such as α-galactosylceramide (αGalCer, KRN7000). An additional advantage is that the quality of NKT cell help is sufficient to overcome the need for an extra immune adjuvant. However, while initial studies with αGalCer-hapten conjugate vaccines report strong and rapid antihapten antibody responses, they can fail to generate lasting memory. Here, we show that antibody responses to the hapten 4-hydoxy-3-nitrophenyl acetyl (NP) can be improved through additional attachment of a fusion peptide containing a promiscuous helper T cell epitope (Pan DR epitope, PADRE) that binds diverse MHC class II molecules. Such αGalCer-hapten-peptide tricomponent vaccines generate strong and sustained anti-NP antibody titers with increased hapten affinity compared to vaccines without the helper epitope. The tricomponent vaccine platform is therefore suitable for further exploration in the pursuit of efficacious antihapten immunotherapies.


Asunto(s)
Haptenos , Vacunas Conjugadas , Animales , Haptenos/inmunología , Haptenos/química , Ratones , Vacunas Conjugadas/inmunología , Péptidos/inmunología , Péptidos/química , Formación de Anticuerpos/inmunología , Ratones Endogámicos C57BL , Galactosilceramidas/inmunología , Galactosilceramidas/química , Femenino , Células T Asesinas Naturales/inmunología , Glucolípidos/inmunología , Glucolípidos/química
18.
J Immunother Cancer ; 12(4)2024 Apr 17.
Artículo en Inglés | MEDLINE | ID: mdl-38631706

RESUMEN

BACKGROUND: Tumor-targeted therapy causes impressive tumor regression, but the emergence of resistance limits long-term survival benefits in patients. Little information is available on the role of the myeloid cell network, especially dendritic cells (DC) during tumor-targeted therapy. METHODS: Here, we investigated therapy-mediated immunological alterations in the tumor microenvironment (TME) and tumor-draining lymph nodes (LN) in the D4M.3A preclinical melanoma mouse model (harboring the V-Raf murine sarcoma viral oncogene homolog B (BRAF)V600E mutation) by using high-dimensional multicolor flow cytometry in combination with multiplex immunohistochemistry. This was complemented with RNA sequencing and cytokine quantification to characterize the immune status of the tumors. The importance of T cells during tumor-targeted therapy was investigated by depleting CD4+ or CD8+ T cells in tumor-bearing mice. Tumor antigen-specific T-cell responses were characterized by performing in vivo T-cell proliferation assays and the contribution of conventional type 1 DC (cDC1) to T-cell immunity during tumor-targeted therapy was assessed using Batf3-/- mice lacking cDC1. RESULTS: Our findings reveal that BRAF-inhibitor therapy increased tumor immunogenicity, reflected by an upregulation of genes associated with immune activation. The T cell-inflamed TME contained higher numbers of activated cDC1 and cDC2 but also inflammatory CCR2-expressing monocytes. At the same time, tumor-targeted therapy enhanced the frequency of migratory, activated DC subsets in tumor-draining LN. Even more, we identified a cDC2 population expressing the Fc gamma receptor I (FcγRI)/CD64 in tumors and LN that displayed high levels of CD40 and CCR7 indicating involvement in T cell-mediated tumor immunity. The importance of cDC2 is underlined by just a partial loss of therapy response in a cDC1-deficient mouse model. Both CD4+ and CD8+ T cells were essential for therapy response as their respective depletion impaired therapy success. On resistance development, the tumors reverted to an immunologically inert state with a loss of DC and inflammatory monocytes together with the accumulation of regulatory T cells. Moreover, tumor antigen-specific CD8+ T cells were compromised in proliferation and interferon-γ-production. CONCLUSION: Our results give novel insights into the remodeling of the myeloid landscape by tumor-targeted therapy. We demonstrate that the transient immunogenic tumor milieu contains more activated DC. This knowledge has important implications for the development of future combinatorial therapies.


Asunto(s)
Melanoma , Humanos , Animales , Ratones , Melanoma/metabolismo , Linfocitos T CD8-positivos , Proteínas Proto-Oncogénicas B-raf/genética , Células Dendríticas , Antígenos de Neoplasias , Microambiente Tumoral
19.
JHEP Rep ; 6(5): 101038, 2024 May.
Artículo en Inglés | MEDLINE | ID: mdl-38694959

RESUMEN

Background & Aims: Liver diseases resulting from chronic HBV infection are a significant cause of morbidity and mortality. Vaccines that elicit T-cell responses capable of controlling the virus represent a treatment strategy with potential for long-term effects. Here, we evaluated vaccines that induce the activity of type I natural killer T (NKT) cells to limit viral replication and license stimulation of conventional antiviral T-cells. Methods: Vaccines were prepared by conjugating peptide epitopes to an NKT-cell agonist to promote co-delivery to antigen-presenting cells, encouraging NKT-cell licensing and stimulation of T cells. Activity of the conjugate vaccines was assessed in transgenic mice expressing the complete HBV genome, administered intravenously to maximise access to NKT cell-rich tissues. Results: The vaccines induced only limited antiviral activity in unmanipulated transgenic hosts, likely attributable to NKT-cell activation as T-cell tolerance to viral antigens is strong. However, in a model of chronic hepatitis B involving transfer of naive HBcAg-specific CD8+ T cells into the transgenic mice, which typically results in specific T-cell dysfunction without virus control, vaccines containing the targeted HBcAg epitope induced prolonged antiviral activity because of qualitatively improved T-cell stimulation. In a step towards a clinical product, vaccines were prepared using synthetic long peptides covering clusters of known HLA-binding epitopes and shown to be immunogenic in HLA transgenic mice. Predictions based on HLA distribution suggest a product containing three selected SLP-based vaccines could give >90 % worldwide coverage, with an average of 3.38 epitopes targeted per individual. Conclusions: The novel vaccines described show promise for further clinical development as a treatment for chronic hepatitis B. Impact and Implications: Although there are effective prophylactic vaccines for HBV infection, it is estimated that 350-400 million people worldwide have chronic hepatitis B, putting these individuals at significant risk of life-threatening liver diseases. Therapeutic vaccination aimed at activating or boosting HBV-specific T-cell responses holds potential as a strategy for treating chronic infection, but has so far met with limited success. Here, we show that a glycolipid-peptide conjugate vaccine designed to coordinate activity of type I NKT cells alongside conventional antiviral T cells has antiviral activity in a mouse model of chronic infection. It is anticipated that a product based on a combination of three such conjugates, each prepared using long peptides covering clusters of known HLA-binding epitopes, could be developed further as a treatment for chronic hepatitis B with broad global HLA coverage.

20.
Haematologica ; 98(3): 376-84, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23065503

RESUMEN

Invariant natural killer T cells recognize glycolipid antigens such as α-galactosylceramide presented by CD1d. In preclinical models of B-cell malignancies, α-galactosylceramide is an adjuvant to tumor vaccination, enhancing tumor-specific T-cell responses and prolonging survival. However, numerical and functional invariant natural killer T-cell defects exist in patients with some cancers. Our aim was to assess this axis in patients with chronic lymphocytic leukemia. The numbers of circulating invariant natural killer T cells and the expression of CD1d on antigen-presenting cells were evaluated in patients with chronic lymphocytic leukemia and age-matched controls. Cytokine profile and in vitro proliferative capacity were determined. Patient- and control-derived invariant natural killer T-cell lines were generated and characterized, and allogeneic and autologous responses to α-galactosylce-ramide-treated leukemia cells were assessed. Absolute numbers and phenotype of invariant natural killer T cells were normal in patients with untreated chronic lymphocytic leukemia, and cytokine profile and proliferative capacity were intact. Chemotherapy-treated patients had reduced numbers of invariant natural killer T cells and myeloid dendritic cells, but α-galactosylceramide-induced proliferation was preserved. Invariant natural killer T-cell lines from patients lysed CD1d-expressing targets. Irradiated α-galactosylceramide-treated leukemic cells elicited allogeneic and autologous invariant natural killer T-cell proliferation, and α-galactosylceramide treatment led to increased proliferation of conventional T cells in response to tumor. In conclusion, the invariant natural killer T-cell and CD1d axis is fundamentally intact in patients with early-stage chronic lymphocytic leukemia and, despite reduced circulating numbers, function is retained in fludarabine-treated patients. Immunotherapies exploiting the adjuvant effect of α-galactosylceramide may be feasible.


Asunto(s)
Antígenos CD1d/metabolismo , Leucemia Linfocítica Crónica de Células B/inmunología , Leucemia Linfocítica Crónica de Células B/metabolismo , Células T Asesinas Naturales/inmunología , Células T Asesinas Naturales/metabolismo , Línea Celular , Proliferación Celular , Citocinas/biosíntesis , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Galactosilceramidas/farmacología , Humanos , Inmunofenotipificación , Inmunoterapia , Leucemia Linfocítica Crónica de Células B/terapia , Células Mieloides/inmunología , Células Mieloides/metabolismo , Células T Asesinas Naturales/efectos de los fármacos , Fenotipo
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