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Anticuerpos Monoclonales Humanizados/uso terapéutico , Interferón gamma/antagonistas & inhibidores , Psoriasis/tratamiento farmacológico , Anticuerpos Monoclonales Humanizados/administración & dosificación , Anticuerpos Monoclonales Humanizados/efectos adversos , Humanos , Resultado del TratamientoRESUMEN
BACKGROUND: Alefacept treatment is highly effective in a select group patients with moderate-to-severe psoriasis, and is an ideal candidate to develop systems to predict who will respond to therapy. A clinical trial of 22 patients with moderate to severe psoriasis treated with alefacept was conducted in 2002-2003, as a mechanism of action study. Patients were classified as responders or non-responders to alefacept based on histological criteria. Results of the original mechanism of action study have been published. Peripheral blood was collected at the start of this clinical trial, and a prior analysis demonstrated that gene expression in PBMCs differed between responders and non-responders, however, the analysis performed could not be used to predict response. METHODS: Microarray data from PBMCs of 16 of these patients was analyzed to generate a treatment response classifier. We used a discriminant analysis method that performs sample classification from gene expression data, via "nearest shrunken centroid method". Centroids are the average gene expression for each gene in each class divided by the within-class standard deviation for that gene. RESULTS: A disease response classifier using 23 genes was created to accurately predict response to alefacept (12.3% error rate). While the genes in this classifier should be considered as a group, some of the individual genes are of great interest, for example, cAMP response element modulator (CREM), v-MAF avian musculoaponeurotic fibrosarcoma oncogene family (MAFF), chloride intracellular channel protein 1 (CLIC1, also called NCC27), NLR family, pyrin domain-containing 1 (NLRP1), and CCL5 (chemokine, cc motif, ligand 5, also called regulated upon activation, normally T expressed, and presumably secreted/RANTES). CONCLUSIONS: Although this study is small, and based on analysis of existing microarray data, we demonstrate that a treatment response classifier for alefacept can be created using gene expression of PBMCs in psoriasis. This preliminary study may provide a useful tool to predict response of psoriatic patients to alefacept.
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Fármacos Dermatológicos/uso terapéutico , Genómica , Medicina de Precisión/métodos , Psoriasis/tratamiento farmacológico , Psoriasis/genética , Proteínas Recombinantes de Fusión/uso terapéutico , Adulto , Anciano , Alefacept , Resistencia a Medicamentos/genética , Femenino , Genoma Humano , Humanos , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Valor Predictivo de las Pruebas , Psoriasis/patología , Índice de Severidad de la EnfermedadRESUMEN
Using high-density oligonucleotide arrays, we measured expression of >12,000 genes in surgical excisions of invasive human squamous cell carcinomas (SCCs) versus site-matched control skin. This analysis defined >1,900 genes with altered expression in SCCs that were statistically different from controls. As SCCs are composed of epithelial cells, which are both hyperplastic and invasive, we sought to define gene sets associated with these biologic processes by comparing gene expression to psoriasis vulgaris, which is a condition of benign keratinocyte hyperplasia without invasiveness or pre-malignant potential. Through this analysis, we found genes that were commonly upregulated in both conditions and unique genes with increased expression in SCCs. Differential gene regulation in these two conditions was confirmed by real-time reverse transcription-PCR and immunohistochemistry. We found that benign hyperplasia is associated with upregulation of genes including DEFB4 (defensin B4), SERPINB3 (serine proteinase inhibitor, member 3), STAT1 (signal transducer and activator of transcription 1), K16 (keratin 16), CEACAMs (carcinoembryonic antigen-related cell adhesion molecules), and WNT 5A (wingless-type MMTV integration site family, member 5A). WNT receptor frizzled homolog 6 (FZD6) and prostaglandin-metabolizing enzyme hydroxyprostaglandin dehydrogenase were increased in SCC alone. Growth factor pleiotrophin (PTN) was expressed at higher levels in non-tumor-bearing skin adjacent to excised SCC. SCC was further characterized by upregulation of matrix metalloproteinases 1, 10, and 13, cathepsin L2, cystatin E/M as well as STAT3 and microseminoprotein, beta (MSMB), and downregulation of inducible nitric oxide synthase, granzyme B, CD8, and CD83. The current study defines a unique gene expression signature for cutaneous SCC in humans and suggests potential roles for WNT, FZD, and PTN in the pathogenesis of SCC.
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Carcinoma de Células Escamosas/diagnóstico , Regulación Neoplásica de la Expresión Génica , Genes Relacionados con las Neoplasias , Neoplasias Cutáneas/diagnóstico , Proliferación Celular , Diagnóstico Diferencial , Genómica , Humanos , Hiperplasia/diagnóstico , Análisis de Secuencia por Matrices de OligonucleótidosRESUMEN
We assessed expression of IL-20 and its receptors in psoriasis, given the recent implication of IL-20 in epidermal hyperplasia. Psoriatic lesional (LS) skin consistently expressed more IL-20 mRNA than nonlesional (NL) skin. Immunoreactivity to IL-20 protein was greater in LS tissue and mainly localized to infiltrating CD68+/CD11c+ (myeloid-derived) dermal leukocytes. Because this contrasted with earlier reports of a keratinocyte source, we assessed IL-20 mRNA expression in a variety of cells in vitro, and confirmed a myeloid-derived cellular source (monocytes). Plastic adhesion, activation of beta2 integrins, and incubation with tumor necrosis factor-alpha stimulated expression in these cells. IL-20 receptor (IL-20R)alpha and IL-20Rbeta mRNA was decreased in LS versus NL skin, which also contrasted with earlier findings. To investigate the relationship between IL-20 and disease activity, we examined psoriasis patients treated with the CD2-targeted agent alefacept. In therapeutic responders, lesional IL-20 mRNA decreased to NL levels, suggesting that CD2+ leukocytes may proximally regulate IL-20. Finally, to assess IL-20 function, we used microarrays to screen IL-20-treated keratinocytes, which demonstrated upregulation of disease-related and IFN-gamma-induced genes. Hence, IL-20 may influence inflammation through IFN-like effects. Together, these data indicate that IL-20 may be an important effector cytokine in psoriasis, and that its inhibition may represent a potential therapeutic target.
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Antígenos CD/análisis , Antígenos de Diferenciación Mielomonocítica/análisis , Antígeno CD11c/análisis , Regulación de la Expresión Génica/fisiología , Interleucinas/metabolismo , Leucocitos/inmunología , Leucocitos/metabolismo , Psoriasis/genética , Psoriasis/metabolismo , Adulto , Alefacept , Antígenos CD2/análisis , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Inflamación/inmunología , Cadenas beta de Integrinas/fisiología , Interferón gamma/fisiología , Interleucinas/análisis , Interleucinas/genética , Interleucinas/farmacología , Queratinocitos/química , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Leucocitos/química , Leucocitos/efectos de los fármacos , Monocitos/química , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Psoriasis/tratamiento farmacológico , Psoriasis/fisiopatología , ARN Mensajero/análisis , ARN Mensajero/genética , Receptores de Interleucina/análisis , Receptores de Interleucina/genética , Receptores de Interleucina/fisiología , Proteínas Recombinantes de Fusión/farmacología , Proteínas Recombinantes de Fusión/uso terapéutico , Piel/química , Factor de Necrosis Tumoral alfa/farmacologíaRESUMEN
BACKGROUND: Microscopists are familiar with many blemishes that fluorescence images can have due to dust and debris, glass flaws, uneven distribution of fluids or surface coatings, etc. Microarray scans show similar artefacts, which affect the analysis, particularly when one tries to detect subtle changes. However, most blemishes are hard to find by the unaided eye, particularly in high-density oligonucleotide arrays (HDONAs). RESULTS: We present a method that harnesses the statistical power provided by having several HDONAs available, which are obtained under similar conditions except for the experimental factor. This method "harshlights" blemishes and renders them evident. We find empirically that about 25% of our chips are blemished, and we analyze the impact of masking them on screening for differentially expressed genes. CONCLUSION: Experiments attempting to assess subtle expression changes should be carefully screened for blemishes on the chips. The proposed method provides investigators with a novel robust approach to improve the sensitivity of microarray analyses. By utilizing topological information to identify and mask blemishes prior to model based analyses, the method prevents artefacts from confounding the process of background correction, normalization, and summarization.
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Biología Computacional/métodos , Análisis de Secuencia por Matrices de Oligonucleótidos/instrumentación , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Algoritmos , Artefactos , Simulación por Computador , Interpretación Estadística de Datos , Perfilación de la Expresión Génica , Humanos , Análisis por Micromatrices , Microscopía Fluorescente/métodos , Modelos Genéticos , Modelos Estadísticos , Análisis Multivariante , Hibridación de Ácido Nucleico , Análisis de Componente Principal , Psoriasis/genética , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
BACKGROUND: Cyclooxygenase (COX)-2 contributes to vascular smooth muscle cell (VSMC) proliferation induced by tumor necrosis factor (TNF) and angiotensin II. The present study demonstrates, however, that depending on prevailing conditions, COX-2-derived prostanoids may also inhibit VSMC proliferation. METHODS AND RESULTS: TNF-alpha stimulated proliferation of VSMCs by shortening the G1 phase of the cell cycle. This effect was abolished by NS-398, a selective COX-2 inhibitor. Addition of TNF did not affect the protein-to-DNA ratio, measured by flow cytometry, suggesting that TNF does not induce VSMC hypertrophy. Inhibition of nitric oxide synthase (NOS) activity attenuated TNF-mediated increases in prostaglandin (PG) I2 synthesis, whereas thromboxane (TX) A2 production and COX-2 protein expression were unaffected. Moreover, inhibition of NOS activity increased TNF-mediated proliferation by approximately 23%. Thus, NO preferentially stimulates PGI2 production, suggesting that production of NO by VSMCs challenged with TNF limits the ability of the cytokine to increase proliferation. NO donors increased COX-2 protein expression and PGI2 synthesis, had no effect on TXA2 production, and decreased cell numbers by 50%, indicating that expression of COX-2 per se might not be sufficient to support proliferation. The effects of NO donors were prevented when COX-2 activity was inhibited with NS-398. CONCLUSIONS: The COX-2-dependent proliferative response of VSMCs to TNF was modulated in an NO-dependent manner, and PGI2 derived from COX-2 might contribute to the antiproliferative effect of NO donors.
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Fase G1/fisiología , Isoenzimas/fisiología , Músculo Liso Vascular/metabolismo , Óxido Nítrico/fisiología , Prostaglandina-Endoperóxido Sintasas/fisiología , Factor de Necrosis Tumoral alfa/fisiología , Animales , Proteínas de Ciclo Celular/metabolismo , División Celular/efectos de los fármacos , División Celular/fisiología , Células Cultivadas , Ciclina D1/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa 2 , Inhibidores de la Ciclooxigenasa/farmacología , ADN/metabolismo , Inhibidores Enzimáticos/farmacología , Epoprostenol/metabolismo , Fase G1/efectos de los fármacos , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Masculino , Músculo Liso Vascular/citología , Músculo Liso Vascular/efectos de los fármacos , Óxido Nítrico/metabolismo , Donantes de Óxido Nítrico/farmacología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Prostaglandina-Endoperóxido Sintasas/metabolismo , Proteínas/metabolismo , Ratas , Ratas Sprague-Dawley , Tromboxano A2/metabolismo , Factor de Necrosis Tumoral alfa/farmacología , Proteínas Supresoras de Tumor/metabolismoRESUMEN
PURPOSE: Distinct molecular subgroups of medulloblastoma, including hedgehog (Hh) pathway-activated disease, have been reported. We identified and clinically validated a five-gene Hh signature assay that can be used to preselect patients with Hh pathway-activated medulloblastoma. EXPERIMENTAL DESIGN: Gene characteristics of the Hh medulloblastoma subgroup were identified through published bioinformatic analyses. Thirty-two genes shown to be differentially expressed in fresh-frozen and formalin-fixed paraffin-embedded tumor samples and reproducibly analyzed by RT-PCR were measured in matched samples. These data formed the basis for building a multi-gene logistic regression model derived through elastic net methods from which the five-gene Hh signature emerged after multiple iterations. On the basis of signature gene expression levels, the model computed a propensity score to determine Hh activation using a threshold set a priori. The association between Hh activation status and tumor response to the Hh pathway inhibitor sonidegib (LDE225) was analyzed. RESULTS: Five differentially expressed genes in medulloblastoma (GLI1, SPHK1, SHROOM2, PDLIM3, and OTX2) were found to associate with Hh pathway activation status. In an independent validation study, Hh activation status of 25 medulloblastoma samples showed 100% concordance between the five-gene signature and Affymetrix profiling. Further, in medulloblastoma samples from 50 patients treated with sonidegib, all 6 patients who responded were found to have Hh-activated tumors. Three patients with Hh-activated tumors had stable or progressive disease. No patients with Hh-nonactivated tumors responded. CONCLUSIONS: This five-gene Hh signature can robustly identify Hh-activated medulloblastoma and may be used to preselect patients who might benefit from sonidegib treatment.
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Neoplasias Cerebelosas/genética , Proteínas Hedgehog/antagonistas & inhibidores , Meduloblastoma/genética , Selección de Paciente , Transcriptoma , Adolescente , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Compuestos de Bifenilo/farmacología , Compuestos de Bifenilo/uso terapéutico , Neoplasias Cerebelosas/tratamiento farmacológico , Neoplasias Cerebelosas/metabolismo , Niño , Preescolar , Biología Computacional , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Proteínas Hedgehog/metabolismo , Humanos , Lactante , Meduloblastoma/tratamiento farmacológico , Meduloblastoma/metabolismo , Modelos Biológicos , Pronóstico , Piridinas/farmacología , Piridinas/uso terapéutico , Reproducibilidad de los Resultados , Transducción de Señal/efectos de los fármacos , Resultado del TratamientoRESUMEN
Psoriasis is a chronic, debilitating, immune-mediated inflammatory skin disease. As IFN-γ is involved in many cellular processes, including activation of dendritic cells (DCs), antigen processing and presentation, cell adhesion and trafficking, and cytokine and chemokine production, IFN-γ-producing Th1 cells were proposed to be integral to the pathogenesis of psoriasis. Recently, IFN-γ was shown to enhance IL-23 and IL-1 production by DCs and subsequently induce Th17 cells, which are important contributors to the inflammatory cascade in psoriatic lesions. To determine whether IFN-γ indeed induces the pathways expressed in psoriatic lesions, a single intradermal injection of IFN-γ was administered to an area of clinically normal, non-lesional (NL) skin of psoriasis patients and biopsies were collected 24 hours later. Although there were no visible changes in the skin, IFN-γ induced many molecular and histological features characteristic of psoriatic lesions. IFN-γ increased a number of differentially expressed genes in the skin, including many chemokines concomitant with an influx of T cells and inflammatory DCs. Furthermore, inflammatory DC products tumor necrosis factor (TNF), inducible nitric oxide synthase, IL-23, and TNF-related apoptosis-inducing ligand were present in IFN-γ-treated skin. Thus, IFN-γ, which is significantly elevated in NL skin compared with healthy skin, appears to be a key pathogenic cytokine that can induce many features of the inflammatory cascade of psoriasis.
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Inflamación/metabolismo , Inflamación/patología , Interferón gamma/farmacología , Psoriasis/metabolismo , Psoriasis/patología , Piel/efectos de los fármacos , Piel/metabolismo , Biopsia , Estudios de Casos y Controles , Movimiento Celular , Células Dendríticas/patología , Humanos , Inflamación/inducido químicamente , Inyecciones Intradérmicas , Interferón gamma/administración & dosificación , Interferón gamma/efectos adversos , Interleucina-23/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Fenotipo , Piel/patología , Linfocitos T/patología , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoAsunto(s)
Proteínas Proto-Oncogénicas c-jun/metabolismo , Psoriasis/metabolismo , Regulación hacia Arriba , Expresión Génica , Humanos , Inmunohistoquímica , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Proto-Oncogénicas c-jun/análisis , Psoriasis/genética , ARN Mensajero/análisis , ARN Mensajero/metabolismoRESUMEN
Interleukin-17A (IL-17A) is elaborated by the T helper 17 (T(H)17) subset of T(H) cells and exhibits potent proinflammatory properties in animal models of autoimmunity, including collagen-induced arthritis, experimental autoimmune encephalomyelitis, and experimental autoimmune uveitis. To determine whether IL-17A mediates human inflammatory diseases, we investigated the efficacy and safety of AIN457, a human antibody to IL-17A, in patients with psoriasis, rheumatoid arthritis, and chronic noninfectious uveitis. Patients with chronic plaque-type psoriasis (n = 36), rheumatoid arthritis (n = 52), or chronic noninfectious uveitis (n = 16) were enrolled in clinical trials to evaluate the effects of neutralizing IL-17A by AIN457 at doses of 3 to 10 mg/kg, given intravenously. We evaluated efficacy by measuring the psoriasis area and severity index (PASI), the American College of Rheumatology 20% response (ACR20) for rheumatoid arthritis, or the number of responders for uveitis, as defined by either vision improvement or reduction in ocular inflammation or corticosteroid dose. AIN457 treatment induced clinically relevant responses of variable magnitude in patients suffering from each of these diverse immune-mediated diseases. Variable response rates may be due to heterogeneity in small patient populations, differential pathogenic roles of IL-17A in these diseases, and the different involvement or activation of IL-17A-producing cells. The rates of adverse events, including infections, were similar in the AIN457 and placebo groups. These results support a role for IL-17A in the pathophysiology of diverse inflammatory diseases including psoriasis, rheumatoid arthritis, and noninfectious uveitis.
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Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/uso terapéutico , Anticuerpos , Artritis Reumatoide/tratamiento farmacológico , Interleucina-17/inmunología , Psoriasis/tratamiento farmacológico , Uveítis/tratamiento farmacológico , Adolescente , Adulto , Anciano , Animales , Anticuerpos/inmunología , Anticuerpos/uso terapéutico , Anticuerpos Monoclonales Humanizados , Artritis Reumatoide/inmunología , Artritis Reumatoide/patología , Método Doble Ciego , Humanos , Persona de Mediana Edad , Placebos/uso terapéutico , Psoriasis/inmunología , Psoriasis/patología , Resultado del Tratamiento , Uveítis/inmunología , Uveítis/patología , Adulto JovenRESUMEN
Genomic signature maps of different cell types can aid in the interpretation of genomic data of specimens collected during disease states. We have defined "lineage-specific" genes, as well as "activation" genes, for cellular components of the skin: keratinocytes, fibroblasts, macrophages, monocytes, T cells, immature, and mature dendritic cells (DCs). Re-analysis of a previously published gene set of psoriasis then provided a model for assessing the usefulness of these maps. We were able to ascribe over 90% of these genes to specific cell types, and there was a surprisingly large contribution from DCs. This shows the utility of such cellular gene maps.
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Mapeo Cromosómico , Genómica , Enfermedades de la Piel/genética , Enfermedades de la Piel/patología , Piel/patología , Biopsia , Linaje de la Célula/fisiología , Células Dendríticas/patología , Células Dendríticas/fisiología , Fibroblastos/patología , Fibroblastos/fisiología , Humanos , Queratinocitos/patología , Queratinocitos/fisiología , Macrófagos/patología , Macrófagos/fisiología , Monocitos/patología , Monocitos/fisiología , Análisis de Secuencia por Matrices de Oligonucleótidos , Piel/inmunología , Enfermedades de la Piel/inmunología , Linfocitos T/patología , Linfocitos T/fisiologíaRESUMEN
Therapeutic antibodies against tumor necrosis factor (TNF) (infliximab) and IFNgamma (fontolizumab) have been developed to treat autoimmune diseases. While the primary targets of these antibodies are clearly defined, the set of inflammatory molecules, which is altered by use of these inhibitors, is poorly understood. We elucidate the target genes of these antibodies in activated human peripheral blood mononuclear cells from healthy volunteers. While genes suppressed by fontolizumab overlap with known IFNgamma-induced genes, majority of genes suppressed by infliximab have previously not been traced to TNF signaling. With this approach we were able to extrapolate new TNF-associated genes to be upregulated in psoriasis vulgaris, an "autoimmune" disease effectively treated with TNF antagonists. These genes represent potential therapeutic targets of TNF antagonists in psoriasis. Furthermore, these data establish an unexpected effect of TNF blockade on IFNgamma synthesis by T cells. Synthesis of IFNgamma, a cytokine of Th1-polarized T cells, is suppressed by 8.1-fold (P<0.01) at the mRNA level, while synthesis of IFNgamma is eliminated in >60% of individual T cells. These data suggest that TNF blockade with infliximab can suppress a major pathway of the adaptive immune response and this observation provides a key rationale for targeting TNF in "Type-1" T-cell-mediated autoimmune diseases.
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Anticuerpos Monoclonales/farmacología , Fármacos Dermatológicos/farmacología , Perfilación de la Expresión Génica , Psoriasis/tratamiento farmacológico , Psoriasis/genética , Anticuerpos Monoclonales Humanizados , Linfocitos B/efectos de los fármacos , Linfocitos B/fisiología , Expresión Génica/efectos de los fármacos , Expresión Génica/inmunología , Humanos , Técnicas In Vitro , Infliximab , Interferón gamma/inmunología , Interferón gamma/metabolismo , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/fisiología , Psoriasis/patología , Células TH1/efectos de los fármacos , Células TH1/fisiología , Factor de Necrosis Tumoral alfa/inmunología , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
The importance of T helper 17 (Th17) cells in inflammation and autoimmunity is now being appreciated. We analyzed psoriasis skin lesions and peripheral blood for the presence of IL-17-producing T cells. We localized Th17 cells predominantly to the dermis of psoriasis skin lesions, confirmed that IL-17 mRNA increased with disease activity, and demonstrated that IL-17 mRNA expression normalized with cyclosporine therapy. IL-22 mRNA expression mirrored IL-17 and both were downregulated in parallel with keratin 16. Th17 cells are a discrete population, separate from Th1 cells (which are also in psoriasis lesions), and Th2 cells. Our findings suggest that psoriasis is a mixed Th1 and Th17 inflammatory environment. Th17 cells may be proximal regulators of psoriatic skin inflammation, and warrant further attention as therapeutic targets.
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Dermis/patología , Psoriasis/inmunología , Psoriasis/patología , Células TH1/clasificación , Células TH1/patología , Animales , Anticuerpos Monoclonales/farmacología , Células Cultivadas , Ciclosporina/uso terapéutico , Dermis/inmunología , Expresión Génica/efectos de los fármacos , Expresión Génica/inmunología , Humanos , Inmunosupresores/uso terapéutico , Interferón gamma/genética , Interleucina-17/genética , Interleucina-17/inmunología , Interleucina-17/metabolismo , Interleucinas/genética , Queratina-16/genética , Psoriasis/tratamiento farmacológico , ARN Mensajero/metabolismo , Ratas , Células TH1/inmunología , Interleucina-22RESUMEN
Therapeutic modulation of psoriasis with targeted immunosuppressive agents defines inflammatory genes associated with disease activity and may be extrapolated to a wide range of autoimmune diseases. Cyclosporine A (CSA) is considered a "gold standard" therapy for moderate-to-severe psoriasis. We conducted a clinical trial with CSA and analyzed the treatment outcome in blood and skin of 11 responding patients. In the skin, as expected, CSA modulated genes from activated T cells and the "type 1" pathway (p40, IFN-gamma, and STAT-1-regulated genes). However, CSA also modulated genes from the newly described Th17 pathway (IL-17, IL-22, and downstream genes S100A12, DEFB-2, IL-1beta, SEPRINB3, LCN2, and CCL20). CSA also affected dendritic cells, reducing TNF and inducible NO synthase (products of inflammatory TNF- and inducible NO synthase-producing dendritic cells), CD83, and IL-23p19. We detected 220 early response genes (day 14 posttreatment) that were down-regulated by CSA. We classified >95% into proinflammatory or skin resident cells. More myeloid-derived than activated T cell genes were modulated by CSA (54 myeloid genes compared with 11 lymphocyte genes), supporting the hypothesis that myeloid derived genes contribute to pathogenic inflammation in psoriasis. In circulating mononuclear leukocytes, in stark contrast, no inflammatory gene activity was detected. Thus, we have constructed a genomic signature of successful treatment of psoriasis which may serve as a reference to guide development of other new therapies. In addition, these data also identify new gene targets for therapeutic modulation and may be applied to wide range of autoimmune diseases.
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Enfermedades Autoinmunes/genética , Ciclosporina/administración & dosificación , Células Dendríticas/efectos de los fármacos , Perfilación de la Expresión Génica , Inmunosupresores/administración & dosificación , Psoriasis/genética , Linfocitos T Colaboradores-Inductores/efectos de los fármacos , Antígenos CD/análisis , Enfermedades Autoinmunes/tratamiento farmacológico , Enfermedades Autoinmunes/inmunología , Complejo CD3/análisis , Células Dendríticas/inmunología , Expresión Génica/efectos de los fármacos , Humanos , Inmunoglobulinas/análisis , Interleucina-17/genética , Subunidad alfa del Receptor de Interleucina-2/análisis , Glicoproteínas de Membrana/análisis , Óxido Nítrico Sintasa de Tipo II/genética , Farmacogenética , Psoriasis/tratamiento farmacológico , Psoriasis/inmunología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Piel/efectos de los fármacos , Piel/inmunología , Piel/patología , Linfocitos T Colaboradores-Inductores/inmunología , Factor de Necrosis Tumoral alfa/genética , Antígeno CD83RESUMEN
Proinflammatory diseases like rheumatoid arthritis, Crohn's disease, and psoriasis have been treated by the tumor necrosis factor (TNF) antagonists infliximab and etanercept with different degrees of success. Although these agents are widely used in humans, little is known about their mechanisms of action or why etanercept and infliximab have differences in clinical activity. In this study, we define leukocyte genes that are suppressed by etanercept within 24 hours of exposure. Compared to previous work with infliximab, fewer immune-related genes are suppressed by etanercept. Importantly, the range of genes suppressed by these alternative TNF inhibitors is only partially overlapping, suggesting each has unique immune modulating effects. In sharp contrast to etanercept, infliximab strongly suppresses genes associated with "Type 1" immune responses (IFN-gamma and the IL-12-receptor beta 2 subunit), providing a clear mechanism for clinically relevant immune suppression.
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Anticuerpos Monoclonales/farmacología , Inmunoglobulina G/farmacología , Inflamación/tratamiento farmacológico , Inflamación/genética , Leucocitos/efectos de los fármacos , Secuencia de Bases , Cartilla de ADN/genética , Regulación hacia Abajo/efectos de los fármacos , Etanercept , Humanos , Técnicas In Vitro , Inflamación/inmunología , Infliximab , Leucocitos/inmunología , Análisis de Secuencia por Matrices de Oligonucleótidos , Farmacogenética , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores del Factor de Necrosis Tumoral , Factor de Necrosis Tumoral alfa/antagonistas & inhibidoresRESUMEN
BACKGROUND: Atopic dermatitis (AD) and psoriasis represent contrasting poles of the T(H)1 versus T(H)2 paradigm. Both diseases have been associated with increased numbers of dendritic cells (DCs) in the skin, but the similarities and differences in DC populations need to be established. OBJECTIVE: We aimed to characterize the specific DC subsets, as well as chemokine and cytokine environment in chronic AD compared with psoriasis. METHODS: Skin biopsies were obtained from patients with acute exacerbation of chronic AD (n = 18), psoriasis (n = 15), and healthy volunteers (n = 15) for microarray analysis, RT-PCR, immunohistochemistry, and double-label immunofluorescence. RESULTS: Myeloid DCs upregulate CCL17 and CCL18 in AD, as opposed to TNF-alpha and inducible nitric oxide synthase (iNOS) in psoriasis. In our study, we identified cells phenotypically identical to the inflammatory dendritic epidermal cells in the dermis in both diseases, although to a lesser extent in psoriasis. We found substantially higher numbers of dermal CCL22 producing plasmacytoid DCs in AD. The thymic stromal lymphopoietin receptor showed significantly higher expression in AD, whereas the thymic stromal lymphopoietin ligand was upregulated more in psoriasis. CONCLUSION: There are major differences in myeloid and plasmacytoid subsets of cutaneous DCs and the chemokine/cytokine environment between AD and psoriasis. Distinct subsets within the CD11c(+) population may influence polarization through the production of regulatory mediators, including iNOS, TNF, CCL17, and CCL18. Plasmacytoid DCs may also influence T(H)2 polarization, having a more important role in AD than previously appreciated. CLINICAL IMPLICATIONS: Dermal inflammatory dendritic cells in AD and TNF and iNOS-producing DCs in psoriasis, and/or their regulatory products, may be potential targets for future therapeutic interventions.
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Células Dendríticas/inmunología , Dermatitis Atópica/inmunología , Inflamación/inmunología , Psoriasis/inmunología , Adolescente , Adulto , Anciano , Citocinas/biosíntesis , Células Dendríticas/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Piel/citología , Piel/inmunología , Linfocitos T/inmunologíaRESUMEN
Alefacept is an LFA3-Ig fusion protein that binds to CD2 and is thought to inhibit T cell activation by antagonism of CD2 signaling or by lysis of CD2(+) cells. Alefacept is potential future therapeutic for organ transplant recipients or graft-vs-host disease and is an approved therapeutic for psoriasis vulgaris, which is a T cell-mediated inflammatory disease. However, alefacept improves psoriasis in only approximately 50% of patients treated for 12 wk. We studied the immunologic effects of alefacept in a group of psoriasis patients during treatment. We found that T cells, especially CD8(+) T cells, were rapidly decreased in the peripheral circulation. Decreases in circulating T cells were not associated with induced apoptosis. Unexpectedly, in addition to suppression of inflammatory genes, we found a marked induction of mRNAs for STAT1, IL-8, and monokine induced by IFN-gamma during the first day of treatment in PBMC. We confirmed the agonistic effects of alefacept in PBMC in vitro, which were similar to CD3/CD28 ligation on T cells. These data establish that alefacept activates gene expression programs in leukocytes and suggest that its therapeutic action may be as a mixed agonist/antagonist. Furthermore, responding patients to alefacept treatment show unique patterns of gene modulation. Whereas alefacept down-regulated TCRs CD3D and CD2 in responders, nonresponders reveal a higher expression of T cell activation genes such as CD69 in pretreatment PBMC. These finding suggest a potential basis for categorizing responders vs nonresponders at an early time point in treatment or before treatment of a broad range of proinflammatory diseases. This study 1) establishes alefacept as a novel CD2 agonist molecule for induction of leukocyte activation genes (prior work proposed its mechanism as a CD2 antagonist) and 2) that differential activation of genes may categorize clinical responders to this agent, critical for cost-effective use of this drug.
Asunto(s)
Regulación de la Expresión Génica/inmunología , Psoriasis/inmunología , Psoriasis/terapia , Proteínas Recombinantes de Fusión/administración & dosificación , Proteínas Recombinantes de Fusión/agonistas , Adulto , Anciano , Alefacept , Apoptosis/genética , Apoptosis/inmunología , Biomarcadores/sangre , Antígenos CD2/biosíntesis , Antígenos CD2/genética , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Regulación hacia Abajo/genética , Regulación hacia Abajo/inmunología , Femenino , Humanos , Memoria Inmunológica/genética , Activación de Linfocitos/genética , Linfopenia/genética , Linfopenia/inmunología , Linfopenia/patología , Masculino , Persona de Mediana Edad , Unión Proteica/genética , Unión Proteica/inmunología , Psoriasis/genética , Psoriasis/patología , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Recombinantes de Fusión/uso terapéutico , Fase de Descanso del Ciclo Celular/genética , Fase de Descanso del Ciclo Celular/inmunología , Piel/inmunología , Piel/metabolismo , Piel/patología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Subgrupos de Linfocitos T/patología , Regulación hacia Arriba/genética , Regulación hacia Arriba/inmunologíaRESUMEN
Basal cell carcinoma (BCC), the most common human cancer, undergoes spontaneous regression in certain circumstances, which is potentially immune-mediated. To understand the immune response surrounding BCCs, we characterized the genomic, protein, and cellular microenvironment associated with BCC in comparison to normal skin. Our results demonstrated the following: (1) CD4+ CD25+ Foxp3+ surround epithelial tumor aggregates; (2) Immature dendritic cells (DCs) were abundant in the tumor microenvironment; (3) BCC showed increased expression of IL-4, IL-10, and CCL22 and increased expression of interferon-associated genes (IFI27, IRF1, IRF7, and G1P2) and IL-12/23, gene indicating a Th2 dominant microenvironment. Our findings suggest a dynamic state within the immune microenvironment associated with BCC. The finding of phenotypic T regs, in conjunction with immature DCs and Th2 cytokines, suggests an attenuated state of immunity to human BCC. In contrast, abundant CD8+ T cells, an interferon signal, and IL-12/23 suggest partial host antitumor response. A better understanding of these opposing forces within the immune microenvironment may facilitate development of more potent immune-based treatment for BCC and other human carcinomas.
Asunto(s)
Carcinoma Basocelular/patología , Factores de Transcripción Forkhead/metabolismo , Neoplasias Cutáneas/patología , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Células Th2/patología , Biopsia , Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/metabolismo , Linfocitos T CD8-positivos/patología , Carcinoma Basocelular/inmunología , Células Dendríticas/inmunología , Células Dendríticas/patología , Humanos , Factor 1 Regulador del Interferón/metabolismo , Factor 7 Regulador del Interferón/metabolismo , Interleucina-10/metabolismo , Interleucina-12/metabolismo , Interleucina-4/metabolismo , Neoplasias Cutáneas/inmunología , Linfocitos T Reguladores/patología , Células Th2/inmunologíaRESUMEN
It is rare that a single gene is sufficient to represent all aspects of genomic activity. Similarly, most common diseases cannot be explained by a mutations at a single locus. Since complex systems tend to be neither linear nor hierarchical in nature, but to have correlated components of unknown relative importance, the assumptions of traditional (parametric) multivariate statistical methods can rarely be justified on theoretical grounds. Empirical "validation" is not only problematic, but also time consuming. Here we demonstrates how bioinformatics tools, ranging from spreadsheets to grids, can enable u-statistics as a non-parametric alternative for scoring multivariate ordinal data. Applications are shown to improve assessment of genetic risk factors, quality control of microarrays and signal value estimation, scoring genomic profiles that best correlated with complex risk factors (cardiovascular diseases), and complex responses to an intervention (treatment of psoriasis).
Asunto(s)
Biología Computacional , Análisis por Micromatrices/estadística & datos numéricos , Ingeniería Biomédica , Enfermedades Cardiovasculares/genética , Femenino , Perfilación de la Expresión Génica/estadística & datos numéricos , Humanos , Masculino , Análisis por Micromatrices/normas , Modelos Estadísticos , Psoriasis/patología , Control de Calidad , Factores de Riesgo , Estadísticas no ParamétricasRESUMEN
Heme oxygenase (HO) is a microsomal enzyme that oxidatively cleaves heme to form biliverdin, with the release of iron and carbon monoxide (CO). HO not only controls the availability of heme for the synthesis of heme proteins but also is responsible for the generation of CO, which binds to the heme moiety of heme proteins thus affecting their enzymatic activity. Cyclooxygenase (COX) is a heme protein that catalyzes the conversion of arachidonic acid to prostaglandin H(2), the precursor of prostanoids that participate in the regulation of vascular function. The goal of the present study was to determine whether the heme-HO system regulates COX enzyme expression and activity in vascular endothelial cells. Endothelial cells stably transfected with the human HO-1 gene exhibited a severalfold increase in human HO-1 mRNA levels, which was accompanied by an increase in HO activity and a marked decrease in prostaglandin (PG) E(2) and 6-keto-PGF(1alpha) levels. Exposure of cells to CoCl(2), an inducer of HO-1 gene expression, resulted in increases in HO-1 protein levels and HO activity. The increase in HO activity was associated with a subsequent decrease in COX activity, which returned to normal levels following normalization of HO activity. The addition of heme resulted in an increase in COX activity with an increase in PGE(2) and 6-keto-PGF(1alpha) levels. The degree of HO-1 expression and, consequently, the level of cellular heme, were directly related to COX activity. These results demonstrate that the heme-HO system can function as a cellular regulator of the expression of vascular COX, thus influencing the generation of prostanoids, PGE(2) and PGI(2), known to play a role in vascular homeostasis.