RESUMEN
Immunological memory is a defining feature of vertebrate physiology, allowing rapid responses to repeat infections. However, the molecular mechanisms required for its establishment and maintenance remain poorly understood. Here, we demonstrated that the first steps in the acquisition of T-cell memory occurred during the initial activation phase of naïve T cells by an antigenic stimulus. This event initiated extensive chromatin remodeling that reprogrammed immune response genes toward a stably maintained primed state, prior to terminal differentiation. Activation induced the transcription factors NFAT and AP-1 which created thousands of new DNase I-hypersensitive sites (DHSs), enabling ETS-1 and RUNX1 recruitment to previously inaccessible sites. Significantly, these DHSs remained stable long after activation ceased, were preserved following replication, and were maintained in memory-phenotype cells. We show that primed DHSs maintain regions of active chromatin in the vicinity of inducible genes and enhancers that regulate immune responses. We suggest that this priming mechanism may contribute to immunological memory in T cells by facilitating the induction of nearby inducible regulatory elements in previously activated T cells.
Asunto(s)
Cromatina/metabolismo , Memoria Inmunológica , Linfocitos T/metabolismo , Animales , Células Cultivadas , Quimiocina CCL1/genética , Subunidad alfa 2 del Factor de Unión al Sitio Principal/genética , Desoxirribonucleasa I/metabolismo , Expresión Génica , Factor Estimulante de Colonias de Granulocitos y Macrófagos/genética , Humanos , Interleucina-3/genética , Células Jurkat , Ratones Transgénicos , Factores de Transcripción NFATC/genética , Proteína Proto-Oncogénica c-ets-1/genética , ARN Mensajero/metabolismo , Bazo/inmunología , Linfocitos T/inmunología , Factor de Transcripción AP-1/genéticaRESUMEN
Mutation of the MYD88 gene has recently been identified in activated B-cell-like diffuse cell lymphoma and enhanced Janus kinase/signal transducer and activator of transcription (JAK-STAT) and nuclear factor κB (NF-κB) signaling pathways. A whole exome-sequencing study of Waldenstrom macroglobulinemia (WM) suggested a high frequency of MYD88 L265P mutation in WM. The genetic background is not fully deciphered in WM, although the role of NF-κB and JAK-STAT has been demonstrated. We analyzed MYD88 mutation in exon 5 and characterized the clinical significance of this genetic alteration in 67 WM patients. Clinical features; immunophenotypic markers; and conventional cytogenetic, fluorescence in situ hybridization, and single nucleotide polymorphism array data were analyzed. MYD88 L265P mutation was acquired in 79% of patients. Overall, we have identified alteration of the MYD88 locus in 91% of WM patients, including 12% with gain on chromosome 3 at the 3p22 locus that included the MYD88 gene. Patients with absence of MYD88 mutation were WM characterized with a female predominance, a splenomegaly, gain of chromosome 3, and CD27 expression. Importantly, inhibition of MYD88 signaling induced cytotoxicity and inhibited cell growth of cell lines issued from patients with WM. In conclusion, these results confirm a high frequency of MYD88 L265P mutation in WM. The discovery of MYD88 L265P mutation may contribute to a better understanding of the physiopathogeny of WM.
Asunto(s)
Factor 88 de Diferenciación Mieloide/genética , Mutación Puntual , Macroglobulinemia de Waldenström/genética , Adulto , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Proliferación Celular , Femenino , Citometría de Flujo , Humanos , Masculino , Persona de Mediana Edad , FN-kappa B/genética , FN-kappa B/metabolismo , Polimorfismo de Nucleótido Simple , Transducción de Señal/fisiología , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/genética , Miembro 7 de la Superfamilia de Receptores de Factores de Necrosis Tumoral/metabolismo , Macroglobulinemia de Waldenström/metabolismo , Macroglobulinemia de Waldenström/terapiaRESUMEN
Bing-Neel syndrome (BNS), a rare neurological syndrome associated with Waldenström macroglobulinaemia (WM), is a direct involvement of the central nervous system by lymphoplasmacytoid cells characterized with an adverse prognostic. The MYD88 L265P mutation has been identified in the vast majority of patients with WM. The diagnosis of BNS is often challenging because of the variety of clinical presentations associated with difficult histological techniques. We hypothesized that identification of MYD88 L265P mutation in the cerebrospinal fluid (CSF) would contribute to the diagnosis of BNS in addition to imaging, flow cytometry and cytology. We identified MYD88 L265P mutation in the CSF and the bone marrow of all cases of BNS using quantitative polymerase chain reaction qPCR and Sanger sequencing. Copy neutral loss of heterozygosity including MYD88 was observed in one case. No mutation of CXCR4, CD79A and CD79B was observed in parallel. We further showed that monitoring the quantitative expression of MYD88 L265P mutation might be a useful molecular tool to monitor response to chemotherapy using qPCR. In conclusion, identification of MYD88 L265P mutation might be a new molecular-based biomarker tool to add to the diagnostic and monitoring armamentarium for BNS.
Asunto(s)
Heterocigoto , Mutación Missense , Factor 88 de Diferenciación Mieloide/genética , Proteínas de Neoplasias/genética , Enfermedades del Sistema Nervioso/diagnóstico , Enfermedades del Sistema Nervioso/genética , Macroglobulinemia de Waldenström/genética , Adulto , Anciano , Médula Ósea/metabolismo , Médula Ósea/patología , Humanos , Masculino , Persona de Mediana Edad , Factor 88 de Diferenciación Mieloide/metabolismo , Proteínas de Neoplasias/metabolismo , Enfermedades del Sistema Nervioso/líquido cefalorraquídeo , Enfermedades del Sistema Nervioso/patología , Estudios Prospectivos , Síndrome , Macroglobulinemia de Waldenström/líquido cefalorraquídeo , Macroglobulinemia de Waldenström/patologíaRESUMEN
The closely linked human IL-3 and GM-CSF genes are tightly regulated and are expressed in activated T cells and mast cells. In this study, we used transgenic mice to study the developmental regulation of this locus and to identify DNA elements required for its correct activity in vivo. Because these two genes are separated by a CTCF-dependent insulator, and the GM-CSF gene is regulated primarily by its own upstream enhancer, the main objective in this study was to identify regions of the locus required for correct IL-3 gene expression. We initially found that the previously identified proximal upstream IL-3 enhancers were insufficient to account for the in vivo activity of the IL-3 gene. However, an extended analysis of DNase I-hypersensitive sites (DHSs) spanning the entire upstream IL-3 intergenic region revealed the existence of a complex cluster of both constitutive and inducible DHSs spanning the -34- to -40-kb region. The tissue specificity of these DHSs mirrored the activity of the IL-3 gene, and included a highly inducible cyclosporin A-sensitive enhancer at -37 kb that increased IL-3 promoter activity 40-fold. Significantly, inclusion of this region enabled correct in vivo regulation of IL-3 gene expression in T cells, mast cells, and myeloid progenitor cells.
Asunto(s)
Elementos de Facilitación Genéticos/inmunología , Regulación del Desarrollo de la Expresión Génica/inmunología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/biosíntesis , Factor Estimulante de Colonias de Granulocitos y Macrófagos/genética , Interleucina-3/biosíntesis , Interleucina-3/genética , Animales , Línea Celular , Línea Celular Tumoral , Células Cultivadas , Desoxirribonucleasa I/genética , Elementos de Facilitación Genéticos/genética , Sitios Genéticos/inmunología , Humanos , Células Jurkat , Ratones , Ratones Transgénicos , Distribución Tisular/genética , Distribución Tisular/inmunologíaRESUMEN
SNP array (SNPa) was developed to detect copy number alteration (CNA) and loss of heterozygosity (LOH) without copy number changes, CN-LOH. We aimed to identify novel genomic aberrations using SNPa in 31 WM with paired samples. Methylation status and mutation were analyzed on target genes. A total of 61 genetic aberrations were observed, 58 CNA (33 gains, 25 losses) in 58% of patients and CN-LOH in 6% of patients. The CNA were widely distributed throughout the genome, including 12 recurrent regions and identified new cryptic clonal chromosomal lesions that were mapped. Gene set expression analysis demonstrated a relationship between either deletion 6q or gain of chromosome 4 and alteration of gene expression profiling. We then studied methylation status and sought for mutations in altered regions on target genes. We observed methylation of DLEU7 on chromosome 13 in all patients (n = 12) with WM, and mutations of CD79B/CD79A genes (17q region), a key component of the BCR pathway, in 15% of cases. Most importantly, higher frequency of ≥3 CNA was observed in symptomatic WM. In conclusion, this study expands the view of the genomic complexity of WM, especially in symptomatic WM, including a potentially new mechanism of gene dysfunction, acquired uniparental disomy/CN-LOH. Finally, we have identified new potential target genes in WM, such as DLEU7 and CD79A/B.
Asunto(s)
Aberraciones Cromosómicas , Variaciones en el Número de Copia de ADN , Metilación de ADN , Regulación de la Expresión Génica , Pérdida de Heterocigocidad , Mutación , Macroglobulinemia de Waldenström/genética , Adulto , Anciano , Anciano de 80 o más Años , Antígenos CD79/genética , Antígenos CD79/metabolismo , Deleción Cromosómica , Duplicación Cromosómica , Estudios de Cohortes , Femenino , Francia , Estudio de Asociación del Genoma Completo , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Macroglobulinemia de Waldenström/metabolismoRESUMEN
Allogeneic hematopoietic cell transplantation (HCT) is part of the standard of care for many hematological diseases. Over the last decades, significant advances in patient and donor selection, conditioning regimens as well as supportive care of patients undergoing allogeneic HCT leading to improved overall survival have been made. In view of many new treatment options in cellular and molecular targeted therapies, the place of allogeneic transplantation in therapy concepts must be reviewed. Most aspects of HCT are well standardized by national guidelines or laws as well as by certification labels such as FACT-JACIE. However, the requirements for human resources, construction and layout of a unit treating patients during the transplantation procedure and for different complications are not well defined. Here, we describe the process of planning a transplant unit in order to open a discussion that could lead to more precise guidelines in the field of personnel and infrastructural requirements for hospitals caring for people with severe immunosuppression.
Asunto(s)
Trasplante de Médula Ósea/normas , Ambiente de Instituciones de Salud/normas , Enfermedades Hematológicas/terapia , Trasplante de Células Madre Hematopoyéticas/normas , Aire/normas , Tratamiento Basado en Trasplante de Células y Tejidos/normas , Dieta Saludable/normas , Selección de Donante/normas , Francia , Personal de Salud/normas , Unidades Hospitalarias/normas , Humanos , Higiene , Terapia de Inmunosupresión/normas , Monitoreo Fisiológico/métodos , Ropa de Protección/normas , Sociedades Médicas , Esterilización/normas , Trasplante Homólogo/normas , Visitas a PacientesRESUMEN
BACKGROUND: Immunomodulatory drugs, IMid compounds, are active in Waldenström's macroglobulinemia (WM), although in a lesser extent than multiple myeloma, where it was initially developed. We hypothesized WM tumour cells might develop mechanisms of resistance, and sought to identify and describe these mechanisms. MATERIAL AND METHOD: MM and WM-derived cell lines, and Waldenström's CD19+ cells were treated using both lenalidomide and pomalidomide. Stable CRBN expressing cells were generated. RESULTS: WM-derived cells were resistant to IMid compounds. We demonstrated a modulation of the downstream targets of IRF4, despite low expression of cereblon, and hypothesized IRF4 was the cause for resistance to IMid compounds. We ruled out the role of various IRF4 regulatory mechanisms, and other pathways activating WM tumor cells, such as B cell activators. CONCLUSION: This study demonstrated that mechanisms of resistance to IMid compounds could be not related to cereblon. IRF4 was identified as the potential mechanism of resistance to lenalidomide and pomalidomide in WM. It potentially explains the lesser activity observed in the clinic in WM. Interestingly, some WM patients benefited strongly to lenalidomide and pomalidomide, and future studies will have to describe the indirect mechanisms of IMid compounds in WM, possibly related to an immune-mediated process.
RESUMEN
Purpose:TP53 is a tumor-suppressor gene that functions as a regulator influencing cellular responses to DNA damage, and TP53 alterations are associated with pejorative outcome in most B-lymphoid disorders. Little is known regarding TP53 alteration in Waldenstrom's macroglobulinemia (WM).Experimental Design: Here, we have explored the incidence of TP53 alteration using Sanger sequencing and ultradeep-targeted sequencing in 125 WM and 10 immunoglobulin M (IgM) monoclonal gammopathy of undetermined significance (MGUS), along with the clinical features and the associated genomic landscape using single-nucleotide polymorphism array and mutational landscape in an integrative study.Results: Overall, we have identified alteration of TP53 locus including mutation, deletion, and copy-neutral LOH in 11.2% of WM. TP53 mutation was acquired in 7.3% of patients with WM at diagnosis, being absent in IgM MGUS, and was highly correlated to deletion 17p. No correlation with CXCR4 mutations was observed. Patients with TP53 alteration had a greater number of genomic abnormalities. Importantly, WM with TP53 alteration had a significantly shorter overall survival, particularly in symptomatic WM, and independently of the international prognostic scoring system for Waldenstrom macroglobulinemia (IPSSWM) score. Specific treatment for WM with TP53 may have to be studied. Nutlin-3a-targeted p53 signaling induced cytotoxicity preclinically, along with new compounds such as ibrutinib, PrimaMet, or CP31398 that bypass p53 pathway in WM, paving the path for future treatment-tailored options.Conclusions: Our results highlight the clinical significance of detection of TP53 alteration in WM to determine the prognosis of WM and guide the treatment choice. Clin Cancer Res; 23(20); 6325-35. ©2017 AACR.
Asunto(s)
Mutación , Proteína p53 Supresora de Tumor/genética , Macroglobulinemia de Waldenström/diagnóstico , Macroglobulinemia de Waldenström/genética , Adulto , Anciano , Anciano de 80 o más Años , Apoptosis , Supervivencia Celular/genética , Deleción Cromosómica , Cromosomas Humanos Par 17 , Femenino , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Unión Proteica , Dominios y Motivos de Interacción de Proteínas/genética , Análisis de Supervivencia , Proteína p53 Supresora de Tumor/química , Proteína p53 Supresora de Tumor/metabolismo , Macroglobulinemia de Waldenström/mortalidadRESUMEN
Approximately 30% of the patients who fulfil the criteria of Waldenström's macroglobulinemia (WM) are diagnosed while asymptomatic (indolent), and will not require immediate therapy. Conversely, patients with a disease-related event will be considered for therapy. The physiopathology of these 2 groups remains unclear, and the mechanisms of progression from indolent to symptomatic WM have yet to be fully understood. Seventeen patients diagnosed with WM were included in this study, 8 asymptomatic WM (A-WM) and 9 symptomatic WM (S-WM). A differential analysis was performed on a first series of 11 patients and identified 48 genes whose expression separated samples from A- to S-WM. This gene signature was then confirmed on a second independent validation set of 6 WM. Within this expression profile, BACH2, a B-cell transcription factor known to be a tumor suppressor gene, was found to be over-expressed in A-MW relatively to S-MW. We specifically over-expressed BACH2 in a WM-related cell line and observed a significant reduction of the clonogenic activity. To the best of our knowledge, we report for the first time a specific gene expression signature that differentiates A-WM and S-WM. Within this expression profile, BACH2 was identified as a candidate gene that may help to understand better the behavior of tumor cells in indolent WM.
RESUMEN
PURPOSE: Whole-genome sequencing has revealed MYD88 L265P and CXCR4 mutations (CXCR4(mut)) as the most prevalent somatic mutations in Waldenström macroglobulinemia. CXCR4 mutation has proved to be of critical importance in Waldenström macroglobulinemia, in part due to its role as a mechanism of resistance to several agents. We have therefore sought to unravel the different aspects of CXCR4 mutations in Waldenström macroglobulinemia. EXPERIMENTAL DESIGN: We have scanned the two coding exons of CXCR4 in Waldenström macroglobulinemia using deep next-generation sequencing and Sanger sequencing in 98 patients with Waldenström macroglobulinemia and correlated with SNP array landscape and mutational spectrum of eight candidate genes involved in TLR, RAS, and BCR pathway in an integrative study. RESULTS: We found all mutations to be heterozygous, somatic, and located in the C-terminal domain of CXCR4 in 25% of the Waldenström macroglobulinemia. CXCR4 mutations led to a truncated receptor protein associated with a higher expression of CXCR4. CXCR4 mutations pertain to the same clone as to MYD88 L265P mutations but were mutually exclusive to CD79A/CD79B mutations (BCR pathway). We identified a genomic signature in CXCR4(mut) Waldenström macroglobulinemia traducing a more complex genome. CXCR4 mutations were also associated with gain of chromosome 4, gain of Xq, and deletion 6q. CONCLUSIONS: Our study panned out new CXCR4 mutations in Waldenström macroglobulinemia and identified a specific signature associated to CXCR4(mut), characterized with complex genomic aberrations among MYD88L265P Waldenström macroglobulinemia. Our results suggest the existence of various genomic subgroups in Waldenström macroglobulinemia.
Asunto(s)
Estudio de Asociación del Genoma Completo , Genómica , Mutación , Receptores CXCR4/genética , Macroglobulinemia de Waldenström/genética , Alelos , Sustitución de Aminoácidos , Biomarcadores , Análisis por Conglomerados , Análisis Citogenético , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Estudio de Asociación del Genoma Completo/métodos , Genómica/métodos , Genotipo , Humanos , Inmunofenotipificación , Masculino , Fenotipo , Pronóstico , Receptores CXCR4/química , Receptores CXCR4/metabolismo , Transcriptoma , Macroglobulinemia de Waldenström/metabolismo , Macroglobulinemia de Waldenström/mortalidadRESUMEN
The pathophysiology of Waldenström macroglobulinemia (WM), a lymphoproliferative disorder characterized by lymphoplasmacytic bone marrow infiltration associated with serum IgM paraprotein, is rather unclear; however, progress has been made in recent years to better determine the genetic profile of WM tumor cells. Studies based on high-throughput genomic analyses-including single-nucleotide polymorphism array (SNPa), array-based comparative genomic hybridization, and, recently, whole-genome sequencing--have improved deciphering some of the key molecular pathways associated with WM. Beyond the discovery of the myeloid differentiation primary response gene 88 (MYD88) L265P mutation, which will help greatly in the differential characterization of WM from other B-cell low-grade lymphomas, several other mechanisms of gene deregulation were identified and mapped that recurrently pointed out nuclear factor-kappa B (NF-κB), breakpoint cluster region (BCR), and Toll-like receptor (TLR) signaling pathways as potential targets for a better understanding of the physiopathology of WM and for future drug development. Herein, we summarize the current knowledge of the genomic patterns of WM to highlight its complexity.
Asunto(s)
Genómica , Macroglobulinemia de Waldenström/genética , Epigénesis Genética , Estudio de Asociación del Genoma Completo , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Mutación , Factor 88 de Diferenciación Mieloide/genética , Disomía UniparentalRESUMEN
Immunomodulators have been used in recent years to reactivate host anti-tumor immunity in several hematological malignancies. This report describes the effect of activating natural killer T (NKT) cells by α-Galactosylceramide (α-GalCer) in the 5T33MM model of multiple myeloma (MM). NKT cells are T lymphocytes, co-expressing T and NK receptors, while invariant NKT cells (iNKTs) also express a unique semi-invariant TCR α-chain. We followed iNKT numbers during the development of the disease in both 5T33MM mice and MM patients and found that their numbers dropped dramatically at the end stage of the disease, leading to a loss of total IFN-γ secretion. We furthermore observed that α-GalCer treatment significantly increased the survival of 5T33MM diseased mice. Taken together, our data demonstrate for the first time the possibility of using a preclinical murine MM model to study the effects of α-GalCer and show promising results of α-GalCer treatment in a low tumor burden setting.
Asunto(s)
Mieloma Múltiple/inmunología , Células T Asesinas Naturales/inmunología , Animales , Antígenos CD1d/metabolismo , Modelos Animales de Enfermedad , Galactosilceramidas/administración & dosificación , Galactosilceramidas/farmacología , Humanos , Interferón gamma/metabolismo , Recuento de Linfocitos , Ratones , Mieloma Múltiple/tratamiento farmacológico , Mieloma Múltiple/metabolismo , Mieloma Múltiple/mortalidad , Células T Asesinas Naturales/efectos de los fármacos , Células T Asesinas Naturales/metabolismoRESUMEN
Although several studies have evaluated the frequency and consequences of medication errors, few have explored their causes. In particular, nurses' knowledge regarding medications has been evaluated minimally. This survey was conducted to determine how nurses master medications prescribed to their patients to determine problems nurses may have with prescribed drugs and identify possible support tools. A questionnaire was created and presented to nurses in a French cancer center. A majority of the respondents correctly identified pharmaceutical classes and medications, as well as administration and storage conditions. However, side effects, contraindications, and drug-drug interactions were not adequately identified. Nurses reported facing problems mainly related to drug administration, drug storage, and generic drugs and their therapeutic equivalence. Multiple tools are in development to help nurses in these areas. This collaborative study between pharmacy and care wards identifies some difficulties nurses have regarding drugs and will help to establish improvement measures within the hospital.