Estudio de perfiles moleculares en pacientes con síndrome mielodisplásico

Los SÍNDROMES MIELODISPLÁSICOS (SMD) son una enfermedad heterogénea clonal de células madre hematopoyéticas. Se caracteriza por una medula ósea ineficaz que produce citopenias en sangre periférica. La severidad de estas citopenias y el riesgo incrementado a progresar a una leucemia mieloide aguda determinarán la evolución de la enfermedad y del paciente. Actualmente estos pacientes se clasifican de acuerdo con 7 subcategorías que establece la revisión del 2008 de la clasificación de la WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues. La alta heterogeneidad de cada subcategoría ha hecho necesarios sistemas de puntuación, el más utilizado es el IPSS-R (International Prognostic Scoring System) e incluye 3 variables, las citopenias en sangre periférica, el porcentaje de blastos en médula ósea y el cariotipo. Nos permite evaluar la supervivencia global (SG) y la probabilidad de progresión a leucemia mieloide aguda. La hipótesis global del trabajo es que alteraciones detectadas mediante técnicas moleculares permiten mejorar la definición del pronóstico de pacientes con SMD. Para dilucidar la hipótesis general, se han estudiado tres grupos de pacientes con cariotipos distintos: pacientes con SMD y cariotipo monosómico y/o complejo, pacientes con el cromosoma 7 alterado mediante FISH y pacientes con deleción 5q [del(5q)]. Pacientes con cariotipo monosómico y con alteraciones en el cromosoma 7 se incluyen en el grupo de SMD de alto riesgo. En cambio, pacientes con del(5q), cuando esta se detecta de manera aislada, se incluyen en el grupo de SMD de bajo riesgo. La del(5q) en un cariotipo complejo pasa a ser de pronóstico desfavorable. Primero analizamos el efecto del cariotipo monosómico (MK). Estudiamos 1.054 pacientes con SMD y cariotipo alterado con el objetivo de describir la incidencia, características y pronóstico del MK, y su relación con la SG y el riesgo de progresión a LMA. Este estudio ha permitido determinar que el mal pronóstico asociado a los pacientes con MK, en el contexto de un cariotipo complejo, se debe a la complejidad en el cariotipo lo que les confiere un peor pronóstico, es decir peor SG y mayor riesgo de progresión a LMA. Segundo, estudiamos 820 pacientes por FISH con SMD, su cariotipo no mostró la monosomía 7 o la deleción 7q. El objetivo del estudio era determinar si la detección mediante FISH de estas alteraciones afectaba a su clasificación. El 5,2% de los casos fueron positivos, esto es de importante relevancia ya que la -7 o la del(7q) se relacionan con un impacto pronóstico negativo. Este estudio nos permitió detectar diferencias significativas en la SG de pacientes con riesgo morfológico intermedio (WHO 2008) FISH positivos y negativos. Debido a la relación de estas alteraciones del cromosoma 7 y un peor pronóstico, sería altamente recomendable analizar mediante FISH las alteraciones que no cumplan criterios de clonalidad.a El último grupo de pacientes que estudiamos presentaban la del(5q). Estudiamos un total de 228 pacientes con neoplasias mieloides (SMD y LMA) y del(5q). Mediante el estudio por secuenciación masiva del exoma pudimos determinar los 6 genes más mutados en nuestra serie: TP53, DNMT3A, CSNK1A1, SF3B1, PRPF8 y ASXL1. TP53 y DNMT3A presentaron un efecto negativo en la SG de los pacientes, pero sólo DNMT3A, retuvo la influencia independiente en la SG, en el análisis multivariado. El perfil mutacional de pacientes con del(5q) no se ha podido determinar. Estudios iniciales en pacientes con CSNK1A1 mutado, por su relación con una buena respuesta a la lenalidomida. Los pacientes con TP53 deberían ser controlados de manera frecuente, por el impacto negativo en la SG de mutaciones en este gen. Y, por último, mutaciones en DNMT3A deberían estudiarse en más pacientes con del(5q) para esclarecer su relación.MYELODYSPLASTIC SYNDROMES are clonal hematopoietic stem cell disorders highly heterogeneous. This disease is characterized by bone marrow failure that led to peripheral blood citopenias. The outcome of these patients is closely related to cytopenias and to an increased risk to acute myeloid leukemia (AML) progression. Nowadays, these patients are classified according to the revised version (2008) of the WHO Classification of Tumours of Haematopoietic and Lymphoid Tissues, which includes 7 subcategories. Due to high heterogeneity, the well-known scoring system IPSS-R (International Prognostic Scoring System) have been developed to better assess the overall survival (OS) their risk to progress. IPSS-R includes 3 variables, cytopenias at peripheral blood, bone marrow blast percentage and karyotype. The last one has shown a high influence in the IPSS-R. We hypothesize that alterations detected by molecular techniques could help us to better define patients with MDS. To elucidate this hypothesis, we studied three groups of MDS patients with different karyotypic alterations, patients with MDS and monosomal karyotype (MK), patients with alterations in chromosome 7 by FISH and patients with MDS or AML and del(5q). MK and alterations of chromosome 7 are included in high risk MDS, whether patients with isolated del(5q) are included in low risk MDS. However, when the alteration is detected in a complex karyotype, the outcome is significantly worse. These three group of patients were studied separately. First we studied the effect of MK. We studied a total of 1,054 adult patients with MDS and altered karyotype. Our main objective was to describe the incidence, characteristics and prognosis of patients with MK and its relation with OS and the progression risk to AML. This study helped us to determine that the worse prognosis related to MK (included in a complex karyotype) is karyotype complexity. This is the one that have a negative impact on patient's prognosis with lower OS and higher risk to AML progression. Second, we studied by FISH a total of 820 patients with MDS without monosomy 7 or 7q deletion by conventional cytogenetics. Our objective was to define the impact of FISH detection (7- or 7q-) in the outcome of MDS patients. A total of 5.2% of cases were positive by FISH. This is of note because -7 and 7q- are related with worse outcome and more aggressive treatments. Moreover patients with an intermediate morphologic risk (WHO 2008) would benefit from a FISH study in order to better classify their prognosis. Due to chromosome 7 alterations worse prognosis, it would be highly recommended to apply FISH 7q in patients with chromosome 7 alterations by conventional cytogenetics who doesn't fulfill clonally criteria. Last group studied were del(5q) patients, this alteration is one of the most common in MDS. We collected 228 patients with del(5q) with myeloid neoplasms (MDS and AML) by applying next-generation sequencing techniques. We were able to determine the 6 genes most commonly mutated: TP53, DNMT3A, CSNK1A1, SF3B1, PRPF8 and ASXL1. Both TP53 and DNMT3A were related to worse prognosis. TP53 and DNMT3A had a negative impact on the OS, but only DNMT3A retained its prognostic impact on the multivariate analysis. Although we couldn't define a mutational profile for patients with del(5q), it would be important to perform initial studies to define patients with a better outcome. Especially when CSNK1A1 is mutated for its correlation with a good lenalidomide's treatment response. In contrast, patients with mutations in TP53 should be closely followed for its well-known negative impact on OS and lenalidomide response. Moreover, due to the high negative impact of DNMT3A mutations in our series. Further studies are needed to clarify its impact on del(5q) patients

The MicroRNA-200 Family Is Upregulated in Endometrial Carcinoma

BACKGROUND: MicroRNAs (miRNAs, miRs) are small non-coding RNAs that negatively regulate gene expression at the post-transcriptional level. MicroRNAs are dysregulated in cancer and may play essential roles in tumorigenesis. Additionally, miRNAs have been shown to have prognostic and diagnostic value in certain types of cancer. The objective of this study was to identify dysregulated miRNAs in endometrioid endometrial adenocarcinoma (EEC) and the precursor lesion, complex atypical hyperplasia (CAH). METHODOLOGY: We compared the expression profiles of 723 human miRNAs from 14 cases of EEC, 10 cases of CAH, and 10 normal proliferative endometria controls using Agilent Human miRNA arrays following RNA extraction from formalin-fixed paraffin-embedded (FFPE) tissues. The expression of 4 dysregulated miRNAs was validated using real time reverse transcription-PCR. RESULTS: Forty-three miRNAs were dysregulated in EEC and CAH compared to normal controls (p<0.05). The entire miR-200 family (miR-200a/b/c, miR-141, and miR-429) was up-regulated in cases of EEC. CONCLUSIONS: This information contributes to the candidate miRNA expression profile that has been generated for EEC and shows that certain miRNAs are dysregulated in the precursor lesion, CAH. These miRNAs in particular may play important roles in tumorigenesis. Examination of miRNAs that are consistently dysregulated in various studies of EEC, like the miR-200 family, will aid in the understanding of the role that miRNAs play in tumorigenesis in this tumour type

The Lipopolysaccharide Core of Brucella abortus Acts as a Shield Against Innate Immunity Recognition

Innate immunity recognizes bacterial molecules bearing pathogen-associated molecular patterns to launch inflammatory responses leading to the activation of adaptive immunity. However, the lipopolysaccharide (LPS) of the gram-negative bacterium Brucella lacks a marked pathogen-associated molecular pattern, and it has been postulated that this delays the development of immunity, creating a gap that is critical for the bacterium to reach the intracellular replicative niche. We found that a B. abortus mutant in the wadC gene displayed a disrupted LPS core while keeping both the LPS O-polysaccharide and lipid A. In mice, the wadC mutant induced proinflammatory responses and was attenuated. In addition, it was sensitive to killing by non-immune serum and bactericidal peptides and did not multiply in dendritic cells being targeted to lysosomal compartments. In contrast to wild type B. abortus, the wadC mutant induced dendritic cell maturation and secretion of pro-inflammatory cytokines. All these properties were reproduced by the wadC mutant purified LPS in a TLR4-dependent manner. Moreover, the core-mutated LPS displayed an increased binding to MD-2, the TLR4 co-receptor leading to subsequent increase in intracellular signaling. Here we show that Brucella escapes recognition in early stages of infection by expressing a shield against recognition by innate immunity in its LPS core and identify a novel virulence mechanism in intracellular pathogenic gram-negative bacteria. These results also encourage for an improvement in the generation of novel bacterial vaccines

Diversity and dynamics of rare and of resident bacterial populations in coastal sands

Coastal sands filter and accumulate organic and inorganic materials from the terrestrial and marine environment, and thus provide a high diversity of microbial niches. Sands of temperate climate zones represent a temporally and spatially highly dynamic marine environment characterized by strong physical mixing and seasonal variation. Yet little is known about the temporal fluctuations of resident and rare members of bacterial communities in this environment. By combining community fingerprinting via pyrosequencing of ribosomal genes with the characterization of multiple environmental parameters, we disentangled the effects of seasonality, environmental heterogeneity, sediment depth and biogeochemical gradients on the fluctuations of bacterial communities of marine sands. Surprisingly, only 3–5% of all bacterial types of a given depth zone were present at all times, but 50–80% of them belonged to the most abundant types in the data set. About 60–70% of the bacterial types consisted of tag sequences occurring only once over a period of 1 year. Most members of the rare biosphere did not become abundant at any time or at any sediment depth, but varied significantly with environmental parameters associated with nutritional stress. Despite the large proportion and turnover of rare organisms, the overall community patterns were driven by deterministic relationships associated with seasonal fluctuations in key biogeochemical parameters related to primary productivity. The maintenance of major biogeochemical functions throughout the observation period suggests that the small proportion of resident bacterial types in sands perform the key biogeochemical processes, with minimal effects from the rare fraction of the communities

Real-time plasma state monitoring and supervisory control on TCV

In ITER and DEMO, various control objectives related to plasma control must be simultaneously achieved by the plasma control system (PCS), in both normal operation as well as off-normal conditions. The PCS must act on off-normal events and deviations from the target scenario, since certain sequences (chains) of events can precede disruptions. It is important that these decisions are made while maintaining a coherent prioritization between the real-time control tasks to ensure high-performance operation. In this paper, a generic architecture for task-based integrated plasma control is proposed. The architecture is characterized by the separation of state estimation, event detection, decisions and task execution among different algorithms, with standardized signal interfaces. Central to the architecture are a plasma state monitor and supervisory controller. In the plasma state monitor, discrete events in the continuous-valued plasma state are modeled using finite state machines. This provides a high-level representation of the plasma state. The supervisory controller coordinates the execution of multiple plasma control tasks by assigning task priorities, based on the finite states of the plasma and the pulse schedule. These algorithms were implemented on the TCV digital control system and integrated with actuator resource management and existing state estimation algorithms and controllers. The plasma state monitor on TCV can track a multitude of plasma events, related to plasma current, rotating and locked neoclassical tearing modes, and position displacements. In TCV experiments on simultaneous control of plasma pressure, safety factor profile and NTMs using electron cyclotron heating (ECH) and current drive (ECCD), the supervisory controller assigns priorities to the relevant control tasks. The tasks are then executed by feedback controllers and actuator allocation management. This work forms a significant step forward in the ongoing integration of control capabilities in experiments on TCV, in support of tokamak reactor operation

Langmuir probe electronics upgrade on the tokamak a configuration variable

A detailed description of the Langmuir probe electronics upgrade for TCV (Tokamak a Configuration Variable) is presented. The number of amplifiers and corresponding electronics has been increased from 48 to 120 in order to simultaneously connect all of the 114 Langmuir probes currently mounted in the TCV divertor and main-wall tiles. Another set of 108 amplifiers is ready to be installed in order to connect 80 new probes, built in the frame of the TCV divertor upgrade. Technical details of the amplifier circuitry are discussed as well as improvements over the first generation of amplifiers developed at SPC (formerly CRPP) in 1993/1994 and over the second generation developed in 2012/2013. While the new amplifiers have been operated successfully for over a year, it was found that their silicon power transistors can be damaged during some off-normal plasma events. Possible solutions are discussed. (C) 2019 Author(s)

Why Are Outcomes Different for Registry Patients Enrolled Prospectively and Retrospectively? Insights from the Global Anticoagulant Registry in the FIELD-Atrial Fibrillation (GARFIELD-AF).

Background: Retrospective and prospective observational studies are designed to reflect real-world evidence on clinical practice, but can yield conflicting results. The GARFIELD-AF Registry includes both methods of enrolment and allows analysis of differences in patient characteristics and outcomes that may result. Methods and Results: Patients with atrial fibrillation (AF) and ≥1 risk factor for stroke at diagnosis of AF were recruited either retrospectively (n = 5069) or prospectively (n = 5501) from 19 countries and then followed prospectively. The retrospectively enrolled cohort comprised patients with established AF (for a least 6, and up to 24 months before enrolment), who were identified retrospectively (and baseline and partial follow-up data were collected from the emedical records) and then followed prospectively between 0-18 months (such that the total time of follow-up was 24 months; data collection Dec-2009 and Oct-2010). In the prospectively enrolled cohort, patients with newly diagnosed AF (≤6 weeks after diagnosis) were recruited between Mar-2010 and Oct-2011 and were followed for 24 months after enrolment. Differences between the cohorts were observed in clinical characteristics, including type of AF, stroke prevention strategies, and event rates. More patients in the retrospectively identified cohort received vitamin K antagonists (62.1% vs. 53.2%) and fewer received non-vitamin K oral anticoagulants (1.8% vs . 4.2%). All-cause mortality rates per 100 person-years during the prospective follow-up (starting the first study visit up to 1 year) were significantly lower in the retrospective than prospectively identified cohort (3.04 [95% CI 2.51 to 3.67] vs . 4.05 [95% CI 3.53 to 4.63]; p = 0.016). Conclusions: Interpretations of data from registries that aim to evaluate the characteristics and outcomes of patients with AF must take account of differences in registry design and the impact of recall bias and survivorship bias that is incurred with retrospective enrolment. Clinical Trial Registration: - URL: http://www.clinicaltrials.gov . Unique identifier for GARFIELD-AF (NCT01090362)