An Efficient Protocol For The Generation Of Monocyte Derived Dendritic Cells Using Serum-free Media For Clinical Applications In Post Remission Aml Patients.

Protocols for the generation of dendritic cells (DCs) using serum as a supplementation of culture media leads to reactions due to animal proteins and disease transmissions. Several types of serum-free media (SFM), based on good manufacture practices (GMP), have recently been used and seem to be a viable option. The aim of this study was to evaluate the results of the differentiation, maturation, and function of DCs from Acute Myeloid Leukemia patients (AML), generated in SFM and medium supplemented with autologous serum (AS). DCs were analyzed by phenotype characteristics, viability, and functionality. The results showed the possibility of generating viable DCs in all the conditions tested. In patients, the X-VIVO 15 medium was more efficient than the other media tested in the generation of DCs producing IL-12p70 (p=0.05). Moreover, the presence of AS led to a significant increase of IL-10 by DCs as compared with CellGro (p=0.05) and X-Vivo15 (p=0.05) media, both in patients and donors. We concluded that SFM was efficient in the production of DCs for immunotherapy in AML patients. However, the use of AS appears to interfere with the functional capacity of the generated DCs.

Cartografia geológica do Mesozóico na AML e aspectos relativos à normalização

Cartografia Geológica Aplicada a Áreas Urbanas

Critérios para o planeamento de equipamentos de saúde. Análise de Caso de Estudo no contexto urbano da AML

Dissertação para a obtenção do Grau de Mestre em Engenharia Civil – Perfil Construção

Influência dos tarifários de transportes colectivos na repartição modal dos transportes na AML

Dissertação para obtenção do Grau de Mestre em Engenharia do Ambiente, Perfil de Ordenamento do Território e Impactes Ambientais

Biologische Charakterisierung von unterschiedlichen FLT3 internen Tandemduplikationen (ITD) in der akuten myeloischen Leukämie (AML)

Magdeburg, Univ., Fak. für Naturwiss., Diss., 2014

IDH1/IDH2 mutations predict survival in glioma and AML

Mutations in the isocitrate dehydrogenase family genes 1 or 2 (IDH1/2) have been discovered by high through put sequencing approaches inglioma and acute myeloid leukemia (AML) and related myeloproliferativeneoplasms. In both diseases, the discovery of IDH mutations has identifieda prognostically new subtype with distinct pathogenetic evolution. Ingliomas mutations are mostly found in IDH1 (>90%). They are infrequent inprimary glioblastoma (GBM) (<10%), but common in secondary GBM thatevolve from lower grade glioma (60−90%). Mutations in IDH1 precede p53mutations or 1p/19q co-deletions in sporadic low grade glioma, hence arean early evant. Co-deletions of 1p/19q, characteristic for oligodenroglioma,are highly associated with IDH1/2 mutations, while they are mutuallyexclusive with EGFR amplifications, a hall mark of primary GBM. IDH1 or 2mutations are associated with younger patient age, but absent in childhoodgliomas, and have a better prognosis that seems to be consistent in gradeII through IV gliomas. In myeloid malignancies mutations are more likelyin IDH2 and are found in de novo and secondary AML (12−18%) andpre-leukemic clonal malignancies (5% chronic; 20% transformed). IDH1/2mutations are strongly associated with NPM1 mutations that are found in30% of novo cytogenetically normal AML. In CN-AML with mutated NPM1,without FLT3 internal tandem duplication (ITD) IDH mutations constitutean adverse prognostic factor. Mutations in the metabolic enzymes IDH1 or2 result in a neomorphic reaction, generating high levels of the metabolite2-hydroxyglutarate (2-HG). IDH mutations are mutually exclusive with TET2mutations in myeloid malignancies that led to the discovery that high levelsof 2-HG inhibit the a-KG dependent dioxygenase TET2. TET2 is involved inepigenetic regulation and mediates demethylation of DNA. This mechanismis in accordance with the association of a methylator phenotype with loss offunction of TET2 by mutation or indirectly by mutation of IDH1/2 in myeloidmalignancies and gliomas, respectively.Metabolism meets Epigenetics. These discoveries will have importantclinical implications: IDH1/2 mutants may serve as unique targets fortherapy. Further, the high concentrations of the onco-metabolite 2-HGgenerated by IDH1/2 mutants, may serve as biomarker in the serum ofpatients with myeloid malignancies and may be amenable by magneticresonance spectroscopy in glioma patients.

Programa Helios, AML de Oviedo : informe de proceso, sondeo de los materiales

Valorar el grado de conocimiento, disponibilidad y uso de los materiales por los profesionales de las diferentes instituciones. Animar al registro de datos en los protocolos de evaluaci??n. Organismos: Centro de Servicios Sociales del Ayuntamiento de Oviedo, Instituto de Servicios Sociales, 4 Asociaciones de Padres, 5 Equipos psicopedag??gicos, 5 Servicios de Minusv??lidos y 6 de otros Servicios de la Consejer??a de Sanidad y Servicios Sociales, 28 centros p??blicos de educaci??n y 14 privados, Unidad Pedag??gica del hospital. Tres meses despu??s del env??o de los materiales desarrollados por la AML de Oviedo ('Anexo al documento de salud infantil' y 'Gu??a de Recursos'), se realiza un sondeo de utilizaci??n de los mismos para saber si se utilizan, si se dispone de los protocolos de evaluaci??n y en caso afirmativo, con que frecuencia se utilizan. Protocolos de evaluaci??n de materiales. La informaci??n sobre el 'Anexo al documento de salud infantil' la proporcionan 23 servicios, de los cuales 8 no lo hab??an recibido y 16 no lo utilizaban. El protocolo de evaluaci??n de este material no lo hab??an recibido 15 servicios, y de los 8 restantes s??lo 1 lo hab??a utilizado. La distribuci??n del 'Anexo al documento de salud infantil' parece haber tenido dos niveles: uno de divulgaci??n hacia todos los equipos y servicios que atienden a las personas discapacitadas y otro, m??s espec??fico, de puesta en funcionamiento. Los servicios que no utilizan el anexo, en su mayor??a, desconocen qu?? han de hacer con ??l. Con respecto al protocolo, parece que no ha sido distribuido conjuntamente con el anexo, ya que hay servicios que dicen haber recibido ??ste y no el protocolo. La informaci??n sobre la 'Gu??a de recursos' ha sido recogida en 65 servicios, equipos y centros educativos, de los cuales 23 dicen no haber recibido el material y 18 no utilizarlo. El protocolo de evaluaci??n no hab??a sido recibido por 40 de ellos y de los 25 restantes s??lo 2 dicen estar registrando datos en ??l. Parece ser que el criterio de distribuci??n de la Gu??a no est?? muy claro ya que algunos centros espec??ficos de Educaci??n Especial o Integraci??n no disponen de la Gu??a; en tanto, otros centros sin alumnado con Necesidades Educativas Especiales s?? la han recibido. Los autores creen que deber??an adoptarse las siguientes medidas: 1) reponer urgentemente los materiales en aquellos centros, equipos o servicios que no lo han recibido; 2) enviar los protocolos de evaluaci??n correspondientes; 3) proceder a una informaci??n m??s directa sobre el uso y la utilidad de estos materiales Helios; 4) instar al INSALUD a que los profesionales de este organismos pongan en marcha los materiales Helios.

Proyecto Helios, AML Oviedo : informe evaluador de la fase de contexto

Analizar la interconexi??n de centros, servicios, equipos e instituciones. 29 centros escolares, 6 asociaciones, 3 equipos psicopedag??gicos y 5 servicios de salud. En cada uno de los centros se analizan: 1) los niveles de atencion al minusv??lido en cuanto a prevenci??n, valoraci??n y tratamiento; 2) los niveles de informaci??n de entrada, proceso y salida; 3) los niveles de coordinaci??n intra e intersectorial. Entrevista realizada por encuestadores previamente preparados por el equipo evaluador. Se recoge la informaci??n siguiendo la pauta y el protocolo conformado en un cuestionario. 1) Centros escolares: en el nivel de atenci??n se observa cierta carencia de motivaci??n de cara a la intervenci??n; se detecta una carencia general en la sistematizaci??n de la informaci??n interna y la coordinaci??n; tambi??n se observan problemas de coordinaci??n inter e intrasector. 2) Asociaciones: la atenci??n preventiva que ofrecen es b??sicamente asistencial y la atenci??n valorativa y de intervenci??n son diferentes seg??n los recursos de que disponen; a nivel de informaci??n destacan el asesoramiento y gesti??n de recursos, el control de datos y la custodia de documentaci??n y casu??stica; la principal coordinaci??n detectada es la de cada asociaci??n con las instituciones que le aportan recursos econ??micos y con las federaciones o confederaciones que se ocupan de las mismas minusval??as. 3) Equipos psicopedag??gicos: a nivel de atenci??n se centran en el apoyo y asesoramiento t??cnico al profesorado, as?? como en la valoraci??n diagn??stica y en la elaboraci??n de informes de identificaci??n de Necesidades Educativas Especiales y/o minusval??as; se aprecia un nivel de informaci??n amplia en los tres momentos del proceso; la coordinaci??n, si bien existe y es considerable, funciona a nivel de contactos personales y de buena voluntad por parte de los integrantes de los equipos. 4) Servicios de salud: todos ponen en marcha programas de prevenci??n y detecci??n precoz de minusval??as pero no en todos se identifica un mismo nivel de tratamiento y seguimiento; el nivel de informaci??n es muy variable; no existe un programa de atenci??n coordinada, las v??as de comunicaci??n no est??n establecidas y las relaciones se establecen puntualmente. Para configurar una red de coordinaci??n integral operativa es fundamental establecer unos cauces de comunicaci??n y discusi??n permanentes que consigan mantener una v??a abierta a la informaci??n y formaci??n de todas aquellas personas que van a estar en contacto con el minusv??lido. Para ello ser?? preciso considerar los tres niveles de atenci??n (familiar-escolar-ocupacional), que delimitan el campo de intervenci??n y las acciones de todos los sectores implicados.

Evidence for complex multigenic inheritance of radiation AML susceptibility in mice revealed using a surrogate phenotypic assay

The mapping of genes which affect individual cancer risk is an important but complex challenge. A surrogate assay of susceptibility to radiation-induced acute myeloid leukaemia (AML) in the mouse based on chromosomal radiosensitivity has been developed and validated. This assay was applied to the mapping of radiation-induced AML risk modifier loci by association with microsatellite markers. A region on chromosome (chr) 18 with strong association is identified and confirmed by backcross analysis. Additional loci on chrs 8 and 13 show significant association. A key candidate gene Rbbp8 on chr18 is identified. Rbbp8 is shown to be upregulated in response to X-irradiation in the AML sensitive CBA strain but not AML resistant C57BL/6 strain. This study demonstrates the strength of utilizing surrogate endpoints of cancer susceptibility in the mapping of mouse loci and identifies additional loci that may affect radiation cancer risk.

In vitro characterization of human AML-reactive CTL clones generated from the naive subset of healthy donors and adoptive transfer into leukemia-engrafted NSG mice

Acute myeloid leukemia (AML) is a very aggressive cancer of the hematopoietic system. Chemotherapy and immunotherapeutical approaches including hematopoietic stem cell transplantation (HSCT) and donor lymphocyte infusion (DLI) are the only curative options available. The beneficial graft-versus-leukemia (GVL) effect of cellular immunotherapy is mostly mediated by donor-derived CD8+ T lymphocytes that recognize minor histocompatibility antigens (mHags) and leukemia-associated antigens (LAAs) presented on the surface of AML blasts (Falkenburg et al. 2008; Kolb 2008). A main complication is graft-versus-host disease (GVHD) that can be induced when cytotoxic T lymphocytes (CTLs) recognize broadly expressed antigens. To reduce the risk of GVHD, specific allogeneic T-cell therapy inducing selective GVL responses could be an option (Barrett &amp; Le Blanc 2010; Parmar et al. 2011; Smits et al. 2011). This requires efficient in vitro strategies to generate AML-reactive T cells with an early differentiation phenotype as well as vigorous effector functions and humanized mouse models to analyze the anti-leukemic potential of adoptively transferred T cells in vivo. In this study, AML-reactive CTL clones and oligoclonal T-cell lines could be reliably generated from the naive subset of healthy HLA-class I-identical donors by stimulation with primary AML blasts in mini-mixed-lymphocyte / leukemia cultures (MLLCs) in eight different patient / donor pairs. These CTLs were promising candidates for cellular immunotherapy because of their relatively early differentiation phenotype and strong proliferative and lytic capabilities. The addition of the common γ-chain cytokine IL-21 to the stimulation protocol enabled more precursors to develop into potent leukemia-reactive CTLs, presumably by its beneficial effects on cell survival and antigen-specific proliferation during the first weeks of cultures. It also strengthened the early-stage phenotype. Three long-term cultured CTLs exemplarily transferred into leukemia-engrafted immunodeficient NSG mice mediated a significant reduction of the leukemic burden after a single transfusion. These results demonstrate that CTL clones with reactivity to patient-derived AML blasts can be isolated from the naive compartment of healthy donors and show potent anti-leukemic effects in vivo. The herein described allo-MLLC approach with in vitro “programmed” naive CTL precursors independent of a HSCT setting is a valuable alternative to the conventional method of isolating in vivo primed donor CTLs out of patients after transplantation (Kloosterboer et al. 2004; Warren et al. 2010). This would make leukemia-reactive CTLs already available at the time point of HSCT, when residual leukemia disease is minimal and the chances for complete leukemia eradication are high. Furthermore, leukemia-reactive CTLs effectively expanded by this in vitro protocol can be used as screening populations to identify novel candidate LAAs and mHags for antigen-specific immunotherapy.

Complexity of miR-223 regulation by CEBPA in human AML

microRNA-223 (miR-223) can trigger normal granulopoiesis. miR-223 expression is regulated by two distinct CEBPA (CCAAT/enhancer binding protein-alpha) sites. Here, we report that miR-223 is largely suppressed in cells from acute myeloid leukemia (AML) patients. By sequencing, we found that miR-223 suppression in AML is not caused by DNA sequence alterations, nor is it mediated by promoter hypermethylation. The analysis of the individual contribution of both CEBPA sites to miR-223 regulation identified the site upstream of the miR-223 primary transcript as the predominant regulatory element. Our results suggest that miR-223 suppression in AML is caused by impaired miR-223 upstream factors.

Heterozygous deletion of the PU.1 locus in human AML

The transcription factor PU.1 is essential for myeloid development. Targeted disruption of an upstream regulatory element (URE) decreases PU.1 expression by 80% and leads to acute myeloid leukemia (AML) in mice. Here, we sequenced the URE sequences of PU.1 in 120 AML patients. Four polymorphisms (single nucleotide polymorphisms [SNPs]) in the URE were observed, with homozygosity in all SNPs in 37 patients. Among them, we compared samples at diagnosis and remission, and one patient with cytogenetically normal acute myeloid leukemia M2 was identified with heterozygosity in 3 of the SNPs in the URE at remission. Loss of heterozygosity was further found in this patient at 2 polymorphic sites in the 5' promoter region and in 2 intronic sites flanking exon 4, thus suggesting loss of heterozygosity covering at least 40 kb of the PU.1 locus. Consistently, PU.1 expression in this patient was markedly reduced. Our study suggests that heterozygous deletion of the PU.1 locus can be associated with human AML.

Lineage-specific transcription factor aberrations in AML

Transcription factors play a key role in the commitment of hematopoietic stem cells to differentiate into specific lineages [78]. This is particularly important in that a block in terminal differentiation is the key contributing factor in acute leukemias. This general theme of the role of transcription factors in differentiation may also extend to other tissues, both in terms of normal development and cancer. Consistent with the role of transcription factors in hematopoietic lineage commitment is the frequent finding of aberrations in transcription factors in AML patients. Here, we intend to review recent findings on aberrations in lineage-restricted transcription factors as observed in patients with acute myeloid leukemia (AML).