Optimized design and data analysis of tag-based cytosine methylation assays

Genome-wide, tag-based cytosine methylation analysis is optimized

Estudio epigenético del asma alérgica mediante el análisis de patrones de metilación

[ES] En esta tesis se estudia el patrón epigenético, en términos de metilación de citosinas, en los linfocitos B de pacientes con asma alérgica e individuos sanos de una forma global, describiendo todo el epigenoma e identificando loci que presenten patrones de metilación diferencial en los linfocitos B de los pacientes alérgicos respecto a los individuos del grupo control. Además se analizan posibles discrepancias en los niveles de expresión de los genes que presenten patrones de metilación diferencial entre los individuos alérgicos y los individuos sanos. Para terminar se correlaciona el estado de metilación de los genes que presenten metilación diferencial con los niveles de expresión de los mismos para identificar la presencia de posibles marcadores epigenéticos de la enfermedad.[EN] In this thesis we study the epigenetic pattern in terms of cytosine methylation in B cells of patients with allergic asthma and healthy individuals in a comprehensive, describing all the epigenome and identifying loci showing differential methylation patterns in B lymphocytes of allergic patients respect to control subjects. Furthermore, we analyze possible differences in the levels of expression of genes showing differential methylation patterns between individuals allergic and healthy individuals. Finally correlate methylation status of genes showing differential methylation levels of expression of these to identify the presence of possible epigenetic markers of the disease

Late-replicating heterochromatin is characterized by decreased cytosine methylation in the human genome

Heterochromatin is believed to be associated with increased levels of cytosine methylation. With the recent availability of genome-wide, high-resolution molecular data reflecting chromatin organization and methylation, such relationships can be explored systematically. As well-defined surrogates for heterochromatin, we tested the relationship between DNA replication timing and DNase hypersensitivity with cytosine methylation in two human cell types, unexpectedly finding the later-replicating, more heterochromatic regions to be less methylated than early replicating regions. When we integrated gene-expression data into the study, we found that regions of increased gene expression were earlier replicating, as previously identified, and that transcription-targeted cytosine methylation in gene bodies contributes to the positive correlation with early replication. A self-organizing map (SOM) approach was able to identify genomic regions with early replication and increased methylation, but lacking annotated transcripts, loci missed in simple two variable analyses, possibly encoding unrecognized intergenic transcripts. We conclude that the relationship of cytosine methylation with heterochromatin is not simple and depends on whether the genomic context is tandemly repetitive sequences often found near centromeres, which are known to be heterochromatic and methylated, or the remaining majority of the genome, where cytosine methylation is targeted preferentially to the transcriptionally active, euchromatic compartment of the genome

Venetoclax improves CD20 immunotherapy in a mouse model of MYC/BCL2 double-expressor diffuse large B-cell lymphoma

BackgroundApproximately one-third of diffuse large B cell lymphoma (DLBCL) patients exhibit co-expression of MYC and BCL2 (double-expressor lymphoma, DEL) and have a dismal prognosis. Targeted inhibition of the anti-apoptotic protein BCL2 with venetoclax (ABT-199) has been approved in multiple B-cell malignancies and is currently being investigated in clinical trials for DLBCL. Whether BCL2 anti-apoptotic function represents a multifaceted vulnerability for DEL-DLBCL, affecting both lymphoma B cells and T cells within the tumor microenvironment, remains to be elucidated.MethodsHere, we present novel genetically engineered mice that preclinically recapitulate DEL-DLBCL lymphomagenesis, and evaluate their sensitivity ex vivo and in vivo to the promising combination of venetoclax with anti-CD20-based standard immunotherapy.ResultsVenetoclax treatment demonstrated specific killing of MYC+/BCL2(+) lymphoma cells by licensing their intrinsically primed apoptosis, and showed previously unrecognized immunomodulatory activity by specifically enriching antigen-activated effector CD8 T cells infiltrating the tumors. Whereas DEL-DLBCL mice were refractory to venetoclax alone, inhibition of BCL2 significantly extended overall survival of mice that were simultaneously treated with a murine surrogate for anti-CD20 rituximab.ConclusionsThese results suggest that the combination of anti-CD20-based immunotherapy and BCL2 inhibition leads to cooperative immunomodulatory effects and improved preclinical responses, which may offer promising therapeutic opportunities for DEL-DLBCL patients