Publications

<p>No change in memory naive proportions and their DNA methylation in relevant genes</p>

<p>Sensitivity to ibrutinib was evaluated by MTT assay (72h). Karpas1718 parental (top) or resistant (bottom) were exposed to 100nM decitabine (A), 5µM tazemetostat (B), or DMSO (control, black) given concomitantly (continuous lines) or 72 hours before ibrutinib (pre DMSO in dashed black or pre decitabine/tazemetostat in dashed red). Error bars correspond to standard deviation of the mean. Data derived from two independent experiments. Statistically significant differences were evaluated by moderated t-test comparing each treatment to control (DMSO).</p>

The use of epithelial cell adhesion molecule (EPCAM) immunohistochemistry (IHC) is not included in the colorectal cancer (CRC) screening algorithm to detect Lynch syndrome (LS) patients. The aim of the present study was to demonstrate that EPCAM IHC is a useful tool to guide the LS germ-line analysis when a loss of MSH2 expression was present. We retrospectively studied MSH2 and EPCAM IHC in a large series of 190 lesions composed of malignant neoplasms (102), precursor lesions of gastrointestinal (71) and extra-gastrointestinal origin (9), and benign neoplasms (8) from different organs of 71 patients suspicious of being LS due to MSH2 alterations. LS was confirmed in 68 patients, 53 with MSH2 mutations and 15 with EPCAM 3′-end deletions. Tissue microarrays were constructed with human normal tissues and their malignant counterparts to assist in the evaluation of EPCAM staining. Among 154 MSH2-negative lesions, 17 were EPCAM-negative, including 10 CRC and 7 colorectal polyps, and 5 of them showed only isolated negative glands. All lesions showing a lack of EPCAM expression belonged to patients with EPCAM 3′-end deletions. EPCAM IHC is a useful screening tool, with 100% specificity to identify LS patients due to EPCAM 3′-end deletions in MSH2-negative CRC and MSH2-negative colorectal polyps.

Introduction Loss-of-function TLR7 variants have been recently reported in a small number of males to underlie strong predisposition to severe COVID-19. We aimed to determine the presence of these rare variants in young men with severe COVID-19. Methods We prospectively studied males between 18 and 50 years-old without predisposing comorbidities that required at least high-flow nasal oxygen to treat COVID-19. The coding region of TLR7 was sequenced to assess the presence of potentially deleterious variants. Results TLR7 missense variants were identified in two out of 14 patients (14.3%). Overall, the median age was 38 (IQR 30-45) years. Both variants were not previously reported in population control databases and were predicted to be damaging by in silico predictors. In a 30-year-old patient a maternally inherited variant [c.644A>G; p.(Asn215Ser)] was identified, co-segregating in his 27-year-old brother who also contracted severe COVID-19. A second variant [c.2797T>C; p.(Trp933Arg)] was found in a 28-year-old patient, co-segregating in his 24-year-old brother who developed mild COVID-19. Functional testing of this variant revealed decreased type I and II interferon responses in peripheral mononuclear blood cells upon stimulation with the TLR7 agonist imiquimod, confirming a loss-of-function effect. Conclusions This study supports a rationale for the genetic screening for TLR7 variants in young men with severe COVID-19 in the absence of other relevant risk factors. A diagnosis of TLR7 deficiency could not only inform on treatment options for the patient, but also enables pre-symptomatic testing of at-risk male relatives with the possibility of instituting early preventive and therapeutic interventions.

Abstract Cardiac angiosarcoma (CAS) is a rare malignant tumour whose genetic basis is unknown. Here we show, by whole-exome sequencing of a TP53 -negative Li–Fraumeni-like (LFL) family including CAS cases, that a missense variant (p.R117C) in POT1 ( protection of telomeres 1 ) gene is responsible for CAS. The same gene alteration is found in two other LFL families with CAS, supporting the causal effect of the identified mutation. We extend the analysis to TP53 -negative LFL families with no CAS and find the same mutation in a breast AS family. The mutation is recently found once in 121,324 studied alleles in ExAC server but it is not described in any other database or found in 1,520 Spanish controls. In silico structural analysis suggests how the mutation disrupts POT1 structure. Functional and in vitro studies demonstrate that carriers of the mutation show reduced telomere-bound POT1 levels, abnormally long telomeres and increased telomere fragility.

Table S11. Table of pairwise Pearson correlations between CpG methylation values of pre-selected markers. (XLSX 63â kb)

<p>Differentially methylated levels in neighboring CpGs on the selected candidate genes.</p>

There are few validated biomarkers that can be used to predict outcomes for patients with colorectal cancer. Part of the challenge is the genetic and molecular heterogeneity of colorectal tumors not only among patients, but also within tumors. We have explored intratumor heterogeneity at the epigenetic level, due to its dynamic nature. We analyzed DNA methylation profiles of the digestive tract surface and the central bulk and invasive front regions of colorectal tumors.We determined the DNA methylation profiles of >450,000 CpG sites in 3 macrodissected regions of 79 colorectal tumors and 23 associated liver metastases, obtained from 2 hospitals in Spain. We also analyzed samples for KRAS and BRAF mutations, 499,170 single nucleotide polymorphisms, and performed immunohistochemical analyses.We observed differences in DNA methylation among the 3 tumor sections; regions of tumor-host interface differed the most from the other tumor sections. Interestingly, tumor samples collected from areas closer to the gastrointestinal transit most frequently shared methylation events with metastases. When we calculated individual coefficients to quantify heterogeneity, we found that epigenetic homogeneity was significantly associated with short time of relapse-free survival (log-rank P = .037) and short time of overall survival (log-rank P = .026) in patients with locoregional colorectal cancer.In an analysis of 79 colorectal tumors, we found significant heterogeneity in patterns of DNA methylation within each tumor; the level of heterogeneity correlates with times of relapse-free and overall survival.

Multivariant analysis. Identification of factors/conditions that influence expression levels between IUGR and control placenta samples. (XLSX 12 kb)

Abstract MeCP2 is a general regulator of transcription involved in the repression/activation of genes depending on the local epigenetic context. It acts as a chromatin regulator and binds with exquisite specificity to gene promoters. The set of epigenetic marks recognized by MeCP2 has been already established (mainly, cytosine modifications in CpG and CpA), as well as many of the constituents of its interactome. We unveil a new set of interactions for MeCP2 with the four canonical nucleosomal histones. MeCP2 interacts with high affinity with H2A, H2B, H3 and H4. In addition, Rett syndrome associated mutations in MeCP2 and histone epigenetic marks modulate these interactions. Given the abundance and the structural/functional relevance of histones and their involvement in epigenetic regulation, this new set of interactions and its modulating elements provide a new addition to the ‘alphabet’ for this epigenetic reader.

Abstract Cancer metastasis could be defined as “the point of no return” in the majority of cancers. The emergence of metastasis is still associated with the worst prognosis and high morbidity. Given the importance of the disease, researchers in this field are focusing on the discovery of new molecular biomarkers able to predict tumor progression and the development of metastasis. Thus, deciphering the molecular and cellular alterations implied in the origin of the metastasis would permit us to develop novel therapies that make a significant contribution to cancer treatment. In this line, the extant variability and versatility of microarray technology provides us a good chance to identify targets involved in cancer progression, to distinguish between cancer types, to identify the tumor type of origin, or to predict the outcome of the disease. According to the successful results obtained applying these kind of techniques, we proposed a genome-wide approach to elucidate the methylation changes associated with cancer progression. Once the first candidates were validated, we assessed the gene function and the consequences that the epigenetic silencing has in the metastatic cascade. The study was carried out in four different paired tumor cell types (11A/11B, MDA-MB-468PT/468LN, 1590PT/1590LN, and SW480/SW620) derived from primary tumor and the corresponding lymph node metastases. For that purpose, we used the new Illumina Infinium Human Methylation 450K array. This powerful platform, mapping 485,577 CpGs, has been successfully validated in our lab and has extensively demonstrated its huge resolution. In order to spot those epigenetic alterations with functional effect on gene activity, we combined these methylation data with the expression data obtained from the Human Genome U133 Plus 2.0 gene expression array performed in the same cell lines. The promoter methylation pattern of candidate genes in cell lines was validated by PCR analysis of bisulfite-modified genomic DNA. In addition, quantitative RT-PCR and Western Blot were used to make sure that the methylation change was indeed associated with a deviation in the gene expression level. DNA methylation status of candidate genes in primary and metastatic tumors was determined by methylation-specific PCR. In our preliminary results we detected several metastasis related genes methylated. Some of them have already been described in colon, breast, esophagus and head and neck cancer metastases but others were new candidates, including both protein-coding genes and microRNAs. Among these, we frequently found ontologies related to metastasis progression such as cell adhesion, cell motility and migration, protein-to-protein interaction with histone deacetylases complexes (HDAC, linked to gene transcription repression), potential members of HDAC, sponge proteins that recruit HDACs or histone acetyltransferases (HATs), and potential suppressor tumor genes. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4091. doi:1538-7445.AM2012-4091

<div>Abstract<p>Epithelial-to-mesenchymal transition (EMT) is a plastic process in which fully differentiated epithelial cells are converted into poorly differentiated, migratory and invasive mesenchymal cells, and it has been related to the metastasis potential of tumors. This is a reversible process and cells can also eventually undergo mesenchymal-to-epithelial transition. The existence of a dynamic EMT process suggests the involvement of epigenetic shifts in the phenotype. Herein, we obtained the DNA methylomes at single-base resolution of Madin–Darby canine kidney cells undergoing EMT and translated the identified differentially methylated regions to human breast cancer cells undergoing a gain of migratory and invasive capabilities associated with the EMT phenotype. We noticed dynamic and reversible changes of DNA methylation, both on promoter sequences and gene-bodies in association with transcription regulation of EMT-related genes. Most importantly, the identified DNA methylation markers of EMT were present in primary mammary tumors in association with the epithelial or the mesenchymal phenotype of the studied breast cancer samples. <i>Cancer Res; 74(19); 5608–19. ©2014 AACR</i>.</p></div>