Publications

<p>Supplementary Figure 1. KAT6B copy number and DNA methylation status in SCLC cell lines</p>

Abstract β Amyloid, present in senile plaques, has been related largely to neuronal loss in the brain of patients with Alzheimer's disease. However, how neurons respond to β amyloid insults is still poorly understood. Here we show that β amyloid increases somatostatin and cortistatin gene expression mainly through an increase in histone 3 lysine 4 methylation (H3K4me3), a modification associated with transcriptional activation. Somatostatin and cortistatin partially decreased β amyloid toxicity in primary cortical neurons in culture. Thus we suggest that neurons respond to β amyloid insults by releasing somatostatin and cortistatin, which will act as a protective agent against β amyloid toxicity. Our results suggest a relevant function for both neuropeptides against β amyloid toxicity, providing new insights into Alzheimer's disease. © 2011 Wiley Periodicals, Inc.

DNA methylation is an important component of the epigenetic machinery that regulates the malignancy of Ewing sarcoma (EWS), the second most common primary bone tumor in children and adolescents. Coordination of DNA methylation and DNA replication is critical for maintaining epigenetic programming and the DNMT1 enzyme has been demonstrated to have an important role in both maintaining the epigenome and controlling cell cycle. Here, we showed that the novel nonnucleoside DNMT inhibitor (DNMTi) MC3343 induces a specific depletion of DNMT1 and affects EWS tumor proliferation through a mechanism that is independent on DNA methylation. Depletion of DNMT1 causes perturbation of the cell cycle, with an accumulation of cells in the G1 phase, and DNA damage, as revealed by the induction of γH2AX foci. These effects elicited activation of p53-dependent signaling and apoptosis in p53wt cells, while in p53 mutated cells, persistent micronuclei and increased DNA instability was observed. Treatment with MC3343 potentiates the efficacy of DNA damaging agents such as doxorubicin and PARP-inhibitors (PARPi). This effect correlates with increased DNA damage and synergistic tumor cytotoxicity, supporting the use of the DNMTi MC3343 as an adjuvant agent in treating EWS.

The Barcelona Conference on Epigenetics and Cancer (BCEC) was held in Barcelona, Spain, on October 1st and 2nd, 2014. The meeting was co-organized by the Cancer Epigenetics and Biology Program (PEBC-IDIBELL) and B·Debate, an initiative of Biocat, with the support of "la Caixa" Foundation. The scientific committee was comprised of leading scientists in the field of epigenetics: Dr. Manel Esteller, director of PEBC-IDIBELL, Dr. Alejandro Vaquero and Dr. Esteban Ballestar, from PEBC-IDIBELL, Juan Ausió from the University of Victoria (Canada), and Marcus Buschbeck, from the Institute of Predictive and Personalized Medicine of Cancer (IMPPC), as BCEC series coordinator. This meeting was the second edition of the BCEC series, which was launched by 5 leading Barcelonan institutes to bring together leading investigators in the fields of epigenetics and chromatin research. The topics discussed during the meeting included the current challenges, opportunities, and perspectives surrounding the study of histone modifications (focusing in acetylation), chromatin structure and gene expression, and the involvement of histone acetylation in physiology and diseases, such as cancer or neurological diseases.

<p>Cell viability was evaluated by MTT assay (72h). Karpas1718 parental (A) or resistant (B) were exposed to DMSO (control, black) or decitabine (100nM, red), given concomitantly (continuous lines) or 72 hours before idelalisib (pre DMSO in dashed black or pre decitabine 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>

We found aberrant DNA methylation of the WNT10B promoter region in 46% of primary hepatocellular carcinoma (HCC) and 15% of colon cancer samples. Three of 10 HCC and one of two colon cancer cell lines demonstrated low or no expression, and 5-aza-2'deoxycytidine reactivated WNT10B expression with the induction of demethylation, indicating that WNT10B is silenced by DNA methylation in some cancers, whereas WNT10B expression is up-regulated in seven of the 10 HCC cell lines and a colon cancer cell line. These results indicate that WNT10B can be deregulated by either overexpression or silencing in cancer. We found that WNT10B up-regulated beta-catenin/Tcf activity. However, WNT10B-overexpressing cells demonstrated a reduced growth rate and anchorage-independent growth that is independent of the beta-catenin/Tcf activation, because mutant beta-catenin-transduced cells did not suppress growth, and dominant-negative hTcf-4 failed to alleviate the growth suppression by WNT10B. Although WNT10B expression alone inhibits cell growth, it acts synergistically with the fibroblast growth factor (FGF) to stimulate cell growth. WNT10B is bifunctional, one function of which is involved in beta-catenin/Tcf activation, and the other function is related to the down-regulation of cell growth through a different mechanism. We suggest that FGF switches WNT10B from a negative to a positive cell growth regulator.

<p>Excel file, with all expression related data and its correlation with methylation</p>

<p>Distribution of mutations in CHIP-associated genes at diagnosis according to response status</p>

<div>AbstractPurpose:<p>The limited knowledge of the molecular alterations that characterize poorly differentiated neuroendocrine carcinomas has limited the clinical development of targeted agents directed to driver mutations. Here we aim to identify new molecular targets in colon neuroendocrine carcinomas (co-NEC) and proof the efficacy of matching drugs.</p>Experimental Design:<p>We performed a multi-omic analysis of co-NEC to identify genetic or epigenetic alterations that could be exploited as effective drug targets. We compared co-NEC samples with colorectal carcinomas (CRC) to identify neuroendocrine-specific traits. Patients with co-NEC and patient-derived xenografts were treated with a BRAFV600E-blocking drug to demonstrate sensitivity.</p>Results:<p>co-NEC and CRC are similar in their mutational repertoire, although co-NECs are particularly enriched in <i>BRAFV600E</i> mutations. We report for the first time that <i>V600EBRAF</i>-mutant co-NECs may benefit from BRAF inhibition in monotherapy and how EGFR status is essential to predict innate sensitivity and acquired resistance by a differential methylation of its gene regulatory regions.</p>Conclusions:<p>The identification of <i>V600E BRAF</i> mutations in high-grade co-NECs has allowed the description of radiological responses to combination therapy of BRAF and MEK inhibitors in basket clinical trials. However, the molecular rationale for this treatment combination was based on the presence of the BRAF mutation and the efficacy observed in other cancer types such as melanoma. Future drug development in this setting should test BRAF inhibitors upfront and the addition of anti-EGFR antibodies instead of MEK inhibitors for an efficient blockade of acquired resistance.</p></div>

<p>Expression levels of surface ERBB4 and pERBB4 was measured by FACS (A) and immunoblotting (B) in Karpas1718 parental and resistant. (A) Two independent experiments were performed: exp1 (left panel) and exp2 (right panel). Density plots show the median MFI values of two replicates from each experiment: negative control (dotted black), parental (grey), resistant (red). Results are presented from Welch Two Sample test performed on MFI values of the two experiments. (B) Immunobloting for total (grey) and phospho (red) ERBB4 in parental (left) and resistant (right) Karpas1718 cells. Representative image of two independent experiments. Data on the barplot represents the mean of two experiments, errors bars represent standard deviation of the mean. * for p<0.05 from a t-test.</p>

<p>Supplementary Table 2. Genes that significantly change expression upon KAT6B depletion. Normalized intensities from the array in each sample and log Fold Changes from the analysis are depicted.</p>

Germline mutations in the BRCA1 or BRCA2 genes are associated with an increased risk of breast and ovarian cancer development. Both genes are involved in DNA repair, and tumors harboring genetic defects in them are thought to be more sensitive to DNA-damaging agents used in chemotherapy. However, as only a minority of breast and ovarian cancer patients carry BRCA1 or BRCA2 mutations, few patients are likely to benefit from these pharmacogenetic biomarkers. Herein, we show that, in cancer cell lines and xenografted tumors, BRCA1 CpG island promoter hypermethylation-associated silencing also predicts enhanced sensitivity to platinum-derived drugs to the same extent as BRCA1 mutations. Most importantly, BRCA1 hypermethylation proves to be a predictor of longer time to relapse and improved overall survival in ovarian cancer patients undergoing chemotherapy with cisplatin.

<p>Ridgeplot on surface levels of ERBB4 by FACS in Karpas1718 resistant cells upon genetic silencing (siRNA) of ERBB2 (yellow), ERBB4 (red) or concomitant ERBB2 and ERBB4 (brown). Silencing control in grey and negative control in black. Data derived from two independent experiments. Statistically significant differences (p<0.05) were evaluated by t-test performed on mean MFI values.</p>