Publications by authors named "Fernando Setien"

47 Publications

Impact of the Epigenetically Regulated Hoxa-5 Gene in Neural Differentiation from Human Adipose-Derived Stem Cells.

Biology (Basel) 2021 Aug 19;10(8). Epub 2021 Aug 19.

Center of Biomedical Research (CIBM), Institute of Biopathology and Regenerative Medicine (IBIMER), University of Granada, 18100 Granada, Spain.

Human adipose-derived mesenchymal stem cells (hASCs) may be used in some nervous system pathologies, although obtaining an adequate degree of neuronal differentiation is an important barrier to their applicability. This requires a deep understanding of the expression and epigenetic changes of the most important genes involved in their differentiation. We used hASCs from human lipoaspirates to induce neuronal-like cells through three protocols (Neu1, 2, and 3), determined the degree of neuronal differentiation using specific biomarkers in culture cells and neurospheres, and analyzed epigenetic changes of genes involved in this differentiation. Furthermore, we selected the gene to determine its potential to improve neuronal differentiation. Our results showed that an excellent hASC neuronal differentiation process using Neu1 which efficiently modulated NES, CHAT, SNAP25, or SCN9A neuronal marker expression. In addition, epigenetic studies showed relevant changes in , , , , , and genes. Functional studies of the gene using CRISPR/dCas9 and lentiviral systems showed that its overexpression induced hASCs neuronal differentiation that was accelerated with the exposure to Neu1. These results suggest that is an essential gene in hASCs neuronal differentiation and therefore, a potential candidate for the development of cell therapy strategies in neurological disorders.
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http://dx.doi.org/10.3390/biology10080802DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8389620PMC
August 2021

Genetic Screening for Variants in Young and Previously Healthy Men With Severe COVID-19.

Front Immunol 2021 23;12:719115. Epub 2021 Jul 23.

Hereditary Cancer Program, Catalan Institute of Oncology, Program in Molecular Mechanisms and Experimental Therapy in Oncology (Oncobell), Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet de Llobregat, Barcelona, Spain.

Introduction: Loss-of-function 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 was sequenced to assess the presence of potentially deleterious variants.

Results: 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 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 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.
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http://dx.doi.org/10.3389/fimmu.2021.719115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8343010PMC
August 2021

SMARCA4 deficient tumours are vulnerable to KDM6A/UTX and KDM6B/JMJD3 blockade.

Nat Commun 2021 07 14;12(1):4319. Epub 2021 Jul 14.

Cancer Genetics Group, Josep Carreras Leukaemia Research Institute (IJC), Badalona, Barcelona, Spain.

Despite the genetic inactivation of SMARCA4, a core component of the SWI/SNF-complex commonly found in cancer, there are no therapies that effectively target SMARCA4-deficient tumours. Here, we show that, unlike the cells with activated MYC oncogene, cells with SMARCA4 inactivation are refractory to the histone deacetylase inhibitor, SAHA, leading to the aberrant accumulation of H3K27me3. SMARCA4-mutant cells also show an impaired transactivation and significantly reduced levels of the histone demethylases KDM6A/UTX and KDM6B/JMJD3, and a strong dependency on these histone demethylases, so that its inhibition compromises cell viability. Administering the KDM6 inhibitor GSK-J4 to mice orthotopically implanted with SMARCA4-mutant lung cancer cells or primary small cell carcinoma of the ovary, hypercalcaemic type (SCCOHT), had strong anti-tumour effects. In this work we highlight the vulnerability of KDM6 inhibitors as a characteristic that could be exploited for treating SMARCA4-mutant cancer patients.
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http://dx.doi.org/10.1038/s41467-021-24618-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8280185PMC
July 2021

Gene Amplification-Associated Overexpression of the Selenoprotein tRNA Enzyme TRIT1 Confers Sensitivity to Arsenic Trioxide in Small-Cell Lung Cancer.

Cancers (Basel) 2021 Apr 14;13(8). Epub 2021 Apr 14.

Josep Carreras Leukaemia Research Institute (IJC), 08916 Barcelona, Spain.

The alteration of RNA modification patterns is emerging as a common feature of human malignancies. If these changes affect key RNA molecules for mRNA translation, such as transfer RNA, they can have important consequences for cell transformation. TRIT1 is the enzyme responsible for the hypermodification of adenosine 37 in the anticodon region of human tRNAs containing serine and selenocysteine. Herein, we show that TRIT1 undergoes gene amplification-associated overexpression in cancer cell lines and primary samples of small-cell lung cancer. From growth and functional standpoints, the induced depletion of TRIT1 expression in amplified cells reduces their tumorigenic potential and downregulates the selenoprotein transcripts. We observed that TRIT1-amplified cells are sensitive to arsenic trioxide, a compound that regulates selenoproteins, whereas reduction of TRIT1 levels confers loss of sensitivity to the drug. Overall, our results indicate a role for TRIT1 as a small-cell lung cancer-relevant gene that, when undergoing gene amplification-associated activation, can be targeted with the differentiation agent arsenic trioxide.
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http://dx.doi.org/10.3390/cancers13081869DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8070726PMC
April 2021

Epigenetic loss of the transfer RNA-modifying enzyme TYW2 induces ribosome frameshifts in colon cancer.

Proc Natl Acad Sci U S A 2020 08 10;117(34):20785-20793. Epub 2020 Aug 10.

Cancer Epigenetics Group, Josep Carreras Leukaemia Research Institute, 08916, Badalona, Barcelona, Catalonia, Spain;

Transfer RNA (tRNA) activity is tightly regulated to provide a physiological protein translation, and tRNA chemical modifications control its function in a complex with ribosomes and messenger RNAs (mRNAs). In this regard, the correct hypermodification of position G37 of phenylalanine-tRNA, adjacent to the anticodon, is critical to prevent ribosome frameshifting events. Here we report that the tRNA-yW Synthesizing Protein 2 (TYW2) undergoes promoter hypermethylation-associated transcriptional silencing in human cancer, particularly in colorectal tumors. The epigenetic loss of TYW2 induces guanosine hypomodification in phenylalanine-tRNA, an increase in -1 ribosome frameshift events, and down-regulation of transcripts by mRNA decay, such as of the key cancer gene ROBO1. Importantly, TYW2 epigenetic inactivation is linked to poor overall survival in patients with early-stage colorectal cancer, a finding that could be related to the observed acquisition of enhanced migration properties and epithelial-to-mesenchymal features in the colon cancer cells that harbor TYW2 DNA methylation-associated loss. These findings provide an illustrative example of how epigenetic changes can modify the epitranscriptome and further support a role for tRNA modifications in cancer biology.
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http://dx.doi.org/10.1073/pnas.2003358117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7456070PMC
August 2020

Epigenetic inactivation of the splicing RNA-binding protein CELF2 in human breast cancer.

Oncogene 2019 11 13;38(45):7106-7112. Epub 2019 Aug 13.

Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet, Barcelona, Catalonia, Spain.

Human tumors show altered patterns of protein isoforms that can be related to the dysregulation of messenger RNA alternative splicing also observed in transformed cells. Although somatic mutations in core spliceosome components and their associated factors have been described in some cases, almost nothing is known about the contribution of distorted epigenetic patterns to aberrant splicing. Herein, we show that the splicing RNA-binding protein CELF2 is targeted by promoter hypermethylation-associated transcriptional silencing in human cancer. Focusing on the context of breast cancer, we also demonstrate that CELF2 restoration has growth-inhibitory effects and that its epigenetic loss induces an aberrant downstream pattern of alternative splicing, affecting key genes in breast cancer biology such as the autophagy factor ULK1 and the apoptotic protein CARD10. Furthermore, the presence of CELF2 hypermethylation in the clinical setting is associated with shorter overall survival of the breast cancer patients carrying this epigenetic lesion.
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http://dx.doi.org/10.1038/s41388-019-0936-xDOI Listing
November 2019

Epigenetic silencing of TGFBI confers resistance to trastuzumab in human breast cancer.

Breast Cancer Res 2019 07 5;21(1):79. Epub 2019 Jul 5.

New Therapeutics Targets Lab (TargetsLab), Department of Medical Sciences, University of Girona, E-17071, Girona, Catalonia, Spain.

Background: Acquired resistance to trastuzumab is a major clinical problem in the treatment of HER2-positive (HER2+) breast cancer patients. The selection of trastuzumab-resistant patients is a great challenge of precision oncology. The aim of this study was to identify novel epigenetic biomarkers associated to trastuzumab resistance in HER2+ BC patients.

Methods: We performed a genome-wide DNA methylation (450K array) and a transcriptomic analysis (RNA-Seq) comparing trastuzumab-sensitive (SK) and trastuzumab-resistant (SKTR) HER2+ human breast cancer cell models. The methylation and expression levels of candidate genes were validated by bisulfite pyrosequencing and qRT-PCR, respectively. Functional assays were conducted in the SK and SKTR models by gene silencing and overexpression. Methylation analysis in 24 HER2+ human BC samples with complete response or non-response to trastuzumab-based treatment was conducted by bisulfite pyrosequencing.

Results: Epigenomic and transcriptomic analysis revealed the consistent hypermethylation and downregulation of TGFBI, CXCL2, and SLC38A1 genes in association with trastuzumab resistance. The DNA methylation and expression levels of these genes were validated in both sensitive and resistant models analyzed. Of the genes, TGFBI presented the highest hypermethylation-associated silencing both at the transcriptional and protein level. Ectopic expression of TGFBI in the SKTR model suggest an increased sensitivity to trastuzumab treatment. In primary tumors, TGFBI hypermethylation was significantly associated with trastuzumab resistance in HER2+ breast cancer patients.

Conclusions: Our results suggest for the first time an association between the epigenetic silencing of TGFBI by DNA methylation and trastuzumab resistance in HER2+ cell models. These results provide the basis for further clinical studies to validate the hypermethylation of TGFBI promoter as a biomarker of trastuzumab resistance in HER2+ breast cancer patients.
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http://dx.doi.org/10.1186/s13058-019-1160-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6612099PMC
July 2019

Epigenetic loss of the endoplasmic reticulum-associated degradation inhibitor SVIP induces cancer cell metabolic reprogramming.

JCI Insight 2019 03 7;5. Epub 2019 Mar 7.

Cancer Epigenetics Group, Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain.

The endoplasmic reticulum (ER) of cancer cells needs to adapt to the enhanced proteotoxic stress associated with the accumulation of unfolded, misfolded and transformation-associated proteins. One way by which tumors thrive in the context of ER stress is by promoting ER-Associated Degradation (ERAD), although the mechanisms are poorly understood. Here, we show that the Small p97/VCP Interacting Protein (SVIP), an endogenous inhibitor of ERAD, undergoes DNA hypermethylation-associated silencing in tumorigenesis to achieve this goal. SVIP exhibits tumor suppressor features and its recovery is associated with increased ER stress and growth inhibition. Proteomic and metabolomic analyses show that cancer cells with epigenetic loss of SVIP are depleted in mitochondrial enzymes and oxidative respiration activity. This phenotype is reverted upon SVIP restoration. The dependence of SVIP hypermethylated cancer cells on aerobic glycolysis and glucose was also associated with sensitivity to an inhibitor of the glucose transporter GLUT1. This could be relevant to the management of tumors carrying SVIP epigenetic loss, because these occur in high-risk patients who manifest poor clinical outcomes. Overall, our study provides insights into how epigenetics helps deal with ER stress and how SVIP epigenetic loss in cancer may be amenable to therapies that target glucose transporters.
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http://dx.doi.org/10.1172/jci.insight.125888DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6538322PMC
March 2019

Germline variation in O-methylguanine-DNA methyltransferase (MGMT) as cause of hereditary colorectal cancer.

Cancer Lett 2019 04 21;447:86-92. Epub 2019 Jan 21.

Hereditary Cancer Program, Catalan Institute of Oncology, IDIBELL, Hospitalet de Llobregat, Barcelona, Spain; Program in Molecular Mechanisms and Experimental Therapy in Oncology (Oncobell), IDIBELL, Hospitalet de Llobregat, Barcelona, Spain; Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Spain. Electronic address:

Somatic epigenetic inactivation of the DNA repair protein O6-methylguanine DNA methyltransferase (MGMT) is frequent in colorectal cancer (CRC); however, its involvement in CRC predisposition remains unexplored. We assessed the role and relevance of MGMT germline mutations and epimutations in familial and early-onset CRC. Mutation and promoter methylation screenings were performed in 473 familial and/or early-onset mismatch repair-proficient nonpolyposis CRC cases. No constitutional MGMT inactivation by promoter methylation was observed. Of six rare heterozygous germline variants identified, c.346C > T (p.H116Y) and c.476G > A (p.R159Q), detected in three and one families respectively, affected highly conserved residues and showed segregation with cancer in available family members. In vitro, neither p.H116Y nor p.R159Q caused statistically significant reduction of MGMT repair activity. No evidence of somatic second hits was found in the studied tumors. Case-control data showed over-representation of c.346C > T (p.H116Y) in familial CRC compared to controls, but no overall association of MGMT mutations with CRC predisposition. In conclusion, germline mutations and constitutional epimutations in MGMT are not major players in hereditary CRC. Nevertheless, the over-representation of c.346C > T (p.H116Y) in our familial CRC cohort warrants further research.
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http://dx.doi.org/10.1016/j.canlet.2019.01.019DOI Listing
April 2019

Circular RNA CpG island hypermethylation-associated silencing in human cancer.

Oncotarget 2018 Jun 26;9(49):29208-29219. Epub 2018 Jun 26.

Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain.

Noncoding RNAs (ncRNAs), such as microRNAs and long noncoding RNAs (lncRNAs), participate in cellular transformation. Work done in the last decade has also demonstrated that ncRNAs with growth-inhibitory functions can undergo promoter CpG island hypermethylation-associated silencing in tumorigenesis. Herein, we wondered whether circular RNAs (circRNAs), a type of RNA transcripts lacking 5'-3' ends and forming closed loops that are gaining relevance in cancer biology, are also a target of epigenetic inactivation in tumors. To tackle this issue, we have used cancer cells genetically deficient for the DNA methyltransferase enzymes in conjuction with circRNA expression microarrays. We have found that the loss of DNA methylation provokes a release of circRNA silencing. In particular, we have identified that promoter CpG island hypermethylation of the genes TUSC3 (tumor suppressor candidate 3), POMT1 (protein O-mannosyltransferase 1), ATRNL1 (attractin-like 1) and SAMD4A (sterile alpha motif domain containing 4A) is linked to the transcriptional downregulation of both linear mRNA and the hosted circRNA. Although some circRNAs regulate the linear transcript, we did not observe changes in TUSC3 mRNA levels upon TUSC3 circ104557 overexpression. Interestingly, we found circRNA-mediated regulation of target miRNAs and an growth inhibitory effect upon TUSC3 circ104557 transduction. Data mining for 5'-end CpG island methylation of TUSC3, ATRNL1, POMT1 and SAMD4A in cancer cell lines and primary tumors showed that the epigenetic defect was commonly observed among different tumor types in association with the diminished expression of the corresponding transcript. Our findings support a role for circRNA DNA methylation-associated loss in human cancer.
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http://dx.doi.org/10.18632/oncotarget.25673DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6044373PMC
June 2018

Germline mutations in the spindle assembly checkpoint genes BUB1 and BUB3 are infrequent in familial colorectal cancer and polyposis.

Mol Cancer 2018 02 15;17(1):23. Epub 2018 Feb 15.

Hereditary Cancer Program, Catalan Institute of Oncology, IDIBELL, Hospitalet de Llobregat, Av. Gran Via 199-203, 08908, Barcelona, Spain.

Germline mutations in BUB1 and BUB3 have been reported to increase the risk of developing colorectal cancer (CRC) at young age, in presence of variegated aneuploidy and reminiscent dysmorphic traits of mosaic variegated aneuploidy syndrome. We performed a mutational analysis of BUB1 and BUB3 in 456 uncharacterized mismatch repair-proficient hereditary non-polyposis CRC families and 88 polyposis cases. Four novel or rare germline variants, one splice-site and three missense, were identified in four families. Neither variegated aneuploidy nor dysmorphic traits were observed in carriers. Evident functional effects in the heterozygous form were observed for c.1965-1G>A, but not for c.2296G>A (p.E766K), in spite of the positive co-segregation in the family. BUB1 c.2473C>T (p.P825S) and BUB3 c.77C>T (p.T26I) remained as variants of uncertain significance. As of today, the rarity of functionally relevant mutations identified in familial and/or early onset series does not support the inclusion of BUB1 and BUB3 testing in routine genetic diagnostics of familial CRC.
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http://dx.doi.org/10.1186/s12943-018-0762-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5815240PMC
February 2018

DNA Methylomes Reveal Biological Networks Involved in Human Eye Development, Functions and Associated Disorders.

Sci Rep 2017 09 18;7(1):11762. Epub 2017 Sep 18.

Cancer Epigenetics Group; Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain.

This work provides a comprehensive CpG methylation landscape of the different layers of the human eye that unveils the gene networks associated with their biological functions and how these are disrupted in common visual disorders. Herein, we firstly determined the role of CpG methylation in the regulation of ocular tissue-specification and described hypermethylation of retinal transcription factors (i.e., PAX6, RAX, SIX6) in a tissue-dependent manner. Second, we have characterized the DNA methylome of visual disorders linked to internal and external environmental factors. Main conclusions allow certifying that crucial pathways related to Wnt-MAPK signaling pathways or neuroinflammation are epigenetically controlled in the fibrotic disorders involved in retinal detachment, but results also reinforced the contribution of neurovascularization (ETS1, HES5, PRDM16) in diabetic retinopathy. Finally, we had studied the methylome in the most frequent intraocular tumors in adults and children (uveal melanoma and retinoblastoma, respectively). We observed that hypermethylation of tumor suppressor genes is a frequent event in ocular tumors, but also unmethylation is associated with tumorogenesis. Interestingly, unmethylation of the proto-oncogen RAB31 was a predictor of metastasis risk in uveal melanoma. Loss of methylation of the oncogenic mir-17-92 cluster was detected in primary tissues but also in blood from patients.
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http://dx.doi.org/10.1038/s41598-017-12084-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5603607PMC
September 2017

Bromodomain inhibition shows antitumoral activity in mice and human luminal breast cancer.

Oncotarget 2017 Aug 29;8(31):51621-51629. Epub 2017 May 29.

Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain.

BET bromodomain inhibitors, which have an antitumoral effect against various solid cancer tumor types, have not been studied in detail in luminal breast cancer, despite the prevalence of this subtype of mammary malignancy. Here we demonstrate that the BET bromodomain inhibitor JQ1 exerts growth-inhibitory activity in human luminal breast cancer cell lines associated with a depletion of the C-MYC oncogene, but does not alter the expression levels of the BRD4 bromodomain protein. Interestingly, expression microarray analyses indicate that, upon JQ1 administration, the antitumoral phenotype also involves downregulation of relevant breast cancer oncogenes such as the Breast Carcinoma-Amplified Sequence 1 (BCAS1) and the PDZ Domain-Containing 1 (PDZK1). We have also applied these findings in an model by studying a transgenic mouse model representing the luminal B subtype of breast cancer, the MMTV-PyMT, in which the mouse mammary tumor virus promoter is used to drive the expression of the polyoma virus middle T-antigen to the mammary gland. We have observed that the use of the BET bromodomain inhibitor for the treatment of established breast neoplasms developed in the MMTV-PyMT model shows antitumor potential. Most importantly, if JQ1 is given before the expected time of tumor detection in the MMTV-PyMT mice, it retards the onset of the disease and increases the survival of these animals. Thus, our findings indicate that the use of bromodomain inhibitors is of great potential in the treatment of luminal breast cancer and merits further investigation.
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http://dx.doi.org/10.18632/oncotarget.18255DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5584274PMC
August 2017

Epigenetic inactivation of the p53-induced long noncoding RNA TP53 target 1 in human cancer.

Proc Natl Acad Sci U S A 2016 11 7;113(47):E7535-E7544. Epub 2016 Nov 7.

Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), L'Hospitalet de Llobregat, 08908 Barcelona, Catalonia, Spain;

Long noncoding RNAs (lncRNAs) are important regulators of cellular homeostasis. However, their contribution to the cancer phenotype still needs to be established. Herein, we have identified a p53-induced lncRNA, TP53TG1, that undergoes cancer-specific promoter hypermethylation-associated silencing. In vitro and in vivo assays identify a tumor-suppressor activity for TP53TG1 and a role in the p53 response to DNA damage. Importantly, we show that TP53TG1 binds to the multifaceted DNA/RNA binding protein YBX1 to prevent its nuclear localization and thus the YBX1-mediated activation of oncogenes. TP53TG1 epigenetic inactivation in cancer cells releases the transcriptional repression of YBX1-targeted growth-promoting genes and creates a chemoresistant tumor. TP53TG1 hypermethylation in primary tumors is shown to be associated with poor outcome. The epigenetic loss of TP53TG1 therefore represents an altered event in an lncRNA that is linked to classical tumoral pathways, such as p53 signaling, but is also connected to regulatory networks of the cancer cell.
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http://dx.doi.org/10.1073/pnas.1608585113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5127373PMC
November 2016

Germline missense pathogenic variants in the BRCA1 BRCT domain, p.Gly1706Glu and p.Ala1708Glu, increase cellular sensitivity to PARP inhibitor olaparib by a dominant negative effect.

Hum Mol Genet 2016 12;25(24):5287-5299

Human Genetics Group, Human Cancer Genetics Programme, Spanish National Cancer Research Centre (CNIO), Madrid, Spain.

BRCA1-deficient cells show defects in DNA repair and rely on other members of the DNA repair machinery, which makes them sensitive to PARP inhibitors (PARPi). Although carrying a germline pathogenic variant in BRCA1/2 is the best determinant of response to PARPi, a significant percentage of the patients do not show sensitivity and/or display increased toxicity to the agent. Considering previously suggested mutation-specific BRCA1 haploinsufficiency, we aimed to investigate whether there are any differences in cellular response to PARPi olaparib depending on the BRCA1 mutation type. Lymphoblastoid cell lines derived from carriers of missense pathogenic variants in the BRCA1 BRCT domain (c.5117G > A, p.Gly1706Glu and c.5123C > A, p.Ala1708Glu) showed higher sensitivity to olaparib than cells with truncating variants or wild types (WT). Response to olaparib depended on a basal PARP enzymatic activity, but did not correlate with PARP1 expression. Interestingly, cellular sensitivity to the agent was associated with the level of BRCA1 recruitment into γH2AX foci, being the lowest in cells with missense variants. Since these variants lead to partially stable protein mutants, we propose a model in which the mutant protein acts in a dominant negative manner on the WT BRCA1, impairing the recruitment of BRCA1 into DNA damage sites and, consequently, increasing cellular sensitivity to PARPi. Taken together, our results indicate that carriers of different BRCA1 mutations could benefit from olaparib in a distinct way and show different toxicities to the agent, which could be especially relevant for a potential future use of PARPi as prophylactic agents in BRCA1 mutation carriers.
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http://dx.doi.org/10.1093/hmg/ddw343DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5815624PMC
December 2016

A mutation in the POT1 gene is responsible for cardiac angiosarcoma in TP53-negative Li-Fraumeni-like families.

Nat Commun 2015 Sep 25;6:8383. Epub 2015 Sep 25.

Human Genetics Group, Spanish National Cancer Research Center (CNIO), Melchor Fernandez Almagro 3, Madrid 28029, Spain.

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.
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http://dx.doi.org/10.1038/ncomms9383DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4598567PMC
September 2015

KAT6B Is a Tumor Suppressor Histone H3 Lysine 23 Acetyltransferase Undergoing Genomic Loss in Small Cell Lung Cancer.

Cancer Res 2015 Sep 24;75(18):3936-45. Epub 2015 Jul 24.

Cancer Epigenetics Group, Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain. Department of Physiological Sciences II, School of Medicine, University of Barcelona, Barcelona, Catalonia, Spain. Institucio Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Catalonia, Spain.

Recent efforts to sequence human cancer genomes have highlighted that point mutations in genes involved in the epigenetic setting occur in tumor cells. Small cell lung cancer (SCLC) is an aggressive tumor with poor prognosis, where little is known about the genetic events related to its development. Herein, we have identified the presence of homozygous deletions of the candidate histone acetyltransferase KAT6B, and the loss of the corresponding transcript, in SCLC cell lines and primary tumors. Furthermore, we show, in vitro and in vivo, that the depletion of KAT6B expression enhances cancer growth, while its restoration induces tumor suppressor-like features. Most importantly, we demonstrate that KAT6B exerts its tumor-inhibitory role through a newly defined type of histone H3 Lys23 acetyltransferase activity.
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http://dx.doi.org/10.1158/0008-5472.CAN-14-3702DOI Listing
September 2015

DNA repair capacity is impaired in healthy BRCA1 heterozygous mutation carriers.

Breast Cancer Res Treat 2015 Jul 14;152(2):271-82. Epub 2015 Jun 14.

Human Genetics Group, Human Cancer Genetics Programme, Spanish National Cancer Research Centre (CNIO), C/Melchor Fernández Almagro 3, 28029, Madrid, Spain.

BRCA1 germline mutations increase the lifetime risk of developing breast and ovarian cancers. However, taking into account the differences in disease manifestation among mutation carriers, it is probable that different BRCA1 mutations have distinct haploinsufficiency effects and lead to the formation of different phenotypes. Using lymphoblastoid cell lines derived from heterozygous BRCA1 mutation carriers and non-carriers, we investigated the haploinsufficiency effects of various mutation types using qPCR, immunofluorescence, and microarray technology. Lymphoblastoid cell lines carrying a truncating mutation showed significantly lower BRCA1 mRNA and protein levels and higher levels of gamma-H2AX than control cells or those harboring a missense mutation, indicating greater spontaneous DNA damage. Cells carrying either BRCA1 mutation type showed impaired RAD51 foci formation, suggesting defective repair in mutated cells. Moreover, compared to controls, cell lines carrying missense mutations displayed a more distinct expression profile than cells with truncating mutations, which is consistent with different mutations giving rise to distinct phenotypes. Alterations in the immune response pathway in cells harboring missense mutations point to possible mechanisms of breast cancer initiation in carriers of these mutations. Our findings offer insight into how various heterozygous mutations in BRCA1 could lead to impairment of BRCA1 function and provide strong evidence of haploinsufficiency in BRCA1 mutation carriers.
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http://dx.doi.org/10.1007/s10549-015-3459-3DOI Listing
July 2015

A novel Werner Syndrome mutation: pharmacological treatment by read-through of nonsense mutations and epigenetic therapies.

Epigenetics 2015 ;10(4):329-41

a Epigenetics of Cancer and Aging Laboratory Institut Pasteur de Montevideo (IPMON) ; Montevideo , Uruguay.

Werner Syndrome (WS) is a rare inherited disease characterized by premature aging and increased propensity for cancer. Mutations in the WRN gene can be of several types, including nonsense mutations, leading to a truncated protein form. WRN is a RecQ family member with both helicase and exonuclease activities, and it participates in several cell metabolic pathways, including DNA replication, DNA repair, and telomere maintenance. Here, we reported a novel homozygous WS mutation (c.3767 C > G) in 2 Argentinian brothers, which resulted in a stop codon and a truncated protein (p.S1256X). We also observed increased WRN promoter methylation in the cells of patients and decreased messenger WRN RNA (WRN mRNA) expression. Finally, we showed that the read-through of nonsense mutation pharmacologic treatment with both aminoglycosides (AGs) and ataluren (PTC-124) in these cells restores full-length protein expression and WRN functionality.
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http://dx.doi.org/10.1080/15592294.2015.1027853DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4622951PMC
November 2015

Whole-exome sequencing identifies MDH2 as a new familial paraganglioma gene.

J Natl Cancer Inst 2015 Mar 11;107(5). Epub 2015 Mar 11.

: Hereditary Endocrine Cancer Group, Spanish National Cancer Research Centre, Madrid, Spain (AC, ICM, MCF, AAdC, VM, LIP, RL, AGG, MAR, CRA, MR); Centro de Investigación Biomédica en Red de Enfermedades Raras, Madrid, Spain (AC, LC, LIP, CRA, JS, MU, MR); Departamento de Biología Molecular, Centro de Biología Molecular Severo Ochoa UAM-CSIC, Universidad Autónoma de Madrid and Instituto de Investigación Sanitaria Fundación Jiménez Díaz, Madrid, Spain (LC, JS); Institute of Clinical Chemistry and Laboratory Medicine, University Hospital Carl Gustav Carus, Medical Faculty Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany (SR, MP, GE); Departments of Pathology (APB) and Endocrinology and Nutrition Service (MC), Hospital 12 de Octubre, Madrid, Spain; Endocrinology Service, Hospital Infanta Sofía, San Sebastián de los Reyes, Spain (SA); Department of Endocrinology and Nutrition Service, Hospital de Fuenlabrada, Madrid, Spain (RVV); Endocrinology Service, Hospital Puerta de Hierro, Majadahonda, Madrid, Spain (JA); Cancer Epigenetics and Biology Program, Bellvitge Biomedical Research Institute, L'Hospitalet, Barcelona, Spain (FS, SM, ME); Department of Pathology, MD Anderson Cancer Center Madrid, Madrid, Spain (JFG); Molecular Cytogenetics Group (ARM), Monoclonal Antibodies Unit, Biotechnology Programme (GR), and Familial Cancer Clinical Unit (MU), Spanish National Cancer Research Centre, Madrid, Spain.

Disruption of the Krebs cycle is a hallmark of cancer. IDH1 and IDH2 mutations are found in many neoplasms, and germline alterations in SDH genes and FH predispose to pheochromocytoma/paraganglioma and other cancers. We describe a paraganglioma family carrying a germline mutation in MDH2, which encodes a Krebs cycle enzyme. Whole-exome sequencing was applied to tumor DNA obtained from a man age 55 years diagnosed with multiple malignant paragangliomas. Data were analyzed with the two-sided Student's t and Mann-Whitney U tests with Bonferroni correction for multiple comparisons. Between six- and 14-fold lower levels of MDH2 expression were observed in MDH2-mutated tumors compared with control patients. Knockdown (KD) of MDH2 in HeLa cells by shRNA triggered the accumulation of both malate (mean ± SD: wild-type [WT] = 1±0.18; KD = 2.24±0.17, P = .043) and fumarate (WT = 1±0.06; KD = 2.6±0.25, P = .033), which was reversed by transient introduction of WT MDH2 cDNA. Segregation of the mutation with disease and absence of MDH2 in mutated tumors revealed MDH2 as a novel pheochromocytoma/paraganglioma susceptibility gene.
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http://dx.doi.org/10.1093/jnci/djv053DOI Listing
March 2015

A demonstration of the Storz C-CAM in endoscopic imagery projection for far forward battlefield telemetric support and training.

Stud Health Technol Inform 2013 ;184:51-5

VA Nebraska-Western Iowa Healthcare System, Omaha, NE, USA.

Airway management skills are essential for healthcare providers within military and civilian settings. To maintain competency in these skills, it is crucial for the provider to have opportunities for review and retraining. Virtual airway training or telementoring can be an effective means to fulfilling these requirements for healthcare providers located in remote sites. The projection of high quality imagery to far forward locations is essential for health care practitioners in the provision of telemedicine and distance training. The Storz C-CAM was developed to interface with existing endoscopy equipment to facilitate implementation of telemetric devices in remote locations. This work describes the use of the Storz C-CAM in providing medical device training to deployed medical personnel at a far forward location.
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July 2013

Virtual simulation training using the Storz C-HUB to support distance airway training for the Spanish Medical Corps and NATO partners.

Stud Health Technol Inform 2012 ;182:1-9

Hospital Central de la Defensa, Madrid, Spain.

In medicine, the advancement of new technologies creates challenges to providers both in learning and in maintaining competency in required skills. For those medical providers located in remote environments, access to learning can be even more formidable. This work describes a collaboration created to facilitate the use of new communication technologies in providing distance training and support to health care personnel deployed in remote areas.
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April 2013

DNA methylation plasticity of human adipose-derived stem cells in lineage commitment.

Am J Pathol 2012 Dec 29;181(6):2079-93. Epub 2012 Sep 29.

Cancer Epigenetics Group, Cancer Epigenetics and Biology Program, Bellvitge Biomedical Biomedical Research Institute, Barcelona, Spain.

Adult stem cells have an enormous potential for clinical use in regenerative medicine that avoids many of the drawbacks characteristic of embryonic stem cells and induced pluripotent stem cells. In this context, easily obtainable human adipose-derived stem cells offer an interesting option for future strategies in regenerative medicine. However, little is known about their repertoire of differentiation capacities, how closely they resemble the target primary tissues, and the potential safety issues associated with their use. DNA methylation is one of the most widely recognized epigenetic factors involved in cellular identity, prompting us to consider how the analyses of 27,578 CpG sites in the genome of these cells under different conditions reflect their different natural history. We show that human adipose-derived stem cells generate myogenic and osteogenic lineages that share much of the DNA methylation landscape characteristic of primary myocytes and osteocytes. Most important, adult stem cells and in vitro-generated myocytes and osteocytes display a significantly different DNA methylome from that observed in transformed cells from these tissue types, such as rhabdomyosarcoma and osteosarcoma. These results suggest that the plasticity of the DNA methylation patterns plays an important role in lineage commitment of adult stem cells and that it could be used for clinical purposes as a biomarker of efficient and safely differentiated cells.
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http://dx.doi.org/10.1016/j.ajpath.2012.08.016DOI Listing
December 2012

Distinct DNA methylomes of newborns and centenarians.

Proc Natl Acad Sci U S A 2012 Jun 11;109(26):10522-7. Epub 2012 Jun 11.

Cancer Epigenetics and Biology Program, Spanish Biomedical Research Centre Network for Epidemiology and Public Health, Catalan Institute of Oncology, Bellvitge Biomedical Research Institute, L’Hospitalet, Barcelona, Catalonia 08908, Spain.

Human aging cannot be fully understood in terms of the constrained genetic setting. Epigenetic drift is an alternative means of explaining age-associated alterations. To address this issue, we performed whole-genome bisulfite sequencing (WGBS) of newborn and centenarian genomes. The centenarian DNA had a lower DNA methylation content and a reduced correlation in the methylation status of neighboring cytosine--phosphate--guanine (CpGs) throughout the genome in comparison with the more homogeneously methylated newborn DNA. The more hypomethylated CpGs observed in the centenarian DNA compared with the neonate covered all genomic compartments, such as promoters, exonic, intronic, and intergenic regions. For regulatory regions, the most hypomethylated sequences in the centenarian DNA were present mainly at CpG-poor promoters and in tissue-specific genes, whereas a greater level of DNA methylation was observed in CpG island promoters. We extended the study to a larger cohort of newborn and nonagenarian samples using a 450,000 CpG-site DNA methylation microarray that reinforced the observation of more hypomethylated DNA sequences in the advanced age group. WGBS and 450,000 analyses of middle-age individuals demonstrated DNA methylomes in the crossroad between the newborn and the nonagenarian/centenarian groups. Our study constitutes a unique DNA methylation analysis of the extreme points of human life at a single-nucleotide resolution level.
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http://dx.doi.org/10.1073/pnas.1120658109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3387108PMC
June 2012

The tumour suppressor and chromatin-remodelling factor BRG1 antagonizes Myc activity and promotes cell differentiation in human cancer.

EMBO Mol Med 2012 Jul 11;4(7):603-16. Epub 2012 Apr 11.

Genes and Cancer Group, Cancer Epigenetics and Biology Program-PEBC, Bellvitge Biomedical Research Institute-IDIBELL, Hospitalet de Llobregat, Barcelona, Spain.

BRG1, a member of the SWI/SNF complex, is mutated in cancer, but it is unclear how it promotes tumourigenesis. We report that re-expression of BRG1 in lung cancer cells up-regulates lung-specific transcripts, restoring the gene expression signature of normal lung. Using cell lines from several cancer types we found that those lacking BRG1 do not respond to retinoic acid (RA) or glucocorticoids (GC), while restoration of BRG1 restores sensitivity. Conversely, in SH-SY5Y cells, a paradigm of RA-dependent differentiation, abrogation of BRG1 prevented the response to RA. Further, our data suggest an antagonistic functional connection between BRG1 and MYC, whereby, refractoriness to RA and GC by BRG1 inactivation involves deregulation of MYC activity. Mechanistically, some of these effects are mediated by BRG1 binding to MYC and MYC-target promoters. The BRG1-MYC antagonism was also evident in primary tumours. Finally, BRG1 restoration significantly dampened invasion and progression and decreased MYC in lung cancer cells orthotopically implanted in nude mice. Thus, BRG1 inactivation enables cancer cells to sustain undifferentiated gene expression programs and prevent its response to environmental stimuli.
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http://dx.doi.org/10.1002/emmm.201200236DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3407948PMC
July 2012

Regulation of SNAIL1 and E-cadherin function by DNMT1 in a DNA methylation-independent context.

Nucleic Acids Res 2011 Nov 16;39(21):9194-205. Epub 2011 Aug 16.

Instituto de Investigaciones Biomédicas Alberto Sols, CSIC-UAM, Instituto de Investigación Hospital Universitario La Paz, Madrid, Spain.

Mammalian DNA methyltransferase 1 (DNMT1) is essential for maintaining DNA methylation patterns after cell division. Disruption of DNMT1 catalytic activity results in whole genome cytosine demethylation of CpG dinucleotides, promoting severe dysfunctions in somatic cells and during embryonic development. While these observations indicate that DNMT1-dependent DNA methylation is required for proper cell function, the possibility that DNMT1 has a role independent of its catalytic activity is a matter of controversy. Here, we provide evidence that DNMT1 can support cell functions that do not require the C-terminal catalytic domain. We report that PCNA and DMAP1 domains in the N-terminal region of DNMT1 are sufficient to modulate E-cadherin expression in the absence of noticeable changes in DNA methylation patterns in the gene promoters involved. Changes in E-cadherin expression are directly associated with regulation of β-catenin-dependent transcription. Present evidence suggests that the DNMT1 acts on E-cadherin expression through its direct interaction with the E-cadherin transcriptional repressor SNAIL1.
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http://dx.doi.org/10.1093/nar/gkr658DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3241660PMC
November 2011

Small molecule enoxacin is a cancer-specific growth inhibitor that acts by enhancing TAR RNA-binding protein 2-mediated microRNA processing.

Proc Natl Acad Sci U S A 2011 Mar 28;108(11):4394-9. Epub 2011 Feb 28.

Cancer Epigenetics and Biology Program, Bellvitge Biomedical Research Institute, 08908 L'Hospitalet, Barcelona, Catalonia, Spain.

MicroRNAs (miRNAs) are small RNA molecules that regulate gene expression at the posttranscriptional level and are critical for many cellular pathways. The disruption of miRNAs and their processing machineries also contributes to the development of human tumors. A common scenario for miRNA expression in carcinogenesis is emerging that shows that impaired miRNA production and/or down-regulation of these transcripts occurs in many neoplasms. Several of these lost miRNAs have tumor-suppressor features, so strategies to restore their expression globally in malignancies would be a welcome addition to the current therapeutic arsenal against cancer. Herein, we show that the small molecule enoxacin, a fluoroquinolone used as an antibacterial compound, enhances the production of miRNAs with tumor suppressor functions by binding to the miRNA biosynthesis protein TAR RNA-binding protein 2 (TRBP). The use of enoxacin in human cell cultures and xenografted, orthotopic, and metastatic mouse models reveals a TRBP-dependent and cancer-specific growth-inhibitory effect of the drug. These results highlight the key role of disrupted miRNA expression patterns in tumorigenesis, and suggest a unique strategy for restoring the distorted microRNAome of cancer cells to a more physiological setting.
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http://dx.doi.org/10.1073/pnas.1014720108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3060242PMC
March 2011

The Δ4-desaturation pathway for DHA biosynthesis is operative in the human species: differences between normal controls and children with the Zellweger syndrome.

Lipids Health Dis 2010 Sep 9;9:98. Epub 2010 Sep 9.

Manuela Martinez Foundation for Children with Metabolic Diseases, Research Laboratory, Plaza Karl Marx 1, Barcelona 08042, Spain.

Background: Docosahexaenoic acid (DHA, 22:6ω3) is a fundamental component of cell membranes, especially in the brain and retina. In the experimental animal, DHA deficiency leads to suboptimal neurological performance and visual deficiencies. Children with the Zellweger syndrome (ZS) have a profound DHA deficiency and symptoms that can be attributed to their extremely low DHA levels. These children seem to have a metabolic defect in DHA biosynthesis, which has never been totally elucidated. Treatment with DHA ethyl ester greatly improves these patients, but if we could normalize their endogenous DHA production we could get additional benefits. We examined whether DHA biosynthesis by Δ4-desaturation could be enhanced in the human species by transfecting the enzyme, and if this could normalize the DHA levels in cells from ZS patients.

Results: We showed that the Δ4-desaturase gene (Fad4) from Thraustochytrium sp, which can be expressed by heterologous transfection in other plant and yeast cells, can also be transfected into human lymphocytes, and that it expresses the enzyme (FAD4, Δ4-desaturase) by producing DHA from direct Δ4-desaturation of 22:5ω3. We also found that the other substrate for Δ4-desaturase, 22:4ω6, was parallely desaturated to 22:5ω6.

Conclusions: The present "in vitro" study demonstrates that Δ4-desaturase can be transfected into human cells and synthesize DHA (as well as 22:5ω6, DPA) from 22:5ω3 and 22:4ω6, respectively, by putative Δ4-desaturation. Even if this pathway may not be the physiological route for DHA biosynthesis "in vivo", the present study opens new perspectives for the treatment of patients within the ZS spectrum.
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http://dx.doi.org/10.1186/1476-511X-9-98DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2949860PMC
September 2010
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