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There are no targeted therapeutic modalities for triple-negative breast cancer (TNBC), thus it is associated with poor prognosis and worst clinical outcome. Here, our aim was to identify deregulated proteins in TNBC with potential therapeutic applications. Proteomics profiling of TNBC and normal breast tissues through two-dimensional electrophoresis and ESI-MS/MS mass spectrometry revealed the existence of 16 proteins (RhoGDI-2, HSP27, SOD1, DJ1, UBE2N, PSME1, FTL, SH3BGRL, and eIF5A-1) with increased abundance in carcinomas. We also evidenced for the first time the deregulation of COX5, MTPN and DB1 proteins in TNBC that may represent novel tumor markers. Particularly, we confirmed the overexpression of the Rho-GDP dissociation inhibitor 2 (RhoGDI-2) in distinct breast cancer subtypes, as well as in metastatic cell lines derived from lung, prostate, and breast cancer. Remarkably, targeted disruption of RhoGDI-2 by RNA interference induced mitochondrial dysfunction, and facilitated caspase-3 and -9 activation in two breast cancer cell lines. Moreover, suppression of RhoGDI-2 resulted in a robust sensitization of breast cancer cells to cisplatin therapy. In conclusion, we identified novel proteins deregulated in TNBC, and confirmed the overexpression of RhoGDI-2. We propose that RhoGDI-2 inhibition may be exploited as a potential therapeutic strategy along cisplatin-based chemotherapy in breast cancer. Biological significance There are no useful biomarkers neither targeted therapeutic modalities for triple-negative breast cancer, which highly contributes to the poor prognosis of this breast cancer subtype. In this work, we used two-dimensional electrophoresis and ESI-MS/MS spectrometry to identify novel deregulated proteins in breast cancer tissues. Particularly, our results showed that RhoGDI-2, a protein that has been associated to metastasis and poor survival in human cancers, is overexpressed in different subtypes of breast tumors, as well as in metastatic cell lines derived from lung, prostate, and breast cancer. Our data also provided novel insights about the role of RhoGDI-2 in apoptosis through intrinsic pathway inhibition. Importantly, they suggested that targeted modulation of RhoGDI-2 levels might be a useful strategy for breast cancer therapy. This article is part of a Special Issue entitled: Proteomics, mass spectrometry and peptidomics, Cancun 2013. Guest Editors: Cesar Lopez-Camarillo, Victoria Pando-Robles and Bronwyn Jane Barkla.

We report the epidemiological impact of carbapenemase-producing Enterobacteriaceae (CPE) in Spain in 2012. Of the 237 carbapenemases detected, 163 were from the OXA-48 group, 60 were from VIM-1, 8 were from KPC-2, 5 were from IMP, and 1 was from NDM-1. Interhospital spread of carbapenemase-producing Klebsiella pneumoniae was due to a limited number of multilocus sequence types (MLST) and carbapenemase types, including ST15-VIM-1, ST11-OXA-48, ST405-OXA-48, ST101-KPC-2, and ST11-VIM-1. The number of CPE cases in Spain has increased sharply in recent years, due mainly to the emergence of OXA-48. This study was supported by the Antibiotic Resistance Surveillance Program of the Spanish Centro Nacional de Microbiología of the Instituto de Salud Carlos III. Verónica Bautista was supported by the Ministerio de Economía y Competitividad, Instituto de Salud Carlos III, cofinanced by the European Development Regional Fund “A way to achieve Europe” ERDF, Spanish Network for the Research in Infectious Diseases (REIPI RD12/0015). Sí

microRNAs are small non-coding RNAs of ~22 nucleotides that function at post-transcriptional level as negative regulators of gene expression. Aberrant expression of microRNAs could promote uncontrolled proliferation, migration and invasion of human cancer cells. In this study, we analyzed the expression of microRNA-18b (miR-18b) in breast cancer cell lines and in a set of clinical specimens. Our results showed that miR-18b was upregulated in four out of five breast cancer cell lines and also in breast tumors. In order to identify potential gene targets, we carried out transcriptional profiling of MDA-MB-231 breast cancer cells that ectopically expressed miR-18b. Our results showed that 263 genes were significantly modulated in miR-18b-deficient cells (fold change >1.5; P≤0.05). We found that knock-down of miR-18b induced the upregulation of 55 olfactory receptor (OR) genes and nine genes (NLRP7, KLK3, OLFM3, POSTN, MAGED4B, KIR3DL3, CRX, SEMG1 and CEACAM5) with key roles in cell migration and metastasis. Consistently, we found that ectopic inhibition of miR-18b suppressed the migration of two breast cancer cell models in vitro. In conclusion, we have uncovered genes directly or indirectly modulated by miR-18b which may represent potential therapeutic targets in breast cancer. Our data also pointed out a role of miR-18b in migration of breast cancer cells.

Benign and malignant pathology can develop in ectopic axillary breast tissue, such as fibroadenomas, phyllodes tumors, and breast cancer. We present a rare case of an asymptomatic 43-year-old woman with an axillary nodule which was identified during screening mammography within ectopic axillary breast tissue, initially considered as a suspicious lymph node. Radiologic studies were considered as Breast Imaging-Reporting Data System (BI-RADS) 4. A hyperdense, lobular, and well-circumscribed nodule was identified in mammogram while the nodule by ultrasound (US) was hypoechoic with indistinct microlobular margins, without vascularity by Doppler, and measuring 1.26×1 cm. Core-needle biopsy reported a fibroepithelial neoplasm. The patient was submitted to local wide-needle excision located in intraoperative radiography of the surgical specimen and margin evaluation. Final histopathological study reported a 1.8×1.2 cm benign phyllodes tumor, with irregular, pushing, and clear wide margins within normal ectopic breast tissue. The patient without surgical complications continued annual screening without recurrence during a follow-up that took place 24 months later.

Deregulated expression of microRNAs has been associated with angiogenesis. Studying the miRNome of locally advanced breast tumors we unsuspectedly found a dramatically repression of miR-204, a small non-coding RNA with no previous involvement in tumor angiogenesis. Downregulation of miR-204 was confirmed in an independent cohort of patients and breast cancer cell lines. Gain-of-function analysis indicates that ectopic expression of miR-204 impairs cell proliferation, anchorage-independent growth, migration, invasion, and the formation of 3D capillary networks in vitro. Likewise, in vivo vascularization and angiogenesis were suppressed by miR-204 in a nu/nu mice model. Genome-wide profiling of MDA-MB-231 cells expressing miR-204 revealed changes in the expression of hundred cancer-related genes. Of these, we focused on the study of pro-angiogenic ANGPT1 and TGFβR2. Functional analysis using luciferase reporter and rescue assays confirmed that ANGPT1 and TGFβR2 are novel effectors downstream of miR-204. Accordingly, an inverse correlation between miR-204 and ANGPT1/TGFβR2 expression was found in breast tumors. Knockdown of TGFβR2, but not ANGPT1, impairs cell proliferation and migration whereas inhibition of both genes inhibits angiogenesis. Taken altogether, our findings reveal a novel role for miR-204/ANGPT1/TGFβR2 axis in tumor angiogenesis. We propose that therapeutic manipulation of miR-204 levels may represent a promising approach in breast cancer.

Objective To show the incidence, as well as the clinical and histopathological characteristics, of patients diagnosed with mammary salivary gland-like carcinomas at our institution. Materials and methods A retrospective study was conducted in all women diagnosed with breast cancer at our institution from January 2005 to February 2016. Patients with diagnosis of salivary gland-like breast carcinomas were included. Results In this period, 6384 patients were diagnosed with breast cancer at our institution; salivary gland-like carcinomas were found in 7 patients (0.1%), adenoid cystic carcinoma was diagnosed in 5 patients (0.07%), acinic cell carcinoma in 1 patient (0.015%) and mucoepidermoid carcinoma in 1 patient (0.015%). The triple-negative subtype was found in all of the tumors. Median follow-up was 66.3 months (range, 1–108 months). No patient developed local or distant recurrence. Conclusions Salivary gland-like breast tumors are extremely rare. We found a global incidence of 0.1%. Adenoid cystic, acinic cell and mucoepidermoid carcinomas were the three histologic types diagnosed. Although the triple-negative subtype is mainly found, good prognosis is expected.

Heterogeneity of breast cancer can be classified according to different molecular signatures, including gene expression profiles that differentiate breast cancer into 5 clinically relevant subtypes: luminal A, luminal B, Her2-like, basal-like and claudin-low. Additionally the expression patterns of gene expression regulators like microRNAs can separate normal tissue from breast tumors, however there is currently limited information about the potential differences in microRNA expression profiles between the different tumor subtypes defined by gene expression. In order to determine microRNA expression profiles in breast tumors, and explore differences in the expression patterns of these molecules between different cancer subtypes, we analyzed the expression of 664 microRNAs with the TaqMan low-density array platform in 40 breast tumors from different subtypes. Tumor sub-typing was carried out with the PAM50 algorithm in expression data obtained with the Affymetrix Human Gene ST 1.0 array, obtaining 8 Luminal A tumors, 9 luminal B, 8 Her2-like and 3 Triple negative basal-like tumors, along with a set of microRNAs that distinguish each tumor subtype. Bioinformatic analysis of mRNA targets of the differentially expressed miRNAs, identified oncogenes like ERBB2, YY1, several MAP kinases, and known tumor-suppressors like FOXA1 and SMAD4. Pathway analysis identified that some biological process that are important in breast carcinogenesis are affected by the altered miRNA expression, including signaling through MAP kinases, RAS, programmed cell death and ERBB2-ERBB3 signaling. Given the clinical importance of the triple negative tumors, we included 48 additional triple-negative tumors defined by immunohistochemistry, to analyze the intra-heterogeneity of these phenotype based in the microRNA expression profiles. After the bioinformatics analysis with the algorithm Consensus Clustering we determined 6 sub-groups of triple negative tumors with different molecular, inmunophenotypical and clinical issues. We also compared the microRNA expression patterns between triple negative and other inmunophenotype (ER+, PR+, Her2+) tumors, detecting 56 differentially expressed microRNAs. Transcriptional targets of these microRNAs include genes involved in the carcinogenesis of triple negative tumors, like PARP1, or in the microRNA biogenesis machinery, like Dicer. Enrichment ontology analysis of the microRNAs differentially expressed in the triple negative tumors, detected pathways like p53 and focal adhesion whose role in cancer development, invasion and metastasis might be crucial; and MAPK, which has been related to recurrence of TN tumors. Immunochemistry analysis on the microRNA biogenesis machinery including Dicer and Argonaute2 revels a down-regulation (approximately 20% less) of both proteins, mainly in TN tumors. Our data identified the altered expression of microRNAs whose aberrant expression might have an important impact on cancer-related cellular pathways and whose role in breast cancer has not been previously described. microRNAs expression is also capable to discriminate between different tumor subtypes and to determine microRNAs that classifies triple negative tumors into different subgroups. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P5-10-12.

Breast carcinoma is the leading cause of cancer-related mortality in women worldwide with an estimated 1.38 million new cases and 458,000 deaths in 2008 alone1. This malignancy represents a heterogeneous group of tumours with characteristic molecular features, prognosis, and responses to available therapy2–4. Recurrent somatic alterations in breast cancer have been described including mutations and copy number alterations, notably ERBB2 amplifications, the first successful therapy target defined by a genomic aberration5. Prior DNA sequencing studies of breast cancer genomes have revealed additional candidate mutations and gene rearrangements 6–10. Here we report the whole-exome sequences of DNA from 103 human breast cancers of diverse subtypes from patients in Mexico and Vietnam compared to matched-normal DNA, together with whole-genome sequences of 22 breast cancer/normal pairs. Beyond confirming recurrent somatic mutations in PIK3CA11, TP536, AKT112, GATA313, and MAP3K110, we discovered recurrent mutations in the CBFB transcription factor gene and deletions of its partner RUNX1. Furthermore, we have identified a recurrent MAGI3-AKT3 fusion enriched in triple-negative breast cancer lacking estrogen and progesterone receptors and ERBB2 expression. The Magi3-Akt3 fusion leads to constitutive activation of Akt kinase, which is abolished by treatment with an ATP-competitive Akt small-molecule inhibitor.

Little information is available about pediatric infections caused by extended-spectrum-β-lactamase (ESBL)-producing Escherichia coli . We characterized an outbreak caused by a CTX-M-14-producing E. coli isolate in a neonatal intensive care unit (NICU) and studied other infections caused by ESBL-producing E. coli in non-NICU pediatric units. All children ≤4 years old who were infected or colonized by ESBL-producing E. coli isolates between January 2009 and September 2010 were included. Molecular epidemiology was studied by phylogroup analysis, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing. Antibiotic resistance genes were analyzed by PCR and sequencing. Plasmids were studied by PFGE with S1 nuclease digestion and by incompatibility group analysis using a PCR-based replicon-typing scheme. Of the ESBL-producing E. coli isolates colonizing or infecting the 30 newborns, identical PFGE results were observed for 21 (70%) isolates, which were classified as CTX-M-14-producing E. coli of ST23 phylogroup A. bla CTX-M-14a was linked to IS Ecp1 and was carried on an ∼80-bp IncK plasmid. A smaller ongoing outbreak due to SHV-12-producing ST131 E. coli was also identified in the same NICU. Fifteen additional infections with ESBL-producing E. coli were identified in non-NICU pediatric units, but none was caused by the CTX-M-14-producing E. coli epidemic clone. Overall, CTX-M-14 (71.1%), CTX-M-15 (13.3%), and SHV-12 (13.3%) were the most important ESBLs causing pediatric infections in this study. Infections of newborns with CTX-M-14-producing E. coli were caused by both clonal and nonclonal isolates.

Staphylococcus aureus subsp. anaerobius, a microaerophilic, catalase-negative bacteria, is the etiological agent of abscess disease, a specific chronic condition of sheep and goats, characterized by the formation of necrotic lesions that are typically located in superficial lymph nodes. In this study, molecular analysis including pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and accessory gene regulator (agr) typing was carried out on 94 S. aureus subsp. anaerobius strains isolated in different countries (79 were isolated from 35 outbreaks of the disease in Spain from 1981 to 2009, 9 were isolated in Italy, 3 in Denmark and 3 in Sudan). All of the 94 S. aureus subsp. anaerobius isolates examined belonged to one PFGE type, within which four minority subtypes were identified. Representative isolates of all PFGE subtypes as well of all countries belonged to the same sequence type (ST), ST1464, which was a singleton, and to the agr type II. Our results support the view that abscess disease is caused by a single bacterial clone worldwide. This bacterium has existed for at least a century and, thus, has undergone long-term small ruminant host restriction.

Los hemangiomas espinales epidurales; son entidades raras. La mayoría de estas lesiones afectan los cuerpos vertebrales y tienen extensión ocasional al espacio epidural. La ocurrencia pura (espinal) de hemangioma epidural es poco común, localizados con más frecuencia en nivel torácico, puede presentarse con síntomas sobre raíz nerviosa y/o compresión medular. Presentamos un caso de hemangioma cavernoso espinal epidural puro a nivel lumbar, manifestado con radiculopatía crónica, con mejoría posterior a la resección quirúrgica total.

To document fosfomycin susceptibility of extended-spectrum β-lactamase-producing Escherichia coli (ESBL-EC), analyse trends in fosfomycin use and investigate fosfomycin resistance in ESBL-EC isolated from urinary tract infections (UTIs).Twenty-seven Spanish hospitals participating in the European Antimicrobial Resistance Surveillance Network were requested to collect up to 10 sequential ESBL-EC for centralized susceptibility testing and typing. EUCAST guidelines were followed for antibiotic susceptibility testing, and bla(ESBL) type, phylogroups and O25b serotype were determined by PCR and sequencing. In addition, the trend in fosfomycin resistance among ESBL-EC causing UTIs was determined in 9 of the 27 hospitals. Total fosfomycin use for ambulatory care was established by WHO-recommended methods.A total of 231 ESBL-EC (42.4% CTX-M-15, 34.2% SHV-12 and 23.4% CTX-M-14) were collected. The overall rate of fosfomycin resistance was 9.1%, but varied according to ESBL type (5.6% of CTX-M-14 isolates, 5.1% of SHV-12 and 15.3% of CTX-M-15). Of 67 O25b/B2 isolates, 11 (16.4%) were fosfomycin resistant. Predictors of infection with fosfomycin-resistant ESBL-EC were O25b/phylogroup B2 isolates, female gender and nursing home residence. Among 114 197 UTIs caused by E. coli 4740 (4.2%) were due to ESBL-EC. Fosfomycin resistance increased in these isolates from 4.4% (2005) to 11.4% (2009). The use of fosfomycin grew from 0.05 defined daily doses per 1000 inhabitants per day (1997) to 0.22 (2008), a 340% increase.Key factors related to increased fosfomycin resistance in ESBL-EC causing UTIs could be the rapid growth in community use of fosfomycin, the widespread distribution of the 025b/B2 E. coli clone and the existence of a susceptible population comprising women residing in nursing home facilities.

Cephalosporins resistance is increasing in Escherichia coli in Spain. We characterize infections by E. coli with reduced susceptibility to third-generation cephalosporins (3GCs) with the AmpC phenotype. Between January 2004 and March 2007, 121 E. coli isolates with the AmpC phenotype were collected (4.8% of all the 2538 E. coli isolates with reduced susceptibility to 3GCs). These isolates were further characterized by clinical and molecular analysis. Plasmid-encoded ampC genes were detected in 46 (38%) isolates (43 CMY-2); 75 isolates (62%) had modifications in the chromosomal ampC promoter region (c-AmpC). CMY-2 producers belonged primarily to the more virulent phylogroup D (48.4%), whereas most isolates of c-AmpC belonged to phylogroup A (56.4%). Bacteremia and infections in children were more frequently produced by CMY-2 producers. CMY-2-producing phylogroup D E. coli belonged to 8 multilocus sequence typing types. Three CMY-2 producers belonged to O25b/ST131/B2 clone. Infections caused by E. coli with the AmpC phenotype may be spreading primarily because of CMY-2-producing phylogroup D isolates, although this enzyme was also detected in the O25b/ST131/B2 clone.

In this study, we investigated the population structure of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli in Spain and determined possible associations between specific multilocus sequence typing (MLST) types and ESBL types. Ninety-two ESBL-producing E. coli isolates from 11 Spanish hospitals were studied. The predominant ESBLs in this collection were CTX-M-14 (45.7%), SHV-12 (21.7%) and CTX-M-9 (20.6%). Phylogenetic groups and MLST types were studied. Thirty-seven isolates (40.2%) belonged to phylogroup A, 26 (28.3%) to group B1, 13 (14.1%) to group B2 and 16 (17.4%) to group D. Fifty-six sequence types (STs) were identified, but ST131 (eight isolates) and ST167 (five isolates) were the most prevalent. The most common ST complexes were ST10 (13 isolates; 14.3%) and ST23 (10 isolates; 11%). Escherichia coli ST131 carried six different ESBLs (CTX-M-1, CTX-M-9, CTX-M-10, CTX-M-14, CTX-M-15 and SHV-12), E. coli ST10 complex carried five ESBLs and E. coli ST23 complex carried four ESBLs. A great diversity of MLST types was observed among Spanish ESBL-producing E. coli isolates.

OBJECTIVES: To characterize the population structure and resistance mechanisms of Klebsiella pneumoniae isolates that are highly resistant to third-generation cephalosporins, collected from five Spanish hospitals.METHODS: A total of 162 K. pneumoniae isolates from five hospitals located in three geographical areas of Spain were characterized. The number of isolates from each hospital ranged from 3 to 82. The genetic relationship between isolates was established by PFGE and multilocus sequence typing (MLST). bla(ESBL) types and other antibiotic resistance genes were analysed by PCR and sequencing. Plasmids were classified according to their incompatibility group by a PCR-based replicon-typing scheme.RESULTS: All 162 isolates carried the bla(CTX-15) gene. Fifty-eight isolates (35.8%) caused clinical infections and 104 (64.2%) were colonizers. Sixty-nine (42.6%) isolates were collected from newborns and 93 (57.4%) from adults. Using PGFE, the 162 isolates were grouped into seven clusters that were further identified as members of the MLST types 1, 11, 14, 17, 20, 35 and 36. Two hospitals each had two different clones and the remaining three hospitals had a single CTX-M-15-producing K. pneumoniae clone. All clones carried different antibiotic resistance genes, including bla(OXA-1), aac(3)-IIa, aac(6')-Ib-cr, qnrS1 and qnrB. In four of the seven (57.1%) clones the bla(CTX-M-15) gene was transferred by conjugation; in all cases plasmids of the incompatibility group IncF were identified by PCR.CONCLUSIONS: This study shows that multiresistant K. pneumoniae producing CTX-M-15 of MLST types 1, 11, 14, 17, 20, 35 and 36 are spreading as pathogens and colonizers among newborns and adult patients in Spain.

OBJECTIVES: To describe trends in fosfomycin resistance in urinary isolates of Escherichia coli producing extended-spectrum beta-lactamases (ESBLs) in relation to fosfomycin consumption and to characterize representative fosfomycin-resistant isolates.METHODS: In 2007-08, an unexpected increase in fosfomycin resistance in ESBL-producing urinary E. coli was observed. Laboratory records were reviewed and a prospective surveillance study was initiated on all urinary tract infections caused by ESBL-producing, fosfomycin-resistant E. coli. bla(ESBL) types, phylogroups, genetic environment and afa/dra operon were determined by PCR and sequencing. Molecular epidemiology was analysed by PFGE and multilocus sequence typing. To elucidate possible mechanisms of fosfomycin resistance, uhpT, glpT, uhpA, ptsI, cyaA and murA genes were analysed. Fosfomycin consumption was determined as recommended by WHO.RESULTS: From 2004 to 2008, fosfomycin consumption increased by 50%, while fosfomycin resistance in ESBL producers increased from 2.2% to 21.7%. Of 26 isolates studied, 24 produced CTX-M-15 and belonged to the O25b-ST131-phylogroup B2 clonal strain. PFGE revealed two clusters. Cluster I included 18 isolates, 16 of them indistinguishable from strains producing CTX-M-15 previously described in Madrid. The five isolates of Cluster II had the IS26 linked to bla(CTX-M-15) and the afa/dra operon. In Cluster I isolates, no mutations in glpT, uhpT, uhpA, ptsI, cyaA and murA were detected. Cluster II isolates showed a 15 bp deletion (A(169)-C(183)) in uhpA.CONCLUSIONS: Fosfomycin resistance in urinary E. coli has increased due to the acquisition of this resistance by a previously circulating CTX-M-15-producing E. coli O25b-ST131-phylogroup B2 strain. This happened during a period when the use of fosfomycin increased by 50%.

The molecular epidemiology and the mechanisms of resistance of Escherichia coli isolated from two patients infected by imipenem-resistant strains are reported in this study. From one patient, three closely related consecutive isolates of E. coli were recovered; the first was carbapenem-susceptible but acquired imipenem resistance after treatment with ertapenem, and the third isolate was again imipenem-susceptible. An additional imipenem-resistant isolate was recovered from another patient who received imipenem. The genetic relatedness of the E. coli isolates was determined by pulsed-field gel electrophoresis (PFGE) after digestion with XbaI. Standard polymerase chain reaction (PCR) conditions were used to amplify several beta-lactamase genes coding for carbapenemases, extended-spectrum beta-lactamases (ESBLs) and plasmid-mediated AmpC; the E. coli ampC gene promoter was also amplified and sequenced. Primers OmpF-F/OmpF-R and OmpC-F/OmpC-R were used to amplify the ompF and ompC genes. The outer membrane protein (OMP) profiles were studied by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Imipenem-resistant E. coli isolates did not produce carbapenemases but lacked the two major OMPs OmpF and OmpC and had ampC promoter mutations; in addition, one of the imipenem-resistant isolates produced the CMY-2 cephalosporinase, whilst the other produced the new CTX-M-67 ESBL. Carbapenem resistance in this study was associated with lack of expression of OmpF and OmpC porins. Additional mechanisms of beta-lactam resistance, such as plasmid-mediated AmpC and ESBL production, were also found. Development of carbapenem resistance in a CTX-M-67-producing E. coli is first described in this study.

Summary Background Most available information on carbapenemase-producing Enterobacteriaceae (CPE) is usually associated with specific types of infection or patient or with descriptions of outbreaks. The aim of this study was to comprehensively analyse the clinical epidemiology, clinical features and outcomes of colonisation and infections due to CPE in Spain. Methods A multicentre prospective cohort study was carried out in 34 Spanish hospitals from February to May 2013. All new patients testing positive for CPE in clinical samples were included. Logistic regression was used to identify predictors of mortality. Results Overall, 245 cases were included. The most frequent organism was Klebsiella pneumoniae (74%) and the carbapenemases belonged to the OXA-48 (74%), metallo-β-lactamase (MBL) (24%) and KPC (2%) groups. Acquisition was nosocomial in 145 cases (60%) and healthcare-associated (HCA) in 91 (37%); 42% of the latter were nursing home residents, in whom OXA-48-producing K. pneumoniae ST405 predominated. MBLs and OXA-48 predominated in ICU and medical patients, respectively. Overall, 67% of patients had infections. The most frequent infections identified in this study were urinary tract (43%) and skin structure (21%) infections, and 10% of infections were bacteraemic. Crude mortality was 20%. Inappropriate antibiotic therapy was independently associated with an increased risk of death (OR = 3.30; 95% CI: 1.34–8.11). Conclusions We found some differences in the epidemiology of CPE depending on the type of carbapenemase produced. Although a low proportion of CPE infections were bacteraemic, active antibiotic therapy was a protective factor for reducing mortality.

This paper aims to study the perceptions of students at Secondary School about the transfer of values in Internet. This is a descriptive research which has involved 94 students on the 3rd and 4th courses. Results have been obtained regarding the identification of values present in the network, the main agents of transmission of values, as well as the positive and negative aspects that ICT provides to transmit values. In addition, we analyzed the frequency with which the students put into practice some values through telematic networks and tools they use to do that. Conclusions, show up, the consistency between the common practices in the network and the most frequent values in these contexts (creativity, dialogue and collaboration). They also point to the increasing role of the family and the peer group in the transmission of values in the network., El presente trabajo tiene como propósito conocer la percepción del alumnado de Educación Secundaria sobre la transmisión de valores en Internet. Se trata de una investigación descriptiva en la que han participado 94 alumnos de 3.º y 4.º de ESO, en ella se han obtenido resultados respecto a la identificación de valores presentes en la Red, sobre los principales agentes de transmisión de valores, así como sobre los aspectos positivos y negativos que proporcionan las TIC para transmitir dichos valores. Además, se ha analizado la frecuencia con la que el alumnado pone en práctica algunos valores a través de las redes telemáticas y las herramientas que utiliza para hacerlo. Entre sus conclusiones destacan, por un lado, la coherencia entre las prácticas habituales en la Red y los valores más frecuentes en estos contextos (creatividad, diálogo y colaboración) y, por otro, el mayor protagonismo de la familia y el grupo de iguales en la transmisión de valores en la Red., Ce document vise à étudier la perception des élèves dans le secondaire sur le transfert des valeurs mobilières de l’Internet. Ce est une recherche descriptive qui a impliqué 94 étudiants. Les résultats ont été obtenus en ce qui concerne l’identification des valeurs présentes dans le réseau, les principaux agents de transmission des valeurs, ainsi que les aspects positifs et négatifs qui offrent les TIC pour transmettre des valeurs. En outre, nous avons analysé la fréquence avec laquelle les élèves mettent en pratique certaines valeurs à travers des réseaux télématiques et les outils qu’ils utilisent pour le faire. Parmi ses conclusions en évidence d’une part, la cohérence entre les pratiques courantes dans le réseau et les valeurs les plus fréquentes dans ces contextes (créativité, dialogue et collaboration), et d’autre part, le rôle croissant de la famille et le groupe de pairs dans le transmission des valeurs dans le réseau.

A total of 151 Escherichia coli strains resistant to cefotaxime and ceftazidime were isolated during a prospective surveillance study. These strains were characterized by clinical, microbiological, and molecular analyses and were distributed into four clusters of 103, 11, 6, and 5 isolates, along with 25 unrelated strains. The principal cluster was isolated from urine, wound, blood, and other samples in three hospitals, eight nursing homes, and a community healthcare center. This cluster was associated with both nosocomial (65%) and community-acquired (35%) infections. Most strains were resistant to ciprofloxacin, gentamicin, tobramycin, cefepime, amoxicillin-clavulanic acid, and trimethoprim-sulfamethoxazole but were susceptible to imipenem. All isolates from the four clusters expressed the extended-spectrum β-lactamase (ESBL) CTX-M-15. This enzyme was also present in 8 (30.8%) of the 26 unrelated isolates. The other ESBLs, CTX-M-14 and CTX-M-32, were detected in five and seven cases, respectively, but they were detected in individual E. coli isolates only. In three clusters, bla CTX-M-15 alleles were linked to an IS Ecp1 -like element, while in eight strains of cluster II an IS 26 element preceded the bla CTX-M-15 allele. An additional pool of resistance genes included tetA , drfA14 or dfrA17 , sul1 or sul2 , aac(6′)Ib , and aac(3)IIb . All except one of the 27 isolates tested for genetic virulence markers harbored the same three virulence genes: iutA and fyuA (siderophores), and traT (serum survival factor). Epidemic or occasional isolates of cefotaxime- and ceftazidime-resistant E. coli can spread between distinct health facilities including hospitals, community health centers, and long-term care centers.

Background: Asymptomatic cases of phyllodes tumor (PT) are extremely rare. The objective of the study was to describe the radiological features and to identify clinical differences between asymptomatic and symptomatic patients with PT in a Mexican population. Methods: A retrospective study was conducted including only surgically treated patients. Asymptomatic and symptomatic cases were compared. Descriptive statistical techniques were used. Radiological features were reported in asymptomatic cases. Results: Over 10 years, 305 women with PT were included (307 tumors). The 25 (8.1%) asymptomatic patients were 6 years older than symptomatic patients (47.6 vs . 41.2 years; P=0.005), with more pregnancies (3 vs . 2; P=0.013), more breastfeeding (80% vs . 61%; P=0.042), with smaller tumor size in radiological, and pathology studies (P vs . 68%; P=0.005), and more benign PT (88%). Mammographic images in asymptomatic cases showed a nodule, mostly isodense, and with well-circumscribed margins. Ultrasound (US) demonstrated macrolobular nodules, with hypoechoic pattern and well-circumscribed margins. Percutaneous core-needle biopsies reported 45.5% as PT in asymptomatic cases. Conclusions: As we expected, asymptomatic PTs were smaller than symptomatic cases. Even asymptomatic PT was not to high suspicious nodules in older patients, we recommend excisional biopsy because radiological features are indistinguishable from those of fibroadenomas, and the only

Sentinel lymph node biopsy in patients with ductal carcinoma in situ still controversial, with positive lymph node in range of 1.4-12.5% due occult invasive breast carcinoma in surgical specimen.To know the frequency of sentimel node metastases in patients with ductal carcinoma in situ, identify differences between positive and negative cases.Retrospective study of patients with ductal carcinoma in situ treated with sentinel lymph node biopsy because mastectomy indication, palpable tumor, radiological lesion = 5 cm, non-favorable breast-tumor relation and/or patients whom surgery could affect lymphatic flow drainage.Of 168 in situ carcinomas, 50 cases with ductal carcinoma in situ and sentinel lymph node biopsy were included, with a mean age of 51.6 years, 30 (60%) asymptomatic. The most common symptoms were palpable nodule (18%), nipple discharge (12%), or both (8%). Microcalcifications were common (72%), comedonecrosis pattern (62%), grade-2 histology (44%), and 28% negative hormonal receptors. Four (8%) cases had intra-operatory positive sentinel lymph node and one patient at final histo-pathological study (60% micrometastases, 40% macrometastases), all with invasive carcinoma in surgical specimen. Patients with intra-operatory positive sentinel lymph node where younger (44.5 vs 51 years), with more palpable tumors (50% vs 23.1%), and bigger (3.5 vs 2 cm), more comedonecrosis pattern (75% vs 60.8%), more indifferent tumors (75% vs 39.1%), and less cases with hormonal receptors (50% vs 73.9%), compared with negative sentinel lymph node cases, all these differences without statistic significance.One of each 12 patients with ductal carcinoma in situ had affection in sentinel lymph node, so we recommend continue doing this procedure to avoid second surgeries due the presence of occult invasive carcinoma.Antecedentes: en pacientes con carcinoma ductal in situ la biopsia de ganglio centinela es motivo de controversia porque se reportan ganglios positivos en 1.4-12.5% debido al carcinoma invasor oculto en la pieza quirúrgica. Objetivo: conocer la frecuencia de metástasis en ganglio centinela en pacientes con carcinoma ductal in situ e identificar las diferencias entre los casos positivos y negativos. Material y métodos: estudio retrospectivo, transversal, analítico de pacientes con carcinoma ductal in situ a quienes se realizó una biopsia de ganglio centinela por requerir mastectomía, tener un tumor palpable, lesión radiológica = 5 cm, inadecuada relación mama-tumor o porque la escisión pudiera afectar el flujo linfático. Resultados: de 168 carcinomas in situ, se incluyeron 50 casos con carcinoma ductal in situ y biopsia de ganglio centinela, de pacientes con edad promedio de 51.6 años, 30 (60%) de ellas asintomáticas. Los signos reportados fueron: nódulo palpable (18%), secreción por el pezón (12%) o ambos (8%). Predominaron las microcalcificaciones (72%), comedonecrosis (62%) y grado histológico -2 (44%) con 28% de receptores hormonales negativos. En el estudio transoperatorio 4 (8%) pacientes tuvieron ganglio centinela positivo y un caso en estudio histopatológico definitivo (60% micrometástasis, 40% macrometástasis), todos con carcinoma invasor en la pieza quirúrgica. Las pacientes con ganglio centinela transoperatorio positivo eran más jóvenes (44.5 vs 51 años), con más tumores palpables (50 vs 23.1%), más grandes (3.5 vs 2 cm), más comedonecrosis (75 vs 60.8%), más indiferenciados (75% vs 39.1%) y menos receptores hormonales (50 vs 73.9%), que las que tenían ganglio centinela negativo, sin que estas diferencias tuvieran significación estadística. Conclusiones: puesto que 1 de cada 12 pacientes con carcinoma ductal in situ tiene afectación ganglionar en el ganglio centinela, se recomienda seguir tomando la biopsia para evitar segundas cirugías por un carcinoma invasor oculto.

Several isolates of four different carbapenemase-producing Enterobacteriaceae species were recovered from a patient hospitalized for 4 months in a teaching hospital in Madrid. These species comprised seven Klebsiella pneumoniae belonging to ST15, four Escherichia coli belonging to ST2531, two Serratia marcescens and one Citrobacter freundii. This patient was the index case of a small outbreak of four patients infected and/or colonized by carbapenemase-producing K. pneumoniae. Molecular results identified the blaOXA-48 gene in all Enterobacteriaceae isolates from the index case and in all isolates from the other three patients, suggesting intra- and interpatient dissemination. Our results highlight the great ability of OXA-48 carbapenemase to spread among different enterobacterial species by both clonal and nonclonal dissemination.

In tumor cells the effectiveness of anti-neoplastic agents that cause cell death by induction of DNA damage is influenced by DNA repair activity. RAD50 protein plays key roles in DNA double strand breaks repair (DSBs), which is crucial to safeguard genome integrity and sustain tumor suppression. However, its role as a potential therapeutic target has not been addressed in breast cancer. Our aim in the present study was to analyze the expression of RAD50 protein in breast tumors, and evaluate the effects of RAD50-targeted inhibition on the cytotoxicity exerted by cisplatin and anthracycline and taxane-based therapies in breast cancer cells. Immunohistochemistry assays on tissue microarrays indicate that the strong staining intensity of RAD50 was reduced in 14% of breast carcinomas in comparison with normal tissues. Remarkably, RAD50 silencing by RNA interference significantly enhanced the cytotoxicity of cisplatin. Combinations of cisplatin with doxorubicin and paclitaxel drugs induced synergistic effects in early cell death of RAD50-deficient MCF-7, SKBR3, and T47D breast cancer cells. Furthermore, we found an increase in the number of DSBs, and delayed phosphorylation of histone H2AX after cisplatin treatment in RAD50-silenced cells. These cellular events were associated to a dramatical increase in the frequency of chromosomal aberrations and a decrease of cell number in metaphase. In conclusion, our data showed that RAD50 abrogation impairs DNA damage response and sensitizes breast cancer cells to cisplatin-combined therapies. We propose that the development and use of inhibitors to manipulate RAD50 levels might represent a promising strategy to sensitize breast cancer cells to DNA damaging agents.

Pediatric patients are rarely infected with metallo-β-lactamase-producing Enterobacteriaceae. We describe 3 cases of children infected with VIM-1-producing clonal Enterobacter cloacae. Patients were treated with amikacin and cotrimoxazole. The blaVIM-1 gene was carried into a class 1 integron and an IncHI2 incompatibility group plasmid. Emergence of pediatric infections caused by carbapenemases-producing Enterobacteriaceae is a critical issue as they are resistant to most β-lactam antibiotics.

Triple negative breast cancer (TNBC) represents a challenging tumor type due to their poor prognosis and limited treatment options. It is well recognize that clinical and molecular heterogeneity of TNBC is driven in part by post-transcriptional regulators such as miRNAs. To stratify TNBCs, we profiled 1050 miRNAs in 132 adjuvant TNBC tumors and 40 tumors from other immunophenotypes using an Affymetrix microarray platform. A NMF clustering analysis allowed us to identify 4 TNBC subtypes featuring unique miRNA expression patterns, disease free and overall survival rates and particular gene ontology enrichments (performed with GSEA algorithm). Our agglomerative approach was cross-validated by using two other clustering algorithms (k-means and consensus clustering). TNBC miRNAs subgroups were also correlated with the Lehmann intrinsic subtypes, finding a significant enrichment of immmnomodulartory and basal 1 subtypes in our high risk miRNA subgroups. 3 cell line models (MDA MB 468, MDA MB 231 and HS578T) were classified according to our miRNA signature, recapitulating two different miRNA subgroups. The TNBC tumors were compared against other phenotypes identifying differentially expressed miRNAs that together with the altered miRNAs within the subgroups allowed us to define interesting miRNAs for further functional analysis. We found low expression levels of miR-342-3p in TNBC tumors compared with other breast cancer phenotypes, and this down-regulation characterizes one of our miRNA subgroups with high risk to relapse. To characterize the functional its functional role, miR-342-3p was transiently transfected in the cell line MDA-MB-468, showing a decrease in cell proliferation, viability and migration rates. A gene expression profile (Human gene st, Affymetrix) revealed 140 altered mRNAs, from which 35 are potential direct targets of miR-342-3p defined by an in-silico analysis. The monocarboxylate transporter 1(MCT1), was confirmed as one target of miR-342-3p by a luciferase assay and western blot analysis. MCT1 repression by the miRNA promotes lactate efflux changes in the tumor cells, reflected in the accumulation of exogenous lactate and the increase in levels of extracellular endogenous lactate together with a decrease level of intra and inter cellular glucose concentration. We also explored other mechanisms that can contribute to the modulation of lactate and glucose levels such as the expression of MCT4, HIF1alpha and Glut1, but not significant changes were observed, pointing out the important role of the regulatory circuit of miR-342-3p and MCT1 in a particular TNBC tumor cells metabolism. These data suggest a metabolic change that favors a more glycolytic environment, which lead to a glucose deprivation context that may contribute to the reduction in proliferation, viability and migration capabilities already described. Furthermore, to define the main transcriptional modulator of the intronic miR-342-3p in TNBC, we evaluated the correlation between the expression of estrogen receptor, miR-342-3p and its host gene EVL in TNBC tumors from TCGA data. A significant correlation between these transcripts was observed. These results, together with information from other studies and Chip-seq analysis suggest that the ER activates the transcription of EVL and consequently the intronic miR-342-3p. We validated these correlations in our cellular model by transiently silencing of the ER. Our data provides evidence that miRNAs sub-classification signature can recapitulate the biological and clinical heterogeneity of TNBC. Moreover, show that miRNAs regulate important oncogenic pathways in TNBC cells such as: proliferation, cell movement and apoptosis in the different subgroups. Citation Format: Sandra L. Romero-Cordoba, Sergio Rodriguez-Cuevas, Rosa Rebollar-Vega, Veronica Bautista-Pina, Antonio Maffuz-Aziz, Elda Tagliabue, Marilena Iorio, Elvira D'Ippolito, Sara Baroni, Ilaria Plantamura, Alfredo Hidalgo-Miranda. A microRNA signature identifies subtypes of triple-negative breast cancer and reveals miR-342-3p as regulator of a lactate metabolic pathway through silencing monocarboxylate transporter 1. [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer: Mechanisms to Medicines ; 2015 Dec 4-7; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2016;76(6 Suppl):Abstract nr A47.

Newborns are rarely infected by extended-spectrum β-lactamase (ESBL)-producing members of the Enterobacteriaceae. In a neonatal intensive care unit, 14 newborns were infected or colonized by CTX-M-15-producing Enterobacter cloacae. All seven infected patients had underlying medical conditions, and five of them were treated successfully with meropenem, whilst one untreated patient died. Paediatric infections caused by multidrug-resistant ESBL-producing Enterobacter cloacae constitute a critical clinical and epidemiological issue.

Objectives To describe the molecular and population-level characterization of a selected group of OXA-48-like-producing Klebsiella pneumoniae isolates collected in Spain between January 2011 and May 2012. Methods During the study period, 151 OXA-48-like-producing K. pneumoniae isolates were collected from 10 hospitals in six different Spanish regions. From these, a representative sample of 21 isolates that caused hospital outbreaks and single infections was selected for further in-depth analysis. Molecular epidemiology was investigated using PFGE and multilocus sequence typing (MLST). Resistance genes were characterized by PCR and sequencing. Plasmids carrying bla(OXA-48-like) were studied by PFGE with S1 nuclease digestion. Results All 21 isolates had ertapenem MICs ≥ 1 mg/L, but 47.6% remained susceptible to imipenem and meropenem; bla(OXA-48) was identified in 19 isolates (90.5%) and the novel bla(OXA-244) and bla(OXA-245) genes were detected in 1 isolate each. With one exception, all isolates that contained bla(OXA-48-like) also contained bla(CTX-M-15). PFGE typing revealed six clusters comprising isolates that belonged to MLST types ST11, ST16, ST392, ST405, ST437 and ST663, respectively. Two main clusters were identified: PFGE cluster 1 (12 isolates, belonging either to ST405 or ST663, from seven hospitals), and PFGE cluster 2 (4 ST16 isolates from two hospitals). Six of seven donor isolates conjugated successfully; bla(OXA-48-like) (but not bla(CTX-M-15)) was carried on ≈ 60 kb Inc L/M plasmids. Conclusions Multidrug-resistant K. pneumoniae producing OXA-48-like carbapenemase are emerging as important pathogens in Spain due to intra- and inter-hospital, clonal and non-clonal dissemination.

Breast cancer is the neoplasia with the highest incidence in women worldwide. Proteomics approaches have accelerated the discovery of diagnostic and prognostic biomarkers. Here, we compared the proteomic profiles of breast tumors versus non-tumoral tissues in order to identify modulated proteins, which could represent potential markers associated to clinical features. By two-dimensional electrophoresis, we detected 28 differentially expressed proteins. Among these, 21 proteins were up-regulated and 7 were down-regulated in tumors (p0.05). Proteins were identified using LC/ESI-MS/MS tandem mass spectrometry. One protein was identified as glyoxalase 1 (GLO1), an enzyme involved in detoxification of methylglyoxal, a cytotoxic product of glycolysis. GLO1 overexpression was confirmed by western blot assays in paired normal and tumor breast tissues in clinical stages I-III, and by immunohistochemistry on tissue microarrays (TMA) comprising a cohort of 98 breast tumors and 20 healthy specimens. Results from TMA demonstrated that GLO1 is overexpressed in 79% of tumors. Interestingly, GLO1 up-regulation correlates with advanced tumor grade (p0.05). These findings demonstrate the association of GLO1 overexpression with tumor grade and pointed out for additional studies to establish the importance of GLO1 in breast cancer.

Fibromatosis is a term used to describe a group of lesions characterized by well-differentiated fibroblast proliferation with an usually benign biological behavior but with an infiltrative pattern of growth, frequently recurrent and locally invasive. It is generally localized in the retroperitoneal area, neck and extremities. The presence of this entity in breast tissue is an uncommon clinical situation, comprising approximately 0.2% of breast tumors and very often misdiagnosed as malignant disease or phyllodes tumor. In this rare condition, immunohistochemistry is an important diagnostic tool in anatomopathological differential diagnosis with other spindle cell tumors of the breast. The purpose of this paper is to report the experience with two cases of this rare condition.We present two cases of breast fibromatosis confirmed immunohistochemically and also by biopsy. One case had the clinical and imaging appearance of breast carcinoma with the classic irregular mass presentation and the other case was misdiagnosed as phyllodes tumor because of the size and density of the tumor and the cytology.Approximately 83 cases of breast fibromatosis have been reported during the last 30 years, including one male patient. We reviewed clinicopathological features of two cases of fibromatosis of the breast treated at our institute.

nonconcrete the mammary injuries are frequent in programs of detection of breast cancer, estereotaxic or ecographic marking is required to realize its split. The intrasurgical radiation control of the surgical piece is indispensable to evaluate the margins of the mammary cancer.to determine the effectiveness of the intrasurgical radiation control of the surgical piece in nonconcrete mammary injuries to diminish the surgical reinterventions to extend margins.women with nonconcrete mammary injuries to those who biopsy by split became, previous marking and intraoperating radiation control of the surgical piece to value margins (suitable margin the same or major of 10 mm, smaller inadequate margin of 10 mm). Intrasurgical reesicion in inadequate radiological margins became. The demographic characteristics, masto-ecographics images, histopathology of the injuries and the radiological-histopatol6gica correlation of the margins studied. Cross-sectional, prospective and descriptive study.103 patients with 113 nonconcrete mammary injuries included themselves, with age average of 51,35 (32-73) years. In all the injuries the intrasurgical radiation control became of the surgical piece. The prevalence of mammary cancer was of 28.3% (32/113), that corresponds to stellar images (42.8%), suspicious microcalcifications with density (39.2%), microcalcifications (31.2%) and nodules (20%). Of the 32 cancers, 16 had inadequate radiological margins that required intraoperating reescision; suitable histopatologic margins in 100% were obtained (16/16). The 16 (62.5%) cancers without intraoperating reescisi6n by suitable radiological margins had suitable histopatologic margins and 37.5% (6/16) inadequate ones that required surgical reinterventionn to control the margins. The discrepancy between margins was related to microcalcifications in 83.3% of the injuries.the intrasurgical radiation control of the surgical piece is effective to evaluate margins; the intrasurgical reescisión changed inadequate margins to suitable in 50% (16/32) of the cancers; only 18.7% (6/32) of the total of cases required another surgery to control the margins.

In a point-prevalence study performed in 145 Spanish hospitals in 2006, we collected 463 isolates of Staphylococcus aureus in a single day. Of these, 135 (29.2%) were methicillin (meticillin)-resistant S. aureus (MRSA) isolates. Susceptibility testing was performed by a microdilution method, and mecA was detected by PCR. The isolates were analyzed by pulsed-field gel electrophoresis (PFGE) after SmaI digestion, staphylococcal chromosomal cassette mec (SCC mec ) typing, agr typing, spa typing with BURP (based-upon-repeat-pattern) analysis, and multilocus sequence typing (MLST). The 135 MRSA isolates showed resistance to ciprofloxacin (93.3%), tobramycin (72.6%), gentamicin (20.0%), erythromycin (66.7%), and clindamycin (39.3%). Among the isolates resistant to erythromycin, 27.4% showed the M phenotype. All of the isolates were susceptible to glycopeptides. Twelve resistance patterns were found, of which four accounted for 65% of the isolates. PFGE revealed 36 different patterns, with 13 major clones (including 2 predominant clones with various antibiotypes that accounted for 52.5% of the MRSA isolates) and 23 sporadic profiles. Two genotypes were observed for the first time in Spain. SCC mec type IV accounted for 6.7% of the isolates (70.1% were type IVa, 23.9% were type IVc, 0.9% were type IVd, and 5.1% were type IVh), and SCC mec type I and SCC mec type II accounted for 7.4% and 5.2% of the isolates, respectively. One isolate was nontypeable. Only one of the isolates produced the Panton-Valentine leukocidin. The isolates presented agr type 2 (82.2%), type 1 (14.8%), and type 3 (3.0%). spa typing revealed 32 different types, the predominant ones being t067 (48.9%) and t002 (14.8%), as well as clonal complex 067 (78%) by BURP analysis. The MRSA clone of sequence type 125 and SCC mec type IV was the most prevalent throughout Spain. In our experience, PFGE, spa typing, SCC mec typing, and MLST presented good correlations for the majority of the MRSA strains; we suggest the use of spa typing and PFGE typing for epidemiological surveillance, since this combination is useful for both long-term and short-term studies.

We carried out a nationwide study aimed at the determination of the molecular epidemiology and antibiotic resistance mechanisms of invasive Staphylococcus aureus in 21 Spanish hospitals.The distributions of molecular markers, including antibiotic resistance genes, were investigated in 203 S. aureus, comprising 90 methicillin-resistant S. aureus (MRSA) and 113 methicillin-susceptible S. aureus (MSSA). Antimicrobial susceptibility was determined by standard methods. Panton-Valentine leucocidin (PVL) detection, staphylococcal cassette chromosome mec (SCCmec) types and agr types were performed/determined by PCR. All isolates were genotyped by PFGE after digestion of chromosomal DNA with SmaI. Multilocus sequence typing and spa-typing were also performed.In MRSA isolates, 74.4% were agr allotype II and were positive for SCCmec IV. Sixty-nine spa-types were identified, 18 in MRSA and 57 in MSSA. Both MRSA and MSSA variants were detected in six spa-types (8.7%). The majority of S. aureus (51.2%) were grouped into four spa-types (t067, t002, t012 and t008). The spa-type t067 was detected in 18 of the 21 (85.7%) participating hospitals, including both MRSA and MSSA in six of them; in total, 25.9% of our isolates were spa-type t067 (49% in MRSA) in comparison with 0.6% in a central spa-typing database. The prevalence of the ant(4')-Ia and msrA/msrB genes was significantly higher in the MRSA spa-type t067 than in the other MRSA spa-types. Association between spa-type t067 and ST125 is described here for the first time. A high prevalence (36.4%) of PVL-positive MSSA was detected.A higher than expected prevalence of spa-type t067 isolates was found among invasive MRSA in Spain. The oxacillin, tobramycin, erythromycin and ciprofloxacin resistance profile of spa-type t067 isolates was linked to the presence of ant(4')-Ia and msrA or msrB genes.

83 Background: Immune response seems to improve outcome in women with Triple-Negative Breast Cancer (TNBC). Recent data suggests that the presence of Tumor-Infiltrating Lymphocytes is an independent factor associated with better prognosis. In this study, we evaluate the prognostic impact of both TIL´s and SIL´s in patients with TNBC. Methods: Data on women diagnosed with TNBC between 2005 and 2013, was collected by retrospectively reviewing at our institute. The rate of intratumoral and/or stromal lymphocytes was evaluated in all hematoxylin and eosin-stained histopathologic sections according to Denkert et al. The five-year disease-free survival (DFS) and overall survival (OS) were compared between groups with the presence or absence of TIL´s or SIL´s. Demographic and clinical characteristics were assessed, variables with a statistical significance between groups were analyzed in a multivariate analysis. Results: A total of 172 patients with TNBC treated at this institution were included with a mean age of 49.8 years. A complete absence of tumor lymphocytes was found in 88 patients while the presence of intratumoral, stromal or both was found in 84 (48.8%). A mean follow-up of 46.12 months showed significantly higher rates of both DFS and OS in women with SIL´s and TIL´s (p = 0.014 and 0.042 respectively) in locally advanced stages (LAS), regardless the rate of infiltrating lymphocytes found [Table 1]. SIL´s are correlated with a better prognosis compared with TIL´s (p = 0.028 and 0.091 respectively). Non-significant differences were found in early stages (p= 0.255). Conclusions: These results show that the presence of SIL´s or TIL´s are strongly associated with higher rates of DFS and OS in LAS, especially when SIL´s are found, suggesting that immunity seems to play a key role regarding the outcome in women with TNBC. Independent of the rate of lymphocytic infiltrate, its presence has a statistical significance. Because it is a feasible and inexpensive test that can be used as a prognostic predictor, we suggest assessing both SIL's and TIL's in histopathologic biopsies of patients with TNBC. [Table: see text]

Heterogeneity of breast cancer, specifically in triple negative (TN) tumors represents a clinical challenge which deserves further research in order to understand tumor biology and improve treatment options. MicroRNAs are small non-coding RNAs that regulates gene expression and play crucial roles in breast carcinogenesis. In order to analyze the heterogeneity and identify different subgroups of TN tumors, we analyzed the genome-wide microRNA expression patterns of 1105 mature miRNAs in 98 FFPE TN and 15 additional tumors with other immunophenotypes, using the Affymetrix Gene Expression miRNA microarray v2.0. Bioinformatic analysis with the non-supervised Consensus Clustering algorithm defined 4 groups, each of them showing association with clinically relevant parameters like the presence of metastases and survival. Several cellular pathways like cell proliferation, programmed cell death and motility are enriched in each of the microRNA expression defined groups. Based on miRNAs expression profiles between TN tumors and other inmunophenotypes (N=15 tumors), 3 miRNAs (miR-125b-2-3p, miR-342-3p -both of them down-regulated in TN tumors- and miR-660 -over-expressed-) were chosen for further functional studies in cellular models to better define their relationship with the biology and phenotype of triple negative tumors. After transfections in TN cell lines (MDA MB 231 and 468) a genomic evaluation of their transcriptional targets was made with the Human Gene St 1.0 array (Affymetrix), this data, together with in silico analysis, defined at least 25 mRNAs that are regulated by those miRNAs. Through this approach, we found that different patterns of mRNA targets emerge for each cellular model, reflecting their distinctive molecular and cellular features. The transcriptional targets identified, are involved in different tumorigenic pathways, such as activation of immune responses, regulation of apoptosis, cell motility and adhesion. Some of these results have been validated by other methodologies like measurement of Ki67 for proliferation, Annexin V for apoptosis and cell cycle status. Our data identified the altered expression of microRNAs that reveals the existence of 4 subgroups with different prognostic values among TN tumors. Furthermore, we performed the characterization of 3 miRNAs with aberrant expression in TN tumors that regulate cancer-related mRNAs and whose role in TN breast cancer has not been previously described. Together, these findings confirm the participation of miRNAs as regulators of cancer programs in triple negative tumors. Citation Format: Sandra L. Romero-Cordoba, Rosa Rebollar-Vega, Valeria Quintanar-Jurado, Alfredo Hidalgo-Miranda, Sergio Rodriguez-Cuevas, Veronica Bautista-Pina, Antonio Maffuz-Aziz. miRNA profiles identify different subgroups of triple negative tumors and reveal novel miRNA-mRNA interactions in breast cancer tumorigenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4370. doi:10.1158/1538-7445.AM2014-4370

Sir, Since the first report in 1988, vancomycin-resistant enterococci (VRE) have emerged as an important cause of hospital-acquired infections, particularly in the United States. However, infections with VRE are still relatively uncommon in many European hospitals. The aim of this study was to characterize isolates from a prolonged outbreak of vancomycin-resistant and teicoplaninsusceptible (VanB phenotype) Enterococcus faecium that occurred in a Spanish hospital. Between January 2004 and July 2006, 107 E. faecium isolates were isolated from clinical samples in the Hospital General de Soria (HGS), Spain (estimated catchment population of 92 700): 30 were isolated in 2004, 44 in 2005 and 33 in 2006. Of these, 34 (31.8%) isolates from separate patients showed VanB phenotypes, for which the annual distribution was 23.2% (7 isolates) in 2004, 29.5% (13 isolates) in 2005 and 42.4% (14 isolates) in 2006. The remaining 73 E. faecium isolates were susceptible to both vancomycin and teicoplanin. From March 2004 to October 2005, only sporadic cases appeared; however, in November 2005 three cases were detected, peaking in March and April 2006 with five and four cases, respectively. Most VanB isolates were from males (58.8%), age . 65 years (84.4%), admitted to ICU (32.4%) and were isolated from wounds (35.3%) or the urinary tract (23.6%). Twenty-eight patients (82.4%) had predisposing underlying conditions, principally respiratory/cardiac diseases (27.3%), neurological diseases (27.3%), tumoral pathologies (27.3%) and urinary tract pathologies (21.2%). Thirty-two patients were treated with antibiotics within the month prior to infection; the most frequent agents were cephalosporins (47.1%) and vancomycin (23.5%). Statistical comparisons of demographic data, clinical characteristics, microbiological features and clinical outcomes of the 34 patients infected by VanB E. faecium with 50 patients infected by vancomycin-susceptible E. faecium showed that only previous treatment with cephalosporins and/or vancomycin was associated with the emergence of VanB E. faecium (P , 0.05). All 34 VanB isolates were also resistant to ampicillin (MICs . 8 mg/L), erythromycin (MICs . 4 mg/L) and levofloxacin (MICs . 2 mg/L), but were susceptible to high concentrations of gentamicin (MICs 4–8 mg/L). Seventeen of these isolates were randomly selected and subjected to further molecular and epidemiological studies. A vanB gene was detected by PCR in all 17 isolates, and sequencing confirmed that this was identical to the vanB2 allele. In addition, the intergenic region vanSB–vanYB, sequenced in four isolates, showed previously reported point mutations and a 5 bp deletion in vanB2. Amplification and posterior sequencing of vanXB–ORFC amplicons demonstrated genetic linkage of the vanB2 operon to a Tn5382-like element. The pbp5 gene was detected by PCR in the four isolates, but we did not find genetic linkage of pbp5 and the vanB2-containing Tn5382. This latter finding is in accordance with the high rate of ampicillin resistance (82.6%; 60/73 isolates) among our vancomycin-susceptible E. faecium isolates. The 17 VanB isolates harboured the enterococcal surface protein gene, esp, but not the hyl, gelE, cylA or asa1 virulence genes. By PFGE, the VanB2 isolates exhibited a genetic relatedness of 92–100%, and 12 were indistinguishable (Figure 1) indicating an outbreak strain. Five VanB isolates from other hospitals and one vancomycin-susceptible strain from HGS were used as controls

Today, more than 55% of the world's breast cancer cases are diagnosed in low and middle-income countries and in 2020, more that 70% of the cases will come from the developing nations. In Mexico, breast cancer-specific mortality doubled during the past 20 years, representing the second-leading cause of death in women between 30 and 59 years and the leading cause of cancer related death in the female population. According to statistics, in Mexico a woman dies due to breast cancer every two hours. Even though breast cancer represents a major public health problem in the developing world, knowledge about the genetic and genomic structure of breast tumors in Mexican or Latin American populations is very limited. In the past four years, we have participated in the Slim Initiative of Genomic Medicine (SIGMA) Project, a collaboration between the Carlos Slim Institute of Health, the Broad Institute, and the National Institute of Genomic Medicine in Mexico city. The goal of the SIGMA project is to characterize the genomic basis of common diseases, including several types of cancer. This effort has focused on the application of whole genome and whole exome sequencing of human tumors. In the case of breast cancer, we have analyzed the whole genomes of 22 tumor/normal tissue pairs and the whole exomes of 103 tumor/normal tissues from Mexican and Vietnamese patients. Sequence analysis led to the novel identification of potential loss of function mutations of the CBFB transcription factor, and deletions of its partner RUNX1, an event which has never been previously reported in breast tumors or in any other epithelial tumor. Of clinical relevance, we also identified a somatic translocation involving MAGI3 and AKT3 in a triple negative breast tumor. Ectopic expression of the fusion transcrip leads to constitutive phosphorylation of downstream GSK and loss of contact inhibition. Most importantly, the activity of the fusion protein can be abrogated by an ATP-competitive small molecule inhibitor of AKT, potentially representing a new therapeutic avenue for these patients. In parallel with sequencing, we have also been working on the analysis of somatic DNA copy number aberrations, messenger RNA expression, and microRNA expression patterns in tumors from Mexican patients. Intrinsic breast cancer sub-typing in 125 tumors from Mexican patients showed that 13.6% of the tumors were basal-like, 16.8% were Her2-enriched, 24.8% Luminal A, 34.4% Luminal B and 10.4 normal-like. With microRNA expression, we have identified a group of microRNAs whose role in breast cancer has not been previously described and are currently analyzing differential microRNA expression across tumor sub-types, in particular triple negative tumors, where we have been able to identify at least three different tumor sub-groups based on microRNA expression patterns. Citation Format: Shantanu Banerji, Kristian Cibulskis, Claudia Rangel-Escareño, Kristin K. Brown, Scott L. Carter, Abbie M. Frederick, Michael S. Lawrence, Andrey Y. Sivachenko, Carrie Sougnez, Lihua Zou, Maria L. Cortes, Juan C. Fernandez-Lopez, Shouyong Peng, Kristin G. Ardlie, Daniel Auclair, Veronica Bautista-Piña, Fujiko Duke, Joshua Francis, Joonil Jung, Antonio Maffuz-Aziz, Robert C. Onofrio, Melissa Parkin, Nam H. Pho, Valeria Quintanar-Jurado, Alex H. Ramos, Rosa Rebollar-Vega, Sergio A. Rodríguez-Cuevas, Sandra L. Romero-Cordoba, Steven E. Schumacher, Nicolas Stransky, Kristin M. Thompson, Laura Uribe-Figueroa, Jose Baselga, Rameen Beroukhim, Kornelia Polyak, Dennis C. Sgroi, Andrea L. Richardson, Gerardo Jimenez-Sánchez, Eric S. Lander, Stacey B. Gabriel, Levi A. Garraway, Todd R. Golub, Jorge Meléndez-Zajgla, Alex Toker, Gad Getz, Matthew Meyerson, Alfredo Hidalgo-Miranda. Molecular profiling of breast cancer in Mexico: Identification of novel therapeutic targets through whole genome sequencing analysis. [abstract]. In: Proceedings of the Fifth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2012 Oct 27-30; San Diego, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2012;21(10 Suppl):Abstract nr PL07-01.

microRNA expression signatures can differentiate normal and breast cancer tissues and can define specific clinico-pathological phenotypes in breast tumors. In order to further evaluate the microRNA expression profile in breast cancer, we analyzed the expression of 667 microRNAs in 29 tumors and 21 adjacent normal tissues using TaqMan Low-density arrays. 130 miRNAs showed significant differential expression (adjusted P value = 0.05, Fold Change = 2) in breast tumors compared to the normal adjacent tissue. Importantly, the role of 43 of these microRNAs has not been previously reported in breast cancer, including several evolutionary conserved microRNA*, showing similar expression rates to that of their corresponding leading strand. The expression of 14 microRNAs was replicated in an independent set of 55 tumors. Bioinformatic analysis of mRNA targets of the altered miRNAs, identified oncogenes like ERBB2, YY1, several MAP kinases, and known tumor-suppressors like FOXA1 and SMAD4. Pathway analysis identified that some biological process which are important in breast carcinogenesis are affected by the altered microRNA expression, including signaling through MAP kinases and TP53 pathways, as well as biological processes like cell death and communication, focal adhesion and ERBB2-ERBB3 signaling. Our data identified the altered expression of several microRNAs whose aberrant expression might have an important impact on cancer-related cellular pathways and whose role in breast cancer has not been previously described.

Breast cancer can be classified according to the gene expression signatures of the tumors into four clinically relevant sub-types, luminal A, luminal B, Her2-like and basal-like. Additionally microRNA expression patterns can separate normal tissue from breast tumors, but there is currently limited information about the potential differences in microRNA expression profiles between the different tumor subtypes, defined by gene expression. In order to determine the microRNA expression profiles in breast tumors, and explore the differences in the expression patterns of these molecules between different cancer subtypes, we analyzed the expression of 664 microRNAs with the TaqMan low-density array platform in 40 breast tumors from different subtypes, and in 21 adjacent normal breast tissues. Tumor sub-typing was carried out with the PAM50 algorithm in expression data obtained with the Affymetrix Human Gene ST 1.0 array, obtaining 8 Luminal A tumors, 9 luminal B, 8 Her2-like and 3 Triple negative basal-like tumors. Given the low number of basal-like tumors, for the microRNA expression analysis, we included 12 additional triple-negative tumors defined by immunohistochemistry. Finally, we analyzed the expression of DICER and Ago2, involved in the biogenesis of microRNAs in the breast tumors. 131 microRNAs showed significant differential expression (adjusted P value=0.05, Fold Change=2) in breast tumors compared to the normal adjacent tissue. The role of 25% of these microRNAs has not been previously reported in breast cancer. 10 microRNAs showed differential expression between basal/triple negative tumors compared to hormone receptor and HER2 positive tumors. Transcriptional targets of these microRNAs include genes involved in the carcinogenesis of triple negative tumors, like PARP1, or in the microRNA biogenesis machinery, like DICER. Enrichment ontology analysis of the microRNAs differentially expressed in the TN tumors, detected pathways like p53 and focal adhesion whose role in cancer development, invasion and metastasis might be crucial; and MAPK, which has been related to recurrence of TN tumors. Regarding the expression of Ago2 and DICER, we detected down-regulation (approximately 20% less) of both proteins, mainly in TN tumors. Down-regulation in the triple negative tumors was further validated at the mRNA level by RT-qPCR assays. Our data identified the altered expression of several microRNAs whose aberrant expression might have an important impact on cancer-related cellular pathways and whose role in breast cancer has not been previously described. microRNAs expression is also capable to discriminate between different tumor subtypes and the expression of proteins involved in microRNA biogenesis might play a role in breast carcinogenesis, specifically in the triple negative subtype. Citation Format: Sandra L. Romero-Cordoba, Rosa G. Rebollar-Vega, Valeria Quintanar-Jurado, Veronica Bautista-Pina, Sergio Rodriguez-Cuevas, Antonio Maffuz-Aziz, Alfredo Hidalgo-Miranda. Differences in microRNA expression patterns in breast cancer subtypes [abstract]. In: Proceedings of the AACR Special Conference on Noncoding RNAs and Cancer; 2012 Jan 8-11; Miami Beach, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(2 Suppl):Abstract nr A25.

Background Aberrant expression of microRNAs has been associated with migration of tumor cells. In this study, we aimed to investigate the biological significance of miR-944 whose function is unknown in breast cancer. Methods MiR-944 expression in breast cancer cells and tumors was evaluated by Taqman qRT-PCR assays. Transcriptional profiling of MDA-MB-231 cells expressing miR-944 was performed using DNA microarrays. Cell viability, migration and invasion were assessed by MTT, scratch/wound-healing and transwell chamber assays, respectively. The luciferase reporter assay was used to evaluate targeting of SIAH1, PTP4A1 and PRKCA genes by miR-944. SIAH1 protein levels were measured by Western blot. Silencing of SIAH1 gene was performed by RNA interference using shRNAs. Results Our data showed that miR-944 expression was severely repressed in clinical specimens and breast cancer cell lines. Suppression of miR-944 levels was independent of hormonal status and metastatic potential of breast cancer cells. Gain-of-function analysis indicated that miR-944 altered the actin cytoskeleton dynamics and impaired cell migration and invasion. Genome-wide transcriptional profiling of MDA-MB-231 cells that ectopically express miR-944 showed that 15 genes involved in migration were significantly repressed. Notably, luciferase reporter assays confirmed the ability of miR-944 to bind the 3´UTR of SIAH1 and PTP4A1 genes, but not PRKCA gene. Congruently, an inverse correlation between miR-944 and SIAH1 protein expression was found in breast cancer cells. Moreover, SIAH1 was upregulated in 75 % of miR-944-deficient breast tumors. Finally, SIAH1 gene silencing by RNA interference significantly impaired cell migration of breast cancer cells. Conclusions Our results pointed out that miR-944 is a novel upstream negative regulator of SIAH1 and PTP4A1 genes and provided for the first time evidence for its functional role in migration and invasion of breast cancer cells. They also suggest that miR-944 restoration may represent a potential strategy for breast cancer therapy. Electronic supplementary material The online version of this article (doi:10.1186/s12885-016-2470-3) contains supplementary material, which is available to authorized users.