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Somatic mutation signatures may represent footprints of genetic and environmental exposures that cause different cancer. Few studies have comprehensively examined their association with germline variants, and none in an indigenous African population. SomaticSignatures was employed to extract mutation signatures based on whole‐genome or whole‐exome sequencing data from female patients with breast cancer (TCGA, training set, n = 1,011; Nigerian samples, validation set, n = 170), and to estimate contributions of signatures in each sample. Association between somatic signatures and common single nucleotide polymorphisms (SNPs) or rare deleterious variants were examined using linear regression. Nine stable signatures were inferred, and four signatures (APOBEC C>T, APOBEC C>G, aging and homologous recombination deficiency) were highly similar to known COSMIC signatures and explained the majority (60–85%) of signature contributions. There were significant heritable components associated with APOBEC C>T signature (h 2 = 0.575, p = 0.010) and the combined APOBEC signatures (h 2 = 0.432, p = 0.042). In TCGA dataset, seven common SNPs within or near GNB5 were significantly associated with an increased proportion (beta = 0.33, 95% CI = 0.21–0.45) of APOBEC signature contribution at genome‐wide significance, while rare germline mutations in MTCL1 was also significantly associated with a higher contribution of this signature (p = 6.1 × 10−6). This is the first study to identify associations between germline variants and mutational patterns in breast cancer across diverse populations and geography. The findings provide evidence to substantiate causal links between germline genetic risk variants and carcinogenesis., What's new? Women of African ancestry are more likely to be diagnosed with clinically aggressive breast cancer than women of European or Asian ancestry. Here, the authors examined associations between germline variants and mutational signatures in breast cancers across different ethnicities, especially in a unique sample set of indigenous African women. They identified four stable mutational signatures that explained the majority of tumor mutations, leading to a better understanding of the complex interplay between germline genetics and somatic mutations across different ethnicities.

Cromosomes; Marcadors pronòstics Cromosomas; Marcadores de pronóstico Chromosomes; Prognostic markers Oxidation of H3 at lysine 4 (H3K4ox) by lysyl oxidase-like 2 (LOXL2) generates an H3 modification with an unknown physiological function. We find that LOXL2 and H3K4ox are higher in triple-negative breast cancer (TNBC) cell lines and patient-derived xenografts (PDXs) than those from other breast cancer subtypes. ChIP-seq revealed that H3K4ox is located primarily in heterochromatin, where it is involved in chromatin compaction. Knocking down LOXL2 reduces H3K4ox levels and causes chromatin decompaction, resulting in a sustained activation of the DNA damage response (DDR) and increased susceptibility to anticancer agents. This critical role that LOXL2 and oxidized H3 play in chromatin compaction and DDR suggests that functionally targeting LOXL2 could be a way to sensitize TNBC cells to conventional therapy. This work was supported by grants from Instituto de Salud Carlos III (ISCIII) FIS/FEDER (PI12/01250; CP08/00223; PI16/00253; and CB16/12/00449), MINECO (SAF2013-48849-C2-1-R) to SP, BFU2015-68354 to THS, Breast Cancer Research Foundation (BCRF-17-008) to JA, AGL2014-52395-C2-2-R to DA, Worldwide Cancer Research, Red Temática de Investigación Cooperativa en Cáncer (RD012/0036/005), Fundación Científica de la Asociación Española contra el Cáncer, and Fundació La Marató TV3. THS was supported by institutional funding (MINECO) through the Centres of Excellence Severo Ochoa award and the CERCA Programme of the Catalan Government, and SS-B, by a Fundació La Caixa fellowship. We thank La Caixa Foundation and Cellex Foundation for provide research facilities and equipment. GV has received funding from the MINECO (a “Juan de la Cierva Incorporation” fellowship; IJCI-2014-20723). SP was a recipient of a Miguel Servet contract (ISCIII/FIS), and AI, JPC-C, LP-G, and GS-B are supported by contracts from Worldwide Cancer Research, Fundació La Marató TV3, Fundació FERO, and a FI Fellowship from the Generalitat de Catalunya, respectively.

The mechanisms of chemotherapy resistance in triple negative breast cancer remain unclear, and so, new molecules which might mediate this resistance could optimize treatment response. Here we analyzed the involvement of the miRNA-449 family in the response to doxorubicin. The cell viability, cell-cycle phases, and the expression of in silico target genes and proteins of sensitive/resistant triple negative breast cancer cell lines were evaluated in response to doxorubicin treatment and after gain/loss of miRNAs-449 function achieved by transient transfection. Triple negative breast cancer patients were selected for ex vivo experiments and to evaluate gene and miRNAs expression changes after treatment, as well as survival analysis by Kaplan-Meier. Doxorubicin treatment upregulated miRNAs-449 and DNA-damage responder factors E2F1 and E2F3 in triple negative breast cancer sensitive breast cancer cells, while expression remained unaltered in resistant ones. In vitro overexpression of miRNAs-449 sensitized cells to the treatment and significantly reduced the resistance to doxorubicin. These changes showed also a strong effect on cell cycle regulation. Finally, elevated levels of miRNA-449a associated significantly with better survival in chemotherapy-treated triple negative breast cancer patients. These results reveal for the first time the involvement of the miRNA-449 family in doxorubicin resistance and their predictive and prognostic value in triple negative breast cancer patients.

Phototype has been associated with an increased risk of prostate cancer, and it is yet unknown if it is related to other hormone-dependent cancers, such as breast cancer or whether this association could be considered causal. Methods We examined the association between the phototype and breast and prostate cancers using a Mendelian randomization analysis. We studied 1,738 incident cases of breast cancer and another 817 cases of prostate cancer. To perform a Mendelian randomization analysis on the phototype—cancer relationship, a genetic pigmentation score was required that met the following criteria: (1) the genetic pigmentation score was associated with phototype in controls; (2) the genetic pigmentation score was not associated with confounders in the relationship between phototype and cancer, and (3) the genetic pigmentation score was associated with cancer only through its association with phototype. Once this genetic score is available, the association between genetic pigmentation score and cancer can be identified as the association between phototype and cancer. Results The association between the genetic pigmentation score and phototype in controls showed that a higher genetic pigmentation score was associated with fair skin, blond hair, blue eyes and the presence of freckles. Applying the Mendelian randomization analysis, we verified that there was no association between the genetic pigmentation score and cancers of the breast and prostate. Conclusions Phototype is not associated with breast or prostate cancer., This work was partially funded by the ªAccion Transversal del Cáncer", approved by the Spanish Ministry Council on the 11th October 2007; The Instituto de Salud Carlos III-FEDER [PI08/1770, PI08/0533, PI08/1359, PI09/00773-Cantabria, PI09/01286-León, PI09/01903-Valencia, PI09/02078-Huelva, PI09/01662-Granada, PI11/ 01403, PI11/01889-FEDER, PI11/00226, PI11/ 01810, PI11/02213, PI12/00488, PI12/00265, PI12/01270, PI12/00715, PI12/00150, PI14/01219, PI14/0613, PI15/00069, PI15/00914, PI15/01032]; The Fundación Marqués de Valdecilla [API 10/09]; The ICGC International Cancer Genome Consortium CLL (The ICGC CLL-Genome Project is funded by Spanish Ministerio de Economía y Competitividad (MINECO) through the Instituto de Salud Carlos III (ISCIII) and Red Temática de Investigación del Cáncer (RTICC) del ISCIII (RD12/ 0036/0036)); The Junta de Castilla y León [LE22A10-2]; The Consejería de Salud of the Junta de Andalucía [2009-S0143]; The Conselleria de Sanitat of the Generalitat Valenciana [AP_061/10]; The Recercaixa [2010ACUP 00310]; The Regional Government of the Basque Country; The Consejería de Sanidad de la Región de Murcia; The European Commission [grants FOOD-CT-2006-036224- HIWATE]; The Spanish Association Against Cancer (AECC) Scientific Foundation; The Catalan Government DURSI [grant 2014SGR647]; The Fundación Caja de Ahorros de Asturias; and the University of Oviedo.

Biological changes that occur during metastatic progression of breast cancer are still incompletely characterized. In this study, we compared intrinsic molecular subtypes and gene expression in 123 paired primary and metastatic tissues from breast cancer patients. Intrinsic subtype was identified using a PAM50 classifier and χ2 tests determined the differences in variable distribution. The rate of subtype conversion was 0% in basal-like tumors, 23.1% in HER2-enriched (HER2-E) tumors, 30.0% in luminal B tumors, and 55.3% in luminal A tumors. In 40.2% of cases, luminal A tumors converted to luminal B tumors, whereas in 14.3% of cases luminal A and B tumors converted to HER2-E tumors. We identified 47 genes that were expressed differentially in metastatic versus primary disease. Metastatic tumors were enriched for proliferation-related and migration-related genes and diminished for luminal-related genes. Expression of proliferation-related genes were better at predicting overall survival in metastatic disease (OSmet) when analyzed in metastatic tissue rather than primary tissue. In contrast, a basal-like gene expression signature was better at predicting OSmet in primary disease compared with metastatic tissue. We observed correlations between time to tumor relapse and the magnitude of changes of proliferation, luminal B, or HER2-E signatures in metastatic versus primary disease. Although the intrinsic subtype was largely maintained during metastatic progression, luminal/HER2-negative tumors acquired a luminal B or HER2-E profile during metastatic progression, likely reflecting tumor evolution or acquisition of estrogen independence. Overall, our analysis revealed the value of stratifying gene expression by both cancer subtype and tissue type, providing clinicians more refined tools to evaluate prognosis and treatment. This work was supported by funds from Instituto de Salud Carlos III - PI13/01718 (A.P.), by a Career Catalyst Grant from the Susan Komen Foundation (A.P.), by Banco Bilbao Vizcaya Argentaria (BBVA) Foundation (A.P.) and by the Breast Cancer Research Foundation. This work was also supported by funds from FEDER (RETICC): RD12/0036/0076 (JA), RD12/0036/0051 (JA), RD12/0036/0070 (AL) and RD12/0036/0076 (MM). JMC holds a fellowship from “PhD4MD”, a Collaborative Research Training Programme for Medical Doctors (IDIBAPS, August Pi i Sunyer Institute for Biomedical Research and IRB Barcelona, Institute for Research in Biomedicine) partially funded by Instituto de Salud Carlos III, (ISCIII, project: II14/00019). JMC and RRG research support is provided by the Spanish Ministry of Science and Innovation grant SAF2013-46196 (FEDER Funds) and the Generalitat de Catalunya AGAUR 2014-SGR grant 535.

A subset of HER2+ breast cancer patients manifest clinical resistance to trastuzumab. Recently, miR-26a and miR-30b have been identified as trastuzumab response regulators, and their target gene CCNE2 seems to play an important role in resistance to trastuzumab therapy. Cell viability was evaluated in trastuzumab treated HER2+ BT474 wt (sensitive), BT474r (acquired resistance), HCC1954 (innate resistance), and MDA-MB-231 (HER2−) cell lines, and the expression of miR-26a, miR-30b, and their target genes was measured. BT474 wt cell viability decreased by 60% and miR-26a and miR-30b were significantly overexpressed (~3-fold, p = 0.003 and p = 0.002, respectively) after trastuzumab treatment, but no differences were observed in resistant and control cell lines. Overexpression of miR-30b sensitized BT474r cells to trastuzumab (p = 0.01) and CCNE2, was significantly overexpressed after trastuzumab treatment in BT474r cells (p = 0.032), but no significant changes were observed in sensitive cell line. When CCNE2 was silenced BT474r cell sensitivity to trastuzumab increased (p = 0.03). Thus, the molecular mechanism of trastuzumab action in BT474 cell line may be regulated by miR-26a and miR-30b and CCNE2 overexpression might play an important role in acquired trastuzumab resistance in HER2+ breast cancer given that resistance was diminished when CCNE2 was silenced.

Background: Our aim was to assess the cumulative risk of false-positive screening results, screen-detected cancer, and interval breast cancer in mammography screening among women with and without a previous benign breast disease and a family history of breast cancer. Methods: The cohort included 42 928 women first screened at the age of 50-51 years at three areas of the Spanish Screening Programme (Girona, and two areas in Barcelona) between 1996 and 2011, and followed up until December 2012. We used discrete-time survival models to estimate the cumulative risk of each screening outcome over 10 biennial screening exams. Results: The cumulative risk of false-positive results, screen-detected breast cancer, and interval cancer was 36.6, 5.3, and 1.4 for women with a previous benign breast disease, 24.1, 6.8, and 1.6% for women with a family history of breast cancer, 37.9, 9.0, and 3.2%; for women with both a previous benign breast disease and a family history, and 23.1, 3.2, and 0.9% for women without either of these antecedents, respectively. Conclusions: Women with a benign breast disease or a family history of breast cancer had an increased cumulative risk of favourable and unfavourable screening outcomes than women without these characteristics. A family history of breast cancer did not increase the cumulative risk of false-positive results. Identifying different risk profiles among screening participants provides useful information to stratify women according to their individualised risk when personalised screening strategies are discussed.

// Elisabet Cuyas 1, 2, * , Salvador Fernandez-Arroyo 3, * , Tomas Alarcon 4, 5, 6, 7 , Ruth Lupu 8, 9 , Jorge Joven 3 , Javier A. Menendez 1, 2 1 ProCURE (Program Against Cancer Therapeutic Resistance), Metabolism and Cancer Group, Catalan Institute of Oncology, Girona, Catalonia, Spain 2 Molecular Oncology Group, Girona Biomedical Research Institute (IDIBGI), Girona, Catalonia, Spain 3 Unitat de Recerca Biomedica, Hospital Universitari de Sant Joan, IISPV, Universitat Rovira i Virgili, Campus of International Excellence Southern Catalonia, Reus, Spain 4 Institucio Catalana d'Estudis i Recerca Avancats (ICREA), Barcelona, Spain 5 Computational and Mathematical Biology Research Group, Centre de Recerca Matematic (CRM), Barcelona, Spain 6 Departament de Matematiques, Universitat Autonoma de Barcelona, Barcelona, Spain 7 Barcelona Graduate School of Mathematics (BGSMath), Barcelona, Spain 8 Mayo Clinic, Department of Laboratory Medicine and Pathology, Division of Experimental Pathology, Rochester, MN, USA 9 Mayo Clinic Cancer Center, Rochester, MN, USA * These authors contributed equally to this work Correspondence to: Javier A. Menendez, email: jmenendez@iconcologia.net , jmenendez@idibgi.org Keywords: BRCA1, hereditary breast cancer, breast cancer susceptibility, bioenergetics, glutamine Received: January 22, 2016 Accepted: May 12, 2016 Published: May 31, 2016 ABSTRACT We hypothesized that women inheriting one germline mutation of the BRCA1 gene (“one-hit”) undergo cell-type-specific metabolic reprogramming that supports the high biosynthetic requirements of breast epithelial cells to progress to a fully malignant phenotype. Targeted metabolomic analysis was performed in isogenic pairs of nontumorigenic human breast epithelial cells in which the knock-in of 185delAG mutation in a single BRCA1 allele leads to genomic instability. Mutant BRCA1 one-hit epithelial cells displayed constitutively enhanced activation of biosynthetic nodes within mitochondria. This metabolic rewiring involved the increased incorporation of glutamine- and glucose-dependent carbon into tricarboxylic acid (TCA) cycle metabolite pools to ultimately generate elevated levels of acetyl-CoA and malonyl-CoA, the major building blocks for lipid biosynthesis. The significant increase of branched-chain amino acids (BCAAs) including the anabolic trigger leucine, which can not only promote protein translation via mTOR but also feed into the TCA cycle via succinyl-CoA, further underscored the anabolic reprogramming of BRCA1 haploinsufficient cells. The anti-diabetic biguanide metformin “reversed” the metabolomic signature and anabolic phenotype of BRCA1 one-hit cells by shutting down mitochondria-driven generation of precursors for lipogenic pathways and reducing the BCAA pool for protein synthesis and TCA fueling. Metformin-induced restriction of mitochondrial biosynthetic capacity was sufficient to impair the tumor-initiating capacity of BRCA1 one-hit cells in mammosphere assays. Metabolic rewiring of the breast epithelium towards increased anabolism might constitute an unanticipated and inherited form of metabolic reprogramming linked to increased risk of oncogenesis in women bearing pathogenic germline BRCA1 mutations. The ability of metformin to constrain the production of mitochondrial-dependent biosynthetic intermediates might open a new avenue for “starvation” chemopreventive strategies in BRCA1 carriers.

BACKGROUND: There are currently no biomarkers for early breast cancer patient populations at risk of bone metastasis. Identification of mediators of bone metastasis could be of clinical interest. METHODS: A de novo unbiased screening approach based on selection of highly bone metastatic breast cancer cells in vivo was used to determine copy number aberrations (CNAs) associated with bone metastasis. The CNAs associated with bone metastasis were examined in independent primary breast cancer datasets with annotated clinical follow-up. The MAF gene encoded within the CNA associated with bone metastasis was subjected to gain and loss of function validation in breast cancer cells (MCF7, T47D, ZR-75, and 4T1), its downstream mechanism validated, and tested in clinical samples. A multivariable Cox cause-specific hazard model with competing events (death) was used to test the association between 16q23 or MAF and bone metastasis. All statistical tests were two-sided. RESULTS: 16q23 gain CNA encoding the transcription factor MAF mediates breast cancer bone metastasis through the control of PTHrP. 16q23 gain (hazard ratio (HR) for bone metastasis = 14.5, 95% confidence interval (CI) = 6.4 to 32.9, P < .001) as well as MAF overexpression (HR for bone metastasis = 2.5, 95% CI = 1.7 to 3.8, P < .001) in primary breast tumors were specifically associated with risk of metastasis to bone but not to other organs. CONCLUSIONS: These results suggest that MAF is a mediator of breast cancer bone metastasis. 16q23 gain or MAF protein overexpression in tumors may help to select patients at risk of bone relapse. MP and SG is supported by “La Caixa” PhD fellowship program. AAE and AB hold PhD fellowships from the Spanish Ministerio de Ciencia e Innovación (MICINN). MTS is supported by the IRB Barcelona PhD program. JU is a “Juan de la Cierva” Researcher (MICINN). FIS PI12/00680, PI12/01552, and PI12/01421 supported JA, ALl, and FR, respectively. JA, ALl, and AP were part of RD12/0036/0051, RD12/0036/0070, and RD12/0036/0042, and FR is part of biobanc RD/09/0076/00101. JA and FR are recipients of intensification grant ISCIII, 2009SGR321, XBTC, MARBiobanc, and Cellex. RRG was supported by the Institució Catalana de Recerca i Estudis Avançats. Support and structural funds were provided by the BBVA Foundation, the Generalitat de Catalunya (2014 SGR 535), and the Spanish Ministerio de Ciencia e Innovación (MICINN) (SAF2013-46196) to RRG.

Seven linker histone H1 variants are present in human somatic cells with distinct prevalence across cell types. Despite being key structural components of chromatin, it is not known whether the different variants have specific roles in the regulation of nuclear processes or are differentially distributed throughout the genome. Using variant-specific antibodies to H1 and hemagglutinin (HA)-tagged recombinant H1 variants expressed in breast cancer cells, we have investigated the distribution of six H1 variants in promoters and genome-wide. H1 is depleted at promoters depending on its transcriptional status and differs between variants. Notably, H1.2 is less abundant than other variants at the transcription start sites of inactive genes, and promoters enriched in H1.2 are different from those enriched in other variants and tend to be repressed. Additionally, H1.2 is enriched at chromosomal domains characterized by low guanine-cytosine (GC) content and is associated with lamina-associated domains. Meanwhile, other variants are associated with higher GC content, CpG islands and gene-rich domains. For instance, H1.0 and H1X are enriched at gene-rich chromosomes, whereas H1.2 is depleted. In short, histone H1 is not uniformly distributed along the genome and there are differences between variants, H1.2 being the one showing the most specific pattern and strongest correlation with low gene expression. © 2014 The Author(s). Published by Oxford University Press., Ministerio de Ciencia e Innovación of Spain (MICINN) and FEDER [BFU2011-23057 to A.J.]; Ministerio de Economía y Competitividad [SAF2012-36199 to N.L.-B.]; Generalitat de Catalunya [2009-SGR-1222 to A.J.]; JAE-Doc contract from CSIC-MICINN (to J.-M.T.); TA contract from CSIC-MICINN (to R.M.); FPU predoctoral fellowship from MICINN (to L.M.-A.). Funding for open access charge: Ministerio de Ciencia e Innovación of Spain (MICINN) and FEDER [BFU2011-23057 to A.J.]

Blocking specific protein interactions can lead to human diseases. Accordingly, protein interactions and the structural knowledge on interacting surfaces of proteins (interfaces) have an important role in predicting the genotype-phenotype relationship. We have built the phenotype specific sub-networks of protein-protein interactions (PPIs) involving the relevant genes responsible for lung and brain metastasis from primary tumor in breast cancer. First, we selected the PPIs most relevant to metastasis causing genes (seed genes), by using the ""guilt-by-association"" principle. Then, we modeled structures of the interactions whose complex forms are not available in Protein Databank (PDB). Finally, we mapped mutations to interface structures (real and modeled), in order to spot the interactions that might be manipulated by these mutations. Functional analyses performed on these sub-networks revealed the potential relationship between immune system-infectious diseases and lung metastasis progression, but this connection was not observed significantly in the brain metastasis. Besides, structural analyses showed that some PPI interfaces in both metastasis sub-networks are originating from microbial proteins, which in turn were mostly related with cell adhesion. Cell adhesion is a key mechanism in metastasis, therefore these PPIs may be involved in similar molecular pathways that are shared by infectious disease and metastasis. Finally, by mapping the mutations and amino acid variations on the interface regions of the proteins in the metastasis sub-networks we found evidence for some mutations to be involved in the mechanisms differentiating the type of the metastasis., Spanish Government (MINECO); Scientific and Technological Research Council of Turkey (TÜBİTAK)

Genetic analysis identifies the HMMR gene as a modifier of the breast cancer risk associated with BRCA1 gene mutation, while cell biological analysis of the protein product suggests a function in regulating development of the mammary gland., Differentiated mammary epithelium shows apicobasal polarity, and loss of tissue organization is an early hallmark of breast carcinogenesis. In BRCA1 mutation carriers, accumulation of stem and progenitor cells in normal breast tissue and increased risk of developing tumors of basal-like type suggest that BRCA1 regulates stem/progenitor cell proliferation and differentiation. However, the function of BRCA1 in this process and its link to carcinogenesis remain unknown. Here we depict a molecular mechanism involving BRCA1 and RHAMM that regulates apicobasal polarity and, when perturbed, may increase risk of breast cancer. Starting from complementary genetic analyses across families and populations, we identified common genetic variation at the low-penetrance susceptibility HMMR locus (encoding for RHAMM) that modifies breast cancer risk among BRCA1, but probably not BRCA2, mutation carriers: n = 7,584, weighted hazard ratio (wHR) = 1.09 (95% CI 1.02–1.16), ptrend = 0.017; and n = 3,965, wHR = 1.04 (95% CI 0.94–1.16), ptrend = 0.43; respectively. Subsequently, studies of MCF10A apicobasal polarization revealed a central role for BRCA1 and RHAMM, together with AURKA and TPX2, in essential reorganization of microtubules. Mechanistically, reorganization is facilitated by BRCA1 and impaired by AURKA, which is regulated by negative feedback involving RHAMM and TPX2. Taken together, our data provide fundamental insight into apicobasal polarization through BRCA1 function, which may explain the expanded cell subsets and characteristic tumor type accompanying BRCA1 mutation, while also linking this process to sporadic breast cancer through perturbation of HMMR/RHAMM., Author Summary Mutations in two genes that were initially identified as predisposing carriers to early-onset breast cancer, BRCA1 and BRCA2, cause similar perturbations in cellular responses to DNA damage but predispose carriers to distinct tumor types. Thus, the two genes may trigger different carcinogenic processes. We have used genetic analyses of affected families to uncover additional genetic variation that is linked to the risk of developing cancer for carriers of BRCA1 mutations. This variation falls within a centrosomal gene, named HMMR. The protein product of HMMR, which is called RHAMM, works in concert with BRCA1 to regulate the structure of normal breast cells as they grow and become polarized. This polarization process depends upon a balance between the activities of BRCA1 and the Aurora kinase A, with the kinase opposing BRCA1 function and promoting growth. Our findings provide new insights into the mechanism through which BRCA1 may promote commitment of initially bipotent mammary cells towards the luminal lineage, and how loss of this function may predispose cells to become breast tumors of a basal-like type.

In their report, Gotte and coworkers [1] analyzed the expression of c-Met in 200 patients with ductal carcinoma in situ. They concluded that c-Met could be related to angiogenic and lymphangiogenic factors in ductal carcinoma in situ. On the other hand, Greenberg and coworkers [2] studied 31 patients with ductal infiltrating carcinoma (DIC) to detect c-Met expression in their axillary fluids. They observed a correlation of c-Met expression with increasing tumor size and grade, capillary and lymphatic invasion and lymph node metastasis. We applied the fluorescent in situ hybridization (FISH) technique using the LSI D7S486/CEP7 commercial probe (Abbott Molecular Inc., Des Plaines, IL, USA), which includes the MET gene, and immunohistochemistry using c-Met monoclonal antibody clone 3D4 (Invitrogen, Carlsbad, CA, USA) to 184 archival invasive breast tumors (93 DIC and 91 lobular carcinomas). We constructed ten tissue microarrays with three replicates per sample. Pearson's chi-squared and Fisher's exact test were used to analyze the results. None of the 155 breast tumors analyzed by FISH presented amplification of MET and 35 cases (22%) had a low grade of polysomy (three to five copies) of chromosome 7. Polysomy was more frequently observed in DIC (25%; P = 0.001). We tried to correlate polysomy of MET in the DIC group with grade, tumor size, lymph node status, clinical stage and expression of HER2, P53, estrogen receptor (ER) and progesterone receptor (PR). We observed that the absence of expression of PR was the unique statistically significant variable (P = 0.001). Moreover, the ER+/PR- samples presented the highest rate of polysomy (38%) compared to ER+/PR+ tumors (15%) (Table ​(Table11). Table 1 Results of IHC of c-Met and FISH of LSI D7S486/CEP7 applied to lobular and ductal carcinomas Out of 168 tumors analyzed by immunohistochemistry, 65 (38.7%) presented expression of c-Met. When histological types were compared, the DIC group also showed the highest number of c-Met-positive samples (48%; P = 0.001). From the analysis with the clinico-pathological variables, the negativity for PR was again statistically significant (P = 0.001). The ER+/PR- tumors presented more frequent expression of c-Met (68%) compared to ER+/PR+ tumors (32%) and were correlated with polysomy (P = 0.020) (Table ​(Table22). Table 2 IHC and FISH results of MET according to the status of PR receptor in DIC carcinomas We can conclude that amplification of MET in breast cancer is not a common event, as opposed to other cancer subtypes (renal, gastric and lung carcinomas). Although found in breast tumors, it seems that overexpression of c-Met is not mainly due to increassed gene copy number of MET/polysomy7. However, polysomy in the ER+/PR- group could be an important mechanism – although not the only one – responsible for the differential expression observed in this type of DIC. This c-Met overexpression and the presence of polysomy 7 could be important events to be considered with regard to the known poor response to endocrine therapies of ER+/PR- breast tumors. Lack of PR expression in ER+ tumors may be a surrogate marker of aberrant growth factor signaling [3] that could be associated with their more aggressive outcome, as has already been described [4]. Our study suggests that it would be interesting to investigate new therapeutic options for ER+/PR- DIC, which may include c-Met inhibitors.

14-3-3σ is frequently lost in human breast cancers by genetic deletion or promoter methylation. We have now investigated the involvement of 14-3-3σ in the termination of NF-κB signal in mammary cells and its putative role in cancer relapse and metastasis. Our results show that 14-3-3σ regulates nuclear export of p65-NF-κB following chronic TNFα stimulation. Restoration of 14-3-3σ in breast cancer cells reduces migration capacity and metastatic abilities in vivo. By microarray analysis, we have identified a genetic signature that responds to TNFα in a 14-3-3σ-dependent manner and significantly associates with different breast and other types of cancer. By interrogating public databases, we have found that over-expression of this signature correlates with poor relapse-free survival in breast cancer patients. Finally, screening of 96 human breast tumors showed that NF-κB activation strictly correlates with the absence of 14-3-3σ and it is significantly associated with worse prognosis in the multivariate analysis. Our findings identify a genetic signature that is important for breast cancer prognosis and for future personalized treatments based on NF-κB targeting. This work was funded by ISCIII/FEDER-Subdirección General de Evaluación y Fomento de la Investigación (PI07/0778, PI10/01128, PS09/1296 and PS09/01285), AGAUR (2009SGR23 and 2009SGR321) and Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica (I+D+I), iniciativa Ingenio 2010, Programa Consolider and Instituto de Salud Carlos III (ISCIII)/FEDER (RD06/0020/0098 and RD06/0020/0109). Financial support was provided to RRG by BBVA foundation and Asociación Española Contra el Cáncer. NLB acknowledges funding from the Spanish Ministry of Science and Technology (SAF2009-06954). AJ-S was funded by Science and Education Spanish Ministry (MEC) FPI predoctoral fellowship (BES-2008-001850). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript