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The HER2 oncogene is overexpressed or amplified in 20% of breast cancers. HER2-positive cancer historically portends a poor prognosis, but the HER2-targeted therapy trastuzumab mitigates this otherwise ominous distinction. Nevertheless, some patients suffer disease recurrence despite trastuzumab, and metastatic disease remains largely incurable due to innate and acquired resistance. Thus, understanding trastuzumab resistance remains an unmet medical need. Through RNA interference screening, we discovered that knockdown of the serine/threonine phosphatase PPM1H confers trastuzumab resistance via reduction in protein levels of the tumor suppressor p27. PPM1H dephosphorylates p27 at threonine 187, thus removing a signal for proteasomal degradation. We further determined that patients whose tumors express low levels of PPM1H trend towards worse clinical outcome on trastuzumab. Identifying PPM1H as a novel p27 phosphatase reveals new insight into how cancer cells destabilize a well-recognized tumor suppressor. Furthermore, low PPM1H expression may identify a subset of HER2-positive tumors that are harder to treat., (©2011 AACR.)

A long-sought milestone in microfluidics research has been the development of integrated technology for scalable analysis of transcription in single cells. Here we present a fully integrated microfluidic device capable of performing high-precision RT-qPCR measurements of gene expression from hundreds of single cells per run. Our device executes all steps of single-cell processing, including cell capture, cell lysis, reverse transcription, and quantitative PCR. In addition to higher throughput and reduced cost, we show that nanoliter volume processing reduced measurement noise, increased sensitivity, and provided single nucleotide specificity. We apply this technology to 3,300 single-cell measurements of (i) miRNA expression in K562 cells, (ii) coregulation of a miRNA and one of its target transcripts during differentiation in embryonic stem cells, and (iii) single nucleotide variant detection in primary lobular breast cancer cells. The core functionality established here provides the foundation from which a variety of on-chip single-cell transcription analyses will be developed.

We present a method to perform absolute quantification of glycogen in human embryonic stem cells (hESCs) in situ based on the use of Raman microspectroscopy. The proposed quantification method was validated by comparison to a commonly used commercial glycogen assay kit. With Raman microspectroscopy, we could obtain the glycogen content of hESCs faster and apparently more accurately than with the kit. In addition, glycogen distributions across a colony could be obtained. Raman spectroscopy can provide reliable estimates of the in situ glycogen content in hESCs, and this approach should also be extensible to their other biochemical constituents as well as to other cell types.

Objective: The molecular mechanisms that maintain human pluripotent stem (PS) cells are not completely understood. Here we sought to identify new candidate PS cell regulators to facilitate future improvements in their generation, expansion, and differentiation., Materials and Methods: We used bioinformatic analyses of multiple serial-analysis-of-gene-expression libraries (generated from human PS cells and their differentiated derivatives), together with small interfering RNA (siRNA) screening to identify candidate pluripotency regulators. Validation of candidate regulators involved promoter analyses, Affymetrix profiling, real-time PCR, and immunoprecipitation., Results: Promoter analysis of genes differentially expressed across multiple serial-analysis-of-gene-expression libraries identified E2F motifs in the promoters of many PS cell-specific genes (e.g., POU5F1, NANOG, SOX2, FOXD3). siRNA analyses identified two retinoblastoma binding proteins (RBBP4, RBBP9) as required for maintenance of multiple human PS cell types. Both RBBPs were bound to RB in human PS cells, and E2F motifs were present in the promoters of genes whose expression was altered by decreasing RBBP4 and RBBP9 expression. Affymetrix and real-time PCR studies of siRNA-treated human PS cells showed that reduced RBBP4 or RBBP9 expression concomitantly decreased expression of POU5F1, NANOG, SOX2, and/or FOXD3 plus certain cell cycle genes (e.g., CCNA2, CCNB1), while increasing expression of genes involved in organogenesis (particularly neurogenesis)., Conclusions: These results reveal new candidate positive regulators of human PS cells, providing evidence of their ability to regulate expression of pluripotency, cell cycle, and differentiation genes in human PS cells. These data provide valuable new leads for further elucidating mechanisms of human pluripotency., (Copyright © 2011 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.)

Background: Humans and mice with loss of function mutations in GPR54 (KISS1R) or kisspeptin do not progress through puberty, caused by a failure to release GnRH. The transcriptional networks regulated by these proteins in the hypothalamus have yet to be explored by genome-wide methods., Results: We show here, using 1 million exon mouse arrays (Exon 1.0 Affymetrix) and quantitative polymerase chain reaction (QPCR) validation to analyse microdissected hypothalamic tissue from Gpr54 and Kiss1 knockout mice, the extent of transcriptional regulation in the hypothalamus. The sensitivity to detect important transcript differences in microdissected RNA was confirmed by the observation of counter-regulation of Kiss1 expression in Gpr54 knockouts and confirmed by immunohistochemistry (IHC). Since Gpr54 and Kiss1 knockout animals are effectively pre-pubertal with low testosterone (T) levels, we also determined which of the validated transcripts were T-responsive and which varied according to genotype alone. We observed four types of transcriptional regulation (i) genotype only dependent regulation, (ii) T only dependent regulation, (iii) genotype and T-dependent regulation with interaction between these variables, (iv) genotype and T-dependent regulation with no interaction between these variables. The results implicate for the first time several transcription factors (e.g. Npas4, Esr2), proteases (Klk1b22), and the orphan 10-transmembrane transporter TMEM144 in the biology of GPR54/kisspeptin function in the hypothalamus. We show for the neuronal activity regulated transcription factor NPAS4, that distinct protein over-expression is seen in the hypothalamus and hippocampus in Gpr54 knockout mice. This links for the first time the hypothalamic-gonadal axis with this important regulator of inhibitory synapse formation. Similarly we confirm TMEM144 up-regulation in the hypothalamus by RNA in situ hybridization and western blot., Conclusions: Taken together, global transcriptional profiling shows that loss of GPR54 and kisspeptin are not fully equivalent in the mouse hypothalamus.

The telomeric amplicon at 8p12 is common in oestrogen receptor-positive (ER+) breast cancers. Array-CGH and expression analyses of 1172 primary breast tumours revealed that ZNF703 was the single gene within the minimal amplicon and was amplified predominantly in the Luminal B subtype. Amplification was shown to correlate with increased gene and protein expression and was associated with a distinct expression signature and poor clinical outcome. ZNF703 transformed NIH 3T3 fibroblasts, behaving as a classical oncogene, and regulated proliferation in human luminal breast cancer cell lines and immortalized human mammary epithelial cells. Manipulation of ZNF703 expression in the luminal MCF7 cell line modified the effects of TGFβ on proliferation. Overexpression of ZNF703 in normal human breast epithelial cells enhanced the frequency of in vitro colony-forming cells from luminal progenitors. Taken together, these data strongly point to ZNF703 as a novel oncogene in Luminal B breast cancer., (Copyright © 2011 EMBO Molecular Medicine.)

Next-generation sequencing of follicular lymphoma and diffuse-large B-cell lymphoma has revealed frequent somatic, heterozygous Y641 mutations in the histone methyltransferase EZH2. Heterozygosity and the presence of equal quantities of both mutant and wild-type mRNA and expressed protein suggest a dominant mode of action. Surprisingly, B-cell lymphoma cell lines and lymphoma samples harboring heterozygous EZH2(Y641) mutations have increased levels of histone H3 Lys-27-specific trimethylation (H3K27me3). Expression of EZH2(Y641F/N) mutants in cells with EZH2(WT) resulted in an increase of H3K27me3 levels in vivo. Structural modeling of EZH2(Y641) mutants suggests a "Tyr/Phe switch" model whereby structurally neutral, nontyrosine residues at position 641 would decrease affinity for unmethylated and monomethylated H3K27 substrates and potentially favor trimethylation. We demonstrate, using in vitro enzyme assays of reconstituted PRC2 complexes, that Y641 mutations result in a decrease in monomethylation and an increase in trimethylation activity of the enzyme relative to the wild-type enzyme. This represents the first example of a disease-associated gain-of-function mutation in a histone methyltransferase, whereby somatic EZH2 Y641 mutations in lymphoma act dominantly to increase, rather than decrease, histone methylation. The dominant mode of action suggests that allele-specific EZH2 inhibitors should be a future therapeutic strategy for this disease.

Background: Mll5 is currently a member of the Mll family of SET domain histone methyltransferase proteins but studies have also showed that it could be part of the SET3 branch of proteins. Recently, constitutive knock out animal studies have shown that Mll5 is required for proper haematopoietic stem cell differentiation, and loss of Mll5 results in synthetic lethality for genome de-methylation. Mll5 deficient male mice are infertile and here we analyse the consequences of Mll5 deficiency for spermatogenesis., Methodology/principal Findings: Mll5 deficient male mice, but not female mice, are infertile. Here we show using RNA in-situ hybridization that Mll5 is expressed in the germ cells of the testes of wild type mice. Consistent with the expression of Mll5, we demonstrate by electron microscopy, video microscopy and in vitro fertilisation techniques that Mll5 deficient mice have defects in terminal maturation and packaging of sperm. The defects seen include detachment of the acrosomal cap and impaired excess cytoplasm removal. Functional tests of sperm motility show a lack of progressive motility of spermatozoa from Mll5 deficient animals. None of these defects could be rescued by in vitro fertilization. Using microarray analysis we show that transcripts implicated in spermatogenesis are dysregulated., Conclusions/significance: Our data demonstrate a clear role of Mll5 in mammalian spermatogenesis at the level of terminal differentiation providing further support for its classification in the SET3 branch of proteins. Moreover, this study identifies Tlk2, Utx, Gpr64, Sult4a1, Rap2ip, Vstm2 and HoxA10 as possible Mll5 targets that together may account for the observed spermatozoa maturation defects.

P-cadherin is a calcium-dependent cell-cell adhesion glycoprotein. P-cadherin expression is restricted to the myoepithelial cells in normal breast tissue, and aberrant staining has also been described in invasive tumors. Several small studies have reported P-cadherin as a marker of poor outcome in breast cancer patients but its prognostic significance in relation to other variables has not been established in a large series of breast cancers. A tissue microarray was constructed from 3992 cases of invasive breast carcinoma, and P-cadherin expression was evaluated using immunohistochemistry. Median follow-up was 12.5 years. The immunohistochemistry-based definitions of cancer subtypes were luminal (ER+ or PR+/HER2-), luminal/HER2+ (ER+ or PR+/HER2+), HER2+ (ER-/PR-/HER2+), and basal (ER-/PR-/HER2-/CK5/6+ or EGFR+). Clinical covariate and biomarker associations were assessed using contingency tables, and Pearson's χ(2) or Fisher's exact test. Survival associations were assessed using Kaplan-Meier plots, logrank and Breslow tests, and Cox proportional hazards regression analysis. P-cadherin was expressed in 34.8% (1290/3710, 50% cut point) of cases. P-cadherin staining was strongly associated with HER2+ and basal carcinoma subtypes (P<0.0005). P-cadherin-positive patients showed significantly poorer short-term (0-10 years) overall survival, disease-specific survival, distant relapse-free interval, and locoregional relapse-free interval in univariable models (P<0.05). In multivariable Cox models containing standard clinical covariates and cancer subtypes, P-cadherin did not show independent prognostic value. P-cadherin expression was positively associated with histological grade, chemotherapy, Ki-67, EGFR, CK5/6, p53, YB-1, and HER2 expression (P<0.002), and negatively associated with age at diagnosis, ER, PR, and Bcl-2 expression (P<0.0005). This study shows the value of P-cadherin as a marker of poor prognosis. The large sample size of this series clarifies contradictory findings of many smaller studies. P-cadherin positivity is associated with high-grade tumor subtypes and well-established markers of poor prognosis, and may represent a promising antibody therapeutic target.

Adhesion-GPCRs provide essential cell-cell and cell-matrix interactions in development, and have been implicated in inherited human diseases like Usher Syndrome and bilateral frontoparietal polymicrogyria. They are the second largest subfamily of seven-transmembrane spanning proteins in vertebrates, but the function of most of these receptors is still not understood. The orphan Adhesion-GPCR GPR126 has recently been shown to play an essential role in the myelination of peripheral nerves in zebrafish. In parallel, whole-genome association studies have implicated variation at the GPR126 locus as a determinant of body height in the human population. The physiological function of GPR126 in mammals is still unknown. We describe a targeted mutation of GPR126 in the mouse, and show that GPR126 is required for embryonic viability and cardiovascular development.

The insulin-like growth factor-1 receptor (IGF1R) is emerging as a promising therapeutic target in human cancers. In the high-risk childhood sarcomas Ewing family tumor and rhabdomyosarcoma, IGF1R-blocking antibodies show impressive antitumor activity in some but not all patients, and acquired resistance is observed. Because tumor IGF1R mutations are not described, the basis of IGF1R inhibitor resistance remains unknown. We hypothesized that compensatory signaling cascades bypassing targeted IGF1R inhibition might be involved. To test this systematically, we performed small interfering RNA (siRNA) screens in sarcoma cell lines to identify IGF1R pathway components or related protein tyrosine kinase (PTK) networks that modulate the antitumor efficacy of the BMS-536924 IGF1R kinase inhibitor. This strategy revealed (a) that sarcoma cells are exquisitely sensitive to loss of distal rather than proximal IGF1R signaling components, such as ribosomal protein S6 (RPS6); (b) that BMS-536924 fails to block RPS6 activation in resistant sarcoma cell lines; and (c) that siRNA knockdown of the macrophage-stimulating 1 receptor tyrosine kinase (MST1R; also known as RON) restores BMS-536924 efficacy, even in highly drug-resistant cell lines. We confirmed MST1R expression across a broad panel of childhood sarcomas, and found that loss of MST1R by RNA interference blocks downstream RPS6 activation when combined with BMS-536924 in vitro. These findings underscore the importance of fully understanding PTK networks for successful clinical implementation of kinase inhibitor strategies., (©2010 AACR.)

Background: Ovarian clear-cell and endometrioid carcinomas may arise from endometriosis, but the molecular events involved in this transformation have not been described., Methods: We sequenced the whole transcriptomes of 18 ovarian clear-cell carcinomas and 1 ovarian clear-cell carcinoma cell line and found somatic mutations in ARID1A (the AT-rich interactive domain 1A [SWI-like] gene) in 6 of the samples. ARID1A encodes BAF250a, a key component of the SWI–SNF chromatin remodeling complex. We sequenced ARID1A in an additional 210 ovarian carcinomas and a second ovarian clear-cell carcinoma cell line and measured BAF250a expression by means of immunohistochemical analysis in an additional 455 ovarian carcinomas., Results: ARID1A mutations were seen in 55 of 119 ovarian clear-cell carcinomas (46%), 10 of 33 endometrioid carcinomas (30%), and none of the 76 high-grade serous ovarian carcinomas. Seventeen carcinomas had two somatic mutations each. Loss of the BAF250a protein correlated strongly with the ovarian clear-cell carcinoma and endometrioid carcinoma subtypes and the presence of ARID1A mutations. In two patients, ARID1A mutations and loss of BAF250a expression were evident in the tumor and contiguous atypical endometriosis but not in distant endometriotic lesions., Conclusions: These data implicate ARID1A as a tumor-suppressor gene frequently disrupted in ovarian clear-cell and endometrioid carcinomas. Since ARID1A mutation and loss of BAF250a can be seen in the preneoplastic lesions, we speculate that this is an early event in the transformation of endometriosis into cancer. (Funded by the British Columbia Cancer Foundation and the Vancouver General Hospital–University of British Columbia Hospital Foundation.).

The International Cancer Genome Consortium (ICGC) was launched to coordinate large-scale cancer genome studies in tumours from 50 different cancer types and/or subtypes that are of clinical and societal importance across the globe. Systematic studies of more than 25,000 cancer genomes at the genomic, epigenomic and transcriptomic levels will reveal the repertoire of oncogenic mutations, uncover traces of the mutagenic influences, define clinically relevant subtypes for prognosis and therapeutic management, and enable the development of new cancer therapies.

Follicular lymphoma (FL) and the GCB subtype of diffuse large B-cell lymphoma (DLBCL) derive from germinal center B cells. Targeted resequencing studies have revealed mutations in various genes encoding proteins in the NF-kappaB pathway that contribute to the activated B-cell (ABC) DLBCL subtype, but thus far few GCB-specific mutations have been identified. Here we report recurrent somatic mutations affecting the polycomb-group oncogene EZH2, which encodes a histone methyltransferase responsible for trimethylating Lys27 of histone H3 (H3K27). After the recent discovery of mutations in KDM6A (UTX), which encodes the histone H3K27me3 demethylase UTX, in several cancer types, EZH2 is the second histone methyltransferase gene found to be mutated in cancer. These mutations, which result in the replacement of a single tyrosine in the SET domain of the EZH2 protein (Tyr641), occur in 21.7% of GCB DLBCLs and 7.2% of FLs and are absent from ABC DLBCLs. Our data are consistent with the notion that EZH2 proteins with mutant Tyr641 have reduced enzymatic activity in vitro.

Introduction: Breast cancer is genetically and clinically a heterogeneous disease. However, the exact contribution of different cell types and oncogenic mutations to this heterogeneity are not well understood. Recently, we discovered an interaction between Wnt and integrin-linked kinase (ILK) within the signaling cascade that regulates cell growth and survival. Interestingly, mammary-specific expression of either one of these proteins has been shown to promote mammary tumorigenesis. In light of our recent findings and to investigate the potential interaction between Wnt and ILK proteins during mammary tumor formation and progression, we established a transgenic mouse model that expresses both Wnt and ILK in mammary epithelial cells., Methods: A novel transgenic mouse model with mammary-specific expression of both Wnt1 and ILK was generated by crossing the two previously characterized mouse models, MMTV-Wnt1 and MMTV-ILK. The resulting MMTV-Wnt/ILK mice were closely monitored for tumor development and growth, as well as for the tumor onset. The molecular phenotypes of both tumors and premalignant mammary glands were investigated by using biochemical and global gene-expression analysis approaches., Results: A significant acceleration in mammary tumor incidence and growth was observed in the MMTV-Wnt/ILK mice. Pre-neoplastic mammary glands also display lobuloalveolar hyperplasia and an increase in ductal epithelium proliferation. Apart from elevated expression of Wnt/ILK targets, such as beta-catenin and cyclin D1, gene-expression profiling identified the surprising activation of the FOXA1 transcription factor. Upregulation of FOXA1, which is also known as the molecular marker of differentiated mammary luminal cells, was consistent with the expansion of the enriched luminal progenitor population or CD29loCD24hiCD61+ cells in MMTV-Wnt/ILK tumors., Conclusions: These results show cooperation between Wnt1 and ILK transgenes during mammary carcinogenesis, leading to changes in a transcriptional network, which could dictate a specific breast cancer phenotype with enhanced growth dynamics. The MMTV-Wnt/ILK can be used as a model to identify further the genes downstream of the estrogen receptor-beta/FOXA1 and to investigate the mechanisms targeting the expansion of the luminal progenitor cells leading to hyperplasia and tumorigenesis.

Identifying new regulators of the stem cell state offers potential for future gains in biomedicine. Evidence that the tumor suppressor p53 is a key regulator of the stem cell state (Cicalese et al., 2009) suggests a broad role for this protein and its pathways in the control of normal tissue homeostasis and tumor formation.

Background: Granulosa-cell tumors (GCTs) are the most common type of malignant ovarian sex cord-stromal tumor (SCST). The pathogenesis of these tumors is unknown. Moreover, their histopathological diagnosis can be challenging, and there is no curative treatment beyond surgery., Methods: We analyzed four adult-type GCTs using whole-transcriptome paired-end RNA sequencing. We identified putative GCT-specific mutations that were present in at least three of these samples but were absent from the transcriptomes of 11 epithelial ovarian tumors, published human genomes, and databases of single-nucleotide polymorphisms. We confirmed these variants by direct sequencing of complementary DNA and genomic DNA. We then analyzed additional tumors and matched normal genomic DNA, using a combination of direct sequencing, analyses of restriction-fragment-length polymorphisms, and TaqMan assays., Results: All four index GCTs had a missense point mutation, 402C-->G (C134W), in FOXL2, a gene encoding a transcription factor known to be critical for granulosa-cell development. The FOXL2 mutation was present in 86 of 89 additional adult-type GCTs (97%), in 3 of 14 thecomas (21%), and in 1 of 10 juvenile-type GCTs (10%). The mutation was absent in 49 SCSTs of other types and in 329 unrelated ovarian or breast tumors., Conclusions: Whole-transcriptome sequencing of four GCTs identified a single, recurrent somatic mutation (402C-->G) in FOXL2 that was present in almost all morphologically identified adult-type GCTs. Mutant FOXL2 is a potential driver in the pathogenesis of adult-type GCTs., (2009 Massachusetts Medical Society)

Introduction: We have previously identified a rare subpopulation of variant human mammary epithelial cells (vHMEC) with repressed p16INK4A that exist in disease-free women yet display premalignant properties, suggesting that they have engaged the process of malignant transformation. In order to gain insight into the molecular alterations required for vHMEC to progress to malignancy, and to characterize the epigenetic events associated with early progression, we examined the effect of oncogenic stress on the behavior of these cells., Methods: HMEC that express p16INK4A and vHMEC that do not, were transduced with constitutively active Ha-rasV12 and subsequently exposed to serum to determine whether signals from the cellular microenvironment could cooperate with ras to promote the malignant transformation of vHMEC. Epigenetic alterations were assessed using methylation-specific polymerase chain reaction (PCR)., Results: vHMEC expressing Ha-rasV12 (vHMEC-ras) bypassed the classic proliferative arrest that has been previously documented in normal fibroblasts following oncogenic stress, and that we also observe here in normal HMEC. Moreover, vHMEC-ras cells exhibited many additional alterations that are observed during progression to malignancy such as the generation of chromosomal abnormalities, upregulation of telomerase activity, immortalization following exposure to serum, and anchorage-independent growth, but they did not form tumors following orthotopic injection in vivo. Associated with their early progression to malignancy was an increase in the number of genes methylated, two of which (RASSF1A and SFRP1) were also methylated in other immortalized mammary cell lines as well as in breast cancer cells and tissues., Conclusions: We have characterized a mammary progression model that recapitulates molecular and methylation alterations observed in many breast cancers. Our data suggest that concomitant methylation of RASSF1A and SFRP1 marks an early event in mammary transformation and may thus have prognostic potential.

Recent genetic evidence in humans and from mouse knockouts has linked kisspeptin-driven GPR54 signaling to the regulation of GnRH release from the hypothalamus. These molecules appear to represent a previously unsuspected control point for GnRH secretion, with important implications for the biology and pathology of the sex steroid axis.

Genetic approaches to validating G protein-coupled receptors (GPCRs) have proven to be a powerful research tool, especially knockout studies in rodents. To date, data related to in vivo function have been published on approximately half of the human rhodopsin-like family-1 GPCRs, which can be attributed to the use of mouse knockouts. It is likely that many currently unknown yet important therapeutic mechanisms will be uncovered through knockout screens in mice. One such recent discovery is the elucidation of the in vivo function of the GPCR GPR54 through mouse genetics, and its subsequent validation in human populations. Although previously unsuspected, GPR54 has been found to be a master-regulator of the hypothalamic-pituitary-gonadal axis.

With about 24,000 extant species, teleosts are the largest group of vertebrates. They constitute more than 99% of the ray-finned fishes (Actinopterygii) that diverged from the lobe-finned fish lineage (Sarcopterygii) about 450 MYA. Although the role of genome duplication in the evolution of vertebrates is now established, its role in structuring the teleost genomes has been controversial. At least two hypotheses have been proposed: a whole-genome duplication in an ancient ray-finned fish and independent gene duplications in different lineages. These hypotheses are, however, based on small data sets and lack adequate statistical and phylogenetic support. In this study, we have made a systematic comparison of the draft genome sequences of Fugu and humans to identify paralogous chromosomal regions ("paralogons") in the Fugu that arose in the ray-finned fish lineage ("fish-specific"). We identified duplicate genes in the Fugu by phylogenetic analyses of the Fugu, human, and invertebrate sequences. Our analyses provide evidence for 425 fish-specific duplicate genes in the Fugu and show that at least 6.6% of the genome is represented by fish-specific paralogons. We estimated the ages of Fugu duplicate genes and paralogons using the molecular clock. Remarkably, the ages of duplicate genes and paralogons are clustered, with a peak around 350 MYA. These data strongly suggest a whole-genome duplication event early during the evolution of ray-finned fishes, probably before the origin of teleosts.

The BRCA2 gene is mutated in familial breast and ovarian cancer, and its product is implicated in DNA repair and transcriptional regulation. Here we identify a protein, EMSY, which binds BRCA2 within a region (exon 3) deleted in cancer. EMSY is capable of silencing the activation potential of BRCA2 exon 3, associates with chromatin regulators HP1beta and BS69, and localizes to sites of repair following DNA damage. EMSY maps to chromosome 11q13.5, a region known to be involved in breast and ovarian cancer. We show that the EMSY gene is amplified almost exclusively in sporadic breast cancer (13%) and higher-grade ovarian cancer (17%). In addition, EMSY amplification is associated with worse survival, particularly in node-negative breast cancer, suggesting that it may be of prognostic value. The remarkable clinical overlap between sporadic EMSY amplification and familial BRCA2 deletion implicates a BRCA2 pathway in sporadic breast and ovarian cancer.