Characterisation of multifocal breast cancer using the 70-gene signature in clinical low-risk patients enrolled in the EORTC 10041/BIG 03-04 MINDACT trial.

BACKGROUND: In multifocal breast cancer, guidelines recommend basing adjuvant systemic treatment decisions on characteristics of the largest lesion, disregarding multifocality as an independent prognosticator. We assessed the association between multifocal disease and both the 70-gene signature (70-GS), and distant metastasis-free survival (DMFS) in clinical low-risk breast cancer patients enrolled in the European Organisation for Research and Treatment of Cancer 10041/BIG 03-04 Microarray In Node-negative and 1 to 3 positive lymph node Disease may Avoid ChemoTherapy (MINDACT) trial. PATIENTS AND METHODS: The analysed population consisted of enrolled patients in the MINDACT trial with clinical low-risk disease, defined by a modified Adjuvant! Online cut-off for the 10-year risk of recurrent disease or death. Eligibility criteria of MINDACT dictate that patients with multifocal disease could be included if the different lesions had similar pathological characteristics. The presence of multifocal disease was deducted from the case report form (CRF)-question for sum of diameter for all invasive tumour foci. Clinicopathological characteristics and gene expression of patients with unifocal and multifocal (largest lesion) disease were compared. Subsequently, the association between multifocal disease and the 70-GS was evaluated as well as the association between multifocality and 5-year DMFS. RESULTS: The study included 3090 clinical low-risk patients with unifocal and 238 patients with multifocal disease. Apart from a higher prevalence of lobular tumours (21.8% versus 10.8%, by local pathology), we did not observe differences in baseline characteristics between multifocal and unifocal tumours. Patients with multifocal tumours were more likely to be at high genomic risk as compared to patients with unifocal tumours (22.7% versus 17.3%, odds ratio [OR] 1.45, 95% confidence interval [CI] 1.02-2.07, P = 0.038). We did not find a significant association between tumour focality and DMFS (97.1% for unifocal versus 96.9% for multifocal, hazard ratio [HR] = 1.55, 95% CI 0.68-3.46, P = 0.172), nor a signal for a potential interaction between the prognostic effect of the 70-GS and focality of the tumour regarding DMFS. CONCLUSION: In the group of clinical low-risk MINDACT patients, multifocal tumours were more likely to have a high-risk 70-GS profile compared to unifocal tumours. We did not observe a significant interaction between multifocality and the 70-GS with respect to survival without distant metastasis in these patients.

Prediction of overall survival in stage II and III colon cancer beyond TNM system: a retrospective, pooled biomarker study.

Background: TNM staging alone does not accurately predict outcome in colon cancer (CC) patients who may be eligible for adjuvant chemotherapy. It is unknown to what extent the molecular markers microsatellite instability (MSI) and mutations in BRAF or KRAS improve prognostic estimation in multivariable models that include detailed clinicopathological annotation. Patients and methods: After imputation of missing at random data, a subset of patients accrued in phase 3 trials with adjuvant chemotherapy (n = 3016)-N0147 (NCT00079274) and PETACC3 (NCT00026273)-was aggregated to construct multivariable Cox models for 5-year overall survival that were subsequently validated internally in the remaining clinical trial samples (n = 1499), and also externally in different population cohorts of chemotherapy-treated (n = 949) or -untreated (n = 1080) CC patients, and an additional series without treatment annotation (n = 782). Results: TNM staging, MSI and BRAFV600E mutation status remained independent prognostic factors in multivariable models across clinical trials cohorts and observational studies. Concordance indices increased from 0.61-0.68 in the TNM alone model to 0.63-0.71 in models with added molecular markers, 0.65-0.73 with clinicopathological features and 0.66-0.74 with all covariates. In validation cohorts with complete annotation, the integrated time-dependent AUC rose from 0.64 for the TNM alone model to 0.67 for models that included clinicopathological features, with or without molecular markers. In patient cohorts that received adjuvant chemotherapy, the relative proportion of variance explained (R2) by TNM, clinicopathological features and molecular markers was on an average 65%, 25% and 10%, respectively. Conclusions: Incorporation of MSI, BRAFV600E and KRAS mutation status to overall survival models with TNM staging improves the ability to precisely prognosticate in stage II and III CC patients, but only modestly increases prediction accuracy in multivariable models that include clinicopathological features, particularly in chemotherapy-treated patients.

Prognosis of stage II and III colon cancer treated with adjuvant 5-fluorouracil or FOLFIRI in relation to microsatellite status: results of the PETACC-3 trial.

BACKGROUND: Although colon cancer (CC) with microsatellite instability (MSI) has a more favorable prognosis than microsatellite stable (MSS) CC, the impact varies according to clinicopathological parameters. We studied how MSI status affects prognosis in a trial-based cohort of stage II and III CC patients treated with 5-fluorouracil (5-FU)/leucovorin or FOLFIRI. MATERIALS AND METHODS: Tissue specimens of 1254 patients were tested for 10 different loci and were classified as MSI-high (MSI-H) when three or more loci were unstable and MSS otherwise. Study end points were overall survival (OS) and relapse-free survival (RFS). RESULTS: In stage II, RFS and OS were better for patients with MSI-H than with MSS CC [hazard ratio (HR) 0.26, 95% CI 0.10-0.65, P = 0.004 and 0.16, 95% CI 0.04-0.64, P = 0.01). In stage III, RFS was slightly better for patients with MSI-H CC (HR 0.67, 95% CI 0.46-0.99, P = 0.04), but the difference was not statistically significant for OS (HR 0.70, 95% CI 0.44-1.09, P = 0.11). Outcomes for patients with MSI-H CC were not different between the two treatment arms. RFS was better for patients with MSI-H than with MSS CC in the right and left colon, whereas for OS this was significant only in the right colon. For patients with KRAS- and BRAF-mutated CC, but not for double wild-type patients, RFS and OS were significantly better when the tumors were also MSI-H. An interaction test was statistically significant for KRAS and MSI status (P = 0.005), but not for BRAF status (P = 0.14). CONCLUSIONS: Our results confirm that for patients with stage II CC but less so for those with stage III MSI-H is strongly prognostic for RFS and OS. In the presence of 5-FU treatment, stage II patients with MSI-H tumors maintain their survival advantage in comparison with MSS patients and adding irinotecan has no added benefit. CLINICALTRIALS.GOV IDENTIFIER: NCT00026273.

Distal and proximal colon cancers differ in terms of molecular, pathological, and clinical features.

BACKGROUND: Differences exist between the proximal and distal colon in terms of developmental origin, exposure to patterning genes, environmental mutagens, and gut flora. Little is known on how these differences may affect mechanisms of tumorigenesis, side-specific therapy response or prognosis. We explored systematic differences in pathway activation and their clinical implications. MATERIALS AND METHODS: Detailed clinicopathological data for 3045 colon carcinoma patients enrolled in the PETACC3 adjuvant chemotherapy trial were available for analysis. A subset of 1404 samples had molecular data, including gene expression and DNA copy number profiles for 589 and 199 samples, respectively. In addition, 413 colon adenocarcinoma from TCGA collection were also analyzed. Tumor side-effect on anti-epidermal growth factor receptor (EGFR) therapy was assessed in a cohort of 325 metastatic patients. Outcome variables considered were relapse-free survival and survival after relapse (SAR). RESULTS: Proximal carcinomas were more often mucinous, microsatellite instable (MSI)-high, mutated in key tumorigenic pathways, expressed a B-Raf proto-oncogene, serine/threonine kinase (BRAF)-like and a serrated pathway signature, regardless of histological type. Distal carcinomas were more often chromosome instable and EGFR or human epidermal growth factor receptor 2 (HER2) amplified, and more frequently overexpressed epiregulin. While risk of relapse was not different per side, SAR was much poorer for proximal than for distal stage III carcinomas in a multivariable model including BRAF mutation status [N = 285; HR 1.95, 95% CI (1.6-2.4), P < 0.001]. Only patients with metastases from a distal carcinoma responded to anti-EGFR therapy, in line with the predictions of our pathway enrichment analysis. CONCLUSIONS: Colorectal carcinoma side is associated with differences in key molecular features, some immediately druggable, with important prognostic effects which are maintained in metastatic lesions. Although within side significant molecular heterogeneity remains, our findings justify stratification of patients by side for retrospective and prospective analyses of drug efficacy and prognosis.

Identification of MAGI1 as a tumor-suppressor protein induced by cyclooxygenase-2 inhibitors in colorectal cancer cells.

Cyclooxyganase-2 (COX-2), a rate-limiting enzyme in the prostaglandin synthesis pathway, is overexpressed in many cancers and contributes to cancer progression through tumor cell-autonomous and paracrine effects. Regular use of non-steroidal anti-inflammatory drugs or selective COX-2 inhibitors (COXIBs) reduces the risk of cancer development and progression, in particular of the colon. The COXIB celecoxib is approved for adjunct therapy in patients with Familial adenomatous polyposis at high risk for colorectal cancer (CRC) formation. Long-term use of COXIBs, however, is associated with potentially severe cardiovascular complications, which hampers their broader use as preventive anticancer agents. In an effort to better understand the tumor-suppressive mechanisms of COXIBs, we identified MAGUK with Inverted domain structure-1 (MAGI1), a scaffolding protein implicated in the stabilization of adherens junctions, as a gene upregulated by COXIB in CRC cells and acting as tumor suppressor. Overexpression of MAGI1 in CRC cell lines SW480 and HCT116 induced an epithelial-like morphology; stabilized E-cadherin and ß-catenin localization at cell-cell junctions; enhanced actin stress fiber and focal adhesion formation; increased cell adhesion to matrix proteins and suppressed Wnt signaling, anchorage-independent growth, migration and invasion in vitro. Conversely, MAGI1 silencing decreased E-cadherin and ß-catenin localization at cell-cell junctions; disrupted actin stress fiber and focal adhesion formation; and enhanced Wnt signaling, anchorage-independent growth, migration and invasion in vitro. MAGI1 overexpression suppressed SW480 and HCT116 subcutaneous primary tumor growth, attenuated primary tumor growth and spontaneous lung metastasis in an orthotopic model of CRC, and decreased the number and size of metastatic nodules in an experimental model of lung metastasis. Collectively, these results identify MAG1 as a COXIB-induced inhibitor of the Wnt/ß-catenin signaling pathway, with tumor-suppressive and anti-metastatic activity in experimental colon cancer.

MFG-E8/lactadherin regulates cyclins D1/D3 expression and enhances the tumorigenic potential of mammary epithelial cells.

Milk fat globule-EGF factor 8 (MFG-E8) is a glycoprotein highly expressed in breast cancer that contributes to tumor progression through largely undefined mechanisms. By analyzing publicly available gene expression profiles of breast carcinomas, we found that MFG-E8 is highly expressed in primary and metastatic breast carcinomas, associated with absent estrogen receptor expression. Immunohistochemistry analysis of breast cancer biopsies revealed that MFG-E8 is expressed on the cell membrane as well as in the cytoplasm and nucleus. We also show that increased expression of MFG-E8 in mammary carcinoma cells increases their tumorigenicity in immunodeficient mice, and conversely, its downregulation reduces their in vivo growth. Moreover, expression of MFG-E8 in immortalized mammary epithelial cells promotes their growth and branching in three-dimensional collagen matrices and induces the expression of cyclins D1/D3 and N-cadherin. A mutant protein unable to bind integrins can in part exert these effects, indicating that MFG-E8 function is only partially dependent on integrin activation. We conclude that MFG-E8-dependent signaling stimulates cell proliferation and the acquisition of mesenchymal properties and contributes to mammary carcinoma development.

p-Medicine: From data sharing and integration via VPH models to personalized medicine.

The Worldwide innovative Networking in personalized cancer medicine (WIN) initiated by the Institute Gustave Roussy (France) and The University of Texas MD Anderson Cancer Center (USA) has dedicated its 3rd symposium (Paris, 6-8 July 2011) to discussion on gateways to increase the efficacy of cancer diagnostics and therapeutics (http://www.winconsortium.org/symposium.html).Speakers ranged from clinical oncologist to researchers, industrial partners, and tools developers; a famous patient was present: Janelle Hail, a 30-year breast cancer survivor, founder and CEO of the National Breast Cancer Foundation, Inc. (NBCF).The p-medicine consortium found this venue a perfect occasion to present a poster about its activities that are in accordance with the take home message of the symposium.In this communication, we summarize what we presented with particular attention to the interaction between the symposium's topic and content and our project.

Non-invasive gene-expression-based detection of well-developed collateral function in individuals with and without coronary artery disease.

BACKGROUND: In patients with coronary artery disease (CAD), a well grown collateral circulation has been shown to be important. The aim of this prospective study using peripheral blood monocytes was to identify marker genes for an extensively grown coronary collateral circulation. METHODS: Collateral flow index (CFI) was obtained invasively by angioplasty pressure sensor guidewire in 160 individuals (110 patients with CAD, and 50 individuals without CAD). RNA was extracted from monocytes followed by microarray-based gene-expression analysis. 76 selected genes were analysed by real-time polymerase chain reaction (PCR). A receiver operating characteristics analysis based on differential gene expression was then performed to separate individuals with poor (CFI<0.21) and well-developed collaterals (CFI>or=0.21) Thereafter, the influence of the chemokine MCP-1 on the expression of six selected genes was tested by PCR. RESULTS: The expression of 203 genes significantly correlated with CFI (p = 0.000002-0.00267) in patients with CAD and 56 genes in individuals without CAD (p = 00079-0.0430). Biological pathway analysis revealed 76 of those genes belonging to four different pathways: angiogenesis, integrin-, platelet-derived growth factor-, and transforming growth factor beta-signalling. Three genes in each subgroup differentiated with high specificity among individuals with low and high CFI (>or=0.21). Two out of these genes showed pronounced differential expression between the two groups after cell stimulation with MCP-1. CONCLUSIONS: Genetic factors play a role in the formation and the preformation of the coronary collateral circulation. Gene expression analysis in peripheral blood monocytes can be used for non-invasive differentiation between individuals with poorly and with well grown collaterals. MCP-1 can influence the arteriogenic potential of monocytes.

A gene expression signature that distinguishes desmoid tumours from nodular fasciitis.

Nodular fasciitis (NF) is a rapidly growing cellular mass composed of fibroblasts/myofibroblasts, usually localized in subcutaneous tissues, that typically undergoes fibrosis and almost never recurs. Desmoid tumours (DTs) are rare forms of fibroblastic/myofibroblastic growth that arise in deep soft tissues, display a propensity for local infiltration and recurrence, but fail to metastasize. Given that both entities are primarily fibroblastic/myofibroblastic lesions with overlapping histological features, their gene expression profiles were compared to identify differentially expressed genes that may provide not only potential diagnostic markers, but also clues as to the pathogenesis of each disorder. Differentially expressed transcripts (89 clones displaying increased expression in DTs and 246 clones displaying increased expression in NF) included genes encoding several receptor and non-receptor tyrosine kinases (EPHB3, PTPRF, GNAZ, SYK, LYN, EPHA4, BIRC3), transcription factors (TWIST1, PITX2, EYA2, OAS1, MITF, TCF20), and members of the Wnt signalling pathway (AXIN2, WISP1, SFRP). Remarkably, almost one-quarter of the differentially expressed genes encode proteins associated with inflammation and tissue remodelling, including members of the interferon (IFN), tumour necrosis factor (TNF), and transforming growth factor beta (TGF-beta) signalling pathways as well as metalloproteinases (MMP1, 9, 13, 23), urokinase plasminogen activator (PLAU), and cathepsins. The observations provide the first comparative molecular characterization of desmoid tumours and nodular fasciitis and suggest that selected tyrosine kinases, transcription factors, and members of the Wnt, TGF-beta, IFN, and TNF signalling pathways may be implicated in influencing and distinguishing their fate.

Aplicación de un programa de acreditación en servicios de farmacia hospitalarios en argentina / Implementing and Accreditation program for in-hospital Pharmacy Departments in Argentina

Objetivo: Desarrollar un programa de Acreditación de Servicios de Farmacia hospitalarios que involucre diferentes alternativas de evaluación, para analizar la calidad de las prestaciones brindadas con la finalidad de promover su mejora continua. Método: Análisis de programas de acreditación ya implementados; diseño de un programa adecuado al desarrollo de los servicios en Argentina; aplicación y análisis de los resultados obtenidos. Resultados: El programa considera todas las actividades posibles de ser desarrolladas, mediante un proceso evaluativo que incluye el uso de un Cuestionario Guía, indicadores específicos con valores estándar asignados, inspección del servicio y documentación. Conclusión: El presente Programa es una propuesta para iniciar a los Servicios de Farmacia Hospitalaria en el proceso de acreditación. El propio programa debe ser corregido y optimizado en base a los resultados que se obtengan y a la evolución de las técnicas de evaluación que se proponen a nivel internacional (AU)

Identification of proteins from Plasmodium falciparum that are homologous to reticulocyte binding proteins in Plasmodium vivax.

Plasmodium falciparum infections can be fatal, while P. vivax infections usually are not. A possible factor involved in the greater virulence of P. falciparum is that this parasite grows in red blood cells (RBCs) of all maturities whereas P. vivax is restricted to growth in reticulocytes, which represent only approximately 1% of total RBCs in the periphery. Two proteins, expressed at the apical end of the invasive merozoite stage from P. vivax, have been implicated in the targeting of reticulocytes for invasion by this parasite. A search of the P. falciparum genome databases has identified genes that are homologous to the P. vivax rbp-1 and -2 genes. Two of these genes are virtually identical over a large region of the 5' end but are highly divergent at the 3' end. They encode high-molecular-mass proteins of >300 kDa that are expressed in late schizonts and localized to the apical end of the merozoite. To test a potential role in merozoite invasion of RBCs, we analyzed the ability of these proteins to bind to mature RBCs and reticulocytes. No binding to mature RBCs or cell preparations enriched for reticulocytes was detected. We identified a parasite clone that lacks the gene for one of these proteins, showing that the gene is not required for normal in vitro growth. Antibodies to these proteins can inhibit merozoite invasion of RBCs.

Expression of Abdominal-B homeoproteins in Drosophila embryos.

The Abdominal-B (Abd-B) gene determines development of the posteriormost segments in Drosophila. Genetic and molecular analysis suggested that it consists of two genetically separable functions that are conferred by two related homeoproteins termed m and r. We have raised an antiserum against Abd-B protein to describe the patterns of Abd-B protein expression during embryonic development. The pattern of r protein expression, as deduced by analysis of Abd-B mutants, is restricted to ps14 and 15 in all germ layers and observes a parasegmental boundary at its anterior margin of expression. In contrast, the pattern of m protein expression is unusual as its level in the ectoderm increases from ps10 to ps13 in parasegmental steps. Its anterior margin of expression is highly dynamic shifting anteriorly across more than 3 parasegments during midembryonic development. Evidently, the control mechanisms of m and r protein expression are considerably different. Moreover, an antibody-positive Abd-B mutant suggests that these differ, in the case of m protein expression, to some extent in individual germ layers.

Drosophila homoeotic genes encode transcriptional activators similar to mammalian OTF-2.

Homoeotic genes in Drosophila melanogaster are active in spatially restricted metameric domains and control the morphogenesis of segment-specific features such as legs or wings within these domains. They exert their function, according to the 'selector gene' hypothesis, by regulating the expression of subordinate genes. Homoeotic genes also control their own expression and the expression of each other. The proteins encoded by these genes contain a domain, called a homoeodomain, that is strongly conserved, and that shows homologies to proteins that bind DNA and regulate transcription. Homoeoproteins have been shown to bind specific DNA sequences. We show here that the Drosophila homoeotic genes Ultrabithorax (Ubx) and Abdominal-B (Abd-B) code for proteins that are capable of activating transcription of reporter genes linked to specific cis-regulatory target sequences in transfected mammalian cells. Their activity, as well as their target specificity, is similar to that of a mammalian lymphoid-specific octamer transcription factor, OTF-2, which was recently found to contain a homoeodomain.

Evidence that the Abdominal-B r element function is conferred by a trans-regulatory homeoprotein.

The Abdominal-B gene is a homeotic gene located in the distal-most region of the bithorax complex (BX-C). Based on complementation analysis it has been proposed that the gene contains two separable genetic elements, called the m and r elements. The r element has a chiefly regulatory function confined to parasegment 14. In a reverse Northern screen of the distal-most BX-C DNA, we found four distinct areas that are transcribed in the embryo. One of the transcripts spans a large genomic region which, upon disruption by rearrangement breakpoints, causes loss of r element function. This transcript is expressed in the early embryo in a single domain apparently corresponding to parasegment 14. We propose that the small homeoprotein encoded by the transcript acts as a trans-regulator to confer r element function.