Identification of a robust gene signature that predicts breast cancer outcome in independent data sets.

BACKGROUND: Breast cancer is a heterogeneous disease, presenting with a wide range of histologic, clinical, and genetic features. Microarray technology has shown promise in predicting outcome in these patients. METHODS: We profiled 162 breast tumors using expression microarrays to stratify tumors based on gene expression. A subset of 55 tumors with extensive follow-up was used to identify gene sets that predicted outcome. The predictive gene set was further tested in previously published data sets. RESULTS: We used different statistical methods to identify three gene sets associated with disease free survival. A fourth gene set, consisting of 21 genes in common to all three sets, also had the ability to predict patient outcome. To validate the predictive utility of this derived gene set, it was tested in two published data sets from other groups. This gene set resulted in significant separation of patients on the basis of survival in these data sets, correctly predicting outcome in 62-65% of patients. By comparing outcome prediction within subgroups based on ER status, grade, and nodal status, we found that our gene set was most effective in predicting outcome in ER positive and node negative tumors. CONCLUSION: This robust gene selection with extensive validation has identified a predictive gene set that may have clinical utility for outcome prediction in breast cancer patients.

Genomic alterations in human mesothelioma including high resolution mapping of common regions of DNA loss in chromosome arm 6q.

BACKGROUND: Molecular genetic analysis of 14 freshly resected human mesotheliomas was used to identify regions in the tumor genomes that display DNA copy number alterations, especially the regions that may harbor tumor suppressor genes. MATERIALS AND METHODS: Three methods for molecular analysis were used, comparative genomic hybridization (CGH), loss of heterozygosity (LOH) and a new method, quantitative microsatellite analysis (QuMA). RESULTS: The most frequent alteration detected by CGH and LOH was deletion in chromosome 6q, for which QuMA was performed at 30 different loci so as to define the region(s) of common deletion(s). Our data indicates that there are three independent regions of common deletion, one of size 8.4 Mb located at 6q14, a second of size 15.9 Mb at 6q22 and a third of size 12.0 Mb at 6q24. CONCLUSION: This suggests that at least 3 tumor suppressor genes mapped to chromosome 6q are commonly involved in the pathogenesis of mesothelioma.

Differential protein expression in MCF7 breast cancer cells transfected with ErbB2, neomycin resistance and luciferase plus yellow fluorescent protein.

Gene transfection is frequently used to explore the molecular and phenotypic consequences of introduced genes. Breast cancer cell lines transfected with genes for growth factor receptors, intracellular signaling molecules or genes that generate luminescent signals are widely used in basic science and preclinical studies. Typically, a target gene of interest is co-transfected with selectable markers that are generally assumed to be innocuous. Perturbations of the cellular genome by transfected sequences may induce subtle and/or unexpected modulations in protein expression, only some of which may be attributable to the target gene of interest. In this study, we show that neomycin resistant MCF7 cells (MCF7 Neo(r)) proliferate twice as rapidly in nude mice as do the untransfected parent cells, but show similar growth rates in vitro. MCF7 transfected with the ErbB2 gene shows minimal alteration in growth rate in vitro, and approximately a threefold increased growth rate in vivo. MCF7 cells that express luciferase and yellow fluorescent protein proliferate slowly in vitro and show essentially no growth in vivo suggesting that overexpression of these tracking proteins adversely affects cellular proliferative capacity. The molecular basis for alterations in proliferative capacity of the transfected sub-lines is poorly understood. We performed two-dimensional gel electrophoresis (2-DE) to compare relative protein expression among the cell lines. Relative to the parental MCF7, transfected cell lines displayed numerous differentially expressed proteins (69 to 149), relative to parental MCF7. Twenty-one of these differentially expressed proteins were identified by mass spectrometry, and included metabolic, structural, and signaling proteins. Possible roles of differentially expressed proteins in altering cellular proliferation are discussed.

Genetic aberrations in prostate carcinoma detected by comparative genomic hybridization and microsatellite analysis: association with progression and angiogenesis.

BACKGROUND: In spite of increasing knowledge about the tumor biology of prostate cancer (PC), molecular events involved in tumor progression are not well characterized. There is evidence that a number of genetic alterations play a role in tumor progression and in addition, angiogenesis also contributes. In this study, comparative genomic hybridization (CGH), a sensitive method for detecting regional DNA copy number abnormalities, and microsatellite analysis was used to identify frequent genome changes in PC. Correlation of these data with microvessel density (MVD) and clinical follow-up data was performed to determine genetic alterations that are associated with angiogenesis and subsequent tumor progression. METHODS: Fifty-seven paraffin embedded radical prostatectomy (RP) specimens were microdissected. DNA from the microdissected PC tissue was amplified by degenerate oligonucleoitide primed (DOP)-polymerase chain reaction (PCR), and CGH was performed on the PCR product. Quantitative analyses of the CGH profiles were performed using a t-statistic. Additionally, a microsatellite analysis of chromosome 13q was performed on a subgroup of 31 of the tumors. Using a polyclonal antibody against factor VIII, MVD was determined for all RP specimens. The results of CGH and microsatellite analysis were correlated with the clinical data of the patients and with MVD. RESULTS: Forty-two of the tumors (75%) showed one or more gains while 39 (70%) showed one or more losses per tumor. The most frequent DNA copy number gains were on chromosome 3, 4, 7, 8, 10, 11, 12, 13, and X. The most frequent losses were on chromosomes 2, 5, 6, 8, 10, 13, 15, and 16. Cancer recurrence occurred in 15 patients. The total number of DNA copy number losses was significantly higher in patients with this progression (86%) than without (52%) (P < 0.001). There was no significant difference in the number of gains in patients with or without progression. Contingency table analysis showed a significant correlation between progression and losses in regions of chromosomes 6q and 13q and a gain of chromosome 7q. In multivariate analysis, only loss of chromosome 6 was independently prognostic. The gains that correlated most closely with MVD > 35 were on chromosomes 2q, 7q, and Xq, while the losses most closely associated with MVD > 35 were on chromosomes 8q, 10q, and 13q. However, only the association between loss of chromosome 13q and MVD > 35 was statistically significant. Microsatellite analysis revealed a statistically significant correlation between MVD and instability of locus 171. CONCLUSIONS: This study indicates that the frequency of genetic alterations in PC as detected by CGH correlates with clinical outcome, and that losses of DNA from chromosomes 6q and 13q are important events that correlate with tumor progression, with loss of 13q, especially instability of locus 171, also associated with angiogenesis.