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Translating the cancer genome: Going beyond p values

Description: Cancer cells are endowed with diverse biological capabilities driven by myriad inherited and somatic genetic and epigenetic aberrations that commandeer key cancer-relevant pathways. Efforts to elucidate these aberrations began with Boveri's hypothesis of aberrant mitoses causing cancer and continue today with a suite of powerful high-resolution technologies that enable detailed catalogues of genomic aberrations and epigenomic modifications. Tomorrow will likely bring the complete atlas of reversible and irreversible alteration in individual cancers. The challenge now is to discern causal molecular abnormalities from genomic and epigenomic 'noise', to understand how the ensemble of these aberrations collaborate to drive cancer pathophysiology. Here, we highlight lessons learned from now classical examples of successful translation of genomic discoveries into clinical practice, lessons that may be used to guide and accelerate translation of emerging genomic insights into practical clinical endpoints that can impact on practice of cancer medicine.
Date: April 3, 2008
Creator: Chin, Lynda; Chin, Lynda & Gray, Joe W.
Partner: UNT Libraries Government Documents Department

Chapter 27 -- Breast Cancer Genomics, Section VI, Pathology and Biological Markers of Invasive Breast Cancer

Description: Breast cancer is predominantly a disease of the genome with cancers arising and progressing through accumulation of aberrations that alter the genome - by changing DNA sequence, copy number, and structure in ways that that contribute to diverse aspects of cancer pathophysiology. Classic examples of genomic events that contribute to breast cancer pathophysiology include inherited mutations in BRCA1, BRCA2, TP53, and CHK2 that contribute to the initiation of breast cancer, amplification of ERBB2 (formerly HER2) and mutations of elements of the PI3-kinase pathway that activate aspects of epidermal growth factor receptor (EGFR) signaling and deletion of CDKN2A/B that contributes to cell cycle deregulation and genome instability. It is now apparent that accumulation of these aberrations is a time-dependent process that accelerates with age. Although American women living to an age of 85 have a 1 in 8 chance of developing breast cancer, the incidence of cancer in women younger than 30 years is uncommon. This is consistent with a multistep cancer progression model whereby mutation and selection drive the tumor's development, analogous to traditional Darwinian evolution. In the case of cancer, the driving events are changes in sequence, copy number, and structure of DNA and alterations in chromatin structure or other epigenetic marks. Our understanding of the genetic, genomic, and epigenomic events that influence the development and progression of breast cancer is increasing at a remarkable rate through application of powerful analysis tools that enable genome-wide analysis of DNA sequence and structure, copy number, allelic loss, and epigenomic modification. Application of these techniques to elucidation of the nature and timing of these events is enriching our understanding of mechanisms that increase breast cancer susceptibility, enable tumor initiation and progression to metastatic disease, and determine therapeutic response or resistance. These studies also reveal the molecular differences between cancer and normal that ...
Date: June 18, 2009
Creator: Spellman, Paul T.; Heiser, Laura & Gray, Joe W.
Partner: UNT Libraries Government Documents Department

A primer on regression methods for decoding cis-regulatory logic

Description: The rapidly emerging field of systems biology is helping us to understand the molecular determinants of phenotype on a genomic scale [1]. Cis-regulatory elements are major sequence-based determinants of biological processes in cells and tissues [2]. For instance, during transcriptional regulation, transcription factors (TFs) bind to very specific regions on the promoter DNA [2,3] and recruit the basal transcriptional machinery, which ultimately initiates mRNA transcription (Figure 1A). Learning cis-Regulatory Elements from Omics Data A vast amount of work over the past decade has shown that omics data can be used to learn cis-regulatory logic on a genome-wide scale [4-6]--in particular, by integrating sequence data with mRNA expression profiles. The most popular approach has been to identify over-represented motifs in promoters of genes that are coexpressed [4,7,8]. Though widely used, such an approach can be limiting for a variety of reasons. First, the combinatorial nature of gene regulation is difficult to explicitly model in this framework. Moreover, in many applications of this approach, expression data from multiple conditions are necessary to obtain reliable predictions. This can potentially limit the use of this method to only large data sets [9]. Although these methods can be adapted to analyze mRNA expression data from a pair of biological conditions, such comparisons are often confounded by the fact that primary and secondary response genes are clustered together--whereas only the primary response genes are expected to contain the functional motifs [10]. A set of approaches based on regression has been developed to overcome the above limitations [11-32]. These approaches have their foundations in certain biophysical aspects of gene regulation [26,33-35]. That is, the models are motivated by the expected transcriptional response of genes due to the binding of TFs to their promoters. While such methods have gathered popularity in the computational domain, they remain largely obscure ...
Date: March 3, 2009
Creator: Das, Debopriya; Pellegrini, Matteo & Gray, Joe W.
Partner: UNT Libraries Government Documents Department

HER2 signaling pathway activation and response of breast cancer cells to HER2-targeting agents is dependent strongly on the 3D microenvironment

Description: Development of effective and durable breast cancer treatment strategies requires a mechanistic understanding of the influence of the microenvironment on response. Previous work has shown that cellular signaling pathways and cell morphology are dramatically influenced by three-dimensional (3D) cultures as opposed to traditional two-dimensional (2D) monolayers. Here, we compared 2D and 3D culture models to determine the impact of 3D architecture and extracellular matrix (ECM) on HER2 signaling and on the response of HER2-amplified breast cancer cell lines to the HER2-targeting agents Trastuzumab, Pertuzumab and Lapatinib. We show that the response of the HER2-amplified AU565, SKBR3 and HCC1569 cells to these anti-HER2 agents was highly dependent on whether the cells were cultured in 2D monolayer or 3D laminin-rich ECM gels. Inhibition of {beta}1 integrin, a major cell-ECM receptor subunit, significantly increased the sensitivity of the HER2-amplified breast cancer cell lines to the humanized monoclonal antibodies Trastuzumab and Pertuzumab when grown in a 3D environment. Finally, in the absence of inhibitors, 3D cultures had substantial impact on HER2 downstream signaling and induced a switch between PI3K-AKT- and RAS-MAPKpathway activation in all cell lines studied, including cells lacking HER2 amplification and overexpression. Our data provide direct evidence that breast cancer cells are able to rapidly adapt to different environments and signaling cues by activating alternative pathways that regulate proliferation and cell survival, events that may play a significant role in the acquisition of resistance to targeted therapies.
Date: July 27, 2009
Creator: Weigelt, Britta; Lo, Alvin T; Park, Catherine C; Gray, Joe W & Bissell, Mina J
Partner: UNT Libraries Government Documents Department

Quantum Dots-based Reverse Phase Protein Microarray

Description: CdSe nanocrystals, also called quantum dots (Qdots) are a novel class of fluorophores, which have a diameter of a few nanometers and possess high quantum yield, tunable emission wavelength and photostability. They are an attractive alternative to conventional fluorescent dyes. Quantum dots can be silanized to be soluble in aqueous solution under biological conditions, and thus be used in bio-detection. In this study, we established a novel Qdot-based technology platform that can perform accurate and reproducible quantification of protein concentration in a crude cell lysate background. Protein lysates have been spiked with a target protein, and a dilution series of the cell lysate with a dynamic range of three orders of magnitude has been used for this proof-of-concept study. The dilution series has been spotted in microarray format, and protein detection has been achieved with a sensitivity that is at least comparable to standard commercial assays, which are based on horseradish peroxidase (HRP) catalyzed diaminobenzidine (DAB) chromogenesis. The data obtained through the Qdot method has shown a close linear correlation between relative fluorescence unit and relative protein concentration. The Qdot results are in almost complete agreement with data we obtained with the well-established HRP-DAB colorimetric array (R{sup 2} = 0.986). This suggests that Qdots can be used for protein quantification in microarray format, using the platform presented here.
Date: July 15, 2005
Creator: Shingyoji, Masato; Gerion, Daniele; Pinkel, Dan; Gray, Joe W. & Chen, Fanqing
Partner: UNT Libraries Government Documents Department

A Fluid Membrane-Based Soluble Ligand Display System for Live CellAssays

Description: Cell communication modulates numerous biological processes including proliferation, apoptosis, motility, invasion and differentiation. Correspondingly, there has been significant interest in the development of surface display strategies for the presentation of signaling molecules to living cells. This effort has primarily focused on naturally surface-bound ligands, such as extracellular matrix components and cell membranes. Soluble ligands (e.g. growth factors and cytokines) play an important role in intercellular communications, and their display in a surface-bound format would be of great utility in the design of array-based live cell assays. Recently, several cell microarray systems that display cDNA, RNAi, or small molecules in a surface array format were proven to be useful in accelerating high-throughput functional genetic studies and screening therapeutic agents. These surface display methods provide a flexible platform for the systematic, combinatorial investigation of genes and small molecules affecting cellular processes and phenotypes of interest. In an analogous sense, it would be an important advance if one could display soluble signaling ligands in a surface assay format that allows for systematic, patterned presentation of soluble ligands to live cells. Such a technique would make it possible to examine cellular phenotypes of interest in a parallel format with soluble signaling ligands as one of the display parameters. Herein we report a ligand-modified fluid supported lipid bilayer (SLB) assay system that can be used to functionally display soluble ligands to cells in situ (Figure 1A). By displaying soluble ligands on a SLB surface, both solution behavior (the ability to become locally enriched by reaction-diffusion processes) and solid behavior (the ability to control the spatial location of the ligands in an open system) could be combined. The method reported herein benefits from the naturally fluid state of the supported membrane, which allows surface-linked ligands to diffuse freely in two dimensions. Ligands can become reorganized beneath ...
Date: October 14, 2005
Creator: Nam, Jwa-Min; Nair, Pradeep N.; Neve, Richard M.; Gray, Joe W. & Groves, Jay T.
Partner: UNT Libraries Government Documents Department

Restriction of Receptor Movement Alters Cellular Response: Physical Force Sensing by EphA2

Description: Activation of the EphA2 receptor tyrosine kinase by ephrin-A1 ligands presented on apposed cell surfaces plays important roles in development and exhibits poorly understood functional alterations in cancer. We reconstituted this intermembrane signaling geometry between live EphA2-expressing human breast cancer cells and supported membranes displaying laterally mobile ephrin-A1. Receptor-ligand binding, clustering, and subsequent lateral transport within this junction were observed. EphA2 transport can be blocked by physical barriers nanofabricated onto the underlying substrate. This physical reorganization of EphA2 alters the cellular response to ephrin-A1, as observed by changes in cytoskeleton morphology and recruitment of a disintegrin and metalloprotease 10. Quantitative analysis of receptor-ligand spatial organization across a library of 26 mammary epithelial cell lines reveals characteristic differences that strongly correlate with invasion potential. These observations reveal a mechanism for spatio-mechanical regulation of EphA2 signaling pathways.
Date: September 9, 2009
Creator: Salaita, Khalid; Nair, Pradeep M; Petit, Rebecca S; Neve, Richard M; Das, Debopriya; Gray, Joe W et al.
Partner: UNT Libraries Government Documents Department

TRWG developmental pathway for biospecimen-based assessment modalities

Description: The Translational Research Working Group (TRWG) was created as a national initiative to evaluate the current status of NCI's investment in translational research and envision its future. The TRWG conceptualized translational research as a set of six developmental processes or pathways focused on various clinical goals. One of those pathways describes the development of biospecimen-based assays that utilize biomarkers for the detection, diagnosis, prognosis, and assessment of response to cancer treatment. The biospecimen-based assessment modality (BM) pathway was conceived not as comprehensive description of the corresponding real-world processes, but rather as a tool designed to facilitate movement of a candidate assay through the translational process to the point where it can be handed off for definitive clinical testing. This paper introduces the pathway in the context of prior work and discusses key challenges associated with the biomarker development process in light of the pathway.
Date: September 3, 2008
Creator: Group, Translational Research Working; Srivastava, Sudhir; Gray, Joe W.; Reid, Brian J.; Grad, Oren; Greenwood, Addison et al.
Partner: UNT Libraries Government Documents Department

The eukaryotic translation elongation factor eEF1A2 induces neoplastic properties and mediates tumorigenic effects of ZNF217 in precursor cells of human ovarian carcinomas

Description: Ovarian epithelial carcinomas (OEC) frequently exhibit amplifications at the 20q13 locus which is the site of several oncogenes, including the eukaryotic elongation factor EEF1A2 and the transcription factor ZNF217. We reported previously that overexpressed ZNF217 induces neoplastic characteristics in precursor cells of OEC. Unexpectedly, ZNF217, which is a transcriptional repressor, enhanced expression of eEF1A2. In this study, array comparative genomic hybridization, single nucleotide polymorphism and Affymetrix analysis of ZNF217-overexpressing cell lines confirmed consistently increased expression of eEF1A2 but not of other oncogenes, and revealed early changes in EEF1A2 gene copy numbers and increased expression at crisis during immortalization. We defined the influence of eEF1A2 overexpression on immortalized ovarian surface epithelial cells, and investigated interrelationships between effects of ZNF217 and eEF1A2 on cellular phenotypes. Lentivirally induced eEF1A2 overexpression caused delayed crisis, apoptosis resistance and increases in serum-independence, saturation densities, and anchorage independence. siRNA to eEF1A2 reversed apoptosis resistance and reduced anchorage independence in eEF1A2-overexpressing lines. Remarkably, siRNA to eEF1A2 was equally efficient in inhibiting both anchorage independence and resistance to apoptosis conferred by ZNF217 overexpression. Our data define neoplastic properties that are caused by eEF1A2 in nontumorigenic ovarian cancer precursor cells, and suggest that eEF1A2 plays a role in mediating ZNF217-induced neoplastic progression.
Date: April 25, 2008
Creator: Sun, Yu; Wong, Nicholas; Guan, Yinghui; Salamanca, Clara M.; Cheng, Jung Chien; Lee, Jonathan M. et al.
Partner: UNT Libraries Government Documents Department

MORPHOMETRIC SUBTYPING FOR A PANEL OF BREAST CANCER CELL LINES

Description: A panel of cell lines of diverse molecular background offers an improved model system for high-content screening, comparative analysis, and cell systems biology. A computational pipeline has been developed to collect images from cell-based assays, segment individual cells and colonies, represent segmented objects in a multidimensional space, and cluster them for identifying distinct subpopulations. While each segmentation strategy can vary for different imaging assays, representation and subpopulation analysis share a common thread. Application of this pipeline to a library of 41 breast cancer cell lines is demonstrated. These cell lines are grown in 2D and imaged through immunofluorescence microscopy. Subpopulations in this panel are identified and shown to correlate with previous subtyping literature that was derived from transcript data.
Date: May 8, 2009
Creator: Han, Ju; Chang, Hang; Fontenay, Gerald; Wang, Nicholas J.; Gray, Joe W. & Parvin, Bahram
Partner: UNT Libraries Government Documents Department

Amplification of PVT1 contributes to the pathophysiology of ovarian and breast cancer

Description: Purpose. This study was designed to elucidate the role of amplification at 8q24 in the pathophysiology of ovarian and breast cancer since increased copy number at this locus is one of the most frequent genomic abnormalities in these cancers. Experimental Design. To accomplish this, we assessed the association of amplification at 8q24 with outcome in ovarian cancers using FISH to tissue microarrays and measured responses of ovarian and breast cancer cell lines to specific small interfering RNAs (siRNA) against the oncogene, MYC, and a putative noncoding RNA, PVT1, both of which map to 8q24. Results. Amplification of 8q24 was associated with significantly reduced survival duration. In addition, siRNA-mediated reduction in either PVT1 or MYC expression inhibited proliferation in breast and ovarian cancer cell lines in which they were both amplified and over expressed but not in lines in which they were not amplified/over expressed. Inhibition of PVT1 expression also induced a strong apoptotic response in cell lines in which it was over expressed but not in lines in which it was not amplified/over expressed. Inhibition of MYC, on the other hand, did not induce an apoptotic response in cell lines in which MYC was amplified and over expressed. Conclusions. These results suggest that MYC and PVT1 contribute independently to ovarian and breast pathogenesis when over expressed because of genomic abnormalities. They also suggest that PVT1 mediated inhibition of apoptosis may explain why amplification of 8q24 is associated with reduced survival duration in patients treated with agents that act through apoptotic mechanisms.
Date: October 9, 2007
Creator: Guan, Yinghui; Kuo, Wen-Lin; Stilwell, Jackie; Takano, Hirokuni; Lapuk, Anna; Fridlyand, Jane et al.
Partner: UNT Libraries Government Documents Department

Aging Impacts Transcriptome but not Genome of Hormone-dependentBreast Cancers

Description: Age is one of the most important risk factors for human malignancies, including breast cancer; in addition, age-at-diagnosis has been shown to be an independent indicator of breast cancer prognosis. However, except for inherited forms of breast cancer, there is little genetic or epigenetic understanding of the biological basis linking aging with sporadic breast cancer incidence and its clinical behavior.
Date: October 9, 2007
Creator: Yau, Christina; Fedele, Vita; Roydasgupta, Ritu; Fridlyand, Jane; Hubbard, Alan; Gray, Joe W. et al.
Partner: UNT Libraries Government Documents Department

Performance of Molecular Inversion Probes (MIP) in Allele CopyNumber Determination

Description: We have developed a new protocol for using MolecularInversion Probes (MIP) to accurately and specifically measure allele copynumber (ACN). The new protocol provides for significant improvementsincluding the reduction of input DNA (from 2?g) by more than 25 fold (to75ng total genomic DNA), higher overall precision resulting in one orderof magnitude lower false positive rate, and greater dynamic range withaccurate absolute copy number up to 60 copies.
Date: May 14, 2007
Creator: Wang, Yuker; Moorhead, Martin; Karlin-Neumann, George; Wang,Nicolas J.; Ireland, James; Lin, Steven et al.
Partner: UNT Libraries Government Documents Department

Radiosensitivity profiles from a panel of ovarian cancer cell lines exhibiting genetic alterations in p53 and disparate DNA-dependent protein kinase activities

Description: The variability of radiation responses in ovarian tumors and tumor-derived cell lines is poorly understood. Since both DNA repair capacity and p53 status can significantly alter radiation sensitivity, we evaluated these factors along with radiation sensitivity in a panel of sporadic human ovarian carcinoma cell lines. We observed a gradation of radiation sensitivity among these sixteen lines, with a five-fold difference in the LD50 between the most radiosensitive and the most radioresistant cells. The DNA-dependent protein kinase (DNA-PK) is essential for the repair of radiation induced DNA double-strand breaks in human somatic cells. Therefore, we measured gene copy number, expression levels, protein abundance, genomic copy and kinase activity for DNA-PK in all of our cell lines. While there were detectable differences in DNA-PK between the cell lines, there was no clear correlation with any of these differences and radiation sensitivity. In contrast, p53 function as determined by two independent methods, correlated well with radiation sensitivity, indicating p53 mutant ovarian cancer cells are typically radioresistant relative to p53 wild-type lines. These data suggest that the activity of regulatory molecules such as p53 may be better indicators of radiation sensitivity than DNA repair enzymes such as DNAPK in ovarian cancer.
Date: September 7, 2009
Creator: Langland, Gregory T.; Yannone, Steven M.; Langland, Rachel A.; Nakao, Aki; Guan, Yinghui; Long, Sydney B.T. et al.
Partner: UNT Libraries Government Documents Department

The expression level of HJURP has an independent prognostic impact and predicts the sensitivity to radiotherapy in breast cancer

Description: Introduction: HJURP (Holliday Junction Recognition Protein) is a newly discovered gene reported to function at centromeres and to interact with CENPA. However its role in tumor development remains largely unknown. The goal of this study was to investigate the clinical significance of HJURP in breast cancer and its correlation with radiotherapeutic outcome. Methods: We measured HJURP expression level in human breast cancer cell lines and primary breast cancers by Western blot and/or by Affymetrix Microarray; and determined its associations with clinical variables using standard statistical methods. Validation was performed with the use of published microarray data. We assessed cell growth and apoptosis of breast cancer cells after radiation using high-content image analysis. Results: HJURP was expressed at higher level in breast cancer than in normal breast tissue. HJURP mRNA levels were significantly associated with estrogen receptor (ER), progesterone receptor (PR), Scarff-Bloom-Richardson (SBR) grade, age and Ki67 proliferation indices, but not with pathologic stage, ERBB2, tumor size, or lymph node status. Higher HJURP mRNA levels significantly decreased disease-free and overall survival. HJURP mRNA levels predicted the prognosis better than Ki67 proliferation indices. In a multivariate Cox proportional-hazard regression, including clinical variables as covariates, HJURP mRNA levels remained an independent prognostic factor for disease-free and overall survival. In addition HJURP mRNA levels were an independent prognostic factor over molecular subtypes (normal like, luminal, Erbb2 and basal). Poor clinical outcomes among patients with high HJURP expression werevalidated in five additional breast cancer cohorts. Furthermore, the patients with high HJURP levels were much more sensitive to radiotherapy. In vitro studies in breast cancer cell lines showed that cells with high HJURP levels were more sensitive to radiation treatment and had a higher rate of apoptosis than those with low levels. Knock down of HJURP in human breast cancer cells using shRNA reduced the sensitivity ...
Date: June 25, 2010
Creator: Hu, Zhi; Huang, Ge; Sadanandam, Anguraj; Gu, Shenda; Lenburg, Marc E; Pai, Melody et al.
Partner: UNT Libraries Government Documents Department

Overexpression of SnoN/SkiL, amplified at the 3q26.2 locus, in ovarian cancers: A role in ovarian pathogenesis

Description: High-resolution array comparative genomic hybridization of 235 serous epithelial ovarian cancers demonstrated a regional increase at 3q26.2 encompassing SnoN/SkiL, a coregulator of SMAD/TGF{beta} signaling. SnoN RNA transcripts were elevated in {approx}80% of advanced stage serous epithelial ovarian cancers. In both immortalized normal (TIOSE) and ovarian carcinoma cell lines (OVCA), SnoN RNA levels were increased by TGF{beta} stimulation and altered by LY294002 and JNK II inhibitor treatment suggesting that the PI3K and JNK signaling pathways may regulate TGF{beta}-induced increases in SnoN RNA. In TIOSE, SnoN protein levels were reduced 15min post TGF{beta}-stimulation, likely by proteosome-mediated degradation. In contrast, in OVCA, SnoN levels were elevated 3h post-stimulation potentially as a result of inhibition of the proteosome. To elucidate the role of SnoN in ovarian tumorigenesis, we explored the effects of both increasing and decreasing SnoN levels. In both TIOSE and OVCA, SnoN siRNA decreased cell growth between 20 and 50% concurrent with increased p21 levels. In TIOSE, transient expression of SnoN repressed TGF{beta} induction of PAI-1 promoters with little effect on the p21 promoter or resultant cell growth. In contrast to the effects of transient expression, stable expression of SnoN in TIOSE led to growth arrest through induction of senescence. Collectively, these results implicate SnoN levels in multiple roles during ovarian carcinogenesis: promoting cellular proliferation in ovarian cancer cells and as a positive mediator of cell cycle arrest and senescence in non-transformed ovarian epithelial cells.
Date: July 18, 2008
Creator: Nanjundan, Meera; Cheng, Kwai Wa; Zhang, Fan; Lahad, John; Kuo, Wen-Lin; Schmandt, Rosemarie et al.
Partner: UNT Libraries Government Documents Department

Prediction of epigenetically regulated genes in breast cancer cell lines

Description: Methylation of CpG islands within the DNA promoter regions is one mechanism that leads to aberrant gene expression in cancer. In particular, the abnormal methylation of CpG islands may silence associated genes. Therefore, using high-throughput microarrays to measure CpG island methylation will lead to better understanding of tumor pathobiology and progression, while revealing potentially new biomarkers. We have examined a recently developed high-throughput technology for measuring genome-wide methylation patterns called mTACL. Here, we propose a computational pipeline for integrating gene expression and CpG island methylation profles to identify epigenetically regulated genes for a panel of 45 breast cancer cell lines, which is widely used in the Integrative Cancer Biology Program (ICBP). The pipeline (i) reduces the dimensionality of the methylation data, (ii) associates the reduced methylation data with gene expression data, and (iii) ranks methylation-expression associations according to their epigenetic regulation. Dimensionality reduction is performed in two steps: (i) methylation sites are grouped across the genome to identify regions of interest, and (ii) methylation profles are clustered within each region. Associations between the clustered methylation and the gene expression data sets generate candidate matches within a fxed neighborhood around each gene. Finally, the methylation-expression associations are ranked through a logistic regression, and their significance is quantified through permutation analysis. Our two-step dimensionality reduction compressed 90% of the original data, reducing 137,688 methylation sites to 14,505 clusters. Methylation-expression associations produced 18,312 correspondences, which were used to further analyze epigenetic regulation. Logistic regression was used to identify 58 genes from these correspondences that showed a statistically signifcant negative correlation between methylation profles and gene expression in the panel of breast cancer cell lines. Subnetwork enrichment of these genes has identifed 35 common regulators with 6 or more predicted markers. In addition to identifying epigenetically regulated genes, we show evidence of differentially expressed ...
Date: May 4, 2010
Creator: Loss, Leandro A; Sadanandam, Anguraj; Durinck, Steffen; Nautiyal, Shivani; Flaucher, Diane; Carlton, Victoria EH et al.
Partner: UNT Libraries Government Documents Department

In situ quantification of genomic instability in breast cancer progression

Description: Genomic instability is a hallmark of breast and other solid cancers. Presumably caused by critical telomere reduction, GI is responsible for providing the genetic diversity required in the multi-step progression of the disease. We have used multicolor fluorescence in situ hybridization and 3D image analysis to quantify genomic instability cell-by-cell in thick, intact tissue sections of normal breast epithelium, preneoplastic lesions (usual ductal hyperplasia), ductal carcinona is situ or invasive carcinoma of the breast. Our in situ-cell by cell-analysis of genomic instability shows an important increase of genomic instability in the transition from hyperplasia to in situ carcinoma, followed by a reduction of instability in invasive carcinoma. This pattern suggests that the transition from hyperplasia to in situ carcinoma corresponds to telomere crisis and invasive carcinoma is a consequence of telomerase reactivation afertelomere crisis.
Date: May 15, 2003
Creator: Ortiz de Solorzano, Carlos; Chin, Koei; Gray, Joe W. & Lockett, Stephen J.
Partner: UNT Libraries Government Documents Department

The morphologies of breast cancer cell lines in three-dimensionalassays correlate with their profiles of gene expression

Description: 3D cell cultures are rapidly becoming the method of choice for the physiologically relevant modeling of many aspects of non-malignant and malignant cell behavior ex vivo. Nevertheless, only a limited number of distinct cell types have been evaluated in this assay to date. Here we report the first large scale comparison of the transcriptional profiles and 3D cell culture phenotypes of a substantial panel of human breast cancer cell lines. Each cell line adopts a colony morphology of one of four main classes in 3D culture. These morphologies reflect, at least in part, the underlying gene expression profile and protein expression patterns of the cell lines, and distinct morphologies were also associated with tumor cell invasiveness and with cell lines originating from metastases. We further demonstrate that consistent differences in genes encoding signal transduction proteins emerge when even tumor cells are cultured in 3D microenvironments.
Date: January 31, 2007
Creator: Kenny, Paraic A.; Lee, Genee Y.; Myers, Connie A.; Neve, RichardM.; Semeiks, Jeremy R.; Spellman, Paul T. et al.
Partner: UNT Libraries Government Documents Department

Decreased expression of RNA interference machinery, Dicer and Drosha, is associated with poor outcome in ovarian cancer patients

Description: The clinical and functional significance of RNA interference (RNAi) machinery, Dicer and Drosha, in ovarian cancer is not known and was examined. Dicer and Drosha expression was measured in ovarian cancer cell lines (n=8) and invasive epithelial ovarian cancer specimens (n=111) and correlated with clinical outcome. Validation was performed with previously published cohorts of ovarian, breast, and lung cancer patients. Anti-Galectin-3 siRNA and shRNA transfections were used for in vitro functional studies. Dicer and Drosha mRNA and protein levels were decreased in 37% to 63% of ovarian cancer cell lines and in 60% and 51% of human ovarian cancer specimens, respectively. Low Dicer was significantly associated with advanced tumor stage (p=0.007), and low Drosha with suboptimal surgical cytoreduction (p=0.02). Tumors with both high Dicer and Drosha were associated with increased median patient survival (>11 years vs. 2.66 years for other groups; p<0.001). In multivariate analysis, high Dicer (HR=0.48; p=0.02), high-grade histology (HR=2.46; p=0.03), and poor chemoresponse (HR=3.95; p<0.001) were identified as independent predictors of disease-specific survival. Findings of poor clinical outcome with low Dicer expression were validated in separate cohorts of cancer patients. Galectin-3 silencing with siRNA transfection was superior to shRNA in cell lines with low Dicer (78-95% vs. 4-8% compared to non-targeting sequences), and similar in cell lines with high Dicer. Our findings demonstrate the clinical and functional impact of RNAi machinery alterations in ovarian carcinoma and support the use of siRNA constructs that do not require endogenous Dicer and Drosha for therapeutic applications.
Date: May 6, 2008
Creator: Merritt, William M.; Lin, Yvonne G.; Han, Liz Y.; Kamat, Aparna A.; Spannuth, Whitney A.; Schmandt, Rosemarie et al.
Partner: UNT Libraries Government Documents Department

Activity of the kinesin spindle protein inhibitor ispinesib (SB-715992) in models of breast cancer

Description: Ispinesib (SB-715992) is a potent inhibitor of kinesin spindle protein (KSP), a kinesin motor protein essential for the formation of a bipolar mitotic spindle and cell cycle progression through mitosis. Clinical studies of ispinesib have demonstrated a 9% response rate in patients with locally advanced or metastatic breast cancer, and a favorable safety profile without significant neurotoxicities, gastrointestinal toxicities or hair loss. To better understand the potential of ispinesib in the treatment of breast cancer we explored the activity of ispinesib alone and in combination several therapies approved for the treatment of breast cancer. We measured the ispinesib sensitivity and pharmacodynamic response of breast cancer cell lines representative of various subtypes in vitro and as xenografts in vivo, and tested the ability of ispinesib to enhance the anti-tumor activity of approved therapies. In vitro, ispinesib displayed broad anti-proliferative activity against a panel of 53 breast cell-lines. In vivo, ispinesib produced regressions in each of five breast cancer models, and tumor free survivors in three of these models. The effects of ispinesib treatment on pharmacodynamic markers of mitosis and apoptosis were examined in vitro and in vivo, revealing a greater increase in both mitotic and apoptotic markers in the MDA-MB-468 model than in the less sensitive BT-474 model. In vivo, ispinesib enhanced the anti-tumor activity of trastuzumab, lapatinib, doxorubicin, and capecitabine, and exhibited activity comparable to paclitaxel and ixabepilone. These findings support further clinical exploration of KSP inhibitors for the treatment of breast cancer.
Date: June 10, 2009
Creator: Purcell, James W; Davis, Jefferson; Reddy, Mamatha; Martin, Shamra; Samayoa, Kimberly; Vo, Hung et al.
Partner: UNT Libraries Government Documents Department

Comparative analyses of gene copy number and mRNA expression in GBM tumors and GBM xenografts

Description: Development of model systems that recapitulate the molecular heterogeneity observed among glioblastoma multiforme (GBM) tumors will expedite the testing of targeted molecular therapeutic strategies for GBM treatment. In this study, we profiled DNA copy number and mRNA expression in 21 independent GBM tumor lines maintained as subcutaneous xenografts (GBMX), and compared GBMX molecular signatures to those observed in GBM clinical specimens derived from the Cancer Genome Atlas (TCGA). The predominant copy number signature in both tumor groups was defined by chromosome-7 gain/chromosome-10 loss, a poor-prognosis genetic signature. We also observed, at frequencies similar to that detected in TCGA GBM tumors, genomic amplification and overexpression of known GBM oncogenes, such as EGFR, MDM2, CDK6, and MYCN, and novel genes, including NUP107, SLC35E3, MMP1, MMP13, and DDX1. The transcriptional signature of GBMX tumors, which was stable over multiple subcutaneous passages, was defined by overexpression of genes involved in M phase, DNA replication, and chromosome organization (MRC) and was highly similar to the poor-prognosis mitosis and cell-cycle module (MCM) in GBM. Assessment of gene expression in TCGA-derived GBMs revealed overexpression of MRC cancer genes AURKB, BIRC5, CCNB1, CCNB2, CDC2, CDK2, and FOXM1, which form a transcriptional network important for G2/M progression and/or checkpoint activation. Our study supports propagation of GBM tumors as subcutaneous xenografts as a useful approach for sustaining key molecular characteristics of patient tumors, and highlights therapeutic opportunities conferred by this GBMX tumor panel for testing targeted therapeutic strategies for GBM treatment.
Date: April 3, 2009
Creator: Hodgson, J. Graeme; Yeh, Ru-Fang; Ray, Amrita; Wang, Nicholas J.; Smirnov, Ivan; Yu, Mamie et al.
Partner: UNT Libraries Government Documents Department

Basal-subtype and MEK-Pl3K feedback signaling determine susceptibility of breast cancer cells to MEK inhibition

Description: Specific inhibitors of MEK have been developed that efficiently inhibit the oncogenic RAF-MEK-ERK pathway. We employed a systems-based approach to identify breast cancer subtypes particularly susceptible to MEK inhibitors and to understand molecular mechanisms conferring resistance to such compounds. Basal-type breast cancer cells were found to be particularly susceptible to growth-inhibition by small-molecule MEK inhibitors. Activation of the PI3 kinase pathway in response to MEK inhibition through a negative MEK-EGFR-PI3 kinase feedback loop was found to limit efficacy. Interruption of this feedback mechanism by targeting MEK and PI3 kinase produced synergistic effects, including induction of apoptosis and, in some cell lines, cell cycle arrest and protection from apoptosis induced by proapoptotic agents. These findings enhance our understanding of the interconnectivity of oncogenic signal transduction circuits and have implications for the design of future clinical trials of MEK inhibitors in breast cancer by guiding patient selection and suggesting rational combination therapies.
Date: January 23, 2009
Creator: Mirzoeva, Olga K.; Das, Debopriya; Heiser, Laura M.; Bhattacharya, Sanchita; Siwak, Doris; Gendelman, Rina et al.
Partner: UNT Libraries Government Documents Department

A systems analysis of the chemosensitivity of breast cancer cells to the polyamine analogue PG-11047

Description: Polyamines regulate important cellular functions and polyamine dysregulation frequently occurs in cancer. The objective of this study was to use a systems approach to study the relative effects of PG-11047, a polyamine analogue, across breast cancer cells derived from different patients and to identify genetic markers associated with differential cytotoxicity. A panel of 48 breast cell lines that mirror many transcriptional and genomic features present in primary human breast tumours were used to study the antiproliferative activity of PG-11047. Sensitive cell lines were further examined for cell cycle distribution and apoptotic response. Cell line responses, quantified by the GI50 (dose required for 50% relative growth inhibition) were correlated with the omic profiles of the cell lines to identify markers that predict response and cellular functions associated with drug sensitivity. The concentrations of PG-11047 needed to inhibit growth of members of the panel of breast cell lines varied over a wide range, with basal-like cell lines being inhibited at lower concentrations than the luminal cell lines. Sensitive cell lines showed a significant decrease in S phase fraction at doses that produced little apoptosis. Correlation of the GI50 values with the omic profiles of the cell lines identified genomic, transcriptional and proteomic variables associated with response. A 13-gene transcriptional marker set was developed as a predictor of response to PG-11047 that warrants clinical evaluation. Analyses of the pathways, networks and genes associated with response to PG-11047 suggest that response may be influenced by interferon signaling and differential inhibition of aspects of motility and epithelial to mesenchymal transition.
Date: November 14, 2009
Creator: Kuo, Wen-Lin; Das, Debopriya; Ziyad, Safiyyah; Bhattacharya, Sanchita; Gibb, William J.; Heiser, Laura M. et al.
Partner: UNT Libraries Government Documents Department