bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3257: NEDD9 is required for initiation of HER2/Neu-dependent mammary tumor development
Authors: Little Joy L. , Izumchenko Eugene , Singh Mahendra K. , Seo Sachiko , Kurokawa Mineo , Egleston Brian L. , Klein-Szanto Andres , Golemis Erica A.
Published: February 2011

Neural precursor cell-Expressed Developmentally Downregulated gene 9 (NEDD9; HEF1, CAS-L) is a scaffolding protein regulating mitosis, survival, and migration. Within the last 3 years, overexpression of NEDD9 has been implicated in the progression and metastasis of several cancers. Using a Nedd9 knockout strain, we have evaluated the in vivo requirement for NEDD9 in mammary tumorigenesis. Loss of Nedd9 increased the latency of tumor formation in the aggressive MMTV-Polyoma Virus middle T antigen (PyV(m)T) model of mammary tumorigenesis (Izumchenko et al. Cancer Res, 2009). We and others have shown that NEDD9 interacts with and positively regulates activity of key oncogenic signaling proteins such as Aurora-A kinase (AURKA) and Src in cell culture. Tumors harvested from MMTV-PyV(m)T;Nedd9−/− mice had greatly reduced activation of AURKA, SRC, and also FAK and AKT, with the lowest levels of activation correlating with the greatest delays in tumor onset.

To explore the role of NEDD9 in a second highly physiological model of mammary tumorigenesis, Nedd9−/− mice were crossed to MMTV-neu (HER2; ErbB2) transgenic mice, and compared with MMTV-neu;Nedd9+/+ counterparts. Strikingly, after over 18 months, 89% of MMTV-neu;Nedd9+/+ mice developed tumors, in contrast to only 29% of MMTV-neu;Nedd9−/− mice. Between genotypes, there is no statistical difference between tumor growth rates or metastasis. Cell lines derived from tumors of both MMTV-neu;Nedd9+/+ and MMTV-neu;Nedd9−/− animals were then used for xenograft tumor growth and experimental metastasis assays. MMTV-neu;Nedd9−/− tumor-derived cell lines grew similarly to MMTV-neu;Nedd9+/+ counterparts as xenografts in the mammary fat pads of SCID mice, but metastasized to the lungs of SCID mice less readily following tail vein injection. These and other data indicate a novel role for NEDD9 in mammary tumor initiation distinct from the role controlling metastasis and progression of tumors in cooperation with the oncogenes FAK and SRC. Further, the work suggests that HER2-driven mammary tumorigenesis is dependent on NEDD9 at initial stages, but NEDD9 is not required for tumor maintenance, progression, or metastasis from primary organ sites.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3257.
Abstract 3260: Allele-specific tumor spectrum in Pten knockin mice
Authors: Wang Hui , Karikomi Matt , Naidu Shan , Rajmohan Ravi , Caserta Enrico , Chen Hui-Zi , Rawahneh Maysoon , Moffitt Julie , Stephens Julie A. , Fernandez Soledad A. , Weinstein Michael , Perle Krista La , Stromberg Paul , Rosol Thomas J. , Eng Charis , Ostrowski Michael C. , Leone Gustavo
Published: February 2011

Germline mutations in the PTEN tumor suppressor cause Cowden and Bannayan-Riley-Ruvalcaba syndromes, two dominantly inherited disorders characterized by mental retardation, multiple hamartomas and variable cancer risk. Here we modeled three sentinel mutant alleles of PTEN identified in Cowden patients and show that the nonsense PtenΔ4–5 and missense PtenC124R and PtenG129E alleles lacking lipid phosphatase activity cause similar developmental abnormalities but distinct tumor spectra with varying severity and age-of-onset. Allele-specific differences may be accounted by loss-of-function for Pten Δ4-5, hypomorphic function for PtenC124R and gain-of-function for PtenG129E. These data demonstrate that the variable tumor phenotypes observed in Cowden and Bannayan-Riley-Ruvalcaba syndrome patients can be attributed to specific mutations in PTEN that alter protein function through distinct mechanisms.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3260.
Abstract 3265: Immortalization and comprehensive characterization of a human ovarian adenocarcinoma cell line derived from an Ashkenazi Jewish patient
Authors: Rozenblum Ester , Sotelo Jose , Kim Gina Y. , Korolevich Susana , Liao Hongling , Cherry James , Munroe David J. , Roschke Anna
Published: February 2011

We have established and characterized a new ovarian cancer cell line derived from stage IV serous ovarian adenocarcinoma metastatic tissue from a 47 year old Ashkenazi Jewish patient. The patient developed ovarian cancer at age 44 and died of the disease at age 48. The tissue was obtained after an extensive secondary cytoreductive intraperitoneal surgery. Prior to the surgery the patient underwent primary cytoreduction followed by several systemic therapies with recurrences. A culture of the metastatic tissue was transfected with Simian Virus 40 Large T-Antigen and telomerase. The cell line has been in culture on and off during 3 years.

The identity of the cell line was confirmed by Short Tandem Repeat genotyping on genomic DNA extracted from early and late passages as well as from frozen metastatic tissue. The cell line was further characterized by array-Comparative Genomic Hybridization and gene expression microarray; and by Spectral Karyotyping and Fluorescent In Situ Hybridization. The results of these analyses were compared, when appropriate, with those of frozen metastatic tissue. A low level of genetic instability was detected in the cell line, but the karyotypes seem to be near diploid and relatively stable, after several years of cultivation.

We believe that this novel ovarian cancer cell line could contribute to the understanding of the molecular genetic basis of the disease in this advanced stage, and be representative of the tumors of a great number of patients that become resistant to the majority if not all known cancer therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3265.
Abstract 3267: Human population based engineered breast tumor model for in vivo biomarker discovery
Authors: Wu Min , Clark Kimberly , Deng Nanhua , Cai Zuhua , Serpa Megan E. , Zi Tong , Sun Xiaojian , Nicoletti Richard , Sun Kathryn , Feng Bing , Lin Jie , Heyer Joerg , Chiu M Isabel , Robinson Murray O.
Published: February 2011

Human breast cancer development is a complex pathobiological process driven by genetic changes in normal epithelial cells which lead to uncontrollable growth in a permissive microenvironment. Therefore, it is not surprising that tumors from different patients exhibit variable responses to standard of care therapy with unfortunately only a small percentage of patients benefitting from therapy. It has therefore become a priority in oncology and personalized medicine to match patients to drugs that will result in a favorable treatment outcome. In this report, we describe a population based approach for response prediction featuring naturally occurring variation in tumors derived from genetically defined human-in-mouse models of cancer.

De novo human breast tumors were generated by genetically engineering normal primary human breast epithelial cells with HER2 and SV40 early region (HER2/SV40er) or KRAS and SV40 early region (KRAS/SV40er) in an in vivo Human_In_Mouse (HIM) tissue transgenic model. The HER2/SV40er and the KRAS/SV40er HIM tumors develop as human breast adenocarcinoma that are histologically similar to those observed in patients. Also similar to that observed in human tumors, microarray profiling demonstrated significant inter-tumor variation among the established tumors. Moreover, the KRAS/SV40er tumors could be clustered with basal type breast cancers from patients, a poor prognosis human breast cancer subtype. Both HER2/SV40er and KRAS/SV40er tumors exhibited variable responses to treatments with the potent selective triple VEGFR inhibitor, tivozanib. Further characterization of those tumors, both pre- and post- treatment, identified potential biomarkers for tumor response to tivozanib. This population-based approach enables us to identify and validate biomarkers of therapeutic response in an in vivo human tumor model.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3267.
Abstract 3268: Alcohol, obesity and estrogen modulate mammary tumor growth through adiposity and angiogenensis
Authors: Hong Jina , Holcomb Valerie B. , Nunez Nomeli P.
Published: February 2011

Alcohol, obesity and estrogen modulate mammary tumor growth through adiposity and angiogenensis

Jina Hong, Valerie B. Holcomb, and Nomelí P Núñez

Background: Alcohol consumption increases breast cancer risk in women. Postmenopausal women who consume alcohol come in all shapes and sizes, including overweight and obese; some of these women may also take estrogen to prevent the side effects of menopause. It is unclear whether the effects of alcohol on breast cancer development are modified by body weight or exogenous estrogen. In this study, we will determine if the effects alcohol on breast cancer are modified by body weight and exogenous estrogen.

Methods: To determine whether the effects of alcohol on mammary cancer are modified by body weight, we injected subcutaneously Met-1 mouse mammary cancer cells to lean, overweight, and obese female mice consuming water or alcohol. Likewise, to determine if the effects of alcohol on mammary tumorigenesis are modified by exogenous estrogen, we implanted estrogen pellets (0.18mg/day or 0.72mg/day) into ovariectomized female mice; subsequently, we injected Met-1 cancer cells into these mice and then measured tumor growth.

Results: Results show that alcohol-consuming mice were more insulin sensitive than water-consuming mice; also, alcohol-consuming mice developed earlier and larger tumors than water-consuming mice (p<0.05). Our data show that obese mice developed larger tumors than lean mice. Obese mice had higher levels of insulin, IGF-1, and leptin. The only mammary tumor growth factor that was increased by alcohol consumption was leptin (p<0.05). Additionally, alcohol intake increased the pro-angiogenesis factor VEGF (p<0.05).

Conclusion: Results suggest that alcohol consumption, obesity, and estrogen treatment modulate mammary tumorigenesis through adipose tissue and angiogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3268.
Abstract 3270: Environmental stress results in an earlier onset of tumors in a HER2/Neu breast cancer model
Authors: Bovbjerg Dana H. , Henning Jill D. , Gutkin Dmitriy W. , Potter Douglas M. , McCarty Kenneth S. , Jenkins Frank J.
Published: February 2011

Introduction

Possible environmental effects on the development of HER2+ breast cancer have received relatively little research attention. Here we used the well-established MMTV-neu mouse model, which overexpresses mammary ErbB2 (HER2-neu) and stochastically develops primary tumors over 6-12 months, to test the hypothesis that chronic exposure to environmental stressors may result in an earlier onset of tumors.

Experimental Procedures

Female mice (FVB/N-Tg(MMTVneu)202Mul/J) were purchased from Jackson Labs at age 4-5 weeks, housed in standard cages in an isolation chamber in a controlled-access animal room, and randomly assigned to one of four experimental exposure conditions in a factorial design that was maintained for the course of the study: 1) Restraint stress (RS) 90 min - 1x/wk (n=24); 2) Social disruption stress (SD) (via cage mate switching across home cages) 2x/wk (n=23); 3) RS&SD (n=22); 4) home cage control (HC)(n=20). Mice were visually inspected and palpated for tumor development 1x/wk (blinded as to treatment group). All mammary glands from each mouse were processed for histologic evaluation one week following positive tumor palpation. All palpated tumors were histologically confirmed by a pathologist's evaluation of H&E sections (blinded as to treatment group).

Results

Consistent with the study hypothesis, the results indicated that mice chronically exposed to the two environmental stressors (RS&SD) developed primary tumors at a significantly (p<0.002, Cox regression) younger age (mean +/− se: 29.8 +/− 0.7 wk) than the controls (HC) (34.4 +/− 0.8 wk). The mice chronically exposed to just one of the stressors developed tumors at an intermediate age (RS = 32.0 +/− 0.8; SD = 32.3 +/− 0.8 wk), suggesting a possible additive pattern of stress responses. Consistent with stress effects early in the process of tumorigenesis, evidence of mammary hyperplasia in the H&E sections was also significantly (p<0.04) more frequent in mice exposed to both stressors (RS&SD) compared with the control group (HC). Body weights increased significantly over the entire course of the study and did not differ across groups, suggesting that the group differences in tumor development were not due to a failure to thrive.

Conclusions

Exposure to environmental stressors may speed the development of tumors in this transgenic model of HER2+ breast cancer. These novel findings suggest the importance of further investigation to determine the mechanisms responsible for these effects of exposure to environmental stressors.

(Support: CA120795)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3270.
Abstract 3272: Chromosome engineered mouse model for human 8q24 amplicon observed in breast cancer patients
Authors: Tseng Yuen-Yi , Schwertfeger Kathryn L. , Bagchi Anindya
Published: February 2011

Chromosomal abnormalities like deletion, amplification and translocation are hallmarks of almost all the cancers. Amplification of human 8q24, a locus that contains the proto-oncogene C-MYC has been implicated in many human cancers, particularly in solid tumors. Importantly, this amplification is often associated with poor progonosis of breast cancer. Mouse models for studying the effects of c-Myc overexpression have been helpful in elucidating its in neoplastic transformation. However, the mammary tumors arising in these transgenic animals are focal and form with variable latency, strongly suggesting the need for additional genetic lesion to mirror the highly penetrant and aggressive tumors associated with C-MYC amplification. Moreover, in many cancers, C-MYC is co-amplified with the neighboring genetic elements which have also been implicated in tumorigenesis in multiple studies. We argue that overexpression of C-MYC alone is not sufficient to result in the poor prognosis seen in the breast cancer patients with 8q24 amplification and hypothesize that additional genes in the 8q24 amplicon contribute substantially to the mammary tumor initiation and/or aggressive progression of neoplasia. To test this hypothesis, we are modeling this genetic lesion in mice by using chromosome engineering to generate an altered allele containing a 4.5 Mb amplification in mouse chromosome 15 syntenic to Hu 8q24. The tumor susceptibility of these mice are compared with the ones that harbor an amplification of c-myc gene alone. Also, human breast tumors arising from BRCA1 mutations have been reported to have higher incidence of 8q24 amplifications (53%) than are usually found in sporadic breast cancers (10-15%). We hypothesize that a 4.5 Mb amplification of 8q24 has deleterious effects on BRCA1 linked breast cancer, resulting in poor outcome in these patients. To test this hypothesis, we will cross the mice carrying amplification of the 4.5 Mb c-myc locus with brca1 mutant mice. The effects of these amplifications on mammary tumorigenesis will be assessed by scoring the latency of mammary tumors arising in these mice.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3272.
Abstract 3273: Prolactin promotes breast lesion progression and alters tumor histotype
Authors: O'Leary Kathleen A. , Moser Amy R. , Schuler Linda A.
Published: February 2011

Prolactin (PRL) is critical for mammary development and lactation. Epidemiological studies also support a role for PRL in breast cancer (1). In order to study the contributions of PRL to tumor development and progression, we developed a transgenic mouse model that expresses PRL in the mammary epithelial cells (NRL-PRL), mimicking the mammary PRL production in women (2). To explore the interactions of PRL and Wnts in mammary tumorigenesis, NRL-PRL mice were crossed with APCmin/+ mice, which have elevated canonical Wnt signals secondary to reduced β-catenin degradation. Females were administered a single ip dose of ENU (50 mg/kg body weight) at 35-40 days of age. This paradigm in the F1 (FVB/N x C57BL/6J) strain background resulted in few mammary tumors (14%) in NRL-PRL nonparous females. APCmin/+ animals developed tumors at a slightly higher incidence (27%), as previously reported (3). However, penetrance rose to 66% in NRL-PRL/APCmin/+ females, indicating that PRL potentiates APCmin/+-induced tumorigenesis (P=0.008). The presence of PRL also strongly impacted the tumor histotype (P<0.0001). The majority of tumors in APCmin/+ animals were composed of squamous nodules with pilar lesions (63%), while tumors in PRL/ APCmin/+ animals were most often adenocarcinomas of varying histotype (75%). Glands from younger age-matched nonparous females (120 d), prior to palpable lesions, were examined to begin to determine the mechanism of PRL actions. Analyses of “whole mounted” glands showed that both APCmin/+ and PRL/ APCmin/+ animals exhibited multiple epithelial hyperplasias. However, although the numbers of hyperplasias were similar between the two genotypes, the lesions were significantly larger in the PRL/ APCmin/+ glands (P<0.05). This correlated with an increase in the proliferation rate (P<0.01), but no change in apoptosis. Thus, PRL appears to promote tumor progression and affect factors influencing tumor histotype in the APCmin/+ model. The NRL-PRL mouse provides a means of studying how PRL promotes breast cancer alone and in conjunction with other oncogenes.

1Tworoger SS, Hankinson SE. (2008) J Mammary Gland Biol Neoplasia. 13:41-53.

2Arendt LM, Schuler LA. (2008) J Mammary Gland Biol Neoplasia. 13:29-40.

3Moser AR, Hegge LF, Cardiff RD. (2001) Cancer Res. 61: 3480-3485.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3273.
Abstract 3274: Loss of interleukin 10 (IL-10) decreases mouse mammary gland development and lactogenic differentiation
Authors: Masso-Welch Patricia , Merhige Patricia M. , Veeranki Omkara Lakshmi M. , Kuo Shiu-Ming
Published: February 2011

Increased expression of IL-10 has been associated with a decreased breast cancer risk but the mechanism is not clear. The purpose of this study was to examine the effects of IL-10 loss on mouse mammary gland development. Female wild-type and IL-10 knockout mouse littermates born to and raised by IL-10 +/− mothers were maintained under specific pathogen-free (SPF) condition with NIH31 diet. They were sacrificed at days 21, 50-55, 80, and 150 of postnatal age, and at day 2 of lactation. IL-10 loss had no effect on body weight at any age. Whole mounts and paraffin sections of mammary glands from wild type and IL-10 knockout mice were compared for changes in development, differentiation, and the presence of any preneoplastic histologic changes. Interactions between genotype and estrus stage were also examined. Despite its role as an anti-inflammatory immune regulator, IL-10 loss did not result in the enlargement of the intramammary lymph node or frank leukocyte infiltration under SPF condition. No significant differences in mammary gland development were observed in prepubertal glands (day 21). In the pubertal gland at day 50-55 of age, loss of IL-10 was associated with significantly decreased epithelial area, decreased terminal end buds, and decreased adipose tissue. This decrease in peripheral structures and terminal end buds was maintained at day 80 and day 150 of age. Pregnancy and lactogenesis was unable to rescue this phenotype: at day 2 of lactation, the IL-10 knockout mouse maintained a decreased lobular complexity and glandular area, accompanied by abnormal retention of large adipocytes, compared to wild-type mice, consistent with published reports of decreased pup growth in pups born to and raised by IL-10 knock-out mother. Lactating IL-10 knockout mice had a significant increase in abundance of cysts, but no other signs of hyperplasia or neoplasia were present. In conclusion, IL-10 loss decreases epithelial branching in the resting gland, resulting in a compromised differentiation during lactogenesis with decreased glandular area. Although the loss of IL-10 alone was not sufficient to induce frank neoplastic changes in the resting mouse mammary gland, our observation suggests a epithelial differentiation-promoting role of IL-10 which is consistent with its association with a decreased breast cancer risk.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3274.
Abstract 328: Complex oligonucleotide libraries enable high-resolution cytogenetic analysis of human and mouse genomes with fluorescence in situ hybridization (FISH)
Authors: Ach Robert A. , Yamada N. Alice , Tsang Peter , Aleksic Kristina , Begus-Nahrmann Yvonne , Scheffer-Wong Alicia , Laderman Stephen , Lechel Andre , Rudolph K. Lenhard , Speicher Michael , Bruhn Laurakay
Published: February 2011

Cytogenetic analysis of chromosomes using Fluorescence in situ Hybridization (FISH) is a powerful method for the study of chromosome structure, organization, and stability. However, to date, the utility of FISH has largely depended upon access to cloned template DNA used to generate probes. Clones for specific regions are often unavailable, or the genomic template may contain repetitive and other non-informative sequences which can confound FISH analysis. We have leveraged our ability to chemically synthesize oligonucleotides in massively parallel reactions to produce DNA libraries that can be used to generate FISH probes. The sequences of the oligonucleotides in these libraries are selected in silico using empirically determined criteria so as to target only the most informative elements in the genomic region to be analyzed and avoiding any repetitive elements or regions homologous to other non-targeted loci. Our FISH protocol is similar to ones used for FISH with BAC probes. Using our methods, human chromosomal regions as small as 1.8 kb and as large as chromosome arms were successfully visualized in both metaphase and interphase cells. Using our probe design methods, we readily obtained specific and robust signals for chromosomal regions rich in repeats and/or GC content.

We also investigated the utility of these probes for studies of mouse chromosomes by generating probes for over ten loci in the mouse genome. All the probes hybridized well using cytologic and cryosectioned samples. In addition, FFPE samples showed promising preliminary results. We have also applied our FISH methods to develop chromosome-specific enumeration probes for the mouse genome at pericentromeric regions. These enumeration probes should be useful for detecting aneuploidies, polyploidies, and other genomic rearrangements that were previously difficult to study at single-cell resolution, particularly in tissue samples. Successful co-hybridization of chromosome enumeration probes and loci-specific probes in mouse intestinal crypt cells allowed for accurate copy number analyses in tissue samples. We are also continuing to explore other breakthrough applications that are enabled by our oligonucleotide-based FISH methodology.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 328.
Abstract 329: Identification of the breakpoints of two chromosome translocations involving chromosomes 4 and 7 observed in two murine mammary carcinomas
Authors: Pampena María B. , Merani Susana , Lanari Claudia , Fabris Victoria T.
Published: February 2011

The translocation between chromosomes 4 and 7, T(4;7), was observed in two different mammary carcinomas from the medroxyprogesterone acetate (MPA) mouse breast cancer model which have acquired a hormone independent (HI) phenotype (C7-2-HI and C4-HI). We had developed a cell line from the C7-2-HI tumor, the MC7-L2A cells, which retain the chromosome translocation T(4;7), and provide an easy tool to obtain metaphases. G-banding analysis showed that the breakpoint on chromosome 4 is located on the band 4E in the C7-2-HI tumor, whereas in the C4-HI tumor, it is located on the region 4C4-4D3. However, the breakpoint on chromosome 7 is located near the centromere in both tumors. Further experiments using fluorescence in situ hybridization (FISH) in MC7-L2A cells have shown that the breakpoint of the translocation involved the chromosome bands 4E2 and 7A1-A2. The aim of this project is to elucidate whether these breakpoints are involving new gene rearrangements that might participate in the acquisition of hormone independency. In this study we focused in determining if the break in chromosome 7 is located on the same band in both HI tumors. A detailed analysis in MC7-L2A cells using FISH with different DNA probes, spanning the region 7A1-7A2, showed that the breakpoint is located on chromosome 7A1 and not on band 7A2. However, a different picture was observed in metaphases obtained from C4-HI tumor cells, in which the probes corresponding to region 7A1-7B1 hybridized only to normal chromosome 7, but not to the translocation T(4;7). These results indicate that the breakpoints on chromosomes 4 and 7, although involving the same chromosomes, are located on different regions in the T(4;7) translocations, suggesting that they may be random breakpoints. Even though we could not associate this breakpoint with the hormone-independent phenotype, the identification of the sites of chromosome breaks may be relevant to understand the biology of each HI tumor.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 329.
Abstract 3291: Upregulation of Insulin Receptor Substrate-2 (IRS-2) expression by hypoxia
Authors: Mardilovich Katerina , Shaw Leslie M.
Published: February 2011

Insulin Receptor Substrate-2 (IRS-2) belongs to the IRS family of cytoplasmic adaptor proteins that mediate signaling from cell surface receptors, including growth factor, cytokine and integrin receptors. Many of these receptors have been implicated in cancer. Although the IRS proteins are highly homologous in structure and have some complementary functions, growing evidence supports that the IRS proteins have unique roles in cancer. IRS-1 has been shown to promote tumor cell proliferation, while IRS-2 has been positively associated with tumor cell motility and invasion. Previously, our lab demonstrated that Polyoma Middle T-derived Irs-2−/− mouse mammary tumors are significantly less metastatic, and Irs-1−/− tumors significantly more metastatic, than their wild-type counterparts. In recent studies, we observed that IRS-2 expression increases in response to hypoxia in both metastatic mouse mammary tumor cells and poorly differentiated ER-negative metastatic human breast carcinoma cells. In contrast, IRS-1 expression does not increase in response to hypoxia, supporting the notion of their non-overlapping functions. Hypoxia promotes the adaptation and resistance of cancer cells to chemo- and radiation therapy, and also promotes tumor cell survival, invasion and metastasis by selecting for aggressive tumor cells that can survive under stressful low oxygen conditions. We have also shown that IRS-2 upregulation in response to hypoxia promotes tumor cell viability and invasion, selecting for tumor cells with increased metastatic potential. To understand how IRS-2 is regulated by hypoxia, we identified a minimal 200 bp region of the IRS-2 promoter located 700 bp upstream from the translation start site that is hypoxia-responsive. Mutation of an E-box binding site within this region that has been previously implicated in regulating IRS-2 in response to hormone stimulation inhibits hypoxic upregulation of IRS-2. We are currently investigating the factors that bind to this E-box to regulate IRS-2 expression. In addition, we have established that HIF is required for IRS-2 upregulation in hypoxic environments, and we are investigating if HIF acts in a direct or indirect manner to regulate IRS-2 expression. Identifying the mechanism by which IRS-2 expression is regulated by hypoxia is essential for developing novel methods to inhibit IRS-2 expression to suppress metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3291.
Abstract 3295: The type I insulin-like growth factor receptor (IGF1R) pathway driven metastasis signature correlates with poor prognosis and decreased distant metastasis-free survival in human breast cancer
Authors: Sachdev Deepali , Sarver Aaron
Published: February 2011

The insulin-like growth factors (IGFs) acting via the type I IGF receptor (IGF1R) regulate growth and metastasis of multiple types of cancers. Agents targeting IGF1R are currently being tested in phase II/III clinical trials. We have previously shown that IGF1R can regulate metastasis independently of primary tumor growth. In a model of high risk metastatic cancer using the MDA435/LCC6 metastatic cancer cells, we have shown that disruption of IGF signaling either with an antibody against IGF1R, AVE1642, or a dominant negative IGF-1R construct, does not affect growth in the mammary fat pad but inhibits pulmonary metastases and colonization of the lungs. Regulation of such a phenotype will not be measured in clinical trials and these findings imply that clinical trials of this therapeutic strategy may need to adjust their endpoints for assessing benefit by taking into account that inhibition of IGF1R signaling may inhibit metastasis but not primary tumor growth.

Here, we sought to develop an IGF1R driven metastasis signature that may be useful in selecting patients that could benefit from IGF1R targeted therapy and determine if changes in expression of the genes in the signature can be useful in monitoring response. Serum starved LCC6-WT cells (with wild-type functional IGF1R) and LCC6-DN (cells with dominant negative IGF1R) were treated with IGF-I, AVE1642 (antibody against IGF1R that inhibits metastasis but not tumor growth in the mammary fat pad), or AVE1642+ IGF-I for 4 and 24 hours and microarray analysis was performed using the U133 Plus 2.0 human arrays with 47,000 transcripts. Array data were analyzed to determine genes regulated by IGF1R signaling in these cells and if disruption of signaling with AVE1642 affected genes whose expression was regulated by IGF-I signaling. A total of 206 transcripts were regulated by IGF signaling in LCC6-WT cells and AVE1642 treatment reversed the effects of IGF1R signaling. 53 of the most significantly regulated genes were used to query two different human breast tumor datasets to determine if genes regulated by activation of IGF1R are associated with lymph node metastasis and poor prognosis in human tumors. IGF1R regulated metastasis signature genes were expressed in human tumors and were correlated to decreased survival and decreased distant metastasis free survival in breast cancer patients. Our results suggest that presence of IGF1R metastasis signature in patients confers prognostic value and identifying IGF1R driven metastasis signatures may be useful to identify and monitor patients who could benefit from IGF1R targeted therapy for inhibition of metastatic disease.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3295.
Abstract 3296: Cell survival in colon cancer metastatic progression
Authors: Wang Wang , Brattain Michael G.
Published: February 2011

Colon cancer is the second most lethal malignancy in the United States. The majority of deaths are due to metastasis, for which no effective therapies are currently available. While much is known about oncogenic initiation of colon cancer, the mechanisms by which cancer cells survive and metastasize in spite of the cellular stresses associated with this process require further investigation. Resent studies indicated that deregulation of apoptosis contributes to metastasis and deregulation of Bcl-2 family members are correlated with cancer progression. We have initiated studies with a unique othotopic implant model based on closely related colon cancer cell lines. HCT116 and HCT116b cells were isolated from the same primary colon tumor. They both bear the H1047R PIK3CA mutation and form progressively growing invasive orthotopic primary tumors. However, only HCT116 cells exhibit robust metastatic potential. Interestingly, we found that Bcl-2 is highly expressed in HCT116 cells, but not in HCT116b cells. Overexpression of exogenous Bcl-2 in HCT116b cells enhanced their ability to withstand external stress, indicated by reduced levels of DNA fragmentation, reduced cytochrome c release from the mitochondria, and increased caspase activation. Immunoprecipitation assays revealed the formation of Bcl-2: Bax heterodimers, which has been shown to exert the anti-apoptotic property of Bcl-2. In complementary experiments, both shRNA-mediated stable knockdown and overexpression of dominant negative forms of Bcl-2 in metastatic HCT116 cells abolish its survival advantage against stress. These results indicate that modulation of Bcl-2 protein alone can reverse the sensitivity of HCT116 and HCT116b cells to stress-induced apoptosis. Current studies in our orthotopic model will determine whether alterations in Bcl-2 expression as well as complex formation of Bcl-2/Bax affect the metastatic potential of colon cancer in vivo, and consequently provide direct evidence as to whether cell survival control determines metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3296.
Abstract 3304: EAPII/TTRAP is overexpressed in HNSCC
Authors: Li Chunyang , Fan Songqing , Sun Shi-yong , Khuri Fadlo R. , Li Runzhao
Published: February 2011

EAPII/TTRAP, a novel protein we previously identified, was most recently recognized as the first enzyme that removes topoisomerase-mediated adducts at the 5′-phosphotyrosyl bond. EAPII/TTRAP restores 5′phosphate termini at double strand breaks preparing them for ligation. However, the physiological and pathological roles of EAPI I/TTRAP in human cells are elusive; in particular, it is unknown whether EAPII/TTRAP is involved in cancer. To determine the EAPII expression profile in the development of the Head and Neck Squamous Cell Carcinoma (HNSCC), we have developed a monoclonal antibody against EAPII suitable for immunohistochemistry (IHC). IHC was carried out in an HNSCC tissue array containing 34 cases of HNSCC and 12 peri-cancer or normal tissues. EAPII is not detectable in normal human oral squamous cells where HNSCC arises. With increased cell proliferation, weak staining of EAPII in the nucleus occurs. However, EAPII expression is significantly observed in HNSCC: Compared to 8% (1/12) in peri-cancer tissue, 82% (28/34) of HNSCC showed strong cytoplasmic-positive staining while nuclear staining reduced from 75% (9/12) to 35% (12/34). This result indicates that EAPII translocation to the cytoplasm might be an oncogenic factor that contributes to the progression of HNSCC. Consistent with the IHC study, high levels of EAPII expression are observed in many HNSCC cancer cell lines. Furthermore, lentivirus-delivered EAPII small hairpin RNA (shRNA) was employed to knockdown EAPII expression in HSNCC 686Ln-M4E cells. Cell growth curves were analyzed by SRB method. Lower cell growth was observed in the cells with reduced EAPII protein levels compared with cells infected with vector control or scramble shRNA.

Altogether, our data support the hypothesis that EAPII is a potential oncogenic factor and that the elevated level of EAPII may be involved in the HNSCC development. The identification of the role of EAPII in the HNSCC development using animal models will provide further insight into the biological significance of EAPII in cancer development and progression. This work was supported in part by National Cancer Institute award 5 P50 CA128613 SPORE in Head and Neck Cancer (SYS) and Startup Fund from the Department of Hematology and Medical Oncology (RL).

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3304.
Abstract 3307: Cell-surface glycoproteomics identifies CUB-domain containing protein 1 to be associated with prostate cancer metastasis
Authors: Yang Lifang , Troyer Dean A. , Nyalwidhe Julius O. , Drake Richard R. , Semmes Oliver J.
Published: February 2011

Prostate cancer is still the most common and second leading cause of cancer death in men in the United States and is overwhelmingly lethal when metastatic. Evidence from histochemical analysis and experimental tumor models indicates alterations in cell-surface sialylation as a general observation associated with the metastatic potential of tumor cells. Our preliminary data from lectin staining also demonstrates changes of cell-surface sialylation in a syngeneic prostate cancer cell model, which includes two PC3 sublines, N2 (non-metastatic) and ML2 (highly metastatic). Characterization of cell-surface sialylated glycoproteins in this model system could provide new information related to the metastatic phenotype in prostate cancer. Thus, cell-surface sialoglycoproteins in the N2 and ML2 cell lines were metabolicly labeled with with an unnatural mannose analogue (AC4ManNAz); This was followed by chemoselective conjugation of the azido sugars with a biotinylated alkyne capture reagent via Cu (I) catalyzed click chemistry and affinity detection and enrichment by streptavidin-conjugated beads (or fluorescein). Captured glycoproteins were separated by 1D gel electrophoresis, in-gel proteolytic digestion, and protein identification by means of tandem mass spectrometry (LC-MS/MS). The expression of the targeted sialoglycoproteins was verified in our prostate cancer cell lines and clinical samples by means of Western blot, confocal microscopy and fluorescent immunohistochemistry. Forty three cell-surface N-linked glycoproteins were identified as differentially expressed between the PC3 metastasis variants, including CUB-domain containing protein 1 (CDCP1). Western blot analysis of the cell-surface glycoprotein enriched fractions and total cell lysate confirmed that CDCP1 was over-expressed in the metastatic ML2 cells. Expression of CDCP1 is dysregulated in prostate cancer tissues, with malignant glands showing a more intense cell surface staining pattern, whereas normal glands demonstrated weak membrane and intracellular granular staining on the basal side. We conclude that glycan metabolic labeling technology combined with mass spectrometry analysis provides a powerful tool for the identification of specific cell-surface glycoproteins that may contribute to prostate cancer metastasis, like CDCP1. Further studies are warranted to evaluate the clinical value of CDCP1 and to understand its function in prostate cancer progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3307.
Abstract 3309: Identifying glycoproteins in ER+ and ER- human breast cancer tissue samples
Authors: Semaan Suzan , Wang Xu Eddy , Emmett R. , Marshall Alan G. , Sang Qing-Xiang Amy
Published: February 2011

Breast cancer is the second leading cause of death from cancer in women. New biomarkers for cancer detection and treatment are needed to increase the life expectancy of breast cancer patients. Protein glycosylation is a post translational modification that gives a protein a new function. A change in the glycosylation on a glycoprotein often causes a change in function of that glycoprotein and such phenomenon has been correlated with cancerous transformation. Thus, glycoproteins in human breast cancer estrogen receptor positive (ER+) tissue and those in estrogen receptor negative (ER-) tissue, the more aggressive stage in breast cancer, are compared and a list of glycoproteins possibly showing a difference in their glycosylation were identified using double staining (glyco- and total protein staining) of the 2-DE gels followed by ESI-FTICR analysis and gel spot count was done using ImageQuant TL. Results show that 30% of the total lysate proteins are glycoproteins, where the total proteins in ER+ are slightly higher than that of the ER-, yet the difference between the glycoprotein in the ER+ and ER- is not significant. Alpha-1-acid glycoprotein, Alpha-1-antitrypsin glycoprotein, Annexin A1, Malate dehydrogenase mitochondrial precursor, Myosin-reactive immunoglobulin kappa chain variable region, IGKV1-5, Peroxiredoxin-1, and Superoxide dismutase mitochondrial precursor are the identified glycoproteins in both ER+ and ER- human breast tissue samples. Alpha-1-acid glycoprotein and Alpha-1-antitrypsin glycoprotein are mostly N-glycosylated proteins. Annexin A1, Malate dehydrogenase mitochondrial precursor, and Superoxide dismutase mitochondrial precursor are possibly N- or O-glycoslated proteins, while Myosin-reactive immunoglobulin kappa chain variable region, IGKV1-5, and Peroxiredoxin-1 are possibly mostly O-glycosylated proteins. Among those glycoproteins Alpha-1-acid glycoprotein, Alpha-1-antitrypsin glycoprotein, Annexin A1, Peroxiredoxin-1, Immunoglobulin gamma-1C region, and Superoxide dismutase mitochondrial precursor are important glycoproteins in breast cancer and possible biomarkers during the progression of human breast cancer. (This work was supported in part by grant BCTR0504465 from the Susan G. Komen for the Cure Breast Cancer Foundation, grants from the Florida Breast Cancer Coalition Research Foundation, the Elsa U. Pardee Foundation, and the Florida State University to Prof. Q.-X. Sang)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3309.
Abstract 3310: Ezrin is an independent prognostic factor in extremity soft tissue sarcomas
Authors: Carneiro Ana , Jönsson Mats , Rambech Eva , Rissler Per , Rydholm Anders , Nilbert Mef
Published: February 2011

Ezrin is a cytoskeleton protein actively involved in regulating the growth and metastatic capacity of cancer cells. It has recently been reported to be to be associated with poor prognosis high grade soft tissue sarcoma (STS) and to mediate growth and survival through AKT pathway.

To further validate the prognostic value of ezrin in STS, we evaluated 229 extremity primary STS using tissue microarrays and immunohistochemistry. Ezrin expression was correlated with metastasis during follow-up for at least 5 years or until death.185 of the 229 cases analyzed (80%) showed ezrin immunoreactivity in the membrane and cytoplasm of the tumor cells. Multivariate analysis with other known risk factors, including size, vascular invasion and necrosis showed that ezrin expression was significantly associated with development of distant metastasis during follow-up (P = 0.01, 95%CI 1.4-3.4). Ezrin expression was also correlated with AKT pathway changes, evaluated by gene expression.

Our findings suggest that ezrin is a promissing prognostic marker in highly malignant STS and support an interaction between ezrin and AKT in the genesis of STS metastasis.

Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3310.