bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4612: Analysis of transcriptional regulation by progesterone receptor in ER-positive breast cancer cells by ChIP-Seq
Authors: Yang Fan , Bilke Sven , Pineda Marbin , Walker Bob , Zhu Jack , Meltzer Paul
Published: February 2011

Breast cancer is among the most common malignancies in women. It is well established that progesterone receptor (PR) is a major determinant in the development and progression of breast tumor, as well as in diagnosis and treatment of breast cancer. However, it is still not fully understood how PR affects downstream signaling pathways and cross talks with other intracellular signal transduction pathways. To better define the role played by PR in estrogen receptor (ER)-dependent gene regulation, we have used chromatin immunoprecipitation coupled with next generation sequencing (ChIP-Seq) to examine the in vivo DNA binding by PR. Whole-Genome ChIP-Seq analysis has identified large number of PR binding sites in T-47D breast cancer cells, which are ER-positive and PR-positive. PR binding sites were detected in proximal promoter regions as well as distal regions, with most located in non-proximal regions. The comparison of PR and ER binding sites revealed that a significant of portion of the PR binding sites overlaps with the ER binding sites, indicating a close relationship between PR and ER in ER-dependent gene regulation. The PR motifs were found to be among the most enriched sequence motifs within the PR binding sites. Further analysis using the chromosome conformation capture assay revealed that progesterone induces changes to the chromatin structures of known PR-regulated genes. The ChIP-Seq data, together with the expression profiling and other results, indicate that PR may cooperate with ER and play an important role in ER-dependent gene regulation, mammary epithelial differentiation and breast cancer development.

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 4612.
Abstract 4615: Mechanism of inhibition of MMTV-neu, -wnt1 and -Myc induced mammary oncogenesis by RARα agonist Am580
Authors: Farias Eduardo F. , Lu Yongke , Bertran Silvina , Garcia Avalon , Jones Alexis , Samuels Tracey-Ann
Published: February 2011

We tested whether specific activation of a single retinoic acid receptor, RAR alpha (RARα), without direct activation of RAR beta (RARβ) and gamma (RARγ), will inhibit mammary tumor oncogenesis in murine models relevant to human cancer. Fifty MMTV-neu (neu), 30 MMTV-myc (myc), and 50 MMTV-wnt1 (wnt1) transgenic mice were treated with 0.3mg/Kg/day RARα agonist, (retinobenzoic acid, Am580), added to the diet for 40 (neu), 50 (myc) and 35 weeks (wnt1), respectively. Different responses to the treatment were observed in the 3 mouse models, the wnt1 the more responsive, the neu with a mild response and a subset of the myc refractory to the treatment. Among the anti-tumor effects observed in the wnt1 and neu models was the inhibition of epithelial hyperplasia, a significant increase (p<0.05) in tumor-free survival, a reduction in tumor incidence and in growth of established tumors and lung metastasis dissemination (neu model, p<0.001). In both models the mechanisms responsible for these effects involved inhibition of proliferation and survival pathways, and induction of apoptosis. The stronger wnt1 response to Am580 also induced differentiation, both in vivo, and in 3D cultures. In these tumors Am580 inhibited the wnt-pathway, measured by loss of nuclear b-catenin, suggesting partial oncogene-dependence of therapy. Am580 treatment increased RARβ and lowered the level of RARγ, an isotype whose expression we linked to tumor proliferation. Interestingly, the myc mice showed a more complex response in which no significant effect was observed on tumor development but a clear reduction on tumor growth (p<0.001) and lung metastasis dissemination (p<0.01) in a subset of mice (63%). Immunohistological analysis showed scattered necrotic foci, induction of Cyp26A1, E-cadherin, CRBP1 and p27, known RARα response genes, suggesting that cell death, growth inhibition and differentiation are involved in the anti-tumor effect of Am580 in this model. As in the neu and wnt1 models, RARγ was overexpressed in the myc tumors, genetic and pharmacological approaches inhibiting RARγ expression and activity resulted in growth inhibition and differentiation in vitro (2D and 3D cultures). Finally, forced expression of CRBP1 in myc tumor derived cells inhibited tumor growth in xenografts (60%, p<0.05) suggesting that the induction of CRBP1 expression by Am580 can be, in part, responsible for the anti-tumor effect. The anti-cancer effect of RARα, together with the newly discovered pro-proliferative role of RARγ, suggests that specific activation of RARα and inhibition of RARγ might be effective in breast cancer therapy.

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 4615.
Abstract 4616: XR5944 interacts with diverse EREs differently, which may influence ER inhibition by XR5944
Authors: Zhang Zhenjiang , Shu Feng-jue , Chen Xi , Carver Megan , Kallen Caleb , Sidell Neil , Yang Danzhou
Published: February 2011

Since the discovery of the estrogen receptor (ER) as the modulator of estrogen action, ER has evolved to be one of the most effective targets for breast cancer therapy. However, the current ER-targeting therapies have a main limitation related to resistance. Our previous studies have shown that XR5944, a DNA bis-intercalator with potent anticancer activity, is capable of inhibiting the ER activity by its ability to specifically block the binding of ER to its consensus DNA sequence ERE compared to other transcription factor-target DNA interactions. This novel mechanism of ER inhibition by a DNA bis-intercalator may be useful in overcoming resistance to current antiestrogens. The consensus ERE is an inverted repeat comprised of two ERE half-sites separated by three bases: AGGTCAnnnTGACCT where nnn is known as the tri-nucleotide spacer. ER can also bind to non-consensus EREs, i.e. natural EREs that differ from the consensus ERE by one or more base pairs but nevertheless serve as potent transactivation sites for estrogen target genes Therefore, to further understand the mechanism and specificity of the inhibition of ER by XR5944, we tested the specificity of XR5944 to block interactions with consensus and non-consensus ERE sequences by 1D 1H NMR titration studies. Our NMR studies of XR5944-DNA interactions suggested that the consensus ERE spacer plays a significant role in determining the binding characteristics of the complex. Of those tested, the CGG/GCC and AGG/CCT spacers showed the best binding, while the CGT/ACG and TTT/AAA spacers showed the worst binding with XR5944. The binding stoichiometry of XR5944 with ERE sequences appears to be 2:1, which explains why the spacer region can affect the drug-DNA binding. To validate our NMR results, we have also conducted functional studies by using reporter constructs containing EREs with tri-nucleotide spacers CGG and TTT. Results of luciferase reporter assays in MCF-7 cells indicated that XR5944 was significantly more potent in inhibiting the activity of reporters containing ERE-CGG than those containing ERE-TTT, consistent with our NMR results. Together with our NMR studies, these reporter gene experiments indicate that the tri-nucleotide spacer plays a significant role in the binding characteristics and functional consequence of XR5944 interactions with the consensus ERE.

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 4616.
Abstract 462: Targeting hepatocellular carcinoma by suppressing HIF-1 alpha alone and in combination with therapeutic drugs
Authors: Xie Zhi , Wu Xin , Wang Kai , Pocalyko David , Yeom Young-II , Yoo Hyang-Sook
Published: February 2011

Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide and the third most common cause of cancer related death. HIF-1 alpha (HIF1α) is overexpressed in many human cancers as a result of intratumoral hypoxia as well as genetic alterations, such as gain-of-function mutations in oncogenes and loss-of-function mutations in tumour-suppressor genes. It has many important functions including angiogenesis, survival, chemo-resistance metastasis, and glucose metabolism. The overexpression of HIF1α is associated with poor prognosis in many types of cancer. To understand the role of HIF1α in HCC, we analyzed the HIF1α gene expression in the clinical tumor tissues and found that HIF1α mRNA was upregulated compared with the non-tumor adjacent tissue. This upregulation was positively correlated with the poor overall survival and disease free survival of the HCC patients. In addition, we knocked down the HIF1α expression by siRNA and examined its impact on the growth of a panel of HCC cell lines under both hypoxia and normaxia conditions. HIF1α siRNA suppressed 90% of HIF1α mRNA and 50% of its protein expression without affecting the expression of HIF1β and HIF2α. Hep1 and Snu182 cell lines were sensitive to HIF1α suppression (>70%) while Huh 7.5 and SNU398 showed little response. The effect of HIF1α on cell growth inhibition is similar under both hypoxia and normoxia conditions. PDGFR and mTOR were reported to regulate HIF1α synthesis. We examined the efficacy of rapamycin and PDGFR inhibitors in combination with HIF1α siRNA. We found differential response in different HCC cell lines. Rapamycin had limited suppression on both Hep1 and SNU182 cell growth. When combined with HIF1α siRNA, we observed significantly enhanced suppression. PDGFR inhibitor suppressed Hep1 and SNU182 in a dose-dependant manner and the efficacy was further enhanced when combined with HIF1α. To understand the molecular mechanism of HIF1α function, we carried out the gene expression analysis on these “sensitive” and “resistant” cell lines under different treatment conditions. The differential expression signature will provide important insight to the function of HIF1α and the other targeted therapeutic drugs in these HCC cell lines. It will also provide understanding of possible resistance mechanisms and combinational drug strategy to overcome these resistances.

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 462.
Abstract 4625: A biotin-label-based antibody array for high-content profiling of protein expression
Authors: Huang Ruochun , Jiang Weidong , Yang Jean , Mao Ying Qing , Zhang Ying , Yang Weiming , Yang Dongzi , Burkholder Brett , Huang Rani Yan , Huang Ruo-Pan
Published: February 2011

Abstract:

Profiling protein expression on a global scale will have significant impact on biomedical research, particularly in the discovery and development of drugs and biomarkers. Through the years, several antibody array systems have been invented and developed for multiple protein detection. However, a reliable and high-content system for protein profiling from many biological samples has yet developed. To address this problem, we have developed a high density antibody array and used this technology to uncover the potential biomarkers of ovarian cancer. In this approach, biological samples are conjugated with biotin. Antibody arrays are then used to capture the biotinylated proteins. The presence of proteins captured by the antibody chip is detected using streptavidin-conjugated fluorescent dye (Cy3 equivalent) as a reporter. The signals, which are visualized by laser scanning, are normalized using positive, negative, and internal controls. Using this biotin-label-based antibody array technology, the expression levels of 507 human, 308 mouse, 90 rat target proteins can be simultaneously detected, including cytokines, chemokines, adipokines, growth factors, angiogenic factors, proteases, soluble receptors, soluble adhesion molecules, and other proteins in a variety of samples. Most proteins can be detected at pg/ml and ng/ml levels with a CV of less than 20%. Using human biotin-based antibody arrays, we screened the serum expression profiles of 507 proteins in 47 ovarian cancer patients and 39 normal subjects. A panel of protein expression showed significant difference between normal and cancer (P<0.05). By Classification Analysis and Split-Point Score Analysis of these two groups, a small group of proteins can be used to distinguish ovarian cancer patients from normal subjects and between ovarian cancer patients and normal subjects. Our results suggest the biotin label-based antibody arrays that we have developed have great potential in applications for biomarker discovery.

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 4625.
Abstract 4629: Expression of cancer-associated proliferating cell nuclear antigen (caPCNA) in various stages and types of neoplasia (breast, cervix, colon, gastric, lung, ovary, thyroid, and uterus)
Authors: Sandusky George E. , John Karla , Gulley Patty , Carr Katie , Cheng Liang , Malkas Linda H. , Hickey Robert
Published: February 2011

Novel immunohistochemistry biomarkers that target specific cellular proteins that are either uniquely or primarily expressed by cancer cells are being developed. One of these biomarkers, caPCNA, [the cancer associated isoform of PCNA] may be useful as an aide for distinguishing early stage neoplasia from hyperplasia in clinical cases that are often difficult to interpret. The caPCNA isoform appears to be expressed almost exclusively in a large variety of cancer cells and at all Stages [1 to 4] of the disease. It is not consistently observed in most normal adjacent tissues. This study analyzed the expression of caPCNA, by immunohistochemistry (IHC) using an automated Ventana immunostaining platform. In the present study, IHC was performed on neutral buffered formalin fixed paraffin embedded tissues obtained from various pathology labs. Evaluation of carcinomas (120 cases) from various organs and clinical stages as well as normal adjacent tissue was performed. Immunostaining was localized in tumor cells only to the cell nucleus. CaPCNA expression was seen in 10 of 10 cervical Ca, 20 of 20 colon Ca, 10 of 10 thyroid Ca, 10 of 10 gastric Ca, 10 of 10 ovarian Ca, 25 of 25 breast Ca, 10 of 10 lung Ca, 11 of 12 uterine Ca, and 15 of 15 DCIS Ca. On a normal tissue panel, caPCNA immunostaining was seen in macrophages, syncytial trophoblasts in the placenta, GI tract crypt epithelial cells, and scattered staining in the basilar layer of the skin. Immunostaining of normal macrophages and most of the other reactive cell types was abolished by removal of the heavy chain of the primary antibody during creation of an Fab2' fragment of the anti-caPCNA antibody. In summary, the present results illustrate the localization of caPCNA at all stages, in many different types of neoplasms. CaPCNA may therefore be a useful biomarker for early stage cancer diagnosis and potentially maybe a predictor of patient outcome.

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 4629.
Abstract 463: Hypoxia leads to impaired differentiation and sustained global histone acetylation in immortalized mammary epithelial cells cultured as acini
Authors: Vaapil Marica , Helczynska Karolina , Beckman Siv , Påhlman Sven , Jögi Annika
Published: February 2011

We study the effect of hypoxia on mammary epithelial cell differentiation at a non-malignant stage as a putative step in early malignant progression. Due to inadequate vasculature solid tumors often contain regions with low oxygen levels (hypoxia). Hypoxia is associated with increased metastatic potential and worse patient outcome. The hypoxic response is in large driven by the two transcription factors hypoxia inducible factor (HIF) 1 and HIF 2.

We have reported that hypoxia leads to a less differentiated phenotype in breast cancer and in neuroblastoma, and in the clinic low tumor cell differentiation is related to poor prognosis in these diseases. To investigate the effect of hypoxia on mammary epithelial cell differentiation we employed the model of acinar morphogenesis. In this model immortalized mammary epithelial cells in three-dimensional (3D) culture on a laminin-rich matrix differentiate and form acini-like structures with evacuated lumen surrounded by polarized cells. We have previously seen that hypoxia (1% oxygen) impairs the process of acinar morphogenesis. The hypoxic cells failed to obtain polarity and markers of differentiation were affected at expression level as well as cellular localization. Addressing the mechanism behind the hypoxia-impaired differentiation, we find that hypoxia impairs the global histone deacetylation that is associated with changes in cell morphology and differentiation of the mammary epithelial cells. Three days post-seeding, during the proliferation stage, both normoxic and hypoxic cells stained positive for acetylated histone 4 (AcH4). When the normoxic cells go through morphological changes and acinar formation the acetylation of histone 4 is reduced whereas it is maintained in the hypoxic cell-clusters 10 days after seeding. Transfer of acini from normoxia to hypoxia at this time-point leads to an increase in AcH4. Thus, the hypoxic condition prevents global deacetylation, which is generally associated with high level of transcription and an undifferentiated proliferative cell state, rendering the cells an open chromatin state and hindering acinar formation. These data indicate that hypoxia promotes loss of differentiation also at a non-malignant stage.

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 463.
Abstract 4634: Decreased infiltration of macrophage scavenger receptor in non-malignant biopsy specimens is associated with detection of prostate cancer
Authors: Takayama Hitoshi , Hatano Koji , Kawashima Atsunari , Mukai Masatoshi , Nagahara Akira , Nakai Yasutomo , Nakayama Masashi , Nishimura Kazuo , Okuyama Akihiko , Nonomura Norio
Published: February 2011

INTRODUCTION AND OBJECTIVE: Screening for prostate cancer (PCA) has become widespread practice. Many men with negative biopsy findings undergo repeat biopsies to rule out PCA. It should be long enough to avoid unnecessary biopsies and costs. The discovery of new predictors is essential to establish appropriate repeat biopsies. Several predictors of subsequent PCA detection have been described. Especially, chronic inflammation is considered to be a risk factor for PCA. The macrophage scavenger receptor (MSR) has been reported to play a role in recognizing foreign pathogenic substances. MSR-positive inflammatory cells are often seen in solid tumors. Tumor-associated macrophages (TAM) have been shown to positively or negatively regulate the growth of various malignant cells. However, the role of MSR and TAM in PCA carcinogenesis is unclear. In the present study, infiltration of MSR and TAM in non-malignant biopsy specimens was evaluated as a possible predictive value for PCA.

METHODS: These reviews were evaluated for 92 patients ranging from 51 to 82 years (median, 69 years), who received repeat needle biopsies of the prostate and were diagnosed as non-malignant during first biopsy. Of these, 30 (32.6%) were positive for cancer during second or third biopsy. TAM and MSR were immunohistochemically labeled using a monoclonal (CD68 and CD204) antibody. For systematic counting, 6 ocular measuring fields were randomly chosen under a microscope at a power of × 400 within first negative biopsy specimens.

RESULTS: There were no significant differences in PSA, Age and Interval of previous and subsequent biopsies, between negative and positive cases for cancer. Median count of MSR and TAM in each case was 32 and 16. MSR count in patients with cancer was lower than in patients without cancer (P < 0.001). However, in TAM count there were no significant differences between patients with or without cancer. CONCLUSION: Decreased infiltration of MSR in non-malignant biopsy is associated with detection of PCA. MSR is predictor for appropriate repeat biopsies.

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 4634.
Abstract 4637: Mini-chromosome maintenance protein 7 as a potential theranostic biomarker in epithelial ovarian cancer
Authors: Ota Takayo , Clayton Amy C. , Minot Douglas M. , Hartmann Lynn C. , Shridhar Viji , Gilks Blake , Chien Jeremy R.
Published: February 2011

Mini-Chromosome Maintenance protein 7 (MCM7) is involved in replicative licensing and synthesis of DNA. It is previously identified as over-expressed gene in high-grade serous carcinomas compared to serous borderline tumors of the ovary in cDNA microarray studies. In this study, we sought to validate MCM7 expression in ovarian tumors by applying immunohistochemical analysis on tissue microarrays consisting of 445 ovarian tumors. MCM7 expression was quantified as MCM7 labeling index by determining the tumor nuclei positive for MCM7 staining as a percentage of total tumor nuclei. The labeling index scores were independently generated by two methods: PATH score provided by a manual review of each samples by a pathologist and ACIS score provided by Automated Cell Imaging System. Analyses of both scores indicated high degree of concordance in score distribution between PATH and ACIS. MCM7 expression is significantly higher in high-grade serous carcinomas compared to serous borderline tumors, confirming our previous cDNA microarray studies. In both PATH and ACIS analysis, MCM7 expression is also significantly higher in high-grade serous carcinomas compared to other histology subtypes of ovarian cancer. Finally, to identify the clinical significance of MCM7 expression, we determined the association between MCM7 expression and disease-free survival. MCM7 labeling index was dichotomized into two groups (high and low) using median labeling index as cut-off. For both PATH or ACIS-derived scores, univariate analyses indicate significant association of high MCM7 labeling index with better disease-free survival in high-grade serous carcinomas. These results suggest the clinical significance of MCM7 expression in high-grade serous carcinomas of the ovary and the need for further evaluation of MCM7 as a potential theranostic biomarker.

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 4637.
Abstract 4640: High expression of the stem cell marker LGR5 is associated with a poor clinical outcome in patients with Ewing's sarcoma
Authors: Scannell Christopher A. , Abdueva Diana , Ji Lingyun , von Levetzow Cornelia , Sposto Richard , Triche Timothy J. , Lawlor Elizabeth R.
Published: February 2011

Ewing's sarcoma family tumors (ESFT) are aggressive bone and soft tissue tumors that are most commonly diagnosed in childhood and adolescence. Unfortunately, drug-resistant relapse is not uncommon and is associated with very poor outcomes. No clinical or pathologic features currently exist that reliably predict chemosensitivity or relapse in newly diagnosed patients.

In this study we used whole genome expression microarray profiling to identify potential markers of drug resistance and relapse in ESFT.

Affymetrix human exon (HuEx) arrays were used to profile a highly drug resistant case of primary, localized ESFT before and after induction therapy. Comparison of gene summarized data from pre- and post-therapy samples with 10 chemosensitive tumors identified 309 genes to be differentially expressed in the resistant case at diagnosis (fold change>2). Of the 309 differentially expressed genes at diagnosis, 130 demonstrated an even greater change in expression post-induction therapy suggesting that these genes might be markers of the most highly drug-resistant cells. Intriguingly, two stem cell associated genes, PROM1 (prominin 1) and LGR5 (leucine-rich-repeat containing G-protein-coupled receptor 5), were included in the list of putative chemoresistance markers. Expression of PROM1 and LGR5 were 18-fold and 17-fold higher, respectively, at diagnosis and 31-fold and 21-fold higher post-induction therapy in the drug resistant case. To determine if either gene is associated with chemoresistant disease or relapse in ESFT we next evaluated whether their expression correlated with outcome in a large cohort of institutional and Children's Oncology Group patients. Clinical correlates analysis of HuEx gene expression data obtained from 53 tumors revealed no correlation between PROM1 expression and outcome. In contrast, increased expression of LGR5 was associated with worse event-free (p=0.008) and overall survival (p=0.046). Variable but detectable expression of LGR5 in ESFT tumors and cell lines was validated using quantitative RT-PCR. Finally, LGR5 expression in ESFT cells was compared to human embryonic and somatic stem cells and fibroblasts. Significantly, LGR5 was found to be highly expressed by neural crest stem cells but not by bone-marrow derived mesenchymal stem cells, differentiated neural crest cells, or embryonic fibroblasts. These data suggest that LGR5 might be useful as a predictor of outcome in ESFT and implicate a primitive neural crest stem cell phenotype in the developmental origins of drug-resistant and relapsed 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 4640.
Abstract 4641: Distinctive expression of polycomb group proteins Bmi1 and EZH2 in non-small cell lung cancers and their clinical and clinicopathological significance
Authors: Kikuchi Junko , Kinoshita Ichiro , Kikuchi Eiki , Konishi Jun , Shimizu Yasushi , Oizumi Satoshi , Nishimura Masaharu , Dosaka-Akita Hirotoshi
Published: February 2011

Background: The polycomb group genes, Bmi1 and EZH2, function as transcriptional repressors involved in gene silencing. They are involved in the malignant transformation and biological aggressiveness of several human carcinomas. In the present study, we evaluated Bmi1 and EZH2 protein expression in specimens of human non-small cell lung cancers (NSCLCs).

Methods: We conducted an immunohistochemical assessment of 157 surgically resected NSCLCs to evaluate the correlation between Bmi1 and EZH2 expression and various features, including clinical, clinicopathological and biological characteristics.

Results: Normal bronchial epithelia showed abundant expression of Bmi1 and sporadic expression of EZH2. NSCLC patients with high EZH2 expression in tumor cells showed poorer prognosis than those with low EZH2 expression in all pStages (p = 0.001) and in pStage I (p = 0.006). Multivariate analysis showed that high EZH2 expression was a independent unfavorable prognostic factor in pStage I patients (p = 0.048). High EZH2 expression was significantly correlated with non-adenocarcinoma histology (p = 0.001), moderate and poor differentiation (p = 0.001), advanced pT stage (p = 0.02), and high Ki-67 and cyclin E labeling index (p < 0.001). Bmi1 expression, in contrast, was not a significant prognostic factor, and was not correlated with any clinicopathological factors other than early pT stage.

Conclusion: Bmi1 and EZH2 show characteristic and distinctive expression in NSCLCs. High EZH2 expression is correlated with tumor aggressiveness, and may provide a novel prognostic marker for NSCLCs.

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 4641.
Abstract 4647: Clinical Implications of Rabphillin-3A-Like (RPH3AL/NOC2) Gene Alterations in Breast Cancer
Authors: Salih Chura , Jia Xu , Shanmugam Chandrakumar , Katkoori Venkat R. , Callens Tom , Messiaen Ludwine M. , Hameed Omar , Manne Upender
Published: February 2011

Background: The Rabphillin-3A-like (RPH3AL) gene, located at the 17p13.3 locus, has been reported to be involved in progression and metastasis of colorectal cancer. Since there have been no studies of this gene in breast cancer (BC), mutations and loss of heterozygosity (LOH) of RPH3AL in BCs were evaluated.

Methods: BCs (n=104) and matching benign tissues were evaluated for LOH by use of two locus-specific polymorphic markers (D17S1866 and D17S643) and ABI-3100 sequencing technology. Of 104 cases, 16 had corresponding snap-frozen BCs and benign epithelial tissues. The cDNA samples of these frozen cases were analyzed for RPH3AL mutations by RT-PCR and direct sequencing. Since most of the missense mutations were located in the exon-6 region, the genomic DNA samples of 68 (of 104) BCs and their corresponding benign tissues were assessed only for exon 6 by the use of exon-6 specific primers. Additionally, 96 (of 104) BCs and the corresponding control tissues were analyzed for germ line mutations at 43 base pairs downstream from exon-6. Mutations and LOH status were correlated with clinicopathologic features of BCs and patient overall survival by chi-square and Kaplan-Meier analyses, respectively.

Results: LOH analysis demonstrated that 69 of 99 (70%) informative cases exhibited LOH in at least one marker. In total, 19% (13 of 68) of BCs exhibited missense point mutations in the coding region (exon-6) of RPH3AL. Six BCs exhibited mutations at codon 175, two BCs at codon 200, one BC had double mutations at codon 178 and 189, and four BCs had single mutations at codons 165, 174, 181 and 196. There was a germ-line mutation (G>T) at 43 base pairs downstream from exon-6. Genotyping of this alteration showed that 31% (30 of 96) of BCs were homozygous for the G/G allele, 56% (54 of 96) were heterozygous for G/T, and 13% (12 of 96) were homozygous for T/T. In our correlative analysis, most BCs with mutations were associated with lymph-node metastases (9 of 13, 69%) and with an allelic loss (8 of 13, 62%). The G/G alleles were more commonly found in African-Americans than Caucasians (χ2, p=0.003), in ER/PR positive BCs than in ER/PR negative BCs (χ2, p=0.03), and in BCs with advanced stage (χ2, p=0.02). The incidence of LOH increased with nodal metastasis (χ2, p=0.04) and tumor size (χ2, p=0.05) of the BCs. Furthermore, LOH was significantly associated with poor patient survival (log rank, p=0.002).

Conclusions: The results suggest that genetic alterations in the RPH3AL gene are associated with aggressive behavior of breast cancer. (Grant Support: UAB-Breast SPORE-Pilot and Susan G. Komen Breast Cancer Foundation, POP138306).

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 4647.
Abstract 4651: Elevated serum tissue inhibitor of metalloproteinases-1 (TIMP-1) level predicts shorter survival in ovarian cancer
Authors: Jamshidi Navid , Shrivastava Vikas , Leitzel Kim , Ali Suhail , Podczask Edward , Macneill Colin , Deimling Timothy , Brown-Shimer Sheryl , Pierce Karen , Carney Walter , Rizvi Syed , Abbi Kamal , Lipton Allan
Published: February 2011

Body: Background: Matrix metalloproteinases (MMPs) play a key role in invasion and metastasis of cancer cells. MMPs are able to degrade extracellular matrix and aid formation of new blood vessels, thus supporting metastatic tumor growth and angiogenesis. The MMP family can be regulated by their inhibitors, the tissue inhibitor of metalloproteinases (TIMPs), which bind MMPs noncovalently. TIMP-1 has paradoxical multifunctional roles in tumorigenesis, independent of its original MMP-inhibitory role: growth promotion, inhibition of apoptosis, and regulation of angiogenesis. Overexpression of TIMP-1 confers resistance to chemotherapy in cell culture, and increased TIMP-1 has been associated with an unfavorable prognosis in multiple cancer types. TIMP-1 has recently been reported to be a predictive biomarker in large human breast cancer trials. The purpose of this study was to determine the correlation between serum TIMP-1 levels at the time of diagnosis in patients with ovarian cancer and correlate this biomarker with overall survival.

Methods: Serum TIMP-1 was assayed from 36 patients diagnosed with ovarian cancer between 1982 and 1987. Blood was collected at the time of diagnosis and serum frozen. Serum TIMP-1 levels were measured using a TIMP-1 ELISA from Oncogene Science / Siemens Healthcare Diagnostics (Cambridge, MA). Clinical outcome data were correlated with serum TIMP1 levels.

Results: TIMP-1 levels from 36 patients had a mean of 559 ng/ml with SD of + 296 (range 145 to 1460 ng/ml). 35 of 36 patients had clinical follow up data available. 82.14% of patients had distant metastasis, 7.14 % had localized tumor, and 10.71 % had regional involvement. Elevated serum TIMP-1 level predicted shorter survival in ovarian cancer by quartile analysis (p=0.022). Median survival was 4079 days for patients with the lowest serum TIMP-1 quartile, and 106 days for those with the highest serum TIMP-1 quartile. On a continuous basis, higher serum TIMP-1 trended to predict significant shorter survival (p=0.063).

Conclusions: Elevated serum TIMP-1 levels at the time of diagnosis predicted shorter survival in ovarian cancer. Circulating TIMP-1 deserves further study as a predictive and prognostic factor in ovarian cancer.

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 4651.
Abstract 4654: Mitochondrial variants and clinical outcome of oral squamous cell carcinoma in Taiwan
Authors: Lai Chih Husiung , Huang Shiang-Fu , Liao Chun-Ta , Chen I-How , Wang Hung-Ming , Hsieh Ling-Ling
Published: February 2011

Mitochondria are essential cytoplasmic organelles, generating cellular energy in the form of adenosine triphosphate by oxidative phosphorylation. A large volume of studies have indicated that polymorphisms in the mitochondrial genome are associated with risk of various diseases, including cancer. However, only few studies examined the effect of polymorphisms on cancer outcome. To explore the possible role of mitochondrial polymorphisms on clinical outcome of oral squamous cell carcinomas (OSCCs) in Taiwan, 7 common length polymorphism (mtLP) sites and the control D-loop region of mitochondrial genome were genotyped or screened by DNA direct sequencing in 298 OSCCs. One hundred and seventy-eight single-nucleotide polymorphisms (SNPs) were found in the D-loop region and 19 SNPs were presented with a high frequency (> 10%) of variant form. The frequency of variant form in the mtLP sites were quite various ranging from 3.7% to 62.8%. In total, 7 variants from 6 polymorphism sites (2 mtLPs and 4 SNPs) were associated with overall survival. Four subgroups (clusters 1 - 4) were identified by hierarchical clustering of the OSCCs based on the genotypes of these 6 polymorphism sites. We found that the overall survival was significantly different among these 4 OSCC clusters. An index score designated as the total effects of the genotypes of these 6 polymorphism sites on survival for each individual was also developed using the estimates from the Cox model. We found that individual with the higher the index score had the poorer disease-free and overall survival. In summary, the findings provide the first evidence pointing toward mitochondrial variants affect the clinical outcome in OSCC and the underlying mechanisms are worthy to investigate.

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 4654.
Abstract 4655: Gene expression signatures of microsatellite-stable stage III colorectal adenocarcinomas based on the p53 status
Authors: Katkoori Venkat R. , Shanmugam Chandra K. , Chen Dongquan , Bumper Harvey L. , Collens Tom , Messiaen Ludwine , Manne Upender
Published: February 2011

Background: Microsatellite (MS) instability and p53 mutations have been implicated in development, progression and metastasis of colorectal cancer (CRCs). Thus, to identify aggressive tumor subsets, the gene expression profiles of Stage- III MS-stable (MSS), sporadic CRCs were analyzed based on their p53 gene status.

Methods: DNA samples extracted from 136 Stage-III formalin-fixed paraffin embedded CRC tissues were analyzed for MSS and for p53 mutations. Further, mRNA samples extracted from 20 (10 each of p53-mutated and wild-type) corresponding snap-frozen Stage-III-MSS-CRCs were profiled for differential gene expression. Affymetrix Human Genome U133 Plus 2.0 arrays were used for gene expression profiling; and Affymetrix Microarray Suite (MAS 5.0), gene ontology annotation, and KEGG pathways were used for data analysis. Differentially expressed (DE) genes were confirmed by measuring mRNA levels by RT-PCR and by the quantitative nuclease protection assay.

Results: A higher incidence of p53 mutations was found in MSS (65 of 113, 58%) than in MSI (6 of 23, 30%) phenotypes. Kaplan-Meier analyses demonstrated that patients with MSS-CRCs exhibiting p53 mutations had poor overall survival when compared to p53-wt-phenotypes (log rank, p=0.031). Gene expression analyses identified 116 DE genes (73 down-regulated and 43 up-regulated). All of these passed the stringent background filter (intensity p-value ≤0.001 and intensity ≥3 SD above mean background) and DE ratio (χ2 p-value ≤0.01 & fold change ≥2.0). GBP1, PSMB9, TRIM29, BST2 and TFF1, that have prognostic value in CRCs, were up-regulated in p53-mutated-MSS-CRCs but down-regulated in p53-wt-MSS-CRCs. Further, genes that regulate cell-cycle arrest (CDKN1B) and tumor growth suppression (ACVR1B, SESN1, WFDC1 and IRF7) were down-regulated in p53-mutated-MSS-CRCs, but up-regulated in p53-wt-MSS-CRCs. The genes presented here are the top-ranked (top-5) genes in their respective groups. The mRNA expression levels of these DE genes in all 136 CRCs were confirmed on two different platforms.

Conclusion: Our findings suggest that mutant- and wt-p53 Stage-III-MSS-CRCs have distinct gene expression profiles and appear to be useful in identifying aggressive Stage III subsets and for the assessment of clinical outcomes. These studies are supported by grants from the National Institute of Health/National Cancer Institute (U54-CA118948, RO1-CA98932, and R03CA139629).

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 4655.
Abstract 4656: DNA repair polymorphisms as predictors of Graft-versus-Host Disease after allogeneic hematopoietic cell transplants
Authors: Thyagarajan Bharat , Lindgren Bruce , Basu Saonli , Nagaraj Sriharsha , Gross Myron , Wesidorf Daniel , Arora Mukta
Published: February 2011

Graft versus host disease (GVHD) remains a major source of morbidity and mortality among patients undergoing hematopoietic cell transplants (HCT). Tissue damage as a result of treatment is the initiating event in the pathogenesis of GVHD and variations in DNA repair can influence the amount of tissue damage in response to treatment modalities used in HCT. Since DNA damage caused by therapeutic agents such as alkylating agents and ionizing radiation used frequently in pre- HCT conditioning regimens is commonly repaired by the base excision repair (BER) pathway we hypothesized that genetic polymorphisms in the BER pathway will be associated with GVHD after HCT. We evaluated the association between single nucleotide polymorphisms (SNPs) (n= 179) in the BER pathway with acute GVHD and chronic GVHD in a cohort of 470 recipients who underwent allogeneic HCT for treatment of hematologic malignancies at the University of Minnesota. After adjustment for donor type, diagnosis, disease status at transplant and gender mismatch, two SNPs in RFC1 (Replication Factor C) gene (rs1057807 and rs4975003) were associated with a decreased risk of grade II-IV acute GVHD (HR:0.74-0.77; p≤0.01) and one SNP, rs6844176, in RFC1 gene was associated with increased risk of grade II-IV acute GVHD (HR:1.39, p=0.001). The rs3730914 SNP in the LIG1 (Ligase I) gene was also associated with a decreased risk of grade III-IV acute GVHD (HR: 0.38; p=0.005). One SNP, rs1805410, in the PARP1 (poly ADP-ribose polymerase 1) gene was associated with increased risk of chronic GVHD (HR: 1.44; p=0.01) post HCT. These findings suggest that SNPs in the BER pathway can be used as genetic biomarkers to predict individuals at high risk for GVHD. If these findings are confirmed, modulation of pre-transplant conditioning may alter risk in these patients.

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 4656.
Abstract 466: Does body weight influence tumor-infiltrating B cells in murine pancreatic cancer
Authors: White Patrick B. , Dalbec Kathryn M. , True Eben M. , Wang Sue S. , Swartz-Basile Deborah A. , Pitt Henry A. , Zyromski Nicholas J.
Published: February 2011

Background: We have developed a murine model of pancreatic cancer in congenitally obese (LepOb and LepDb) mice where tumors grow larger, metastasize, and decrease survival. Obese mice also had altered splenic T and B cells as well as downregulation of B cell and immunoglobulin genes and reduced numbers of B cells in the tumors. The question remained, however, whether alterations in tumor-infiltrating lymphocytes were a reflection of body weight or leptin metabolism. Therefore, we designed an experiment to determine whether diet-induced weight gain would alter systemic and tumor-infiltrating lymphocytes.

Methods: Thirty lean C57 female mice were fed either a control 10% fat (n=10) or a 60% fat diet (n=20) starting at 6 weeks of age. At 11 weeks, all mice were inoculated with 2.5x105 PAN02 murine pancreatic cancer cells. After 6 weeks tumors that developed in 18 mice, the spleen (n=12), and the sera were harvested. Mice were categorized as Lean or Overweight if they weighed less or more than the mean final weight of the 60% fat animals (23.1g). Tumors in Lean (n=10) and Overweight (n=8) mice were sectioned and stained for B and T cells. Two observers blinded to animal weight graded cell density on a scale of 0-4 in 10 high-powered fields. Flow cytometry was performed on the spleen from Lean and Overweight mice for B, T, activated T and B (ACT), natural killer (NK) cells, and macrophages (MACRO). Serum leptin was measured. ANOVA and Student's t-test were applied.

Results: Tumors were larger (1.33 vs. 0.54g, p=0.03) and leptin levels were higher (3.1 vs. 1.4ng/ml, p=0.05) in Overweight mice. Mean tumor-infiltrating lymphocyte (TIL) scores and spleen lymphocyte data are presented in the table.

Conclusion: The data suggest that 1) both B and T cells are present in the pancreatic cancer microenvironment and 2) TIL and splenic lymphocyte populations do not differ between lean and overweight mice. We conclude that obesity, as opposed to overweight, is required to alter systemic and tumor microenvironment lymphocyte populations. B Cell TILT Cell TILB Cell (%) SpleenT Cell (%) SpleenACT (%) SpleenNK (%) SpleenMACRO (%) SpleenLean0.7±0.12.2±0.264±244±32.3±0.72.3±0.71.6±0.2Overweight0.7±0.12.0±0.258±139±51.5±0.31.4±0.41.4±0.3

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 466.
Abstract 4661: Decreased epidermal growth factor receptor plasma levels as an indicator for head and neck squamous cell carcinoma case status and disease progression
Authors: Egloff Ann Marie , Kelly Lori , Panelli Monica , Davis Autumn Gaither , Siegfried Jill M. , Grandis Jennifer R. , Argiris Athanassios
Published: February 2011

The epidermal growth factor receptor (EGFR), c-Met and G-protein-coupled receptor signaling pathways contribute to head and neck squamous cell carcinoma (HNSCC) growth. To assess these pathways in HNSCC patients, we measured plasma levels of EGFR, the EGFR ligands transforming growth factor alpha (TGF-α) and amphiregulin (AR), the c-Met ligand hepatocyte growth factor (HGF) and COX-2, a component of prostaglandin production, in HNSCC patients (n=22) and cancer-free controls (n=32) enrolled through an Early Detection Research Network- (EDRN-) sponsored effort. Analytes were measured using a multiplex enzyme-linked immunosorbent assay. Differences in analyte levels between cases and controls and the prognostic potential of each analyte were evaluated.

EGFR plasma levels were significantly decreased (P=0.04, rank sum test (RST)) and HGF plasma levels significantly increased (P<0.01, RST) in HNSCC cases compared to cancer-free controls. TGF-α levels tended to be higher in HNSCC cases (P=0.07, RST) while AR and COX-2 levels did not differ between cases and controls (P=0.32 and 0.32, respectively RST). Area under the Receiver Operating Characteristic (ROC) curve for HGF levels in subjects with low EGFR plasma levels was 0.92. EGFR plasma levels were significantly reduced in HNSCC cases who were former or active smokers compared to never smokers (P = 0.009, Kruskal-Wallis test (KWT)), but no difference in EGFR plasma levels by smoking status was observed among cancer-free controls (P = 0.97, KWT). HGF plasma levels tended to be lower in HNSCC cases who were never smokers (P=0.06 KWT) but did not differ by smoking status among cancer-free controls (P=0.16, KWT). Low EGFR plasma levels were associated with significantly reduced progression-free survival (PFS) in EDRN HNSCC cases (P = 0.03, log rank test); no other analyte tested was associated with PFS. We noted a negative correlation between EGFR plasma levels in EDRN HNSCC cases and previously reported EGFR tumor levels assessed by immunohistochemical staining that was not significant (Spearman's rho=-0.41, P=0.10, n=17).

Plasma EGFR and HGF levels may provide indications of HNSCC cancer status, and EGFR plasma levels may be a valuable prognostic indicator for patients with HNSCC. Decreased plasma EGFR levels in HNSCC patients were likely not due to general effects related to smoking and instead may reflect tissue characteristics specific to tobacco-related cancers. We found EGFR plasma levels in HNSCC patients were not positively correlated with EGFR tumor levels.

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 4661.