bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract LB-128: The N-terminus of the ASPP2 tumor suppressor binds Ras and enhances Ras/Raf/MEK activation to promote oncogene-induced cellular senescence
Authors: Wang Zhiping , Stork Philip J. , Lopez Charles D.
Published: June 2012

The ASPP2 tumor suppressor is a pro-apoptotic member of a family of p53 binding proteins. It functions in part by selectively enhancing p53 transcriptional activation of pro-apoptotic target genes. Although the ASPP2 C-terminal domain binds the p53 core domain, little is known about the physical and functional interactions of other proteins to the highly structured ASPP2 N-terminus. Moreover, mounting evidence from our lab and others demonstrate that ASPP2 also mediates non-apoptotic p53-independent functions. However, the molecular mechanisms remain poorly understood. Previous reports have found that ASPP2 can localize to the cytoplasm and have identified a potential Ras-association domain in the ASPP2 N-terminus. Therefore, we wondered if novel ASPP2 functions could be mediated by ASPP2 N-terminal interaction with cytoplasmic Ras. Using co-immunoprecipitation and immunofluorescence microscopy, we identified ASPP2 in a complex with active Ras on the plasma membrane. Moreover, we also found that this interaction required the ASPP2 N-terminus since BBP, a naturally occurring alternatively spliced ASPP2 isoform lacking the N-terminus, showed markedly reduced ability to bind Ras. To determine if ASPP2 could modulate Ras function, we ectopically expressed ASPP2 in cells and found enhancement of Ras activation and subsequent stimulation of ERK phosphorylation. Conversely, ASPP2 knockdown by siRNA drastically reduced ERK phosphorylation and activation by EGF. Although it is oncogenic in tumor cells, constitutive Ras activation in normal cells leads to premature senescence in a Raf-MEK-ERK activation dependent manner. To explore if ASPP2 enhanced senescence, we knocked down ASPP2 expression with siRNA and found that senescence induced by RasV12 lentiviral infection in normal human fibroblasts was significantly attenuated. This was similar to the attenuation of senescence achieved by the MEK inhibitor PD98059. Together, our results demonstrate a novel function of the ASPP2 N-terminus in promoting cellular senescence via Ras-interaction and stimulation of Raf-MEK-ERK activity. These findings suggest a novel mechanism by which ASPP2-induced senescence may mediate its tumor suppression ability, and opens new avenues for investigation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-128. doi:10.1158/1538-7445.AM2011-LB-128
Abstract LB-16: Inhibition of RalA as a novel strategy for targeting ovarian cancer
Authors: Farassati Faris
Published: June 2012

Abstract: The Ral (Ras like) GTPase guanyl nucleotide-binding proteins, RalA and RalB are direct effectors of pro-oncogene Ras. RalA and RalB are implicated in tumorigenesis and play distinct roles in mediating carcinogenesis. The critical role of RalA in the initiation and progression of ovarian cancer is entirely unclear. Here, we report that RalA is overactivated in human ovarian cancer cells and human ovarian cancer tissues. The RalA downstream effectors (RalBP-1 and CDC42) and the regulators (GalGDS, PP2A Aα and β, as well as Aurora Kinase) that modulate RalA activation during the post-translational process were found to be de-regulated in ovarian cancer cells. ShRNA-mediated knockdown of RalA inhibited ovarian cancer cell proliferation and invasion. Geranyl-Geranyl transferase inhibitors (GGTI-2147) or Aurora Kinase Inhibitor (AKI) that inhibit RalA activation at post-translational levels suppressed ovarian cancer cell growth and invasion in-vitro. Additionally, subcutaneous (SC) mouse model with these two inhibitors also showed effective inhibition of tumor growth in-vivo. Therefore, RalA is a key player in the biology and dispersal of ovarian cancer cells and is a promising target for the development of novel therapeutics. To the best of our knowledge, the role of RalA in ovarian cancer tumorigenesis has not been documented before, therefore our strategy in targeting RalA for treatment of ovarian cancer merits highly in terms of novelty.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-16. doi:10.1158/1538-7445.AM2011-LB-16
Abstract LB-160: Dual application of TLR5 agonist in cancer treatment: A single agent combines supporting care and antitumor activities
Authors: Burdelya Lyudmila , Gleiberman Anatoly , Gitlin Ilya , Aygun-Sunar Semra , Rustum Youcef , Gudkov Andrei V.
Published: June 2012

We have previously shown the strong radioprotective properties of TLR5 agonist flagellin in both mouse and non-human primate models (Burdelya et al., 2008, Science 320: 226–230). A pharmacologically optimized flagellin derivative, Protectan CBLB502, is currently at an advanced stage of development as a prospective radiation antidote for biodefense applications. By testing CBLB502 in combination with radiation treatment of experimental mouse tumors in vivo, we demonstrated that its tissue protection properties are limited to normal tissues with no tumor protection detected in any of numerous mouse tumor models. For example, injections of CBLB502 significantly reduced the severity of mucositis and dermatitis associated with local fractioned irradiation of the head and neck area of mice with syngeneic orthotopically grown head and neck carcinoma with no reduction of antitumor effect of experimental radiotherapy. On the contrary, use of CBLB502 led to enhanced tumor suppression by radiation and produced a detectable tumor suppressive effect even without irradiation. Similar results were obtained in s.c. grown syngeneic colon Wart tumors in rats. In this model, administration of CBLB502 reduced the severity of irinotecan-induced diarrhea, but did not interfere with the anti-tumor effect of the drug. Moreover, use of CBLB502 as a single agent had a pronounced antitumor effect in the majority of Wart tumor-carrying rats. Comparison of the effect of CBLB502 on in vivo growth of isogenic pairs of tumor cell lines differing in their TLR5 status showed that the antitumor effect of the TLR5 agonist is TLR5 dependent and is associated with tumor infiltration by immunocytes, presumably attracted following activation of TLR5 signaling in the tumor cells. Remarkably, CBLB502 caused an immunotherapeutic effect in TLR5-negative tumors growing as experimental metastases in the liver which was found to be the primary target of TLR5 agonist. The liver responds to CBLB502 injection by massive and rapid activation of NF-kB in all hepatocytes. This results in dramatic changes in the content of secreted factors, attraction of immunocytes and a strong reversible increase in liver resistance to a variety of toxic agents. Based on these results, we project clinical applications of CBLB502 as a bi-functional therapeutic agent with supporting care (protection of normal tissues from radio- and chemotherapy side effects) and immunotherapeutic activities.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-160. doi:10.1158/1538-7445.AM2011-LB-160
Abstract LB-226: Celecoxib synergizes human pancreatic ductal adenocarcinoma cells to sorafenib-induced growth inhibition involving p21 suppression
Authors: Rosendahl Ann H. , Gundewar Chinmay , Said Katarzyna , Andersson Roland
Published: June 2012

Purpose: The aim of this study was to evaluate the potential for combining the multikinase inhibitor sorafenib and the specific cyclo-oxygenase 2 (COX-2) inhibitor celecoxib as therapy in pancreatic adenocarcinoma cells and to test the hypothesis that a synergistic or additive effect on the Ras/MAPK/ERK and related pathways might be obtained.

Experimental design: COX-2 positive (BxPC-1) and low/negative (MIAPaCa-2, PANC-1 and AsPC-1) human pancreatic adenocarcinoma cells were exposed to sorafenib and celecoxib combined treatment in vitro, after which cell viability and various growth promoting and survival signaling pathways were monitored by MTT, flow cytometry and Western blotting.

Results: Combined treatment with sorafenib and celecoxib resulted in synergistic inhibition of pancreatic adenocarcinoma cell proliferation through COX-2 independent mechanisms. This regimen produced combination index (CI) values between 0.67 and 0.92 for the various cell lines, suggesting significant synergistic interactions between sorafenib and celecoxib, which also markedly inhibited the migratory capacity. The growth inhibition was associated with a reduction of basal ERK1/2 activity, accumulation of cells in the G0/G1 phase of the cell cycle, induction of apoptosis and poly(ADP-ribose) polymerase cleavage. These changes were accompanied by a significant reduction of p21WAF1/Cip1 levels, where celecoxib sensitized the cells to sorafenib-mediated p21WAF1/Cip1 suppression.

Conclusion: p21WAF1/Cip1 levels are elevated frequently in human pancreatic adenocarcinoma and increases with disease progression. In this study we demonstrate that combined treatment with sorafenib and celecoxib synergistically induce growth inhibition and apoptosis through a process involving phospho-ERK1/2 and p21WAF1/Cip1 suppression. These data suggest the combined treatment of sorafenib and celecoxib as potential therapy for clinical testing in patients with pancreatic adenocarcinoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-226. doi:10.1158/1538-7445.AM2011-LB-226
Abstract LB-26: A mathematical methodology for determining the temporal order of pathway alterations in cancer
Authors: Cheng Yu-Kang , Beroukhim Rameen , Levine Ross L. , Mellinghoff Ingo K. , Holland Eric C. , Michor Franziska
Published: June 2012

Human cancer is caused by the accumulation of genetic alterations in cells. Of special importance are changes that occur early during malignant transformation because they may result in oncogene addiction and represent promising targets for therapeutic intervention. We have previously described a computational approach, called Retracing the Evolutionary Steps in Cancer (RESIC), to determine the temporal sequence of genetic alterations during tumorigenesis from cross-sectional genomic data of tumors at their fully transformed stage (1). This approach was based on data of alterations in individual genes. Since specific mutational events may not be necessary to cause an oncogenic phenotype and alterations within the set of genes belonging to a particular signaling pathway may have similar or equivalent effects, we apply a pathway-based systems biology approach to the RESIC methodology. This methodology is used to determine whether alterations in specific pathways develop early or late in malignant transformation. When applied to glioblastoma multiforme copy number data from The Cancer Genome Atlas project (2), we find a consistent temporal order of common genetic alterations and the pathways to which the genes belong. These results are also consistent with experimental results that high level EGFR amplification occurs late during gliomagenesis (3–4). This extension to the RESIC methodology provides an evolutionary mathematical approach to identify the temporal sequence of pathway changes driving tumorigenesis and may be useful in guiding the understanding of signaling rearrangements in cancer development.

1. Attolini CS-O, et al. (2010) A mathematical framework to determine the temporal sequence of somatic genetic events in cancer. Proceedings of the National Academy of Sciences 107:17604–17609.

2. TCGA (2008) Comprehensive genomic characterization defines human glioblastoma genes and core pathways. Nature 455(7216):1061–1068.

3. Bachoo RM, et al. (2002) Epidermal growth factor receptor and Ink4a/Arf: convergent mechanisms governing terminal differentiation and transformation along the neural stem cell to astrocyte axis. Cancer Cell 1(3):269–277.

4. Zhu H, et al. (2009) Oncogenic EGFR signaling cooperates with loss of tumor suppressor gene functions in gliomagenesis. Proc Natl Acad Sci U S A 106(8):2712–2716.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-26. doi:10.1158/1538-7445.AM2011-LB-26
Abstract LB-261: Maintenance of aerobic glycolysis provides selective protection from apoptosis upon loss of growth signals or inhibition of BCR-Abl through suppression of p53 activation
Authors: Mason Emily F. , Zhao Yuxing , Goraksha-Hicks Pankuri , Coloff Jonathan L. , Gannon Hugh , Jones Stephen N. , Rathmell Jeffrey C.
Published: June 2012

Healthy cells require input from growth factor signaling pathways to maintain cell metabolism and survival. In contrast, cancer cells can maintain growth factor-independent glycolysis and survival through expression of oncogenic kinases, such as BCR-Abl. While disruption of these growth signals through targeted kinase inhibition can promote cancer cell death, therapeutic resistance frequently develops, and further mechanistic understanding is needed. Cell metabolism may play a central role in this cell death pathway, as we have shown that growth factor deprivation leads to decreased glycolysis that promotes apoptosis via p53 activation and induction of the pro-apoptotic protein Puma. However, it has remained unclear how cell metabolism regulates p53 activation. To investigate this, we have utilized a system in which stable overexpression of the glucose transporter Glut1 and hexokinase 1 in hematopoietic cells drives growth factor-independent glycolysis. This system allows us to examine the effects of altered glucose metabolism in the absence of other signaling events activated downstream of growth factor receptors or oncogenic kinases. Here, we extend our findings to demonstrate that elevated glucose metabolism, characteristic of cancer cells, can suppress Protein Kinase C delta-dependent p53 activation to maintain cell survival after growth factor withdrawal. In contrast, DNA damage-induced p53 activation was independent of Protein Kinase C delta and was not metabolically sensitive. Both stresses required phosphorylation of p53 at serine 18 for maximal activity, but each led to a unique pattern of p53 target gene expression, demonstrating distinct activation and response pathways for p53 that were differentially regulated by metabolism. Consistent with oncogenic kinases acting to replace growth factors, treatment of BCR-Abl-expressing cells with the kinase inhibitor imatinib led to reduced glycolysis and p53- and Puma-dependent cell death. Accordingly, maintenance of glucose metabolism inhibited p53 activation and promoted imatinib resistance. Furthermore, inhibition of glycolysis enhanced imatinib sensitivity in BCR-Abl-positive cells expressing wild type p53 but had little effect on p53-null cells. These data demonstrate that distinct pathways regulate p53 after DNA damage and metabolic stress and that inhibition of glucose metabolism may enhance the efficacy of and overcome resistance to targeted molecular cancer therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-261. doi:10.1158/1538-7445.AM2011-LB-261
Abstract LB-276: Fusion transcripts in breast cancer cell lines and tumors
Authors: Thompson E. Aubrey , Asmann Yan W. , Necela Brian M. , Kalari Krishna R. , Williamson David W. , Carr Jennifer M. , Baker Tiffany R. , Schroth Gary P. , Kocher Jean-Pierre A. , Perez Edith A.
Published: June 2012

The role of fusion gene transcripts such as BRC-ABL1 has been long appreciated in hematopoietic malignancies. Recent evidence from next generation sequencing projects has suggested that gene fusion events and the resultant chimaeric transcripts may be expressed in solid tumors. We have developed a novel bioinformatics analytical pipeline to detect fusion gene transcripts from paired-end mRNA-seq data. The bioinformatics tool used for fusion transcript discovery employs multiple steps of false positive filtering and nominates the fusion transcript candidates with high confidence (approaching 100%). The screening and validation of the fusion candidates were quickly carried out using the recommended primer design regions as one of the outputs of the SnowShoes-FD pipeline. This pipeline was used to analyze the transcriptome of 22 breast cancer cell lines and 9 non-transformed breast cell populations. Fifty-four fusion candidates were nominated, all of which were validated using reverse transcription PCR and Sanger sequencing. Five fusion products were predicted to have a second fusion junction point between two fusion partners. These 54 fusion transcripts consist of 103 partner genes that are involved in cell cycle and/or nuclear receptor signaling. No fusion transcripts were identified from the non-transformed breast cell lines. We subsequently analyzed a panel of primary breast tumors, 8 each HER2+, triple negative, and ER+. Fifteen novel fusion transcript candidates have been nominated. Validation and functional analysis of these candidates is in progress.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-276. doi:10.1158/1538-7445.AM2011-LB-276
Abstract SY34-01: Biomarkers for predicting response to PI3K inhibitors
Authors: Cantley Lewis C.
Published: June 2012

Phosphoinositide 3-kinase (PI 3-kinase) is a central enzyme in a signaling pathway that mediates cellular responses to growth factors. The pathway is highly conserved from worms and flies to humans and genetic analysis of the pathway has revealed a conserved role in regulating glucose metabolism, cell growth and organism longevity. Based on deletion of genes encoding the catalytic or regulatory subunits of PI 3-kinase in the mouse, PI 3-kinase mediates insulin-dependent regulation of glucose metabolism, and defects in activation of this pathway result in insulin resistance and type 2 diabetes. In contrast, mutational events that lead to hyperactivation of PI 3-kinase result in cancers. Activating mutations in PIK3CA, encoding the p110alpha catalytic subunit of PI 3-kinase or inactivating mutations in PTEN, a phosphoinositide 3-phosphatases that reverses the effects of PI 3-kinase, are among the most common events in solid tumors. We, and others have generated mouse models in which a mutated form of the PIK3CA gene is expressed in a tissue-specific and reversibly inducible manner. These mice develop cancers that are dependent on continuous expression of the mutant PIK3CA gene. The PIK3CA driven tumors are FDG-PET positive and turning off PI 3-kinase with drugs results in an acute decline in FDG-PET signal that precedes tumor shrinkage. These results suggest that the ability of PI 3-kinase to stimulate high rates of glucose uptake and metabolism may be critical for the survival of PIK3CA mutant tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr SY34-01. doi:10.1158/1538-7445.AM2011-SY34-01
Abstract A26: Multi-Reg: An integrative parallel approach to uncover drivers of cancer
Authors: Akavia Uri David , Danussi Carla , Sanchez-Garcia Felix , Kotliar Dylan , Iavarone Antonio , Pe'er Dana
Published: June 2012

Tumor samples harbor a vast number of genomic alterations of various kinds, and it is not easy to distinguish driver alterations that contribute to oncogenesis from passenger alterations. Most computational methods attempt to identify driver alternations by focusing on the most frequent alterations. Previous work in our lab has led to the development of CONEXIC, a Bayesian framework for integrating copy number and gene expression to identify candidate driver genes in cancer and to link them to gene expression signatures they regulate (Akavia et al, Cell, 2010). This framework was applied to data from melanoma cell lines, where it correctly identified known drivers (MITF) and connected them to their known targets. In addition, it predicted novel tumor dependencies not previously implicated in melanoma, which were confirmed experimentally.

In general, current algorithms identify only one driver (the strongest) controlling a gene expression signature. However, drivers may act in parallel, where not only the strongest one is important. For example, either PTEN deletion or AKT activation can lead to a similar expression signature and phenotype. Therefore, we developed a new algorithm, based on the same principles as CONEXIC with multiple improvements. The new algorithm (called Multi-Reg) is capable of detecting multiple candidate regulators that can all act in parallel to regulate an expression signature. This algorithm can integrate mutations in addition to copy number and expression. Finally, we have designed Multi-Reg to be easier and quicker to run and more robust.

We applied Multi-Reg to glioblastoma data from The Cancer Genome Atlas (TCGA). This data includes copy number, gene expression and mutations for hundreds of primary tumor samples. We found 84 candidate regulators that were missed by CONEXIC, but discovered by Multi-Reg. Because of Multi-Reg's ability to search for genes working in parallel, it identified FGFR3, PDGFRA and NF1, in addition to EGFR & MET (identified by CONEXIC), as important candidate drivers. Additionally, Multi-Reg results identified RHPN2 as a novel oncogenic factor controlling a gene expression signature related to invasion and migration. Validation has shown that RHPN2 has limited effect on cell proliferation but induces invasiveness in glioblastoma cell lines.

Our results correctly identify known drivers of glioblastoma progression, including oncogenes such as EGFR, MET, CEBPB and tumor suppressors such as p16 and NF1. Multi-Reg also has the capability to identify more regulators than CONEXIC, and has correctly linked RHPN2 to the oncogenic phenomena it controls.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A26.
Abstract A31: The Myb transcription factor links the ERα and EGFR pathways in breast cancer cells
Authors: Bi Jing , Gilbert Nick
Published: June 2012

Drug resistance in breast cancer remains a big challenge, with many patients relapsing on endocrine therapies. To investigate growth-dependent signaling pathways in cancer we have generated a novel long-term estrogen-deprived cell line in serum-free conditions called DH as a model for antiestrogen treatment. DH cells are sensitive to ICI 182780, showing they are estrogen receptor (ERα)-dependent, they upregulate estrogen (E2)-dependent genes, indicating they are E2 responsive and are highly tamoxifen sensitive. This led us to hypothesize that in the total absence of E2 these cells might evolve to become highly E2 responsive and be capable of activating other growth-promoting pathways. We find that unliganded ERα is a negative regulator of EGFR expression but E2 stimulation triggers a slow upregulation of EGFR and a rapid reduction in ERα levels. After initial stimulation, EGFR expression is maintained despite E2 withdrawal, indicating a stable epigenetic modification at the EGFR promoter. E2 activates the growth-promoting MAPK pathway in an EGFR-dependent manner and we show a 5-day E2 pulse substantially increases the resistance of DH, MCF7, and ZR75-1 cells to tamoxifen and is accompanied by EGFR upregulation and phosphorylation. The cells remain sensitive to AG1478, an EGFR inhibitor, demonstrating a surprising functional switch between ERα and EGFR survival pathways, triggered by E2. To further investigate the relationship between ERα and EGFR, we used expression microarrays to identify MYB, a transcription factor, which is upregulated in response to E2. Upon E2 treatment, ERα binds to MYB intron 1, promoting MYB expression and consequently MYB is transiently located upstream of the EGFR promoter. MYB depletion by siRNA abrogates E2-dependent EGFR expression showing a direct link between these two pathways. These findings maybe significant for understanding acquired drug resistance on endocrine therapies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A31.
Abstract A38: Overexpression of FAT1 in human GBM (glioblastoma multiforme) and high-grade glioma cell lines
Authors: Dikshit Bhawana , Chosdol Kunzang , Sinha Subrata
Published: June 2012

FAT1, a human homologue of Drosophila tumor suppressor gene fat, is located on chromosome 4q35 in humans. In Drosophila, fat gene is known to be acting as an apical regulator of the tumor suppressor signaling, Salvador-Warts-Hippo (SWH) pathway. In mammalian cells, FAT1 is known to have a role in cell migration. The role of FAT1 in cancer is not clear and not much literature is available. In this study, we analyzed the expression of FAT1 in GBM (glioblastoma multiforme, grade IV glioma) samples at both mRNA and protein levels by means of q-PCR and immunohistochemistry. Herein, we demonstrate that 51.4% (18/35) of GBM samples had high expression of FAT1 mRNA, and 53.33% (8/15) of GBM samples had high FAT1 protein expression. We also checked the expression of FAT1 in glioma cell lines by q-PCR. Of the six glioma cell lines analyzed, four were GBM cell lines (U87MG, A172, U373MG and T98G) and two were astrocytoma grade 3 cell lines (GOS3 and SW1088). All GBM cell lines had high FAT1 expression while grade 3 cell lines had no or very low FAT1 expression. Since U87MG had maximum FAT1 expression we chose this cell line to study the effects of FAT1 knockdown by siRNA. FAT1 knockdown resulted in altered morphology and reduced migration of U87MG cells as compared to control siRNA treated cells. Since FAT1 is known to be an apical regulator of hippo pathway in Drosophila, we checked the expression of hippo pathway molecules by q-PCR and found that Mst2 (mammalian homolog of Drosophila hippo) expression was reduced up to 65% while other molecules like LATS and YAP were not altered at mRNA level. Considering the fact that regulation of SWH pathway primarily occurs at posttranslational level and depends upon phosphorylation status of the molecules, we are in process of analyzing the expression of these molecules at protein level and their phosphorylation status using phosphospecific antibodies, which would provide better understanding of the role of FAT1 in influencing SWH pathway. Since FAT1 is overexpressed in 50% of GBM samples studied and also found to have high expression in high-grade glioma cell lines as compared to low-grade glioma cell lines, it might act as an oncogene and may be involved in progression from low-grade to high-grade glioma. Thus, we propose FAT1 as a novel molecular target for therapeutic intervention in glioma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A38.
Abstract A40: Investigating the functional response of the ARF tumor suppressor to acute p53 loss
Authors: Forys Jason T. , Kladney Raleigh D. , Weber Jason D.
Published: June 2012

The tumor suppressor, p19ARF, is classically considered to function in response to hyerproliferative signals by binding and sequestering MDM2 in the nucleolus, thereby activating p53. Steady-state levels of ARF are kept low to prevent activation of p53 and are thought to monitor nucleolar function. Interestingly, p53 itself inhibits the Arf locus by recruiting proteins involved in heterochromatin formation. Cell lines that lack functional p53, therefore, accumulate enormous amounts of ARF protein but proliferate relatively well. Given that ARF is known to have functions independent of p53 both in vitro and in vivo, we wanted to investigate the function of ARF in the setting of acute p53 loss using conditional deletion of p53 from p53Flox/Flox mouse embryonic fibroblasts (MEFs). Cre-mediated excision of p53 leads to an induction of ARF by 4 days post infection, but levels do not peak until 10 days. Adeno-Cre infected MEFs have a higher proliferation rate than Adeno-LacZ controls, and do not undergo replicative senescence. However, knockdown of Arf in p53Δ/Δ MEFs using lentiviral shRNAs results in an increase in proliferation and enhanced growth in semisolid medium following RasI/12 transformation. These findings suggest that the induced ARF levels are acting to inhibit p53-null cells in some capacity. Loss of p53 also leads to a dramatic downregulation of MDM2 protein levels. Immunoprecipitation experiments indicate that ARF predominantly interacts with MDM2 in LacZ-infected cells, but binds the ribosomal chaperone nucleophosmin (NPM) more readily in p53Δ/Δ MEFs. Since inhibition of NPM by ARF has been shown to negatively affect overall protein synthesis, we hypothesize that in the absence of p53, ARF functions to negatively regulate overall protein synthesis through NPM in an effort to slow proliferation. Indeed, p53Δ/Δ MEFs contain less total protein per cell, copies of 47S rRNA, and have reduced active translation compared to their LacZ counterparts. We are currently testing whether knockdown Arf in p53Δ/Δ MEFs can rescue protein synthesis, thus allowing these cells to proliferate at an even faster rate. These findings could explain why certain cancers (i.e., pancreatic cancer) preferentially inactivate both ARF and p53. We are also attempting to understand why the high levels of endogenous ARF seen in the absence of p53 are not able to induce a cell cycle arrest when ARF is clearly capable of eliciting such a response independently of p53. As p53 mutation occurs in over 50% of all cancers it would be of interest to know how we could activate this function of ARF in those patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A40.
Abstract A47: Metastatic development of prostate cancer and multidrug resistance
Authors: Hsiao Pei-Wen , Tsai Chin-Hsien , Yang Yu-Chih , Lee Ming-Ting , Tsai Mong-Hsun
Published: June 2012

Signaling pathway of androgen receptor plays a central role in the prostate cancer progression. Accordingly, approaches and molecular mechanisms to ablate the androgen receptor function in prostate cancer have been actively studied to improve the disease treatment. However, the disease inevitably develops resistance to all forms of hormonal therapies. To further understand metastatic prostate cancer, we established orthotopic xenograft model using various human cell lines expressing Luc2 reporter by lentivirus-mediated infection. Growth of these orthotopic xenografts develops a wide spectrum of spontaneous metastases in vivo, from which we also obtained cells from more aggressive and metastasized tumors. Indeed, cancer cells isolated from larger primary tumors and metastatic tumors are more invasive than those isolated from smaller primary tumors. Genome-wide RNA expression (transcriptomes) in cancer cells from metastases against primary large and small tumors were compared to reveal the difference between the different cancer stages. Real-time RT-PCR confirmation and Ingenuity Pathways Analysis (IPA) software analysis reveal activation of NFkB pathways and P-glycoprotein, so-called multidrug resistance-associated proteins in metastasis. Our study established useful cells and indicates a potential molecular basis of the drug resistance along the in vivo development of human prostate cancer.

Supported by Academia Sinica, Career Development Award.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A47.
Abstract A49: The ERK/MAPK scaffold IQGAP1 is a tumor-selective target
Authors: Jameson Katherine L. , Zehnder Ashley M. , Khavari Paul A.
Published: June 2012

To advance insight into skin biology and pathophysiology, our lab has focused on understanding how the normal processes of cell division and migration are regulated and how they become dysregulated in pathological states, including cancer. One particular pathway of interest in the lab is the extracellular-regulated kinase/mitogen-activated protein kinase (Erk/MAPK) pathway. This highly conserved pathway is critical to epidermal homeostasis and tumorigenesis, but its mechanisms of action remain unclear. This research explores how the Erk/MAPK-interacting protein IQGAP1 mediates signaling in tumorigenesis. IQGAP1 physically interacts with canonical Erk/MAPK pathway members in order to precisely modulate their signal output and in its absence, signaling through this pathway is severely diminished. We recently found that depleting expression of IQGAP1, a Mek- and Erk-binding MAPK-IP, exerted no effect on normal tissue but effectively blocked invasive neoplasia. We are currently working to define the functional domains of IQGAP1 required for Erk MAPK-driven epidermal neoplasia. These efforts will help identify potentially important and therapeutically relevant neoplasia-selective interactions of IQGAP1.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A49.
Abstract A60: Kras-transformed Caco-2 cells establish a 3-D model of colon cancer
Authors: Magudia Kirti , Lahoz-Ruano Aurelia , Hall Alan
Published: June 2012

While Caco-2 cells derived from a human colorectal carcinoma have historically been used to study differentiated colonic epithelium in 2-D cultures, recent work has used a 3-D cell culture system to model epithelial morphogenesis. We describe here the use of 3-D Caco-2 cell cultures to model colon cancer progression. Caco-2 cells harbor APC, p53, and SMAD4 mutations, but are wild type for Kras, the most frequently mutated gene in colon cancer. Control Caco-2 cells seeded in extracellular matrix generate hollow, polarized structures after 10 days. In contrast, Caco-2 cells expressing oncogenic Kras form much larger and filled structures that lack a central lumen. Furthermore, Western blot analysis of protein lysates made from day 1 and day 10 control structures reveals a strong decrease in cyclin B, suggesting a reduction in the proportion of cells undergoing mitosis. Kras structures, however, maintain cyclin B levels, providing evidence of prolonged proliferation. Immunofluorescence of the apical marker aPKC and the tight junction protein Z0–1 shows that Kras structures have disrupted polarity. Interestingly, Kras-expressing Caco-2 cells do not invade through a matrigel-based matrix. Thus we will investigate which other factors cooperate with oncogenic Kras to promote tumor progression in addition to which pathways downstream of Kras contribute to this transformed acinar phenotype. As most colon cancer research is conducted either with cell lines in 2-D culture or in models, this 3-D model has the potential to provide both relevance to human tumors and ease of mechanistic inquiry.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A60.
Abstract B28: Mitochondrial dysfunction is associated with cancer progression in ulcerative colitis
Authors: Risques Rosa Ana , Ussakli Cigdem Himmetoglu , Ebaee Anoosheh , Brentnall Teresa A. , Rabinovitch Peter S.
Published: June 2012

Ulcerative colitis (UC) is an inflammatory disease of the colon that predisposes to colorectal cancer. Dysplasia and cancer arise in pre-cancerous fields of inflammed epithelium, which harbor molecular alterations in spite of histologically normal appearance. We hypothesized that these pre-cancerous fields might contain dysfunctional mitochondria as a consequence of oxidative stress associated with the inflammatory environment. If so, mitochondrial alterations could be used to identify UC patients at risk for cancer progression. To test this hypothesis we stained colon biopsies with Cytochrome C oxidase subunit I (COX), a respiratory transport chain protein whose loss is attributed to mitochondrial DNA mutations. Four histologically normal biopsies randomly collected from the colon of 9 UC patients with high-grade dysplasia or cancer (Progressors) and 9 patients free of dysplasia (Non-progressors) were stained with COX and quantified using multispectral imaging. Low and high grade dysplasia (LGD, HGD) and cancer biopsies were also included in the analysis. Subsequent to COX staining, a subset of the biopsies were also stained for Prohibitin, Opal, and Mfn1, which are nuclear encoded mitochondrial proteins. On average, UC progressors had 40% less COX staining than UC non-progressors (p<0.0001). The mean COX staining intensity of 4 biopsies could distinguish UC progressors from non-progressors with 100% sensitivity and 77.8% specificity. Moreover, in UC progressors, the closer the biopsies were located to dysplasia the lower the amount of COX staining (p=0.003), suggesting that mitochondrial abnormalities increase progressively in more advanced preneoplastic fields. Complete COX loss was observed in some non-dysplastic biopsies <10cm to dysplasia and in LGD biopsies. Interestingly, HGD and cancer showed areas with very intense COX staining, suggesting a recovery of mitochondrial function later in neoplastic progression. Prohibitin, Opa1, and Mfn1 showed similar trends of loss in early progression and gain in later progression, although the patterns of staining did not always overlap with COX. Thus, we have shown that mitochondrial dysfunction, measured as the loss of COX staining, precedes the development of dysplasia and cancer and may be useful as a biomarker for cancer risk in UC patients. Intriguingly, mitochondrial function seems to be recovered in HGD and cancer, suggesting that epigenetic mechanisms underlie the pre-neoplastic loss of mitochondria. These results are currently under investigation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr B28.
Abstract B29: Female reproductive cancers incidence in Bihar, India with the risk of xenoestrogenic arsenic in serum and in drinking water sources
Authors: Nath Akhileshwari , Singh Jitendra Kumar , Vendan S. Ezhil , K Priyanka
Published: June 2012

Reproductive cancers are the most prevalent cancers among the women worldwide. Breast, ovary, uterine and cervix cancers are leading cancers in India. Hormonal carcinogens also known as endocrine disturbing chemicals (EDCs) are predominant factors causing reproductive cancer risks in women. Heavy metal Arsenic (As) is naturally found in Northern India. It is popularly known as EDC especially mimicking as estrogen (i.e., xenoestrogen). The present study aimed to assess the hormonal carcinogenic risk of As among women in Bihar, India. Cancer incidences in Bihar were recorded from the patient's clinical registry in Mahavir Cancer Sansthan hospital. Serum was collected from the different cancer patients and the level of As in serum was estimated by Atomic Absorption Spectroscopy. Drinking water samples were collected from different regions of Bihar and As was estimated by As kit. During the last ten years cervix and breast cancer incidence were very high followed by ovary, uterine and other cancers. Among the 38 districts of Bihar, those cancers incidence were highly marked in the Bhagalpur, Bhojpur, Chapra, East Champran, Gaya, Gopalanj, Jahanabad, Lakhisarai, Madobani, Muzaffarpur, Nalanda, Patna, Samastipur, Saran, Siwan and Vaishali districts. Maximum incidences were highly recorded in the age group of 36 to 45 years. More than 10 ppb level of As in serum was recorded in all the tested breast, ovary, uterine and cervix cancer cases. Highest concentration of As was recorded in cervix (17.6 ppb) followed by breast, uterine and ovary cases (13.3, 11.3 and 4.9 ppb respectively). WHO permits 10 ppb of As as maximum in drinking water sources, whereas in the present study we found that near to Gangetic region As level was high (more than 10 ppb) than the non-Gangetic region. Together, the results indicate that As contamination in drinking water shows a relationship between female reproductive cancers incidence and As occurrence in patients serum. In conclusion, As might be suppress female reproductive function by as environmental toxicant that is contra-mechanistic to that of estrogen.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr B29.
Abstract B35: Genetic and epigenetic alterations in colorectal cancer among atomic bomb survivors
Authors: Ito Reiko , Eguchi Hideaki , Hamatani Kiyohiro , Taga Masataka , Oue Naohide , Yasui Wataru , Nakachi Kei , Kusunoki Yoichiro
Published: June 2012

Colon cancer among atomic bomb survivors has been shown at a significantly high excess risk attributed to atomic radiation exposure. In this study, we focused on microsatellite instability (MSI)-related colorectal carcinogenesis and analyzed both epigenetic and genetic alterations of MLH1 and Ras-signaling related genes, as well as CpG island methylator phenotype (CIMP).

Study subjects were 35 colorectal cancer cases from the RERF cohort study (Life Span Study). The MSI-high (H) was found in five cases with use of six microsatellite markers, showing a significantly higher median radiation dose than that of microsatellite stable (MSS) and MSI-low (L) cases. All five MSI-H cases carried LOH of MLH1, three of which had methylated MLH1 in the remaining alleles. Two cases with unmethylated MLH1 were found to have point mutations in MLH1, presumably causing truncated protein and splicing abnormality. Of 30 MSS and MSI-L cases, two methylated MLH1 cases were identified, i.e., a total of five methylated MLH1 cases in all study cases, and these five cases showed a higher radiation dose than that of unmethylated cases. Undetected levels of MLH1 protein expression were confirmed with four of the five MSI-High cases. Next, we analyzed the gene alterations (BRAF and K-RAS mutations and RASSF2 methylation) in the Ras-signaling pathway, which is thought to be a preceding event of MLH1 methylation. All five MSI-H cases carried one or two alterations of these Ras-signaling-related genes, and a total of 18 cases were found to have altered Ras-signaling-related genes with a significantly higher radiation dose than that of 17 cases without the alternations. When the study cases were grouped by the numbers (2, 1, and 0) of alternated Ras-signaling-related genes, three groups revealed a remarkably increasing trend in radiation dose with increased numbers of altered genes. Although CIMP status significantly correlated with MLH1 methylation, no association was found with CIMP status and radiation dose.

The results obtained thus far suggest that radiation exposure may influence MSI-related epigenetic/genetic alterations in colorectal carcinogenesis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr B35.
Abstract B36: Glutathione S-transferase pi modulates DNA damage responses
Authors: Kang Bong-su , Lee Beom Jun , Kim Jong-Soo
Published: June 2012

Glutathione S-transferase pi (GSTP1 in human) has been considered to be a marker for cancer development, because of high levels of expression in variety of cancers and preneoplastic lesions, because in most normal tissues, its expression is very low. High level of GSTP1 expression and its nuclear localization have been shown to correlate with drug resistance and also with tumor increased tumor grade.

To demonstrate the function of GST-pi in DNA damage response, using the GST-pi over expressing stable cells ((HEK293-GST-pi) we examined protein interactions

We found that phosphorylation levels of p53 at Ser-15 induce by non-lethal concentration of hydrogen peroxide (0.25∼0.5mM) were lower in HEK293-GST-pi cells than those in wild-type cells. p53 is phosphorylated at Ser-15 in response to cellular stress, DNA damage, by ATM (ataxia-telangiectasia mutated), ATR (ataxia-telangiectasia mutated and Rad3-related), and DNA-PK (DNA-dependent protein kinase).

On the other hand, elevated nuclear Mre11 (DNA double strand break sensor) level was observed following exposure to hydrogen peroxide in HEKk293-GST-pi cells compared with wild-type cells, whereas the expression level of Mre11 in HEK293-GST-pi cells was similar to or even lower than that of wild-type cells in the absence of hydrogen peroxide treatment.

In HEK293-GST-pi cells, hydrogen peroxide treatment induced predominant translocation of nuclear GST-pi to cytoplasm. Similar results were obtained in UV irradiated experiments.

Since p53 and Mre11/Rad50/NBS1 complex are downstream regulators of ATM/ATR, GST-pi may play a role in modulating ATM/ATR signaling in cellular stress condition.

Further observations are undergoing.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr B36.