bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3049: Overexpression of miR-17 family miRNAs in prostate cancer biopsies: Evidence for a stem-cell-like miRNA profile in high grade/high stage tumors
Authors: Gaston Sandra M. , Guerra Andrew L. , Nadadur Rangarajan C. , Meadows Julie , Kearney Michael C. , Sorbara Lynn , Srivastava Sudhir , Meltzer Stephen J. , Pinsky Paul , Kearney Gary P.
Published: February 2011

Complex changes in microRNA (miRNA) expression patterns have been observed in many different types of cancer, including prostate cancer. Although the consequences of deregulation of miRNA expression in human cancer are still largely unknown, the use of miRNAs for cancer classification and prognosis has emerged as an important new area of biomarker research. We compared the tissue expression of miRNAs in prostate biopsies from patients with high grade/high stage tumors with biopsies from patients with no evidence of malignancy. Snap frozen prostate biopsy samples were prospectively collected using tissue printing techniques and total RNA was isolated for miRNA analysis using Agilent miRNA microarrays (Version 1, containing 470 human tissue miRNAs). The prostate cancer cases included in the microarray profiling study had at least 5 of 12 cores positive for cancer and significant areas of Gleason 7 or more. The no-cancer controls showed no prostate cancer, HGPIN or suspicious histology in any of the 12 biopsy cores. In our analysis of miRNA expression patterns, “significant change in expression” was defined as at least 2 fold change and F test (p<0.05). Three different internal reference normalization methods were used in the data analysis; here we classify miRNAs as “overexpressed” or “underexpressed” only when all three methods agree. In high grade/high stage biopsy cancer, 64 of the 470 human tissue miRNAs on the microarray (13.6%) were found to be significantly overexpressed when compared with miRNA expression in biopsies from no-cancer controls. This set of overexpressed miRNAs includes 4 of the 5 miR-200 family miRNAs that have been implicated in the regulation of the epithelial-to-mesenchymal transition (EMT). The cancer-overexpressed miRs also include 6 of the 8 miR-17 family genes; this miRNA family has been implicated in the control of stem-cell differentiation during embryogenesis. Interestingly, the 6 miR-17 family miRs that are overexpressed in high grade/high stage prostate cancer include 2 from each of the 3 polycistronic clusters that encode this highly conserved miR family, an expression pattern similar to the coordinated expression observed in early embryonic development. This robust stem-cell-like miRNA profile offers an interesting new candidate biomarker that may be particularly sensitive to an aggressive prostate cancer phenotype.

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 3049.
Abstract 3050: MicroRNA expression profiling of human Barrett's carcinogenesis
Authors: Fassan Matteo , Volinia Stefano , Palatini Jeff , Pizzi Marco , Baffa Raffaele , Clemente Roberto , Rizzetto Christian , Croce Carlo M. , Zaninotto Giovanni , Ancona Ermanno , Rugge Massimo
Published: February 2011

Barrett's Mucosa (BM) is a metaplastic replacement of the native esophageal (squamous) epithelium (ESq) by columnar-intestinalized mucosa. BM is the main risk factor for Barrett's adenocarcinoma (BAc). MicroRNAs (miRNAs) are a class of small non-coding RNAs that control gene expression by targeting mRNAs; miRNAs deregulation has been associated with Barrett's oncogenesis. To explore the hypothesis that a specific miRNAs signature is associated with BM (and its associated cancer risk) a miRNA microarray analysis (OSU-CCC version 4.0; Ohio State University) compared ESq versus all the spectrum of the phenotypic lesions of the Barrett's carcinogenesis. Specimens were collected at Department of Pathology of Padova University from 14 patients undergone esophagectomy for BAc or High-grade non-invasive neoplasia (HG-NiN). Tissues samples for the microarray study included: ESq= 14 samples; BM= 14; Low-grade non-invasive neoplasia (LG-NiN)= 7; HG-NiN= 5; BAc= 11. The miRNA-profiling study consistently disclosed increased expression of 6 miRNAs (hsa-miR-215, hsa-miR-560, hsa-miR-615-3p, hsa-miR-192, hsa-miR-326, hsa-miR-147), and decreased expression of 7 others (hsa-miR-100, hsa-miR-23a, hsa-miR-605, hsa-miR-99a, hsa-miR-205, hsa-let-7c, hsa-miR-203). Microarray results were further validated by qRT-PCR, using a different series of 10 consecutive BAc cases (Esq= 10 samples; BM= 6 samples; BAc= 10 samples). Interestingly, some of the miRNAs found deregulated in this series are already known as suitable markers of cancer progression in other models of epithelial oncogenesis (i.e.: hsa-let-7c, hsa-miR-192, hsa-miR-203, hsa-miR-205, hsa-miR-215). The results achieved by the miRNA profiling study were further validated by the immunohistochemical (IHC) analysis on a well-defined miRNA gene target (HMGA2 [hsa-let-7c]). The IHC expression of the protein product was consistent with the corresponding miRNA's deregulation. In fact, HMGA2 IHC expression was up-regulated in HG-NiN and BAc. The achieved results confirm that specific miRNAs are involved in BM carcinogenesis and that they may represent a novel diagnostic/prognostic tool in the characterization of BAc gene targets.

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 3050.
Abstract 3057: The serine protease inhibitor, elafin, exhibits novel tumor suppressor functions in the context of breast cancer
Authors: Caruso Joseph A. , Hunt Kelly K. , Keyomarsi Khandan
Published: February 2011

Serine proteases and their inhibitors have an established role in cancer progression, invasion, and metastasis. Elafin is an endogenously expressed inhibitor of human neutrophil elastase. In mammary epithelial cells elafin is highly expressed; however in breast cancer it is transcripitionally downregulated. We hypothesize that the loss of elafin is critical to the development of the malignant phenotype in breast cancer. In human mammary epithelial cells (HMECs) and breast cancer cell lines, the regulation and activity of elafin is dependent on the critical tumor suppressor retinoblastoma (Rb). Elafin is dramatically upregulated in quiescent HMECs following growth factor deprivation. This upregulation is characterized by an Rb dependent increase in elafin stability and cytoplasmic accumulation. Knockdown of elafin by shRNA in HMECs causes them to be refractory to growth factor deprivation induced quiescence; suggesting that elafin upregulation is a key regulator of cellular quiescence. Overexpression of elafin by adenovirus in both HMECs and breast cancer cell lines lacking Rb causes caspase-3 dependent, apoptotic cell death. Additionally, adenoviral elafin causes Rb-dependent cell cycle arrest in tumor cells with functional Rb. The Rb pathway is lost or inactivated in the majority of cancers. The design of a therapeutic strategy capable of specifically targeting tumor cells with Rb inactivation may prove to have significant utility in the treatment of cancer. The data summarized here demonstrates that overexpression of elafin is cytostatic in Rb-positive breast cancer cells and cytotoxic in Rb-negative breast cancer cells expressing caspase 3; suggesting that elafin could be engineered into a therapeutic modality in the treatment of breast 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 3057.
Abstract 3062: Identification and functional analysis of NOL7 tumor suppressor gene nuclear and nucleolar localization signals
Authors: Lingen Mark W. , Zhou Guolin , Doci Colleen
Published: February 2011

NOL7 is candidate tumor suppressor found within the nucleus and nucleolus. However, the mechanisms regulating its subcellular localization have not been elucidated. An in vitro import assay demonstrated that NOL7 requires cytosolic machinery for active nuclear transport. Using NOL7 deletion constructs and cytoplasmic pyruvate kinase (PK) fusion proteins, three nuclear localization signals (NLSs) were identified. Site-directed mutagenesis of PK fusions and full-length NOL7 defined the functional regions within each NLS. Further characterization revealed that NLS2 and NLS3 were critical for both the rate and efficiency of nuclear targeting. In addition, four basic clusters within NLS2 and NLS3 were independently capable of nucleolar targeting. The dynamics of nucleolar localization were investigated by FRAP and specific nucleic acid depletion. NOL7 is relatively immobile within the nucleolus, but shuttles rapidly between the nucleolus and nucleoplasm. In addition, targeting to the nucleolar compartment was dependent on the presence of RNA, as depletion of total RNA or rRNA resulted in a nucleoplasmic shift of NOL7. Take together, these results identify the minimal sequences required for the regulation of NOL7 subcellular localization, characterizes the contribution of each sequence to NOL7 nuclear and nucleolar dynamics and suggests a functional role for NOL7 in both compartments.

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 3062.
Abstract 3063: Novel putative tumor suppressor LZAP binds and regulates Wip1
Authors: An Hanbing , Lu Xinyuan , Yarbrough Wendell G.
Published: February 2011

Our group has shown that LZAP is lost in ∼30% of head and neck squamous cell carcinomas and that inhibition of LZAP expression is associated cellular transformation and xenograft tumor growth. LZAP increases invasion and inhibits cellular proliferation and clonogenic growth mediated correlating with inhibition of NF-kB and activation of p53. Other researchers have found that LZAP promotes cell death in response to cytotoxic therapy at least partially dependent on activation of Cdk1 through Chk1/2 inhibition. We have also found that LZAP binds and inhibits p38 and this inhibition does not depend on upstream regulators MKK3 and MKK6. The activity of MAPKs reflects a balance between the upstream activating kinases and inactivating protein phosphatases. Since LZAP inhibition of p38 is not through upstream kinases, we determined if phosphatase activity targeting p38 was regulated by LZAP.

Wip1 is a major phosphatase regulating p38 phosphorylation and activation. Wip1 is a member of the PP2C family of serine/threonine phosphatases and was identified in a screen to detect p53 responsive genes. We noted that LZAP and Wip1 had many binding partners/targets including RelA, p38, and Chk1/Chk2 in common. Mechanisms of LZAP inhibition of RelA, p38 and Chk1/Chk2 are not well described, but expression of LZAP is associated with decreased phosphorylation of all three target proteins. To determine if LZAP could regulate Wip1 activity, we first evaluated LZAP and Wip1 interaction through immuno- or affinity precipitation. LZAP and Wip1 co-precipitated after expression in mammalian cells and bacterially expressed LZAP pulled down Wip1 generated through in vitro transcription and translation. After transient expression in mammalian cells, Wip1 and LZAP co-localized by indirect immunofluorescence staining. When expressed singly, Flag-tagged Wip1 localized to the nucleus and LZAP was excluded from the nucleolus, but localized to both cytoplasm and nucleus. When Wip1 and LZAP were co-expressed LZAP localization was altered so that nucleolar LZAP was observed. These findings indicate that co-expression of LZAP and Wip1 results in alternation of LZAP subcellular localization with resultant co-localization of these two proteins to nucleus. Functionally, co-expression of LZAP with Wip1 increased Wip1 content within p38 immunocomplexes which was associated with decreased p38 phosphorylation.

Together, our findings indicate LZAP binds and co-localizes with Wip1 and increases activity of Wip1 towards p38. Wip1 may play a common mediator for LZAP activities.

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 3063.
Abstract 3066: YPEL3, a novel inducer of senescence, is inhibited by estrogen in ER+ mammary tumor cells
Authors: Tuttle Rebecca M. , Miller Kelly L. , Termulhen Paula M. , Berberich Steven J.
Published: February 2011

It was recently discovered by this laboratory that Yippie-like 3 (YPEL3) is a novel p53 regulated gene that when induced triggers premature senescence in primary and tumor cell lines. Initial screens of human tumor samples uncovered significant decreases in YPEL3 gene expression in ER- verses ER+ breast cancers leading us to believe that estrogen may have a regulatory effect on YPEL3 expression. Using ER+ MCF7 cells, we determined that estrogen is a significant inhibitor of YPEL3 gene expression in cells grown in media with 10% serum verses cells grown in media with charcoal stripped serum (CSS) from which estrogen has been removed. The increase in YPEL3 mRNA expression demonstrated in CSS can be blocked by the addition of estrogen, and as expected tamoxifen competes with this estrogen repressive effect. The effects of estrogen on YPEL3 expression are not observed in MCF10A mammary epithelial cells which are ER negative demonstrating the requirement for the estrogen receptor in the observed effect. Given our observation that YPEL3 induction triggers senescence in breast cancer cells, we employed beta-galactosidase activity as a measurement of senescence and observed an 8-fold increase in senescence in MCF7 cells grown in CSS media. The increase is reduced to less than 3.5-fold with the addition of estrogen to the CSS media, and the increase in cellular senescence is blocked in MCF7 cells stably selected for the loss of YPEL3 (MCF7-shYPEL3) suggesting the need for the absence of estrogen and the presence of YPEL3 in senescence induction. We are currently testing the effects of estrogen on MCF7 cells where p53 has been knocked down using RNAi approaches and are planning to test the effects of RNAi targeting ER-alpha. We believe estrogen is inhibiting YPEL3 expression through its association with the estrogen receptor and their association with p53 thereby inhibiting p53 transactivation of YPEL3. These findings suggest that modulation of YPEL3 by estrogen and its growth suppressive effects may provide a target for future breast cancer therapies.

Note: First two authors contributed equally.

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 3066.
Abstract 3069: Importance of zinc-binding domain of matrix metalloproteinase 19 (MMP19) in tumor suppression and anti-angiogenesis of nasopharyngeal carcinoma
Authors: Chi Chan King , Ko Josephine M.Y. , Lung Hong Lok , Sedlacek Radislav , Lung Maria Li
Published: February 2011

Background and aims: Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases consisting of more than 20 enzymes. MMP19 shares the typical MMP structural domains. It degrades various ECM components including collagen type IV, nidogen-1, fibronectin, tenascin-C isoform, aggrecan, and laminin-5-gamma-2-chain. In our previous studies, MMP19 was identified as a candidate tumor suppressor gene (TSG) in nasopharyngeal carcinoma (NPC). It was down-regulated in NPC cell lines and NPC biopsies, when compared to an immortalized normal nasopharyngeal epithelial cell line and corresponding non-tumor nasopharyngeal tissues. Transfection of wild-type (WT) MMP19 into the HONE1-2 NPC cell line significantly suppressed tumor formation in the nude mouse tumorigenicity assay and decreased the tube-forming ability of both human umbilical vein endothelial cells (HUVEC) and human microvascular endothelial cells (HMEC-1). In this study, we aimed to validate the importance of the zinc-binding domain of MMP19 in NPC. Methods: We transfected the mutant MMP19, which contained a mutation at the zinc-binding domain, into HONE1-2 to study the effect of mutant MMP19 in tumor suppression and anti-angiogenesis compared to WT MMP19. Results: Mutant MMP19 was not tumor-suppressive, as is the WT MMP19, and aggressive tumors appeared with the same growth kinetics as the vector-alone control. The anti-angiogenesis effect was also not observed in endothelial cells incubated with conditioned medium from mutant MMP19 clones compared to WT MMP19 clones. Conclusions: Previous studies showed mutation of the zinc-binding domain in MMP19 leads to loss of MMP19 function in cell proliferation, migration, and protein degradation in a human keratinocyte cell model. This study demonstrates loss of tumor suppressive and anti-angiogenesis function of mutant MMP19 in NPC and indicates the importance of the zinc-binding motif for WT MMP19 in suppression of tumor formation and in inhibition of tube-forming ability of endothelial cells in NPC.

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 3069.
Abstract 307: EphB4 promotes or suppresses Ras/ERK pathway depending on cellular contexts: Implications for EphB4 as a cancer target
Authors: Xiao Zhan , Carrasco Rosa A. , Kinneer Krista , Sabol Darrin , Coats Steve , Jallal Bahija , Tice David
Published: February 2011

EphB4 is a member of the Eph RTK family. Its classical role has been shown to act in neuronal guidance and mediate venal/arterial separation. In contrast to these more established roles, EphB4's function in cancer is controversial since both tumor promoting as well as tumor suppressing functions have been ascribed to it. Here we provide a vivid illustration of this intriguing dichotomy by showing that EphB4 activation resulted in inhibition of the Ras/ERK pathway in endothelial cells but induction of the same pathway in MCF-7 breast cancer cells. This was true if EphB4 was stimulated with ephrinB2, its natural ligand, or an EphB4-specific fully human monoclonal antibody. The reason for these dramatic differences is due to potential functional coupling of EphB4 to different downstream effectors. In HUVEC cells, the RNAi knockdown of p120 RasGAP, a Ras inhibitor, abrogated the inhibitory effect on Ras/ERK pathway. However, in MCF7 cells, RNAi knockdown of PP2A, which has been shown to activate the Ras/ERK pathway through removal of the inhibitory phosphorylation on c-Raf, negated EphB4-driven activation of the Ras/ERK pathway. This represents the first time a functional coupling between Eph receptor and PP2A has been established, and this interaction appears to confer activation but not suppression of an oncogenic pathway. Our study also demonstrates the caveats and potential challenges of targeting EphB4 for cancer therapy due to the presumed conflicting effects on cancer cell and endothelial cell compartments.

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 307.
Abstract 3074: Neutrophil gelatinase-associated lipocalin inhibits proliferation of human esophageal adenocarcinoma cells both in vitro and in vivo
Authors: Tong Zhimin , Koolwal Pooja , DeMars Catherine J. , Buttar Navtej S. , Guha Sushovan
Published: February 2011

Background: Neutrophil gelatinase-associated lipocalin (NGAL), a member of lipocalin family, is a 25-kDa secreted glycoprotein. NGAL modulates multiple biological functions including cell proliferation, invasion, and metastasis. NGAL is upregulated in multiple gastrointestinal cancers including colorectal, pancreatic, and esophageal adenocarcinoma (EAC). Our group recently showed that NGAL over-expression significantly blocked metastasis of pancreatic adenocarcinoma cells [Cancer Research 68(15): 6100-6108, 2008]. However, the biological role of NGAL in EAC is not known.

Aim: To examine the biological role of NGAL in human EAC cells both in vitro and in vivo.

Methods and Results: We used OE33 (well-differentiated) and Flo-1 (poorly differentiated) EAC cell lines as our model system. We observed that OE33 cells have significantly higher levels of NGAL expression than Flo-1 cells both by qPCR and western blot analyses. To further study the biological role of NGAL in EAC, we created stable NGAL over-expression clones in Flo-1 (Flo1-NGAL) and NGAL under-expression clones in OE33 (OE33-shNGAL) using lentiviral system. We observed that under-expression of NGAL in OE33 significantly increased cell proliferation as determined by clonogenic assays. In contrast, NGAL over-expression in Flo-1 significantly suppressed cell proliferation. We also observed that NGAL under-expression increased cyclin D1 expression, a known cell cycle regulator. This, in part, explained increased EAC cell proliferation secondary to NGAL under-expression. In addition, NGAL over-expression potently suppressed and conversely, NGAL under-expression significantly increased invasion of EAC clones through MatrigelTM in vitro. Furthermore, to confirm our in vitro findings, we observed that OE33-shNGAL clones strikingly increased tumor volume in a subcutaneouos heterotopic xenograft model in nude mice.

Conclusion: We demonstrated for the first time that NGAL under-expression increased cell proliferation and invasion in vitro, which could be in part secondary to regulation of cyclin D1 expression. NGAL under-expression also enhanced tumor growth in vivo. Thus, modulation of NGAL can be a novel therapeutic target in progression of EAC.

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 3074.
Abstract 308: Differential roles of AP2 in p38 MAPK- and ligand-induced EGFR internalization
Authors: Grandal Michael V. , Andersen Mette H. , Holst Mikkel , Grøvdal Lene M. , van Deurs Bo
Published: February 2011

The epidermal growth factor receptor (EGFR) is an important regulator of normal growth and differentiation, and it is involved in the pathogenesis of many cancers. Endocytic downregulation is central in terminating EGFR signalling after ligand stimulation, and this downregulation is fairly well known. Recently, it has been shown that also p38 MAPK activation can stimulate EGFR endocytosis. This endocytosis differs from the ligand-induced internalization by lacking many of the hallmarks normally necessary for endocytosis of EGFR. We compared the two kinds of endocytosis with regard to the requirements for proteins in the internalization machinery. Both kinds of endocytosis require clathrin heavy chain, but whereas EGF-induced EGFR internalization required Grb2, p38 MAPK-induced internalization did not. Interestingly, AP2 knock down blocked p38 MAPK-induced EGFR internalization, but only mildly affected EGF-induced internalization. The involvement of AP2 in EGFR endocytosis was verified with at Eps15 mutant that sequesters AP2 in the cytosol and abolishes AP2 function. The functional difference of AP2 in the two kinds of EGFR internalization is currently investigated.

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 308.
Abstract 3083: Regulation of novel tumor suppressor gene FUS1 expression through 5’- and 3’-UTRs and in response to DNA damage in human lung cancer cells
Authors: Lin Jing , Xu Kai , Roth Jack A. , Ji Lin
Published: February 2011

FUS1 is a novel tumor suppressor gene located in the human chromosome 3p21.3 region. FUS1 mRNA transcripts could be detected on Northern blots in both normal lung and some lung cancer cell lines, but no endogenous FUS1 protein could be detected in a majority of lung cancer cell lines. Loss or reduction of FUS1 expression was observed in all small cell lung cancer and in more than 70 % of non-small cell lung carcinomas and was associated with significantly worse overall patient survival. However, mechanisms regulating FUS1 protein expression and its inactivation in primary lung cancer cells are largely unknown. In this study, we investigated the role of the 5’- and 3’-untranslated regions (UTRs) of the FUS1 gene in the regulation of FUS1 protein expression and determined changes in FUS1 gene and protein expression in response to oncogenic stress in normal lung and cancer cell lines. We constructed a series of expression plasmids containing the FUS1 coding region with or without the intrinsic UTRs and transfected them into normal lung bronchial epithelial and tumor cells to assess their roles in regulation of FUS1 protein expression. We identified some critical structure elements in FUS1 UTRs that could significantly regulate FUS1 protein expression. We found that two small upstream open-reading frames (uORFs) encoding an 8-amino-acid peptide and another 6-amino-acid peptide, respectively, in the 5’UTR of FUS1 could markedly inhibit the translational initiation of FUS1 protein by interfering with the “scanning” of the ribosome initiation complexes. Several secondary RNA structural elements/motifs on the 3’UTR of FUS1 also showed a significant inhibitory effect on FUS1 protein expression. The 3’UTR-mediated regulatory effect on FUS1 proteins expression was differentially detected in normal lung epithelial and fibroblast cells compared to lung cancer cells. We found that FUS1 expression could be induced or activated in the presence of DNA damaging agents, such as UV irradiation, etoposide, and cisplatin. UV irradiation could induce FUS1 protein and mRNA expression independent of the status of FUS1 UTRs. The FUS1 expression activated by DNA damaging agents such as etoposide or cisplatin facilitated FUS1-induced apoptosis, prevented cells from dividing and accelerated cell death. Our results provide new insight into the molecular mechanisms in the regulation of FUS1 expression in lung cancer cells. (Supported by NIH SPORE P50CA70907 and RO1CA116322 grants)

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 3083.
Abstract 3090: RREB1 is necessary for the proliferation of bladder cancer cell lines
Authors: Nitz Matthew D. , Harding Michael A. , Theodorescu Dan
Published: February 2011

It is estimated that in 2009 nearly 71,000 Americans will be diagnosed with bladder cancer and over 14,000 will die of the disease (SEER Database). With few treatment advances we studied the Ral GTPases as possible therapeutic targets, finding that the Ral paralogs are necessary for cell migration, anchorage independent growth, and metastasis (Gildea et al., Cancer Res. 2002 Feb 15;62(4):982-5)(unpublished data). To investigate possible transcription factors downstream of Ral in the bladder cancer cell line UMUC3 we performed gene expression profiling on cells with siRNA to both Ral paralogs. Computational analysis identified a statistical over representation of RREB1 binding sites in the promoters of differentially expressed genes. Further analysis confirmed RREB1 to be a novel transcriptional effector of Ral (Oxford et al., Oncogene. 2007 Nov 1;26(50):7143-52.) However, the role of RREB1 in bladder cancer has yet to be studied. Genetic depletion of RREB1 in the bladder cancer cell lines UMUC3, KU7 and J82 show a statistically significant decrease in growth in vitro. Furthermore, we identified two previously unreported RREB1 isoforms in addition to the two identified in NCBI genbank. Stable expression of each RREB1 isoform individually in the UMUC3 cell line resulted in significantly faster growth rates. Our data is consistent with the recent identification of RREB1 as an oncogene in the Moloney Murine Leukemia Virus (MuLV) tumor model (Uren et al., Cell 2008;133:727-41.) Our preliminary findings suggest that RREB1 is necessary for the survival of bladder cancer cell lines.

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 3090.
Abstract 3093: Overexpression of transcriptional coactivator AIB1 promotes hepatocellular carcinoma progression by enhancing cell proliferation and invasiveness
Authors: Xu Yixiang , Chen Qiang , Li Wengang , Su Xinhui , Chen Tenghui , Liu Yonghong , Zhao Yang , O'Malley Bert W. , Yu Chundong
Published: February 2011

Amplified in breast cancer 1 (AIB1; SRC-3) is a transcriptional coactivator for nuclear receptors and other transcription factors. AIB1 has been identified to play an important role in malignancy of several cancers such as breast and prostate cancers; however, its involvement in the progression of hepatocellular carcinoma (HCC) remains unclear. The aim of this study is to investigate the expression and possible role of AIB1 in HCC. Western blot analysis revealed that AIB1 was significantly increased in 15 of 22 HCC samples (68%), whereas only trace amounts of AIB1 were detected in the most matched nontumorous liver tissues. Functional studies on HCC cells demonstrated that AIB1 specific small interfering RNA mediated down-regulation of AIB1 resulted in decreased cell proliferation rate, cell migration/invasion ability, colony formation ability, and tumorigenic potential in nude mice. Marked increase of cell cycle inhibitor p21Cip1/Waf1 expression and dramatic decrease of the expression of phosphorylated Akt, proliferating cell nuclear antigen (PCNA), and matrix metallopeptidase MMP-9 in AIB1 knockdown cells contributed to these phenotypic changes. Consistent with these results, clinical AIB1-positive HCC exhibited higher levels of PCNA protein expression compared with AIB1-negative HCC. Further statistical calculation revealed a positive and tight correlation between the levels of AIB1 protein and PCNA protein in HCC, indicating that AIB1 contributes significantly to HCC cell proliferation. Conclusion: our results demonstrate that AIB1 promotes the progression of human HCC by enhancing cell proliferation and invasiveness, and could be a potential therapeutic target molecule for HCC treatment.

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 3093.
Abstract 310: The role of c-Jun N-terminal kinase 2alpha in non-small cell lung carcinoma tumorigenesis
Authors: Nitta Ryan T. , Chu Albert , Mitra Siddhartha , Godwin Andrew , Wong Albert
Published: February 2011

c-Jun N-terminal kinases (JNKs) are members of the mitogen activated protein kinase (MAPK) family and have been implicated in tumorigenesis. One isoform in particular, JNK2alpha, has been shown to be frequently activated in primary brain tumors, to enhance several tumorigenic phenotypes, and to increase tumor formation in mice. It was reported that there is frequent activation of JNK in non-small cell lung carcinoma (NSCLC). In this study, we investigated the role of the JNK2alpha isoform in NSCLC formation by examining its expression in primary tumors and by modulating its expression in cultured cell lines. We discovered 70% of the tested primary NSCLC tumors had 2 to 3-fold higher JNK2alpha protein and mRNA expression compared to normal lung tissue indicating a possible role of JNK2alpha in NSCLC tumorigenesis. To determine the importance of JNK2alpha in NSCLC progression, we reduced JNK2α in multiple NSCLC cell lines using short hairpin RNA. Cell lines deficient in JNK2alpha had decreased cellular growth and anchorage-independent growth, and the tumors were 4-fold smaller in mass. To elucidate the mechanism by which JNK2alpha induces NSCLC growth, we analyzed the JNK substrate, STAT3. Our data show that JNK2alpha can regulate the transcriptional activity of STAT3 by phosphorylating the Ser727 residue of STAT3 thereby regulating expression of oncogenic genes, such as c-Myc protein. Our studies revealed a novel mechanism in which phosphorylation of STAT3 is mediated by a constitutively active JNK2 isoform, JNK2alpha.

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 310.
Abstract 3100: The RNA-binding protein insulin-like growth factor 2 mRNA-binding protein 3 is oncogenic in liposarcoma
Authors: Angeles Christina V. , Hafner Markus , Socci Nicholas D. , DeCarolis Penelope , Tuschl Thomas , Singer Samuel
Published: February 2011

Introduction: The insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3) is an oncofetal mRNA binding protein that controls migration and proliferation through post-transcriptional regulation of target genes. It is aberrantly expressed in various cancers. We sought to determine the alterations in IGF2BP3 expression and copy number in well-differentiated (WD) and dedifferentiated (DD) liposarcoma (LS) and elucidate its role in driving oncogenesis in WD and DD LS cell lines.

Methods: U133A microarray analysis was performed on 61 DD and 52 WD LS as well as 17 normal fat (NF) tissue samples. CGH Agilent arrays were used to determine IGF2BP3 amplification status. An shRNA lentiviral system was used to knockdown (KD) IGF2BP3 in DD (DDLS8817, LPS141) and WD (RWD3051) cell lines. Cell proliferation, apoptosis, and mRNA expression were analyzed by Cyquant, annexin V, and qRT-PCR, respectively.

Results: CGH analysis showed IGF2BP3 amplification in 19% of DDLS (n=36) and 5% of WDLS (n=36) tumor samples. On U133A analysis IGF2BP3 was 2.5-fold and 1.2-fold upregulated in DDLS and WDLS, respectively, compared to NF. IGF2BP3 was overexpressed in 49% of DDLS and 33% of WDLS tumor samples. Gene expression was increased in DDLS8817 (3-fold), LPS141 (6-fold) and RWD3051 (6-fold) compared to adipose-derived stem cells. IGF2BP3 KD reduced expression >95%(p<0.05) compared to empty vector control for each cell line. IGF2BP3 KD decreased proliferation by 43%, 31%, and 36%(p<0.001) in DDLS8817, LPS141 and RWD3051, respectively, compared to control. Apoptosis was increased by 34%, 55%, and 58%(p<0.0001) compared to control in DDLS8817, LPS141, and RWD3051, respectively, and correlated with level of IGF2BP3 upregulation. p27, a cell cycle inhibitor, was identified as a potential target of IGF2BP3 by photoreactive crosslinking and immunoprecipitation. On Western blot analysis p27 was increased, cyclin D1 was decreased, and p53 was decreased in IGF2BP3 KD compared to control in LPS141, consistent with the functional effects observed.

Conclusions: IGF2BP3 overexpression is essential for proliferation and survival of LS cells. The cell cycle regulator, p27 was identified as an IGF2BP3 target mediating these functional effects. Identification of additional IGF2BP3 targets may lead to new therapeutic options for patients with DD and WD liposarcoma.

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 3100.
Abstract 311: Phosphorylation of RalB by PKC regulates its intracellular distribution and promotion of bladder tumor growth
Authors: Wang Hong , Wang Xuejiao , Owens Charles , Chandra Nidhi , Conaway Mark R. , Brautigan David L. , Theodorescu Dan
Published: February 2011

RalA and RalB are members of the Ras family of monomeric G proteins that are 83% identical in amino acid sequence, but have paralog-specific effects on cancer cell proliferation, metastasis and apoptosis. Using in vitro kinase assays and phosphosite specific antibodies, here we show paralog-specific phosphorylation of RalA by PKA and RalB by PKC. Mutagenesis identified Ser194 as the single PKA phosphorylation site in RalA and S198 as the primary PKC phosphorylation site in RalB. Phorbol ester treatment of human bladder carcinoma cells induced phosphorylation of S198 in both stably expressed FLAG-RalB and endogenous RalB. Compared to activated RalB(G23V) that localized at plasma membrane, phosphosite mutant RalB(G23V-S198A) colocalized with GFP-Rab11, a recycling endosome marker. Phorbol ester treatment enhanced anchorage independent growth of UMUC3 human bladder carcinoma cells stably transfected with wild type RalB or activated RalB(G23V), but not with phosphosite mutants RalB(S198A) or RalB(G23V-S198A). Furthermore, UMUC3 cells transfected with RalB(G23V) exhibited enhanced subcutaneous tumor growth compared to vector transfected cells, but cells expressing RalB(G23V-S198A) had growth indistinguishable from the controls. Our data demonstrate that RalA and RalB are specifically phosphorylated by different kinases, and RalB phosphorylation regulates its intracellular localization and in vivo tumor growth promotion. Phosphorylation of RalA and RalB by different kinases may contribute to the biological specificity of these closely related proteins.

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 311.
Abstract 3111: The ETS factor ETV1 is an oncogene in melanoma
Authors: Jane-Valbuena Judit , Widlund Hans , Perner Sven , Johnson Laura , Dibner Aurora , Lin William , Baker Alissa , Nazarian Rosalynn , Vijayendran Krishna , Sellers William , Hahn William , Duncan Lyn , Rubin Mark , Fisher David , Garraway Levi A.
Published: February 2011

Chromosome 7 copy gains are a recurrent genomic alteration in melanoma. Previous genomic studies identified a common peak of gain within 7p, highlighting it as a region likely to contain “driver” cancer genes. In this study, we identified several melanoma samples with focal amplification events located within this peak, and found that the overlapping region of these amplicons contained the locus of the ETS transcription factor and oncogene, ETV1. Fluorescent in situ hybridization analysis of a progressive melanoma tissue microarray revealed ETV1 amplification in 13% of the primary tumors and 18%of the metastatic melanomas, whereas no amplification in nevus samples. Silencing of ETV1 expression demonstrated dependency on ETV1 expression for proliferation and anchorage-independent growth in some melanoma cell lines. Moreover, over-expression of ETV1 in combination with oncogenic NRASG12D transformed primary melanocytes and promoted tumor formation in mice. Interestingly, overexpression of ETV1 increased MITF expression, a known oncogene in melanoma. Together, these observations suggest an oncogenic role for ETS transcription factors in general and ETV1 in particular in melanoma.

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 3111.
Abstract 3119: Phosphorylation of epidermal growth factor receptor at Ser1046/1047 by p38 mitogen-activated protein kinase plays a critical role in its downregulation in cancer cells
Authors: Adachi Seiji , Nakashima Masanori , Yamauchi Takahiro , Shirakami Yohei , Shimizu Masahito , Yasuda Ichiro , Moriwaki Hisataka , Kozawa Osamu
Published: February 2011

The mechanism underlying downregulation of EGF receptor (EGFR) is important for the regulation of cancer cells. Although the phosphorylation of EGFR at Tyr1045 and Ser1046/1047 reportedly account for its downregulation, the exact mechanism remains unclear. We recently reported that p38 mitogen-activated protein kinase (MAPK), a stress-inducible kinase believed to negatively regulate tumorigenesis, controls EGFR downregulation via its phosphorylation at Ser1046/1047 (Cancer Lett 2009;277:108-13). To further investigate the role of phosphorylation of p38 MAPK in the action of anti-cancer drug, we examined the effects of (−)-epigallocatechin gallate (EGCG) and several HSP90 inhibitors (17-AAG, geldanamycin and 17-DMAG) on p38 MAPK in colon and pancreatic cancer cells. While EGCG and HSP90 inhibitors caused downregulation of EGFR and subsequent cell death, they induced phosphorylation of p38 MAPK, and inhibition of p38 MAPK suppressed the internalization and subsequent degradation of EGFR. Moreover, we found that they caused phosphorylation of EGFR at Ser1046/1047, not Tyr1045, and that they were also suppressed by the inhibition of p38 MAPK. Taken together, our results strongly suggest that phosphorylation of EGFR at Ser 1046/1047 via activation of p38 MAPK plays a pivotal role in anti-cancer drug-induced downregulation of EGFR. These findings may indicate new therapeutic strategies for inhibiting the proliferation of human cancers that are highly dependent on the activation of EGFR. (Adachi S et al., Carcinogenesis 2009;30:1544-52)

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 3119.
Abstract 312: P190-B RhoGAP overexpression disrupts mitosis, alters cell polarity, and increases adhesion and invasion in mammary epithelial cells
Authors: McHenry Peter R. , Park Clara , Chang Peggy , Zeng Erliang , Sears James , Vargo-Gogola Tracy
Published: February 2011

Breast cancer causes approximately 40,000 deaths each year in the United States alone. Collective epithelial invasion is an important process in tumor progression, and it also occurs in the normal mammary gland (MG) through specialized structures called terminal end buds (TEBs) that proliferate and migrate to drive ductal outgrowth during MG development. P190-B RhoGAP is highly expressed in TEBs, suggesting it may regulate the invasive process of MG development. Indeed, tetracycline-inducible p190-B overexpression causes perturbations in TEB morphology, stromal composition, and MG branching morphogenesis. In the current study, we analyzed the gene expression patterns of MECs isolated from p190-B overexpressing mice to determine how p190-B regulates epithelial morphogenesis as well as stromal-epithelial interactions. We investigated the expression patterns of p190-B and investigated the effects of overexpression in MCF-7 breast cancer cells. We further investigated the effects of p190-B overexpression on primary MECs in a three-dimensional (3D) culture MEC morphogenesis assay in reconstituted basement membrane (rBM) and in adhesion assays. Microarray results demonstrated that a large number of genes involved in regulating mitotic cell division and the PI3K pathway of proliferation/survival signaling are altered in p190-B overexpressing primary MECs. Real-time PCR confirmed a number of genes are significantly up- or down-regulated in these pathways. Consistent with these findings, p190-B localizes to the mitotic spindle and kinetochores during metaphase of mitosis and to the midbody at cytokinesis. Furthermore, overexpression of p190-B caused abnormal centrosome formation, multipolar spindles, failed cytokinesis, and multinucleation in MCF-7 cells. In addition, primary MECs overexpressing p190-B demonstrated increased adhesion to laminin and type IV collagen, components of the basement membrane, and primary MECs overexpressing p190-B cultured in rBM showed increased disrupted acinar formation, loss of cell polarity, and invasion. P190-B overexpression alters cell polarity and mitosis and regulates adhesive and invasive properties in MECs. These phenotypes may be critical for TEB structure and function during MG branching morphogenesis. We propose that p190-B overexpression may contribute to tumorigenesis by disrupting mitosis and promoting invasion.

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 312.
Abstract 3120: Aberrant activation of ERBB3 oncogenic signaling in human hepatocellular carcinoma
Authors: Hsieh Sen-Yung , He Jung-Ru , Hsu Chih-Yun , Yu Ming-Chin , Weng Wen-Hui , Huang Shiu-Fen
Published: February 2011

Background & aims: Recent evidence that ERBB3 is responsible for tumor resistance to EGFR or ERBB2 targeted therapies highlighted its crucial role in the oncogenic signaling of EGFR family. However, its roles in hepatocelluar carcinoma (HCC) remain unclear. Methods: Expression and phosphorylation of ERBB3 was examined in HCC tissues and cells. Gene number was determined using fluorescence in situ hybridization. Roles of EGEF, ERBB2, and neuregulin in phosphorylation of ERBB3 were evaluated via small-interference RNAs to silence their expression. The effects of ERBB3 on cell proliferation, invasion and tumorigenesis were assayed in vitro and in vivo. Results: Overexpression of ERBB3 was associated with a higher recurrence (P=0.000), and a worse prognosis (P=0.004). Gain of ERBB3 gene number was found in most HCC cells. EGFR, ERBB2, neuregulin and phosphorylated ERBB3 were detected in HCC cells. Phosphorylation of ERBB3 was induced by treatment with the conditioned media of HCC cells, while it was abolished by pre-treatment of the conditioned media with the anti-neuregulin antibodies or by silencing neuregulin expression. Phosphorylation of ERBB3 was suppressed by silencing the expression of ERBB2 but not EGFR. Induction of ERBB3 phosphorylation promoted cell proliferation, invasion and colony formation, while silencing ERBB3 expression suppressed xenograft tumor formation and growth in nude mice. Conclusions: Overexpression of ERBB3, primarily attributable to gene amplification, was associated with a poor prognosis of HCC. Aberrant activation of ERBB3, via an autocrine mechanism by dimerization with ERBB2, enhanced tumorigenesis in vitro and in vivo. The oncogenic signaling of ERBB3 can be the target for anti-HCC 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 3120.