bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3114: MLL2 overexpression and incomplete proteolytic processing in human cell lines from invasive breast and colon cancers
Authors: Natarajan Thanemozhi G. , Kallakury Bhaskar V. , Sheehan Christine , Bartlett Margaret B. , Ganesan Natarajan , Preet Anju , Ross Jeffrey , FitzGerald Kevin T.
Published: February 2011

Background: MLL2 is an epigenetic regulator in mammalian cells and mediates histone 3 lysine 4 tri-methylation (H3K4me3) at specific gene loci, through formation of a multiprotein complex. MLL2 shares a high degree of structural similarity with MLL1, which frequently undergoes translocations resulting in leukemogenic fusion oncoproteins. Despite the structural similarity with MLL1, MLL2 is not involved in leukemias. Earlier we reported aberrant MLL2 expression in human breast and colonic tumor tissues compared to normal tissues. In continuation with the above study, we now present the results of MLL2 investigation in cell lines from human breast and colon cancers.

Methods: Established cell lines from human mammary and colonic tissues were cultured under optimal conditions. Cells were processed for nuclear and cytoplasmic fractionation as well as DNA and total RNA extraction. Nuclear and cytoplasmic extracts were separated on polyacrylamide gels and immunoblotted for MLL2. The RNA extracts were used to determine MLL2 transcript levels by one-step reverse transcriptase-PCR using appropriate primers. The DNA extracts were used to amplify MLL2 specific sequences using appropriate primers, followed by sequencing reaction.

Result: Examination of cell lines revealed 2-3 fold increase in MLL2 transcript levels in the highly invasive tumor cell lines of both breast and colon. Expression of MLL2 at protein level was consistent with the transcript levels, indicating MLL2 overexpression as the cause of increased protein levels in the tumor cells. Additionally, we also observed incomplete proteolytic processing of MLL2 in the highly invasive cell lines from either tissue types, compared to the corresponding non-tumor/less invasive tumor cell lines. Interestingly, the unprocessed full length MLL2 was significantly abundant in the nuclear compartments of the highly invasive tumor cell lines. Analysis of the coding sequences for the putative cleavage site did not reveal any alterations.

Conclusion: We conclude that MLL2 expression and proteolytic processing are both deregulated in breast and colon cancers. This deregulation may be associated with mammary and colonic tumor growth/progression. Given the epigenetic regulatory role, further investigation is required to study MLL2 involvement in tumorigenesis.

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 3114.
Abstract 3115: Ephrin-dependent and -independent activities of the receptor tyrosine kinase EphB4
Authors: Yang Nai-Ying Michelle , Hooten Nikki Noren , Pasquale Elena B.
Published: February 2011

Recent evidence has shown that the EphB4 receptor tyrosine kinase is upregulated in various types of cancer. However, it is unclear how the EphB4 receptor may contribute to oncogenesis because its activation by its preferred ligand, ephrin-B2, appears to be low in most cancer cell types. We have found that activation of EphB4 in breast cancer cells by stimulation with exogenous ephrin-B2 Fc ligand activates a tumor suppressor pathway that inhibits cell growth and migration. Our laboratory has also recently demonstrated an ephrin-independent activity of EphB4 in breast cancer cells, which may play a role in the tumors where the ligand is absent. By using RNA interference to downregulate EphB4 in MCF-7 and MDA-MB-435 cells, we found that EphB4 inhibits integrin-mediated substrate adhesion, spreading and migration, and reduces β1-integrin protein levels. Through these effects, EphB4 expression would be expected to inhibit tumor development and growth. Ephrin binding does not appear to be important for this EphB4-dependent regulation of β1-integrin levels. Here, we show that EphB4 knock down also dramatically increases β1-integrin protein levels in PC3 prostate cancer cells. Stimulation with a soluble form of the ligand, ephrin-B2 Fc, also reduces EphB4 expression. Therefore, experiments are in progress to determine if EphB4 downregulation by ligand also results in decreased β1-integrin protein levels, besides the inhibition of integrin-mediated cell adhesion induced by EphB4 ligand-dependent signaling. Experiments are also in progress to investigate the mechanism by which EphB4 regulates integrin levels, and whether it involves regulation of mRNA levels or protein stability. Interestingly, a recent report shows that the decreased expression of EphB4 in intestinal tumors from APCmin/+ mice that lack one EphB4 allele causes increased transcription of genes involved in cell attachment to the extracellular matrix and invasion (Dopeso et al., 2009, Cancer Res 69:7430). However, an increase in β1-integrin mRNA level was not detected in this model. Therefore, it will be interesting to investigate whether EphB4 regulates integrins differently in different cancer cell types, the mechanisms involved, and the effects on cell migration and 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 3115.
Abstract 3116: Serine protease inhibitor Kazal type 1 plays a role of colon carcinogenesis through the epidermal growth factor receptor
Authors: Ida Satoshi , Ohmuraya Masaki , Tanaka Hiroshi , Watanabe Masayuki , Takamori Hiroshi , Beppu Tooru , Baba Hideo
Published: February 2011

Serine protease inhibitor Kazal type 1 (SPINK1: mouse homolog Spink3), which is also known as tumor associated trypsin inhibitor (TATI), is mainly produced in the acinar cells. The role of SPINK1 has been postulated to be the prevention of inadvertent proteolysis in the pancreas caused by intra-acinar premature activation of trypsinogen. SPINK1 is expressed not only in normal human pancreatic acinar cells, but also in a various cancers, including colon cancer. In the colon cancer, previous reports said high expression of SPINK1 correlates with metastasis and poor prognosis. However, the role of SPINK1 in malignant tumors remains to be elucidated. We previously demonstrated that SPINK1/Spink3 can bind to EGFR and work as a growth factor through EGFR in the pancreatic cancer, even the chronic pancreatitis. Hence, we hypothesized that SPINK1 plays a role as the growth factor for tissue repair in the inflammatory state, and if prolonged, it works to carcinogenesis in the colon tissue. We first showed that SPINK1 induced proliferation of colon cancer cell lines examined. To clarify the role in the colon cancer of SPINK1 in vivo, we use an efficient animal model for colitis-related carcinogenesis, male C57BL/6 mice were given a single intraperitoneal administration of azoxymethane (AOM), followed by a 1-week oral exposure to dextran sodium sulfate (DSS). We showed that expression of EGFR and Spink3 (its ligand) were observed in DSS induced colitis state. Sixteen weeks after injected AOM, multiple colon cancer occurred in all treated mice. In the Western blot, EGFR and Spink3 overexpressed in the whole colon tissue (cancer tissue and non-cancerous lesion), but Spink3 slightly decreased in the cancer. To determine whether Spink3 has an essential role of carcinogenesis, we further analyzed the tumor incidence of Spink3+/−mice. At 16 weeks, tumor number was 4.6 ± 0.8 (mean ± standard error) in Spink3+/+ mice and 1 ± 0.4 in Spink3+/− mice, and a significant difference was observed between the 2 groups (p<0.05). To further analyze the clinical importance of SPINK1 in the development of colon cancer, we examined the expression of SPINK1 and EGFR in adenoma and adenocarcinoma. SPINK1 express in adenoma and well differentiated adenocarcinoma, but not expressed in poorly differentiated adenocarcinoma. Taken together, these results suggest that SPINK1 stimulates the proliferation of colon cancer cells through the EGFR, and especially role in early carcinogenesis in colon 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 3116.
Abstract 3117: COX-2 is a novel transcriptional target of the nuclear EGFR-STAT3 and EGFRvIII-STAT3 signaling axes
Authors: Lo Hui-Wen , Cao Xinyu , Zhu Hu , Ali-Osman Francis
Published: February 2011

Emerging evidence indicate novel modes of EGFR signaling, notably, one involves EGFR nuclear translocalization and subsequent gene activation. To date, however, the significance of the nuclear EGFR pathway in glioblastoma (GBM), the most frequent and deadliest brain malignancy in adults, is unknown. Here, we report that EGFR and its constitutively activated variant EGFRvIII undergo nuclear translocalization in GBM cells, in which the former event requires EGF stimulation and the latter is constitutive. To gain insight into the impact of nuclear EGFR on gene expression in GBM, we created isogenic GBM cell lines, namely, U87MG-vector, U87MG-EGFR and U87MG-EGFRdNLS that, respectively, express the control vector, EGFR and nuclear-entry defective EGFR with a deletion of the nuclear localization signal (NLS). Microarray analysis shows that 19 genes, including, cyclooxygenase-2 (COX-2), to be activated in U87MG-EGFR cells but not in U87MG-EGFRdNLS and U87MG-vector cells. Subsequent validation studies indicate that COX-2 gene is expressed at higher levels in cells with EGFR and EGFRvIII than those with EGFRdNLS and EGFRvIIIdNLS. Nuclear EGFR and its transcriptional co-factor STAT3 associate with the COX-2 promoter. Increased expression of EGFR/EGFRvIII and activated STAT3 leads to synergistic activation of the COX-2 promoter. Promoter mutational analysis identified a proximal STAT3-binding site that is required for EGFR/EGFRvIII-STAT3 mediated COX-2 gene activation. In GBM tumors, an association exists between levels of COX-2, EGFR/EGFRvIII and activated STAT3. Together, these findings indicate the existence of the nuclear EGFR/EGFRvIII signaling pathway in GBM and its functional interaction with STAT3 to activate COX-2 gene expression, thus linking EGFR-STAT3 and EGFRvIII-STAT3 signaling axes to pro-inflammatory COX-2 mediated pathway.

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 3117.
Abstract 3118: Platelet-derived growth factor-D contributes to aggressiveness of breast cancer cells by up-regulation of Notch and NF-κB signaling pathways
Authors: Ahmad Aamir , Kong Dejuan , Wang Zhiwei , Ali Raza , Banerjee Sanjeev , Sarkar Fazlul H.
Published: February 2011

Platelet-derived growth factor-D (PDGF-D) has been suggested to be associated with progression of several human malignancies; however, its role in breast cancer progression has not been investigated. We found that PDGF-D expressing breast cancer cell lines MDA-MB-231 and SUM-149 are more aggressive compared to the cell lines with little or no expression of PDGF-D, such as MDA-MB-468 and MCF-7 cells. In order to test our hypothesis that PDGF-D influences cell proliferation, invasion and metastasis mediated by the activation of Notch and NF-κB signaling, we over-expressed PDGF-D in MDA-MB-468 and MCF-7 by cDNA transfection as well as silenced it in MDA-MB-231 and SUM-149 cells using PDGF-D siRNA, for mechanistic studies. We found that PDGF-D over-expression increased cell proliferation while its silencing resulted in decreased cell proliferation and increased induction of apoptosis. PDGF-D expression correlated with the activated expression of several members of Notch-signaling, namely, Notch-1, Notch-4 and Jagged-1, and attenuation of PDGF-D levels led to the inactivation of Notch signaling. PDGF-D over-expression was also found to induce the expression of mesenchymal markers (Vimentin and ZEB-2) concomitant with decreased expression of epithelial marker E-cadherin, which appears to be one of the mechanism by which PDGF-D may contribute to the aggressiveness of breast cancer cells. Since NF-κB plays a crucial role in Notch signaling as well as in the processes of epithelial-mesenchymal transition (EMT), we studied the effect of PDGF-D signaling on the DNA binding activity of NF-κB. We found that PDGF-D over-expression led to increased expression of Notch-1 and increased the DNA binding activity of NF- κB whereas the silencing of PDGF-D reduced the expression of Notch-1 and also reduced the DNA binding activity of NF-κB, suggesting that PDGF-D is upstream of Notch and NF-κB signaling pathways. These results were also consistent with the expression and activity of MMP-9 and VEGF, as well as invasive characteristics. From these results, we conclude that PDGF-D plays an important role in breast tumor aggressiveness and this process is mechanistically linked with Notch-1 and NF-κB signaling.

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 3118.
Abstract 3121: A novel molecular mechanism for HER2-induced sustained signaling and transformation
Authors: Zhou Xiandong , Ahmed Naushin , Agazie Yehenew M.
Published: February 2011

HER2 overexpression in breast and other tumors leads to an aggressive disease state and poor prognosis for patient survival. It is believed that increased cell growth and survival signaling induced by HER2 is responsible for the aggressive disease state and unfavorable outcome. The peculiarity of HER2 among the four EGFR family members is that even a modest expression leads to cell transformation, while similar expression of the other members does not. But how HER2 overcomes the receptor regulatory mechanism to induce constitutive signaling and cell transformation is virtually unknown. Here, we show that HER2 induces distinctive EGFR autophosphorylation that disfavors c-Cbl and RasGAP, but favors Gab1/2, PLCγ, SHP2 and PI3K interactions. We further show that HER2 expression leads to reduced interaction of Grb2 with EGFR-HER2 complexes, an effect that further reduces the interaction c-Cbl. The inefficient interaction of c-Cbl confers receptor resistance to EGF-induced degradation, while the reduced interaction with RasGAP leads to an increase in the half live activated Ras. Therefore, the molecular mechanism by which HER2 confers sustained signaling and cell transformation is by inducing distinctive receptor autophosphorylation that favors receptor recycling and prolon

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 3121.
Abstract 3126: Tumor heterogeneity in glioblastoma is an active process driven by a mutant EGFR-induced paracrine circuit
Authors: Inda Maria-del-Mar , Bonavia Rudy , Sah Dinah WY , Vandenberg Scott , Brennan Cameron , Johns Terrance G. , Hadwiger Philipp , Tan Pamela , Cavenee Webster K. , Furnari Frank B.
Published: February 2011

Amplification and mutation of the epidermal growth factor receptor gene (EGFR), resulting in the common and oncogenic EGFRvIII (ΔEGFR) variant, is a signature pathogenetic event in GBM. Strikingly, despite its greater biological activity than wildtype EGFR (wtEGFR), only a minority of cancer cells in the primary tumor possess the hallmark ΔEGFR lesion, while the remainder of cancer cells express wtEGFR. We hypothesized that the ΔEGFR-expressing subpopulation provides enhanced tumorigenicity to the entire tumor cell population through a paracrine mechanism. Using a combination of mixed tumor engraftments and biochemical analysis of paracrine factors and signaling pathways activation, we determined that human glioma tissues, glioma cell lines, glioma stem cells and primary mouse astrocytes, that express ΔEGFR each secrete IL-6 and/or LIF cytokines. This then prompts a novel interaction between the receptor that is common to these cytokines, gp130, and wtEGFR in neighboring cells that express amplified levels of EGFR, resulting in co-receptor activation and tumor growth enhancement. Ablating IL-6, LIF or gp130 uncouples this cellular cross-talk and potently attenuates tumor growth enhancement. These findings demonstrate that the heterogeneity which characterizes GBM, and perhaps other tumors with this feature, does not occur stochastically, but instead can be an actively maintained feature and illuminates a heterotypic cancer cell interaction of potential therapeutic significance.

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 3126.
Abstract 3128: mTOR expression and activity patterns in gastroenteropancreatic neuroendocrine tumors
Authors: Kasajima Atsuko , Pavel Marianne , Darb-Esfahani Silvia , Noske Aurelia , Stenzinger Albrecht , Sasano Hironobu , Dietel Manfred , Denkert Carsten , Röcken Christoph , Wiedenmann Bertram , Weichert Wilko
Published: February 2011

Recently, novel small molecules targeting the mammalian target of rapamycin (mTOR) have been successfully tested for the treatment of endocrine neoplasias. Since these drugs will certainly be used as routine therapeutics in the near future, it is surprising that information on the exact expression patterns of mTOR and its downstream target 4EBP1 in endocrine tumors is still lacking.

We investigated expression of mTOR and total as well as phosphorylated 4EBP4 in specimen of endocrine tumors by immunohistochemistry and correlated our findings with clinicopathologic variables and patient prognosis.

We found that 61 (61.6%), 92 (92.2%), 79 (79.8%) and 69 (69.7%) out of 99 neuroendocrine foregut and midgut tumors were positive for mTOR, 4EBP1, cytoplasmic phospho-4EBP1 (p-4EBP1) and nuclear phospho-4EBP1, respectively. mTOR, 4EBP1 and p-4EBP1 (either cytoplasmic or nuclear) expression was higher in foregut tumors than in midgut tumors. Expression of mTOR was higher in foregut tumors with distant metastases (p=0.035) when compared to their locally restricted counterparts. 4EBP1 as well as p-4EBP1 (either cytoplasmic or nuclear) expression was associated with higher Ki-67 positivity and thereby proliferative activity.

mTOR expression and activity as measured by phosphorylation of its downstream target 4EBP1 vary considerably in dependence of tumor location and metastatic status. We hypothesize that differences in mTOR expression patterns and/or activity might predict response to mTOR inhibitors which should be taken into account when future clinical trials with these substances are being planned.

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 3128.
Abstract 3131: IGF-I regulates Cyr61-induced breast cancer cell growth and migration
Authors: Sarkissyan Suren , Sarkissyan Marianna , Wu Yanyuan , Koeffler H Phillip , Vadgama Jaydutt V.
Published: February 2011

Cysteine-rich 61 (Cyr61) is a protein involved in wound healing, regulation of extracellular matrix microenvironment, and in tumor invasion. Data from our laboratory and others have shown that highly invasive cancer cells have high Cyr61 transcriptional levels as compared with non-invasive cells. Recent studies from our laboratory have shown that IGF-I (insulin growth factor I) is involved in activating the PI3/Akt pathway, and this may increase Cyr61 expression. We have hypothesized that IGF-I induces tumor invasion by upregulating Cyr61 through Akt phosphorylation (pAkt). In the current study, several breast cancer cell lines, MDA-MB231, BT474, MCF-7, SKBR3 were used; two of which were stably transfected with active Akt1(SKBR3/AA and MCF-7/AA) and dominant negative Akt1 (SKBR3/DN and MCF-7/DN). To determine Cyr61 expression in response to IGF-I, cells were treated with IGF-I and Cyr61 expression was examined by RT-Real time PCR. To determine cell invasiveness in relation to Cyr61 expressions and IGF-I induction, a migration assay was used. Our result showed that Cyr61 expression was significantly higher in MDA-MB231 cells. SKBR3/AA and MCF-7/AA had increased levels of Cyr61, while SKBR3/DN and MCF-7/DN had decreased levels of Cyr61 compared to SKBR3 and MCF-7. Furthermore, IGF-I (100ng/ml) increased Cyr61 expression significantly and cells with high level of Cyr61 increased invasiveness. In summary, active Akt was associated with the IGF-I induced Cyr61, which caused breast cancer cell migration and invasion. Hence, Cyr61 may be a potential therapeutic target for treatment of invasive breast tumors.

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 3131.
Abstract 3132: Proliferative effects of FGF1 on ovarian cancer cells correlate with FGFR4 over expression and are mediated by MAPK/ERK signaling
Authors: Zaid Tarrik , Thompson Melissa S. , Wong Kwong-kwok , Yeung Tsz-Lun Yeung , Yeung Zachary , Yates Melinda S. , Birrer Michael J. , Mok Samuel C.
Published: February 2011

Objectives: To identify the effect of FGF1 on cell proliferation in ovarian cancer and endothelial cells, its relation to FGFR4 over expression and signaling pathways involved

Methods: Amplification of FGFR4 on 5q31-35, which was previously shown to be associated with poor survival in patients with advanced stage high-grade serous ovarian cancer (HGSC), was validated by quantitative PCR in 52 HGSC. Association between DNA copy numbers of FGFR4 and FGF1 located on the same segment was performed by Spearman correlation analysis. Over expression of FGFR4 protein was confirmed in ovarian cancer cell lines and tissue by western blot analysis and immunohistochemistry, respectively. Endogenous FGF1 production by several ovarian cancer cell lines in conditioned medium was quantified using ELISA. Effect of exogenous FGF1 on cell proliferation of high and low-FGFR4 expressing ovarian cancer cell lines and an immortalized microvascular endothelial cell line was determined by WST-1 assay. Signaling pathways activation was screened by treating ovarian cancer cell lines and endothelial cells transduced with six different transcription response elements using the Cignal-Lenti reporter system with exogenous FGF1.

Results: FGFR4 was amplified and over-expressed in a majority of ovarian cancer cell lines and tumor tissue and FGFR4 DNA copy number was significantly correlated with that of FGF1 (r=0.4, p=0.04). Endogenous FGF1 production was detected by ELISA in 2 of 5 screened ovarian cell lines. FGF1 significantly induced cell proliferation in high FGFR4 expressing ovarian cancer cells (20%, p<0.001) and in endothelial cells (15%, p<0.001); but not in low FGFR4 expressers. Significant activation of MAPK/ERK signal transduction pathways was noted after treatment of ovarian(1.43 fold, P < 0.001) and endothelial(1.33 fold, P < 0.001) cell lines transduced with the Cignal-Lenti reporter system.

Conclusions: Our report provides preliminary evidence that poor survival associated with FGF1 amplification and over expression appears to be mediated through FGFR4, which is also amplified and over-expressed in ovarian cancer cell. In addition, FGF1 produced by ovarian cancer cells may exert both autocrine and paracrine effect on cancer cells and endothelial cells through the activation of MAPK/ERK pathway. These data suggests that new regimens targeting FGF1, FGFR4 and downstream signaling pathways may improve ovarian cancer patient survival.

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 3132.
Abstract 3135: Segregation of non-R132H mutations in IDH1 in distinct molecular subtypes of glioma
Authors: Bralten Linda B. , Gravendeel Lonneke AM , Kloosterhof Nanne K. , van Marion Ronald , Dubbink Hendrikus J. , Dinjens Winand , Bleeker Fonnet E. , Hoogenraad Casper C. , Michiels Erna , Kros Johan M. , van den Bent Martin , Smitt Peter AE Sillevis , French Pim J.
Published: February 2011

Mutations in the gene encoding the isocitrate dehydrogenase 1 gene (IDH1) occur at a high frequency (up to 80%) in many different subtypes of glioma. In this study, we have screened for IDH1 mutations in a cohort of 496 gliomas. IDH1 mutations were most frequently observed in low grade gliomas with 395G>A (R132H) representing >90% of all IDH1 mutations. Interestingly, non-R132H mutations segregate in distinct histological and molecular subtypes of glioma. Histologically, they occur sporadically in classic oligodendrogliomas and at significantly higher frequency in other grade II and III gliomas. Genetically, non-R132H mutations occur in tumors with TP53 mutation, are virtually absent in tumors with loss of heterozygosity on 1p and 19q and accumulate in distinct (gene-expression profiling based) intrinsic molecular subtypes. The IDH1 mutation type does not affect patient survival. Our results were validated on an independent sample cohort, indicating that the IDH1 mutation spectrum may aid glioma subtype classification. Functional differences between R132H and non-R132H mutated IDH1 may explain the segregation in distinct glioma subtypes.

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 3135.
Abstract 3136: Activated Src increases total Rac levels and requires Rac and IL6 for full neoplastic transformation
Authors: D'Abreo Carmeline , Arulanandam Rozanne , Geletu Mulu , Raptis Leda H.
Published: February 2011

Activated Src (vSrc) was previously shown to activate Rac through the exchange factors Vav2 and Tiam, and to require Rac for neoplastic transformation, but the exact mechanism remains unclear. Since Src requires the function of the Signal transducer and activator of transcription-3 (Stat3), a Rac target for transformation, we explored the potential role of the Rac/Stat3 axis in transformation by vSrc. In fact, we recently demonstrated a dramatic increase in the activity of Stat3 in breast carcinoma as well as normal epithelial cells and fibroblasts, as a consequence of cell to cell adhesion (Oncogene 23:2600).

In light of these findings, we revisited the question of the role of Rac and Stat3 in vSrc transformation. The results showed that, as previously demonstrated, vSrc expressing mouse fibroblasts had higher Rac activity, at all densities examined. Unexpectedly, our results also demonstrate for the first time a dramatic surge in total Rac upon Src expression, which was at least in part responsible for the Stat3 activity increase. Moreover, Rac activation caused an increase in levels of cytokines of the IL6 family, which was required for Stat3 activation, as shown by the fact that knockdown of gp130, the common receptor subunit of this family, reduced Stat3 activation and neoplastic transformation. Besides transformation, the Rac/IL6/Stat3 axis was required for apoptosis inhibition upon vSrc expression. (supported by CIHR, CBCF-Ontario chapter, US Army breast cancer program, NSERC and Breast Cancer Action Kingston).

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 3136.
Abstract 3138: Stat3 signaling in thyroid cancer
Authors: Silva Joana , Soares Paula , Trovisco Vitor , Knauf Jeffrey , Fagin James , Bromberg Jacqueline
Published: February 2011

In this study, we addressed the relevance and regulation of Stat3 activation in thyroid cancer. Papillary Thyroid Carcinoma (PTC) comprises almost 80% of thyroid neoplasias and 70% of PTC present activating alterations of components of the ERK-MAPK pathway, either BRAF mutations, RET/PTC rearrangements or RAS mutations. PTCs often display a strong desmoplastic reaction, where tumor is surrounded by a dense stroma and infiltrated with inflammatory cells. To examine the role of Stat3 in thyroid tumours and its putative relation to the associated inflammatory reaction, we analysed Stat3 activation (i.e. Y705-phosphorylation) by IHC in 58 samples of PTC. Approximately 50% of the tumors express pStat3. Frequently, nuclear positivity is only observed in the edge of the tumor as well as in the thyroid cells within tumor stroma. The inner part of the tumor is generally negative. Using multiple cell line models of PTC we observe that both RET/PTC and BRAFV600E expression can lead to Stat3 activation. Similarly, transgenic models of both BRAFV600E and RET/PTC also demonstrate high levels of pStat3 in particular on the inflammatory/leading edge of the tumor. We hypothesize that Stat3 activation, particularly in BRAFV600E tumors, may result from a paracrine effect of stromal cells-derived factors in the adjacent tumor cells. Stat3 is principally activated by autocrine/paracrine IL-6/Jak signaling in lung and breast tumors. Similarly, here we demonstrate that pan-Jak inhibitors block Stat3 activation in PTC cell lines. Furthermore, IL-6 signaling appears to mediate Stat3 activation in RET/PTC, BRAFV600E and RAS expressing cell lines. BRAFV600E knockdown led to a significant decrease of IL-6 levels, as well as of pStat3. In conclusion, we have found that Stat3 is activated in PTC, and that, particularly in BRAFV600E tumors, such activation may result from a paracrine effect of stromal cells through upregulation of IL-6/Jak pathway.

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 3138.
Abstract 3143: PKD as a downstream target for K-ras in pancreatic cancer
Authors: Storz Peter
Published: February 2011

Growth factor and K-ras-induced generation of reactive oxygen species have important roles in mediating pancreatic ductal adenocarcinoma. Pancreatic cancer cells also express high levels of active transcription factor Nuclear Factor-κB (NF-κB), which is responsible for cell survival and resistance to chemotherapeutics. The serine/threonine kinase Protein Kinase D1 (PKD1) is a sensor for oxidative stress and protects cancer cells by inducing anti-apoptotic and anti-oxidant genes. Here we provide evidence that PKD1 is the means of how high NF-κB activity levels are achieved in pancreatic cancer cells. We also show the contribution of K-ras, which is upregulated in its activity in over 90% of PDAC (pancreatic ductal adenocarcinoma). We found that K-ras in pancreatic ductal HPDE cells increases PKD1 activity and that the depletion of the pancreatic ductal adenocarcinoma cell line Panc-1 from K-ras leads to decreased PKD1 activity and NF-κB activation. As a mechanism we found that K-ras mediates activation of PKD1 via Src and that these signaling events relay to high NF-κB levels. Our data reveal K-ras as an activator of PKD1 and show that both oncogenes, Src and K-ras cooperate to activate NF-κB in PDAC through PKD1. This suggests the K-ras/Src/PKD1/NF-κB pathway as a new potential drug target for pancreatic 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 3143.
Abstract 3144: Tgfbr1 haploinsufficiency inhibits the development of murine mutant Kras-induced pancreatic precancer
Authors: Adrian Kevin , Strouch Matthew , Zeng Qinghua , Barron Morgan , Cheon Eric , Honasage Akilesh , Xu Yanfei , Phukan Sharbani , Sadim Maureen , Bentrem David , Pasche Boris , Grippo Paul
Published: February 2011

Abstract

Transforming Growth Factor Beta (TGFβ) signaling can play dual roles in pancreatic tumor development. In human pancreatic cancer, loss of DPC4/smad4 has been observed in more than half of all pancreatic cancers. In vivo loss of either Smad4 or Tgfbr2 promotes pancreatic adenocarcinoma in mice expressing mutant Kras. Conversely, pharmacologically-induced reduction in Tgfbr1 kinase activity attenuates growth and metastasis of Panc1 cells orthotopically injected into mouse pancreas. To dissect the role of constitutively altered Tgfbr1 signaling in pancreatic cancer development, we crossed Elastase-KrasG12D (EL-Kras) mice with Tgfbr1 haploinsufficient mice to generate EL-Kras/Tgfbr1+/− mice. Mice were euthanized at 6-9 months to compare the incidence, frequency, and size of precancerous lesions in the pancreas. Only 50% of all EL-Kras/Tgfbr1+/− mice developed preinvasive lesions compared to 100% of EL-Kras (wild type Tgfbr1) mice. The frequency of precancerous lesions was 4-fold lower in haploinsufficient than in control mice. The mitotic index of precancerous cells and the observable levels of fibrosis, lipoatrophy, and lymphocytic infiltration were reduced in EL-Kras/Tgfbr1+/− mice. Tgfbr1 haploinsufficiency was demonstrated by decreased levels of psmad2, psmad3, and nuclear localization of psmad4 and was culminated by a reduction in the Tgfbr1: Tgfbr2 ratio in whole mouse pancreas, which was predominantly caused by increased Tgfbr2. Paradoxically, the precancerous lesions of EL-Kras/Tgfbr1+/− mice were considerably larger than those in EL-Kras mice, which was due, in part, to a reduced level of apoptosis. We conclude that Tgfbr1 signaling promotes the development of precancerous lesions in mice. These findings correlate with TGF-β signals that increase ECM deposition and augment immune responses and strongly suggest that Tgfbr1 haploinsufficiency may have a protective role with respect to early pancreatic cancer development. It is also possible that individuals with constitutively decreased TGFBR1 expression may have a decreased risk of pancreatic 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 3144.
Abstract 3149: 4E-BP1 loss of function in pancreatic carcinogenesis
Authors: Azar Rania , Susini Christiane , Bousquet Corinne , Pyronnet Stephane
Published: February 2011

Inactivation of the transcription factor Smad4/DPC4 gene is a frequent event that correlates with aggressiveness of pancreatic adenocarcinomas. We have previously shown that the stress-inducible translational inhibitor 4E-BP1 is a target of Smad4 essential for TGFbeta-mediated inhibition of cell proliferation. Here, we show that 4E-BP1 expression parallels that of Smad4 through the graded stages of pancreatic cancer progression. 4E-BP1 amount transiently increases in human PanIN-1 and PanIN-2 lesions, but is dramatically diminished in PanIN-3 and invasive carcinomas. A similar pattern of 4E-BP1 expression is seen in PanINs from pancreas of mice that carry a pancreas-specific mutated Kras allele (Kras-Pdx1/cre mice). Furthermore, we show in vitro that 4E-BP1 expression is efficiently induced under hypoxia in Smad4+/+, but not in Smad4−/− pancreatic duct cells. We further reveal that, at the molecular level, Smad4 cooperates with HIF1-alpha to induce 4E-BP1 gene transcription in hypoxic cells. Thus, it appears that pancreatic duct cells which carry Smad4 loss are no longer capable of inducing 4E-BP1 under stress conditions. This may have important consequences in pancreatic carcinogenesis as 4E-BP1 is viewed as a stress-inducible factor whose function is to block cellular protein synthesis to concentrate resources on an appropriate stress response aimed at restraining carcinogenesis.

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 3149.
Abstract 3151: Posttranscriptional regulation of p21 by MCG10, an RNA-binding protein and a target of tumor suppressor p53
Authors: Scoumanne Ariane , Cho Seong Jun , Chen Xinbin
Published: February 2011

The tumor suppressor p53 plays a crucial role in regulating many cellular processes, such as cell cycle arrest, apoptosis, senescence and cell metabolism, in response to stress signals. As a transcription factor, p53 modulates the expression of a plethora of genes, including p21, Killer/DR5, GADD45, TIGAR and MDM2. The tumor suppressor function of p53 is further underscored by the high propensity of p53 mutations found in human cancer. RNA-binding proteins are key modulators in RNA metabolism involved in alternative splicing, mRNA stability, mRNA transport as well as translation. Abnormal expression of RNA-binding proteins is implicated in a growing number of human diseases ranging from neurological disorders to cancer. Here, we showed that MCG10, an RNA-binding protein and a target of p53, regulates the stability of cell cycle inhibitor p21 transcript. Indeed, we found that MCG10 decreases p21 levels induced upon DNA damage. In addition, we showed that MCG10 directly binds to the 3′untranslated region of p21 transcript to destabilize it. Furthermore, we generated MCF7 breast adenocarcinoma and RKO colon carcinoma cell lines in which MCG10 is inducibly knocked down. We found that MCG10 is required for cell proliferation. In addition, we showed that deficiency in MCG10 leads to an increase in basal and DNA damage-induced p21 levels. Taken together, we uncovered a role for RNA-binding protein MCG10 in modulating the p53 pathway through posttranscriptional regulation of p21.

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 3151.
Abstract 3155: The SCFFBW7 E3 ubiquitin ligase targets Krüppel-like factor 5 (KLF5) for proteolysis in breast cancer
Authors: Zhao Dong , Zheng Han-Qiu , Zhou Zhongmei , Chen Ceshi
Published: February 2011

Krüppel-like factor 5 (KLF5) is an evolutionarily conserved zinc finger-containing transcription factor promoting breast cell proliferation, survival, and tumor growth through upregulating its target gene FGF-BP. The protein abundance of KLF5 has been shown to be strictly regulated at transcriptional and posttranslational levels. Our previous studies suggest that the KLF5 protein degradation is through the ubiquitin proteasome pathway. The WWP1 E3 ligase can target KLF5 for ubiquitin-mediated proteasomal degradation. In this study, we demonstrated the F-box protein FBW7 targets KLF5 for proteolysis and suppresses KLF5's functions in breast cancer. The SCFFBW7 (SKP1-Cullin1-F-box) E3 ligase complex has been shown to target several oncoproteins, including MYC and CyclinE, for proteolysis. FBW7 could be a tumor suppressor due to its frequent mutations in multiple cancers. Here, we first demonstrated that inactivation of endogenous FBW7 by gene knockout, knockdown, or mutation remarkably increases the endogenous KLF5 protein abundances through blocking its degradation in multiple cell lines. Second, we found that overexpression of FBW7 decreases the exogenous and endogenous KLF5 protein levels in HEK293T and SUM149PT cell lines. Functionally, overexpression of FBW7 in the endogenous FBW7 mutated SUM149PT cells decreases the FGF-BP gene expression and the DNA synthesis. Importantly, knockdown of endogenous Fbw7 increases the FGF-BP gene expression, cell proliferation, and survival through upregulating the KLF5 expression in breast cells. These findings suggest that the Fbw7 tumor suppressor inhibit breast cancer cell proliferation and survival at least partially through targeting KLF5 for ubiquitin-mediated degradation. This new regulatory mechanism of KLF5 degradation may result in useful diagnostic and therapeutic targets for breast cancer and other cancers.

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 3155.
Abstract 3156: eIF4E in human breast cancer and its role in regulating translation of splice variants of breast cancer genes
Authors: Cheung Jenny Yuen-Nei , Wong Ashley San-Yu , Chan Kelvin Yuen-Kwong , Khoo Ui-Soon
Published: February 2011

In eukaryotes, most of mRNAs are translated by the scanning mechanism. The trimeric complex eIF4F (consisting of eukaryotic initiation factor eIF4E, eIF4G and eIF4A) binds to ribosomes and the 5′end of mRNA, unwinding the secondary structures for scanning. eIF4E is the least abundant initiation factor, and may therefore determine the rate of translation. Overexpression of eIF4E can cause malignant transformation of immortalized human cells. Anti-sense oligonucleotides of eIF4E partially reversed the malignant phenotypes, suggesting that overexpression of eIF4E is the direct cause of transformation. Two- to threefold higher eIF4E levels have been detected in breast cancer using immunohistochemistry. Aabout 10% of cellular mRNAs contain atypically long 5’UTR and many of these encode proto-oncogenes and growth factors. It is believed that these long 5’UTR sequences tend to form stable secondary structure, requiring higher levels or activities of eIF4F for their translation.

Over 60% of human genes are alternatively spliced. These may produce variants with different degrees of secondary structure at their 5’-UTR. We hypothesized that the elevated eIF4E level in breast cancer cells allows more effective translation of breast cancer gene variants with extensive 5’UTR secondary structure. These changes in variants profile may be important for breast cancer development and progression. We first analyzed the expression levels of eIF4E in human breast cancer and non-tumor biopsies in tissue microarray. Our preliminary results showed that there was significantly higher eIF4E expression in tumor versus non-tumor and eIF4E expression was positively correlated to estrogen receptor (ER) expression (OR=4.4, p=0.006). Although eIF4E expression levels were independent of tumor grade and size, it was strongly associated with overall clinical ER and progesterone receptor status (OR=6.3, p=0.003 and OR=6.5, p=0.002). The differential translation of breast cancer gene splice variants in breast cancer cell lines is being analyzed and their correlation with the extent of secondary structure of their 5’UTRs will be determined.

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 3156.
Abstract 3157: Subcellular localization and mRNA targets of a novel human RNA-binding protein, KIAA0020
Authors: Yang Ching-Wen , Cheng Shu-Yu , Chang Christina L.
Published: February 2011

Human KIAA0020 belongs to the PUF family of RNA-binding proteins. Most of family members contain eight RNA-binding Pumilio repeats and are known to regulate mRNA stability and translation. In fact, hunchback, Cyclin B, and HO endonuclease mRNAs are known targets for certain PUF family members in human and other species. Currently little is known about human KIAA0020, which contains six instead of eight Pumilio repeats and shares low homology with its family members. In this study, we characterized this protein and found that wild-type KIAA0020 was localized to multiple foci within the nucleus of HeLa and HEK293T cells. Deletion of C- terminus and, to a less extent, N-terminus of KIAA0020, permitted partial cytosolic localization. Furthermore, we have found a number of mRNA targets that were regulated by KIAA0020 in cells containing ectopic overexpression of KIAA0020 variants or shRNA-mediated knockdown. Based on microarray data analysis from Oncomine, KIAA0020 appears to be deregulated in a variety of human cancer types. Understanding the subcellular localization and mRNA targets of KIAA0020 will improve our knowledge of the function of this protein and its relevance to 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 3157.