bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1070: Pancreatic Ductal Adenocarcinoma (PDA) mice lacking Mucin 1 has a profound defect in tumor growth and metastasis
Authors: Besmer Dahlia , Roy Lopamudra Das , Curry Jennifer , Tinder Teresa , Manouchehrabadi Mahnaz , Schettini Jorge , Hwang Sunil , Lee Yong-Yook , Gendler Sandra , Mukherjee Pinku
Published: June 2012

MUC1 is over expressed and aberrantly glycosolated in >60% of pancreatic ductal adenocarcinomas. Over expression of MUC1 in pancreatic cancer is known, however the functional role has yet to be fully elucidated due to a dearth of appropriate models. In the present study, we have generated mouse models that spontaneously develop pancreatic ductal adenocarcinoma (KC), which are either Muc1 null (KCKO) or express human MUC1 as a self antigen (KCM). We show that PDA mice lacking Muc1 have significantly slower tumor progression, with tumor weight reaching up to a third of the KCM tumor and half of KC tumor. One out of 10 KCKO mice developed secondary metastasis versus 6 out of 10 and 3 out of 10 KCM and KC respectively. Cell lines derived from KCKO tumors have significantly lower tumorigenic capacity compared to cells derived from KCM tumors. Therefore, mice with KCKO tumors had a significant survival benefit compared to mice bearing the KCM tumors. In vitro, the KCKO cells have reduced proliferation and invasion compared to KCM cells and failed to respond to exogenous epidermal growth factor (EGF), platelet-derived growth factor (PDGF), or matrix metalloproteinase-9 (MMP9) stimuli. Further, significantly fewer KCKO cells entered the G2M phase of the cell cycle compared to the KCM cells, consequently more KCKO cells were observed at the S-phase of the cell cycle. Proteomics and western blotting analysis revealed a complete loss of cdc-25c expression, phosphorylation of MAPK, as well as a significant decrease in Nestin and Tubulin α-2 chain expression in KCKO cells. Treatment of the KCM cells with a MEK1/2 inhibitor, U0126, completely abrogated the enhanced proliferation of the KCM cells but had minimal effect on KCKO cells, strongly suggesting that MUC1 is necessary for MAPK activity and downstream oncogenic signaling. This is the first study to utilize a Muc1-null PDA mouse in order to fully elucidate the oncogenic role of MUC1, both in vivo and in vitro.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1070. doi:10.1158/1538-7445.AM2011-1070
Abstract 1071: Repression of ERK1/2 signaling by RanBPM
Authors: Atabakhsh Elnaz , Schild-Poulter Caroline
Published: June 2012

Ran binding protein-M (RanBPM, also called RanBP9) is a nucleo-cytoplasmic protein that has been implicated in a multitude of cellular roles, including the regulation of transcription, protein modifications and stability, and extracellular receptor signaling pathways. However the precise mechanism by which RanBPM functions within the cell remains largely unknown. Previous studies in our lab have identified RanBPM as a pro-apoptotic protein that mediates its effects through modulation of Bcl-2 levels. Here we present evidence that the effect of RanBPM on apoptosis is mediated, at least in part, through regulation of the ERK1/2 signaling pathway.

First, we determined that in addition to Bcl-2, RanBPM also regulated the expression of a second anti-apoptotic factor, Bcl-XL. siRNA-mediated knockdown of RanBPM expression led to elevated Bcl-2 and Bcl-XL mRNA expression, suggesting regulation by RanBPM at the transcriptional level. Conversely, we found that expression of RanBPM also affected ectopically expressed Bcl-2 protein levels. Together, these findings suggested that RanBPM regulated the expression of Bcl-2 and Bcl-XL at both the transcriptional and post-translational levels.

One pathway that has previously been shown to regulate the expression of Bcl-2 and Bcl-XL at both the transcriptional and post-translational levels is the ERK1/2 signaling pathway. We found that RanBPM-deficient cells exhibited enhanced ERK1/2 phosphorylation and activation, while re-expression of RanBPM led to a marked decrease in the levels of phosphorylated ERK1/2. Further analyses indicated that enhanced ERK1/2 activation in RanBPM-deficient cells contributed to the elevated levels of Bcl-2 expression observed in these cells, as inhibition of ERK1/2 activation correlated with decreased Bcl-2 protein expression. These findings suggested that RanBPM acts as a novel negative regulator of ERK1/2 signaling.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1071. doi:10.1158/1538-7445.AM2011-1071
Abstract 1085: Nuclear localization and function of EphB4 in prostate cancer cells
Authors: Mertens-Walker Inga , Burke Leslie , Herington Adrian C. , Stephenson Sally-Anne
Published: June 2012

EphB4 is a member of the largest group of receptor tyrosine kinases. EphB4 is significantly overexpressed in prostate cancer where it contributes to tumour progression as demonstrated through knock down of EphB4 in both in vitro and in vivo cell models. EphB4's interaction with its preferred ligand ephrin-B2 may lead, in some cases, to tumour suppression, suggesting therefore that ligand-independent effects are oncogenic. EphB4 is usually located to the plasma membrane, but using an in-house specific monoclonal antibody we have identified EphB4 in the nucleus of prostate cancer cell lines and patient tumour samples. Accordingly, we hypothesise that EphB4 can directly regulate gene expression in prostate cancer cells and that this may be at least one mechanism through which the ligand-independent effects on tumour progression may be manifested. Using chromatin immunoprecipitation with the specific EphB4 antibody we were able to precipitate numerous DNA sequences that are currently being validated. Using confocal microscopy we have discovered that EphB4 colocalises with SUMO-1 (small ubiquitin-like modifier-1). SUMOylation is a posttranslational modification by which one or multiple SUMO molecules are attached to certain lysine residues of a target protein and this can result in cytoplasmic-nuclear shuttling. In silico analysis of EphB4 revealed that it contains a highly conserved potential sumoylation site, K616. This is a highly novel and significant finding in the Eph receptor field and may provide a mechanism by which EphB4 localises to the nucleus. Further characterisation of EphB4 sumoylation and nuclear translocation is currently being undertaken. This study provides novel insight into Eph receptor biology and may identify a major new mechanism by which EphB4 can control gene expression and potentially influence cancer progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1085. doi:10.1158/1538-7445.AM2011-1085
Abstract 1099: Aza-derivatives of resveratrol are potent inhibitors of macrophage migration inhibitory factor (MIF)
Authors: Fujita Yoshihiko , Islam Rafiqul Islam , Kimura Hideharu , Matsumoto Kazuko , Furuta Kazuyuki , Nagai Tomoyuki , Kudo Kanae , Kaneda Hiroyasu , Tamura Daisuke , Sakai Kazuko , Aomatsu Keiichi , Arao Tokuzo , Okawara Tadashi , Nishio Kazuto
Published: June 2012

Resveratrol (3, 4’, 5-trihydroxy-trans-stilbene), a natural phytoalexin found in grapes and wine, has an anti-proliferative activity in human-derived cancer cells. In our study, we used a conventional condensation reaction between aldehydes and amines to provide a number of aza-resveratrol (3, 4’, 5-trihydroxy-trans- aza-stilbene) derivatives in an attempt to screen for compounds with resveratrol's action but with increased potency. Aza-resveratrol and its hydroxylated derivative (3, 4, 4’, 5-tetrahydroxy-trans- aza-stilbene) showed a more enhanced anti-proliferative effect than resveratrol in an MCF-7 breast carcinoma cell line. To identify the cellular targets of the aza derivatives of resveratrol, we conjugated the latter aza-stilbene compound with epoxy-activated agarose and performed an affinity purification. Macrophage migration inhibitory factor (MIF), a proinflammatory cytokine, was identified as a major target protein in MCF-7 cell lysates using a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF MS). The aza-resveratrol and its hydroxylated derivative, but not resveratrol, were also found to be potent inhibitors of MIF tautomerase activity, which may be associated with their inhibitory effects on MIF bioactivity for cell growth. Such inhibitors may lead to the development of drugs for the treatment of cancer and, possibly, inflammatory and autoimmune diseases.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1099. doi:10.1158/1538-7445.AM2011-1099
Abstract 1102: c-Kit regulates stem-like phenotype and chemoresistance in ovarian cancer cells
Authors: Chau Wing Ka , Wong Alice Sze Tsai
Published: June 2012

One emerging concept of the development of drug-resistant tumors involves a pool of self-renewing malignant progenitors. However, the factors and molecular mechanisms that govern this process remain largely elusive. It has been reported that the high malignancy of these cells is associated with a more efficient cytokine network. In this study, we characterized tumor-initiating cells (TIC) from human ovarian cancer cell line as a model. In contrast to non-TIC, TIC had higher colony formation ability in vitro and greater tumorigenicity in vivo, suggesting that TIC enrich cancer stem-like cells. Notably, the c-Kit receptor tyrosine kinase and its ligand stem cell factor were significantly elevated in TIC. The use of c-Kit siRNA or small molecule inhibitor imatinib to inhibit c-Kit expression and kinase activity, respectively, in TIC resulted in diminished cell proliferation and survival and in vivo tumorigenicity, confirming that the effect was c-Kit specific. Furthermore, we showed that while the two front-line chemotherapeutic drugs cisplatin and paclitaxel could eliminate non-TIC, it had no effect on TIC. Combining cisplatin/paclitaxel with c-Kit siRNA or imatinib prevented the growth of both tumor cell populations. These effects were found to be dependent on signaling of phosphatidylinositol 3-kinase/Akt, but not extracellular signal-regulated kinase 1/2. These results reveal an important role for c-Kit in regulation of self-renewal and proliferation of ovarian cancer cells, which could be selectively targeted for improved treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1102. doi:10.1158/1538-7445.AM2011-1102
Abstract 1104: Insulin-like growth factor-1 is elevated in viscerally obese oesophageal adenocarcinoma patients and expression of its receptor correlates with tumor invasiveness
Authors: Doyle Suzanne L. , Donohoe Claire L. , Lithander Fiona E. , Reynolds John V. , Pidgeon Graham P. , Lysaght Joanne
Published: June 2012

The insulin-like growth factor (IGF) pathway and visceral obesity have been independently linked with esophageal cancer development. This study aimed to delineate the differential and interlinked role of visceral obesity and the IGF-1 system in esophageal adenocarcinoma and squamous cell carcinoma (SCC) progression.

IGF-1 receptor (IGF-1R) mRNA and protein were examined in esophageal SCC (KYSE410, OE21) and adenocarcinoma (OE19, OE33) cell lines by Western Blotting. Tumor cell proliferation in response to IGF-1 was assessed by BrdU incorporation assay. Circulating levels of IGF-1 were measured in 99 serum samples by ELISA and IGF-1R expression was assessed in 50 resected tumor specimens by immunohistochemistry. Results were related to tumor type, gender and visceral adiposity.

Higher IGF-1R mRNA and protein expression were observed in SCC cells, however only adenocarcinoma cell lines showed significantly increased cell proliferation in response to IGF-1 (p<0.01). Significantly higher circulating levels of IGF-1 were detected in patients who had adenocarcinoma (p<0.05) and who were viscerally obese (p<0.05). In resected esophageal adenocarcinoma, IGF-1R expression significantly increased as the tissue progressed along the malignant sequence, with the highest expression observed at the invasive leading edge of the tumor. Uniform IGF-1R expression was observed in SCC tumors.

This novel study is the first of its kind to examine both the differential role of the IGF system in esophageal adenocarcinoma and SCC, and also its association with visceral obesity. These data indicate that in esophageal adenocarcinoma, the IGF-1 axis plays a key role in malignant progression and represents a plausible mechanism by which visceral obesity increases esophageal adenocarcinoma risk.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1104. doi:10.1158/1538-7445.AM2011-1104
Abstract 1113: TGFβ pathway activation limits hematopoietic recovery from chemotherapy
Authors: Brenet Fabienne , Scandura Joseph M.
Published: June 2012

Hematopoietic stem cells (HSCs) are capable of self-renewal and govern the long-term repopulation of all cell types of the blooding forming system. During normal hematopoiesis, there is a tight balance maintained between HSC self-renewal and differentiation. This balance is partially regulated through cell cycle regulators such as cyclin-dependent-kinases (CDKs) and CDK inhibitors (CKDIs); crucial positive and negative regulators of cell-cycle progression. We previously identified the CDKI p57Kip2 (Cdkn1c, p57) as an early transcriptional target of the tumor suppressor TGFβ that is required for TGFβ-induced cytostasis of human CD34+ hematopoietic progenitor/stem cells; functions that provide a mechanistic basis for its expression being silenced in many aggressive human myeloid and lymphoid malignancies. P57 is essential during late embryogenesis, functioning in tissue-specific developmental programs to coordinate proliferation, differentiation and apoptosis. We, and others, have found that p57 is highly expressed in deeply quiescent HSCs yet no role for p57 in hematopoiesis has been assigned. Utilizing a murine p57 knockout (KO) model, we studied the function of p57 in steady-state and stress hematopoiesis. Using limiting-dilution and competitive repopulation HSC assays, we found fewer long-term HSCs in the fetal liver of p57-KO animals demonstrating a role for p57 in HSCs development. In adult hematopoiesis, we found that p57 restrains the hematopoietic response to stresses imposed by serial-HSCs transplantation, hemolysis or myelotoxic chemotherapy. As a consequence, animal with p57-KO hematopoiesis tolerate higher doses of chemotherapeutics due to their more robust recovery from chemotherapy-induced myelosuppression. Using phospho-specific SMAD2 antibodies we found that the TGFβ pathway is activated in immature hematopoietic cells, in vivo, following chemotherapy-induced hematopoietic stress. Normally this promotes p57-induced cytostasis but p57-KO HSCs escape this restraint. We used 9-color flow cytometry to investigate how p57 affects hematopoietic recovery in long-term HSCs (LKSCD34- Flk2-), short-term HSCs (LKSCD34+ Flk2-), multipotent progenitors (LKSCD34+ Flk2+), common myeloid progenitors (LKSca-CD34+ FcRlo), granulocyte-monocyte progenitors (LKSca- CD34+ FcR+) and megakaryocyte erythroid progenitors (LKSca- CD34- FcR-). These results show that the most deeply quiescent HSCs undergo more robust self-renewal during stress in the absence of p57. Our results suggest that myelotoxic chemotherapy blockade of TGFβ-induced cytostasis, normally mediated by p57, could be used to limit chemotherapy-induced myelosuppression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1113. doi:10.1158/1538-7445.AM2011-1113
Abstract 1124: IGFBP-4 activates the Wnt/beta-catenin signaling pathway and induces M-CAM expression in human renal cell carcinoma
Authors: Ueno Koji , Hirata Hiroshi , Tabatabai Z. Laura , Hinoda Yuji , Dahiya Rajvir
Published: June 2012

Introduction and Objective : The Wnt/beta-catenin signaling pathway is inactivated by Wnt antagonists in most cancers. IGFBP-4 is an antagonist of the Wnt/beta-catenin signaling pathway, however its function is not currently understood in renal cell carcinoma (RCC).

Methods : The expression levels of IGFBP-4 protein in kidney tissue microarray were examined by immunostaining using antibody to IGFBP-4. The IGFBP-4 mRNA expression level was analyzed in a normal kidney cell line (HK-2), primary renal cancer cell lines (A498, 769-P) and metastatic renal cancer cell lines (ACHN, Hs891.T). To examine the function of IGFBP4, MTS, matrigel invasion, colony formation and wound healing assays were performed using 769-P mock and stable IGFBP-4 transfected cells. To screen for metastatic related genes in IGFBP-4 stable cells, cDNA PCR array was performed with a Human RT2 Profiler PCR Array. We investigated mRNA expression of the MMP family and measured Tcf transcriptional activity to monitor the Wnt/beta-catenin dependent pathway in 769-P mock and IGFBP-4 stable transfected cells. To analyze the role of IGFBP-4 in metastatic RCC cell lines, IGFBP-4 mRNA was knocked down using a siRNA technique.

Results : We found that the expression of IGFBP-4 was significantly lower in primary RCC and higher in metastatic RCC compared to normal human kidney tissues. To assess the function of IGFBP4, we established IGFBP4 transfectants (primary renal cancer cell line) and performed functional analyses including Tcf reporter assays, cell viability, invasive capability, mortality, and in vivo tumor growth. Interestingly IGFBP-4 transfectants promoted cell growth (in vitro and in vivo), invasion, and motility in 769-P cancer cells. Tcf transcriptional activity was significantly increased in IGFBP-4 transfectants compared to mock cells and beta-catenin expression was increased. Also the beta-catenin downstream effector, MT1-MMP showed increased expression in IGFBP4 transfectants. Additionally IGFBP4 induced the expression of M-CAM, a marker of tumor progression. In order to assess the role of IGFBP4 in metastatic renal cancer, IGFBP-4 mRNA in a metastatic renal cancer cell line (ACHN) was knocked-down using a siRNA technique. Cell growth and motility was decreased in si-IGFBP4 transfected ACHN cells compared to cells transfected with control siRNA.

Conclusions : This is the first report documenting that IGFBP-4 expression in RCC activates cell growth, metastasis, Wnt/beta-catenin signaling and may be involved in RCC metastasis.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1124. doi:10.1158/1538-7445.AM2011-1124
Abstract 1129: Function of mutant EGFR in maintaining glioblastoma growth and heterogeneity
Authors: Inda Maria Del Mar , Bonavia Rudy , Vandenberg Scott , Cavenee Webster , Furnari Frank
Published: June 2012

Glioblastoma Multiforme (GBM) exhibits intratumoral heterogeneity on the morphological and molecular levels, a hallmark feature which may contribute to its poor response to targeted therapy. A powerful oncogenic event in GBM is amplification and rearrangement of the epidermal growth factor receptor (EGFRvIII variant, deleted for exons 2-7, hereafter ΔEGFR), 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 observed that the tumorigenicy of wtEGFR-expressing cells can be significantly enhanced by the presence of ΔEGFR-expressing cells within the tumor, suggesting that soluble factors produced by the latter cells might mediate this potent tumor enhancement. 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. In addition, over-expression of both receptors in the same cell potently enhances tumorigenicity in vivo and colony formation in vitro through the activation of wtEGFR, as well as Akt and STAT3 pathways, suggesting that the secretion of IL-6/LIF cytokines can act not only in a paracrine fashion, but also in an autrocrine fashion when the cell express both receptors. 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 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1129. doi:10.1158/1538-7445.AM2011-1129
Abstract 1132: Oncogenic role of EAPII/TTRAP in lung cancer development and its activation of MAPK-ERK pathway
Authors: Li Chunyang , Fan Songqing , Owonikoko Taofeek K. , Khuri Fadlo R. , Sun Shi-yong , Li Runzhao
Published: June 2012

Cancer progression involves multiple complex and interdependent steps including progressive proliferation, angiogenesis and metastases. The complexity of these processes requires a comprehensive elucidation of the integrated signaling networks for better understanding. EAPII/TTRAP interacts with multiple cancer-related proteins, but its biological significance in cancer development remains unknown. In this report we identified the elevated level of EAPII/TTRAP protein in non-small cell lung carcinoma (NSCLC) patients and NSCLC cell lines in culture. The oncogenic role of EAPII in lung cancer development was demonstrated using NSCLC cells with genetic manipulations that influence EAPII expression: EAPII overexpression increases proliferation of NSCLC cells with an accelerated transition of cell cycle and facilitates xenograft tumor growth in vivo; EAPII knockdown results in apoptosis of NSCLC cells and reduces xenograft tumor formation. To further explore the mechanism of EAPII's oncogenic role in lung cancer development and to elucidate the potential signaling pathway(s) that EAPII may impact, we employed antibody array to investigate the alternation of the major signaling pathways in NSCLC cells with altered EAPII level. We found that EAPII overexpression significantly activated Raf-1 and ERK1/2, but not JNK and p38 pathways. Consistently, the protein and mRNA levels of MYC and cyclin D1, which are targets of the mitogen-activated protein kinases/extracellular signal-regulated kinases (MAPK-ERK) pathway, are significantly increased by EAPII overexpression. Taken together, we demonstrated that EAPII is an oncogenic factor and the activation of MAPK-ERK signaling pathway by EAPII may contribute to lung cancer development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1132. doi:10.1158/1538-7445.AM2011-1132
Abstract 1146: Multiple HDAC inhibitors (HDACi) reduce the expression of the oncogenic miR-17∼92 cluster; a pan HDACi class effect
Authors: Talbert Dominique R. , Peng Xiaodong , Shell Scott A. , Bacus Sarah
Published: June 2012

miR-17∼92 is an oncogenic polycistronic cluster encoding six mature miRNAs (MIR-17, MIR-20A, MIR-18A, MIR-19A, MIR-19B, and MIR-92A). This oncogenic cluster is driven by MYC activity and is highly-expressed in many tumor types where it has been shown to promote tumorigenesis through the suppression of oncogene-induced senescence. We recently observed reduced expression of the miR-17∼92 miRNA cluster in a miRNA microarray analysis of MCF-7 and MDA-MB-231 breast cancer cells treated with the pan-HDAC inhibitors (HDACi's) Vorinostat, MS-275, and LAQ824. Treatment with all three HDACi's, at concentrations shown to induce the expression of p21 and acetylated histone-H3, demonstrated this effect which was further validated by qRT-PCR. Furthermore, significant cell cycle arrest and induction of apoptosis and/or senescence were shown with treatment of both MCF-7 and MDA-MB-231 cells with the HDACi's. There was also significant reduction of MYC expression upon treatment with HDACi's which is consistent with studies showing that the miR-17∼92 cluster is driven by MYC. The miR-17∼92 cluster has also been shown to suppress the expression of several cellular targets important in tumor suppression including p21, PTEN, E2F1, and BIM. The expression of these proteins will also be assessed in MCF-7 and MDA-MB-231 cells after treatment with HDACi. The modulation of MYC, the miR-17∼92 cluster and its oncogenic targets will also be confirmed in MDA-MB-231 and MCF-7 tumor xenograft mice treated with the HDACi's. Lastly, antagomirs will be used to inhibit the expression of the miR-17∼92 cluster in MCF-7 and MDA-MB-231 cells in order to compare the functional (apoptosis, senescence, and cell cycle arrest) and cellular (repression of target proteins) effects elicited by HDACi treatment. Together, we have shown that pan-HDAC inhibitors inhibit the expression of MYC and the oncogenic miR-17-92 cluster. This finding could have important utility in the clinical development of pan-HDACi's by identifying MYC and/or the miR-17-92 cluster as potential biomarkers to select patients more likely to respond to pan-HDACi treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1146. doi:10.1158/1538-7445.AM2011-1146
Abstract 1150: Genetic variations in microRNA-related genes as predictors for recurrence and progression in patients with non-muscle invasive bladder cancer
Authors: Ke Hung-Lung , Chen Meng , Dinney Colin P. , Wu Xifeng
Published: June 2012

MicroRNAs (miRNAs) are a group of short non-protein-coding RNAs regulating gene expression in eukaryotes, which can act as oncogenes or tumor suppressors. To test the hypothesis that genetic variations in the miRNA biogenesis genes are associated with the prognosis of bladder cancer, we genotyped 76 single nucleotide polymorphisms (SNPs) in 8 genes (DDX20, DGCR8, DICER1, EIF2C1, GEMIN4, RAN, RNASEN and XPO5) from the miRNA biogenesis pathway in 421 patients with non-muscle invasive bladder cancer. Eight SNPs reached significant association with cancer recurrence in patients who received trans-urethral resection (TUR) only, and three SNPs in patients who received intravesical Bacillus Calmette-Guérin (BCG) instillation. The most significant SNP in TUR group was rs197412 in DDX20, with the variant allele conferring a 42% decreased risk of recurrence (HR=0.58, 95% CI=0.40-0.82, P=0.002). The most significant SNP in BCG group was rs563002 in DDX20, with the variant allele conferring an increased risk of recurrence (HR=2.80, 95% CI=1.30-6.04, P=0.009). In addition, seven SNPs showed significant association with cancer progression. The most significant SNP associated with progression was rs3087833 in GEMIN4, with the variant allele conferring an increased risk of progression (HR=9.73, 95% CI=2.31-40.92, P=0.002). Finally, we performed cumulative effect analysis for the number of unfavorable genotype on bladder cancer recurrence and prognosis. These data demonstrated that miRNA-related SNPs may impact the recurrence and progression in bladder cancer patients. Further study in independent large population and functional characterization are needed to validate these results.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1150. doi:10.1158/1538-7445.AM2011-1150
Abstract 1154: MicroRNA-221, -222 and -181 regulates tamoxifen resistance in breast cancer by downregulating TIMP3 and facilitating growth factor signaling
Authors: Lu Yuanzhi , Roy Satavisha , Miller Tyler , Ghoshal Kalpana , Ramaswamy Bhuvaneswari , Shapiro Charlie , Jacob Samson T , Majumder Sarmila
Published: June 2012

We have shown earlier that microRNA (miR)-221 and -222 are upregulated in tamoxifen (TAM) resistant breast cancer cell line MCF-7 (OHTR), and in Her2 positive primary human breast tumors compared to the Her2 negative tumors1. In addition, expression of miR-181b was 3.5-fold higher in OHTR cells compared to the MCF-7 cells and endocrine resistant primary tumors. To determine the underlying mechanism of miR-mediated regulation of TAM resistance, we focused on identification of target genes regulated by these miRs. Bioinformatics search revealed presence of seed sequences for all three miRs on 3’-UTR of TIMP3, a tissue metalloprotease inhibitor. Luciferase assay confirmed TIMP3 to be a direct target of these miRs and was found to be regulated at both RNA and protein levels. In situ hybridization and immunohistochemical analysis demonstrated reciprocal relationship between TIMP3 and miR 221/222 expression in primary human tumors, where invasive ductal carcinomas expressed high level of the miRs and normal or non-invasive cancers expressed TIMP3, but not the miRs. Cell proliferation and wound healing assay demonstrated increased sensitization of the TAM resistant cells to the drug upon ectopic expression of TIMP3. In addition, mouse mammary tumors induced with TIMP3 overexpressing cells were found to be more sensitive to TAM compared to the tumors initiated with control cells. Conversely, knocking down TIMP3 by siRNA reduced sensitivity to TAM, both in cell cultures and in mouse mammary tumors.

As TIMP3 is an inhibitor of matrix metalloproteases involved in positive regulation of growth factor signaling, we speculated that reduction in TIMP3 due to high miR-221/222/181 expression could facilitate growth factor signaling in OHTR cells. Indeed, EGF-induced MAPK phosphorylation was significantly higher in OHTR cells compared to MCF-7 cells and miR-221/222 overexpressing MCF-7 cells mimicked this phenomenon. On the contrary, phosphMAPK level was reduced when TIMP3 was overexpressed in OHTR cells and increased upon knocking down of TIMP3 in MCF-7 cells. Similar differences in MAPK phosphorylation was also observed in cells treated with low level estrogen or TAM, agonists for TAM resistant cells.

Based on this data we conclude that overexpression of miR-221/222 and -181b facilitates growth factor signaling in TAM resistant cells by downregulating TIMP3.

1. Miller et. al. J. Biol. Chem 2008; 283 (44): 29897

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1154. doi:10.1158/1538-7445.AM2011-1154
Abstract 1156: MicroRNA-21 in gliomas: An in situ hybridization study
Authors: Hermansen Simon K. , Dahlrot Rikke H. , Nielsen Boye S. , Hansen Steinbjørn , Kristensen Bjarne W.
Published: June 2012

High-grade gliomas are some of the most lethal forms of human cancers. In glioblastomas, which are the most frequent and malignant type of glioma, microRNA-21 (miR21) has recently been suggested to be strongly upregulated. Until now, miR21 expression analysis has mainly been carried out using PCR methods, hence little is known about the cellular localization of miR21 in gliomas. Studies of other cancers have demonstrated miR21's prognostic value as well as roles in invasion, growth and anti-apoptosis. This suggests that miR21 is a potential therapeutic target as well as a promising novel prognostic marker in gliomas. The aim of the present study was therefore to investigate the expression of miR21 in gliomas and correlate it with tumor grade and overall survival using the new possibilities of locked nucleic acid (LNA) based in situ hybridization. We used 193 formalin-fixed paraffin embedded astrocytic and oligodendroglial brain tumors of increasing grade (WHO I-IV). Sections from these tumors were stained by a robust in situ hybridization assay using high-affinity LNA probes that specifically detect miR21. The expression of miR21 was revealed as a blue chromogenic staining reaction. Using advanced image analysis, the area of the staining reaction was measured by a trained pixel classifier on images recorded by stereological principles. The optimal sampling fraction was estimated to be 10%. The results showed that the miR21 signal was localized to blood vessels and populations of dispersed single cells. Moreover, the characteristic gemistocytic tumor cells seemed to display a distinct staining reaction. The miR21 signal increased with tumor grade and a very high individual signal variation was observed, especially among glioblastomas. When comparing the miR21 signal with overall survival, a high signal was associated with shorter survival, but in multivariate analysis this was not independent of grade. In conclusion miR21 is localized in both tumor cells and blood vessels in gliomas. A high miR21 expression appears to be associated with tumor grade and shorter survival underlining the importance of miR21 in glioma biology.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1156. doi:10.1158/1538-7445.AM2011-1156
Abstract 1161: Detection and identification of a miRNA expression profile from cell-free urine: Potential utility in bladder cancer
Authors: Holway Antonia H. , DeLong Jessica , Harty Niall , Kozinn Spencer , Summerhayes Kelly , Summerhayes Ian C. , Libertino John A. , Rieger-Christ Kimberly M.
Published: June 2012

Introduction/Objective: MicroRNAs are small, non-coding RNAs that have been shown to play an important role in tumorigenesis. There is differential expression of miRNA in cancer progression, and profiling of miRNA is promising for both diagnosis and treatment of malignant tumors. In this study we isolated RNA from cell-free urine in an attempt to characterize miRNA profiles indicating the presence of urothelial carcinoma and its potential use as a non-invasive assay to identify patients with cancer progression.

Methods: Urine was collected from patients diagnosed with bladder cancer and control patients with no history of cancer under an IRB-approved protocol. Urine was centrifuged and total RNA was isolated from the supernatants using the mirVana Paris™ kit. A total of 178 samples were grouped according to grade and stage (healthy controls (35), TaG1 (19), T1G3 (16), ≥T2 (30), carcinoma in situ (CIS; 28) and no evidence of disease following therapy (50). Seven hundred and thirty miRNAs were profiled by qRT-PCR on pooled samples within each group. Validation of selected miRNAs was performed on individual samples using qRT-PCR.

Results: Cell-free RNA was isolated from urine of 35 healthy controls and 143 patients with bladder cancer. Of the 730 miRNAs tested, 236 were detected in at least one of the pooled samples using a Ct cutoff of 35. The number of miRNAs detected in the pooled samples correlated with disease progression where the healthy control group and the ≥T2 group expressed 8 and 228 miRNAs, respectively. qRT-PCR of individual samples revealed a gradual increase of some miRNAs with disease progression. Statistical analysis adjusted for multiple comparisons demonstrated differences between groups based on miRNA expression levels. In addition, a panel of miRNAs was identified which discriminated between cancer and cancer-free patients.

Conclusion: This study demonstrates the successful isolation of miRNAs from cell-free urine. Utilizing non-invasive urine based assays, we identified a miRNA panel that can discriminate between cancer-free patients and patients with urinary carcinoma of the bladder. These findings provide evidence that profiling of miRNAs from cell-free urine holds the promise for the development of valuable clinical tools.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1161. doi:10.1158/1538-7445.AM2011-1161
Abstract 1166: Inhibition of tumor growth in mouse models of cancer by systemic delivery of tumor suppressor miRNAs
Authors: Brown David , Daige Chris , Wiggins Jason , Kelnar Kevin , Ruffino Lynnsie , Omotola Michael , Zhou Jane , Bader Andreas
Published: June 2012

Tumor suppressor miRNAs represent an intriguing new class of biomolecules that might be developed into therapeutic candidates for cancer. Based upon its involvement in the p53 pathway and its capacity to regulate cell cycle and apoptosis via multiple oncogenic pathways, miR-34 might be the most intriguing of the tumor suppressor miRNAs. We have developed mimics of the miR-34 tumor suppressor that have been tested for efficacy using xenograft and transgenic mouse models of cancer. When complexed with an effective delivery agent, tail-vein injected miR-34 mimics have proven capable of inhibiting the growth and metastasis of mature tumors. When delivered systemically to BalbC mice, the same complexed miR-34 mimics fail to incease the serum levels of liver, kidney, and heart toxicity markers. Systemically delivered miR-34 mimics also failed to induce cytokines in mice. The development of miR-34 mimics with enhanced pharmacokinetic profiles and a systemic delivery formulation that can be manufactured under cGMP conditions are underway. Preclinical efficacy, PK/PD, and rodent safety data will be presented.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1166. doi:10.1158/1538-7445.AM2011-1166
Abstract 1181: Expression studies from the PVT1 region of 8q24
Authors: Huppi Konrad , Pitt Jason , Wahlberg Brady , Jones Tamara L. , Mushinski J. Frederic , Janz Siegfried , Caplen Natasha J.
Published: June 2012

Several SNPs in the area surrounding the PVT1 noncoding gene region on human chromosome 8q24 have recently been found by Genome Wide Association (GWA) to be associated with susceptibility to a number of malignancies including Hodgkin's Lymphoma and other diseases including Type II Diabetes and End Stage Renal Disease in Type I Diabetes. This genetically unstable region is one of the most frequent targets of retroviral integration of the Human Papilloma Virus Type 18 (HPV18 in Cervical Carcinoma) and is the frequent target of chromosomal translocation in lymphoid malignancies such as Non-Hodgkin's Lymphoma [incl. Burkitt's Lymphoma (BL) and Diffuse Large B Cell Lymphoma (DLBL)]. Gene amplification in carcinomas such as breast, prostate, ovarian, colon, pancreatic, esophageal, as well as osteogenic sarcoma usually encompasses the proximal region of PVT1 along with its well-known neighboring transcript, MYC, whereas chromosomal deletion (in hematodermic NK lymphoma) affects the distal segment of PVT1. In a search for functionality to PVT1, many laboratories (including our own) have cloned and sequenced transcripts from the PVT1 region from many species in addition to human, with the conclusion that PVT1 is a noncoding or lincRNA with many alternatively spliced isoforms. We have systematically documented these isoforms with a particular focus on promoter and regulatory elements that are found in the region. Our laboratory has also identified a cluster of at least 6 miRNAs (miR-1204∼1208) that span the PVT1 region adding yet another level of complexity to the locus. To begin to assign some function to PVT1 based transcripts in these disease phenotypes, we have made 4th generation (CMV promoter with WPE enhancer) over-expressing lentiviral constructs containing either a large segment of PVT1 (Exons 1b-10) or the most proximal and abundant miRNA, miR-1204. These constructs follow earlier studies with a 3rd generation lentivirus (CMV promoter) containing miR-1204 that rapidly generated DLBL in cooperation with MYC or IL6 transgenes (∼16 days or 82 days respectively). While these results pointed to over-expression of miR-1204 playing a specific role in the development of B cell malignancy, additional studies with the LentiCMV-miR-1204 construct in other tumor cell lines (prostate and breast) also revealed potential phenotypic changes in cell proliferation and migration. With the new 4th generation constructs, experiments are now underway in cell lines and mouse models to determine if over-expression of miR-1204 or PVT1 alone is sufficient to induce phenotypic changes.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1181. doi:10.1158/1538-7445.AM2011-1181
Abstract 1192: Regulation of CCND2, HMGA2, and the IGF pathway by LIN28B in HPV negative head and neck squamous cell carcinoma (HNSCC)
Authors: Alajez Nehad M. , Wong Dennis , Shi Wei , Lenarduzzi Michelle , Liu Fei-Fei
Published: June 2012

Background: Lin28 and Lin28B have recently been implicated in the post-transcriptional regulation of Let-7/miR-98 biogenesis. While Lin28B over-expression has been associated with worse clinical outcome in several human cancers, the prognostic significance of Lin28B expression and the molecular mechanism(s) by which it drives tumour progression remains largely unknown.

Methods: Let-7-Resist Lin28B-expression plasmid was stably transfected into the FaDu human hypopharyngeal squamous cell carcinoma line. MicoRNA expression profiling was conducted using the Nanostring nCounter Analysis System. Genome-wide micro-array and in silico analyses were utilized to identify Let-7/miR-98 targets. Cell viability and apoptosis were measured using MTS assay and flow cytometry, respectively. In vivo experiments were conducted using SCID mice.

Results: qRT-PCR analysis of genes involved in the miRNA biogenesis pathway (DICER, LIN28, LIN28B, and TUT4) suggested an inverse relationship between the expression of Lin28B and mature Let-7 miRNA. Over-expression of Lin28B mediated significant increase in the migration of FaDu cells in vitro, and tumor growth in SCID mice, which in turn, was associated with marked reduction in the Let-7 family of microRNAs. Using a tri-Pronged approach by combining prediction algorithms, experimentally-derived mRNA profiling, along with gene expression data in primary HNSCC samples identified several upregulated Let-7/miR98 gene targets in Lin28B-FaDu cells, including CCND2, HMGA2, and regulators of the insulin growth factor (IGF) mitogenic pathway. Concordantly, treating FaDu-Lin28B cells with IGF (10-50 nM) led to a 26% and 50% increase in cell viability under stress conditions (serum starvation), whereas treating FaDu-GFP cells with IGF had no effect on viability, suggesting the relevance of the IGF axis in FaDu-Lin28B cells. Furthermore, analyzing publicly available data sets on primary HPV positive vs. HPV negative HNSCC samples revealed a significantly higher expression of HMGA2, and genes that regulate the IGF pathway in the latter group of tumours, suggesting a possible role for Lin28B, Let-7/miR-98 and their downstream targets in mediating the more aggressive behavior of HPV negative HNSCC.

Conclusions: Our data report for the first time, a potentially important role for Lin28B in regulating the IGF mitogenic pathway, as one possible mechanism in driving the biological behaviour of HPV negative HNSCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1192. doi:10.1158/1538-7445.AM2011-1192