bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5084: Treadmill exercise modulates AMP-activated protein kinase and downstream signaling in mammary gland of p53+/−, but not p53+/+ mice
Authors: Wheatley Karrie E. , Nogueira Leticia M. , Perkins Susan N. , Hursting Stephen D.
Published: February 2011

Epidemiological data show that exercise is generally protective against breast cancer. Inhibition of mammalian target of rapamyocin (mTOR) through activation of AMP-activated protein kinase (AMPK) has been shown to be responsive to exercise. Decreased activation of ribosomal protein S6, a marker for mTOR inhibition, is associated with decreased growth and proliferation. Recently, this energy-sensing pathway has been shown to be inhibited by increased production of reactive oxygen species (ROS) leading to AMPK activation. ROS levels are maintained by p53-inducible anti-oxidant enzymes. Therefore, we hypothesized that energy sensing pathways that mediate the protective effects of exercise in breast cancer are modulated by p53 gene dosage. To test this hypothesis, we fed ovariectomized mice a high-fat diet (60% kcal) ad libitum for 10 weeks to model post-menopausal obesity. Mice were then randomized to a sedentary or exercise regimen (n=15/group). Exercised mice ran on a variable speed treadmill (5% incline, 20 m/min, 40 min/day, 5 days/week for 22 weeks). Changes in percent body fat were measured every 2 weeks using quantitative magnetic resonance. Immunoblot analysis was performed to measure phosphorylation of AMPK (Thr 172) and ribosomal protein S6 (Ser 240/244) relative to total levels in the mammary fat pad (MFP). Also, qRT-PCR was performed to measure mRNA expression of the p53-inducible anti-oxidant enzyme sestrin 1. At the end of the diet-induced obesity regimen, there was no effect of p53 genotype on percent body fat (54.4 ± 0.7 in p53+/+ versus 55.9 ± 1.1 in p53+/−). After 22 weeks of exercise, the EX mice displayed signficantly less percent body fat than sedentary controls, with no effect of genotype (44.4 ± 0.6 versus 47.8 ± 0.8 % respectively, p<0.05). Immunoblot analysis revealed that in p53+/− mice, but not p53+/+ mice, exercise increased AMPK phosphorylation. This increased AMPK phosphorylation was associated with decreased phosphorylation of ribosomal protein S6, a marker of mTOR pathway activation (n=3/group). The observed increase in AMPK phosphorylation in p53+/− exercised mice relative to sedentary p53+/− was associated with decreased mRNA levels of sestrin 1 (n=4-6/group), indicating that the p53+/− mice may have higher levels of ROS in the MFP. In conclusion, exercise may increase AMPK activation in MFP of p53+/− mice relative to sedentary p53+/− mice through an ROS-mediated mechanism. Current analysis is focused on understanding how exercise affects AMPK activation in the mammary fat pad of MMTV-Wnt-1 transgenic mice heterozygous for p53, and how this impacts mammary 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 5084.
Abstract 509: Novel role of MeCP2 in response to EGFR targeted therapy
Authors: Ramanand Susmita G. , Ahsan Aarif , Ao Xiaoping , Lubman David M. , Ray Dipankar , Lawrence Theodore S. , Nyati Mukesh K.
Published: February 2011

One of the targets towards the development of new agents to combat cancer is the inhibition of epidermal growth factor receptor (EGFR). A therapeutic antibody to EGFR, cetuximab, is seen to be effective in combination with cisplatin or radiation. However, only one out of four patients treated benefit from this therapy. It is therefore important to identify patients who can gain from this treatment. We hypothesize that through phosphoproteome analysis, we can identify alterations in novel signal transduction molecule(s) induced by cetuximab treatment in sensitive cell lines or TMA that will predict response to treatment. Phosphoproteomic changes upon cetuximab treatment in UMSCC-1 (cetuximab-responsive) and UMSCC-74B (cetuximab-non-responsive) cell lines using nonporous silica reverse-phase high performance liquid chromatography (NPS-RP-HPLC) were assessed. To confirm the results of phosphoproteomics data, three proteins that were seen to be affected in the sensitive cell line were investigated by immunoprecipitating total proteins followed by immunoblotting with phospho-specific antibodies. We found pharmacodynamic changes in 12 novel proteins upon cetuximab treatment in UMSCC-1 cell line. By Immunoblotting, we confirmed that these three proteins (methyl-CpG binding proteins) are indeed phosphorylated at serine, threonine and tyrosine sites. Cetuximab treatment in UMSCC-1, induced phosphorylation of NCoR1 and MeCP2 (which cause transcriptional repression) and decreased phosphorylation of MBD2 (which causes transcriptional activation), thus confirming the phosphoproteomic data. In contrast, the phosphoproteomics data in UMSCC-74B indicated either no effect or an opposing effect to that seen in the sensitive cells. These data suggest that we have a powerful strategy to identify novel phosphoproteins that are affected by EGFR inhibition and thus, novel biomarkers of response to cetuximab. These and additional biomarkers that we hope to discover will undergo rigorous in vitro and in vivo testing before employing them on TMA from xenografts.

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 509.
Abstract 5099: Adiponectin inhibits oxidative stress in prostate cancer cells
Authors: Hou ZheFeng , Lu JianPing , Pinthus Jehonathan H.
Published: February 2011

INTRODUCTION AND OBJECTIVES: Emerging data suggest that obesity increases the risk of aggressive prostate cancer (PC) but the mechanisms underlying this relationship remain to be fully elucidated. Adiponectin, an adipocyte-specific hormone, circulates at relatively high levels in healthy human but its levels are reduced in obesity. Case control studies have documented an association between lower adiponectin blood levels and PC implying that obese individuals with low adiponectin levels are more susceptible in developing PC. In this study we examined the effect of adiponectin on the generation and reduction of oxidative stress (OS) in PC cells, a process that intimately relates to the initiation and development of PC.

METHOD: As models we used two human PC cell lines (22rv1 and DU145) expressing the adiponectin receptors AdipoR1, AdipoR2 and T-cadherin. Cells were treated with increasing doses of human recombinant adiponectin (0-40μg/ml) and were subjected to the following tests: Quantification of cellular OS (NBT reduction assay), expression of catalase and MnSOD (Western blot analysis), comparison of transcript levels of NADPH oxidases (NOX) enzymes- a major source of reactive oxygen species in PC cells (RT-PCR) and determination of the global cellular anti-oxidative capacity and catalase activity (colorimetric assays).

RESULT: Adiponectin significantly inhibited OS in both cell types in a dose dependent manner by inducing the expression of the two key anti-oxidative enzymes; catalase and MnSOD, leading to an increase in the anti-oxidative capacity of the cells. Treatment with adiponectin reduced Nox 2,4 mRNA expression in Du145 cells but increased it in 22RV1 cells.

CONCLUSIONS: These results suggest a protective role for adiponectin against the initiation and development of PC, through OS inhibition.

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 5099.
Abstract 5101: Acute inhibition of myostatin-family proteins preserves muscle in cancer cachexia
Authors: Klimek Margaret E. Benny , Aydogdu Tufan , Link Majik , Pons Marianne , Koniaris Leonidas G. , Zimmers Teresa A.
Published: February 2011

Cachexia, progressive loss of fat and muscle mass despite adequate nutrition, is a devastating complication of cancer associated with poor quality of life and increased mortality. Myostatin is a potent tonic muscle growth inhibitor. We tested how myostatin inhibition might influence cancer cachexia using genetic and pharmacological approaches in mice. First, myostatin null mice were challenged with Lewis lung carcinoma or B16F10 melanoma. Although hypermuscular at baseline, myostatin null mice were more sensitive to tumor-induced cachexia, losing more absolute mass and proportionately more muscle mass than wild-type mice. Because myostatin null mice lack expression from development, however, we also sought to manipulate myostatin acutely. The histone deacetylase inhibitor Trichostatin A has been shown to increase muscle mass in normal and dystrophic mice by inducing the myostatin inhibitor, Follistatin. Although Trichostatin A administration induced muscle growth in normal mice, it failed to preserve muscle in colon-26 cancer cachexia. Finally we sought to inhibit myostatin and related ligands by administration of the Activin receptor extracellular domain/Fc fusion protein, ACVR2B-Fc, which inhibits myostatin activity and induces dramatic muscle growth in normal mice. Systemic administration of ACVR2B-Fc potently inhibited muscle wasting and protected adipose stores in both colon-26 and Lewis lung carcinoma cachexia, without affecting tumor growth. Enhanced cachexia in myostatin knockouts indicates that host-derived myostatin is not the sole mediator of muscle wasting in cancer. More importantly, skeletal muscle preservation with ACVR2B-Fc establishes that targeting myostatin family ligands using ACVR2B-Fc or related molecules is an important and potent therapeutic avenue in cancer cachexia.

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 5101.
Abstract 5102: In vitro hedgehog signaling pathway alterations after acquired-platinum resistance in small cell lung cancer
Authors: Ranade Aarati , Weiss Glen J.
Published: February 2011

Background: The Hedgehog (HH) signaling pathway regulates cell growth and differentiation during embryonic tissue development and may play a role of ‘stem cell’ maintenance in adults. Studies have shown that SCLC platinum resistance and cell death is associated with HH signaling pathway maintenance, potentiating SCLC re-growth after chemotherapy. We hypothesized that acquired-platinum resistance would alter HH pathway signaling.

Methods: SCLC cell lines, H69 and H146 along with their acquired cisplatin (H69Cis1) and carboplatin resistant (H69Car1, H146Car1, and H146Car2) versions were used. mRNA expression of genes in HH pathway were measured. Fold changes in mRNA expression in resistant cells (RCs) compared to parent cells were calculated. A two-sided t-test was used to compute the p-values for the fold changes. Proteins from these cells were probed for SHH, SUFU, and GLI1 by immunoblot.

Results: The mRNA expression of genes in HH pathway was altered in platinum RCs compared to their parental cells (Table 1). As with mRNA expression, SHH protein is decreased in both H69 cisplatin and carboplatin RCs (by 93% in H69Cis1 and by 76% in H69Car1) and by 62% in H146 carboplatin RCs. SUFU mRNA and protein expression are not significantly different in the platinum RCs compared to their parental cells. GLI1 protein was not expressed in vitro in these cell lines, but there was a significant increase in GLI1 mRNA expression in platinum RCs.

Conclusions: Our results indicate that the genes in the HH pathway such as GLI1, GLI2, PTCH1, PTCH2, SHH, and SMO are altered in vitro in cisplatin and carboplatin H69 and H146 acquired RCs compared to respective parent cell lines. Detailed studies with other SCLC cells lines with variable sensitivity to platinum and the impact of HH pathway inhibitors on the platinum resistant cell lines in vivo are worth further exploration.Table 1:Relative Fold Changes in mRNA Expression of HH Pathway Genes in Acquired-Platinum Resistant SCLC Cell Lines GLI1GLI2HHIPPTCH1PTCH2SHHSMOSuFuH6911111111H69Car13.17 (0.29)3.46 (0.51)1.21 (0.14)1.52 (0.048*)1.05 (0.93)88.54 (0.17)2.13 (0.037*)0.9 (0.80)H69Cis113.49 (0.028*)3.71 (<0.001*)2.8 (0.018*)1.37 (0.07)1.34 (0.20)55.26 (0.009*)26.01 (<0.001*)1.16 (0.57)H14611111111H146Car16.62 (0.02*)8.49 (<0.001*)0.71 (0.43)0.24 (0.05)0.17 (0.043*)8.49 (<0.001*)0.11 (0.011*)0.71 (0.15)H146Car23.45 (0.05)11.71 (<0.001*)1.13 (0.62)0.17 (0.045*)0.45 (0.15)11.71 (<0.001*)0.13 (0.001*)1.10 (0.49)The values in the table indicate mean (p-value)*Significant p<0.05Fold changes in mRNA expression in resistant cells (RCs) compared to parent cells were calculated.(Supported by the TGen Foundation and SHC Foundation)

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 5102.
Abstract 5104: Cell-surface receptor expression in osteosarcoma
Authors: Geller David S. , Hassan Sheref E. , Gorlick Richard
Published: February 2011

Purpose: Although numerous cell-signaling receptors have been described in the setting of osteosarcoma, the comprehensive characterization of receptors on the cell-surface has not been described. The goal of the current study was to characterize and quantify the expression of cell-surface receptors across multiple standard osteosarcoma cell lines and patient samples placed in culture.

Methods: Both standard (n=4) and patient derived (n=10) osteosarcoma cell lines were cultured and labelled with a panel of commercially available flurophore-bound antibodies including antibodies to EGFR, HER-2, HER-3, HER-4, IGF-1R, IGF-2R, IR, VEGFR-1, VEGFR-2, VEGFR-3, CMET, FGFR-2, FGFR-3, PDGF alpha, and PDGF beta. Standard cell lines utilized were 143b, HOS, Saos, and U205. Patient samples included samples 229, 231, 232, 238, 252, 293, 308, 311, 242, and 290. Background fluorescence was accounted for using either a known negative control cell line if available or the antibody-specific IgG isotype. Unlabelled cells were also evaluated for inherent fluorescence. Cell analysis was performed using flow cytometry (FACS) and data was analyzed using FloJo software. The geometric fluorescent means were calculated and compared against positive controls.

Results: Although a degree of variability in cell-surface receptor expression existed between the different cell lines, preliminary results show high expression of PDGFR-beta in all 14 cell lines, with an average geometric mean of 1376 (range 93.8 - 4496). A higher expression of IGF-1R was also noted across all cell lines, with an average geometric mean of 881 (range 122 - 3191). In depth analysis of receptor expression and the interrelationship of these various receptors is ongoing.

Conclusion: Preliminary results demonstrate PDGFR-Beta and to a lesser extent IGF-1R to be consistently expressed on the cell-surface in all tested osteosarcomas cell lines. Data analysis of receptor expression and interrelationships between receptor expression is ongoing.

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 5104.
Abstract 5122: Regulation of integrin trafficking by Rab proteins
Authors: Hasan Nazarul , Hu Chuan
Published: February 2011

Integrins, the major receptors for cell adhesion to the extracellular matrix, are synthesized in the endoplasmic reticulum, transported in vesicles, and delivered to the plasma membrane or the cell surface by exocytosis. Like other receptors, integrins at the plasma membrane are constantly endocytosed, transported into endosomal compartments and then recycled back to the surface. Therefore, the density of integrins at the cell surface is ultimately determined by the balance between two vesicle trafficking events: endocytosis and exocytosis. Rab proteins are small (20-29 kDa) monomeric Ras-like GTPases that regulate vesicle budding, transport and tethering to the target membranes. Rab4, Rab11 and Rab25 function in the endocytic and recycling pathway. In this study, we examined roles of the Rab proteins in integrin trafficking. Silencing of Rab11 in HeLa cells had neither obvious effects on cell surface or total cellular expression of α3, α5 and β1 integrins nor on cell migration. Silencing of Rab25 reduced the surface expression of all three integrins and inhibited chemotactic migration. Surprisingly, Rab4 silencing dramatically increased both cell surface and total cellular expression of the integrins, and inhibited cell migration as well. These data suggest that Rab25 is required in integrin trafficking to the cell surface, and that Rab4 negatively regulates the expression and trafficking of integrins. These data support the notion that reduced or excessive integrins at the cell surface diminish cell motility.

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 5122.
Abstract 5127: Prosaposin down-modulation decreases metastatic prostate cancer cell adhesion, migration, and invasion
Authors: Hu SiYi , Delorme Nathalie , Liu Zhenzhen , Liu Tao , Velasco-Gonzalez Cruz , Garai Jone , Pullikuth Ashok , Koochekpour Shahriar
Published: February 2011

Factors responsible for invasive and metastatic progression of prostate cancer (PCa) remain largely unknown. Previously, we reported cloning of prosaposin (PSAP) and its genomic amplification and/or overexpression in several androgen-independent metastatic PCa cell lines and lymph node metastases. PSAP is the lysosomal precursor of saposins, which serve as activators for lysosomal hydrolases involved in the degradation of ceramide (Cer) and other sphingolipids. Our current data show that, in metastatic PCa cells, stable down-modulation of PSAP by RNA-interference via a lysosomal proteolysis-dependent pathway decreased β1A-integrin expression, its cell-surface clustering, and adhesion to basement membrane proteins; led to disassembly of focal adhesion complex; and decreased phosphorylative activity of focal adhesion kinase and its downstream adaptor molecule, paxillin. Cathepsin D (CathD) expression and proteolytic activity, migration, and invasion were also significantly decreased in PSAP knock-down cells. Transient-transfection studies with β1A integrin- or CathD-siRNA oligos confirmed the cause and effect relationship between PSAP and CathD or PSAP and Cer-β1A integrin, regulating PCa cell migration and invasion. Our findings suggest that by a coordinated regulation of Cer levels, CathD and β1A-integrin expression, and attenuation of “inside-out” integrin-signaling pathway, PSAP is involved in PCa invasion and therefore might be used as a molecular target for PCa therapy.

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 5127.
Abstract 513: Inhibition of histone deacetylase activity down-regulates urokinase plasminogen activator and matrix metalloproteinase-9 expression in gastric cancer cell lines
Authors: Lee Kyung Hee , Kim Min Kyung , Kim Jae Ryong , Kim Sang Woon , Song Sun Kyo
Published: February 2011

Histone acetylation plays an important role in chromatin remodeling and gene expression. An imbalance of this reaction leads to aberrant behavior of cells in the cell cycle, i.e., carcinogenesis. Histone deacetylase (HDAC) inhibitors have been shown to have anti-tumor effects in clinical trials. However, the exact mechanisms by which HDAC inhibitors exert anti-tumor effects and modulate gene expression are not completely understood, and thus remain a subject of intense investigation. In the current study, we determined whether HDACs regulate urokinase plasminogen activator (uPA), matrix metalloproteinase (MMP)-9, and tumor invasion. Using cDNA microarray analysis, we showed that hepatocyte growth factor (HGF) induced HDAC-5 expression in both gastric cancer cell lines. TSA, a HDAC inhibitor, decreased HGF-induced HADC-5 expression and also repressed uPA and MMP-9 expression. We confirmed these results using HDAC-5 gene overexpressing cell lines. The number of viable cells in the cultures with or without TSA treatment was determined by the MTT assay. TSA inhibited cell proliferation in both cell lines. We characterized the effects of TSA on MMP-9-mediated proteolysis as assessed by zymography. In vitro Matrigel invasion assays showed that the HDAC inhibitor resulted in a significant decrease in cancer cell invasion. Furthermore, Gö6976, a PKC inhibitor, significantly inhibited HGF-induced HDAC-5 expression and invasion. These results demonstrated that HDACs regulate HGF-induced uPA and MMP-9 expression of gastric cancer cells, and the PKC pathway may play an important role in HGF-mediated cell invasion in gastric cancer cells, suggesting that such activities serve as anti-tumor mechanisms of the HDAC inhibitor.

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 513.
Abstract 5139: Trop2 modulates beta1 integrin-mediated adhesion and migration of prostate cancer cells
Authors: Trerotola Marco , Alberti Saverio , Languino Lucia R.
Published: February 2011

Trop1 and Trop2 are highly related transmembrane glycoproteins, which are overexpressed in many human carcinomas and have been suggested to contribute to cancer progression. Recently, Trop2 has been discovered as an important marker for identification of candidate tumor-initiating cells in a mouse model of prostate cancer (PrCa), suggesting that this protein is a novel regulator of PrCa progression.

In this study, we show that Trop2 was expressed at high levels in aggressive human PrCa cells, and was co-localized with beta1 integrins, which are known to modulate cell adhesion and invasion and to be deregulated in PrCa. We demonstrate that Trop2 inhibited adhesion of PC3 PrCa cells on fibronectin (the major ligand for beta1), but strongly enhanced their ability to migrate on fibronectin as compared to controls. This effect was specific for cells seeded on fibronectin, since no effect was observed on vitronectin.

Confocal microscopy studies using PC3 cells seeded on fibronectin showed strong co-localization of Trop2 with beta1 in leading edges. On the other hand, Trop2 did not co-localize with beta5, an integrin that did not affect migration of PC3 cells on fibronectin. We did not observe any change of beta1 expression in Trop2-overexpressing PC3 cells compared to controls, suggesting that affinity for fibronectin or clustering in focal complexes might be modulated by Trop2. Our studies on Trop2-overexpressing PC3 cells seeded on fibronectin demonstrated that beta1 was in fact displaced from focal adhesions to relocate to Trop2-containing leading edges.

Our findings demonstrate a novel cross-talk between Trop2 and beta1 integrins in PrCa cells. We hypothesize that this interplay can result in destabilization of firm adhesion to fibronectin and accelerated re-organization of dynamic structures responsible for migration/invasion of PrCa cells.

MT is recipient of a fellowship from the Italian Foundation for Cancer Research (FIRC).

Grant Support: NIH R01 CA109874 (LRL).

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 5139.
Abstract 5149: Nucleostemin is a key factor in maintenance of human glioblastoma-derived cancer stem cells
Authors: Izquierdo Marta L. , Gil Jon
Published: February 2011

Glioblastomas (GBM) may contain a variable proportion of active cancer stem cells (CSCs) capable of self-renewal, of aggregating into CD133+ neurospheres, of retaining their multi-lineage potential over time, and to develop intracranial tumors that phenocopy the original ones. We hypothesized that nucleostemin may contribute to cancer stem cell biology as these cells share characteristics with normal stem cells. We will report that nucleostemin is expressed in GBM-CSCs isolated from patient samples. The significance of its expression was addressed by targeting the corresponding mRNA using lentivirally transduced short hairpin RNA (shRNA). Knockdown of nucleostemin in GBM-CSC abolishes proliferation and induces apoptosis. Furthermore, GBM-CSCs with decreased nucleostemin levels failed to form neurospheres in vitro or tumors when xenotransplanted into the brains of immunocompromised rats. These findings suggest a central role of nucleostemin in regulating proliferation, self-renewal and survival of GBM-CSC, and targeting its expression may offer a new therapeutic approach for this incurable type of brain 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 5149.
Abstract 5154: Human umbilical cord matrix stem cells transfected with the interferon-β gene markedly attenuated the growth of bronchioloalveolar carcinoma xenografts in mice
Authors: Kawabata Atsushi , Doi Chiyo , Matsuzuka Takaya , Rachakatla Raja Shekar , Ohta Naomi , Pyle Marla , Marini Frank , Troyer Deryl , Tamura Masaaki
Published: February 2011

Mesenchymal stem cells derived from the human umbilical cord matrix (UCMSC) have great potential for therapeutic use in multiple diseases including cancer. UCMSC can be used as carriers for targeted delivery of genes or therapeutics and for local production of biologic agents. The therapeutic strategy using UCMSC as cellular vehicles for targeted gene delivery can avoid the problem of short life or excessive toxicity of a particular gene product in vivo. Interferon-β (IFN-β) has demonstrated a potent anti-tumor effect on many tumor cell lines in vitro. However, in vivo inhibition of tumor cell growth by IFN-β required blood concentrations much higher than the maximally tolerated dose. Therefore, clinical therapy by systemic administration of IFN-β was not possible. The aim of this study was to determine the anti-cancer effect of IFN-β gene transfected-UCMSC (IFN-UCMSC) on bronchioloalveolar carcinoma (BAC) cell lines in vitro and in vivo. The conditioned medium derived from IFN-UCMSC significantly attenuated cell growth of H358 and SW1573 BAC cell lines in vitro compared to the medium derived from non-transfected UCMSC. The percentage of dead cells, as measured by the trypan blue exclusion test, was significantly higher in the cells treated with the medium derived from IFN-UCMSC than UCMSC alone. The co-culture of IFN-UCMSC with cancer cells inhibited cell growth significantly through stimulation of apoptosis. Finally, systemic administration of IFN-UCMSC (0.3 million cells/injection, 5 day interval for four times) markedly attenuated tumor multiplicity and size of orthotopic H358 BAC xenografts in SCID mouse lungs. Histochemical analysis of the tumors indicated tumor-targeted localization of UCMSC and significant reduction of tumor cell proliferation index as measured by Ki-67 staining. These results clearly indicated that IFN-β production by IFN-UCMSC attenuated growth of BAC cells by inducing programmed cell death both in vitro and in vivo. This research clearly demonstrated that IFN-UCMSC are a powerful anti-cancer therapeutic tool as delivery vehicles for IFN-β. This work was supported by the KSU Terry C. Johnson Center for Basic Cancer Research, KSU Provost's Fund, KSU CVM Dean's Fund, NIH P20 RR017686 and a Joan's Legacy Research Grant.

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 5154.
Abstract 5166: Sex determining region Y-box 2 (SOX2) is a potential cell-lineage gene highly expressed in the pathogenesis of squamous cell carcinomas of the lung
Authors: Yuan Ping , Kadara Humam , Behrens Carmen , Tang Ximing , Woods Denise , Solis Luisa M. , Huang Jiaoti , Spinola Monica , Dong Wenli , Yin Guosheng , Fujimoto Junya , Kim Edward , Xie Yang , Girard Luc , Moran Cesar , Hong Waun Ki , Minna John D. , Wistuba Ignacio Ivan
Published: February 2011

Non-small cell lung cancer (NSCLC) represents the majority (85%) of lung cancers and is comprised mainly of adenocarcinomas and squamous cell carcinomas (SCCs). The sequential pathogenesis of lung adenocarcinomas and SCCs occurs through dissimilar phases as the former tumors typically arise in the lung periphery whereas the latter normally arise near the central airway. We assessed the expression of SOX2, an embryonic stem cell transcriptional factor that also plays important roles in the proliferation of basal tracheal cells and whose expression is restricted to the main and central airways and bronchioles of the developing and adult mouse lung, in NSCLC by various methodologies. Here, we found that SOX2 mRNA levels, from various published datasets, were significantly elevated in lung SCCs compared to adenocarcinomas (all p<0.001). Moreover, a previously characterized OCT4/SOX2/NANOG signature effectively separated lung SCCs from adenocarcinomas following integration with two independent publicly available gene expression microarray datasets and which correlated with increased SOX2 mRNA in SCCs. Immunohistochemical analysis of various histological lung tissue specimens demonstrated marked nuclear SOX2 protein expression in all normal bronchial epithelia, alveolar bronchiolization structures and premalignant lesions in SCC development (hyperplasia, dysplasia and carcinoma in situ) and absence of expression in all normal alveoli and atypical adenomatous hyperplasias. Moreover, SOX2 protein expression was greatly higher in lung SCCs compared to adenocarcinomas following analyses in two independent large tissue microarray (TMA) sets (TMA set I, n=287; TMA set II, n=511 both p<0.001). Furthermore, amplification of SOX2 DNA was detected in 20% of lung SCCs tested (n=40) and in none of the adenocarcinomas (n=17). Our findings highlight a cell-lineage gene expression pattern for the stem cell transcriptional factor SOX2 in the pathogenesis of lung SCCs and raise the intriguing possibility of the growth addiction of lung SCCs specifically to SOX2-dependent pathways. Supported in part by grants from the Department of Defense (W81XWH-04-1-0142 and W81XWH-07-1-03060), and the Specialized Program of Research Excellence in Lung Cancer grant P50CA70907.

Running title: SOX2 abnormalities in NSCLC

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 5166.
Abstract 5167: Notch1 is required for endothelial cell lineage differentiation from adult bone marrow stem cells in vitro
Authors: Shao Hongwei , Xiao Min , Cai Long , Radtke Freddy , Velazquez Omaida C. , Liu Zhao-Jun
Published: February 2011

Background and Objectives. Adult bone-marrow stem cells (BMSCs) are able to differentiate into endothelial progenitor cells (EPCs) and subsequently endothelial cells (ECs) under appropriate conditions. BMSCs-derived EPCs/ECs contribute to post-natal neovascularization and are important tools for regenerative medicine. However, the molecular mechanism determining specific differentiation of endothelial lineage remains unknown. The Notch signaling pathway plays an integral role in the differentiation of a variety of cell lineages. We hypothesize that Notch signaling is involved in controlling endothelial lineage differentiation from BMSCs and examined the role of Notch1 signaling in this process.

Methods and Results. An in vitro method for differentiation of endothelial cell lineage from murine BMSCs was developed. Mononuclear cells were isolated from mouse whole BM cells by light density separation using Ficoll-Paque® Plus, and cultured in the differentiation medium to differentiate endothelial cells. The endothelial phenotype of differentiated cells was characterized by immunostaining and flow cytometry analyses. Those fractions of cells with VEGFR2+/CD31+ or Tie2+/CD31+ were isolated and tested. VEGFR2+/CD31+ cells could form networks on the MatrigelTM, indicating that these cells are ECs. mRNA expression of Notch pathway components in VEGFR2+/CD31+ cells was examined by real-time RT-PCR. Multiple components of the Notch signaling pathway were expressed in the differentiated ECs. The effect of loss-of-function in Notch1 on the differentiation of endothelial cell lineage was examined using BMSCs from Notch1Flox/Flox mice. Cells expressing VEGFR2 were significantly decreased when Notch1 is deleted.

Conclusions. A novel and reliable method has been developed to differentiate murine adult bone marrow stem cells into endothelial cell lineage in vitro. The role of Notch1 in determining endothelial cell differentiation was tested on this platform. Our results implicated that Notch1 is required for the differentiation of endothelial cell lineage from BMSCs.

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 5167.
Abstract 5168: Interaction of sonic hedgehog and TP53/TP63 networks in lymphoma cells
Authors: Chari Nikhil S. , Teo Albert E.K. , Medeiros L. Jeffrey , McDonnell Timothy J.
Published: February 2011

Cellular homeostasis requires a delicate balance between proliferation, apoptosis and differentiation be maintained. These processes are regulated by a number of tumor suppressors and proto-oncogenes that are frequently deregulated during tumor progression. Amongst these are the TP53/TP63 family of tumor suppressors and the sonic hedgehog (SHH) pathway. The SHH network associated with proliferative stem cell niches of many organs has been shown to affect disease progression and metastasis in diverse cancers. We investigated the possibility of interactions between the SHH and the TP53 networks in cancer cells. Using a Ponasterone A inducible TP53 H1299 cell line as a model system we were, able to show alterations in RNA and protein levels of SHH signaling intermediates on TP53 activation suggesting a regulation at the transcriptional level. A preliminary screening of a number of hematological cancer cell lines showed a majority of cell lines having aberrant SHH activation. A panel of diffuse large B-cell lymphoma (DLBCL) cell lines that included activated B-cell (ABC) like DLBCL and germinal center B-cell (GCB) like DLBCL subgroups were selected for further study. These cell lines also varied from p53-deficient to wild type p53. Treatment of these cell lines with nutlin, an HDM2 antagonist showed alterations in the SHH pathway at the protein and transcript levels. In silico analysis of the promoters of genes in the SHH network reveal multiple potential TP53/TP63 binding sites. Luciferase promoter constructs of SHH pathway genes were generated including SHH-luciferase and SUFU-luciferase promoter constructs and confirmed that these promoters were regulated by both TP63 and TP53. Further validation was provided using in vivo chromatin immunoprecipitation (ChIP) studies and site-directed mutational analysis. These findings suggest that there are multiple interacting nodes between the components of the SHH network and TP53/TP63 family of transcription networks. Additionally, a therapeutic strategy involving specific and simultaneous targeting of these networks in DLBCL cells would be expected to result in a more robust therapeutic response.

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 5168.
Abstract 5171: EGFR induction of SOX9 is required in a cellular migration program induced by urothelial injury and maintained in cancer
Authors: Ling Shizhang , Chang Xiaofei , Lee Thomas , Marchionni Luigi , Netto Georges , Sidransky David , Berman David M.
Published: February 2011

Data from a variety of tissue types and experimental systems indicate that the transcriptional regulator SRY-related HMG box 9 (SOX9) can be induced by injury and promote migration as well as tumor growth. We hypothesize that in cancers associated with chronic injury, such as urothelial carcinoma (UroCa), SOX9 may mediate epithelial injury repair. We found that urothelial SOX9 was induced during times of rapid proliferation during bladder organogenesis and injury repair. High levels of epidermal growth factor (EGF) ligands have been noted in urine and represent a potential trigger for urothelial injury repair programs. Using specific antagonists, we found that EGF receptor activation by cognate ligands heparain binding EGF-like growth factor (HB-EGF) and/or amphiregulin was responsible for Sox9 induction in urothelial cells and tissues. As expected, EGFR activation occurred through the mitogen associated protein kinase (MAPK) pathway. With chronic breaches of urothelial integrity, EGFR-Sox9 stimulation may promote urothelial carcinogenesis, as Sox9 expression was significantly upregulated in invasive human UroCa samples and cell lines. When expressed in established tumors, SOX9 may confer a selective advantage to cancer cells by promoting their ability to invade and colonize new sites. As support for this idea, small interfering RNA inhibition of SOX9 levels abrogated in vitro wound healing and invasion, suggesting a role for SOX9 in tumor spread. These results indicate a potential mechanism for urothelial carcinogenesis through an EGFR-MAPK-SOX9 cascade, and a further function for this axis in established cancers. Future studies will examine the potential mechanisms supporting perseverative SOX9 expression in cancer and opportunities for disrupting this pathway as a therapeutic strategy.

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 5171.
Abstract 5173: Adipocytes attract leukemia cells
Authors: Pramanik Rocky , Sheng Xia , Mittelman Steven D.
Published: February 2011

Obesity is associated with increased progression and relapse of various types of cancer. We have previously found that obesity impairs leukemia treatment by mitigating chemotherapeutic drug. In addition, we observed the presence of acute lymphoblastic leukemia (ALL) cells in adipose tissue during chemotherapy treatment. Therefore, we have investigated whether adipocytes stimulate leukemia cell migration in addition to exerting their protective effect against chemotherapies. We used a cell migration assay in which murine ALL cells were cultured in 5μm pore Transwell inserts in an upper chamber, and test medium or adipocyte-conditioned medium (ACM) was used in the bottom chamber. The number of cells which migrated into the lower chamber in 90 minutes was counted using trypan blue exclusion and expressed as % of total cells initially plated in the transwell. When the upper Transwell inserts contained regular medium and the lower chamber contained ACM, 31.2±5.8% of cells migrated to the lower chamber. By contrast, when the lower chamber contained regular medium, only 0.3±0.2% of cells migrated (p<0.01, vs ACM, n=4). Preadipocyte-conditioned medium also stimulated migration, but less than ACM (18.2±4.8% cells migrated, p<0.05 vs. ACM, n=4). The ACM-stimulated migration had both directional and non-directional components, in that when both the upper and lower chamber contained ACM, 13.2±0.2% of cells migrated to the lower chamber, which was significantly more than with regular media (p<0.05), but less than with ACM (p<0.01). The above results suggest that adipocyte-secreted molecules increase ALL cell motility and attract leukemia cells in a directional fashion. It is plausible that adipose tissue, particularly in obese people, might attract leukemia cells and act as a safe heaven for leukemia cells against chemotherapy, thereby promoting leukemia relapse.

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 5173.
Abstract 5179: Inflammatory breast cancer cell motility: Understanding the role of E-cadherin and gli1 in IBC migration and the development of a novel high content assay to assess cell migration
Authors: Thomas Zainab , Sexton Jonathan Z. , Devi Gayathri R. , Williams Kevin P.
Published: February 2011

Inflammatory Breast Cancer (IBC) is a highly aggressive subtype of breast cancer and is now regarded as clinicopathologically distinct from neglected locally advanced breast cancer (LABC). In addition, African-Americans, native Americans and younger women are disproportionately affected with IBC having the worse survival. The molecular mechanisms underlying the invasiveness of IBC are not clear. Interestingly, upregulation of the integral membrane protein E-cadherin is the marker reported to be seen most consistently in IBC. In contrast to IBC, E-cadherin is typically reduced in most cancers and the subsequent loss in cell-cell adhesion results in the rapid progression to an invasive and metastatic phenotype. Inactivation of the E-cadherin cell adhesion system can occur by a number of genetic and epigenetic mechanisms. Gli1, the main transcriptional activator of hedgehog signaling, can promote loss of E-cadherin. Gli1 induces expression of Snail which is then found to downregulate E-cadherin expression.

In an IBC cellular model, SUM149, we have identified elevated levels of Gli1. Further, inhibition of Gli1 using siRNA caused apoptosis in these cells. We hypothesize that the invasiveness of IBC cells is modulated in part by a connection between E-cadherin and Gli1 levels in IBC and that modulating this pathway will lead to changes in IBC cell motility. This project has two main objectives: 1), Investigate the expression levels of E-cadherin and Gli1 in IBC cells and their roles in IBC motility, and 2), Develop and validate a high content microscope-based assay for measuring motility of IBC cells. Cell motility assays were performed in six-well plates with Gli1 siRNA treated SUM149 cells that resulted in reduced motility and viability of these cells compared to control siRNA treated cells. These results suggest that targeting Gli1 can reduce IBC's invasiveness. This pathway provides opportunities for identifying molecules that might affect tumor invasiveness.

We have also developed a novel live cell high content phenotypic screen for modulators of IBC cell motility that may potentially lead to new therapeutics for IBC. An understanding of how any of these molecules affect the Gli-Snail-E-cadherin system would provide useful tools for better probing the role of this pathway in IBC. Furthermore, the high content migration phenotypic assay should be generally applicable to investigating a range of invasive cancer types.

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 5179.
Abstract 5187: Migration and proliferation of colon carcinoma cells are importantly affected by leptin and adiponectin via diverse pathways
Authors: Ratke Janina , Niggemann Bernd , Zänker Kurt S. , Dittmar Thomas , Lang Kerstin
Published: February 2011

Obesity is a dramatically increasing public health problem worldwide. Traditionally, fat tissue was considered to be solely an energy storage depot. However, recent studies have shown that adipose tissue exerts important endocrine functions, which are predominantly mediated by a network of various soluble factors derived from adipocytes. New evidence has come to light elucidating a modulatory role of this adipocytokines in the regulation of cancer development. The most prominent adipocytokines are leptin and adiponectin, which are predominantly secreted by differentiated adipocytes. Depending on the energy status of adipocytes, the secreted concentrations of leptin and adiponectin vary drastically. In obesity, serum level of leptin is considerable increased (greater than or equal to 100 nanogram/ml), whereas the adiponectin concentration is very low. In contrast, adiponectin concentration accelerates by weight reduction (greater than or equal to 10 microgram/ml). The aim of our study was to investigate the influence of adipocytokines on cancer cells. Using our three-dimensional, collagen-based migration assay we found a strong impact of leptin and adiponectin on the migration of colon cancer cells. Both adipocytokines alone significantly enhanced the migratory activity of SW480 colon carcinoma cells from 30.3±7.4% spontaneously locomoting cells to 50.9±8.2%. In contrast, both adipocytokines in combination reduced the migration of colon carcinoma cells to control level. Moreover, treatment of the cells with leptin alone enhanced the proliferation of SW480 cells up to 50%, whereas adiponectin mediated an anti-proliferative function by 30%. Surprisingly, addition of leptin and adiponectin together did not have any effect on the proliferation. On the molecular level, leptin-induced migration was mediated by phosphorylation of focal adhesion kinase (FAK) and activation of phosphatidyl-inositol-3-kinase (PI3K). The pro-migratory effect of adiponectin was accompanied by an activation of PI3K/AKT and various transcription factors. In contrast, incubation with both adipocytokines in combination led to a decreased phosphorylation of FAK and AKT. Understanding the impact of leptin and adiponectin alone and their interplay on the carcinogenesis of colon cancer as well as the underlying molecular mechanisms have a major clinical significance, and provide the basis for the development of novel therapeutics to treat obesity-associated colorectal cancer.

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 5187.