bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1280: Nano-scale phospho-proteomic analysis to define diagnostic signatures and biomarkers of therapeutic activity in cancer
Authors: Fan Alice C. , Banerjee Prajna , Xu Liwen , Kong Christina , Sabatti Chiara , Wilhelm Francois , Greenberg Peter , Felsher Dean W.
Published: August 2012

The ability to quantify changes in protein activity in a clinical setting is important for the development of therapeutics that target cancer signaling pathways. We have developed the use of a highly sensitive microfluidic nano-immunoassay (NIA, Nanopro1000) to quantify unphosphorylated, single- and multi- phosphorylated isoforms of proteins in clinical specimens including blood, bone marrow, and fine needle aspirates (FNAs). Using as little as 2 nanoliters of cellular lysate, we can measure over 40 proteomic parameters, and assess changes in proteins that mediate signal transduction, cell cycle progression and apoptosis. Using NIA to resolve different phosphorylated isoforms of individual proteins and directly measure percent phosphorylation has led to new hypotheses and approaches to biomarker development. To develop novel diagnostic biomarkers, we have analyzed MAP Kinase phospho-protein profiles for over 200 specimens collected from patients with hematopoietic and solid tumors. We found that quantifying even subtle differences in protein signaling between tumor calls and adjacent non-tumor tissue can distinguish various malignancies. We will also present preclinical and clinical data using NIA to develop biomarkers of therapeutic activity biologic activity of tyrosine kinase inhibitors (TKIs) in myelodysplastic syndrome and leukemia. One unanticipated finding has been that TKIs might preferentially inhibit or activate specific phospho-isoforms of proteins. Our NIA studies also suggest that dynamic changes in protein signaling may better predict therapeutic outcome than profiles from a single, pre-treatment time point. One example is that the decrease in single phospho-ERK2 (p-ERK2) is associated with response to TKIs in chronic myelogenous leukemia. Notably, measurements of the phospho-isoform at a single time point were not sufficient to distinguish the two groups, which may reflect that individual tumors may have different initial “set-points” for the pathway. In contrast, comparison of proteomic signatures serially sampled from patients before and during treatment can provide valuable information regarding degree of target kinase inhibition and also compensatory signaling that can not be gleaned from analysis of a single time point. Our studies using nanoscale proteomic analysis to profile tumor signatures and characterize biologic response to therapies have led to new potential strategies for developing biomarkers, moving us closer to individualized cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1280. doi:1538-7445.AM2012-1280
Abstract 1287: High mobility of triple-negative breast cancer cells is due to repression of plakoglobin gene by SLUG
Authors: Bailey Charvann K. , Mittal Mukul K. , Misra Smita , Chaudhuri Gautam
Published: August 2012

One of highly pathogenic breast cancer cell types are the SLUG-high claudin-low triple negative (negative in the expression of estrogen, progesterone and ERBB2 receptors) breast cancer (TNBC) cells. These cells are highly metastatic and have low levels of the motility regulatory catenin plakoglobin. In order to link high levels of the transcriptional repressor SLUG in the TNBC cells with low levels of plakoglobin we found that SLUG inhibits the expression of plakoglobin gene directly in these cells and thus, among other downstream effects, help disseminating these tumor cells. Overexpression of SLUG in the SLUG-deficient cancer cells significantly decreased the levels of mRNA and protein of plakoglobin. On the contrary, knockdown of SLUG in SLUG-high cancer cells elevated the levels of plakoglobin. Overexpression of SLUG in the SLUG-deficient cells elevated the invasiveness and motility of these cells. On the other hand, knockdown of plakoglobin in these low motility non-invasive breast cancer cells did not affect the ability of the cancer cells to penetrate Matrigel matrix but increase the growth and migration rates of these cells. This study thus implicates high levels of SLUG and low levels of plakoglobin as determinants in the progression of highly disseminating breast cancer. Supported in parts by the DOD-CDMRP grants BC050641, BC086542 and BC103645 to GC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1287. doi:1538-7445.AM2012-1287
Abstract 1289: Cooperation between beta-catenin, Snail, Slug, and Twist is required for maximum repression of the human vascular endothelial-cadherin promoter
Authors: Lopez Dayami , Carter W. Bradford
Published: August 2012

Vascular endothelial (VE)-cadherin is a key protein in the adherens junctions. This protein forms homophilic intercellular interactions and is required for the integrity of the endothelial monolayer, endothelial permeability, and control of cell growth. We have previously demonstrated that exposure of endothelial cells to breast cancer cell-conditioned media results in upregulation of Twist, Slug, and Snail expression, which in turn downregulate the activity of the human VE-cadherin promoter through at least two E-box motifs. Since these repressors have been linked to the β-catenin signaling pathway, we chose to determine whether β-catenin could regulate the activity of the human VE-cadherin promoter. Herein, we demonstrate that β-catenin represses the VE-cadherin promoter either alone or in coordination with the repressors Twist, Slug, and Snail. Electrophoretic mobility shift assays showed that the VE-cadherin promoter contains four β-catenin/LEF-1 binding motifs (designated Site-1-4) that appear to be involved in this regulatory process. In addition, we identified TGFβ as one of the factors released by breast cancer cells that could be involved in the regulation of the VE-cadherin gene when HDMEC are exposed to breast cancer cell conditioned medium.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1289. doi:1538-7445.AM2012-1289
Abstract 1290: The ability of the transcription factor p63 to induce selected gene expression modules associated with mesenchymal to epithelial transition of prostate cells
Authors: Olsen Jan Roger , Ke Xi-Song , Rostad Kari , Hellem Margrete R. , Qu Yi , Lin Biaoyang , Lorens James B. , Micklem David R. , Haugen Hallvard , Gravdal Karsten , Halvorsen Ole Johan , Akslen Lars A. , Rotter Varda , Oyan Anne Margrete , Kalland Karl H.
Published: August 2012

Genome-wide gene expression analysis identified p63 as one of the most consistently under-expressed transcription factors in prostate cancer samples compared to matched benign prostate tissue. Two alternative p63 promoters generate two different N-terminal variants, TA or ΔN which contains and lacks the transactivating domain, respectively. Differential splicing generates multiple additional isoforms. The activity and interactions of the different isoforms remain unresolved. Isoform-specific real-time quantitative PCR assays revealed that ΔNp63α is the predominant isoform in epithelial prostate tissues, but other isoforms are detectable. Our group has established an experimental cell culture model based upon primary, immortalized prostate epithelial cells (EP156T cells), where p63 was shut down when EP156T cells underwent epithelial-to-mesenchymal transition (EMT) to become EPT1 cells. Subsequently, several additional mesenchymal like cell subtypes with additional malignant features, including anchorage independent growth in soft agar and ability to grow at much higher density in monolayers, were derived from EPT1 cells. All of these subtypes exhibited very low p63 expression. To examine the potential of p63 to induce mesenchymal to epithelial transition (MET) in this EMT model, we have stably re-expressed the ΔNp63α and TAp63α isoforms in different mesenchymal like cells of the model using retroviral/lentiviral vector transductions. Genome-wide gene expression analyses and phenotypic cellular assays revealed that the ΔNp63α isoform induced re-expression of multiple genes involved in cell adhesion. But this was not associated with complete MET induction. Cells re-expressing ΔNp63α obtained an intermediate morphology with both epithelial and mesenchymal traits, and with actin filaments organized in stress fibers. ΔNp63α re-expression significantly compromised migratory but not invasive ability of the cells. Based upon these findings we want to investigate whether p63 may compromise epithelial to mesenchymal plasticity in an animal model of tumorigenic cells and if there is differential effect on primary tumor growth compared to metastatic growth.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1290. doi:1538-7445.AM2012-1290
Abstract 1293: Galectin-3 facilitates the stability of p21 protein through carbohydrate recognition domain in prostate cancer cells
Authors: Wang Yi , Raz Avraham
Published: August 2012

Galectin-3 (Gal-3), a β-galactoside-binding protein, has multiple functions such as regulating cell growth, apoptosis, angiogenesis, tumor metastasis and so on. p21cip/waf-1, a cyclin-dependent kinase inhibitor, also regulates apoptosis and cell growth as well. Gal-3 and p21 have some similar functions, and both of them play important roles in tumor progression. Here, we try to understand how Gal-3 regulates p21 protein in prostate cancer cells. Our study showed that the changes of expressions of Gal-3 and p21 in prostate cancer cells are parallel. Functions of Gal-3 in regulating apoptosis and cell growth are partially mediated by p21. Gal-3 protein helps to prevent the degradation of p21 protein and delay the turnover of p21 in prostate cancer cells. The protective effect of Gal-3 on the stability of p21 protein is attenuated by Gal-3 inhibitor lactose and modified citrus pectin, which suggests that carbohydrate recognition domain of Gal-3 is responsible for its effect on the stability of p21. Our study is helpful to further understand the molecular mechanisms of Gal-3 functions and demonstrate the crosslink between Gal-3 and p21 pathways.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1293. doi:1538-7445.AM2012-1293
Abstract 1297: Engrailed-2 (EN2) represses the transcription of the CST6 tumour suppressor gene in prostate cancer
Authors: Morgan Richard , Simpson Guy , Michael Agnieszka , Pandha Hardev S.
Published: August 2012

Engrailed-2 (EN2) is a homeodomain-containing transcription factor that is expressed in prostate and breast tumours but not in normal adult tissue. Although EN2 protein is known to be secreted by prostate tumours and is a potential urine based biomarker of this disease, its function in prostate tumours is unknown. In this study we have used siRNA-mediated knock down of EN2 as well as forced over expression in prostate cancer cells. Although neither treatment has a significant effect on cell survival, we identify two key transcriptional targets of EN2, the tumour suppressor gene CST6 and the Death Receptor ligand TRAIL, the transcription of which are repressed and activated by EN2, respectively. EN2 synergises with epigenetic changes to silence CST6, and may function to block CST6 expression very early in the development of malignancy. Further, we show that EN2 protein can regulate gene transcription in non-malignant prostate cells when supplied externally, indicating that EN2 secretion by prostate tumour cells may be able to modify its microenvironment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1297. doi:1538-7445.AM2012-1297
Abstract 1420: Microsomal prostaglandin E2 synthase 1 (mPGES-1) is expressed in neuroblastoma and may represent a novel target for therapy
Authors: Kock Anna , Rasmuson Agnes , Idborg Helena , Johnsen John Inge , Kogner Per
Published: August 2012

Neuroblastoma (NB) is an embryonic tumor of the sympathetic nervous system and it is the most common extracranial tumor affecting young children. Despite a very intensive treatment regimen high-risk neuroblastoma patients still have a poor prognosis. Neuroblastoma cells are enriched in arachidonic acid, over express COX-2 and produce prostaglandin E2 (PGE2) that acts as an autocrine or paracrine survival and proliferation factor by induction of Akt signaling. Downstream of the COX enzymes, specific synthases converts PGH2 into the different prostaglandins. Microsomal prostaglandin E2 synthase-1 (mPGES-1) specifically converts PGH2 to PGE2 and is thought to primarily couple to COX-2. The aim of this study is to investigate if inhibition of mPGES-1 could represent an alternative therapeutic approach to COX- inhibition in neuroblastoma. We could detect expression of mPGES-1 in a panel of seven human neuroblastoma cell lines. Stable knock down of mPGES-1 expression using shRNA significantly reduced the clonogenic capacity and treatment with the mPGES-1 inhibitor CAY1052 inhibited neuroblastoma cell growth in vitro. By immunohistochemistry and western blot we could also detect expression of COX-1, COX-2 and mPGES-1 in tumors from the transgenic TH-MYCN mouse model of neuroblastoma. Treatment of homozygous TH-MYCN mice with 10 mg/L diclofenac for two weeks, starting at the age of four weeks, significantly reduced tumour weight compared to untreated animals. Ex vivo analysis with LC-MS/MS of tumor tissues from diclofenac treated animals revealed a significant decreased level of COX metabolites compared to control. The expression of mPGES-1was not affected by the treatment. Furthermore the number of cells staining positive for cleaved caspase-3 were higher in the treated tumors indicating induced apoptosis. In conclusion, our results show that mPGES-1 is expressed in neuroblastoma therefore mPGES-1 might represent an alternative way of reducing NB growth through specific PGE2 inhibition.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1420. doi:1538-7445.AM2012-1420
Abstract 1439: Loss of expression of SWI/SNF subunits in high risk Wilms tumor is accompanied by global gene expression changes
Authors: Gadd Samantha L. , Gerhard Daniela S. , Smith Malcom A. , Auvil Jaime Guidry , Huff Vicki , Geller James I. , Dome Jeffrey , Huang Chiang-Ching , Perlman Elizabeth J.
Published: August 2012

Introduction: The TARGET initiative seeks to identify therapeutic targets through genomic sequencing supported by copy number and gene expression analysis. The High Risk Wilms Tumor TARGET project, currently in progress, includes the analysis of favorable histology (FH) tumors that relapsed and tumors with anaplasia (unfavorable histology). We report the identification of a group of Wilms tumors (WT) identified by a unique gene expression pattern. Methods and Results: 102 high risk WT (43 anaplastic, 59 relapsed FH) were analyzed using Affymetrix U133+2 chips. Unsupervised hierarchical analysis consistently selected 9 tumors (5 anaplastic, 4 FH) that shared a striking gene expression pattern. These patients presented at 15-96 months (median 51); 7 had a triphasic histology and 2 were blastemal predominate; 4 died of their disease. When the top differentially expressed genes (P<0.0001) were analyzed with PANTHER, significant repression (p=5x10−7) of genes responsible for the regulation of chromatin architecture was apparent. Among the most significantly down-regulated were ARID1A (fold change (FC) 0.2, p=5.2x10−8) and SMARCA4 (BRG1, FC 0.16 p=9x10−6), among others. Transcriptomic analysis using hierarchical clustering based on chromosomal position displayed a unique pattern of alternating increased and decreased expression of small genetic regions for all chromosomes in this group. These findings may represent long-range genomic regulation that has been attributed to histone modifications and are in keeping with the role of the SWI/SNF chromatin remodeling complex. Analysis using the Affymetrix 6.0 SNP chip demonstrated LOH for 1p31 (ARID1A) and 19p13 (BRG1) in 2 and 1 of the 9 tumors, respectively. Global methylation analysis is underway. Conclusion: Mutations in several SWI/SNF subunits (including ARID1A, BRG1, SMARCB1, PBRM1) have been implicated in the development of a wide range of neoplasias. In normal cells, SWI/SNF subunit protein levels are tightly controlled in a coordinated fashion in order to maintain constant cellular levels. Little is known about the transcriptional regulation of these subunits. This report suggests that a subset of Wilms tumors show striking repression of many SWI/SNF subunits, resulting in a unique globally abnormal gene expression pattern that has not been previously described in tumors with ARID1A or SMARCB1 loss. Whole genomic or exomic sequencing of our tumor set is in progress.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1439. doi:1538-7445.AM2012-1439
Abstract 1458: Hyperthermia and radiotherapy in combination with curcumin prevent tumor cell migration and progression of breast cancer
Authors: Natarajan Mohan , Aravindan Sheeja , Herman Terence S. , Aravindan Natarajan
Published: August 2012

We have demonstrated that hyperthermia (HT)-induced nitric oxide regulates NF-kB, and that potentiates radiotherapy (RT)-induced breast cancer cell death. As a follow-up, in this study we investigated the efficacy of pharmacologically-safe bioactive curcumin (CUR) in combination with HT and therapeutic doses of radiation in regulating breast cancer progression. Human breast adenocarcinoma (MCF-7) cells were either mock-irradiated, exposed to 2Gy (IR), exposed to HT (43oC for 1 h) and /or treated with CUR in the presence or absence of radiation exposure. The treated/irradiated cells were examined for (i) transcriptional modulations of ninety three molecules that are directly involved in tumor invasion and metastasis; (ii) cellular localization of cMYC, p65, p53, MMP2 and MMP9; (iii) cellular migration; (iv) metabolic activity and (v) cell death. Compared to mock-IR control, QPCR profiling demonstrated a significant modulation of tumor invasion and metastasis molecules in IR exposed cells. Conversely, combinations like ‘HT with IR’, ‘CUR with IR’ or ‘CUR with HT’ robustly inhibited IR-induced and in parallel conferred IR suppressed tumor invasion and metastasis molecules in this setting. More importantly, CUR combined with both HT and IR significantly inhibited (through blocking seventy four genes) tumor invasion and metastasis molecules. Scratch wound assay demonstrated complete inhibition of tumor cell migration in CUR-HT-IR exposed cells as opposed to every stand alone and combinations investigated. Likewise, cytotoxic assay, MTT showed a significant cell killing with CUR-HT-IR exposure compared to all other groups. Furthermore, we observed significant and robust apoptotic characteristics in CUR-HT-IR exposed cells. These data together with our earlier report clearly demonstrate that CUR in combination with HT and IR may significantly inhibit breast cancer progression through combinatorial regulation of tumor invasion and metastasis molecules. The findings further imply that CUR may significantly enhance the IR dose reduction factor at least for the treatment of breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1458. doi:1538-7445.AM2012-1458
Abstract 1460: Comparison of radiosensitizing effects of several PARP inhibitors under clinical investigation
Authors: Yoo Stephen S. , Cerna David , Wilsker Deborah , Teicher Beverly A. , Coleman C. Norman
Published: August 2012

Targeting tumor DNA repair is an attractive strategy in selectively killing tumors while sparing normal tissues. Inhibiting Poly (ADP-ribose) polymerase (PARP) in cancers with BRCA1 and BRCA2 mutations has been shown to be a promising approach either as a single agent or in combination therapy with chemotherapeutics or ionizing radiation. To systematically compare the radiosensitizing efficacy of various PARP inhibitors we have evaluated the long-term cytotoxicity of ABT-888, AZD2281, MK4827, and BSI-201 in clonogenic survival assays in combination with ionizing radiation. For these studies we used the breast cancer cell lines MDA-MB 231 BR, a subline of MDA-MB-231 selected for its tendency to metastasize to brain (BRCA1+/−) and MX-1 (BRCA1 deletion and BRCA2 mutation). Furthermore, we have evaluated PARP inhibitors under normoxic and hypoxic conditions (21% O2 versus 0.1% O2). In MDA-MB 231 BR cells, at a clinically relevant and achievable concentration for ABT-888 (J Clin Oncol. 2009; 27(16)) and AZD2281(N Engl J Med. 2009; 361(2)), and at 1 uM MK4827 showed radiosensitizing effects under both normoxic and hypoxic conditions. BSI-201 at a clinically achievable concentration (ASCO 2010, Abstract 2012) did not exhibit radiosensitizing effects under the same conditions. When MX-1 cells were treated with BSI-201 under hypoxic conditions (0.1% O2) the cytotoxicity of BSI-201 was evident. Taken together, the results show that ABT-888, AZD2281 and MK4827 show promising radiosensitizing effects under both normoxic and hypoxic conditions whereas the cytotoxicity of BSI-201 was observed only under hypoxic conditions in BRCA1/2 mutant carrier MX-1cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1460. doi:1538-7445.AM2012-1460
Abstract 1472: Comprehensive evaluation of PEGylated gold nanorods for two photon photoluminescence image guided radiation therapy enhancement
Authors: Kumar Rajiv , Patel Janki , Korideck Houari , Berbeco Ross I. , Makrigiorgos Mike G. , Sridhar Srinivas
Published: August 2012

Nanoparticle formulations of gold have shown a tremendous potential in various biomedical applications. The use of high atomic number (Z) materials presents an attractive approach in enhancing the therapeutic efficacy of the radiation therapy. The high Z number for gold (Z=79) makes them an ideal candidate for radiosensitization enhancement. The ability of the anisotropic gold nanorods to sustain the resonating surface plasmon with minimal damping results in highly efficient two-photon induced photoluminescence imaging. Here we present the synthesis and in vitro characterization of PEGylated gold nanorods as efficient radiosensitizing agents which can be imaged using inherent two-photon photoluminescence without a conjugated fluorophore. We have synthesized gold nanorods with an aspect ratio of 2.5 and functionalized the surface with different ratios of methoxy and amine PEG for modulating the charge and imparting the functional groups. The cellular uptake behavior in prostate cancer cell line PC3 was studied and the results indicated a robust uptake of the amine functionalized nanorods. The quantitative estimation of the nanorods uptake was performed with BCA assay as amount of nanorods present in per microgram of protein. Two photon photoluminescence imaging also confirmed a robust cellular uptake of the amine functionalized nanorods. The radiation damage enhancement of the gold nanorods was confirmed using PC3 cells. The cells treated with gold nanorods were irradiated with kilovoltage X-rays. A γH2AX assay was used for quantitation of the DNA damage with and without nanorods as controls. The results indicated an increased highly efficient DNA damage in cells treated with gold nanorods. The outcome of this research will enable further application of targeted nanorods in advanced animal models as efficient radiation dose enhancement agents wherein two-photon photoluminescence imaging will provide a real time assessment of the therapeutic response and disease progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1472. doi:1538-7445.AM2012-1472
Abstract 1481: microRNA mediated reprograming of fibroblasts into cancer associated fibroblasts in ovarian cancer
Authors: Mitra Anirban K. , Zillhardt Marion , Hua You-Jia , Tiwari Payal , Murmann Andrea E. , Peter Marcus E. , Lengyel Ernst
Published: August 2012

Cancer associated fibroblasts (CAFs) are a major constituent of the tumor stroma and promote the invasion and growth of cancer cells. However, the mechanism by which cancer cells transform normal fibroblasts into CAFs is unclear. While it is clearly established that microRNAs are deregulated in cancer and are involved in cellular reprograming, it is not known if they play a role in the transformation of normal fibroblasts into CAFs. Therefore, we investigated the novel possibility of microRNA mediated reprogramming of normal fibroblasts into CAFs. Primary CAFs extracted from the metastasis of patients with ovarian cancer and normal omental fibroblasts (NOFs) extracted from non-cancer patients were profiled for microRNA expression. The functional role of microRNAs in CAFs was established by assessing their ability to promote ovarian cancer co-invasion, colony formation and tumor growth in mice. The microRNA targets were identified using gene array analysis, validated by 3′UTR luciferase reporter assay, and functionally verified both in vitro and in mouse xenografts. Comparing CAFs and NOFs we found that, in ovarian cancer CAFs miR-31 and miR-214 were downregulated while miR-155 was upregulated as compared to normal or tumor-adjacent fibroblasts. Mimicking this deregulation by transfecting microRNAs and microRNA inhibitors induced a phenotypic and functional conversion of normal fibroblasts into CAFs, and the reverse experiment resulted in the reprograming of CAFs into normal fibroblasts. The microRNA induced CAFs and patient-derived CAFs shared multiple upregulated genes highly enriched in chemokines important for CAF function. The most highly upregulated chemokine, CCL5/RANTES, was found to be a direct target of miR-214. CCL5 was functionally important for both CAFs and normal fibroblasts reprogrammed into CAFs, in vitro and in mouse xenografts. These results indicate that tumor cells can reprogram fibroblasts to become CAFs through the regulation of their microRNAs, suggesting that targeting microRNA in stromal cells, as well as in tumor cells, could be beneficial for cancer treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1481. doi:1538-7445.AM2012-1481
Abstract 1484: Functional and molecular characterization of macrophage-tumor cell communication
Authors: Schmall Anja , Herold Susanne , Vipotnik Natasha , Al-tamari Hamza M , Pullamsetti Soni Savai , Grimminger Friedrich , Seeger Werner , Savai Rajkumar
Published: August 2012

Lung Cancer is the leading cause of cancer deaths worldwide. It is increasingly appreciated that the tumor stroma, consistent of a heterogeneous population of non-neoplastic host cells, is an essential part of cancer initiation, growth and progression. Tumor-associated macrophages (TAMs) can influence cancer progression and metastasis, but the responsible mechanisms remain unclear. Here, we focus on the involvement of bone-marrow derived macrophages (BM- MM) and two of their major chemotactic pathways (CX3CR1-CX3CL1 and CCR2-CCL2 axis) in cancer proliferation and migration. We demonstrate that conditioned medium (CM) derived from co-cultures of lewis lung carcinoma (LLC1) cells with BM- MM (purity >99%) enhances proliferation of LLC1-cells significantly from 1.79±0.1717 to 2.43±0.2067 in an absorbance based BrdU-Assay and a higher colony-number was found in a colony-formation-assay compared to control. In addition LLC1-migration in presence of CM was 5 fold increased compared to controls (172.9±2.18 versus 33.93±2.29). Further, analysis of the cytokine profile of co-culture-derived CM showed regulation of pro-inflammatory cytokines (Interleukin-6 and Interleukin-1α), anti-inflammatory cytokines (Interleukin-10 and Interleukin-1RA) and several chemokines (CCL5 and CCL2). Importantly, evaluation of tumor growth in chemokine receptor knockout mice, CX3CR1-KO and CCR2-KO demonstrated reduced tumor size compared to wild type mice. CX3CR1-KO and CCR2-KO tumor showed reduced in vivo proliferation as assessed by PCNA immunostaining (12.46%±3.188 versus 46.45%±2.922). Further analysis of the tumor microenvironment in CX3CR1-KO and CCR2-KO demonstrated a significant decrease in monocyte/ macrophage accumulation compared to control tumors as assessed by FACS. Compared to the percentage of macrophages in control tumor (7.518±0.78%), a significant decrease was observed in CCR2-KO (2.96± 0.233%) and CX3CR1-KO (3.975±0.6019%) tumors. Based on these findings, we conclude that macrophage plays a crucial role in lung cancer progression and the knowledge of tumor-host interactions can provide novel therapeutic approaches for lung cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1484. doi:1538-7445.AM2012-1484
Abstract 1487: Tumor-derived microvesicles induce proangiogenic phenotype in endothelial cells via endocytosis
Authors: Kawamoto Taisuke , Ohga Noritaka , Akiyama Kosuke , Hirata Naoya , Maishi Nako , Osawa Takahiro , Yamamoto Kazuyuki , Kondoh Miyako , Shindoh Masanobu , Hida Yasuhiro , Hida Kyoko
Published: August 2012

Background: Increasing evidence indicates that tumor endothelial cells (TEC) differ from normal endothelial cells (NEC). Our previous reports also showed that TEC were different from NEC. For example, TEC have chromosomal abnormality and proangiogenic properties such as high motility and proliferative activity. However, the mechanism by which TEC acquire a specific character remains unclear. To investigate this mechanism, we focused on tumor-derived microvesicles (TMV). Recent studies have shown that TMV contain numerous types of bioactive molecules and affect normal stromal cells in the tumor microenvironment. However, most of the functional mechanisms of TMV remain unclear. Methodology/Principal Findings: Here we showed that TMV isolated from tumor cells were taken up by NEC through endocytosis. In addition, we found that TMV promoted random motility and tube formation through the activation of the phosphoinositide 3-kinase/Akt pathway in NEC. Moreover, the effects induced by TMV were inhibited by the endocytosis inhibitor dynasore. Our results indicate that TMV could confer proangiogenic properties to NEC partly via endocytosis. Conclusion: We for the first time showed that endocytosis of TMV contributes to tumor angiogenesis. These findings offer new insights into cancer therapies and the crosstalk between tumor and endothelial cells mediated by TMV in the tumor microenvironment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1487. doi:1538-7445.AM2012-1487
Abstract 1499: Periprostatic fat from obese patients promotes prostate cancer growth
Authors: Venkatasubramanian Palamadai N. , Crawford Susan E. , Fitchev Philip S. , Brendler Charles B. , Plunkett Beth A. , Abroe Betsy , Mafi Massa , Morgan Gina , Cornwell Mona L. , O'Leary Judy , Pruitt Jacklyn , Stadnik Agnieszka , Burright Marna J. , McGuire Michael S. , Wyrwicz Alice M. , Doll Jennifer A.
Published: August 2012

Background: Obesity, particularly visceral obesity, increases the risk of prostate cancer (PCa) progression. However, the mechanism of this remains unclear. Here, we hypothesized that the visceral periprostatic fat (PPF) from obese PCa patients stimulates progression as compared to subcutaneous fat (SQF) or PPF from lean patients. Methods: PPF and SQF were collected from PCa patients. MTT assays were performed on endothelial and PC-3 PCa cells to test proliferative activity in explant culture tissue conditioned media (CM). Angiogenesis was evaluated ex vivo using MR to generate T2 relaxation times from a series of T2-weighted spin-echo images (Bruker 14.1 T imager). Results: PPF CM from an obese patient increased endothelial cell proliferation ∼5 times that of PPF CM from a lean patient (P<0.01). In contrast, SQF CM, from lean or obese patients, showed only minimal activity. When tested on PC-3 cells, SQF CM from obese or lean patients only weakly promoted proliferation. In contrast, PPF CM significantly induced proliferation; however, obese PPF CM was significantly higher than that of lean PPF CM (P<0.05). Interestingly, the MR data revealed key differences between PPF and SQF and between lean and obese fat (Table 1). In all patients, the PPF T2 was lower than that of the SQF (P<0.01). And, the PPF T2 from obese patients was lower than that of lean patients. A lower T2 reflects elevated hemoglobin levels and thus could serve as an indirect measure of vascularity. And, our data suggest that vascularity is increased in obese PPF as compared to SQF and lean PPF tissues. Conclusions: Our data reveal that PPF has increased angiogenic and tumorigenic capacity as compared to SQF and this potential is markedly increased by the presence of obesity. Adipokines and/or other molecules expressed by the PPF may modify the PCa microenvironment to potentiate tumor growth. Our results also support the potential use of MR clinically as a non-invasive test of the angiogenic potential of PPF in PCa patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1499. doi:1538-7445.AM2012-1499
Abstract 1513: Functional analysis of TGFBR1 mutations in breast cancer associated fibroblasts
Authors: Moore-Smith Lakisha D. , Manne Kanika , Caldwell Charisma , Frost Andra , Pasche Boris
Published: August 2012

The TGFBR1*6A variant consists of a GCG repeat deletion resulting in a three-alanine deletion in the TGFBR1 coding region which has been linked to breast cancer risk. We have screened 30 breast cancer cell lines including: 20 epithelial (2 from normal breast) and 10 CAFs (2 from normal breast) for the presence of this mutation. Initial screening indicates that 45% of the epithelial cell lines screened harbor the *6A mutation. Of the 8 fibroblast screened 67% harbored TGFBR1*6A. The following CAF cell lines were used for the remainder of our functional studies: Hs748 (9A/9A), Hs841.T (9A/6A) and Hs742.T (6A/6A) genotypes. CAFs were treated with 100pM TGF-β resulting in decreased growth inhibitory response of 20% for Hs742.T (*6A/*6A) cells when compared to Hs841.T (9A/*6A) cells with 42% inhibition. Invasion assays, using a matrigel-coated Boyden Chamber, indicated a higher invasive capacity for Hs841.T (9A/6A) fibroblast cell line than in the Hs742.T (6A/6A) or Hs748.T (9A/9A). Western blot analysis was used to determine cell differentiation status. Cells which harbor TGFBR1*6A showed an increased expression of α-sma when compared to the WT fibroblasts indicating progression to a more aggressive myofibroblast phenotype. Studies are underway to reproduce these data in additional CAF cell lines, however, this preliminary data suggest that TGFBR1*6A enhances breast cancer development by enhancing tumor-associated fibroblast oncogenic properties in addition to its direct effect on breast tumor cells. TGF-β signaling was altered in TGFBR1*6A cells which may alter normal tumor-stromal signaling pathways. Future studies, using primary cell lines derived from a transgenic mouse model harboring the TGFBR1*6A allele will provide an optimal system to dissect the functional effects of this mutation in tumor and stromal cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1513. doi:1538-7445.AM2012-1513
Abstract 1518: Multiple myeloma cells cross talk with bone marrow stroma lead to induction of DKK1 expression and produce bortezomib resistance
Authors: Ryu Jiyeon , Jung Woo-june , Lee Chansu , Park Soyoung , Park Juwon , Bae Eunkyung , Ahn Kwang-Sung , Yoon Sung-Soo
Published: August 2012

The importance of the micro-environment in tumor progression is now well established. Dynamic interaction between multiple myeloma (MM) cells and bone marrow stromal cells (BMSCs) plays critical role in the progression of MM including drug response and bone erosion. Firstly, we found that DKK1 expression was statistically increased in MM cells (MOLP8, KMS12BM, KMS12PE, NCIH929, LP1, MOLP2, EJM, U266, RPMI8226, and 536MM, respectively) co-cultured with BMSCs obtained from MM patients when compared to MM cells alone (p<0.05). Also, MM cells co-cultured with BMSCs were significantly more resistant against 25 nM bortezomib than MM cells alone (p<0.05). We, therefore, established a bortezomib-resistant cell line using U266 cells (U266/velR), and explored the characteristics of U266/velR cells. Cytotoxic effect of bortezomib in U266/velR was 1.5 folds lower than U266 cells, and the cross-resistance against thalidomide was observed. DKK1 expression in U266/velR was higher than that in parental cells. We found that elevated levels of p-p65 were detected in U266/velR, and the degree of p65 and I-κB expression levels reduced by bortezomib was different between U266/velR and U266. Elevated levels of p-p65/p65 were effectively suppressed by treating with NF-κB activator inhibitor (6-aminoquinazoline). Also, combined treatment of bortezomib and 6-aminoquinazoline reduced the expression of DKK1 in co-culture of U266 cells and BMSCs. The expression of HGF in CD138− fractions of MM bone marrow cells was higher than that in its CD138+ fractions. Whereas, IL-6 and OPN in CD138+ fraction of MM bone marrow cells was higher compared to its CD138− fractions. In co-culture of U266 and BMSCs, IL-6, OPN and HGF was dramatically increased compared to U266 alone and BMSC alone (p<0.01; p<0.05). Taken together, cytokines and growth factor regulated by dynamic interplay between MM cells and BMSCs contributes the bortezomib response and bone erosion.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1518. doi:1538-7445.AM2012-1518
Abstract 1556: α-mannosylceramide, a new NKT cell agonist, induces tumor immunity without anergy induction in NKT cells
Authors: O'Konek Jessica , Takao Satomi , Izhak Liat , Illarinov Petr , Besra Gurdyal , Berzofsky Jay , Terabe Masaki
Published: August 2012

NKT cells are CD1d-restricted T cells recognizing lipid antigens. When NKT cells are activated with a prototypic antigen, α-galactosylceramide (α-GalCer), they induce strong tumor immunity. Strong preclinical evidence for α-GalCer's ability to induce tumor immunity through NKT cell-activation led to translation into therapy of cancer patients. However, so far minimal success has been achieved in patients. We reasoned that having an α-linked sugar moiety in the structure of α-GalCer can be a target for anti-α-linked galactose natural antibodies, which are a cause of acute xenograft rejection, and which might abrogate the biological activity of α-GalCer. After testing multiple glycosylceramides with potential agonistic activity for NKT cells, we discovered that α-mannosylceramide (α-ManCer) has strong activity to induce tumor immunity through a TNF-α- and NOS-dependent mechanism that is distinct from the IFN-γ dependent protection induced by α-GalCer. Another downside of α-GalCer treatment is induction of long-lasting anergy in activated NKT cells. Since we did not observe massive cytokine production with α-ManCer, in contrast to α-GalCer, we asked whether this new compound activates NKT cells without inducing long-lasting anergy in vivo. In contrast to α-GalCer, α-ManCer did not induce long-lasting anergy of activated NKT cells. This was correlated with the observation that α-ManCer induced only a short term expression of PD-1 on activated NKT cells, which has been reported to be responsible for α-GalCer-induced long-term anergy. Furthermore, while prior treatment with α-GalCer abrogated the protective effect of either α-GalCer or α-ManCer against CT26 colon carcinoma tumor challenge for at least two months, the pre-treatment with α-ManCer had no effect on the protection induced by either NKT cell agonist. Since α-ManCer can activate human NKT cells, this new NKT cell agonist may provide a better opportunity for NKT cell targeted immunotherapy in cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1556. doi:1538-7445.AM2012-1556
Abstract 1557: High mobility group box 1 secretion supports tumor progression of human malignant mesothelioma
Authors: Jube Sandro , Rivera Zeyana , Bianchi Marco , Powers Amy , Wang Ena , Pagano Ian , Pass Harvey I. , Gaudino Giovanni , Carbone Michele , Yang Haining
Published: August 2012

Human malignant mesothelioma (MM) is an aggressive and highly lethal cancer, often associated with exposure to asbestos and erionite. Prognosis is poor, due to late-stage diagnosis and resistance to current conventional therapies. We previously showed that high-mobility group box-1 protein (HMGB1), a damage-associated molecular pattern (DAMP) protein, is involved in the early stages of mesothelial cell transformation. Here we show that HMGB1 establishes an autocrine circuit in MM cells influencing tumor cell proliferation and survival. MM cells express HMGB1 at high levels and secrete HMGB1 into the extracellular space. Accordingly, HMGB1 levels in MM patients’ sera are significantly higher than in those of healthy individuals. In addition, motility, survival and anchorage-independent growth of HMGB1-secreting MM cells were inhibited in vitro by a monoclonal antibody (mAb) against HMGB1, by the recombinant HMGB1 competitive antagonist BoxA and by antibodies against the receptor for advanced glycation end products (RAGE). Inhibition of HMGB1 reduced the growth of MM xenografts in SCID mice and extended survival. Our findings indicate that MM cells become “addicted” to HMGB1 and that targeting HMGB1 can be a promising novel therapeutic approach for MM.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1557. doi:1538-7445.AM2012-1557
Abstract 1813: Evaluation of TBK1 as a novel cancer target in the K-Ras pathway
Authors: Vangamudi Bhava , Ayres Aimee E. , Burke Jason P. , Waterson Alex G. , Rossanese Olivia W. , Fesik Stephen W.
Published: August 2012

TBK1 (TANK-binding kinase 1) is a serine/threonine kinase best known for its role in mediating the innate immune response through modulation of the interferon and NF-kB pathways. TBK1 has also been implicated in supporting cancer cell survival and transformation as the downstream effector of the small GTPase RalB, and is overexpressed in lung, breast and colon tumors. Recently, a high-throughput RNAi screen identified TBK1 as a synthetic lethal partner of oncogenic K-Ras, suggesting that small molecule inhibition of TBK1 may provide a novel therapeutic strategy for K-Ras mutant tumors. Given the high tractability of kinases as drug targets, we sought to confirm that kinase inhibition recapitulated the synthetic lethal effects observed with shRNA silencing. We examined two small molecule inhibitors of TBK1, BX795 (6 nM) and GSK2292978A (4 nM), in a panel of lung, pancreas and colon cancer cell lines. We found only a small subset of lines that were sensitive to TBK1 inhibition and this sensitivity did not correlate with K-Ras dependence. We then used K-Ras and TBK1 siRNA in a similar panel of cell lines. As anticipated, knockdown of K-Ras using siRNA produced significant growth and signaling defects in K-Ras dependent cell lines. In contrast, knockdown of TBK1 did not result in a significant alteration of growth in any of our cell lines. Neither TBK1 siRNA nor TBK1 inhibitors revealed a sensitivity that correlated with K-Ras dependence in our lung, pancreatic, or colorectal cancer cell lines. Our data disputes the proposed synthetic lethal relationship of oncogenic K-Ras and TBK1 and does not support development of TBK1 inhibitors for treatment of K-Ras driven tumors.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1813. doi:1538-7445.AM2012-1813