bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4254: Human neuroendocrine Bon 1 cells as a three-dimensional model for the study of human neuroendocrine tumor therapy
Authors: Xu Eugenia Y. , Wong Chung , Levine Arnold J. , Vosburgh Evan
Published: August 2012

Pancreatic neuroendocrine tumors are rare tumors, with an incidence of one to two per 100000 individuals per year, and they account for 1-2% of all pancreatic neoplasms. Currently, much needed is to develop the therapeutical drugs for patients with pancreatic neuroendocrine tumors. Human pancreatic neuroendocrine BON 1 cell line is widely used for testing the therapeutic drugs in vitro. Here, we develop an alternative research model that better mimics the in vivo tumor tissue organization and microenvironments. This method produces in vitro three-dimensional (3D) tumor spheroids from human pancreatic BON1 cells. The 3D tumor spheroids generated in vitro exhibit the characteristics of avascular tumors. Structurally and genetically the 3D spheroids show similarity to the xenograft BON1 tumor and are more tumorigenic than 2D monolayer BON1 cells. We also present how to use these 3D spheroids to screen the therapeutic drugs. This method will help us establish human neuroendocrine tumor spheroids for 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 4254. doi:1538-7445.AM2012-4254
Abstract 4268: Gene signature of host irradiation from Trp53 null mammary tumors detects radiation-preceded human cancers and has prognostic value for sporadic human breast cancer
Authors: Nguyen David H. , Fredlund Erik , Zhao Wei , Perou Charles M. , Balmain Allan , Mao Jian-Hua , Barcellos-Hoff Mary Helen
Published: August 2012

The microenvironment mediates many important aspects of tumorigenesis, providing complementary mechanisms that work in concert with mutagenic challenges to initiate and promote tumor development. To test the effect of ionizing radiation, a carcinogen known to act via the microenvironment, we employed a radiation-genetic chimera in which an irradiated mouse was transplanted with unirradiated Trp53 null mammary epithelium. Host irradiation not only decreased tumor latency, but unexpectedly enriched for tumors that were negative for estrogen receptor-α (Cancer Cell 19:40-51, 2011). Using gene expression microarrays we identified a profile containing 323 genes (323-IHC) that could separate the tumors that arose in irradiated hosts from those that arose in naive mice. To evaluate the relevance of an irradiated microenvironment in human cancer, the 323-gene expression signature was applied to expression profiles of sporadic or radiation-preceded human cancers. Clustering using the 323-IHC was able to detect radiation history in thyroid carcinomas from Chernobyl and radiotherapy associated sarcomas even though there was no overlap with the differential gene profiles published for either data set, demonstrating the relevance of microenvironment reprogramming as one of the fundamental facets of radiation carcinogenesis. We next utilized a gene expression data set representing 1,608 human breast cancers compiled from 10 independent studies. Clustering of these samples, using genes from our mouse-derived radiation signature that were also present on the breast cancer gene expression platform, segregated four subgroups that were enriched for intrinsic subtypes, and two major groups with differential relapse-free and overall survival. A 72-gene centroid derived from the 323-IHC classified the breast cancers as either similar to tumors from host-irradiation, or similar to tumors from non-irradiated hosts. Classification of the human breast cancer expression profiles revealed an association between host-irradiation and estrogen receptor negative, basal-like breast cancers. Thus, the biology represented in the 323-IHC signature is present in human cancers and associates with etiology and clinical outcome.

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 4268. doi:1538-7445.AM2012-4268
Abstract 428: Post-translational phosphorylation is essential for nanog-mediated tumorigenesis
Authors: Xie Xiujie , Piao Longzhu , Cavey Greg S. , Teknos Theodoros N. , Pan Quintin
Published: August 2012

Objectives: Nanog is a transcription factor intimately involved in embryonic stem cell pluripotency. Emerging evidence indicates that Nanog plays a critical role in oncogenesis, in part through regulation of the cancer initiating cell population. We investigated the role of post-translational phosphorylation as a regulatory mechanism to control Nanog function in head and neck squamous cell carcinoma (HNSCC). Methods: Liquid chromatography-mass spectrometry/mass spectrometry was used to identify the PKC∈-mediated phosphorylation sites on Nanog. Phosphorylation-insensitive Nanog mutants (T200A and T280A) were generated and assessed for in vitro and in vivo phenotype. Chromatin immunoprecipitation (ChIP) and co-immunoprecipitation experiments were performed to determine Nanog recruitment and association with p300. ChIP, electrophorectic mobility shift assay, and luciferase reporter assay were used to assess Nanog regulation of Bmi-1. Forward and reverse genetic experiments were carried out to determine the role of the Nanog-Bmi-1 signaling node in HNSCC. Results: Phosphopeptide mapping identified that Nanog is phosphorylated by PKC∈ at multiple resides, including T200 and T280. Phosphorylation-insensitive Nanog mutants (T200A and T280A) showed a loss-of-function phenotype due to impaired p300, a transcriptional co-activator, recruitment resulting in a defect in target gene promoter activation. Ectopic expression of phosphorylation-insensitive Nanog mutants in HNSCC cells significantly reduced cell proliferation, colony formation, invasion, migration, and the cancer initiating cell population. Additionally, in vivo tumor initiating ability was severely compromised in HNSCC cells expressing phosphorylation-insensitive Nanog mutants; 87.5% (14/16), 12.5% (1/8), and 0% (0/8) for empty vector, T200A, and T280A, respectively. Nanog occupied the promoter of Bmi-1 at two distinct regions to directly regulate Bmi-1 levels. Genetic ablation and rescue experiments demonstrate that Bmi-1 is a critical downstream signaling node for the pleotropic pro-oncogenic effects of Nanog. Conclusion:Taken together, our study revealed, for the first time, that post-translational phosphorylation of Nanog is essential to directly regulate Bmi-1 and promote tumorigenesis.

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 428. doi:1538-7445.AM2012-428
Abstract 4290: Visualizing immune surveillance in lung metastasis progression
Authors: Zal M. Anna , Bartkowiak Todd , Chodaczek Grzegorz , Papanna Veena , Shanmugasundaram Meenakshi , Zal Tomasz
Published: August 2012

As stipulated by the Paget and Fuchs’ “seed and soil” analogy, the establishment of metastases depends on the microenvironment of target tissue; as such, it also depends on the tissue immunosurveillance system. Anti-tumor immune responses are subdued in advanced tumors by various immune suppressory mechanisms, such as by tumor associated CD4 FoxP3 regulatory T-reg cells. But how immune suppression sets in at the sites of metastasis, and how it could be overcome there remains unclear. Using multiphoton and confocal microscopy, we have visualized the immune cell's dynamics that is triggered by various metastatic cancers in the lungs. We found that single, blood-carried cancer cells that settle in the capillary bed triggered strikingly rapid recruitment of CD11c dendritic cells (DCs), which were heterogeneous. Some of those DCs wrapped around the singular cancer cells while other probed their intracellular content by acquiring tumor-derived vesicles. These phagocytic DCs then carried away their cargo towards the draining lymph nodes, thereby initiating tumor antigen cross-presentation. After 24-48 hours, micrometastases became surrounded by T cells that were highly enriched in T-reg. The motility patterns and cell depletion experiments showed that T-reg largely ignored tumor cells but were attracted by the phagocytic tumor-associated DCs. In result of these interactions, the phagocytic DCs became the foci of highly dynamic three-cell interactions between T-reg and the effector T cells, sequestering most T cells at the tumor margin. In this context, we tested the impact of immunomodulatory therapies. The immune cell dynamics that we uncovered in metastases was responsive to therapeutic immune modulation aimed at the tumor-associated phagocytic DCs and also at Programmed Death (PD-1) pathways. Taken together, our intravital dynamic imaging results highlight the duality of metastasis-associated phagocytic DCs. On the one hand, these cells provide tumor antigen cross-presentation and recruit T cells to nascent secondary tumors, but on the other hand, they aid metastasis progression by promoting early on the contact of tumor-infiltrating T cells with T-reg.

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 4290. doi:1538-7445.AM2012-4290
Abstract 4292: In vivo and in vitro imaging of Cyclooxygenase-2 in tumor
Authors: Wong Kenneth , Urban Konnie , Ang Angel , Zhang Ning , Singh Rajendra , Kim Jae-Beom
Published: August 2012

Cyclooxygenase-2 (Cox-2) is a biomarker for tumor progression and inflammatory diseases. It is known that normal tissues express minimal amounts of Cox-2 proteins while inflammatory and malignant tumor cells express high levels. Thus, inhibition of Cox-2 is a strategy in the treatment of tumor progression and inflammatory diseases. Recently, many different kinds of molecular imaging reagents for Cox-2 have been developed. Among them, an indomethacin-based Cox-2 probe, fluorocoxib, has been shown to bind to Cox-2 in vitro and in vivo. The binding specificity to Cox-2 was shown using cell culture and tumor xenograph models. In our present study, in order to optimize fluorocoxib imaging conditions, we utilized HCT116 and HT29 cells as low and high Cox-2 expressers, respectively. First, cultured cells were grown overnight on cover slips. Cells were then fixed with paraformaldehyde and incubated with fluorocoxib probe. Multispectral images were taken using a camera equipped with liquid crystal tunable filters. Spectral unmixing was applied to these images to enhance fluorocoxib signals. Our results indicate that HCT116 cells express fair amounts of Cox-2 protein. We also transplanted HCT116 and HT29 cells into the flank regions of nu/nu mice subcutaneously. After tumors grew to sufficient size (∼100 mm3) we injected fluorocoxib intravenously and monitored probe binding to the tumors. Whole animal in vivo imaging indicated that fluorocoxib binds to HT29 tumors after 3 hours. All the probes were eliminated from the animals by 24 hours. On the other hand, HCT116 tumors did not show strong binding of fluorocoxib as expected. However, HCT116 tumors did show some weak binding at the 3 and 6 hr time points. Once there was no longer any probe detected in vivo, we dissected the primary tumors from the HCT116 and HT29 implantations. Tissue sections were made with paraffin embedded tumors. After deparaffinization, tissue sections were incubated with fluorocoxib and fluorescent images were taken using multispectral imaging technology. Spectral unmixing data confirmed that HCT116 tumor sections showed fair amounts of Cox-2 probe expression. The amount of binding was also quantitated using a corresponding software package. Our results demonstrated that fluorescent biomarker probes can be imaged in vitro and in vivo non-invasively. Furthermore, we showed that the expression level can be quantitated using spectral unmixing imaging technology.

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 4292. doi:1538-7445.AM2012-4292
Abstract 4296: Ultrasmall gadolinium-loaded polymer nanoparticles for imaging in glioblastoma
Authors: Himes Benjamin , Zhou Jiangbing , Patel Toral , Coman Daniel , Hyder Fahmeed , Saltzman Mark
Published: August 2012

Glioblastoma multiforme (GBM) is a devastating disease for which prognosis remains exceptionally poor even with maximal intervention. A central challenge in the treatment of GBM is realizing effective drug delivery to the tumor and surrounding brain tissue, due to the inability of many drugs to cross the blood-brain barrier (BBB). The use of nanoparticles comprised of biodegradeable polymer as vehicles for intracranial drug delivery circumvents the BBB and allows for sustained, controlled release of drug in a targeted area. Nanoparticle encapsulation also protects therapeutic payloads from premature degradation in the microenvironment. However, effective distribution of nanoparticles in brain tissue has emerged as a potential hurdle to therapeutic efficacy. To this end we have synthesized ultrasmall poly(lactic-co-glycolic acid) (PLGA) nanoparticles 60-80nm in size, which demonstrate pronounced improvement in volume of distribution relative to larger particles. This enables particle diffusion into normal brain tissue, where the pore size of the extracellular matrix is limited to 38-64nm, allowing for drug delivery to the infiltrative edge to the tumor. Further, as the utility of these nanoparticles lies in their ability to deliver drug in a clinically relevant area, it is vital to develop a translatable means of imaging intracranial particle delivery to ensure adequate distribution. To address this issue we have developed gadolinium-loaded ultrasmall PLGA nanoparticles. These particles are detectable by MRI and distribute over a wide area when administered with the aid of convection-enhanced delivery (CED). These particles, composed entirely of FDA-approved materials, provide a diagnostic means to visualize distribution of nanoparticles in a clinically applicable capacity.

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 4296. doi:1538-7445.AM2012-4296
Abstract 430: MiRNA profiling of prostate cancer cell-derived large oncosomes identifies a signature of invasion and metastasis
Authors: Morello Matteo , Dudley Andrew C. , Schow Peter , Adam Rosalyn M. , Morley Samantha , Mulholland David , Insabato Luigi , Wu Hong , Klagsbrun Michael , Bhowmick Neil A. , Rubin Mark A. , D'Souza-Schorey Crislyn , Freeman Michael R. , Vizio Dolores Di
Published: August 2012

Introduction: We recently demonstrated that amoeboid prostate cancer (PCa) cells shed large (1-10µm diameter), bioactive membrane particles, representing a new class of tumor microvesicles termed large oncosomes (LO). LO can transfer oncogenic signals horizontally within the tumor microenvironment, as demonstrated by activation of phospho-tyrosines and Akt in recipient cells. Methods: We used differential ultracentrifugation and immuno-flow cytometry with size beads to isolate oncosomes in the 1-10µm diameter range. The method was applied to culture medium as well as to the plasma of mice with prostate cancer. In order to identify oncosome miRNAs relevant to metastasis, we performed a GeneChip® miRNA 2.0 Array screen of oncosomes shed by tumorigenic RWPE-2 PCa cells. Non-tumorigenic RWPE-1 cells were also analyzed. Results: LO contained protease and significantly stimulated migration of tumor and endothelial cells. LO also contained Caveolin-1, the GTPase ARF6 and other proteins that we used as markers to identify them in the circulation of mice with PCa, and in murine and human tissues with metastatic disease. Oncosomes also contained RNA. The miRNA profile of the oncosomes was compared to the miRNA profile of the cells of origin. While the miRNA profile of RWPE-2 almost completely matched the profile of RWPE-1 cells, the profiles differed significantly between LO isolated from the two cell lines. In particular, RWPE-2-derived LO contained miR-1228*, miR-150*, miR-373*, miR-135a*, miR-34b and miR-491-3p, which have been implicated in cancer cell proliferation and migration. In particular, the miRNAs miR-135a* and miR-373* target and inhibit the focal adhesion kinase (FAK), which plays a role in promoting a mesenchymal to amoeboid transition. miR-135a* and miR-373* have also been demonstrated to stimulate cell invasion, and are up-regulated in metastatic prostate and breast cancer. Target prediction analysis identified members of pathways involved in metabolic deregulation, and in oncogenic intercellular signaling and cell-cell interaction specifically in the LO purified from the tumorigenic cell line. Conclusions: Collectively, our results indicate that tumor cells can use oncosome formation to actively export pro-oncogenic miRNAs and that oncosomes therefore serve as a potential source of biomarkers of tumor 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 430. doi:1538-7445.AM2012-430
Abstract 4304: A microfluidic invasion assay to investigate the mechanisms involved in prostate cancer metastasis to bone
Authors: Bischel Lauren L. , Casavant Benjamin P. , Beebe David J.
Published: August 2012

Background: The majority of cancer patient mortality can be attributed to the spread of the cancer to secondary sites. Bone represents one of the most preferential sites for metastases from prostate and other cancers. While there has been much investigation into the underlying mechanisms involved in the homing of prostate cancer metastases to bone, there is still much that remains to be elucidated. Methods: We present a microfluidic invasion assay that can be used to investigate the mechanisms by which prostate cancer cells metastasize to bone. The assay uses a controlled microenvironment with a simple readout to observe the formation of cellular protrusions during invasion. By flowing fluid through an extracellular matrix (ECM) hydrogel in a microfluidic channel, an ECM hydrogel with a continuous lumen patterned through the microfluidic channel can be formed. Cancer cells can then be seeded into the lumen and invasion of the cells into the ECM hydrogel can be quantitatively analyzed using microscopy after 24 hours. The assay can be altered to model invasion in different microenvironments by altering the ECM hydrogel composition and pre-seeding the channels with cell representative of a secondary site before the ECM coating is formed. Results: In cocultures of LNCaPs (a prostate cancer cell line) or C4-2B (a bone metastatic LNCaP derived prostate cancer cell line) in the lumen with MC3T3s (osteoblast cell line) pre-seeded on the side of the microchannel, the C4-2Bs invaded more readily than LNCaPs. When C4-2Bs in the lumen are cultured with MC3T3-E1s on the side of the microchannel an 8-fold increase in the percentage of invasive C4-2Bs was observed compared to C4-2B alone. A 5-fold increase in invasiveness was observed when C4-2Bs were cultured with conditioned media from a mixed culture of C4-2Bs and MC3T3-E1s. Conclusions: These results indicate that cross-talk between prostate cancer cells and osteoblasts could play a role in the targeting of prostate cancer metastases to bone. This assay offers several advantages over traditional macroscale invasion assays (e.g. transwell assays) including the capability to control and vary the matrix composition and to perform the assay with a smaller number of cells. The microchannels can be arrayed allowing many different conditions to be tested in a single platform. Additionally, the assay set-up can be performed with automated liquid handling systems further increasing the ability to assay different conditions in a high-throughput manner. These capabilities will allow in vitro studies not previously possible which could yield new insights into the mechanisms controlling prostate cancer cell homing to bone.

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 4304. doi:1538-7445.AM2012-4304
Abstract 4306: Lack of TGF-β signaling induces a switch from single to collective cell migration in vivo
Authors: Matise Lauren A. , Palmer Trenis D. , Ashby William J. , Nashabi Abudi , Chytil Anna , Aakre Mary , Gorska Agnieszka , Pickup Michael W. , Zijlstra Andries , Moses Harold L.
Published: August 2012

Transforming growth factor-beta (TGF-β) has a dual role during tumorigenesis initially as a suppressor and then as a promoter. In stromal fibroblasts, TGF-β signaling suppresses tumorigenesis in adjacent epithelia, while its ablation potentiates tumor formation. Concurrently, epithelial TGF-β signaling regulates fibroblast recruitment and activation. Yet, the role of TGF-β in stromal-epithelial migration during tumorigenesis is unknown. We hypothesize that TGF-β is a critical regulator of tumor-stromal interactions that promote mammary tumor cell migration and invasion. We are using the chicken embryo chorioallantoic membrane (CAM) model to study this interplay by grafting both breast cancer epithelial cells, isolated from either TGF-β receptor II knockout (TβRII-deficient) or TβRII floxed (control) mice, and fibroblasts onto the CAM. Intravital microscopy revealed fibroblast stimulation of either a single cell/strand migration of control cells or collective migration of TβRII-deficient cells, both along the vasculature. Epithelial clusters at the epithelial-stromal boundaries of TβRII-deficient tumors express E-cadherin, p120, and β-catenin, while control epithelia in these regions express αSMA. TβRII-deficient tumors also exhibit a two-fold greater local CAM metastasis than control tumors. Epithelial tumor RNA isolated by laser capture microdissection was analyzed on a qPCR array. In TβRII-deficient epithelium compared to control epithelium, we found upregulation of Igfbp4 and Tspan13 and downregulation of Col1α2, Bmp7, Wnt11, Gng11, Vcan, Tmeff1, and Dsc2. Western and qPCR analyses validated these targets in vitro. For current and future studies determining the relevance of these targets to migration patterning, we achieved Wnt11 overexpression in TβRII-deficient cells, as well as knockdown of Wnt11 and Tmeff1 in control cells. Our findings about TGF-β signaling in stromal-epithelial interactions are important in identifying migratory mechanisms that can be targeted as recourse for breast 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 4306. doi:1538-7445.AM2012-4306
Abstract 4307: The effect of ablating ADF/cofilin pathway during skin homeostasis and tumorigenesis
Authors: Zhou Jing , Patel Hitesh , Brunton Valerie G. , Frame Margaret C.
Published: August 2012

Proteins of the cofilin pathway are key regulators of actin dynamics. There are three cofilin isoforms encoded by human and mice. Of these, ADF-cofilin(ADF) and cofilin1(COF) are expressed in most epithelial tissues while muscle-cofilin is not. Recent data has indicated that components of the cofilin pathway(ADF, COF) are frequently mis-regulated in cancer cells, but there are conflicting reports on how these cofilins and the upstream regulatory LIM kinases contribute to the malignant phenotype by their over-expression, or suppression. The data suggest that it is the balance of the cofilin pathway regulators, and the cofilin pathway “output” which determines the invasiveness of tumor cells. The cofilin pathway must therefore be considered as a whole. Therefore, we will address the role of the cofilin pathway in skin homeostasis and cancer by deletion of ADF and COF in the skin. We generated K14-Cre-ER/ADF(-/-)/COF(fl/fl) mice, which are nullizygous for ADF and have floxed COF under control of keratin14 Cre recombinase, as ubiquitous cofilin deletion is embryonic lethal. After 4-OHT-induced COF deletion, we examined the effects on skin homeostasis and skin tumorigenesis. However, the homozygous ADF and COF knockout mice died within a few days following induction of Cre. There was a thickening of the epidermis due to hyperplasia of the spinous layer of cells. A marked increase in apoptosis of epithelial cells was also evident, predominantly affecting cells of the basal layer, resulting in ulceration of the skin and oral mucosa. These animals become acutely sick due to the oral ulceration leading to reduced food and water intake. Immunohistochemical analysis showed that the “stress inducible” keratin6, which is usually observed in hyperproliferative conditions, was aberrantly expressed in the epidermis of the homozygous ADF and COF knock out mice. Both in vitro and in vivo experiments indicated that E-cadherin was lost from cell-cell contacts in double ADF/COF knockout keratinocytes. In addition, the double ADF/COF knockout keratinocytes migrated in a mesenchymal manner and their movement was 4 times faster than wild type keratinocytes, which retained cell-cell contacts and moved in collective manner. All these data indicate that the output of the cofilin pathway is likely to be involved in cancer cell processes, including migration, proliferation, differentiation and apoptosis. We are currently generating tumor-derived cell lines from invasive squamous cell carcinomas in which both ADF and COF have been deleted and analyzing cancer-associated behaviour both in vitro and in vivo.

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 4307. doi:1538-7445.AM2012-4307
Abstract 4321: EGFR signaling in glioblastoma regulates the expression of a brain specific isoform of alphaPIX/Arhgef6 to promote invasion
Authors: Babic Ivan , Gini Beatrice , Tanaka Kazuhiro , Jacobs Carolina , Yong William H. , Cloughesy Timothy F. , Mischel Paul S.
Published: August 2012

The gene Arhgef6 encodes the protein alphaPIX/Cool-2, a guanine nucleotide exchange factor specific for the Rho GTPases Rac1 and Cdc42. AlphaPIX is a protein of 90 kDa predominantly expressed in haemopoietic cells where it has been shown to have a role in cytoskeletal reorganization. However, Arhgef6 is also expressed in the brain and mutations in Arhgef6 have been linked to intellectual disability (ID) in humans. AlphaPIX mRNA was reported to be upregulated in glioblastoma multiforme (GBM), the most lethal form of brain cancer. We examined alphaPIX expression in several glioma cell lines and observed a smaller isoform migrating at 72 kDa. To characterize this isoform we performed 5′RACE from several glioma cell lines and show the smaller isoform is an amino-terminal truncated form of alphaPIX lacking the calponin homology (CH) domain. We demonstrate this isoform is expressed during post-natal brain development and its expression coincides with activation of mTOR. Examination of glioblastoma from both autopsy and biopsy clinical samples shows alphaPIX expression correlates with EGFR/PI3K/Akt/mTOR activation. EGF stimulation is shown to upregulate alphaPIX, and pharmacological inhibition of Akt inhibits alphaPIX expression. To examine the function of alphaPIX in glioblastoma we performed siRNA-mediated knockdown and demonstrate increased cell adhesion upon alphaPIX loss. In addition, knockdown of alphaPIX in glioma cells decreases invasive capacity and decreases the formation of invadopodia. These data demonstrate a previously uncharacterized isoform of alphaPIX is expressed during post-natal brain development, and is highly upregulated in GBM with activated EGFR signaling. A functional consequence of EGFR-mediated alphaPIX expression in glioma is increased invasion as a result of dysregulated cytoskeletal organization.

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 4321. doi:1538-7445.AM2012-4321
Abstract 4323: MED28 (magicin/EG-1) modulates MEK1 (MAP2K1)-dependent cellular migration in human breast cancer cells
Authors: Huang Chun-Yin , Hsieh Nien-Tsu , Chou Yu-Hsuan , Lee Ming-Fen
Published: August 2012

The objective of this study is to understand the role of MED28 (magicin/EG-1) in the aspect of cellular migration in human breast cancer cells. MED28 exhibits several cellular roles, including a mammalian Mediator subunit (MED28), a Grb2 and cytoskeleton-associated protein, a repressor of smooth muscle cell differentiation, and an endothelial-derived gene (EG-1). The association of MED28 with multiple proteins, including Grb2, Src, merlin, and actin, strongly suggests that MED28 is involved in many cellular signaling pathways. Specifically, several tumors, including breast cancer, highly express MED28; however, the fundamental mechanism is unclear at present. Using RNA interference (RNAi) technology, we found that suppression of MED28 blocked cellular migration and invasion with concomitant reduced expression levels of matrix metalloproteinase-2 (MMP2) and mitogen-activated protein kinase kinase 1 (MEK1; MAP2K1). In contrast, overexpression of MED28 enhanced cellular migration and up-regulated MMP2 and MEK1 expression. Moreover, MEK1 knockdown blocked MED28-induced MMP2 activation, cellular migration, and invasion as shown by gelatin zymography, wound-healing, and Matrigel invasion assays. Our data indicate that MED28 modulates cellular migration in a MEK1-dependent manner in human breast cancer cells, reinforcing the important cellular function of MED28. Funding: This work was supported by the National Science Council grant, NSC 98-2320-B-309-002-MY3, to M.-F.L.

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 4323. doi:1538-7445.AM2012-4323
Abstract 4326: The potential role of IMP-3 and PDPN in invadopodia formation and collective cancer invasion: Anti-invadopodia therapy as a therapeutic target
Authors: Hwang Young Sun , Park Kwang-Kyun , Chung Won-Yoon
Published: August 2012

Better understanding the mechanisms underlying the metastatic process is essential to developing novel targeted therapeutics. Recently, invadopodia have been increasingly recognized as important drivers of local invasion in metastasis. Invadopodia are actin-rich structures that have the ability to degrade the underlying extracellular matrix (ECM) and promote cell invasion. However, many questions concerning the mechanisms of invadopodia formation, regulation and function remain open to debate. Meanwhile, invasion of cells into the surrounding tissue and destruction of normal tissue architecture are two hallmarks of malignant tumors. Morphologically, two patterns of tumor invasion can be distinguished; single cell and collective cell invasion. The invasion of single cells and collective cells is often correlated with dramatic changes in the expression and function of adhesive and regulatory proteins. These changes are reminiscent of early developmental processes when cells acquire a migratory, mesenchymal phenotype. Currently, the role of invadopodia in collective cancer invasion is completely unknown. In the present study, we reveal that insuline-like growth factor-II mRNA-binding protein-3 (IMP-3) associated with invadopodial infiltration and collective cancer invasion through podoplanin (PDPN) mRNA stabilization as a component of invadopodia, suggesting that invadopodia plays a pivotal role in invasive front of collective cancer invasion and thereby involved the cancer metastasis. This finding, combined with other investigations into the mechanisms of invadopodia formation, reveal clinically-relevant targets for intervention in invadopodia, including IMP-3 and PDPN.

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 4326. doi:1538-7445.AM2012-4326
Abstract 4336: Macrophage migration inhibitory factor (MIF) plays roles in proliferation, survival and migration of Ewing tumor cells by activation of a CD74-CD44 receptor complex
Authors: Kang Hyung-Gyoo , Triche Timothy J.
Published: August 2012

The deregulated expression of cytokines and the aberrant activation of their downstream signaling pathways would give tumor cells the abnormal proliferation and survival. Therefore identification and understanding of deregulated cytokines in certain tumor would provide an opportunity of better targeted therapies of the cancer. MIF, a proinflammatory cytokine has been known to play a role in proliferation, survival and apoptosis in several tumors. In our studies it was found that MIF expression in Ewing tumors is pretty high and it is secreted out of the cells. Ewing tumor cell lines deficient of MIF showed morphological change, deregulation of differentiation, decrease of proliferation and increase of apoptosis. Through the investigation of several signaling pathways it was found that JAK2/STATs pathways are involved in the MIF dependent survival and proliferation of Ewing tumors and in addition, MIF is involved in the autophagy of Ewing tumors through the activation of AMPK. It has been reported that CD74 is the main receptor of MIF and in our study it was found that CD74 is expressed in Ewing tumors even though the expression is much less than MIF. The decreased expression of CD74 in Ewing tumor cell lines caused the morphological change and inhibition of proliferation, which are the similar phenotypes of MIF deficient Ewing tumor cell lines. It is known that CD74 is associated with CD44 which is involved in the signaling pathways of cellular survival and proliferation. In our study it was found that decreased expression of CD44 in Ewing tumor cell lines caused the inhibition of cell survival and proliferation. In conclusion, our data implies that MIF and its downstream signaling pathways such as JAK/STATs, AMPK, CD74 and CD44 might be involved in growth, survival, proliferation and metastasis of Ewing tumors. Therefore MIF and its downstream signaling pathways could be promising targets of Ewing tumors 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 4336. doi:1538-7445.AM2012-4336
Abstract 4341: A novel sonodynamic therapy using hydroxyl radical generated from ultrasound activated TiO2 for human epithelial carcinoma cells
Authors: Itano Osamu , Onishi Tatsuya , Matsuda Sachiko , Fujimura Tomonori , Jinno Hiromitsu , Kitagou Minoru , Shinoda Masahiro , Kawachi Shigeyuki , Tanabe Minoru , Ogino Chiaki , Shimizu Nobuaki , Katagawa Yuko
Published: August 2012

 Radical oxygen species, which are produced by the action of some chemotherapy drugs, radiation and some types of anticancer agent, are known to mediate cytotoxic activity by modifying the cellular homeostatic redox balance. Photodynamic therapy uses photosensitizers activated by light to induce cell death via formation of free radicals and is actually used in clinical setting. However the limited tissue penetrability of light has been a crucial problem because most of cancers often develop in deep tissue. The generation of hydroxyl radical was investigated during ultrasonic (US) irradiation and in the presence of titanium dioxide (TiO2) (Shimizu, et al. Ultrason. Sonochem. 2008). In this study, the effect of US combined with TiO2 particles on human epithelial carcinoma cell (A431) was investigated in vitro and in vivo.  TiO2 nanoparticles used were uniform in size of about 200 nm in diameter. A431 (1.0x106) were irradiated with US in the various density of TiO2 particles (0 to 10mg/ml). Cell viability was measured immediately after US irradiation (1 MHz, 2.0 W/cm2 for 30, 60, 120 and 300 sec) by using trypan blue exclusion assay. The effect of the combination of TiO2 particle injection (0.1, 1.0 and 10mg, day5) and US exposure (1 MHz, 2.0 W/cm2, 5 min duration, day 5, 7, 9, 11, 13) on subcutaneously implanted A431 solid tumors in female BALB c nu/nu mice were investigated by measuring tumor volume.  The cell viability was significantly decreased after US irradiation in the presence of high dose of TiO2. In vivo results also showed significant inhibition of tumor growth in groups treated with high dose of TiO2 particles and US. The efficacies of US could not be demonstrated combined with low dose of TiO2 particles. Our results demonstrate the potent toxicity of TiO2 combined with US irradiation against tumor in vitro and also the growth inhibition of tumors in a mouse xenograft model. Moreover the cytotoxic efficacy of TiO2 was dependent on the amount of TiO2 particles attaching to cancer cells. These results suggest that the effective and low toxic method for accumulation of TiO2 in the tumor (injection, arterial injection, DDS, etc.) should be necessary for clinical use.

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 4341. doi:1538-7445.AM2012-4341
Abstract 4342: Inhibition of radiation-enhanced cell adhesion in breast cancer cells by ROS scanvenger
Authors: Cheng Huiwen , Lee Shinhee , Wu Shiyong
Published: August 2012

Ionizing radiation (IR) therapy is one of the main therapies to treat breast cancer, but accumulating evidence suggests that IR is potentially associated with increased invasion and metastasis. A great increase in breast cancer cell adhesion to fibronectin after high dose α-radiation was observed in our group, and it accompanied with ROS boost. However it is not known whether ROS increase related to the cell adhesion increase after α-radiation. In this report, we showed that the increased cell adhesion to fibronectin was reduced by ROS scavenger L-NAC by 21.8% after α-radiation, but was not affected by H2O2 scavenger, catalase or by cNOS inhibitor, L-NAME with 24 h treatment. We then determined that the cell adhesion inhibition after α-radiation involve reduced ROS, and reduced vinculin expression and integrin activation after treating with L-NAC for 24 h. However, a short-term, 1 h pre-IR followed by 2 h post-IR, treatment of L-NAC promotes ROS production and has no effect on cell adhesion at 24 h post-IR. These results suggest that L-NAC long-term treatment could potentially have an impact on breast cancer metastasis by inhibiting the ROS and cell adhesion. These results suggest that to inhibit IR-induced breast cancer adhesion, an uninterrupted treatment of L-NAC may be required.

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 4342. doi:1538-7445.AM2012-4342
Abstract 4346: Influence of ionizing radiation on transcription of human endogenous retrovirus-K (HERV-K) in breast cancer cell lines
Authors: Agoni Lorenzo , Lenz Jack , Guha Chandan
Published: August 2012

Ionizing Radiation (IR) has been reported to increase the transcriptional activity of many viruses, including retroviruses. We hypothesized that IR could reactivate subsets of Human Endogenous Retroviruses (HERVs), which are retroviruses integrated into the human genome, remnants of ancient retroviral infections, and that HERV-derived peptides, the final product of HERVs transcription, will increase the diversity of the antigen repertoire presented by irradiated cancer cells, thereby inducing anti-tumor immune responses. IR has been shown to increase the immunogenicity of cancer cells by modulating various cellular processes, including antigen presentation by MHC. HERVs are known to be transcribed in numerous diseases including cancer, HIV infection, and autoimmune disorders. HERV-K is the most recently integrated and thus intact HERV family, with approximately 350 loci in the human genome, 98 to 99% identical. Previous experiments in our laboratory have shown IR-dependent increases in HERV-K transcription, plus increased HERV-K-derived peptides in irradiated prostate carcinoma cell lines upon proteomic LC-MS/MS analysis of peptides eluted from MHC-I complexes. HERV-K activation has been reported in breast cancer. To test the influence of IR on HERV-K transcription, three breast cancer cell lines (MDA-MB-468, MDA-MB-231 and MCF7) were analyzed by RT-PCR and qRT-PCR at specific time-points, from 1 to 72 hours, after different doses of α-irradiation (0, 2.5, 5, 10 and 20 Gy). HERV-K transcription was detected in all cell lines studied. After IR, each cell line showed distinct dose-response curves of HERV-K transcription during the time-frame of observation. Overall, the peak of HERV-K transcription in irradiated samples, when compared to non-irradiated samples, was increased by an average of 2- to 5-fold. Multiple, active, individual HERV-K loci were detected. Cloning and sequencing are ongoing to identify specific active loci for each cell line. These findings show that HERV-K is actively transcribed in breast carcinoma cell lines. The IR-dependent increase in HERV-K transcription may enhance antigen repertoire diversity, contribute to the immunomodulatory properties of IR, and offers attractive targets for the development of novel IR-enhanced immunotherapies for 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 4346. doi:1538-7445.AM2012-4346
Abstract 4353: Tristetraprolin (TTP) mediates radiation-induced TNF-α production in lung macrophages
Authors: Ray Dipankar , Helman Abigail , Shukla Shirish , Ramanand Susmita Gurjar , Tran Linda , Bassetti Michael , Nyati Mukesh K. , Zhang Ming , Lawrence Theodore S.
Published: August 2012

The efficacy of radiation therapy for lung cancer is limited by radiation-induced lung toxicity (RILT). Although tumor necrosis factor-alpha (TNF-α) signaling plays a critical role in RILT, the molecular regulators of radiation-induced TNF-α production remain unknown. We investigated the role of a major TNF-α regulator, Tristetraprolin (TTP), in radiation-induced TNF-α production by lung macrophages. Tristetraprolin (TTP), an adenosine-uridine (AU) rich element (ARE) associated RNA binding protein, is known to destabilize TNF-α mRNA during lipopolysaccharide-induced inflammatory response and now we have identified that radiation causes TTP inactivation via inhibitory (Ser178) phosphorylation and induction of proteasome-mediated degradation, leading to increased TNF-α mRNA stability and secretion. Our preliminary studies indicate possible involvements of PI3K and p38 signaling in radiation-induced inactivating phosphorylation of TTP and we further identify an ubiquitin ligase [Skp-Cullin-F-box (SCFα-TrCP)] responsible for controlling radiation-induced TTP protein stability. These findings suggest that agents capable of blocking TTP phosphorylation or stabilizing TTP after irradiation could decrease RILT.

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 4353. doi:1538-7445.AM2012-4353
Abstract 4355: Use of MnSOD tet on transgenic mice as a model for evaluating irradiation protection and mitigation
Authors: Epperly Michael W. , Chaillet J. Richard , Cao Shaonan , Zhang Xichen , Greenberger Joel S.
Published: August 2012

Manganese superoxide dismutase (MnSOD) is a mitochondrial enzyme responsible for the dismutation or reduction of superoxide produced by cellular respiration. Absence of MnSOD is lethal two to three days after birth. Following irradiation there is an increase in superoxide production which combined with nitric oxide can result in the formation of the highly reactive radical peroxynitrite. Increased expression of MnSOD in a cell increases survival by the reduction of superoxide produced following irradiation. Decreased MnSOD production has resulted in decreased survival following irradiation. To better understand the role of MnSOD in cellular respiration as well as protecting the cell from irradiation damage, we have developed a MnSOD tet on mouse which has no MnSOD activity. In the absence of doxycycline the homozygous MnSOD tet on mouse does not survive past 2 or 3 days after birth. If doxycycline (2 µg/ml) is added to the water of heterozygous mothers, the MnSOD tet on mice develop normally and are fertile as long as they remain on the doxycycline treated water. Long term bone marrow cultures (LTBMCs) were established from homozygous MnSOD tet on mice, heterozygous tet on mice and control C57BL/6J mice. The cultures from the control mice had more hematopoietic cell activity than the MnSOD tet on cultures. To determine when the MnSOD gene is turned on after addition of doxocycline, bone marrow stromal cell lines were developed from the LTBMCs were incubated in 1 µg doxocycline/ml media for time points up to 24 hr, RNA was extracted and RT-PCR for the MnSOD transgene was performed. By 12 hours there was an increase in MnSOD RNA expression in the MnSOD tet on cells. Removal of the doxycycline resulted in a decrease in MnSOD RNA by 1 hr in the MnSOD tet on cells as well as the heteroxyzous cells. This mouse model will be useful for testing new compounds for radiation protection and mitigation.

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 4355. doi:1538-7445.AM2012-4355