bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 4193: TNF-α dynamically modulates genome-wide cross-regulation of cRel, ΔNp63 and TAp73 promoter binding and gene expression in head and neck cancer: TP53 and NFκB ChIP-Seq in HNSCC
Authors: Si Han , Lu Hai , Yang Xinping , Jang Minyoung , Burnett Jeffery , Davis Sean , Sun Hong-wei , Xiao Wenming , Wei Lai , Meltzer Paul , Waes Carter Van , Chen Zhong
Published: August 2012

We previously showed that cRel, ΔNp63, and TAp73, members of NF-kappaB and TP53 transcription factor (TF) families, together regulate broad biological processes critical for inflammation and cancer survival. However, the underlying molecular mechanisms explaining the global cross-regulation of transcriptome and interactive signal network have not been revealed. Here, we demonstrate that inflammatory cytokine TNF-α induces a dynamic, genome-wide increase in cRel and ΔNp63 chromatin occupancy after replacing TAp73 binding activity, using Illumina Next Generation ChIP-sequencing and fitting the data into a dynamic Poisson distribution model integrated in MACS. Globally, significant binding peaks (p<10-5) were highly enriched in promoter regions (∼10%). These binding peaks further showed significant enrichment around transcription starting sites (±2000bp), and co-localized peaks for any of the two TFs were prevalent within a distance of ∼200 bp. De novo motif search using Gibbs Sampler indicated that cRel and ΔNp63 bound to p53 motifs while TAp73 binding switched from p53 to AP1 motifs when treated with TNF-α. Motifs from these TFs also co-localized within ∼200 bp. Binding activities of individual transcription factors were validated by ChIP-PCR and oliogo-ELISA based binding assays. In addition, TNF-α modulated 1050 differentially expressed genes detected by microarray, of which one-third exhibited binding activity for at least one TF, and 73 genes were identified with co-binding activities for the three TFs simultaneously. Ingenuity Systems Pathway Analysis (IPA) and Gene Ontology revealed that these genes are enriched in cancer-related transcription factors and signaling pathways. Our data suggest a novel mechanism of genome-wide replacement of TAp73 by cRel and ΔNp63, supporting cross-regulation and interactive network between NF-kappaB and TP53 families.

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 4193. doi:1538-7445.AM2012-4193
Abstract 4198: Transcriptional regulation of breast cancer microRNAs by GATA3 and TFAP2A
Authors: Huang Simo , Yee Kathy , Bockhorn Jessica , Kim Sun , Huo Dezheng , Olopade Olufunmilayo , Liu Huiping , Greene Geoffrey
Published: August 2012

Metastasis remains a major obstacle in the successful treatment of breast cancer because it is responsible for an estimated ninety percent of breast cancer related deaths. An emerging field in the understanding of breast cancer metastasis is the study of microRNAs (miRNAs) which are short nucleotide sequences shown to be involved in the regulation of cell self-renewal, epithelial-to-mesenchymal transition (EMT) and cancer progression [1]. Our previous data suggested that miR-200 and other miRNAs regulate EMT and breast cancer metastasis [2]. Given the significant roles of miRNAs and their modulated levels in cancer, it is important to understand their regulatory mechanisms. We hypothesize that miRNAs are transcriptionally regulated in breast cancer. Our preliminary data with clinical breast cancer specimens suggests correlative relationships between transcription factors TFAP2A and GATA3 with mir-200a and other miRNAs. The goal of this project is to determine whether TFAP2A and GATA3 regulate these miRNA levels in breast cancer cells. Using quantitative real-time PCR, the miRNA levels were measured in MDA-MB 231 breast cancer cells with modulated expression of GATA3 or TFAP2A. The results show that overexpression of GATA3 in MDA-MB 231 cancer cells significantly increased the expression of miRNAs aberrantly downregulated in ER- breast tumors, such as miR-138. Chromatin-immunoprecipitation and reporter luciferase assays will be used to determine whether the regulation of miRNAs by GATA3 and TFAP2A is a direct transcriptional regulation. 1. Mani, S.A. et al., 2008. The epithelial-mesenchymal transition generates cells with properties of stem cells. Cell 133, 70415. 2. Park, S.M. et al., 2008. The miR-200 family determines the epithelial phenotype of cancer cells by targeting the E-cadherin repressors ZEB1 and ZEB2. Genes Dev 22, 894-907.

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 4198. doi:1538-7445.AM2012-4198
Abstract 4200: Translation initiation factor eIF3b expression and its role in cancer cell growth and metastases
Authors: Wang Hong , Wang Xuejiao , Ru Yuanbin , Conaway Mark R. , Kieft Jeffery S. , Theodorescu Dan
Published: August 2012

The initiation of translation in eukaryotes is promoted by multi-protein eukaryotic initiation factor complexes (eIFs). While several eIF subunits are overexpressed in cancer, very few have been conclusively implicated as drivers of human cancer phenotypes. Through a transcriptomic analysis of human bladder cancer our studies suggested that eIF3b, considered to be the major scaffolding subunit within the eIF3 complex, is elevated compared to normal urothelium. We went on to functionally evaluate eIF3b in human bladder cancer by siRNA depletion in UMUC3 and lul2 human bladder cancer lines. This reveled that expression of eIF3b was important for anchorage dependent and independent cell growth and G1 to S cell cycle transition. Depletion of eIF3b also inhibited cell migration in vitro and this was associated with disruption of actin cytoskeleton organization and focal adhesion formation through integrin [[Unsupported Character - Symbol Font ]]5, which in turn decreased FAK phosphorylation and downstream Akt phosphorylation. We also observed the similar results from PC3 prostate cancer cells. In vivo, UMUC3 and lul2 cells depleted of eIF3b formed smaller tumors that grew more slowly following subcutaneous inoculation and these cells formed fewer lung metastases following intravenous inoculation. This data provides the first demonstration of a causal role of eIF3b in human cancer cell phenotypes 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 4200. doi:1538-7445.AM2012-4200
Abstract 4214: Identifying cell surface receptor VAP targets to sensitize melanoma to chemotherapy
Authors: Sun Xiaowen , DeMali Kris A.
Published: August 2012

Melanoma accounts for 79% of all skin cancer deaths. The incidence rate has doubled throughout the past three decades, while the survival rate remains consistently low. Melanoma is unusually lethal as it metastasizes early and is highly resistance to all available chemotherapeutics. The newest treatment utilizes a small inhibitor targeting the BRAF pathway, considering the BRAF V600E mutation occurs in 65% of clinical melanoma cases. Unfortunately, it is clear that there is both primary and acquired resistance to the drug. Recent studies demonstrate that engaging defective cell surface adhesion receptors known as integrins increases the sensitivity of melanoma to traditional DNA damaging agents. Our laboratory has identified a novel peptide denoted vinculin activating peptide (VAP) that increases integrin engagement from within the cell by targeting the integrin cytoplasmic binding partner, vinculin. Furthermore, VAP sensitizes melanoma to DNA damaging agents in vitro and in vivo. The goal of this project is to identify the cell surface target of VAP. Both α1and β3 integrins are highly over-expressed in melanoma, suggesting that VAP targets one or both of these defective integrins. To test this possibility, melanoma cells lacking β1 or β3 integrins were generated. In the absence of β1 integrins, VAP did not induce elevations in p53 levels or increase apoptosis in melanoma cell lines treated with cytotoxic agents. In addition, its ability to sensitize melanoma cells to therapeutics in biological assays was ablated. Furthermore, the β1 integrin-dependency of VAP's effect was confirmed by examining melanoma cells lacking β3 integrins. Ongoing experiments are testing the requirement for β1 integrin in mouse models of melanoma. Taken together, these findings suggest that β1 integrins are required for VAP to sensitize melanomas to chemotherapy agents. The successful completion of these studies will provide the mechanistic underpinning for a new class of therapeutics that sensitize melanomas to chemotherapy.

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 4214. doi:1538-7445.AM2012-4214
Abstract 4373: Targeting FGFR system in a model of HCC
Authors: Lang Sven A. , Scheller Tobias , Moser Christian , Mycielska Maria , Hellerbrand Claus , Schlitt Hans J. , Geissler Edward K.
Published: August 2012

Activation of the fibroblast growth factor receptor (FGF/FGFR) system has been reported in a large number of human cancers including human hepatocellular carcinoma (HCC). Furthermore, expression of FGFR2 has been associated with aggressive tumor growth and poor prognosis in this tumor entity. Moreover, HCC is known to be a hypervascularized tumor and activation of the FGF/FGFR system is a crucial event in tumor angiogenesis. Hence, we addressed the issue of targeting the FGFR system in HCC (Huh-7, HepG2) and stromal cells (endothelial cells (EC), vascular smooth muscle cells (VSMC), hepatic stellate cells (HSC)). The FGFR inhibitor BGJ398 (Novartis Oncology, Basel, Switzerland) was used to inhibit FGFR activation. Effects of targeting FGFR on growth of cells were determined by MTT assays. Inhibition of constitutive and growth factor induced cell motility was investigated in modified Boyden Chamber assays. Activation of signaling pathways upon treatment with BGJ398 was assessed by Western Blot analyses whereas effects on mediators of tumor angiogenesis (VEGF-A, PDGF-B) where measured by RT-PCR and ELISA, respectively. Results show that inhibition of the FGF/FGFR system with BGJ398 led to dose-dependent inhibition of growth in tumor cells and ECs whereas only minor effects were observed on VSMC and HSCs. Moreover, motility of tumor and stromal cells (ECs, VSMCs, HSCs) was significantly impaired upon constitutive conditions (P<0.05 for all). In addition, FGFR inhibition led to decrease in HSC motility even when cell were incubated with conditioned media from cancer cells (P<0.05). With regards to activation of signaling intermediates, FGFR inhibition impaired constitutive and growth factor induced phosphorylation of Akt and ERK in HCC cell lines, ECs and HSCs whereas only minor effect was observed in VSMCs. Moreover, treatment with BGJ398 led to inhibition of c-Myc expression and significantly reduced VEGF-A production from cancer cells, as determined by Western blotting and ELISA, respectively. Evaluation of effectiveness of BGJ398 in animal models is currently ongoing. However, results so far indicate that targeting FGFR impairs angiogenic signaling and motility in HCC cell lines and stromal cells. Therefore, inhibition of FGFR warrants further evaluation and might be an interesting novel approach for anti-neoplastic treatment protocols in human HCC.

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 4373. doi:1538-7445.AM2012-4373
Abstract 4379: Activating the prostaglandin I2-IP signaling suppresses metastasis in lung cancer
Authors: Minami Yoshinori , Endo Satoshi , Okumura Shunsuke , Shibukawa Kiyoko , Sasaki Takaaki , Ohsaki Yoshinobu
Published: August 2012

Background: The mechanism by which bone marrow (BM)-derived cells might contribute to tumor metastasis and angiogenesis is controversial. We have reported that bone marrow derived angiogenic progenitor cells (Lin- cKit+ Flk-1+ cells) express prostaglandin I2 (PGI2) specific receptor; IP, and the PGI2-IP system is necessary for vascular remodeling and angiogenesis. Additionally, we have reported that the specific knockdown of IP in bone marrow derived cells (BMDCs) increases tumor metastasis in mouse models. Objectives: The purpose of this study was to test the hypothesis that activating the PGI2-IP signaling suppresses tumor metastasis. Materials & Methods: We employed both a mouse lung metastatic model and a mouse bone marrow transplanted model. Mouse-derived Lewis lung carcinoma (LLC) cells labeled with DsRed were used for a mouse lung metastatic model to distinguish cancer cells from BMDCs. The LLCs were injected into the tail vein of mice (c57BL/6J), which bone marrow cells were transplanted from GFP-labeled mice. Beraprost sodium (BPS; IP receptor agonist) or PBS control was continuously administered for 3 weeks by subcutaneous osmotic pumps. Tumor metastasis to lung was assessed by using hematoxylin-eosin staining. The a-SMA as a pericyte marker and the CD31 as an endothelial cell marker were studied by immunofluorescence to evaluate angiogenesis in metastatic lung tumors. Results: The size and number of tumor metastasis were significantly decreased in BPS group compared with in PBS control group assessed by the mice lung metastasis model. Immunofluorescence analysis revealed that the pericytes derived from bone marrow were scattered within tumors in PBS control group, while those were observed along with tumor micro vessels by supporting the endothelial cells within tumors in BPS group. Conclusions: The present study demonstrated that activating the PGI2-IP signaling suppresses metastasis in lung cancer. These results also suggested that the maturation of tumor angiogenesis by pericytes may decrease tumor metastasis. Finally, we propose that PGI2-IP system would be a novel therapeutic target in lung metastasis.

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 4379. doi:1538-7445.AM2012-4379
Abstract 4388: Combination therapy with anti-CTLA-4 plus leuprolide acetate in the pre-surgical setting of patients with regional, high-risk prostate cancer
Authors: Gao Jianjun , Sun Jingjing , Ward John F. , Rao Priya , Troncoso Patricia , Araujo John C. , Tang Derek N. , Chen Hong , Thall Peter , Wen Sijin , Sharma Padmanee
Published: August 2012

Anti-CTLA-4 is a monoclonal antibody (Ipilimumab or Yervoy, Bristol-Myers Squibb) that was recently FDA-approved for the treatment of patients with metastatic melanoma. Since anti-CTLA-4 targets a T cell specific molecule, as opposed to a tumor-specific molecule, it is being tested for therapeutic efficacy in multiple malignancies, including prostate cancer. In addition, since anti-CTLA-4 enhances anti-tumor T cell responses by blocking an inhibitory signal on activated T cells, it has been postulated that traditional therapies that lead to tumor cell death, thereby releasing tumor antigens to prime and potentially activate T cell responses, can be used in combination with CTLA-4 blockade to sustain activated T cell responses and enhance anti-tumor immune responses. Pre-clinical mouse models have shown this to be true. To determine the impact of combination therapy on human immune responses, we designed a pre-surgical clinical trial with a hormonal agent consisting of leuprolide acetate, a standard agent that has therapeutic efficacy against prostate cancer, plus anti-CTLA-4 (ipilimumab), in the setting of patients with regional, high-risk prostate cancer. As of October 2011, we have treated 4 out of a planned 20 patients on trial. Patients receive one injection of leuprolide acetate (Lupron, Tap Pharmaceuticals) at a dose of 22.5 mg on week 0 and ipilimumab at 10 mg/kg/dose on weeks 1 and 4. Patients then undergo surgery at week 8. Tumor tissues are collected prior to any therapy and then at the time of surgery. Blood is collected throughout the study prior to each therapeutic intervention and after surgery. Immunologic analyses are ongoing on all collected patients’ samples. These data will be compared to those obtained from previously completed trials consisting of a trial whereby patients were treated with ipilimumab monotherapy and a trial whereby patients were treated with hormonal therapy prior to surgery. Our preliminary data indicate that combination therapy impacts T cell responses including a measurable increase in the frequency of T cells expressing the inducible costimulator (ICOS) molecule. In addition, four patients have completed the trial, including surgery, without any serious adverse events related to the combination 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 4388. doi:1538-7445.AM2012-4388
Abstract 4410: Neutralizing Endosialin antibody, MORAb-004, enhances efficacy of TAXOTERE in a metastatic melanoma model
Authors: Krauthauser Candice , Grasso Luigi , Nicolaides Nicholas , Lin JianMin
Published: August 2012

Endosialin/Tumor Endothelial Marker-1 (TEM1) is a cell surface glycoprotein that is expressed on cells involved in the development of tumor vasculature, primarily pericytes and stromal fibroblasts. Studies have found it to play a key role in tumor growth and neo-vessel formation in numerous cancer types. We are currently evaluating the safety and therapeutic potential of MORAb-004, a humanized monoclonal IgG targeting endosialin/TEM-1, in clinical trials. Because MORAb-004 only recognizes human antigen and has no cross-reactivity with rodent endosialin/TEM-1, we generated a human endosialin/TEM-1 knock-in mouse in order to assess its activity in vivo. Previously, in this transgenic model, we found that MORAb-004 significantly inhibited subcutaneous B16-F10 melanoma progression by ∼70% when administered intravenously for 5 consecutive days. Moreover, using the Lewis Lung Carcinoma model of metastatic disease our results showed that MORAb-004 had a significant inhibitory effect on the number of lung metastases. For these studies, supporting clinical trials for using MORAb-004 in combination with standard of care chemotherapeutics, we developed a B16-F10 melanoma lung colonization model. In this model, the human endosialin/TEM-1 knock-in mice were implanted intravenously with B16F10 cells adapted for in vivo lung colonization. These mice were treated with Docetaxel, MORAb-004, or the combination of the two agents, respectively. A group of the tumor-bearing mice receiving PBS served as control. At the end of the study, the lungs of each mouse were harvested. A ratio of the area of tumor nodules versus total lung surface area is calculated as tumor burden. MORAb-004, at 50 mg/kg three times weekly for a total of 7 i.v. injections reduced tumor burden by 45%. Treatment of Docetaxel, at 3mg/kg twice weekly for a total of 5 i.p. injections, reduced less than 20% of tumor burden in the lungs. When MORAb004 was added to Docetaxel treatment, a significant enhancement of efficacy was observed. The reduction of tumor nodules in the lungs was increased to approximately 70%. A similar trend is seen in combination therapy where MORAb-004 was added to gemcitabine regimen. Our results demonstrate the ability of MORAb-004 to inhibit tumor growth in a murine model. Moreover, when added to the chemotherapeutic regimen, it significantly enhances the therapeutic effect of traditional chemotherapy.

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 4410. doi:1538-7445.AM2012-4410
Abstract 4489: Clinical specimen-based investigatory study of biomarkers for gastric cancer
Authors: Oshima Takashi , Kunisaki Chikara , Hasegawa Shinichi , Ono Hidetaka , Fujii Shinichi , Sato Tsutomu , Yamada Roppei , Yukawa Norio , Rino Yasushi , Masuda Munetaka , Imada Toshio
Published: August 2012

Objectives: We have identified potential biomarkers for gastric cancer using a cDNA bank on the basis of mRNA extracted from frozen specimens of gastric cancer tissue and adjacent normal gastric mucosa. We report currently available results. Methods: The study group comprised 227 patients in whom 5 or more years had elapsed since gastrectomy. The disease stage was IB in 29 patients, II in 66, III in 103, and IV in 29. Among the 139 patients who postoperatively received fluoropyrimidines, 97 with stage II or III disease were given adjuvant chemotherapy with S-1. A total 104 genes were selected on the basis of the results of comprehensive DNA microarray analysis, extraction from serial analysis of gene expression (SAGE) libraries, and preclinical and other studies of potential targets for molecular-targeted therapy. The relative expression levels of these genes in gastric cancer tissue and adjacent normal mucosa from each patient were measured by a quantitative polymerase chain reaction assay using the cDNA bank described above, and the relations between clinicopathological factors and outcomes were examined. Results: In patients who underwent gastrectomy, high expression of matrix metalloproteinase-9 (MMP9), insulin-like growth factor receptor 1 (IGF-R1), and apolipoprotein E (APOE) genes were associated with significantly worse outcomes. In patients with stage II or III disease who received adjuvant chemotherapy with S-1, high expression of IGF-R1, thymidylate synthase (TS), and regenerating IV (Reg IV) genes were associated with significantly worse outcomes. Conclusions: MMP9, IGFR-1, and APOE gene expression levels were suggested to be useful prognostic factors in patients who underwent gastrectomy for gastric cancer. In patients with stage II or III disease who received adjuvant chemotherapy with S-1, IGF-1R, TS, and Reg IV gene expression levels were suggested to be useful prognostic factors.

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 4489. doi:1538-7445.AM2012-4489
Abstract 4621: EMP2 regulates the tumor microenvironment and is a novel therapeutic target for triple negative breast cancer
Authors: Goodglick Lee , Fu Maoyong , Braun Jonathan , Wadehra Madhuri
Published: August 2012

Breast cancer remains the most common malignancy in women, with the aggressive triple negative subtype accounting for nearly half of all breast cancer deaths. In this abstract, we describe a novel tetraspan protein, epithelial membrane protein-2 (EMP2) that is highly expressed in the majority of breast cancer tumors examined compared to healthy mammary epithelium. In particular, high levels of EMP2 were observed in over 70% of triple negative breast cancer cases examined. In tumor cells, EMP2 altered the integrin expression profile as well as promoted endothelial cell migration and tube formation through upregulation of VEGF. We produced a fully human intact IgG1 to test their effect on EMP2 expressing breast cancer cell lines. In vitro treatment with anti-EMP2 IgG1 decreased Src and FAK activation and resulted in cell death. Moreover, cultured supernatant from treated cells reduced endothelial cell migration and capillary-like tube formation. To determine the in vivo efficacy of EMP2 treatment, we first characterized potential toxicity from systemic administration. Anti-EMP2 IgG1 produced no observable toxicity (weight, CBC counts, tissue pathology) when administered at 10mg/kg over 7 weeks. Using similar conditions in preclinical xenograft and syngeneic mouse models, anti-EMP2 IgG1 decreased tumor growth with induction of necrosis. This effect was specific as no difference in tumor size was observed in EMP2 negative (Ramos) cell tumors. As in vitro and in vivo cytotoxicity was observed in both triple negative cell lines as well as cells expressing estrogen and progesterone receptors and/or Her-2/neu, these results present compelling evidence that EMP2 is a novel therapeutic target in triple negative breast cancers as well as other variants.

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 4621. doi:1538-7445.AM2012-4621
Abstract 4662: Shikonin causes cancer cell death by inducing mitochondrial dysfunction
Authors: Wiench Benjamin , Eichhorn Tolga , Efferth Thomas
Published: August 2012

Shikonin, a naturally occurring napthoquinone, has been used in herbal formulations for the treatment of several inflammatory diseases in Traditional Chinese Medicine since decades. In recent studies, shikonin revealed remarkable anticancer activities and thereby is a promising candidate for cancer chemotherapy. However, the underlying cellular mechanisms and targets of shikonin are still unknown. Here, we showed that shikonin indeed exhibits strong cytotoxic effects on a panel of 15 different cancer cell lines also containing multi-drug resistant cells. The strongest effects were obtained on U937 leukemia cells. To better understand the underlying mechanisms, we performed a whole genome expression analysis of these cells after shikonin treatment. Thereby novel molecular key players and genetic networks responsible for the effects of shikonin were identified. Induction of cell cycle arrest, apoptosis and increased levels of reactive oxygen species and DNA damage were revealed as striking cellular effects of shikonin. Furthermore, whole sets of genes responsible for cytoskeletal formation and mitochondrial energy metabolism were deregulated after shikonin treatment. Using cell based assays we showed that shikonin indeed induces G1/S phase cell cycle arrest and caspase 3/7 dependent apoptosis. Furthermore we confirmed the induction of reactive oxygen species and quantified the subsequent DNA damage. In addition shikonin significantly prevents cancer cell migration and cytoskeleton formation by inhibiting the dynamics of the microtubule system. The reason for all this secondary effects was revealed by real-time kinetic uptake experiments based on the intrinsic fluorescence of shikonin. Thereby a rapid cellular uptake and a direct accumulation of the drug in the mitochondria were observed. This caused a breakdown of the mitochondrial membrane potential within the same time range of drug uptake, resulting in a failure of the respiratory chain and the cellular energy production. As the mitochondrial structure and metabolism differs strongly between cancer and normal cells, targeting of mitochondria is a very recent and promising approach in cancer therapy. Our results showed that mitochondria are the primary targets of shikonin in cancer cells and thus we believe that shikonin has the potential in future clinical application.

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 4662. doi:1538-7445.AM2012-4662
Abstract 492: Identification of a novel complex MUC1/CIN85 involved in invasion and migration of breast cancer cells
Authors: Cascio Sandra , Finn Olivera
Published: August 2012

MUC1 is a transmembrane glycoprotein abnormally expressed in human adenocarcinoma. The extracellular domain of MUC1 contains a variable number of tandem repeats 20aa in length (VNTR region) which is fully glycosylated in normal epithelia and underglycosylated in tumor cells. We recently demonstrated that tumor form of MUC1 increases the accessibility of the VNTR peptide backbone to other proteins thus affecting protein-protein interactions and intracellular signaling. By using MALDI-TOF, we identified CIN85 as one of the proteins that bind the extracellular domain of MUC1. CIN85 is an adaptor protein involved in multiple cellular processes including signal transduction, vesicle-mediated transport, cytoskeleton remodelling, invasion and migration of cancer cells. Structurally, CIN85 contains three SH3 domains, a proline-rich region and a C-terminal coiled-coil region. The SH3 domain of CIN85 recognizes a specific sequence PXXAP that corresponds to the PDTRP sequence in the 20 amino acid tandem repeat in the MUC1 VNTR. Co-immunoprecipitation assay and confocal immunofluorescence confirmed that MUC1 and CIN85 co-localize in MUC1-transfected RMA (mouse T cell lymphoma) and IG10 (mouse ovarian carcinoma) cells and in human breast cancer cells MDA-MB-231 and MDA-MB-435 that express endogenous MUC1 and CIN85. Subcellular protein fractionation revealed that CIN85 and MUC1 co-localize mainly at plasma membrane but also in the cytosol. To better understand the function of CIN85/ MUC1 complex we analyzed their expression by immunohistochemistry and confocal immunofluorescence in human tissue microarray (TMA) composed of 31 breast cancer samples and 5 adjacent normal tissue. Hypoglycosylated MUC1 and CIN85 are over-expressed and co-localize in early and advanced clinical stage of breast cancer. Interestingly, CIN85 and MUC1 were localized to invadopodia-like structures suggesting a potential role in cancer cell invasion. siRNA-mediated silencing of CIN85 and/or MUC1 revealed that MUC1 enhances CIN85-dependent cell invasion and migration of breast cancer cells. In contrast, ectopic expression of MUC1 reduces the motility induced by CIN85. In conclusion, our finding suggest that the novel complex CIN85/MUC1 play a critical role in invadopodia formation and have control on invasive and metastatic potential of breast cancer 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 492. doi:1538-7445.AM2012-492
Abstract 4945: Targeting ErbB3-addicted cancers across the HER2 spectrum
Authors: Onsum Matthew , Yarar Defne , Paragas Violette , Zhang Bo , Nguyen Stephanie , Schoeberl Birgit , McDonagh Charlotte , MacBeath Gavin , Nielsen Ulrik
Published: August 2012

ErbB3 is a critical activator of phosphoinositide 3-kinase (PI3K) signaling in EGFR-, HER2-, and MET-addicted cancers, and ligand-driven activation of ErbB3 is a prominent method by which cancers become resistant to targeted inhibitors of ErbB signaling, as well as anti-hormonal and chemotherapeutic agents. With the goals of inhibiting survival signaling in cancer cells, delaying the onset of resistance to standard-of-care therapies, and re-sensitizing cancer cells to prior therapies, we developed an anti-ErbB3 monoclonal antibody, MM-121, that effectively blocks ligand-dependent activation of ErbB3 and induces internalization and degradation of the receptor. The activity of single-agent MM-121 was examined in xenograft models and a machine-learning algorithm was developed that accurately predicts which preclinical models of cancer will respond to MM-121 based on five protein-based biomarkers. By simulating ErbB3 inhibition using a computational model of ErbB signaling, we predicted that a bispecific molecule that docks onto HER2 and subsequently binds to ErbB3 would effectively block ErbB3-driven signaling in HER2-amplified tumors. We therefore developed a bispecific antibody comprising two human scFv antibodies linked by a modified form of human serum albumin. The resulting molecule, MM-111, forms a trimeric complex with HER2 and ErbB3, effectively inhibiting ErbB3 signaling and demonstrating antitumor activity in preclinical models that are dependent on HER2 overexpression. Both MM-121 and MM-111 are now in early stages of clinical development.

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 4945. doi:1538-7445.AM2012-4945
Abstract 4961: In human breast cancer cells thyroid hormone signaling overshadows estrogen signaling on SMP30 gene leading to induction of apoptosis
Authors: Mishra Sandip K. , Sar Pranati , Rath Bandita , Bhargava Dharmendra , Sengupta Debomita , Chaudhary Sanjib
Published: August 2012

Crosstalk between thyroid and estrogen hormones actions have been documented with regard to a variety physiological functions. Both of these hormones regulate transcription of target genes by binding to their nuclear receptors that interact with specific responsive elements (estrogen and thyroid hormone response elements, i.e. ERE and TRE, respectively. Recently, we have demonstrated that 3, 3′5 Triiodo L Thyronine (T3) can induce apoptosis in ER+ve highly metastatic breast cancer MCF-7 cells through downregulating Senescence marker protein-30 (SMP30) gene. SMP30 has also been reported to be downregulated by 17β-Estradiol (E2) in prostrate gland and mammary epithelial cells. Interestingly, thyroid receptors (TRs) and estrogen receptor (ER) share a common response element, so we hypothesised a possible competition between both the receptors in SMP30 promoter under different hormone responses. To prove this hypothesis, gel retardation and luciferase assays were conducted by taking hSMP30 promoter reporter construct validated our findings of the putative ERE site. Competition Chromatin immunoprecipitation assay (ChIP) showed prominent TRβ binding irrespective of thyroid and estrogen hormone treatment. As a result, there was a significant repression found in whole promoter activity in presence of T3 and also in presence of both E2 and T3 treatment. Although the SMP30 promoter activity was same in response to E2 and T3 but the functional consequences after downregulation of SMP30 in human breast cancer cells were different. Overall our studies indicate that estrogen signalling/downstream target molecules of E2 weren't affected by estrogen mediated SMP30 downregulation however, SMP30 downregulation by thyroid hormone synergized the effect of T3 by further inducing apoptosis in human breast cancer cells. Thyroid hormone induced apoptosis in human breast cancer cells was through activated p53 molecule.

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 4961. doi:1538-7445.AM2012-4961
Abstract 5071: The genomic landscape of pancreatic cancer: from discovery to patient care
Authors: Biankin Andrew V. , Grimmond Sean M. ,   Australian Pancreatic Cancer Genome Initiative
Published: August 2012

Pancreatic Cancer is the fourth leading cause of cancer death in our society, with a mortality that virtually parallels its incidence, a median survival of < 12 months even with maximal therapy, and a 5-year survival rate of < 5%. The diversity of clinical outcomes and the molecular heterogeneity of histopathologically similar cancer types, incomplete knowledge of the genomic aberrations that drive carcinogenesis and the lack of therapeutics that specifically target most known genomic aberrations necessitates large scale detailed analysis of cancer genomes to identify novel potential therapeutic strategies. As part of the International Cancer Genome Consortium (ICGC), the Australian Pancreatic Cancer Genome Initiative (APGI), together with the Baylor College of Medicine and the Ontario Institute of Cancer Research used exomic sequencing and copy number analysis to define genomic aberrations that characterise a large clinically focused prospectively accrued cohort of patients with pancreatic cancer (n = 142). The cohort consists of early (clinical stages I and II) non-pretreated patients with pancreatic ductal adenocarcinoma who underwent operative resection with curative intent. We devise approaches to adjust for low epithelial content in primary tumours and define the genomic landscape of pancreatic cancer to identify novel candidate driver genes and mechanisms. We develop stratified, molecular phenotype guided therapeutic strategies using existing therapeutics that are either rescued, repurposed, in development, or are known to be effective in an undefined subgroup of PC patients. These are then tested in primary patient derived xenografts and cell lines from the above deeply characterised cohort. In addition, we are launching a clinical trial named IMPaCT (Individualised Molecular Pancreatic Cancer Therapy). This trial randomises patients with metastatic disease to either standard first line therapy with gemcitabine, or a molecular phenotype guided approach using next generation sequencing strategies to screen for actionable mutations defined through the ICGC effort.

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 5071. doi:1538-7445.AM2012-5071
Abstract 5091: Proteomic study using a newly established metastatic lung cancer model
Authors: Wang Dongsheng , Hu Zhongliang , Koenig Lydia , Nannapaneni Sreenivas , Shin Dong M. , Chen Zhuo (Georgia)
Published: August 2012

To understand metastasis of lung cancer, we established a highly metastatic pulmonary large cell carcinoma cell line (801BL) through 2 rounds of in vivo selection using a nude mouse xenograft model. In the first round, low-metastatic human large cell lung cancer cells (801D cell line) were subcutaneously injected into the flank of the mice. Metastatic tumor cells were collected from mouse brain and re-injected into the mice flank in the second round. A highly metastatic cell line 801BL was established from the metastatic cells in the lung. A PCR-based satellite tandem repeat (STR) analysis confirmed the same genomic background of the newly established metastatic cell line 801BL as the non-metastatic counterpart 801C and low-metastatic counterpart 801D cell lines. A xenografted mouse model showed 100% of the mice (8 out of 8 mice) injected subcutaneously with 801BL developed lung metastatic tumors, while none of the mice injected with 801C cells developed lung metastasis (p<0.0001). The metastatic cells present a different cancer stem cell-like morphological feature compared with the 801C and 801D cell lines. Their invasive capability is also increased by 3.47±0.44 fold compared to the non-metastatic counterpart (801C) in a matrigel invasion assay. A stable isotope labeling by amino acids in cell culture (SILAC)-based quantitative proteomic study was conducted by comparing the isotopic labeled 801BL cells with non-labeled 801C cells. Bioinformatic analysis using Ingenuity software (http://ingenuity.com) revealed significant alterations in several dominant cell signal networks including cell morphology (p-value:2.80E-09 - 6.83E-03), cellular function and maintenance (p-value:1.08E-08 - 6.83E-03), DNA replication, recombination, and repair (p-value:1.08E-08 - 6.83E-03), and cell death (p-value: 1.88E-08 - 6.83E-03), between the highly metastatic cell line (801BL) and its non-metastatic counterpart (801C). Western blot assay of several proteins from each signaling network confirmed the proteomic findings. Since the highly metastatic cell line and its non-metastatic counterpart share the identical genetic background, the model based on this cell line provides a powerful tool for the study of the mechanisms underlying lung cancer metastasis. (This work is supported by grant from DOD W81XWH-07-1-0306).

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 5091. doi:1538-7445.AM2012-5091
Abstract 5128: Identification of osteosarcoma driver genes by integrative analysis of copy number and gene expression data
Authors: Kuijjer Marieke L. , Rydbeck Halfdan , Kresse Stine H. , Buddingh Emilie P. , Roelofs Helene , Bürger Horst , Myklebost Ola , Hogendoorn Pancras CW , Meza-Zepeda Leonardo A. , Cleton-Jansen Anne-Marie
Published: August 2012

High-grade osteosarcoma is a tumor with a complex genomic profile, occurring primarily in adolescents. The extensive genomic alterations obscure the identification of genes driving tumorigenesis in osteosarcoma progenitor cells. In order to identify such driver genes, we integrated DNA copy number profiles (Affymetrix SNP 6.0) of 32 diagnostic biopsies with 84 expression profiles (Illumina Human-6 v2.0) of osteosarcoma as compared with its putative progenitor cells, i.e. mesenchymal stem cells (n=12) or osteoblasts (n=3). In addition, we performed paired analyses between copy number and expression profiles of a subset of 29 patients for which we had both DNA and mRNA profiles. Integrative analyses were performed in Nexus Copy Number Software and statistical language R. Paired analyses were performed on all probes which were significantly differentially expressed in corresponding LIMMA analyses. For both non-paired and paired analyses, copy number aberration frequency was set to >35%. Non-paired integrative analyses resulted in 45 genes that were present in both analyses using different control sets, i.e. MSCs and osteoblasts. 101 genes overlapped between the two paired integrative analyses. Paired analyses detected >90% of all genes found with the corresponding non-paired analyses. Remarkably, approximately twice as many genes as found in the corresponding non-paired analyses were detected. Affected genes were compared with differential expression in osteosarcoma cell lines. An overrepresentation of altered cell division related genes was found, such as MCM4 and LATS2, suggesting that deregulation of the cell cycle is an initial driver of osteosarcomagenesis.

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 5128. doi:1538-7445.AM2012-5128
Abstract 5151: Transcriptional regulation of the pyruvate dehydrogenase kinase Pdk2 by tumor suppressors
Authors: Contractor Tanupriya , Harris Chris
Published: August 2012

Conversion of pyruvate into lactate despite the presence of oxygen is a hallmark of the Warburg effect, and distinguishes the metabolism of many tumors from the metabolism of many normal cell types. Pdk2, a pyruvate dehydrogenase kinase, facilitates conversion of pyruvate into lactate by inhibiting the pyruvate dehydrogenase complex, which would otherwise convert pyruvate into acetyl-CoA. Previously, Pdk2 activity was thought to be controlled exclusively in a post-translational fashion, through inhibition or activation by metabolites. But in this study we show that two important tumor suppressor pathways, p53 and retinoblastoma, prevent the Warburg effect, and do so through repression of pdk2 transcription. Importantly, a heterologous promoter that drives physiological expression of Pdk2, but disallows repression by p53 or Rb, prevents these tumor suppressors from blocking the Warburg effect. Another consequence of pdk2 repression is increased production of reactive oxygen species, which stimulates p53-dependent apoptosis. We further show that when unable to downregulate Pdk2 expression, p53 can no longer cause apoptosis. Thus two critical features of tumor suppression, apoptosis and pyruvate metabolism, are mediated by repression of pdk2 transcription. This study proves that proper transcriptional regulation of the pdk2 gene is critical for tumor suppression.

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 5151. doi:1538-7445.AM2012-5151
Abstract 5153: Steroid receptor coactivator-2 mediates oncogenic reprogramming of cancer cell metabolism
Authors: Dasgupta Subhamoy , Zhang Bin , Louet Jean-Francois , O'Malley Bert W.
Published: August 2012

Steroid receptor coactivators/p160 (SRC) are frequently amplified and overexpressed in various human cancers which mediate transcriptional functions of nuclear receptors and other transcription factors, providing survival advantage to cancer cells. Recent findings from genomic profiling of human prostate tumors have identified SRC-2 (also known as NCOA2/TIF2), one of the SRC family member as the major oncogene. SRC-2 was found to be significantly enhanced in 8% of primary tumors and 37% of metastatic tumors, and its expression positively correlated with disease recurrence. This suggests SRC-2 oncogene may play crucial role in the development and metastatic progression of prostate cancer. The current study is aimed to understand the molecular function of SRC-2 in the pathogenesis of prostate cancer. RNAi mediated knockdown of SRC-2 in cultured prostate cancer cell lines significantly reduced the cell viability and this phenotype was rescued by ectopic overexpression of SRC-2. Since SRC-2 coactivator is a master regulator of key metabolic pathways in normal cellular physiology, we investigated its role in cancer cell metabolism. Depletion of SRC-2 severely impairs accumulation of lipid droplets, suggesting SRC-2 regulates lipogenesis in cancer cells. Lipid biosynthetic pathways are integral part of cancer cell growth generating fatty acids and cholesterol required for the biogenesis of membrane. Lipids derive majority of carbon chain from glucose flux by diverting pyruvate into acetyl coenzyme A, an important precursor for lipid biosynthesis. Initial screening identified SRC-2 regulates expression of key metabolic enzymes 3-hydroxy-3-methylglutaryl-CoA reductase (HMGCR), pyruvate dehydrogenase (PDH), fatty acid synthase (FASN), and stearoyl-CoA desaturase (SCD1) involved in fatty acid and cholesterol biosynthesis. Interestingly we found expression of HMGCR, FASN and PDH is dramatically increased in metastatic C4-2 cells compared to parental LNCaP cells, and their expression can be reversed by SRC-2 depletion. Ongoing studies will further examine the regulation of glucose and pyruvate metabolism by SRC-2 and its importance in cancer cell metastasis. Although it is well known that many tumours exhibit increased glycolysis, less clear is how associated metabolic pathways are affected and whether they contribute to cancer cell proliferation. These analyses will allow us to identify unpredictable nodes of metabolic regulation by SRC-2 oncogene and may provide rational treatment approaches for targeting tumor metabolism.

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 5153. doi:1538-7445.AM2012-5153