bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3957: Cardiac metastasis in mice with primary breast carcinoma.
Authors: Zhang Yong , Zhang Nan , Hoffman Robert M. , Zhao Ming
Published: August 2014

The incidence of cardiac metastases reported in the literature ranges from 2.3% to 18.3%. Cancer cells invade the heart either directly or through the blood stream or lymphatics. This report describes cardiac metastasis from primary breast carcinoma in an orthotopic nude mouse model. 4T1 mammary carcinoma cells expressing FP were orthotopically implanted into the right second mammary fat pad of BALB/c mice. When the average tumor volume reached approximate 500-600 mm3, the 4T1 primary tumors were removed at approximately day-18 after tumor implantation. At autopsy, 2 of 12 mice were found to have metastasized to the heart, as brightly visualized by RFP fluorescence, when the mice were sacrificed 6-8 weeks after cell implantation. Post-mortem histological and frozen sample sections showed extensive tumor infiltration into the pericardium, epicardium, and myocardium. The novel mouse model of cardiac metastasis can be used to screen for effective agents.

Citation Format: Yong Zhang, Nan Zhang, Robert M. Hoffman, Ming Zhao. Cardiac metastasis in mice with primary breast carcinoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3957. doi:10.1158/1538-7445.AM2013-3957
Abstract 3962: SGN-LIV1A: a development stage antibody drug-conjugate targeting LIV-1 for the treatment of metastatic breast cancer.
Authors: Sussman Django , Smith Leia M. , Anderson Martha E. , Duniho Steve , Hunter Joshua H. , Kostner Heather , Miyamoto Jamie B. , Nesterova Albina , Westendorf Lori , Van Epps Heather A. , Whiting Nancy , Benjamin Dennis R.
Published: August 2014

LIV-1, also known as SLC39A6 or ZIP6, is a member of the zinc transporter family and was first identified as an estrogen-inducible gene in breast cancer derived cell lines. LIV-1, as a downstream target of STAT3, promotes the epithelial to mesenchymal transition that is important in the malignant progression to metastasis. Consistent with its role in cancer, we determined by immunohistochemical (IHC) analysis that LIV-1 is expressed in subtypes of metastatic breast cancers (ER+/HER2-, HER2+ and triple negative). In healthy human tissues, LIV-1 expression is limited to four hormonally-regulated organs. The broad expression of LIV-1 in metastatic breast cancer in combination with the limited expression in vital organs makes LIV-1 an excellent target for an antibody-drug conjugate (ADC). SGN-LIV1A is an ADC consisting of a humanized anti-LIV-1 mAb conjugated to the microtubule-disrupting agent, monomethyl auristatin E, via a protease-cleavable linker. In vitro, SGN-LIV1A shows target specific internalization and cytotoxic activity against a breast cancer cell line. In vivo studies also demonstrate antitumor activity of SGN-LIV1A in preclinical xenograft models with significant delay of tumor growth compared to control groups. These findings support further evaluation and development of SGN-LIVA as a therapeutic for the treatment of metastatic breast cancer.

Citation Format: Django Sussman, Leia M. Smith, Martha E. Anderson, Steve Duniho, Joshua H. Hunter, Heather Kostner, Jamie B. Miyamoto, Albina Nesterova, Lori Westendorf, Heather A. Van Epps, Nancy Whiting, Dennis R. Benjamin. SGN-LIV1A: a development stage antibody drug-conjugate targeting LIV-1 for the treatment of metastatic breast cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3962. doi:10.1158/1538-7445.AM2013-3962
Abstract 401: Gene expression signatures differentiate primary and metastatic uterine leiomyosarcoma.
Authors: Davidson Ben , Abeler Vera Maria , Førsund Mette , Holth Arild , Shih Ie-Ming , Kristensen Gunnar B. , Chen Li , Yang Yanqin , Wang Tian-Li
Published: August 2014

Leiomyosarcoma (LMS) is the most common uterine sarcoma, but is a rare cancer. The genetic events related to LMS metastasis are unknown to date. The aim of the present study was to compare the global gene expression patterns of primary uterine LMS and LMS metastases in order to identify potential biological targets. Gene expression profiles of 13 primary uterine LMS and 15 LMS metastases were analyzed using the HumanRef-8 BeadChip from Illumina. Differentially expressed candidate genes were validated using quantitative real-time PCR and immunohistochemistry. Unsupervised hierarchical clustering using all 54,675 genes in the array separated primary from metastatic LMS samples. We identified 203 unique probes that were significantly differentially expressed in the two tumor groups by greater than 2-fold with 1% FDR cutoff using one-way ANOVA with Benjamini-Hochberg correction, of which 94 and 109 were overexpressed in primary and metastatic LMS, respectively. Genes overexpressed in primary uterine LMS included OSTN, NLGN4X, NLGN1, SLITRK4, MASP1, XRN2, ASS1, RORB, HRASLS and TSPAN7. Genes overexpressed in LMS metastases included FOLR3, TDO2, CRYM, GJA1, TSPAN10, THBS1, SGK1, SHMT1, EGR2 and AGT. Results for selected genes were validated by quantitative real-time PCR and immunohistochemistry. We present the first study in which gene expression profiling was shown to distinguish between primary uterine LMS and LMS metastases. The molecular signatures unique to primary and metastatic LMS may aid in understanding tumor progression in this cancer and in providing a molecular basis for prognostic studies and therapeutic target discovery.

Citation Format: Ben Davidson, Vera Maria Abeler, Mette Førsund, Arild Holth, Ie-Ming Shih, Gunnar B. Kristensen, Li Chen, Yanqin Yang, Tian-Li Wang. Gene expression signatures differentiate primary and metastatic uterine leiomyosarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 401. doi:10.1158/1538-7445.AM2013-401
Abstract 4032: Transcriptomics analyses of normal and transformed keratinocyte lines generated novel biomarkers for prognosticating outcome in head and neck cancer.
Authors: Grafstrom Roland C. , Merne Marina , Kohonen Pekka , Jerhammar Fredrik , Nees Matthias , Roberg Karin , Ceder Rebecca
Published: August 2014

We tested the hypothesis that characterization of differentiation-related features in human cell lines might reveal prognosticating genes undetected in cancer tissue analyses. Serum-driven changes in transcripts were assessed in otherwise serum-free models of oral epithelium, including normal oral keratinocytes and the transformed keratinocyte lines SVpgC2a, SqCC/Y1 and LK0412. Bioinformatics-driven analyses implicated altered expression of many differentation-related functions, including unique sets of gene ontologies, molecular networks and key regulator genes for each line. Analysis of the complete in vitro/serum exposure-derived gene set relative an information-rich tumor data set identified 17 survival-associated genes, several of which had no previous association to head and neck squamous cell carcinoma (HNSCC). Multi-step validation of these genes relative to independent clinical microarray data sets, including the Human Gene Expression Map and Human Protein Atlas databases, confirmed novel association or predictive correlation with poor outcome in HNSCC for a subset of the genes. The results agreed with the hypothesis under testing, pointing to that cell line studies under a systems biology concept might couple new and prognosis-relevant genes to diverse cancers.

Citation Format: Roland C. Grafstrom, Marina Merne, Pekka Kohonen, Fredrik Jerhammar, Matthias Nees, Karin Roberg, Rebecca Ceder. Transcriptomics analyses of normal and transformed keratinocyte lines generated novel biomarkers for prognosticating outcome in head and neck cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4032. doi:10.1158/1538-7445.AM2013-4032
Abstract 4038: Investigating the effect of glucose-regulated protein 78 (GRP78) inhibition in pancreatic cancer chemoresistance.
Authors: Richards Katherine , Gunn Ellen , Hill Reginald
Published: August 2014

Pancreatic ductal adenocarcinoma (PDAC) is the 4th leading cause of cancer-related deaths in the United States with a 5-year survival rate of 6%. Current therapeutic regimens have done little to halt the progression of this disease. This demonstrates an urgent need for novel therapeutic strategies which address the mechanisms underlying the tumor cells’ ability to develop resistance to drug treatment. Increased expression of GRP78, a member of the Hsp70 protein family, is associated with poor prognosis in a number of human cancers. This prompted us to explore the role of GRP78 expression in pancreatic tumorigenesis.

Our data show that while no GRP78 expression was observed in the ducts of wild-type pancreata, GRP78 expression is significantly increased in the ductal cells of PDACs in our murine models. Moreover, this upregulated GRP78 expression, especially its membrane-associated form, corresponded to P-AKT activation. Our previous results showed that increased P-AKT signaling is significantly associated with distinct pancreatic cancer patient subgroups with more aggressive disease progression. Taken together, these data raised the question of whether GRP78 is essential for pancreatic chemoresistance.

GRP78 expression was inhibited in several gemcitabine resistant PDAC cell lines using siRNA. GRP78 knockdown produced no measurable levels of cell death in any cell lines tested. Likewise, treatment with gemcitabine alone produced very little cell death. However, gemcitabine treatment in cells with GRP78 knockdown showed significantly increased cell death. Examination of cells treated with GRP78 siRNA showed decreased P-AKT activation. These data suggest GRP78 could serve as a potential therapeutic target for chemoresistant pancreatic cancers which show PI3K/AKT pathway activation.

Citation Format: Katherine Richards, Ellen Gunn, Reginald Hill. Investigating the effect of glucose-regulated protein 78 (GRP78) inhibition in pancreatic cancer chemoresistance. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4038. doi:10.1158/1538-7445.AM2013-4038
Abstract 4040: Age-related effects following exposure of bone marrow-derived mesenchymal stem cells to etoposide .
Authors: Hare Ian , Gencheva Marieta , Gibson Laura
Published: August 2014

Mesenchymal stem cells (MSCs) have been shown to be one of the populations that support hematopoietic stem cell (HSC) development within the bone marrow microenvironment. High dose myelosuppressive or ablative chemotherapy regimens that are utilized prior to bone marrow transplantation have the potential to damage MSCs, consistent with damage which we have previously reported for more differentiated stromal and osteoblast cells. As with more differentiated progenitor cells, interactions between MSCs and HSCs are vital to hematopoietic homeostasis with alterations of these interactions having the potential to hamper hematopoietic reconstitution following treatment. In the current study we begin to investigate age associated differences in the vulnerability of the microenvironment to stress utilizing primary human MSCs from passages 6 to 10, characterized functionally by adipocytic and osteogenic differentiation and surface expression profiling (CD105+CD44+CD166+CD45−). We have shown that treatment of MSCs with 25μM etoposide for 24 hours in-vitro causes the occurrence of dsDNA breaks that recover within 48 hours following treatment. In addition, chemotherapy induced stress was followed by an induction of p21. Homologous recombination associated transcripts (XRCC2, XRCC3, Rad51) were reduced following etoposide treatment relative to non-homologous end joining associated transcripts (Ku70, Ku80, DNA-PK), and the magnitude of these differences was augmented with age in-vitro. Pro-apoptotic transcripts Puma and Noxa and the anti-apoptosis associated Bcl-XL showed induction with treatment while Bcl-2 decreased, suggesting the wide range of genes that are responsive to chemotherapy exposure. These results provide insight into the damage responses of MSCs to etoposide and how DNA repair capabilities following chemotherapy could be influenced by patient age. A more mechanistic understanding of the recovery pathways that are altered with age may be valuable for innovative approaches to the treatment of elderly patients for whom escalated chemotherapy is required in an effort to optimize microenvironment supported immune system recovery.

Citation Format: Ian Hare, Marieta Gencheva, Laura Gibson. Age-related effects following exposure of bone marrow-derived mesenchymal stem cells to etoposide . [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4040. doi:10.1158/1538-7445.AM2013-4040
Abstract 4091: RapidCap: A new generation of mouse models for prostate cancer.
Authors: Cho Hyejin , Herzka Tali , Zheng Wu , Castillo-Martin Mireia , Cordon-Cardo Carlos , Trotman Lloyd C.
Published: August 2014

Genetically Engineered Mouse Models are the gold standard for functional cancer research. However, the associated time and cost requirements severely limit their application. As a consequence, projects carry typically a high risk, are lengthy, and scientists become ‘locked in’ with a few chosen candidate alterations. To realize the full potential of mouse modeling technology we have developed RapidCaP, a system that relies on surgical gene transfer. Through prostate speciﬁc delivery of transgenic virus we can 1) reduce model generation times from several years to a few weeks, 2) test various genetic alterations such as loss or gain of function, alone or in combination, and 3) use non-invasive imaging to monitor disease progression. Using RapidCaP, we show that focal loss of Pten and Trp53 genes triggers prostate lesions within 2 months. Furthermore, we ﬁnd that this disease responds to castration by strong regression within several weeks, but it later relapses to produce lethal hormone refractory disease. Taken together, our approach establishes a novel platform for basic prostate cancer research and it realizes the goal of carrying out pre-clinical studies in genetically engineered mice.

Citation Format: Hyejin Cho, Tali Herzka, Wu Zheng, Mireia Castillo-Martin, Carlos Cordon-Cardo, Lloyd C. Trotman. RapidCap: A new generation of mouse models for prostate cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4091. doi:10.1158/1538-7445.AM2013-4091
Abstract 4131: Alternative splicing changes during the acquisition of anoikis resistance.
Authors: Huynh Quoc , Chalfant Charles E. , Park Margaret A.
Published: August 2014

Breast cancer is the most common cancer in women and the second leading cause of cancer-related death among women in the U.S. Metastases from the primary tumor are common and are the main cause of mortality in this population. Therefore, it is imperative to identify therapeutic molecules that target mechanisms specific to metastatic cells. One such mechanism to target is anoikis, a specialized form of programmed cell death that prevents epithelial cells shed from basement membranes from colonizing distant organs. Circulating tumor cells (CTCs) become anoikis-resistant, which is an essential early step in the development of distant metastases in many types of cancer including breast cancer. Recent studies have implicated RNA splicing dysregulation in a number of cancer phenotypes (e.g. oncogenesis).

In these studies, we have developed anoikis-resistant (AnR) breast cancer cell lines, which grow unattached to hydrophilic surfaces. We then performed a preliminary intron / exon array screen comparing parental and AnR breast cancer cell lines. This approach identified a novel splice variant of Non-catalytic region of tyrosine kinase associated protein 1 (NckAP1), in which the 15-18 cassette is deleted. This splice variant is dysregulated in both AnR cells and in relevant samples from breast cancer patients. NckAP1, along with Nck and Ras-related C3 botulinum toxin substrate (Rac), are found in a complex with the actin-polymerization protein Wiskott Aldrich syndrome protein verprolin homologous 1 (WAVE1). After dissociation from this complex, WAVE1 is able to induce polymerization of actin during lamellipodia formation, and is thus a mediator of migration. Hence, splicing of NckAP1 may help mediate migratory capacity of breast cancer cells. These studies also suggest a major role for splicing regulation in metastasis.

Citation Format: Quoc Huynh, Charles E. Chalfant, Margaret A. Park. Alternative splicing changes during the acquisition of anoikis resistance. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4131. doi:10.1158/1538-7445.AM2013-4131
Abstract 4136: Development of a new and sensitive detection technology for galectin cancer proteins.
Authors: van Hattum Hilde , Ruijtenbeek Rob , Hilhorst Riet , Pieters Roland
Published: August 2014

The galectin protein family consists of fifteen galactoside-binding proteins that are present in all classes of life where they function as cell-cell and cell-matrix interaction mediators. Galectins are however also highly overexpressed in various processes related to cancer, including apoptosis, angiogenesis and metastasis. Galectins are therefore excellent targets for tumor diagnostics and medical prognostic purposes. Current proteomic tools, e.g. Polymerase Chain Reaction (PCR) and Enzyme-linked Immunosorbent Assay (ELISA), are based on protein abundance rather than protein activity. In contrast, we have developed a galectin detection assay that visualizes galactoside-binding, i.e. functional, galectins using PamChip® microarray technology. Firstly, antibodies directed against different galectins are efficiently immobilized on the microarray surface by the Fc-binding protein A/G. Then galectin-1 was captured from a spiked cell lysate that is added to the microarray and the unbound proteins are washed away. The galectin presence is subsequently quantified with a general fluorescent ligand that is only bound by active galectins. Several ligands based on lactose and lactosamine were designed and tested. With the best ligands it was possible to detect galectin quantities in the low nanogram range using only 1 μg or less of precious anti-galectin antibodies per measurement. These results are truly promising for the use of the newly developed galectin detection technology in a clinical setting.

Citation Format: Hilde van Hattum, Rob Ruijtenbeek, Riet Hilhorst, Roland Pieters. Development of a new and sensitive detection technology for galectin cancer proteins. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4136. doi:10.1158/1538-7445.AM2013-4136
Abstract 4140: Negative enrichment of circulating tumor cells using geometrically activated surface interaction (GASI) chip.
Authors: Jung Hyo-Il , Hyun Kyung-A , Lee Tae Yoon
Published: August 2014

Circulating tumor cells (CTCs), existing extremely rare in peripheral blood (the order of one cell per 1* 106 nucleated cells), have attracted a lot of attention as they can be exploited to investigate metastasis. The isolation of intact CTCs is challenging owing to their rarity and heterogeneous features.

A lot of methodologies for CTCs enrichment in blood were focused using antibodies to epithelial cell adhesion molecule (EpCAM). However, the recent studies found that the CTCs undergo a process of epithelial-mesenchymal transition (EMT) to adapt oneself to new circumstances which is blood stream and down regulated EpCAM expression on their surface. It is thought that the many CTCs may escape detection by conventional detection methods. So, we decided to use the negative enrichment (i.e. capturing WBCs which are non-target cells and eluting CTCs which are target cells) for isolation of intact and heterogeneous CTCs.

To capture a large number of WBCs in the channel, we developed a geometrically activated surface interaction (GASI)-Chip which is based on the asymmetric herringbone that generate the enhanced mixing flows increasing the surface interaction between the WBCs and CD45 antibody-conjugated channel surface. Using the optimized experimental conditions, we isolated 94.67% of MCF-7 cells (breast cancer cell line) and eliminated 98.94% of Jurkat cells (human T cell lymphoblast like cell line) at 20 μL/min flow rate.

Based on these results, we have attempted to isolate CTCs from patients with several kinds of metastatic cancer patients under the same conditions. The blood samples (1ml) were collected from breast, gastric and lung cancer patients, respectively. After blood samples passed through the GASI-Chip, collected cells were fixed on a slide glass and stained with DAPI for DNA content, PE conjugated Cytokeratin for CTC, and FITC conjugated CD45 for WBC. Following imaging the slide glass, captured images were examined carefully with predefined criteria (DAPI positive, Cytokeratin positive, CD45 negative). The number of isolated CTCs varied from 1 to 51 in 1 ml of blood. Although this device was not intended to process full clinical blood draws, which typically yield 7.5 mL, quite a number of CTCs can be detected from only 1ml of blood compared with other approaches based on the characters of CTCs such as size. Furthermore, the intact and heterogeneous CTCs can be isolated regardless of EpCAM expression because our devices do not require any labeling processes (e.g., EpCAM antibody for CTC).

Citation Format: Hyo-Il Jung, Kyung-A Hyun, Tae Yoon Lee. Negative enrichment of circulating tumor cells using geometrically activated surface interaction (GASI) chip. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4140. doi:10.1158/1538-7445.AM2013-4140
Abstract 4169: Iodine regulates miR-19-mediated inhibition of Smad4 and restores TGFBeta responsiveness in BRAF-activated thyroid cells.
Authors: Fuziwara Cesar S. , Kimura Edna T.
Published: August 2014

Background: BRAF mutation (T1799A) is the most prevalent genetic alteration in thyroid malignancy, associated with aggressive and poor prognosis thyroid cancer. Iodine excess exerts anti-proliferative effects in thyroid cells associated to TGFβ pathway enhancement, although refractoriness to TGFβ inhibitory signal is observed in thyroid cancer. We previously showed that high iodine dose protects thyroid cells from BRAF-induced genomic instability and also, delays oncogenic effects of RET/PTC3 oncogene. MicroRNAs are small non-coding RNAs that regulate protein levels by pairing to 3-UTR of mRNA and blocking translation. MiR-19 belongs to cluster miR-17-92 which attenuates several tumor-suppressor genes and is predicted to target TGFβ pathway components. Therefore, we aim to analyze the influence of iodine on microRNA-mediated TGFβ signaling in BRAFT1799A activated thyroid follicular cells. Methods: BRAF9-6 cells, derived from normal rat thyroid follicular cell, which express BRAFT1799A under doxacyclin stimulation were treated with iodine-containing medium at 10-5M (NaI group) for 5 days. Dox treatment was performed for 2 days (Dox-group), in the absence or presence of iodine (Dox+NaI-group). Control was performed without Dox/NaI. MiR-19 expression was performed by real-time qPCR and Smad4 target validation was performed by Western-blotting and luciferase-assay. TGFβ responsiveness was evaluated by analyzing cell cycle of TGFβ-treated cells using flow-cytometry. Results: BRAFT1799A induction in thyroid cells increased miR-19a and miR-19b levels in 634.3-fold/141.6-fold, respectively (Dox vs Control), while adding iodine prior to BRAFT1799A induction resulted in 2.36-fold/0.66-fold levels (Dox+NaI vs Control). Iodine per se treatment did not change miR-19 levels. BRAFT1799A induction reduced Smad4 protein which was restored by iodine treatment. In parallel to the low Smad4 levels during BRAFT1799A-activation, we observed an impairment of responsiveness to recombinant TGFβ treatment. Moreover, the iodine-containing medium restored the inhibitory signal and rendered cells arrested in G1-phase. Bioinformatic search for miR-19 potential targets revealed Smad4 as a predicted target which was validated by luciferase-assay that showed reduced luminescence of Smad4 3’-UTR plasmid in the presence miR-19. Conclusion: Iodine is a modulator of oncogene activated mir19 expression, which target Smad4 and regulates the efficiency of TGFβ anti-mitogenic action on the BRAFT1799A induced thyroid follicular cells (Supported by FAPESP and CNPq Grant).

Citation Format: Cesar S. Fuziwara, Edna T. Kimura. Iodine regulates miR-19-mediated inhibition of Smad4 and restores TGFBeta responsiveness in BRAF-activated thyroid cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4169. doi:10.1158/1538-7445.AM2013-4169
Abstract 4176: MicroRNA-193b is growth-inhibitory in Ewing sarcoma, and represses ErbB4.
Authors: Moore Colin , Jedlicka Paul
Published: August 2014

Ewing sarcoma, the second most common cancer of bone and soft tissue in adolescents and young adults, is an aggressive malignancy with poor long-term outcome. Ewing sarcoma is driven primarily by the EWS/Fli-1 fusion oncoprotein, which arises from the recurrent t(11,22) translocation. Our laboratory has been interested in identifying microRNAs (miRs) with growth-modulating properties in Ewing sarcoma, manipulation of which could potentially be used for novel therapies, as shown in preclinical studies for other cancers. We have previously shown that replacement of select miRs, normally repressed by EWS/Fli1, can inhibit the growth of Ewing sarcoma cells. To identify additional, and more potent, miRs with growth-suppressive properties in EWS, we have performed a phenotype-based, secondary miniscreen in Ewing sarcoma cell lines. This screen, using transfection of miR mimics into the EWS cell lines A673 and SK-ES-1, followed by growth assays as read-out, identified miR-193b as a strong candidate growth suppressive miR in Ewing sarcoma. Subsequent cloning and stable overexpression of endogenous miR-193b verified the growth suppressive phenotype. Examination of candidate targets further identified ErbB4 as potently repressed upon miR-193b overexpression. ErbB4 has recently been shown to play a key role in Ewing sarcoma, specifically as a driver of metastasis and as a mediator of resistance to cytotoxic chemotherapy, key biological parameters that define high-risk disease. With the need for therapy expansion in pediatric EWS, miR-193b replacement, through ErbB4 inhibition, may be a way to help control high-risk disease.

Citation Format: Colin Moore, Paul Jedlicka. MicroRNA-193b is growth-inhibitory in Ewing sarcoma, and represses ErbB4. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4176. doi:10.1158/1538-7445.AM2013-4176
Abstract 4190: miR-30 as a tumor suppressor connects EGF/Src signal to EMT and ERG.
Authors: Kao Chiao-Jung , Martiniez Anthony , Shi Xu-Bao , Dobi Albert , de Vere White Ralph W. , Kung Hsing-Jien
Published: August 2014

Src tyrosine kinase (Src) is implicated in the development of bone metastasis and castration-resistance of prostate cancer. Src inhibitors are currently being tested in clinical trials for such diseases. Understanding the molecular and cellular actions of Src inhibitors holds the key to future improvement of this line of therapy. Here we describe the microRNA expression profiles modulated by two Src inhibitors and demonstrate that the miR-30 family members are the most prominently induced species. Consistent with its tumor suppressor role, miR-30 is downmodulated by oncogenic signals such as epidermal growth factor (EGF) and hepatocyte growth factor (HGF), and is generally underexpressed in prostate cancer specimens. A number of epithelial to mesenchymal transition (EMT)-associated genes are predicted targets of miR-30. Among these genes the Ets Related Gene (ERG) is the most frequently overexpressed-oncogene in prostate cancer activated by genomic fusion events between promoter upstream sequences of the TMPRSS2 and coding sequences of ERG. We showed by ERG 3’UTR-reporter and mutagenesis assays that ERG is a direct target of miR-30. Overexpression of miR-30 in prostate cancer cells suppresses EMT phenotypes and inhibits cell migration and invasion. It also inhibits the in vitro and in vivo growth of VCaP cells, which depends on TMPRSS2-ERG for proliferation. TMPRSS2-ERG is generally regulated by androgen at the transcriptional level. Our finding reveals a new post-transcriptional mechanism of TMPRSS2-ERG regulation by Src and growth signals via miR-30 providing a rationale for targeting ERG positive castration resistant tumors with Src inhibitors.

Citation Format: Chiao-Jung Kao, Anthony Martiniez, Xu-Bao Shi, Albert Dobi, Ralph W. deVere White, Hsing-Jien Kung. miR-30 as a tumor suppressor connects EGF/Src signal to EMT and ERG. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4190. doi:10.1158/1538-7445.AM2013-4190
Abstract 4198: MiR-17-92 cluster promotes human cholangiocarcinoma growth upon the activation of STAT3 signaling pathway.
Authors: Zhu Hanqing , Han Chang , Wu Tong
Published: August 2014

MiR-17-92 cluster is a potent oncogenic miRNA which has been shown to promote cell proliferation, increase angiogenesis and sustain cell survival in several cancer types. However, the potential role of miR-17-92 cluster in hepatic carcinogenesis has not been investigated. This study was designed to examine the impact of miR-17-92 in hepatic cancer cells. qRT-PCR analysis showed that the expression of miR-17-92 cluster is increased in several human hepatic cancer cell lines compared to the transformed, non-cancerous hepatic cell line, THLE-2. In addition to that, paraffin slides from human hepatic cancer sample also show higher expression of miR-19a as a result of in situ hybridization. The miR-17-92 inhibitors (blocking each individual miRNAs within the cluster) prevented hepatic cancer cell growth. In contrast, stable transfection of pri-miR-17-92 cluster increased cholangiocarcinoma cell (CCLP-1) proliferation. IL-6 treatment greatly upregulates the miR-17-92 cluster mRNA and each individual miRNA level. However, blocking IL-6/STAT3 signaling by STAT3 siRNA or STAT3 phosphorylation inhibitor both inhibit upregulation of miR-17-92 by IL-6 stimulation. Since there is a reported binding site on promoter region of miR-17-92 cluster for STAT3, we used synthesized DNA probe to precipitate STAT3, and found IL-6 treatment increases the STAT3 binding to the promoter of miR-17-92. IL-6 stimulates CCLP-1cell growth, but blocking miR-17-92 will abolish this effect. Targetscan and other prediction tools were used to predict many possible targets of miR-17-92, such as PTEN. Western blot analysis showed decreased PTEN protein level in CCLP-1 cells transfected with the pri-miR-17-92 cluster, suggesting that PTEN may represent one of the miR-17-92 downstream targets. Further experiments are undergoing to show the IL-6 activates STAT3 and thus promote cholangiocarcinoma cell growth through increasing miR-17-92 cluster level.

Citation Format: Hanqing Zhu, Chang Han, Tong Wu. MiR-17-92 cluster promotes human cholangiocarcinoma growth upon the activation of STAT3 signaling pathway. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4198. doi:10.1158/1538-7445.AM2013-4198
Abstract 4203: Overexpression of miR-26a-2 in human liposarcoma is correlated with poor patient survival.
Authors: Lee Dhong Hyun Tony , Amanat Soroosh , Goff Catherine , Weiss Lawrence , Cutter Suzanne , Said Jonathan , Doan Ngan , Ogawa Seishi , Matsubara Aiko , Forscher Charles , Koeffler H.Phillip
Published: August 2014

Approximately 90% of well-differentiated/de-differentiated LPS (WDLPS/DDLPS), the most common LPS subtype, have chromosomal amplification at 12q13-q15. Many protein-coding genes in the region, such as MDM2 and CDK4, have been studied as potential therapeutic targets for LPS treatment with minimal success. Within this amplicon near the MDM2 gene, our SNP array analysis identified frequent amplification of miR-26a-2. Besides being in the amplicon, we found that miR-26a-2 is overexpressed significantly in WDLPS/DDLPS, as well as in myxoid/round cell LPS (MRC). Furthermore, Kaplan-Meier survival analysis showed that overexpression of miR-26a-2 significantly correlated with poor patient survival in both types of LPS.

Based on these findings, we hypothesized that miR-26a-2 plays an important role in LPS tumorigenesis, regardless of LPS subtypes. Overexpression of miR-26a-2 in LPS cells could help the growth and survival of cancer cells in a cell type-specific manner, including faster cell proliferation, faster cell migration, enhanced clonogenicity, suppressed adipocyte differentiation, and/or resistance to apoptosis. Inhibition of miR-26a-2 in LPS cells using anti-miR-26a-2 resulted in the opposite responses.

To explain further the effect of miR-26a-2 overexpression in LPS cells, we performed in sillico analysis and identified 93 candidate targets of miR-26a-2. Among these candidate genes, we found that RCBTB1 (RCC1 and BTB domain containing protein 1) was located at 13q12.3-q14.3, a recurrent region of loss of heterozygosity (LOH) in human LPS. Indeed, overexpression of RCBTB1 made cells more susceptible to apoptosis. Likewise, inhibition of RCBTB1 made cells more resistant to apoptosis.

In conclusion, our study reveals, for the first time, the contribution that miR-26a-2 makes to LPS tumorigenesis, partly through inhibiting RCBTB1. Our study shows that miR-26a-2 is a novel therapeutic target for human LPS.

Citation Format: Dhong Hyun Tony Lee, Soroosh Amanat, Catherine Goff, Lawrence Weiss, Suzanne Cutter, Jonathan Said, Ngan Doan, Seishi Ogawa, Aiko Matsubara, Charles Forscher, H.Phillip Koeffler. Overexpression of miR-26a-2 in human liposarcoma is correlated with poor patient survival. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4203. doi:10.1158/1538-7445.AM2013-4203
Abstract 4243: Regulation of H3K9 methylation by the demethyltransferase gene JMJD1C in multiple myeloma.
Authors: LoBello Janine , DiPerna Danielle , Gale Moly , Watanabe Aprill , Hostetter Galen , Fonseca Rafael , Salhia Bodour
Published: August 2014

Increasing evidence points towards the importance of epigenetic changes in the pathogenesis of multiple myeloma (MM). The biochemical modifications that govern epigenetics are DNA methylation, and post-translational modifications of histone proteins. Histone methylation is catalyzed by histone methyltransferases (HMT) and histone demethylases (HDMT). The purpose of our study was to interrogate genomics data to identify HMT or HDMT genes that are altered in MM. We mined copy number, gene expression and whole genome sequencing data generated as part of the Multiple Myeloma Research Consortium Genomics Initiative. Collectively, the datasets converged on numerous alterations involving histone methylation of lysine 9 (H3K9). One of these genes, the HDMT JMJD1C, was over-expressed in approximately 15% of MM samples examined. We validated over-expression of JMJD1C expression by RT-PCR in over 50 clinical MM samples and 10 human myeloma cell lines (HMCLs). We also confirmed over-expression of JMJD1C by immunohistochemistry (IHC) in a tissue microarray (TMA) consisting of over 60 MM samples. Next we demonstrated a negative correlation between JMJD1C expression and H3K9 methylation by IHC and western blot in 6/10 HMCL and in over 60% of clinical samples examined on the TMA. This validates the H3K9 demethylase activity of JMJD1C. Furthermore, we demonstrated that, in general, H3K9 methylation, a repressive mark associated with heterochromatin, is low in a large percentage of samples. To study the functional significance, we generated a double knockout isogenic cell line pair in U266 cells using zinc finger nuclease technology. We performed ChIP-seq and RNA-seq analyses to identify genes affected by JMJD1C binding to H3K9. Our data indicate that loss of JMJD1C expression alleviates the repression conferred by H3K9 methylation in MM. Studies are ongoing to realize JMJD1C as a potential and novel therapeutic target in MM.

Citation Format: Janine LoBello, Danielle DiPerna, Moly Gale, Aprill Watanabe, Galen Hostetter, Rafael Fonseca, Bodour Salhia. Regulation of H3K9 methylation by the demethyltransferase gene JMJD1C in multiple myeloma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4243. doi:10.1158/1538-7445.AM2013-4243
Abstract 4250: DNA methylation portraits of breast cancer in a population-based cohort.
Authors: Gindin Yevgeniy , Davis Sean , Walker Robert L. , Zhu Yuelin J. , Bilke Sven , Sherman Mark E. , Figueroa Jonine , Meltzer Paul S.
Published: August 2014

Genome scale molecular profiling methods have had a major impact on the classification, diagnosis, and management of breast cancer. However, the biological basis and fundamental consequences of differential DNA methylation across molecular breast cancer subtypes is incompletely understood. In this study, we used gene expression and DNA methylation data from 208 unselected breast cancer samples obtained from a Polish population-based study. We used a network biology approach to identify gene co-methylation modules - sets of highly correlated DNA methylation probes. We found two gene co-methylation modules strongly correlated with Basal (hypermethylated in Basal) and Luminal B (hypermethylated in Luminal B) breast cancer subtypes. Functional analysis of the Basal gene co-methylation module shows that it significantly overlaps with genes silenced during epithelial to mesenchymal transition. The Luminal B gene co-methylation module significantly overlaps with gene targets of the Polycomb repression complex 2. Kaplan-Meier survival analysis revealed poor prognosis associated with hypermethylation of the Luminal B gene co-methylation module. This study offers new biological perspectives of the vital role of DNA methylation in breast cancer.

Citation Format: Yevgeniy Gindin, Sean Davis, Robert L. Walker, Yuelin J. Zhu, Sven Bilke, Mark E. Sherman, Jonine Figueroa, Paul S. Meltzer. DNA methylation portraits of breast cancer in a population-based cohort. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4250. doi:10.1158/1538-7445.AM2013-4250
Abstract 4251: Genomewide methylation and expression profiles in cervical cancer.
Authors: Yongil Kwon , Lee Nakwoo , Lee Sungjong
Published: August 2014

Cervical carcinoma is one of the most common cancers in females. This cancer begins with precancerous stages and can evolves to carcinoma over 10 years or more. Cervical cancer is a multistep process with accumulation of genetic and epigenetic alterations in regulatory genes, leading to activation of oncogenes and inactivation or loss of tumor suppressor genes. We aimed to identify genetic alterations associated with pre-cancerous stage and also with cervical cancer. For this purpose, microarray comparative genomic hybridization(Illumina 450K and Illumina Expression HT12 Human CGH Whole-Genome Arrays) was performed with 16 cases of cervical intraepithelial neoplasia(CIN) of different stages and 3 cases of carcinoma. We identified epigenetic inactivation of TSGs by hyper- or hypo-methylation which has been recognized as an important and alternative mechanism in tumorigenesis. Here we discuss these epigenetic alterations in cervical intrapeithelial neoplasia and cervical cancer focusing on DNA methylation.

Citation Format: Kwon Yongil, Nakwoo Lee, Sungjong Lee. Genomewide methylation and expression profiles in cervical cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4251. doi:10.1158/1538-7445.AM2013-4251
Abstract 4252: Redefining the DNA methylome of prostate cancer using DNA methylation arrays.
Authors: Ramalho-Carvalho Joao , Henrique Rui , Sayols Sergi , Gomez Antonio , Jerónimo Carmen , Esteller Manel
Published: August 2014

Prostate cancer (PCa) is the most common noncutaneous malignancy and a leading cause of cancer death among men. Clinically localized disease might be successfully treated whereas disseminated disease remains mostly lethal. During prostate cancer development and progression, tumor cells undergo abnormal epigenetic modifications, including DNA methylation alterations. DNA methylation is the most studied epigenetic alteration in human cancer, and is associated with transcriptional repression of both coding and non-coding regions of the genome. Aberrant DNA methylation is gaining force in the fields of cancer risk assessment, diagnosis and therapy monitoring in different cancer types, including prostate cancer. Importantly, aberrant hypermethylation is widespread during neoplastic alteration of prostate cells, suggesting that restoration of a normal prostate epigenome through treatment with demethylating drugs could be clinically beneficial.

The main aim of this work was to identify new epigenetically downregulated CpG's in PCa, using the 450K cytosine microarray-based approach, followed by validation in a larger set of clinical samples.

We used the Infinium HumanMethylation450 BeadChip (Illumina) to consistently and significantly detect CpG methylation changes in prostate cancer tissues in comparison with morphological normal prostate tissue. The selected genomic regions were validated by pyrosequencing. The methylation data were subsequently confirmed at mRNA level using qRT-PCR. To prove the functional role of aberrant DNA methylation, we also performed a treatment with 5-Aza-2′-Deoxycytidine in PCa cell lines.

Our work might provide new translational applications to the clinical management of PCa patients, based on of DNA methylation changes in PCa.

Citation Format: Joao Ramalho-Carvalho, Rui Henrique, Sergi Sayols, Antonio Gomez, Carmen Jerónimo, Manel Esteller. Redefining the DNA methylome of prostate cancer using DNA methylation arrays. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4252. doi:10.1158/1538-7445.AM2013-4252
Abstract 4264: PI3K/Akt/mTOR signaling modulation in solar UV-induced skin carcinogenesis.
Authors: Bermudez Yira , Stratton Steve P. , Curiel-Lewandrowski Clara , Hu Chengcheng , Bowden George T. , Saboda Kathylynn , Alberts David S. , Einspahr Janine G.
Published: August 2014

Basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) are the most common types of cancers. Since most BCC and SCC develop on sun-exposed areas of the body, chronic sun exposure is considered the main etiologic factor of these tumors. In vitro and in vivo studies have identified PI3K/Akt/mTOR signaling pathway as a solar light-activated pathway. However, little work has been done on studying the expression profiles of this pathway in humans. In the study herein, we evaluate the modulation of solar light-activated PI3K/Akt/mTOR signaling pathway in normal human skin and solar simulated light (SSL)-irradiated human skin to determine the expression profiles of upstream and downstream phosphorylated proteins Akt (Ser473), mTOR (Ser2448), S6 ribosomal protein (Ser235/236), and 4E-BP1 (Thr37/46). The study population consisted of individuals 18 years or older in general good health with skin types II-III. SSL irradiation at 2, 2.5, and 3 times minimal erythema dose (MED) were applied to unexposed skin. Punch biopsies were taken at baseline, 5 minutes, 1 h, 5 h, and 24 h post-irradiation and immediately fixed in 10% formalin. Tissues were evaluated by immunohistochemistry for the expression of phosphorylated proteins. Proliferating cell nuclear antigen (PCNA) and cleaved caspase 3 (Asp175) were evaluated as markers for keratinocyte proliferation and apoptosis, respectively. Induction of phosphorylated Akt was seen at 5 hours with a 2-fold increase in expression when compared to baseline. Expression of phosphorylated mTOR gradually increased with a peak at 24 hours (3-fold). Significant induction of phosphorylated S6 ribosomal protein was seen at 1 h (3-fold), 5 h (7-fold), and 24 h (10-fold) post SSL irradiation. Induction of phosphorylated 4E-BP1 was limited to the 24 h time point with an increase of 2-fold when compared to baseline. Our data corroborate in vitro and in vivo studies finding, which allow for better understanding of the expression profile patterns of phosphorylated proteins in the PI-3 kinase/Akt/mTOR pathway during exposure of physiological relevant solar-simulated light. Identification of key proteins in this pathway could lead to the use of these proteins as biomarkers and/or specific targets for the prevention and control of skin tumors.

Citation Format: Yira Bermudez, Steve P. Stratton, Clara Curiel-Lewandrowski, Chengcheng Hu, George T. Bowden, Kathylynn Saboda, David S. Alberts, Janine G. Einspahr. PI3K/Akt/mTOR signaling modulation in solar UV-induced skin carcinogenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4264. doi:10.1158/1538-7445.AM2013-4264