bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 295: PKCε phosphorylates and modulates the cell membrane translocation of RhoA
Authors: Su Ti-Zhi , Straight Samuel , Bao Li-Wei , Cavey Greg , Pan Quintin 
Published: February 2011

Abstract
Accumulating evidence has clearly defined the importance of PKCε and small Rho GTPases in tumor metastasis. Previous studies in our laboratory have demonstrated that PKCε signals through RhoA to modulate tumor cell invasion and motility in head and neck squamous cell carcinoma. In this study, we further delineated the link between PKCε and RhoA. Recombinant PKCε directly phosphorylated recombinant RhoA using an in vitro kinase assay. Phosphopeptide mapping with LC-MSE identified T127 and S188 as confident phosphorylation sites and T19 and S26 as tentative phosphorylation sites on RhoA. Interestingly, recombinant PKCε was able to bind to recombinant RhoA in the absence of PKC activators and ATP. This observation demonstrates that the binding interaction between PKCε and RhoA does not require an active PKCε conformation and thus, the interaction between these proteins may be more complex than a typical substrate-kinase event. N-terminus fluorophore-tagged PKCε (CherryFP or CFP) and RhoA (GFP or YFP) were co-expressed in HEK293 cells to examine the interaction between these two proteins in live-cells. Live-cell fluorescence microscopy followed by FRET stoichiometric analysis confirmed the interaction between PKCε and RhoA in the cytoplasm of non-stimulated cells. Stimulation of the cells with PMA to activate PKCε resulted in the translocation of PKCε and RhoA from the cytoplasm to the cell membrane and increased the binding of PKCε to RhoA at the cell membrane by three-fold as measured by FRET. Taken together, our results demonstrate, for the first time, that PKCε phosphorylates, co-localizes, and regulates the cell membrane translocation of RhoA.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 295.
Abstract 2953: Evaluating gene expression in FFPE breast cancer tissues using DASL®
Authors: Burr Tom , Dixon Rick , Green Andy , Ellis Ian , Murray Cliff 
Published: February 2011

Abstract
The study of gene expression in conventionally processed tissues is hampered by degradation of mRNA. Expensive, low multiplex, quantitative PCR methods can be unreliable due to the limited template sizes. One way to overcome this problem is to use array-based methods. DASL® technology relies on random priming for production of cDNA, in concert with universal bead-arrays to allow the detection and relative quantitation of expression of specific gene subsets.
Using the Illumina DASL® Cancer Panel (500 cancer-associated genes on one array), we evaluated the expression of key genes in archival FFPE tissue samples from 80 breast cancer patients with well characterised pathological and clinical features. We first assessed transcript integrity in the samples on the basis of levels of mRNA encoding RPL13A, prior to running the Cancer Panel. A subset of genes of interest was then assessed by qPCR to confirm the relative levels observed using the DASL® assay. Finally the expression of the same subset of genes was evaluated at the protein level by immunohistochemistry.
We were able to predict, with good accuracy based upon RPL13A assays, those samples unsuitable for DASL® analysis. Furthermore, the results of DASL® analysis showed good correlation with protein levels, as measured by immunohistochemistry, for a number of key genes including ERBB2 (HER2) and ESR1 (ER). We conclude that DASL® represents a powerful tool for assessing expression of multiple genes in archival tissue.
Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2953.
Abstract 2954: Overexpression of specific genes in surgical margins of head and neck squamous cell carcinoma patients may predict a significantly increased risk of recurrence
Authors: de Carvalho Ana Carolina , Carvalho André L. , Soares Fernando A. , Kowalski Luiz Paulo , Maia Danielle C C , Rettori Marianna M. , Vettore André L. 
Published: February 2011

Abstract
Head and Neck Squamous Cell Carcinoma (HNSCC) is a high incidence and mortality disease. Despite significant improvements in therapeutic strategies, the overall survival rate in 5 years is around 50%. The presence of microscopic tumor in surgical margins is associated with treatment failure, locoregional recurrence and reduction of overall survival. For this reason, it is vital to ensure tumor-free surgical margins in these patients. Commonly, the extension of surgical resection is determined during the surgery through a low sensitivity histological exam of frozen tissue stained with Hematoxylin and Eosin. Since genetic alterations precede histological alterations, molecular markers may be useful to identify cells that have already started malignant transformation without having developed a histological phenotype that could be detected through conventional histopathological techniques. Also, novel molecular techniques make possible to obtain molecular data of surgical margins in a period of time compatible with the surgery, helping the surgeon in the resection of positive molecular margins in the same surgery and thereby reducing the risk of local recurrence. This study aimed to determine the gene expression pattern of specific markers in HNSCC surgical margins and verify the utility of these markers in detecting the presence of tumor cells in these tissues and predict tumor recurrence. Nine selected genes were evaluated and a panel of 5 genes showed a high specificity and good sensibility. Overexpression of at least one of these genes was found in 50% (23/46) of histologically free surgical margins analyzed and was associated with local recurrence. Patients with positive molecular margins were 7.3-fold more likely to have local recurrence. These preliminary results suggest that the approach used in this analysis is more sensible than traditional histopathological exams and could be useful for the intraoperative diagnosis of the presence of tumor cells in HNSCC surgical margins and as an informative tool to identify and monitor subsets of HNSCC patients with an increased risk of recurrence and consequently reduced survival.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2954.
Abstract 296: Protein kinase D regulates Rho GTPase activity through rhotekin
Authors: Pusapati Ganesh Varma , Rykx An , Vandoninck Sandy , van Lint Johan , Adler Guido , Seufferlein Thomas 
Published: February 2011

Abstract
The Protein Kinase D (PKD) family of serine / threonine kinases comprises PKD1,PKD2, and PKD3 and is grouped under the calcium / calmodulin - dependent protein kinase (CAMK) superfamily. PKDs play an important role in various biological processes including proliferation, adhesion, migration, cell shape, survival and apoptosis. Identification of the putative target substrates for this kinase family would provide a deeper insight into their role in the physiological and pathological processes. In the present study, we identified rhotekin (RTKN), an effector protein of small GTPase Rho, as a potential substrate of PKDs. Using the consensus PKD substrate sequence motif scan approach, we identified Ser435 in the C-terminal region of rhotekin as the potential phosphorylatable residue. We generated a phospho-specific antibody corresponding to Ser435 and could validate that Ser435 is indeed the critical amino acid targeted by PKD2 in vivo in HEK-293T cells. The phospho-mimecking mutant of rhotekin (Ser435Glu) when transfected into HEK-293T cells resulted in the increase of endogenous active RhoA GTPase levels, as determined from the Rho activation assay. Rho GTPases were shown to be an important upstream molecules regulating PKD activity in the process of cancer cell migration. However, our results demonstrate for the first time the role of PKDs in the regulation of RhoGTP activity levels. Considering the role of RhoGTPases in various cellular processes, our results provide a hint to the potential regulation of various cellular processes by PKDs through phosphorylation of rhotekin and thereby regulating the activity of RhoGTPases.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 296.
Abstract 2960: TRIM29 is a novel biomarker for basal-like breast cancer
Authors: Liu Jin , Bernard Philip 
Published: February 2011

Abstract
Basal-like breast cancer is a subtype of tumors associated with aggressive clinical behavior and poor prognosis. The genes responsible for their aggressive phenotype have not been well characterized and may provide an opportunity for targeted therapies. Tripartite motif-containing 29 (TRIM29) is a member of the TRIM protein family. Microarray and RT-qPCR analyses shows TRIM29 overexpressed in Basal-like breast tumors and it is highly correlated to other basal tumor markers such as CK5/6, CK17, EGFR, and FOXC1. We recently observed using immunohistochemistry that TRIM29 predominantly localizes to the nucleus in basal tumor epithelial cells, in contrast to a cytoplasmic localization seen in ER-positive tumors. Thus, TRIM29 may act as a transcriptional regulatory factor involved in carcinogenesis. In order to investigate the role of TRIM29 in the initiation and progression of Basal-like breast cancer, we employed chromatin immunoprecipitation and massively parallel sequencing (ChIP-seq) to create a genome-wide profile of in vivo TRIM29-binding sites in MDA-MB-468 cells, a human breast cancer cell line. ChIP-seq analysis revealed 94 putative TRIM29 binding regions in the cancer genome of MDA-MB-468 cells with an estimated false discovery rate of less than 5%. One TRIM29 binding motif was found in the TRIM29 binding regions with a core consensus sequence of CTGTCTCC. We also identified some known cancer genes in close proximity to TRIM29 binding sites, including JUND, FBXO31, CCND3, EGFR, and FOXC1. In addition to mapping TRIM29-DNA binding sites, we used co-immunoprecipitation (co-IP) coupled with tandem mass spectrometry to identify proteins that interact with TRIM29 in MDA-MB-468 cells. Proteins that co-precipitated with TRIM29 included splicing factor proline/glutamine-rich (SFPQ, a transcriptional co-repressor), poly(A) binding protein (PABPC4); FUS and TAF15 RNA polymerase II (both are proto-oncoproteins and transcriptional activators), and A kinase anchor protein 13 (AKAP13, an adaptor protein involved in protein kinase A signaling pathway). Confirmation of the initial co-IP results was achieved through several independent techniques, including detection of binding partners in TRIM29 precipitates by Western blot and reciprocal co-IP using antibodies to partner proteins and detection of TRIM29 in the precipitates. Overall, results from our studies indicated that besides functioning as a transcription factor, TRIM29 may also participate in signal transduction pathways by forming protein complexes. Our studies have identified new biomarkers for basal-like breast cancers and revealed kinases that could potentially be targeted by biologic therapies.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2960.
Abstract 2962: Role of Cyr61 in the progression of pancreatic cancer
Authors: Mehta Smita , Dhar Kakali , Haque Inamul , Majumder Monami , McGregor Douglas H. , Banerjee Sushanta K. , Banerjee Snigdha 
Published: February 2011

Abstract
Pancreatic Ductal Adenocarcinoma (PDAC) is the fourth most common cause of cancer in men and fifth in women in the United States. The overall survival rate is very poor in PDAC due to late presentation, tumor aggressiveness and lack of effective treatment strategy. The objective of this study is to explore the role of Cyr61 in the progression of the PDAC. Cyr61/CCN1 is a secreted, cysteine-rich heparin-binding protein, a member of CCN family and also plays a role in cell proliferation, adhesion, migration and differentiation during development but very little is known about cancer biology. We studied Cyr61 mRNA expression and protein level in tissue samples (archival and tissue array samples) using In Situ hybridization and immunohistochemistry. Its expression was also studied in different cell lines using Northern Blotting, quantitative real time PCR and Western blot analysis. The studies revealed that Cyr61 mRNA and protein was highly expressed in PDAC as compared to normal and chronic pancreatitis sections. The expression profile of Cyr61 increases as the disease progressed from Grade I to Grade III. Interestingly, in some PDAC sections Cyr61 expression was undetected or barely detected. Cyr61 was highly expressed in Panc-1, an invasive cell line, as compared to less invasive cell lines (i.e., BxPC3 and Capan-1) where expression of Cyr61 is very minimal. In order to evaluate the functional role of Cyr61, we nullified Cyr61 in Panc-1 cells using retroviral-mediated shRNA. We determined the migration of these cells and the effect of the conditioned media of these cells on in vitro angiogenesis. We found that Cyr61 is essential for in vitro migration and induction of angiogenesis. Furthermore, these studies reveal that Cyr61 contributes in the development of drug resistance.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2962.
Abstract 2963: The Foxp3 transcription factor expression in human gynecologic cancer cell lines
Authors: Ran Park Yu , Yong-Man Kim , Wha Lee Shin , Dae-Yeon Kim , Jong-Hyeok Kim , Joo-Hyun Nam , Young-Tak Kim 
Published: February 2011

Abstract
Objectives: The forkhead box protein 3 (Foxp3) transcription factor is considered as a master control gene for the function of regulatory T cells. However, recently some publications revealed that Foxp3 in cancer expressed higher than other tissues. A very recent publication describes the expression of Foxp3 in pancreatic carcinoma cells providing evidence that this could be an important tumor escape mechanism. As a result, some research considered that Foxp3 is associated with carcinogenesis, and cancer progression. To this end, This study was scheduled to investigate whether the expression of Foxp3 transcription factor can occur in gynecologic tumor types.
Materials and methods: Ten tumor cell lines of cervical, ovarian, breast cancer origins were selected. The cell lines of cervical cancer were SiHa, HeLa, and Caski. Ovarian cancers were OV-CAR-3, SK-OV-3, SNU-8, and SNU-251. MDA-MB-231, MCF-7,and T47-D were selected as breast cancer cell lines. They were maintained in specific medium (RPM1640, DMEM (Dulbecco's modified Eagle's medium)). The cell lines were cultured for a period of 2 weeks. Detection of Foxp3 mRNA was performed using both conventional quantitative real-time PCR, the RT2 PCR Primer Set for Foxp3 (SuperArray, USA, forward 5’ GGCATCGTGATGGACTCCG 3’ and reverse 5’ GCTGGAAGGTGGACAGCGA 3′). β-Actin was used as a reference gene. All experiments were repeated three times.
Results: The expression of Foxp3 mRNA was revealed in all tumor cell lines studied. This expression did not relate to the tissue of origin of each line. Relative expression values for tumor cells varied widely between 0.21 and 4.97 (1.71±0.09). Cervical cancer cell lines expressed highly Foxp3 mRNA in relative expression values for tumor cells. The highest expression level of Foxp3 in cervical cancer cell lines were with the SiHa cell lines. OV-CAR-3 cell lines expressed highly in ovarian cancer cell lines. In breast cancer cell lines, Foxp3 mRNA expression revealed the highest levels in T47-D cell lines.
Conclusion: Foxp3 mRNA was detected in all tumor cell lines, albeit in variable levels, not related to the origins of tissue. We offer evidence that Foxp3 mRNA expression, characterizes tumor cells of various tissue origins. The biological significance of these findings warrants further investigation in the context of tumor immune escape, and especially under the light of current anti-cancer efforts interfering with Foxp3 expression.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2963.
Abstract 2964: YB-1 expression in early hematopoiesis and leukemic cells
Authors: Bhullar Jasjeet , Sollars Vincent E. 
Published: February 2011

Abstract
Hematopoietic transcription factors play a critical role in directing the commitment and differentiation of hematopoietic stem cells (HSC) along a particular lineage. Y box protein (YB-1), a cold shock family protein is a transcription factor which is widely expressed throughout development and has been implicated as a cell survival factor that regulates transcription and translation. YB-1 is known to be involved in erythroid cell development by interacting with Globin Transcription Factor (GATA); this study aims to investigate YB-1 expression in normal hematopoietic differentiation and leukemia. EML-clone 1 cells, a murine hematopoietic stem cell line, was used as a model for looking at the expression of YB-1 during myeloid differentiation by western blotting and quantitative RT-PCR. Fluorescence activated cell sorting was conducted to isolate lineage——/IL-7R—/c-kit+/Sca1+ (LKS) hematopoietic stem, lineage——/IL-7R—/c-kit+/Sca1— myeloid progenitor cells and granulocytes from mouse bone marrow to assess the YB-1 expression in vivo. YB-1 protein levels were analyzed in a panel of myeloid leukemic cell lines by immunoblotting. YB-1 mRNA and protein levels were high in the EML cells but the expression goes down in RA/IL-3 treated EML-clone 1 cells (myeloid progenitors) and was even more dramatically down-regulated in GM-CSF treated EML cells during the course of myeloid differentiation. Interestingly, LKS (enriched fraction of hematopoietic stem cells) and myeloid progenitor cells showed high level of YB-1 expression as compared to the differentiated cells like granulocytes. Further, we observed that YB-1 protein was expressed in several myeloid leukemic cell lines blocked at different stages of myeloid development. Thus, our data suggest that YB-1 is down-regulated during myeloid differentiation and it might be involved in hematopoietic differentiation. Aberrant YB-1 expression could be a contributing factor in the development of leukemia thus making it an excellent molecular target for therapy in myeloproliferative disorders and leukemia.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2964.
Abstract 2965: Dissecting the molecular mechanism of TLX1-mediated T-cell acute lymphoblastic leukemia development using an in vitro immortalization model
Authors: Zweier-Renn Lynnsey A. , Hawley Teresa S. , Ramezani Ali , Riz Irene , Hawley Robert G. 
Published: February 2011

Abstract
Inappropriate activation of the TLX1 homeobox gene (previously known as HOX11) by chromosomal translocation is a recurrent event in human T-cell acute lymphoblastic leukemia (T-ALL). Enforced expression of TLX1 in murine hematopoietic stem/progenitor cells using a conventional retroviral vector yields immortalized hematopoietic progenitor cell lines. Recent clinical gene therapy trials have shown that retroviral vectors can cause insertional activation of proto-oncogenes which has resulted in the development of hematopoietic clonal dominance or T-ALL-like diseases. Our lab has developed an insulated self-inactivating retroviral vector, RMSinOFB, which has been safety-modified to virtually eliminate insertional genotoxicity during transgene delivery. To observe the effect of reducing retroviral genotoxicity during TLX1 gene transfer, we subcloned the TLX1 gene into the RMSinOFB vector for ectopic expression in murine hematopoietic stem/progenitor cells. Post transduction, TLX1-expressing bone marrow cells had a growth/survival advantage, and readily gave rise to immortalized cell lines. To determine whether insertional mutagenesis played a role in TLX1-mediated immortalization, we identified the integration sites in 15 TLX1-immortalized hematopoietic cell lines by LM-PCR. No common immortalizing genes, such as Evi1 or Prdm16, were identified among the cell lines which underlines the importance of TLX1 transcriptional properties for cell immortalization. Additionally, when tested by RT-PCR no differential vector-influenced expression was observed for candidate genes nearest the integration sites compared to the cell lines lacking such insertions. Furthermore, gene ontology analysis of genes within 100-kb windows of the integration sites did not reveal overrepresentation of a particular functional group. Although several laboratories recently reported the generation of hematopoietic progenitor cell lines due to insertional mutagenesis by retroviral vector backbones, these results confirm and extend our previous findings suggesting that constitutive TLX1 expression alone is sufficient to initiate the series of events culminating in hematopoietic progenitor cell immortalization. We are currently investigating whether non insertion site-related mutations, such as inactivation of the Cdkn2a tumor suppressor locus which collaborates with TLX1 during human T-cell leukemogenesis, may contribute to the process. Investigating the molecular mechanism of TLX1-mediated cellular immortalization in our in vitro model should provide a better understanding of the early stages of TLX1-associated T-ALL development.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2965.