bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 4851: A single-tube protocol for next gen library construction increases complexity and simplifies parallel sample handling
Authors: Park Kyusung , Duan Xiaoping , Meredith Rhonda , Kilzer Jennifer M. , White Eric , Ye Delia , Duncan Cisilya , Sokolsky Tanya , Harris Adam N. , Bennett Rob 
Published: June 2012

Abstract
With the advent of second-generation sequencing technologies, exome- and transcriptome- and whole genome sequencing are tools of choice in studying cancer to detect alterations in the genome correlating with tumorigenesis. It is essential to compare genomic alterations in cancer with matched normal DNA from the same individual largely due to incomplete knowledge of the “normal” variations. There have been concerted efforts for targeted sequencing of cancer-related exomes in a high throughput manner. However, the needs to process multiple samples simultaneously while maintaining the complexity of libraries have not been addressed adequately.
Fragment library generation for single- and paired-end reads is typically performed by a series of enzymatic steps following gDNA shearing by ultrasonication. These may include end-repair, A-tailing, and ligation, each followed by bead- or column-based purification. The clean-ups are necessary to prevent enzyme activity carryover from one step to another.
We have developed a single-tube method to take sheared, bead size-selected DNA through end-repair, A-tailing, and ligation without intermittent purifications. The A-tailing is done by a thermophilic polymerase at a temperature sufficient to denature the end-repair enzymes. Ligation is performed by adding ligase and adaptors after cooling the reaction. A bead- or column-based purification is then performed after the ligation reaction to produce template for amplification.
With the removal of the clean-up steps, the workflow is simplified, allowing faster processing of samples with less hands-on time. We also reduce loss of sample associated with the purifications, increasing the complexity of the output library or enabling a lower sample input concentration. Our method is well suited for labs which are producing multiple libraries each week but have not scaled up to full automation.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4851. doi:10.1158/1538-7445.AM2011-4851
Abstract 4857: Improved technology for ribosomal RNA (rRNA) removal from formalin-fixed paraffin-embedded (FFPE) total RNA
Authors: Sooknanan Roy , Radek Agnes , Hitchen John , Khanna Anupama 
Published: June 2012

Abstract
Deep, massively parallel sequencing of cDNA generated from RNA (“RNA-Seq”) is rapidly gaining momentum for transcript profiling, discovery of novel transcripts, and identification of alternative splicing events. Current methods for making sequencer-specific di-tagged DNA fragment libraries for RNA-Seq typically comprise first, preparing rRNA-depleted RNA from total RNA samples that are usually of good quality followed by the synthesis of the di-tagged cDNA sequencing templates. In the past few years however, it has also become evident from microarray and qPCR studies that formalin-fixed paraffin-embedded (FFPE) cancer tissues hold valuable secrets about diseased states. However, RNA-Seq libraries prepared from such highly fragmented FFPE RNA cancer samples yield limited information since these libraries contain a majority of rRNA reads, which decreases sequencing depth and coverage. Current commercially available rRNA-removal kits are not designed to remove fragments of rRNA, which poses a significant limitation for preparing highly informative RNA-Seq libraries from FFPE RNA samples.
Here, we present RNA-Seq results obtained using a novel rRNA removal technology (“Ribo-ZERO™”) and a novel, ligation-free process for preparing directional di-tagged DNA fragment libraries (“ScriptSeq™ Technology”) for RNA-Seq. Using these methods, directional di-tagged DNA fragment libraries can be prepared in about 6 hours from either intact or fragmented (e.g., FFPE) total RNA samples (as little as 100 ng FFPE total RNA sample required). Less than 2 % of the sequence reads from libraries generated from total RNA from either intact or FFPE samples map to rRNA sequences (28S, 18S, 5.8S and 5S). This reduction in rRNA sequence reads from FFPE RNA samples improves sequence depth and coverage, and increases the percentage of uniquely mapped reads, increasing the information obtained from these diseased samples.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4857. doi:10.1158/1538-7445.AM2011-4857
Abstract 4859: Tumor and normal classification of formalin-fixed, paraffin-embedded (FFPE) specimens by transcriptome RNA-seq
Authors: Qu Kunbin , Morlan John , Collin Francois , Millward Carl , Stephans James , Liu Mei-Lan , Jeong Jennie , Baker Joffre , Sinicropi Dominick 
Published: June 2012

Abstract
We have used RNA-seq to profile and compare normal and cancerous human breast tissue. FFPE breast specimens from a total of 24 patients, 12 normal (N) and 12 tumor (T) specimens from surgical resections, were analyzed on an Illumina's GA IIx sequencer. Whole transcriptome RNA-Seq libraries were prepared after depletion of ribosomal RNA by a protocol developed at Genomic Health Inc. (GHI). The analysis was multiplexed across two flow cells using barcoding, with two specimens per sequencing lane (1 T and 1 closely age-matched N library from a different patient). FFPE tissue archive times ranged from 10 to 13 years and they were also closely matched within each lane. To evaluate reproducibility, triplicate libraries were created from 4 of the specimens and analyzed within and across flow cells. Libraries yielded, on average, 19 million 51 bp sequences. R2 values obtained from replicate libraries prepared from the same patient RNA were &gt; 0.9 within and between flow cells. More than 80% of known genes in the human genome were detected in all patients. Several thousand intergenic transcripts were identified by an algorithm developed at GHI. A negative binomial model with tag-wise estimates of dispersion was applied to the known genes and intergenic regions. Inter-patient count variance is generally higher in the set of intergenic sequences than in the set of gene (RefSeq) sequences. Thousands of gene (RefSeq) and intergenic sequences were found to be differentially expressed between T and N tissues. We sought to build classifiers based on flow cell #1 data that could stratify T and N tissues when applied to flow cell #2 data. Sets of genes and intergenic regions were selected for analysis based on high inter-patient count variance. Support vector machine classifiers were trained and then applied to the data from flow cell #2, and also to another GHI tumor/normal RNA-Seq study. Either a set of 100 genes (RefSeq), or a set of 70 intergenic sequences accurately distinguished tumor and normal tissues. Our results offer further evidence of the potential of RNA-Seq for discovery of biomarkers.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4859. doi:10.1158/1538-7445.AM2011-4859
Abstract 4869: p27kip1 is a metastasis-associated antigen that promotes cell migration in osteosarcoma
Authors: Li Yiting , Flores Ricardo , Hicks John , Chintagumpala Murali , Lau Ching , Man Tsz Kwong 
Published: June 2012

Abstract
Metastasis is the leading cause of death in osteosarcoma (OS). Despite the advent of cancer therapy, patients with metastasis still have a dismal outcome. Hence, identification and characterization of new cellular targets that are critical to the metastatic process are urgently needed in order to improve both the treatment and detection of metastatic patients. Previous evidence has indicated that cancer patients produce autoantibodies. Many of these tumor-associated autoantibodies target cellular proteins that are essential for tumor formation. In this study, we used a high-density Human Protein Array (HPA) that contains over 8,000 purified proteins, to identify tumor-associated antigens that are associated with metastatic OS at the time of diagnosis. Plasma samples from patients with metastatic (n=10) or localized (n=16) OS, as well as hospitalized children with non-cancerous diseases (n=21) were pooled separately and analyzed using HPA. Totally, 16 autoantibodies were found to be reactive in metastatic OS. Of those, 6 of them showed higher prevalence in the metastatic cases relative to the localized cases and the control cases (p&lt;0.01). One of these autoantibodies targets a cell cycle inhibitor protein, p27kip1, which is involved in tumor suppression and invasion. The higher prevalence of p27kip1 autoantibody in metastatic OS was subsequently validated using a novel HaloLink assay and individual plasma samples. To test if p27kip1 plays a role in metastatic OS, immunohistochemistry was performed on three pairs of metastatic OS cell lines and their parental, non-metastatic cell lines. We found that the p27kip1 protein, which is normally localized in the nucleus, was mislocalized in the cytoplasm in all three metastatic OS cell lines. Functional studies demonstrated that the wild type or CDK- mutant of p27kip1 harboring a nuclear export signal significantly induced OS cell migration in vitro. In summary, our results indicate that mislocalization of p27kip1 can promote metastatic potential of OS. Further characterization of p27kip1 and its related pathways may lead to the development of a novel therapeutic strategy for treating metastatic OS and other sarcomas.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4869. doi:10.1158/1538-7445.AM2011-4869
Abstract 4871: Photoreactive stapled peptides to capture cancer proteins and define their sites of interaction
Authors: Braun Craig R. , Mintseris Julian , Gavathiotis Evripidis , Bird Gregory H. , Gygi Steven P. , Walensky Loren D. 
Published: June 2012

Abstract
Protein interactions mediate the pathologic cellular activities that contribute to the development, maintenance, and recurrence of cancer. Whether fleeting or stable, homeostatic or aberrant, protein partnerships and their sites of contact form the basis for discovery of biological pathways, disease mechanisms, and opportunities for therapeutic intervention. When the biological activity of cancer-causing proteins is assigned to discrete structural subdomains, their mode of action, menu of protein targets, and explicit sites of target engagement are frequently unknown. Ideally, the evolutionarily honed substructures could be used to capture and thereby catalogue their protein targets; however, out of context from the whole protein, bioactive subdomains often unfold, resulting in loss of biological shape, potency, and specificity. In an effort to reclaim the enormous capacity of structured peptides to selectively bind and capture their protein targets, we combined peptide alpha-helix stabilization, photoaffinity labeling, proteomic analysis, and structure calculation to generate a new strategy for both (1) expanding our grasp of the static and dynamic alpha-helical interactome of cancer proteins and (2) localizing and targeting alpha-helix/protein interaction sites. We piloted our approach using photoreactive stabilized alpha-helices of BCL-2 domains or pSAHBs and demonstrate that they serve as high fidelity research tools capable of efficiently, selectively, and irreversibly crosslinking both static and dynamic protein targets for proteomic and mechanistic analyses. In addition, the compounds were applied to rapidly and correctly locate protein interaction sites on the anti-apoptotic targets BCL-XL and BCL-w. A unique and compelling feature of this approach is the ability to conduct rigorous validation studies for the identified targets and their sites of interaction. Because stapled peptides can penetrate intact cells and access their intracellular targets, stapled peptide derivatives of the very reagents used for covalent capture can be applied to vet and potentially drug the cancer-causing interactions in vivo.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4871. doi:10.1158/1538-7445.AM2011-4871
Abstract 488: Reprogramming cancer cells into terminal differentiation via induced pluripotency
Authors: Matushansky Igor , Zhang Xi 
Published: June 2012

Abstract
Induced pluripotent stem cells (iPSCs) are defined as somatic cells that have been “reprogrammed” (using either the four transcriptional factors Oct4, Sox2, Klf4 and c-Myc or Oct4, Sox2, Nanog, and Lin28) to exhibit embryonic stem cell-like pluripotent differentiation properties. The generation of iPSCs was a great achievement for the field of cell transplantation and tissue engineering. More recently multiple cancer cell lines (e.g., leukemic cells, melanoma cells & gastric cancer cells) have been successfully reprogrammed to the iPSC-state, at least as defined by (1) expression of genes specific to undifferentiated embryonic stem cells; and (2) pluripotency as defined by embryoid body formation in vitro and early markers of commitment to various differentiation lineages.
However, the question that remains unanswered is: can the cancer derived iPSCs complete maturation along a given lineage, be it the lineage from which it was derived (i.e., originally prematurely blocked) or an alternative lineage? If the former were demonstrated it would in essence answer the question of whether global changes in chromatin architecture or ‘epigenetics’ (which are believed to underlie the mechanism of nuclear iPSC reprogramming) can supersede the specific genetic changes (i.e., mutations) in DNA that resulted in the lineage specific differentiation block and subsequent tumorigenesis.
To attempt to answer these questions we generated iPSC-cancer cells from sarcoma cell lines representing undifferentiated sarcomas, osteosarcoma cells, liposarcoma cells, and sarcomas of unknown lineage. While the original sarcoma cell lines could not be differentiated in vitro into mature connective tissue cells, their iPSC-counterparts readily differentiated into multiple mature connective tissue lineages. Furthermore, assessment of proliferation indicated that acquisition of the mature phenotype was concurrent with cessation of proliferation; and thus abrogation of tumorigenicity.
We are currently (1) performing gene expression, miRNA, and global DNA promoter methylation profiling in order to assess the mechanism by which iPSC-based reprogramming of cancer cells over-rides the inherent genetic mutations in these cells and restores the capacity for terminal differentiation; and (2) extending our work to carcinomas.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 488. doi:10.1158/1538-7445.AM2011-488
Abstract 4886: Investigation of cytoskeletal dynamics in real-time during carcinoma cell migration and invasion in vitro
Authors: Conant Carolyn G. , Schwartz Michael , Ionescu-Zanetti Cristian 
Published: June 2012

Abstract
Understanding the fine details of events leading to the establishment of secondary metastatic foci is complicated by methods that are either endpoint or in vivo. We have devised a unique method to investigate real-time events of cell migration and invasion in vitro. We utilized laminar flow cells with a two-inlet geometry to create compartments of either matrigel or chemotactic stimuli. We then introduced carcinoma cells (HT1080) into opposing neutral compartments and imaged the real-time events associated with cell motility. Use of transient transduction of actin-GFP, the myristolyation/palmitoylation sequence from Lck tyrosine kinase-CFP (MPS) and SV40 nuclear-localization sequence-RFP (BacMam) allowed live investigation of cell motility during chemotaxis and invasion. We found that membrane ruffling occurred within minutes of cell adhesion in the neutral compartment. Chemotaxis and invasion occurred within 4 hours of cell introduction, concurrent with massive and dynamic cytoskeletal reorganization. Migration and cell proliferation continued for the entire duration of the data capture up to 24-hours. This method is further extensible to understanding detailed information regarding signaling, inhibition and fluid dynamics on processes of metastatic cell travel through the vasculature, cell adhesion and extravasation.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4886. doi:10.1158/1538-7445.AM2011-4886
Abstract 4893: Microfluidic biochip for the label-free isolation of viablecirculating tumor cells from cancer patients
Authors: Tan Swee Jin , Chen Peng Fei , Lim Wan Teck , Lim Chwee Teck 
Published: June 2012

Abstract
Circulating tumor cells (CTCs) found in the peripheral blood of metastatic cancer patients are rare and this makes them hard to detect. However, the detection and enumeration of CTCs can potentially be used to indicate the severity of the disease or for prognostic and treatment monitoring purposes. Retrieval of viable CTCs can also aid in the study of the phenotypic and genotypic expressions of cancer cells.
We developed an effective label-free detection and isolation of viable CTCs using a microfluidic device that utilizes the unique differences in size and deformability between cancer cells and blood cells. The use of microfluidics also allows the flow conditions to be accurately controlled to achieve an effective and efficient isolation of CTCs. By placing crescent shaped microstructures, termed cell traps, in the path of blood flow in a microchannel, the larger and stiffer CTCs are easily trapped while the blood cells are quickly removed. By reversing the flow, we are able to dislodge and retrieve the trapped CTCs. Using whole blood samples obtained from metastatic lung and kidney cancer patients, we achieved successful detection and separation of CTCs. Here, there is no need for pre-preparation such as staining, tagging or dilution of blood samples as whole blood taken from patient is directly processed in the biochip. The biochip is optically transparent and allows for direct observation using an inverted microscope and for in situ immunofluorescence staining to confirm the presence of CTCs.
Here, we successfully demonstrated the detection and isolation of viable CTCs from cancer patients using the differences in size and deformability of cancer cells and blood cells. This CTC separation process is also label free and will address concerns linked to affinity based isolation methods.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4893. doi:10.1158/1538-7445.AM2011-4893
Abstract 4897: A robust 384-well cell migration assay for high content analysis (HCA) of cells treated with anti-cancer therapeutics
Authors: Fronczak Jennifer A. , Hulkower Keren I. , Vogt Andreas 
Published: June 2012

Abstract
Platypus Technologies has developed a first-in-class 384-well cell migration assay to enable high throughput screening (HTS) of anti-cancer compounds and wound healing agents on adherent tumor and endothelial cell lines. The assay is completely automatable with liquid handling systems and high content analysis (HCA) instrumentation. This assay format utilizes a centrally located self-dissolving, non-toxic biocompatible gel (BCG) to form a uniformly sized, cell-free detection zone on tissue culture treated or collagen I coated cell culture surfaces. Cells are seeded into 384-well plates and pattern in an annular monolayer surrounding the BCG. Once the BCG dissolves, cells can migrate into the detection zone previously occupied by the BCG. Dose-dependent inhibition of migration was observed using the actin polymerization inhibitors Cytochalasin D and Latrunculin A, the kinase inhibitors Dasatinib and Sorafenib on HT-1080, and human umbilical vein endothelial cells (HUVECs), and the phosphatase inhibitors NSC95397 and BCI on MDA-MB231 human breast cancer cells. While migration of cells was faster on collagen coated surfaces than on tissue culture treated surfaces, the IC 50 values obtained for the compounds and rank order of potency were similar on both surfaces. This assay format allows an unobstructed view of cell motility throughout the duration of the experiment. Cells may be fixed and treated with multiple stains, including Hoechst or DAPI to visualize nuclei and TRITC-phalloidin to observe F-actin, to enable flexible data capture by either enumerating migrating cells or by calculating the area of closure within the detection zone. The 384-well assay was successfully automated using a Biomek 2000 robotic liquid handler and a Cellomics ArrayScan II high content imaging (HCI) instrument.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4897. doi:10.1158/1538-7445.AM2011-4897