bluuebunny/crossref_metadata_2025_split huggingface dataset:
Abstract 422: Early growth response 1 is required for radiation-induced apoptosis in normal tissues.
Authors: Zhao Yi D. , Jacobs Keith M. , Karvas Rowan M. , Hallahan Dennis E. , Thotala Dinesh 
Published: August 2014

Abstract
Although radiation therapy is a mainstay of cancer treatments, it can have deleterious effects on normal tissues, leading to poor quality of life for cancer survivors. Radiation can cause depletion of stem cells in normal tissues that can result in late sequelae, including poor learning, cognitive disorders, intestinal malabsorption, infertility, and skin injuries. Our laboratory has shown that low doses of radiation can induce apoptosis selectively in undifferentiated normal stem cells compared to adjacent differentiated cells in multiple niches including the brain and intestine. To elucidate the molecular basis underlying stem cell sensitivity to radiation, we performed discovery research to detect genomic differences between stem cells and differentiated cells. Among the candidates, the Early growth response 1 (Egr1) gene was expressed at high levels in stem cells, but not in isogenic differentiated cells. Egr1 is a zinc-finger transcription factor that is rapidly induced in response to radiation and activates the transcription of pro-apoptotic genes. Since cancer survivors can suffer from cognitive difficulties and intestinal malabsorption, we investigated the role of Egr1 in neuronal and intestinal cell death following irradiation. Knockdown of Egr1 using shRNA increased clonogenic survival in irradiated HT-22 hippocampal neuronal cells and IEC-6 intestinal epithelial cells. This increased radioprotection was largely due to reduced apoptosis in Egr1 knockdown cells. Additionally, suppression of Egr1 promoter binding by mithramycin A and chromomycin A3 also conferred radioprotection to HT-22 and IEC-6 cells. Mouse models further supported Egr1’s role in radiation-induced apoptosis. Egr1 null mice had significantly fewer apoptotic cells in the hippocampus and small intestinal crypts following irradiation, as compared to their wild-type littermates. Finally, we examined mechanisms by which Egr1 regulates radiation-induced apoptosis. Irradiation of HT-22 and IEC-6 cells led to a G2/M cell cycle arrest, which was abrogated by the knockdown of Egr1. Also, knockdown of Egr1 by shRNA and inhibition of Egr1 promoter binding both attenuated radiation induction of the pro-apoptotic factors p53, Bim, and Bax, and upregulated the anti-apoptotic protein Bcl-2. Thus, this study establishes a crucial role for Egr1 in the radiation-induced apoptosis of hippocampal and intestinal tissues. Egr1 could be a potential molecular target to minimize the normal tissue complications associated with radiation therapy.
Citation Format: Yi D. Zhao, Keith M. Jacobs, Rowan M. Karvas, Dennis E. Hallahan, Dinesh Thotala. Early growth response 1 is required for radiation-induced apoptosis in normal tissues. [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 422. doi:10.1158/1538-7445.AM2013-422
Abstract 4222: Ion AmpliSeq Panel for the DNA sequencing of BRCA1 and BRCA2 genes on Ion Torrent PGM.
Authors: Costa Jose L. , Mensenkamp Arjen , Rico Alain , Liu Guoying , Petraroli Rosella , Ligtenberg Marjolijn , Machado Jose Carlos 
Published: August 2014

Abstract
The development of massive parallel sequencing (MPS) has revolutionized the field of genomics and created new avenues for basic research. The introduction of the benchtop sequencers enables the development of methodologies for genetic screening that are faster, more comprehensive and more cost-effective than conventional Sanger sequencing. In this study we describe a collaborative effort for the validation of a strategy for the screening of the breast cancer susceptibility genes- BRCA1 and BRCA2, making use of the most recent advances on Ion AmpliSeq™ multiplex PCR technology combined with Ion PGM™ System.
Strict criteria were set to the design of the amplification primers: 1) 100% coverage of all coding exons and exon-intron boundaries; 2) amplicons covering exons must be overlapping; 3) primers must not overlap; 4) no SNPs in the last five nucleotides of primer; and 5) maximum of three non-validated SNPs per primer. Constitutional genomic DNA of 50 BRCA1 or BRCA2 mutated cases are used. The cases are divided in two groups. The first 20 cases are used to test and optimize the methodology and are selected based on the presence of mutations in or within close proximity to homopolymer regions. The additional 30 cases are used for an independent and blind evaluation and power estimation of the new methodology. All samples are studied using both “gold standard” Sanger sequencing and Ion PGM System. For data-analysis various software packages are evaluated.
In the first 19 cases all SNPs and mutations, identified by Sanger sequencing were detected with our Ion AmpliSeq™ based approach. The disease-causing mutations identified included missense, nonsense and indel mutations. The number false positive results were always less than five per sample and reproducible. This work demonstrates the potential for mutational screening of BRCA1 and BRCA2 using the Ion AmpliSeq™ technology combined with the Ion Torrent PGM.
Citation Format: Jose L. Costa, Arjen Mensenkamp, Alain Rico, Guoying Liu, Rosella Petraroli, Marjolijn Ligtenberg, Jose Carlos Machado. Ion AmpliSeq Panel for the DNA sequencing of BRCA1 and BRCA2 genes on Ion Torrent PGM. [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 4222. doi:10.1158/1538-7445.AM2013-4222
Abstract 4223: Combining the CEE-selector assay with sequencing for sensitive mutation detection in cancer.
Authors: Alexiadis Vassilios , Watanaskul Tim , Arnold Lyle 
Published: August 2014

Abstract
We previously described the utility of the CEE-Selector assay for the sensitive detection of the EGFR T790M mutation in plasma samples from lung cancer patients. The assay combined the CEE-Selector assay with melt curve analysis of the mutant PCR product, followed by Sanger sequencing to verify the presence of the mutation. We showed that the assay selectively amplified the mutant sequence compared to wild type by &gt;100,000 fold. We have now expanded the CEE-Selector assay to include additional mutations relevant to patient care (KRAS, BRAF V600E, EGFR L858R). We are presenting data of allele/mutation-specific as well as “footprint” versions of the CEE-Selector assay. The “footprint” version of the assay provides for the interrogation of “hot spot” regions of the genome where multiple cancer relevant mutations are located within a small region, as in the case of KRAS codons 12 and 13 where the CEE-Selector assay is sensitive to all known mutations. Combining the “footprint” version of the CEE-Selector assay with next-generation sequencing, allows us to simultaneously detect, with extremely high sensitivity, a broad array of mutations of interest. Additionally, when used with barcoding, samples from multiple patients can be interrogated simultaneously. The combination of the CEE-Selector assay with next-generation sequencing increases the sensitivity of sequencing by a factor of &gt;1000 and allows for simultaneous sensitive detection of multiple mutations from complex samples, like plasma, where the mutations may be very rare. With the increases in sensitivity made possible using the CEE-Selector assay, the limiting factor for the sensitive detection of mutations/alleles is determined fundamentally by the ability to obtain at least a single copy of the mutant allele within the sample under interrogation.
Citation Format: Vassilios Alexiadis, Tim Watanaskul, Lyle Arnold. Combining the CEE-selector assay with sequencing for sensitive mutation detection in 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 4223. doi:10.1158/1538-7445.AM2013-4223
Abstract 4224: A quick and easy way of assembling TALE/TALEN modules for genetic modifications in mammalian cells .
Authors: Zhang Juan , Mo Yin-Yuan 
Published: August 2014

Abstract
The transcription activator-like effectors (TALEs) are a family of virulence factors produced by plant bacterial pathogens, and they are able to regulate expression of specific host genes by binding to the corresponding promoter regions in a sequence-specific manner. The transcription activator-like effector nuclease (TALEN) is made by fusing a nuclease to the DNA binding modules of TALE, which would allow for generation of DNA deletions, knockouts or knockins. Since their discovery, genes from different organisms have been successfully targeted using designer TALEs through assembling sequence specific modules, which is a critical step to make TALE/TALEN vectors. However, current methods of assembling DNA binding modules, such as the golden-gate ligation, are still time consuming and labor intensive. Moreover, they are also technically challenging. In the present study, we adopted a single step approach to assemble TALE modules. For proof of principle, we used p53 binding site (Tgacttgcatgtacatgcc) derived from the lncRNA-RoR promoter for proof of principle. First, we designed four gBlock DNA oligos which carried 30 bp overlapping between two adjacent oligos. Then, we linked them together by PCR and subsequently directly cloned into an expression vector which carries two flanking TALE sequences. To determine the function of this construct, we generated a reporter vector carrying this p53 binding site and mCherry and showed that this newly made construct was able to activate the mCherry reporter. Experiments are underway to test if the p53 specific TALE is able to activate the endogenous lncRNA-RoR.
Citation Format: Juan Zhang, Yin-Yuan Mo. A quick and easy way of assembling TALE/TALEN modules for genetic modifications in mammalian 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 4224. doi:10.1158/1538-7445.AM2013-4224
Abstract 4227: Simple identification of specific mutations using allele-specific PCR primers and micro capillary electrophoresis (WAVE MCE).
Authors: Richardson Katherine A. , Wu Grant , Lin Rui , Shi Yanggu , Eastlake Phil , Legendre Benjamin 
Published: August 2014

Abstract
Transgenomic, Inc. has developed a size-based analysis for the detection of all EGFR Exon 19 deletions (including complex mutation) greater than 6 base pairs (ACE™ Kit EGFR Exon 19). Currently there are over 70 known EGFR Exon 19 deletions in the region important for response of non-small cell lung cancer (NSCLC) to thymidine kinase inhibitors (TKIs). The majority of these deletions are rare, but the patients may benefit from TKI treatment if the deletion is present. In most current testing paradigms, a NSCLC patient with one of these rare mutations would be considered EGFR mutation negative and not receive TKI therapy. We have designed an easy, size-based PCR assay to detect any of the deletions present in any sample after a single PCR reaction. The assay is based on the separation of PCR fragments of different sizes using microcapillary electrophyoresis (MCE). Transgenomic's WAVE® MCE platform can discriminate the deletions from the wild-type sequence in EGFR Exon 19, however the exact nature of the mutation cannot be determined. If the exact nature of the mutation is required, the PCR fragment can be sequenced.
To confirm that the platform can detect common and rare mutations, plasmids containing various EGFR Exon 19 deletions were synthesized, mixed with EGFR wild-type plasmids and analyzed on the WAVE® MCE platform. The Limit of detection for the plasmid mixing experiments is between 0.5 and 1% for all plasmid constructs tested. These plasmids include 6, 9, 15, 21, 24, and 25 bp deletions . As all samples tested have some wild-type sequence present, 6 base pair deletions cannot be resolved from the wild-type fragment. Several of the deletion constructs contain those not detected by any allele-specific kit, however patients with these deletions, as determined by Sanger sequence analysis, have benefitted from TKI therapy. In addition cell line DNA and DNA isolated from FFPE sections have been analyzed for the detection of EGFR Exon 19 deletions and similar limits of detection (0.5-1.0%) were seen.
This simple, size-based analysis for EGFR Exon deletions will result in NSCLC patients with both common and rare EGFR Exon 19 deletions being able to receive the treatment option best suited for their molecular profile.
Citation Format: Katherine A. Richardson, Grant Wu, Rui Lin, Yanggu Shi, Phil Eastlake, Benjamin Legendre. Simple identification of specific mutations using allele-specific PCR primers and micro capillary electrophoresis (WAVE MCE). [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 4227. doi:10.1158/1538-7445.AM2013-4227
Abstract 4235: Site specific profiling of histone methyltransferases in cancer cells using histone peptide microarray containing a comprehensive set of histone peptides.
Authors: Zhu Bing , Hong Ailing , Hebel Chris , Zhou Xiaochuan , Gao Xiaolian 
Published: August 2014

Abstract
Post-translational modifications (PTMs) of histones play a critical role in diverse biological processes including chromatin compaction, gene expression and cell differentiation. Among a myriad of PMTs, histone methylation catalyzed by histone methyltransferases (HMTs) has been increasingly recognized as an important player responsible for a major signaling mechanism in eukaryotic cells. This suite of epigenetic modifiers represents a new and promising class of therapeutic targets. In cancer, there is a growing body of evidence that suggests changes in the activity of HMTs (a class of chromatin-modifying enzymes) contribute to the uncontrolled cell proliferation that is a hallmark of this devastating disease. The sequence specificity of the substrates of HMTs under a cellular condition are largely unknown but known targets have been mostly identified through a conventional candidate-based approach by using purified HMTs. However, such an experiment frequently does not reflect what could be occurring in cellular contexts or in vivo. In this study, we designed and synthesized a comprehensive histone peptide microarray (PepArray) on a microfluidic chip which contains 3,919 peptides. The peptides contain nine residues with the methylation sites and mutant sites situated in the middle of the sequence so that each peptide has a unique possibility for modification such as methylation or acetylation. We obtained nuclear extract from the breast cancer cell line T47D, and applied the protein lysates to the histone methylation PepArray chip. After incubation of the chip with a methyl-specific antibody, significant signals were detected at the sites containing peptides corresponding to H2AK74, H3K122, and H4K59. We found null signal at mutant sites where the target lysine(K) was replaced with alanine(A). These results reveal the specific activity profiles of HMTs at defined histone sites in a cellular system. Planned further investigation will compare the different histone methylation or acetylation profiles in the various cellular systems, especially in different cancer systems. The current experiment demonstrates an effective solution to comprehensive studies of epigenetic modification. This information may be translated into therapeutic targets of histone methylation inhibition by focusing on identifying inhibitors of specific HMTs as targets for a new generation of therapeutics.
Citation Format: Bing Zhu, Ailing Hong, Chris Hebel, Xiaochuan Zhou, Xiaolian Gao. Site specific profiling of histone methyltransferases in cancer cells using histone peptide microarray containing a comprehensive set of histone peptides. [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 4235. doi:10.1158/1538-7445.AM2013-4235
Abstract 4236: Universal homogenous bioluminescent assay to monitor the effect of modulators on various classes of methyltransferases in vitro.
Authors: Goueli Said A. , Hsiao Kevin , Zegzouti Hicham 
Published: August 2014

Abstract
Methylation/demethylation of DNA and Proteins play major roles in modulation of the epigenome and has been implicated in a wide variety of human diseases. Recent biochemical and biological data suggest that the enzymatic activities of several of these enzymes have pathogenic roles in cancer, inflammation, and neurodegenerative diseases. Thus, pharmacological modulation of these enzymes by small molecules will be beneficial in developing novel therapeutics for multiple unmet medical needs. Towards this goal of searching for activators/inhibitors of these enzymes for the development of next generation of drugs, screening assays for these modulators are urgently needed. To address these unmet needs, we have developed a novel assay that monitors the activities of these enzymes and their modulation by small molecules. The assay is bioluminescent based, HTS formatted and highly sensitive. The assay is universal since it is based on monitoring the formation of the universal product S-adenosylhomocysteine (SAH), i.e., capable of detecting changes in activity of a broad range of methyltransferases such as DNA, protein, and small molecules methyltransferases. In addition, the assay has been validated for all classes of protein methyltransferases (Lysine and Arginine), and with different types of substrates (small peptides, large proteins, or even nucleosomes). This enables determining the specificity of these enzymes and their substrate requirements. The assay has high signal to background and low C.V. The assay is robust (Z’ value &gt; 0.7) and has been validated using various plate densities such as 96-, 384, and 1536-well plates. A strong feature of this assay is its utility with broad range of substrates with no limitations on the use of high concentrations of substrates or the composition of the substrates (short vs. long peptides), thus enabling the generation of kinetic data and determining the mechanism of action of various modulators of methyltransferases of interest.
Citation Format: Said A. Goueli, Kevin Hsiao, Hicham Zegzouti. Universal homogenous bioluminescent assay to monitor the effect of modulators on various classes of methyltransferases in vitro. [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 4236. doi:10.1158/1538-7445.AM2013-4236
Abstract 4241: Isozyme-selective intracellular binding assay for histone deacetylases .
Authors: Woodroofe Carolyn C. , Robers Matthew B. , Kirkland Thomas A. , Kupcho Kevin , Niles Andrew L. , Cong Mei , Wood Keith V. 
Published: August 2014

Abstract
The precise modulation of histone acetylation is crucial to cellular survival and control of reproduction.Misregulation of acetylation is often correlated with uncontrolled cellular proliferation, and general inhibition of histone deacetylases has proven to be an effective strategy in the treatment of some cancers. The underlying biology of histone deacetylases is however quite complex, with four classes of isozymes and a large number of interacting proteins that modulate the activity of each. The general scarcity of potent isozyme-selective HDAC inhibitors further complicates the task of deciphering individual isozyme activity in a cellular context. We now report a novel approach to monitoring the intracellular inhibition of single isozymes based on bioluminescence resonance energy transfer from NanoLucTM, a small and exceedingly bright bioluminescent reporter. In our approach, a NanoLucTM fusion of the isozyme of interest is used in combination with an optimized fluorescent tracer. When bound, energy transfer occurs from the bioluminescent enzyme to the fluorescent probe. Displacement of the probe by a competitive inhibitor is then measured as a decrease in the energy transfer ratio, yielding IC50 data specific to the chosen isozyme. We demonstrate the ability to measure intracellular IC50 values of inhibitors against individual isozymes of class I and class IIb based on choice of the NanoLucTM fusion.
Citation Format: Carolyn C. Woodroofe, Matthew B. Robers, Thomas A. Kirkland, Kevin Kupcho, Andrew L. Niles, Mei Cong, Keith V. Wood. Isozyme-selective intracellular binding assay for histone deacetylases . [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 4241. doi:10.1158/1538-7445.AM2013-4241
Abstract 4274: Prolidase directly binds and activates epidermal growth factor receptor and stimulates downstream signaling.
Authors: Yang Lu 
Published: August 2014

Abstract
Prolidase, also known as Xaa-Pro dipeptidase or peptidase D (PEPD), is a ubiquitously expressed enzyme that hydrolyzes dipeptides with proline or hydroxyproline at the carboxy terminus. Unexpectedly, we have found that recombinant human PEPD directly binds and activates epidermal growth factor receptor (EGFR), and that EGFR activation by PEPD in turn leads to activation of multiple signaling molecules downstream of EGFR, including but may not be limited to AKT, ERK and STAT3. EGFR is a cell surface receptor that plays a major role in a variety of cellular responses, and EGFR activation is known to result in increase in DNA synthesis, cell growth, cell proliferation and cell migration, which perhaps are most clearly demonstrated in cancer cells. The ability of PEPD to activate EGFR signaling is neither cell-specific nor dependent on its enzymatic activity; in the latter case, an enzymatically inactive PEPD mutant was similarly effective in activating EGFR, AKT, ERK and STAT3. PEPD is also a novel inducer of cyclooxygenase-2 (Cox-2), known to play an important role in cancer, inflammation and other diseases, but Cox-2 induction by PEPD also depends on EGFR activation. In line with the pro-survival and pro-proliferation activities of EGFR and Cox-2, PEPD stimulates cell proliferation and DNA synthesis in cultured cells. However, PEPD could activate EGFR only when it was present in the extracellular space, as forced expression of human PEPD via transient gene transfection failed to activate EGFR and the downstream signaling proteins. This result is nonetheless consistent with EGFR being a cell surface receptor. Although PEPD is normally a cytosolic protein, it is apparently released from injured cells and tissues in vivo, and such release appears to result in EGFR activation and activation of its downstream signals. PEPD differs from all known EGFR ligands in that it does not possess an epidermal growth factor (EGF) motif and is not synthesized as a transmembrane precursor. Indeed, there are several notable differences between PEPD and EGF with regard to binding and activation of EGFR and activation of downstream signaling molecules. In conclusion, PEPD is a new ligand of EGFR and presents a novel mechanism of EGFR activation.
Citation Format: Lu Yang. Prolidase directly binds and activates epidermal growth factor receptor and stimulates downstream signaling. [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 4274. doi:10.1158/1538-7445.AM2013-4274