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Sample records for affinity responsive target

  1. Identification of Multiple Cryptococcal Fungicidal Drug Targets by Combined Gene Dosing and Drug Affinity Responsive Target Stability Screening

    Directory of Open Access Journals (Sweden)

    Yoon-Dong Park

    2016-08-01

    Full Text Available Cryptococcus neoformans is a pathogenic fungus that is responsible for up to half a million cases of meningitis globally, especially in immunocompromised individuals. Common fungistatic drugs, such as fluconazole, are less toxic for patients but have low efficacy for initial therapy of the disease. Effective therapy against the disease is provided by the fungicidal drug amphotericin B; however, due to its high toxicity and the difficulty in administering its intravenous formulation, it is imperative to find new therapies targeting the fungus. The antiparasitic drug bithionol has been recently identified as having potent fungicidal activity. In this study, we used a combined gene dosing and drug affinity responsive target stability (GD-DARTS screen as well as protein modeling to identify a common drug binding site of bithionol within multiple NAD-dependent dehydrogenase drug targets. This combination genetic and proteomic method thus provides a powerful method for identifying novel fungicidal drug targets for further development.

  2. Specific capture of uranyl protein targets by metal affinity chromatography

    International Nuclear Information System (INIS)

    Basset, C.; Dedieu, A.; Guerin, P.; Quemeneur, E.; Meyer, D.; Vidaud, C.

    2008-01-01

    To improve general understanding of biochemical mechanisms in the field of uranium toxicology, the identification of protein targets needs to be intensified. Immobilized metal affinity chromatography (IMAC) has been widely developed as a powerful tool for capturing metal binding proteins from biological extracts. However uranyl cations (UO 2 2+ ) have particular physico-chemical characteristics which prevent them from being immobilized on classical metal chelating supports. We report here on the first development of an immobilized uranyl affinity chromatography method, based on the cation-exchange properties of amino-phosphonate groups for uranyl binding. The cation distribution coefficient and loading capacity on the support were determined. Then the stability of the uranyl-bonded phase under our chromatographic conditions was optimized to promote affinity mechanisms. The successful enrichment of uranyl binding proteins from human serum was then proven using proteomic and mass spectral analysis. (authors)

  3. Affinity resins as new tools for identifying target proteins of ascorbic acid.

    Science.gov (United States)

    Iwaoka, Yuji; Nishino, Kohei; Ishikawa, Takahiro; Ito, Hideyuki; Sawa, Yoshihiro; Tai, Akihiro

    2018-02-12

    l-Ascorbic acid (AA) has diverse physiological functions, but little is known about the functional mechanisms of AA. In this study, we synthesized two types of affinity resin on which AA is immobilized in a stable form to identify new AA-targeted proteins, which can provide important clues for elucidating unknown functional mechanisms of AA. To our knowledge, an affinity resin on which AA as a ligand is immobilized has not been prepared, because AA is very unstable and rapidly degraded in an aqueous solution. By using the affinity resins, cytochrome c (cyt c) was identified as an AA-targeted protein, and we showed that oxidized cyt c exhibits specific affinity for AA. These results suggest that two kinds of AA-affinity resin can be powerful tools to identify new target proteins of AA.

  4. Supramolecular Affinity Chromatography for Methylation-Targeted Proteomics.

    Science.gov (United States)

    Garnett, Graham A E; Starke, Melissa J; Shaurya, Alok; Li, Janessa; Hof, Fraser

    2016-04-05

    Proteome-wide studies of post-translationally methylated species using mass spectrometry are complicated by high sample diversity, competition for ionization among peptides, and mass redundancies. Antibody-based enrichment has powered methylation proteomics until now, but the reliability, pan-specificity, polyclonal nature, and stability of the available pan-specific antibodies are problematic and do not provide a standard, reliable platform for investigators. We have invented an anionic supramolecular host that can form host-guest complexes selectively with methyllysine-containing peptides and used it to create a methylysine-affinity column. The column resolves peptides on the basis of methylation-a feat impossible with a comparable commercial cation-exchange column. A proteolyzed nuclear extract was separated on the methyl-affinity column prior to standard proteomics analysis. This experiment demonstrates that such chemical methyl-affinity columns are capable of enriching and improving the analysis of methyllysine residues from complex protein mixtures. We discuss the importance of this advance in the context of biomolecule-driven enrichment methods.

  5. Identification of Thioredoxin Target Disulfides Using Isotope-Coded Affinity Tags

    DEFF Research Database (Denmark)

    Hägglund, Per; Bunkenborg, Jakob; Maeda, Kenji

    2014-01-01

    Thioredoxins (Trx) are small redox proteins that reduce disulfide bonds in various target proteins and maintain cellular thiol redox control. Here, a thiol-specific labeling and affinity enrichment approach for identification and relative quantification of Trx target disulfides in complex protein...... reduction is determined by LC-MS/MS-based quantification of tryptic peptides labeled with "light" (12C) and "heavy" (13C) ICAT reagents. The methodology can be adapted to monitor the effect of different reductants or oxidants on the redox status of thiol/disulfide proteomes in biological systems....... extracts is described. The procedure utilizes the isotope-coded affinity tag (ICAT) reagents containing a thiol reactive iodoacetamide group and a biotin affinity tag to target peptides containing reduced cysteine residues. The identification of substrates for Trx and the extent of target disulfide...

  6. Targeting Anti-Cancer Active Compounds: Affinity-Based Chromatographic Assays

    OpenAIRE

    de Moraes, Marcela Cristina; Cardoso, Carmen Lucia; Seidl, Claudia; Moaddel, Ruin; Cass, Quezia Bezerra

    2016-01-01

    Affinity-based chromatography assays encompass the use of solid supports containing immobilized biological targets to monitor binding events in the isolation , identification and/or characterization of bioactive compounds. This powerful bioanalytical technique allows the screening of potential binders through fast analyses that can be directly performed using isolated substances or complex matrices. An overview of the recent researches in frontal and zonal affinity-based chromatography screen...

  7. Targeting Anti-Cancer Active Compounds: Affinity-Based Chromatographic Assays

    Science.gov (United States)

    de Moraes, Marcela Cristina; Cardoso, Carmen Lucia; Seidl, Claudia; Moaddel, Ruin; Cass, Quezia Bezerra

    2016-01-01

    Affinity-based chromatography assays encompass the use of solid supports containing immobilized biological targets to monitor binding events in the isolation , identification and/or characterization of bioactive compounds. This powerful bioanalytical technique allows the screening of potential binders through fast analyses that can be directly performed using isolated substances or complex matrices. An overview of the recent researches in frontal and zonal affinity-based chromatography screening assays, which has been used as a tool in the identification and characterization of new anti-cancer agents, is discussed. In addition, a critical evaluation of the recently emerged ligands fishing assays in complex mixtures is also discussed. PMID:27306095

  8. Endothelial targeting of high-affinity multivalent polymer nanocarriers directed to intercellular adhesion molecule 1.

    Science.gov (United States)

    Muro, Silvia; Dziubla, Thomas; Qiu, Weining; Leferovich, John; Cui, Xiumin; Berk, Erik; Muzykantov, Vladimir R

    2006-06-01

    Targeting of diagnostic and therapeutic agents to endothelial cells (ECs) provides an avenue to improve treatment of many maladies. For example, intercellular adhesion molecule 1 (ICAM-1), a constitutive endothelial cell adhesion molecule up-regulated in many diseases, is a good determinant for endothelial targeting of therapeutic enzymes and polymer nanocarriers (PNCs) conjugated with anti-ICAM (anti-ICAM/PNCs). However, intrinsic and extrinsic factors that control targeting of anti-ICAM/PNCs to ECs (e.g., anti-ICAM affinity and PNC valency and flow) have not been defined. In this study we tested in vitro and in vivo parameters of targeting to ECs of anti-ICAM/PNCs consisting of either prototype polystyrene or biodegradable poly(lactic-coglycolic) acid polymers (approximately 200 nm diameter spheres carrying approximately 200 anti-ICAM molecules). Anti-ICAM/PNCs, but not control IgG/PNCs 1) rapidly (t1/2 approximately 5 min) and specifically bound to tumor necrosis factor-activated ECs in a dose-dependent manner (Bmax approximately 350 PNC/cell) at both static and physiological shear stress conditions and 2) bound to ECs and accumulated in the pulmonary vasculature after i.v. injection in mice. Anti-ICAM/PNCs displayed markedly higher EC affinity versus naked anti-ICAM (Kd approximately 80 pM versus approximately 8 nM) in cell culture and, probably because of this factor, higher value (185.3 +/- 24.2 versus 50.5 +/- 1.5% injected dose/g) and selectivity (lung/blood ratio 81.0 +/- 10.9 versus 2.1 +/- 0.02, in part due to faster blood clearance) of pulmonary targeting. These results 1) show that reformatting monomolecular anti-ICAM into high-affinity multivalent PNCs boosts their vascular immuno-targeting, which withstands physiological hydrodynamics and 2) support potential anti-ICAM/PNCs utility for medical applications.

  9. Direct molecular mimicry enables off-target cardiovascular toxicity by an enhanced affinity TCR designed for cancer immunotherapy.

    Science.gov (United States)

    Raman, Marine C C; Rizkallah, Pierre J; Simmons, Ruth; Donnellan, Zoe; Dukes, Joseph; Bossi, Giovanna; Le Provost, Gabrielle S; Todorov, Penio; Baston, Emma; Hickman, Emma; Mahon, Tara; Hassan, Namir; Vuidepot, Annelise; Sami, Malkit; Cole, David K; Jakobsen, Bent K

    2016-01-13

    Natural T-cell responses generally lack the potency to eradicate cancer. Enhanced affinity T-cell receptors (TCRs) provide an ideal approach to target cancer cells, with emerging clinical data showing significant promise. Nevertheless, the risk of off target reactivity remains a key concern, as exemplified in a recent clinical report describing fatal cardiac toxicity, following administration of MAGE-A3 specific TCR-engineered T-cells, mediated through cross-reactivity with an unrelated epitope from the Titin protein presented on cardiac tissue. Here, we investigated the structural mechanism enabling TCR cross-recognition of MAGE-A3 and Titin, and applied the resulting data to rationally design mutants with improved antigen discrimination, providing a proof-of-concept strategy for altering the fine specificity of a TCR towards an intended target antigen. This study represents the first example of direct molecular mimicry leading to clinically relevant fatal toxicity, mediated by a modified enhanced affinity TCR designed for cancer immunotherapy. Furthermore, these data demonstrate that self-antigens that are expressed at high levels on healthy tissue should be treated with extreme caution when designing immuno-therapeutics.

  10. Characterization of the ER-Targeted Low Affinity Ca2+ Probe D4ER

    Directory of Open Access Journals (Sweden)

    Elisa Greotti

    2016-09-01

    Full Text Available Calcium ion (Ca2+ is a ubiquitous intracellular messenger and changes in its concentration impact on nearly every aspect of cell life. Endoplasmic reticulum (ER represents the major intracellular Ca2+ store and the free Ca2+ concentration ([Ca2+] within its lumen ([Ca2+]ER can reach levels higher than 1 mM. Several genetically-encoded ER-targeted Ca2+ sensors have been developed over the last years. However, most of them are non-ratiometric and, thus, their signal is difficult to calibrate in live cells and is affected by shifts in the focal plane and artifactual movements of the sample. On the other hand, existing ratiometric Ca2+ probes are plagued by different drawbacks, such as a double dissociation constant (Kd for Ca2+, low dynamic range, and an affinity for the cation that is too high for the levels of [Ca2+] in the ER lumen. Here, we report the characterization of a recently generated ER-targeted, Förster resonance energy transfer (FRET-based, Cameleon probe, named D4ER, characterized by suitable Ca2+ affinity and dynamic range for monitoring [Ca2+] variations within the ER. As an example, resting [Ca2+]ER have been evaluated in a known paradigm of altered ER Ca2+ homeostasis, i.e., in cells expressing a mutated form of the familial Alzheimer’s Disease-linked protein Presenilin 2 (PS2. The lower Ca2+ affinity of the D4ER probe, compared to that of the previously generated D1ER, allowed the detection of a conspicuous, more clear-cut, reduction in ER Ca2+ content in cells expressing mutated PS2, compared to controls.

  11. Enhanced membrane pore formation through high-affinity targeted antimicrobial peptides.

    Directory of Open Access Journals (Sweden)

    Christopher J Arnusch

    Full Text Available Many cationic antimicrobial peptides (AMPs target the unique lipid composition of the prokaryotic cell membrane. However, the micromolar activities common for these peptides are considered weak in comparison to nisin, which follows a targeted, pore-forming mode of action. Here we show that AMPs can be modified with a high-affinity targeting module, which enables membrane permeabilization at low concentration. Magainin 2 and a truncated peptide analog were conjugated to vancomycin using click chemistry, and could be directed towards specific membrane embedded receptors both in model membrane systems and whole cells. Compared with untargeted vesicles, a gain in permeabilization efficacy of two orders of magnitude was reached with large unilamellar vesicles that included lipid II, the target of vancomycin. The truncated vancomycin-peptide conjugate showed an increased activity against vancomycin resistant Enterococci, whereas the full-length conjugate was more active against a targeted eukaryotic cell model: lipid II containing erythrocytes. This study highlights that AMPs can be made more selective and more potent against biological membranes that contain structures that can be targeted.

  12. Development of immobilized membrane-based affinity columns for use in the online characterization of membrane bound proteins and for targeted affinity isolations

    International Nuclear Information System (INIS)

    Moaddel, Ruin; Wainer, Irving W.

    2006-01-01

    Membranes obtained from cell lines that express or do not express a target membrane bound protein have been immobilized on a silica-based liquid chromatographic support or on the surface of an activated glass capillary. The resulting chromatographic columns have been placed in liquid chromatographic systems and used to characterize the target proteins and to identify small molecules that bind to the target. Membranes containing ligand gated ion channels, G-protein coupled receptors and drug transporters have been prepared and characterized. If a marker ligand has been identified for the target protein, frontal or zonal displacement chromatographic techniques can be used to determine binding affinities (K d values) and non-linear chromatography can be used to assess the association (k on ) and dissociation (k off ) rate constants and the thermodynamics of the binding process. Membrane-based affinity columns have been created using membranes from a cell line that does not express the target protein (control) and the same cell line that expresses the target protein (experimental) after genomic transfection. The resulting columns can be placed in a parallel chromatography system and the differential retention between the control and experimental columns can be used to identify small molecules and protein that bind to the target protein. These applications will be illustrated using columns created using cellular membranes containing nicotinic acetylcholine receptors and the drug transporter P-glycoprotein

  13. Development of immobilized membrane-based affinity columns for use in the online characterization of membrane bound proteins and for targeted affinity isolations

    Energy Technology Data Exchange (ETDEWEB)

    Moaddel, Ruin [Gerontology Research Center, National Institute on Aging, National Institutes of Health, 5600 Nathan Shock Drive, Baltimore, MD 21224-6825 (United States); Wainer, Irving W. [Gerontology Research Center, National Institute on Aging, National Institutes of Health, 5600 Nathan Shock Drive, Baltimore, MD 21224-6825 (United States)]. E-mail: Wainerir@grc.nia.nih.gov

    2006-03-30

    Membranes obtained from cell lines that express or do not express a target membrane bound protein have been immobilized on a silica-based liquid chromatographic support or on the surface of an activated glass capillary. The resulting chromatographic columns have been placed in liquid chromatographic systems and used to characterize the target proteins and to identify small molecules that bind to the target. Membranes containing ligand gated ion channels, G-protein coupled receptors and drug transporters have been prepared and characterized. If a marker ligand has been identified for the target protein, frontal or zonal displacement chromatographic techniques can be used to determine binding affinities (K {sub d} values) and non-linear chromatography can be used to assess the association (k {sub on}) and dissociation (k {sub off}) rate constants and the thermodynamics of the binding process. Membrane-based affinity columns have been created using membranes from a cell line that does not express the target protein (control) and the same cell line that expresses the target protein (experimental) after genomic transfection. The resulting columns can be placed in a parallel chromatography system and the differential retention between the control and experimental columns can be used to identify small molecules and protein that bind to the target protein. These applications will be illustrated using columns created using cellular membranes containing nicotinic acetylcholine receptors and the drug transporter P-glycoprotein.

  14. Radiosensitizing and cytotoxic properties of DNA targeted phenanthridine-linked nitroheterocycles of varying electron affinities

    International Nuclear Information System (INIS)

    Cowan, D.S.M.; Rauth, A.M.; Toronto Univ., ON; Matejovic, J.F.; McClelland, R.A.; Wardman, P.

    1994-01-01

    2-Nitroimidazoles targeted to DNA via intercalation have previously been shown to be as much as 10-100 times more efficient on a molar basis than the untargeted nitroimidazole, misonidazole, in vitro as hypoxic cell selective radiosensitizers and cytotoxins based on extracellular concentrations. In this work the effect of varying the nitroaromatic group has been examined through the preparation of a DNA-targeted 4-nitroimidazole (4-MeNLP-3), a 5-nitroimidazole (5-NLP-3) and a 5-nitrofuran (FEP-2) linked to phenanthridinium ions. With the previously synthesized 2-nitroimidazoles, this provides a series of DNA targeted compounds of varying electron affinity as well as structure at the nitroaromatic position. The present series of compounds was tested for partition coefficient, DNA binding ability, reduction potentials and in vitro radiosensitizing and cytotoxic abilities. The results obtained indicate that targeting such compounds to DNA diminishes the dependency of radiosensitizing and cytotoxic properties on reduction potential and may allow significant uncoupling of toxicity from radiosensitizing ability. (author)

  15. Accurate and Reliable Prediction of the Binding Affinities of Macrocycles to Their Protein Targets.

    Science.gov (United States)

    Yu, Haoyu S; Deng, Yuqing; Wu, Yujie; Sindhikara, Dan; Rask, Amy R; Kimura, Takayuki; Abel, Robert; Wang, Lingle

    2017-12-12

    Macrocycles have been emerging as a very important drug class in the past few decades largely due to their expanded chemical diversity benefiting from advances in synthetic methods. Macrocyclization has been recognized as an effective way to restrict the conformational space of acyclic small molecule inhibitors with the hope of improving potency, selectivity, and metabolic stability. Because of their relatively larger size as compared to typical small molecule drugs and the complexity of the structures, efficient sampling of the accessible macrocycle conformational space and accurate prediction of their binding affinities to their target protein receptors poses a great challenge of central importance in computational macrocycle drug design. In this article, we present a novel method for relative binding free energy calculations between macrocycles with different ring sizes and between the macrocycles and their corresponding acyclic counterparts. We have applied the method to seven pharmaceutically interesting data sets taken from recent drug discovery projects including 33 macrocyclic ligands covering a diverse chemical space. The predicted binding free energies are in good agreement with experimental data with an overall root-mean-square error (RMSE) of 0.94 kcal/mol. This is to our knowledge the first time where the free energy of the macrocyclization of linear molecules has been directly calculated with rigorous physics-based free energy calculation methods, and we anticipate the outstanding accuracy demonstrated here across a broad range of target classes may have significant implications for macrocycle drug discovery.

  16. Microtubule affinity-regulating kinases are potential druggable targets for Alzheimer's disease.

    Science.gov (United States)

    Annadurai, Narendran; Agrawal, Khushboo; Džubák, Petr; Hajdúch, Marián; Das, Viswanath

    2017-11-01

    Alzheimer's disease (AD) is a progressive neurodegenerative disorder that affects normal functions of the brain. Currently, AD is one of the leading causes of death in developed countries and the only one of the top ten diseases without a means to prevent, cure, or significantly slow down its progression. Therefore, newer therapeutic concepts are urgently needed to improve survival and the quality of life of AD patients. Microtubule affinity-regulating kinases (MARKs) regulate tau-microtubule binding and play a crucial role in neurons. However, their role in hyperphosphorylation of tau makes them potential druggable target for AD therapy. Despite the relevance of MARKs in AD pathogenesis, only a few small molecules are known to have anti-MARK activity and not much has been done to progress these compounds into therapeutic candidates. But given the diverse role of MARKs, the specificity of novel inhibitors is imperative for their successful translation from bench to bedside. In this regard, a recent co-crystal structure of MARK4 in association with a pyrazolopyrimidine-based inhibitor offers a potential scaffold for the development of more specific MARK inhibitors. In this manuscript, we review the biological role of MARKs in health and disease, and draw attention to the largely unexplored area of MARK inhibitors for AD.

  17. High affinity calmodulin target sequence in the signalling molecule PI 3-kinase

    DEFF Research Database (Denmark)

    Fischer, R; Julsgart, J; Berchtold, M W

    1998-01-01

    M-binding peptide derived from the p110gamma isoform interacts with CaM in a calcium-dependent way. Using gel shift analysis and fluorescence spectrophotometry we discovered that the peptide forms a high affinity complex with CaM. Titration experiments using dansylated CaM gave an affinity constant of 5 n...

  18. Relative binding affinity of carboxylate-, phosphonate-, and bisphosphonate-functionalized gold nanoparticles targeted to damaged bone tissue

    Energy Technology Data Exchange (ETDEWEB)

    Ross, Ryan D. [Rush University Medical Center, Department of Anatomy and Cell Biology (United States); Cole, Lisa E.; Roeder, Ryan K., E-mail: rroeder@nd.edu [University of Notre Dame, Department of Aerospace and Mechanical Engineering Bioengineering Graduate Program (United States)

    2012-10-15

    Functionalized Au NPs have received considerable recent interest for targeting and labeling cells and tissues. Damaged bone tissue can be targeted by functionalizing Au NPs with molecules exhibiting affinity for calcium. Therefore, the relative binding affinity of Au NPs surface functionalized with either carboxylate (l-glutamic acid), phosphonate (2-aminoethylphosphonic acid), or bisphosphonate (alendronate) was investigated for targeted labeling of damaged bone tissue in vitro. Targeted labeling of damaged bone tissue was qualitatively verified by visual observation and backscattered electron microscopy, and quantitatively measured by the surface density of Au NPs using field-emission scanning electron microscopy. The surface density of functionalized Au NPs was significantly greater within damaged tissue compared to undamaged tissue for each functional group. Bisphosphonate-functionalized Au NPs exhibited a greater surface density labeling damaged tissue compared to glutamic acid- and phosphonic acid-functionalized Au NPs, which was consistent with the results of previous work comparing the binding affinity of the same functionalized Au NPs to synthetic hydroxyapatite crystals. Targeted labeling was enabled not only by the functional groups but also by the colloidal stability in solution. Functionalized Au NPs were stabilized by the presence of the functional groups, and were shown to remain well dispersed in ionic (phosphate buffered saline) and serum (fetal bovine serum) solutions for up to 1 week. Therefore, the results of this study suggest that bisphosphonate-functionalized Au NPs have potential for targeted delivery to damaged bone tissue in vitro and provide motivation for in vivo investigation.

  19. Maximizing in vivo target clearance by design of pH-dependent target binding antibodies with altered affinity to FcRn.

    Science.gov (United States)

    Yang, Danlin; Giragossian, Craig; Castellano, Steven; Lasaro, Marcio; Xiao, Haiguang; Saraf, Himanshu; Hess Kenny, Cynthia; Rybina, Irina; Huang, Zhong-Fu; Ahlberg, Jennifer; Bigwarfe, Tammy; Myzithras, Maria; Waltz, Erica; Roberts, Simon; Kroe-Barrett, Rachel; Singh, Sanjaya

    2017-10-01

    Antibodies with pH-dependent binding to both target antigens and neonatal Fc receptor (FcRn) provide an alternative tool to conventional neutralizing antibodies, particularly for therapies where reduction in antigen level is challenging due to high target burden. However, the requirements for optimal binding kinetic framework and extent of pH dependence for these antibodies to maximize target clearance from circulation are not well understood. We have identified a series of naturally-occurring high affinity antibodies with pH-dependent target binding properties. By in vivo studies in cynomolgus monkeys, we show that pH-dependent binding to the target alone is not sufficient for effective target removal from circulation, but requires Fc mutations that increase antibody binding to FcRn. Affinity-enhanced pH-dependent FcRn binding that is double-digit nM at pH 7.4 and single-digit nM at pH 6 achieved maximal target reduction when combined with similar target binding affinities in reverse pH directions. Sustained target clearance below the baseline level was achieved 3 weeks after single-dose administration at 1.5 mg/kg. Using the experimentally derived mechanistic model, we demonstrate the essential kinetic interplay between target turnover and antibody pH-dependent binding during the FcRn recycling, and identify the key components for achieving maximal target clearance. These results bridge the demand for improved patient dosing convenience with the "know-how" of therapeutic modality by design.

  20. Viral Escape Mutant Epitope Maintains TCR Affinity for Antigen yet Curtails CD8 T Cell Responses.

    Directory of Open Access Journals (Sweden)

    Shayla K Shorter

    Full Text Available T cells have the remarkable ability to recognize antigen with great specificity and in turn mount an appropriate and robust immune response. Critical to this process is the initial T cell antigen recognition and subsequent signal transduction events. This antigen recognition can be modulated at the site of TCR interaction with peptide:major histocompatibility (pMHC or peptide interaction with the MHC molecule. Both events could have a range of effects on T cell fate. Though responses to antigens that bind sub-optimally to TCR, known as altered peptide ligands (APL, have been studied extensively, the impact of disrupting antigen binding to MHC has been highlighted to a lesser extent and is usually considered to result in complete loss of epitope recognition. Here we present a model of viral evasion from CD8 T cell immuno-surveillance by a lymphocytic choriomeningitis virus (LCMV escape mutant with an epitope for which TCR affinity for pMHC remains high but where the antigenic peptide binds sub optimally to MHC. Despite high TCR affinity for variant epitope, levels of interferon regulatory factor-4 (IRF4 are not sustained in response to the variant indicating differences in perceived TCR signal strength. The CD8+ T cell response to the variant epitope is characterized by early proliferation and up-regulation of activation markers. Interestingly, this response is not maintained and is characterized by a lack in IL-2 and IFNγ production, increased apoptosis and an abrogated glycolytic response. We show that disrupting the stability of peptide in MHC can effectively disrupt TCR signal strength despite unchanged affinity for TCR and can significantly impact the CD8+ T cell response to a viral escape mutant.

  1. Shark immunity bites back: affinity maturation and memory response in the nurse shark, Ginglymostoma cirratum.

    Science.gov (United States)

    Dooley, Helen; Flajnik, Martin F

    2005-03-01

    The cartilaginous fish are the oldest phylogenetic group in which all of the molecular components of the adaptive immune system have been found. Although early studies clearly showed that sharks could produce an IgM-based response following immunization, evidence for memory, affinity maturation and roles for the other isotypes (notably IgNAR) in this group remained inconclusive. The data presented here illustrate that the nurse shark (Ginglymostoma cirratum) is able to produce not only an IgM response, but we also show for the first time a highly antigen-specific IgNAR response. Additionally, under appropriate conditions, a memory response for both isotypes can be elicited. Analysis of the response shows differential expression of pentameric and monomeric IgM. Pentameric IgM provides the 'first line of defense' through high-avidity, low-affinity interaction with antigen. In contrast, monomeric IgM and IgNAR seem responsible for the specific, antigen-driven response. We propose the presence of distinct lineages of B cells in sharks. As there is no conventional isotype switching, each lineage seems pre-determined to express a single isotype (IgM versus IgNAR). However, our data suggest that there may also be specific lineages for the different forms (pentameric versus monomeric) of the IgM isotype.

  2. Locked nucleic acid (LNA): High affinity targeting of RNA for diagnostics and therapeutics

    DEFF Research Database (Denmark)

    Kauppinen, S.; Vester, Birte; Wengel, Jesper

    2005-01-01

    Locked nucleic acid (LNA) is a nucleic acid analogue containing one or more LNA nucleotide monomers with a bicyclic furanose unit locked in an RNA mimicking sugar conformation. This conformational restriction results in unprecedented hybridization affinity towards complementary single stranded RN...

  3. Identification of Thioredoxin Disulfide Targets Using a Quantitative Proteomics Approach Based on Isotope-Coded Affinity Tags

    DEFF Research Database (Denmark)

    Hägglund, Per; Bunkenborg, Jakob; Maeda, Kenji

    2008-01-01

    Thioredoxin (Trx) is a ubiquitous protein disulfide reductase involved in a wide range of cellular redox processes. A large number of putative target proteins have been identified using proteomics approaches, but insight into target specificity at the molecular level is lacking since the reactivity...... of Trx toward individual disulfides has not been quantified. Here, a novel proteomics procedure is described for quantification of Trx-mediated target disulfide reduction based on thiol-specific differential labeling with the iodoacetamide-based isotope-coded affinity tag (ICAT) reagents. Briefly......, protein extract of embryos from germinated barley seeds was treated +/- Trx, and thiols released from target protein disulfides were irreversibly blocked with iodoacetamide. The remaining cysteine residues in the Trx-treated and the control (-Trx) samples were then chemically reduced and labeled...

  4. In Vitro and In Vivo Evaluations of A High Affinity and Specificity Photoacoustic Nanoparticle Targeting to Cancer

    DEFF Research Database (Denmark)

    Ma, Lixin; Xu, Hang; Lee, Li Ean

    oxide (SIO) nanoparticle as a potent cancer cell selective PA contrast agent, with a high binding affinity and selectivity to the gastrin releasing peptide receptor (GRPR) which is overexpressed in many human cancers including prostate cancer, breast cancer and small cell lung cancer etc.......Photoacoustic (PA) imaging uses a short-pulsed laser to create ultrasound signals for the high resolution acoustic imaging. The development of targeting PA contrast agents offers a unique opportunity to improve the early detection of cancer cells. This work aims to develop a silica coated iron...

  5. Affinity-Mediated Homogeneous Electrochemical Aptasensor on a Graphene Platform for Ultrasensitive Biomolecule Detection via Exonuclease-Assisted Target-Analog Recycling Amplification.

    Science.gov (United States)

    Ge, Lei; Wang, Wenxiao; Sun, Ximei; Hou, Ting; Li, Feng

    2016-02-16

    As is well-known, graphene shows a remarkable difference in affinity toward nonstructured single-stranded (ss) DNA and double-stranded (ds) DNA. This property makes it popular to prepare DNA-based optical sensors. In this work, taking this unique property of graphene in combination with the sensitive electrochemical transducer, we report a novel affinity-mediated homogeneous electrochemical aptasensor using graphene modified glassy carbon electrode (GCE) as the sensing platform. In this approach, the specific aptamer-target recognition is converted into an ultrasensitive electrochemical signal output with the aid of a novel T7 exonuclease (T7Exo)-assisted target-analog recycling amplification strategy, in which the ingeniously designed methylene blue (MB)-labeled hairpin DNA reporters are digested in the presence of target and, then, converted to numerous MB-labeled long ssDNAs. The distinct difference in differential pulse voltammetry response between the designed hairpin reporters and the generated long ssDNAs on the graphene/GCE allows ultrasensitive detection of target biomolecules. Herein, the design and working principle of this homogeneous electrochemical aptasensor were elucidated, and the working conditions were optimized. The gel electrophoresis results further demonstrate that the designed T7Exo-assisted target-analog recycling amplification strategy can work well. This electrochemical aptasensor realizes the detection of biomolecule in a homogeneous solution without immobilization of any bioprobe on electrode surface. Moreover, this versatile homogeneous electrochemical sensing system was used for the determination of biomolecules in real serum samples with satisfying results.

  6. High affinity γPNA sandwich hybridization assay for rapid detection of short nucleic acid targets with single mismatch discrimination.

    Science.gov (United States)

    Goldman, Johnathan M; Zhang, Li Ang; Manna, Arunava; Armitage, Bruce A; Ly, Danith H; Schneider, James W

    2013-07-08

    Hybridization analysis of short DNA and RNA targets presents many challenges for detection. The commonly employed sandwich hybridization approach cannot be implemented for these short targets due to insufficient probe-target binding strengths for unmodified DNA probes. Here, we present a method capable of rapid and stable sandwich hybridization detection for 22 nucleotide DNA and RNA targets. Stable hybridization is achieved using an n-alkylated, polyethylene glycol γ-carbon modified peptide nucleic acid (γPNA) amphiphile. The γPNA's exceptionally high affinity enables stable hybridization of a second DNA-based probe to the remaining bases of the short target. Upon hybridization of both probes, an electrophoretic mobility shift is measured via interaction of the n-alkane modification on the γPNA with capillary electrophoresis running buffer containing nonionic surfactant micelles. We find that sandwich hybridization of both probes is stable under multiple binding configurations and demonstrate single base mismatch discrimination. The binding strength of both probes is also stabilized via coaxial stacking on adjacent hybridization to targets. We conclude with a discussion on the implementation of the proposed sandwich hybridization assay as a high-throughput microRNA detection method.

  7. Spectral response variation of a negative-electron-affinity photocathode in the preparation process

    International Nuclear Information System (INIS)

    Liu Lei; Du Yujie; Chang Benkang; Yunsheng Qian

    2006-01-01

    In order to research the spectral response variation of a negative electron affinity (NEA) photocathode in the preparation process, we have done two experiments on a transmission-type GaAs photocathode.First, an automatic spectral response recording system is described, which is used to take spectral response curves during the activation procedure of the photocathode. By this system, the spectral response curves of a GaAs:Cs-Ophotocathode measured in situ are presented. Then, after the cathode is sealed with a microchannel plate and a fluorescence screen into the image tube, we measure the spectral response of the tube by another measurement instrument. By way of comparing and analyzing these curves, we can find the typical variation in spectral-responses.The reasons for the variation are discussed. Based on these curves, spectral matching factors of a GaAs cathode for green vegetation and rough concrete are calculated. The visual ranges of night-vision goggles under specific circumstances are estimated. The results show that the spectral response of the NEA photocathode degraded in the sealing process, especially at long wavelengths. The variation has also influenced the whole performance of the intensifier tube

  8. Affine-response model of molecular solvation of ions: Accurate predictions of asymmetric charging free energies.

    Science.gov (United States)

    Bardhan, Jaydeep P; Jungwirth, Pavel; Makowski, Lee

    2012-09-28

    Two mechanisms have been proposed to drive asymmetric solvent response to a solute charge: a static potential contribution similar to the liquid-vapor potential, and a steric contribution associated with a water molecule's structure and charge distribution. In this work, we use free-energy perturbation molecular-dynamics calculations in explicit water to show that these mechanisms act in complementary regimes; the large static potential (∼44 kJ/mol/e) dominates asymmetric response for deeply buried charges, and the steric contribution dominates for charges near the solute-solvent interface. Therefore, both mechanisms must be included in order to fully account for asymmetric solvation in general. Our calculations suggest that the steric contribution leads to a remarkable deviation from the popular "linear response" model in which the reaction potential changes linearly as a function of charge. In fact, the potential varies in a piecewise-linear fashion, i.e., with different proportionality constants depending on the sign of the charge. This discrepancy is significant even when the charge is completely buried, and holds for solutes larger than single atoms. Together, these mechanisms suggest that implicit-solvent models can be improved using a combination of affine response (an offset due to the static potential) and piecewise-linear response (due to the steric contribution).

  9. Affine-response model of molecular solvation of ions: Accurate predictions of asymmetric charging free energies

    Science.gov (United States)

    Bardhan, Jaydeep P.; Jungwirth, Pavel; Makowski, Lee

    2012-01-01

    Two mechanisms have been proposed to drive asymmetric solvent response to a solute charge: a static potential contribution similar to the liquid-vapor potential, and a steric contribution associated with a water molecule's structure and charge distribution. In this work, we use free-energy perturbation molecular-dynamics calculations in explicit water to show that these mechanisms act in complementary regimes; the large static potential (∼44 kJ/mol/e) dominates asymmetric response for deeply buried charges, and the steric contribution dominates for charges near the solute-solvent interface. Therefore, both mechanisms must be included in order to fully account for asymmetric solvation in general. Our calculations suggest that the steric contribution leads to a remarkable deviation from the popular “linear response” model in which the reaction potential changes linearly as a function of charge. In fact, the potential varies in a piecewise-linear fashion, i.e., with different proportionality constants depending on the sign of the charge. This discrepancy is significant even when the charge is completely buried, and holds for solutes larger than single atoms. Together, these mechanisms suggest that implicit-solvent models can be improved using a combination of affine response (an offset due to the static potential) and piecewise-linear response (due to the steric contribution). PMID:23020318

  10. High-affinity triplex targeting of double stranded DNA using chemically modified peptide nucleic acid oligomers

    DEFF Research Database (Denmark)

    Hansen, Mads E; Bentin, Thomas; Nielsen, Peter E

    2009-01-01

    While sequence-selective dsDNA targeting by triplex forming oligonucleotides has been studied extensively, only very little is known about the properties of PNA-dsDNA triplexes-mainly due to the competing invasion process. Here we show that when appropriately modified using pseudoisocytosine subs...

  11. Target Response Adaptation for Correlation Filter Tracking

    KAUST Repository

    Bibi, Adel Aamer; Mueller, Matthias; Ghanem, Bernard

    2016-01-01

    Most correlation filter (CF) based trackers utilize the circulant structure of the training data to learn a linear filter that best regresses this data to a hand-crafted target response. These circularly shifted patches are only approximations

  12. Melanoma affinity in mice and immunosuppressed sheep of [125I]N-(4-dipropylaminobutyl)-4-iodobenzamide, a new targeting agent

    International Nuclear Information System (INIS)

    Labarre, Pierre; Papon, Janine; Rose, Alison H.; Guerquin-Kern, Jean-Luc; Morandeau, Laurence; Wu, Ting-di; Moreau, Marie-France; Bayle, Martine; Chezal, Jean-Michel; Croisy, Alain; Madelmont, Jean-Claude; Turner, Harvey; Moins, Nicole

    2008-01-01

    The increasing incidence of melanoma and the lack of effective therapy have prompted the development of new vectors, more specific to the pigmented tumor, for early detection and treatment. Targeted agents have to exhibit a rapid, high tumor uptake, long tumor retention and rapid clearance from nontarget organs. This joint work presents results obtained with a new melanoma targeting agent, [ 125 I]-N-(4-dipropylaminobutyl)-4-iodobenzamide or [ 125 I]BZ18. After labeling with a high specific activity, the biodistribution of the compound was investigated in two animal models, the mouse and the sheep. Melanotic tumor retention was observed lasting several days. We visualized the internalization of the agent inside the melanosomes by secondary ion mass spectroscopy imaging, we measured the affinity constants of [ 125 I]BZ18 on a synthetic melanin model and we demonstrated a radiotoxic effect of this labeled agent on B16F0 melanoma cell culture due to its cellular internalization. From this work, [ 125 I]BZ18 appeared a promising melanoma targeting agent in the nuclear medicine field

  13. Change in Allosteric Network Affects Binding Affinities of PDZ Domains: Analysis through Perturbation Response Scanning

    Science.gov (United States)

    Gerek, Z. Nevin; Ozkan, S. Banu

    2011-01-01

    The allosteric mechanism plays a key role in cellular functions of several PDZ domain proteins (PDZs) and is directly linked to pharmaceutical applications; however, it is a challenge to elaborate the nature and extent of these allosteric interactions. One solution to this problem is to explore the dynamics of PDZs, which may provide insights about how intramolecular communication occurs within a single domain. Here, we develop an advancement of perturbation response scanning (PRS) that couples elastic network models with linear response theory (LRT) to predict key residues in allosteric transitions of the two most studied PDZs (PSD-95 PDZ3 domain and hPTP1E PDZ2 domain). With PRS, we first identify the residues that give the highest mean square fluctuation response upon perturbing the binding sites. Strikingly, we observe that the residues with the highest mean square fluctuation response agree with experimentally determined residues involved in allosteric transitions. Second, we construct the allosteric pathways by linking the residues giving the same directional response upon perturbation of the binding sites. The predicted intramolecular communication pathways reveal that PSD-95 and hPTP1E have different pathways through the dynamic coupling of different residue pairs. Moreover, our analysis provides a molecular understanding of experimentally observed hidden allostery of PSD-95. We show that removing the distal third alpha helix from the binding site alters the allosteric pathway and decreases the binding affinity. Overall, these results indicate that (i) dynamics plays a key role in allosteric regulations of PDZs, (ii) the local changes in the residue interactions can lead to significant changes in the dynamics of allosteric regulations, and (iii) this might be the mechanism that each PDZ uses to tailor their binding specificities regulation. PMID:21998559

  14. Use of thermodynamic coupling between antibody-antigen binding and phospholipid acyl chain phase transition energetics to predict immunoliposome targeting affinity.

    Science.gov (United States)

    Klegerman, Melvin E; Zou, Yuejiao; Golunski, Eva; Peng, Tao; Huang, Shao-Ling; McPherson, David D

    2014-09-01

    Thermodynamic analysis of ligand-target binding has been a useful tool for dissecting the nature of the binding mechanism and, therefore, potentially can provide valuable information regarding the utility of targeted formulations. Based on a consistent coupling of antibody-antigen binding and gel-liquid crystal transition energetics observed for antibody-phosphatidylethanolamine (Ab-PE) conjugates, we hypothesized that the thermodynamic parameters and the affinity for antigen of the Ab-PE conjugates could be effectively predicted once the corresponding information for the unconjugated antibody is determined. This hypothesis has now been tested in nine different antibody-targeted echogenic liposome (ELIP) preparations, where antibody is conjugated to dipalmitoylphosphatidylethanolamine (DPPE) head groups through a thioether linkage. Predictions were satisfactory (affinity not significantly different from the population of values found) in five cases (55.6%), but the affinity of the unconjugated antibody was not significantly different from the population of values found in six cases (66.7%), indicating that the affinities of the conjugated antibody tended not to deviate appreciably from those of the free antibody. While knowledge of the affinities of free antibodies may be sufficient to judge their suitability as targeting agents, thermodynamic analysis may still provide valuable information regarding their usefulness for specific applications.

  15. Target Response Adaptation for Correlation Filter Tracking

    KAUST Repository

    Bibi, Adel Aamer

    2016-09-16

    Most correlation filter (CF) based trackers utilize the circulant structure of the training data to learn a linear filter that best regresses this data to a hand-crafted target response. These circularly shifted patches are only approximations to actual translations in the image, which become unreliable in many realistic tracking scenarios including fast motion, occlusion, etc. In these cases, the traditional use of a single centered Gaussian as the target response impedes tracker performance and can lead to unrecoverable drift. To circumvent this major drawback, we propose a generic framework that can adaptively change the target response from frame to frame, so that the tracker is less sensitive to the cases where circular shifts do not reliably approximate translations. To do that, we reformulate the underlying optimization to solve for both the filter and target response jointly, where the latter is regularized by measurements made using actual translations. This joint problem has a closed form solution and thus allows for multiple templates, kernels, and multi-dimensional features. Extensive experiments on the popular OTB100 benchmark show that our target adaptive framework can be combined with many CF trackers to realize significant overall performance improvement (ranging from 3 %-13.5% in precision and 3.2 %-13% in accuracy), especially in categories where this adaptation is necessary (e.g. fast motion, motion blur, etc.). © Springer International Publishing AG 2016.

  16. Oxygen affinity and Bohr effect responses to 2,3-diphosphoglycerate in equine and human blood.

    Science.gov (United States)

    diBella, G; Scandariato, G; Suriano, O; Rizzo, A

    1996-05-01

    The dependence of blood oxygen affinity and the Bohr effect on the concentration of 2,3-diphosphoglycerate (DPG) in erythrocytes was investigated in 24 trotter horses and 24 healthy men. The oxygen tension at half saturation and standard conditions (P50st at pH 7.4, PCO2(40) mmHg and 37 degrees C) and the carbon dioxide or fixed-acid-induced Bohr effect (dlogP50/dpH) were determined. Samples of fresh blood and blood depleted of or enriched with DPG were studied. In the absence of measurable DPG, the equine and human blood had similar mean (SD) values of P50st (16.6 [0.6] and 16.2 [0.7] mmHg, respectively). In both species these values increased with increasing DPG, but the response of equine blood was significantly lower, at least up to physiological values (P50st = 24.6 [0.6] and 26.2 [0.7]) mmHg; DPG = 14([1.8] and 12.8 [1.2] mumol gHb-1, respectively, in fresh blood). For concentrations above 20 to 25 mumol gHb-1 of DPG the difference between the values of P50st in the two species tended to decrease because the response in human blood reached a plateau. The interactions between the Bohr effect and the concentration of DPG showed that in the horses, as in the men, the level of DPG played an important role in governing the relative magnitude of carbon dioxide and fixed acid factors. The difference between them, which is associated with the oxylabile carbamino binding, was greatest in DPG-depleted blood, but whereas in the men the difference was suppressed by an above normal DPG concentration, in the horses it was still measurable.

  17. Development of a nonlinear model for the prediction of response times of glucose affinity sensors using concanavalin A and dextran and the development of a differential osmotic glucose affinity sensor

    Science.gov (United States)

    Reis, Louis G.

    With the increasing prevalence of diabetes in the United States and worldwide, blood glucose monitoring must be accurate and reliable. Current enzymatic sensors have numerous disadvantages that make them unreliable and unfavorable among patients. Recent research in glucose affinity sensors correct some of the problems that enzymatic sensors experience. Dextran and concanavalin A are two of the more common components used in glucose affinity sensors. When these sensors were first explored, a model was derived to predict the response time of a glucose affinity sensor using concanavalin A and dextran. However, the model assumed the system was linear and fell short of calculating times representative of the response times determined through experimental tests with the sensors. In this work, a new model that uses the Stokes-Einstein Equation to demonstrate the nonlinear behavior of the glucose affinity assay was developed to predict the response times of similar glucose affinity sensors. In addition to the device tested by the original linear model, additional devices were identified and tested with the proposed model. The nonlinear model was designed to accommodate the many different variations between systems. The proposed model was able to accurately calculate response times for sensors using the concanavalin A-dextran affinity assay with respect to the experimentally reported times by the independent research groups. Parameter studies using the nonlinear model were able to identify possible setbacks that could compromise the response of thesystem. Specifically, the model showed that the improper use of asymmetrical membranes could increase the response time by as little as 20% or more as the device is miniaturized. The model also demonstrated that systems using the concanavalin Adextran assay would experience higher response times in the hypoglycemic range. This work attempted to replicate and improve an osmotic glucose affinity sensor. The system was designed to

  18. Molecularly Imprinted Polymers with Stimuli-Responsive Affinity: Progress and Perspectives

    OpenAIRE

    Wei Chen; Yue Ma; Jianmin Pan; Zihui Meng; Guoqing Pan; Börje Sellergren

    2015-01-01

    Intelligent stimuli-responsive molecularly imprinted polymers (SR-MIPs) have attracted considerable research interest in recent years due to the potential applications in drug delivery, biotechnology and separation sciences. This review comprehensively summarizes various SR-MIPs, including the design and applications of thermo-responsive MIPs, pH-responsive MIPs, photo-responsive MIPs, biomolecule-responsive MIPs and ion-responsive MIPs. Besides the development of current SR-MIPs, the advanta...

  19. Smart multifunctional nanoagents for in situ monitoring of small molecules with a switchable affinity towards biomedical targets

    Science.gov (United States)

    Shevchenko, Konstantin G.; Cherkasov, Vladimir R.; Nikitina, Irina L.; Babenyshev, Andrey V.; Nikitin, Maxim P.

    2018-02-01

    The great diversity of nanomaterials provides ample opportunities for constructing effective agents for biomedical applications ranging from biosensing to drug delivery. Multifunctional nanoagents that combine several features in a single particle are of special interest due to capabilities that substantially exceed those of molecular drugs. An ideal theranostic agent should simultaneously be an advanced biosensor to identify a disease and report the diagnosis and a biomedical actuator to treat the disease. While many approaches were developed to load a nanoparticle with various drugs for actuation of the diseased cells (e.g., to kill them), the nanoparticle-based approaches for the localized biosensing with real-time reporting of the marker concentration severely lag behind. Here, we show a smart in situ nanoparticle-based biosensor/actuator system that dynamically and reversibly changes its structural and optical properties in response to a small molecule marker to allow real-time monitoring of the marker concentration and adjustment of the system ability to bind its biomedical target. Using the synergistic combination of signal readout based on the localized surface plasmon resonance and an original method of fabrication of smart ON/OFF-switchable nanoagents, we demonstrate reversible responsiveness of the system to a model small molecule marker (antibiotic chloramphenicol) in a wide concentration range. The proposed approach can be used for the development of advanced multifunctional nanoagents for theranostic applications.

  20. Insights from the Fungus Fusarium oxysporum Point to High Affinity Glucose Transporters as Targets for Enhancing Ethanol Production from Lignocellulose

    Science.gov (United States)

    Ali, Shahin S.; Nugent, Brian; Mullins, Ewen; Doohan, Fiona M.

    2013-01-01

    Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt) from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km(glucose) was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing. PMID:23382943

  1. Insights from the fungus Fusarium oxysporum point to high affinity glucose transporters as targets for enhancing ethanol production from lignocellulose.

    Directory of Open Access Journals (Sweden)

    Shahin S Ali

    Full Text Available Ethanol is the most-widely used biofuel in the world today. Lignocellulosic plant biomass derived from agricultural residue can be converted to ethanol via microbial bioprocessing. Fungi such as Fusarium oxysporum can simultaneously saccharify straw to sugars and ferment sugars to ethanol. But there are many bottlenecks that need to be overcome to increase the efficacy of microbial production of ethanol from straw, not least enhancement of the rate of fermentation of both hexose and pentose sugars. This research tested the hypothesis that the rate of sugar uptake by F. oxysporum would enhance the ethanol yields from lignocellulosic straw and that high affinity glucose transporters can enhance ethanol yields from this substrate. We characterized a novel hexose transporter (Hxt from this fungus. The F. oxysporum Hxt represents a novel transporter with homology to yeast glucose signaling/transporter proteins Rgt2 and Snf3, but it lacks their C-terminal domain which is necessary for glucose signalling. Its expression level decreased with increasing glucose concentration in the medium and in a glucose uptake study the Km((glucose was 0.9 mM, which indicated that the protein is a high affinity glucose transporter. Post-translational gene silencing or over expression of the Hxt in F. oxysporum directly affected the glucose and xylose transport capacity and ethanol yielded by F. oxysporum from straw, glucose and xylose. Thus we conclude that this Hxt has the capacity to transport both C5 and C6 sugars and to enhance ethanol yields from lignocellulosic material. This study has confirmed that high affinity glucose transporters are ideal candidates for improving ethanol yields from lignocellulose because their activity and level of expression is high in low glucose concentrations, which is very common during the process of consolidated processing.

  2. Generation of high-affinity, internalizing anti-FGFR2 single-chain variable antibody fragment fused with Fc for targeting gastrointestinal cancers.

    Science.gov (United States)

    Borek, Aleksandra; Sokolowska-Wedzina, Aleksandra; Chodaczek, Grzegorz; Otlewski, Jacek

    2018-01-01

    Fibroblast growth factor receptors (FGFRs) are promising targets for antibody-based cancer therapies, as their substantial overexpression has been found in various tumor cells. Aberrant activation of FGF receptor 2 (FGFR2) signaling through overexpression of FGFR2 and/or its ligands, mutations, or receptor amplification has been reported in multiple cancer types, including gastric, colorectal, endometrial, ovarian, breast and lung cancer. In this paper, we describe application of the phage display technology to produce a panel of high affinity single chain variable antibody fragments (scFvs) against the extracellular ligand-binding domain of FGFR2 (ECD_FGFR2). The binders were selected from the human single chain variable fragment scFv phage display libraries Tomlinson I + J and showed high specificity and binding affinity towards human FGFR2 with nanomolar KD values. To improve the affinity of the best binder selected, scFvF7, we reformatted it to a bivalent diabody format, or fused it with the Fc region (scFvF7-Fc). The scFvF7-Fc antibody construct presented the highest affinity for FGFR2, with a KD of 0.76 nM, and was selectively internalized into cancer cells overexpressing FGFR2, Snu-16 and NCI-H716. Finally, we prepared a conjugate of scFvF7-Fc with the cytotoxic drug monomethyl-auristatin E (MMAE) and evaluated its cytotoxicity. The conjugate delivered MMAE selectively to FGFR2-positive tumor cells. These results indicate that scFvF7-Fc-vcMMAE is a highly potent molecule for the treatment of cancers with FGFR2 overexpression.

  3. Responses and mechanisms of positive electron affinity molecules in the N2 mode of the thermionic ionization detector and the electron-capture detector

    International Nuclear Information System (INIS)

    Jones, C.S.

    1989-01-01

    Very little knowledge has been acquired in the past on the mechanistic pathway by which molecules respond in the N 2 mode of the thermionic ionization detector. An attempt is made here to elucidate the response mechanism of the detector. The basic response mechanisms are known for the electron capture detector, and an attempt is made to identify the certain mechanism by which selected molecules respond. The resonance electron capture rate constant has been believed to be temperature independent, and investigations of the temperature dependence of electron capture responses are presented. Mechanisms for the N 2 mode of the thermionic ionization detector have been proposed by examining the detector response to positive electron affinity molecules and by measurement of the ions produced by the detector. Electron capture mechanisms for selected molecules have been proposed by examining their temperature dependent responses in the electron capture detector and negative ion mass spectra of the samples. In studies of the resonance electron capture rate constant, the relative responses of selected positive electron affinity molecules and their temperature dependent responses were investigated. Positive electron affinity did not guarantee large responses in the N 2 mode thermionic ionization detector. High mass ions were measured following ionization of samples in the detector. Responses in the electron capture detector varied with temperature and electron affinity

  4. Direct binding of radioiodinated monoclonal antibody to tumor cells: significance of antibody purity and affinity for drug targeting or tumor imaging

    International Nuclear Information System (INIS)

    Kennel, S.J.; Foote, L.J.; Lankford, P.K.; Johnson, M.; Mitchell, T.; Braslawsky, G.R.

    1983-01-01

    For MoAb to be used efficiently for drug targeting and tumor imaging, the fraction of antibody binding to tumor cells must be maximized. The authors have studied the binding of 125 I MoAb in three different tumor systems. The fraction of antibody that could be bound to the cell surface was directly proportional to the antibody purity. The affinity constant also limits the fraction of antibody that can bind to cells at a given antigen concentration. Rearrangement of the standard expression for univalent equilibrium binding between two reactants shows that in antigen excess, the maximum fraction of antibody that can bind =Ka[Ag total]/1 + Ka[Ag total]. Binding data using four different MoAb with three cell systems confirm this relationship. Estimates for reasonable concentrations of tumor antigens in vivo indicate that antibodies with binding constants less than 10 8 M -1 are not likely to be useful for drug targeting or tumor imaging

  5. Lectin affinity electrophoresis.

    Science.gov (United States)

    Kobayashi, Yuka

    2014-01-01

    An interaction or a binding event typically changes the electrophoretic properties of a molecule. Affinity electrophoresis methods detect changes in the electrophoretic pattern of molecules (mainly macromolecules) that occur as a result of biospecific interactions or complex formation. Lectin affinity electrophoresis is a very effective method for the detection and analysis of trace amounts of glycobiological substances. It is particularly useful for isolating and separating the glycoisomers of target molecules. Here, we describe a sensitive technique for the detection of glycoproteins separated by agarose gel-lectin affinity electrophoresis that uses antibody-affinity blotting. The technique is tested using α-fetoprotein with lectin (Lens culinaris agglutinin and Phaseolus vulgaris agglutinin)-agarose gels.

  6. Consumer responses to ethnic targeted marketing

    NARCIS (Netherlands)

    A.-S.I.A. Lenoir (Anne-Sophie); S. Puntoni (Stefano)

    2014-01-01

    markdownabstract__Abstract__ Marketing is impacted more than ever by demographic change, to the extent that practitioners targeting ethnic groups should re-think their approach depending upon the strength with which different generations identify with their cultural heritage.

  7. Affine-response model of molecular solvation of ions: Accurate predictions of asymmetric charging free energies

    Czech Academy of Sciences Publication Activity Database

    Bardhan, J. P.; Jungwirth, Pavel; Makowski, L.

    Roč. 137, č. 12 ( 2012 ), 124101/1-124101/6 ISSN 0021-9606 R&D Projects: GA MŠk LH12001 Institutional research plan: CEZ:AV0Z40550506 Keywords : ion solvation * continuum models * linear response Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 3.164, year: 2012

  8. Preclinical evaluation of multistep targeting of diasialoganglioside GD2 using a IgG-scFv bispecific antibody with high affinity for GD2 and DOTA metal complex

    Science.gov (United States)

    Cheal, Sarah M.; Xu, Hong; Guo, Hong-fen; Zanzonico, Pat B.; Larson, Steven M.; Cheung, Nai-Kong

    2014-01-01

    Bispecific antibodies (BsAb) have proven to be useful targeting vectors for pretargeted radioimmunotherapy (PRIT). We sought to overcome key PRIT limitations such as high renal radiation exposure and immunogenicity (e.g. of streptavidin-antibody fusions), to advance clinical translation of this PRIT strategy for diasialoganglioside GD2-positive (GD2(+)) tumors. For this purpose, a IgG-scFv BsAb was engineered using the sequences for the anti-GD2 humanized monoclonal antibody hu3F8 (1) and C825, a murine scFv antibody with high affinity for the chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) complexed with beta-particle emitting radiometals such as 177Lu and 90Y (2, 3). A three-step regimen including hu3F8-C825, a dextran-based clearing agent, and p-aminobenzyl-DOTA radiolabeled with 177Lu (as 177Lu-DOTA-Bn; t1/2 = 6.71 days (d)) was optimized in immunocompromised mice carrying subcutaneous (s.c.) human GD2(+) neuroblastoma (NB) xenografts. Absorbed doses for tumor and normal tissues were ∼85 cGy/MBq and ≤3.7 cGy/MBq, respectively, with therapeutic indicies (TI) of 142 for blood and 23 for kidney. A therapy study (n = 5 per group; tumor volume: 240 ± 160 mm3) with three successive PRIT cycles (total 177Lu: ∼33 MBq; tumor dose ∼3400 cGy), revealed complete tumor response in 5/5 animals, with no recurrence up to 28 d post-treatment. Tumor ablation was confirmed histologically in 4/5 mice, and normal organs showed minimal overall toxicities. All non-treated mice required sacrifice within 12 d (>1.0 cm3 tumor volume). We conclude that this novel anti-GD2 PRIT approach has sufficient TI to successfully ablate s.c. GD2(+)–NB in mice while sparing kidney and bone marrow. PMID:24944121

  9. Blue light-induced LOV domain dimerization enhances the affinity of Aureochrome 1a for its target DNA sequence

    Science.gov (United States)

    Heintz, Udo; Schlichting, Ilme

    2016-01-01

    The design of synthetic optogenetic tools that allow precise spatiotemporal control of biological processes previously inaccessible to optogenetic control has developed rapidly over the last years. Rational design of such tools requires detailed knowledge of allosteric light signaling in natural photoreceptors. To understand allosteric communication between sensor and effector domains, characterization of all relevant signaling states is required. Here, we describe the mechanism of light-dependent DNA binding of the light-oxygen-voltage (LOV) transcription factor Aureochrome 1a from Phaeodactylum tricornutum (PtAu1a) and present crystal structures of a dark state LOV monomer and a fully light-adapted LOV dimer. In combination with hydrogen/deuterium-exchange, solution scattering data and DNA-binding experiments, our studies reveal a light-sensitive interaction between the LOV and basic region leucine zipper DNA-binding domain that together with LOV dimerization results in modulation of the DNA affinity of PtAu1a. We discuss the implications of these results for the design of synthetic LOV-based photosensors with application in optogenetics. DOI: http://dx.doi.org/10.7554/eLife.11860.001 PMID:26754770

  10. Manipulating the in vivo immune response by targeted gene knockdown.

    Science.gov (United States)

    Lieberman, Judy

    2015-08-01

    Aptamers, nucleic acids selected for high affinity binding to proteins, can be used to activate or antagonize immune mediators or receptors in a location and cell-type specific manner and to enhance antigen presentation. They can also be linked to other molecules (other aptamers, siRNAs or miRNAs, proteins, toxins) to produce multifunctional compounds for targeted immune modulation in vivo. Aptamer-siRNA chimeras (AsiCs) that induce efficient cell-specific knockdown in immune cells in vitro and in vivo can be used as an immunological research tool or potentially as an immunomodulating therapeutic. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. A Dualistic Conformational Response to Substrate Binding in the Human Serotonin Transporter Reveals a High Affinity State for Serotonin*

    Science.gov (United States)

    Bjerregaard, Henriette; Severinsen, Kasper; Said, Saida; Wiborg, Ove; Sinning, Steffen

    2015-01-01

    Serotonergic neurotransmission is modulated by the membrane-embedded serotonin transporter (SERT). SERT mediates the reuptake of serotonin into the presynaptic neurons. Conformational changes in SERT occur upon binding of ions and substrate and are crucial for translocation of serotonin across the membrane. Our understanding of these conformational changes is mainly based on crystal structures of a bacterial homolog in various conformations, derived homology models of eukaryotic neurotransmitter transporters, and substituted cysteine accessibility method of SERT. However, the dynamic changes that occur in the human SERT upon binding of ions, the translocation of substrate, and the role of cholesterol in this interplay are not fully elucidated. Here we show that serotonin induces a dualistic conformational response in SERT. We exploited the substituted cysteine scanning method under conditions that were sensitized to detect a more outward-facing conformation of SERT. We found a novel high affinity outward-facing conformational state of the human SERT induced by serotonin. The ionic requirements for this new conformational response to serotonin mirror the ionic requirements for translocation. Furthermore, we found that membrane cholesterol plays a role in the dualistic conformational response in SERT induced by serotonin. Our results indicate the existence of a subpopulation of SERT responding differently to serotonin binding than hitherto believed and that membrane cholesterol plays a role in this subpopulation of SERT. PMID:25614630

  12. Architecture of a minimal signaling pathway explains the T-cell response to a 1 million-fold variation in antigen affinity and dose

    Science.gov (United States)

    Lever, Melissa; Lim, Hong-Sheng; Kruger, Philipp; Nguyen, John; Trendel, Nicola; Abu-Shah, Enas; Maini, Philip Kumar; van der Merwe, Philip Anton

    2016-01-01

    T cells must respond differently to antigens of varying affinity presented at different doses. Previous attempts to map peptide MHC (pMHC) affinity onto T-cell responses have produced inconsistent patterns of responses, preventing formulations of canonical models of T-cell signaling. Here, a systematic analysis of T-cell responses to 1 million-fold variations in both pMHC affinity and dose produced bell-shaped dose–response curves and different optimal pMHC affinities at different pMHC doses. Using sequential model rejection/identification algorithms, we identified a unique, minimal model of cellular signaling incorporating kinetic proofreading with limited signaling coupled to an incoherent feed-forward loop (KPL-IFF) that reproduces these observations. We show that the KPL-IFF model correctly predicts the T-cell response to antigen copresentation. Our work offers a general approach for studying cellular signaling that does not require full details of biochemical pathways. PMID:27702900

  13. Online manual movement adjustments in response to target position changes and apparent target motion

    NARCIS (Netherlands)

    Oostwoud Wijdenes, L.; Brenner, E.; Smeets, J.B.J.

    2014-01-01

    This study set out to determine whether the fastest online hand movement corrections are only responses to changing judgments of the targets' position or whether they are also influenced by the apparent target motion. Introducing a gap between when a target disappears and when it reappears at a new

  14. Everyday stress response targets in the science of behavior change.

    Science.gov (United States)

    Smyth, Joshua M; Sliwinski, Martin J; Zawadzki, Matthew J; Scott, Stacey B; Conroy, David E; Lanza, Stephanie T; Marcusson-Clavertz, David; Kim, Jinhyuk; Stawski, Robert S; Stoney, Catherine M; Buxton, Orfeu M; Sciamanna, Christopher N; Green, Paige M; Almeida, David M

    2018-02-01

    Stress is an established risk factor for negative health outcomes, and responses to everyday stress can interfere with health behaviors such as exercise and sleep. In accordance with the Science of Behavior Change (SOBC) program, we apply an experimental medicine approach to identifying stress response targets, developing stress response assays, intervening upon these targets, and testing intervention effectiveness. We evaluate an ecologically valid, within-person approach to measuring the deleterious effects of everyday stress on physical activity and sleep patterns, examining multiple stress response components (i.e., stress reactivity, stress recovery, and stress pile-up) as indexed by two key response indicators (negative affect and perseverative cognition). Our everyday stress response assay thus measures multiple malleable stress response targets that putatively shape daily health behaviors (physical activity and sleep). We hypothesize that larger reactivity, incomplete recovery, and more frequent stress responses (pile-up) will negatively impact health behavior enactment in daily life. We will identify stress-related reactivity, recovery, and response in the indicators using coordinated analyses across multiple naturalistic studies. These results are the basis for developing a new stress assay and replicating the initial findings in a new sample. This approach will advance our understanding of how specific aspects of everyday stress responses influence health behaviors, and can be used to develop and test an innovative ambulatory intervention for stress reduction in daily life to enhance health behaviors. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Insulation and wiring specificity of BceR-like response regulators and their target promoters in Bacillus subtilis.

    Science.gov (United States)

    Fang, Chong; Nagy-Staroń, Anna; Grafe, Martin; Heermann, Ralf; Jung, Kirsten; Gebhard, Susanne; Mascher, Thorsten

    2017-04-01

    BceRS and PsdRS are paralogous two-component systems in Bacillus subtilis controlling the response to antimicrobial peptides. In the presence of extracellular bacitracin and nisin, respectively, the two response regulators (RRs) bind their target promoters, P bceA or P psdA , resulting in a strong up-regulation of target gene expression and ultimately antibiotic resistance. Despite high sequence similarity between the RRs BceR and PsdR and their known binding sites, no cross-regulation has been observed between them. We therefore investigated the specificity determinants of P bceA and P psdA that ensure the insulation of these two paralogous pathways at the RR-promoter interface. In vivo and in vitro analyses demonstrate that the regulatory regions within these two promoters contain three important elements: in addition to the known (main) binding site, we identified a linker region and a secondary binding site that are crucial for functionality. Initial binding to the high-affinity, low-specificity main binding site is a prerequisite for the subsequent highly specific binding of a second RR dimer to the low-affinity secondary binding site. In addition to this hierarchical cooperative binding, discrimination requires a competition of the two RRs for their respective binding site mediated by only slight differences in binding affinities. © 2016 John Wiley & Sons Ltd.

  16. Providing affinity

    DEFF Research Database (Denmark)

    Guglielmi, Michel; Johannesen, Hl

    2004-01-01

    , Essex, Hertfordshire, Norfolk and Suffolk. Research found that there was a lack of identity or sense of belonging and nothing anchoring people to the region as a whole. Common affinity is somehow forced to the people of East England and thereby we came to the conclusion that a single landmark...... and potential situations but also virtual events that calls for an undeterminated process of resolution. This process is activated by the user who co-produces the actualisation as an answer to a virtual reality that we defined at the first place. The potential situations or the possible it is a fantomatic real....... The possible is like the real. It is determinated and it only lakes existence. While the possible is already made, the virtual is like a problematic which needs to be resolved and actualized. Our installations are based on high tech interactivity where we use sensors and remote communication to offer a sense...

  17. Functional SNPs of INCENP Affect Semen Quality by Alternative Splicing Mode and Binding Affinity with the Target Bta-miR-378 in Chinese Holstein Bulls.

    Directory of Open Access Journals (Sweden)

    Juan Liu

    Full Text Available Inner centromere protein (INCENP plays an important role in mitosis and meiosis as the main member of chromosomal passenger protein complex (CPC. To investigate the functional markers of the INCENP gene associated with semen quality, the single nucleotide polymorphisms (SNPs g.19970 A>G and g.34078 T>G were identified and analyzed. The new splice variant INCENP-TV is characterized by the deletion of exon 12. The g.19970 A>G in the exonic splicing enhancer (ESE motif region results in an aberrant splice variant by constructing two minigene expression vectors using the pSPL3 exon capturing vector and transfecting vectors into MLTC-1 cells. INCENP-TV was more highly expressed than INCENP-reference in adult bull testes. The g.34078 T>G located in the binding region of bta-miR-378 could affect the expression of INCENP, which was verified by luciferase assay. To analyze comprehensively the correlation of SNPs with sperm quality, haplotype combinations constructed by g.19970 A>G and g.34078 T>G, as well as g.-692 C>T and g.-556 G>T reported in our previous studies, were analyzed. The bulls with H1H12 and H2H2 exhibited a higher ejaculate volume than those with H2H10 and H9H12, respectively (P G and g.34078 T>G in INCENP both of which appear to change the molecular and biological characteristics of the mRNA transcribed from the locus may serve as a biomarkers of male bovine fertility by affecting alternative splicing mode and binding affinity with the target bta-miR-378.

  18. Mapping Affinities in Academic Organizations

    Directory of Open Access Journals (Sweden)

    Dario Rodighiero

    2018-02-01

    Full Text Available Scholarly affinities are one of the most fundamental hidden dynamics that drive scientific development. Some affinities are actual, and consequently can be measured through classical academic metrics such as co-authoring. Other affinities are potential, and therefore do not leave visible traces in information systems; for instance, some peers may share interests without actually knowing it. This article illustrates the development of a map of affinities for academic collectives, designed to be relevant to three audiences: the management, the scholars themselves, and the external public. Our case study involves the School of Architecture, Civil and Environmental Engineering of EPFL, hereinafter ENAC. The school consists of around 1,000 scholars, 70 laboratories, and 3 institutes. The actual affinities are modeled using the data available from the information systems reporting publications, teaching, and advising scholars, whereas the potential affinities are addressed through text mining of the publications. The major challenge for designing such a map is to represent the multi-dimensionality and multi-scale nature of the information. The affinities are not limited to the computation of heterogeneous sources of information; they also apply at different scales. The map, thus, shows local affinities inside a given laboratory, as well as global affinities among laboratories. This article presents a graphical grammar to represent affinities. Its effectiveness is illustrated by two actualizations of the design proposal: an interactive online system in which the map can be parameterized, and a large-scale carpet of 250 square meters. In both cases, we discuss how the materiality influences the representation of data, in particular the way key questions could be appropriately addressed considering the three target audiences: the insights gained by the management and their consequences in terms of governance, the understanding of the scholars’ own

  19. Radiation responses of stem cells: targeted and non-targeted effects

    International Nuclear Information System (INIS)

    Kavanagh, J.N.; Waring, E.J.; Prise, K.M.

    2015-01-01

    Stem cells are fundamental to the development of any tissue or organism via their ability to self-renew, which is aided by their unlimited proliferative capacity and their ability to produce fully differentiated offspring, often from multiple lineages. Stems cells are long lived and have the potential to accumulate mutations, including in response to radiation exposure. It is thought that stem cells have the potential to be induced into a cancer stem cell phenotype and that these may play an important role in resistance to radiotherapy. For radiation-induced carcinogenesis, the role of targeted and non-targeted effects is unclear with tissue or origin being important. Studies of genomic instability and bystander responses have shown consistent effects in haematopoietic models. Several models of radiation have predicted that stem cells play an important role in tumour initiation and that bystander responses could play a role in proliferation and self-renewal. (authors)

  20. Report: Affinity Chromatography.

    Science.gov (United States)

    Walters, Rodney R.

    1985-01-01

    Supports, affinity ligands, immobilization, elution methods, and a number of applications are among the topics considered in this discussion of affinity chromatography. An outline of the basic principles of affinity chromatography is included. (JN)

  1. 2017 Guralp Affinity Digitizer Evaluation.

    Energy Technology Data Exchange (ETDEWEB)

    Merchant, Bion J.

    2018-03-01

    Sandia National Laboratories has tested and evaluated two Guralp Affinity digitizers. The Affinity digitizers are intended to record sensor output for seismic and infrasound monitoring applications. The purpose of this digitizer evaluation is to measure the performance characteristics in such areas as power consumption, input impedance, sensitivity, full scale, self- noise, dynamic range, system noise, response, passband, and timing. The Affinity digitizers are being evaluated for potential use in the International Monitoring System (IMS) of the Comprehensive Nuclear Test-Ban-Treaty Organization (CTBTO).

  2. Anti-HIV-1 B cell responses are dependent on B cell precursor frequency and antigen-binding affinity.

    Science.gov (United States)

    Dosenovic, Pia; Kara, Ervin E; Pettersson, Anna-Klara; McGuire, Andrew T; Gray, Matthew; Hartweger, Harald; Thientosapol, Eddy S; Stamatatos, Leonidas; Nussenzweig, Michel C

    2018-04-16

    The discovery that humans can produce potent broadly neutralizing antibodies (bNAbs) to several different epitopes on the HIV-1 spike has reinvigorated efforts to develop an antibody-based HIV-1 vaccine. Antibody cloning from single cells revealed that nearly all bNAbs show unusual features that could help explain why it has not been possible to elicit them by traditional vaccination and instead would require a sequence of different immunogens. This idea is supported by experiments with genetically modified immunoglobulin (Ig) knock-in mice. Sequential immunization with a series of specifically designed immunogens was required to shepherd the development of bNAbs. However, knock-in mice contain superphysiologic numbers of bNAb precursor-expressing B cells, and therefore how these results can be translated to a more physiologic setting remains to be determined. Here we make use of adoptive transfer experiments using knock-in B cells that carry a synthetic intermediate in the pathway to anti-HIV-1 bNAb development to examine how the relationship between B cell receptor affinity and precursor frequency affects germinal center (GC) B cell recruitment and clonal expansion. Immunization with soluble HIV-1 antigens can recruit bNAb precursor B cells to the GC when there are as few as 10 such cells per mouse. However, at low precursor frequencies, the extent of clonal expansion is directly proportional to the affinity of the antigen for the B cell receptor, and recruitment to GCs is variable and dependent on recirculation.

  3. Stimuli-responsive Smart Liposomes in Cancer Targeting.

    Science.gov (United States)

    Jain, Ankit; Jain, Sanjay K

    2018-02-08

    Liposomes are vesicular carriers which possess aqueous core entrapped within the lipid bilayer. These are carriers of choice because of biocompatible and biodegradable features in addition to flexibility of surface modifications at surface and lipid compositions of lipid bilayers. Liposomes have been reported well for cancer treatment using both passive and active targeting approaches however tumor microenvironment is still the biggest hurdle for safe and effective delivery of anticancer agents. To overcome this problem, stimuli-responsive smart liposomes have emerged as promising cargoes pioneered to anomalous tumor milieu in response to pH, temperature, and enzymes etc. as internal triggers, and magnetic field, ultrasound, and redox potential as external guides for enhancement of drug delivery to tumors. This review focuses on all such stimuli-responsive approaches using fabrication potentiality of liposomes in combination to various ligands, linkers, and PEGylation etc. Scientists engaged in cancer targeting approaches can get benefited greatly with this knowledgeable assemblage of advances in liposomal nanovectors. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  4. Quinolone Resistance Reversion by Targeting the SOS Response.

    Science.gov (United States)

    Recacha, E; Machuca, J; Díaz de Alba, P; Ramos-Güelfo, M; Docobo-Pérez, F; Rodriguez-Beltrán, J; Blázquez, J; Pascual, A; Rodríguez-Martínez, J M

    2017-10-10

    Suppression of the SOS response has been postulated as a therapeutic strategy for potentiating antimicrobial agents. We aimed to evaluate the impact of its suppression on reversing resistance using a model of isogenic strains of Escherichia coli representing multiple levels of quinolone resistance. E. coli mutants exhibiting a spectrum of SOS activity were constructed from isogenic strains carrying quinolone resistance mechanisms with susceptible and resistant phenotypes. Changes in susceptibility were evaluated by static (MICs) and dynamic (killing curves or flow cytometry) methodologies. A peritoneal sepsis murine model was used to evaluate in vivo impact. Suppression of the SOS response was capable of resensitizing mutant strains with genes encoding three or four different resistance mechanisms (up to 15-fold reductions in MICs). Killing curve assays showed a clear disadvantage for survival (Δlog 10 CFU per milliliter [CFU/ml] of 8 log units after 24 h), and the in vivo efficacy of ciprofloxacin was significantly enhanced (Δlog 10 CFU/g of 1.76 log units) in resistant strains with a suppressed SOS response. This effect was evident even after short periods (60 min) of exposure. Suppression of the SOS response reverses antimicrobial resistance across a range of E. coli phenotypes from reduced susceptibility to highly resistant, playing a significant role in increasing the in vivo efficacy. IMPORTANCE The rapid rise of antibiotic resistance in bacterial pathogens is now considered a major global health crisis. New strategies are needed to block the development of resistance and to extend the life of antibiotics. The SOS response is a promising target for developing therapeutics to reduce the acquisition of antibiotic resistance and enhance the bactericidal activity of antimicrobial agents such as quinolones. Significant questions remain regarding its impact as a strategy for the reversion or resensitization of antibiotic-resistant bacteria. To address this

  5. PIRATE: pediatric imaging response assessment and targeting environment

    Science.gov (United States)

    Glenn, Russell; Zhang, Yong; Krasin, Matthew; Hua, Chiaho

    2010-02-01

    By combining the strengths of various imaging modalities, the multimodality imaging approach has potential to improve tumor staging, delineation of tumor boundaries, chemo-radiotherapy regime design, and treatment response assessment in cancer management. To address the urgent needs for efficient tools to analyze large-scale clinical trial data, we have developed an integrated multimodality, functional and anatomical imaging analysis software package for target definition and therapy response assessment in pediatric radiotherapy (RT) patients. Our software provides quantitative tools for automated image segmentation, region-of-interest (ROI) histogram analysis, spatial volume-of-interest (VOI) analysis, and voxel-wise correlation across modalities. To demonstrate the clinical applicability of this software, histogram analyses were performed on baseline and follow-up 18F-fluorodeoxyglucose (18F-FDG) PET images of nine patients with rhabdomyosarcoma enrolled in an institutional clinical trial at St. Jude Children's Research Hospital. In addition, we combined 18F-FDG PET, dynamic-contrast-enhanced (DCE) MR, and anatomical MR data to visualize the heterogeneity in tumor pathophysiology with the ultimate goal of adaptive targeting of regions with high tumor burden. Our software is able to simultaneously analyze multimodality images across multiple time points, which could greatly speed up the analysis of large-scale clinical trial data and validation of potential imaging biomarkers.

  6. Vibrio elicits targeted transcriptional responses from copepod hosts.

    Science.gov (United States)

    Almada, Amalia A; Tarrant, Ann M

    2016-06-01

    Copepods are abundant crustaceans that harbor diverse bacterial communities, yet the nature of their interactions with microbiota are poorly understood. Here, we report that Vibrio elicits targeted transcriptional responses in the estuarine copepod Eurytemora affinis We pre-treated E. affinis with an antibiotic cocktail and exposed them to either a zooplankton specialist (Vibrio sp. F10 9ZB36) or a free-living species (Vibrio ordalii 12B09) for 24 h. We then identified via RNA-Seq a total of 78 genes that were differentially expressed following Vibrio exposure, including homologs of C-type lectins, chitin-binding proteins and saposins. The response differed between the two Vibrio treatments, with the greatest changes elicited upon inoculation with V. sp. F10 We suggest that these differentially regulated genes play important roles in cuticle integrity, the innate immune response, and general stress response, and that their expression may enable E. affinis to recognize and regulate symbiotic vibrios. We further report that V. sp. F10 culturability is specifically altered upon colonization of E. affinis These findings suggest that rather than acting as passive environmental vectors, copepods discriminately interact with vibrios, which may ultimately impact the abundance and activity of copepod-associated bacteria. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  7. In silico binding affinity studies of N-9 substituted 6-(4-(4-propoxyphenylpiperazin-1-yl-9H-purine derivatives-Target for P70-S6K1 & PI3K-δ kinases

    Directory of Open Access Journals (Sweden)

    Manjunath G. Sunagar

    2018-03-01

    Full Text Available P70-S6K1 & PI3K-δ kinases are identified to be involved in many physiological processes associated with cancer, therefore many of the inhibitors being designed to target these kinases are in clinical trials. In the current study we have exploited the N-9 substituted 6-(4-(4-propoxyphenyl piperazin-1-yl-9H-purine derivatives for their inhibitory properties with the above kinases. We have used an in silico docking study with seventeen purine derivatives for their binding affinity calculations. The binding affinities of these small molecules with P70-S6K1 & PI3K-δ were performed using AutoDock Vina. Among all the compounds, PP16 showed highest binding affinity of −14.7 kcal/mol with P70-S6K1 kinase & −17.2 kcal/mol with PI3K-δ kinases as compared to the molecules under clinical trials (PF-4708671 & IC-87114. Docking studies revealed that N-9 coumarine substituted purine derivative could be one of the potential ligands for the inhibition of P70-S6K1 & PI3K-δ kinases. Hence, this compound can be further investigated by in vitro and in vivo experiments for further validation.

  8. HAMS: High-Affinity Mass Spectrometry Screening. A High-Throughput Screening Method for Identifying the Tightest-Binding Lead Compounds for Target Proteins with No False Positive Identifications.

    Science.gov (United States)

    Imaduwage, Kasun P; Go, Eden P; Zhu, Zhikai; Desaire, Heather

    2016-11-01

    A major challenge in drug discovery is the identification of high affinity lead compounds that bind a particular target protein; these leads are typically identified by high throughput screens. Mass spectrometry has become a detection method of choice in drug screening assays because the target and the ligand need not be modified. Label-free assays are advantageous because they can be developed more rapidly than assays requiring labels, and they eliminate the risk of the label interfering with the binding event. However, in commonly used MS-based screening methods, detection of false positives is a major challenge. Here, we describe a detection strategy designed to eliminate false positives. In this approach, the protein and the ligands are incubated together, and the non-binders are separated for detection. Hits (protein binders) are not detectable by MS after incubation with the protein, but readily identifiable by MS when the target protein is not present in the incubation media. The assay was demonstrated using three different proteins and hundreds of non-inhibitors; no false positive hits were identified in any experiment. The assay can be tuned to select for ligands of a particular binding affinity by varying the quantity of protein used and the immobilization method. As examples, the method selectively detected inhibitors that have K i values of 0.2 μM, 50 pM, and 700 pM. These findings demonstrate that the approach described here compares favorably with traditional MS-based screening methods. Graphical Abstract ᅟ.

  9. Specificity of Bacillus thuringiensis endotoxins is correlated with the presence of high-affinity binding sites in the brush border membrane of target insect midguts

    International Nuclear Information System (INIS)

    Hofmann, C.; Vanderbruggen, H.; Hoefte, H.; Van Rie, J.; Jansens, S.; Van Mellaert, H.

    1988-01-01

    Binding studies were performed with two 125 I-labeled Bacillus thuringiensis δ-endotoxins on brush border membrane vesicles prepared from the larval midgut of the tobacco hornworm Manduca sexta or the cabbage butterfly Pieris brassicae. One δ-endotoxin, Bt2-protoxin, is a 130-kDa recombinant crystalline protein from B. thuringiensis subsp. berliner. It kills larvae of both insect species. The active Bt2-toxin is a 60-kDa proteolytic fragment of the Bt2-protoxin. It binds saturably and with high affinity to brush border membrane vesicles from the midgut of both species. The other δ-endotoxin, Bt4412-protoxin, is a 136-kDa crystalline protein from B. thuringiensis subsp. thuringiensis, which is highly toxic for P. brassicae, but not for M. sexta larvae. Bt4412-toxin, obtained after proteolytic activation of Bt4412-protoxin, shows high-affinity saturable binding to P. brassicae vesicles but not to M. sexta vesicles. The correlation between toxicity and specific binding is further strengthened by competition studies. Other B. thuringiensis δ-endotoxins active against M. sexta compete for binding of 125 I-labeled Bt2-toxin to M. sexta vesicles, whereas toxins active against dipteran or coleopteran larvae do not compete. Bt2-toxin and Bt4412-toxin bind to different sites on P. brassicae vesicles

  10. Stimuli-responsive PEGylated prodrugs for targeted doxorubicin delivery

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Minghui; Qian, Junmin, E-mail: jmqian@mail.xjtu.edu.cn; Liu, Xuefeng; Liu, Ting; Wang, Hongjie

    2015-05-01

    In recent years, stimuli-sensitive prodrugs have been extensively studied for the rapid “burst” release of antitumor drugs to enhance chemotherapeutic efficiency. In this study, a novel stimuli-sensitive prodrug containing galactosamine as a targeting moiety, poly(ethylene glycol)–doxorubicin (PEG–DOX) conjugate, was developed for targeting HepG2 human liver cancer cells. To obtain the PEG–DOX conjugate, both galactosamine-decorated poly(ethylene glycol) aldehyde (Gal-PEG-CHO) and methoxy poly(ethylene glycol) aldehyde (mPEG-CHO) were firstly synthesized and functionalized with dithiodipropionate dihydrazide (TPH) through direct reductive amination via Schiff's base formation, and then DOX molecules were chemically conjugated to the hydrazide end groups of TPH-functionalized Gal-/m-PEG chains via pH-sensitive hydrazone linkages. The chemical structures of TPH-functionalized PEG and PEG–DOX prodrug were confirmed by {sup 1}H NMR analysis. The PEG–DOX conjugate could self-assemble into spherical nanomicelles with a mean diameter of 140 nm, as indicated by transmission electron microscopy and dynamic light scattering. The drug loading content and loading efficiency in the prodrug nanomicelles were as high as 20 wt.% and 75 wt.%, respectively. In vitro drug release studies showed that DOX was released rapidly from the prodrug nanomicelles at the intracellular levels of pH and reducing agent. Cellular uptake and MTT experiments demonstrated that the galactosamine-decorated prodrug nanomicelles were more efficiently internalized into HepG2 cells via a receptor-mediated endocytosis process and exhibited a higher toxicity, compared with pristine prodrug nanomicelles. These results suggest that the novel Gal-PEG–DOX prodrug nanomicelles have tremendous potential for targeted liver cancer therapy. - Highlights: • A novel stimuli-responsive PEGylated prodrugs is synthesized. • PEGylated prodrugs can self-assemble into spherical nanoparticles (140 nm

  11. Poly(vinyl chloride) catheters modified with pH-responsive poly(methacrylic acid) with affinity for antimicrobial agents

    Science.gov (United States)

    Zuñiga-Zamorano, Ivette; Meléndez-Ortiz, H. Iván; Costoya, Alejandro; Alvarez-Lorenzo, Carmen; Concheiro, Angel; Bucio, Emilio

    2018-01-01

    Radiation-grafting of pH-responsive methacrylic acid (MAA) onto poly(vinyl chloride) (PVC) was carried out by the pre-irradiation method using gamma rays, which demonstrated to be an efficient and fast procedure for obtaining PVC-g-MAA copolymers. The influence of preparation conditions, such as absorbed dose, monomer concentration, reaction time, and reaction temperature on the grafting yield was studied. The grafting of MAA onto PVC catheters was confirmed by means of Fourier transform infrared spectroscopy (FT-IR), thermogravimetry analysis (TGA), and differential scanning calorimetry (DSC). The pH-responsiveness of the grafted copolymers (critical point 8.5) was measured by swelling under cyclic changes in the pH of the medium. Interestingly, PVC-g-MAA showed enhanced capability to immobilize benzalkonium chloride and, particularly, ciprofloxacin and to sustain the release this antimicrobial agent at both acid and alkaline pH. Tests carried out with Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus point out that the developed functionalized catheters may play a role in the prevention/management of urinary tract infections.

  12. Response to platelet-activating factor in human platelets stored and aged in plasma. Decrease in aggregation, phosphoinositide turnover, and receptor affinity

    International Nuclear Information System (INIS)

    Shukla, S.D.; Morrison, W.J.; Klachko, D.M.

    1989-01-01

    Human platelet concentrates were stored in polyolefin bags at 22 to 24 degrees C on a horizontal shaker for up to 8 days. At different intervals, aliquots of platelet-rich plasma (PRP) were removed aseptically and five variables, i.e., platelet counts, morphology, platelet-activating factor (PAF)-stimulated aggregation, phosphoinositide turnover, and [3H]PAF binding to platelet receptors, were studied. The number of platelets did not change during the 8 days of storage. Scanning electron microscopy of the platelets revealed a gradual morphologic change from biconcave flat discs to irregular, crenated forms. The PAF-induced aggregation of platelets declined with time of storage. A decrease to 50 percent of the Day 1 aggregatory response to PAF was evident on Day 2, and there was a further decline to about 20 percent by Day 6. Similarly, PAF receptor-coupled phosphoinositide turnover, as monitored by 32P incorporation into individual phosphoinositides, decreased dramatically with storage. After 2 to 3 days of storage, the phosphoinositide turnover was reduced to 50 percent of the original response, and it continued to decline to about 25 percent of original response by Day 5 or 6. The binding of [3H]PAF to washed human platelets indicated subtle changes between Days 2 and 4, which became more noticeable by Day 6. These results have raised the possibility of changes in the number of the receptors and/or their affinity for the ligand during storage. We conclude that although the number of platelets was maintained during storage for 8 days, a general deterioration of their responses to PAF occurred at the levels of cell surface receptor, transmembrane signaling (phosphoinositide turnover), and response (aggregation)

  13. Quantitative modeling of responses to chronic ionizing radiation exposure using targeted and non-targeted effects.

    Directory of Open Access Journals (Sweden)

    Igor Shuryak

    Full Text Available The biological effects of chronic ionizing radiation exposure can be difficult to study, but important to understand in order to protect the health of occupationally-exposed persons and victims of radiological accidents or malicious events. They include targeted effects (TE caused by ionizations within/close to nuclear DNA, and non-targeted effects (NTE caused by damage to other cell structures and/or activation of stress-signaling pathways in distant cells. Data on radiation damage in animal populations exposed over multiple generations to wide ranges of dose rates after the Chernobyl nuclear-power-plant accident are very useful for enhancing our understanding of these processes. We used a mechanistically-motivated mathematical model which includes TE and NTE to analyze a large published data set on chromosomal aberrations in pond snail (Lymnaea stagnalis embryos collected over 16 years from water bodies contaminated by Chernobyl fallout, and from control locations. The fraction of embryo cells with aberrations increased dramatically (>10-fold and non-linearly over a dose rate range of 0.03-420 μGy/h (0.00026-3.7 Gy/year. NTE were very important for describing the non-linearity of this radiation response: the TE-only model (without NTE performed dramatically worse than the TE+NTE model. NTE were predicted to reach ½ of maximal intensity at 2.5 μGy/h (0.022 Gy/year and to contribute >90% to the radiation response slope at dose rates <11 μGy/h (0.1 Gy/year. Internally-incorporated 90Sr was possibly more effective per unit dose than other radionuclides. The radiation response shape for chromosomal aberrations in snail embryos was consistent with data for a different endpoint: the fraction of young amoebocytes in adult snail haemolymph. Therefore, radiation may affect different snail life stages by similar mechanisms. The importance of NTE in our model-based analysis suggests that the search for modulators of NTE-related signaling pathways

  14. Molecular electron affinities

    International Nuclear Information System (INIS)

    Fukuda, E.K.

    1983-01-01

    Molecular electron affinities have historically been difficult quantities to measure accurately. These difficulties arise from differences in structure between the ion and neutral as well as the existence of excited negative ion states. To circumvent these problems, relative electron affinities were determined in this dissertation by studying equilibrium electron transfer reactions using a pulsed ion cyclotron resonance (ICR) spectrometer. Direct measurement of ion and neutral concentrations for reactions of the general type, A - + B = B - + A, allow calculation of the equilibrium constant and, therefore, the free energy change. The free energy difference is related to the difference in electron affinities between A and B. A relative electron affinity scale covering a range of about 45 kcal/mol was constructed with various substituted p-benzoquinones, nitrobenzenes, anhydrides, and benzophenones. To assign absolute electron affinities, various species with accurately known electron affinities are tied to the scale via ion-cyclotron double resonance bracketing techniques. After the relative scale is anchored to these species with well-known electron affinities, the scale is then used as a check on other electron affinity values as well as generating new electron affinity values. Many discrepancies were found between the electron affinities measured using the ICR technique and previous literature determinations

  15. Innate Immune Response and Off-Target Mis-splicing Are Common Morpholino-Induced Side Effects in Xenopus.

    Science.gov (United States)

    Gentsch, George E; Spruce, Thomas; Monteiro, Rita S; Owens, Nick D L; Martin, Stephen R; Smith, James C

    2018-03-12

    Antisense morpholino oligomers (MOs) have been indispensable tools for developmental biologists to transiently knock down (KD) genes rather than to knock them out (KO). Here we report on the implications of genetic KO versus MO-mediated KD of the mesoderm-specifying Brachyury paralogs in the frog Xenopus tropicalis. While both KO and KD embryos fail to activate the same core gene regulatory network, resulting in virtually identical morphological defects, embryos injected with control or target MOs also show a systemic GC content-dependent immune response and many off-target splicing defects. Optimization of MO dosage and increasing incubation temperatures can mitigate, but not eliminate, these MO side effects, which are consistent with the high affinity measured between MO and off-target sequence in vitro. We conclude that while MOs can be useful to profile loss-of-function phenotypes at a molecular level, careful attention must be paid to their immunogenic and off-target side effects. Copyright © 2018 The Francis Crick Institute. Published by Elsevier Inc. All rights reserved.

  16. Immunotherapeutic strategies targeting Natural killer T cell responses in cancer

    Science.gov (United States)

    Shissler, Susannah C.; Bollino, Dominique R.; Tiper, Irina V.; Bates, Joshua; Derakhshandeh, Roshanak; Webb, Tonya J.

    2017-01-01

    Natural killer T (NKT) cells are a unique subset of lymphocytes that bridge the innate and adaptive immune system. NKT cells possess a classic αβ T-cell receptor (TCR) that is able to recognize self and foreign glycolipid antigens presented by the nonclassical class I major histocompatibility complex (MHC) molecule, CD1d. Type I NKT cells (referred to as invariant NKT cells) express a semi-invariant Vα14Jα18 TCR in mice and Vα24Jα18 TCR in humans. Type II NKT cells are CD1d-restricted T cells that express a more diverse set of TCR α chains. The two types of NKT cells often exert opposing effects especially in tumor immunity, where Type II cells generally suppress tumor immunity while Type I NKT cells can enhance antitumor immune responses. In this review, we focus on the role of NKT cells in cancer. We discuss their effector and suppressive functions, as well as describe preclinical and clinical studies utilizing therapeutic strategies focused on harnessing their potent anti-tumor effector functions, and conclude with a discussion on potential next steps for the utilization of NKT cell targeted therapies for the treatment of cancer. PMID:27393665

  17. Magnetically responsive microparticles for targeted drug and radionuclide delivery

    International Nuclear Information System (INIS)

    Kaminski, M. D.; Ghebremeskel, A. N.; Nunez, L.; Kasza, K. E.; Chang, F.; Chien, T.-H.; Fisher, P. F.; Eastman, J. A.; Rosengart, A. J.; McDonald, L.; Xie, Y.; Johns, L.; Pytel, P.; Hafeli, U. O.

    2004-01-01

    We are currently investigating the use of magnetic particles--polymeric-based spheres containing dispersed magnetic nanocrystalline phases--for the precise delivery of drugs via the human vasculature. According to this review, meticulously prepared magnetic drug targeting holds promise as a safe and effective method of delivering drugs to specific organ, tissue or cellular targets. We have critically examined the wide range of approaches in the design and implementation of magnetic-particle-based drug delivery systems to date, including magnetic particle preparation, drug encapsulation, biostability, biocompatibility, toxicity, magnetic field designs, and clinical trials. However, we strongly believe that there are several limitations with past developments that need to be addressed to enable significant strides in the field. First, particle size has to be carefully chosen. Micrometer-sized magnetic particles are better attracted over a distance than nanometer sized magnetic particles by a constant magnetic field gradient, and particle sizes up to 1 (micro)m show a much better accumulation with no apparent side effects in small animal models, since the smallest blood vessels have an inner diameter of 5-7 (micro)m. Nanometer-sized particles <70 nm will accumulate in organ fenestrations despite an effective surface stabilizer. To be suitable for future human applications, our experimental approach synthesizes the magnetic drug carrier according to specific predefined outcome metrics: monodisperse population in a size range of 100 nm to 1.0 (micro)m, non-toxic, with appropriate magnetic properties, and demonstrating successful in vitro and in vivo tests. Another important variable offering possible improvement is surface polarity, which is expected to prolong particle half-life in circulation and modify biodistribution and stability of drugs in the body. The molecules in the blood that are responsible for enhancing the uptake of particles by the reticuloendothelial

  18. Affinity-tuning leukocyte integrin for development of safe therapeutics

    Science.gov (United States)

    Park, Spencer

    Much attention has been given to the molecular and cellular pathways linking inflammation with cancer and the local tumor environment to identify new target molecules that could lead to improved diagnosis and treatment. Among the many molecular players involved in the complex response, central to the induction of inflammation is intercellular adhesion molecule (ICAM)-1, which is of particular interest for its highly sensitive and localized expression in response to inflammatory signals. ICAM-1, which has been implicated to play a critical role in tumor progression in various types of cancer, has also been linked to cancer metastases, where ICAM-1 facilitates the spread of metastatic cancer cells to secondary sites. This unique expression profile of ICAM-1 throughout solid tumor microenvironment makes ICAM-1 an intriguing molecular target, which holds great potential as an important diagnostic and therapeutic tool. Herein, we have engineered the ligand binding domain, or the inserted (I) domain of a leukocyte integrin, to exhibit a wide range of monovalent affinities to the natural ligand, ICAM-1. Using the resulting I domain variants, we have created drug and gene delivery nanoparticles, as well as targeted immunotherapeutics that have the ability to bind and migrate to inflammatory sites prevalent in tumors and the associated microenvironment. Through the delivery of diagnostic agents, chemotherapeutics, and immunotherapeutics, the following chapters demonstrate that the affinity enhancements achieved by directed evolution bring the affinity of I domains into the range optimal for numerous applications.

  19. Experimental validation of plant peroxisomal targeting prediction algorithms by systematic comparison of in vivo import efficiency and in vitro PTS1 binding affinity.

    Science.gov (United States)

    Skoulding, Nicola S; Chowdhary, Gopal; Deus, Mara J; Baker, Alison; Reumann, Sigrun; Warriner, Stuart L

    2015-03-13

    Most peroxisomal matrix proteins possess a C-terminal targeting signal type 1 (PTS1). Accurate prediction of functional PTS1 sequences and their relative strength by computational methods is essential for determination of peroxisomal proteomes in silico but has proved challenging due to high levels of sequence variability of non-canonical targeting signals, particularly in higher plants, and low levels of availability of experimentally validated non-canonical examples. In this study, in silico predictions were compared with in vivo targeting analyses and in vitro thermodynamic binding of mutated variants within the context of one model targeting sequence. There was broad agreement between the methods for entire PTS1 domains and position-specific single amino acid residues, including residues upstream of the PTS1 tripeptide. The hierarchy Leu>Met>Ile>Val at the C-terminal position was determined for all methods but both experimental approaches suggest that Tyr is underweighted in the prediction algorithm due to the absence of this residue in the positive training dataset. A combination of methods better defines the score range that discriminates a functional PTS1. In vitro binding to the PEX5 receptor could discriminate among strong targeting signals while in vivo targeting assays were more sensitive, allowing detection of weak functional import signals that were below the limit of detection in the binding assay. Together, the data provide a comprehensive assessment of the factors driving PTS1 efficacy and provide a framework for the more quantitative assessment of the protein import pathway in higher plants. Copyright © 2014 Elsevier Ltd. All rights reserved.

  20. Helper T cell epitope-mapping reveals MHC-peptide binding affinities that correlate with T helper cell responses to pneumococcal surface protein A.

    Directory of Open Access Journals (Sweden)

    Rajesh Singh

    2010-02-01

    Full Text Available Understanding the requirements for protection against pneumococcal carriage and pneumonia will greatly benefit efforts in controlling these diseases. Several proteins and polysaccharide capsule have recently been implicated in the virulence of and protective immunity against Streptococcus pneumonia. Pneumococcal surface protein A (PspA is highly conserved among S. pneumonia strains, inhibits complement activation, binds lactoferrin, elicits protective systemic immunity against pneumococcal infection, and is necessary for full pneumococcal virulence. Identification of PspA peptides that optimally bind human leukocyte antigen (HLA would greatly contribute to global vaccine efforts, but this is hindered by the multitude of HLA polymorphisms. Here, we have used an experimental data set of 54 PspA peptides and in silico methods to predict peptide binding to HLA and murine major histocompatibility complex (MHC class II. We also characterized spleen- and cervical lymph node (CLN-derived helper T lymphocyte (HTL cytokine responses to these peptides after S. pneumonia strain EF3030-challenge in mice. Individual, yet overlapping peptides, 15 amino acids in length revealed residues 199 to 246 of PspA (PspA(199-246 consistently caused the greatest IFN-gamma, IL-2, IL-5 and proliferation as well as moderate IL-10 and IL-4 responses by ex vivo stimulated splenic and CLN CD4(+ T cells isolated from S. pneumonia strain EF3030-challeged F(1 (B6xBALB/c mice. IEDB, RANKPEP, SVMHC, MHCPred, and SYFPEITHI in silico analysis tools revealed peptides in PspA(199-246 also interact with a broad range of HLA-DR, -DQ, and -DP allelles. These data suggest that predicted MHC class II-peptide binding affinities do not always correlate with T helper (Th cytokine or proliferative responses to PspA peptides, but when used together with in vivo validation can be a useful tool to choose candidate pneumococcal HTL epitopes.

  1. Dynamic response of the target container under pulsed heating

    Energy Technology Data Exchange (ETDEWEB)

    Liping Ni [Paul Scherrer Inst. (PSI), Villigen (Switzerland)

    1997-09-01

    The structural mechanics of a liquid target container for pulsed spallation sources have been simulated using both a commercial code and a PSI-developed program. Results from the transient thermal-structural analysis showed that, due to inertia effects, the dynamic stress in the target container is contributed mainly from direct heating in the initial time stage, and later from the pressure wave in the target liquid once it reaches the wall. (author) figs., tab., refs.

  2. INVESTIGATION ON THE RESPONSE OF SEGMENTED CONCRETE TARGETS TO PROJECTILE IMPACTS

    Energy Technology Data Exchange (ETDEWEB)

    Booker, Paul M.; Cargile, James D.; Kistler, Bruce L.; La Saponara, Valeria

    2009-07-19

    The study of penetrator performance without free-surface effects can require prohibitively large monolithic targets. One alternative to monolithic targets is to use segmented targets made by stacking multiple concrete slabs in series. This paper presents an experimental investigation on the performance of segmented concrete targets. Six experiments were carried out on available small scale segmented and monolithic targets using instrumented projectiles. In all but one experiment using stacked slabs, the gap between slabs remained open. In the final experiment design, grout was inserted between the slabs, and this modification produced a target response that more closely represents that of the monolithic target. The results from this study suggest that further research on segmented targets is justified, to explore in more detail the response of segmented targets and the results of large scale tests when using segmented targets versus monolithic targets.

  3. Structural dynamic response of target container against proton beam

    Energy Technology Data Exchange (ETDEWEB)

    Kikuchi, Kenji; Ishikura, Syuichi; Futakawa, Masatoshi; Hino, Ryutaro [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment

    1997-11-01

    Stress waves were analyzed for a target container of neutron science research project using a high-intensity proton accelerator that generates high energy and high current proton beam. In the mercury target, the pulsed proton beam generates intense power density in the course of spallation reaction and causes pressure wave in the mercury and stress wave in the target container due to a sudden temperature change. Structural integrity of the target container depends on the power intensity at a maximum energy deposit. A broad proton profile is favorable to the structural assessment of the container rather than narrow one. Stress wave have propagated in the target container at a speed of sound. It only takes 0.1 ms for the size of 40 cm length stainless steel container. Further assessment is necessary to optimize a geometry of the container and establish a method to evaluate a life time. (author)

  4. Structural dynamic response of target container against proton beam

    International Nuclear Information System (INIS)

    Kikuchi, Kenji; Ishikura, Syuichi; Futakawa, Masatoshi; Hino, Ryutaro

    1997-01-01

    Stress waves were analyzed for a target container of neutron science research project using a high-intensity proton accelerator that generates high energy and high current proton beam. In the mercury target, the pulsed proton beam generates intense power density in the course of spallation reaction and causes pressure wave in the mercury and stress wave in the target container due to a sudden temperature change. Structural integrity of the target container depends on the power intensity at a maximum energy deposit. A broad proton profile is favorable to the structural assessment of the container rather than narrow one. Stress wave have propagated in the target container at a speed of sound. It only takes 0.1 ms for the size of 40 cm length stainless steel container. Further assessment is necessary to optimize a geometry of the container and establish a method to evaluate a life time. (author)

  5. Role of DNA Repair Factor Xeroderma Pigmentosum Protein Group C in Response to Replication Stress As Revealed by DNA Fragile Site Affinity Chromatography and Quantitative Proteomics.

    Science.gov (United States)

    Beresova, Lucie; Vesela, Eva; Chamrad, Ivo; Voller, Jiri; Yamada, Masayuki; Furst, Tomas; Lenobel, Rene; Chroma, Katarina; Gursky, Jan; Krizova, Katerina; Mistrik, Martin; Bartek, Jiri

    2016-12-02

    Replication stress (RS) fuels genomic instability and cancer development and may contribute to aging, raising the need to identify factors involved in cellular responses to such stress. Here, we present a strategy for identification of factors affecting the maintenance of common fragile sites (CFSs), which are genomic loci that are particularly sensitive to RS and suffer from increased breakage and rearrangements in tumors. A DNA probe designed to match the high flexibility island sequence typical for the commonly expressed CFS (FRA16D) was used as specific DNA affinity bait. Proteins significantly enriched at the FRA16D fragment under normal and replication stress conditions were identified using stable isotope labeling of amino acids in cell culture-based quantitative mass spectrometry. The identified proteins interacting with the FRA16D fragment included some known CFS stabilizers, thereby validating this screening approach. Among the hits from our screen so far not implicated in CFS maintenance, we chose Xeroderma pigmentosum protein group C (XPC) for further characterization. XPC is a key factor in the DNA repair pathway known as global genomic nucleotide excision repair (GG-NER), a mechanism whose several components were enriched at the FRA16D fragment in our screen. Functional experiments revealed defective checkpoint signaling and escape of DNA replication intermediates into mitosis and the next generation of XPC-depleted cells exposed to RS. Overall, our results provide insights into an unexpected biological role of XPC in response to replication stress and document the power of proteomics-based screening strategies to elucidate mechanisms of pathophysiological significance.

  6. Continuous affine processes

    DEFF Research Database (Denmark)

    Buchardt, Kristian

    2016-01-01

    Affine processes possess the property that expectations of exponential affine transformations are given by a set of Riccati differential equations, which is the main feature of this popular class of processes. In this paper we generalise these results for expectations of more general transformati...

  7. Fluctuations in Echo Level Associated with Changes in Target Aspect and Target Frequency Response (abstract only)

    NARCIS (Netherlands)

    Zampolli, M.; Ainslie, M.A.; Zon, T. van

    2011-01-01

    Ping-to-ping variations in echo level can be caused by time variations in sonar parameters (source level, orientation), target aspect, relative and absolute motion of sonar and target, and time varying environment (e.g. surface waves). Quantifying and understanding such fluctuations are important,

  8. C. difficile 630Δerm Spo0A regulates sporulation, but does not contribute to toxin production, by direct high-affinity binding to target DNA.

    Directory of Open Access Journals (Sweden)

    Katharina E Rosenbusch

    Full Text Available Clostridium difficile is a Gram positive, anaerobic bacterium that can form highly resistant endospores. The bacterium is the causative agent of C. difficile infection (CDI, for which the symptoms can range from a mild diarrhea to potentially fatal pseudomembranous colitis and toxic megacolon. Endospore formation in Firmicutes, including C. difficile, is governed by the key regulator for sporulation, Spo0A. In Bacillus subtilis, this transcription factor is also directly or indirectly involved in various other cellular processes. Here, we report that C. difficile Spo0A shows a high degree of similarity to the well characterized B. subtilis protein and recognizes a similar binding sequence. We find that the laboratory strain C. difficile 630Δerm contains an 18bp-duplication near the DNA-binding domain compared to its ancestral strain 630. In vitro binding assays using purified C-terminal DNA binding domain of the C. difficile Spo0A protein demonstrate direct binding to DNA upstream of spo0A and sigH, early sporulation genes and several other putative targets. In vitro binding assays suggest that the gene encoding the major clostridial toxin TcdB may be a direct target of Spo0A, but supernatant derived from a spo0A negative strain was no less toxic towards Vero cells than that obtained from a wild type strain, in contrast to previous reports. These results identify for the first time direct (putative targets of the Spo0A protein in C. difficile and make a positive effect of Spo0A on production of the large clostridial toxins unlikely.

  9. Affinity Strings: Enterprise Data for Resource Recommendations

    Directory of Open Access Journals (Sweden)

    Shane Nackerud

    2008-12-01

    Full Text Available The University of Minnesota Libraries have created a MyLibrary portal, with databases and e-journals targeted to users, based on their affiliations. The University's enterprise authentication system provides an "affinity string", now used to personalize the MyLibrary portal. This affinity string automates discovery of a user's relationship to the University--describing a user's academic department and degree program or position at the University. Affinity strings also provide the Libraries with an anonymized view of resource usage, allowing data collection that respects users' privacy and lays the groundwork for automated recommendation of relevant resources based on the practices and habits of their peers.

  10. Synthesis and biological evaluation of copper-64 radiolabeled [DUPA-6-Ahx-(NODAGA)-5-Ava-BBN(7-14)NH2], a novel bivalent targeting vector having affinity for two distinct biomarkers (GRPr/PSMA) of prostate cancer

    International Nuclear Information System (INIS)

    Bandari, Rajendra Prasad; Jiang, Zongrun; Reynolds, Tamila Stott; Bernskoetter, Nicole E.; Szczodroski, Ashley F.; Bassuner, Kurt J.; Kirkpatrick, Daniel L.; Rold, Tammy L.; Sieckman, Gary L.; Hoffman, Timothy J.; Connors, James P.; Smith, Charles J.

    2014-01-01

    Gastrin-releasing peptide receptors (GRPr) and prostate-specific membrane antigen (PSMA) are two identifying biomarkers expressed in very high numbers on prostate cancer cells and could serve as a useful tool for molecular targeting and diagnosis of disease via positron-emission tomography (PET). The aim of this study was to produce the multipurpose, bivalent [DUPA-6-Ahx-( 64 Cu-NODAGA)-5-Ava-BBN(7-14)NH 2 ] radioligand for prostate cancer imaging, where DUPA = (2-[3-(1,3-dicarboxypropyl)-ureido]pentanedioic acid), a small-molecule, PSMA-targeting probe, 6Ahx = 6-aminohexanoic acid, 5-Ava = 5-aminovaleric acid, NODAGA = [2-(4,7-biscarboxymethyl)-1,4,7-(triazonan-1-yl)pentanedioic acid] (a derivative of NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid)), and BBN(7-14)NH 2 = bombesin, a GRPr-specific peptide targeting probe. Methods: The PSMA/GRPr dual targeting ligand precursor [DUPA-6-Ahx-K-5-Ava-BBN(7-14)NH 2 ], was synthesized by solid-phase and manual peptide synthesis, after which NODAGA was added via manual conjugation to the ε-amine of lysine (K). The new bivalent GRPr/PSMA targeting vector was purified by reversed-phase high performance liquid chromatography (RP-HPLC), characterized by electrospray-ionization mass spectrometry (ESI-MS), and metallated with 64 CuCl 2 and nat CuCl 2 . The receptor binding affinity was evaluated in human, prostate, PC-3 (GRPr-positive) and LNCaP (PSMA-positive) cells and the tumor-targeting efficacy determined in severe combined immunodeficient (SCID) and athymic nude mice bearing PC-3 and LNCaP tumors. Whole-body maximum intensity microPET/CT images of PC-3/LNCaP tumor-bearing mice were obtained 18 h post-injection (p.i.). Results: Competitive binding assays in PC-3 and LNCaP cells indicated high receptor binding affinity for the [DUPA-6-Ahx-( nat Cu-NODAGA)-5-Ava-BBN(7-14)NH 2 ] conjugate. MicroPET scintigraphy in PC-3/LNCaP tumor-bearing mice indicated that xenografted tumors were visible at 18 h p.i. with collateral

  11. Synthesis and biological evaluation of copper-64 radiolabeled [DUPA-6-Ahx-(NODAGA)-5-Ava-BBN(7-14)NH2], a novel bivalent targeting vector having affinity for two distinct biomarkers (GRPr/PSMA) of prostate cancer.

    Science.gov (United States)

    Bandari, Rajendra Prasad; Jiang, Zongrun; Reynolds, Tamila Stott; Bernskoetter, Nicole E; Szczodroski, Ashley F; Bassuner, Kurt J; Kirkpatrick, Daniel L; Rold, Tammy L; Sieckman, Gary L; Hoffman, Timothy J; Connors, James P; Smith, Charles J

    2014-04-01

    Gastrin-releasing peptide receptors (GRPr) and prostate-specific membrane antigen (PSMA) are two identifying biomarkers expressed in very high numbers on prostate cancer cells and could serve as a useful tool for molecular targeting and diagnosis of disease via positron-emission tomography (PET). The aim of this study was to produce the multipurpose, bivalent [DUPA-6-Ahx-((64)Cu-NODAGA)-5-Ava-BBN(7-14)NH2] radioligand for prostate cancer imaging, where DUPA = (2-[3-(1,3-dicarboxypropyl)-ureido]pentanedioic acid), a small-molecule, PSMA-targeting probe, 6Ahx = 6-aminohexanoic acid, 5-Ava = 5-aminovaleric acid, NODAGA = [2-(4,7-biscarboxymethyl)-1,4,7-(triazonan-1-yl)pentanedioic acid] (a derivative of NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid)), and BBN(7-14)NH2 = bombesin, a GRPr-specific peptide targeting probe. The PSMA/GRPr dual targeting ligand precursor [DUPA-6-Ahx-K-5-Ava-BBN(7-14)NH2], was synthesized by solid-phase and manual peptide synthesis, after which NODAGA was added via manual conjugation to the ε-amine of lysine (K). The new bivalent GRPr/PSMA targeting vector was purified by reversed-phase high performance liquid chromatography (RP-HPLC), characterized by electrospray-ionization mass spectrometry (ESI-MS), and metallated with (64)CuCl2 and (nat)CuCl2. The receptor binding affinity was evaluated in human, prostate, PC-3 (GRPr-positive) and LNCaP (PSMA-positive) cells and the tumor-targeting efficacy determined in severe combined immunodeficient (SCID) and athymic nude mice bearing PC-3 and LNCaP tumors. Whole-body maximum intensity microPET/CT images of PC-3/LNCaP tumor-bearing mice were obtained 18 h post-injection (p.i.). Competitive binding assays in PC-3 and LNCaP cells indicated high receptor binding affinity for the [DUPA-6-Ahx-((nat)Cu-NODAGA)-5-Ava-BBN(7-14)NH2] conjugate. MicroPET scintigraphy in PC-3/LNCaP tumor-bearing mice indicated that xenografted tumors were visible at 18h p.i. with collateral, background radiation also

  12. Response surface methodology optimization of partitioning of xylanase form Aspergillus Niger by metal affinity polymer-salt aqueous two-phase systems.

    Science.gov (United States)

    Fakhari, Mohamad Ali; Rahimpour, Farshad; Taran, Mojtaba

    2017-09-15

    Aqueous two phase affinity partitioning system using metal ligands was applied for partitioning and purification of xylanase produced by Aspergillus Niger. To minimization the number of experiments for the design parameters and develop predictive models for optimization of the purification process, response surface methodology (RSM) with a face-centered central composite design (CCF) has been used. Polyethylene glycol (PEG) 6000 was activated using epichlorohydrin, covalently linked to iminodiacetic acid (IDA), and the specific metal ligand Cu was attached to the polyethylene glycol-iminodiacetic acid (PEG-IDA). The influence of some experimental variables such as PEG (10-18%w/w), sodium sulfate (8-12%), PEG-IDA-Cu 2+ concentration (0-50% w/w of total PEG), pH of system (4-8) and crude enzyme loading (6-18%w/w) on xylanase and total protein partitioning coefficient, enzyme yield and enzyme specific activity were systematically evaluated. Two optimal point with high enzyme partitioning factor 10.97 and yield 79.95 (including 10% PEG, 12% Na 2 SO 4 , 50% ligand, pH 8 and 6% crude enzyme loading) and high specific activity in top phase 42.21 (including 14.73% PEG, 8.02% Na 2 SO 4 , 28.43% ligand, pH 7.7 and 6.08% crude enzyme loading) were attained. The adequacy of the RSM models was verified by a good agreement between experimental and predicted results. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Affinity in electrophoresis.

    Science.gov (United States)

    Heegaard, Niels H H

    2009-06-01

    The journal Electrophoresis has greatly influenced my approaches to biomolecular affinity studies. The methods that I have chosen as my main tools to study interacting biomolecules--native gel and later capillary zone electrophoresis--have been the topic of numerous articles in Electrophoresis. Below, the role of the journal in the development and dissemination of these techniques and applications reviewed. Many exhaustive reviews on affinity electrophoresis and affinity CE have been published in the last few years and are not in any way replaced by the present deliberations that are focused on papers published by the journal.

  14. In Silico Characterization of the Binding Affinity of Dendrimers to Penicillin-Binding Proteins (PBPs): Can PBPs be Potential Targets for Antibacterial Dendrimers?

    Science.gov (United States)

    Ahmed, Shaimaa; Vepuri, Suresh B; Ramesh, Muthusamy; Kalhapure, Rahul; Suleman, Nadia; Govender, Thirumala

    2016-04-01

    We have shown that novel silver salts of poly (propyl ether) imine (PETIM) dendron and dendrimers developed in our group exhibit preferential antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus. This led us to examine whether molecular modeling methods could be used to identify the key structural design principles for a bioactive lead molecule, explore the mechanism of binding with biological targets, and explain their preferential antibacterial activity. The current article reports the conformational landscape as well as mechanism of binding of generation 1 PETIM dendron and dendrimers to penicillin-binding proteins (PBPs) in order to understand the antibacterial activity profiles of their silver salts. Molecular dynamics at different simulation protocols and conformational analysis were performed to elaborate on the conformational features of the studied dendrimers, as well as to create the initial structure for further binding studies. The results showed that for all compounds, there were no significant conformational changes due to variation in simulation conditions. Molecular docking calculations were performed to investigate the binding theme between the studied dendrimers and PBPs. Interestingly, in significant accordance with the experimental data, dendron and dendrimer with aliphatic cores were found to show higher activity against S. aureus than the dendrimer with an aromatic core. The latter showed higher activity against MRSA. The findings from this computational and molecular modeling report together with the experimental results serve as a road map toward designing more potent antibacterial dendrimers against resistant bacterial strains.

  15. Examination of Studies Targeting Social Skills with Pivotal Response Treatment

    Science.gov (United States)

    Bozkus Genc, Gulden; Vuran, Sezgin

    2013-01-01

    In early education, especially in effective teaching to children with autism spectrum disorders, the teaching methods which are applicable in natural settings like pivotal response treatment (PRT) are commonly used. It is one of the naturalistic intervention models aiming to facilitate the stimulant-response generalization, decrease the dependency…

  16. A Generalized Affine Isoperimetric Inequality

    OpenAIRE

    Chen, Wenxiong; Howard, Ralph; Lutwak, Erwin; Yang, Deane; Zhang, Gaoyong

    2004-01-01

    A purely analytic proof is given for an inequality that has as a direct consequence the two most important affine isoperimetric inequalities of plane convex geometry: The Blaschke-Santalo inequality and the affine isoperimetric inequality of affine differential geometry.

  17. Electron affinities: theoretical

    International Nuclear Information System (INIS)

    Kaufman, J.J.

    1976-01-01

    A brief description is given of the conceptual background and formalism of the various ab-initio and semi-ab-initio quantum computational techniques for calculating atomic and molecular electron affinities: Hartree--Fock--Roothaan SCF, configuration interaction (CI), multiconfiguration SCF (MC-SCF), Bethe--Goldstone, superposition of configurations (SOC), ab-initio effective core model potentials, Xα-MS, plus other less common methods. Illustrative and comparative examples of electron affinities calculated by these various methods are presented

  18. In vitro and in vivo evaluation of a (18F-labeled high affinity NOTA conjugated bombesin antagonist as a PET ligand for GRPR-targeted tumor imaging.

    Directory of Open Access Journals (Sweden)

    Zohreh Varasteh

    Full Text Available Expression of the gastrin-releasing peptide receptor (GRPR in prostate cancer suggests that this receptor can be used as a potential molecular target to visualize and treat these tumors. We have previously investigated an antagonist analog of bombesin (D-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2, RM26 conjugated to 1,4,7-triazacyclononane-N,N',N''-triacetic acid (NOTA via a diethylene glycol (PEG2 spacer (NOTA-P2-RM26 labeled with (68Ga and (111In. We found that this conjugate has favorable properties for in vivo imaging of GRPR-expression. The focus of this study was to develop a (18F-labelled PET agent to visualize GRPR. NOTA-P2-RM26 was labeled with (18F using aluminum-fluoride chelation. Stability, in vitro binding specificity and cellular processing tests were performed. The inhibition efficiency (IC50 of the [(natF]AlF-NOTA-P2-RM26 was compared to that of the (natGa-loaded peptide using (125I-Tyr(4-BBN as the displacement radioligand. The pharmacokinetics and in vivo binding specificity of the compound were studied. NOTA-P2-RM26 was labeled with (18F within 1 h (60-65% decay corrected radiochemical yield, 55 GBq/µmol. The radiopeptide was stable in murine serum and showed high specific binding to PC-3 cells. [(natF]AlF-NOTA-P2-RM26 showed a low nanomolar inhibition efficiency (IC50=4.4±0.8 nM. The internalization rate of the tracer was low. Less than 14% of the cell-bound radioactivity was internalized after 4 h. The biodistribution of [(18F]AlF-NOTA-P2-RM26 demonstrated rapid blood clearance, low liver uptake and low kidney retention. The tumor uptake at 3 h p.i. was 5.5±0.7 %ID/g, and the tumor-to-blood, -muscle and -bone ratios were 87±42, 159±47, 38±16, respectively. The uptake in tumors, pancreas and other GRPR-expressing organs was significantly reduced when excess amount of non-labeled peptide was co-injected. The low uptake in bone suggests a high in vivo stability of the Al-F bond. High contrast PET image was obtained 3 h p

  19. Assessing the Response to Targeted Therapies in Renal Cell Carcinoma: Technical Insights and Practical Considerations

    NARCIS (Netherlands)

    Bex, A.; Fournier, L.; Lassau, N.; Mulders, P.F.A.; Nathan, P.; Oyen, W.J.G.; Powles, T.

    2014-01-01

    CONTEXT: The introduction of targeted agents for the treatment of renal cell carcinoma (RCC) has resulted in new challenges for assessing response to therapy, and conventional response criteria using computed tomography (CT) are limited. It is widely recognised that targeted therapies may lead to

  20. Therapeutic responses in systematic targeted alpha therapy trial for melanoma

    International Nuclear Information System (INIS)

    Raja, C.; Rizvi, S.M.A.; Song, E.Y.; Graham, P.; Kearsley, J.H.; Goldsmith, H.; Bosserhoff, A.; Morgenstern, A.; Apostolidis, C.; Reisfeld, R.

    2006-01-01

    Full text: The therapeutic response of melanoma patients after systemic alpha therapy has been investigated in an open-labeled Phase 1 dose escalation study to establish the effective dose of alpha-immunoconjugate 2l3 Bi-cDTPA-9.2.27 mAb (AIC). The tools used to investigate the effects were physical examination; the images of the tumours, pathology comparisons over 12 weeks; CT comparisons and changes in tumour marker over 8 weeks. The physical examination indicated varied tumour responses. One patient showed complete response at 12 weeks post-TAT, with 20 of 21 tumours completely regressing, the last reduced by 80%. The tumour beds were biopsied for staining; S l OO was negative and no viable cells were observed. Most patients showed stable disease at 2 weeks. In one patient the CT comparison of 8 weeks with baseline showed marked reduction in three lung lesions. At least 4/21 patients showed partial response at 4 to 8 weeks and the same number showed stable disease. The disease progressed in 7 patients. The tumour marker melanoma inhibitory activity protein (MIA) showed reductions over 8 weeks, and was consistent with observations in most patients. Complete and partial responses were observed in systemic TAT for stage IV melanoma, but there was no dose-response relationship

  1. High-affinity human leucocyte antigen class I binding variola-derived peptides induce CD4(+) T cell responses more than 30 years post-vaccinia virus vaccination

    DEFF Research Database (Denmark)

    Wang, M.; Tang, Sheila Tuyet; Lund, Ole

    2009-01-01

    Interferon-gamma secreting T lymphocytes against pox virus-derived synthetic 9-mer peptides were tested by enzyme-linked immunospot in peripheral blood of individuals vaccinated with vaccinia virus more than 30 years ago. The peptides were characterized biochemically as high-affinity human leucoc...

  2. Opposing effects of actin signaling and LFA-1 on establishing the affinity threshold for inducing effector T-cell responses in mice

    Czech Academy of Sciences Publication Activity Database

    Palmer, E.; Drobek, Aleš; Štěpánek, Ondřej

    2016-01-01

    Roč. 46, č. 8 (2016), s. 1887-1901 ISSN 0014-2980 R&D Projects: GA ČR GJ16-09208Y Institutional support: RVO:68378050 Keywords : Actin cytoskeleton * Antigen affinity treshold * LFA-1 * Rap1 * Rho-family GTPases * T-cell receptor signaling Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.227, year: 2016

  3. Know your epidemic, know your response: targeting HIV in Asia

    DEFF Research Database (Denmark)

    Lazarus, Jeff; Curth, Nadja; Bridge, Jamie

    2010-01-01

    This article provides an overview of the HIV epidemic in Asia, the context within which the epidemic is evolving, and the key actions to address the challenges faced by countries and risk groups. HIV epidemics across Asia are predominantly concentrated among most-at-risk populations. Although...... prevention and treatment services. In order to reach the Millennium Development Goal of halting and reversing the spread of HIV by 2015 and to achieve universal access to HIV treatment, these barriers must be overcome across Asia. High-impact programs must be targeted at those in need, with continuous...

  4. Nitric oxide mediated bystander responses induced by microbeam targeted cells

    International Nuclear Information System (INIS)

    Shao, C.; Prise, K.M.; Folkard, M.; Michael, B.D.

    2003-01-01

    Considerable evidence has recently been accumulated in support of the existence of a 'bystander effect', which cells having received no irradiation show biological consequences from their vicinal irradiated cells. The application of microbeams is providing new insights into the radiation-induced bystander effect. The present study found that when a fraction of radioresistant human glioblastoma cells were individually targeted with a precise number of helium ions generated from the Gray Cancer Institute Charged Particle Microbeam, micronucleus (MN) induction significantly exceeded the expected value that was calculated from the number of MN observed when all of the cells were targeted assuming no bystander effect occurring. Even when only a single cell within a population was hit by one helium ion, the MN induction in the population could be increased by 16%. These results provide direct evidence of radiation-induced bystander effect. Moreover, MN was effectively induced in the unirradiated primary human fibroblasts and glioblastoma cells either co-cultured with irradiated cells or treated with the medium harvested from irradiated cells, indicating a signal molecule was produced from the irradiated cells. However, when c-PTIO, a nitric oxide (NO)-specific scavenger, was present in the medium during and after irradiation until MN analysis, the production of MN in all of the above cases was reduced to low levels. Consequently, NO plays an important role in the radiation-induced bystander effect

  5. Quinolone Resistance Reversion by Targeting the SOS Response

    Directory of Open Access Journals (Sweden)

    E. Recacha

    2017-10-01

    Full Text Available Suppression of the SOS response has been postulated as a therapeutic strategy for potentiating antimicrobial agents. We aimed to evaluate the impact of its suppression on reversing resistance using a model of isogenic strains of Escherichia coli representing multiple levels of quinolone resistance. E. coli mutants exhibiting a spectrum of SOS activity were constructed from isogenic strains carrying quinolone resistance mechanisms with susceptible and resistant phenotypes. Changes in susceptibility were evaluated by static (MICs and dynamic (killing curves or flow cytometry methodologies. A peritoneal sepsis murine model was used to evaluate in vivo impact. Suppression of the SOS response was capable of resensitizing mutant strains with genes encoding three or four different resistance mechanisms (up to 15-fold reductions in MICs. Killing curve assays showed a clear disadvantage for survival (Δlog10 CFU per milliliter [CFU/ml] of 8 log units after 24 h, and the in vivo efficacy of ciprofloxacin was significantly enhanced (Δlog10 CFU/g of 1.76 log units in resistant strains with a suppressed SOS response. This effect was evident even after short periods (60 min of exposure. Suppression of the SOS response reverses antimicrobial resistance across a range of E. coli phenotypes from reduced susceptibility to highly resistant, playing a significant role in increasing the in vivo efficacy.

  6. Functionalized mesoporous silica nanoparticles for stimuli-responsive and targeted

    Energy Technology Data Exchange (ETDEWEB)

    Knezevic, Nikola [Iowa State Univ., Ames, IA (United States)

    2009-12-15

    Construction of functional supramolecular nanoassemblies has attracted great deal of attention in recent years for their wide spectrum of practical applications. Mesoporous silica nanoparticles (MSN) in particular were shown to be effective scaffolds for the construction of drug carriers, sensors and catalysts. Herein, we describe the synthesis and characterization of stimuli-responsive, controlled release MSN-based assemblies for drug delivery.

  7. Dual responsive PNIPAM–chitosan targeted magnetic nanopolymers for targeted drug delivery

    Energy Technology Data Exchange (ETDEWEB)

    Yadavalli, Tejabhiram, E-mail: tejabhiram@gmail.com [Nanotechnology Research Centre, SRM University, Chennai 603203 (India); Ramasamy, Shivaraman [Nanotechnology Research Centre, SRM University, Chennai 603203 (India); School of Physics, The University of Western Australia, 35 Stirling Hwy, Crawley, WA 6009 (Australia); Chandrasekaran, Gopalakrishnan; Michael, Isaac; Therese, Helen Annal [Nanotechnology Research Centre, SRM University, Chennai 603203 (India); Chennakesavulu, Ramasamy [Department of Pharmacy practice, SRM College of Pharmacy, Chennai 603203 (India)

    2015-04-15

    A dual stimuli sensitive magnetic hyperthermia based drug delivery system has been developed for targeted cancer treatment. Thermosensitive amine terminated poly-N-isopropylacrylamide complexed with pH sensitive chitosan nanoparticles was prepared as the drug carrier. Folic acid and fluorescein were tagged to the nanopolymer complex via N-hydroxysuccinimide and ethyl-3-(3-dimethylaminopropyl)carbodiimide reaction to form a fluorescent and cancer targeting magnetic carrier system. The formation of the polymer complex was confirmed using infrared spectroscopy. Gadolinium doped nickel ferrite nanoparticles prepared by a hydrothermal method were encapsulated in the polymer complex to form a magnetic drug carrier system. The proton relaxation studies on the magnetic carrier system revealed a 200% increase in the T1 proton relaxation rate. These magnetic carriers were loaded with curcumin using solvent evaporation method with a drug loading efficiency of 86%. Drug loaded nanoparticles were tested for their targeting and anticancer properties on four cancer cell lines with the help of MTT assay. The results indicated apoptosis of cancer cell lines within 3 h of incubation. - Highlights: • The use of gadolinium doped nickel ferrite with the suggested doping level. • The use of PNIPMA–chitosan polymer with folic acid and fluorescein as a drug carrier complex. • Magnetic hyperthermia studies of gadolinium doped nickel ferrites are being reported for the first time. • Proton relaxivity studies which indicate the MRI contrasting properties on the reported system are new. • Use of curcumin, a hydrophobic Indian spice as a cancer killing agent inside the reported magnetic polymer complex.

  8. A self-affine multi-fractal wave turbulence discrimination method using data from single point fast response sensors in a nocturnal atmospheric boundary layer

    OpenAIRE

    Kamada, Ray; Decaria, Alex Joseph

    1992-01-01

    We present DA, a self-affine, multi-fractal which may become the first routine wave/turbulence discriminant for time series data. Using nocturnal atmospheric data, we show the advantages of D A over self-similar fractals and standard turbulence measures such as FFTs, Richardson number, Brunt-Vaisala frequency, buoyancy length scale, variances, turbulent kinetic energy, and phase averaging. DA also shows promise in resolving "wave-break" events. Since it uses local basis functions, DA may be...

  9. Affine stochastic mortality

    NARCIS (Netherlands)

    Schrager, D.F.

    2006-01-01

    We propose a new model for stochastic mortality. The model is based on the literature on affine term structure models. It satisfies three important requirements for application in practice: analytical tractibility, clear interpretation of the factors and compatibility with financial option pricing

  10. Affine pairings on ARM

    NARCIS (Netherlands)

    Acar, T.; Lauter, K.; Naehrig, M.; Shumow, D.

    2011-01-01

    Pairings on elliptic curves are being used in an increasing number of cryptographic applications on many different devices and platforms, but few performance numbers for cryptographic pairings have been reported on embedded and mobile devices. In this paper we give performance numbers for affine and

  11. Affine pairings on ARM

    NARCIS (Netherlands)

    Acar, T.; Lauter, K.; Naehrig, M.; Shumow, D.; Abdalla, M.; Lange, T.

    2013-01-01

    We report on relative performance numbers for affine and projective pairings on a dual-core Cortex A9 ARM processor. Using a fast inversion in the base field and doing inversion in extension fields by using the norm map to reduce to inversions in smaller fields, we find a very low ratio of

  12. 3D modelling of the electromagnetic response of geophysical targets using the FDTD method

    Energy Technology Data Exchange (ETDEWEB)

    Debroux, P.S. [Univ. of Arizona, Tucson, AZ (United States). Mining and Geological Engineering Dept.

    1996-05-01

    A publicly available and maintained electromagnetic finite-difference time domain (FDTD) code has been applied to the forward modelling of the response of 1D, 2D and 3D geophysical targets to a vertical magnetic dipole excitation. The FDTD method is used to analyze target responses in the 1 MHz to 100 MHz range, where either conduction or displacement currents may have the controlling role. The response of the geophysical target to the excitation is presented as changes in the magnetic field ellipticity. The results of the FDTD code compare favorably with previously published integral equation solutions of the response of 1D targets, and FDTD models calculated with different finite-difference cell sizes are compared to find the effect of model discretization on the solution. The discretization errors, calculated as absolute error in ellipticity, are presented for the different ground geometry models considered, and are, for the most part, below 10% of the integral equation solutions. Finally, the FDTD code is used to calculate the magnetic ellipticity response of a 2D survey and a 3D sounding of complicated geophysical targets. The response of these 2D and 3D targets are too complicated to be verified with integral equation solutions, but show the proper low- and high-frequency responses.

  13. It's the peptide-MHC affinity, stupid.

    Science.gov (United States)

    Kammertoens, Thomas; Blankenstein, Thomas

    2013-04-15

    Adoptively transferred T cells can reject large established tumors, but recurrence due to escape variants frequently occurs. In this issue of Cancer Cell, Engels et al. demonstrate that the affinity of the target peptide to the MHC molecule determines whether large tumors will relapse following adoptive T cell therapy. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. Affine field theories

    International Nuclear Information System (INIS)

    Cadavid, A.C.

    1989-01-01

    The author constructs a non-Abelian field theory by gauging a Kac-Moody algebra, obtaining an infinite tower of interacting vector fields and associated ghosts, that obey slightly modified Feynman rules. She discusses the spontaneous symmetry breaking of such theory via the Higgs mechanism. If the Higgs particle lies in the Cartan subalgebra of the Kac-Moody algebra, the previously massless vectors acquire a mass spectrum that is linear in the Kac-Moody index and has additional fine structure depending on the associated Lie algebra. She proceeds to show that there is no obstacle in implementing the affine extension of supersymmetric Yang-Mills theories. The result is valid in four, six and ten space-time dimensions. Then the affine extension of supergravity is investigated. She discusses only the loop algebra since the affine extension of the super-Poincare algebra appears inconsistent. The construction of the affine supergravity theory is carried out by the group manifold method and leads to an action describing infinite towers of spin 2 and spin 3/2 fields that interact subject to the symmetries of the loop algebra. The equations of motion satisfy the usual consistency check. Finally, she postulates a theory in which both the vector and scalar fields lie in the loop algebra of SO(3). This theory has an expanded soliton sector, and corresponding to the original 't Hooft-Polyakov solitonic solutions she now finds an infinite family of exact, special solutions of the new equations. She also proposes a perturbation method for obtaining an arbitrary solution of those equations for each level of the affine index

  15. Enhancing Natural Killer Cell Mediated Targeting and Responses to Myeloid Leukemias

    Science.gov (United States)

    2017-10-01

    AWARD NUMBER: W81XWH-16-1-0380 TITLE: Enhancing Natural Killer Cell Mediated Targeting and Responses to Myeloid Leukemias PRINCIPAL...2017 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Enhancing Natural Killer Cell Mediated Targeting and Responses to Myeloid Leukemias 5b. GRANT NUMBER...leukemias still have poor prognosis, particularly in the elderly, and require hematopoietic cell transplants to fully kill the tumor, which is both

  16. Electromagnetic modelling of Ground Penetrating Radar responses to complex targets

    Science.gov (United States)

    Pajewski, Lara; Giannopoulos, Antonis

    2014-05-01

    defined through a constant real value, or else its frequency-dispersion properties can be taken into account by incorporating into the model Debye approximations. The electromagnetic source can be represented as a simple line of current (in the case of two-dimensional models), a Hertzian dipole, a bow tie antenna, or else, the realistic description of a commercial antenna can be included in the model [2]. Preliminary results for some of the proposed cells are presented, obtained by using GprMax [3], a freeware tool which solves Maxwell's equations by using a second order in space and time Finite-Difference Time-Domain algorithm. B-Scans and A-Scans are calculated at 1.5 GHz, for the total electric field and for the field back-scattered by targets embedded in the cells. A detailed description of the structures, together with the relevant numerical results obtained to date, are available for the scientific community on the website of COST Action TU1208, www.GPRadar.eu. Research groups working on the development of electromagnetic forward- and inverse-scattering techniques, as well as on imaging methods, might test and compare the accuracy and applicability of their approaches on the proposed set of scenarios. The aim of this initiative is not that of identifying the best methods, but more properly to indicate the range of reliability of each approach, highlighting its advantages and drawbacks. In the future, the realisation of the proposed concrete cells and the acquisition of GPR experimental data would allow a very effective benchmark for forward and inverse scattering methods. References [1] R. Yelf, A. Ward, "Nine steps to concrete wisdom." Proc. 13th International Conference on Ground Penetrating Radar, Lecce, Italy, 21-25 June 2010, pp. 1-8. [2] C. Warren, A. Giannopoulos, "Creating FDTD models of commercial GPR antennas using Taguchi's optimisation method." Geophysics (2011), 76, article ID G37. [3] A. Giannopoulos, "Modelling ground penetrating radar by GPRMAX

  17. Bone metastasis target redox-responsive micell for the treatment of lung cancer bone metastasis and anti-bone resorption.

    Science.gov (United States)

    Ye, Wei-Liang; Zhao, Yi-Pu; Cheng, Ying; Liu, Dao-Zhou; Cui, Han; Liu, Miao; Zhang, Bang-Le; Mei, Qi-Bing; Zhou, Si-Yuan

    2018-01-16

    In order to inhibit the growth of lung cancer bone metastasis and reduce the bone resorption at bone metastasis sites, a bone metastasis target micelle DOX@DBMs-ALN was prepared. The size and the zeta potential of DOX@DBNs-ALN were about 60 nm and -15 mV, respectively. DOX@DBMs-ALN exhibited high binding affinity with hydroxyapatite and released DOX in redox-responsive manner. DOX@DBMs-ALN was effectively up taken by A549 cells and delivered DOX to the nucleus of A549 cells, which resulted in strong cytotoxicity on A549 cells. The in vivo experimental results indicated that DOX@DBMs-ALN specifically delivered DOX to bone metastasis site and obviously prolonged the retention time of DOX in bone metastasis site. Moreover, DOX@DBMs-ALN not only significantly inhibited the growth of bone metastasis tumour but also obviously reduced the bone resorption at bone metastasis sites without causing marked systemic toxicity. Thus, DOX@DBMs-ALN has great potential in the treatment of lung cancer bone metastasis.

  18. The specificity of targeted vaccines for APC surface molecules influences the immune response phenotype.

    Directory of Open Access Journals (Sweden)

    Gunnveig Grødeland

    Full Text Available Different diseases require different immune responses for efficient protection. Thus, prophylactic vaccines should prime the immune system for the particular type of response needed for protection against a given infectious agent. We have here tested fusion DNA vaccines which encode proteins that bivalently target influenza hemagglutinins (HA to different surface molecules on antigen presenting cells (APC. We demonstrate that targeting to MHC class II molecules predominantly induced an antibody/Th2 response, whereas targeting to CCR1/3/5 predominantly induced a CD8(+/Th1 T cell response. With respect to antibodies, the polarizing effect was even more pronounced upon intramuscular (i.m delivery as compared to intradermal (i.d. vaccination. Despite these differences in induced immune responses, both vaccines protected against a viral challenge with influenza H1N1. Substitution of HA with ovalbumin (OVA demonstrated that polarization of immune responses, as a consequence of APC targeting specificity, could be extended to other antigens. Taken together, the results demonstrate that vaccination can be tailor-made to induce a particular phenotype of adaptive immune responses by specifically targeting different surface molecules on APCs.

  19. Bidimensional microdosimetry as a tool for evaluating biological response and target structure

    International Nuclear Information System (INIS)

    Booz, J.; Schmitz, Th.; Feinendegen, L.E.; Olko, P.

    1992-01-01

    The paper addresses the issue of the relevance of microdosimetric spectra for quantifying the effects of low-level exposures to radiation. Biological response functions derived to date from numerical analyses of radiobiological and microdosimetric observations refer to uniform targets of a preassumed size. The characteristic two-modal shape of functions obtained for several endpoints reflects the importance of two different pathways of damage formation, each of them related in fact to different target sizes. The correlated energy deposition distributions in such a bidimensional system are suggested as a more appropriate physical input for analysing biological response and target structure. (author)

  20. Dynamics behind affinity maturation of an anti-HCMV antibody family influencing antigen binding

    KAUST Repository

    Di Palma, Francesco; Tramontano, Anna

    2017-01-01

    The investigation of antibody affinity maturation and its effects on antigen binding is important with respect to understanding the regulation of the immune response. To shed light on this crucial process, we analyzed two Igs neutralizing the human cytomegalovirus: the primary germline antibody M2J1 and its related mature antibody 8F9. Both antibodies target the AD-2S1 epitope of the gB envelope protein and are considered to establish similar interactions with the cognate antigen. We used molecular dynamics simulations to understand the effect of mutations on the antibody–antigen interactions. The results provide a qualitative explanation for the increased 8F9 peptide affinity compared with that of M2J1. The emerging atomistic-detailed description of these complexes reveals the molecular effects of the somatic hypermutations occurring during affinity maturation.

  1. Dynamics behind affinity maturation of an anti-HCMV antibody family influencing antigen binding

    KAUST Repository

    Di Palma, Francesco

    2017-08-03

    The investigation of antibody affinity maturation and its effects on antigen binding is important with respect to understanding the regulation of the immune response. To shed light on this crucial process, we analyzed two Igs neutralizing the human cytomegalovirus: the primary germline antibody M2J1 and its related mature antibody 8F9. Both antibodies target the AD-2S1 epitope of the gB envelope protein and are considered to establish similar interactions with the cognate antigen. We used molecular dynamics simulations to understand the effect of mutations on the antibody–antigen interactions. The results provide a qualitative explanation for the increased 8F9 peptide affinity compared with that of M2J1. The emerging atomistic-detailed description of these complexes reveals the molecular effects of the somatic hypermutations occurring during affinity maturation.

  2. Compound immobilization and drug-affinity chromatography.

    Science.gov (United States)

    Rix, Uwe; Gridling, Manuela; Superti-Furga, Giulio

    2012-01-01

    Bioactive small molecules act through modulating a yet unpredictable number of targets. It is therefore of critical importance to define the cellular target proteins of a compound as an entry point to understanding its mechanism of action. Often, this can be achieved in a direct fashion by chemical proteomics. As with any affinity chromatography, immobilization of the bait to a solid support is one of the earliest and most crucial steps in the process. Interfering with structural features that are important for identification of a target protein will be detrimental to binding affinity. Also, many molecules are sensitive to heat or to certain chemicals, such as acid or base, and might be destroyed during the process of immobilization, which therefore needs to be not only efficient, but also mild. The subsequent affinity chromatography step needs to preserve molecular and conformational integrity of both bait compound and proteins in order to result in the desired specific enrichment while ensuring a high level of compatibility with downstream analysis by mass spectrometry. Thus, the right choice of detergent, buffer, and protease inhibitors is also essential. This chapter describes a widely applicable procedure for the immobilization of small molecule drugs and for drug-affinity chromatography with subsequent protein identification by mass spectrometry.

  3. Quantotypic Properties of QconCAT Peptides Targeting Bovine Host Response to Streptococcus uberis

    DEFF Research Database (Denmark)

    Bislev, Stine Lønnerup; Kusebauch, Ulrike; Codrea, Marius Cosmin

    2012-01-01

    with host response to pathogens remains a challenging task. In this paper we present a targeted proteome analysis of a panel of 20 proteins that are widely believed to be key players and indicators of bovine host response to mastitis pathogens. Stable isotope labeled variants of two concordant proteotypic...

  4. Affine and quasi-affine frames for rational dilations

    DEFF Research Database (Denmark)

    Bownik, Marcin; Lemvig, Jakob

    2011-01-01

    In this paper we extend the investigation of quasi-affine systems, which were originally introduced by Ron and Shen [J. Funct. Anal. 148 (1997), 408-447] for integer, expansive dilations, to the class of rational, expansive dilations. We show that an affine system is a frame if, and only if......, the corresponding family of quasi-affine systems are frames with uniform frame bounds. We also prove a similar equivalence result between pairs of dual affine frames and dual quasi-affine frames. Finally, we uncover some fundamental differences between the integer and rational settings by exhibiting an example...

  5. Structural Basis of Low-Affinity Nickel Binding to the Nickel-Responsive Transcription Factor NikR from Escherichia coli

    International Nuclear Information System (INIS)

    Phillips, C.; Schreiter, E.; Stultz, C.; Drennan, C.

    2010-01-01

    Escherichia coli NikR regulates cellular nickel uptake by binding to the nik operon in the presence of nickel and blocking transcription of genes encoding the nickel uptake transporter. NikR has two binding affinities for the nik operon: a nanomolar dissociation constant with stoichiometric nickel and a picomolar dissociation constant with excess nickel (Bloom, S. L., and Zamble, D. B. (2004) Biochemistry 43, 10029-10038; Chivers, P. T., and Sauer, R. T. (2002) Chem. Biol. 9, 1141-1148). While it is known that the stoichiometric nickel ions bind at the NikR tetrameric interface (Schreiter, E. R., et al. (2003) Nat. Struct. Biol. 10, 794-799; Schreiter, E. R., et al. (2006) Proc. Natl. Acad. Sci. U.S.A. 103, 13676-13681), the binding sites for excess nickel ions have not been fully described. Here we have determined the crystal structure of NikR in the presence of excess nickel to 2.6 (angstrom) resolution and have obtained nickel anomalous data (1.4845 (angstrom)) in the presence of excess nickel for both NikR alone and NikR cocrystallized with a 30-nucleotide piece of double-stranded DNA containing the nik operon. These anomalous data show that excess nickel ions do not bind to a single location on NikR but instead reveal a total of 22 possible low-affinity nickel sites on the NikR tetramer. These sites, for which there are six different types, are all on the surface of NikR, and most are found in both the NikR alone and NikR-DNA structures. Using a combination of crystallographic data and molecular dynamics simulations, the nickel sites can be described as preferring octahedral geometry, utilizing one to three protein ligands (typically histidine) and at least two water molecules.

  6. Responses of female New World screwworm flies to coloured targets in the laboratory

    International Nuclear Information System (INIS)

    Green, C.H.; Warnes, M.L.

    1992-01-01

    The responses of unmated female New World screwworm flies, Cochliomyia hominivorax, to visual targets were studied in a wind-tunnel. Both activity and frequency of contacts with targets increased greatly when the screwworm attractant mixture swormlure-4 was added to the airstream. Target-orientated responses depended on target colour, with red and black targets being preferred over blue, white and yellow ones; this preference was much greater in the presence of odour than in its absence. No preference was detected for different shapes and orientations of red targets, all of equivalent surface area. Omitting different components from swormlure-4 generally resulted in a large reduction in activation and target contacts. Attempts to substitute 1-octen-3-o1 for the butanol fraction were unsuccessful, but skatole may substitute to some extent for indole; the two isomers of butanol normally present in swormlure-4 may substitute partly or completely for each other. This type of measurement forms a suitable bioassay in the development of attractive targets for monitoring and control of wild adult screwworm populations. 4 figs, 1 tab

  7. Engineering of bispecific affinity proteins with high affinity for ERBB2 and adaptable binding to albumin.

    Directory of Open Access Journals (Sweden)

    Johan Nilvebrant

    Full Text Available The epidermal growth factor receptor 2, ERBB2, is a well-validated target for cancer diagnostics and therapy. Recent studies suggest that the over-expression of this receptor in various cancers might also be exploited for antibody-based payload delivery, e.g. antibody drug conjugates. In such strategies, the full-length antibody format is probably not required for therapeutic effect and smaller tumor-specific affinity proteins might be an alternative. However, small proteins and peptides generally suffer from fast excretion through the kidneys, and thereby require frequent administration in order to maintain a therapeutic concentration. In an attempt aimed at combining ERBB2-targeting with antibody-like pharmacokinetic properties in a small protein format, we have engineered bispecific ERBB2-binding proteins that are based on a small albumin-binding domain. Phage display selection against ERBB2 was used for identification of a lead candidate, followed by affinity maturation using second-generation libraries. Cell surface display and flow-cytometric sorting allowed stringent selection of top candidates from pools pre-enriched by phage display. Several affinity-matured molecules were shown to bind human ERBB2 with sub-nanomolar affinity while retaining the interaction with human serum albumin. Moreover, parallel selections against ERBB2 in the presence of human serum albumin identified several amino acid substitutions that dramatically modulate the albumin affinity, which could provide a convenient means to control the pharmacokinetics. The new affinity proteins competed for ERBB2-binding with the monoclonal antibody trastuzumab and recognized the native receptor on a human cancer cell line. Hence, high affinity tumor targeting and tunable albumin binding were combined in one small adaptable protein.

  8. Targeting Plant Ethylene Responses by Controlling Essential Protein-Protein Interactions in the Ethylene Pathway.

    Science.gov (United States)

    Bisson, Melanie M A; Groth, Georg

    2015-08-01

    The gaseous plant hormone ethylene regulates many processes of high agronomic relevance throughout the life span of plants. A central element in ethylene signaling is the endoplasmic reticulum (ER)-localized membrane protein ethylene insensitive2 (EIN2). Recent studies indicate that in response to ethylene, the extra-membranous C-terminal end of EIN2 is proteolytically processed and translocated from the ER to the nucleus. Here, we report that the conserved nuclear localization signal (NLS) mediating nuclear import of the EIN2 C-terminus provides an important domain for complex formation with ethylene receptor ethylene response1 (ETR1). EIN2 lacking the NLS domain shows strongly reduced affinity for the receptor. Interaction of EIN2 and ETR1 is also blocked by a synthetic peptide of the NLS motif. The corresponding peptide substantially reduces ethylene responses in planta. Our results uncover a novel mechanism and type of inhibitor interfering with ethylene signal transduction and ethylene responses in plants. Disruption of essential protein-protein interactions in the ethylene signaling pathway as shown in our study for the EIN2-ETR1 complex has the potential to guide the development of innovative ethylene antagonists for modern agriculture and horticulture. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

  9. Antibody affinity maturation

    DEFF Research Database (Denmark)

    Skjødt, Mette Louise

    Yeast surface display is an effective tool for antibody affinity maturation because yeast can be used as an all-in-one workhorse to assemble, display and screen diversified antibody libraries. By employing the natural ability of yeast Saccharomyces cerevisiae to efficiently recombine multiple DNA...... laboratory conditions. A particular emphasis was put on using molecular techniques in conjunction with microenvironmental measurements (O2, pH, irradiance), a combination that is rarely found but provides a much more detailed understanding of “cause and effect” in complex natural systems...

  10. Using Culturally Responsive Stories in Mathematics: Responses from the Target Audience

    Science.gov (United States)

    Corp, Amy

    2017-01-01

    This study examined how Black students responded to the utilization of culturally responsive stories in their mathematics class. All students in the two classes participated in mathematics lessons that began with an African American story (culturally responsive to this population), followed by mathematical discussion and concluded with solving…

  11. Individualized therapies in colorectal cancer: KRAS as a marker for response to EGFR-targeted therapy

    Directory of Open Access Journals (Sweden)

    Li Kuiyuan

    2009-04-01

    Full Text Available Abstract Individualized therapies that are tailored to a patient's genetic composition will be of tremendous value for treatment of cancer. Recently, Kirsten ras (KRAS status has emerged as a predictor of response to epidermal growth factor receptor (EGFR targeted therapies. In this article, we will discuss targeted therapies for colorectal cancers (CRC based on EGFR signaling pathway and review published data about the potential usefulness of KRAS as a biological marker for response to these therapies. Results from relevant studies published since 2005 and unpublished results presented at national meetings were retrieved and summarized. These studies reflected response (or lack of response to EGFR-targeted therapies in patients with metastatic CRC as a function of KRAS status. It has become clear that patients with colorectal cancer whose tumor has an activating mutation in KRAS do not respond to monoclonal antibody therapies targeting EGFR. It should now become a standard practice that any patients being considered for EGFR targeted therapies have their tumors tested for KRAS status and only those with wild-type KRAS being offered such therapies.

  12. Impact velocity vs. target hardness relationships for equivalent response of cask structures

    International Nuclear Information System (INIS)

    Chen, T.F.; Chen, J.C.; Witte, M.C.; Fischer, L.E.

    1993-01-01

    In this paper, impact velocity vs. target hardness relationships for cask structures are reviewed. The relationships are based on equivalent cask responses in terms of equal deceleration or similar cask damages. By examining several past cask or container tests as well as some analytical results, some conclusions can be drawn about the relationship between target hardness and equivalent impact velocities. This relationship clearly shows that the cask response to impact is cask-dependent and that the rigid sphere impact model results in an unconservative estimate of equivalent velocity

  13. The Potential of Stimuli-Responsive Nanogels in Drug and Active Molecule Delivery for Targeted Therapy

    Directory of Open Access Journals (Sweden)

    Marta Vicario-de-la-Torre

    2017-05-01

    Full Text Available Nanogels (NGs are currently under extensive investigation due to their unique properties, such as small particle size, high encapsulation efficiency and protection of active agents from degradation, which make them ideal candidates as drug delivery systems (DDS. Stimuli-responsive NGs are cross-linked nanoparticles (NPs, composed of polymers, natural, synthetic, or a combination thereof that can swell by absorption (uptake of large amounts of solvent, but not dissolve due to the constituent structure of the polymeric network. NGs can undergo change from a polymeric solution (swell form to a hard particle (collapsed form in response to (i physical stimuli such as temperature, ionic strength, magnetic or electric fields; (ii chemical stimuli such as pH, ions, specific molecules or (iii biochemical stimuli such as enzymatic substrates or affinity ligands. The interest in NGs comes from their multi-stimuli nature involving reversible phase transitions in response to changes in the external media in a faster way than macroscopic gels or hydrogels due to their nanometric size. NGs have a porous structure able to encapsulate small molecules such as drugs and genes, then releasing them by changing their volume when external stimuli are applied.

  14. Paired hormone response elements predict caveolin-1 as a glucocorticoid target gene.

    Directory of Open Access Journals (Sweden)

    Marinus F van Batenburg

    2010-01-01

    Full Text Available Glucocorticoids act in part via glucocorticoid receptor binding to hormone response elements (HREs, but their direct target genes in vivo are still largely unknown. We developed the criterion that genomic occurrence of paired HREs at an inter-HRE distance less than 200 bp predicts hormone responsiveness, based on synergy of multiple HREs, and HRE information from known target genes. This criterion predicts a substantial number of novel responsive genes, when applied to genomic regions 10 kb upstream of genes. Multiple-tissue in situ hybridization showed that mRNA expression of 6 out of 10 selected genes was induced in a tissue-specific manner in mice treated with a single dose of corticosterone, with the spleen being the most responsive organ. Caveolin-1 was strongly responsive in several organs, and the HRE pair in its upstream region showed increased occupancy by glucocorticoid receptor in response to corticosterone. Our approach allowed for discovery of novel tissue specific glucocorticoid target genes, which may exemplify responses underlying the permissive actions of glucocorticoids.

  15. Antigen-targeting strategies using single-domain antibody fragments

    NARCIS (Netherlands)

    Duarte, Joao Nuno Silva

    2017-01-01

    Antibodies display high selectivity and affinity and have been the preferred platform for antigen targeting. Despite the development of antigen-delivery systems that enable T cell activation, targeting approaches that enhance antibody responses need improvement. This need specially applies to poorly

  16. The solutions of affine and conformal affine Toda field theory

    International Nuclear Information System (INIS)

    Papadopoulos, G.; Spence, B.

    1994-02-01

    We give new formulations of the solutions of the field equations of the affine Toda and conformal affine Toda theories on a cylinder and two-dimensional Minkowski space-time. These solutions are parameterised in terms of initial data and the resulting covariant phase spaces are diffeomorphic to the Hamiltonian ones. We derive the fundamental Poisson brackets of the parameters of the solutions and give the general static solutions for the affine theory. (authors). 10 refs

  17. Effect of a viscoelastic target on the impact response of a flat-nosed projectile

    Science.gov (United States)

    Liu, Hu; Yang, Jialing; Liu, Hua

    2018-02-01

    Taylor impact is a widely used strategy in which a flat-nosed projectile is fired onto a rigid anvil directly to determine the dynamic strength of rod specimens. Nowadays, the rigid anvil is often replaced by an output target bar to ensure the accuracy of measurement via recording strain signals in the output bar. For testing the dynamic strength of low-density materials, a low-impedance target bar, which exhibits viscoelastic characteristics is often employed. In this paper, an extended Taylor model is proposed to improve the idealization of treating the target bar as perfectly rigid material in the classic Taylor model, and the viscoelastic effect of the target bar is incorporated. The viscoelastic target bar is depicted by two elastic springs and one dashpot. Based on the plastic shock wave theory in the flat-nosed projectile associated with the viscoelastic wave analysis in the target bar, the viscoelastic effect of the target bar on the impact response of the flat-nosed projectile is investigated. The finite element simulation is also carried out to verify the theoretical model, and good agreement is found. The present theoretical model is also called the Taylor-cylinder Hopkinson impact, which provides a more accurate way to identify the dynamic material parameters. The dynamic responses of the present model are further compared with previous elastic and rigid target bar models. It is found that the viscoelastic effect of the target bar should be taken into consideration in the Taylor-cylinder Hopkinson impact test for low-impedance materials.

  18. MicroRNA-target gene responses to lead-induced stress in cotton (Gossypium hirsutum L.).

    Science.gov (United States)

    He, Qiuling; Zhu, Shuijin; Zhang, Baohong

    2014-09-01

    MicroRNAs (miRNAs) play key roles in plant responses to various metal stresses. To investigate the miRNA-mediated plant response to heavy metals, cotton (Gossypium hirsutum L.), the most important fiber crop in the world, was exposed to different concentrations (0, 25, 50, 100, and 200 µM) of lead (Pb) and then the toxicological effects were investigated. The expression patterns of 16 stress-responsive miRNAs and 10 target genes were monitored in cotton leaves and roots by quantitative real-time PCR (qRT-PCR); of these selected genes, several miRNAs and their target genes are involved in root development. The results show a reciprocal regulation of cotton response to lead stress by miRNAs. The characterization of the miRNAs and the associated target genes in response to lead exposure would help in defining the potential roles of miRNAs in plant adaptation to heavy metal stress and further understanding miRNA regulation in response to abiotic stress.

  19. Response of Solid and Liquid Targets to High Power Proton Beams for Neutrino Factories

    CERN Document Server

    Sievers, P

    2000-01-01

    The response of solid and liquid targets to rapid heating by the incident proton beam is assessed in a classical way, among other things by solving the wave equation under linear conditions and in cylindrical symmetry. This study provides bench mark values and allows to identify critical issues and limiting factors which can help to guide further investigations with more sophisticated means.

  20. Doing worse, but feeling happy : Social comparison and identification in response to upward and downward targets

    NARCIS (Netherlands)

    Groothof, H.; Siero, F.W.; Buunk, Abraham (Bram)

    2007-01-01

    We investigated people's responses to exposure to downward and upward targets. In Study 1, among 197 participants, it was predicted and found that such exposure led to a contrast effect on self-evaluation, and to an assimilation effect on affect. In Study 2, among 148 participants, it was predicted

  1. Fundamentals of affinity cell separations.

    Science.gov (United States)

    Zhang, Ye; Lyons, Veronica; Pappas, Dimitri

    2018-03-01

    Cell separations using affinity methods continue to be an enabling science for a wide variety of applications. In this review, we discuss the fundamental aspects of affinity separation, including the competing forces for cell capture and elution, cell-surface interactions, and models for cell adhesion. Factors affecting separation performance such as bond affinity, contact area, and temperature are presented. We also discuss and demonstrate the effects of nonspecific binding on separation performance. Metrics for evaluating cell separations are presented, along with methods of comparing separation techniques for cell isolation using affinity capture. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Assessing the response to targeted therapies in renal cell carcinoma: technical insights and practical considerations.

    Science.gov (United States)

    Bex, Axel; Fournier, Laure; Lassau, Nathalie; Mulders, Peter; Nathan, Paul; Oyen, Wim J G; Powles, Thomas

    2014-04-01

    The introduction of targeted agents for the treatment of renal cell carcinoma (RCC) has resulted in new challenges for assessing response to therapy, and conventional response criteria using computed tomography (CT) are limited. It is widely recognised that targeted therapies may lead to significant necrosis without significant reduction in tumour size. In addition, the vascular effects of antiangiogenic therapy may occur long before there is any reduction in tumour size. To perform a systematic review of conventional and novel imaging methods for the assessment of response to targeted agents in RCC and to discuss their use from a clinical perspective. Relevant databases covering the period January 2006 to April 2013 were searched for studies reporting on the use of anatomic and functional imaging techniques to predict response to targeted therapy in RCC. Inclusion criteria were randomised trials, nonrandomised controlled studies, retrospective case series, and cohort studies. Reviews, animal and preclinical studies, case reports, and commentaries were excluded. A narrative synthesis of the evidence is presented. A total of 331 abstracts and 76 full-text articles were assessed; 34 studies met the inclusion criteria. Current methods of response assessment in RCC include anatomic methods--based on various criteria including Choi, size and attenuation CT, and morphology, attenuation, size, and structure--and functional techniques including dynamic contrast-enhanced (DCE) CT, DCE-magnetic resonance imaging, DCE-ultrasonography, positron emission tomography, and approaches utilising radiolabelled monoclonal antibodies. Functional imaging techniques are promising surrogate biomarkers of response in RCC and may be more appropriate than anatomic CT-based methods. By enabling quantification of tumour vascularisation, functional techniques can directly and rapidly detect the biologic effects of antiangiogenic therapies compared with the indirect detection of belated effects

  3. CERN antiproton target: Hydrocode analysis of its core material dynamic response under proton beam impact

    Directory of Open Access Journals (Sweden)

    Claudio Torregrosa Martin

    2016-07-01

    Full Text Available Antiprotons are produced at CERN by colliding a 26  GeV/c proton beam with a fixed target made of a 3 mm diameter, 55 mm length iridium core. The inherent characteristics of antiproton production involve extremely high energy depositions inside the target when impacted by each primary proton beam, making it one of the most dynamically demanding among high energy solid targets in the world, with a rise temperature above 2000 °C after each pulse impact and successive dynamic pressure waves of the order of GPa’s. An optimized redesign of the current target is foreseen for the next 20 years of operation. As a first step in the design procedure, this numerical study delves into the fundamental phenomena present in the target material core under proton pulse impact and subsequent pressure wave propagation by the use of hydrocodes. Three major phenomena have been identified, (i the dominance of a high frequency radial wave which produces destructive compressive-to-tensile pressure response (ii The existence of end-of-pulse tensile waves and its relevance on the overall response (iii A reduction of 44% in tensile pressure could be obtained by the use of a high density tantalum cladding.

  4. Artificial ground motion compatible with specified peak ground displacement and target multi-damping response spectra

    International Nuclear Information System (INIS)

    Zhang Yushan; Zhao Fengxin

    2010-01-01

    With respect to the design ground motion of nuclear power plant (NPP), the Regular Guide 1.60 of the US not only defined the standard multi-damping response spectra, i.e. the RG1.60 spectra, but also definitely prescribed the peak ground displacement (PGD) value corresponding to the standard spectra. However, in the engineering practice of generating multi-damping-spectra-compatible artificial ground motion for the seismic design of NPP, the PGD value had been neglected. Addressing this issue, this paper proposed a synthesizing method which generates the artificial ground motion compatible with not only the target multi-damping response spectra but also the specified PGD value. Firstly, by the transfer formula between the power spectrum and the response spectrum, an initial uniformly modulated acceleration time history is synthesized by multiplying the stationary Gaussian process with the prescribed intensity envelope to simulate the amplitude-non-stationarity of earthquake ground motion. And then by superimposing a series of narrow-band time histories in the time domain, the initial time history is modified in the iterative manner to match the target PGD as well as the target multi-damping spectra with the pre-specified matching precisions. Numerical examples are provided to demonstrate the matching precisions of the proposed method to the target values.

  5. Dynamic Nucleolar Targeting of Dengue Virus Polymerase NS5 in Response to Extracellular pH

    Science.gov (United States)

    Fraser, Johanna E.; Rawlinson, Stephen M.; Heaton, Steven M.

    2016-01-01

    ABSTRACT The nucleolar subcompartment of the nucleus is increasingly recognized as an important target of RNA viruses. Here we document for the first time the ability of dengue virus (DENV) polymerase, nonstructural protein 5 (NS5), to accumulate within the nucleolus of infected cells and to target green fluorescent protein (GFP) to the nucleolus of live transfected cells. Intriguingly, NS5 exchange between the nucleus and nucleolus is dynamically modulated by extracellular pH, responding rapidly and reversibly to pH change, in contrast to GFP alone or other nucleolar and non-nucleolar targeted protein controls. The minimal pH-sensitive nucleolar targeting region (pHNTR), sufficient to target GFP to the nucleolus in a pH-sensitive fashion, was mapped to NS5 residues 1 to 244, with mutation of key hydrophobic residues, Leu-165, Leu-167, and Val-168, abolishing pHNTR function in NS5-transfected cells, and severely attenuating DENV growth in infected cells. This is the first report of a viral protein whose nucleolar targeting ability is rapidly modulated by extracellular stimuli, suggesting that DENV has the ability to detect and respond dynamically to the extracellular environment. IMPORTANCE Infections by dengue virus (DENV) threaten 40% of the world's population yet there is no approved vaccine or antiviral therapeutic to treat infections. Understanding the molecular details that govern effective viral replication is key for the development of novel antiviral strategies. Here, we describe for the first time dynamic trafficking of DENV nonstructural protein 5 (NS5) to the subnuclear compartment, the nucleolus. We demonstrate that NS5's targeting to the nucleolus occurs in response to acidic pH, identify the key amino acid residues within NS5 that are responsible, and demonstrate that their mutation severely impairs production of infectious DENV. Overall, this study identifies a unique subcellular trafficking event and suggests that DENV is able to detect and respond

  6. Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

    Energy Technology Data Exchange (ETDEWEB)

    Muhsain, Siti Nur Fadzilah, E-mail: sitinurfadzilah077@ppinang.uitm.edu.my [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Faculty of Pharmacy, University Teknologi Mara (Malaysia); Lang, Matti A., E-mail: m.lang@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Abu-Bakar, A' edah, E-mail: a.abubakar@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia)

    2015-01-01

    The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

  7. Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

    International Nuclear Information System (INIS)

    Muhsain, Siti Nur Fadzilah; Lang, Matti A.; Abu-Bakar, A'edah

    2015-01-01

    The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

  8. Age-related changes in gait adaptability in response to unpredictable obstacles and stepping targets.

    Science.gov (United States)

    Caetano, Maria Joana D; Lord, Stephen R; Schoene, Daniel; Pelicioni, Paulo H S; Sturnieks, Daina L; Menant, Jasmine C

    2016-05-01

    A large proportion of falls in older people occur when walking. Limitations in gait adaptability might contribute to tripping; a frequently reported cause of falls in this group. To evaluate age-related changes in gait adaptability in response to obstacles or stepping targets presented at short notice, i.e.: approximately two steps ahead. Fifty older adults (aged 74±7 years; 34 females) and 21 young adults (aged 26±4 years; 12 females) completed 3 usual gait speed (baseline) trials. They then completed the following randomly presented gait adaptability trials: obstacle avoidance, short stepping target, long stepping target and no target/obstacle (3 trials of each). Compared with the young, the older adults slowed significantly in no target/obstacle trials compared with the baseline trials. They took more steps and spent more time in double support while approaching the obstacle and stepping targets, demonstrated poorer stepping accuracy and made more stepping errors (failed to hit the stepping targets/avoid the obstacle). The older adults also reduced velocity of the two preceding steps and shortened the previous step in the long stepping target condition and in the obstacle avoidance condition. Compared with their younger counterparts, the older adults exhibited a more conservative adaptation strategy characterised by slow, short and multiple steps with longer time in double support. Even so, they demonstrated poorer stepping accuracy and made more stepping errors. This reduced gait adaptability may place older adults at increased risk of falling when negotiating unexpected hazards. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Hemoglobin affinity in Andean rodents

    Directory of Open Access Journals (Sweden)

    HRVOJ OSTOJIC

    2002-01-01

    Full Text Available Blood hemoglobin oxygen affinity (P50 was measured in three Andean species and in the laboratory rat (control, all raised near sea level. Chinchilla lanigera (Molina, 1792 has an altitudinal habitat range from low Andean slopes up to 3000 m., while Chinchilla brevicaudata (Waterhouse, 1848 has an altitudinal range from 3000 to 5000 m. The laboratory type guinea pig, wild type guinea pig (Cavia porcellus, (Waterhouse, 1748, and laboratory rat (Rattus norvegicus were also raised at sea level. The Andean species had high hemoglobin oxygen affinities (low P50 compared with the rat. Chinchilla brevicaudata had a higher affinity than Chinchilla lanigera. The wild type guinea pig had a higher affinity than the laboratory type. As has been shown in other species, this is another example of an inverse correlation between the altitude level and the P50 values. This is the first hemoglobin oxygen affinity study in Chinchilla brevicaudata.

  10. Vascular endothelium as a target of immune response in renal transplant rejection

    Directory of Open Access Journals (Sweden)

    Giovanni ePiotti

    2014-10-01

    Full Text Available This review of clinical and experimental studies aims at analysing the interplay between graft endothelium and host immune system in renal transplantation, and how it affects the survival of the graft. Graft endothelium is indeed the first barrier between self and non-self that is encountered by host lymphocytes upon reperfusion of vascularised solid transplants. Endothelial cells express all the major sets of antigens that elicit host immune response, and therefore represent a preferential target in organ rejection.Some of the antigens expressed by endothelial cells are target of the antibody-mediated response, such as the AB0 blood group system, the HLA and MICA systems, and the endothelial cell-restricted antigens; for each of these systems, the mechanisms of interaction and damage of both preformed and de novo donor-specific antibodies are reviewed along with their impact on renal graft survival. Moreover the rejection process can force injured endothelial cells to expose cryptic self-antigens, toward which an auto-immune response mounts, overlapping to the allo-immune response in the damaging of the graft. Not only are endothelial cells a passive target of the host immune response, but also an active player in lymphocyte activation; therefore their interaction with allogenic T-cells is analysed on the basis of experimental in vitro and in vivo studies, according to the patterns of expression of the HLA class I and II and the co-stimulatory molecules specific for cytotoxic and helper T-cells.Finally, as the response that follows transplantation has proven to be not necessarily destructive, the factors that foster graft endothelium functioning in spite of rejection, and how they could be therapeutically harnessed to promote long-term graft acceptance, are described: accommodation that is resistance of endothelial cells to donor-specific antibodies, and endothelial cell ability to induce Foxp3+ Regulatory T-cells, that are crucial mediators of

  11. Advances in Targeted Pesticides with Environmentally Responsive Controlled Release by Nanotechnology

    Directory of Open Access Journals (Sweden)

    Bingna Huang

    2018-02-01

    Full Text Available Pesticides are the basis for defending against major biological disasters and important for ensuring national food security. Biocompatible, biodegradable, intelligent, and responsive materials are currently an emerging area of interest in the field of efficient, safe, and green pesticide formulation. Using nanotechnology to design and prepare targeted pesticides with environmentally responsive controlled release via compound and chemical modifications has also shown great potential in creating novel formulations. In this review, special attention has been paid to intelligent pesticides with precise controlled release modes that can respond to micro-ecological environment changes such as light-sensitivity, thermo-sensitivity, humidity sensitivity, soil pH, and enzyme activity. Moreover, establishing intelligent and controlled pesticide release technologies using nanomaterials are reported. These technologies could increase pesticide-loading, improve the dispersibility and stability of active ingredients, and promote target ability.

  12. Identification of treatment response predictors and potential molecular targets for chemo preventive and antiangiogenic therapies

    International Nuclear Information System (INIS)

    Pfeffer, U.; Albini, A.

    2009-01-01

    The aims of the project were: To evaluate the cellular responses to anti-inflammatory and anti-angiogenic natural or synthetic compounds (chemo preventives, inhibitors of cell survival and inflammation related signal transduction). To identify bio markers for treatment response through the selection of targets that are common to or specific for anti-inflammatory and anti-angiogenic activities. To analyze the regulation of the key tumor-promotion pathways Akt, HIF1α, NFκB. We focused our studies on the antiapoptotic role of the AKT survival pathway, which is involved in prostate tumor progression to an androgen-independent phenotype

  13. Modifying chemotherapy response by targeted inhibition of eukaryotic initiation factor 4A

    International Nuclear Information System (INIS)

    Cencic, R; Robert, F; Galicia-Vázquez, G; Malina, A; Ravindar, K; Somaiah, R; Pierre, P; Tanaka, J; Deslongchamps, P; Pelletier, J

    2013-01-01

    Translation is regulated predominantly at the initiation phase by several signal transduction pathways that are often usurped in human cancers, including the PI3K/Akt/mTOR axis. mTOR exerts unique administration over translation by regulating assembly of eukaryotic initiation factor (eIF) 4F, a heterotrimeric complex responsible for recruiting 40S ribosomes (and associated factors) to mRNA 5′ cap structures. Hence, there is much interest in targeted therapies that block eIF4F activity to assess the consequences on tumor cell growth and chemotherapy response. We report here that hippuristanol (Hipp), a translation initiation inhibitor that selectively inhibits the eIF4F RNA helicase subunit, eIF4A, resensitizes Eμ-Myc lymphomas to DNA damaging agents, including those that overexpress eIF4E—a modifier of rapamycin responsiveness. As Mcl-1 levels are significantly affected by Hipp, combining its use with the Bcl-2 family inhibitor, ABT-737, leads to a potent synergistic response in triggering cell death in mouse and human lymphoma and leukemia cells. Suppression of eIF4AI using RNA interference also synergized with ABT-737 in murine lymphomas, highlighting eIF4AI as a therapeutic target for modulating tumor cell response to chemotherapy

  14. Developing Demand-Response Based Solutions for Hawaii’s 100% Renewable Energy Target

    OpenAIRE

    Kansal, Rachit

    2017-01-01

    The State of Hawaii has set a target to achieve a 100% Renewables by 2045. Due to the State’s high electricity prices and dependence on imported oil, renewables are seen as an environmental and economic solution to the problem. While the state has seen substantial renewables growth in the last few years, a truly transformative system is needed to push for a fully renewable future. This system would be likely to include Demand Response (DR) capability, Distributed Energy Reso...

  15. A target based approach identifies genomic predictors of breast cancer patient response to chemotherapy

    Directory of Open Access Journals (Sweden)

    Hallett Robin M

    2012-05-01

    Full Text Available Abstract Background The efficacy of chemotherapy regimens in breast cancer patients is variable and unpredictable. Whether individual patients either achieve long-term remission or suffer recurrence after therapy may be dictated by intrinsic properties of their breast tumors including genetic lesions and consequent aberrant transcriptional programs. Global gene expression profiling provides a powerful tool to identify such tumor-intrinsic transcriptional programs, whose analyses provide insight into the underlying biology of individual patient tumors. For example, multi-gene expression signatures have been identified that can predict the likelihood of disease reccurrence, and thus guide patient prognosis. Whereas such prognostic signatures are being introduced in the clinical setting, similar signatures that predict sensitivity or resistance to chemotherapy are not currently clinically available. Methods We used gene expression profiling to identify genes that were co-expressed with genes whose transcripts encode the protein targets of commonly used chemotherapeutic agents. Results Here, we present target based expression indices that predict breast tumor response to anthracycline and taxane based chemotherapy. Indeed, these signatures were independently predictive of chemotherapy response after adjusting for standard clinic-pathological variables such as age, grade, and estrogen receptor status in a cohort of 488 breast cancer patients treated with adriamycin and taxotere/taxol. Conclusions Importantly, our findings suggest the practicality of developing target based indices that predict response to therapeutics, as well as highlight the possibility of using gene signatures to guide the use of chemotherapy during treatment of breast cancer patients.

  16. Surface Modified Multifunctional and Stimuli Responsive Nanoparticles for Drug Targeting: Current Status and Uses

    Directory of Open Access Journals (Sweden)

    Panoraia I. Siafaka

    2016-08-01

    Full Text Available Nanocarriers, due to their unique features, are of increased interest among researchers working with pharmaceutical formulations. Polymeric nanoparticles and nanocapsules, involving non-toxic biodegradable polymers, liposomes, solid lipid nanoparticles, and inorganic–organic nanomaterials, are among the most used carriers for drugs for a broad spectrum of targeted diseases. In fact, oral, injectable, transdermal-dermal and ocular formulations mainly consist of the aforementioned nanomaterials demonstrating promising characteristics such as long circulation, specific targeting, high drug loading capacity, enhanced intracellular penetration, and so on. Over the last decade, huge advances in the development of novel, safer and less toxic nanocarriers with amended properties have been made. In addition, multifunctional nanocarriers combining chemical substances, vitamins and peptides via coupling chemistry, inorganic particles coated by biocompatible materials seem to play a key role considering that functionalization can enhance characteristics such as biocompatibility, targetability, environmental friendliness, and intracellular penetration while also have limited side effects. This review aims to summarize the “state of the art” of drug delivery carriers in nanosize, paying attention to their surface functionalization with ligands and other small or polymeric compounds so as to upgrade active and passive targeting, different release patterns as well as cell targeting and stimuli responsibility. Lastly, future aspects and potential uses of nanoparticulated drug systems are outlined.

  17. Reaching Hard-to-Reach Individuals: Nonselective Versus Targeted Outbreak Response Vaccination for Measles

    Science.gov (United States)

    Minetti, Andrea; Hurtado, Northan; Grais, Rebecca F.; Ferrari, Matthew

    2014-01-01

    Current mass vaccination campaigns in measles outbreak response are nonselective with respect to the immune status of individuals. However, the heterogeneity in immunity, due to previous vaccination coverage or infection, may lead to potential bias of such campaigns toward those with previous high access to vaccination and may result in a lower-than-expected effective impact. During the 2010 measles outbreak in Malawi, only 3 of the 8 districts where vaccination occurred achieved a measureable effective campaign impact (i.e., a reduction in measles cases in the targeted age groups greater than that observed in nonvaccinated districts). Simulation models suggest that selective campaigns targeting hard-to-reach individuals are of greater benefit, particularly in highly vaccinated populations, even for low target coverage and with late implementation. However, the choice between targeted and nonselective campaigns should be context specific, achieving a reasonable balance of feasibility, cost, and expected impact. In addition, it is critical to develop operational strategies to identify and target hard-to-reach individuals. PMID:24131555

  18. Molecular Imaging to Predict Response to Targeted Therapies in Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Ingrid Leguerney

    2017-01-01

    Full Text Available Molecular magnetic resonance imaging targeted to an endothelial integrin involved in neoangiogenesis was compared to DCE-US and immunochemistry to assess the early response of three different therapeutic agents in renal cell carcinoma. Human A498 renal cells carcinoma was subcutaneously inoculated into 24 nude mice. Mice received either phosphate-buffered saline solution, sunitinib, everolimus, or bevacizumab during 4 days. DCE-US and molecular MRI targeting αvβ3 were performed at baseline and 4 days after treatment initiation. PI, AUC, relaxation rate variations ΔR2⁎, and percentage of vessels area quantified on CD31-stained microvessels were compared. Significant decreases were observed for PI and AUC parameters measured by DCE-US for bevacizumab group as early as 4 days, whereas molecular αvβ3-targeted MRI was able to detect significant changes in both bevacizumab and everolimus groups. Percentage of CD31-stained microvessels was significantly correlated with DCE-US parameters, PI (R=0.87, p=0.0003 and AUC (R=0.81, p=0.0013. The percentage of vessel tissue area was significantly reduced (p<0.01 in both sunitinib and bevacizumab groups. We report an early detection of neoangiogenesis modification after induction of targeted therapies, using DCE-US or αvβ3-targeted MRI. We consider these outcomes should encourage clinical trial developments to further evaluate the potential of this molecular MRI technique.

  19. The CERN antiproton target: hydrocode analysis of its core material dynamic response under proton beam impact

    CERN Document Server

    Martin, Claudio Torregrosa; Calviani, Marco; Muñoz-Cobo, José-Luis

    2016-01-01

    Antiprotons are produced at CERN by colliding a 26 GeV/c proton beam with a fixed target made of a 3 mm diameter, 55 mm length iridium core. The inherent characteristics of antiproton production involve extremely high energy depositions inside the target when impacted by each primary proton beam, making it one of the most dynamically demanding among high energy solid targets in the world, with a rise temperature above 2000 {\\deg}C after each pulse impact and successive dynamic pressure waves of the order of GPa's. An optimized redesign of the current target is foreseen for the next 20 years of operation. As a first step in the design procedure, this numerical study delves into the fundamental phenomena present in the target material core under proton pulse impact and subsequent pressure wave propagation by the use of hydrocodes. Three major phenomena have been identified, (i) the dominance of a high frequency radial wave which produces destructive compressive-to-tensile pressure response (ii) The existence of...

  20. Profiling the Targets of Protective CD8+ T Cell Responses to Infection

    Directory of Open Access Journals (Sweden)

    Joseph T. Bruder

    2017-12-01

    Full Text Available T cells are critical effectors of host immunity that target intracellular pathogens, such as the causative agents of HIV, tuberculosis, and malaria. The development of vaccines that induce effective cell-mediated immunity against such pathogens has proved challenging; for tuberculosis and malaria, many of the antigens targeted by protective T cells are not known. Here, we report a novel approach for screening large numbers of antigens as potential targets of T cells. Malaria provides an excellent model to test this antigen discovery platform because T cells are critical mediators of protection following immunization with live sporozoite vaccines and the specific antigen targets are unknown. We generated an adenovirus array by cloning 312 highly expressed pre-erythrocytic Plasmodium yoelii antigens into adenovirus vectors using high-throughput methodologies. The array was screened to identify antigen-specific CD8+ T cells induced by a live sporozoite vaccine regimen known to provide high levels of sterile protection mediated by CD8+ T cells. We identified 69 antigens that were targeted by CD8+ T cells induced by this vaccine regimen. The antigen that recalled the highest frequency of CD8+ T cells, PY02605, induced protective responses in mice, demonstrating proof of principle for this approach in identifying antigens for vaccine development.

  1. Circular revisit orbits design for responsive mission over a single target

    Science.gov (United States)

    Li, Taibo; Xiang, Junhua; Wang, Zhaokui; Zhang, Yulin

    2016-10-01

    The responsive orbits play a key role in addressing the mission of Operationally Responsive Space (ORS) because of their capabilities. These capabilities are usually focused on supporting specific targets as opposed to providing global coverage. One subtype of responsive orbits is repeat coverage orbit which is nearly circular in most remote sensing applications. This paper deals with a special kind of repeating ground track orbit, referred to as circular revisit orbit. Different from traditional repeat coverage orbits, a satellite on circular revisit orbit can visit a target site at both the ascending and descending stages in one revisit cycle. This typology of trajectory allows a halving of the traditional revisit time and does a favor to get useful information for responsive applications. However the previous reported numerical methods in some references often cost lots of computation or fail to obtain such orbits. To overcome this difficulty, an analytical method to determine the existence conditions of the solutions to revisit orbits is presented in this paper. To this end, the mathematical model of circular revisit orbit is established under the central gravity model and the J2 perturbation. A constraint function of the circular revisit orbit is introduced, and the monotonicity of that function has been studied. The existent conditions and the number of such orbits are naturally worked out. Taking the launch cost into consideration, optimal design model of circular revisit orbit is established to achieve a best orbit which visits a target twice a day in the morning and in the afternoon respectively for several days. The result shows that it is effective to apply circular revisit orbits in responsive application such as reconnoiter of natural disaster.

  2. Non-proinflammatory and responsive nanoplatforms for targeted treatment of atherosclerosis.

    Science.gov (United States)

    Dou, Yin; Chen, Yue; Zhang, Xiangjun; Xu, Xiaoqiu; Chen, Yidan; Guo, Jiawei; Zhang, Dinglin; Wang, Ruibing; Li, Xiaohui; Zhang, Jianxiang

    2017-10-01

    Atherosclerosis is the leading cause of many fatal cardiovascular and cerebrovascular diseases. Whereas nanomedicines are promising for targeted therapy of atherosclerosis, great challenges remain in development of effective, safe, and translational nanotherapies for its treatment. Herein we hypothesize that non-proinflammatory nanomaterials sensitive to low pH or high reactive oxygen species (ROS) may serve as effective platforms for triggerable delivery of anti-atherosclerotic therapeutics in cellular and tissue microenvironments of inflammation. To demonstrate this hypothesis, an acid-labile material of acetalated β-cyclodextrin (β-CD) (Ac-bCD) and a ROS-sensitive β-CD material (Ox-bCD) were separately synthesized by chemical modification of β-CD, which were formed into responsive nanoparticles (NPs). Ac-bCD NP was rapidly hydrolyzed in mildly acidic buffers, while hydrolysis of Ox-bCD NP was selectively accelerated by H 2 O 2 . Using an anti-atherosclerotic drug rapamycin (RAP), we found stimuli-responsive release of therapeutic molecules from Ac-bCD and Ox-bCD nanotherapies. Compared with non-responsive poly(lactide-co-glycolide) (PLGA)-based NP, Ac-bCD and Ox-bCD NPs showed negligible inflammatory responses in vitro and in vivo. By endocytosis in cells and intracellularly releasing cargo molecules in macrophages, responsive nanotherapies effectively inhibited macrophage proliferation and suppressed foam cell formation. After intraperitoneal (i.p.) delivery in apolipoprotein E-deficient (ApoE -/- ) mice, fluorescence imaging showed accumulation of NPs in atherosclerotic plaques. Flow cytometry analysis indicated that the lymphatic translocation mediated by neutrophils and monocytes/macrophages may contribute to atherosclerosis targeting of i.p. administered NPs, in addition to targeting via the leaky blood vessels. Correspondingly, i.p. treatment with different nanotherapies afforded desirable efficacies. Particularly, both pH and ROS-responsive

  3. Lp-dual affine surface area

    Science.gov (United States)

    Wei, Wang; Binwu, He

    2008-12-01

    According to the notion of Lp-affine surface area by Lutwak, in this paper, we introduce the concept of Lp-dual affine surface area. Further, we establish the affine isoperimetric inequality and the Blaschke-Santaló inequality for Lp-dual affine surface area. Besides, the dual Brunn-Minkowski inequality for Lp-dual affine surface area is presented.

  4. Affinity Purification and Comparative Biosensor Analysis of Citrulline-Peptide-Specific Antibodies in Rheumatoid Arthritis

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    Eszter Szarka

    2018-01-01

    Full Text Available Background: In rheumatoid arthritis (RA, anti-citrullinated protein/peptide antibodies (ACPAs are responsible for disease onset and progression, however, our knowledge is limited on ligand binding affinities of autoantibodies with different citrulline-peptide specificity. Methods: Citrulline-peptide-specific ACPA IgGs were affinity purified and tested by ELISA. Binding affinities of ACPA IgGs and serum antibodies were compared by surface plasmon resonance (SPR analysis. Bifunctional nanoparticles harboring a multi-epitope citrulline-peptide and a complement-activating peptide were used to induce selective depletion of ACPA-producing B cells. Results: KD values of affinity-purified ACPA IgGs varied between 10−6 and 10−8 M and inversely correlated with disease activity. Based on their cross-reaction with citrulline-peptides, we designed a novel multi-epitope peptide, containing Cit-Gly and Ala-Cit motifs in two–two copies, separated with a short, neutral spacer. This peptide detected antibodies in RA sera with 66% sensitivity and 98% specificity in ELISA and was recognized by 90% of RA sera, while none of the healthy samples in SPR. When coupled to nanoparticles, the multi-epitope peptide specifically targeted and depleted ACPA-producing B cells ex vivo. Conclusions: The unique multi-epitope peptide designed based on ACPA cross-reactivity might be suitable to develop better diagnostics and novel therapies for RA.

  5. The utility of affine variables and affine coherent states

    International Nuclear Information System (INIS)

    Klauder, John R

    2012-01-01

    Affine coherent states are generated by affine kinematical variables much like canonical coherent states are generated by canonical kinematical variables. Although all classical and quantum formalisms normally entail canonical variables, it is shown that affine variables can serve equally well for many classical and quantum studies. This general purpose analysis provides tools to discuss two major applications: (1) the completely successful quantization of a nonrenormalizable scalar quantum field theory by affine techniques, in complete contrast to canonical techniques which only offer triviality; and (2) a formulation of the kinematical portion of quantum gravity that favors affine kinematical variables over canonical kinematical variables, and which generates a framework in which a favorable analysis of the constrained dynamical issues can take place. All this is possible because of the close connection between the affine and the canonical stories, while the few distinctions can be used to advantage when appropriate. This article is part of a special issue of Journal of Physics A: Mathematical and Theoretical devoted to ‘Coherent states: mathematical and physical aspects’. (review)

  6. The effects of incidentally learned temporal and spatial predictability on response times and visual fixations during target detection and discrimination.

    Directory of Open Access Journals (Sweden)

    Melissa R Beck

    Full Text Available Responses are quicker to predictable stimuli than if the time and place of appearance is uncertain. Studies that manipulate target predictability often involve overt cues to speed up response times. However, less is known about whether individuals will exhibit faster response times when target predictability is embedded within the inter-trial relationships. The current research examined the combined effects of spatial and temporal target predictability on reaction time (RT and allocation of overt attention in a sustained attention task. Participants responded as quickly as possible to stimuli while their RT and eye movements were measured. Target temporal and spatial predictability were manipulated by altering the number of: 1 different time intervals between a response and the next target; and 2 possible spatial locations of the target. The effects of target predictability on target detection (Experiment 1 and target discrimination (Experiment 2 were tested. For both experiments, shorter RTs as target predictability increased across both space and time were found. In addition, the influences of spatial and temporal target predictability on RT and the overt allocation of attention were task dependent; suggesting that effective orienting of attention relies on both spatial and temporal predictability. These results indicate that stimulus predictability can be increased without overt cues and detected purely through inter-trial relationships over the course of repeated stimulus presentations.

  7. Chamber wall response to target implosion in inertial fusion reactors : new and critical assessments

    International Nuclear Information System (INIS)

    Hassanein, A.; Morozov, V.

    2002-01-01

    The chamber walls in inertial fusion energy (IFE) reactors are exposed to harsh conditions following each target implosion. Key issues of the cyclic IFE operation include intense photon and ion deposition, wall thermal and hydrodynamic evolution, wall erosion and fatigue lifetime, and chamber clearing and evacuation to ensure desirable conditions prior to target implosion. Several methods for wall protection have been proposed in the past, each having its own advantages and disadvantages. These methods include use of solid bare walls, gas-filled cavities, and liquid walls/jets. Detailed models have been developed for reflected laser light, emitted photons, and target debris deposition and interaction with chamber components and have been implemented in the comprehensive HEIGHTS software package. The hydrodynamic response of gas filled cavities and photon radiation transport of the deposited energy has been calculated by means of new and advanced numerical techniques. Fragmentation models of liquid jets as a result of the deposited energy have also been developed, and the impact on chamber clearing dynamics has been evaluated. Th focus of this study is to critically assess the reliability and the dynamic response of chamber walls in various proposed protection methods for IFE systems. Of particular concern is the effect on wall erosion lifetime of various erosion mechanisms, such as vaporization, chemical and physical sputtering, melt/liquid splashing and explosive erosion, and fragmentation of liquid walls

  8. Hyaluronic acid oligosaccharide modified redox-responsive mesoporous silica nanoparticles for targeted drug delivery.

    Science.gov (United States)

    Zhao, Qinfu; Geng, Hongjian; Wang, Ying; Gao, Yikun; Huang, Jiahao; Wang, Yan; Zhang, Jinghai; Wang, Siling

    2014-11-26

    A redox-responsive delivery system based on colloidal mesoporous silica (CMS) has been developed, in which 6-mercaptopurine (6-MP) was conjugated to vehicles by cleavable disulfide bonds. The oligosaccharide of hyaluronic acid (oHA) was modified on the surface of CMS by disulfide bonds as a targeting ligand and was able to increase the stability and biocompatibility of CMS under physiological conditions. In vitro release studies indicated that the cumulative release of 6-MP was less than 3% in the absence of glutathione (GSH), and reached nearly 80% within 2 h in the presence of 3 mM GSH. Confocal microscopy and fluorescence-activated cell sorter (FACS) methods were used to evaluate the cellular uptake performance of fluorescein isothiocyanate (FITC) labeled CMS, with and without oHA modification. The CMS-SS-oHA exhibited a higher cellular uptake performance via CD44 receptor-mediated endocytosis in HCT-116 (CD44 receptor-positive) cells than in NIH-3T3 (CD44 receptor-negative) cells. 6-MP loaded CMS-SS-oHA exhibited greater cytotoxicity against HCT-116 cells than NIH-3T3 cells due to the enhanced cell uptake behavior of CMS-SS-oHA. This study provides a novel strategy to covalently link bioactive drug and targeting ligand to the interiors and exteriors of mesoporous silica to construct a stimulus-responsive targeted drug delivery system.

  9. Representations of affine Hecke algebras

    CERN Document Server

    Xi, Nanhua

    1994-01-01

    Kazhdan and Lusztig classified the simple modules of an affine Hecke algebra Hq (q E C*) provided that q is not a root of 1 (Invent. Math. 1987). Ginzburg had some very interesting work on affine Hecke algebras. Combining these results simple Hq-modules can be classified provided that the order of q is not too small. These Lecture Notes of N. Xi show that the classification of simple Hq-modules is essentially different from general cases when q is a root of 1 of certain orders. In addition the based rings of affine Weyl groups are shown to be of interest in understanding irreducible representations of affine Hecke algebras. Basic knowledge of abstract algebra is enough to read one third of the book. Some knowledge of K-theory, algebraic group, and Kazhdan-Lusztig cell of Cexeter group is useful for the rest

  10. Preclinical Evaluation and Monitoring of the Therapeutic Response of a Dual Targeted Hyaluronic Acid Nanodrug

    Directory of Open Access Journals (Sweden)

    Minglong Chen

    2017-01-01

    Full Text Available Chemotherapy is a powerful cancer treatment but suffers from poor biocompatibility and a lack of tumor targeting. Here, we developed a CD44-targeted polymeric nanocomplex by encapsulating 10-hydroxycamptothecin (HCPT into hyaluronic acid nanoparticles (HANP for targeted cancer therapy. In vitro, the HANP/HCPT showed improved cytotoxicity to five cancer cell lines including HT29, A549, MDA-MB-231, HepG2, and MDA-MB-435 versus free HCPT. After systemic administration into MDA-MB-231 breast cancer xenograft, tumor growth was significantly inhibited 5.25 ± 0.21 times in the HANP/HCPT treated group relative to the nontreated group. In addition, the treatment response was also accessed and confirmed by 18F-fluoro-2-deoxy-D-glucose ([18F] FDG positron emission tomography (PET. The MDA-MB-231 tumors responded to HANP/HCPT 7 days after the first treatment, which benefits treatment strategy adjustment and personalization. No apparent systemic toxic effects were seen in mice treated with HANP/HCPT. In summary, the HANPs have great promise as a targeted drug carrier for cancer chemotherapy. Our HANP platform can also deliver other hydrophobic chemotherapy agents.

  11. MRI-guided targeting delivery of doxorubicin with reduction-responsive lipid-polymer hybrid nanoparticles

    Directory of Open Access Journals (Sweden)

    Wu B

    2017-09-01

    Full Text Available Bo Wu,1,2 Shu-Ting Lu,1 Kai Deng,2 Hui Yu,2 Can Cui,2 Yang Zhang,2 Ming Wu,2 Ren-Xi Zhuo,2 Hai-Bo Xu,1 Shi-Wen Huang2 1Department of Radiology, Zhongnan Hospital of Wuhan University, 2Key Laboratory of Biomedical Polymers, Ministry of Education, Department of Chemistry, Wuhan University, Wuhan, People’s Republic of China Abstract: In recent years, there has been increasing interest in developing a multifunctional nanoscale platform for cancer monitoring and chemotherapy. However, there is still a big challenge for current clinic contrast agents to improve their poor tumor selectivity and response. Herein, we report a new kind of Gd complex and folate-coated redox-sensitive lipid-polymer hybrid nanoparticle (Gd-FLPNP for tumor-targeted magnetic resonance imaging and therapy. Gd-FLPNPs can simultaneously accomplish diagnostic imaging, and specific targeting and controlled release of doxorubicin (DOX. They exhibit good monodispersity, excellent size stability, and a well-defined core-shell structure. Paramagnetic nanoparticles based on gadolinium-diethylenetriaminepentaacetic acid-bis-cetylamine have paramagnetic properties with an approximately two-fold enhancement in the longitudinal relaxivity compared to clinical used Magnevist. For targeted and reduction-sensitive drug delivery, Gd-FLPNPs released DOX faster and enhanced cell uptake in vitro, and exhibited better antitumor effect both in vitro and in vivo. Keywords: redox-sensitive, tumor-targeted, gadolinium, contrast agents, PLGA

  12. Contractions of affine spherical varieties

    International Nuclear Information System (INIS)

    Arzhantsev, I V

    1999-01-01

    The language of filtrations and contractions is used to describe the class of G-varieties obtainable as the total spaces of the construction of contraction applied to affine spherical varieties, which is well-known in invariant theory. These varieties are local models for arbitrary affine G-varieties of complexity 1 with a one-dimensional categorical quotient. As examples, reductive algebraic semigroups and three-dimensional SL 2 -varieties are considered

  13. The application of magnetic force differentiation for the measurement of the affinity of peptide libraries

    International Nuclear Information System (INIS)

    Shang Hao; Kirkham, Perry M.; Myers, Tina M.; Cassell, Gail H.; Lee, Gil U.

    2005-01-01

    A new method has been developed for measuring the binding affinity of phage displayed peptides and a target protein using magnetic particles. The specific interaction between the phage displayed peptides and the target protein was subject to a force generated by the magnetic particle. The binding affinity was obtained by analyzing the force-bond lifetime

  14. Cellular Pathways in Response to Ionizing Radiation and Their Targetability for Tumor Radiosensitization

    Directory of Open Access Journals (Sweden)

    Patrick Maier

    2016-01-01

    Full Text Available During the last few decades, improvements in the planning and application of radiotherapy in combination with surgery and chemotherapy resulted in increased survival rates of tumor patients. However, the success of radiotherapy is impaired by two reasons: firstly, the radioresistance of tumor cells and, secondly, the radiation-induced damage of normal tissue cells located in the field of ionizing radiation. These limitations demand the development of drugs for either radiosensitization of tumor cells or radioprotection of normal tissue cells. In order to identify potential targets, a detailed understanding of the cellular pathways involved in radiation response is an absolute requirement. This review describes the most important pathways of radioresponse and several key target proteins for radiosensitization.

  15. Target of Rapamycin (TOR) Regulates Growth in Response to Nutritional Signals.

    Science.gov (United States)

    Weisman, Ronit

    2016-10-01

    All organisms can respond to the availability of nutrients by regulating their metabolism, growth, and cell division. Central to the regulation of growth in response to nutrient availability is the target of rapamycin (TOR) signaling that is composed of two structurally distinct complexes: TOR complex 1 (TORC1) and TOR complex 2 (TORC2). The TOR genes were first identified in yeast as target of rapamycin, a natural product of a soil bacterium, which proved beneficial as an immunosuppressive and anticancer drug and is currently being tested for a handful of other pathological conditions including diabetes, neurodegeneration, and age-related diseases. Studies of the TOR pathway unraveled a complex growth-regulating network. TOR regulates nutrient uptake, transcription, protein synthesis and degradation, as well as metabolic pathways, in a coordinated manner that ensures that cells grow or cease growth in response to nutrient availability. The identification of specific signals and mechanisms that stimulate TOR signaling is an active and exciting field of research that has already identified nitrogen and amino acids as key regulators of TORC1 activity. The signals, as well as the cellular functions of TORC2, are far less well understood. Additional open questions in the field concern the relationships between TORC1 and TORC2, as well as the links with other nutrient-responsive pathways. Here I review the main features of TORC1 and TORC2, with a particular focus on yeasts as model organisms.

  16. A gene expression profile indicative of early stage HER2 targeted therapy response.

    Science.gov (United States)

    O'Neill, Fiona; Madden, Stephen F; Clynes, Martin; Crown, John; Doolan, Padraig; Aherne, Sinéad T; O'Connor, Robert

    2013-07-01

    Efficacious application of HER2-targetting agents requires the identification of novel predictive biomarkers. Lapatinib, afatinib and neratinib are tyrosine kinase inhibitors (TKIs) of HER2 and EGFR growth factor receptors. A panel of breast cancer cell lines was treated with these agents, trastuzumab, gefitinib and cytotoxic therapies and the expression pattern of a specific panel of genes using RT-PCR was investigated as a potential marker of early drug response to HER2-targeting therapies. Treatment of HER2 TKI-sensitive SKBR3 and BT474 cell lines with lapatinib, afatinib and neratinib induced an increase in the expression of RB1CC1, ERBB3, FOXO3a and NR3C1. The response directly correlated with the degree of sensitivity. This expression pattern switched from up-regulated to down-regulated in the HER2 expressing, HER2-TKI insensitive cell line MDAMB453. Expression of the CCND1 gene demonstrated an inversely proportional response to drug exposure. A similar expression pattern was observed following the treatment with both neratinib and afatinib. These patterns were retained following exposure to traztuzumab and lapatinib plus capecitabine. In contrast, gefitinib, dasatinib and epirubicin treatment resulted in a completely different expression pattern change. In these HER2-expressing cell line models, lapatinib, neratinib, afatinib and trastuzumab treatment generated a characteristic and specific gene expression response, proportionate to the sensitivity of the cell lines to the HER2 inhibitor.Characterisation of the induced changes in expression levels of these genes may therefore give a valuable, very early predictor of the likely extent and specificity of tumour HER2 inhibitor response in patients, potentially guiding more specific use of these agents.

  17. Weak affinity chromatography for evaluation of stereoisomers in early drug discovery.

    Science.gov (United States)

    Duong-Thi, Minh-Dao; Bergström, Maria; Fex, Tomas; Svensson, Susanne; Ohlson, Sten; Isaksson, Roland

    2013-07-01

    In early drug discovery (e.g., in fragment screening), recognition of stereoisomeric structures is valuable and guides medicinal chemists to focus only on useful configurations. In this work, we concurrently screened mixtures of stereoisomers and estimated their affinities to a protein target (thrombin) using weak affinity chromatography-mass spectrometry (WAC-MS). Affinity determinations by WAC showed that minor changes in stereoisomeric configuration could have a major impact on affinity. The ability of WAC-MS to provide instant information about stereoselectivity and binding affinities directly from analyte mixtures is a great advantage in fragment library screening and drug lead development.

  18. Targeted Proteomics to Assess the Response to Anti-Angiogenic Treatment in Human Glioblastoma (GBM).

    Science.gov (United States)

    Demeure, Kevin; Fack, Fred; Duriez, Elodie; Tiemann, Katja; Bernard, Amandine; Golebiewska, Anna; Bougnaud, Sébastien; Bjerkvig, Rolf; Domon, Bruno; Niclou, Simone P

    2016-02-01

    Glioblastoma (GBM) is a highly aggressive primary brain tumor with dismal outcome for affected patients. Because of the significant neo-angiogenesis exhibited by GBMs, anti-angiogenic therapies have been intensively evaluated during the past years. Recent clinical studies were however disappointing, although a subpopulation of patients may benefit from such treatment. We have previously shown that anti-angiogenic targeting in GBM increases hypoxia and leads to a metabolic adaptation toward glycolysis, suggesting that combination treatments also targeting the glycolytic phenotype may be effective in GBM patients. The aim of this study was to identify marker proteins that are altered by treatment and may serve as a short term readout of anti-angiogenic therapy. Ultimately such proteins could be tested as markers of efficacy able to identify patient subpopulations responsive to the treatment. We applied a proteomics approach based on selected reaction monitoring (SRM) to precisely quantify targeted protein candidates, selected from pathways related to metabolism, apoptosis and angiogenesis. The workflow was developed in the context of patient-derived intracranial GBM xenografts developed in rodents and ensured the specific identification of human tumor versus rodent stroma-derived proteins. Quality control experiments were applied to assess sample heterogeneity and reproducibility of SRM assays at different levels. The data demonstrate that tumor specific proteins can be precisely quantified within complex biological samples, reliably identifying small concentration differences induced by the treatment. In line with previous work, we identified decreased levels of TCA cycle enzymes, including isocitrate dehydrogenase, whereas malectin, calnexin, and lactate dehydrogenase A were augmented after treatment. We propose the most responsive proteins of our subset as potential novel biomarkers to assess treatment response after anti-angiogenic therapy that warrant future

  19. Training motor responses to food: A novel treatment for obesity targeting implicit processes.

    Science.gov (United States)

    Stice, Eric; Lawrence, Natalia S; Kemps, Eva; Veling, Harm

    2016-11-01

    The present review first summarizes results from prospective brain imaging studies focused on identifying neural vulnerability factors that predict excessive weight gain. Next, findings from cognitive psychology experiments evaluating various interventions involving food response inhibition training or food response facilitation training are reviewed that appear to target these neural vulnerability factors and that have produced encouraging weight loss effects. Findings from both of these reviewed research fields suggest that interventions that reduce reward and attention region responses to high calorie food cues and increase inhibitory region responses to high calorie food cues could prove useful in the treatment of obesity. Based on this review, a new conceptual model is presented to describe how different cognitive training procedures may contribute to modifying eating behavior and important directions for future research are offered. It is concluded that there is a need for evaluating the effectiveness of more intensive food response training interventions and testing whether adding such training to extant weight loss interventions increases their efficacy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Methods for quantifying T cell receptor binding affinities and thermodynamics

    Science.gov (United States)

    Piepenbrink, Kurt H.; Gloor, Brian E.; Armstrong, Kathryn M.; Baker, Brian M.

    2013-01-01

    αβ T cell receptors (TCRs) recognize peptide antigens bound and presented by class I or class II major histocompatibility complex (MHC) proteins. Recognition of a peptide/MHC complex is required for initiation and propagation of a cellular immune response, as well as the development and maintenance of the T cell repertoire. Here we discuss methods to quantify the affinities and thermodynamics of interactions between soluble ectodomains of TCRs and their peptide/MHC ligands, focusing on titration calorimetry, surface plasmon resonance, and fluorescence anisotropy. As TCRs typically bind ligand with weak-to-moderate affinities, we focus the discussion on means to enhance the accuracy and precision of low affinity measurements. In addition to further elucidating the biology of the T cell mediated immune response, more reliable low affinity measurements will aid with more probing studies with mutants or altered peptides that can help illuminate the physical underpinnings of how TCRs achieve their remarkable recognition properties. PMID:21609868

  1. DNA demethylases target promoter transposable elements to positively regulate stress responsive genes in Arabidopsis.

    Science.gov (United States)

    Le, Tuan-Ngoc; Schumann, Ulrike; Smith, Neil A; Tiwari, Sameer; Au, Phil Chi Khang; Zhu, Qian-Hao; Taylor, Jennifer M; Kazan, Kemal; Llewellyn, Danny J; Zhang, Ren; Dennis, Elizabeth S; Wang, Ming-Bo

    2014-09-17

    DNA demethylases regulate DNA methylation levels in eukaryotes. Arabidopsis encodes four DNA demethylases, DEMETER (DME), REPRESSOR OF SILENCING 1 (ROS1), DEMETER-LIKE 2 (DML2), and DML3. While DME is involved in maternal specific gene expression during seed development, the biological function of the remaining DNA demethylases remains unclear. We show that ROS1, DML2, and DML3 play a role in fungal disease resistance in Arabidopsis. A triple DNA demethylase mutant, rdd (ros1 dml2 dml3), shows increased susceptibility to the fungal pathogen Fusarium oxysporum. We identify 348 genes differentially expressed in rdd relative to wild type, and a significant proportion of these genes are downregulated in rdd and have functions in stress response, suggesting that DNA demethylases maintain or positively regulate the expression of stress response genes required for F. oxysporum resistance. The rdd-downregulated stress response genes are enriched for short transposable element sequences in their promoters. Many of these transposable elements and their surrounding sequences show localized DNA methylation changes in rdd, and a general reduction in CHH methylation, suggesting that RNA-directed DNA methylation (RdDM), responsible for CHH methylation, may participate in DNA demethylase-mediated regulation of stress response genes. Many of the rdd-downregulated stress response genes are downregulated in the RdDM mutants nrpd1 and nrpe1, and the RdDM mutants nrpe1 and ago4 show enhanced susceptibility to F. oxysporum infection. Our results suggest that a primary function of DNA demethylases in plants is to regulate the expression of stress response genes by targeting promoter transposable element sequences.

  2. Genome-wide dynamics of a bacterial response to antibiotics that target the cell envelope

    Directory of Open Access Journals (Sweden)

    Tran Ngat

    2011-05-01

    Full Text Available Abstract Background A decline in the discovery of new antibacterial drugs, coupled with a persistent rise in the occurrence of drug-resistant bacteria, has highlighted antibiotics as a diminishing resource. The future development of new drugs with novel antibacterial activities requires a detailed understanding of adaptive responses to existing compounds. This study uses Streptomyces coelicolor A3(2 as a model system to determine the genome-wide transcriptional response following exposure to three antibiotics (vancomycin, moenomycin A and bacitracin that target distinct stages of cell wall biosynthesis. Results A generalised response to all three antibiotics was identified which involves activation of transcription of the cell envelope stress sigma factor σE, together with elements of the stringent response, and of the heat, osmotic and oxidative stress regulons. Attenuation of this system by deletion of genes encoding the osmotic stress sigma factor σB or the ppGpp synthetase RelA reduced resistance to both vancomycin and bacitracin. Many antibiotic-specific transcriptional changes were identified, representing cellular processes potentially important for tolerance to each antibiotic. Sensitivity studies using mutants constructed on the basis of the transcriptome profiling confirmed a role for several such genes in antibiotic resistance, validating the usefulness of the approach. Conclusions Antibiotic inhibition of bacterial cell wall biosynthesis induces both common and compound-specific transcriptional responses. Both can be exploited to increase antibiotic susceptibility. Regulatory networks known to govern responses to environmental and nutritional stresses are also at the core of the common antibiotic response, and likely help cells survive until any specific resistance mechanisms are fully functional.

  3. Epitope characterization of the ADA response directed against a targeted immunocytokine.

    Science.gov (United States)

    Stubenrauch, Kay; Künzel, Christian; Vogel, Rudolf; Tuerck, Dietrich; Schick, Eginhard; Heinrich, Julia

    2015-10-10

    Targeted immunocytokines (TICs) display potent activity in selective tumor suppression. This class of multi domain biotherapeutics (MDBs) is composed of the three major domains Fab, Fc, and a cytokine which may induce a complex polyclonal anti-drug antibody (ADA) response. However, classical ADA assays usually are not suitable to specify ADAs and to identify the immunogenic domains of a TIC. The purpose of the present study was to establish epitope characterization of ADA responses in order to specify immunogenic responses against a TIC and their direct impact on the pharmacokinetic profile, safety, and efficacy. Based on standard ADA screening and confirmation assays, respectively, domain detection assays (DDAs) and domain competition assays (DCAs) were established and compared by the use of 12 ADA-positive samples obtained from a cynomolgus monkey study in early development. Both domain-specific assays were sensitive enough to preserve the positive screening assay result and revealed an overall accordance for the evaluation of domain-specific ADA responses. About half of the samples displayed one ADA specificity, either for the Fab or for the cytokine (Cy) domain, and the remaining samples showed a combination of Fab-specific and Cy-specific ADA fractions. Fc-specific ADAs occurred in only one sample. In-depth comparison of DCAs and DDAs showed that both assays appeared to be appropriate to assess multi-specific ADA responses as well as minor ADA fractions. An advantage of DCAs is typically a fast and easy assay establishment, whereas, DDAs in some cases may be superior to assess low abundant ADAs in multi-specific responses. Our results reveal that both approaches benefit from thorough reagent development as an essential precondition for reliable epitope characterization of ADA responses. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Viral Response to Specifically Targeted Antiviral Therapy for Hepatitis C and the Implications for Treatment Success

    Directory of Open Access Journals (Sweden)

    Curtis L Cooper

    2010-01-01

    Full Text Available Currently, hepatitis C virus (HCV antiviral therapy is characterized by long duration, a multitude of side effects, difficult administration and suboptimal success; clearly, alternatives are needed. Collectively, specifically targeted antiviral therapy for HCV (STAT-C molecules achieve rapid viral suppression and very high rapid virological response rates, and improve sustained virological response rates. The attrition rate of agents within this class has been high due to various toxicities. Regardless, several STAT-C molecules are poised to become the standard of care for HCV treatment in the foreseeable future. Optimism must be tempered with concerns related to the rapid development of drug resistance with resulting HCV rebound. Strategies including induction dosing with interferon and ribavirin, use of combination high-potency STAT-C molecules and an intensive emphasis on adherence to HCV antiviral therapy will be critical to the success of this promising advance in HCV therapy.

  5. Dose response of bone-targeted enzyme replacement for murine hypophosphatasia.

    Science.gov (United States)

    Yadav, Manisha C; Lemire, Isabelle; Leonard, Pierre; Boileau, Guy; Blond, Laurent; Beliveau, Martin; Cory, Esther; Sah, Robert L; Whyte, Michael P; Crine, Philippe; Millán, José Luis

    2011-08-01

    Hypophosphatasia (HPP) features rickets or osteomalacia from tissue-nonspecific alkaline phosphatase (TNSALP) deficiency due to deactivating mutations within the ALPL gene. Enzyme replacement therapy with a bone-targeted, recombinant TNSALP (sALP-FcD(10), renamed ENB-0040) prevents manifestations of HPP when initiated at birth in TNSALP knockout (Akp2(-/-)) mice. Here, we evaluated the dose-response relationship of ENB-0040 to various phenotypic traits of Akp2(-/-) mice receiving daily subcutaneous (SC) injections of ENB-0040 from birth at 0.5, 2.0, or 8.2mg/kg for 43days. Radiographs, μCT, and histomorphometric analyses documented better bone mineralization with increasing doses of ENB-0040. We found a clear, positive correlation between ENB-0040 dose and prevention of mineralization defects of the feet, rib cage, lower limbs, and jaw bones. According to a dose-response model, the ED(80) (the dose that prevents bone defects in 80% of mice) was 3.2, 2.8 and 2.9mg/kg/day for these sites, respectively. Long bones seemed to respond to lower daily doses of ENB-0040. There was also a positive relationship between ENB-0040 dose and survival. Median survival, body weight, and bone length all improved with increasing doses of ENB-0040. Urinary PP(i) concentrations remained elevated in all treatment groups, indicating that while this parameter is a good biochemical marker for diagnosing HPP in patients, it may not be a good follow up marker for evaluating response to treatment when administering bone-targeted TNSALP to mice. These dose-response relationships strongly support the pharmacological efficacy of ENB-0040 for HPP, and provide the experimental basis for the therapeutic range of ENB-0040 chosen for clinical trials. Copyright © 2011 Elsevier Inc. All rights reserved.

  6. Augmentation of radiation response with the vascular targeting agent ZD6126

    International Nuclear Information System (INIS)

    Hoang Tien; Huang Shyhmin; Armstrong, Eric; Eickhoff, Jens C.; Harari, Paul M.

    2006-01-01

    Purpose: To examine the antivascular and antitumor activity of the vascular targeting agent ZD6126 in combination with radiation in lung and head-and-neck (H and N) cancer models. The overall hypothesis was that simultaneous targeting of tumor cells (radiation) and tumor vasculature (ZD6126) might enhance tumor cell killing. Methods and Materials: A series of in vitro studies using human umbilical vein endothelial cells (HUVEC) and in vivo studies in athymic mice bearing human lung (H226) and H and N (squamous cell carcinoma [SCC]1, SCC6) tumor xenografts treated with ZD6126 and/or radiation were performed. Results: ZD6126 inhibited the capillary-like network formation in HUVEC. Treatment of HUVEC with ZD6126 resulted in cell cycle arrest in G2/M, with decrease of cells in S phase and proliferation inhibition in a dose-dependent manner. ZD6126 augmented the cell-killing effect of radiation and radiation-induced apoptosis in HUVEC. The combination of ZD6126 and radiation further decreased tumor vascularization in an in vivo Matrigel angiogenesis assay. In tumor xenografts, ZD6126 enhanced the antitumor activity of radiation, resulting in tumor growth delay. Conclusions: These preclinical studies suggest that ZD6126 can augment the radiation response of proliferating endothelial H and N and lung cancer cells. These results complement recent reports suggesting the potential value of combining radiation with vascular targeting/antiangiogenic agents

  7. Mechanisms and biomaterials in pH-responsive tumour targeted drug delivery: A review.

    Science.gov (United States)

    Kanamala, Manju; Wilson, William R; Yang, Mimi; Palmer, Brian D; Wu, Zimei

    2016-04-01

    As the mainstay in the treatment of various cancers, chemotherapy plays a vital role, but still faces many challenges, such as poor tumour selectivity and multidrug resistance (MDR). Targeted drug delivery using nanotechnology has provided a new strategy for addressing the limitations of the conventional chemotherapy. In the last decade, the volume of research published in this area has increased tremendously, especially with functional nano drug delivery systems (nanocarriers). Coupling a specific stimuli-triggered drug release mechanism with these delivery systems is one of the most prevalent approaches for improving therapeutic outcomes. Among the various stimuli, pH triggered delivery is regarded as the most general strategy, targeting the acidic extracellular microenvironment and intracellular organelles of solid tumours. In this review, we discuss recent advances in the development of pH-sensitive nanocarriers for tumour-targeted drug delivery. The review focuses on the chemical design of pH-sensitive biomaterials, which are used to fabricate nanocarriers for extracellular and/or intracellular tumour site-specific drug release. The pH-responsive biomaterials bring forth conformational changes in these nanocarriers through various mechanisms such as protonation, charge reversal or cleavage of a chemical bond, facilitating tumour specific cell uptake or drug release. A greater understanding of these mechanisms will help to design more efficient drug delivery systems to address the challenges encountered in conventional chemotherapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. MicroRNA-302 Cluster Downregulates Enterovirus 71-Induced Innate Immune Response by Targeting KPNA2.

    Science.gov (United States)

    Peng, Nanfang; Yang, Xuecheng; Zhu, Chengliang; Zhou, Li; Yu, Haisheng; Li, Mengqi; Lin, Yong; Wang, Xueyu; Li, Qian; She, Yinglong; Wang, Jun; Zhao, Qian; Lu, Mengji; Zhu, Ying; Liu, Shi

    2018-05-18

    Enterovirus 71 (EV71) induces significantly elevated levels of cytokines and chemokines, leading to local or systemic inflammation and severe complications. As shown in our previous study, microRNA (miR) 302c regulates influenza A virus-induced IFN expression by targeting NF-κB-inducing kinase. However, little is known about the role of the miR-302 cluster in EV71-mediated proinflammatory responses. In this study, we found that the miR-302 cluster controls EV71-induced cytokine expression. Further studies demonstrated that karyopherin α2 (KPNA2) is a direct target of the miR-302 cluster. Interestingly, we also found that EV71 infection upregulates KPNA2 expression by downregulating miR-302 cluster expression. Upon investigating the mechanisms behind this event, we found that KPNA2 intracellularly associates with JNK1/JNK2 and p38, leading to translocation of those transcription factors from the cytosol into the nucleus. In EV71-infected patients, miR-302 cluster expression was downregulated and KPNA2 expression was upregulated compared with controls, and their expression levels were closely correlated. Taken together, our work establishes a link between the miR-302/ KPNA2 axis and EV71-induced cytokine expression and represents a promising target for future antiviral therapy. Copyright © 2018 by The American Association of Immunologists, Inc.

  9. Quantitation without Calibration: Response Profile as an Indicator of Target Amount.

    Science.gov (United States)

    Debnath, Mrittika; Farace, Jessica M; Johnson, Kristopher D; Nesterova, Irina V

    2018-06-21

    Quantitative assessment of biomarkers is essential in numerous contexts from decision-making in clinical situations to food quality monitoring to interpretation of life-science research findings. However, appropriate quantitation techniques are not as widely addressed as detection methods. One of the major challenges in biomarker's quantitation is the need to have a calibration for correlating a measured signal to a target amount. The step complicates the methodologies and makes them less sustainable. In this work we address the issue via a new strategy: relying on position of response profile rather than on an absolute signal value for assessment of a target's amount. In order to enable the capability we develop a target-probe binding mechanism based on a negative cooperativity effect. A proof-of-concept example demonstrates that the model is suitable for quantitative analysis of nucleic acids over a wide concentration range. The general principles of the platform will be applicable toward a variety of biomarkers such as nucleic acids, proteins, peptides, and others.

  10. Chamber wall response to target implosion in inertial fusion reactors: new and critical assessments

    International Nuclear Information System (INIS)

    Hassanein, A.; Morozov, V.

    2002-01-01

    The chamber walls in inertial fusion energy (IFE) reactors are exposed to harsh conditions following each target implosion. Key issues of the cyclic IFE operation include intense photon and ion deposition, wall thermal and hydrodynamic evolution, wall erosion and fatigue lifetime, and chamber clearing and evacuation to ensure desirable conditions prior to next target implosion. Several methods for wall protection have been proposed in the past, each having its own advantages and disadvantages. These methods include use of solid bare walls, gas-filled cavities, and liquid walls/jets. Detailed models have been developed for reflected laser light, emitted photons, and target debris deposition and interaction with chamber components and have been implemented in the comprehensive HEIGHTS software package. The focus of this study is to critically assess the reliability and the dynamic response of chamber walls in IFE systems. Of particular concern is the effect on wall erosion lifetime due to various erosion mechanisms, such as vaporization, chemical and physical sputtering, melt/liquid splashing and explosive erosion, and fragmentation of liquid walls

  11. MRI-guided targeting delivery of doxorubicin with reduction-responsive lipid-polymer hybrid nanoparticles.

    Science.gov (United States)

    Wu, Bo; Lu, Shu-Ting; Deng, Kai; Yu, Hui; Cui, Can; Zhang, Yang; Wu, Ming; Zhuo, Ren-Xi; Xu, Hai-Bo; Huang, Shi-Wen

    2017-01-01

    In recent years, there has been increasing interest in developing a multifunctional nanoscale platform for cancer monitoring and chemotherapy. However, there is still a big challenge for current clinic contrast agents to improve their poor tumor selectivity and response. Herein, we report a new kind of Gd complex and folate-coated redox-sensitive lipid-polymer hybrid nanoparticle (Gd-FLPNP) for tumor-targeted magnetic resonance imaging and therapy. Gd-FLPNPs can simultaneously accomplish diagnostic imaging, and specific targeting and controlled release of doxorubicin (DOX). They exhibit good monodispersity, excellent size stability, and a well-defined core-shell structure. Paramagnetic nanoparticles based on gadolinium-diethylenetriaminepentaacetic acid-bis-cetylamine have paramagnetic properties with an approximately two-fold enhancement in the longitudinal relaxivity compared to clinical used Magnevist. For targeted and reduction-sensitive drug delivery, Gd-FLPNPs released DOX faster and enhanced cell uptake in vitro, and exhibited better antitumor effect both in vitro and in vivo.

  12. Epigenetic Mechanisms Regulating Adaptive Responses to Targeted Kinase Inhibitors in Cancer.

    Science.gov (United States)

    Angus, Steven P; Zawistowski, Jon S; Johnson, Gary L

    2018-01-06

    Although targeted inhibition of oncogenic kinase drivers has achieved remarkable patient responses in many cancers, the development of resistance has remained a significant challenge. Numerous mechanisms have been identified, including the acquisition of gatekeeper mutations, activating pathway mutations, and copy number loss or gain of the driver or alternate nodes. These changes have prompted the development of kinase inhibitors with increased selectivity, use of second-line therapeutics to overcome primary resistance, and combination treatment to forestall resistance. In addition to genomic resistance mechanisms, adaptive transcriptional and signaling responses seen in tumors are gaining appreciation as alterations that lead to a phenotypic state change-often observed as an epithelial-to-mesenchymal shift or reversion to a cancer stem cell-like phenotype underpinned by remodeling of the epigenetic landscape. This epigenomic modulation driving cell state change is multifaceted and includes modulation of repressive and activating histone modifications, DNA methylation, enhancer remodeling, and noncoding RNA species. Consequently, the combination of kinase inhibitors with drugs targeting components of the transcriptional machinery and histone-modifying enzymes has shown promise in preclinical and clinical studies. Here, we review mechanisms of resistance to kinase inhibition in cancer, with special emphasis on the rewired kinome and transcriptional signaling networks and the potential vulnerabilities that may be exploited to overcome these adaptive signaling changes.

  13. Specific Nongluten Proteins of Wheat Are Novel Target Antigens in Celiac Disease Humoral Response

    Science.gov (United States)

    2014-01-01

    While the antigenic specificity and pathogenic relevance of immunologic reactivity to gluten in celiac disease have been extensively researched, the immune response to nongluten proteins of wheat has not been characterized. We aimed to investigate the level and molecular specificity of antibody response to wheat nongluten proteins in celiac disease. Serum samples from patients and controls were screened for IgG and IgA antibody reactivity to a nongluten protein extract from the wheat cultivar Triticum aestivum Butte 86. Antibodies were further analyzed for reactivity to specific nongluten proteins by two-dimensional gel electrophoresis and immunoblotting. Immunoreactive molecules were identified by tandem mass spectrometry. Compared with healthy controls, patients exhibited significantly higher levels of antibody reactivity to nongluten proteins. The main immunoreactive nongluten antibody target proteins were identified as serpins, purinins, α-amylase/protease inhibitors, globulins, and farinins. Assessment of reactivity toward purified recombinant proteins further confirmed the presence of antibody response to specific antigens. The results demonstrate that, in addition to the well-recognized immune reaction to gluten, celiac disease is associated with a robust humoral response directed at a specific subset of the nongluten proteins of wheat. PMID:25329597

  14. Structural insights into a high affinity nanobody:antigen complex by homology modelling

    DEFF Research Database (Denmark)

    Skottrup, Peter Durand

    2017-01-01

    Porphyromonas gingivalis is a major periodontitis-causing pathogens. P. gingivalis secrete a cysteine protease termed RgpB, which is specific for Arg-Xaa bonds in substrates. Recently, a nanobody-based assay was used to demonstrate that RgpB could represent a novel diagnostic target, thereby...... simplifying. P. gingivalis detection. The nanobody, VHH7, had a high binding affinity and was specific for RgpB, when tested towards the highly identical RgpA. In this study a homology model of VHH7 was build. The complementarity determining regions (CDR) comprising the paratope residues responsible for Rgp...

  15. Target-responsive aptamer release from manganese dioxide nanosheets for electrochemical sensing of cocaine with target recycling amplification.

    Science.gov (United States)

    Chen, Zongbao; Lu, Minghua

    2016-11-01

    A novel electrochemical sensing platform based on manganese dioxide (MnO2) nanosheets was developed for sensitive screening of target cocaine with the signal amplification. Ferrocene-labeled cocaine aptamers were initially immobilized onto MnO2 nanosheets-modified screen-printed carbon electrode because of π-stacking interaction between nucleobases and nanosheets. The immobilized ferrocene-aptamer activated the electrical contact with the electrode, thereby resulting in the sensor circuit to switch on. Upon target cocaine introduction, the analyte reacted with the aptamer and caused the dissociation of ferrocene-aptamer from the electrode, thus giving rise to the detection circuit to switch off. The released aptamer was cleaved by DNase I with target recycling. Under optimal conditions, the decreasing percentage of the electronic signal relative to background current increased with the increasing cocaine concentration in the dynamic range of 0.1-20nM, and the detection limit was 32pM. The reproducibility, selectivity and method accuracy were acceptable. Importantly, this concept offers promise for rapid, simple, and cost-effective analysis of cocaine biological samples without the needs of sample separation and multiple washing steps. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. The Structure of Affine Buildings

    CERN Document Server

    Weiss, Richard M

    2009-01-01

    In The Structure of Affine Buildings, Richard Weiss gives a detailed presentation of the complete proof of the classification of Bruhat-Tits buildings first completed by Jacques Tits in 1986. The book includes numerous results about automorphisms, completions, and residues of these buildings. It also includes tables correlating the results in the locally finite case with the results of Tits's classification of absolutely simple algebraic groups defined over a local field. A companion to Weiss's The Structure of Spherical Buildings, The Structure of Affine Buildings is organized around the clas

  17. Targeting the cell stress response of Plasmodium falciparum to overcome artemisinin resistance.

    Directory of Open Access Journals (Sweden)

    Con Dogovski

    2015-04-01

    Full Text Available Successful control of falciparum malaria depends greatly on treatment with artemisinin combination therapies. Thus, reports that resistance to artemisinins (ARTs has emerged, and that the prevalence of this resistance is increasing, are alarming. ART resistance has recently been linked to mutations in the K13 propeller protein. We undertook a detailed kinetic analysis of the drug responses of K13 wild-type and mutant isolates of Plasmodium falciparum sourced from a region in Cambodia (Pailin. We demonstrate that ART treatment induces growth retardation and an accumulation of ubiquitinated proteins, indicative of a cellular stress response that engages the ubiquitin/proteasome system. We show that resistant parasites exhibit lower levels of ubiquitinated proteins and delayed onset of cell death, indicating an enhanced cell stress response. We found that the stress response can be targeted by inhibiting the proteasome. Accordingly, clinically used proteasome inhibitors strongly synergize ART activity against both sensitive and resistant parasites, including isogenic lines expressing mutant or wild-type K13. Synergy is also observed against Plasmodium berghei in vivo. We developed a detailed model of parasite responses that enables us to infer, for the first time, in vivo parasite clearance profiles from in vitro assessments of ART sensitivity. We provide evidence that the clinical marker of resistance (delayed parasite clearance is an indirect measure of drug efficacy because of the persistence of unviable parasites with unchanged morphology in the circulation, and we suggest alternative approaches for the direct measurement of viability. Our model predicts that extending current three-day ART treatment courses to four days, or splitting the doses, will efficiently clear resistant parasite infections. This work provides a rationale for improving the detection of ART resistance in the field and for treatment strategies that can be employed in areas

  18. Targeting the oxidative stress response system of fungi with safe, redox-potent chemosensitizing agents

    Directory of Open Access Journals (Sweden)

    Jong H. eKim

    2012-03-01

    Full Text Available The cellular antioxidation system is a target in the antifungal action of amphotericin B (AMB and itraconazole (ITZ, in filamentous fungi. The sakAΔ mutant of Aspergillus fumigatus, a mitogen-activated protein kinase (MAPK gene deletion mutant in the antioxidation system, was found to be more sensitive to AMB or ITZ than other A. fumigatus strains, a wild type and a mpkCΔ mutant (MAPK gene deletion mutant in polyalcohol sugar utilization system. The sakAΔ mutant showed no growth at 0.5 μg mL-1 of ITZ or reduced growth at 1.0 to 2.0 μg mL-1 of AMB, while the other strains exhibited robust growth. Complete fungal kill (≥ 99.9% by ITZ or AMB was achieved by much lower dosages for the sakAΔ mutant than for the other strains. SakA and MpkC appear to have overlapping roles in marshalling the oxidative stress response under treatment by an organic peroxide, tert-butyl hydroperoxide (t-BuOOH, or hydrogen peroxide (H2O2. The SakA signalling pathway was found to be responsible for fungal tolerance to AMB or ITZ toxicity. It appears msnA, an Aspergillus ortholog to Saccharomyces cerevisiae MSN2 (encoding a stress-responsive C2H2-type zinc-finger regulator and sakA and/or mpkC (upstream MAPKs are in the same stress response network under t-BuOOH-, H2O2- or AMB-triggered toxicity. Of note is that ITZ-sensitive yeast pathogens (Candida krusei and Cryptococcus neoformans were also sensitive to t-BuOOH, showing a connection between ITZ toxicity and oxidative stress response. This was shown by enhanced antifungal activity of AMB or ITZ when co-applied with redox-potent natural compounds, 2,3-dihydroxybenzaldehyde, thymol or salicylaldehyde, as chemosensitizing agents. Hence, redox compounds, which target the antioxidation system in fungi, possess a potent chemosensitizing capacity to enhance efficacy of conventional drugs inducing oxidative stress. Such chemosensitization can reduce costs and alleviate negative side effects associated with current

  19. Identification of a New Target slr0946 of the Response Regulator Sll0649 Involving Cadmium Tolerance in Synechocystis sp. PCC 6803

    Directory of Open Access Journals (Sweden)

    Tao Sun

    2017-08-01

    Full Text Available Survival of photosynthetic cyanobacteria is challenged by environmental contaminations like heavy metals. Among them, deciphering the regulatory mechanisms for cadmium (Cd in cyanobacteria would facilitate the construction of Cd-resistant strains. In this study, the DNA-Affinity-Purified-chromatin immunoprecipitation assay was employed to identify the direct targets of Sll0649, which was a Cd2+-related response regulator identified in our previous work in model cyanobacteria Synechocystis sp. PCC 6803. As a result, the promoter region of slr0946 encoding the arsenate reductase was enriched fourfolds by quantitative real time PCR analysis. Further, deletion of slr0946 led to a sensitive phenotype to Cd2+ stress compared with the wild type (WT and the sensitive phenotype of Δslr0946 could be rescued by complementation assay via introducing slr0946 back into Δslr0946. Finally, individually overexpression of slr0946 as well as two Cd2+-related genes identified priviously (i.e., sll1598 and slr0798 in WT could significantly improve the tolerance of Synechocystis sp. PCC 6803 to Cd2+. This study provided a better understanding of the tolerance mechanism to Cd2+ in cyanobacteria and also feasible strategies for tolerance modifications to heavy metals in the future.

  20. Enhanced radiation response in radioresistant MCF-7 cells by targeting peroxiredoxin II

    Directory of Open Access Journals (Sweden)

    Diaz AJG

    2013-10-01

    Full Text Available Anthony Joseph Gomez Diaz,1 Daniel Tamae,2 Yun Yen,3 JianJian Li,4 Tieli Wang1 1Department of Chemistry and Biochemistry, California State University at Dominguez Hills, Carson, CA, 2Center of Excellence in Environmental Toxicology, Department of Pharmacology, University of Pennsylvania, Philadelphia, PA, 3Department of Clinical and Molecular Pharmacology, Beckman Research Institute of City of Hope National Medical Center, Duarte, CA, 4Department of Radiation Oncology, University of California Davis, Sacramento, CA, USA Abstract: In our previous study, we identified that a protein target, peroxiredoxin II (PrxII, is overexpressed in radioresistant MCF+FIR3 breast-cancer cells and found that its expression and function is associated with breast-cancer radiation sensitivity or resistance. Small interference RNA (siRNA targeting PrxII gene expression was able to sensitize MCF+FIR3 radioresistant breast-cancer cells to ionizing radiation. The major focus of this work was to investigate how the radiation response of MCF+FIR3 radioresistant cells was affected by the siRNA that inhibits PrxII gene expression. Our results, presented here, show that silencing PrxII gene expression increased cellular toxicity by altering cellular thiol status, inhibiting Ca2+ efflux from the cells, and perturbing the intracellular Ca2+ homeostasis. By combining radiotherapy and siRNA technology, we hope to develop new therapeutic strategies that may have potential to enhance the efficacy of chemotherapeutic agents due to this technology's property of targeting to specific cancer-related genes. Keywords: siRNA, PrxII, radiation resistance, Ca2+, MCF+FIR3

  1. Perceptions of similarity and response to selected comparison targets in type 2 diabetes.

    Science.gov (United States)

    Arigo, Danielle; Smyth, Joshua M; Suls, Jerry M

    2015-01-01

    Social comparisons (i.e. self-evaluations relative to others) may affect motivation for diabetes self-care behaviours. Comparisons can have either positive or negative effects, but it is not clear what differentiates these responses. This study tested the effect of a patient's perceived similarity to a comparison target on motivation for self-care. Individuals with type 2 diabetes (n = 180, MA1c = 7.59%) selected to read one of four brief descriptions of a patient with diabetes. Participants rated their motivation for self-care behaviours prior and subsequent to reading and reported the extent to which they focused on similarities between the self and the selected patient while reading. Perceived similarity moderated the effect of selection on motivation for self-care (p = .01, η2 = .06). Increased motivation was observed if participants focused on similarities with patients 'doing better' (i.e. high coping effectiveness/low symptom severity) and decreased motivation if they focused on similarities with patients 'doing worse' (low coping effectiveness/high symptom severity). Providing social comparison information in diabetes management (and perhaps other chronic diseases) may improve motivation for self-care among some patients. A subset of patients, however, may benefit from guidance to focus on similarities with certain targets.

  2. Nanomedicine targeting the tumor microenvironment: Therapeutic strategies to inhibit angiogenesis, remodel matrix, and modulate immune responses

    Directory of Open Access Journals (Sweden)

    Elizabeth L. Siegler

    2016-11-01

    Full Text Available Increasing attention has been given to the tumor microenvironment (TME, which includes cellular and structural components such as fibroblasts, immune cells, vasculature, and extracellular matrix (ECM that surround tumor sites. These components contribute to tumor growth and metastasis and are one reason why traditional chemotherapy often is insufficient to eradicate the tumor completely. Newer treatments that target aspects of the TME, such as antiangiogenic and immunostimulatory therapies, have seen limited clinical success despite promising preclinical results. This can be attributed to a number of reasons, including a lack of drug penetration deeper into the necrotic tumor core, nonspecific delivery, rapid clearance from serum, or toxic side effects at high doses. Nanoparticles offer a potential solution to all of these obstacles, and many recent studies have shown encouraging results using nanomedicine to target TME vasculature, ECM, and immune response. While few of these platforms have made it to clinical trials to date, these strategies are relatively new and may offer a way to improve the effects of anticancer therapies.

  3. Affinity biosensors: techniques and protocols

    National Research Council Canada - National Science Library

    Rogers, Kim R; Mulchandani, Ashok

    1998-01-01

    ..., and government to begin or expand their biosensors research. This volume, Methods in Biotechnology vol. 7: Affinity Biosensors: Techniques and Protocols, describes a variety of classical and emerging transduction technologies that have been interfaced to bioaffinity elements (e.g., antibodies and receptors). Some of the reas...

  4. Targeting the Oxidative Stress Response System of Fungi with Redox-Potent Chemosensitizing Agents

    Science.gov (United States)

    Kim, Jong H.; Chan, Kathleen L.; Faria, Natália C. G.; Martins, M. de L.; Campbell, Bruce C.

    2012-01-01

    The cellular antioxidant system is a target in the antifungal action of amphotericin B (AMB) and itraconazole (ITZ), in filamentous fungi. The sakAΔ mutant of Aspergillus fumigatus, a mitogen-activated protein kinase (MAPK) gene deletion mutant in the antioxidant system, was found to be more sensitive to AMB or ITZ than other A. fumigatus strains, a wild type and a mpkCΔ mutant (a MAPK gene deletion mutant in the polyalcohol sugar utilization system). Complete fungal kill (≥99.9%) by ITZ or AMB was also achieved by much lower dosages for the sakAΔ mutant than for the other strains. It appears msnA, an Aspergillus ortholog to Saccharomyces cerevisiae MSN2 (encoding a stress-responsive C2H2-type zinc-finger regulator) and sakA and/or mpkC (upstream MAPKs) are in the same stress response network under tert-butyl hydroperoxide (t-BuOOH)-, hydrogen peroxide (H2O2)- or AMB-triggered toxicity. Of note is that ITZ-sensitive yeast pathogens were also sensitive to t-BuOOH, showing a connection between ITZ sensitivity and antioxidant capacity of fungi. Enhanced antifungal activity of AMB or ITZ was achieved when these drugs were co-applied with redox-potent natural compounds, 2,3-dihydroxybenzaldehyde, thymol or salicylaldehyde, as chemosensitizing agents. We concluded that redox-potent compounds, which target the antioxidant system in fungi, possess a chemosensitizing capacity to enhance efficacy of conventional drugs. PMID:22438852

  5. Tumor responsive targeted multifunctional nanosystems for cancer imaging, chemo- and siRNA therapy

    Science.gov (United States)

    Savla, Ronak

    Cancer is one of the most insidious diseases. Compromising of over 100 different types and sharing the unifying factors of uncontrolled growth and metastasis, unmet clinical needs in terms of cancer diagnosis and treatment continue to exist. It is widely accepted that most forms of cancer are treatable or even curable if detected before widespread metastasis occurs. Nearly a quarter of deaths in the United States is the result of cancer and it only trails heart disease in terms of annual mortality. Surgery, chemotherapy, and radiation therapy are the primary treatment modalities for cancer. Research in these procedures has resulted in substantial benefits for cancer patients, but there is still room for an improvement. However, a time has been reached at which it appears that the benefits from these modalities have been reached the maximum. Therefore, it is vital to develop new strategies for the diagnosis and treatment of cancer. The field of nanotechnology is concerned with structures in the nanometer size range and holds the potential to drastically impact and improve the lives of patients suffering from cancer. Not only can nanotechnology improve current methods of diagnosis and treatment, it has a possibility of introducing newer and better modalities. The overall purpose of this work is to develop novel nanotechnology-based methodologies for the diagnosis and treatment of various forms of cancers. The first aim of the project is the development of a multifunctional targeted nanosystem for the delivery of siRNA to overcome drug resistance. The second aspect is the synthesis of a quantum dot-based delivery system that releases drug in response to pH changes. The third aim is the development of a targeted, tumor environment responsive magnetic resonance nanoparticle contrast agent coupled with a nanoparticle-based treatment.

  6. The innate immune response in fetal lung mesenchymal cells targets VEGFR2 expression and activity.

    Science.gov (United States)

    Medal, Rachel M; Im, Amanda M; Yamamoto, Yasutoshi; Lakhdari, Omar; Blackwell, Timothy S; Hoffman, Hal M; Sahoo, Debashis; Prince, Lawrence S

    2017-06-01

    In preterm infants, soluble inflammatory mediators target lung mesenchymal cells, disrupting airway and alveolar morphogenesis. However, how mesenchymal cells respond directly to microbial stimuli remains poorly characterized. Our objective was to measure the genome-wide innate immune response in fetal lung mesenchymal cells exposed to the bacterial endotoxin lipopolysaccharide (LPS). With the use of Affymetrix MoGene 1.0st arrays, we showed that LPS induced expression of unique innate immune transcripts heavily weighted toward CC and CXC family chemokines. The transcriptional response was different between cells from E11, E15, and E18 mouse lungs. In all cells tested, LPS inhibited expression of a small core group of genes including the VEGF receptor Vegfr2 Although best characterized in vascular endothelial populations, we demonstrated here that fetal mouse lung mesenchymal cells express Vegfr2 and respond to VEGF-A stimulation. In mesenchymal cells, VEGF-A increased cell migration, activated the ERK/AKT pathway, and promoted FOXO3A nuclear exclusion. With the use of an experimental coculture model of epithelial-mesenchymal interactions, we also showed that VEGFR2 inhibition prevented formation of three-dimensional structures. Both LPS and tyrosine kinase inhibition reduced three-dimensional structure formation. Our data suggest a novel mechanism for inflammation-mediated defects in lung development involving reduced VEGF signaling in lung mesenchyme. Copyright © 2017 the American Physiological Society.

  7. Syk/Src Pathway-Targeted Inhibition of Skin Inflammatory Responses by Carnosic Acid

    Directory of Open Access Journals (Sweden)

    Jueun Oh

    2012-01-01

    Full Text Available Carnosic acid (CA is a diterpene compound exhibiting antioxidative, anticancer, anti-angiogenic, anti-inflammatory, anti-metabolic disorder, and hepatoprotective and neuroprotective activities. In this study, the effect of CA on various skin inflammatory responses and its inhibitory mechanism were examined. CA strongly suppressed the production of IL-6, IL-8, and MCP-1 from keratinocyte HaCaT cells stimulated with sodium lauryl sulfate (SLS and retinoic acid (RA. In addition, CA blocked the release of nitric oxide (NO, tumor necrosis factor (TNF-α, and prostaglandin E2 (PGE2 from RAW264.7 cells activated by the toll-like receptor (TLR-2 ligands, Gram-positive bacterium-derived peptidoglycan (PGN and pam3CSK, and the TLR4 ligand, Gram-negative bacterium-derived lipopolysaccharide (LPS. CA arrested the growth of dermatitis-inducing Gram-positive and Gram-negative microorganisms such Propionibacterium acnes, Pseudomonas aeruginosa, and Staphylococcus aureus. CA also blocked the nuclear translocation of nuclear factor (NF-κB and its upstream signaling including Syk/Src, phosphoinositide 3-kinase (PI3K, Akt, inhibitor of κBα (IκBα kinase (IKK, and IκBα for NF-κB activation. Kinase assays revealed that Syk could be direct enzymatic target of CA in its anti-inflammatory action. Therefore, our data strongly suggest the potential of CA as an anti-inflammatory drug against skin inflammatory responses with Src/NF-κB inhibitory properties.

  8. GTPase ROP2 binds and promotes activation of target of rapamycin, TOR, in response to auxin.

    Science.gov (United States)

    Schepetilnikov, Mikhail; Makarian, Joelle; Srour, Ola; Geldreich, Angèle; Yang, Zhenbiao; Chicher, Johana; Hammann, Philippe; Ryabova, Lyubov A

    2017-04-03

    Target of rapamycin (TOR) promotes reinitiation at upstream ORFs (uORFs) in genes that play important roles in stem cell regulation and organogenesis in plants. Here, we report that the small GTPase ROP2, if activated by the phytohormone auxin, promotes activation of TOR, and thus translation reinitiation of uORF-containing mRNAs. Plants with high levels of active ROP2, including those expressing constitutively active ROP2 (CA-ROP2), contain high levels of active TOR ROP2 physically interacts with and, when GTP-bound, activates TOR in vitro TOR activation in response to auxin is abolished in ROP-deficient rop2 rop6 ROP4 RNAi plants. GFP-TOR can associate with endosome-like structures in ROP2-overexpressing plants, indicating that endosomes mediate ROP2 effects on TOR activation. CA-ROP2 is efficient in loading uORF-containing mRNAs onto polysomes and stimulates translation in protoplasts, and both processes are sensitive to TOR inhibitor AZD-8055. TOR inactivation abolishes ROP2 regulation of translation reinitiation, but not its effects on cytoskeleton or intracellular trafficking. These findings imply a mode of translation control whereby, as an upstream effector of TOR, ROP2 coordinates TOR function in translation reinitiation pathways in response to auxin. © 2017 The Authors.

  9. Apolipoprotein E-specific innate immune response in astrocytes from targeted replacement mice

    Directory of Open Access Journals (Sweden)

    Montine Thomas J

    2006-04-01

    Full Text Available Abstract Background Inheritance of the three different alleles of the human apolipoprotein (apo E gene (APOE are associated with varying risk or clinical outcome from a variety of neurologic diseases. ApoE isoform-specific modulation of several pathogenic processes, in addition to amyloid β metabolism in Alzheimer's disease, have been proposed: one of these is innate immune response by glia. Previously we have shown that primary microglia cultures from targeted replacement (TR APOE mice have apoE isoform-dependent innate immune activation and paracrine damage to neurons that is greatest with TR by the ε4 allele (TR APOE4 and that derives from p38 mitogen-activated protein kinase (p38MAPK activity. Methods Primary cultures of TR APOE2, TR APOE3 and TR APOE4 astrocytes were stimulated with lipopolysaccharide (LPS. ApoE secretion, cytokine production, and nuclear factor-kappa B (NF-κB subunit activity were measured and compared. Results Here we showed that activation of primary astrocytes from TR APOE mice with LPS led to TR APOE-dependent differences in cytokine secretion that were greatest in TR APOE2 and that were associated with differences in NF-κB subunit activity. Conclusion Our results suggest that LPS activation of innate immune response in TR APOE glia results in opposing outcomes from microglia and astrocytes as a result of TR APOE-dependent activation of p38MAPK or NF-κB signaling in these two cell types.

  10. Intelligently targeted drug delivery and enhanced antitumor effect by gelatinase-responsive nanoparticles.

    Directory of Open Access Journals (Sweden)

    Rutian Li

    Full Text Available AIMS: The matrix metalloproteinase (MMP 2/9, also known as collagenases IV and gelatinases A/B, play a key role in cancer invasion and metastasis. However, the clinical trials of the MMP inhibitors (MMPIs ended up with disappointing results. In this paper, we synthesized a gelatinase-responsive copolymer (mPEG-PCL by inserting a gelatinase cleavable peptide (PVGLIG between mPEG and PCL blocks of mPEG-PCL for anticancer drug delivery to make use of MMP2/9 as an intelligent target for drug delivery. MATERIALS AND METHODS: mPEG-pep-PCL copolymer was synthesized via ring-opening copolymerization and double-amidation. To evaluate whether Nanoparticles (NPs prepared from this copolymer are superior to NPs prepared from mPEG-PCL, NPs prepared from mPEG-PCL copolymer were used as positive control. Docetaxel-loading NPs using mPEG-pep-PCL and mPEG-PCL were prepared by nano-precipitation method, mentioned as Gel-NPs and Con-NPs, respectively. The morphologic changes of the NPs after treatment with gelatinases were observed macroscopically by spectrophotometer and microscopically by transmission electron microscopy (TEM and atomic force microscopy (AFM. The cellular uptake amount and cytotoxicity of Gel-NPs and Con-NPs, respectively, in cell lines with different levels of gelatinase expression were studied. Moreover, the cytotoxicity study on the primary cancer cells isolated from pericardial fluids from a patient with late-stage lung cancer was conducted. RESULTS: The Gel-NPs aggregated in response to gelatinases, which was confirmed macroscopically and microscopically. The cellular uptake amount of Gel-NPs was correlated with the level of gelatinases. The in vitro antitumor effect of Gel-NPs was also correlated with the level of gelatinases and was superior to Taxotere (commercially available docetaxel as well as the Con-NPs. The cytotoxicity study on the primary lung cancer cells also confirmed the effectiveness of Gel-NPs. CONCLUSION: The results in

  11. Targeting of non-dominant antigens as a vaccine strategy to broaden T-cell responses during chronic viral infection

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Jensen, Benjamin Anderschou Holbech; Ragonnaud, Emeline

    2015-01-01

    In this study, we compared adenoviral vaccine vectors with the capacity to induce equally potent immune responses against non-dominant and immunodominant epitopes of murine lymphocytic choriomeningitis virus (LCMV). Our results demonstrate that vaccination targeting non-dominant epitopes facilita......In this study, we compared adenoviral vaccine vectors with the capacity to induce equally potent immune responses against non-dominant and immunodominant epitopes of murine lymphocytic choriomeningitis virus (LCMV). Our results demonstrate that vaccination targeting non-dominant epitopes...... was lost over time in T cells specific for the dominant T cell epitopes, and these cells were fully capable of expanding in response to a new viral challenge. Overall, our data suggests a potential for broadening of the antiviral CD8+ T-cell response by selecting non-dominant antigens to be targeted...

  12. In Vitro Drug Sensitivity Tests to Predict Molecular Target Drug Responses in Surgically Resected Lung Cancer.

    Directory of Open Access Journals (Sweden)

    Ryohei Miyazaki

    Full Text Available Epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs and anaplastic lymphoma kinase (ALK inhibitors have dramatically changed the strategy of medical treatment of lung cancer. Patients should be screened for the presence of the EGFR mutation or echinoderm microtubule-associated protein-like 4 (EML4-ALK fusion gene prior to chemotherapy to predict their clinical response. The succinate dehydrogenase inhibition (SDI test and collagen gel droplet embedded culture drug sensitivity test (CD-DST are established in vitro drug sensitivity tests, which may predict the sensitivity of patients to cytotoxic anticancer drugs. We applied in vitro drug sensitivity tests for cyclopedic prediction of clinical responses to different molecular targeting drugs.The growth inhibitory effects of erlotinib and crizotinib were confirmed for lung cancer cell lines using SDI and CD-DST. The sensitivity of 35 cases of surgically resected lung cancer to erlotinib was examined using SDI or CD-DST, and compared with EGFR mutation status.HCC827 (Exon19: E746-A750 del and H3122 (EML4-ALK cells were inhibited by lower concentrations of erlotinib and crizotinib, respectively than A549, H460, and H1975 (L858R+T790M cells were. The viability of the surgically resected lung cancer was 60.0 ± 9.8 and 86.8 ± 13.9% in EGFR-mutants vs. wild types in the SDI (p = 0.0003. The cell viability was 33.5 ± 21.2 and 79.0 ± 18.6% in EGFR mutants vs. wild-type cases (p = 0.026 in CD-DST.In vitro drug sensitivity evaluated by either SDI or CD-DST correlated with EGFR gene status. Therefore, SDI and CD-DST may be useful predictors of potential clinical responses to the molecular anticancer drugs, cyclopedically.

  13. Food and Natural Materials Target Mechanisms to Effectively Regulate Allergic Responses.

    Science.gov (United States)

    Shin, Hee Soon; Shon, Dong-Hwa

    2015-01-01

    An immune hypersensitivity disorder called allergy is caused by diverse allergens entering the body via skin contact, injection, ingestion, and/or inhalation. These allergic responses may develop into allergic disorders, including inflammations such as atopic dermatitis, asthma, anaphylaxis, food allergies, and allergic rhinitis. Several drugs have been developed to treat these allergic disorders; however, long-term intake of these drugs could have adverse effects. As an alternative to these medicines, food and natural materials that ameliorate allergic disorder symptoms without producing any side effects can be consumed. Food and natural materials can effectively regulate successive allergic responses in an allergic chain-reaction mechanism in the following ways: [1] Inhibition of allergen permeation via paracellular diffusion into epithelial cells, [2] suppression of type 2 T-helper (Th) cell-related cytokine production by regulating Th1/Th2 balance, [3] inhibition of pathogenic effector CD4(+) T cell differentiation by inducing regulatory T cells (Treg), and [4] inhibition of degranulation in mast cells. The immunomodulatory effects of food and natural materials on each target mechanism were scientifically verified and shown to alleviate allergic disorder symptoms. Furthermore, consumption of certain food and natural materials such as fenugreek, skullcap, chitin/chitosan, and cheonggukjang as anti-allergics have merits such as safety (no adverse side effects), multiple suppressive effects (as a mixture would contain various components that are active against allergic responses), and ease of consumption when required. These merits and anti-allergic properties of food and natural materials help control various allergic disorders.

  14. Allergen and Epitope Targets of Mouse-Specific T Cell Responses in Allergy and Asthma

    Directory of Open Access Journals (Sweden)

    Véronique Schulten

    2018-02-01

    Full Text Available Mouse allergy has become increasingly common, mainly affecting laboratory workers and inner-city households. To date, only one major allergen, namely Mus m 1, has been described. We sought to identify T cell targets in mouse allergic patients. PBMC from allergic donors were expanded with either murine urine or epithelial extract and subsequently screened for cytokine production (IL-5 and IFNγ in response to overlapping peptides spanning the entire Mus m 1 sequence, peptides from various Mus m 1 isoforms [major urinary proteins (MUPs], peptides from mouse orthologs of known allergens from other mammalian species and peptides from proteins identified by immunoproteomic analysis of IgE/IgG immunoblots of mouse urine and epithelial extracts. This approach let to the identification of 106 non-redundant T cell epitopes derived from 35 antigens. Three major T cell-activating regions were defined in Mus m 1 alone. Moreover, our data show that immunodominant epitopes were largely shared between Mus m 1 and other MUPs even from different species, suggesting that sequence conservation in different allergens is a determinant for immunodominance. We further identified several novel mouse T cell antigens based on their homology to known mammalian allergens. Analysis of cohort-specific T cell responses revealed that rhinitis and asthmatic patients recognized different epitope repertoires. Epitopes defined herein can be formulated into an epitope “megapool” used to diagnose mouse allergy and study mouse-specific T cell responses directly ex vivo. This analysis of T cell epitopes provides a good basis for future studies to increase our understanding of the immunopathology associated with MO-allergy and asthma.

  15. ErbB polymorphisms: Insights and implications for response to targeted cancer therapeutics

    Directory of Open Access Journals (Sweden)

    Moulay A Alaoui-Jamali

    2015-02-01

    Full Text Available Advances in high-throughput genomic-scanning have expanded the repertory of genetic variations in DNA sequences encoding ErbB tyrosine kinase receptors in humans, including single nucleotide polymorphisms (SNPs, polymorphic repetitive elements, microsatellite variations, small-scale insertions and deletions. The ErbB family members: EGFR, ErbB2, ErbB3 and ErbB4 receptors are established as drivers of many aspects of tumor initiation and progression to metastasis. This knowledge has provided rationales for the development of an arsenal of anti-ErbB therapeutics, ranging from small molecule kinase inhibitors to monoclonal antibodies. Anti-ErbB agents are becoming the cornerstone therapeutics for the management of cancers that overexpress hyperactive variants of ErbB receptors, in particular ErbB2-positive breast cancer and non-small cell lung carcinomas. However, their clinical benefit has been limited to a subset of patients due to a wide heterogeneity in drug response despite the expression of the ErbB targets, attributed to intrinsic (primary and to acquired (secondary resistance. Somatic mutations in ErbB tyrosine kinase domains have been extensively investigated in preclinical and clinical setting as determinants for either high sensitivity or resistance to anti-ErbB therapeutics. In contrast, only scant information is available on the impact of SNPs, which are widespread in genes encoding ErbB receptors, on receptor structure and activity, and their predictive values for drug susceptibility. This review aims to briefly update polymorphic variations in genes encoding ErbB receptors based on recent advances in deep sequencing technologies, and to address challenging issues for a better understanding of the functional impact of single versus combined SNPs in ErbB genes to receptor topology, receptor-drug interaction, and drug susceptibility. The potential of exploiting SNPs in the era of stratified targeted therapeutics is discussed.

  16. The distribution of BRAF gene fusions in solid tumors and response to targeted therapy.

    Science.gov (United States)

    Ross, Jeffrey S; Wang, Kai; Chmielecki, Juliann; Gay, Laurie; Johnson, Adrienne; Chudnovsky, Jacob; Yelensky, Roman; Lipson, Doron; Ali, Siraj M; Elvin, Julia A; Vergilio, Jo-Anne; Roels, Steven; Miller, Vincent A; Nakamura, Brooke N; Gray, Adam; Wong, Michael K; Stephens, Philip J

    2016-02-15

    Although the BRAF V600E base substitution is an approved target for the BRAF inhibitors in melanoma, BRAF gene fusions have not been investigated as anticancer drug targets. In our study, a wide variety of tumors underwent comprehensive genomic profiling for hundreds of known cancer genes using the FoundationOne™ or FoundationOne Heme™ comprehensive genomic profiling assays. BRAF fusions involving the intact in-frame BRAF kinase domain were observed in 55 (0.3%) of 20,573 tumors, across 12 distinct tumor types, including 20 novel BRAF fusions. These comprised 29 unique 5' fusion partners, of which 31% (9) were known and 69% (20) were novel. BRAF fusions included 3% (14/531) of melanomas; 2% (15/701) of gliomas; 1.0% (3/294) of thyroid cancers; 0.3% (3/1,062) pancreatic carcinomas; 0.2% (8/4,013) nonsmall-cell lung cancers and 0.2% (4/2,154) of colorectal cancers, and were enriched in pilocytic (30%) vs. nonpilocytic gliomas (1%; p < 0.0001), Spitzoid (75%) vs. nonSpitzoid melanomas (1%; p = 0.0001), acinar (67%) vs. nonacinar pancreatic cancers (<1%; p < 0.0001) and papillary (3%) vs. nonpapillary thyroid cancers (0%; p < 0.03). Clinical responses to trametinib and sorafenib are presented. In conclusion, BRAF fusions are rare driver alterations in a wide variety of malignant neoplasms, but enriched in Spitzoid melanoma, pilocytic astrocytomas, pancreatic acinar and papillary thyroid cancers. © 2015 The Authors. Published by Wiley Periodicals, Inc. on behalf of UICC.

  17. Fluorescence and computational studies of thymidine phosphorylase affinity toward lipidated 5-FU derivatives

    Science.gov (United States)

    Lettieri, R.; D'Abramo, M.; Stella, L.; La Bella, A.; Leonelli, F.; Giansanti, L.; Venanzi, M.; Gatto, E.

    2018-04-01

    Thymidine phosphorylase (TP) is an enzyme that is up-regulated in a wide variety of solid tumors, including breast and colorectal cancers. It is involved in tumor growth and metastasis, for this reason it is one of the key enzyme to be inhibited, in an attempt to prevent tumor proliferation. However, it also plays an active role in cancer treatment, through its contribution in the conversion of the anti-cancer drug 5-fluorouracil (5-FU) to an irreversible inhibitor of thymidylate synthase (TS), responsible of the inhibition of the DNA synthesis. In this work, the intrinsic TP fluorescence has been investigated for the first time and exploited to study TP binding affinity for the unsubstituted 5-FU and for two 5-FU derivatives, designed to expose this molecule on liposomal membranes. These molecules were obtained by functionalizing the nitrogen atom with a chain consisting of six (1) or seven (2) units of glycol, linked to an alkyl moiety of 12 carbon atoms. Derivatives (1) and (2) exhibited an affinity for TP in the micromolar range, 10 times higher than the parent compound, irrespective of the length of the polyoxyethylenic spacer. This high affinity was maintained also when the compounds were anchored in liposomal membranes. Experimental results were supported by molecular dynamics simulations and docking calculations, supporting a feasible application of the designed supramolecular lipid structure in selective targeting of TP, to be potentially used as a drug delivery system or sensor device.

  18. The affine quantum gravity programme

    CERN Document Server

    Klauder, J R

    2002-01-01

    The central principle of affine quantum gravity is securing and maintaining the strict positivity of the matrix left brace g-hat sub a sub b (x)right brace composed of the spatial components of the local metric operator. On spectral grounds, canonical commutation relations are incompatible with this principle, and they must be replaced by noncanonical, affine commutation relations. Due to the partial second-class nature of the quantum gravitational constraints, it is advantageous to use the recently developed projection operator method, which treats all quantum constraints on an equal footing. Using this method, enforcement of regularized versions of the gravitational operator constraints is formulated quite naturally by means of a novel and relatively well-defined functional integral involving only the same set of variables that appears in the usual classical formulation. It is anticipated that skills and insight to study this formulation can be developed by studying special, reduced-variable models that sti...

  19. Affine invariants of convex polygons.

    Science.gov (United States)

    Flusser, Jan

    2002-01-01

    In this correspondence, we prove that the affine invariants, for image registration and object recognition, proposed recently by Yang and Cohen (see ibid., vol.8, no.7, p.934-46, July 1999) are algebraically dependent. We show how to select an independent and complete set of the invariants. The use of this new set leads to a significant reduction of the computing complexity without decreasing the discrimination power.

  20. Explaining the effects of targeted online advertising on children's cognitive, affective, and behavioral brand responses

    NARCIS (Netherlands)

    van Reijmersdal, E.; Rozendaal, E.; Smink, N.; van Noort, G.; Buijzen, M.

    2013-01-01

    Increasingly, information from children's profile pages on social network sites is used to target online advertising. This practice has raised concerns in society and academia, however, effects of profile targeting on children remained unstudied. Therefore, this study focused on children's

  1. Targeted Drug Delivery and Treatment of Endoparasites with Biocompatible Particles of pH-Responsive Structure.

    Science.gov (United States)

    Mathews, Patrick D; Fernandes Patta, Ana C M; Gonçalves, Joao V; Gama, Gabriella Dos Santos; Garcia, Irene Teresinha Santos; Mertins, Omar

    2018-02-12

    Biomaterials conceived for vectorization of bioactives are currently considered for biomedical, biological, and environmental applications. We have produced a pH-sensitive biomaterial composed of natural source alginate and chitosan polysaccharides for application as a drug delivery system via oral administration. The composite particle preparation was in situ monitored by means of isothermal titration calorimetry. The strong interaction established between the macromolecules during particle assembly led to 0.60 alginate/chitosan effective binding sites with an intense exothermic effect and negative enthalpy variation on the order of a thousand kcal/mol. In the presence of model drugs mebendazole and ivermectin, with relatively small and large structures, respectively, mebendazole reduced the amount of chitosan monomers available to interact with alginate by 27%, which was not observed for ivermectin. Nevertheless, a state of intense negative Gibbs energy and large entropic decrease was achieved, providing evidence that formation of particles is thermodynamically driven and favored. Small-angle X-ray scattering provided further evidence of similar surface aspects independent of the presence of drug. The physical responses of the particles to pH variation comprise partial hydration, swelling, and the predominance of positive surface charge in strong acid medium, whereas ionization followed by deprotonation leads to compaction and charge reversal rather than new swelling in mild and slightly acidic mediums, respectively. In vivo performance was evaluated in the treatment of endoparasites in Corydoras fish. Systematically with a daily base oral administration, particles significantly reduced the infections over 15 days of treatment. The experiments provide evidence that utilizing particles granted and boosted the action of the antiparasitic drugs, leading to substantial reduction or elimination of infection. Hence, the pH-responsive particles represent a biomaterial

  2. Targeting brains, producing responsibilities: the use of neuroscience within British social policy.

    Science.gov (United States)

    Broer, Tineke; Pickersgill, Martyn

    2015-05-01

    Concepts and findings 'translated' from neuroscientific research are finding their way into UK health and social policy discourse. Critical scholars have begun to analyse how policies tend to 'misuse' the neurosciences and, further, how these discourses produce unwarranted and individualizing effects, rooted in middle-class values and inducing guilt and anxiety. In this article, we extend such work while simultaneously departing from the normative assumptions implied in the concept of 'misuse'. Through a documentary analysis of UK policy reports focused on the early years, adolescence and older adults, we examine how these employ neuroscientific concepts and consequently (re)define responsibility. In the documents analysed, responsibility was produced in three different but intersecting ways: through a focus on optimisation, self-governance, and vulnerability. Our work thereby adds to social scientific examinations of neuroscience in society that show how neurobiological terms and concepts can be used to construct and support a particular imaginary of citizenship and the role of the state. Neuroscience may be leveraged by policy makers in ways that (potentially) reduce the target of their intervention to the soma, but do so in order to expand the outcome of the intervention to include the enhancement of society writ large. By attending as well to more critical engagements with neuroscience in policy documents, our analysis demonstrates the importance of being mindful of the limits to the deployment of a neurobiological idiom within policy settings. Accordingly, we contribute to increased empirical specificity concerning the impacts and translation of neuroscientific knowledge in contemporary society whilst refusing to take for granted the idea that the neurosciences necessarily have a dominant role (to play). Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  3. Rank Two Affine Manifolds in Genus 3

    OpenAIRE

    Aulicino, David; Nguyen, Duc-Manh

    2016-01-01

    We complete the classification of rank two affine manifolds in the moduli space of translation surfaces in genus three. Combined with a recent result of Mirzakhani and Wright, this completes the classification of higher rank affine manifolds in genus three.

  4. Alternative affinity tools: more attractive than antibodies?

    NARCIS (Netherlands)

    Ruigrok, V.J.B.; Levisson, M.; Eppink, M.H.M.; Smidt, H.; Oost, van der J.

    2011-01-01

    Antibodies are the most successful affinity tools used today, in both fundamental and applied research (diagnostics, purification and therapeutics). Nonetheless, antibodies do have their limitations, including high production costs and low stability. Alternative affinity tools based on nucleic acids

  5. Spectral affinity in protein networks.

    Science.gov (United States)

    Voevodski, Konstantin; Teng, Shang-Hua; Xia, Yu

    2009-11-29

    Protein-protein interaction (PPI) networks enable us to better understand the functional organization of the proteome. We can learn a lot about a particular protein by querying its neighborhood in a PPI network to find proteins with similar function. A spectral approach that considers random walks between nodes of interest is particularly useful in evaluating closeness in PPI networks. Spectral measures of closeness are more robust to noise in the data and are more precise than simpler methods based on edge density and shortest path length. We develop a novel affinity measure for pairs of proteins in PPI networks, which uses personalized PageRank, a random walk based method used in context-sensitive search on the Web. Our measure of closeness, which we call PageRank Affinity, is proportional to the number of times the smaller-degree protein is visited in a random walk that restarts at the larger-degree protein. PageRank considers paths of all lengths in a network, therefore PageRank Affinity is a precise measure that is robust to noise in the data. PageRank Affinity is also provably related to cluster co-membership, making it a meaningful measure. In our experiments on protein networks we find that our measure is better at predicting co-complex membership and finding functionally related proteins than other commonly used measures of closeness. Moreover, our experiments indicate that PageRank Affinity is very resilient to noise in the network. In addition, based on our method we build a tool that quickly finds nodes closest to a queried protein in any protein network, and easily scales to much larger biological networks. We define a meaningful way to assess the closeness of two proteins in a PPI network, and show that our closeness measure is more biologically significant than other commonly used methods. We also develop a tool, accessible at http://xialab.bu.edu/resources/pnns, that allows the user to quickly find nodes closest to a queried vertex in any protein

  6. Spectral affinity in protein networks

    Directory of Open Access Journals (Sweden)

    Teng Shang-Hua

    2009-11-01

    Full Text Available Abstract Background Protein-protein interaction (PPI networks enable us to better understand the functional organization of the proteome. We can learn a lot about a particular protein by querying its neighborhood in a PPI network to find proteins with similar function. A spectral approach that considers random walks between nodes of interest is particularly useful in evaluating closeness in PPI networks. Spectral measures of closeness are more robust to noise in the data and are more precise than simpler methods based on edge density and shortest path length. Results We develop a novel affinity measure for pairs of proteins in PPI networks, which uses personalized PageRank, a random walk based method used in context-sensitive search on the Web. Our measure of closeness, which we call PageRank Affinity, is proportional to the number of times the smaller-degree protein is visited in a random walk that restarts at the larger-degree protein. PageRank considers paths of all lengths in a network, therefore PageRank Affinity is a precise measure that is robust to noise in the data. PageRank Affinity is also provably related to cluster co-membership, making it a meaningful measure. In our experiments on protein networks we find that our measure is better at predicting co-complex membership and finding functionally related proteins than other commonly used measures of closeness. Moreover, our experiments indicate that PageRank Affinity is very resilient to noise in the network. In addition, based on our method we build a tool that quickly finds nodes closest to a queried protein in any protein network, and easily scales to much larger biological networks. Conclusion We define a meaningful way to assess the closeness of two proteins in a PPI network, and show that our closeness measure is more biologically significant than other commonly used methods. We also develop a tool, accessible at http://xialab.bu.edu/resources/pnns, that allows the user to

  7. Inflammatory and immune responses in the cochlea: potential therapeutic targets for sensorineural hearing loss

    Directory of Open Access Journals (Sweden)

    Masato eFujioka

    2014-12-01

    Full Text Available The inner ear was previously assumed to be an immune-privileged organ due to the existence of its tight junction-based blood-labyrinth barrier. However, studies performed during the past decade revealed that the mesenchymal region of the cochlea, including its lateral wall, is a common site of inflammation. Neutrophils do not enter this region, which is consistent with the old dogma; however, bone marrow-derived resident macrophages are always present in the spiral ligament of the lateral wall and are activated in response to various types of insults, including noise exposure, ischemia, mitochondrial damage and surgical stress. Recent studies have also revealed another type of immune cell, called perivascular melanocyte-like macrophages (PVM/Ms, in the stria vascularis. These dedicated antigen-presenting cells also control vascular contraction and permeability. This review discusses a series of reports regarding inflammatory/immune cells in the cochlear lateral wall, the pathways involved in cochlear damage and their potential as therapeutic targets.

  8. New therapeutic targets in idiopathic pulmonary fibrosis. Aiming to rein in runaway wound-healing responses.

    Science.gov (United States)

    Ahluwalia, Neil; Shea, Barry S; Tager, Andrew M

    2014-10-15

    Idiopathic pulmonary fibrosis (IPF) is a devastating disease, with a median survival as short as 3 years from the time of diagnosis and no pharmacological therapies yet approved by the U.S. Food and Drug Administration. To address the great unmet need for effective IPF therapy, a number of new drugs have recently been, or are now being, evaluated in clinical trials. The rationales for most of these therapeutic candidates are based on the current paradigm of IPF pathogenesis, in which recurrent injury to the alveolar epithelium is believed to drive aberrant wound healing responses, resulting in fibrosis rather than repair. Here we discuss drugs in recently completed or currently ongoing phase II and III IPF clinical trials in the context of their putative mechanisms of action and the aberrant repair processes they are believed to target: innate immune activation and polarization, fibroblast accumulation and myofibroblast differentiation, or extracellular matrix deposition and stiffening. Placed in this context, the positive results of recently completed trials of pirfenidone and nintedanib, and results that will come from ongoing trials of other agents, should provide valuable insights into the still-enigmatic pathogenesis of this disease, in addition to providing benefits to patients with IPF.

  9. Targeting Oxidatively Induced DNA Damage Response in Cancer: Opportunities for Novel Cancer Therapies

    Directory of Open Access Journals (Sweden)

    Pierpaola Davalli

    2018-01-01

    Full Text Available Cancer is a death cause in economically developed countries that results growing also in developing countries. Improved outcome through targeted interventions faces the scarce selectivity of the therapies and the development of resistance to them that compromise the therapeutic effects. Genomic instability is a typical cancer hallmark due to DNA damage by genetic mutations, reactive oxygen and nitrogen species, ionizing radiation, and chemotherapeutic agents. DNA lesions can induce and/or support various diseases, including cancer. The DNA damage response (DDR is a crucial signaling-transduction network that promotes cell cycle arrest or cell death to repair DNA lesions. DDR dysregulation favors tumor growth as downregulated or defective DDR generates genomic instability, while upregulated DDR may confer treatment resistance. Redox homeostasis deeply and capillary affects DDR as ROS activate/inhibit proteins and enzymes integral to DDR both in healthy and cancer cells, although by different routes. DDR regulation through modulating ROS homeostasis is under investigation as anticancer opportunity, also in combination with other treatments since ROS affect DDR differently in the patients during cancer development and treatment. Here, we highlight ROS-sensitive proteins whose regulation in oxidatively induced DDR might allow for selective strategies against cancer that are better tailored to the patients.

  10. New Therapeutic Targets in Idiopathic Pulmonary Fibrosis. Aiming to Rein in Runaway Wound-Healing Responses

    Science.gov (United States)

    Ahluwalia, Neil; Shea, Barry S.

    2014-01-01

    Idiopathic pulmonary fibrosis (IPF) is a devastating disease, with a median survival as short as 3 years from the time of diagnosis and no pharmacological therapies yet approved by the U.S. Food and Drug Administration. To address the great unmet need for effective IPF therapy, a number of new drugs have recently been, or are now being, evaluated in clinical trials. The rationales for most of these therapeutic candidates are based on the current paradigm of IPF pathogenesis, in which recurrent injury to the alveolar epithelium is believed to drive aberrant wound healing responses, resulting in fibrosis rather than repair. Here we discuss drugs in recently completed or currently ongoing phase II and III IPF clinical trials in the context of their putative mechanisms of action and the aberrant repair processes they are believed to target: innate immune activation and polarization, fibroblast accumulation and myofibroblast differentiation, or extracellular matrix deposition and stiffening. Placed in this context, the positive results of recently completed trials of pirfenidone and nintedanib, and results that will come from ongoing trials of other agents, should provide valuable insights into the still-enigmatic pathogenesis of this disease, in addition to providing benefits to patients with IPF. PMID:25090037

  11. Lp-mixed affine surface area

    Science.gov (United States)

    Wang, Weidong; Leng, Gangsong

    2007-11-01

    According to the three notions of mixed affine surface area, Lp-affine surface area and Lp-mixed affine surface area proposed by Lutwak, in this article, we give the concept of ith Lp-mixed affine surface area such that the first and second notions of Lutwak are its special cases. Further, some Lutwak's results are extended associated with this concept. Besides, applying this concept, we establish an inequality for the volumes and dual quermassintegrals of a class of star bodies.

  12. Manifolds with integrable affine shape operator

    Directory of Open Access Journals (Sweden)

    Daniel A. Joaquín

    2005-05-01

    Full Text Available This work establishes the conditions for the existence of vector fields with the property that theirs covariant derivative, with respect to the affine normal connection, be the affine shape operatorS in hypersurfaces. Some results are obtained from this property and, in particular, for some kind of affine decomposable hypersurfaces we explicitely get the actual vector fields.

  13. Affinity Spaces and 21st Century Learning

    Science.gov (United States)

    Gee, James Paul

    2017-01-01

    This article discusses video games as "attractors" to "affinity spaces." It argues that affinity spaces are key sites today where people teach and learn 21st Century skills. While affinity spaces are proliferating on the Internet as interest-and-passion-driven sites devoted to a common set of endeavors, they are not new, just…

  14. Using Affinity Diagrams to Evaluate Interactive Prototypes

    DEFF Research Database (Denmark)

    Lucero, Andrés

    2015-01-01

    our particular use of affinity diagramming in prototype evaluations. We reflect on a decade’s experience using affinity diagramming across a number of projects, both in industry and academia. Our affinity diagramming process in interaction design has been tailored and consists of four stages: creating...

  15. In vitro evolution and affinity-maturation with Coliphage qβ display.

    Directory of Open Access Journals (Sweden)

    Claudia Skamel

    Full Text Available The Escherichia coli bacteriophage, Qβ (Coliphage Qβ, offers a favorable alternative to M13 for in vitro evolution of displayed peptides and proteins due to high mutagenesis rates in Qβ RNA replication that better simulate the affinity maturation processes of the immune response. We describe a benchtop in vitro evolution system using Qβ display of the VP1 G-H loop peptide of foot-and-mouth disease virus (FMDV. DNA encoding the G-H loop was fused to the A1 minor coat protein of Qβ resulting in a replication-competent hybrid phage that efficiently displayed the FMDV peptide. The surface-localized FMDV VP1 G-H loop cross-reacted with the anti-FMDV monoclonal antibody (mAb SD6 and was found to decorate the corners of the Qβ icosahedral shell by electron microscopy. Evolution of Qβ-displayed peptides, starting from fully degenerate coding sequences corresponding to the immunodominant region of VP1, allowed rapid in vitro affinity maturation to SD6 mAb. Qβ selected under evolutionary pressure revealed a non-canonical, but essential epitope for mAb SD6 recognition consisting of an Arg-Gly tandem pair. Finally, the selected hybrid phages induced polyclonal antibodies in guinea pigs with good affinity to both FMDV and hybrid Qβ-G-H loop, validating the requirement of the tandem pair epitope. Qβ-display emerges as a novel framework for rapid in vitro evolution with affinity-maturation to molecular targets.

  16. Optimizing prognosis-related key miRNA-target interactions responsible for cancer metastasis.

    Science.gov (United States)

    Zhao, Hongying; Yuan, Huating; Hu, Jing; Xu, Chaohan; Liao, Gaoming; Yin, Wenkang; Xu, Liwen; Wang, Li; Zhang, Xinxin; Shi, Aiai; Li, Jing; Xiao, Yun

    2017-12-12

    Increasing evidence suggests that the abnormality of microRNAs (miRNAs) and their downstream targets is frequently implicated in the pathogenesis of human cancers, however, the clinical benefit of causal miRNA-target interactions has been seldom studied. Here, we proposed a computational method to optimize prognosis-related key miRNA-target interactions by combining transcriptome and clinical data from thousands of TCGA tumors across 16 cancer types. We obtained a total of 1,956 prognosis-related key miRNA-target interactions between 112 miRNAs and 1,443 their targets. Interestingly, these key target genes are specifically involved in tumor progression-related functions, such as 'cell adhesion' and 'cell migration'. Furthermore, they are most significantly correlated with 'tissue invasion and metastasis', a hallmark of metastasis, in ten distinct types of cancer through the hallmark analysis. These results implicated that the prognosis-related key miRNA-target interactions were highly associated with cancer metastasis. Finally, we observed that the combination of these key miRNA-target interactions allowed to distinguish patients with good prognosis from those with poor prognosis both in most TCGA cancer types and independent validation sets, highlighting their roles in cancer metastasis. We provided a user-friendly database named miRNATarget (freely available at http://biocc.hrbmu.edu.cn/miRNATar/), which provides an overview of the prognosis-related key miRNA-target interactions across 16 cancer types.

  17. National responses to global health targets: exploring policy transfer in the context of the UNAIDS '90-90-90' treatment targets in Ghana and Uganda.

    Science.gov (United States)

    McRobie, Ellen; Matovu, Fred; Nanyiti, Aisha; Nonvignon, Justice; Abankwah, Daniel Nana Yaw; Case, Kelsey K; Hallett, Timothy B; Hanefeld, Johanna; Conteh, Lesong

    2018-01-01

    Global health organizations frequently set disease-specific targets with the goal of eliciting adoption at the national-level; consideration of the influence of target setting on national policies, programme and health budgets is of benefit to those setting targets and those intended to respond. In 2014, the Joint United Nations Programme on HIV/AIDS set 'ambitious' treatment targets for country adoption: 90% of HIV-positive persons should know their status; 90% of those on treatment; 90% of those achieving viral suppression. Using case studies from Ghana and Uganda, we explore how the target and its associated policy content have been adopted at the national level. That is whether adoption is in rhetoric only or supported by programme, policy or budgetary changes. We review 23 (14 from Ghana, 9 from Uganda) national policy, operational and strategic documents for the HIV response and assess commitments to '90-90-90'. In-person semi-structured interviews were conducted with purposively sampled key informants (17 in Ghana, 20 in Uganda) involved in programme-planning and resource allocation within HIV to gain insight into factors facilitating adoption of 90-90-90. Interviews were transcribed and analysed thematically, inductively and deductively, guided by pre-existing policy theories, including Dolowitz and Marsh's policy transfer framework to describe features of the transfer and the Global Health Advocacy and Policy Project framework to explain observations. Regardless of notable resource constraints, transfer of the 90-90-90 targets was evident beyond rhetoric with substantial shifts in policy and programme activities. In both countries, there was evidence of attempts to minimize resource constraints by seeking programme efficiencies, prioritization of programme activities and devising domestic financing mechanisms; however, significant resource gaps persist. An effective health network, comprised of global and local actors, mediated the adoption and adaptation

  18. Does corporate social responsibility put reputation at risk by inviting activist targeting? An empirical test among European SMEs.

    NARCIS (Netherlands)

    Graafland, Johan

    Corporate social responsibility (CSR) is believed to improve a company’s reputation. However,CSR may also put reputation at risk by making the company a more attractive target for activists’campaigns. We test this effect on a sample of 1355 European small and medium-sized enterprises(SMEs). We find

  19. Discovery of PF-06928215 as a high affinity inhibitor of cGAS enabled by a novel fluorescence polarization assay.

    Science.gov (United States)

    Hall, Justin; Brault, Amy; Vincent, Fabien; Weng, Shawn; Wang, Hong; Dumlao, Darren; Aulabaugh, Ann; Aivazian, Dikran; Castro, Dana; Chen, Ming; Culp, Jeffrey; Dower, Ken; Gardner, Joseph; Hawrylik, Steven; Golenbock, Douglas; Hepworth, David; Horn, Mark; Jones, Lyn; Jones, Peter; Latz, Eicke; Li, Jing; Lin, Lih-Ling; Lin, Wen; Lin, David; Lovering, Frank; Niljanskul, Nootaree; Nistler, Ryan; Pierce, Betsy; Plotnikova, Olga; Schmitt, Daniel; Shanker, Suman; Smith, James; Snyder, William; Subashi, Timothy; Trujillo, John; Tyminski, Edyta; Wang, Guoxing; Wong, Jimson; Lefker, Bruce; Dakin, Leslie; Leach, Karen

    2017-01-01

    Cyclic GMP-AMP synthase (cGAS) initiates the innate immune system in response to cytosolic dsDNA. After binding and activation from dsDNA, cGAS uses ATP and GTP to synthesize 2', 3' -cGAMP (cGAMP), a cyclic dinucleotide second messenger with mixed 2'-5' and 3'-5' phosphodiester bonds. Inappropriate stimulation of cGAS has been implicated in autoimmune disease such as systemic lupus erythematosus, thus inhibition of cGAS may be of therapeutic benefit in some diseases; however, the size and polarity of the cGAS active site makes it a challenging target for the development of conventional substrate-competitive inhibitors. We report here the development of a high affinity (KD = 200 nM) inhibitor from a low affinity fragment hit with supporting biochemical and structural data showing these molecules bind to the cGAS active site. We also report a new high throughput cGAS fluorescence polarization (FP)-based assay to enable the rapid identification and optimization of cGAS inhibitors. This FP assay uses Cy5-labelled cGAMP in combination with a novel high affinity monoclonal antibody that specifically recognizes cGAMP with no cross reactivity to cAMP, cGMP, ATP, or GTP. Given its role in the innate immune response, cGAS is a promising therapeutic target for autoinflammatory disease. Our results demonstrate its druggability, provide a high affinity tool compound, and establish a high throughput assay for the identification of next generation cGAS inhibitors.

  20. Discovery of PF-06928215 as a high affinity inhibitor of cGAS enabled by a novel fluorescence polarization assay

    Energy Technology Data Exchange (ETDEWEB)

    Hall, Justin; Brault, Amy; Vincent, Fabien; Weng, Shawn; Wang, Hong; Dumlao, Darren; Aulabaugh, Ann; Aivazian, Dikran; Castro, Dana; Chen, Ming; Culp, Jeffrey; Dower, Ken; Gardner, Joseph; Hawrylik, Steven; Golenbock, Douglas; Hepworth, David; Horn, Mark; Jones, Lyn; Jones, Peter; Latz, Eicke; Li, Jing; Lin, Lih-Ling; Lin, Wen; Lin, David; Lovering, Frank; Niljanskul, Nootaree; Nistler, Ryan; Pierce, Betsy; Plotnikova, Olga; Schmitt, Daniel; Shanker, Suman; Smith, James; Snyder, William; Subashi, Timothy; Trujillo, John; Tyminski, Edyta; Wang, Guoxing; Wong, Jimson; Lefker, Bruce; Dakin, Leslie; Leach, Karen (UMASS, MED); (Pfizer)

    2017-09-21

    Cyclic GMP-AMP synthase (cGAS) initiates the innate immune system in response to cytosolic dsDNA. After binding and activation from dsDNA, cGAS uses ATP and GTP to synthesize 2', 3' -cGAMP (cGAMP), a cyclic dinucleotide second messenger with mixed 2'-5' and 3'-5' phosphodiester bonds. Inappropriate stimulation of cGAS has been implicated in autoimmune disease such as systemic lupus erythematosus, thus inhibition of cGAS may be of therapeutic benefit in some diseases; however, the size and polarity of the cGAS active site makes it a challenging target for the development of conventional substrate-competitive inhibitors. We report here the development of a high affinity (KD = 200 nM) inhibitor from a low affinity fragment hit with supporting biochemical and structural data showing these molecules bind to the cGAS active site. We also report a new high throughput cGAS fluorescence polarization (FP)-based assay to enable the rapid identification and optimization of cGAS inhibitors. This FP assay uses Cy5-labelled cGAMP in combination with a novel high affinity monoclonal antibody that specifically recognizes cGAMP with no cross reactivity to cAMP, cGMP, ATP, or GTP. Given its role in the innate immune response, cGAS is a promising therapeutic target for autoinflammatory disease. Our results demonstrate its druggability, provide a high affinity tool compound, and establish a high throughput assay for the identification of next generation cGAS inhibitors.

  1. Quiet Eye Duration Is Responsive to Variability of Practice and to the Axis of Target Changes

    Science.gov (United States)

    Horn, Robert R.; Okumura, Michelle S.; Alexander, Melissa G. F.; Gardin, Fredrick A.; Sylvester, Curtis T.

    2012-01-01

    We tested the hypothesis that quiet eye, the final fixation before the initiation of a movement in aiming tasks, is used to scale the movement's parameters. Two groups of 12 participants (N = 24) threw darts to targets in the horizontal and vertical axes under conditions of higher (random) or lower (blocked) target variability. Supporting our…

  2. The affine quantum gravity programme

    International Nuclear Information System (INIS)

    Klauder, John R

    2002-01-01

    The central principle of affine quantum gravity is securing and maintaining the strict positivity of the matrix { g-hat ab (x)} composed of the spatial components of the local metric operator. On spectral grounds, canonical commutation relations are incompatible with this principle, and they must be replaced by noncanonical, affine commutation relations. Due to the partial second-class nature of the quantum gravitational constraints, it is advantageous to use the recently developed projection operator method, which treats all quantum constraints on an equal footing. Using this method, enforcement of regularized versions of the gravitational operator constraints is formulated quite naturally by means of a novel and relatively well-defined functional integral involving only the same set of variables that appears in the usual classical formulation. It is anticipated that skills and insight to study this formulation can be developed by studying special, reduced-variable models that still retain some basic characteristics of gravity, specifically a partial second-class constraint operator structure. Although perturbatively nonrenormalizable, gravity may possibly be understood nonperturbatively from a hard-core perspective that has proved valuable for specialized models. Finally, developing a procedure to pass to the genuine physical Hilbert space involves several interconnected steps that require careful coordination

  3. Affine-projective field laws

    International Nuclear Information System (INIS)

    Murphy, G.L.

    1975-01-01

    The general topic of geometric unified field theories is discussed in the first section. Some reasons are given for pursuing such theories, and some criticisms are considered. The second section develops the fundamental equations of a purely affine theory which is invariant under projective transformations of the affine connection. This theory is a generalization of that of Schrodinger. Possible identifications for the space-time metric are considered in Sec. III. Sections IV and V deal with the limits of pure gravitation and electrodynamics. In the symmetric limit, Einstein's vacuum equations with cosmological term are recovered. The theory also contains a generalized electrodynamic set of equations which is very similar to the Born-Infeld set. In the weak-field approximation, a finite mass must be attributed to the photon. The problem of motion for charges is discussed here, and it is argued that criticisms of unified field theories because of a supposed inability to produce the Lorentz force law are probably not justified. Three more speculative sections deal with possible explanations of nuclear forces, the spin-torsion relation, and particle structure

  4. Specificity and affinity quantification of protein-protein interactions.

    Science.gov (United States)

    Yan, Zhiqiang; Guo, Liyong; Hu, Liang; Wang, Jin

    2013-05-01

    Most biological processes are mediated by the protein-protein interactions. Determination of the protein-protein structures and insight into their interactions are vital to understand the mechanisms of protein functions. Currently, compared with the isolated protein structures, only a small fraction of protein-protein structures are experimentally solved. Therefore, the computational docking methods play an increasing role in predicting the structures and interactions of protein-protein complexes. The scoring function of protein-protein interactions is the key responsible for the accuracy of the computational docking. Previous scoring functions were mostly developed by optimizing the binding affinity which determines the stability of the protein-protein complex, but they are often lack of the consideration of specificity which determines the discrimination of native protein-protein complex against competitive ones. We developed a scoring function (named as SPA-PP, specificity and affinity of the protein-protein interactions) by incorporating both the specificity and affinity into the optimization strategy. The testing results and comparisons with other scoring functions show that SPA-PP performs remarkably on both predictions of binding pose and binding affinity. Thus, SPA-PP is a promising quantification of protein-protein interactions, which can be implemented into the protein docking tools and applied for the predictions of protein-protein structure and affinity. The algorithm is implemented in C language, and the code can be downloaded from http://dl.dropbox.com/u/1865642/Optimization.cpp.

  5. Structural insights into a high affinity nanobody:antigen complex by homology modelling.

    Science.gov (United States)

    Skottrup, Peter Durand

    2017-09-01

    Porphyromonas gingivalis is a major periodontitis-causing pathogens. P. gingivalis secrete a cysteine protease termed RgpB, which is specific for Arg-Xaa bonds in substrates. Recently, a nanobody-based assay was used to demonstrate that RgpB could represent a novel diagnostic target, thereby simplifying. P. gingivalis detection. The nanobody, VHH7, had a high binding affinity and was specific for RgpB, when tested towards the highly identical RgpA. In this study a homology model of VHH7 was build. The complementarity determining regions (CDR) comprising the paratope residues responsible for RgpB binding were identified and used as input to the docking. Furthermore, residues likely involved in the RgpB epitope was identified based upon RgpB:RgpA alignment and analysis of residue surface accessibility. CDR residues and putitative RgpB epitope residues were used as input to an information-driven flexible docking approach using the HADDOCK server. Analysis of the VHH7:RgpB model demonstrated that the epitope was found in the immunoglobulin-like domain and residue pairs located at the molecular paratope:epitope interface important for complex stability was identified. Collectively, the VHH7 homology model and VHH7:RgpB docking supplies knowledge of the residues involved in the high affinity interaction. This information could prove valuable in the design of an antibody-drug conjugate for specific RgpB targeting. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Tracking targeted bimodal nanovaccines: immune responses and routing in cells, tissue, and whole organism.

    Science.gov (United States)

    Cruz, Luis J; Tacken, Paul J; Zeelenberg, Ingrid S; Srinivas, Mangala; Bonetto, Fernando; Weigelin, Bettina; Eich, Christina; de Vries, I Jolanda; Figdor, Carl G

    2014-12-01

    Dendritic cells (DCs) are the most potent antigen-presenting cells (APCs), involved in the induction of immunity and currently exploited for antitumor immunotherapies. An optimized noninvasive imaging modality capable of determining and quantifying DC-targeted nanoparticle (NP) trajectories could provide valuable information regarding therapeutic vaccine outcome. Here, targeted poly(d,l-lactide-co-glycolide) nanoparticles (PLGA NPs) recognizing DC receptors were equipped with superparamagnetic iron oxide particles (SPIO) or gold nanoparticles with fluorescently labeled antigen. The fluorescent label allowed for rapid analysis and quantification of DC-specific uptake of targeted PLGA NPs in comparison to uptake by other cells. Transmission electron microscopy (TEM) showed that a fraction of the encapsulated antigen reached the lysosomal compartment of DCs, where SPIO and gold were already partially released. However, part of the PLGA NPs localized within the cytoplasm, as confirmed by confocal microscopy. DCs targeted with NPs carrying SPIO or fluorescent antigen were detected within lymph nodes as early as 1 h after injection by magnetic resonance imaging (MRI). Despite the fact that targeting did not markedly affect PLGA NP biodistribution on organism and tissue level, it increased delivery of NPs to DCs residing in peripheral lymph nodes and resulted in enhanced T cell proliferation. In conclusion, two imaging agents within a single carrier allows tracking of targeted PLGA NPs at the subcellular, cellular, and organismal levels, thereby facilitating the rational design of in vivo targeted vaccination strategies.

  7. Target activation and radiological response of ARIES-IFE dry wall chamber

    International Nuclear Information System (INIS)

    El-Guebaly, L.A.; Henderson, D.L.; Wilson, P.P.H.; Abdou, A.E.

    2002-01-01

    The choice of target coating and hohlraum wall materials is among the most critical decisions to be made for inertial fusion energy (IFE) designs. Gold and gold/gadolinium have long been considered to be the coating and hohlraum wall materials of choice for direct drive (DD) and indirect drive (ID) targets, respectively, offering high target performance and low beam energy losses. More recently, a variety of other materials have been considered, including W, Pb, Pt, Pd, and Ag for the DD target coating and Au, W, Pb, Hg, Ta, Cs, and Hf for the hohlraum wall of the ID target. The choice of the coating/hohlraum material is a tradeoff between the target design elements such as safety, target physics, economics, etc. We identified the key safety issues and have investigated the neutron-induced irradiation effects of the candidate coating/hohlraum materials using the radiation chamber conditions of the ARIES-IFE dry wall concept. The safety requirements have specific impacts in terms of the coating/hohlraum materials choice

  8. Are predictions of cancer response to targeted drugs, based on effects in unrelated tissues, the 'Black Swan' events?

    Science.gov (United States)

    Kurbel, Beatrica; Golem, Ante Zvonimir; Kurbel, Sven

    2015-01-01

    Adverse effects of targeted drugs on normal tissues can predict the cancer response. Rash correlates with efficacy of erlotinib, cetuximab and gefitinib and onset of arterial hypertension with response to bevacizumab, sunitinib, axitinib and sorafenib, possible examples of 'Black Swan' events, unexpected scientific observations, as described by Karl Popper in 1935. The proposition is that our patients have individual intrinsic variants of cell growth control, important for tumor response and adverse effects on tumor-unrelated tissue. This means that the lack of predictive side effects in healthy tissue is linked with poor results of tumor therapy when tumor resistance is caused by mechanisms that protect all cells of that patient from the targeted drug effects.

  9. Purification of CD47-streptavidin fusion protein from bacterial lysate using biotin-agarose affinity chromatography.

    Science.gov (United States)

    Salehi, Nasrin; Peng, Ching-An

    2016-07-08

    CD47 is a widely expressed transmembrane glycoprotein that modulates the activity of a plethora of immune cells via its extracellular domain. Therefore, CD47 plays important roles in the regulation of immune responses and may serve as targets for the development of immunotherapeutic agents. To make sure CD47 functionality is intact under the process of protein conjugation, CD47-streptavidin fusion protein was expressed and purified because it can easily bind to biotin-tagged materials via the unique biotin-streptavidin affinity. In this study, gene sequences of CD47 extracellular domain (CD47ECD) and core streptavidin (coreSA) with a total 834 bp were inserted into pET20b plasmid to construct recombinant plasmid encoding CD47-SA fusion gene. After bacteria transformation, the CD47-SA fusion protein was expressed by isopropyl-β-d-thiogalactopyranoside (IPTG) induction. The collected bacteria lysate was loaded on biotinylated agarose to proceed the purification of CD47-SA fusion protein. Due to the unexpected high affinity between biotin and coreSA, standard washing and elution approaches (e.g., varying pH, using biotin, and applying guanidine hydrochloride) reported for biotin-streptavidin affinity chromatography were not able to separate the target fusion protein. Instead, using low concentration of the non-ionic detergent Triton X-100 followed with alkaline buffer could efficiently weaken the binding between biotin and coreSA, thereby eluting out CD47-SA fusion protein from the biotin agarose column. The purified CD47-SA fusion protein was further characterized by molecular biology methods and its antiphagocytic functionality was confirmed by the phagocytosis assay. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:949-958, 2016. © 2016 American Institute of Chemical Engineers.

  10. Large-scale integration of small molecule-induced genome-wide transcriptional responses, Kinome-wide binding affinities and cell-growth inhibition profiles reveal global trends characterizing systems-level drug action

    Directory of Open Access Journals (Sweden)

    Dusica eVidovic

    2014-09-01

    Full Text Available The Library of Integrated Network-based Cellular Signatures (LINCS project is a large-scale coordinated effort to build a comprehensive systems biology reference resource. The goals of the program include the generation of a very large multidimensional data matrix and informatics and computational tools to integrate, analyze, and make the data readily accessible. LINCS data include genome-wide transcriptional signatures, biochemical protein binding profiles, cellular phenotypic response profiles and various other datasets for a wide range of cell model systems and molecular and genetic perturbations. Here we present a partial survey of this data facilitated by data standards and in particular a robust compound standardization workflow; we integrated several types of LINCS signatures and analyzed the results with a focus on mechanism of action and chemical compounds. We illustrate how kinase targets can be related to disease models and relevant drugs. We identified some fundamental trends that appear to link Kinome binding profiles and transcriptional signatures to chemical information and biochemical binding profiles to transcriptional responses independent of chemical similarity. To fill gaps in the datasets we developed and applied predictive models. The results can be interpreted at the systems level as demonstrated based on a large number of signaling pathways. We can identify clear global relationships, suggesting robustness of cellular responses to chemical perturbation. Overall, the results suggest that chemical similarity is a useful measure at the systems level, which would support phenotypic drug optimization efforts. With this study we demonstrate the potential of such integrated analysis approaches and suggest prioritizing further experiments to fill the gaps in the current data.

  11. Geospatial analysis of emergency department visits for targeting community-based responses to the opioid epidemic.

    Directory of Open Access Journals (Sweden)

    Daniel A Dworkis

    Full Text Available The opioid epidemic in the United States carries significant morbidity and mortality and requires a coordinated response among emergency providers, outpatient providers, public health departments, and communities. Anecdotally, providers across the spectrum of care at Massachusetts General Hospital (MGH in Boston, MA have noticed that Charlestown, a community in northeast Boston, has been particularly impacted by the opioid epidemic and needs both emergency and longer-term resources. We hypothesized that geospatial analysis of the home addresses of patients presenting to the MGH emergency department (ED with opioid-related emergencies might identify "hot spots" of opioid-related healthcare needs within Charlestown that could then be targeted for further investigation and resource deployment. Here, we present a geospatial analysis at the United States census tract level of the home addresses of all patients who presented to the MGH ED for opioid-related emergency visits between 7/1/2012 and 6/30/2015, including 191 visits from 100 addresses in Charlestown, MA. Among the six census tracts that comprise Charlestown, we find a 9.5-fold difference in opioid-related ED visits, with 45% of all opioid-related visits from Charlestown originating in tract 040401. The signal from this census tract remains strong after adjusting for population differences between census tracts, and while this tract is one of the higher utilizing census tracts in Charlestown of the MGH ED for all cause visits, it also has a 2.9-fold higher rate of opioid-related visits than the remainder of Charlestown. Identifying this hot spot of opioid-related emergency needs within Charlestown may help re-distribute existing resources efficiently, empower community and ED-based physicians to advocate for their patients, and serve as a catalyst for partnerships between MGH and local community groups. More broadly, this analysis demonstrates that EDs can use geospatial analysis to address

  12. EBV-encoded miRNAs target ATM-mediated response in nasopharyngeal carcinoma.

    Science.gov (United States)

    Lung, Raymond W-M; Hau, Pok-Man; Yu, Ken H-O; Yip, Kevin Y; Tong, Joanna H-M; Chak, Wing-Po; Chan, Anthony W-H; Lam, Ka-Hei; Lo, Angela Kwok-Fung; Tin, Edith K-Y; Chau, Shuk-Ling; Pang, Jesse C-S; Kwan, Johnny S-H; Busson, Pierre; Young, Lawrence S; Yap, Lee-Fah; Tsao, Sai-Wah; To, Ka-Fai; Lo, Kwok-Wai

    2018-04-01

    Nasopharyngeal carcinoma (NPC) is a highly invasive epithelial malignancy that is prevalent in southern China and Southeast Asia. It is consistently associated with latent Epstein-Barr virus (EBV) infection. In NPC, miR-BARTs, the EBV-encoded miRNAs derived from BamH1-A rightward transcripts, are abundantly expressed and contribute to cancer development by targeting various cellular and viral genes. In this study, we establish a comprehensive transcriptional profile of EBV-encoded miRNAs in a panel of NPC patient-derived xenografts and an EBV-positive NPC cell line by small RNA sequencing. Among the 40 miR-BARTs, predominant expression of 22 miRNAs was consistently detected in these tumors. Among the abundantly expressed EBV-miRNAs, BART5-5p, BART7-3p, BART9-3p, and BART14-3p could negatively regulate the expression of a key DNA double-strand break (DSB) repair gene, ataxia telangiectasia mutated (ATM), by binding to multiple sites on its 3'-UTR. Notably, the expression of these four miR-BARTs represented more than 10% of all EBV-encoded miRNAs in tumor cells, while downregulation of ATM expression was commonly detected in all of our tested sequenced samples. In addition, downregulation of ATM was also observed in primary NPC tissues in both qRT-PCR (16 NP and 45 NPC cases) and immunohistochemical staining (35 NP and 46 NPC cases) analysis. Modulation of ATM expression by BART5-5p, BART7-3p, BART9-3p, and BART14-3p was demonstrated in the transient transfection assays. These findings suggest that EBV uses miRNA machinery as a key mechanism to control the ATM signaling pathway in NPC cells. By suppressing these endogenous miR-BARTs in EBV-positive NPC cells, we further demonstrated the novel function of miR-BARTs in inhibiting Zta-induced lytic reactivation. These findings imply that the four viral miRNAs work co-operatively to modulate ATM activity in response to DNA damage and to maintain viral latency, contributing to the tumorigenesis of NPC. © 2017 The Authors

  13. EBV‐encoded miRNAs target ATM‐mediated response in nasopharyngeal carcinoma

    Science.gov (United States)

    Lung, Raymond W‐M; Hau, Pok‐Man; Yu, Ken H‐O; Yip, Kevin Y; Tong, Joanna H‐M; Chak, Wing‐Po; Chan, Anthony W‐H; Lam, Ka‐Hei; Lo, Angela Kwok‐Fung; Tin, Edith K‐Y; Chau, Shuk‐Ling; Pang, Jesse C‐S; Kwan, Johnny S‐H; Busson, Pierre; Young, Lawrence S; Yap, Lee‐Fah; Tsao, Sai‐Wah

    2018-01-01

    Abstract Nasopharyngeal carcinoma (NPC) is a highly invasive epithelial malignancy that is prevalent in southern China and Southeast Asia. It is consistently associated with latent Epstein–Barr virus (EBV) infection. In NPC, miR‐BARTs, the EBV‐encoded miRNAs derived from BamH1‐A rightward transcripts, are abundantly expressed and contribute to cancer development by targeting various cellular and viral genes. In this study, we establish a comprehensive transcriptional profile of EBV‐encoded miRNAs in a panel of NPC patient‐derived xenografts and an EBV‐positive NPC cell line by small RNA sequencing. Among the 40 miR‐BARTs, predominant expression of 22 miRNAs was consistently detected in these tumors. Among the abundantly expressed EBV‐miRNAs, BART5‐5p, BART7‐3p, BART9‐3p, and BART14‐3p could negatively regulate the expression of a key DNA double‐strand break (DSB) repair gene, ataxia telangiectasia mutated (ATM), by binding to multiple sites on its 3'‐UTR. Notably, the expression of these four miR‐BARTs represented more than 10% of all EBV‐encoded miRNAs in tumor cells, while downregulation of ATM expression was commonly detected in all of our tested sequenced samples. In addition, downregulation of ATM was also observed in primary NPC tissues in both qRT‐PCR (16 NP and 45 NPC cases) and immunohistochemical staining (35 NP and 46 NPC cases) analysis. Modulation of ATM expression by BART5‐5p, BART7‐3p, BART9‐3p, and BART14‐3p was demonstrated in the transient transfection assays. These findings suggest that EBV uses miRNA machinery as a key mechanism to control the ATM signaling pathway in NPC cells. By suppressing these endogenous miR‐BARTs in EBV‐positive NPC cells, we further demonstrated the novel function of miR‐BARTs in inhibiting Zta‐induced lytic reactivation. These findings imply that the four viral miRNAs work co‐operatively to modulate ATM activity in response to DNA damage and to maintain viral latency

  14. MicroRNA-125b Affects Vascular Smooth Muscle Cell Function by Targeting Serum Response Factor

    Directory of Open Access Journals (Sweden)

    Zhibo Chen

    2018-04-01

    Full Text Available Background/Aims: Increasing evidence links microRNAs to the pathogenesis of peripheral vascular disease. We recently found microRNA-125b (miR-125b to be one of the most significantly down‑regulated microRNAs in human arteries with arteriosclerosis obliterans (ASO of the lower extremities. However, its function in the process of ASO remains unclear. This study aimed to investigate the expression, regulatory mechanisms, and functions of miR-125b in the process of ASO. Methods: Using the tissue explants adherent method, vascular smooth muscle cells (VSMCs were prepared for this study. A rat carotid artery balloon injury model was constructed to simulate the development of vascular neointima, and a lentiviral transduction system was used to overexpress serum response factor (SRF or miR-125b. Quantitative real‑time PCR (qRT‑PCR was used to detect the expression levels of miR‑125b and SRF mRNA. Western blotting was performed to determine the expression levels of SRF and Ki67. In situ hybridization analysis was used to analyze the location and expression levels of miR-125b. CCK-8 and EdU assays were used to assess cell proliferation, and transwell and wound closure assays were performed to measure cell migration. Flow cytometry was used to evaluate cell apoptosis, and a dual-luciferase reporter assay was conducted to examine the effects of miR‑125b on SRF. Immunohistochemistry and immunofluorescence analyses were performed to analyze the location and expression levels of SRF and Ki67. Results: miR-125b expression was decreased in ASO arteries and platelet-derived growth factor (PDGF-BB-stimulated VSMCs. miR-125b suppressed VSMC proliferation and migration but promoted VSMC apoptosis. SRF was determined to be a direct target of miR-125b. Exogenous miR-125b expression modulated SRF expression and inhibited vascular neointimal formation in balloon-injured rat carotid arteries. Conclusions: These findings demonstrate a specific role of the mi

  15. DFT study on the effect of exocyclic substituents on the proton affinity of 1-methylimidazole

    International Nuclear Information System (INIS)

    Liu, Haining; Bara, Jason E.; Turner, C. Heath

    2013-01-01

    Highlights: • DFT calculations are used to predict the proton affinity of 1-methylimidazoles. • The electron-withdrawing groups dominate the predicted proton affinity. • The effects of multiple substituents on the proton affinity can be accurately predicted. • Large compound libraries can be screened for imidazoles with tailored reactivity. - Abstract: A deeper understanding of the acid/base properties of imidazole derivatives will aid the development of solvents, polymer membranes and other materials that can be used for CO 2 capture and acid gas removal. In this study, we employ density functional theory calculations to investigate the effect of various electron-donating and electron-withdrawing groups on the proton affinity of 1-methylimidazole. We find that electron-donating groups are able to increase the proton affinity relative to 1-methylimidazole, i.e., making the molecule more basic. In contrast, electron-withdrawing groups cause a decrease of the proton affinity. When multiple substituents are present, their effects on the proton affinity were found to be additive. This finding offers a quick approach for predicting and targeting the proton affinities of this series of molecules, and we show the strong correlation between the calculated proton affinities and experimental pK a values

  16. Complementary three-dimensional quantitative structure-activity relationship modeling of binding affinity and functional potency

    DEFF Research Database (Denmark)

    Tosco, Paolo; Ahring, Philip K; Dyhring, Tino

    2009-01-01

    Complementary 3D-QSAR modeling of binding affinity and functional potency is proposed as a tool to pinpoint the molecular features of the ligands, and the corresponding amino acids in the receptor, responsible for high affinity binding vs those driving agonist behavior and receptor activation. Th...

  17. RNF111/Arkadia is a SUMO-targeted ubiquitin ligase that facilitates the DNA damage response

    DEFF Research Database (Denmark)

    Poulsen, Sara L; Hansen, Rebecca K; Wagner, Sebastian A

    2013-01-01

    nonproteolytic, K63-linked ubiquitylation of SUMOylated target proteins. We demonstrate that RNF111 promoted ubiquitylation of SUMOylated XPC (xeroderma pigmentosum C) protein, a central DNA damage recognition factor in nucleotide excision repair (NER) extensively regulated by ultraviolet (UV...

  18. Development of an aptamer-based affinity purification method for vascular endothelial growth factor

    Directory of Open Access Journals (Sweden)

    Maren Lönne

    2015-12-01

    Full Text Available Since aptamers bind their targets with high affinity and specificity, they are promising alternative ligands in protein affinity purification. As aptamers are chemically synthesized oligonucleotides, they can be easily produced in large quantities regarding GMP conditions allowing their application in protein production for therapeutic purposes. Several advantages of aptamers compared to antibodies are described in general within this paper. Here, an aptamer directed against the human Vascular Endothelial Growth Factor (VEGF was used as affinity ligand for establishing a purification platform for VEGF in small scale. The aptamer was covalently immobilized on magnetic beads in a controlled orientation resulting in a functional active affinity matrix. Target binding was optimized by introduction of spacer molecules and variation of aptamer density. Further, salt-induced target elution was demonstrated as well as VEGF purification from a complex protein mixture proving the specificity of protein-aptamer binding.

  19. Platelet microparticle-inspired clot-responsive nanomedicine for targeted fibrinolysis.

    Science.gov (United States)

    Pawlowski, Christa L; Li, Wei; Sun, Michael; Ravichandran, Kavya; Hickman, DaShawn; Kos, Clarissa; Kaur, Gurbani; Sen Gupta, Anirban

    2017-06-01

    Intravascular administration of plasminogen activators is a clinically important thrombolytic strategy to treat occlusive vascular conditions. A major issue with this strategy is the systemic off-target drug action, which affects hemostatic capabilities and causes substantial hemorrhagic risks. This issue can be potentially resolved by designing technologies that allow thrombus-targeted delivery and site-specific action of thrombolytic drugs. To this end, leveraging a liposomal platform, we have developed platelet microparticle (PMP)-inspired nanovesicles (PMINs), that can protect encapsulated thrombolytic drugs in circulation to prevent off-target uptake and action, anchor actively onto thrombus via PMP-relevant molecular mechanisms and allow drug release via thrombus-relevant enzymatic trigger. Specifically, the PMINs can anchor onto thrombus via heteromultivalent ligand-mediated binding to active platelet integrin GPIIb-IIIa and P-selectin, and release the thrombolytic payload due to vesicle destabilization triggered by clot-relevant enzyme phospholipase-A 2 . Here we report on the evaluation of clot-targeting efficacy, lipase-triggered drug release and resultant thrombolytic capability of the PMINs in vitro, and subsequently demonstrate that intravenous delivery of thrombolytic-loaded PMINs can render targeted fibrinolysis without affecting systemic hemostasis, in vivo, in a carotid artery thrombosis model in mice. Our studies establish significant promise of the PMIN technology for safe and site-targeted nanomedicine therapies in the vascular compartment. Copyright © 2017. Published by Elsevier Ltd.

  20. Diversity of the murine antibody response targeting influenza A(H1N1pdm09) hemagglutinin.

    Science.gov (United States)

    Wilson, Jason R; Tzeng, Wen-Pin; Spesock, April; Music, Nedzad; Guo, Zhu; Barrington, Robert; Stevens, James; Donis, Ruben O; Katz, Jacqueline M; York, Ian A

    2014-06-01

    diversity of the anti-hemagglutinin antibody response, as well as characterizing the cross-reactivity, affinity, and molecular nature of the antibody response. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. Novel trends in affinity biosensors: current challenges and perspectives

    International Nuclear Information System (INIS)

    Arugula, Mary A; Simonian, Aleksandr

    2014-01-01

    Molecular biorecognition processes facilitate physical and biochemical interactions between molecules in all crucial metabolic pathways. Perhaps the target analyte and the biorecognition element interactions have the most impactful use in biosensing applications. Traditional analytical sensing systems offer excellent biorecognition elements with the ability to detect and determine the presence of analytes. High affinity antibodies and DNA play an important role in the development of affinity biosensors based on electrochemical, optical and mass sensitive approaches. Advancements in this area routinely employ labels, label free, nanoparticles, multifunctional matrices, carbon nanotubes and other methods to meet the requirements of its own application. However, despite increasing affinity ceilings for conventional biosensors, the field draws back in meeting specifically important demands, such as long-term stability, ultrasensitivity, rapid detection, extreme selectivity, strong biological base, calibration, in vivo measurements, regeneration, satisfactory performance and ease of production. Nevertheless, recent efforts through this line have produced novel high-tech nanosensing systems such as ‘aptamers’ and ‘phages’ which exhibit high-throughput sensing. Aptamers and phages are powerful tools that excel over antibodies in sensibility, stability, multi-detection, in vivo measurements and regeneration. Phages are superior in stability, screening for affinity-based target molecules ranging from small to proteins and even cells, and easy production. In this review, we focus mainly on recent developments in affinity-based biosensors such as immunosensors, DNA sensors, emphasizing aptasensors and phage-based biosensors basing on novel electrochemical, optical and mass sensitive detection techniques. We also address enzyme inhibition-based biosensors and the current problems associated with the above sensors and their future perspectives. (topical review)

  2. Expression Profile of Stress-responsive Arabidopsis thaliana miRNAs and their Target Genes in Response to Inoculation with Pectobacterium carotovorum subsp. carotovorum.

    Science.gov (United States)

    Djami-Tchatchou, A T; Ntushelo, K

    2017-01-01

    Pectobacterium carotovorum subsp. carotovorum (Pcc) is a soft rot bacterium which upon entry into the plant macerates plant tissues by producing plant cell wall degrading enzymes. It has a wide host range which includes carrot, potato, tomato, leafy greens, squash and other cucurbits, onion, green peppers and cassava. During plant-microbe interactions, one of the ways of plant response to pathogen infection is through the small RNA silencing mechanism. Under pathogen attack the plant utilizes microRNAs to regulate gene expression by means of mediating gene silencing at transcriptional and post-transcriptional level. This study aims to assess for the first time, the expression profile of some stress-responsive miRNA and differential expression pattern of their target genes in Arabidopsis thaliana inoculated with Pcc. Leaves of five weeks old Arabidopsis thaliana plants were infected with Pcc and the quantitative real time-PCR, was used to investigate after 0, 24, 48 and 72 h post infection, the expression profiling of the stress-responsive miRNAs which include: miR156, miR159, miR169, miR393, miR396 miR398, miR399 and miR408 along with their target genes which include: Squamosa promoter-binding-like protein, myb domain protein 101, nuclear factor Y subunit A8, concanavalin A-like lectin protein kinase, growth regulating factor 4, copper superoxide dismutase, ubiquitin-protein ligase and plantacyanin respectively. The findings showed that the overexpression of 6 miRNAs at 24, 48 and 72 h after infection resulted in the repression of their target genes and the expression of 2 miRNAs didn't affect their target genes. These results provide the first indication of the miRNAs role in response to the infection of Pcc in A. thaliana and open new vistas for a better understanding of miRNA regulation of plant response to Pcc.

  3. The Salmonella Effector Protein SopA Modulates Innate Immune Responses by Targeting TRIM E3 Ligase Family Members.

    Directory of Open Access Journals (Sweden)

    Jana Kamanova

    2016-04-01

    Full Text Available Salmonella Typhimurium stimulates inflammatory responses in the intestinal epithelium, which are essential for its ability to replicate within the intestinal tract. Stimulation of these responses is strictly dependent on the activity of a type III secretion system encoded within its pathogenicity island 1, which through the delivery of effector proteins, triggers signaling pathways leading to inflammation. One of these effectors is SopA, a HECT-type E3 ligase, which is required for the efficient stimulation of inflammation in an animal model of Salmonella Typhimurium infection. We show here that SopA contributes to the stimulation of innate immune responses by targeting two host E3 ubiquitin ligases, TRIM56 and TRIM65. We also found that TRIM65 interacts with the innate immune receptor MDA5 enhancing its ability to stimulate interferon-β signaling. Therefore, by targeting TRIM56 and TRIM65, SopA can stimulate signaling through two innate immune receptors, RIG-I and MDA5. These findings describe a Salmonella mechanism to modulate inflammatory responses by directly targeting innate immune signaling mechanisms.

  4. Antisymmetric tensor generalizations of affine vector fields.

    Science.gov (United States)

    Houri, Tsuyoshi; Morisawa, Yoshiyuki; Tomoda, Kentaro

    2016-02-01

    Tensor generalizations of affine vector fields called symmetric and antisymmetric affine tensor fields are discussed as symmetry of spacetimes. We review the properties of the symmetric ones, which have been studied in earlier works, and investigate the properties of the antisymmetric ones, which are the main theme in this paper. It is shown that antisymmetric affine tensor fields are closely related to one-lower-rank antisymmetric tensor fields which are parallelly transported along geodesics. It is also shown that the number of linear independent rank- p antisymmetric affine tensor fields in n -dimensions is bounded by ( n + 1)!/ p !( n - p )!. We also derive the integrability conditions for antisymmetric affine tensor fields. Using the integrability conditions, we discuss the existence of antisymmetric affine tensor fields on various spacetimes.

  5. Involvement of miR160/miR393 and their targets in cassava responses to anthracnose disease.

    Science.gov (United States)

    Pinweha, Nattaya; Asvarak, Thipa; Viboonjun, Unchera; Narangajavana, Jarunya

    2015-02-01

    Cassava is a starchy root crop for food and industrial applications in many countries around the world. Among the factors that affect cassava production, diseases remain the major cause of yield loss. Cassava anthracnose disease is caused by the fungus Colletotrichum gloeosporioides. Severe anthracnose attacks can cause tip die-backs and stem cankers, which can affect the availability of planting materials especially in large-scale production systems. Recent studies indicate that plants over- or under-express certain microRNAs (miRNAs) to cope with various stresses. Understanding how a disease-resistant plant protects itself from pathogens should help to uncover the role of miRNAs in the plant immune system. In this study, the disease severity assay revealed different response to C. gloeosporioides infection in two cassava cultivars. Quantitative RT-PCR analysis uncovered the differential expression of the two miRNAs and their target genes in the two cassava cultivars that were subjected to fungal infection. The more resistant cultivar revealed the up-regulation of miR160 and miR393, and consequently led to low transcript levels in their targets, ARF10 and TIR1, respectively. The more susceptible cultivar exhibited the opposite pattern. The cis-regulatory elements relevant to defense and stress responsiveness, fungal elicitor responsiveness and hormonal responses were the most prevalent present in the miRNAs gene promoter regions. The possible dual role of these specific miRNAs and their target genes associated with cassava responses to C. gloeosporioides is discussed. This is the first study to address the molecular events by which miRNAs which might play a role in fungal-infected cassava. A better understanding of the functions of miRNAs target genes should greatly increase our knowledge of the mechanism underlying susceptibility and lead to new strategies to enhance disease tolerance in this economically important crop. Copyright © 2014 Elsevier GmbH. All

  6. An affinity pull-down approach to identify the plant cyclic nucleotide interactome

    KAUST Repository

    Donaldson, Lara Elizabeth; Meier, Stuart Kurt

    2013-01-01

    Cyclic nucleotides (CNs) are intracellular second messengers that play an important role in mediating physiological responses to environmental and developmental signals, in species ranging from bacteria to humans. In response to these signals, CNs are synthesized by nucleotidyl cyclases and then act by binding to and altering the activity of downstream target proteins known as cyclic nucleotide-binding proteins (CNBPs). A number of CNBPs have been identified across kingdoms including transcription factors, protein kinases, phosphodiesterases, and channels, all of which harbor conserved CN-binding domains. In plants however, few CNBPs have been identified as homology searches fail to return plant sequences with significant matches to known CNBPs. Recently, affinity pull-down techniques have been successfully used to identify CNBPs in animals and have provided new insights into CN signaling. The application of these techniques to plants has not yet been extensively explored and offers an alternative approach toward the unbiased discovery of novel CNBP candidates in plants. Here, an affinity pull-down technique for the identification of the plant CN interactome is presented. In summary, the method involves an extraction of plant proteins which is incubated with a CN-bait, followed by a series of increasingly stringent elutions that eliminates proteins in a sequential manner according to their affinity to the bait. The eluted and bait-bound proteins are separated by one-dimensional gel electrophoresis, excised, and digested with trypsin after which the resultant peptides are identified by mass spectrometry - techniques that are commonplace in proteomics experiments. The discovery of plant CNBPs promises to provide valuable insight into the mechanism of CN signal transduction in plants. © Springer Science+Business Media New York 2013.

  7. An affinity pull-down approach to identify the plant cyclic nucleotide interactome

    KAUST Repository

    Donaldson, Lara Elizabeth

    2013-09-03

    Cyclic nucleotides (CNs) are intracellular second messengers that play an important role in mediating physiological responses to environmental and developmental signals, in species ranging from bacteria to humans. In response to these signals, CNs are synthesized by nucleotidyl cyclases and then act by binding to and altering the activity of downstream target proteins known as cyclic nucleotide-binding proteins (CNBPs). A number of CNBPs have been identified across kingdoms including transcription factors, protein kinases, phosphodiesterases, and channels, all of which harbor conserved CN-binding domains. In plants however, few CNBPs have been identified as homology searches fail to return plant sequences with significant matches to known CNBPs. Recently, affinity pull-down techniques have been successfully used to identify CNBPs in animals and have provided new insights into CN signaling. The application of these techniques to plants has not yet been extensively explored and offers an alternative approach toward the unbiased discovery of novel CNBP candidates in plants. Here, an affinity pull-down technique for the identification of the plant CN interactome is presented. In summary, the method involves an extraction of plant proteins which is incubated with a CN-bait, followed by a series of increasingly stringent elutions that eliminates proteins in a sequential manner according to their affinity to the bait. The eluted and bait-bound proteins are separated by one-dimensional gel electrophoresis, excised, and digested with trypsin after which the resultant peptides are identified by mass spectrometry - techniques that are commonplace in proteomics experiments. The discovery of plant CNBPs promises to provide valuable insight into the mechanism of CN signal transduction in plants. © Springer Science+Business Media New York 2013.

  8. Affine LIBOR Models with Multiple Curves

    DEFF Research Database (Denmark)

    Grbac, Zorana; Papapantoleon, Antonis; Schoenmakers, John

    2015-01-01

    are specified following the methodology of the affine LIBOR models and are driven by the wide and flexible class of affine processes. The affine property is preserved under forward measures, which allows us to derive Fourier pricing formulas for caps, swaptions, and basis swaptions. A model specification...... with dependent LIBOR rates is developed that allows for an efficient and accurate calibration to a system of caplet prices....

  9. Selection of imprinted nanoparticles by affinity chromatography.

    Science.gov (United States)

    Guerreiro, António R; Chianella, Iva; Piletska, Elena; Whitcombe, Michael J; Piletsky, Sergey A

    2009-04-15

    Soluble molecularly imprinted nanoparticles were synthesised via iniferter initiated polymerisation and separated by size via gel permeation chromatography. Subsequent fractionation of these particles by affinity chromatography allowed the separation of high affinity fractions from the mixture of nanoparticles. Fractions selected this way possess affinity similar to that of natural antibodies (K(d) 6.6x10(-8)) M and were also able to discriminate between related functional analogues of the template.

  10. Connections between quantized affine algebras and superalgebras

    International Nuclear Information System (INIS)

    Zhang, R.B.

    1992-08-01

    Every affine superalgebra with a symmetrizable Cartan matrix is closely related to an ordinary affine algebra with the same Cartan matrix. It is shown that the quantum supergroup associated with the former is essentially isomorphic to the quantum group associated with the latter in an appropriate class of representations. At the classical level, each integrable irreducible highest weight representation of the affine superalgebra has a corresponding irreducible representation of the affine algebra, which has the same weight space decomposition. (author). 5 refs, 3 tabs

  11. A Novel Vertex Affinity for Community Detection

    Energy Technology Data Exchange (ETDEWEB)

    Yoo, Andy [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Sanders, Geoffrey [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Henson, Van [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Vassilevski, Panayot [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2015-10-05

    We propose a novel vertex affinity measure in this paper. The new vertex affinity quantifies the proximity between two vertices in terms of their clustering strength and is ideal for such graph analytics applications as community detection. We also developed a framework that combines simple graph searches and resistance circuit formulas to compute the vertex affinity efficiently. We study the properties of the new affinity measure empirically in comparison to those of other popular vertex proximity metrics. Our results show that the existing metrics are ill-suited for community detection due to their lack of fundamental properties that are essential for correctly capturing inter- and intra-cluster vertex proximity.

  12. Safety targets and public risk perceptions in the nuclear field - technical treadmill or institutional responses?

    International Nuclear Information System (INIS)

    Wynne, B.

    1989-01-01

    The context of our treatment of risk perceptions and safety targets is the apparently wide gap between expert judgements of 'objective risks' and public perceptions of those risks. In the nuclear field the latter appear to so multiply the objective risks as seen by the experts, as to make safety targets vastly too strict (whether for routine discharges or for large accidents), thus design extravagantly expensive on any 'rational' criteria. In recent years the nuclear industry has come to terms more with the public perceptions problem, and has accepted that it is legitimate to exercise different, more severe and costly safety standards in the nuclear field if that is what society wants, as it appears to do. Whilst retaining the conviction that this is scientifically unwarranted, the industry has therefore reconciled itself somewhat to more stringent technical safety targets. (author)

  13. Molecular strategies targeting the host component of cancer to enhance tumor response to radiation therapy

    International Nuclear Information System (INIS)

    Kim, Dong Wook; Huamani, Jessica; Fu, Allie; Hallahan, Dennis E.

    2006-01-01

    The tumor microenvironment, in particular, the tumor vasculature, as an important target for the cytotoxic effects of radiation therapy is an established paradigm for cancer therapy. We review the evidence that the phosphoinositide 3-kinase (PI3K)/Akt pathway is activated in endothelial cells exposed to ionizing radiation (IR) and is a molecular target for the development of novel radiation sensitizing agents. On the basis of this premise, several promising preclinical studies that targeted the inhibition of the PI3K/Akt activation as a potential method of sensitizing the tumor vasculature to the cytotoxic effects of IR have been conducted. An innovative strategy to guide cytotoxic therapy in tumors treated with radiation and PI3K/Akt inhibitors is presented. The evidence supports a need for further investigation of combined-modality therapy that involves radiation therapy and inhibitors of PI3K/Akt pathway as a promising strategy for improving the treatment of patients with cancer

  14. TARGET COSTING – THE RESPONSE OF THE MANAGERIAL ACCOUNTING TO CHANGES IN THE ENVIRONMENT

    Directory of Open Access Journals (Sweden)

    OFILEANU DIMI

    2014-06-01

    Full Text Available The aim of this article is to highlight the Target Costing concept and the way in which the concept is used by the Romanian entities. The article analyzes the concept starting from the time of its appearance and up to the present moment, presenting the concept’s characteristics and its mode of action. A careful review of the specialized literature will be presented and the fact that this concept represents in fact a method of management. In order to study an eventual correlation between using Target Costing concept and the financial results obtained, an analysis of a sample of Romanian entities which produce shoes will be made. Also, it was analyzed the level of use and how much the Target Costing concept is known.

  15. Affinity of antibody secreted by a single cell

    International Nuclear Information System (INIS)

    Doran, D.M.

    1978-01-01

    It was the intention of this research to measure the affinity of antibody secreted by a single cell, and to describe the spectrum of affinities displayed in response to antigenic stimulation. The single cell secreting specific antibody was isolated by means of the hemolytic plaque assay. The amount of antibody secreted by the cell was to be measured through the use of a solid phase radioimmunoassay. The affinity of the antibody would be estimated by comparing the diameter of the plaque, and the amount of antibody secreted, with a mathematical theory of the formation of a plaque in agar. As a test system, a solid phase radioimmunoassay was developed for human serum albumin using antibody coupled to Sephadex. A sensitivity of 1 nanogram was attained with this assay. A solid phase radioimmunoassay for mouse immunoglobulin M was developed, using antibody coupled to Sepharose. The sensitivity attained with this assay was only on the order of 10 micrograms. The mouse immunoglobulin M radioimmunoassay was not sensitive enough to measure the amount of antibody secreted by a single cell. From a theoretical equation, the relationship between antibody affinity, plaque diameter and antibody secretion rate was calculated for the experimental conditions used in this research. By assuming a constant antibody secretion rate, an effective binding constant for the antibody was estimated from the average plaque diameters. This effective binding constant was observed to increase during the immune response

  16. Molecular targets of omega 3 and conjugated linoleic fatty acids – micromanaging cellular response

    Directory of Open Access Journals (Sweden)

    Francesco eVisioli

    2012-02-01

    Full Text Available Essential fatty acids cannot be synthesized de novo by mammals and need to be ingested either with the diet or through the use of supplements/functional foods to ameliorate cardiovascular prognosis. This review focus on the molecular targets of omega 3 fatty acids and CLA, as paradigmatic molecules that can be explored both as nutrients and as pharmacological agents, especially as related to cardioprotection. In addition, we indicate novel molecular targets, namely microRNAs that might contribute to the observed biological activities of such essential fatty acids.

  17. Real-time multi-target ranging based on chaotic polarization laser radars in the drive-response VCSELs.

    Science.gov (United States)

    Zhong, Dongzhou; Xu, Geliang; Luo, Wei; Xiao, Zhenzhen

    2017-09-04

    According to the principle of complete chaos synchronization and the theory of Hilbert phase transformation, we propose a novel real-time multi-target ranging scheme by using chaotic polarization laser radar in the drive-response vertical-cavity surface-emitting lasers (VCSELs). In the scheme, to ensure each polarization component (PC) of the master VCSEL (MVCSEL) to be synchronized steadily with that of the slave VCSEL, the output x-PC and y-PC from the MVCSEL in the drive system and those in the response system are modulated by the linear electro-optic effect simultaneously. Under this condition, by simulating the influences of some key parameters of the system on the synchronization quality and the relative errors of the two-target ranging, related operating parameters can be optimized. The x-PC and the y-PC, as two chaotic radar sources, are used to implement the real-time ranging for two targets. It is found that the measured distances of the two targets at arbitrary position exhibit strong real-time stability and only slight jitter. Their resolutions are up to millimeters, and their relative errors are very small and less than 2.7%.

  18. Proteome-wide analysis of SUMO2 targets in response to pathological DNA replication stress in human cells.

    Science.gov (United States)

    Bursomanno, Sara; Beli, Petra; Khan, Asif M; Minocherhomji, Sheroy; Wagner, Sebastian A; Bekker-Jensen, Simon; Mailand, Niels; Choudhary, Chunaram; Hickson, Ian D; Liu, Ying

    2015-01-01

    SUMOylation is a form of post-translational modification involving covalent attachment of SUMO (Small Ubiquitin-like Modifier) polypeptides to specific lysine residues in the target protein. In human cells, there are four SUMO proteins, SUMO1-4, with SUMO2 and SUMO3 forming a closely related subfamily. SUMO2/3, in contrast to SUMO1, are predominantly involved in the cellular response to certain stresses, including heat shock. Substantial evidence from studies in yeast has shown that SUMOylation plays an important role in the regulation of DNA replication and repair. Here, we report a proteomic analysis of proteins modified by SUMO2 in response to DNA replication stress in S phase in human cells. We have identified a panel of 22 SUMO2 targets with increased SUMOylation during DNA replication stress, many of which play key functions within the DNA replication machinery and/or in the cellular response to DNA damage. Interestingly, POLD3 was found modified most significantly in response to a low dose aphidicolin treatment protocol that promotes common fragile site (CFS) breakage. POLD3 is the human ortholog of POL32 in budding yeast, and has been shown to act during break-induced recombinational repair. We have also shown that deficiency of POLD3 leads to an increase in RPA-bound ssDNA when cells are under replication stress, suggesting that POLD3 plays a role in the cellular response to DNA replication stress. Considering that DNA replication stress is a source of genome instability, and that excessive replication stress is a hallmark of pre-neoplastic and tumor cells, our characterization of SUMO2 targets during a perturbed S-phase should provide a valuable resource for future functional studies in the fields of DNA metabolism and cancer biology. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. On affine non-negative matrix factorization

    DEFF Research Database (Denmark)

    Laurberg, Hans; Hansen, Lars Kai

    2007-01-01

    We generalize the non-negative matrix factorization (NMF) generative model to incorporate an explicit offset. Multiplicative estimation algorithms are provided for the resulting sparse affine NMF model. We show that the affine model has improved uniqueness properties and leads to more accurate id...

  20. Global affine differential geometry of hypersurfaces

    CERN Document Server

    Li, An-Min; Zhao, Guosong; Hu, Zejun

    2015-01-01

    This book draws a colorful and widespread picture of global affine hypersurface theory up to the most recent state. Moreover, the recent development revealed that affine differential geometry- as differential geometry in general- has an exciting intersection area with other fields of interest, like partial differential equations, global analysis, convex geometry and Riemann surfaces.

  1. Training motor responses to food: A novel treatment for obesity targeting implicit processes

    NARCIS (Netherlands)

    Stice, E.; Lawrence, N.S.; Kemps, E.; Veling, H.P.

    2016-01-01

    The present review first summarizes results from prospective brain imaging studies focused on identifying neural vulnerability factors that predict excessive weight gain. Next, findings from cognitive psychology experiments evaluating various interventions involving food response inhibition training

  2. Identification of hookworm DAF-16/FOXO response elements and direct gene targets.

    Directory of Open Access Journals (Sweden)

    Xin Gao

    2010-08-01

    Full Text Available The infective stage of the parasitic nematode hookworm is developmentally arrested in the environment and needs to infect a specific host to complete its life cycle. The canine hookworm (Ancylostoma caninum is an excellent model for investigating human hookworm infections. The transcription factor of A. caninum, Ac-DAF-16, which has a characteristic fork head or "winged helix" DNA binding domain (DBD, has been implicated in the resumption of hookworm development in the host. However, the precise roles of Ac-DAF-16 in hookworm parasitism and its downstream targets are unknown. In the present study, we combined molecular techniques and bioinformatics to identify a group of Ac-DAF-16 binding sites and target genes.The DNA binding domain of Ac-DAF-16 was used to select genomic fragments by in vitro genomic selection. Twenty four bound genomic fragments were analyzed for the presence of the DAF-16 family binding element (DBE and possible alternative Ac-DAF-16 bind motifs. The 22 genes linked to these genomic fragments were identified using bioinformatics tools and defined as candidate direct gene targets of Ac-DAF-16. Their developmental stage-specific expression patterns were examined. Also, a new putative DAF-16 binding element was identified.Our results show that Ac-DAF-16 is involved in diverse biological processes throughout hookworm development. Further investigation of these target genes will provide insights into the molecular basis by which Ac-DAF-16 regulates its downstream gene network in hookworm infection.

  3. Molecular Mechanisms of Foot-and-Mouth Disease Virus Targeting the Host Antiviral Response.

    Science.gov (United States)

    Rodríguez Pulido, Miguel; Sáiz, Margarita

    2017-01-01

    Foot-and-mouth disease virus (FMDV) is the causative agent of an acute vesicular disease affecting pigs, cattle and other domestic, and wild animals worldwide. The aim of the host interferon (IFN) response is to limit viral replication and spread. Detection of the viral genome and products by specialized cellular sensors initiates a signaling cascade that leads to a rapid antiviral response involving the secretion of type I- and type III-IFNs and other antiviral cytokines with antiproliferative and immunomodulatory functions. During co-evolution with their hosts, viruses have acquired strategies to actively counteract host antiviral responses and the balance between innate response and viral antagonism may determine the outcome of disease and pathogenesis. FMDV proteases Lpro and 3C have been found to antagonize the host IFN response by a repertoire of mechanisms. Moreover, the putative role of other viral proteins in IFN antagonism is being recently unveiled, uncovering sophisticated immune evasion strategies different to those reported to date for other members of the Picornaviridae family. Here, we review the interplay between antiviral responses induced by FMDV infection and viral countermeasures to block them. Research on strategies used by viruses to modulate immunity will provide insights into the function of host pathways involved in defense against pathogens and will also lead to development of new therapeutic strategies to fight virus infections.

  4. Integration of the Pokeweed miRNA and mRNA Transcriptomes Reveals Targeting of Jasmonic Acid-Responsive Genes

    Directory of Open Access Journals (Sweden)

    Kira C. M. Neller

    2018-05-01

    Full Text Available The American pokeweed plant, Phytolacca americana, displays broad-spectrum resistance to plant viruses and is a heavy metal hyperaccumulator. However, little is known about the regulation of biotic and abiotic stress responses in this non-model plant. To investigate the control of miRNAs in gene expression, we sequenced the small RNA transcriptome of pokeweed treated with jasmonic acid (JA, a hormone that mediates pathogen defense and stress tolerance. We predicted 145 miRNAs responsive to JA, most of which were unique to pokeweed. These miRNAs were low in abundance and condition-specific, with discrete expression change. Integration of paired mRNA-Seq expression data enabled us to identify correlated, novel JA-responsive targets that mediate hormone biosynthesis, signal transduction, and pathogen defense. The expression of approximately half the pairs was positively correlated, an uncommon finding that we functionally validated by mRNA cleavage. Importantly, we report that a pokeweed-specific miRNA targets the transcript of OPR3, novel evidence that a miRNA regulates a JA biosynthesis enzyme. This first large-scale small RNA study of a Phytolaccaceae family member shows that miRNA-mediated control is a significant component of the JA response, associated with widespread changes in expression of genes required for stress adaptation.

  5. Nanoparticle-based B-cell targeting vaccines: Tailoring of humoral immune responses by functionalization with different TLR-ligands.

    Science.gov (United States)

    Zilker, Claudia; Kozlova, Diana; Sokolova, Viktoriya; Yan, Huimin; Epple, Matthias; Überla, Klaus; Temchura, Vladimir

    2017-01-01

    Induction of an appropriate type of humoral immune response during vaccination is essential for protection against viral and bacterial infections. We recently observed that biodegradable calcium phosphate (CaP) nanoparticles coated with proteins efficiently targeted and activated naïve antigen-specific B-cells in vitro. We now compared different administration routes for CaP-nanoparticles and demonstrated that intramuscular immunization with such CaP-nanoparticles induced stronger immune responses than immunization with monovalent antigen. Additional functionalization of the CaP-nanoparticles with TRL-ligands allowed modulating the IgG subtype response and the level of mucosal IgA antibodies. CpG-containing CaP-nanoparticles were as immunogenic as a virus-like particle vaccine. Functionalization of CaP-nanoparticles with T-helper cell epitopes or CpG also allowed overcoming lack of T-cell help. Thus, our results indicate that CaP-nanoparticle-based B-cell targeting vaccines functionalized with TLR-ligands can serve as a versatile platform for efficient induction and modulation of humoral immune responses in vivo. Copyright © 2016 Elsevier Inc. All rights reserved.

  6. Improving image segmentation by learning region affinities

    Energy Technology Data Exchange (ETDEWEB)

    Prasad, Lakshman [Los Alamos National Laboratory; Yang, Xingwei [TEMPLE UNIV.; Latecki, Longin J [TEMPLE UNIV.

    2010-11-03

    We utilize the context information of other regions in hierarchical image segmentation to learn new regions affinities. It is well known that a single choice of quantization of an image space is highly unlikely to be a common optimal quantization level for all categories. Each level of quantization has its own benefits. Therefore, we utilize the hierarchical information among different quantizations as well as spatial proximity of their regions. The proposed affinity learning takes into account higher order relations among image regions, both local and long range relations, making it robust to instabilities and errors of the original, pairwise region affinities. Once the learnt affinities are obtained, we use a standard image segmentation algorithm to get the final segmentation. Moreover, the learnt affinities can be naturally unutilized in interactive segmentation. Experimental results on Berkeley Segmentation Dataset and MSRC Object Recognition Dataset are comparable and in some aspects better than the state-of-art methods.

  7. Receptor-targeted metalloradiopharmaceuticals. Final technical report

    International Nuclear Information System (INIS)

    Green, Mark A.

    2000-01-01

    Copper (II) and platinum (II) coordination complexes were prepared and characterized. These complexes were designed to afford structural homology with steroidal and non-steroidal estrogens for possible use as receptor-targeted radiopharmaceuticals. While weak affinity for the estrogen receptor was detectable, none would appear to have sufficient receptor-affinity for estrogen-receptor-targeted imaging or therapy

  8. Dual responsive promoters to target therapeutic gene expression to radiation-resistant hypoxic tumor cells

    International Nuclear Information System (INIS)

    Chadderton, Naomi; Cowen, Rachel L.; Sheppard, Freda C.D.; Robinson, Suzanne; Greco, Olga; Scott, Simon D.; Stratford, Ian J.; Patterson, Adam V.; Williams, Kaye J.

    2005-01-01

    Purpose: Tumor hypoxia is unequivocally linked to poor radiotherapy outcome. This study aimed to identify enhancer sequences that respond maximally to a combination of radiation and hypoxia for use in genetic radiotherapy approaches. Methods and materials: The influence of radiation (5 Gy) and hypoxia (1% O 2 ) on reporter-gene expression driven by hypoxia (HRE) and radiation (Egr-1) responsive elements was evaluated in tumor cells grown as monolayers or multicellular spheroids. Hypoxia-inducible factor-1α (HIF-1α) and HIF-2α protein expression was monitored in parallel. Results: Of the sequences tested, an HRE from the phosphoglycerate kinase-1 gene (PGK-18[5+]) was maximally induced in response to hypoxia plus radiation in all 5 cell lines tested. The additional radiation treatment afforded a significant increase in the induction of PGK-18[5+] compared with hypoxia alone in 3 cell lines. HIF-1α/2α were induced by radiation but combined hypoxia/radiation treatment did not yield a further increase. The dual responsive nature of HREs was maintained when spheroids were irradiated after delivery of HRE constructs in a replication-deficient adenovirus. Conclusions: Hypoxia-responsive enhancer element sequences are dually responsive to combined radiation and hypoxic treatment. Their use in genetic radiotherapy in vivo could maximize expression in the most radio-resistant population at the time of radiation and also exploit microenvironmental changes after radiotherapy to yield additional switch-on

  9. Affinity purification mass spectrometry analysis of PD-1 uncovers SAP as a new checkpoint inhibitor.

    Science.gov (United States)

    Peled, Michael; Tocheva, Anna S; Sandigursky, Sabina; Nayak, Shruti; Philips, Elliot A; Nichols, Kim E; Strazza, Marianne; Azoulay-Alfaguter, Inbar; Askenazi, Manor; Neel, Benjamin G; Pelzek, Adam J; Ueberheide, Beatrix; Mor, Adam

    2018-01-16

    Programmed cell death-1 (PD-1) is an essential inhibitory receptor in T cells. Antibodies targeting PD-1 elicit durable clinical responses in patients with multiple tumor indications. Nevertheless, a significant proportion of patients do not respond to anti-PD-1 treatment, and a better understanding of the signaling pathways downstream of PD-1 could provide biomarkers for those whose tumors respond and new therapeutic approaches for those whose tumors do not. We used affinity purification mass spectrometry to uncover multiple proteins associated with PD-1. Among these proteins, signaling lymphocytic activation molecule-associated protein (SAP) was functionally and mechanistically analyzed for its contribution to PD-1 inhibitory responses. Silencing of SAP augmented and overexpression blocked PD-1 function. T cells from patients with X-linked lymphoproliferative disease (XLP), who lack functional SAP, were hyperresponsive to PD-1 signaling, confirming its inhibitory role downstream of PD-1. Strikingly, signaling downstream of PD-1 in purified T cell subsets did not correlate with PD-1 surface expression but was inversely correlated with intracellular SAP levels. Mechanistically, SAP opposed PD-1 function by acting as a molecular shield of key tyrosine residues that are targets for the tyrosine phosphatase SHP2, which mediates PD-1 inhibitory properties. Our results identify SAP as an inhibitor of PD-1 function and SHP2 as a potential therapeutic target in patients with XLP.

  10. Measuring Response to Therapy by Near-Infrared Imaging of Tumors Using a Phosphatidylserine-Targeting Antibody Fragment

    Directory of Open Access Journals (Sweden)

    Jian Gong

    2013-06-01

    Full Text Available Imaging tumors and their response to treatment could be a valuable biomarker toward early assessment of therapy in patients with cancer. Phosphatidylserine (PS is confined to the inner leaflet of the plasma membrane in normal cells but is externalized on tumor vascular endothelial cells (ECs and tumor cells, and PS exposure is further enhanced in response to radiation and chemotherapy. In the present study, we evaluated the potential of a PS-targeting human F(ab'2 antibody fragment, PGN650, to detect exposure of PS in tumor-bearing mice. Tumor uptake of PGN650 was measured by near-infrared optical imaging in human tumor xenografts in immunodeficient mice. PGN650 specifically targeted tumors and was shown to target CD31-positive ECs and tumor cells. Tumor uptake of PGN650 was significantly higher in animals pretreated with docetaxel. The peak tumor to normal tissue (T/N ratio of probe was observed at 24 hours postinjection of probe, and tumor binding was detected for at least 120 hours. In repeat dose studies, PGN650 uptake in tumors was significantly higher following pretreatment with docetaxel compared to baseline uptake prior to treatment. PGN650 may be a useful probe to detect PS exposed in tumors and to monitor enhanced PS exposure to optimize therapeutic agents to treat tumors.

  11. Adrenergic Signaling: A Targetable Checkpoint Limiting Development of the Antitumor Immune Response

    Science.gov (United States)

    Qiao, Guanxi; Chen, Minhui; Bucsek, Mark J.; Repasky, Elizabeth A.; Hylander, Bonnie L.

    2018-01-01

    An immune response must be tightly controlled so that it will be commensurate with the level of response needed to protect the organism without damaging normal tissue. The roles of cytokines and chemokines in orchestrating these processes are well known, but although stress has long been thought to also affect immune responses, the underlying mechanisms were not as well understood. Recently, the role of nerves and, specifically, the sympathetic nervous system, in regulating immune responses is being revealed. Generally, an acute stress response is beneficial but chronic stress is detrimental because it suppresses the activities of effector immune cells while increasing the activities of immunosuppressive cells. In this review, we first discuss the underlying biology of adrenergic signaling in cells of both the innate and adaptive immune system. We then focus on the effects of chronic adrenergic stress in promoting tumor growth, giving examples of effects on tumor cells and immune cells, explaining the methods commonly used to induce stress in preclinical mouse models. We highlight how this relates to our observations that mandated housing conditions impose baseline chronic stress on mouse models, which is sufficient to cause chronic immunosuppression. This problem is not commonly recognized, but it has been shown to impact conclusions of several studies of mouse physiology and mouse models of disease. Moreover, the fact that preclinical mouse models are chronically immunosuppressed has critical ramifications for analysis of any experiments with an immune component. Our group has found that reducing adrenergic stress by housing mice at thermoneutrality or treating mice housed at cooler temperatures with β-blockers reverses immunosuppression and significantly improves responses to checkpoint inhibitor immunotherapy. These observations are clinically relevant because there are numerous retrospective epidemiological studies concluding that cancer patients who were

  12. Non-MHC Antigenic Targets of the Humoral Immune Response in Transplantation

    OpenAIRE

    Zhang, Qiuheng; Reed, Elaine F.

    2010-01-01

    There is a growing body of data supporting a role for non-HLA antibodies in acute and chronic rejection of solid organ transplants. While many of these non-HLA antigens remain poorly defined, the principle antigenic targets are expressed on cells of the allograft including endothelium and epithelium. These non-HLA antigens are classified as either alloantigens, such as the major histocompatibility complex class I chain-related gene A (MICA) or MICB, or tissue-specific autoantigens such as vim...

  13. An Affine Combination of Adaptive Filters for Channels with Different Sparsity Levels

    Directory of Open Access Journals (Sweden)

    M. Butsenko

    2016-06-01

    Full Text Available In this paper we present an affine combination strategy for two adaptive filters. One filter is designed to handle sparse impulse responses and the other one performs better if impulse response is dispersive. Filter outputs are combined using an adaptive mixing parameter and the resulting output shows a better performance than each of the combining filters separately. We also demonstrate that affine combination results in faster convergence than a convex combination of two adaptive filters.

  14. Synthetic Peptides to Target Stringent Response-Controlled Virulence in a Pseudomonas aeruginosa Murine Cutaneous Infection Model

    Directory of Open Access Journals (Sweden)

    Daniel Pletzer

    2017-09-01

    Full Text Available Microorganisms continuously monitor their surroundings and adaptively respond to environmental cues. One way to cope with various stress-related situations is through the activation of the stringent stress response pathway. In Pseudomonas aeruginosa this pathway is controlled and coordinated by the activity of the RelA and SpoT enzymes that metabolize the small nucleotide secondary messenger molecule (pppGpp. Intracellular ppGpp concentrations are crucial in mediating adaptive responses and virulence. Targeting this cellular stress response has recently been the focus of an alternative approach to fight antibiotic resistant bacteria. Here, we examined the role of the stringent response in the virulence of P. aeruginosa PAO1 and the Liverpool epidemic strain LESB58. A ΔrelA/ΔspoT double mutant showed decreased cytotoxicity toward human epithelial cells, exhibited reduced hemolytic activity, and caused down-regulation of the expression of the alkaline protease aprA gene in stringent response mutants grown on blood agar plates. Promoter fusions of relA or spoT to a bioluminescence reporter gene revealed that both genes were expressed during the formation of cutaneous abscesses in mice. Intriguingly, virulence was attenuated in vivo by the ΔrelA/ΔspoT double mutant, but not the relA mutant nor the ΔrelA/ΔspoT complemented with either gene. Treatment of a cutaneous P. aeruginosa PAO1 infection with anti-biofilm peptides increased animal welfare, decreased dermonecrotic lesion sizes, and reduced bacterial numbers recovered from abscesses, resembling the phenotype of the ΔrelA/ΔspoT infection. It was previously demonstrated by our lab that ppGpp could be targeted by synthetic peptides; here we demonstrated that spoT promoter activity was suppressed during cutaneous abscess formation by treatment with peptides DJK-5 and 1018, and that a peptide-treated relA complemented stringent response double mutant strain exhibited reduced peptide

  15. High affinity soluble ILT2 receptor: a potent inhibitor of CD8(+) T cell activation.

    Science.gov (United States)

    Moysey, Ruth K; Li, Yi; Paston, Samantha J; Baston, Emma E; Sami, Malkit S; Cameron, Brian J; Gavarret, Jessie; Todorov, Penio; Vuidepot, Annelise; Dunn, Steven M; Pumphrey, Nicholas J; Adams, Katherine J; Yuan, Fang; Dennis, Rebecca E; Sutton, Deborah H; Johnson, Andy D; Brewer, Joanna E; Ashfield, Rebecca; Lissin, Nikolai M; Jakobsen, Bent K

    2010-12-01

    Using directed mutagenesis and phage display on a soluble fragment of the human immunoglobulin super-family receptor ILT2 (synonyms: LIR1, MIR7, CD85j), we have selected a range of mutants with binding affinities enhanced by up to 168,000-fold towards the conserved region of major histocompatibility complex (MHC) class I molecules. Produced in a dimeric form, either by chemical cross-linking with bivalent polyethylene glycol (PEG) derivatives or as a genetic fusion with human IgG Fc-fragment, the mutants exhibited a further increase in ligand-binding strength due to the avidity effect, with resident half-times (t(1/2)) on the surface of MHC I-positive cells of many hours. The novel compounds antagonized the interaction of CD8 co-receptor with MHC I in vitro without affecting the peptide-specific binding of T-cell receptors (TCRs). In both cytokine-release assays and cell-killing experiments the engineered receptors inhibited the activation of CD8(+) cytotoxic T lymphocytes (CTLs) in the presence of their target cells, with subnanomolar potency and in a dose-dependent manner. As a selective inhibitor of CD8(+) CTL responses, the engineered high affinity ILT2 receptor presents a new tool for studying the activation mechanism of different subsets of CTLs and could have potential for the development of novel autoimmunity therapies.

  16. Identification of novel human damage response proteins targeted through yeast orthology.

    Directory of Open Access Journals (Sweden)

    J Peter Svensson

    Full Text Available Studies in Saccharomyces cerevisiae show that many proteins influence cellular survival upon exposure to DNA damaging agents. We hypothesized that human orthologs of these S. cerevisiae proteins would also be required for cellular survival after treatment with DNA damaging agents. For this purpose, human homologs of S. cerevisiae proteins were identified and mapped onto the human protein-protein interaction network. The resulting human network was highly modular and a series of selection rules were implemented to identify 45 candidates for human toxicity-modulating proteins. The corresponding transcripts were targeted by RNA interference in human cells. The cell lines with depleted target expression were challenged with three DNA damaging agents: the alkylating agents MMS and 4-NQO, and the oxidizing agent t-BuOOH. A comparison of the survival revealed that the majority (74% of proteins conferred either sensitivity or resistance. The identified human toxicity-modulating proteins represent a variety of biological functions: autophagy, chromatin modifications, RNA and protein metabolism, and telomere maintenance. Further studies revealed that MMS-induced autophagy increase the survival of cells treated with DNA damaging agents. In summary, we show that damage recovery proteins in humans can be identified through homology to S. cerevisiae and that many of the same pathways are represented among the toxicity modulators.

  17. Targeting EGFR with photodynamic therapy in combination with Erbitux enhances in vivo bladder tumor response

    Directory of Open Access Journals (Sweden)

    Soo Khee

    2009-11-01

    Full Text Available Abstract Background Photodynamic therapy (PDT is a promising cancer treatment modality that involves the interaction of the photosensitizer, molecular oxygen and light of specific wavelength to destroy tumor cells. Treatment induced hypoxia is one of the main side effects of PDT and efforts are underway to optimize PDT protocols for improved efficacy. The aim of this study was to investigate the anti-tumor effects of PDT plus Erbitux, an angiogenesis inhibitor that targets epidermal growth factor receptor (EGFR, on human bladder cancer model. Tumor-bearing nude mice were assigned to four groups that included control, PDT, Erbitux and PDT plus Erbitux and tumor volume was charted over 90-day period. Results Our results demonstrate that combination of Erbitux with PDT strongly inhibits tumor growth in the bladder tumor xenograft model when compared to the other groups. Downregulation of EGFR was detected using immunohistochemistry, immunofluorescence and western blotting. Increased apoptosis was associated with tumor inhibition in the combination therapy group. In addition, we identified the dephosphorylation of ErbB4 at tyrosine 1284 site to play a major role in tumor inhibition. Also, at the RNA level downregulation of EGFR target genes cyclin D1 and c-myc was observed in tumors treated with PDT plus Erbitux. Conclusion The combination therapy of PDT and Erbitux effectively inhibits tumor growth and is a promising therapeutic approach in the treatment of bladder tumors.

  18. Near-Infrared Light Responsive Folate Targeted Gold Nanorods for Combined Photothermal-Chemotherapy of Osteosarcoma.

    Science.gov (United States)

    Li Volsi, Anna; Scialabba, Cinzia; Vetri, Valeria; Cavallaro, Gennara; Licciardi, Mariano; Giammona, Gaetano

    2017-04-26

    Folate-targeted gold nanorods (GNRs) are proposed as selective theranostic agents for osteosarcoma treatment. An amphiphilic polysaccharide based graft-copolymer (INU-LA-PEG-FA) and an amino derivative of the α,β-poly(N-2-hydroxyethyl)-d,l-aspartamide functionalized with folic acid (PHEA-EDA-FA), have been synthesized to act as coating agents for GNRs. The obtained polymer-coated GNRs were characterized in terms of size, shape, zeta potential, chemical composition, and aqueous stability. They protected the anticancer drug nutlin-3 and were able to deliver it efficiently in different physiological media. The ability of the proposed systems to selectively kill tumor cells was tested on U2OS cancer cells expressing high levels of FRs and compared with human bronchial epithelial cells (16HBE) and human dermal fibroblasts (HDFa). The property of the nanosystems of efficiently controlling drug release upon NIR laser irradiation and of acting as an excellent hyperthermia agent as well as Two Photon Luminescence imaging contrast agents was demonstrated. The proposed folate-targeted GNRs have also been tested in terms of chemoterapeutic and thermoablation efficacy on tridimensional (3-D) osteosarcoma models.

  19. Biomechanical responses of PMHS in moderate-speed rear impacts and development of response targets for evaluating the internal and external biofidelity of ATDS.

    Science.gov (United States)

    Kang, Yun-Seok; Bolte, John H; Moorhouse, Kevin; Donnelly, Bruce; Herriott, Rodney; Mallory, Ann

    2012-10-01

    The objectives of this study were to obtain biomechanical responses of post mortem human subjects (PMHS) by subjecting them to two moderate-speed rear impact sled test conditions (8.5g, 17 km/h; 10.5g, 24 km/h) while positioned in an experimental seat system, and to create biomechanical targets for internal and external biofidelity evaluation of rear impact ATDs. The experimental seat was designed to measure external loads on the head restraint (4 load cells), seat back (6 load cells), and seat pan (4 load cells) such that subject dynamic interaction with the seat could be evaluated. This seat system was capable of simulating the dynamic characteristics of modern vehicle seat backs by considering the moment-rotation properties of a typical passenger vehicle, thus providing a more realistic test environment than using a rigid seat with a non-rotating seat back as done in previous studies. Instrumentation used to measure biomechanical responses of the PMHS included both accelerometers and angular rate sensors (ARS). A total of fourteen sled tests using eight PMHS (males 175.8 ± 6.2 cm of stature and 78.4 ± 7.2 kg of weight) provided data sets of seven PMHS for both test conditions. The biomechanical responses are described at both speeds, and cervical spine injuries are documented. Biomechanical targets are also created for internal and external biofidelity evaluation of rear impact anthropomorphic test devices (ATDs).

  20. Transcriptional profiling uncovers a network of cholesterol-responsive atherosclerosis target genes.

    Directory of Open Access Journals (Sweden)

    Josefin Skogsberg

    2008-03-01

    Full Text Available Despite the well-documented effects of plasma lipid lowering regimes halting atherosclerosis lesion development and reducing morbidity and mortality of coronary artery disease and stroke, the transcriptional response in the atherosclerotic lesion mediating these beneficial effects has not yet been carefully investigated. We performed transcriptional profiling at 10-week intervals in atherosclerosis-prone mice with human-like hypercholesterolemia and a genetic switch to lower plasma lipoproteins (Ldlr(-/-Apo(100/100Mttp(flox/flox Mx1-Cre. Atherosclerotic lesions progressed slowly at first, then expanded rapidly, and plateaued after advanced lesions formed. Analysis of lesion expression profiles indicated that accumulation of lipid-poor macrophages reached a point that led to the rapid expansion phase with accelerated foam-cell formation and inflammation, an interpretation supported by lesion histology. Genetic lowering of plasma cholesterol (e.g., lipoproteins at this point all together prevented the formation of advanced plaques and parallel transcriptional profiling of the atherosclerotic arterial wall identified 37 cholesterol-responsive genes mediating this effect. Validation by siRNA-inhibition in macrophages incubated with acetylated-LDL revealed a network of eight cholesterol-responsive atherosclerosis genes regulating cholesterol-ester accumulation. Taken together, we have identified a network of atherosclerosis genes that in response to plasma cholesterol-lowering prevents the formation of advanced plaques. This network should be of interest for the development of novel atherosclerosis therapies.

  1. Specific nongluten proteins of wheat are novel target antigens in celiac disease humoral response

    Science.gov (United States)

    Background: Celiac disease is an immune-mediated enteropathy that is generally understood to be triggered by the ingestion of gluten proteins of wheat and related cereals. The skin manifestation of the condition is known as dermatitis herpetiformis. Antibody response to native and deamidated seque...

  2. Early Detection Rapid Response Program Targets New Noxious Weed Species in Washington State

    Science.gov (United States)

    Andreas, Jennifer E.; Halpern, Alison D.; DesCamp, Wendy C.; Miller, Timothy W.

    2015-01-01

    Early detection, rapid response is a critical component of invasive plant management. It can be challenging, however, to detect new invaders before they become established if landowners cannot identify species of concern. In order to increase awareness, eye-catching postcards were developed in Washington State as part of a noxious weed educational…

  3. Identification of Predictive Response Markers and Novel Treatment Targets for Gliomas

    NARCIS (Netherlands)

    L. Erdem-Eraslan (Lale)

    2016-01-01

    markdownabstractGliomas are the most frequent primary brain tumors in adults. Despite multimodality treatment strategies, the survival of patients with a diffuse glioma remains poor. There has been an increasing use of molecular markers to assist diagnosis and predict prognosis and response to

  4. The intestinal T cell response to alpha-gliadin in adult celiac disease is focused on a single deamidated glutamine targeted by tissue transglutaminase

    DEFF Research Database (Denmark)

    Arentz-Hansen, H; Körner, R; Molberg, O

    2000-01-01

    The great majority of patients that are intolerant of wheat gluten protein due to celiac disease (CD) are human histocompatibility leukocyte antigen (HLA)-DQ2(+), and the remaining few normally express HLA-DQ8. These two class II molecules are chiefly responsible for the presentation of gluten...... for T cell recognition. Gluten-specific T cell lines from 16 different adult patients all responded to one or both of these deamidated peptides, indicating that these epitopes are highly relevant to disease pathology. Binding studies showed that the deamidated peptides displayed an increased affinity...

  5. Detection of Waterborne Viruses Using High Affinity Molecularly Imprinted Polymers.

    Science.gov (United States)

    Altintas, Zeynep; Gittens, Micah; Guerreiro, Antonio; Thompson, Katy-Anne; Walker, Jimmy; Piletsky, Sergey; Tothill, Ibtisam E

    2015-07-07

    Molecularly imprinted polymers (MIPs) are artificial receptor ligands which can recognize and specifically bind to a target molecule. They are more resistant to chemical and biological damage and inactivation than antibodies. Therefore, target specific-MIP nanoparticles are aimed to develop and implemented to biosensors for the detection of biological toxic agents such as viruses, bacteria, and fungi toxins that cause many diseases and death due to the environmental contamination. For the first time, a molecularly imprinted polymer (MIP) targeting the bacteriophage MS2 as the template was investigated using a novel solid-phase synthesis method to obtain the artificial affinity ligand for the detection and removal of waterborne viruses through optical-based sensors. A high affinity between the artificial ligand and the target was found, and a regenerative MIP-based virus detection assay was successfully developed using a new surface plasmon resonance (SPR)-biosensor which provides an alternative technology for the specific detection and removal of waterborne viruses that lead to high disease and death rates all over the world.

  6. Targeting Tumor-Associated Macrophages as a Potential Strategy to Enhance the Response to Immune Checkpoint Inhibitors.

    Science.gov (United States)

    Cassetta, Luca; Kitamura, Takanori

    2018-01-01

    Inhibition of immune checkpoint pathways in CD8 + T cell is a promising therapeutic strategy for the treatment of solid tumors that has shown significant anti-tumor effects and is now approved by the FDA to treat patients with melanoma and lung cancer. However the response to this therapy is limited to a certain fraction of patients and tumor types, for reasons still unknown. To ensure success of this treatment, CD8 + T cells, the main target of the checkpoint inhibitors, should exert full cytotoxicity against tumor cells. However recent studies show that tumor-associated macrophages (TAM) can impede this process by different mechanisms. In this mini-review we will summarize recent studies showing the effect of TAM targeting on immune checkpoint inhibitors efficacy. We will also discuss on the limitations of the current strategies as well on the future scientific challenges for the progress of the tumor immunology field.

  7. Ingroup categorization and response conflict: Interactive effects of target race, flanker compatibility, and infrequency on N2 amplitude.

    Science.gov (United States)

    Dickter, Cheryl L; Bartholow, Bruce D

    2010-05-01

    Three largely independent lines of research have investigated experimental manipulations that influence the amplitude of the N2 component of the event-related brain potential (ERP), one linking heightened N2 amplitude to response conflict, another showing that N2 is sensitive to stimulus infrequency, and the third showing larger N2 amplitude during categorization of racial ingroup relative to racial outgroup targets. The purpose of this research was to investigate potential interactions between these three features on the amplitude of the N2. ERPs were recorded while participants completed a modified flanker task using pictures of ingroup and outgroup faces. Results showed a 3-way interaction, indicating that the N2 was largest for ingroup targets on high-conflict trials but only when such trials were relatively infrequent. Implications of these findings for theories of both conflict monitoring and person perception are discussed.

  8. The Cutting Edge of Affinity Electrophoresis Technology

    Science.gov (United States)

    Kinoshita, Eiji; Kinoshita-Kikuta, Emiko; Koike, Tohru

    2015-01-01

    Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years. PMID:28248262

  9. The Cutting Edge of Affinity Electrophoresis Technology.

    Science.gov (United States)

    Kinoshita, Eiji; Kinoshita-Kikuta, Emiko; Koike, Tohru

    2015-03-18

    Affinity electrophoresis is an important technique that is widely used to separate and analyze biomolecules in the fields of biology and medicine. Both quantitative and qualitative information can be gained through affinity electrophoresis. Affinity electrophoresis can be applied through a variety of strategies, such as mobility shift electrophoresis, charge shift electrophoresis or capillary affinity electrophoresis. These strategies are based on changes in the electrophoretic patterns of biological macromolecules that result from interactions or complex-formation processes that induce changes in the size or total charge of the molecules. Nucleic acid fragments can be characterized through their affinity to other molecules, for example transcriptional factor proteins. Hydrophobic membrane proteins can be identified by means of a shift in the mobility induced by a charged detergent. The various strategies have also been used in the estimation of association/disassociation constants. Some of these strategies have similarities to affinity chromatography, in that they use a probe or ligand immobilized on a supported matrix for electrophoresis. Such methods have recently contributed to profiling of major posttranslational modifications of proteins, such as glycosylation or phosphorylation. Here, we describe advances in analytical techniques involving affinity electrophoresis that have appeared during the last five years.

  10. Chamber and Wall Response to Target Implosion in Inertial and Z-Pinch Fusion and Lithography Devices

    International Nuclear Information System (INIS)

    Hassanein, A.; Konkashbaev, I.; Morozov, V.; Sizyuk, V.

    2006-01-01

    The chamber walls, both solid and liquid, in inertial fusion energy (IFE) and Z-pinch reactors and Lithography devices are exposed to harsh conditions following each target implosion or pinching of plasma. Key issues of the cyclic IFE operation include intense photon and ion deposition, wall thermal and hydrodynamic evolution, wall erosion and fatigue lifetime, and chamber clearing and evacuation to ensure desirable conditions prior to target implosion. Detailed models have been developed for reflected laser light, emitted photons, neutrons, and target debris deposition and interaction with chamber components and have been implemented in the comprehensive HEIGHTS software package. The hydrodynamic response of chamber walls in bare or in gas-filled cavities and the photon transport of the deposited energy has been calculated by means of new and advanced numerical techniques for accurate shock treatment and propagation. These models include detail media hydrodynamics, non-LTE multi-group for both continuum and line radiation transport, and dynamics of eroded debris resulting from the intense energy deposition. The focus of this study is to critically assess the reliability and the dynamic response of chamber walls in various proposed protection methods for IFE systems. Key requirements are that: (i) the chamber wall accommodates the cyclic energy deposition while providing the required lifetime due to various erosion mechanisms, such as vaporization, chemical and physical sputtering, melt/liquid splashing and explosive erosion, and fragmentation of liquid walls, and (ii) after each shot the chamber is cleared and returned to a quiescent state in preparation for the target injection and the firing of the driver for the subsequent shot. This paper investigates in details these two important issues and found that the required operating frequency of the IFE reactors for power production may be severely limited due to these two requirements. (author)

  11. Mobile Technology Affinity in Renal Transplant Recipients.

    Science.gov (United States)

    Reber, S; Scheel, J; Stoessel, L; Schieber, K; Jank, S; Lüker, C; Vitinius, F; Grundmann, F; Eckardt, K-U; Prokosch, H-U; Erim, Y

    Medication nonadherence is a common problem in renal transplant recipients (RTRs). Mobile health approaches to improve medication adherence are a current trend, and several medication adherence apps are available. However, it is unknown whether RTRs use these technologies and to what extent. In the present study, the mobile technology affinity of RTRs was analyzed. We hypothesized significant age differences in mobile technology affinity and that mobile technology affinity is associated with better cognitive functioning as well as higher educational level. A total of 109 RTRs (63% male) participated in the cross-sectional study, with an overall mean age of 51.8 ± 14.2 years. The study included the Technology Experience Questionnaire (TEQ) for the assessment of mobile technology affinity, a cognitive test battery, and sociodemographic data. Overall, 57.4% of the patients used a smartphone or tablet and almost 45% used apps. The TEQ sum score was 20.9 in a possible range from 6 (no affinity to technology) to 30 (very high affinity). Younger patients had significantly higher scores in mobile technology affinity. The only significant gender difference was found in having fun with using electronic devices: Men enjoyed technology more than women did. Mobile technology affinity was positively associated with cognitive functioning and educational level. Young adult patients might profit most from mobile health approaches. Furthermore, high educational level and normal cognitive functioning promote mobile technology affinity. This should be kept in mind when designing mobile technology health (mHealth) interventions for RTRs. For beneficial mHealth interventions, further research on potential barriers and desired technologic features is necessary to adapt apps to patients' needs. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Dramatic Response of BRAF V600E Mutant Papillary Craniopharyngioma to Targeted Therapy.

    Science.gov (United States)

    Brastianos, Priscilla K; Shankar, Ganesh M; Gill, Corey M; Taylor-Weiner, Amaro; Nayyar, Naema; Panka, David J; Sullivan, Ryan J; Frederick, Dennie T; Abedalthagafi, Malak; Jones, Pamela S; Dunn, Ian F; Nahed, Brian V; Romero, Javier M; Louis, David N; Getz, Gad; Cahill, Daniel P; Santagata, Sandro; Curry, William T; Barker, Fred G

    2016-02-01

    We recently reported that BRAF V600E is the principal oncogenic driver of papillary craniopharyngioma, a highly morbid intracranial tumor commonly refractory to treatment. Here, we describe our treatment of a man age 39 years with multiply recurrent BRAF V600E craniopharyngioma using dabrafenib (150mg, orally twice daily) and trametinib (2mg, orally twice daily). After 35 days of treatment, tumor volume was reduced by 85%. Mutations that commonly mediate resistance to MAPK pathway inhibition were not detected in a post-treatment sample by whole exome sequencing. A blood-based BRAF V600E assay detected circulating BRAF V600E in the patient's blood. Re-evaluation of the existing management paradigms for craniopharyngioma is warranted, as patient morbidity might be reduced by noninvasive mutation testing and neoadjuvant-targeted treatment. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. A Colon Targeted Delivery System for Resveratrol Enriching in pH Responsive-Model

    DEFF Research Database (Denmark)

    Andishmand, Hashem; Hamishehkar, Hamed; Mohammadifar, Mohammad Amin

    2017-01-01

    Background: Resveratrol effects on the prevention and treatment of colon cancer have been well documented recently, but low solubility, rapid absorption and metabolism of resveratrol limit its beneficial effects on colon cancer. Designing a formulation that enhances the solubility of resveratrol......, protects resveratrol from oxidation and isomerization, and delivers it to the colon is a priority of food and drug industry. In this study, resveratrol-polyethylene glycol (PEG)-loaded pectin-chitosan polyelectrolyte complex was designed as a colon targeted delivery system. Methods: The effects of adding...... PEG, ultra-sonication time, pH, and pectin to chitosan ratio were investigated on particle size, polydispersity index (PDI), zeta potential by particle size analyzer, and scanning electron microscopy (SEM). Encapsulation efficiency (EE), release of resveratrol in simulated gastrointestinal fluid...

  14. Dose Dependent Survival Response in Chronic Myeloid Leukemia under Continuous and Pulsed Targeted Therapy

    International Nuclear Information System (INIS)

    Pizzolato, N.; Valenti, D.; Spagnolo, B.; Persano Adorno, D.

    2010-01-01

    A simulative study of cancer growth dynamics in patients affected by Chronic Myeloid Leukemia (CML), under the effect of a targeted dose dependent continuous or pulsed therapy, is presented. We have developed a model for the dynamics of CML in which the stochastic evolution of white blood cell populations are simulated by adopting a Monte Carlo approach. Several scenarios in the evolutionary dynamics of white blood cells, as a consequence of the efficacy of the different modelled therapies, pulsed or continuous, are described. The best results, in terms of a permanent disappearance of the leukemic phenotype, are achieved with a continuous therapy and higher dosage. However, our findings demonstrate that an intermittent therapy could represent a valid choice in patients with high risk of toxicity, when a long-term therapy is considered. A suitably tuned pulsed therapy can enhance the treatment efficacy and reduce the percentage of patients developing resistance. (authors)

  15. HIV-1 accessory proteins VPR and Vif modulate antiviral response by targeting IRF-3 for degradation

    International Nuclear Information System (INIS)

    Okumura, Atsushi; Alce, Tim; Lubyova, Barbora; Ezelle, Heather; Strebel, Klaus; Pitha, Paula M.

    2008-01-01

    The activation of IRF-3 during the early stages of viral infection is critical for the initiation of the antiviral response; however the activation of IRF-3 in HIV-1 infected cells has not yet been characterized. We demonstrate that the early steps of HIV-1 infection do not lead to the activation and nuclear translocation of IRF-3; instead, the relative levels of IRF-3 protein are decreased due to the ubiquitin-associated proteosome degradation. Addressing the molecular mechanism of this effect we show that the degradation is independent of HIV-1 replication and that virion-associated accessory proteins Vif and Vpr can independently degrade IRF-3. The null mutation of these two genes reduced the capacity of the HIV-1 virus to down modulate IRF-3 levels. The degradation was associated with Vif- and Vpr-mediated ubiquitination of IRF-3 and was independent of the activation of IRF-3. N-terminal lysine residues were shown to play a critical role in the Vif- and Vpr-mediated degradation of IRF-3. These data implicate Vif and Vpr in the disruption of the initial antiviral response and point to the need of HIV-1 to circumvent the antiviral response during the very early phase of replication

  16. First- and Second-Line Targeted Systemic Therapy in Hepatocellular Carcinoma—An Update on Patient Selection and Response Evaluation

    Directory of Open Access Journals (Sweden)

    Johann von Felden

    2016-11-01

    Full Text Available Advanced hepatocellular carcinoma (HCC with vascular invasion and/or extrahepatic spread and preserved liver function, according to stage C of the Barcelona Clinic Liver Cancer (BCLC classification, has a dismal prognosis. The multi-targeted tyrosine-kinase receptor inhibitor (TKI sorafenib is the only proven active substance in systemic HCC therapy for first-line treatment. In this review, we summarize current aspects in patient selection and management of side effects, and provide an update on response evaluation during first-line sorafenib therapy. Since second-line treatment options have been improved with the successful completion of the RESORCE trial, demonstrating a survival benefit for second-line treatment with the TKI regorafenib, response monitoring during first-line therapy will be critical to deliver optimal systemic therapy in HCC. To this regard, specific side effects, in particular worsening of arterial hypertension and diarrhea, might suggest treatment response during first-line sorafenib therapy; however, clear predictive clinical markers, as well as laboratory test or serum markers, are not established. Assessment of radiologic response according to the modified Response Evaluation Criteria in Solid Tumors (mRECIST is helpful to identify patients who do not benefit from sorafenib treatment.

  17. The Swiss Alpine Glacier's Response to the '2°C Target

    Science.gov (United States)

    Salzmann, Nadine; Machguth, Horst

    2010-05-01

    The "2°C target" for global warming (relative to pre-industrial level) became a main focus in the climate change debate since the UN Climate Change Conference in Copenhagen (COP15) in December 2009 at the latest. While this target implies to be a ‘clear' goal for politicians and decision makers, the effective impacts that a global mean air temperature increase of 2°C has on natural and human systems on regional to local scales remain complex. So far, most impact studies use only relative and static 2°C delta change approaches. Here, however, we use results from latest climate model outputs an take into account the warming that has already occurred in a specific region. Global warming is not equally distributed around the globe. Observations show that during the last century air temperature trends significantly differ between regions. In Switzerland, for example, air temperature has increased about twice as much as the global mean during the last century. In glacierised mountain regions, where glaciers represent an important source for fresh water and control a great part of the hydrological cycle, the retreat or disappearance of glaciers as a consequence of climatic changes will have major socio-economical consequences on the people living there and the adjacent lowland. A trend to negative glacier mass balances is observed and well documented for many mountain ranges all over the world. Based on climate model projections it is very likely that this trend continues or even accelerates. Here, we make an effort to assess the impact of a global 2°C (that is about 4°C for Switzerland) air temperature increase compared to pre-industrial conditions for the Swiss Alpine glaciers. We use 12 homogenised long-term climate observations to define the warming that has already taken place. The ‘remaining' temperature increase up to the level of 2°C, is based on results from a selection of Regional Climate Model results that have been simulated in the recently finished

  18. Dual DNA binding property of ABA insensitive 3 like factors targeted to promoters responsive to ABA and auxin.

    Science.gov (United States)

    Nag, Ronita; Maity, Manas Kanti; Dasgupta, Maitrayee

    2005-11-01

    The ABA responsive ABI3 and the auxin responsive ARF family of transcription factors bind the CATGCATG (Sph) and TGTCTC core motifs in ABA and auxin response elements (ABRE and AuxRE), respectively. Several evidences indicate ABI3s to act downstream to auxin too. Because DNA binding domain of ABI3s shows significant overlap with ARFs we enquired whether auxin responsiveness through ABI3s could be mediated by their binding to canonical AuxREs. Investigations were undertaken through in vitro gel mobility shift assays (GMSA) using the DNA binding domain B3 of PvAlf (Phaseolus vulgaris ABI3 like factor) and upstream regions of auxin responsive gene GH3 (-267 to -141) and ABA responsive gene Em (-316 to -146) harboring AuxRE and ABRE, respectively. We demonstrate that B3 domain of PvAlf could bind AuxRE only when B3 was associated with its flanking domain B2 (B2B3). Such strict requirement of B2 domain was not observed with ABRE, where B3 could bind with or without being associated with B2. This dual specificity in DNA binding of ABI3s was also demonstrated with nuclear extracts of cultured cells of Arachis hypogea. Supershift analysis of ABRE and AuxRE bound nuclear proteins with antibodies raised against B2B3 domains of PvAlf revealed that ABI3 associated complexes were detectable in association with both cis elements. Competition GMSA confirmed the same complexes to bind ABRE and AuxRE. This dual specificity of ABI3 like factors in DNA binding targeted to natural promoters responsive to ABA and auxin suggests them to have a potential role in conferring crosstalk between these two phytohormones.

  19. Study of Geological Analogues for Understanding the Radar Sounder Response of the RIME Targets

    Science.gov (United States)

    Thakur, S.; Bruzzone, L.

    2017-12-01

    Radar for Icy Moon Exploration (RIME), the radar sounder onboard the Jupiter Icy Moons Explorer (JUICE), is aimed at characterizing the ice shells of the Jovian moons - Ganymede, Europa and Callisto. RIME is optimized to operate at 9 MHz central frequency with bandwidth of 1 MHz and 2.7 MHz to achieve a penetration depth up to 9 km through ice. We have developed an approach to the definition of a database of simulated RIME radargrams by leveraging the data available from airborne and orbital radar sounder acquisitions over geological analogues of the expected icy moon features. These simulated radargrams are obtained by merging real radar sounder data with models of the subsurface of the Jupiter icy moons. They will be useful for geological interpretation of the RIME radargrams and for better predicting the performance of RIME. The database will also be useful in developing pre-processing and automatic feature extraction algorithms to support data analysis during the mission phase of RIME. Prior to the JUICE mission exploring the Jovian satellites with RIME, there exist radar sounders such as SHARAD (onboard MRO) and MARSIS (onboard MEX) probing Mars, the LRS (onboard SELENE) probing the Moon, and many airborne sounders probing the polar regions of Earth. Analogues have been identified in these places based on similarity in geo-morphological expression. Moreover, other analogues have been identified on the Earth for possible dedicated acquisition campaigns before the RIME operations. By assuming that the subsurface structure of the RIME targets is approximately represented in the analogue radargrams, the difference in composition is accounted for by imposing different dielectric and subsurface attenuation models. The RIME radargrams are simulated from the analogue radargrams using the radar equation and the RIME processing chain and accounting for different possible scenarios in terms of subsurface structure, dielectric properties and instrument parameters. For

  20. Brugia malayi gene expression in response to the targeting of the Wolbachia endosymbiont by tetracycline treatment.

    Directory of Open Access Journals (Sweden)

    Elodie Ghedin

    2009-10-01

    Full Text Available Brugia malayi, like most human filarial parasite species, harbors an endosymbiotic bacterium of the genus Wolbachia. Elimination of the endosymbiont leads to sterilization of the adult female. Previous biochemical and genetic studies have established that communication with its endobacterium is essential for survival of the worm.We used electron microscopy to examine the effects of antibiotic treatment on Wolbachia cell structure. We have also used microarray and quantitative RT-PCR analyses to examine the regulation of the B. malayi transcripts altered in response to the anti-Wolbachia treatment. Microscopy of worms taken from animals treated with tetracycline for 14 and 21 days (14 d and 21 d demonstrated substantial morphologic effects on the Wolbachia endobacterium by 14 d and complete degeneration of the endobacterial structures by 21 d. We observed upregulation of transcripts primarily encoding proteins involved in amino acid synthesis and protein translation, and downregulation of transcripts involved in cuticle biosynthesis after both 7 d and 14 d of treatment. In worms exposed to tetracycline in culture, substantial effects on endobacteria morphology were evident by day 3, and extensive death of the endobacteria was observed by day 5. In a detailed examination of the expression kinetics of selected signaling genes carried out on such cultured worms, a bimodal pattern of regulation was observed. The selected genes were upregulated during the early phase of antibiotic treatment and quickly downregulated in the following days. These same genes were upregulated once more at 6 days post-treatment.Upregulation of protein translation and amino acid synthesis may indicate a generalized stress response induced in B. malayi due to a shortage of essential nutrients/factors that are otherwise supplied by Wolbachia. Downregulation of transcripts involved in cuticle biosynthesis perhaps reflects a disruption in the normal embryogenic program. This is

  1. Targeted Apoptosis of Senescent Cells Restores Tissue Homeostasis in Response to Chemotoxicity and Aging.

    Science.gov (United States)

    Baar, Marjolein P; Brandt, Renata M C; Putavet, Diana A; Klein, Julian D D; Derks, Kasper W J; Bourgeois, Benjamin R M; Stryeck, Sarah; Rijksen, Yvonne; van Willigenburg, Hester; Feijtel, Danny A; van der Pluijm, Ingrid; Essers, Jeroen; van Cappellen, Wiggert A; van IJcken, Wilfred F; Houtsmuller, Adriaan B; Pothof, Joris; de Bruin, Ron W F; Madl, Tobias; Hoeijmakers, Jan H J; Campisi, Judith; de Keizer, Peter L J

    2017-03-23

    The accumulation of irreparable cellular damage restricts healthspan after acute stress or natural aging. Senescent cells are thought to impair tissue function, and their genetic clearance can delay features of aging. Identifying how senescent cells avoid apoptosis allows for the prospective design of anti-senescence compounds to address whether homeostasis can also be restored. Here, we identify FOXO4 as a pivot in senescent cell viability. We designed a FOXO4 peptide that perturbs the FOXO4 interaction with p53. In senescent cells, this selectively causes p53 nuclear exclusion and cell-intrinsic apoptosis. Under conditions where it was well tolerated in vivo, this FOXO4 peptide neutralized doxorubicin-induced chemotoxicity. Moreover, it restored fitness, fur density, and renal function in both fast aging Xpd TTD/TTD and naturally aged mice. Thus, therapeutic targeting of senescent cells is feasible under conditions where loss of health has already occurred, and in doing so tissue homeostasis can effectively be restored. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Critical target and dose and dose-rate responses for the induction of chromosomal instability by ionizing radiation

    Science.gov (United States)

    Limoli, C. L.; Corcoran, J. J.; Milligan, J. R.; Ward, J. F.; Morgan, W. F.

    1999-01-01

    To investigate the critical target, dose response and dose-rate response for the induction of chromosomal instability by ionizing radiation, bromodeoxyuridine (BrdU)-substituted and unsubstituted GM10115 cells were exposed to a range of doses (0.1-10 Gy) and different dose rates (0.092-17.45 Gy min(-1)). The status of chromosomal stability was determined by fluorescence in situ hybridization approximately 20 generations after irradiation in clonal populations derived from single progenitor cells surviving acute exposure. Overall, nearly 700 individual clones representing over 140,000 metaphases were analyzed. In cells unsubstituted with BrdU, a dose response was found, where the probability of observing delayed chromosomal instability in any given clone was 3% per gray of X rays. For cells substituted with 25-66% BrdU, however, a dose response was observed only at low doses (1.0 Gy), the incidence of chromosomal instability leveled off. There was an increase in the frequency and complexity of chromosomal instability per unit dose compared to cells unsubstituted with BrdU. The frequency of chromosomal instability appeared to saturate around approximately 30%, an effect which occurred at much lower doses in the presence of BrdU. Changing the gamma-ray dose rate by a factor of 190 (0.092 to 17.45 Gy min(-1)) produced no significant differences in the frequency of chromosomal instability. The enhancement of chromosomal instability promoted by the presence of the BrdU argues that DNA comprises at least one of the critical targets important for the induction of this end point of genomic instability.

  3. Solute Carrier NTCP Regulates Innate Antiviral Immune Responses Targeting Hepatitis C Virus Infection of Hepatocytes.

    Science.gov (United States)

    Verrier, Eloi R; Colpitts, Che C; Bach, Charlotte; Heydmann, Laura; Zona, Laetitia; Xiao, Fei; Thumann, Christine; Crouchet, Emilie; Gaudin, Raphaël; Sureau, Camille; Cosset, François-Loïc; McKeating, Jane A; Pessaux, Patrick; Hoshida, Yujin; Schuster, Catherine; Zeisel, Mirjam B; Baumert, Thomas F

    2016-10-25

    Chronic hepatitis B, C, and D virus (HBV, HCV, and HDV) infections are the leading causes of liver disease and cancer worldwide. Recently, the solute carrier and sodium taurocholate co-transporter NTCP has been identified as a receptor for HBV and HDV. Here, we uncover NTCP as a host factor regulating HCV infection. Using gain- and loss-of-function studies, we show that NTCP mediates HCV infection of hepatocytes and is relevant for cell-to-cell transmission. NTCP regulates HCV infection by augmenting the bile-acid-mediated repression of interferon-stimulated genes (ISGs), including IFITM3. In conclusion, our results uncover NTCP as a mediator of innate antiviral immune responses in the liver, and they establish a role for NTCP in the infection process of multiple viruses via distinct mechanisms. Collectively, our findings suggest a role for solute carriers in the regulation of innate antiviral responses, and they have potential implications for virus-host interactions and antiviral therapies. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  4. Solute Carrier NTCP Regulates Innate Antiviral Immune Responses Targeting Hepatitis C Virus Infection of Hepatocytes

    Directory of Open Access Journals (Sweden)

    Eloi R. Verrier

    2016-10-01

    Full Text Available Chronic hepatitis B, C, and D virus (HBV, HCV, and HDV infections are the leading causes of liver disease and cancer worldwide. Recently, the solute carrier and sodium taurocholate co-transporter NTCP has been identified as a receptor for HBV and HDV. Here, we uncover NTCP as a host factor regulating HCV infection. Using gain- and loss-of-function studies, we show that NTCP mediates HCV infection of hepatocytes and is relevant for cell-to-cell transmission. NTCP regulates HCV infection by augmenting the bile-acid-mediated repression of interferon-stimulated genes (ISGs, including IFITM3. In conclusion, our results uncover NTCP as a mediator of innate antiviral immune responses in the liver, and they establish a role for NTCP in the infection process of multiple viruses via distinct mechanisms. Collectively, our findings suggest a role for solute carriers in the regulation of innate antiviral responses, and they have potential implications for virus-host interactions and antiviral therapies.

  5. Affinity purification of recombinant human plasminogen activator ...

    African Journals Online (AJOL)

    Affinity purification of recombinant human plasminogen activator from ... Screening antibody was performed using rhPA milk in an ELISA-elution assay. ... useful for purifying other tPA mutants or other novel recombinant milkderived proteins.

  6. PRINCIPLES OF AFFINITY-BASED BIOSENSORS

    Science.gov (United States)

    Despite the amount of resources that have been invested by national and international academic, government, and commercial sectors to develop affinity-based biosensor products, little obvious success has been realized through commercialization of these devices for specific applic...

  7. Quantum deformation of the affine transformation algebra

    International Nuclear Information System (INIS)

    Aizawa, N.; Sato, Haru-Tada

    1994-01-01

    We discuss a quantum deformation of the affine transformation algebra in one-dimensional space. It is shown that the quantum algebra has a non-cocommutative Hopf algebra structure, simple realizations and quantum tensor operators. (orig.)

  8. Dynamics of Open Systems with Affine Maps

    International Nuclear Information System (INIS)

    Zhang Da-Jian; Liu Chong-Long; Tong Dian-Min

    2015-01-01

    Many quantum systems of interest are initially correlated with their environments and the reduced dynamics of open systems are an interesting while challenging topic. Affine maps, as an extension of completely positive maps, are a useful tool to describe the reduced dynamics of open systems with initial correlations. However, it is unclear what kind of initial state shares an affine map. In this study, we give a sufficient condition of initial states, in which the reduced dynamics can always be described by an affine map. Our result shows that if the initial states of the combined system constitute a convex set, and if the correspondence between the initial states of the open system and those of the combined system, defined by taking the partial trace, is a bijection, then the reduced dynamics of the open system can be described by an affine map. (paper)

  9. On the Lp affine isoperimetric inequalities

    Indian Academy of Sciences (India)

    surface area measure on convex bodies. We also establish the reverse version of -Petty projection inequality and an affine isoperimetric inequality of − p K . Author Affiliations. Wuyang Yu1 Gangsong Leng2. Institute of Management Decision ...

  10. Hierarchy and Assortativity as New Tools for Binding-Affinity Investigation: The Case of the TBA Aptamer-Ligand Complex.

    Science.gov (United States)

    Cataldo, Rosella; Alfinito, Eleonora; Reggiani, Lino

    2017-12-01

    Aptamers are single stranded DNA, RNA, or peptide sequences having the ability to bind several specific targets (proteins, molecules as well as ions). Therefore, aptamer production and selection for therapeutic and diagnostic applications is very challenging. Usually, they are generated in vitro, although computational approaches have been recently developed for the in silico production. Despite these efforts, the mechanism of aptamer-ligand formation is not completely clear, and producing high-affinity aptamers is still quite difficult. This paper aims to develop a computational model able to describe aptamer-ligand affinity. Topological tools, such as the conventional degree distribution, the rank-degree distribution (hierarchy), and the node assortativity are employed. In doing so, the macromolecules tertiary-structures are mapped into appropriate graphs. These graphs reproduce the main topological features of the macromolecules, by preserving the distances between amino acids (nucleotides). Calculations are applied to the thrombin binding aptamer (TBA), and the TBA-thrombin complex produced in the presence of Na + or K + . The topological analysis is able to detect several differences between complexes obtained in the presence of the two cations, as expected by previous investigations. These results support graph analysis as a novel computational tool for testing affinity. Otherwise, starting from the graphs, an electrical network can be obtained by using the specific electrical properties of amino acids and nucleobases. Therefore, a further analysis concerns with the electrical response, revealing that the resistance is sensitively affected by the presence of sodium or potassium, thus suggesting resistance as a useful physical parameter for testing binding affinity.

  11. Dual antibody therapy to harness the innate anti-tumor immune response to enhance antibody targeting of tumors.

    Science.gov (United States)

    Chester, Cariad; Marabelle, Aurelien; Houot, Roch; Kohrt, Holbrook E

    2015-04-01

    Cancer immunotherapy is a rapidly evolving field that offers a novel paradigm for cancer treatment: therapies focus on enhancing the immune system's innate and adaptive anti-tumor response. Early immunotherapeutics have achieved impressive clinical outcomes and monoclonal antibodies are now integral to therapeutic strategies in a variety of cancers. However, only recently have antibodies targeting innate immune cells entered clinical development. Innate immune effector cells play important roles in generating and maintaining antitumor immunity. Antibody-dependent cell-mediated cytotoxicity (ADCC) and antibody-dependent cellular phagocytosis (ADCP) are important innate immune mechanisms for tumor eradication. These cytolytic processes are initiated by the detection of a tumor-targeting antibody and can be augmented by activating co-stimulatory pathways or blocking inhibitory signals on innate immune cells. The combination of FDA-approved monoclonal antibodies with innate effector-targeting antibodies has demonstrated potent preclinical therapeutic synergy and early-phase combinatorial clinical trials are ongoing. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. The Development of a 3D LADAR Simulator Based on a Fast Target Impulse Response Generation Approach

    Science.gov (United States)

    Al-Temeemy, Ali Adnan

    2017-09-01

    A new laser detection and ranging (LADAR) simulator has been developed, using MATLAB and its graphical user interface, to simulate direct detection time of flight LADAR systems, and to produce 3D simulated scanning images under a wide variety of conditions. This simulator models each stage from the laser source to data generation and can be considered as an efficient simulation tool to use when developing LADAR systems and their data processing algorithms. The novel approach proposed for this simulator is to generate the actual target impulse response. This approach is fast and able to deal with high scanning requirements without losing the fidelity that accompanies increments in speed. This leads to a more efficient LADAR simulator and opens up the possibility for simulating LADAR beam propagation more accurately by using a large number of laser footprint samples. The approach is to select only the parts of the target that lie in the laser beam angular field by mathematically deriving the required equations and calculating the target angular ranges. The performance of the new simulator has been evaluated under different scanning conditions, the results showing significant increments in processing speeds in comparison to conventional approaches, which are also used in this study as a point of comparison for the results. The results also show the simulator's ability to simulate phenomena related to the scanning process, for example, type of noise, scanning resolution and laser beam width.

  13. microRNAs targeting DEAD-box helicases are involved in salinity stress response in rice (Oryza sativa L.

    Directory of Open Access Journals (Sweden)

    Macovei Anca

    2012-10-01

    Full Text Available Abstract Background Rice (Oryza sativa L., one of the most important food crop in the world, is considered to be a salt-sensitive crop. Excess levels of salt adversely affect all the major metabolic activities, including cell wall damage, cytoplasmic lysis and genomic stability. In order to cope with salt stress, plants have evolved high degrees of developmental plasticity, including adaptation via cascades of molecular networks and changes in gene expression profiles. Posttranscriptional regulation, through the activity of microRNAs, also plays an important role in the plant response to salinity conditions. MicroRNAs are small endogenous RNAs that modulate gene expression and are involved in the most essential physiological processes, including plant development and adaptation to environmental changes. Results In the present study, we investigated the expression profiles of osa-MIR414, osa-MIR408 and osa-MIR164e along with their targeted genes, under salinity stress conditions in wild type and transgenic rice plants ectopically expressing the PDH45 (Pea DNA Helicase gene. The present miRNAs were predicted to target the OsABP (ATP-Binding Protein, OsDSHCT (DOB1/SK12/helY-like DEAD-box Helicase and OsDBH (DEAD-Box Helicase genes, included in the DEAD-box helicase family. An in silico characterization of the proteins was performed and the miRNAs predicted targets were validated by RLM-5′RACE. The qRT-PCR analysis showed that the OsABP, OsDBH and OsDSHCT genes were up-regulated in response to 100 and 200 mM NaCl treatments. The present study also highlighted an increased accumulation of the gene transcripts in wild type plants, with the exception of the OsABP mRNA which showed the highest level (15.1-fold change compared to control in the transgenic plants treated with 200 mM NaCl. Salinity treatments also affected the expression of osa-MIR414, osa-MIR164e and osa-MIR408, found to be significantly down-regulated, although the changes in mi

  14. High-throughput fragment screening by affinity LC-MS.

    Science.gov (United States)

    Duong-Thi, Minh-Dao; Bergström, Maria; Fex, Tomas; Isaksson, Roland; Ohlson, Sten

    2013-02-01

    Fragment screening, an emerging approach for hit finding in drug discovery, has recently been proven effective by its first approved drug, vemurafenib, for cancer treatment. Techniques such as nuclear magnetic resonance, surface plasmon resonance, and isothemal titration calorimetry, with their own pros and cons, have been employed for screening fragment libraries. As an alternative approach, screening based on high-performance liquid chromatography separation has been developed. In this work, we present weak affinity LC/MS as a method to screen fragments under high-throughput conditions. Affinity-based capillary columns with immobilized thrombin were used to screen a collection of 590 compounds from a fragment library. The collection was divided into 11 mixtures (each containing 35 to 65 fragments) and screened by MS detection. The primary screening was performed in 3500 fragments per day). Thirty hits were defined, which subsequently entered a secondary screening using an active site-blocked thrombin column for confirmation of specificity. One hit showed selective binding to thrombin with an estimated dissociation constant (K (D)) in the 0.1 mM range. This study shows that affinity LC/MS is characterized by high throughput, ease of operation, and low consumption of target and fragments, and therefore it promises to be a valuable method for fragment screening.

  15. Affinity chromatography: A versatile technique for antibody purification.

    Science.gov (United States)

    Arora, Sushrut; Saxena, Vikas; Ayyar, B Vijayalakshmi

    2017-03-01

    Antibodies continue to be extremely utilized entities in myriad applications including basic research, imaging, targeted delivery, chromatography, diagnostics, and therapeutics. At production stage, antibodies are generally present in complex matrices and most of their intended applications necessitate purification. Antibody purification has always been a major bottleneck in downstream processing of antibodies, due to the need of high quality products and associated high costs. Over the years, extensive research has focused on finding better purification methodologies to overcome this holdup. Among a plethora of different techniques, affinity chromatography is one of the most selective, rapid and easy method for antibody purification. This review aims to provide a detailed overview on affinity chromatography and the components involved in purification. An array of support matrices along with various classes of affinity ligands detailing their underlying working principles, together with the advantages and limitations of each system in purifying different types of antibodies, accompanying recent developments and important practical methodological considerations to optimize purification procedure are discussed. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Recombinant spider silk genetically functionalized with affinity domains.

    Science.gov (United States)

    Jansson, Ronnie; Thatikonda, Naresh; Lindberg, Diana; Rising, Anna; Johansson, Jan; Nygren, Per-Åke; Hedhammar, My

    2014-05-12

    Functionalization of biocompatible materials for presentation of active protein domains is an area of growing interest. Herein, we describe a strategy for functionalization of recombinant spider silk via gene fusion to affinity domains of broad biotechnological use. Four affinity domains of different origin and structure; the IgG-binding domains Z and C2, the albumin-binding domain ABD, and the biotin-binding domain M4, were all successfully produced as soluble silk fusion proteins under nondenaturing purification conditions. Silk films and fibers produced from the fusion proteins were demonstrated to be chemically and thermally stable. Still, the bioactive domains are concluded to be folded and accessible, since their respective targets could be selectively captured from complex samples, including rabbit serum and human plasma. Interestingly, materials produced from mixtures of two different silk fusion proteins displayed combined binding properties, suggesting that tailor-made materials with desired stoichiometry and surface distributions of several binding domains can be produced. Further, use of the IgG binding ability as a general mean for presentation of desired biomolecules could be demonstrated for a human vascular endothelial growth factor (hVEGF) model system, via a first capture of anti-VEGF IgG to silk containing the Z-domain, followed by incubation with hVEGF. Taken together, this study demonstrates the potential of recombinant silk, genetically functionalized with affinity domains, for construction of biomaterials capable of presentation of almost any desired biomolecule.

  17. RNAseq reveals weed-induced PIF3-like as a candidate target to manipulate weed stress response in soybean.

    Science.gov (United States)

    Horvath, David P; Hansen, Stephanie A; Moriles-Miller, Janet P; Pierik, Ronald; Yan, Changhui; Clay, David E; Scheffler, Brian; Clay, Sharon A

    2015-07-01

    Weeds reduce yield in soybeans (Glycine max) through incompletely defined mechanisms. The effects of weeds on the soybean transcriptome were evaluated in field conditions during four separate growing seasons. RNASeq data were collected from six biological samples of soybeans growing with or without weeds. Weed species and the methods to maintain weed-free controls varied between years to mitigate treatment effects, and to allow detection of general soybean weed responses. Soybean plants were not visibly nutrient- or water-stressed. We identified 55 consistently downregulated genes in weedy plots. Many of the downregulated genes were heat shock genes. Fourteen genes were consistently upregulated. Several transcription factors including a PHYTOCHROME INTERACTING FACTOR 3-like gene (PIF3) were included among the upregulated genes. Gene set enrichment analysis indicated roles for increased oxidative stress and jasmonic acid signaling responses during weed stress. The relationship of this weed-induced PIF3 gene to genes involved in shade avoidance responses in Arabidopsis provide evidence that this gene may be important in the response of soybean to weeds. These results suggest that the weed-induced PIF3 gene will be a target for manipulating weed tolerance in soybean. No claim to original US government works New Phytologist © 2015 New Phytologist Trust.

  18. Aging and Spaceflight: Catalase Targeted to Mitochondria Alters Skeletal Structure and Responses to Musculoskeletal Disuse

    Science.gov (United States)

    Globus, Ruth K.; Tahimic, Candice; Schreurs, Ann-Sofie

    2018-01-01

    Microgravity and ionizing radiation in the spaceflight environment pose multiple challenges to homeostasis and may contribute to cellular stress. Effects may include increased generation of reactive oxygen species (ROS), DNA damage and repair error, cell cycle arrest, cell senescence or death. Our central hypothesis is that prolonged exposure to the spaceflight environment leads to excess production of ROS and oxidative damage, culminating in accelerated tissue degeneration which resembles aging. The main goal of this project is to determine the importance of cellular redox defense for physiological adaptations and tissue degeneration in the space environment. To accomplish this, we will use both wildtype (WT) mice and a well-established, genetically-engineered animal model (mCAT mice) which displays extended lifespan (Schriner et al. 2005). The animal model selected to test these ideas is engineered to quench ROS in mitochondria by targeted over-expression of the human catalase gene to the mitochondrial matrix. We showed previously that mCAT mice express the catalase transgene in skeletal tissues, bone forming osteoblasts, and bone resorbing osteoclasts. In addition, mCAT mice also display increased catalase activity in bone. Our findings revealed that exposure of adult, male, C57Bl/6J mice to simulated spaceflight (hindlimb unloading and gamma radiation) led to an increase in markers of oxidative damage (malondialdehyde, 4-hydroxynonenol) in skeletal tissue of WT mice but not mCAT mice. To extend our hypothesis to other, spaceflight-relevant tissues, we are performing a ground-based study simulating 30 days of spaceflight by hindlimb unloading to determine potential protective effects of mitochondrial catalase activity on aging of multiple tissues (cardiovascular, nervous and skeletal).

  19. A Colon Targeted Delivery System for Resveratrol Enriching in pH Responsive-Model

    Directory of Open Access Journals (Sweden)

    Hashem Andishmand, Hamed Hamishehkar, Afshin Babazadeh, Arezou Taghvimi, Mohammad Amin Mohammadifar, Mahnaz Tabibiazar

    2017-03-01

    Full Text Available Background: Resveratrol effects on the prevention and treatment of colon cancer have been well documented recently, but low solubility, rapid absorption and metabolism of resveratrol limit its beneficial effects on colon cancer. Designing a formulation that enhances the solubility of resveratrol, protects resveratrol from oxidation and isomerization, and delivers it to the colon is a priority of food and drug industry. In this study, resveratrol-polyethylene glycol (PEG-loaded pectin-chitosan polyelectrolyte complex was designed as a colon targeted delivery system. Methods: The effects of adding PEG, ultra-sonication time, pH, and pectin to chitosan ratio were investigated on particle size, polydispersity index (PDI, zeta potential by particle size analyzer, and scanning electron microscopy (SEM. Encapsulation efficiency (EE, release of resveratrol in simulated gastrointestinal fluid, and different pHs were analyzed via High Performance Liquid Chromatography (HPLC. Antioxidant activity was measured by (2, 2-diphenyl-1-picryl-hydrazyl-hydrate DPPH free-radical method. Results: Results showed that colloidal stable micro-particles (725 ± 20 nm with PDI < 0.3 and zeta potential +27 ± 2 mV was formed in the ratio of 5:1 of pectin to chitosan w/v % after a 10-min sonication. Encapsulation efficiency was 81 ± 7 %. The reduction of antioxidant activity of resveratrol loaded micro-particles after one month was less than 13%. Micro-particles released about 33% of resveratrol in the simulated gastric and intestinal fluids. Conclusion: Two-thirds of the loaded resveratrol in Pectin-Chitosan complex reached colon. The developed system had enough specification for enriching fruit based drinks due to remarkable colloidal stability in the pH range of 3.5 to 4.5.

  20. Different endothelin receptor affinities in dog tissues

    International Nuclear Information System (INIS)

    Loeffler, B.M.L.; Loehrer, W.

    1991-01-01

    Endothelin (ET) is a long-lasting potent vasoconstrictor-peptide. Here the authors report different binding affinities of endothelin-1 (ET-1) to ET-receptors of various dog tissues. Crude microsomal fractions were prepared after homogenisation of dog tissues in 50 mM Tris/HCl, 20 mM MnCl2, 1 mM EDTA, pH 7.4 by differential centrifugation. Aliquots of microsomal fractions (70 micrograms of protein) were incubated at 25 degrees C for 180 min in the presence of 20 pM 125I-ET-1 and various concentrations of cold ET-1. Four different ET-1 receptor binding affinities were found: adrenals, cerebrum, liver, heart, skeletal muscle and stomach microsomal membranes contained high affinity binding sites (Kd 50 - 80 pM, Bmax 60 - 250 fmol/mg). In cerebellum and spleen medium affinity ET-1 receptors (Kd 350 pM, Bmax 880 and 1200 fmol/mg respectively) were present. In comparison lung and kidney microsomes contained a low affinity ET-1 receptor (Kd 800 and 880 pM, Bmax 1600 and 350 fmol/mg). Receptors of even lower affinity were present in heart, intestine and liver microsomes with Kd values of 3 - 6 nM

  1. Bananas, pesticides and health in southwestern Ecuador: A scalar narrative approach to targeting public health responses.

    Science.gov (United States)

    Brisbois, Benjamin

    2016-02-01

    Public health responses to agricultural pesticide exposure are often informed by ethnographic or other qualitative studies of pesticide risk perception. In addition to highlighting the importance of structural determinants of exposure, such studies can identify the specific scales at which pesticide-exposed individuals locate responsibility for their health issues, with implications for study and intervention design. In this study, an ethnographic approach was employed to map scalar features within explanatory narratives of pesticides and health in Ecuador's banana-producing El Oro province. Unstructured observation, 14 key informant interviews and 15 in-depth semi-structured interviews were carried out during 8 months of fieldwork in 2011-2013. Analysis of interview data was informed by human geographic literature on the social construction of scale. Individual-focused narratives of some participants highlighted characteristics such as carelessness and ignorance, leading to suggestions for educational interventions. More structural explanations invoked farm-scale processes, such as uncontrolled aerial fumigations on plantations owned by elites. Organization into cooperatives helped to protect small-scale farmers from 'deadly' banana markets, which in turn were linked to the Ecuadorian nation-state and actors in the banana-consuming world. These scalar elements interacted in complex ways that appear linked to social class, as more well-off individuals frequently attributed the health problems of other (poorer) people to individual behaviours, while providing more structural explanations of their own difficulties. Such individualizing narratives may help to stabilize inequitable social structures. Research implications of this study include the possibility of using scale-focused qualitative research to generate theory and candidate levels for multi-level models. Equity implications include a need for public health researchers planning interventions to engage with

  2. A Kinetic Model Explains Why Shorter and Less Affine Enzyme-recruiting Oligonucleotides Can Be More Potent

    Directory of Open Access Journals (Sweden)

    Lykke Pedersen

    2014-01-01

    Full Text Available Antisense oligonucleotides complementary to RNA targets promise generality and ease of drug design. The first systemically administered antisense drug was recently approved for treatment and others are in clinical development. Chemical modifications that increase the hybridization affinity of oligonucleotides are reasoned to confer higher potency, i.e., modified oligonucleotides can be dosed at lower concentrations to achieve the same effect. Surprisingly, shorter and less affine oligonucleotides sometimes display increased potency. To explain this apparent contradiction, increased uptake or decreased propensity to form structures have been suggested as possible mechanisms. Here, we provide an alternative explanation that invokes only the kinetics behind oligonucleotide-mediated cleavage of RNA targets. A model based on the law of mass action predicts, and experiments support, the existence of an optimal binding affinity. Exaggerated affinity, and not length per se, is detrimental to potency. This finding clarifies how to optimally apply high-affinity modifications in the discovery of potent antisense oligonucleotide drugs.

  3. Peptide-targeted, stimuli-responsive polymersomes for delivering a cancer stemness inhibitor to cancer stem cell microtumors.

    Science.gov (United States)

    Karandish, Fataneh; Froberg, James; Borowicz, Pawel; Wilkinson, John C; Choi, Yongki; Mallik, Sanku

    2018-03-01

    Often cancer relapses after an initial response to chemotherapy because of the tumor's heterogeneity and the presence of progenitor stem cells, which can renew. To overcome drug resistance, metastasis, and relapse in cancer, a promising approach is the inhibition of cancer stemness. In this study, the expression of the neuropilin-1 receptor in both pancreatic and prostate cancer stem cells was identified and targeted with a stimuli-responsive, polymeric nanocarrier to deliver a stemness inhibitor (napabucasin) to cancer stem cells. Reduction-sensitive amphiphilic block copolymers PEG 1900 -S-S-PLA 6000 and the N 3 -PEG 1900 -PLA 6000 were synthesized. The tumor penetrating iRGD peptide-hexynoic acid conjugate was linked to the N 3 -PEG 1900 -PLA 6000 polymer via a Cu 2+ catalyzed "Click" reaction. Subsequently, this peptide-polymer conjugate was incorporated into polymersomes for tumor targeting and tissue penetration. We prepared polymersomes containing 85% PEG 1900 -S-S-PLA 6000 , 10% iRGD-polymer conjugate, and 5% DPPE-lissamine rhodamine dye. The iRGD targeted polymersomes encapsulating the cancer stemness inhibitor napabucasin were internalized in both prostate and pancreatic cancer stem cells. The napabucasin encapsulated polymersomes significantly (p < .05) reduced the viability of both prostate and pancreatic cancer stem cells and decreased the stemness protein expression notch-1 and nanog compared to the control and vesicles without any drug. The napabucasin encapsulated polymersome formulations have the potential to lead to a new direction in prostate and pancreatic cancer therapy by penetrating deeply into the tumors, releasing the encapsulated stemness inhibitor, and killing cancer stem cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Expression and affinity purification of recombinant proteins from plants

    Science.gov (United States)

    Desai, Urvee A.; Sur, Gargi; Daunert, Sylvia; Babbitt, Ruth; Li, Qingshun

    2002-01-01

    With recent advances in plant biotechnology, transgenic plants have been targeted as an inexpensive means for the mass production of proteins for biopharmaceutical and industrial uses. However, the current plant purification techniques lack a generally applicable, economic, large-scale strategy. In this study, we demonstrate the purification of a model protein, beta-glucuronidase (GUS), by employing the protein calmodulin (CaM) as an affinity tag. In the proposed system, CaM is fused to GUS. In the presence of calcium, the calmodulin fusion protein binds specifically to a phenothiazine-modified surface of an affinity column. When calcium is removed with a complexing agent, e.g., EDTA, calmodulin undergoes a conformational change allowing the dissociation of the calmodulin-phenothiazine complex and, therefore, permitting the elution of the GUS-CaM fusion protein. The advantages of this approach are the fast, efficient, and economical isolation of the target protein under mild elution conditions, thus preserving the activity of the target protein. Two types of transformation methods were used in this study, namely, the Agrobacterium-mediated system and the viral-vector-mediated transformation system. Copyright 2002 Elsevier Science (USA).

  5. A simple method for affinity purification of radiolabeled monoclonal antibodies

    Energy Technology Data Exchange (ETDEWEB)

    Juweid, M; Sato, J; Paik, C; Onay-Basaran, S; Weinstein, J N; Neumann, R D [National Cancer Inst., Bethesda, MD (United States)

    1993-04-01

    A simple method is described for affinity purification of radiolabeled antibodies using glutaraldehyde-fixed tumor target cells. The cell-bound antibody fraction is removed from the cells by an acid wash and then immediately subjected to buffer-exchange chromatography. The method was applied to the D3 murine monoclonal antibody which binds to a 290 kDa antigen on the surface of Line 10 guinea pig carcinoma cells. No alteration in the molecular size profile was detected after acid washing. Purification resulted in a significant increase in immunoreactivity by an average of 14 [+-] 47% (SD; range 4-30%). (author).

  6. Prostate cancer characteristics associated with response to pre-receptor targeting of the androgen axis.

    Directory of Open Access Journals (Sweden)

    Elahe A Mostaghel

    Full Text Available Factors influencing differential responses of prostate tumors to androgen receptor (AR axis-directed therapeutics are poorly understood, and predictors of treatment efficacy are needed. We hypothesized that the efficacy of inhibiting DHT ligand synthesis would associate with intra-tumoral androgen ratios indicative of relative dependence on DHT-mediated growth.We characterized two androgen-sensitive prostate cancer xenograft models after androgen suppression by castration in combination with the SRD5A inhibitor, dutasteride, as well as a panel of castration resistant metastases obtained via rapid autopsy.In LuCaP35 tumors (intra-tumoral T:DHT ratio 2:1 dutasteride suppressed DHT to 0.02 ng/gm and prolonged survival vs. castration alone (337 vs.152 days, HR 2.8, p = 0.0015. In LuCaP96 tumors (T:DHT 10:1, survival was not improved despite similar DHT reduction (0.02 ng/gm. LuCaP35 demonstrated higher expression of steroid biosynthetic enzymes maintaining DHT levels (5-fold higher SRD5A1, 41 fold higher, 99-fold higher RL-HSD, p<0.0001 for both, reconstitution of intra-tumoral DHT (to ∼30% of untreated tumors, and ∼2 fold increased expression of full length AR. In contrast, LuCaP96 demonstrated higher levels of steroid catabolizing enzymes (6.9-fold higher AKR1C2, 3000-fold higher UGT2B15, p = 0.002 and p<0.0001 respectively, persistent suppression of intra-tumoral DHT, and 6-8 fold induction of full length AR and the ligand independent V7 AR splice variant. Human metastases demonstrated bio-active androgen levels and AR full length and AR splice-variant expression consistent with the range observed in xenografts.Intrinsic differences in basal steroidogenesis, as well as variable expression of full length and splice-variant AR, associate with response and resistance to pre-receptor AR ligand suppression. Expression of steroidogenic enzymes and AR isoforms may serve as potential biomarkers of sensitivity to potent AR-axis inhibition and

  7. The metric-affine gravitational theory as the gauge theory of the affine group

    International Nuclear Information System (INIS)

    Lord, E.A.

    1978-01-01

    The metric-affine gravitational theory is shown to be the gauge theory of the affine group, or equivalently, the gauge theory of the group GL(4,R) of tetrad deformations in a space-time with a locally Minkowskian metric. The identities of the metric-affine theory, and the relationship between them and those of general relativity and Sciama-Kibble theory, are derived. (Auth.)

  8. Targeting tumor antigens to secreted membrane vesicles in vivo induces efficient antitumor immune responses.

    Science.gov (United States)

    Zeelenberg, Ingrid S; Ostrowski, Matias; Krumeich, Sophie; Bobrie, Angélique; Jancic, Carolina; Boissonnas, Alexandre; Delcayre, Alain; Le Pecq, Jean-Bernard; Combadière, Béhazine; Amigorena, Sebastian; Théry, Clotilde

    2008-02-15

    Expression of non-self antigens by tumors can induce activation of T cells in vivo, although this activation can lead to either immunity or tolerance. CD8+ T-cell activation can be direct (if the tumor expresses MHC class I molecules) or indirect (after the capture and cross-presentation of tumor antigens by dendritic cells). The modes of tumor antigen capture by dendritic cells in vivo remain unclear. Here we examine the immunogenicity of the same model antigen secreted by live tumors either in association with membrane vesicles (exosomes) or as a soluble protein. We have artificially addressed the antigen to secreted vesicles by coupling it to the factor VIII-like C1C2 domain of milk fat globule epidermal growth factor-factor VIII (MFG-E8)/lactadherin. We show that murine fibrosarcoma tumor cells that secrete vesicle-bound antigen grow slower than tumors that secrete soluble antigen in immunocompetent, but not in immunodeficient, host mice. This growth difference is due to the induction of a more potent antigen-specific antitumor immune response in vivo by the vesicle-bound than by the soluble antigen. Finally, in vivo secretion of the vesicle-bound antigen either by tumors or by vaccination with naked DNA protects against soluble antigen-secreting tumors. We conclude that the mode of secretion can determine the immunogenicity of tumor antigens and that manipulation of the mode of antigen secretion may be used to optimize antitumor vaccination protocols.

  9. Measurements and simulations of the BLM response to a radiation field inside the CERF target area

    CERN Document Server

    Lebbos, E; Dehning, B; Effinger, E; Ferrari, A; Kramer, D; Nordt, A; Roeed, K; Roesler, S; Sapinski, M; Vlachoudis, V

    2010-01-01

    The CERN-EU high-energy reference field (CERF) facility is installed in one of the secondary beam lines (H6) of the Super Proton Synchrotron (SPS), in the North Experimental Area at CERN. This facility is used as a reference for testing, inter-comparing and calibrating passive and active instruments. In May 2009, the SPS provided a mixed hadron beam (protons, pions and kaons) during a few days, in order to perform several measurements with different devices such as the Radiation Protection Monitor used for residual dose rates due to Induced Radioactivity in the LHC (PMI), the Secondary Emission Monitor used for high beam losses (SEM), the Radiation Monitor for electronics (RadMon), and the Beam Loss Monitor for the LHC (BLM). This report focuses on the measurements of the BLM response during this year’s operation at CERF. The measurements evaluate the sensitivity of the BLM signal to the particle energy spectrum, with special attention to the contribution coming from thermal neutrons. For this purpose, meas...

  10. Using Complementary and Alternative Medicines to Target the Host Response during Severe Influenza

    Directory of Open Access Journals (Sweden)

    Lisa M. Alleva

    2010-01-01

    Full Text Available It is now accepted that an overwhelming inflammatory response is the cause of human deaths from avian H5N1 influenza infection. With this in mind we sought to examine the literature for examples of complementary and alternative medicines that reduce inflammation, and to place the results of this search in the context of our own work in a mouse model of influenza disease, using a pharmaceutical agent with anti-inflammatory properties. Two Chinese herbs, Angelica sinensis (Dang Gui and Salvia miltiorrhiza (Danshen, have been recently shown to protect mice during lethal experimental sepsis via inhibition of the novel inflammatory cytokine High Mobility Group Box 1 protein (HMGB1. Biochanin A, a ligand of the peroxisome proliferator activated receptors (PPAR alpha and gamma and the active isoflavone in Trifolium pratense (red clover, has anti-inflammatory properties, and thus could be used as an influenza treatment. This is of great interest since we have recently shown that gemfibrozil, a drug used to treat hyperlipidemia in humans and a synthetic ligand of PPAR alpha, significantly reduces the mortality associated with influenza infections in mice. The inflammation-modulating abilities of these natural agents should be considered in light of what is now known about the mechanisms of fatal influenza, and tested as potential candidates for influenza treatments in their own right, or as adjunct treatments to antivirals.

  11. Hirota's solitons in the affine and the conformal affine Toda models

    International Nuclear Information System (INIS)

    Aratyn, H.; Constantinidis, C.P.; Ferreira, L.A.; Gomes, J.F.; Zimerman, A.H.

    1993-01-01

    We use Hirota's method formulated as a recursive scheme to construct a complete set of soliton solutions for the affine Toda field theory based on an arbitrary Lie algebra. Our solutions include a new class of solitons connected with two different types of degeneracies encountered in Hirota's perturbation approach. We also derive an universal mass formula for all Hirota's solutions to the affine Toda model valid for all underlying Lie groups. Embedding of the affine Toda model in the conformal affine Toda model plays a crucial role in this analysis. (orig.)

  12. Vaccination targeting human HER3 alters the phenotype of infiltrating T cells and responses to immune checkpoint inhibition.

    Science.gov (United States)

    Osada, Takuya; Morse, Michael A; Hobeika, Amy; Diniz, Marcio A; Gwin, William R; Hartman, Zachary; Wei, Junping; Guo, Hongtao; Yang, Xiao-Yi; Liu, Cong-Xiao; Kaneko, Kensuke; Broadwater, Gloria; Lyerly, H Kim

    2017-01-01

    Expression of human epidermal growth factor family member 3 (HER3), a critical heterodimerization partner with EGFR and HER2, promotes more aggressive biology in breast and other epithelial malignancies. As such, inhibiting HER3 could have broad applicability to the treatment of EGFR- and HER2-driven tumors. Although lack of a functional kinase domain limits the use of receptor tyrosine kinase inhibitors, HER3 contains antigenic targets for T cells and antibodies. Using novel human HER3 transgenic mouse models of breast cancer, we demonstrate that immunization with recombinant adenoviral vectors encoding full length human HER3 (Ad-HER3-FL) induces HER3-specific T cells and antibodies, alters the T cell infiltrate in tumors, and influences responses to immune checkpoint inhibitions. Both preventative and therapeutic Ad-HER3-FL immunization delayed tumor growth but were associated with both intratumoral PD-1 expressing CD8 + T cells and regulatory CD4 + T cell infiltrates. Immune checkpoint inhibition with either anti-PD-1 or anti-PD-L1 antibodies increased intratumoral CD8 + T cell infiltration and eliminated tumor following preventive vaccination with Ad-HER3-FL vaccine. The combination of dual PD-1/PD-L1 and CTLA4 blockade slowed the growth of tumor in response to Ad-HER3-FL in the therapeutic model. We conclude that HER3-targeting vaccines activate HER3-specific T cells and induce anti-HER3 specific antibodies, which alters the intratumoral T cell infiltrate and responses to immune checkpoint inhibition.

  13. Mechanical and IL-1β Responsive miR-365 Contributes to Osteoarthritis Development by Targeting Histone Deacetylase 4

    Directory of Open Access Journals (Sweden)

    Xu Yang

    2016-03-01

    Full Text Available Mechanical stress plays an important role in the initiation and progression of osteoarthritis. Studies show that excessive mechanical stress can directly damage the cartilage extracellular matrix and shift the balance in chondrocytes to favor catabolic activity over anabolism. However, the underlying mechanism remains unknown. MicroRNAs (miRNAs are emerging as important regulators in osteoarthritis pathogenesis. We have found that mechanical loading up-regulated microRNA miR-365 in growth plate chondrocytes, which promotes chondrocyte differentiation. Here, we explored the role of the mechanical responsive microRNA miR-365 in pathogenesis of osteoarthritis (OA. We found that miR-365 was up-regulated by cyclic loading and IL-1β stimulation in articular chondrocytes through a mechanism that involved the transcription factor NF-κB. miR-365 expressed significant higher level in rat anterior cruciate ligament (ACL surgery induced OA cartilage as well as human OA cartilage from primary OA patients and traumatic OA Patients. Overexpression of miR-365 in chondrocytes increases gene expression of matrix degrading enzyme matrix metallopeptidase 13 (MMP13 and collagen type X (Col X. The increase in miR-365 expression in OA cartilage and in response to IL-1 may contribute to the abnormal gene expression pattern characteristic of OA. Inhibition of miR-365 down-regulated IL-1β induced MMP13 and Col X gene expression. We further showed histone deacetylase 4 (HDAC4 is a direct target of miR-365, which mediates mechanical stress and inflammation in OA pathogenesis. Thus, miR-365 is a critical regulator of mechanical stress and pro-inflammatory responses, which contributes cartilage catabolism. Manipulation of the expression of miR-365 in articular chondrocytes by miR-365 inhibitor may be a potent therapeutic target for the prevention and treatment of osteoarthritis.

  14. Opinions of patients with type 2 diabetes about responsibility, setting targets and willingness to take medication: a cross-sectional survey.

    NARCIS (Netherlands)

    Gorter, K.J.; Tuytel, G.J.; Leeuw, R.R. de; Bensing, J.M.; Rutten, G.E.H.M.

    2011-01-01

    OBJECTIVE: To assess opinions and their determinants of patients with type 2 diabetes about responsibility for managing their diabetes, setting treatment targets and willingness taking medication. METHODS: Questionnaire survey carried out in general practices and outpatient clinics across the

  15. The fourth dimension in immunological space: how the struggle for nutrients selects high-affinity lymphocytes.

    Science.gov (United States)

    Wensveen, Felix M; van Gisbergen, Klaas P J M; Eldering, Eric

    2012-09-01

    Lymphocyte activation via the antigen receptor is associated with radical shifts in metabolism and changes in requirements for nutrients and cytokines. Concomitantly, drastic changes occur in the expression of pro-and anti-apoptotic proteins that alter the sensitivity of lymphocytes to limiting concentrations of key survival factors. Antigen affinity is a primary determinant for the capacity of activated lymphocytes to access these vital resources. The shift in metabolic needs and the variable access to key survival factors is used by the immune system to eliminate activated low-affinity cells and to generate an optimal high-affinity response. In this review, we focus on the control of apoptosis regulators in activated lymphocytes by nutrients, cytokines, and costimulation. We propose that the struggle among individual clones that leads to the formation of high-affinity effector cell populations is in effect an 'invisible' fourth signal required for effective immune responses. © 2012 John Wiley & Sons A/S.

  16. Inhibition of glutathione synthesis eliminates the adaptive response of ascitic hepatoma 22 cells to nedaplatin that targets thioredoxin reductase

    International Nuclear Information System (INIS)

    Wang, Yijun; Lu, Hongjuan; Wang, Dongxu; Li, Shengrong; Sun, Kang; Wan, Xiaochun; Taylor, Ethan Will; Zhang, Jinsong

    2012-01-01

    Thioredoxin reductase (TrxR) is a target for cancer therapy and the anticancer mechanism of cisplatin involves TrxR inhibition. We hypothesize that the anticancer drug nedaplatin (NDP), an analogue of cisplatin and a second-generation platinum complex, also targets TrxR. Furthermore, we investigate whether the therapeutic efficacy of NDP can be enhanced by simultaneous modulation of 1) TrxR, via NDP, and 2) glutathione (GSH), via the GSH synthesis inhibitor buthionine sulfoximine (BSO). Mice bearing ascitic hepatoma 22 (H22) cells were treated with NDP alone or NDP plus BSO. TrxR activity of H22 cells was inhibited by NDP in a dose-dependent manner. A high correlation between the inhibition of TrxR activity at 6 h and the inhibition of ascitic fluid volume at 72 h was established (r = 0.978, p < 0.01). As an adaptive response, the viable ascitic cancer cells after NDP treatment displayed an enlarged cell phenotype, assembled with several-fold more antioxidant enzymes and GSH-predominant non-protein free thiols. This adaptive response was largely eliminated when BSO was co-administered with NDP, leading to the decimation of the H22 cell population without enhancing renal toxicity, since at this dose, NDP did not inhibit renal TrxR activity. In conclusion, the pharmacological effect of NDP involves TrxR inhibition, and the adaptive response of NDP-treated ascitic H22 cells can be efficiently counteracted by BSO. Simultaneous modulation of TrxR and GSH on ascitic H22 cells using NDP plus BSO greatly enhances therapeutic efficacy as compared with the single modulation of TrxR using NDP alone. -- Highlights: ► Nedaplatin at a pharmacological dose inhibits TrxR in cancer cells but not in kidney. ► The nedaplatin-treated cancer cells exhibit adaptive response. ► Buthionine sulfoximine inhibits glutathione in both cancer cells and kidney. ► Buthionine sulfoximine counteracts the adaptive response to the nedaplatin treatment. ► Buthionine sulfoximine does not

  17. Inhibition of glutathione synthesis eliminates the adaptive response of ascitic hepatoma 22 cells to nedaplatin that targets thioredoxin reductase

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yijun [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China); Lu, Hongjuan [Productivity Center of Jiangsu Province, Nanjing 210042, Jiangsu (China); Wang, Dongxu; Li, Shengrong; Sun, Kang; Wan, Xiaochun [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China); Taylor, Ethan Will [Department of Nanoscience, Joint School of Nanoscience and Nanoengineering, University of North Carolina at Greensboro, Greensboro, NC 27402 (United States); Zhang, Jinsong, E-mail: zjs@ahau.edu.cn [School of Tea and Food Science, Anhui Agricultural University, Hefei 230036, Anhui (China)

    2012-12-15

    Thioredoxin reductase (TrxR) is a target for cancer therapy and the anticancer mechanism of cisplatin involves TrxR inhibition. We hypothesize that the anticancer drug nedaplatin (NDP), an analogue of cisplatin and a second-generation platinum complex, also targets TrxR. Furthermore, we investigate whether the therapeutic efficacy of NDP can be enhanced by simultaneous modulation of 1) TrxR, via NDP, and 2) glutathione (GSH), via the GSH synthesis inhibitor buthionine sulfoximine (BSO). Mice bearing ascitic hepatoma 22 (H22) cells were treated with NDP alone or NDP plus BSO. TrxR activity of H22 cells was inhibited by NDP in a dose-dependent manner. A high correlation between the inhibition of TrxR activity at 6 h and the inhibition of ascitic fluid volume at 72 h was established (r = 0.978, p < 0.01). As an adaptive response, the viable ascitic cancer cells after NDP treatment displayed an enlarged cell phenotype, assembled with several-fold more antioxidant enzymes and GSH-predominant non-protein free thiols. This adaptive response was largely eliminated when BSO was co-administered with NDP, leading to the decimation of the H22 cell population without enhancing renal toxicity, since at this dose, NDP did not inhibit renal TrxR activity. In conclusion, the pharmacological effect of NDP involves TrxR inhibition, and the adaptive response of NDP-treated ascitic H22 cells can be efficiently counteracted by BSO. Simultaneous modulation of TrxR and GSH on ascitic H22 cells using NDP plus BSO greatly enhances therapeutic efficacy as compared with the single modulation of TrxR using NDP alone. -- Highlights: ► Nedaplatin at a pharmacological dose inhibits TrxR in cancer cells but not in kidney. ► The nedaplatin-treated cancer cells exhibit adaptive response. ► Buthionine sulfoximine inhibits glutathione in both cancer cells and kidney. ► Buthionine sulfoximine counteracts the adaptive response to the nedaplatin treatment. ► Buthionine sulfoximine does not

  18. FBXW7 regulates glucocorticoid response in T-cell acute lymphoblastic leukaemia by targeting the glucocorticoid receptor for degradation.

    Science.gov (United States)

    Malyukova, A; Brown, S; Papa, R; O'Brien, R; Giles, J; Trahair, T N; Dalla Pozza, L; Sutton, R; Liu, T; Haber, M; Norris, M D; Lock, R B; Sangfelt, O; Marshall, G M

    2013-04-01

    Loss of function mutation in FBXW7, an E3 ubiquitin ligase, is associated with good prognosis and early glucocorticoid treatment response in childhood T-cell acute lymphoblastic leukemia (T-ALL) by unknown mechanisms. Here, we show that FBXW7 targets the glucocorticoid receptor α (GRα) for ubiquitylation and proteasomal degradation in a manner dependent on glycogen synthase kinase 3 β-mediated phsophorylation. FBXW7 inactivation caused elevated GRα levels, and enhanced the transcriptional response to glucocorticoids. There was significant enhancement of GR transcriptional responses in FBXW7-deficient cell lines and primary T-ALL samples, in particular, for those pro-apoptotic regulatory proteins, BIM and PUMA. Reduced FBXW7 expression or function promoted glucocorticoid sensitivity, but not sensitivity to other chemotherapeutic agents used in T-ALL. Moreover, this was a general feature of different cancer cell types. Taken together, our work defines GRα as a novel FBXW7 substrate and demonstrates that favorable patient prognosis in T-ALL is associated with FBXW7 mutations due to enhanced GRα levels and steroid sensitivity. These findings suggest that inactivation of FBXW7, a putative tumor suppressor protein, may create a synthetic lethal state in the presence of specific anticancer therapies.

  19. Target or barrier? The cell wall of early- and later- diverging plants vs cadmium toxicity: differences in the response mechanisms

    Directory of Open Access Journals (Sweden)

    Luigi eParrotta

    2015-03-01

    Full Text Available Increasing industrialization and urbanization result in emission of pollutants in the environment including toxic heavy metals, as cadmium and lead. Among the different heavy metals contaminating the environment, cadmium raises great concern, as it is ecotoxic and as such can heavily impact ecosystems. The cell wall is the first structure of plant cells to come in contact with heavy metals. Its composition, characterized by proteins, polysaccharides and in some instances lignin and other phenolic compounds, confers the ability to bind non-covalently and/or covalently heavy metals via functional groups. A strong body of evidence in the literature has shown the role of the cell wall in heavy metal response: it sequesters heavy metals, but at the same time its synthesis and composition can be severely affected. The present review analyzes the dual property of plant cell walls, i.e. barrier and target of heavy metals, by taking Cd toxicity as example. Following a summary of the known physiological and biochemical responses of plants to Cd, the review compares the wall-related mechanisms in early- and later-diverging land plants, by considering the diversity in cell wall composition. By doing so, common as well as unique response mechanisms to metal/cadmium toxicity are identified among plant phyla and discussed. After discussing the role of hyperaccumulators’ cell walls as a particular case, the review concludes by considering important aspects for plant engineering.

  20. Metabolomics reveals metabolic targets and biphasic responses in breast cancer cells treated by curcumin alone and in association with docetaxel.

    Directory of Open Access Journals (Sweden)

    Mathilde Bayet-Robert

    Full Text Available BACKGROUND: Curcumin (CUR has deserved extensive research due to its anti-inflammatory properties, of interest in human diseases including cancer. However, pleiotropic even paradoxical responses of tumor cells have been reported, and the mechanisms of action of CUR remain uncompletely elucidated. METHODOLOGY/PRINCIPAL FINDINGS: (1H-NMR spectroscopy-based metabolomics was applied to get novel insight into responses of MCF7 and MDA-MB-231 breast cancer cells to CUR alone, and MCF7 cells to CUR in cotreatment with docetaxel (DTX. In both cell types, a major target of CUR was glutathione metabolism. Total glutathione (GSx increased at low dose CUR (≤ 10 mg.l(-1-28 µM- (up to +121% in MCF7 cells, P<0.01, and +138% in MDA-MB-231 cells, P<0.01, but decreased at high dose (≥ 25 mg.l(-1 -70 µM- (-49%, in MCF7 cells, P<0.02, and -56% in MDA-MB-231 cells, P<0.025. At high dose, in both cell types, GSx-related metabolites decreased, including homocystein, creatine and taurine (-60 to -80%, all, P<0.05. Together with glutathione-S-transferase actvity, data established that GSx biosynthesis was upregulated at low dose, and GSx consumption activated at high dose. Another major target, in both cell types, was lipid metabolism involving, at high doses, accumulation of polyunsaturated and total free fatty acids (between ×4.5 and ×11, P<0.025, and decrease of glycerophospho-ethanolamine and -choline (about -60%, P<0.025. Multivariate statistical analyses showed a metabolic transition, even a biphasic behavior of some metabolites including GSx, between low and high doses. In addition, CUR at 10 mg.l(-1 in cotreatment with DTX induced modifications in glutathione metabolism, lipid metabolism, and glucose utilization. Some of these changes were biphasic depending on the duration of exposure to CUR. CONCLUSIONS/SIGNIFICANCE: Metabolomics reveals major metabolic targets of CUR in breast cancer cells, and biphasic responses that challenge the widely accepted

  1. Conformal treatment of prostate cancer with improved targeting: superior prostate-specific antigen response compared to standard treatment

    Energy Technology Data Exchange (ETDEWEB)

    Corn, Benjamin W; Hanks, Gerald E; Schultheiss, Timothy E; Hunt, Margie A; Lee, W Robert; Coia, Lawrence R

    1995-05-15

    Purpose: Conformal radiation therapy (CRT) decreases the morbidity of prostate cancer treatment, but no published data attest to the improved ability of CRT to control disease. Therefore, we compared Prostate-Specific Antigen (PSA) response at 1 year among similarly staged patients treated by conformal techniques to those treated with conventional approaches, looking for an early indicator of tumor response. Method and Materials: Patients with locally advanced disease were treated by pelvic fields followed by prostate field conedowns; those with early stage/low grade disease received only prostate field irradiation. Between October, 1987 and November, 1991, conventional treatments used rectangular beams with or without corner blocks. Neither urethrography nor immobilization casts were used for conventionally treated patients. Between April, 1989 and December, 1992, conformal treatments have used rigid immobilization and Computed Tomography-based, beams-eye-view field design. As such, our conformal approach allowed improved targeting. Median prescribed doses (minimal doses to the Planning Target Volume) were 70 Gy (66-73 Gy) and 70.2 Gy (64.8-75 Gy) for conventionally and conformally treated patients, respectively. Median daily fraction size was 1.8 Gy for conventional treatment and 2.0 Gy for conformal therapy. Baseline PSA data were available on 170 consecutive patients treated conformally and 90 consecutive patients treated conventionally. Results: Among those receiving only prostatic field irradiation, 12-month PSA values returned to normal in 96% and 85% of conformally and conventionally treated patients, respectively, when normalization was defined as {<=} 4 ng/ml (p < 0.03) and in 76% vs. 55% of patients when PSA normalization was defined as {<=} 1.5 ng/ml (p < 0.02). Among those receiving pelvic irradiation prior to prostatic conedown, PSA normalization ({<=} 4 ng/ml) occurred in 82% and 61% (p < 0.01) of conformally and conventionally treated patients

  2. Predicting the dynamics of bacterial growth inhibition by ribosome-targeting antibiotics

    Science.gov (United States)

    Greulich, Philip; Doležal, Jakub; Scott, Matthew; Evans, Martin R.; Allen, Rosalind J.

    2017-12-01

    Understanding how antibiotics inhibit bacteria can help to reduce antibiotic use and hence avoid antimicrobial resistance—yet few theoretical models exist for bacterial growth inhibition by a clinically relevant antibiotic treatment regimen. In particular, in the clinic, antibiotic treatment is time-dependent. Here, we use a theoretical model, previously applied to steady-state bacterial growth, to predict the dynamical response of a bacterial cell to a time-dependent dose of ribosome-targeting antibiotic. Our results depend strongly on whether the antibiotic shows reversible transport and/or low-affinity ribosome binding (‘low-affinity antibiotic’) or, in contrast, irreversible transport and/or high affinity ribosome binding (‘high-affinity antibiotic’). For low-affinity antibiotics, our model predicts that growth inhibition depends on the duration of the antibiotic pulse, and can show a transient period of very fast growth following removal of the antibiotic. For high-affinity antibiotics, growth inhibition depends on peak dosage rather than dose duration, and the model predicts a pronounced post-antibiotic effect, due to hysteresis, in which growth can be suppressed for long times after the antibiotic dose has ended. These predictions are experimentally testable and may be of clinical significance.

  3. Targeted Therapies for Myeloma and Metastatic Bone Cancers

    National Research Council Canada - National Science Library

    Vail, Neal

    2008-01-01

    ... done. Quantified functional groups available for ligand conjugation using S35-labeled ligands. Developed alternative assay to confirm affinity of bone-targeting nanoparticles to hydroxyapatite substrates...

  4. Rational design of peptide affinity ligands for the purification of therapeutic enzymes.

    Science.gov (United States)

    Trasatti, John P; Woo, James; Ladiwala, Asif; Cramer, Steven; Karande, Pankaj

    2018-04-25

    Non-mAb biologics represent a growing class of therapeutics under clinical development. Although affinity chromatography is a potentially attractive approach for purification, the development of platform technologies, such as Protein A for mAbs, has been challenging due to the inherent chemical and structural diversity of these molecules. Here, we present our studies on the rapid development of peptide affinity ligands for the purification of biologics using a prototypical enzyme therapeutic in clinical use. Employing a suite of de novo rational and combinatorial design strategies we designed and screened a library of peptides on microarray platforms for their ability to bind to the target with high affinity and selectivity in cell culture fluid. Lead peptides were evaluated on resin in batch conditions and compared with a commercially available resin to evaluate their efficacy. Two lead candidates identified from microarray studies provided high binding capacity to the target while demonstrating high selectivity against culture contaminants and product variants compared to a commercial resin system. These findings provide a proof-of-concept for developing affinity peptide-based bioseparations processes for a target biologic. Peptide affinity ligand design and screening approaches presented in this work can also be easily translated to other biologics of interest. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 2018. © 2018 American Institute of Chemical Engineers.

  5. Role of T cell receptor affinity in the efficacy and specificity of adoptive T cell therapies

    Directory of Open Access Journals (Sweden)

    Jennifer D. Stone

    2013-08-01

    Full Text Available Over the last several years, there has been considerable progress in the treatment of cancer using gene modified adoptive T cell therapies. Two approaches have been used, one involving the introduction of a conventional alpha-beta T cell receptor (TCR against a pepMHC cancer antigen, and the second involving introduction of a chimeric antigen receptor (CAR consisting of a single-chain antibody as an Fv fragment (scFv linked to transmembrane and signaling domains. In this review, we focus on one aspect of TCR-mediated adoptive T cell therapies, the impact of the affinity of the alpha-beta TCR for the pepMHC cancer antigen on both efficacy and specificity. We discuss the advantages of higher affinity TCRs in mediating potent activity of CD4 T cells. This is balanced with the potential disadvantage of higher affinity TCRs in mediating greater self-reactivity against a wider range of structurally similar antigenic peptides, especially in synergy with the CD8 co-receptor. Both TCR affinity and target selection will influence potential safety issues. We suggest pre-clinical strategies that might be used to examine each TCR for possible on-target and off-target side effects due to self-reactivities, and to adjust TCR affinities accordingly.

  6. Transient Expression of CRISPR/Cas9 Machinery Targeting TcNPR3 Enhances Defense Response in Theobroma cacao.

    Science.gov (United States)

    Fister, Andrew S; Landherr, Lena; Maximova, Siela N; Guiltinan, Mark J

    2018-01-01

    Theobroma cacao , the source of cocoa, suffers significant losses to a variety of pathogens resulting in reduced incomes for millions of farmers in developing countries. Development of disease resistant cacao varieties is an essential strategy to combat this threat, but is limited by sources of genetic resistance and the slow generation time of this tropical tree crop. In this study, we present the first application of genome editing technology in cacao, using Agrobacterium-mediated transient transformation to introduce CRISPR/Cas9 components into cacao leaves and cotyledon cells. As a first proof of concept, we targeted the cacao Non-Expressor of Pathogenesis-Related 3 (TcNPR3) gene, a suppressor of the defense response. After demonstrating activity of designed single-guide RNAs (sgRNA) in vitro , we used Agrobacterium to introduce a CRISPR/Cas9 system into leaf tissue, and identified the presence of deletions in 27% of TcNPR3 copies in the treated tissues. The edited tissue exhibited an increased resistance to infection with the cacao pathogen Phytophthora tropicalis and elevated expression of downstream defense genes. Analysis of off-target mutagenesis in sequences similar to sgRNA target sites using high-throughput sequencing did not reveal mutations above background sequencing error rates. These results confirm the function of NPR3 as a repressor of the cacao immune system and demonstrate the application of CRISPR/Cas9 as a powerful functional genomics tool for cacao. Several stably transformed and genome edited somatic embryos were obtained via Agrobacterium -mediated transformation, and ongoing work will test the effectiveness of this approach at a whole plant level.

  7. Cell Connections by Tunneling Nanotubes: Effects of Mitochondrial Trafficking on Target Cell Metabolism, Homeostasis, and Response to Therapy

    Science.gov (United States)

    2017-01-01

    Intercellular communications play a major role in tissue homeostasis and responses to external cues. Novel structures for this communication have recently been described. These tunneling nanotubes (TNTs) consist of thin-extended membrane protrusions that connect cells together. TNTs allow the cell-to-cell transfer of various cellular components, including proteins, RNAs, viruses, and organelles, such as mitochondria. Mesenchymal stem cells (MSCs) are both naturally present and recruited to many different tissues where their interaction with resident cells via secreted factors has been largely documented. Their immunosuppressive and repairing capacities constitute the basis for many current clinical trials. MSCs recruited to the tumor microenvironment also play an important role in tumor progression and resistance to therapy. MSCs are now the focus of intense scrutiny due to their capacity to form TNTs and transfer mitochondria to target cells, either in normal physiological or in pathological conditions, leading to changes in cell energy metabolism and functions, as described in this review. PMID:28659978

  8. Cell Connections by Tunneling Nanotubes: Effects of Mitochondrial Trafficking on Target Cell Metabolism, Homeostasis, and Response to Therapy

    Directory of Open Access Journals (Sweden)

    Marie-Luce Vignais

    2017-01-01

    Full Text Available Intercellular communications play a major role in tissue homeostasis and responses to external cues. Novel structures for this communication have recently been described. These tunneling nanotubes (TNTs consist of thin-extended membrane protrusions that connect cells together. TNTs allow the cell-to-cell transfer of various cellular components, including proteins, RNAs, viruses, and organelles, such as mitochondria. Mesenchymal stem cells (MSCs are both naturally present and recruited to many different tissues where their interaction with resident cells via secreted factors has been largely documented. Their immunosuppressive and repairing capacities constitute the basis for many current clinical trials. MSCs recruited to the tumor microenvironment also play an important role in tumor progression and resistance to therapy. MSCs are now the focus of intense scrutiny due to their capacity to form TNTs and transfer mitochondria to target cells, either in normal physiological or in pathological conditions, leading to changes in cell energy metabolism and functions, as described in this review.

  9. Science to Practice: Killing Dormant Cells-Is Targeting Autophagy the Key to Complete Tumor Response in Transarterial Chemoembolization?

    Science.gov (United States)

    Savic, Lynn Jeanette; Chapiro, Julius; Geschwind, Jean-François

    2017-06-01

    In this issue of Radiology, Gade et al ( 1 ) describe a unique mechanism of hepatocellular carcinoma (HCC) cells for surviving ischemia induced by transarterial embolization (TAE)/transarterial chemoembolization (TACE) in a state of cell cycle arrest-a function that may serve as a defensive shield against conventional chemotherapeutic agents. This finding adds to our knowledge and establishes a previously poorly understood mechanism of chemoresistance in HCC. As the Achilles heel in terms of this process, a concurrent upregulation of autophagic flux as an adaptive response to TAE-like ischemia was found by the authors. This is a targetable mechanism that can potentially be exploited for combined therapeutic approaches of embolotherapy and autophagy inhibition in HCC.

  10. A General Strategy for Targeting Drugs to Bone.

    Science.gov (United States)

    Jahnke, Wolfgang; Bold, Guido; Marzinzik, Andreas L; Ofner, Silvio; Pellé, Xavier; Cotesta, Simona; Bourgier, Emmanuelle; Lehmann, Sylvie; Henry, Chrystelle; Hemmig, René; Stauffer, Frédéric; Hartwieg, J Constanze D; Green, Jonathan R; Rondeau, Jean-Michel

    2015-11-23

    Targeting drugs to their desired site of action can increase their safety and efficacy. Bisphosphonates are prototypical examples of drugs targeted to bone. However, bisphosphonate bone affinity is often considered too strong and cannot be significantly modulated without losing activity on the enzymatic target, farnesyl pyrophosphate synthase (FPPS). Furthermore, bisphosphonate bone affinity comes at the expense of very low and variable oral bioavailability. FPPS inhibitors were developed with a monophosphonate as a bone-affinity tag that confers moderate affinity to bone, which can furthermore be tuned to the desired level, and the relationship between structure and bone affinity was evaluated by using an NMR-based bone-binding assay. The concept of targeting drugs to bone with moderate affinity, while retaining oral bioavailability, has broad application to a variety of other bone-targeted drugs. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Pharmaceutical micelles featured with singlet oxygen-responsive cargo release and mitochondrial targeting for enhanced photodynamic therapy

    Science.gov (United States)

    Zhang, Xin; Yan, Qi; Naer Mulatihan, Di; Zhu, Jundong; Fan, Aiping; Wang, Zheng; Zhao, Yanjun

    2018-06-01

    The efficacy of nanoparticulate photodynamic therapy is often compromised by the short life time and limited diffusion radius of singlet oxygen as well as uncontrolled intracellular distribution of photosensitizer. It was hypothesized that rapid photosensitizer release upon nanoparticle internalization and its preferred accumulation in mitochondria would address the above problems. Hence, the aim of this study was to engineer a multifunctional micellar nanosystem featured with singlet oxygen-responsive cargo release and mitochondria-targeting. An imidazole-bearing amphiphilic copolymer was employed as the micelle building block to encapsulate triphenylphosphonium-pyropheophorbide a (TPP-PPa) conjugate or PPa. Upon laser irradiation, the singlet oxygen produced by TPP-PPa/PPa oxidized the imidazole moiety to produce hydrophilic urea, leading to micelle disassembly and rapid cargo release. The co-localization analysis showed that the TPP moiety significantly enhanced the photosensitizer uptake by mitochondria, improved mitochondria depolarization upon irradiation, and hence boosted the cytotoxicity in 4T1 cells. The targeting strategy also dramatically reduced the intracellular ATP concentration as a consequence of mitochondria injury. The mitochondria damage was accompanied with the activation of the apoptosis signals (caspase 3 and caspase 9), whose level was directly correlated to the apoptosis extent. The current work provides a facile and robust means to enhance the efficacy of photodynamic therapy.

  12. pH-Responsive Hyaluronic Acid-Based Mixed Micelles for the Hepatoma-Targeting Delivery of Doxorubicin

    Directory of Open Access Journals (Sweden)

    Jing-Liang Wu

    2016-03-01

    Full Text Available The tumor targetability and stimulus responsivity of drug delivery systems are crucial in cancer diagnosis and treatment. In this study, hepatoma-targeting mixed micelles composed of a hyaluronic acid–glycyrrhetinic acid conjugate and a hyaluronic acid-l-histidine conjugate (HA–GA/HA–His were prepared through ultrasonic dispersion. The formation and characterization of the mixed micelles were confirmed via 1H-NMR, particle size, and ζ potential measurements. The in vitro cellular uptake of the micelles was evaluated using human liver carcinoma (HepG2 cells. The antitumor effect of doxorubicin (DOX-loaded micelles was investigated in vitro and in vivo. Results indicated that the DOX-loaded HA–GA/HA–His micelles showed a pH-dependent controlled release and were remarkably absorbed by HepG2 cells. Compared with free DOX, the DOX-loaded HA–GA/HA–His micelles showed a higher cytotoxicity to HepG2 cells. Moreover, the micelles effectively inhibited tumor growth in H22 cell-bearing mice. These results suggest that the HA–GA/HA–His mixed micelles are a good candidate for drug delivery in the prevention and treatment of hepatocarcinoma.

  13. Analyses of Potential Predictive Markers and Response to Targeted Therapy in Patients with Advanced Clear-cell Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Yan Song

    2015-01-01

    Full Text Available Background: Vascular endothelial growth factor-targeted agents are standard treatments in advanced clear-cell renal cell carcinoma (ccRCC, but biomarkers of activity are lacking. The aim of this study was to investigate the association of Von Hippel-Lindau (VHL gene status, vascular endothelial growth factor receptor (VEGFR or stem cell factor receptor (KIT expression, and their relationships with characteristics and clinical outcome of advanced ccRCC. Methods: A total of 59 patients who received targeted treatment with sunitinib or pazopanib were evaluated for determination at Cancer Hospital and Institute, Chinese Academy of Medical Sciences between January 2010 and November 2012. Paraffin-embedded tumor samples were collected and status of the VHL gene and expression of VEGFR and KIT were determined by VHL sequence analysis and immunohistochemistry. Clinical-pathological features were collected and efficacy such as response rate and Median progression-free survival (PFS and overall survival (OS were calculated and then compared based on expression status. The Chi-square test, the Kaplan-Meier method, and the Lon-rank test were used for statistical analyses. Results: Of 59 patients, objective responses were observed in 28 patients (47.5%. The median PFS was 13.8 months and median OS was 39.9 months. There was an improved PFS in patients with the following clinical features: Male gender, number of metastatic sites 2 or less, VEGFR-2 positive or KIT positive. Eleven patients (18.6% had evidence of VHL mutation, with an objective response rate of 45.5%, which showed no difference with patients with no VHL mutation (47.9%. VHL mutation status did not correlate with either overall response rate (P = 0.938 or PFS (P = 0.277. The PFS was 17.6 months and 22.2 months in VEGFR-2 positive patients and KIT positive patients, respectively, which was significantly longer than that of VEGFR-2 or KIT negative patients (P = 0.026 and P = 0.043. Conclusion

  14. Current advances in screening for bioactive components from medicinal plants by affinity ultrafiltration mass spectrometry.

    Science.gov (United States)

    Chen, Guilin; Huang, Bill X; Guo, Mingquan

    2018-05-21

    Medicinal plants have played an important role in maintaining human health for thousands of years. However, the interactions between the active components in medicinal plants and some certain biological targets during a disease are still unclear in most cases. To conduct the high-throughput screening for small active molecules that can interact with biological targets, which is of great theoretical significance and practical value. The ultrafiltration mass spectrometry (UF-LC/MS) is a powerful bio-analytical method by combining affinity ultrafiltration and liquid chromatography-mass spectrometry (LC/MS), which could rapidly screen and identify small active molecules that bind to biological targets of interest at the same time. Compared with other analytical methods, affinity UF-LC/MS has the characteristics of fast, sensitive and high throughput, and is especially suitable for the complicated extracts of medicinal plants. In this review, the basic principle, characteristics and some most recent challenges in UF-LC/MS have been demonstrated. Meanwhile, the progress and applications of affinity UF-LC/MS in the discovery of the active components from natural medicinal plants and the interactions between small molecules and biological target proteins are also briefly summarised. In addition, the future directions for UF-LC/MS are also prospected. Affinity UF-LC/MS is a powerful tool in studies on the interactions between small active molecules and biological protein targets, especially in the high-throughput screening of active components from the natural medicinal plants. Copyright © 2018 John Wiley & Sons, Ltd.

  15. Engineering an antibody with picomolar affinity to DOTA chelates of multiple radionuclides for pretargeted radioimmunotherapy and imaging

    Energy Technology Data Exchange (ETDEWEB)

    Orcutt, Kelly Davis; Slusarczyk, Adrian L. [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Cieslewicz, Maryelise [Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Ruiz-Yi, Benjamin [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Bhushan, Kumar R. [Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Frangioni, John V. [Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Department of Radiology, Beth Israel Deaconess Medical Center, Boston, MA 02215 (United States); Wittrup, K. Dane, E-mail: wittrup@mit.ed [Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States); Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139 (United States)

    2011-02-15

    Introduction: In pretargeted radioimmunotherapy (PRIT), a bifunctional antibody is administered and allowed to pre-localize to tumor cells. Subsequently, a chelated radionuclide is administered and captured by cell-bound antibody while unbound hapten clears rapidly from the body. We aim to engineer high-affinity binders to 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelates for use in PRIT applications. Methods: We mathematically modeled antibody and hapten pharmacokinetics to analyze hapten tumor retention as a function of hapten binding affinity. Motivated by model predictions, we used directed evolution and yeast surface display to affinity mature the 2D12.5 antibody to DOTA, reformatted as a single chain variable fragment (scFv). Results: Modeling predicts that for high antigen density and saturating bsAb dose, a hapten-binding affinity of 100 pM is needed for near-maximal hapten retention. We affinity matured 2D12.5 with an initial binding constant of about 10 nM to DOTA-yttrium chelates. Affinity maturation resulted in a 1000-fold affinity improvement to biotinylated DOTA-yttrium, yielding an 8.2{+-}1.9 picomolar binder. The high-affinity scFv binds DOTA complexes of lutetium and gadolinium with similar picomolar affinity and indium chelates with low nanomolar affinity. When engineered into a bispecific antibody construct targeting carcinoembryonic antigen, pretargeted high-affinity scFv results in significantly higher tumor retention of a {sup 111}In-DOTA hapten compared to pretargeted wild-type scFv in a xenograft mouse model. Conclusions: We have engineered a versatile, high-affinity, DOTA-chelate-binding scFv. We anticipate it will prove useful in developing pretargeted imaging and therapy protocols to exploit the potential of a variety of radiometals.

  16. On Affine Fusion and the Phase Model

    Directory of Open Access Journals (Sweden)

    Mark A. Walton

    2012-11-01

    Full Text Available A brief review is given of the integrable realization of affine fusion discovered recently by Korff and Stroppel. They showed that the affine fusion of the su(n Wess-Zumino-Novikov-Witten (WZNW conformal field theories appears in a simple integrable system known as the phase model. The Yang-Baxter equation leads to the construction of commuting operators as Schur polynomials, with noncommuting hopping operators as arguments. The algebraic Bethe ansatz diagonalizes them, revealing a connection to the modular S matrix and fusion of the su(n WZNW model. The noncommutative Schur polynomials play roles similar to those of the primary field operators in the corresponding WZNW model. In particular, their 3-point functions are the su(n fusion multiplicities. We show here how the new phase model realization of affine fusion makes obvious the existence of threshold levels, and how it accommodates higher-genus fusion.

  17. Affine coherent states and Toeplitz operators

    Science.gov (United States)

    Hutníková, Mária; Hutník, Ondrej

    2012-06-01

    We study a parameterized family of Toeplitz operators in the context of affine coherent states based on the Calderón reproducing formula (= resolution of unity on L_2( {R})) and the specific admissible wavelets (= affine coherent states in L_2( {R})) related to Laguerre functions. Symbols of such Calderón-Toeplitz operators as individual coordinates of the affine group (= upper half-plane with the hyperbolic geometry) are considered. In this case, a certain class of pseudo-differential operators, their properties and their operator algebras are investigated. As a result of this study, the Fredholm symbol algebras of the Calderón-Toeplitz operator algebras for these particular cases of symbols are described. This article is part of a special issue of Journal of Physics A: Mathematical and Theoretical devoted to ‘Coherent states: mathematical and physical aspects’.

  18. Differential affinity of mammalian histone H1 somatic subtypes for DNA and chromatin

    Directory of Open Access Journals (Sweden)

    Mora Xavier

    2007-05-01

    chromatin suggest that differential affinity could be functionally relevant and thus contribute to the functional differentiation of the subtypes. The conservation of the relative affinities for SAR and non-SAR DNA, in spite of a strong preference for SAR sequences, indicates that differential affinity alone cannot be responsible for the heterogeneous distribution of some subtypes in cell nuclei.

  19. The dynamics of metric-affine gravity

    International Nuclear Information System (INIS)

    Vitagliano, Vincenzo; Sotiriou, Thomas P.; Liberati, Stefano

    2011-01-01

    Highlights: → The role and the dynamics of the connection in metric-affine theories is explored. → The most general second order action does not lead to a dynamical connection. → Including higher order invariants excites new degrees of freedom in the connection. → f(R) actions are also discussed and shown to be a non- representative class. - Abstract: Metric-affine theories of gravity provide an interesting alternative to general relativity: in such an approach, the metric and the affine (not necessarily symmetric) connection are independent quantities. Furthermore, the action should include covariant derivatives of the matter fields, with the covariant derivative naturally defined using the independent connection. As a result, in metric-affine theories a direct coupling involving matter and connection is also present. The role and the dynamics of the connection in such theories is explored. We employ power counting in order to construct the action and search for the minimal requirements it should satisfy for the connection to be dynamical. We find that for the most general action containing lower order invariants of the curvature and the torsion the independent connection does not carry any dynamics. It actually reduces to the role of an auxiliary field and can be completely eliminated algebraically in favour of the metric and the matter field, introducing extra interactions with respect to general relativity. However, we also show that including higher order terms in the action radically changes this picture and excites new degrees of freedom in the connection, making it (or parts of it) dynamical. Constructing actions that constitute exceptions to this rule requires significant fine tuned and/or extra a priori constraints on the connection. We also consider f(R) actions as a particular example in order to show that they constitute a distinct class of metric-affine theories with special properties, and as such they cannot be used as representative toy

  20. Phosphopeptide enrichment by immobilized metal affinity chromatography

    DEFF Research Database (Denmark)

    Thingholm, Tine E.; Larsen, Martin R.

    2016-01-01

    Immobilized metal affinity chromatography (IMAC) has been the method of choice for phosphopeptide enrichment prior to mass spectrometric analysis for many years and it is still used extensively in many laboratories. Using the affinity of negatively charged phosphate groups towards positively...... charged metal ions such as Fe3+, Ga3+, Al3+, Zr4+, and Ti4+ has made it possible to enrich phosphorylated peptides from peptide samples. However, the selectivity of most of the metal ions is limited, when working with highly complex samples, e.g., whole-cell extracts, resulting in contamination from...

  1. Control and estimation of piecewise affine systems

    CERN Document Server

    Xu, Jun

    2014-01-01

    As a powerful tool to study nonlinear systems and hybrid systems, piecewise affine (PWA) systems have been widely applied to mechanical systems. Control and Estimation of Piecewise Affine Systems presents several research findings relating to the control and estimation of PWA systems in one unified view. Chapters in this title discuss stability results of PWA systems, using piecewise quadratic Lyapunov functions and piecewise homogeneous polynomial Lyapunov functions. Explicit necessary and sufficient conditions for the controllability and reachability of a class of PWA systems are

  2. New unitary affine-Virasoro constructions

    International Nuclear Information System (INIS)

    Halpern, M.B.; Kiritsis, E.; Obers, N.A.; Poratti, M.; Yamron, J.P.

    1990-01-01

    This paper reports on a quasi-systematic investigation of the Virasoro master equation. The space of all affine-Virasoro constructions is organized by K-conjugation into affine-Virasoro nests, and an estimate of the dimension of the space shows that most solutions await discovery. With consistent ansatze for the master equation, large classes of new unitary nests are constructed, including quadratic deformation nests with continuous conformal weights, and unitary irrational central charge nests, which may dominate unitary rational central charge on compact g

  3. Applications of Affine and Weyl geometry

    CERN Document Server

    García-Río, Eduardo; Nikcevic, Stana

    2013-01-01

    Pseudo-Riemannian geometry is, to a large extent, the study of the Levi-Civita connection, which is the unique torsion-free connection compatible with the metric structure. There are, however, other affine connections which arise in different contexts, such as conformal geometry, contact structures, Weyl structures, and almost Hermitian geometry. In this book, we reverse this point of view and instead associate an auxiliary pseudo-Riemannian structure of neutral signature to certain affine connections and use this correspondence to study both geometries. We examine Walker structures, Riemannia

  4. Defective bone formation and anabolic response to exogenous estrogen in mice with targeted disruption of endothelial nitric oxide synthase.

    Science.gov (United States)

    Armour, K E; Armour, K J; Gallagher, M E; Gödecke, A; Helfrich, M H; Reid, D M; Ralston, S H

    2001-02-01

    Nitric oxide (NO) is a pleiotropic signaling molecule that is produced by bone cells constitutively and in response to diverse stimuli such as proinflammatory cytokines, mechanical strain, and sex hormones. Endothelial nitric oxide synthase (eNOS) is the predominant NOS isoform expressed in bone, but its physiological role in regulating bone metabolism remains unclear. Here we studied various aspects of bone metabolism in female mice with targeted disruption of the eNOS gene. Mice with eNOS deficiency (eNOS KO) had reduced bone mineral density, and cortical thinning when compared with WT controls and histomorphometric analysis of bone revealed profound abnormalities of bone formation, with reduced osteoblast numbers, surfaces and mineral apposition rate. Studies in vitro showed that osteoblasts derived from eNOS KO mice had reduced rates of growth when compared with WT and were less well differentiated as reflected by lower levels of alkaline phosphatase activity. Mice with eNOS deficiency lost bone normally following ovariectomy but exhibited a significantly blunted anabolic response to high dose exogenous estrogen. We conclude that the eNOS pathway plays an essential role in regulating bone mass and bone turnover by modulating osteoblast function.

  5. Crossing Chris: Some Markerian Affinities

    Directory of Open Access Journals (Sweden)

    Adrian Martin

    2010-01-01

    -pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}

    Abstract (E: This essay creatively explores a group of artists, writers, and other special individuals whose work or life story can be described as having an intriguing affinity with the protean career of Chris Marker. Avoiding the ‘usual suspects’ (such as Godard or Sebald, it discusses gossip columnist Milt Machlin, record collector Harry Smith, painter Gianfranco Baruchello, writer-filmmaker Edgardo Cozarinsky, and several others. From this constellation, a particular view of Markerian poetics emerges, touching upon the meanings of anonymity, storytelling, history and archiving.

     

    Abstract (F: Cet essai brosse de manière créative le portrait d’un groupe d'artistes, d'écrivains et d'autres personnes particulières dont le travail ou la biographie peuvent être décrits comme montrant une étrange mais certaine connivence avec la carrière protéiforme de Chris Marker. Evitant les lieux communs (comme Godard ou Sebald, cet article trace des références moins attendues :

  6. Affinity Programs and the Real Estate Brokerage Industry

    OpenAIRE

    G Stacy Sirmans; David A. Macpherson

    2001-01-01

    This study surveys active real estate brokers obtaining information on involvement in affinity programs and referral/relocation networks. Some results regarding affinity involvement are: (a) 13% of respondents reported affinity affilliations, 75% reported no affiliations, and 12% indicated plans to become involved within the next year; (b) about half having affinity affiliations were involved with 2-4 groups; (c) affinity relationships were most often with membership organizations, corporatio...

  7. Polynomials associated with equilibria of affine Toda-Sutherland systems

    International Nuclear Information System (INIS)

    Odake, S; Sasaki, R

    2004-01-01

    An affine Toda-Sutherland system is a quasi-exactly solvable multi-particle dynamics based on an affine simple root system. It is a 'cross' between two well-known integrable multi-particle dynamics, an affine Toda molecule (exponential potential, periodic nearest-neighbour interaction) and a Sutherland system (inverse sine-square interaction). Polynomials describing the equilibrium positions of affine Toda-Sutherland systems are determined for all affine simple root systems

  8. High throughput deep degradome sequencing reveals microRNAs and their targets in response to drought stress in mulberry (Morus alba).

    Science.gov (United States)

    Li, Ruixue; Chen, Dandan; Wang, Taichu; Wan, Yizhen; Li, Rongfang; Fang, Rongjun; Wang, Yuting; Hu, Fei; Zhou, Hong; Li, Long; Zhao, Weiguo

    2017-01-01

    MicroRNAs (miRNAs) play important regulatory roles by targeting mRNAs for cleavage or translational repression. Identification of miRNA targets is essential to better understanding the roles of miRNAs. miRNA targets have not been well characterized in mulberry (Morus alba). To anatomize miRNA guided gene regulation under drought stress, transcriptome-wide high throughput degradome sequencing was used in this study to directly detect drought stress responsive miRNA targets in mulberry. A drought library (DL) and a contrast library (CL) were constructed to capture the cleaved mRNAs for sequencing. In CL, 409 target genes of 30 conserved miRNA families and 990 target genes of 199 novel miRNAs were identified. In DL, 373 target genes of 30 conserved miRNA families and 950 target genes of 195 novel miRNAs were identified. Of the conserved miRNA families in DL, mno-miR156, mno-miR172, and mno-miR396 had the highest number of targets with 54, 52 and 41 transcripts, respectively, indicating that these three miRNA families and their target genes might play important functions in response to drought stress in mulberry. Additionally, we found that many of the target genes were transcription factors. By analyzing the miRNA-target molecular network, we found that the DL independent networks consisted of 838 miRNA-mRNA pairs (63.34%). The expression patterns of 11 target genes and 12 correspondent miRNAs were detected using qRT-PCR. Six miRNA targets were further verified by RNA ligase-mediated 5' rapid amplification of cDNA ends (RLM-5' RACE). Gene Ontology (GO) annotations and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that these target transcripts were implicated in a broad range of biological processes and various metabolic pathways. This is the first study to comprehensively characterize target genes and their associated miRNAs in response to drought stress by degradome sequencing in mulberry. This study provides a framework for understanding

  9. Fatty acid and drug binding to a low-affinity component of human serum albumin, purified by affinity chromatography

    DEFF Research Database (Denmark)

    Vorum, H; Pedersen, A O; Honoré, B

    1992-01-01

    Binding equilibria for decanoate to a defatted, commercially available human serum albumin preparation were investigated by dialysis exchange rate determinations. The binding isotherm could not be fitted by the general binding equation. It was necessary to assume that the preparation was a mixture...... of two albumin components about 40% of the albumin having high affinity and about 60% having low affinity. By affinity chromatography we succeeded in purifying the low-affinity component from the mixture. The high-affinity component, however, could not be isolated. We further analyzed the fatty acid...... and drug binding abilities of the low-affinity component. The fatty acids decanoate, laurate, myristate and palmitate were bound with higher affinity to the mixture than to the low-affinity component. Diazepam was bound with nearly the same affinity to the low-affinity component as to the albumin mixture...

  10. Fan Affinity Laws from a Collision Model

    Science.gov (United States)

    Bhattacharjee, Shayak

    2012-01-01

    The performance of a fan is usually estimated using hydrodynamical considerations. The calculations are long and involved and the results are expressed in terms of three affinity laws. In this paper we use kinetic theory to attack this problem. A hard sphere collision model is used, and subsequently a correction to account for the flow behaviour…

  11. General super Virasoro construction on affine G

    International Nuclear Information System (INIS)

    Mohammedi, N.

    1990-10-01

    We consider a bosonic current algebra and a theory of free fermions and construct a general N = 1 super Virasoro current algebra. We obtain a master-set of equations which comprises the bosonic master equation for general Virasoro construction on affine G. As an illustration we study the case of the group SU(2). (author). 13 refs

  12. MIR144* inhibits antimicrobial responses against Mycobacterium tuberculosis in human monocytes and macrophages by targeting the autophagy protein DRAM2.

    Science.gov (United States)

    Kim, Jin Kyung; Lee, Hye-Mi; Park, Ki-Sun; Shin, Dong-Min; Kim, Tae Sung; Kim, Yi Sak; Suh, Hyun-Woo; Kim, Soo Yeon; Kim, In Soo; Kim, Jin-Man; Son, Ji-Woong; Sohn, Kyung Mok; Jung, Sung Soo; Chung, Chaeuk; Han, Sang-Bae; Yang, Chul-Su; Jo, Eun-Kyeong

    2017-02-01

    Autophagy is an important antimicrobial effector process that defends against Mycobacterium tuberculosis (Mtb), the human pathogen causing tuberculosis (TB). MicroRNAs (miRNAs), endogenous noncoding RNAs, are involved in various biological functions and act as post-transcriptional regulators to target mRNAs. The process by which miRNAs affect antibacterial autophagy and host defense mechanisms against Mtb infections in human monocytes and macrophages is largely uncharacterized. In this study, we show that Mtb significantly induces the expression of MIR144*/hsa-miR-144-5p, which targets the 3'-untranslated region of DRAM2 (DNA damage regulated autophagy modulator 2) in human monocytes and macrophages. Mtb infection downregulated, whereas the autophagy activators upregulated, DRAM2 expression in human monocytes and macrophages by activating AMP-activated protein kinase. In addition, overexpression of MIR144* decreased DRAM2 expression and formation of autophagosomes in human monocytes, whereas inhibition of MIR144* had the opposite effect. Moreover, the levels of MIR144* were elevated, whereas DRAM2 levels were reduced, in human peripheral blood cells and tissues in TB patients, indicating the clinical significance of MIR144* and DRAM2 in human TB. Notably, DRAM2 interacted with BECN1 and UVRAG, essential components of the autophagic machinery, leading to displacement of RUBCN from the BECN1 complex and enhancement of Ptdlns3K activity. Furthermore, MIR144* and DRAM2 were critically involved in phagosomal maturation and enhanced antimicrobial effects against Mtb. Our findings identify a previously unrecognized role of human MIR144* in the inhibition of antibacterial autophagy and the innate host immune response to Mtb. Additionally, these data reveal that DRAM2 is a key coordinator of autophagy activation that enhances antimicrobial activity against Mtb.

  13. Specific phosphorylation of histone demethylase KDM3A determines target gene expression in response to heat shock.

    Directory of Open Access Journals (Sweden)

    Mo-bin Cheng

    2014-12-01

    Full Text Available Histone lysine (K residues, which are modified by methyl- and acetyl-transferases, diversely regulate RNA synthesis. Unlike the ubiquitously activating effect of histone K acetylation, the effects of histone K methylation vary with the number of methyl groups added and with the position of these groups in the histone tails. Histone K demethylases (KDMs counteract the activity of methyl-transferases and remove methyl group(s from specific K residues in histones. KDM3A (also known as JHDM2A or JMJD1A is an H3K9me2/1 demethylase. KDM3A performs diverse functions via the regulation of its associated genes, which are involved in spermatogenesis, metabolism, and cell differentiation. However, the mechanism by which the activity of KDM3A is regulated is largely unknown. Here, we demonstrated that mitogen- and stress-activated protein kinase 1 (MSK1 specifically phosphorylates KDM3A at Ser264 (p-KDM3A, which is enriched in the regulatory regions of gene loci in the human genome. p-KDM3A directly interacts with and is recruited by the transcription factor Stat1 to activate p-KDM3A target genes under heat shock conditions. The demethylation of H3K9me2 at the Stat1 binding site specifically depends on the co-expression of p-KDM3A in the heat-shocked cells. In contrast to heat shock, IFN-γ treatment does not phosphorylate KDM3A via MSK1, thereby abrogating its downstream effects. To our knowledge, this is the first evidence that a KDM can be modified via phosphorylation to determine its specific binding to target genes in response to thermal stress.

  14. Specificity and affinity motifs for Grb2 SH2-ligand interactions

    NARCIS (Netherlands)

    Kessels, Helmut W. H. G.; Ward, Alister C.; Schumacher, Ton N. M.

    2002-01-01

    Protein-protein interactions are often mediated by the recognition of short continuous amino acid stretches on target proteins by specific binding domains. Affinity-based selection strategies have successfully been used to define recognition motifs for a large series of such protein domains.

  15. OptMAVEn--a new framework for the de novo design of antibody variable region models targeting specific antigen epitopes.

    Directory of Open Access Journals (Sweden)

    Tong Li

    Full Text Available Antibody-based therapeutics provides novel and efficacious treatments for a number of diseases. Traditional experimental approaches for designing therapeutic antibodies rely on raising antibodies against a target antigen in an immunized animal or directed evolution of antibodies with low affinity for the desired antigen. However, these methods remain time consuming, cannot target a specific epitope and do not lead to broad design principles informing other studies. Computational design methods can overcome some of these limitations by using biophysics models to rationally select antibody parts that maximize affinity for a target antigen epitope. This has been addressed to some extend by OptCDR for the design of complementary determining regions. Here, we extend this earlier contribution by addressing the de novo design of a model of the entire antibody variable region against a given antigen epitope while safeguarding for immunogenicity (Optimal Method for Antibody Variable region Engineering, OptMAVEn. OptMAVEn simulates in silico the in vivo steps of antibody generation and evolution, and is capable of capturing the critical structural features responsible for affinity maturation of antibodies. In addition, a humanization procedure was developed and incorporated into OptMAVEn to minimize the potential immunogenicity of the designed antibody models. As case studies, OptMAVEn was applied to design models of neutralizing antibodies targeting influenza hemagglutinin and HIV gp120. For both HA and gp120, novel computational antibody models with numerous interactions with their target epitopes were generated. The observed rates of mutations and types of amino acid changes during in silico affinity maturation are consistent with what has been observed during in vivo affinity maturation. The results demonstrate that OptMAVEn can efficiently generate diverse computational antibody models with both optimized binding affinity to antigens and reduced

  16. Affinity selection-mass spectrometry and its emerging application to the high throughput screening of G protein-coupled receptors.

    Science.gov (United States)

    Whitehurst, Charles E; Annis, D Allen

    2008-07-01

    Advances in combinatorial chemistry and genomics have inspired the development of novel affinity selection-based screening techniques that rely on mass spectrometry to identify compounds that preferentially bind to a protein target. Of the many affinity selection-mass spectrometry techniques so far documented, only a few solution-based implementations that separate target-ligand complexes away from unbound ligands persist today as routine high throughput screening platforms. Because affinity selection-mass spectrometry techniques do not rely on radioactive or fluorescent reporters or enzyme activities, they can complement traditional biochemical and cell-based screening assays and enable scientists to screen targets that may not be easily amenable to other methods. In addition, by employing mass spectrometry for ligand detection, these techniques enable high throughput screening of massive library collections of pooled compound mixtures, vastly increasing the chemical space that a target can encounter during screening. Of all drug targets, G protein coupled receptors yield the highest percentage of therapeutically effective drugs. In this manuscript, we present the emerging application of affinity selection-mass spectrometry to the high throughput screening of G protein coupled receptors. We also review how affinity selection-mass spectrometry can be used as an analytical tool to guide receptor purification, and further used after screening to characterize target-ligand binding interactions, enabling the classification of orthosteric and allosteric binders.

  17. Selective high-affinity polydentate ligands and methods of making such

    Energy Technology Data Exchange (ETDEWEB)

    Denardo, Sally J.; Denardo, Gerald L.; Balhorn, Rodney L.

    2018-02-06

    This invention provides novel polydentate selective high affinity ligands (SHALs) that can be used in a variety of applications in a manner analogous to the use of antibodies. SHALs typically comprise a multiplicity of ligands that each bind different region son the target molecule. The ligands are joined directly or through a linker thereby forming a polydentate moiety that typically binds the target molecule with high selectivity and avidity.

  18. Targeting HSP90 and monoclonal protein trafficking modulates the unfolded protein response, chaperone regulation and apoptosis in myeloma cells

    International Nuclear Information System (INIS)

    Born, E J; Hartman, S V; Holstein, S A

    2013-01-01

    Multiple myeloma is characterized by the production of substantial quantities of monoclonal protein. We have previously demonstrated that select inhibitors of the isoprenoid biosynthetic pathway (IBP) induce apoptosis of myeloma cells via inhibition of Rab geranylgeranylation, leading to disruption of monoclonal protein trafficking and induction of the unfolded protein response (UPR) pathway. Heat-shock protein 90 (HSP90) inhibitors disrupt protein folding and are currently under clinical investigation in myeloma. The effects of combining IBP and HSP90 inhibitors on cell death, monoclonal protein trafficking, the UPR and chaperone regulation were investigated in monoclonal protein-producing cells. An enhanced induction of cell death was observed following treatment with IBP and HSP90 inhibitors, which occurred through both ER stress and non-ER stress pathways. The HSP90 inhibitor 17-AAG abrogated the effects of the IBP inhibitors on intracellular monoclonal protein levels and localization as well as induction of the UPR in myeloma cells. Disparate effects on chaperone expression were observed in myeloma vs amyloid light chain cells. Here we demonstrate that the novel strategy of targeting MP trafficking in concert with HSP90 enhances myeloma cell death via a complex modulation of ER stress, UPR, and cell death pathways

  19. Gene expression promoted by the SV40 DNA targeting sequence and the hypoxia-responsive element under normoxia and hypoxia

    Directory of Open Access Journals (Sweden)

    C.B. Sacramento

    2010-08-01

    Full Text Available The main objective of the present study was to find suitable DNA-targeting sequences (DTS for the construction of plasmid vectors to be used to treat ischemic diseases. The well-known Simian virus 40 nuclear DTS (SV40-DTS and hypoxia-responsive element (HRE sequences were used to construct plasmid vectors to express the human vascular endothelial growth factor gene (hVEGF. The rate of plasmid nuclear transport and consequent gene expression under normoxia (20% O2 and hypoxia (less than 5% O2 were determined. Plasmids containing the SV40-DTS or HRE sequences were constructed and used to transfect the A293T cell line (a human embryonic kidney cell line in vitro and mouse skeletal muscle cells in vivo. Plasmid transport to the nucleus was monitored by real-time PCR, and the expression level of the hVEGF gene was measured by ELISA. The in vitro nuclear transport efficiency of the SV40-DTS plasmid was about 50% lower under hypoxia, while the HRE plasmid was about 50% higher under hypoxia. Quantitation of reporter gene expression in vitro and in vivo, under hypoxia and normoxia, confirmed that the SV40-DTS plasmid functioned better under normoxia, while the HRE plasmid was superior under hypoxia. These results indicate that the efficiency of gene expression by plasmids containing DNA binding sequences is affected by the concentration of oxygen in the medium.

  20. The nucleolus as a fundamental regulator of the p53 response and a new target for cancer therapy.

    Science.gov (United States)

    Woods, Simone J; Hannan, Katherine M; Pearson, Richard B; Hannan, Ross D

    2015-07-01

    Recent studies have highlighted the fundamental role that key oncogenes such as MYC, RAS and PI3K occupy in driving RNA Polymerase I transcription in the nucleolus. In addition to maintaining essential levels of protein synthesis, hyperactivated ribosome biogenesis and nucleolar function plays a central role in suppressing p53 activation in response to oncogenic stress. Consequently, disruption of ribosome biogenesis by agents such as the small molecule inhibitor of RNA Polymerase I transcription, CX-5461, has shown unexpected, potent, and selective effects in killing tumour cells via disruption of nucleolar function leading to activation of p53, independent of DNA damage. This review will explore the mechanism of DNA damage-independent activation of p53 via the nucleolar surveillance pathway and how this can be utilised to design novel cancer therapies. Non-genotoxic targeting of nucleolar function may provide a new paradigm for treatment of a broad range of oncogene-driven malignancies with improved therapeutic windows. This article is part of a Special Issue entitled: Translation and Cancer. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Targeting the Innate Immune Response to Improve Cardiac Graft Recovery after Heart Transplantation: Implications for the Donation after Cardiac Death

    Directory of Open Access Journals (Sweden)

    Stefano Toldo

    2016-06-01

    Full Text Available Heart transplantation (HTx is the ultimate treatment for end-stage heart failure. The number of patients on waiting lists for heart transplants, however, is much higher than the number of available organs. The shortage of donor hearts is a serious concern since the population affected by heart failure is constantly increasing. Furthermore, the long-term success of HTx poses some challenges despite the improvement in the management of the short-term complications and in the methods to limit graft rejection. Myocardial injury occurs during transplantation. Injury initiated in the donor as result of brain or cardiac death is exacerbated by organ procurement and storage, and is ultimately amplified by reperfusion injury at the time of transplantation. The innate immune system is a mechanism of first-line defense against pathogens and cell injury. Innate immunity is activated during myocardial injury and produces deleterious effects on the heart structure and function. Here, we briefly discuss the role of the innate immunity in the initiation of myocardial injury, with particular focus on the Toll-like receptors and inflammasome, and how to potentially expand the donor population by targeting the innate immune response.

  2. Cis-regulatory element based targeted gene finding: genome-wide identification of abscisic acid- and abiotic stress-responsive genes in Arabidopsis thaliana.

    Science.gov (United States)

    Zhang, Weixiong; Ruan, Jianhua; Ho, Tuan-Hua David; You, Youngsook; Yu, Taotao; Quatrano, Ralph S

    2005-07-15

    A fundamental problem of computational genomics is identifying the genes that respond to certain endogenous cues and environmental stimuli. This problem can be referred to as targeted gene finding. Since gene regulation is mainly determined by the binding of transcription factors and cis-regulatory DNA sequences, most existing gene annotation methods, which exploit the conservation of open reading frames, are not effective in finding target genes. A viable approach to targeted gene finding is to exploit the cis-regulatory elements that are known to be responsible for the transcription of target genes. Given such cis-elements, putative target genes whose promoters contain the elements can be identified. As a case study, we apply the above approach to predict the genes in model plant Arabidopsis thaliana which are inducible by a phytohormone, abscisic acid (ABA), and abiotic stress, such as drought, cold and salinity. We first construct and analyze two ABA specific cis-elements, ABA-responsive element (ABRE) and its coupling element (CE), in A.thaliana, based on their conservation in rice and other cereal plants. We then use the ABRE-CE module to identify putative ABA-responsive genes in A.thaliana. Based on RT-PCR verification and the results from literature, this method has an accuracy rate of 67.5% for the top 40 predictions. The cis-element based targeted gene finding approach is expected to be widely applicable since a large number of cis-elements in many species are available.

  3. Affinity Labeling of Membrane Receptors Using Tissue-Penetrating Radiations

    Directory of Open Access Journals (Sweden)

    Franklin C. Wong

    2013-01-01

    Full Text Available Photoaffinity labeling, a useful in vivo biochemical tool, is limited when applied in vivo because of the poor tissue penetration by ultraviolet (UV photons. This study investigates affinity labeling using tissue-penetrating radiation to overcome the tissue attenuation and irreversibly label membrane receptor proteins. Using X-ray (115 kVp at low doses (<50 cGy or Rad, specific and irreversible binding was found on striatal dopamine transporters with 3 photoaffinity ligands for dopamine transporters, to different extents. Upon X-ray exposure (115 kVp, RTI-38 and RTI-78 ligands showed irreversible and specific binding to the dopamine transporter similar to those seen with UV exposure under other conditions. Similarly, gamma rays at higher energy (662 keV also affect irreversible binding of photoreactive ligands to peripheral benzodiazepine receptors (by PK14105 and to the dopamine (D2 membrane receptors (by azidoclebopride, respectively. This study reports that X-ray and gamma rays induced affinity labeling of membrane receptors in a manner similar to UV with photoreactive ligands of the dopamine transporter, D2 dopamine receptor (D2R, and peripheral benzodiazepine receptor (PBDZR. It may provide specific noninvasive irreversible block or stimulation of a receptor using tissue-penetrating radiation targeting selected anatomic sites.

  4. Rapid purification of circular DNA by triplex-mediated affinity capture

    Science.gov (United States)

    Ji, H.; Smith, L.M.

    1997-01-07

    A single-step capture of a target supercoiled double-stranded DNA molecule is accomplished by forming a local triple-helix among two strands of the supercoiled circular DNA and an oligonucleotide probe. The oligonucleotide is bound to an immobilizing support which facilitates the immobilization and purification of target DNA molecules. Non-target DNA molecules and other contaminating cellular material are easily removed by washing. The triple-helical structure is destabilized by raising the pH, leaving purified target DNA in the supernatant and reusable affinity capture oligonucleotide secured to the immobilizing support. 3 figs.

  5. miR-630 targets IGF1R to regulate response to HER-targeting drugs and overall cancer cell progression in HER2 over-expressing breast cancer.

    Science.gov (United States)

    Corcoran, Claire; Rani, Sweta; Breslin, Susan; Gogarty, Martina; Ghobrial, Irene M; Crown, John; O'Driscoll, Lorraine

    2014-03-24

    While the treatment of HER2 over-expressing breast cancer with recent HER-targeted drugs has been highly effective for some patients, primary (also known as innate) or acquired resistance limits the success of these drugs. microRNAs have potential as diagnostic, prognostic and predictive biomarkers, as well as replacement therapies. Here we investigated the role of microRNA-630 (miR-630) in breast cancer progression and as a predictive biomarker for response to HER-targeting drugs, ultimately yielding potential as a therapeutic approach to add value to these drugs. We investigated the levels of intra- and extracellular miR-630 in cells and conditioned media from breast cancer cell lines with either innate- or acquired- resistance to HER-targeting lapatinib and neratinib, compared to their corresponding drug sensitive cell lines, using qPCR. To support the role of miR-630 in breast cancer, we examined the clinical relevance of this miRNA in breast cancer tumours versus matched peritumours. Transfection of miR-630 mimics and inhibitors was used to manipulate the expression of miR-630 to assess effects on response to HER-targeting drugs (lapatinib, neratinib and afatinib). Other phenotypic changes associated with cellular aggressiveness were evaluated by motility, invasion and anoikis assays. TargetScan prediction software, qPCR, immunoblotting and ELISAs, were used to assess miR-630's regulation of mRNA, proteins and their phosphorylated forms. We established that introducing miR-630 into cells with innate- or acquired- resistance to HER-drugs significantly restored the efficacy of lapatinib, neratinib and afatinib; through a mechanism which we have determined to, at least partly, involve miR-630's regulation of IGF1R. Conversely, we demonstrated that blocking miR-630 induced resistance/insensitivity to these drugs. Cellular motility, invasion, and anoikis were also observed as significantly altered by miR-630 manipulation, whereby introducing miR-630 into cells

  6. Structure of IL-22 Bound to Its High-Affinity IL-22R1 Chain

    Energy Technology Data Exchange (ETDEWEB)

    Jones, B.C.; Logsdon, N.J.; Walter, M.R. (UAB)

    2008-09-29

    IL-22 is an IL-10 family cytokine that initiates innate immune responses against bacterial pathogens and contributes to immune disease. IL-22 biological activity is initiated by binding to a cell-surface complex composed of IL-22R1 and IL-10R2 receptor chains and further regulated by interactions with a soluble binding protein, IL-22BP, which shares sequence similarity with an extracellular region of IL-22R1 (sIL-22R1). IL-22R1 also pairs with the IL-20R2 chain to induce IL-20 and IL-24 signaling. To define the molecular basis of these diverse interactions, we have determined the structure of the IL-22/sIL-22R1 complex. The structure, combined with homology modeling and surface plasmon resonance studies, defines the molecular basis for the distinct affinities and specificities of IL-22 and IL-10 receptor chains that regulate cellular targeting and signal transduction to elicit effective immune responses.

  7. Detection-Guided Fast Affine Projection Channel Estimator for Speech Applications

    Directory of Open Access Journals (Sweden)

    Yan Wu Jennifer

    2007-04-01

    Full Text Available In various adaptive estimation applications, such as acoustic echo cancellation within teleconferencing systems, the input signal is a highly correlated speech. This, in general, leads to extremely slow convergence of the NLMS adaptive FIR estimator. As a result, for such applications, the affine projection algorithm (APA or the low-complexity version, the fast affine projection (FAP algorithm, is commonly employed instead of the NLMS algorithm. In such applications, the signal propagation channel may have a relatively low-dimensional impulse response structure, that is, the number m of active or significant taps within the (discrete-time modelled channel impulse response is much less than the overall tap length n of the channel impulse response. For such cases, we investigate the inclusion of an active-parameter detection-guided concept within the fast affine projection FIR channel estimator. Simulation results indicate that the proposed detection-guided fast affine projection channel estimator has improved convergence speed and has lead to better steady-state performance than the standard fast affine projection channel estimator, especially in the important case of highly correlated speech input signals.

  8. Paper-based immune-affinity arrays for detection of multiple mycotoxins in cereals.

    Science.gov (United States)

    Li, Li; Chen, Hongpu; Lv, Xiaolan; Wang, Min; Jiang, Xizhi; Jiang, Yifei; Wang, Heye; Zhao, Yongfu; Xia, Liru

    2018-03-01

    Mycotoxins produced by different species of fungi may coexist in cereals and feedstuffs, and could be highly toxic for humans and animals. For quantification of multiple mycotoxins in cereals, we developed a paper-based mycotoxin immune-affinity array. First, paper-based microzone arrays were fabricated by photolithography. Then, monoclonal mycotoxin antibodies were added in a copolymerization reaction with a cross-linker to form an immune-affinity monolith on the paper-based microzone array. With use of a competitive immune-response format, paper-based mycotoxin immune-affinity arrays were successfully applied to detect mycotoxins in samples. The detection limits for deoxynivalenol, zearalenone, T-2 toxin, and HT-2 toxin were 62.7, 10.8, 0.36, and 0.23 μg·kg -1 , respectively, which meet relevant requirements for these compounds in food. The recovery rates were 81-86% for deoxynivalenol, 89-117% for zearalenone, 79-86% for T-2 toxin, and 78-83% for HT-2 toxin, and showed the paper-based immune-affinity arrays had good reproducibility. In summary, the paper-based mycotoxin immune-affinity array provides a sensitive, rapid, accurate, stable, and convenient platform for detection of multiple mycotoxins in agro-foods. Graphical abstract Paper-based immune-affinity monolithic array. DON deoxynivalenol, HT-2 HT-2 toxin, T-2 T-2 toxin, PEGDA polyethylene glycol diacrylate, ZEN zearalenone.

  9. Staircase Models from Affine Toda Field Theory

    CERN Document Server

    Dorey, P; Dorey, Patrick; Ravanini, Francesco

    1993-01-01

    We propose a class of purely elastic scattering theories generalising the staircase model of Al. B. Zamolodchikov, based on the affine Toda field theories for simply-laced Lie algebras g=A,D,E at suitable complex values of their coupling constants. Considering their Thermodynamic Bethe Ansatz equations, we give analytic arguments in support of a conjectured renormalisation group flow visiting the neighbourhood of each W_g minimal model in turn.

  10. Transcriptome-Wide Analysis of Botrytis elliptica Responsive microRNAs and Their Targets in Lilium Regale Wilson by High-Throughput Sequencing and Degradome Analysis

    Directory of Open Access Journals (Sweden)

    Xue Gao

    2017-05-01

    Full Text Available MicroRNAs, as master regulators of gene expression, have been widely identified and play crucial roles in plant-pathogen interactions. A fatal pathogen, Botrytis elliptica, causes the serious folia disease of lily, which reduces production because of the high susceptibility of most cultivated species. However, the miRNAs related to Botrytis infection of lily, and the miRNA-mediated gene regulatory networks providing resistance to B. elliptica in lily remain largely unexplored. To systematically dissect B. elliptica-responsive miRNAs and their target genes, three small RNA libraries were constructed from the leaves of Lilium regale, a promising Chinese wild Lilium species, which had been subjected to mock B. elliptica treatment or B. elliptica infection for 6 and 24 h. By high-throughput sequencing, 71 known miRNAs belonging to 47 conserved families and 24 novel miRNA were identified, of which 18 miRNAs were downreguleted and 13 were upregulated in response to B. elliptica. Moreover, based on the lily mRNA transcriptome, 22 targets for 9 known and 1 novel miRNAs were identified by the degradome sequencing approach. Most target genes for elliptica-responsive miRNAs were involved in metabolic processes, few encoding different transcription factors, including ELONGATION FACTOR 1 ALPHA (EF1a and TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR 2 (TCP2. Furthermore, the expression patterns of a set of elliptica-responsive miRNAs and their targets were validated by quantitative real-time PCR. This study represents the first transcriptome-based analysis of miRNAs responsive to B. elliptica and their targets in lily. The results reveal the possible regulatory roles of miRNAs and their targets in B. elliptica interaction, which will extend our understanding of the mechanisms of this disease in lily.

  11. Calculation of protein-ligand binding affinities.

    Science.gov (United States)

    Gilson, Michael K; Zhou, Huan-Xiang

    2007-01-01

    Accurate methods of computing the affinity of a small molecule with a protein are needed to speed the discovery of new medications and biological probes. This paper reviews physics-based models of binding, beginning with a summary of the changes in potential energy, solvation energy, and configurational entropy that influence affinity, and a theoretical overview to frame the discussion of specific computational approaches. Important advances are reported in modeling protein-ligand energetics, such as the incorporation of electronic polarization and the use of quantum mechanical methods. Recent calculations suggest that changes in configurational entropy strongly oppose binding and must be included if accurate affinities are to be obtained. The linear interaction energy (LIE) and molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) methods are analyzed, as are free energy pathway methods, which show promise and may be ready for more extensive testing. Ultimately, major improvements in modeling accuracy will likely require advances on multiple fronts, as well as continued validation against experiment.

  12. 2D Affine and Projective Shape Analysis.

    Science.gov (United States)

    Bryner, Darshan; Klassen, Eric; Huiling Le; Srivastava, Anuj

    2014-05-01

    Current techniques for shape analysis tend to seek invariance to similarity transformations (rotation, translation, and scale), but certain imaging situations require invariance to larger groups, such as affine or projective groups. Here we present a general Riemannian framework for shape analysis of planar objects where metrics and related quantities are invariant to affine and projective groups. Highlighting two possibilities for representing object boundaries-ordered points (or landmarks) and parameterized curves-we study different combinations of these representations (points and curves) and transformations (affine and projective). Specifically, we provide solutions to three out of four situations and develop algorithms for computing geodesics and intrinsic sample statistics, leading up to Gaussian-type statistical models, and classifying test shapes using such models learned from training data. In the case of parameterized curves, we also achieve the desired goal of invariance to re-parameterizations. The geodesics are constructed by particularizing the path-straightening algorithm to geometries of current manifolds and are used, in turn, to compute shape statistics and Gaussian-type shape models. We demonstrate these ideas using a number of examples from shape and activity recognition.

  13. Excited state electron affinity calculations for aluminum

    Science.gov (United States)

    Hussein, Adnan Yousif

    2017-08-01

    Excited states of negative aluminum ion are reviewed, and calculations of electron affinities of the states (3s^23p^2)^1D and (3s3p^3){^5}{S}° relative to the (3s^23p)^2P° and (3s3p^2)^4P respectively of the neutral aluminum atom are reported in the framework of nonrelativistic configuration interaction (CI) method. A priori selected CI (SCI) with truncation energy error (Bunge in J Chem Phys 125:014107, 2006) and CI by parts (Bunge and Carbó-Dorca in J Chem Phys 125:014108, 2006) are used to approximate the valence nonrelativistic energy. Systematic studies of convergence of electron affinity with respect to the CI excitation level are reported. The calculated value of the electron affinity for ^1D state is 78.675(3) meV. Detailed Calculations on the ^5S°c state reveals that is 1216.8166(3) meV below the ^4P state.

  14. Affinity functions for modeling glass dissolution rates

    Energy Technology Data Exchange (ETDEWEB)

    Bourcier, W.L. [Lawrence Livermore National Lab., CA (United States)

    1997-07-01

    Glass dissolution rates decrease dramatically as glass approach ''saturation'' with respect to the leachate solution. Most repository sites are chosen where water fluxes are minimal, and therefore the waste glass is most likely to dissolve under conditions close to ''saturation''. The key term in the rate expression used to predict glass dissolution rates close to ''saturation'' is the affinity term, which accounts for saturation effects on dissolution rates. Interpretations of recent experimental data on the dissolution behaviour of silicate glasses and silicate minerals indicate the following: 1) simple affinity control does not explain the observed dissolution rate for silicate minerals or glasses; 2) dissolution rates can be significantly modified by dissolved cations even under conditions far from saturation where the affinity term is near unity; 3) the effects of dissolved species such as Al and Si on the dissolution rate vary with pH, temperature, and saturation state; and 4) as temperature is increased, the effect of both pH and temperature on glass and mineral dissolution rates decrease, which strongly suggests a switch in rate control from surface reaction-based to diffusion control. Borosilicate glass dissolution models need to be upgraded to account for these recent experimental observations. (A.C.)

  15. T-cell memory responses elicited by yellow fever vaccine are targeted to overlapping epitopes containing multiple HLA-I and -II binding motifs.

    Directory of Open Access Journals (Sweden)

    Andréa Barbosa de Melo

    Full Text Available The yellow fever vaccines (YF-17D-204 and 17DD are considered to be among the safest vaccines and the presence of neutralizing antibodies is correlated with protection, although other immune effector mechanisms are known to be involved. T-cell responses are known to play an important role modulating antibody production and the killing of infected cells. However, little is known about the repertoire of T-cell responses elicited by the YF-17DD vaccine in humans. In this report, a library of 653 partially overlapping 15-mer peptides covering the envelope (Env and nonstructural (NS proteins 1 to 5 of the vaccine was utilized to perform a comprehensive analysis of the virus-specific CD4(+ and CD8(+ T-cell responses. The T-cell responses were screened ex-vivo by IFN-γ ELISPOT assays using blood samples from 220 YF-17DD vaccinees collected two months to four years after immunization. Each peptide was tested in 75 to 208 separate individuals of the cohort. The screening identified sixteen immunodominant antigens that elicited activation of circulating memory T-cells in 10% to 33% of the individuals. Biochemical in-vitro binding assays and immunogenetic and immunogenicity studies indicated that each of the sixteen immunogenic 15-mer peptides contained two or more partially overlapping epitopes that could bind with high affinity to molecules of different HLAs. The prevalence of the immunogenicity of a peptide in the cohort was correlated with the diversity of HLA-II alleles that they could bind. These findings suggest that overlapping of HLA binding motifs within a peptide enhances its T-cell immunogenicity and the prevalence of the response in the population. In summary, the results suggests that in addition to factors of the innate immunity, "promiscuous" T-cell antigens might contribute to the high efficacy of the yellow fever vaccines.

  16. Targeting Homologous Recombination by Pharmacological Inhibitors Enhances the Killing Response of Glioblastoma Cells Treated with Alkylating Drugs.

    Science.gov (United States)

    Berte, Nancy; Piée-Staffa, Andrea; Piecha, Nadine; Wang, Mengwan; Borgmann, Kerstin; Kaina, Bernd; Nikolova, Teodora

    2016-11-01

    Malignant gliomas exhibit a high level of intrinsic and acquired drug resistance and have a dismal prognosis. First- and second-line therapeutics for glioblastomas are alkylating agents, including the chloroethylating nitrosoureas (CNU) lomustine, nimustine, fotemustine, and carmustine. These agents target the tumor DNA, forming O 6 -chloroethylguanine adducts and secondary DNA interstrand cross-links (ICL). These cross-links are supposed to be converted into DNA double-strand breaks, which trigger cell death pathways. Here, we show that lomustine (CCNU) with moderately toxic doses induces ICLs in glioblastoma cells, inhibits DNA replication fork movement, and provokes the formation of DSBs and chromosomal aberrations. Since homologous recombination (HR) is involved in the repair of DSBs formed in response to CNUs, we elucidated whether pharmacologic inhibitors of HR might have impact on these endpoints and enhance the killing effect. We show that the Rad51 inhibitors RI-1 and B02 greatly ameliorate DSBs, chromosomal changes, and the level of apoptosis and necrosis. We also show that an inhibitor of MRE11, mirin, which blocks the formation of the MRN complex and thus the recognition of DSBs, has a sensitizing effect on these endpoints as well. In a glioma xenograft model, the Rad51 inhibitor RI-1 clearly enhanced the effect of CCNU on tumor growth. The data suggest that pharmacologic inhibition of HR, for example by RI-1, is a reasonable strategy for enhancing the anticancer effect of CNUs. Mol Cancer Ther; 15(11); 2665-78. ©2016 AACR. ©2016 American Association for Cancer Research.

  17. Transfer RNA Derived Small RNAs Targeting Defense Responsive Genes Are Induced during Phytophthora capsici Infection in Black Pepper (Piper nigrum L.).

    Science.gov (United States)

    Asha, Srinivasan; Soniya, Eppurath V

    2016-01-01

    Small RNAs derived from transfer RNAs were recently assigned as potential gene regulatory candidates for various stress responses in eukaryotes. In this study, we report on the cloning and identification of tRNA derived small RNAs from black pepper plants in response to the infection of the quick wilt pathogen, Phytophthora capsici. 5'tRFs cloned from black pepper were validated as highly expressed during P. capsici infection. A high-throughput systematic analysis of the small RNAome (sRNAome) revealed the predominance of 5'tRFs in the infected leaf and root. The abundance of 5'tRFs in the sRNAome and the defense responsive genes as their potential targets indicated their regulatory role during stress response in black pepper. The 5'Ala(CGC) tRF mediated cleavage was experimentally mapped at the tRF binding sites on the mRNA targets of Non-expresser of pathogenesis related protein (NPR1), which was down-regulated during pathogen infection. Comparative sRNAome further demonstrated sequence conservation of 5'Ala tRFs across the angiosperm plant groups, and many important genes in the defense response were identified in silico as their potential targets. Our findings uncovered the diversity, differential expression and stress responsive functional role of tRNA-derived small RNAs during Phytophthora infection in black pepper.

  18. The microRNA miR-29 controls innate and adaptive immune responses to intracellular bacterial infection by targeting interferon-γ.

    Science.gov (United States)

    Ma, Feng; Xu, Sheng; Liu, Xingguang; Zhang, Qian; Xu, Xiongfei; Liu, Mofang; Hua, Minmin; Li, Nan; Yao, Hangping; Cao, Xuetao

    2011-07-24

    Interferon-γ (IFN-γ) has a critical role in immune responses to intracellular bacterial infection. MicroRNAs (miRNAs) are important in the regulation of innate and adaptive immunity. However, whether miRNAs can directly target IFN-γ and regulate IFN-γ production post-transcriptionally remains unknown. Here we show that infection of mice with Listeria monocytogenes or Mycobacterium bovis bacillus Calmette-Guérin (BCG) downregulated miR-29 expression in IFN-γ-producing natural killer cells, CD4(+) T cells and CD8(+) T cells. Moreover, miR-29 suppressed IFN-γ production by directly targeting IFN-γ mRNA. We developed mice with transgenic expression of a 'sponge' target to compete with endogenous miR-29 targets (GS29 mice). We found higher serum concentrations of IFN-γ and lower L. monocytogenes burdens in L. monocytogenes-infected GS29 mice than in their littermates. GS29 mice had enhanced T helper type 1 (T(H)1) responses and greater resistance to infection with BCG or Mycobacterium tuberculosis. Therefore, miR-29 suppresses immune responses to intracellular pathogens by targeting IFN-γ.

  19. Development of an affinity-matured humanized anti-epidermal growth factor receptor antibody for cancer immunotherapy.

    Science.gov (United States)

    Nakanishi, Takeshi; Maru, Takamitsu; Tahara, Kazuhiro; Sanada, Hideaki; Umetsu, Mitsuo; Asano, Ryutaro; Kumagai, Izumi

    2013-02-01

    We showed previously that humanization of 528, a murine anti-epidermal growth factor receptor (EGFR) antibody, causes reduced affinity for its target. Here, to improve the affinity of the humanized antibody for use in cancer immunotherapy, we constructed phage display libraries focused on the complementarity-determining regions (CDRs) of the antibody and carried out affinity selection. Two-step selections using libraries constructed in a stepwise manner enabled a 32-fold affinity enhancement of humanized 528 (h528). Thermodynamic analysis of the interactions between the variable domain fragment of h528 (h528Fv) mutants and the soluble extracellular domain of EGFR indicated that the h528Fv mutants obtained from the first selection showed a large increase in negative enthalpy change due to binding, resulting in affinity enhancement. Furthermore, mutants from the second selection showed a decrease in entropy loss, which led to further affinity maturation. These results suggest that a single mutation in the heavy chain variable domain (i.e. Tyr(52) to Trp) enthalpically contributed for overcoming the energetic barrier to the antigen-antibody interaction, which was a major hurdle for the in vitro affinity maturation of h528. We reported previously that the humanized bispecific diabody hEx3 Db, which targets EGFR and CD3, shows strong anti-tumor activity. hEx3 Db mutants, in which the variable domains of h528 were replaced with those of the affinity-enhanced mutants, were prepared and characterized. In a growth inhibition assay of tumor cells, the hEx3 Db mutants showed stronger anti-tumor activity than that of hEx3 Db, suggesting that affinity enhancement of h528Fv enhances the anti-tumor activity of the bispecific diabody.

  20. Survival benefit with proapoptotic molecular and pathologic responses from dual targeting of mammalian target of rapamycin and epidermal growth factor receptor in a preclinical model of pancreatic neuroendocrine carcinogenesis.

    Science.gov (United States)

    Chiu, Christopher W; Nozawa, Hiroaki; Hanahan, Douglas

    2010-10-10

    Pancreatic neuroendocrine tumors (PNETs), although rare, often metastasize, such that surgery, the only potentially curative therapy, is not possible. There is no effective systemic therapy for patients with advanced PNETs. Therefore, new strategies are needed. Toward that end, we investigated the potential benefit of dual therapeutic targeting of the epidermal growth factor receptor (EGFR) and mammalian target of rapamycin (mTOR) kinases, using a preclinical mouse model of PNET. Rapamycin and erlotinib, inhibitors of mTOR and EGFR, respectively, were used to treat RIP-Tag2 transgenic mice bearing advanced multifocal PNET. Tumor growth and survival were monitored, and tumors were surveyed for potential biomarkers of response to the therapeutics. Rapamycin monotherapy was notably efficacious, prolonging survival concomitant with tumor stasis (stable disease). However, the tumors developed resistance, as evidenced by eventual relapse to progressive tumor growth. Erlotinib monotherapy slowed tumor growth and elicited a marginal survival benefit. In combination, there was an unprecedented survival benefit in the face of this aggressive multifocal cancer and, in contrast to either monotherapy, the development of adaptive resistance was not apparent. Additionally, the antiapoptotic protein survivin was implicated as a biomarker of sensitivity and beneficial responses to the dual targeted therapy. Preclinical trials in a mouse model of endogenous PNET suggest that combined targeting of the mTOR and EGFR signaling pathways could have potential clinical benefit in treating PNET. These results have encouraged development of an ongoing phase II clinical trial aimed to evaluate the efficacy of this treatment regimen in human neuroendocrine tumors.

  1. Vaccine Targeting of Subdominant CD8+ T Cell Epitopes Increases the Breadth of the T Cell Response upon Viral Challenge, but May Impair Immediate Virus Control

    DEFF Research Database (Denmark)

    Steffensen, Maria A; Pedersen, Louise Holm; Jahn, Marie Louise

    2016-01-01

    As a result of the difficulties in making efficient vaccines against genetically unstable viruses such as HIV, it has been suggested that future vaccines should preferentially target subdominant epitopes, the idea being that this should allow a greater breadth of the induced T cell response and, ...... a limitation of our model, but clearly our findings underscore the importance of carefully weighing the pros and cons of changes in epitope targeting before any implementation.......As a result of the difficulties in making efficient vaccines against genetically unstable viruses such as HIV, it has been suggested that future vaccines should preferentially target subdominant epitopes, the idea being that this should allow a greater breadth of the induced T cell response and......, hence, a greater efficiency in controlling escape variants. However, to our knowledge the evidence supporting this concept is limited at best. To improve upon this, we used the murine lymphocytic choriomeningitis virus model and adenoviral vectors to compare a vaccine expressing unmodified Ag...

  2. Protein kinase FgSch9 serves as a mediator of the target of rapamycin and high osmolarity glycerol pathways and regulates multiple stress responses and secondary metabolism in Fusarium graminearum.

    Science.gov (United States)

    Gu, Qin; Zhang, Chengqi; Yu, Fangwei; Yin, Yanni; Shim, Won-Bo; Ma, Zhonghua

    2015-08-01

    Saccharomyces cerevisiae protein kinase Sch9 is one of the downstream effectors of the target of rapamycin (TOR) complex 1 and plays multiple roles in stress resistance, longevity and nutrient sensing. However, the functions of Sch9 orthologs in filamentous fungi, particularly in pathogenic species, have not been characterized to date. Here, we investigated biological and genetic functions of FgSch9 in Fusarium graminearum. The FgSCH9 deletion mutant (ΔFgSch9) was defective in aerial hyphal growth, hyphal branching and conidial germination. The mutant exhibited increased sensitivity to osmotic and oxidative stresses, cell wall-damaging agents, and to rapamycin, while showing increased thermal tolerance. We identified FgMaf1 as one of the FgSch9-interacting proteins that plays an important role in regulating mycotoxin biosynthesis and virulence of F. graminearum. Co-immunoprecipitation and affinity capture-mass spectrometry assays showed that FgSch9 also interacts with FgTor and FgHog1. More importantly, both ΔFgSch9 and FgHog1 null mutant (ΔFgHog1) exhibited increased sensitivity to osmotic and oxidative stresses. This defect was more severe in the FgSch9/FgHog1 double mutant. Taken together, we propose that FgSch9 serves as a mediator of the TOR and high osmolarity glycerol pathways, and regulates vegetative differentiation, multiple stress responses and secondary metabolism in F. graminearum. © 2014 Society for Applied Microbiology and John Wiley & Sons Ltd.

  3. Affine fractal functions as bases of continuous funtions | Navascues ...

    African Journals Online (AJOL)

    The objective of the present paper is the study of affine transformations of the plane, which provide self-affine curves as attractors. The properties of these curves depend decisively of the coefficients of the system of affinities involved. The corresponding functions are continuous on a compact interval. If the scale factors are ...

  4. Single-step affinity purification for fungal proteomics.

    Science.gov (United States)

    Liu, Hui-Lin; Osmani, Aysha H; Ukil, Leena; Son, Sunghun; Markossian, Sarine; Shen, Kuo-Fang; Govindaraghavan, Meera; Varadaraj, Archana; Hashmi, Shahr B; De Souza, Colin P; Osmani, Stephen A

    2010-05-01

    A single-step protein affinity purification protocol using Aspergillus nidulans is described. Detailed protocols for cell breakage, affinity purification, and depending on the application, methods for protein release from affinity beads are provided. Examples defining the utility of the approaches, which should be widely applicable, are included.

  5. Single-Step Affinity Purification for Fungal Proteomics ▿ †

    OpenAIRE

    Liu, Hui-Lin; Osmani, Aysha H.; Ukil, Leena; Son, Sunghun; Markossian, Sarine; Shen, Kuo-Fang; Govindaraghavan, Meera; Varadaraj, Archana; Hashmi, Shahr B.; De Souza, Colin P.; Osmani, Stephen A.

    2010-01-01

    A single-step protein affinity purification protocol using Aspergillus nidulans is described. Detailed protocols for cell breakage, affinity purification, and depending on the application, methods for protein release from affinity beads are provided. Examples defining the utility of the approaches, which should be widely applicable, are included.

  6. IA-2 autoantibody affinity in children at risk for type 1 diabetes.

    Science.gov (United States)

    Krause, Stephanie; Chmiel, Ruth; Bonifacio, Ezio; Scholz, Marlon; Powell, Michael; Furmaniak, Jadwiga; Rees Smith, Bernard; Ziegler, Anette-G; Achenbach, Peter

    2012-12-01

    Autoantibodies to insulinoma-associated protein 2 (IA-2A) are associated with increased risk for type 1 diabetes. Here we examined IA-2A affinity and epitope specificity to assess heterogeneity in response intensity in relation to pathogenesis and diabetes risk in 50 children who were prospectively followed from birth. At first IA-2A appearance, affinity ranged from 10(7) to 10(11)L/mol and was high (>1.0×10(9)L/mol) in 41 (82%) children. IA-2A affinity was not associated with epitope specificity or HLA class II haplotype. On follow-up, affinity increased or remained high, and IA-2A were commonly against epitopes within the protein tyrosine phosphatase-like IA-2 domain and the homologue protein IA-2β. IA-2A were preceded or accompanied by other islet autoantibodies in 49 (98%) children, of which 34 progressed to diabetes. IA-2A affinity did not stratify diabetes risk. In conclusion, the IA-2A response in children is intense with rapid maturation against immunogenic epitopes and a strong association with diabetes development. Copyright © 2012 Elsevier Inc. All rights reserved.

  7. Functional Dissociation of Latency-Variable, Stimulus- and Response-Locked Target P3 Sub-components in Task-Switching.

    Science.gov (United States)

    Brydges, Christopher R; Barceló, Francisco

    2018-01-01

    Cognitive control warrants efficient task performance in dynamic and changing environments through adjustments in executive attention, stimulus and response selection. The well-known P300 component of the human event-related potential (ERP) has long been proposed to index "context-updating"-critical for cognitive control-in simple target detection tasks. However, task switching ERP studies have revealed both target P3 (300-350 ms) and later sustained P3-like potentials (400-1,200 ms) to first targets ensuing transition cues, although it remains unclear whether these target P3-like potentials also reflect context updating operations. To address this question, we applied novel single-trial EEG analyses-residue iteration decomposition (RIDE)-in order to disentangle target P3 sub-components in a sample of 22 young adults while they either repeated or switched (updated) task rules. The rationale was to revise the context updating hypothesis of P300 elicitation in the light of new evidence suggesting that "the context" consists of not only the sensory units of stimulation, but also associated motor units, and intermediate low- and high-order sensorimotor units, all of which may need to be dynamically updated on a trial by trial basis. The results showed functionally distinct target P3-like potentials in stimulus-locked, response-locked, and intermediate RIDE component clusters overlying parietal and frontal regions, implying multiple functionally distinct, though temporarily overlapping context updating operations. These findings support a reformulated version of the context updating hypothesis, and reveal a rich family of distinct target P3-like sub-components during the reactive control of target detection in task-switching, plausibly indexing the complex and dynamic workings of frontoparietal cortical networks subserving cognitive control.

  8. Simulating the influence of plasma protein on measured receptor affinity in biochemical assays reveals the utility of Schild analysis for estimating compound affinity for plasma proteins.

    Science.gov (United States)

    Blakeley, D; Sykes, D A; Ensor, P; Bertran, E; Aston, P J; Charlton, S J

    2015-11-01

    Plasma protein binding (PPB) influences the free fraction of drug available to bind to its target and is therefore an important consideration in drug discovery. While traditional methods for assessing PPB (e.g. rapid equilibrium dialysis) are suitable for comparing compounds with relatively weak PPB, they are not able to accurately discriminate between highly bound compounds (typically >99.5%). The aim of the present work was to use mathematical modelling to explore the potential utility of receptor binding and cellular functional assays to estimate the affinity of compounds for plasma proteins. Plasma proteins are routinely added to in vitro assays, so a secondary goal was to investigate the effect of plasma proteins on observed ligand-receptor interactions. Using the principle of conservation of mass and the law of mass action, a cubic equation was derived describing the ligand-receptor complex [LR] in the presence of plasma protein at equilibrium. The model demonstrates the profound influence of PPB on in vitro assays and identifies the utility of Schild analysis, which is usually applied to determine receptor-antagonist affinities, for calculating affinity at plasma proteins (termed KP ). We have also extended this analysis to functional effects using operational modelling and demonstrate that these approaches can also be applied to cell-based assay systems. These mathematical models can potentially be used in conjunction with experimental data to estimate drug-plasma protein affinities in the earliest phases of drug discovery programmes. © 2015 The British Pharmacological Society.

  9. BRCA1, FANCD2 and Chk1 are potential molecular targets for the modulation of a radiation-induced DNA damage response in bystander cells.

    Science.gov (United States)

    Burdak-Rothkamm, Susanne; Rothkamm, Kai; McClelland, Keeva; Al Rashid, Shahnaz T; Prise, Kevin M

    2015-01-28

    Radiotherapy is an important treatment option for many human cancers. Current research is investigating the use of molecular targeted drugs in order to improve responses to radiotherapy in various cancers. The cellular response to irradiation is driven by both direct DNA damage in the targeted cell and intercellular signalling leading to a broad range of bystander effects. This study aims to elucidate radiation-induced DNA damage response signalling in bystander cells and to identify potential molecular targets to modulate the radiation induced bystander response in a therapeutic setting. Stalled replication forks in T98G bystander cells were visualised via bromodeoxyuridine (BrdU) nuclear foci detection at sites of single stranded DNA. γH2AX co-localised with these BrdU foci. BRCA1 and FANCD2 foci formed in T98G bystander cells. Using ATR mutant F02-98 hTERT and ATM deficient GM05849 fibroblasts it could be shown that ATR but not ATM was required for the recruitment of FANCD2 to sites of replication associated DNA damage in bystander cells whereas BRCA1 bystander foci were ATM-dependent. Phospho-Chk1 foci formation was observed in T98G bystander cells. Clonogenic survival assays showed moderate radiosensitisation of directly irradiated cells by the Chk1 inhibitor UCN-01 but increased radioresistance of bystander cells. This study identifies BRCA1, FANCD2 and Chk1 as potential targets for the modulation of radiation response in bystander cells. It adds to our understanding of the key molecular events propagating out-of-field effects of radiation and provides a rationale for the development of novel molecular targeted drugs for radiotherapy optimisation. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  10. Affine Fullerene C60 in a GS-Quasigroup

    Directory of Open Access Journals (Sweden)

    Vladimir Volenec

    2014-01-01

    Full Text Available It will be shown that the affine fullerene C60, which is defined as an affine image of buckminsterfullerene C60, can be obtained only by means of the golden section. The concept of the affine fullerene C60 will be constructed in a general GS-quasigroup using the statements about the relationships between affine regular pentagons and affine regular hexagons. The geometrical interpretation of all discovered relations in a general GS-quasigroup will be given in the GS-quasigroup C(1/2(1+5.

  11. On the structure of self-affine convex bodies

    Energy Technology Data Exchange (ETDEWEB)

    Voynov, A S [M. V. Lomonosov Moscow State University, Faculty of Mechanics and Mathematics, Moscow (Russian Federation)

    2013-08-31

    We study the structure of convex bodies in R{sup d} that can be represented as a union of their affine images with no common interior points. Such bodies are called self-affine. Vallet's conjecture on the structure of self-affine bodies was proved for d = 2 by Richter in 2011. In the present paper we disprove the conjecture for all d≥3 and derive a detailed description of self-affine bodies in R{sup 3}. Also we consider the relation between properties of self-affine bodies and functional equations with a contraction of an argument. Bibliography: 10 titles.

  12. Organelle targeting: third level of drug targeting

    Directory of Open Access Journals (Sweden)

    Sakhrani NM

    2013-07-01

    Full Text Available Niraj M Sakhrani, Harish PadhDepartment of Cell and Molecular Biology, BV Patel Pharmaceutical Education and Research Development (PERD Centre, Gujarat, IndiaAbstract: Drug discovery and drug delivery are two main aspects for treatment of a variety of disorders. However, the real bottleneck associated with systemic drug administration is the lack of target-specific affinity toward a pathological site, resulting in systemic toxicity and innumerable other side effects as well as higher dosage requirement for efficacy. An attractive strategy to increase the therapeutic index of a drug is to specifically deliver the therapeutic molecule in its active form, not only into target tissue, nor even to target cells, but more importantly, into the targeted organelle, ie, to its intracellular therapeutic active site. This would ensure improved efficacy and minimize toxicity. Cancer chemotherapy today faces the major challenge of delivering chemotherapeutic drugs exclusively to tumor cells, while sparing normal proliferating cells. Nanoparticles play a crucial role by acting as a vehicle for delivery of drugs to target sites inside tumor cells. In this review, we spotlight active and passive targeting, followed by discussion of the importance of targeting to specific cell organelles and the potential role of cell-penetrating peptides. Finally, the discussion will address the strategies for drug/DNA targeting to lysosomes, mitochondria, nuclei and Golgi/endoplasmic reticulum.Keywords: intracellular drug delivery, cancer chemotherapy, therapeutic index, cell penetrating peptides

  13. Prenatal smoking predicts non-response to an intervention targeting attention - deficit/hyperactivity problems in elementary schoolchildren

    NARCIS (Netherlands)

    Vuijk, P.J.; van Lier, P.A.C.; Huizink, A.C.; Verhulst, F.C.; Crijnen, A.A.M.

    2006-01-01

    Background: Some evidence suggests that prenatal exposure to maternal smoking contributes to the etiology of Attention-Deficit/Hyperactivity Disorder (ADHD). The present study tested an intervention targeting disruptive behavior to establish whether exposure to maternal smoking during pregnancy

  14. Generation of recombinant antibodies to rat GABAA receptor subunits by affinity selection on synthetic peptides.

    Directory of Open Access Journals (Sweden)

    Sujatha P Koduvayur

    Full Text Available The abundance and physiological importance of GABAA receptors in the central nervous system make this neurotransmitter receptor an attractive target for localizing diagnostic and therapeutic biomolecules. GABAA receptors are expressed within the retina and mediate synaptic signaling at multiple stages of the visual process. To generate monoclonal affinity reagents that can specifically recognize GABAA receptor subunits, we screened two bacteriophage M13 libraries, which displayed human scFvs, by affinity selection with synthetic peptides predicted to correspond to extracellular regions of the rat α1 and β2 GABAA subunits. We isolated three anti-β2 and one anti-α1 subunit specific scFvs. Fluorescence polarization measurements revealed all four scFvs to have low micromolar affinities with their cognate peptide targets. The scFvs were capable of detecting fully folded GABAA receptors heterologously expressed by Xenopus laevis oocytes, while preserving ligand-gated channel activity. Moreover, A10, the anti-α1 subunit-specific scFv, was capable of detecting native GABAA receptors in the mouse retina, as observed by immunofluorescence staining. In order to improve their apparent affinity via avidity, we dimerized the A10 scFv by fusing it to the Fc portion of the IgG. The resulting scFv-Fc construct had a Kd of ∼26 nM, which corresponds to an approximately 135-fold improvement in binding, and a lower detection limit in dot blots, compared to the monomeric scFv. These results strongly support the use of peptides as targets for generating affinity reagents to membrane proteins and encourage investigation of molecular conjugates that use scFvs as anchoring components to localize reagents of interest at GABAA receptors of retina and other neural tissues, for studies of receptor activation and subunit structure.

  15. Knowing where is different from knowing what: Distinct response time profiles and accuracy effects for target location, orientation, and color probability.

    Science.gov (United States)

    Jabar, Syaheed B; Filipowicz, Alex; Anderson, Britt

    2017-11-01

    When a location is cued, targets appearing at that location are detected more quickly. When a target feature is cued, targets bearing that feature are detected more quickly. These attentional cueing effects are only superficially similar. More detailed analyses find distinct temporal and accuracy profiles for the two different types of cues. This pattern parallels work with probability manipulations, where both feature and spatial probability are known to affect detection accuracy and reaction times. However, little has been done by way of comparing these effects. Are probability manipulations on space and features distinct? In a series of five experiments, we systematically varied spatial probability and feature probability along two dimensions (orientation or color). In addition, we decomposed response times into initiation and movement components. Targets appearing at the probable location were reported more quickly and more accurately regardless of whether the report was based on orientation or color. On the other hand, when either color probability or orientation probability was manipulated, response time and accuracy improvements were specific for that probable feature dimension. Decomposition of the response time benefits demonstrated that spatial probability only affected initiation times, whereas manipulations of feature probability affected both initiation and movement times. As detection was made more difficult, the two effects further diverged, with spatial probability disproportionally affecting initiation times and feature probability disproportionately affecting accuracy. In conclusion, all manipulations of probability, whether spatial or featural, affect detection. However, only feature probability affects perceptual precision, and precision effects are specific to the probable attribute.

  16. A potential therapy for chordoma via antibody-dependent cell-mediated cytotoxicity employing NK or high-affinity NK cells in combination with cetuximab.

    Science.gov (United States)

    Fujii, Rika; Schlom, Jeffrey; Hodge, James W

    2018-05-01

    OBJECTIVE Chordoma is a rare bone tumor derived from the notochord and is resistant to conventional therapies such as chemotherapy, radiotherapy, and targeting therapeutics. Expression of epidermal growth factor receptor (EGFR) in a large proportion of chordoma specimens indicates a potential target for therapeutic intervention. In this study the authors investigated the potential role of the anti-EGFR antibody cetuximab in immunotherapy for chordoma. METHODS Since cetuximab is a monoclonal antibody of the IgG1 isotype, it has the potential to mediate antibody-dependent cell-mediated cytotoxicity (ADCC) employing natural killer (NK) cells as effectors. Polymorphisms in the CD16 allele expressed on NK cells have been shown to influence the degree of ADCC of tumor cells, with the high-affinity valine (V)/V allele being responsible for more lysis than the V/phenylalanine (F) or FF allele. Unfortunately, however, only approximately 10% of the population expresses the VV allele on NK cells. An NK cell line, NK-92, has now been engineered to endogenously express IL-2 and the high-affinity CD16 allele. These irradiated high-affinity (ha)NK cells were analyzed for lysis of chordoma cells with and without cetuximab, and the levels of lysis observed in ADCC were compared with those of NK cells from donors expressing the VV, VF, and FF alleles. RESULTS Here the authors demonstrate for the first time 1) that cetuximab in combination with NK cells can mediate ADCC of chordoma cells; 2) the influence of the NK CD16 polymorphism in cetuximab-mediated ADCC for chordoma cell lysis; 3) that engineered haNK cells-that is, cells transduced to express the CD16 V158 FcγRIIIa receptor-bind cetuximab with similar affinity to normal NK cells expressing the high-affinity VV allele; and 4) that irradiated haNK cells induce ADCC with cetuximab in chordoma cells. CONCLUSIONS These studies provide rationale for the use of cetuximab in combination with irradiated haNK cells for therapy for

  17. Fc-Binding Ligands of Immunoglobulin G: An Overview of High Affinity Proteins and Peptides

    Directory of Open Access Journals (Sweden)

    Weonu Choe

    2016-12-01

    Full Text Available The rapidly increasing application of antibodies has inspired the development of several novel methods to isolate and target antibodies using smart biomaterials that mimic the binding of Fc-receptors to antibodies. The Fc-binding domain of antibodies is the primary binding site for e.g., effector proteins and secondary antibodies, whereas antigens bind to the Fab region. Protein A, G, and L, surface proteins expressed by pathogenic bacteria, are well known to bind immunoglobulin and have been widely exploited in antibody purification strategies. Several difficulties are encountered when bacterial proteins are used in antibody research and application. One of the major obstacles hampering the use of bacterial proteins is sample contamination with trace amounts of these proteins, which can invoke an immune response in the host. Many research groups actively develop synthetic ligands that are able to selectively and strongly bind to antibodies. Among the reported ligands, peptides that bind to the Fc-domain of antibodies are attractive tools in antibody research. Besides their use as high affinity ligands in antibody purification chromatography, Fc-binding peptides are applied e.g., to localize antibodies on nanomaterials and to increase the half-life of proteins in serum. In this review, recent developments of Fc-binding peptides are presented and their binding characteristics and diverse applications are discussed.

  18. Immuno and affinity cytochemical analysis of cell wall composition in the moss Physcomitrella patens

    Directory of Open Access Journals (Sweden)

    Elizabeth A. Berry

    2016-03-01

    Full Text Available In contrast to homeohydric vascular plants, mosses employ a poikilohydric strategy for surviving in the dry aerial environment. A detailed understanding of the structure, composition, and development of moss cell walls can contribute to our understanding of not only the evolution of overall cell wall complexity, but also the differences that have evolved in response to selection for different survival strategies. The model moss species Physcomitrella patens has a predominantly haploid lifecycle consisting of protonemal filaments that regenerate from protoplasts and enlarge by tip growth, and leafy gametophores composed of cells that enlarge by diffuse growth and differentiate into several different types. Advantages for genetic studies include methods for efficient targeted gene modification and extensive genomic resources. Immuno and affinity cytochemical labeling were used to examine the distribution of polysaccharides and proteins in regenerated protoplasts, protonemal filaments, rhizoids, and sectioned gametophores of P. patens. The cell wall composition of regenerated protoplasts was also characterized by flow cytometry. Crystalline cellulose was abundant in the cell walls of regenerating protoplasts and protonemal cells that developed on media of high osmolarity, whereas homogalacturonan was detected in the walls of protonemal cells that developed on low osmolarity media and not in regenerating protoplasts. Mannan was the major hemicellulose detected in all tissues tested. Arabinogalactan proteins were detected in different cell types by different probes, consistent with structural heterogeneity. The results reveal developmental and cell type specific differences in cell wall composition and provide a basis for analyzing cell wall phenotypes in knockout mutants.

  19. In situ targeting TEM8 via immune response and polypeptide recognition by wavelength-modulated surface plasmon resonance biosensor

    Science.gov (United States)

    Wang, Yimin; Luo, Zewei; Liu, Kunping; Wang, Jie; Duan, Yixiang

    2016-01-01

    There is an increasing interest in real-time and in situ monitoring of living cell activities in life science and medicine. This paper reports a whole cell sensing protocol over the interface of Au film coupled in a wavelength-modulated surface plasmon resonance (WMSPR) biosensor. With dual parabolic mirrors integrated in the sensor, the compact and miniaturized instrument shows satisfactory refractive index sensitivity (2220 nm/RIU) and a high resolution of resonance wavelength shift of 0.3 nm to liquid samples. The affinity interactions between the biomarker of human tumor endothelial marker 8 (TEM8) and antibody (Ab) or specific polypeptide (PEP) were firstly introduced to WMSPR biosensor analysis. Both the interaction events of Ab-cell and PEP-cell over the Au film interface can be recognized by the sensor and the balance time of interactions is about 20 min. The concentration range of Ab for quantitative monitoring of the TEM8 expression on human colon carcinoma SW620 cells was investigated. The present low-cost and time-saving method provides a time resolution of binding specificity between Ab/PEP and TEM8 for real-time analysis of antigen on living tumor cell surface. PMID:26822761

  20. High affinity hemoglobin and Parkinson's disease.

    Science.gov (United States)

    Graham, Jeffrey; Hobson, Douglas; Ponnampalam, Arjuna

    2014-12-01

    Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra (SN) region of the midbrain. Oxidative damage in this region has been shown to play an important role in the pathogenesis of this disease. Human neurons have been discovered to contain hemoglobin, with an increased concentration seen in the neurons of the SN. High affinity hemoglobin is a clinical entity resulting from mutations that create a functional increase in the binding of hemoglobin to oxygen and an inability to efficiently unload it to tissues. This can result in a number of metabolic compensatory changes, including an elevation in circulating hemoglobin and an increase in the molecule 2,3-diphosphoglycerate (2,3-DPG). Population based studies have revealed that patients with PD have elevated hemoglobin as well as 2,3-DPG levels. Based on these observations, we hypothesize that the oxidative damage seen in PD is related to an underlying high affinity hemoglobin subtype. Copyright © 2014 Elsevier Ltd. All rights reserved.

  1. Aspects of affine Toda field theory

    International Nuclear Information System (INIS)

    Braden, H.W.; Corrigan, E.; Dorey, P.E.; Sasaki, R.

    1990-05-01

    The report is devoted to properties of the affine Toda field theory, the intention being to highlight a selection of curious properties that should be explicable in terms of the underlying group theory but for which in most cases there are no explanation. The motivation for exploring the ideas contained in this report came principally from the recent work of Zamolodchikov concerning the two dimensional Ising model at critical temperature perturbed by a magnetic field. Hollowood and Mansfield pointed out that since Toda field theory is conformal the perturbation considered by Zamolodchikov might well be best regarded as a perturbation of a Toda field theory. This work made it seem plausible that the theory sought by Zamolodchikov was actually affine E 8 Toda field theory. However, this connection required an imaginary value of the coupling constant. Investigations here concerning exact S-matrices use a perturbative approach based on real coupling and the results differ in various ways from those thought to correspond to perturbed conformal field theory. A further motivation is to explore the connection between conformal and perturbed conformal field theories in other contexts using similar ideas. (N.K.)

  2. From affine Hecke algebras to boundary symmetries

    International Nuclear Information System (INIS)

    Doikou, Anastasia

    2005-01-01

    Motivated by earlier works we employ appropriate realizations of the affine Hecke algebra and we recover previously known non-diagonal solutions of the reflection equation for the U q (gl n -bar ) case. The corresponding N site spin chain with open boundary conditions is then constructed and boundary non-local charges associated to the non-diagonal solutions of the reflection equation are derived, as coproduct realizations of the reflection algebra. With the help of linear intertwining relations involving the aforementioned solutions of the reflection equation, the symmetry of the open spin chain with the corresponding boundary conditions is exhibited, being essentially a remnant of the U q (gl n -bar ) algebra. More specifically, we show that representations of certain boundary non-local charges commute with the generators of the affine Hecke algebra and with the local Hamiltonian of the open spin chain for a particular choice of boundary conditions. Furthermore, we are able to show that the transfer matrix of the open spin chain commutes with a certain number of boundary non-local charges, depending on the choice of boundary conditions

  3. Gravitational Goldstone fields from affine gauge theory

    Science.gov (United States)

    Tresguerres, Romualdo; Mielke, Eckehard W.

    2000-08-01

    In order to facilitate the application of standard renormalization techniques, gravitation should be described, in the pure connection formalism, as a Yang-Mills theory of a certain spacetime group, say the Poincaré or the affine group. This embodies the translational as well as the linear connection. However, the coframe is not the standard Yang-Mills-type gauge field of the translations, since it lacks the inhomogeneous gradient term in the gauge transformations. By explicitly restoring this ``hidden'' piece within the framework of nonlinear realizations, the usual geometrical interpretation of the dynamical theory becomes possible, and in addition one can avoid the metric or coframe degeneracy which would otherwise interfere with the integrations within the path integral. We claim that nonlinear realizations provide the general mathematical scheme for the foundation of gauge theories of spacetime symmetries. When applied to construct the Yang-Mills theory of the affine group, tetrads become identified with nonlinear translational connections; the anholonomic metric no longer constitutes an independent gravitational potential, since its degrees of freedom reveal a correspondence to eliminateable Goldstone bosons. This may be an important advantage for quantization.

  4. Detection of aflatoxin B1 in food samples based on target-responsive aptamer-cross-linked hydrogel using a handheld pH meter as readout.

    Science.gov (United States)

    Zhao, Mengmeng; Wang, Peilong; Guo, Yajuan; Wang, Lixu; Luo, Fang; Qiu, Bin; Guo, Longhua; Su, Xiaoou; Lin, Zhenyu; Chen, Guonan

    2018-01-01

    Aflatoxin B 1 (AFB 1 ) can cause great threat to human health, so the development of convenient and portable device for sensitive detection of AFB 1 is highly desired. The portable pH meter has the characters of facile operation, low cost, and easy availability. Therefore, in this study, we investigate the applicability of utilizing a pH meter as the readout to develop a portable sensor for AFB 1 . The specific detection of AFB 1 is realized via the combination of AFB 1 -responsive aptamer-cross-linked hydrogel. Upon the addition of AFB 1 , AFB 1 binds to its aptamer with high affinity in lieu of aptamer/DNA complex, causing the collapse of hydrogel network and results in the releasing of urease into the solution. The released urease can catalyse the hydrolysis of urea and result in the rise of pH value. The change of pH value has a direct relationship to the concentration of AFB 1 in the range of 0.2-20µM with a detection limit of 0.1µM (S/N = 3). The proposed portable device is successfully applied to assay AFB 1 in the food samples with satisfied results. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Affinity Crystallography: A New Approach to Extracting High-Affinity Enzyme Inhibitors from Natural Extracts.

    Science.gov (United States)

    Aguda, Adeleke H; Lavallee, Vincent; Cheng, Ping; Bott, Tina M; Meimetis, Labros G; Law, Simon; Nguyen, Nham T; Williams, David E; Kaleta, Jadwiga; Villanueva, Ivan; Davies, Julian; Andersen, Raymond J; Brayer, Gary D; Brömme, Dieter

    2016-08-26

    Natural products are an important source of novel drug scaffolds. The highly variable and unpredictable timelines associated with isolating novel compounds and elucidating their structures have led to the demise of exploring natural product extract libraries in drug discovery programs. Here we introduce affinity crystallography as a new methodology that significantly shortens the time of the hit to active structure cycle in bioactive natural product discovery research. This affinity crystallography approach is illustrated by using semipure fractions of an actinomycetes culture extract to isolate and identify a cathepsin K inhibitor and to compare the outcome with the traditional assay-guided purification/structural analysis approach. The traditional approach resulted in the identification of the known inhibitor antipain (1) and its new but lower potency dehydration product 2, while the affinity crystallography approach led to the identification of a new high-affinity inhibitor named lichostatinal (3). The structure and potency of lichostatinal (3) was verified by total synthesis and kinetic characterization. To the best of our knowledge, this is the first example of isolating and characterizing a potent enzyme inhibitor from a partially purified crude natural product extract using a protein crystallographic approach.

  6. Affinity-based screening of combinatorial libraries using automated, serial-column chromatography

    Energy Technology Data Exchange (ETDEWEB)

    Evans, D.M.; Williams, K.P.; McGuinness, B. [PerSeptive Biosystems, Framingham, MA (United States)] [and others

    1996-04-01

    The authors have developed an automated serial chromatographic technique for screening a library of compounds based upon their relative affinity for a target molecule. A {open_quotes}target{close_quotes} column containing the immobilized target molecule is set in tandem with a reversed-phase column. A combinatorial peptide library is injected onto the target column. The target-bound peptides are eluted from the first column and transferred automatically to the reversed-phase column. The target-specific peptide peaks from the reversed-phase column are identified and sequenced. Using a monoclonal antibody (3E-7) against {beta}-endorphin as a target, we selected a single peptide with sequence YGGFL from approximately 5800 peptides present in a combinatorial library. We demonstrated the applicability of the technology towards selection of peptides with predetermined affinity for bacterial lipopolysaccharide (LPS, endotoxin). We expect that this technology will have broad applications for high throughput screening of chemical libraries or natural product extracts. 21 refs., 4 figs.

  7. Displacement affinity chromatography of protein phosphatase one (PP1 complexes

    Directory of Open Access Journals (Sweden)

    Gourlay Robert

    2008-11-01

    Full Text Available Abstract Background Protein phosphatase one (PP1 is a ubiquitously expressed, highly conserved protein phosphatase that dephosphorylates target protein serine and threonine residues. PP1 is localized to its site of action by interacting with targeting or regulatory proteins, a majority of which contains a primary docking site referred to as the RVXF/W motif. Results We demonstrate that a peptide based on the RVXF/W motif can effectively displace PP1 bound proteins from PP1 retained on the phosphatase affinity matrix microcystin-Sepharose. Subsequent co-immunoprecipitation experiments confirmed that each identified binding protein was either a direct PP1 interactor or was in a complex that contains PP1. Our results have linked PP1 to numerous new nuclear functions and proteins, including Ki-67, Rif-1, topoisomerase IIα, several nuclear helicases, NUP153 and the TRRAP complex. Conclusion This modification of the microcystin-Sepharose technique offers an effective means of purifying novel PP1 regulatory subunits and associated proteins and provides a simple method to uncover a link between PP1 and additional cellular processes.

  8. Proteome-wide analysis of SUMO2 targets in response to pathological DNA replication stress in human cells

    DEFF Research Database (Denmark)

    Bursomanno, Sara; Beli, Petra; Khan, Asif M

    2015-01-01

    SUMOylation is a form of post-translational modification involving covalent attachment of SUMO (Small Ubiquitin-like Modifier) polypeptides to specific lysine residues in the target protein. In human cells, there are four SUMO proteins, SUMO1-4, with SUMO2 and SUMO3 forming a closely related subf......, and that excessive replication stress is a hallmark of pre-neoplastic and tumor cells, our characterization of SUMO2 targets during a perturbed S-phase should provide a valuable resource for future functional studies in the fields of DNA metabolism and cancer biology....

  9. New approaches for the reliable in vitro assessment of binding affinity based on high-resolution real-time data acquisition of radioligand-receptor binding kinetics.

    Science.gov (United States)

    Zeilinger, Markus; Pichler, Florian; Nics, Lukas; Wadsak, Wolfgang; Spreitzer, Helmut; Hacker, Marcus; Mitterhauser, Markus

    2017-12-01

    Resolving the kinetic mechanisms of biomolecular interactions have become increasingly important in early-phase drug development. Since traditional in vitro methods belong to dose-dependent assessments, binding kinetics is usually overlooked. The present study aimed at the establishment of two novel experimental approaches for the assessment of binding affinity of both, radiolabelled and non-labelled compounds targeting the A 3 R, based on high-resolution real-time data acquisition of radioligand-receptor binding kinetics. A novel time-resolved competition assay was developed and applied to determine the K i of eight different A 3 R antagonists, using CHO-K1 cells stably expressing the hA 3 R. In addition, a new kinetic real-time cell-binding approach was established to quantify the rate constants k on and k off , as well as the dedicated K d of the A 3 R agonist [ 125 I]-AB-MECA. Furthermore, lipophilicity measurements were conducted to control influences due to physicochemical properties of the used compounds. Two novel real-time cell-binding approaches were successfully developed and established. Both experimental procedures were found to visualize the kinetic binding characteristics with high spatial and temporal resolution, resulting in reliable affinity values, which are in good agreement with values previously reported with traditional methods. Taking into account the lipophilicity of the A 3 R antagonists, no influences on the experimental performance and the resulting affinity were investigated. Both kinetic binding approaches comprise tracer administration and subsequent binding to living cells, expressing the dedicated target protein. Therefore, the experiments resemble better the true in vivo physiological conditions and provide important markers of cellular feedback and biological response.

  10. Putative therapeutic targets for symptom subtypes of adult ADHD: D4 receptor agonism and COMT inhibition improve attention and response inhibition in a novel translational animal model.

    Science.gov (United States)

    Tomlinson, Anneka; Grayson, Ben; Marsh, Samuel; Hayward, Andrew; Marshall, Kay M; Neill, Joanna C

    2015-04-01

    Prefrontal cortical dopamine plays an important role in cognitive control, specifically in attention and response inhibition; the core deficits in ADHD. We have previously shown that methylphenidate and atomoxetine differentially improve these deficits dependent on baseline performance. The present study extends this work to investigate the effects of putative therapeutic targets in our model. A selective dopamine D4 receptor agonist (A-412997) and the catechol-O-methyl-transferase (COMT) inhibitor; tolcapone, were investigated in the combined subtype of adult ADHD (ADHD-C). Adult female rats were trained to criterion in the 5C-CPT (5-Choice Continuous Performance Task) and then separated into subgroups according to baseline levels of sustained attention, vigilance, and response disinhibition. The subgroups included: high-attentive (HA) and low-attentive with high response disinhibition (ADHD-C). The ADHD-C subgroup was selected to represent the combined subtype of adult ADHD. Effects of tolcapone (3.0, 10.0, 15.0mg/kg) and A-412997 (0.1, 0.3, 1.0µmol/kg) were tested by increasing the variable inter-trial-interval (ITI) duration in the 5C-CPT. Tolcapone (15mg/kg) significantly increased sustained attention, vigilance and response inhibition in ADHD-C animals, and impaired attention in HA animals. A-412997 (1.0µmol/kg) significantly increased vigilance and response inhibition in ADHD-C animals only, with no effect in HA animals. This is the first study to use the translational 5C-CPT to model the adult ADHD-C subtype in rats and to study new targets in this model. Both tolcapone and A-412997 increased vigilance and response inhibition in the ADHD-C subgroup. D4 and COMT are emerging as important potential therapeutic targets in adult ADHD that warrant further investigation. Copyright © 2014 Elsevier B.V. and ECNP. All rights reserved.

  11. Transcriptome and Degradome of microRNAs and Their Targets in Response to Drought Stress in the Plants of a Diploid and Its Autotetraploid Paulownia australis.

    Directory of Open Access Journals (Sweden)

    Suyan Niu

    Full Text Available MicroRNAs (miRNAs are small, non-coding RNAs that play vital roles in plant growth, development, and stress response. Increasing numbers of studies aimed at discovering miRNAs and analyzing their functions in plants are being reported. In this study, we investigated the effect of drought stress on the expression of miRNAs and their targets in plants of a diploid and derived autotetraploid Paulownia australis. Four small RNA (sRNA libraries and four degradome libraries were constructed from diploid and autotetraploid P. australis plants treated with either 75% or 25% relative soil water content. A total of 33 conserved and 104 novel miRNAs (processing precision value > 0.1 were identified, and 125 target genes were identified for 36 of the miRNAs by using the degradome sequencing. Among the identified miRNAs, 54 and 68 were differentially expressed in diploid and autotetraploid plants under drought stress (25% relative soil water content, respectively. The expressions of miRNAs and target genes were also validated by quantitative real-time PCR. The results showed that the relative expression trends of the randomly selected miRNAs were similar to the trends predicted by Illumina sequencing. And the correlations between miRNAs and their target genes were also analyzed. Furthermore, the functional analysis showed that most of these miRNAs and target genes were associated with plant development and environmental stress response. This study provided molecular evidence for the possible involvement of certain miRNAs in the drought response and/or tolerance in P. australis, and certain level of differential expression between diploid and autotetraploid plants.

  12. Uncovering leaf rust responsive miRNAs in wheat (Triticum aestivum L.) using high-throughput sequencing and prediction of their targets through degradome analysis.

    Science.gov (United States)

    Kumar, Dhananjay; Dutta, Summi; Singh, Dharmendra; Prabhu, Kumble Vinod; Kumar, Manish; Mukhopadhyay, Kunal

    2017-01-01

    Deep sequencing identified 497 conserved and 559 novel miRNAs in wheat, while degradome analysis revealed 701 targets genes. QRT-PCR demonstrated differential expression of miRNAs during stages of leaf rust progression. Bread wheat (Triticum aestivum L.) is an important cereal food crop feeding 30 % of the world population. Major threat to wheat production is the rust epidemics. This study was targeted towards identification and functional characterizations of micro(mi)RNAs and their target genes in wheat in response to leaf rust ingression. High-throughput sequencing was used for transcriptome-wide identification of miRNAs and their expression profiling in retort to leaf rust using mock and pathogen-inoculated resistant and susceptible near-isogenic wheat plants. A total of 1056 mature miRNAs were identified, of which 497 miRNAs were conserved and 559 miRNAs were novel. The pathogen-inoculated resistant plants manifested more miRNAs compared with the pathogen infected susceptible plants. The miRNA counts increased in susceptible isoline due to leaf rust, conversely, the counts decreased in the resistant isoline in response to pathogenesis illustrating precise spatial tuning of miRNAs during compatible and incompatible interaction. Stem-loop quantitative real-time PCR was used to profile 10 highly differentially expressed miRNAs obtained from high-throughput sequencing data. The spatio-temporal profiling validated the differential expression of miRNAs between the isolines as well as in retort to pathogen infection. Degradome analysis provided 701 predicted target genes associated with defense response, signal transduction, development, metabolism, and transcriptional regulation. The obtained results indicate that wheat isolines employ diverse arrays of miRNAs that modulate their target genes during compatible and incompatible interaction. Our findings contribute to increase knowledge on roles of microRNA in wheat-leaf rust interactions and could help in rust

  13. Description of the EuroTARGET cohort: A European collaborative project on TArgeted therapy in renal cell cancer-GEnetic- and tumor-related biomarkers for response and toxicity.

    Science.gov (United States)

    van der Zanden, Loes F M; Vermeulen, Sita H; Oskarsdottir, Arna; Maurits, Jake S F; Diekstra, Meta H M; Ambert, Valentin; Cambon-Thomsen, Anne; Castellano, Daniel; Fritsch, Achim; Garcia Donas, Jesus; Guarch Troyas, Rosa; Guchelaar, Henk-Jan; Hartmann, Arndt; Hulsbergen-van de Kaa, Christina; Jaehde, Ulrich; Junker, Kerstin; Martinez-Cardus, Anna; Masson, Gisli; Oosterwijk-Wakka, Jeannette; Radu, Marius T; Rafnar, Thorunn; Rodriguez-Antona, Cristina; Roessler, Max; Ruijtenbeek, Rob; Stefansson, Kari; Warren, Anne; Wessels, Lodewyk; Eisen, Tim; Kiemeney, Lambertus A L M; Oosterwijk, Egbert

    2017-08-01

    For patients with metastatic renal cell cancer (mRCC), treatment choice is mainly based on clinical parameters. With many treatments available and the limited response to treatment and associated toxicities, there is much interest in identifying better biomarkers for personalized treatment. EuroTARGET aims to identify and characterize host- and tumor-related biomarkers for prediction of response to tyrosine kinase inhibitor therapy in mRCC. Here, we describe the EuroTARGET mRCC patient cohort. EuroTARGET is a European collaborative project designed as an observational study for which patients with mRCC were recruited prospectively in 62 centers. In addition, 462 patients with mRCC from previous studies were included. Detailed clinical information (baseline and follow-up) from all patients was entered in web-based case record forms. Blood was collected for germline DNA and pharmacokinetic/pharmacodynamic analyses and, where available, fresh-frozen tumor material was collected to perform tumor DNA, RNA, kinome, and methylome analyses. In total, 1,210 patients with mRCC were included. Of these, 920 received a tyrosine kinase inhibitor as first-line targeted treatment (sunitinib [N = 713, 78%], sorafenib [N = 41, 4%], or pazopanib [N = 166, 18%]) and had at least 6 months of outcome assessment (median follow-up 15.3 months [interquartile range: 8.5-30.2 months]). Germline DNA samples were available from 824 of these patients, fresh-frozen tumor material from 142 patients, fresh-frozen normal kidney tissue from 95 patients, and tissue microarrays created from formalin-fixed paraffin-embedded tumor material from 247 patients. Of the 920 patients, germline DNA variant chip data were successfully generated for 811 patients (Illumina HumanOmniExpress BeadChip). For 80 patients, next-generation exome sequencing of germline and tumor DNA was performed, tumor RNA sequencing was performed for 124 patients, kinome activity measured and processed for 121 patients (PamChip), and

  14. Genome-wide targeted prediction of ABA responsive genes in rice based on over-represented cis-motif in co-expressed genes.

    Science.gov (United States)

    Lenka, Sangram K; Lohia, Bikash; Kumar, Abhay; Chinnusamy, Viswanathan; Bansal, Kailash C

    2009-02-01

    Abscisic acid (ABA), the popular plant stress hormone, plays a key role in regulation of sub-set of stress responsive genes. These genes respond to ABA through specific transcription factors which bind to cis-regulatory elements present in their promoters. We discovered the ABA Responsive Element (ABRE) core (ACGT) containing CGMCACGTGB motif as over-represented motif among the promoters of ABA responsive co-expressed genes in rice. Targeted gene prediction strategy using this motif led to the identification of 402 protein coding genes potentially regulated by ABA-dependent molecular genetic network. RT-PCR analysis of arbitrarily chosen 45 genes from the predicted 402 genes confirmed 80% accuracy of our prediction. Plant Gene Ontology (GO) analysis of ABA responsive genes showed enrichment of signal transduction and stress related genes among diverse functional categories.

  15. Integrin-Targeted Hybrid Fluorescence Molecular Tomography/X-ray Computed Tomography for Imaging Tumor Progression and Early Response in Non-Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Xiaopeng Ma

    2017-01-01

    Full Text Available Integrins play an important role in tumor progression, invasion and metastasis. Therefore we aimed to evaluate a preclinical imaging approach applying ανβ3 integrin targeted hybrid Fluorescence Molecular Tomography/X-ray Computed Tomography (FMT-XCT for monitoring tumor progression as well as early therapy response in a syngeneic murine Non-Small Cell Lung Cancer (NSCLC model. Lewis Lung Carcinomas were grown orthotopically in C57BL/6 J mice and imaged in-vivo using a ανβ3 targeted near-infrared fluorescence (NIRF probe. ανβ3-targeted FMT-XCT was able to track tumor progression. Cilengitide was able to substantially block the binding of the NIRF probe and suppress the imaging signal. Additionally mice were treated with an established chemotherapy regimen of Cisplatin and Bevacizumab or with a novel MEK inhibitor (Refametinib for 2 weeks. While μCT revealed only a moderate slowdown of tumor growth, ανβ3 dependent signal decreased significantly compared to non-treated mice already at one week post treatment. ανβ3 targeted imaging might therefore become a promising tool for assessment of early therapy response in the future.

  16. Membrane-Dependent Bystander Effect Contributes to Amplification of the Response to Alpha-Particle Irradiation in Targeted and Nontargeted Cells

    International Nuclear Information System (INIS)

    Hanot, Maite; Hoarau, Jim; Carriere, Marie; Angulo, Jaime F.; Khodja, Hicham

    2009-01-01

    Purpose: Free radicals are believed to play an active role in the bystander response. This study investigated their origin as well as their temporal and spatial impacts in the bystander effect. Methods and Materials: We employed a precise alpha-particle microbeam to target a small fraction of subconfluent osteoblastic cells (MC3T3-E1). γH2AX-53BP1 foci, oxidative metabolism changes, and micronuclei induction in targeted and bystander cells were assessed. Results: Cellular membranes and mitochondria were identified as two distinct reactive oxygen species producers. The global oxidative stress observed after irradiation was significantly attenuated after cells were treated with filipin, evidence for the primal role of membrane in the bystander effect. To determine the membrane's impact at a cellular level, micronuclei yield was measured when various fractions of the cell population were individually targeted while the dose per cell remained constant. Induction of micronuclei increased in bystander cells as well as in targeted cells and was attenuated by filipin treatment, demonstrating a role for bystander signals between irradiated cells in an autocrine/paracrine manner. Conclusions: A complex interaction of direct irradiation and bystander signals leads to a membrane-dependent amplification of cell responses that could influence therapeutic outcomes in tissues exposed to low doses or to environmental exposure.

  17. Therapeutic targeting of regulatory T cells enhances tumor-specific CD8+ T cell responses in Epstein–Barr virus associated nasopharyngeal carcinoma

    International Nuclear Information System (INIS)

    Fogg, Mark; Murphy, John R.; Lorch, Jochen; Posner, Marshall; Wang, Fred

    2013-01-01

    Epstein–Barr virus (EBV) is associated with multiple malignancies including nasopharyngeal carcinoma (NPC). In nasopharynx cancer, CD8+ T cells specific for EBV Nuclear Antigen-1 (EBNA-1) and Latent Membrane Protein 2 (LMP2) are important components of anti-tumor immunity since both are consistently expressed in NPC. We have previously shown that EBNA-1-specific CD8+ T cell responses were suppressed in NPC patients compared to healthy controls. We now find that CD8+ T cell responses specific for LMP2 are also abnormal in NPC patients, and both EBNA-1- and LMP2-specific responses are suppressed by regulatory T cells (Treg). EBNA-1 and LMP2-specific CD8+ T cell responses, as well as immune control of EBV-infected cells in vitro, could be restored by the depletion of Tregs and by use of a clinically approved drug targeting Tregs. Thus, in vivo modulation of Tregs may be an effective means of enhancing these anti-tumor immune responses in NPC patients. - Highlights: • Viral proteins are tumor antigens in Epstein–Barr virus associated Nasopharyngeal Carcinoma. • CD8+ T cell responses against EBV proteins EBNA-1 and LMP2 are suppressed in NPC patients. • T regulatory cells are responsible for suppressing EBV immunity in NPC patients. • Depletion of Tregs with Ontak can rescue EBV-specific CD8+ T cell responses in NPC patients. • This clinically approved drug may be effective for enhancing anti-tumor immunity in NPC patients

  18. Therapeutic targeting of regulatory T cells enhances tumor-specific CD8+ T cell responses in Epstein–Barr virus associated nasopharyngeal carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Fogg, Mark [Department of Medicine, Brigham and Women' s Hospital (United States); Murphy, John R. [Departments of Medicine and Microbiology, Boston University School of Medicine, Boston, MA 02118 (United States); Lorch, Jochen; Posner, Marshall [Department of Adult Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115 (United States); Wang, Fred, E-mail: fwang@research.bwh.harvard.edu [Department of Medicine, Brigham and Women' s Hospital (United States); Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115 (United States)

    2013-07-05

    Epstein–Barr virus (EBV) is associated with multiple malignancies including nasopharyngeal carcinoma (NPC). In nasopharynx cancer, CD8+ T cells specific for EBV Nuclear Antigen-1 (EBNA-1) and Latent Membrane Protein 2 (LMP2) are important components of anti-tumor immunity since both are consistently expressed in NPC. We have previously shown that EBNA-1-specific CD8+ T cell responses were suppressed in NPC patients compared to healthy controls. We now find that CD8+ T cell responses specific for LMP2 are also abnormal in NPC patients, and both EBNA-1- and LMP2-specific responses are suppressed by regulatory T cells (Treg). EBNA-1 and LMP2-specific CD8+ T cell responses, as well as immune control of EBV-infected cells in vitro, could be restored by the depletion of Tregs and by use of a clinically approved drug targeting Tregs. Thus, in vivo modulation of Tregs may be an effective means of enhancing these anti-tumor immune responses in NPC patients. - Highlights: • Viral proteins are tumor antigens in Epstein–Barr virus associated Nasopharyngeal Carcinoma. • CD8+ T cell responses against EBV proteins EBNA-1 and LMP2 are suppressed in NPC patients. • T regulatory cells are responsible for suppressing EBV immunity in NPC patients. • Depletion of Tregs with Ontak can rescue EBV-specific CD8+ T cell responses in NPC patients. • This clinically approved drug may be effective for enhancing anti-tumor immunity in NPC patients.

  19. Gene targeting by the vitamin D response element binding protein reveals a role for vitamin D in osteoblast mTOR signaling.

    Science.gov (United States)

    Lisse, Thomas S; Liu, Ting; Irmler, Martin; Beckers, Johannes; Chen, Hong; Adams, John S; Hewison, Martin

    2011-03-01

    Transcriptional regulation by hormonal 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] involves occupancy of vitamin D response elements (VDREs) by the VDRE binding protein (VDRE-BP) or 1,25(OH)(2)D(3)-bound vitamin D receptor (VDR). This relationship is disrupted by elevated VDRE-BP, causing a form of hereditary vitamin D-resistant rickets (HVDRR). DNA array analysis showed that of 114 genes regulated by 1,25(OH)(2)D(3) in control cells, almost all (113) were rendered insensitive to the hormone in VDRE-BP-overexpressing HVDRR cells. Among these was the gene for DNA-damage-inducible transcript 4 (DDIT4), an inhibitor of mammalian target of rapamycin (mTOR) signaling. Chromatin immunoprecipitation PCR using 1,25(OH)(2)D(3)-treated osteoblasts confirmed that VDR and VDRE-BP compete for binding to the DDIT4 gene promoter. Expression of DDIT4 mRNA in these cells was induced (1.6-6 fold) by 1,25(OH)(2)D(3) (10-100 nM), and Western blot and flow cytometry analysis showed that this response involved suppression of phosphorylated S6K1(T389) (a downstream target of mTOR) similar to rapamycin treatment. siRNA knockdown of DDIT4 completely abrogated antiproliferative responses to 1,25(OH)(2)D(3), whereas overexpression of VDRE-BP exerted a dominant-negative effect on transcription of 1,25(OH)(2)D(3)-target genes. DDIT4, an inhibitor of mTOR signaling, is a direct target for 1,25(OH)(2)D(3) and VDRE-BP, and functions to suppress cell proliferation in response to vitamin D.

  20. An off-on fluorescence probe targeting mitochondria based on oxidation-reduction response for tumor cell and tissue imaging

    Science.gov (United States)

    Yao, Hanchun; Cao, Li; Zhao, Weiwei; Zhang, Suge; Zeng, Man; Du, Bin

    2017-10-01

    In this study, a tumor-targeting poly( d, l-lactic-co-glycolic acid) (PLGA) loaded "off-on" fluorescent probe nanoparticle (PFN) delivery system was developed to evaluate the region of tumor by off-on fluorescence. The biodegradability of the nanosize PFN delivery system readily released the probe under tumor acidic conditions. The probe with good biocompatibility was used to monitor the intracellular glutathione (GSH) of cancer cells and selectively localize to mitochondria for tumor imaging. The incorporated tumor-targeting probe was based on the molecular photoinduced electron transfer (PET) mechanism preventing fluorescence ("off" state) and could be easily released under tumor acidic conditions. However, the released tumor-targeting fluorescence probe molecule was selective towards GSH with high selectivity and an ultra-sensitivity for the mitochondria of cancer cells and tissues significantly increasing the probe molecule fluorescence signal ("on" state). The tumor-targeting fluorescence probe showed sensitivity to GSH avoiding interference from cysteine and homocysteine. The PFNs could enable fluorescence-guided cancer imaging during cancer therapy. This work may expand the biological applications of PFNs as a diagnostic reagent, which will be beneficial for fundamental research in tumor imaging. [Figure not available: see fulltext.

  1. Setting ART initiation targets in response to changing guidelines: The importance of addressing both steady-state and backlog.

    Science.gov (United States)

    Martin, Catherine; Naidoo, Nicolette P; Venter, W D Francois; Jaffer, Ambereen; Barker, Pierre M

    2014-05-12

    Target setting is useful in planning, assessing and improving antiretroviral treatment (ART) programmes. In the past 4 years, the ART initiation environment has been transformed due to the change in eligibility criteria (starting ART at a CD4+ count ART. To describe and illustrate the use of a target-setting model for estimating district-based targets in the era of an expanding ART programme and changing CD4+ count thresholds for ART initiation. Using previously described models and data for annual new HIV infections, we estimated both steady-state need for ART initiation and backlog in a North West Province district, accounting for the shift in eligibility. Comparison of actual v. targeted ART initiations was undertaken. The change in CD4+ count threshold adds a once-off group of newly eligible patients to the pool requiring ART - the backlog. The steady-state remains unchanged as it is determined by the annual rate of new HIV infections in previous years. The steady-state need for the district was 639 initiations/month, and the backlog was ~15,388 patients. After the shift in eligibility in September 2011, the steady-state target was exceeded over several months with some backlog addressed. Of the total backlog for this district, 72% remains to be cleared. South Africa has two pools of patients who need ART: the steady-state of HIV-infected patients entering the programme each year, determined by historical infection rates; and the backlog created by the shift in eligibility. The healthcare system needs to build long- term capacity to meet the steady-state need for ART and additional capacity to address the backlog.

  2. Data Stream Clustering With Affinity Propagation

    KAUST Repository

    Zhang, Xiangliang; Furtlehner, Cyril; Germain-Renaud, Cecile; Sebag, Michele

    2014-01-01

    Data stream clustering provides insights into the underlying patterns of data flows. This paper focuses on selecting the best representatives from clusters of streaming data. There are two main challenges: how to cluster with the best representatives and how to handle the evolving patterns that are important characteristics of streaming data with dynamic distributions. We employ the Affinity Propagation (AP) algorithm presented in 2007 by Frey and Dueck for the first challenge, as it offers good guarantees of clustering optimality for selecting exemplars. The second challenging problem is solved by change detection. The presented StrAP algorithm combines AP with a statistical change point detection test; the clustering model is rebuilt whenever the test detects a change in the underlying data distribution. Besides the validation on two benchmark data sets, the presented algorithm is validated on a real-world application, monitoring the data flow of jobs submitted to the EGEE grid.

  3. Data Stream Clustering With Affinity Propagation

    KAUST Repository

    Zhang, Xiangliang

    2014-07-09

    Data stream clustering provides insights into the underlying patterns of data flows. This paper focuses on selecting the best representatives from clusters of streaming data. There are two main challenges: how to cluster with the best representatives and how to handle the evolving patterns that are important characteristics of streaming data with dynamic distributions. We employ the Affinity Propagation (AP) algorithm presented in 2007 by Frey and Dueck for the first challenge, as it offers good guarantees of clustering optimality for selecting exemplars. The second challenging problem is solved by change detection. The presented StrAP algorithm combines AP with a statistical change point detection test; the clustering model is rebuilt whenever the test detects a change in the underlying data distribution. Besides the validation on two benchmark data sets, the presented algorithm is validated on a real-world application, monitoring the data flow of jobs submitted to the EGEE grid.

  4. Self-affinity and nonextensivity of sunspots

    International Nuclear Information System (INIS)

    Moret, M.A.

    2014-01-01

    In this paper we study the time series of sunspots by using two different approaches, analyzing its self-affine behavior and studying its distribution. The long-range correlation exponent α has been calculated via Detrended Fluctuation Analysis and the power law vanishes to values greater than 11 years. On the other hand, the distribution of the sunspots obeys a q-exponential decay that suggests a non-extensive behavior. This observed characteristic seems to take an alternative interpretation of the sunspots dynamics. The present findings suggest us to propose a dynamic model of sunspots formation based on a nonlinear Fokker–Planck equation. Therefore its dynamic process follows the generalized thermostatistical formalism.

  5. Heparin-binding peptide as a novel affinity tag for purification of recombinant proteins.

    Science.gov (United States)

    Morris, Jacqueline; Jayanthi, Srinivas; Langston, Rebekah; Daily, Anna; Kight, Alicia; McNabb, David S; Henry, Ralph; Kumar, Thallapuranam Krishnaswamy Suresh

    2016-10-01

    Purification of recombinant proteins constitutes a significant part of the downstream processing in biopharmaceutical industries. Major costs involved in the production of bio-therapeutics mainly depend on the number of purification steps used during the downstream process. Affinity chromatography is a widely used method for the purification of recombinant proteins expressed in different expression host platforms. Recombinant protein purification is achieved by fusing appropriate affinity tags to either N- or C- terminus of the target recombinant proteins. Currently available protein/peptide affinity tags have proved quite useful in the purification of recombinant proteins. However, these affinity tags suffer from specific limitations in their use under different conditions of purification. In this study, we have designed a novel 34-amino acid heparin-binding affinity tag (HB-tag) for the purification of recombinant proteins expressed in Escherichia coli (E. coli) cells. HB-tag fused recombinant proteins were overexpressed in E. coli in high yields. A one-step heparin-Sepharose-based affinity chromatography protocol was developed to purify HB-fused recombinant proteins to homogeneity using a simple sodium chloride step gradient elution. The HB-tag has also been shown to facilitate the purification of target recombinant proteins from their 8 M urea denatured state(s). The HB-tag has been demonstrated to be successfully released from the fusion protein by an appropriate protease treatment to obtain the recombinant target protein(s) in high yields. Results of the two-dimensional NMR spectroscopy experiments indicate that the purified recombinant target protein(s) exist in the native conformation. Polyclonal antibodies raised against the HB-peptide sequence, exhibited high binding specificity and sensitivity to the HB-fused recombinant proteins (∼10 ng) in different crude cell extracts obtained from diverse expression hosts. In our opinion, the HB-tag provides a

  6. Rapid Diagnostic Assay for Intact Influenza Virus Using a High Affinity Hemagglutinin Binding Protein.

    Science.gov (United States)

    Anderson, Caitlin E; Holstein, Carly A; Strauch, Eva-Maria; Bennett, Steven; Chevalier, Aaron; Nelson, Jorgen; Fu, Elain; Baker, David; Yager, Paul

    2017-06-20

    Influenza is a ubiquitous and recurring infection that results in approximately 500 000 deaths globally each year. Commercially available rapid diagnostic tests are based upon detection of the influenza nucleoprotein, which are limited in that they are unable to differentiate by species and require an additional viral lysis step. Sample preprocessing can be minimized or eliminated by targeting the intact influenza virus, thereby reducing assay complexity and leveraging the large number of hemagglutinin proteins on the surface of each virus. Here, we report the development of a paper-based influenza assay that targets the hemagglutinin protein; the assay employs a combination of antibodies and novel computationally designed, recombinant affinity proteins as the capture and detection agents. This system leverages the customizability of recombinant protein design to target the conserved receptor-binding pocket of the hemagglutinin protein and to match the trimeric nature of hemagglutinin for improved avidity. Using this assay, we demonstrate the first instance of intact influenza virus detection using a combination of antibody and affinity proteins within a porous network. The recombinant head region binder based assays yield superior analytical sensitivity as compared to the antibody based assay, with lower limits of detection of 3.54 × 10 7 and 1.34 × 10 7 CEID 50 /mL for the mixed and all binder stacks, respectively. Not only does this work describe the development of a novel influenza assay, it also demonstrates the power of recombinant affinity proteins for use in rapid diagnostic assays.

  7. Fluorescent carbon dot-gated multifunctional mesoporous silica nanocarriers for redox/enzyme dual-responsive targeted and controlled drug delivery and real-time bioimaging.

    Science.gov (United States)

    Wang, Ying; Cui, Yu; Zhao, Yating; He, Bing; Shi, Xiaoli; Di, Donghua; Zhang, Qiang; Wang, Siling

    2017-08-01

    A distinctive and personalized nanocarrier is described here for controlled and targeted antitumor drug delivery and real-time bioimaging by combining a redox/enzyme dual-responsive disulfide-conjugated carbon dot with mesoporous silica nanoparticles (MSN-SS-CD HA ). The carbon dot with controlling and targeting abilities was prepared through a polymerizing reaction by applying citric acid and HA as starting materials (named CD HA ). The as-prepared MSN-SS-CD HA exhibited not only superior photostability and excellent biocompatibility, but also the ability to target A549 cells with overexpression of CD44 receptors. Upon loading the antitumor drug, doxorubicin (DOX), into the mesoporous channels of MSN nanoparticles, CD HA with a diameter size of 3nm completely blocked the pore entrance of DOX-encapsulated MSN nanoparticles with a pore size of about 3nm, thus preventing the premature leakage of DOX and increasing the antitumor activity until being triggered by specific stimuli in the tumor environment. The results of the cell imaging and cytotoxicity studies demonstrated that the redox/enzyme dual-responsive DOX-encapsulated MSN-SS-CD HA nanoparticles can selectively deliver and control the release of DOX into tumor cells. Ex vivo fluorescence images showed a much stronger fluorescence of MSN-SS-CD HA -DOX in the tumor site than in normal tissues, greatly facilitating the accumulation of DOX in the target tissue. However, its counterpart, MSN-SH-DOX exhibited no or much lower tumor cytotoxicity and drug accumulation in tumor tissue. In addition, MSN-SS-CD was also used as a control to investigate the ability of MSN-SS-CD HA to target A549 cells. The results obtained indicated that MSN-SS-CD HA possessed a higher cellular uptake through the CD44 receptor-mediated endocytosis compared with MSN-SS-CD in the A549 cells. Such specific redox/enzyme dual-responsive targeted nanocarriers are a useful strategy achieving selective controlled and targeted delivery of

  8. Isolation and characterization of mutated alcohol oxidases from the yeast Hansenula polymorpha with decreased affinity toward substrates and their use as selective elements of an amperometric biosensor

    Directory of Open Access Journals (Sweden)

    Schuhmann Wolfgang

    2007-06-01

    Full Text Available Abstract Background Accurate, rapid, and economic on-line analysis of ethanol is very desirable. However, available biosensors achieve saturation at very low ethanol concentrations and thus demand the time and labour consuming procedure of sample dilution. Results Hansenula polymorpha (Pichia angusta mutant strains resistant to allyl alcohol in methanol medium were selected. Such strains possessed decreased affinity of alcohol oxidase (AOX towards methanol: the KM values for AOX of wild type and mutant strains CA2 and CA4 are shown to be 0.62, 2.48 and 1.10 mM, respectively, whereas Vmax values are increased or remain unaffected. The mutant AOX alleles from H. polymorpha mutants CA2 and CA4 were isolated and sequenced. Several point mutations in the AOX gene, mostly different between the two mutant alleles, have been identified. Mutant AOX forms were isolated and purified, and some of their biochemical properties were studied. An amperometric biosensor based on the mutated form of AOX from the strain CA2 was constructed and revealed an extended linear response to the target analytes, ethanol and formaldehyde, as compared to the sensor based on the native AOX. Conclusion The described selection methodology opens up the possibility of isolating modified forms of AOX with further decreased affinity toward substrates without reduction of the maximal velocity of reaction. It can help in creation of improved ethanol biosensors with a prolonged linear response towards ethanol in real samples of wines, beers or fermentation liquids.

  9. The advantages and challenges of using FDG PET/CT for response assessment in melanoma in the era of targeted agents and immunotherapy

    International Nuclear Information System (INIS)

    Wong, Annie N.M.; McArthur, Grant A.; Hofman, Michael S.; Hicks, Rodney J.

    2017-01-01

    The treatment of melanoma has been revolutionised in recent years by advances in the understanding of the genomic landscape of this disease, which has led to the development of new targeted therapeutic agents, and the ability to therapeutically manipulate the immune system through inhibition of cancer cell-T-cell interactions that prevent an adaptive immune response. While these therapeutic interventions have dramatically improved the prospects of survival for patients with advanced melanoma, they bring significant complexity to the interpretation of therapeutic response because their mechanisms and temporal profile of response vary considerably. In this review, we discuss the mode of action of these emerging therapies and their toxicities to provide a framework for the use of FDG PET/CT in therapeutic response assessment. We propose that the greatest utility of PET in assessment of response to agents that abrogate signalling related to BRAF mutation is for early assessment of resistance, while in anti-CTLA4 therapy, immunological flare can compromise early assessment of response but can identify potentially life-threatening autoimmune reactions. For anti-PD1/PDL1 therapy, the role of FDG PET/CT is more akin to its use in other solid malignancies undergoing treatment with conventional chemotherapy. However, further research is required to optimise the timing of scans and response criteria in this disease. (orig.)

  10. The advantages and challenges of using FDG PET/CT for response assessment in melanoma in the era of targeted agents and immunotherapy

    Energy Technology Data Exchange (ETDEWEB)

    Wong, Annie N.M.; McArthur, Grant A. [The Peter MacCallum Cancer Centre, Cancer Medicine, Melbourne (Australia); The University of Melbourne, The Sir Peter MacCallum Department of Oncology, Melbourne (Australia); Hofman, Michael S. [The Peter MacCallum Cancer Centre, Cancer Imaging, Melbourne, VIC (Australia); Hicks, Rodney J. [The University of Melbourne, The Sir Peter MacCallum Department of Oncology, Melbourne (Australia); The Peter MacCallum Cancer Centre, Cancer Imaging, Melbourne, VIC (Australia)

    2017-08-15

    The treatment of melanoma has been revolutionised in recent years by advances in the understanding of the genomic landscape of this disease, which has led to the development of new targeted therapeutic agents, and the ability to therapeutically manipulate the immune system through inhibition of cancer cell-T-cell interactions that prevent an adaptive immune response. While these therapeutic interventions have dramatically improved the prospects of survival for patients with advanced melanoma, they bring significant complexity to the interpretation of therapeutic response because their mechanisms and temporal profile of response vary considerably. In this review, we discuss the mode of action of these emerging therapies and their toxicities to provide a framework for the use of FDG PET/CT in therapeutic response assessment. We propose that the greatest utility of PET in assessment of response to agents that abrogate signalling related to BRAF mutation is for early assessment of resistance, while in anti-CTLA4 therapy, immunological flare can compromise early assessment of response but can identify potentially life-threatening autoimmune reactions. For anti-PD1/PDL1 therapy, the role of FDG PET/CT is more akin to its use in other solid malignancies undergoing treatment with conventional chemotherapy. However, further research is required to optimise the timing of scans and response criteria in this disease. (orig.)

  11. Multiprocessor Real-Time Scheduling with Hierarchical Processor Affinities

    OpenAIRE

    Bonifaci , Vincenzo; Brandenburg , Björn; D'Angelo , Gianlorenzo; Marchetti-Spaccamela , Alberto

    2016-01-01

    International audience; Many multiprocessor real-time operating systems offer the possibility to restrict the migrations of any task to a specified subset of processors by setting affinity masks. A notion of " strong arbitrary processor affinity scheduling " (strong APA scheduling) has been proposed; this notion avoids schedulability losses due to overly simple implementations of processor affinities. Due to potential overheads, strong APA has not been implemented so far in a real-time operat...

  12. Treatment of Renal Cell Carcinoma with 2-Stage Total en bloc Spondylectomy after Marked Response to Molecular Target Drugs

    Directory of Open Access Journals (Sweden)

    Yasuhiro Inoue

    2013-01-01

    Full Text Available Metastatic renal cell carcinoma of the bone occurs at a high rate, and the prognosis is poor. In general, total en bloc spondylectomy is considered when there is only one vertebral metastasis and the primary disease is treated. However, palliative surgery is selected when the primary disease is not being treated or metastasis occurs to an important organ. We encountered a patient in whom lung and vertebra metastases were already present at the time of the first examination at our department and the prognosis was considered poor. However, molecular targeted therapy was markedly effective and enabled 2-stage total en bloc spondylectomy. As of one year after total en bloc spondylectomy, the condition has improved to cane gait, and surgery for lung metastasis is planned. Molecular target drugs might markedly change the current therapeutic strategy for renal cell carcinoma.

  13. The potential impact of case-area targeted interventions in response to cholera outbreaks: A modeling study.

    Science.gov (United States)

    Finger, Flavio; Bertuzzo, Enrico; Luquero, Francisco J; Naibei, Nathan; Touré, Brahima; Allan, Maya; Porten, Klaudia; Lessler, Justin; Rinaldo, Andrea; Azman, Andrew S

    2018-02-01

    Cholera prevention and control interventions targeted to neighbors of cholera cases (case-area targeted interventions [CATIs]), including improved water, sanitation, and hygiene, oral cholera vaccine (OCV), and prophylactic antibiotics, may be able to efficiently avert cholera cases and deaths while saving scarce resources during epidemics. Efforts to quickly target interventions to neighbors of cases have been made in recent outbreaks, but little empirical evidence related to the effectiveness, efficiency, or ideal design of this approach exists. Here, we aim to provide practical guidance on how CATIs might be used by exploring key determinants of intervention impact, including the mix of interventions, "ring" size, and timing, in simulated cholera epidemics fit to data from an urban cholera epidemic in Africa. We developed a micro-simulation model and calibrated it to both the epidemic curve and the small-scale spatiotemporal clustering pattern of case households from a large 2011 cholera outbreak in N'Djamena, Chad (4,352 reported cases over 232 days), and explored the potential impact of CATIs in simulated epidemics. CATIs were implemented with realistic logistical delays after cases presented for care using different combinations of prophylactic antibiotics, OCV, and/or point-of-use water treatment (POUWT) starting at different points during the epidemics and targeting rings of various radii around incident case households. Our findings suggest that CATIs shorten the duration of epidemics and are more resource-efficient than mass campaigns. OCV was predicted to be the most effective single intervention, followed by POUWT and antibiotics. CATIs with OCV started early in an epidemic focusing on a 100-m radius around case households were estimated to shorten epidemics by 68% (IQR 62% to 72%), with an 81% (IQR 69% to 87%) reduction in cases compared to uncontrolled epidemics. These same targeted interventions with OCV led to a 44-fold (IQR 27 to 78) reduction in

  14. Evaluation of Efficiency Activities in the Industrial Sector Undertaken in Response to Greenhouse Gas Emission Reduction Targets

    Energy Technology Data Exchange (ETDEWEB)

    Price, Lynn; de la Rue du Can, Stephane; Lu, Hongyou; Horvath, Arpad

    2010-05-21

    The 2006 California Global Warming Solutions Act calls for reducing greenhouse gas (GHG) emissions to 1990 levels by 2020. Meeting this target will require action from all sectors of the California economy, including industry. The industrial sector consumes 25% of the energy used and emits 28% of the carbon dioxide (CO{sub 2}) produced in the state. Many countries around the world have national-level GHG reduction or energy-efficiency targets, and comprehensive programs focused on implementation of energy efficiency and GHG emissions mitigation measures in the industrial sector are essential for achieving their goals. A combination of targets and industry-focused supporting programs has led to significant investments in energy efficiency as well as reductions in GHG emissions within the industrial sectors in these countries. This project has identified program and policies that have effectively targeted the industrial sector in other countries to achieve real energy and CO{sub 2} savings. Programs in Ireland, France, The Netherlands, Denmark, and the UK were chosen for detailed review. Based on the international experience documented in this report, it is recommended that companies in California's industrial sector be engaged in a program to provide them with support to meet the requirements of AB32, The Global Warming Solution Act. As shown in this review, structured programs that engage industry, require members to evaluate their potential efficiency measures, plan how to meet efficiency or emissions reduction goals, and provide support in achieving the goals, can be quite effective at assisting companies to achieve energy efficiency levels beyond those that can be expected to be achieved autonomously.

  15. Spectral imaging based in vivo model system for characterization of tumor microvessel response to vascular targeting agents

    Science.gov (United States)

    Wankhede, Mamta

    Functional vasculature is vital for tumor growth, proliferation, and metastasis. Many tumor-specific vascular targeting agents (VTAs) aim to destroy this essential tumor vasculature to induce indirect tumor cell death via oxygen and nutrition deprivation. The tumor angiogenesis-inhibiting anti-angiogenics (AIs) and the established tumor vessel targeting vascular disrupting agents (VDAs) are the two major players in the vascular targeting field. Combination of VTAs with conventional therapies or with each other, have been shown to have additive or supra-additive effects on tumor control and treatment. Pathophysiological changes post-VTA treatment in terms of structural and vessel function changes are important parameters to characterize the treatment efficacy. Despite the abundance of information regarding these parameters acquired using various techniques, there remains a need for a quantitative, real-time, and direct observation of these phenomenon in live animals. Through this research we aspired to develop a spectral imaging based mouse tumor system for real-time in vivo microvessel structure and functional measurements for VTA characterization. A model tumor system for window chamber studies was identified, and then combinatorial effects of VDA and AI were characterized in model tumor system. (Full text of this dissertation may be available via the University of Florida Libraries web site. Please check http://www.uflib.ufl.edu/etd.html)

  16. Global mapping of binding sites for Nrf2 identifies novel targets in cell survival response through ChIP-Seq profiling and network analysis

    Science.gov (United States)

    Malhotra, Deepti; Portales-Casamar, Elodie; Singh, Anju; Srivastava, Siddhartha; Arenillas, David; Happel, Christine; Shyr, Casper; Wakabayashi, Nobunao; Kensler, Thomas W.; Wasserman, Wyeth W.; Biswal, Shyam

    2010-01-01

    The Nrf2 (nuclear factor E2 p45-related factor 2) transcription factor responds to diverse oxidative and electrophilic environmental stresses by circumventing repression by Keap1, translocating to the nucleus, and activating cytoprotective genes. Nrf2 responses provide protection against chemical carcinogenesis, chronic inflammation, neurodegeneration, emphysema, asthma and sepsis in murine models. Nrf2 regulates the expression of a plethora of genes that detoxify oxidants and electrophiles and repair or remove damaged macromolecules, such as through proteasomal processing. However, many direct targets of Nrf2 remain undefined. Here, mouse embryonic fibroblasts (MEF) with either constitutive nuclear accumulation (Keap1−/−) or depletion (Nrf2−/−) of Nrf2 were utilized to perform chromatin-immunoprecipitation with parallel sequencing (ChIP-Seq) and global transcription profiling. This unique Nrf2 ChIP-Seq dataset is highly enriched for Nrf2-binding motifs. Integrating ChIP-Seq and microarray analyses, we identified 645 basal and 654 inducible direct targets of Nrf2, with 244 genes at the intersection. Modulated pathways in stress response and cell proliferation distinguish the inducible and basal programs. Results were confirmed in an in vivo stress model of cigarette smoke-exposed mice. This study reveals global circuitry of the Nrf2 stress response emphasizing Nrf2 as a central node in cell survival response. PMID:20460467

  17. Selective induction of high-ouabain-affinity isoform of Na+-K+-ATPase by thyroid hormone

    International Nuclear Information System (INIS)

    Haber, R.S.; Loeb, J.N.

    1988-01-01

    The administration of thyroid hormone is known to result in an induction of the Na + -K + -adenosinetriphosphatase (Na + -K + -ATPase) in rat skeletal muscle and other thyroid hormone-responsive tissues. Since the Na + -K + -ATPase in a variety of mammalian tissues has recently been reported to exist in at least two forms distinguishable by differing affinities for the inhibitory cardiac glycoside ouabain. The authors have studied the effects of 3,3',5-triiodo-L-thyronine (T 3 ) treatment on these two forms of the enzyme in rat diaphragm. The inhibition of Na + -K + -ATPase activity in a crude membrane fraction by varying concentrations of ouabain conformed to a biphasic pattern consistent with the presence of two distinct isoforms with inhibition constants (K I s) for ouabain of ∼10 -7 and 10 -4 M, respectively. Measurement of the specific binding of [ 3 H]ouabain to these membranes confirmed the presence of a class of high-affinity ouabain binding sites with a dissociation constant (K d ) of slightly less than 10 -7 M, whose maximal binding capacity was increased by T 3 treatment by 185%. Binding studies in unfractionated homogenates of diaphragm similarly demonstrated the presence of high-affinity sites whose maximal binding capacity was increased by T 3 treatment. Quantitation of both the high- and low-ouabain-affinity forms of the Na + -K + -ATPase by ouabain-dependent phosphorylation from [ 32 P]orthophosphate confirmed that T 3 treatment markedly increased the number of high-affinity sites while having little effect on the number of low-affinity sites. These observations provide, to our knowledge, the first demonstration that these two forms of the Na + -K + -ATPase are subject to selective hormonal induction

  18. DNA-Damage Response RNA-Binding Proteins (DDRBPs): Perspectives from a New Class of Proteins and Their RNA Targets.

    Science.gov (United States)

    Dutertre, Martin; Vagner, Stéphan

    2017-10-27

    Upon DNA damage, cells trigger an early DNA-damage response (DDR) involving DNA repair and cell cycle checkpoints, and late responses involving gene expression regulation that determine cell fate. Screens for genes involved in the DDR have found many RNA-binding proteins (RBPs), while screens for novel RBPs have identified DDR proteins. An increasing number of RBPs are involved in early and/or late DDR. We propose to call this new class of actors of the DDR, which contain an RNA-binding activity, DNA-damage response RNA-binding proteins (DDRBPs). We then discuss how DDRBPs contribute not only to gene expression regulation in the late DDR but also to early DDR signaling, DNA repair, and chromatin modifications at DNA-damage sites through interactions with both long and short noncoding RNAs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Identification of a novel LMF1 nonsense mutation responsible for severe hypertriglyceridemia by targeted next-generation sequencing.

    Science.gov (United States)

    Cefalù, Angelo B; Spina, Rossella; Noto, Davide; Ingrassia, Valeria; Valenti, Vincenza; Giammanco, Antonina; Fayer, Francesca; Misiano, Gabriella; Cocorullo, Gianfranco; Scrimali, Chiara; Palesano, Ornella; Altieri, Grazia I; Ganci, Antonina; Barbagallo, Carlo M; Averna, Maurizio R

    Severe hypertriglyceridemia (HTG) may result from mutations in genes affecting the intravascular lipolysis of triglyceride (TG)-rich lipoproteins. The aim of this study was to develop a targeted next-generation sequencing panel for the molecular diagnosis of disorders characterized by severe HTG. We developed a targeted customized panel for next-generation sequencing Ion Torrent Personal Genome Machine to capture the coding exons and intron/exon boundaries of 18 genes affecting the main pathways of TG synthesis and metabolism. We sequenced 11 samples of patients with severe HTG (TG>885 mg/dL-10 mmol/L): 4 positive controls in whom pathogenic mutations had previously been identified by Sanger sequencing and 7 patients in whom the molecular defect was still unknown. The customized panel was accurate, and it allowed to confirm genetic variants previously identified in all positive controls with primary severe HTG. Only 1 patient of 7 with HTG was found to be carrier of a homozygous pathogenic mutation of the third novel mutation of LMF1 gene (c.1380C>G-p.Y460X). The clinical and molecular familial cascade screening allowed the identification of 2 additional affected siblings and 7 heterozygous carriers of the mutation. We showed that our targeted resequencing approach for genetic diagnosis of severe HTG appears to be accurate, less time consuming, and more economical compared with traditional Sanger resequencing. The identification of pathogenic mutations in candidate genes remains challenging and clinical resequencing should mainly intended for patients with strong clinical criteria for monogenic severe HTG. Copyright © 2017 National Lipid Association. Published by Elsevier Inc. All rights reserved.

  20. Immune Response to Recombinant Adenovirus in Humans: Capsid Components from Viral Input Are Targets for Vector-Specific Cytotoxic T Lymphocytes

    Science.gov (United States)

    Molinier-Frenkel, Valérie; Gahery-Segard, Hanne; Mehtali, Majid; Le Boulaire, Christophe; Ribault, Sébastien; Boulanger, Pierre; Tursz, Thomas; Guillet, Jean-Gérard; Farace, Françoise

    2000-01-01

    We previously demonstrated that a single injection of 109 PFU of recombinant adenovirus into patients induces strong vector-specific immune responses (H. Gahéry-Ségard, V. Molinier-Frenkel, C. Le Boulaire, P. Saulnier, P. Opolon, R. Lengagne, E. Gautier, A. Le Cesne, L. Zitvogel, A. Venet, C. Schatz, M. Courtney, T. Le Chevalier, T. Tursz, J.-G. Guillet, and F. Farace, J. Clin. Investig. 100:2218–2226, 1997). In the present study we analyzed the mechanism of vector recognition by cytotoxic T lymphocytes (CTL). CD8+ CTL lines were derived from two patients and maintained in long-term cultures. Target cell infections with E1-deleted and E1-plus E2-deleted adenoviruses, as well as transcription-blocking experiments with actinomycin D, revealed that host T-cell recognition did not require viral gene transcription. Target cells treated with brefeldin A were not lysed, indicating that viral input protein-derived peptides are associated with HLA class I molecules. Using recombinant capsid component-loaded targets, we observed that the three major proteins could be recognized. These results raise the question of the use of multideleted adenoviruses for gene therapy in the quest to diminish antivector CTL responses. PMID:10906225

  1. Genomic response to Wnt signalling is highly context-dependent - Evidence from DNA microarray and chromatin immunoprecipitation screens of Wnt/TCF targets

    International Nuclear Information System (INIS)

    Railo, Antti; Pajunen, Antti; Itaeranta, Petri; Naillat, Florence; Vuoristo, Jussi; Kilpelaeinen, Pekka; Vainio, Seppo

    2009-01-01

    Wnt proteins are important regulators of embryonic development, and dysregulated Wnt signalling is involved in the oncogenesis of several human cancers. Our knowledge of the downstream target genes is limited, however. We used a chromatin immunoprecipitation-based assay to isolate and characterize the actual gene segments through which Wnt-activatable transcription factors, TCFs, regulate transcription and an Affymetrix microarray analysis to study the global transcriptional response to the Wnt3a ligand. The anti-β-catenin immunoprecipitation of DNA-protein complexes from mouse NIH3T3 fibroblasts expressing a fusion protein of β-catenin and TCF7 resulted in the identification of 92 genes as putative TCF targets. GeneChip assays of gene expression performed on NIH3T3 cells and the rat pheochromocytoma cell line PC12 revealed 355 genes in NIH3T3 and 129 genes in the PC12 cells with marked changes in expression after Wnt3a stimulus. Only 2 Wnt-regulated genes were shared by both cell lines. Surprisingly, Disabled-2 was the only gene identified by the chromatin immunoprecipitation approach that displayed a marked change in expression in the GeneChip assay. Taken together, our approaches give an insight into the complex context-dependent nature of Wnt pathway transcriptional responses and identify Disabled-2 as a potential new direct target for Wnt signalling.

  2. Genomic Copy Number Dictates a Gene-Independent Cell Response to CRISPR/Cas9 Targeting | Office of Cancer Genomics

    Science.gov (United States)

    The CRISPR/Cas9 system enables genome editing and somatic cell genetic screens in mammalian cells. We performed genome-scale loss-of-function screens in 33 cancer cell lines to identify genes essential for proliferation/survival and found a strong correlation between increased gene copy number and decreased cell viability after genome editing. Within regions of copy-number gain, CRISPR/Cas9 targeting of both expressed and unexpressed genes, as well as intergenic loci, led to significantly decreased cell proliferation through induction of a G2 cell-cycle arrest.

  3. Traceless affinity labeling of endogenous proteins for functional analysis in living cells.

    Science.gov (United States)

    Hayashi, Takahiro; Hamachi, Itaru

    2012-09-18

    Protein labeling and imaging techniques have provided tremendous opportunities to study the structure, function, dynamics, and localization of individual proteins in the complex environment of living cells. Molecular biology-based approaches, such as GFP-fusion tags and monoclonal antibodies, have served as important tools for the visualization of individual proteins in cells. Although these techniques continue to be valuable for live cell imaging, they have a number of limitations that have only been addressed by recent progress in chemistry-based approaches. These chemical approaches benefit greatly from the smaller probe sizes that should result in fewer perturbations to proteins and to biological systems as a whole. Despite the research in this area, so far none of these labeling techniques permit labeling and imaging of selected endogenous proteins in living cells. Researchers have widely used affinity labeling, in which the protein of interest is labeled by a reactive group attached to a ligand, to identify and characterize proteins. Since the first report of af