Sample records for target identification validation

  1. Open Targets: a platform for therapeutic target identification and validation

    PubMed Central

    Koscielny, Gautier; An, Peter; Carvalho-Silva, Denise; Cham, Jennifer A.; Fumis, Luca; Gasparyan, Rippa; Hasan, Samiul; Karamanis, Nikiforos; Maguire, Michael; Papa, Eliseo; Pierleoni, Andrea; Pignatelli, Miguel; Platt, Theo; Rowland, Francis; Wankar, Priyanka; Bento, A. Patrícia; Burdett, Tony; Fabregat, Antonio; Forbes, Simon; Gaulton, Anna; Gonzalez, Cristina Yenyxe; Hermjakob, Henning; Hersey, Anne; Jupe, Steven; Kafkas, Şenay; Keays, Maria; Leroy, Catherine; Lopez, Francisco-Javier; Magarinos, Maria Paula; Malone, James; McEntyre, Johanna; Munoz-Pomer Fuentes, Alfonso; O'Donovan, Claire; Papatheodorou, Irene; Parkinson, Helen; Palka, Barbara; Paschall, Justin; Petryszak, Robert; Pratanwanich, Naruemon; Sarntivijal, Sirarat; Saunders, Gary; Sidiropoulos, Konstantinos; Smith, Thomas; Sondka, Zbyslaw; Stegle, Oliver; Tang, Y. Amy; Turner, Edward; Vaughan, Brendan; Vrousgou, Olga; Watkins, Xavier; Martin, Maria-Jesus; Sanseau, Philippe; Vamathevan, Jessica; Birney, Ewan; Barrett, Jeffrey; Dunham, Ian

    2017-01-01

    We have designed and developed a data integration and visualization platform that provides evidence about the association of known and potential drug targets with diseases. The platform is designed to support identification and prioritization of biological targets for follow-up. Each drug target is linked to a disease using integrated genome-wide data from a broad range of data sources. The platform provides either a target-centric workflow to identify diseases that may be associated with a specific target, or a disease-centric workflow to identify targets that may be associated with a specific disease. Users can easily transition between these target- and disease-centric workflows. The Open Targets Validation Platform is accessible at https://www.targetvalidation.org. PMID:27899665

  2. The druggable genome and support for target identification and validation in drug development.

    PubMed

    Finan, Chris; Gaulton, Anna; Kruger, Felix A; Lumbers, R Thomas; Shah, Tina; Engmann, Jorgen; Galver, Luana; Kelley, Ryan; Karlsson, Anneli; Santos, Rita; Overington, John P; Hingorani, Aroon D; Casas, Juan P

    2017-03-29

    Target identification (determining the correct drug targets for a disease) and target validation (demonstrating an effect of target perturbation on disease biomarkers and disease end points) are important steps in drug development. Clinically relevant associations of variants in genes encoding drug targets model the effect of modifying the same targets pharmacologically. To delineate drug development (including repurposing) opportunities arising from this paradigm, we connected complex disease- and biomarker-associated loci from genome-wide association studies to an updated set of genes encoding druggable human proteins, to agents with bioactivity against these targets, and, where there were licensed drugs, to clinical indications. We used this set of genes to inform the design of a new genotyping array, which will enable association studies of druggable genes for drug target selection and validation in human disease. Copyright © 2017, American Association for the Advancement of Science.

  3. Applications of CRISPR genome editing technology in drug target identification and validation.

    PubMed

    Lu, Quinn; Livi, George P; Modha, Sundip; Yusa, Kosuke; Macarrón, Ricardo; Dow, David J

    2017-06-01

    The analysis of pharmaceutical industry data indicates that the major reason for drug candidates failing in late stage clinical development is lack of efficacy, with a high proportion of these due to erroneous hypotheses about target to disease linkage. More than ever, there is a requirement to better understand potential new drug targets and their role in disease biology in order to reduce attrition in drug development. Genome editing technology enables precise modification of individual protein coding genes, as well as noncoding regulatory sequences, enabling the elucidation of functional effects in human disease relevant cellular systems. Areas covered: This article outlines applications of CRISPR genome editing technology in target identification and target validation studies. Expert opinion: Applications of CRISPR technology in target validation studies are in evidence and gaining momentum. Whilst technical challenges remain, we are on the cusp of CRISPR being applied in complex cell systems such as iPS derived differentiated cells and stem cell derived organoids. In the meantime, our experience to date suggests that precise genome editing of putative targets in primary cell systems is possible, offering more human disease relevant systems than conventional cell lines.

  4. Identification and validation nucleolin as a target of curcumol in nasopharyngeal carcinoma cells.

    PubMed

    Wang, Juan; Wu, Jiacai; Li, Xumei; Liu, Haowei; Qin, Jianli; Bai, Zhun; Chi, Bixia; Chen, Xu

    2018-06-30

    Identification of the specific protein target(s) of a drug is a critical step in unraveling its mechanisms of action (MOA) in many natural products. Curcumol, isolated from well known Chinese medicinal plant Curcuma zedoary, has been shown to possess multiple biological activities. It can inhibit nasopharyngeal carcinoma (NPC) proliferation and induce apoptosis, but its target protein(s) in NPC cells remains unclear. In this study, we employed a mass spectrometry-based chemical proteomics approach reveal the possible protein targets of curcumol in NPC cells. Cellular thermal shift assay (CETSA), molecular docking and cell-based assay was used to validate the binding interactions. Chemical proteomics capturing uncovered that NCL is a target of curcumol in NPC cells, Molecular docking showed that curcumol bound to NCL with an -7.8 kcal/mol binding free energy. Cell function analysis found that curcumol's treatment leads to a degradation of NCL in NPC cells, and it showed slight effects on NP69 cells. In conclusion, our results providing evidences that NCL is a target protein of curcumol. We revealed that the anti-cancer effects of curcumol in NPC cells are mediated, at least in part, by NCL inhibition. Many natural products showed high bioactivity, while their mechanisms of action (MOA) are very poor or completely missed. Understanding the MOA of natural drugs can thoroughly exploit their therapeutic potential and minimize their adverse side effects. Identification of the specific protein target(s) of a drug is a critical step in unraveling its MOA. Compound-centric chemical proteomics is a classic chemical proteomics approach which integrates chemical synthesis with cell biology and mass spectrometry (MS) to identify protein targets of natural products determine the drug mechanism of action, describe its toxicity, and figure out the possible cause of off-target. It is an affinity-based chemical proteomics method to identify small molecule-protein interactions

  5. OPTICAL correlation identification technology applied in underwater laser imaging target identification

    NASA Astrophysics Data System (ADS)

    Yao, Guang-tao; Zhang, Xiao-hui; Ge, Wei-long

    2012-01-01

    the speed and orientation efficiency of target identification effectively, and validate the feasibility of this method primarily.

  6. Literature evidence in open targets - a target validation platform.

    PubMed

    Kafkas, Şenay; Dunham, Ian; McEntyre, Johanna

    2017-06-06

    We present the Europe PMC literature component of Open Targets - a target validation platform that integrates various evidence to aid drug target identification and validation. The component identifies target-disease associations in documents and ranks the documents based on their confidence from the Europe PMC literature database, by using rules utilising expert-provided heuristic information. The confidence score of a given document represents how valuable the document is in the scope of target validation for a given target-disease association by taking into account the credibility of the association based on the properties of the text. The component serves the platform regularly with the up-to-date data since December, 2015. Currently, there are a total number of 1168365 distinct target-disease associations text mined from >26 million PubMed abstracts and >1.2 million Open Access full text articles. Our comparative analyses on the current available evidence data in the platform revealed that 850179 of these associations are exclusively identified by literature mining. This component helps the platform's users by providing the most relevant literature hits for a given target and disease. The text mining evidence along with the other types of evidence can be explored visually through https://www.targetvalidation.org and all the evidence data is available for download in json format from https://www.targetvalidation.org/downloads/data .

  7. Advances in identification and validation of protein targets of natural products without chemical modification.

    PubMed

    Chang, J; Kim, Y; Kwon, H J

    2016-05-04

    Covering: up to February 2016Identification of the target proteins of natural products is pivotal to understanding the mechanisms of action to develop natural products for use as molecular probes and potential therapeutic drugs. Affinity chromatography of immobilized natural products has been conventionally used to identify target proteins, and has yielded good results. However, this method has limitations, in that labeling or tagging for immobilization and affinity purification often result in reduced or altered activity of the natural product. New strategies have recently been developed and applied to identify the target proteins of natural products and synthetic small molecules without chemical modification of the natural product. These direct and indirect methods for target identification of label-free natural products include drug affinity responsive target stability (DARTS), stability of proteins from rates of oxidation (SPROX), cellular thermal shift assay (CETSA), thermal proteome profiling (TPP), and bioinformatics-based analysis of connectivity. This review focuses on and reports case studies of the latest advances in target protein identification methods for label-free natural products. The integration of newly developed technologies will provide new insights and highlight the value of natural products for use as biological probes and new drug candidates.

  8. Strategies for target identification of antimicrobial natural products.

    PubMed

    Farha, Maya A; Brown, Eric D

    2016-05-04

    Covering: 2000 to 2015Despite a pervasive decline in natural product research at many pharmaceutical companies over the last two decades, natural products have undeniably been a prolific and unsurpassed source for new lead antibacterial compounds. Due to their inherent complexity, natural extracts face several hurdles in high-throughout discovery programs, including target identification. Target identification and validation is a crucial process for advancing hits through the discovery pipeline, but has remained a major bottleneck. In the case of natural products, extremely low yields and limited compound supply further impede the process. Here, we review the wealth of target identification strategies that have been proposed and implemented for the characterization of novel antibacterials. Traditionally, these have included genomic and biochemical-based approaches, which, in recent years, have been improved with modern-day technology and better honed for natural product discovery. Further, we discuss the more recent innovative approaches for uncovering the target of new antibacterial natural products, which have resulted from modern advances in chemical biology tools. Finally, we present unique screening platforms implemented to streamline the process of target identification. The different innovative methods to respond to the challenge of characterizing the mode of action for antibacterial natural products have cumulatively built useful frameworks that may advocate a renovated interest in natural product drug discovery programs.

  9. Aptamers as tools for target prioritization and lead identification.

    PubMed

    Burgstaller, Petra; Girod, Anne; Blind, Michael

    2002-12-15

    The increasing number of potential drug target candidates has driven the development of novel technologies designed to identify functionally important targets and enhance the subsequent lead discovery process. Highly specific synthetic nucleic acid ligands--also known as aptamers--offer a new exciting route in the drug discovery process by linking target validation directly with HTS. Recently, aptamers have proven to be valuable tools for modulating the function of endogenous cellular proteins in their natural environment. A set of technologies has been developed to use these sophisticated ligands for the validation of potential drug targets in disease models. Moreover, aptamers that are specific antagonists of protein function can act as substitute interaction partners in HTS assays to facilitate the identification of small-molecule lead compounds.

  10. Probability of identification: a statistical model for the validation of qualitative botanical identification methods.

    PubMed

    LaBudde, Robert A; Harnly, James M

    2012-01-01

    A qualitative botanical identification method (BIM) is an analytical procedure that returns a binary result (1 = Identified, 0 = Not Identified). A BIM may be used by a buyer, manufacturer, or regulator to determine whether a botanical material being tested is the same as the target (desired) material, or whether it contains excessive nontarget (undesirable) material. The report describes the development and validation of studies for a BIM based on the proportion of replicates identified, or probability of identification (POI), as the basic observed statistic. The statistical procedures proposed for data analysis follow closely those of the probability of detection, and harmonize the statistical concepts and parameters between quantitative and qualitative method validation. Use of POI statistics also harmonizes statistical concepts for botanical, microbiological, toxin, and other analyte identification methods that produce binary results. The POI statistical model provides a tool for graphical representation of response curves for qualitative methods, reporting of descriptive statistics, and application of performance requirements. Single collaborator and multicollaborative study examples are given.

  11. IFPTarget: A Customized Virtual Target Identification Method Based on Protein-Ligand Interaction Fingerprinting Analyses.

    PubMed

    Li, Guo-Bo; Yu, Zhu-Jun; Liu, Sha; Huang, Lu-Yi; Yang, Ling-Ling; Lohans, Christopher T; Yang, Sheng-Yong

    2017-07-24

    Small-molecule target identification is an important and challenging task for chemical biology and drug discovery. Structure-based virtual target identification has been widely used, which infers and prioritizes potential protein targets for the molecule of interest (MOI) principally via a scoring function. However, current "universal" scoring functions may not always accurately identify targets to which the MOI binds from the retrieved target database, in part due to a lack of consideration of the important binding features for an individual target. Here, we present IFPTarget, a customized virtual target identification method, which uses an interaction fingerprinting (IFP) method for target-specific interaction analyses and a comprehensive index (Cvalue) for target ranking. Evaluation results indicate that the IFP method enables substantially improved binding pose prediction, and Cvalue has an excellent performance in target ranking for the test set. When applied to screen against our established target library that contains 11,863 protein structures covering 2842 unique targets, IFPTarget could retrieve known targets within the top-ranked list and identified new potential targets for chemically diverse drugs. IFPTarget prediction led to the identification of the metallo-β-lactamase VIM-2 as a target for quercetin as validated by enzymatic inhibition assays. This study provides a new in silico target identification tool and will aid future efforts to develop new target-customized methods for target identification.

  12. Identification of NPM and DDX5 as Therapeutic Targets in TSC

    DTIC Science & Technology

    2017-12-01

    Individuals with TSC develop benign tumors in multiple organs, including the retina, skin, lung, kidney and brain. The identification of valid targets in TSC...individuals. Individuals with TSC develop benign tumors in multiple organs, including the retina, skin, lung, kidney and brain. However, these lesions can

  13. Employing machine learning for reliable miRNA target identification in plants.

    PubMed

    Jha, Ashwani; Shankar, Ravi

    2011-12-29

    miRNAs are ~21 nucleotide long small noncoding RNA molecules, formed endogenously in most of the eukaryotes, which mainly control their target genes post transcriptionally by interacting and silencing them. While a lot of tools has been developed for animal miRNA target system, plant miRNA target identification system has witnessed limited development. Most of them have been centered around exact complementarity match. Very few of them considered other factors like multiple target sites and role of flanking regions. In the present work, a Support Vector Regression (SVR) approach has been implemented for plant miRNA target identification, utilizing position specific dinucleotide density variation information around the target sites, to yield highly reliable result. It has been named as p-TAREF (plant-Target Refiner). Performance comparison for p-TAREF was done with other prediction tools for plants with utmost rigor and where p-TAREF was found better performing in several aspects. Further, p-TAREF was run over the experimentally validated miRNA targets from species like Arabidopsis, Medicago, Rice and Tomato, and detected them accurately, suggesting gross usability of p-TAREF for plant species. Using p-TAREF, target identification was done for the complete Rice transcriptome, supported by expression and degradome based data. miR156 was found as an important component of the Rice regulatory system, where control of genes associated with growth and transcription looked predominant. The entire methodology has been implemented in a multi-threaded parallel architecture in Java, to enable fast processing for web-server version as well as standalone version. This also makes it to run even on a simple desktop computer in concurrent mode. It also provides a facility to gather experimental support for predictions made, through on the spot expression data analysis, in its web-server version. A machine learning multivariate feature tool has been implemented in parallel and

  14. Utilizing random Forest QSAR models with optimized parameters for target identification and its application to target-fishing server.

    PubMed

    Lee, Kyoungyeul; Lee, Minho; Kim, Dongsup

    2017-12-28

    The identification of target molecules is important for understanding the mechanism of "target deconvolution" in phenotypic screening and "polypharmacology" of drugs. Because conventional methods of identifying targets require time and cost, in-silico target identification has been considered an alternative solution. One of the well-known in-silico methods of identifying targets involves structure activity relationships (SARs). SARs have advantages such as low computational cost and high feasibility; however, the data dependency in the SAR approach causes imbalance of active data and ambiguity of inactive data throughout targets. We developed a ligand-based virtual screening model comprising 1121 target SAR models built using a random forest algorithm. The performance of each target model was tested by employing the ROC curve and the mean score using an internal five-fold cross validation. Moreover, recall rates for top-k targets were calculated to assess the performance of target ranking. A benchmark model using an optimized sampling method and parameters was examined via external validation set. The result shows recall rates of 67.6% and 73.9% for top-11 (1% of the total targets) and top-33, respectively. We provide a website for users to search the top-k targets for query ligands available publicly at http://rfqsar.kaist.ac.kr . The target models that we built can be used for both predicting the activity of ligands toward each target and ranking candidate targets for a query ligand using a unified scoring scheme. The scores are additionally fitted to the probability so that users can estimate how likely a ligand-target interaction is active. The user interface of our web site is user friendly and intuitive, offering useful information and cross references.

  15. Target identification of small molecules based on chemical biology approaches.

    PubMed

    Futamura, Yushi; Muroi, Makoto; Osada, Hiroyuki

    2013-05-01

    Recently, a phenotypic approach-screens that assess the effects of compounds on cells, tissues, or whole organisms-has been reconsidered and reintroduced as a complementary strategy of a target-based approach for drug discovery. Although the finding of novel bioactive compounds from large chemical libraries has become routine, the identification of their molecular targets is still a time-consuming and difficult process, making this step rate-limiting in drug development. In the last decade, we and other researchers have amassed a large amount of phenotypic data through progress in omics research and advances in instrumentation. Accordingly, the profiling methodologies using these datasets expertly have emerged to identify and validate specific molecular targets of drug candidates, attaining some progress in current drug discovery (e.g., eribulin). In the case of a compound that shows an unprecedented phenotype likely by inhibiting a first-in-class target, however, such phenotypic profiling is invalid. Under the circumstances, a photo-crosslinking affinity approach should be beneficial. In this review, we describe and summarize recent progress in both affinity-based (direct) and phenotypic profiling (indirect) approaches for chemical biology target identification.

  16. Refining the accuracy of validated target identification through coding variant fine-mapping in type 2 diabetes.

    PubMed

    Mahajan, Anubha; Wessel, Jennifer; Willems, Sara M; Zhao, Wei; Robertson, Neil R; Chu, Audrey Y; Gan, Wei; Kitajima, Hidetoshi; Taliun, Daniel; Rayner, N William; Guo, Xiuqing; Lu, Yingchang; Li, Man; Jensen, Richard A; Hu, Yao; Huo, Shaofeng; Lohman, Kurt K; Zhang, Weihua; Cook, James P; Prins, Bram Peter; Flannick, Jason; Grarup, Niels; Trubetskoy, Vassily Vladimirovich; Kravic, Jasmina; Kim, Young Jin; Rybin, Denis V; Yaghootkar, Hanieh; Müller-Nurasyid, Martina; Meidtner, Karina; Li-Gao, Ruifang; Varga, Tibor V; Marten, Jonathan; Li, Jin; Smith, Albert Vernon; An, Ping; Ligthart, Symen; Gustafsson, Stefan; Malerba, Giovanni; Demirkan, Ayse; Tajes, Juan Fernandez; Steinthorsdottir, Valgerdur; Wuttke, Matthias; Lecoeur, Cécile; Preuss, Michael; Bielak, Lawrence F; Graff, Marielisa; Highland, Heather M; Justice, Anne E; Liu, Dajiang J; Marouli, Eirini; Peloso, Gina Marie; Warren, Helen R; Afaq, Saima; Afzal, Shoaib; Ahlqvist, Emma; Almgren, Peter; Amin, Najaf; Bang, Lia B; Bertoni, Alain G; Bombieri, Cristina; Bork-Jensen, Jette; Brandslund, Ivan; Brody, Jennifer A; Burtt, Noël P; Canouil, Mickaël; Chen, Yii-Der Ida; Cho, Yoon Shin; Christensen, Cramer; Eastwood, Sophie V; Eckardt, Kai-Uwe; Fischer, Krista; Gambaro, Giovanni; Giedraitis, Vilmantas; Grove, Megan L; de Haan, Hugoline G; Hackinger, Sophie; Hai, Yang; Han, Sohee; Tybjærg-Hansen, Anne; Hivert, Marie-France; Isomaa, Bo; Jäger, Susanne; Jørgensen, Marit E; Jørgensen, Torben; Käräjämäki, Annemari; Kim, Bong-Jo; Kim, Sung Soo; Koistinen, Heikki A; Kovacs, Peter; Kriebel, Jennifer; Kronenberg, Florian; Läll, Kristi; Lange, Leslie A; Lee, Jung-Jin; Lehne, Benjamin; Li, Huaixing; Lin, Keng-Hung; Linneberg, Allan; Liu, Ching-Ti; Liu, Jun; Loh, Marie; Mägi, Reedik; Mamakou, Vasiliki; McKean-Cowdin, Roberta; Nadkarni, Girish; Neville, Matt; Nielsen, Sune F; Ntalla, Ioanna; Peyser, Patricia A; Rathmann, Wolfgang; Rice, Kenneth; Rich, Stephen S; Rode, Line; Rolandsson, Olov; Schönherr, Sebastian; Selvin, Elizabeth; Small, Kerrin S; Stančáková, Alena; Surendran, Praveen; Taylor, Kent D; Teslovich, Tanya M; Thorand, Barbara; Thorleifsson, Gudmar; Tin, Adrienne; Tönjes, Anke; Varbo, Anette; Witte, Daniel R; Wood, Andrew R; Yajnik, Pranav; Yao, Jie; Yengo, Loïc; Young, Robin; Amouyel, Philippe; Boeing, Heiner; Boerwinkle, Eric; Bottinger, Erwin P; Chowdhury, Rajiv; Collins, Francis S; Dedoussis, George; Dehghan, Abbas; Deloukas, Panos; Ferrario, Marco M; Ferrières, Jean; Florez, Jose C; Frossard, Philippe; Gudnason, Vilmundur; Harris, Tamara B; Heckbert, Susan R; Howson, Joanna M M; Ingelsson, Martin; Kathiresan, Sekar; Kee, Frank; Kuusisto, Johanna; Langenberg, Claudia; Launer, Lenore J; Lindgren, Cecilia M; Männistö, Satu; Meitinger, Thomas; Melander, Olle; Mohlke, Karen L; Moitry, Marie; Morris, Andrew D; Murray, Alison D; de Mutsert, Renée; Orho-Melander, Marju; Owen, Katharine R; Perola, Markus; Peters, Annette; Province, Michael A; Rasheed, Asif; Ridker, Paul M; Rivadineira, Fernando; Rosendaal, Frits R; Rosengren, Anders H; Salomaa, Veikko; Sheu, Wayne H-H; Sladek, Rob; Smith, Blair H; Strauch, Konstantin; Uitterlinden, André G; Varma, Rohit; Willer, Cristen J; Blüher, Matthias; Butterworth, Adam S; Chambers, John Campbell; Chasman, Daniel I; Danesh, John; van Duijn, Cornelia; Dupuis, Josée; Franco, Oscar H; Franks, Paul W; Froguel, Philippe; Grallert, Harald; Groop, Leif; Han, Bok-Ghee; Hansen, Torben; Hattersley, Andrew T; Hayward, Caroline; Ingelsson, Erik; Kardia, Sharon L R; Karpe, Fredrik; Kooner, Jaspal Singh; Köttgen, Anna; Kuulasmaa, Kari; Laakso, Markku; Lin, Xu; Lind, Lars; Liu, Yongmei; Loos, Ruth J F; Marchini, Jonathan; Metspalu, Andres; Mook-Kanamori, Dennis; Nordestgaard, Børge G; Palmer, Colin N A; Pankow, James S; Pedersen, Oluf; Psaty, Bruce M; Rauramaa, Rainer; Sattar, Naveed; Schulze, Matthias B; Soranzo, Nicole; Spector, Timothy D; Stefansson, Kari; Stumvoll, Michael; Thorsteinsdottir, Unnur; Tuomi, Tiinamaija; Tuomilehto, Jaakko; Wareham, Nicholas J; Wilson, James G; Zeggini, Eleftheria; Scott, Robert A; Barroso, Inês; Frayling, Timothy M; Goodarzi, Mark O; Meigs, James B; Boehnke, Michael; Saleheen, Danish; Morris, Andrew P; Rotter, Jerome I; McCarthy, Mark I

    2018-04-01

    We aggregated coding variant data for 81,412 type 2 diabetes cases and 370,832 controls of diverse ancestry, identifying 40 coding variant association signals (P < 2.2 × 10 -7 ); of these, 16 map outside known risk-associated loci. We make two important observations. First, only five of these signals are driven by low-frequency variants: even for these, effect sizes are modest (odds ratio ≤1.29). Second, when we used large-scale genome-wide association data to fine-map the associated variants in their regional context, accounting for the global enrichment of complex trait associations in coding sequence, compelling evidence for coding variant causality was obtained for only 16 signals. At 13 others, the associated coding variants clearly represent 'false leads' with potential to generate erroneous mechanistic inference. Coding variant associations offer a direct route to biological insight for complex diseases and identification of validated therapeutic targets; however, appropriate mechanistic inference requires careful specification of their causal contribution to disease predisposition.

  17. Employing machine learning for reliable miRNA target identification in plants

    PubMed Central

    2011-01-01

    Background miRNAs are ~21 nucleotide long small noncoding RNA molecules, formed endogenously in most of the eukaryotes, which mainly control their target genes post transcriptionally by interacting and silencing them. While a lot of tools has been developed for animal miRNA target system, plant miRNA target identification system has witnessed limited development. Most of them have been centered around exact complementarity match. Very few of them considered other factors like multiple target sites and role of flanking regions. Result In the present work, a Support Vector Regression (SVR) approach has been implemented for plant miRNA target identification, utilizing position specific dinucleotide density variation information around the target sites, to yield highly reliable result. It has been named as p-TAREF (plant-Target Refiner). Performance comparison for p-TAREF was done with other prediction tools for plants with utmost rigor and where p-TAREF was found better performing in several aspects. Further, p-TAREF was run over the experimentally validated miRNA targets from species like Arabidopsis, Medicago, Rice and Tomato, and detected them accurately, suggesting gross usability of p-TAREF for plant species. Using p-TAREF, target identification was done for the complete Rice transcriptome, supported by expression and degradome based data. miR156 was found as an important component of the Rice regulatory system, where control of genes associated with growth and transcription looked predominant. The entire methodology has been implemented in a multi-threaded parallel architecture in Java, to enable fast processing for web-server version as well as standalone version. This also makes it to run even on a simple desktop computer in concurrent mode. It also provides a facility to gather experimental support for predictions made, through on the spot expression data analysis, in its web-server version. Conclusion A machine learning multivariate feature tool has been

  18. Systematic Identification of Combinatorial Drivers and Targets in Cancer Cell Lines

    PubMed Central

    Tabchy, Adel; Eltonsy, Nevine; Housman, David E.; Mills, Gordon B.

    2013-01-01

    There is an urgent need to elicit and validate highly efficacious targets for combinatorial intervention from large scale ongoing molecular characterization efforts of tumors. We established an in silico bioinformatic platform in concert with a high throughput screening platform evaluating 37 novel targeted agents in 669 extensively characterized cancer cell lines reflecting the genomic and tissue-type diversity of human cancers, to systematically identify combinatorial biomarkers of response and co-actionable targets in cancer. Genomic biomarkers discovered in a 141 cell line training set were validated in an independent 359 cell line test set. We identified co-occurring and mutually exclusive genomic events that represent potential drivers and combinatorial targets in cancer. We demonstrate multiple cooperating genomic events that predict sensitivity to drug intervention independent of tumor lineage. The coupling of scalable in silico and biologic high throughput cancer cell line platforms for the identification of co-events in cancer delivers rational combinatorial targets for synthetic lethal approaches with a high potential to pre-empt the emergence of resistance. PMID:23577104

  19. Systematic identification of combinatorial drivers and targets in cancer cell lines.

    PubMed

    Tabchy, Adel; Eltonsy, Nevine; Housman, David E; Mills, Gordon B

    2013-01-01

    There is an urgent need to elicit and validate highly efficacious targets for combinatorial intervention from large scale ongoing molecular characterization efforts of tumors. We established an in silico bioinformatic platform in concert with a high throughput screening platform evaluating 37 novel targeted agents in 669 extensively characterized cancer cell lines reflecting the genomic and tissue-type diversity of human cancers, to systematically identify combinatorial biomarkers of response and co-actionable targets in cancer. Genomic biomarkers discovered in a 141 cell line training set were validated in an independent 359 cell line test set. We identified co-occurring and mutually exclusive genomic events that represent potential drivers and combinatorial targets in cancer. We demonstrate multiple cooperating genomic events that predict sensitivity to drug intervention independent of tumor lineage. The coupling of scalable in silico and biologic high throughput cancer cell line platforms for the identification of co-events in cancer delivers rational combinatorial targets for synthetic lethal approaches with a high potential to pre-empt the emergence of resistance.

  20. Patterns of Cognitive Strengths and Weaknesses: Identification Rates, Agreement, and Validity for Learning Disabilities Identification

    PubMed Central

    Miciak, Jeremy; Fletcher, Jack M.; Stuebing, Karla; Vaughn, Sharon; Tolar, Tammy D.

    2014-01-01

    Purpose Few empirical investigations have evaluated LD identification methods based on a pattern of cognitive strengths and weaknesses (PSW). This study investigated the reliability and validity of two proposed PSW methods: the concordance/discordance method (C/DM) and cross battery assessment (XBA) method. Methods Cognitive assessment data for 139 adolescents demonstrating inadequate response to intervention was utilized to empirically classify participants as meeting or not meeting PSW LD identification criteria using the two approaches, permitting an analysis of: (1) LD identification rates; (2) agreement between methods; and (3) external validity. Results LD identification rates varied between the two methods depending upon the cut point for low achievement, with low agreement for LD identification decisions. Comparisons of groups that met and did not meet LD identification criteria on external academic variables were largely null, raising questions of external validity. Conclusions This study found low agreement and little evidence of validity for LD identification decisions based on PSW methods. An alternative may be to use multiple measures of academic achievement to guide intervention. PMID:24274155

  1. Recommendations for Improving Identification and Quantification in Non-Targeted, GC-MS-Based Metabolomic Profiling of Human Plasma

    PubMed Central

    Wang, Hanghang; Muehlbauer, Michael J.; O’Neal, Sara K.; Newgard, Christopher B.; Hauser, Elizabeth R.; Shah, Svati H.

    2017-01-01

    The field of metabolomics as applied to human disease and health is rapidly expanding. In recent efforts of metabolomics research, greater emphasis has been placed on quality control and method validation. In this study, we report an experience with quality control and a practical application of method validation. Specifically, we sought to identify and modify steps in gas chromatography-mass spectrometry (GC-MS)-based, non-targeted metabolomic profiling of human plasma that could influence metabolite identification and quantification. Our experimental design included two studies: (1) a limiting-dilution study, which investigated the effects of dilution on analyte identification and quantification; and (2) a concentration-specific study, which compared the optimal plasma extract volume established in the first study with the volume used in the current institutional protocol. We confirmed that contaminants, concentration, repeatability and intermediate precision are major factors influencing metabolite identification and quantification. In addition, we established methods for improved metabolite identification and quantification, which were summarized to provide recommendations for experimental design of GC-MS-based non-targeted profiling of human plasma. PMID:28841195

  2. TargetMiner: microRNA target prediction with systematic identification of tissue-specific negative examples.

    PubMed

    Bandyopadhyay, Sanghamitra; Mitra, Ramkrishna

    2009-10-15

    Prediction of microRNA (miRNA) target mRNAs using machine learning approaches is an important area of research. However, most of the methods suffer from either high false positive or false negative rates. One reason for this is the marked deficiency of negative examples or miRNA non-target pairs. Systematic identification of non-target mRNAs is still not addressed properly, and therefore, current machine learning approaches are compelled to rely on artificially generated negative examples for training. In this article, we have identified approximately 300 tissue-specific negative examples using a novel approach that involves expression profiling of both miRNAs and mRNAs, miRNA-mRNA structural interactions and seed-site conservation. The newly generated negative examples are validated with pSILAC dataset, which elucidate the fact that the identified non-targets are indeed non-targets.These high-throughput tissue-specific negative examples and a set of experimentally verified positive examples are then used to build a system called TargetMiner, a support vector machine (SVM)-based classifier. In addition to assessing the prediction accuracy on cross-validation experiments, TargetMiner has been validated with a completely independent experimental test dataset. Our method outperforms 10 existing target prediction algorithms and provides a good balance between sensitivity and specificity that is not reflected in the existing methods. We achieve a significantly higher sensitivity and specificity of 69% and 67.8% based on a pool of 90 feature set and 76.5% and 66.1% using a set of 30 selected feature set on the completely independent test dataset. In order to establish the effectiveness of the systematically generated negative examples, the SVM is trained using a different set of negative data generated using the method in Yousef et al. A significantly higher false positive rate (70.6%) is observed when tested on the independent set, while all other factors are kept the

  3. 2-Aryl-5-carboxytetrazole as a New Photoaffinity Label for Drug Target Identification

    PubMed Central

    2016-01-01

    Photoaffinity labels are powerful tools for dissecting ligand–protein interactions, and they have a broad utility in medicinal chemistry and drug discovery. Traditional photoaffinity labels work through nonspecific C–H/X–H bond insertion reactions with the protein of interest by the highly reactive photogenerated intermediate. Herein, we report a new photoaffinity label, 2-aryl-5-carboxytetrazole (ACT), that interacts with the target protein via a unique mechanism in which the photogenerated carboxynitrile imine reacts with a proximal nucleophile near the target active site. In two distinct case studies, we demonstrate that the attachment of ACT to a ligand does not significantly alter the binding affinity and specificity of the parent drug. Compared with diazirine and benzophenone, two commonly used photoaffinity labels, in two case studies ACT showed higher photo-cross-linking yields toward their protein targets in vitro based on mass spectrometry analysis. In the in situ target identification studies, ACT successfully captured the desired targets with an efficiency comparable to the diazirine. We expect that further development of this class of photoaffinity labels will lead to a broad range of applications across target identification, and validation and elucidation of the binding site in drug discovery. PMID:27740749

  4. 2-Aryl-5-carboxytetrazole as a New Photoaffinity Label for Drug Target Identification.

    PubMed

    Herner, András; Marjanovic, Jasmina; Lewandowski, Tracey M; Marin, Violeta; Patterson, Melanie; Miesbauer, Laura; Ready, Damien; Williams, Jon; Vasudevan, Anil; Lin, Qing

    2016-11-09

    Photoaffinity labels are powerful tools for dissecting ligand-protein interactions, and they have a broad utility in medicinal chemistry and drug discovery. Traditional photoaffinity labels work through nonspecific C-H/X-H bond insertion reactions with the protein of interest by the highly reactive photogenerated intermediate. Herein, we report a new photoaffinity label, 2-aryl-5-carboxytetrazole (ACT), that interacts with the target protein via a unique mechanism in which the photogenerated carboxynitrile imine reacts with a proximal nucleophile near the target active site. In two distinct case studies, we demonstrate that the attachment of ACT to a ligand does not significantly alter the binding affinity and specificity of the parent drug. Compared with diazirine and benzophenone, two commonly used photoaffinity labels, in two case studies ACT showed higher photo-cross-linking yields toward their protein targets in vitro based on mass spectrometry analysis. In the in situ target identification studies, ACT successfully captured the desired targets with an efficiency comparable to the diazirine. We expect that further development of this class of photoaffinity labels will lead to a broad range of applications across target identification, and validation and elucidation of the binding site in drug discovery.

  5. Identification of control targets in Boolean molecular network models via computational algebra.

    PubMed

    Murrugarra, David; Veliz-Cuba, Alan; Aguilar, Boris; Laubenbacher, Reinhard

    2016-09-23

    Many problems in biomedicine and other areas of the life sciences can be characterized as control problems, with the goal of finding strategies to change a disease or otherwise undesirable state of a biological system into another, more desirable, state through an intervention, such as a drug or other therapeutic treatment. The identification of such strategies is typically based on a mathematical model of the process to be altered through targeted control inputs. This paper focuses on processes at the molecular level that determine the state of an individual cell, involving signaling or gene regulation. The mathematical model type considered is that of Boolean networks. The potential control targets can be represented by a set of nodes and edges that can be manipulated to produce a desired effect on the system. This paper presents a method for the identification of potential intervention targets in Boolean molecular network models using algebraic techniques. The approach exploits an algebraic representation of Boolean networks to encode the control candidates in the network wiring diagram as the solutions of a system of polynomials equations, and then uses computational algebra techniques to find such controllers. The control methods in this paper are validated through the identification of combinatorial interventions in the signaling pathways of previously reported control targets in two well studied systems, a p53-mdm2 network and a blood T cell lymphocyte granular leukemia survival signaling network. Supplementary data is available online and our code in Macaulay2 and Matlab are available via http://www.ms.uky.edu/~dmu228/ControlAlg . This paper presents a novel method for the identification of intervention targets in Boolean network models. The results in this paper show that the proposed methods are useful and efficient for moderately large networks.

  6. Validating models of target acquisition performance in the dismounted soldier context

    NASA Astrophysics Data System (ADS)

    Glaholt, Mackenzie G.; Wong, Rachel K.; Hollands, Justin G.

    2018-04-01

    The problem of predicting real-world operator performance with digital imaging devices is of great interest within the military and commercial domains. There are several approaches to this problem, including: field trials with imaging devices, laboratory experiments using imagery captured from these devices, and models that predict human performance based on imaging device parameters. The modeling approach is desirable, as both field trials and laboratory experiments are costly and time-consuming. However, the data from these experiments is required for model validation. Here we considered this problem in the context of dismounted soldiering, for which detection and identification of human targets are essential tasks. Human performance data were obtained for two-alternative detection and identification decisions in a laboratory experiment in which photographs of human targets were presented on a computer monitor and the images were digitally magnified to simulate range-to-target. We then compared the predictions of different performance models within the NV-IPM software package: Targeting Task Performance (TTP) metric model and the Johnson model. We also introduced a modification to the TTP metric computation that incorporates an additional correction for target angular size. We examined model predictions using NV-IPM default values for a critical model constant, V50, and we also considered predictions when this value was optimized to fit the behavioral data. When using default values, certain model versions produced a reasonably close fit to the human performance data in the detection task, while for the identification task all models substantially overestimated performance. When using fitted V50 values the models produced improved predictions, though the slopes of the performance functions were still shallow compared to the behavioral data. These findings are discussed in relation to the models' designs and parameters, and the characteristics of the behavioral

  7. Automatic target validation based on neuroscientific literature mining for tractography

    PubMed Central

    Vasques, Xavier; Richardet, Renaud; Hill, Sean L.; Slater, David; Chappelier, Jean-Cedric; Pralong, Etienne; Bloch, Jocelyne; Draganski, Bogdan; Cif, Laura

    2015-01-01

    Target identification for tractography studies requires solid anatomical knowledge validated by an extensive literature review across species for each seed structure to be studied. Manual literature review to identify targets for a given seed region is tedious and potentially subjective. Therefore, complementary approaches would be useful. We propose to use text-mining models to automatically suggest potential targets from the neuroscientific literature, full-text articles and abstracts, so that they can be used for anatomical connection studies and more specifically for tractography. We applied text-mining models to three structures: two well-studied structures, since validated deep brain stimulation targets, the internal globus pallidus and the subthalamic nucleus and, the nucleus accumbens, an exploratory target for treating psychiatric disorders. We performed a systematic review of the literature to document the projections of the three selected structures and compared it with the targets proposed by text-mining models, both in rat and primate (including human). We ran probabilistic tractography on the nucleus accumbens and compared the output with the results of the text-mining models and literature review. Overall, text-mining the literature could find three times as many targets as two man-weeks of curation could. The overall efficiency of the text-mining against literature review in our study was 98% recall (at 36% precision), meaning that over all the targets for the three selected seeds, only one target has been missed by text-mining. We demonstrate that connectivity for a structure of interest can be extracted from a very large amount of publications and abstracts. We believe this tool will be useful in helping the neuroscience community to facilitate connectivity studies of particular brain regions. The text mining tools used for the study are part of the HBP Neuroinformatics Platform, publicly available at http://connectivity-brainer.rhcloud.com/. PMID

  8. Detection and Identification of Multiple Stationary Human Targets Via Bio-Radar Based on the Cross-Correlation Method

    PubMed Central

    Zhang, Yang; Chen, Fuming; Xue, Huijun; Li, Zhao; An, Qiang; Wang, Jianqi; Zhang, Yang

    2016-01-01

    Ultra-wideband (UWB) radar has been widely used for detecting human physiological signals (respiration, movement, etc.) in the fields of rescue, security, and medicine owing to its high penetrability and range resolution. In these applications, especially in rescue after disaster (earthquake, collapse, mine accident, etc.), the presence, number, and location of the trapped victims to be detected and rescued are the key issues of concern. Ample research has been done on the first issue, whereas the identification and localization of multi-targets remains a challenge. False positive and negative identification results are two common problems associated with the detection of multiple stationary human targets. This is mainly because the energy of the signal reflected from the target close to the receiving antenna is considerably stronger than those of the targets at further range, often leading to missing or false recognition if the identification method is based on the energy of the respiratory signal. Therefore, a novel method based on cross-correlation is proposed in this paper that is based on the relativity and periodicity of the signals, rather than on the energy. The validity of this method is confirmed through experiments using different scenarios; the results indicate a discernible improvement in the detection precision and identification of the multiple stationary targets. PMID:27801795

  9. Detection and Identification of Multiple Stationary Human Targets Via Bio-Radar Based on the Cross-Correlation Method.

    PubMed

    Zhang, Yang; Chen, Fuming; Xue, Huijun; Li, Zhao; An, Qiang; Wang, Jianqi; Zhang, Yang

    2016-10-27

    Ultra-wideband (UWB) radar has been widely used for detecting human physiological signals (respiration, movement, etc.) in the fields of rescue, security, and medicine owing to its high penetrability and range resolution. In these applications, especially in rescue after disaster (earthquake, collapse, mine accident, etc.), the presence, number, and location of the trapped victims to be detected and rescued are the key issues of concern. Ample research has been done on the first issue, whereas the identification and localization of multi-targets remains a challenge. False positive and negative identification results are two common problems associated with the detection of multiple stationary human targets. This is mainly because the energy of the signal reflected from the target close to the receiving antenna is considerably stronger than those of the targets at further range, often leading to missing or false recognition if the identification method is based on the energy of the respiratory signal. Therefore, a novel method based on cross-correlation is proposed in this paper that is based on the relativity and periodicity of the signals, rather than on the energy. The validity of this method is confirmed through experiments using different scenarios; the results indicate a discernible improvement in the detection precision and identification of the multiple stationary targets.

  10. Network-assisted target identification for haploinsufficiency and homozygous profiling screens

    PubMed Central

    Wang, Sheng

    2017-01-01

    Chemical genomic screens have recently emerged as a systematic approach to drug discovery on a genome-wide scale. Drug target identification and elucidation of the mechanism of action (MoA) of hits from these noisy high-throughput screens remain difficult. Here, we present GIT (Genetic Interaction Network-Assisted Target Identification), a network analysis method for drug target identification in haploinsufficiency profiling (HIP) and homozygous profiling (HOP) screens. With the drug-induced phenotypic fitness defect of the deletion of a gene, GIT also incorporates the fitness defects of the gene’s neighbors in the genetic interaction network. On three genome-scale yeast chemical genomic screens, GIT substantially outperforms previous scoring methods on target identification on HIP and HOP assays, respectively. Finally, we showed that by combining HIP and HOP assays, GIT further boosts target identification and reveals potential drug’s mechanism of action. PMID:28574983

  11. Identification and location tasks rely on different mental processes: a diffusion model account of validity effects in spatial cueing paradigms with emotional stimuli.

    PubMed

    Imhoff, Roland; Lange, Jens; Germar, Markus

    2018-02-22

    Spatial cueing paradigms are popular tools to assess human attention to emotional stimuli, but different variants of these paradigms differ in what participants' primary task is. In one variant, participants indicate the location of the target (location task), whereas in the other they indicate the shape of the target (identification task). In the present paper we test the idea that although these two variants produce seemingly comparable cue validity effects on response times, they rest on different underlying processes. Across four studies (total N = 397; two in the supplement) using both variants and manipulating the motivational relevance of cue content, diffusion model analyses revealed that cue validity effects in location tasks are primarily driven by response biases, whereas the same effect rests on delay due to attention to the cue in identification tasks. Based on this, we predict and empirically support that a symmetrical distribution of valid and invalid cues would reduce cue validity effects in location tasks to a greater extent than in identification tasks. Across all variants of the task, we fail to replicate the effect of greater cue validity effects for arousing (vs. neutral) stimuli. We discuss the implications of these findings for best practice in spatial cueing research.

  12. Camouflage, detection and identification of moving targets

    PubMed Central

    Hall, Joanna R.; Cuthill, Innes C.; Baddeley, Roland; Shohet, Adam J.; Scott-Samuel, Nicholas E.

    2013-01-01

    Nearly all research on camouflage has investigated its effectiveness for concealing stationary objects. However, animals have to move, and patterns that only work when the subject is static will heavily constrain behaviour. We investigated the effects of different camouflages on the three stages of predation—detection, identification and capture—in a computer-based task with humans. An initial experiment tested seven camouflage strategies on static stimuli. In line with previous literature, background-matching and disruptive patterns were found to be most successful. Experiment 2 showed that if stimuli move, an isolated moving object on a stationary background cannot avoid detection or capture regardless of the type of camouflage. Experiment 3 used an identification task and showed that while camouflage is unable to slow detection or capture, camouflaged targets are harder to identify than uncamouflaged targets when similar background objects are present. The specific details of the camouflage patterns have little impact on this effect. If one has to move, camouflage cannot impede detection; but if one is surrounded by similar targets (e.g. other animals in a herd, or moving background distractors), then camouflage can slow identification. Despite previous assumptions, motion does not entirely ‘break’ camouflage. PMID:23486439

  13. Camouflage, detection and identification of moving targets.

    PubMed

    Hall, Joanna R; Cuthill, Innes C; Baddeley, Roland; Shohet, Adam J; Scott-Samuel, Nicholas E

    2013-05-07

    Nearly all research on camouflage has investigated its effectiveness for concealing stationary objects. However, animals have to move, and patterns that only work when the subject is static will heavily constrain behaviour. We investigated the effects of different camouflages on the three stages of predation-detection, identification and capture-in a computer-based task with humans. An initial experiment tested seven camouflage strategies on static stimuli. In line with previous literature, background-matching and disruptive patterns were found to be most successful. Experiment 2 showed that if stimuli move, an isolated moving object on a stationary background cannot avoid detection or capture regardless of the type of camouflage. Experiment 3 used an identification task and showed that while camouflage is unable to slow detection or capture, camouflaged targets are harder to identify than uncamouflaged targets when similar background objects are present. The specific details of the camouflage patterns have little impact on this effect. If one has to move, camouflage cannot impede detection; but if one is surrounded by similar targets (e.g. other animals in a herd, or moving background distractors), then camouflage can slow identification. Despite previous assumptions, motion does not entirely 'break' camouflage.

  14. Categorization and identification of simultaneous targets.

    PubMed

    Theeuwes, J

    1991-02-01

    Early and late selection theories of visual attention disagree about whether identification occurs before or after selection. Studies showing the category effect, i.e., the time to detect a letter is hardly affected by the number of digits present in the display, are taken as evidence for late selection theories since these studies suggest parallel identification of all items in the display. As an extension of previous studies, in the present study two categorically different targets were presented simultaneously among a variable number of nontargets. Subjects were shown brief displays of two target letters among either 2, 4 or 6 nontarget digits. Subjects responded 'same' when the two letters were identical and 'different' otherwise. Since the 'same-different' response reflects the combined outcome of the simultaneous targets, late-selection theory predicts that the time to match the target letters is independent of the number of nontarget digits. Alternatively, early-selection theory predicts a linear increase of reaction time with display size since the presence of more than one target disrupts parallel preattentive processing, leading to a serial search through all items in the display. The results provide evidence for the early-selection view since reaction time increased linearly with the number of categorically different nontargets. A control experiment revealed that none of the alternative explanations could account for the display size effect.

  15. Identification of human microRNA targets from isolated argonaute protein complexes.

    PubMed

    Beitzinger, Michaela; Peters, Lasse; Zhu, Jia Yun; Kremmer, Elisabeth; Meister, Gunter

    2007-06-01

    MicroRNAs (miRNAs) constitute a class of small non-coding RNAs that regulate gene expression on the level of translation and/or mRNA stability. Mammalian miRNAs associate with members of the Argonaute (Ago) protein family and bind to partially complementary sequences in the 3' untranslated region (UTR) of specific target mRNAs. Computer algorithms based on factors such as free binding energy or sequence conservation have been used to predict miRNA target mRNAs. Based on such predictions, up to one third of all mammalian mRNAs seem to be under miRNA regulation. However, due to the low degree of complementarity between the miRNA and its target, such computer programs are often imprecise and therefore not very reliable. Here we report the first biochemical identification approach of miRNA targets from human cells. Using highly specific monoclonal antibodies against members of the Ago protein family, we co-immunoprecipitate Ago-bound mRNAs and identify them by cloning. Interestingly, most of the identified targets are also predicted by different computer programs. Moreover, we randomly analyzed six different target candidates and were able to experimentally validate five as miRNA targets. Our data clearly indicate that miRNA targets can be experimentally identified from Ago complexes and therefore provide a new tool to directly analyze miRNA function.

  16. Atlas-based identification of targets for functional radiosurgery

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Stancanello, Joseph; Romanelli, Pantaleo; Modugno, Nicola

    2006-06-15

    Functional disorders of the brain, such as Parkinson's disease, dystonia, epilepsy, and neuropathic pain, may exhibit poor response to medical therapy. In such cases, surgical intervention may become necessary. Modern surgical approaches to such disorders include radio-frequency lesioning and deep brain stimulation (DBS). The subthalamic nucleus (STN) is one of the most useful stereotactic targets available: STN DBS is known to induce substantial improvement in patients with end-stage Parkinson's disease. Other targets include the Globus Pallidus pars interna (GPi) for dystonia and Parkinson's disease, and the centromedian nucleus of the thalamus (CMN) for neuropathic pain. Radiosurgery is an attractive noninvasivemore » alternative to treat some functional brain disorders. The main technical limitation to radiosurgery is that the target can be selected only on the basis of magnetic resonance anatomy without electrophysiological confirmation. The aim of this work is to provide a method for the correct atlas-based identification of the target to be used in functional neurosurgery treatment planning. The coordinates of STN, CMN, and GPi were identified in the Talairach and Tournoux atlas and transformed to the corresponding regions of the Montreal Neurological Institute (MNI) electronic atlas. Binary masks describing the target nuclei were created. The MNI electronic atlas was deformed onto the patient magnetic resonance imaging-T1 scan by applying an affine transformation followed by a local nonrigid registration. The first transformation was based on normalized cross correlation and the second on optimization of a two-part objective function consisting of similarity criteria and weighted regularization. The obtained deformation field was then applied to the target masks. The minimum distance between the surface of an implanted electrode and the surface of the deformed mask was calculated. The validation of the method consisted of comparing the electrode

  17. Detection and identification of human targets in radar data

    NASA Astrophysics Data System (ADS)

    Gürbüz, Sevgi Z.; Melvin, William L.; Williams, Douglas B.

    2007-04-01

    Radar offers unique advantages over other sensors, such as visual or seismic sensors, for human target detection. Many situations, especially military applications, prevent the placement of video cameras or implantment seismic sensors in the area being observed, because of security or other threats. However, radar can operate far away from potential targets, and functions during daytime as well as nighttime, in virtually all weather conditions. In this paper, we examine the problem of human target detection and identification using single-channel, airborne, synthetic aperture radar (SAR). Human targets are differentiated from other detected slow-moving targets by analyzing the spectrogram of each potential target. Human spectrograms are unique, and can be used not just to identify targets as human, but also to determine features about the human target being observed, such as size, gender, action, and speed. A 12-point human model, together with kinematic equations of motion for each body part, is used to calculate the expected target return and spectrogram. A MATLAB simulation environment is developed including ground clutter, human and non-human targets for the testing of spectrogram-based detection and identification algorithms. Simulations show that spectrograms have some ability to detect and identify human targets in low noise. An example gender discrimination system correctly detected 83.97% of males and 91.11% of females. The problems and limitations of spectrogram-based methods in high clutter environments are discussed. The SNR loss inherent to spectrogram-based methods is quantified. An alternate detection and identification method that will be used as a basis for future work is proposed.

  18. Identification and Validation of ESP Teacher Competencies: A Research Design

    ERIC Educational Resources Information Center

    Venkatraman, G.; Prema, P.

    2013-01-01

    The paper presents the research design used for identifying and validating a set of competencies required of ESP (English for Specific Purposes) teachers. The identification of the competencies and the three-stage validation process are also discussed. The observation of classes of ESP teachers for field-testing the validated competencies and…

  19. Target Abundance-Based Fitness Screening (TAFiS) Facilitates Rapid Identification of Target-Specific and Physiologically Active Chemical Probes

    PubMed Central

    Butts, Arielle; DeJarnette, Christian; Peters, Tracy L.; Parker, Josie E.; Kerns, Morgan E.; Eberle, Karen E.; Kelly, Steve L.

    2017-01-01

    -generation target-based whole-cell screening approach that incorporates the principles of both chemical genetics and competitive fitness, which enables the identification of target-specific and physiologically active compounds from a single screen. We have chosen to validate this approach using the important human fungal pathogen Candida albicans with the intention of pursuing novel antifungal targets. However, this approach is broadly applicable and is expected to dramatically reduce the time and resources required to progress from screening hit to lead compound. PMID:28989971

  20. Target identification for small bioactive molecules: finding the needle in the haystack.

    PubMed

    Ziegler, Slava; Pries, Verena; Hedberg, Christian; Waldmann, Herbert

    2013-03-04

    Identification and confirmation of bioactive small-molecule targets is a crucial, often decisive step both in academic and pharmaceutical research. Through the development and availability of several new experimental techniques, target identification is, in principle, feasible, and the number of successful examples steadily grows. However, a generic methodology that can successfully be applied in the majority of the cases has not yet been established. Herein we summarize current methods for target identification of small molecules, primarily for a chemistry audience but also the biological community, for example, the chemist or biologist attempting to identify the target of a given bioactive compound. We describe the most frequently employed experimental approaches for target identification and provide several representative examples illustrating the state-of-the-art. Among the techniques currently available, protein affinity isolation using suitable small-molecule probes (pulldown) and subsequent mass spectrometric analysis of the isolated proteins appears to be most powerful and most frequently applied. To provide guidance for rapid entry into the field and based on our own experience we propose a typical workflow for target identification, which centers on the application of chemical proteomics as the key step to generate hypotheses for potential target proteins. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Molecular identification of python species: development and validation of a novel assay for forensic investigations.

    PubMed

    Ciavaglia, Sherryn A; Tobe, Shanan S; Donnellan, Stephen C; Henry, Julianne M; Linacre, Adrian M T

    2015-05-01

    Python snake species are often encountered in illegal activities and the question of species identity can be pertinent to such criminal investigations. Morphological identification of species of pythons can be confounded by many issues and molecular examination by DNA analysis can provide an alternative and objective means of identification. Our paper reports on the development and validation of a PCR primer pair that amplifies a segment of the mitochondrial cytochrome b gene that has been suggested previously as a good candidate locus for differentiating python species. We used this DNA region to perform species identification of pythons, even when the template DNA was of poor quality, as might be the case with forensic evidentiary items. Validation tests are presented to demonstrate the characteristics of the assay. Tests involved the cross-species amplification of this marker in non-target species, minimum amount of DNA template required, effects of degradation on product amplification and a blind trial to simulate a casework scenario that provided 100% correct identity. Our results demonstrate that this assay performs reliably and robustly on pythons and can be applied directly to forensic investigations where the presence of a species of python is in question. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  2. Plant microRNA-Target Interaction Identification Model Based on the Integration of Prediction Tools and Support Vector Machine

    PubMed Central

    Meng, Jun; Shi, Lin; Luan, Yushi

    2014-01-01

    Background Confident identification of microRNA-target interactions is significant for studying the function of microRNA (miRNA). Although some computational miRNA target prediction methods have been proposed for plants, results of various methods tend to be inconsistent and usually lead to more false positive. To address these issues, we developed an integrated model for identifying plant miRNA–target interactions. Results Three online miRNA target prediction toolkits and machine learning algorithms were integrated to identify and analyze Arabidopsis thaliana miRNA-target interactions. Principle component analysis (PCA) feature extraction and self-training technology were introduced to improve the performance. Results showed that the proposed model outperformed the previously existing methods. The results were validated by using degradome sequencing supported Arabidopsis thaliana miRNA-target interactions. The proposed model constructed on Arabidopsis thaliana was run over Oryza sativa and Vitis vinifera to demonstrate that our model is effective for other plant species. Conclusions The integrated model of online predictors and local PCA-SVM classifier gained credible and high quality miRNA-target interactions. The supervised learning algorithm of PCA-SVM classifier was employed in plant miRNA target identification for the first time. Its performance can be substantially improved if more experimentally proved training samples are provided. PMID:25051153

  3. Target specific compound identification using a support vector machine.

    PubMed

    Plewczynski, Dariusz; von Grotthuss, Marcin; Spieser, Stephane A H; Rychlewski, Leszek; Wyrwicz, Lucjan S; Ginalski, Krzysztof; Koch, Uwe

    2007-03-01

    In many cases at the beginning of an HTS-campaign, some information about active molecules is already available. Often known active compounds (such as substrate analogues, natural products, inhibitors of a related protein or ligands published by a pharmaceutical company) are identified in low-throughput validation studies of the biochemical target. In this study we evaluate the effectiveness of a support vector machine applied for those compounds and used to classify a collection with unknown activity. This approach was aimed at reducing the number of compounds to be tested against the given target. Our method predicts the biological activity of chemical compounds based on only the atom pairs (AP) two dimensional topological descriptors. The supervised support vector machine (SVM) method herein is trained on compounds from the MDL drug data report (MDDR) known to be active for specific protein target. For detailed analysis, five different biological targets were selected including cyclooxygenase-2, dihydrofolate reductase, thrombin, HIV-reverse transcriptase and antagonists of the estrogen receptor. The accuracy of compound identification was estimated using the recall and precision values. The sensitivities for all protein targets exceeded 80% and the classification performance reached 100% for selected targets. In another application of the method, we addressed the absence of an initial set of active compounds for a selected protein target at the beginning of an HTS-campaign. In such a case, virtual high-throughput screening (vHTS) is usually applied by using a flexible docking procedure. However, the vHTS experiment typically contains a large percentage of false positives that should be verified by costly and time-consuming experimental follow-up assays. The subsequent use of our machine learning method was found to improve the speed (since the docking procedure was not required for all compounds from the database) and also the accuracy of the HTS hit lists (the

  4. Validated MicroRNA Target Databases: An Evaluation.

    PubMed

    Lee, Yun Ji Diana; Kim, Veronica; Muth, Dillon C; Witwer, Kenneth W

    2015-11-01

    Preclinical Research Positive findings from preclinical and clinical studies involving depletion or supplementation of microRNA (miRNA) engender optimism about miRNA-based therapeutics. However, off-target effects must be considered. Predicting these effects is complicated. Each miRNA may target many gene transcripts, and the rules governing imperfectly complementary miRNA: target interactions are incompletely understood. Several databases provide lists of the relatively small number of experimentally confirmed miRNA: target pairs. Although incomplete, this information might allow assessment of at least some of the off-target effects. We evaluated the performance of four databases of experimentally validated miRNA: target interactions (miRWalk 2.0, miRTarBase, miRecords, and TarBase 7.0) using a list of 50 alphabetically consecutive genes. We examined the provided citations to determine the degree to which each interaction was experimentally supported. To assess stability, we tested at the beginning and end of a five-month period. Results varied widely by database. Two of the databases changed significantly over the course of 5 months. Most reported evidence for miRNA: target interactions were indirect or otherwise weak, and relatively few interactions were supported by more than one publication. Some returned results appear to arise from simplistic text searches that offer no insight into the relationship of the search terms, may not even include the reported gene or miRNA, and may thus, be invalid. We conclude that validation databases provide important information, but not all information in all extant databases is up-to-date or accurate. Nevertheless, the more comprehensive validation databases may provide useful starting points for investigation of off-target effects of proposed small RNA therapies. © 2015 Wiley Periodicals, Inc.

  5. Genome-Wide Identification of CBX2 Targets: Insights in the Human Sex Development Network

    PubMed Central

    Eid, Wassim; Opitz, Lennart

    2015-01-01

    Chromobox homolog 2 (CBX2) is a chromatin modifier that plays an important role in sexual development and its disorders (disorders of sex development [DSD]), yet the exact rank and function of human CBX2 in this pathway remains unclear. Here, we performed large-scale mapping and analysis of in vivo target loci of the protein CBX2 in Sertoli-like NT-2D1 cells, using the DNA adenine methyltransferase identification technique. We identified close to 1600 direct targets for CBX2. Intriguingly, validation of selected candidate genes using qRT-PCR in cells overexpressing CBX2 or in which CBX2 has been knocked down indicated that several CBX2-responsive genes encode proteins that are involved in DSD. We further validated these effects on the candidate genes using a mutated CBX2 causing DSD in human patient. Overall, our findings suggest that CBX2 role in the sex development cascade is to stimulate the male pathway and concurrently inhibit the female pathway. These data provide fundamental insights into potential etiology of DSD. PMID:25569159

  6. Identification of Direct Target Genes Using Joint Sequence and Expression Likelihood with Application to DAF-16

    PubMed Central

    Yu, Ron X.; Liu, Jie; True, Nick; Wang, Wei

    2008-01-01

    A major challenge in the post-genome era is to reconstruct regulatory networks from the biological knowledge accumulated up to date. The development of tools for identifying direct target genes of transcription factors (TFs) is critical to this endeavor. Given a set of microarray experiments, a probabilistic model called TRANSMODIS has been developed which can infer the direct targets of a TF by integrating sequence motif, gene expression and ChIP-chip data. The performance of TRANSMODIS was first validated on a set of transcription factor perturbation experiments (TFPEs) involving Pho4p, a well studied TF in Saccharomyces cerevisiae. TRANSMODIS removed elements of arbitrariness in manual target gene selection process and produced results that concur with one's intuition. TRANSMODIS was further validated on a genome-wide scale by comparing it with two other methods in Saccharomyces cerevisiae. The usefulness of TRANSMODIS was then demonstrated by applying it to the identification of direct targets of DAF-16, a critical TF regulating ageing in Caenorhabditis elegans. We found that 189 genes were tightly regulated by DAF-16. In addition, DAF-16 has differential preference for motifs when acting as an activator or repressor, which awaits experimental verification. TRANSMODIS is computationally efficient and robust, making it a useful probabilistic framework for finding immediate targets. PMID:18350157

  7. Treponema pallidum Putative Novel Drug Target Identification and Validation: Rethinking Syphilis Therapeutics with Plant-Derived Terpenoids

    PubMed Central

    Tiwari, Sameeksha; Singh, Priyanka; Singh, Swati; Awasthi, Manika; Pandey, Veda P.

    2015-01-01

    Abstract Syphilis, a slow progressive and the third most common sexually transmitted disease found worldwide, is caused by a spirochete gram negative bacteria Treponema pallidum. Emergence of antibiotic resistant T. pallidum has led to a search for novel drugs and their targets. Subtractive genomics analyses of pathogen T. pallidum and host Homo sapiens resulted in identification of 126 proteins essential for survival and viability of the pathogen. Metabolic pathway analyses of these essential proteins led to discovery of nineteen proteins distributed among six metabolic pathways unique to T. pallidum. One hundred plant-derived terpenoids, as potential therapeutic molecules against T. pallidum, were screened for their drug likeness and ADMET (absorption, distribution, metabolism, and toxicity) properties. Subsequently the resulting nine terpenoids were docked with five unique T. pallidum targets through molecular modeling approaches. Out of five targets analyzed, D-alanine:D-alanine ligase was found to be the most promising target, while terpenoid salvicine was the most potent inhibitor. A comparison of the inhibitory potential of the best docked readily available natural compound, namely pomiferin (flavonoid) with that of the best docked terpenoid salvicine, revealed that salvicine was a more potent inhibitor than that of pomiferin. To the best of our knowledge, this is the first report of a terpenoid as a potential therapeutic molecule against T. pallidum with D-alanine:D-alanine ligase as a novel target. Further studies are warranted to evaluate and explore the potential clinical ramifications of these findings in relation to syphilis that has public health importance worldwide. PMID:25683888

  8. C-mii: a tool for plant miRNA and target identification.

    PubMed

    Numnark, Somrak; Mhuantong, Wuttichai; Ingsriswang, Supawadee; Wichadakul, Duangdao

    2012-01-01

    MicroRNAs (miRNAs) have been known to play an important role in several biological processes in both animals and plants. Although several tools for miRNA and target identification are available, the number of tools tailored towards plants is limited, and those that are available have specific functionality, lack graphical user interfaces, and restrict the number of input sequences. Large-scale computational identifications of miRNAs and/or targets of several plants have been also reported. Their methods, however, are only described as flow diagrams, which require programming skills and the understanding of input and output of the connected programs to reproduce. To overcome these limitations and programming complexities, we proposed C-mii as a ready-made software package for both plant miRNA and target identification. C-mii was designed and implemented based on established computational steps and criteria derived from previous literature with the following distinguishing features. First, software is easy to install with all-in-one programs and packaged databases. Second, it comes with graphical user interfaces (GUIs) for ease of use. Users can identify plant miRNAs and targets via step-by-step execution, explore the detailed results from each step, filter the results according to proposed constraints in plant miRNA and target biogenesis, and export sequences and structures of interest. Third, it supplies bird's eye views of the identification results with infographics and grouping information. Fourth, in terms of functionality, it extends the standard computational steps of miRNA target identification with miRNA-target folding and GO annotation. Fifth, it provides helper functions for the update of pre-installed databases and automatic recovery. Finally, it supports multi-project and multi-thread management. C-mii constitutes the first complete software package with graphical user interfaces enabling computational identification of both plant miRNA genes and mi

  9. C-mii: a tool for plant miRNA and target identification

    PubMed Central

    2012-01-01

    Background MicroRNAs (miRNAs) have been known to play an important role in several biological processes in both animals and plants. Although several tools for miRNA and target identification are available, the number of tools tailored towards plants is limited, and those that are available have specific functionality, lack graphical user interfaces, and restrict the number of input sequences. Large-scale computational identifications of miRNAs and/or targets of several plants have been also reported. Their methods, however, are only described as flow diagrams, which require programming skills and the understanding of input and output of the connected programs to reproduce. Results To overcome these limitations and programming complexities, we proposed C-mii as a ready-made software package for both plant miRNA and target identification. C-mii was designed and implemented based on established computational steps and criteria derived from previous literature with the following distinguishing features. First, software is easy to install with all-in-one programs and packaged databases. Second, it comes with graphical user interfaces (GUIs) for ease of use. Users can identify plant miRNAs and targets via step-by-step execution, explore the detailed results from each step, filter the results according to proposed constraints in plant miRNA and target biogenesis, and export sequences and structures of interest. Third, it supplies bird's eye views of the identification results with infographics and grouping information. Fourth, in terms of functionality, it extends the standard computational steps of miRNA target identification with miRNA-target folding and GO annotation. Fifth, it provides helper functions for the update of pre-installed databases and automatic recovery. Finally, it supports multi-project and multi-thread management. Conclusions C-mii constitutes the first complete software package with graphical user interfaces enabling computational identification of

  10. A novel neural network based image reconstruction model with scale and rotation invariance for target identification and classification for Active millimetre wave imaging

    NASA Astrophysics Data System (ADS)

    Agarwal, Smriti; Bisht, Amit Singh; Singh, Dharmendra; Pathak, Nagendra Prasad

    2014-12-01

    Millimetre wave imaging (MMW) is gaining tremendous interest among researchers, which has potential applications for security check, standoff personal screening, automotive collision-avoidance, and lot more. Current state-of-art imaging techniques viz. microwave and X-ray imaging suffers from lower resolution and harmful ionizing radiation, respectively. In contrast, MMW imaging operates at lower power and is non-ionizing, hence, medically safe. Despite these favourable attributes, MMW imaging encounters various challenges as; still it is very less explored area and lacks suitable imaging methodology for extracting complete target information. Keeping in view of these challenges, a MMW active imaging radar system at 60 GHz was designed for standoff imaging application. A C-scan (horizontal and vertical scanning) methodology was developed that provides cross-range resolution of 8.59 mm. The paper further details a suitable target identification and classification methodology. For identification of regular shape targets: mean-standard deviation based segmentation technique was formulated and further validated using a different target shape. For classification: probability density function based target material discrimination methodology was proposed and further validated on different dataset. Lastly, a novel artificial neural network based scale and rotation invariant, image reconstruction methodology has been proposed to counter the distortions in the image caused due to noise, rotation or scale variations. The designed neural network once trained with sample images, automatically takes care of these deformations and successfully reconstructs the corrected image for the test targets. Techniques developed in this paper are tested and validated using four different regular shapes viz. rectangle, square, triangle and circle.

  11. Limitations of contrast enhancement for infrared target identification

    NASA Astrophysics Data System (ADS)

    Du Bosq, Todd W.; Fanning, Jonathan D.

    2009-05-01

    Contrast enhancement and dynamic range compression are currently being used to improve the performance of infrared imagers by increasing the contrast between the target and the scene content. Automatic contrast enhancement techniques do not always achieve this improvement. In some cases, the contrast can increase to a level of target saturation. This paper assesses the range-performance effects of contrast enhancement for target identification as a function of image saturation. Human perception experiments were performed to determine field performance using contrast enhancement on the U.S. Army RDECOM CERDEC NVESD standard military eight target set using an un-cooled LWIR camera. The experiments compare the identification performance of observers viewing contrast enhancement processed images at various levels of saturation. Contrast enhancement is modeled in the U.S. Army thermal target acquisition model (NVThermIP) by changing the scene contrast temperature. The model predicts improved performance based on any improved target contrast, regardless of specific feature saturation or enhancement. The measured results follow the predicted performance based on the target task difficulty metric used in NVThermIP for the non-saturated cases. The saturated images reduce the information contained in the target and performance suffers. The model treats the contrast of the target as uniform over spatial frequency. As the contrast is enhanced, the model assumes that the contrast is enhanced uniformly over the spatial frequencies. After saturation, the spatial cues that differentiate one tank from another are located in a limited band of spatial frequencies. A frequency dependent treatment of target contrast is needed to predict performance of over-processed images.

  12. Validation of N-myristoyltransferase as an antimalarial drug target using an integrated chemical biology approach

    NASA Astrophysics Data System (ADS)

    Wright, Megan H.; Clough, Barbara; Rackham, Mark D.; Rangachari, Kaveri; Brannigan, James A.; Grainger, Munira; Moss, David K.; Bottrill, Andrew R.; Heal, William P.; Broncel, Malgorzata; Serwa, Remigiusz A.; Brady, Declan; Mann, David J.; Leatherbarrow, Robin J.; Tewari, Rita; Wilkinson, Anthony J.; Holder, Anthony A.; Tate, Edward W.

    2014-02-01

    Malaria is an infectious disease caused by parasites of the genus Plasmodium, which leads to approximately one million deaths per annum worldwide. Chemical validation of new antimalarial targets is urgently required in view of rising resistance to current drugs. One such putative target is the enzyme N-myristoyltransferase, which catalyses the attachment of the fatty acid myristate to protein substrates (N-myristoylation). Here, we report an integrated chemical biology approach to explore protein myristoylation in the major human parasite P. falciparum, combining chemical proteomic tools for identification of the myristoylated and glycosylphosphatidylinositol-anchored proteome with selective small-molecule N-myristoyltransferase inhibitors. We demonstrate that N-myristoyltransferase is an essential and chemically tractable target in malaria parasites both in vitro and in vivo, and show that selective inhibition of N-myristoylation leads to catastrophic and irreversible failure to assemble the inner membrane complex, a critical subcellular organelle in the parasite life cycle. Our studies provide the basis for the development of new antimalarials targeting N-myristoyltransferase.

  13. Identification of Direct Protein Targets of Small Molecules

    PubMed Central

    2010-01-01

    Small-molecule target identification is a vital and daunting task for the chemical biology community as well as for researchers interested in applying the power of chemical genetics to impact biology and medicine. To overcome this “target ID” bottleneck, new technologies are being developed that analyze protein–drug interactions, such as drug affinity responsive target stability (DARTS), which aims to discover the direct binding targets (and off targets) of small molecules on a proteome scale without requiring chemical modification of the compound. Here, we review the DARTS method, discuss why it works, and provide new perspectives for future development in this area. PMID:21077692

  14. Bombing Target Identification from Limited Transect Data

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Roberts, Barry L.; Hathaway, John E.; Pulsipher, Brent A.

    2006-08-07

    A series of sensor data combined with geostatistical techniques were used to determine likely target areas for a historic military aerial bombing range. Primary data consisted of magnetic anomaly information from limited magnetometer transects across the site. Secondary data included airborne LIDAR, orthophotography, and other general site characterization information. Identification of likely target areas relied primarily upon kriging estimates of magnetic anomaly densities across the site. Secondary information, such as impact crater locations, was used to refine the boundary delineations.

  15. A new disaster victim identification management strategy targeting "near identification-threshold" cases: Experiences from the Boxing Day tsunami.

    PubMed

    Wright, Kirsty; Mundorff, Amy; Chaseling, Janet; Forrest, Alexander; Maguire, Christopher; Crane, Denis I

    2015-05-01

    The international disaster victim identification (DVI) response to the Boxing Day tsunami, led by the Royal Thai Police in Phuket, Thailand, was one of the largest and most complex in DVI history. Referred to as the Thai Tsunami Victim Identification operation, the group comprised a multi-national, multi-agency, and multi-disciplinary team. The traditional DVI approach proved successful in identifying a large number of victims quickly. However, the team struggled to identify certain victims due to incomplete or poor quality ante-mortem and post-mortem data. In response to these challenges, a new 'near-threshold' DVI management strategy was implemented to target presumptive identifications and improve operational efficiency. The strategy was implemented by the DNA Team, therefore DNA kinship matches that just failed to reach the reporting threshold of 99.9% were prioritized, however the same approach could be taken by targeting, for example, cases with partial fingerprint matches. The presumptive DNA identifications were progressively filtered through the Investigation, Dental and Fingerprint Teams to add additional information necessary to either strengthen or conclusively exclude the identification. Over a five-month period 111 victims from ten countries were identified using this targeted approach. The new identifications comprised 87 adults, 24 children and included 97 Thai locals. New data from the Fingerprint Team established nearly 60% of the total near-threshold identifications and the combined DNA/Physical method was responsible for over 30%. Implementing the new strategy, targeting near-threshold cases, had positive management implications. The process initiated additional ante-mortem information collections, and established a much-needed, distinct "end-point" for unresolved cases. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  16. Collaborative identification method for sea battlefield target based on deep convolutional neural networks

    NASA Astrophysics Data System (ADS)

    Zheng, Guangdi; Pan, Mingbo; Liu, Wei; Wu, Xuetong

    2018-03-01

    The target identification of the sea battlefield is the prerequisite for the judgment of the enemy in the modern naval battle. In this paper, a collaborative identification method based on convolution neural network is proposed to identify the typical targets of sea battlefields. Different from the traditional single-input/single-output identification method, the proposed method constructs a multi-input/single-output co-identification architecture based on optimized convolution neural network and weighted D-S evidence theory. The simulation results show that

  17. Drug target identification using network analysis: Taking active components in Sini decoction as an example

    NASA Astrophysics Data System (ADS)

    Chen, Si; Jiang, Hailong; Cao, Yan; Wang, Yun; Hu, Ziheng; Zhu, Zhenyu; Chai, Yifeng

    2016-04-01

    Identifying the molecular targets for the beneficial effects of active small-molecule compounds simultaneously is an important and currently unmet challenge. In this study, we firstly proposed network analysis by integrating data from network pharmacology and metabolomics to identify targets of active components in sini decoction (SND) simultaneously against heart failure. To begin with, 48 potential active components in SND against heart failure were predicted by serum pharmacochemistry, text mining and similarity match. Then, we employed network pharmacology including text mining and molecular docking to identify the potential targets of these components. The key enriched processes, pathways and related diseases of these target proteins were analyzed by STRING database. At last, network analysis was conducted to identify most possible targets of components in SND. Among the 25 targets predicted by network analysis, tumor necrosis factor α (TNF-α) was firstly experimentally validated in molecular and cellular level. Results indicated that hypaconitine, mesaconitine, higenamine and quercetin in SND can directly bind to TNF-α, reduce the TNF-α-mediated cytotoxicity on L929 cells and exert anti-myocardial cell apoptosis effects. We envisage that network analysis will also be useful in target identification of a bioactive compound.

  18. Drug target identification using network analysis: Taking active components in Sini decoction as an example

    PubMed Central

    Chen, Si; Jiang, Hailong; Cao, Yan; Wang, Yun; Hu, Ziheng; Zhu, Zhenyu; Chai, Yifeng

    2016-01-01

    Identifying the molecular targets for the beneficial effects of active small-molecule compounds simultaneously is an important and currently unmet challenge. In this study, we firstly proposed network analysis by integrating data from network pharmacology and metabolomics to identify targets of active components in sini decoction (SND) simultaneously against heart failure. To begin with, 48 potential active components in SND against heart failure were predicted by serum pharmacochemistry, text mining and similarity match. Then, we employed network pharmacology including text mining and molecular docking to identify the potential targets of these components. The key enriched processes, pathways and related diseases of these target proteins were analyzed by STRING database. At last, network analysis was conducted to identify most possible targets of components in SND. Among the 25 targets predicted by network analysis, tumor necrosis factor α (TNF-α) was firstly experimentally validated in molecular and cellular level. Results indicated that hypaconitine, mesaconitine, higenamine and quercetin in SND can directly bind to TNF-α, reduce the TNF-α-mediated cytotoxicity on L929 cells and exert anti-myocardial cell apoptosis effects. We envisage that network analysis will also be useful in target identification of a bioactive compound. PMID:27095146

  19. Efficient Identification of Murine M2 Macrophage Peptide Targeting Ligands by Phage Display and Next-Generation Sequencing.

    PubMed

    Liu, Gary W; Livesay, Brynn R; Kacherovsky, Nataly A; Cieslewicz, Maryelise; Lutz, Emi; Waalkes, Adam; Jensen, Michael C; Salipante, Stephen J; Pun, Suzie H

    2015-08-19

    Peptide ligands are used to increase the specificity of drug carriers to their target cells and to facilitate intracellular delivery. One method to identify such peptide ligands, phage display, enables high-throughput screening of peptide libraries for ligands binding to therapeutic targets of interest. However, conventional methods for identifying target binders in a library by Sanger sequencing are low-throughput, labor-intensive, and provide a limited perspective (<0.01%) of the complete sequence space. Moreover, the small sample space can be dominated by nonspecific, preferentially amplifying "parasitic sequences" and plastic-binding sequences, which may lead to the identification of false positives or exclude the identification of target-binding sequences. To overcome these challenges, we employed next-generation Illumina sequencing to couple high-throughput screening and high-throughput sequencing, enabling more comprehensive access to the phage display library sequence space. In this work, we define the hallmarks of binding sequences in next-generation sequencing data, and develop a method that identifies several target-binding phage clones for murine, alternatively activated M2 macrophages with a high (100%) success rate: sequences and binding motifs were reproducibly present across biological replicates; binding motifs were identified across multiple unique sequences; and an unselected, amplified library accurately filtered out parasitic sequences. In addition, we validate the Multiple Em for Motif Elicitation tool as an efficient and principled means of discovering binding sequences.

  20. miRTar2GO: a novel rule-based model learning method for cell line specific microRNA target prediction that integrates Ago2 CLIP-Seq and validated microRNA-target interaction data.

    PubMed

    Ahadi, Alireza; Sablok, Gaurav; Hutvagner, Gyorgy

    2017-04-07

    MicroRNAs (miRNAs) are ∼19-22 nucleotides (nt) long regulatory RNAs that regulate gene expression by recognizing and binding to complementary sequences on mRNAs. The key step in revealing the function of a miRNA, is the identification of miRNA target genes. Recent biochemical advances including PAR-CLIP and HITS-CLIP allow for improved miRNA target predictions and are widely used to validate miRNA targets. Here, we present miRTar2GO, which is a model, trained on the common rules of miRNA-target interactions, Argonaute (Ago) CLIP-Seq data and experimentally validated miRNA target interactions. miRTar2GO is designed to predict miRNA target sites using more relaxed miRNA-target binding characteristics. More importantly, miRTar2GO allows for the prediction of cell-type specific miRNA targets. We have evaluated miRTar2GO against other widely used miRNA target prediction algorithms and demonstrated that miRTar2GO produced significantly higher F1 and G scores. Target predictions, binding specifications, results of the pathway analysis and gene ontology enrichment of miRNA targets are freely available at http://www.mirtar2go.org. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  1. Drug Target Validation Methods in Malaria - Protein Interference Assay (PIA) as a Tool for Highly Specific Drug Target Validation.

    PubMed

    Meissner, Kamila A; Lunev, Sergey; Wang, Yuan-Ze; Linzke, Marleen; de Assis Batista, Fernando; Wrenger, Carsten; Groves, Matthew R

    2017-01-01

    The validation of drug targets in malaria and other human diseases remains a highly difficult and laborious process. In the vast majority of cases, highly specific small molecule tools to inhibit a proteins function in vivo are simply not available. Additionally, the use of genetic tools in the analysis of malarial pathways is challenging. These issues result in difficulties in specifically modulating a hypothetical drug target's function in vivo. The current "toolbox" of various methods and techniques to identify a protein's function in vivo remains very limited and there is a pressing need for expansion. New approaches are urgently required to support target validation in the drug discovery process. Oligomerisation is the natural assembly of multiple copies of a single protein into one object and this self-assembly is present in more than half of all protein structures. Thus, oligomerisation plays a central role in the generation of functional biomolecules. A key feature of oligomerisation is that the oligomeric interfaces between the individual parts of the final assembly are highly specific. However, these interfaces have not yet been systematically explored or exploited to dissect biochemical pathways in vivo. This mini review will describe the current state of the antimalarial toolset as well as the potentially druggable malarial pathways. A specific focus is drawn to the initial efforts to exploit oligomerisation surfaces in drug target validation. As alternative to the conventional methods, Protein Interference Assay (PIA) can be used for specific distortion of the target protein function and pathway assessment in vivo. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  2. Common pitfalls in preclinical cancer target validation.

    PubMed

    Kaelin, William G

    2017-07-01

    An alarming number of papers from laboratories nominating new cancer drug targets contain findings that cannot be reproduced by others or are simply not robust enough to justify drug discovery efforts. This problem probably has many causes, including an underappreciation of the danger of being misled by off-target effects when using pharmacological or genetic perturbants in complex biological assays. This danger is particularly acute when, as is often the case in cancer pharmacology, the biological phenotype being measured is a 'down' readout (such as decreased proliferation, decreased viability or decreased tumour growth) that could simply reflect a nonspecific loss of cellular fitness. These problems are compounded by multiple hypothesis testing, such as when candidate targets emerge from high-throughput screens that interrogate multiple targets in parallel, and by a publication and promotion system that preferentially rewards positive findings. In this Perspective, I outline some of the common pitfalls in preclinical cancer target identification and some potential approaches to mitigate them.

  3. Extended version of the "Sniffin' Sticks" identification test: test-retest reliability and validity.

    PubMed

    Sorokowska, A; Albrecht, E; Haehner, A; Hummel, T

    2015-03-30

    The extended, 32-item version of the Sniffin' Sticks identification test was developed in order to create a precise tool enabling repeated, longitudinal testing of individual olfactory subfunctions. Odors of the previous test version had to be changed for technical reasons, and the odor identification test needed re-investigation in terms of reliability, validity, and normative values. In our study we investigated olfactory abilities of a group of 100 patients with olfactory dysfunction and 100 controls. We reconfirmed the high test-retest reliability of the extended version of the Sniffin' Sticks identification test and high correlations between the new and the original part of this tool. In addition, we confirmed the validity of the test as it discriminated clearly between controls and patients with olfactory loss. The additional set of 16 odor identification sticks can be either included in the current olfactory test, thus creating a more detailed diagnosis tool, or it can be used separately, enabling to follow olfactory function over time. Additionally, the normative values presented in our paper might provide useful guidelines for interpretation of the extended identification test results. The revised version of the Sniffin' Sticks 32-item odor identification test is a reliable and valid tool for the assessment of olfactory function. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. Target-decoy Based False Discovery Rate Estimation for Large-scale Metabolite Identification.

    PubMed

    Wang, Xusheng; Jones, Drew R; Shaw, Timothy I; Cho, Ji-Hoon; Wang, Yuanyuan; Tan, Haiyan; Xie, Boer; Zhou, Suiping; Li, Yuxin; Peng, Junmin

    2018-05-23

    Metabolite identification is a crucial step in mass spectrometry (MS)-based metabolomics. However, it is still challenging to assess the confidence of assigned metabolites. In this study, we report a novel method for estimating false discovery rate (FDR) of metabolite assignment with a target-decoy strategy, in which the decoys are generated through violating the octet rule of chemistry by adding small odd numbers of hydrogen atoms. The target-decoy strategy was integrated into JUMPm, an automated metabolite identification pipeline for large-scale MS analysis, and was also evaluated with two other metabolomics tools, mzMatch and mzMine 2. The reliability of FDR calculation was examined by false datasets, which were simulated by altering MS1 or MS2 spectra. Finally, we used the JUMPm pipeline coupled with the target-decoy strategy to process unlabeled and stable-isotope labeled metabolomic datasets. The results demonstrate that the target-decoy strategy is a simple and effective method for evaluating the confidence of high-throughput metabolite identification.

  5. [Exploring New Drug Targets through the Identification of Target Molecules of Bioactive Natural Products].

    PubMed

    Arai, Masayoshi

    2016-01-01

    With the development of cell biology and microbiology, it has become easy to culture many types of animal cells and microbes, and they are frequently used for phenotypic screening to explore medicinal seeds. On the other hand, it is recognized that cells and pathogenic microbes present in pathologic sites and infected regions of the human body display unique properties different from those under general culture conditions. We isolated several bioactive compounds from marine medicinal resources using constructed bioassay-guided separation focusing on the unique changes in the characteristics of cells and pathogenic microbes (Mycobacterium spp.) in the human body under disease conditions. In addition, we also carried out identification studies of target molecules of the bioactive compounds by methods utilizing the gene expression profile, transformants of cells or microbes, synthetic probe molecules of the isolated compounds, etc., since bioactive compounds isolated from the phenotypic screening system often target new molecules. This review presents our phenotypic screening systems, isolation of bioactive compounds from marine medicinal resources, and target identification of bioactive compounds.

  6. The Influence of Attention and Target Identification on Saccadic Eye Movements Depends on Prior Target Location

    PubMed Central

    Hardwick, David R.; Cutmore, Timothy R. H.; Hine, Trevor J.

    2014-01-01

    Saccadic latency is reduced by a temporal gap between fixation point and target, by identification of a target feature, and by movement in a new direction (inhibition of saccadic return, ISR). A simple additive model was compared with a shared resources model that predicts a three-way interaction. Twenty naïve participants made horizontal saccades to targets left and right of fixation in a randomised block design. There was a significant three-way interaction among the factors on saccade latency. This was revealed in a two-way interaction between feature identification and the gap versus no gap factor which was only apparent when the saccade was in the same direction as the previous saccade. No interaction was apparent when the saccade was in the opposite direction. This result supports an attentional inhibitory effect that is present during ISR to a previous location which is only partly released by the facilitative effect of feature identification and gap. Together, anticipatory error data and saccade latency interactions suggest a source of ISR at a higher level of attention, possibly localised in the dorsolateral prefrontal cortex and involving tonic activation. PMID:24719754

  7. Visually-guided attention enhances target identification in a complex auditory scene.

    PubMed

    Best, Virginia; Ozmeral, Erol J; Shinn-Cunningham, Barbara G

    2007-06-01

    In auditory scenes containing many similar sound sources, sorting of acoustic information into streams becomes difficult, which can lead to disruptions in the identification of behaviorally relevant targets. This study investigated the benefit of providing simple visual cues for when and/or where a target would occur in a complex acoustic mixture. Importantly, the visual cues provided no information about the target content. In separate experiments, human subjects either identified learned birdsongs in the presence of a chorus of unlearned songs or recalled strings of spoken digits in the presence of speech maskers. A visual cue indicating which loudspeaker (from an array of five) would contain the target improved accuracy for both kinds of stimuli. A cue indicating which time segment (out of a possible five) would contain the target also improved accuracy, but much more for birdsong than for speech. These results suggest that in real world situations, information about where a target of interest is located can enhance its identification, while information about when to listen can also be helpful when targets are unfamiliar or extremely similar to their competitors.

  8. In silico target prediction for elucidating the mode of action of herbicides including prospective validation.

    PubMed

    Chiddarwar, Rucha K; Rohrer, Sebastian G; Wolf, Antje; Tresch, Stefan; Wollenhaupt, Sabrina; Bender, Andreas

    2017-01-01

    The rapid emergence of pesticide resistance has given rise to a demand for herbicides with new mode of action (MoA). In the agrochemical sector, with the availability of experimental high throughput screening (HTS) data, it is now possible to utilize in silico target prediction methods in the early discovery phase to suggest the MoA of a compound via data mining of bioactivity data. While having been established in the pharmaceutical context, in the agrochemical area this approach poses rather different challenges, as we have found in this work, partially due to different chemistry, but even more so due to different (usually smaller) amounts of data, and different ways of conducting HTS. With the aim to apply computational methods for facilitating herbicide target identification, 48,000 bioactivity data against 16 herbicide targets were processed to train Laplacian modified Naïve Bayesian (NB) classification models. The herbicide target prediction model ("HerbiMod") is an ensemble of 16 binary classification models which are evaluated by internal, external and prospective validation sets. In addition to the experimental inactives, 10,000 random agrochemical inactives were included in the training process, which showed to improve the overall balanced accuracy of our models up to 40%. For all the models, performance in terms of balanced accuracy of≥80% was achieved in five-fold cross validation. Ranking target predictions was addressed by means of z-scores which improved predictivity over using raw scores alone. An external testset of 247 compounds from ChEMBL and a prospective testset of 394 compounds from BASF SE tested against five well studied herbicide targets (ACC, ALS, HPPD, PDS and PROTOX) were used for further validation. Only 4% of the compounds in the external testset lied in the applicability domain and extrapolation (and correct prediction) was hence impossible, which on one hand was surprising, and on the other hand illustrated the utilization of

  9. Enhancement of Chemical Entity Identification in Text Using Semantic Similarity Validation

    PubMed Central

    Grego, Tiago; Couto, Francisco M.

    2013-01-01

    With the amount of chemical data being produced and reported in the literature growing at a fast pace, it is increasingly important to efficiently retrieve this information. To tackle this issue text mining tools have been applied, but despite their good performance they still provide many errors that we believe can be filtered by using semantic similarity. Thus, this paper proposes a novel method that receives the results of chemical entity identification systems, such as Whatizit, and exploits the semantic relationships in ChEBI to measure the similarity between the entities found in the text. The method assigns a single validation score to each entity based on its similarities with the other entities also identified in the text. Then, by using a given threshold, the method selects a set of validated entities and a set of outlier entities. We evaluated our method using the results of two state-of-the-art chemical entity identification tools, three semantic similarity measures and two text window sizes. The method was able to increase precision without filtering a significant number of correctly identified entities. This means that the method can effectively discriminate the correctly identified chemical entities, while discarding a significant number of identification errors. For example, selecting a validation set with 75% of all identified entities, we were able to increase the precision by 28% for one of the chemical entity identification tools (Whatizit), maintaining in that subset 97% the correctly identified entities. Our method can be directly used as an add-on by any state-of-the-art entity identification tool that provides mappings to a database, in order to improve their results. The proposed method is included in a freely accessible web tool at www.lasige.di.fc.ul.pt/webtools/ice/. PMID:23658791

  10. Challenges in validating candidate therapeutic targets in cancer

    PubMed Central

    Sawyers, Charles L; Hunter, Tony

    2018-01-01

    More than 30 published articles have suggested that a protein kinase called MELK is an attractive therapeutic target in human cancer, but three recent reports describe compelling evidence that it is not. These reports highlight the caveats associated with some of the research tools that are commonly used to validate candidate therapeutic targets in cancer research. PMID:29417929

  11. Prioritization of biomarker targets in human umbilical cord blood: identification of proteins in infant blood serving as validated biomarkers in adults.

    PubMed

    Hansmeier, Nicole; Chao, Tzu-Chiao; Goldman, Lynn R; Witter, Frank R; Halden, Rolf U

    2012-05-01

    Early diagnosis represents one of the best lines of defense in the fight against a wide array of human diseases. Umbilical cord blood (UCB) is one of the first easily available diagnostic biofluids and can inform about the health status of newborns. However, compared with adult blood, its diagnostic potential remains largely untapped. Our goal was to accelerate biomarker research on UCB by exploring its detectable protein content and providing a priority list of potential biomarkers based on known proteins involved in disease pathways. We explored cord blood serum proteins by profiling a UCB pool of 12 neonates with different backgrounds using a combination of isoelectric focusing and liquid chromatography coupled with matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) and by comparing results with information contained in metabolic and disease databases available for adult blood. A total of 1,210 UCB proteins were identified with a protein-level false discovery rate of ~ 5% as estimated by naïve target-decoy and MAYU approaches, signifying a 6-fold increase in the number of UCB proteins described to date. Identified proteins correspond to 138 different metabolic and disease pathways and provide a platform of mechanistically linked biomarker candidates for tracking disruptions in cellular processes. Moreover, among the identified proteins, 38 were found to be approved biomarkers for adult blood. The results of this study advance current knowledge of the human cord blood serum proteome. They showcase the potential of UCB as a diagnostic medium for assessing infant health by detection and identification of candidate biomarkers for known disease pathways using a global, nontargeted approach. These biomarkers may inform about mechanisms of exposure-disease relationships. Furthermore, biomarkers approved by the U.S. Food and Drug Administration for screening in adult blood were detected in UCB and represent high-priority targets for

  12. Reliability and validity of a talent identification test battery for seated and standing Paralympic throws.

    PubMed

    Spathis, Jemima Grace; Connick, Mark James; Beckman, Emma Maree; Newcombe, Peter Anthony; Tweedy, Sean Michael

    2015-01-01

    Paralympic throwing events for athletes with physical impairments comprise seated and standing javelin, shot put, discus and seated club throwing. Identification of talented throwers would enable prediction of future success and promote participation; however, a valid and reliable talent identification battery for Paralympic throwing has not been reported. This study evaluates the reliability and validity of a talent identification battery for Paralympic throws. Participants were non-disabled so that impairment would not confound analyses, and results would provide an indication of normative performance. Twenty-eight non-disabled participants (13 M; 15 F) aged 23.6 years (±5.44) performed five kinematically distinct criterion throws (three seated, two standing) and nine talent identification tests (three anthropometric, six motor); 23 were tested a second time to evaluate test-retest reliability. Talent identification test-retest reliability was evaluated using Intra-class Correlation Coefficient (ICC) and Bland-Altman plots (Limits of Agreement). Spearman's correlation assessed strength of association between criterion throws and talent identification tests. Reliability was generally acceptable (mean ICC = 0.89), but two seated talent identification tests require more extensive familiarisation. Correlation strength (mean rs = 0.76) indicated that the talent identification tests can be used to validly identify individuals with competitively advantageous attributes for each of the five kinematically distinct throwing activities. Results facilitate further research in this understudied area.

  13. Patterns of Cognitive Strengths and Weaknesses: Identification Rates, Agreement, and Validity for Learning Disabilities Identification

    ERIC Educational Resources Information Center

    Miciak, Jeremy; Fletcher, Jack M.; Stuebing, Karla K.; Vaughn, Sharon; Tolar, Tammy D.

    2014-01-01

    Few empirical investigations have evaluated learning disabilities (LD) identification methods based on a pattern of cognitive strengths and weaknesses (PSW). This study investigated the reliability and validity of two proposed PSW methods: the concordance/discordance method (C/DM) and cross battery assessment (XBA) method. Cognitive assessment…

  14. Quantitative and Systems Pharmacology. 1. In Silico Prediction of Drug-Target Interactions of Natural Products Enables New Targeted Cancer Therapy.

    PubMed

    Fang, Jiansong; Wu, Zengrui; Cai, Chuipu; Wang, Qi; Tang, Yun; Cheng, Feixiong

    2017-11-27

    Natural products with diverse chemical scaffolds have been recognized as an invaluable source of compounds in drug discovery and development. However, systematic identification of drug targets for natural products at the human proteome level via various experimental assays is highly expensive and time-consuming. In this study, we proposed a systems pharmacology infrastructure to predict new drug targets and anticancer indications of natural products. Specifically, we reconstructed a global drug-target network with 7,314 interactions connecting 751 targets and 2,388 natural products and built predictive network models via a balanced substructure-drug-target network-based inference approach. A high area under receiver operating characteristic curve of 0.96 was yielded for predicting new targets of natural products during cross-validation. The newly predicted targets of natural products (e.g., resveratrol, genistein, and kaempferol) with high scores were validated by various literature studies. We further built the statistical network models for identification of new anticancer indications of natural products through integration of both experimentally validated and computationally predicted drug-target interactions of natural products with known cancer proteins. We showed that the significantly predicted anticancer indications of multiple natural products (e.g., naringenin, disulfiram, and metformin) with new mechanism-of-action were validated by various published experimental evidence. In summary, this study offers powerful computational systems pharmacology approaches and tools for the development of novel targeted cancer therapies by exploiting the polypharmacology of natural products.

  15. Characterization of anti-leukemia components from Indigo naturalis using comprehensive two-dimensional K562/cell membrane chromatography and in silico target identification.

    PubMed

    Wu, Xunxun; Chen, Xiaofei; Dan, Jia; Cao, Yan; Gao, Shouhong; Guo, Zhiying; Zerbe, Philipp; Chai, Yifeng; Diao, Yong; Zhang, Lei

    2016-05-06

    Traditional Chinese Medicine (TCM) has been developed for thousands of years and has formed an integrated theoretical system based on a large amount of clinical practice. However, essential ingredients in TCM herbs have not been fully identified, and their precise mechanisms and targets are not elucidated. In this study, a new strategy combining comprehensive two-dimensional K562/cell membrane chromatographic system and in silico target identification was established to characterize active components from Indigo naturalis, a famous TCM herb that has been widely used for the treatment of leukemia in China, and their targets. Three active components, indirubin, tryptanthrin and isorhamnetin, were successfully characterized and their anti-leukemia effects were validated by cell viability and cell apoptosis assays. Isorhamnetin, with undefined cancer related targets, was selected for in silico target identification. Proto-oncogene tyrosine-protein kinase (Src) was identified as its membrane target and the dissociation constant (Kd) between Src and isorhamnetin was 3.81 μM. Furthermore, anti-leukemia effects of isorhamnetin were mediated by Src through inducing G2/M cell cycle arrest. The results demonstrated that the integrated strategy could efficiently characterize active components in TCM and their targets, which may bring a new light for a better understanding of the complex mechanism of herbal medicines.

  16. Characterization of anti-leukemia components from Indigo naturalis using comprehensive two-dimensional K562/cell membrane chromatography and in silico target identification

    PubMed Central

    Wu, Xunxun; Chen, Xiaofei; Dan, Jia; Cao, Yan; Gao, Shouhong; Guo, Zhiying; Zerbe, Philipp; Chai, Yifeng; Diao, Yong; Zhang, Lei

    2016-01-01

    Traditional Chinese Medicine (TCM) has been developed for thousands of years and has formed an integrated theoretical system based on a large amount of clinical practice. However, essential ingredients in TCM herbs have not been fully identified, and their precise mechanisms and targets are not elucidated. In this study, a new strategy combining comprehensive two-dimensional K562/cell membrane chromatographic system and in silico target identification was established to characterize active components from Indigo naturalis, a famous TCM herb that has been widely used for the treatment of leukemia in China, and their targets. Three active components, indirubin, tryptanthrin and isorhamnetin, were successfully characterized and their anti-leukemia effects were validated by cell viability and cell apoptosis assays. Isorhamnetin, with undefined cancer related targets, was selected for in silico target identification. Proto-oncogene tyrosine-protein kinase (Src) was identified as its membrane target and the dissociation constant (Kd) between Src and isorhamnetin was 3.81 μM. Furthermore, anti-leukemia effects of isorhamnetin were mediated by Src through inducing G2/M cell cycle arrest. The results demonstrated that the integrated strategy could efficiently characterize active components in TCM and their targets, which may bring a new light for a better understanding of the complex mechanism of herbal medicines. PMID:27150638

  17. Snapshot spectral and polarimetric imaging; target identification with multispectral video

    NASA Astrophysics Data System (ADS)

    Bartlett, Brent D.; Rodriguez, Mikel D.

    2013-05-01

    As the number of pixels continue to grow in consumer and scientific imaging devices, it has become feasible to collect the incident light field. In this paper, an imaging device developed around light field imaging is used to collect multispectral and polarimetric imagery in a snapshot fashion. The sensor is described and a video data set is shown highlighting the advantage of snapshot spectral imaging. Several novel computer vision approaches are applied to the video cubes to perform scene characterization and target identification. It is shown how the addition of spectral and polarimetric data to the video stream allows for multi-target identification and tracking not possible with traditional RGB video collection.

  18. Visually-guided Attention Enhances Target Identification in a Complex Auditory Scene

    PubMed Central

    Ozmeral, Erol J.; Shinn-Cunningham, Barbara G.

    2007-01-01

    In auditory scenes containing many similar sound sources, sorting of acoustic information into streams becomes difficult, which can lead to disruptions in the identification of behaviorally relevant targets. This study investigated the benefit of providing simple visual cues for when and/or where a target would occur in a complex acoustic mixture. Importantly, the visual cues provided no information about the target content. In separate experiments, human subjects either identified learned birdsongs in the presence of a chorus of unlearned songs or recalled strings of spoken digits in the presence of speech maskers. A visual cue indicating which loudspeaker (from an array of five) would contain the target improved accuracy for both kinds of stimuli. A cue indicating which time segment (out of a possible five) would contain the target also improved accuracy, but much more for birdsong than for speech. These results suggest that in real world situations, information about where a target of interest is located can enhance its identification, while information about when to listen can also be helpful when targets are unfamiliar or extremely similar to their competitors. PMID:17453308

  19. Targeted stock identification using multilocus genotype 'familyprinting'

    USGS Publications Warehouse

    Letcher, B.H.; King, T.L.

    1999-01-01

    We present an approach to stock identification of small, targeted populations that uses multilocus microsatellite genotypes of individual mating adults to uniquely identify first- and second-generation offspring in a mixture. We call the approach 'familyprinting'; unlike DNA fingerprinting where tissue samples of individuals are matched, offspring from various families are assigned to pairs of parents or sets of four grandparents with known genotypes. The basic unit of identification is the family, but families can be nested within a variety of stock units ranging from naturally reproducing groups of fish in a small tributary or pond from which mating adults can be sampled to large or small collections of families produced in hatcheries and stocked in specific locations. We show that, with as few as seven alleles per locus using four loci without error, first-generation offspring can be uniquely assigned to the correct family. For second-generation applications in a hatchery more alleles per locus (10) and loci (10) are required for correct assignment of all offspring to the correct set of grandparents. Using microsatellite DNA variation from an Atlantic salmon (Salmo solar) restoration river (Connecticut River, USA), we also show that this population contains sufficient genetic diversity in sea-run returns for 100% correct first, generation assignment and 97% correct second-generation assignment using 14 loci. We are currently using first- and second-generation familyprinting in this population with the ultimate goal of identifying stocking tributary. In addition to within-river familyprinting, there also appears to be sufficient genetic diversity within and between Atlantic salmon populations for identification of 'familyprinted' fish in a mixture of multiple populations. We also suggest that second-generation familyprinting with multiple populations may also provide a tool for examining stock structure. Familyprinting with microsatellite DNA markers is a viable

  20. Development and validation of real-time PCR tests for the identification of four Spodoptera species: Spodoptera eridania, Spodoptera frugiperda, Spodoptera littoralis, and Spodoptera litura (Lepidoptera: Noctuidae).

    PubMed

    Van de Vossenberg, B T L H; Van der Straten, M J

    2014-08-01

    The genus Spodoptera comprises 31 species, 4 of which are listed as quarantine pests for the European Union: Spodoptera eridania (Cramer), Spodoptera frugiperda (Smith), Spodoptera littoralis (Boisduval), and Spodoptera litura (F.). In international trade, the earlier life stages (eggs and larvae) are being intercepted at point of inspection most frequently, challenging the possibilities of morphological identification. To realize a rapid and reliable identification for all stages, we developed and validated four simplex real-time polymerase chain reaction identification tests based on the mitochondrial cytochrome b gene using dual-labeled hydrolysis probes. Method validation on dilutions of extracted DNA of the target organisms showed that low levels of template (up to 0.2-100 pg) can reliably be identified. No cross-reactivity was observed with 14 nontarget Spodoptera and 5 non-Spodoptera species in the specific Spodoptera tests. The tests showed to be repeatable, reproducible (both 100%), and robust. The new Spodoptera tests have proven to be suitable tools for routine identification of all life stages of S. eridania, S. frugiperda, S. littoralis, and S. litura.

  1. Identification of regulatory targets of tissue-specific transcription factors: application to retina-specific gene regulation

    PubMed Central

    Qian, Jiang; Esumi, Noriko; Chen, Yangjian; Wang, Qingliang; Chowers, Itay; Zack, Donald J.

    2005-01-01

    Identification of tissue-specific gene regulatory networks can yield insights into the molecular basis of a tissue's development, function and pathology. Here, we present a computational approach designed to identify potential regulatory target genes of photoreceptor cell-specific transcription factors (TFs). The approach is based on the hypothesis that genes related to the retina in terms of expression, disease and/or function are more likely to be the targets of retina-specific TFs than other genes. A list of genes that are preferentially expressed in retina was obtained by integrating expressed sequence tag, SAGE and microarray datasets. The regulatory targets of retina-specific TFs are enriched in this set of retina-related genes. A Bayesian approach was employed to integrate information about binding site location relative to a gene's transcription start site. Our method was applied to three retina-specific TFs, CRX, NRL and NR2E3, and a number of potential targets were predicted. To experimentally assess the validity of the bioinformatic predictions, mobility shift, transient transfection and chromatin immunoprecipitation assays were performed with five predicted CRX targets, and the results were suggestive of CRX regulation in 5/5, 3/5 and 4/5 cases, respectively. Together, these experiments strongly suggest that RP1, GUCY2D, ABCA4 are novel targets of CRX. PMID:15967807

  2. In silico analysis and experimental validation of azelastine hydrochloride (N4) targeting sodium taurocholate co-transporting polypeptide (NTCP) in HBV therapy.

    PubMed

    Fu, L-L; Liu, J; Chen, Y; Wang, F-T; Wen, X; Liu, H-Q; Wang, M-Y; Ouyang, L; Huang, J; Bao, J-K; Wei, Y-Q

    2014-08-01

    The aim of this study was to explore sodium taurocholate co-transporting polypeptide (NTCP) exerting its function with hepatitis B virus (HBV) and its targeted candidate compounds, in HBV therapy. Identification of NTCP as a novel HBV target for screening candidate small molecules, was used by phylogenetic analysis, network construction, molecular modelling, molecular docking and molecular dynamics (MD) simulation. In vitro virological examination, q-PCR, western blotting and cytotoxicity studies were used for validating efficacy of the candidate compound. We used the phylogenetic analysis of NTCP and constructed its protein-protein network. Also, we screened compounds from Drugbank and ZINC, among which five were validated for their authentication in HepG 2.2.15 cells. Then, we selected compound N4 (azelastine hydrochloride) as the most potent of them. This showed good inhibitory activity against HBsAg (IC50 = 7.5 μm) and HBeAg (IC50 = 3.7 μm), as well as high SI value (SI = 4.68). Further MD simulation results supported good interaction between compound N4 and NTCP. In silico analysis and experimental validation together demonstrated that compound N4 can target NTCP in HepG2.2.15 cells, which may shed light on exploring it as a potential anti-HBV drug. © 2014 John Wiley & Sons Ltd.

  3. Development and validation of a multiplex real-time PCR method to simultaneously detect 47 targets for the identification of genetically modified organisms.

    PubMed

    Cottenet, Geoffrey; Blancpain, Carine; Sonnard, Véronique; Chuah, Poh Fong

    2013-08-01

    Considering the increase of the total cultivated land area dedicated to genetically modified organisms (GMO), the consumers' perception toward GMO and the need to comply with various local GMO legislations, efficient and accurate analytical methods are needed for their detection and identification. Considered as the gold standard for GMO analysis, the real-time polymerase chain reaction (RTi-PCR) technology was optimised to produce a high-throughput GMO screening method. Based on simultaneous 24 multiplex RTi-PCR running on a ready-to-use 384-well plate, this new procedure allows the detection and identification of 47 targets on seven samples in duplicate. To comply with GMO analytical quality requirements, a negative and a positive control were analysed in parallel. In addition, an internal positive control was also included in each reaction well for the detection of potential PCR inhibition. Tested on non-GM materials, on different GM events and on proficiency test samples, the method offered high specificity and sensitivity with an absolute limit of detection between 1 and 16 copies depending on the target. Easy to use, fast and cost efficient, this multiplex approach fits the purpose of GMO testing laboratories.

  4. Validation of the Electromagnetic Code FACETS for Numerical Simulation of Radar Target Images

    DTIC Science & Technology

    2009-12-01

    Validation of the electromagnetic code FACETS for numerical simulation of radar target images S. Wong...Validation of the electromagnetic code FACETS for numerical simulation of radar target images S. Wong DRDC Ottawa...for simulating radar images of a target is obtained, through direct simulation-to-measurement comparisons. A 3-dimensional computer-aided design

  5. Identification and Validation of Ifit1 as an Important Innate Immune Bottleneck

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    McDermott, Jason E.; Vartanian, Keri B.; Mitchell, Hugh D.

    The innate immune system plays important roles in a number of disparate processes. Foremost, innate immunity is a first responder to invasion by pathogens and triggers early defensive responses and recruits the adaptive immune system. The innate immune system also responds to endogenous damage signals that arise from tissue injury. Recently it has been found that innate immunity plays an important role in neuroprotection against ischemic stroke through the activation of the primary innate immune receptors, Toll-like receptors (TLRs). Using several large-scale transcriptomic data sets from mouse and mouse macrophage studies we identified targets predicted to be important in controllingmore » innate immune processes initiated by TLR activation. Targets were identified as genes with high betweenness centrality, so-called bottlenecks, in networks inferred from statistical associations between gene expression patterns. A small set of putative bottlenecks were identified in each of the data sets investigated including interferon-stimulated genes (Ifit1, Ifi47, Tgtp and Oasl2) as well as genes uncharacterized in immune responses (Axud1 and Ppp1r15a). We further validated one of these targets, Ifit1, in mouse macrophages by showing that silencing it suppresses induction of predicted downstream genes by lipopolysaccharide (LPS)-mediated TLR4 activation through an unknown direct or indirect mechanism. Our study demonstrates the utility of network analysis for identification of interesting targets related to innate immune function, and highlights that Ifit1 can exert a positive regulatory effect on downstream genes.« less

  6. Target Acquisition: Human Observer Performance Studies and TARGAC Model Validation

    DTIC Science & Technology

    1994-06-01

    complete Scenario I run were displayed on a monitor. A transparent sheet was attached to the face of the monitor, the approach route was sketched on...track, passed a mark on the monitor face , the target was at a stop-sign location. The IRIG-B time of that instance was noted, and, since the distance of...route right 100- 80- 60 - 40 C.) 20 [- ion, A20 recognitio 0 0 . identification,C.) . 1I . I - - -• , 4) 1000 2000 3000 4000 SP Target G Q Target I0

  7. Accurate and exact CNV identification from targeted high-throughput sequence data.

    PubMed

    Nord, Alex S; Lee, Ming; King, Mary-Claire; Walsh, Tom

    2011-04-12

    Massively parallel sequencing of barcoded DNA samples significantly increases screening efficiency for clinically important genes. Short read aligners are well suited to single nucleotide and indel detection. However, methods for CNV detection from targeted enrichment are lacking. We present a method combining coverage with map information for the identification of deletions and duplications in targeted sequence data. Sequencing data is first scanned for gains and losses using a comparison of normalized coverage data between samples. CNV calls are confirmed by testing for a signature of sequences that span the CNV breakpoint. With our method, CNVs can be identified regardless of whether breakpoints are within regions targeted for sequencing. For CNVs where at least one breakpoint is within targeted sequence, exact CNV breakpoints can be identified. In a test data set of 96 subjects sequenced across ~1 Mb genomic sequence using multiplexing technology, our method detected mutations as small as 31 bp, predicted quantitative copy count, and had a low false-positive rate. Application of this method allows for identification of gains and losses in targeted sequence data, providing comprehensive mutation screening when combined with a short read aligner.

  8. Probability of identification (POI): a statistical model for the validation of qualitative botanical identification methods

    USDA-ARS?s Scientific Manuscript database

    A qualitative botanical identification method (BIM) is an analytical procedure which returns a binary result (1 = Identified, 0 = Not Identified). A BIM may be used by a buyer, manufacturer, or regulator to determine whether a botanical material being tested is the same as the target (desired) mate...

  9. RNAi screen for rapid therapeutic target identification in leukemia patients

    PubMed Central

    Tyner, Jeffrey W.; Deininger, Michael W.; Loriaux, Marc M.; Chang, Bill H.; Gotlib, Jason R.; Willis, Stephanie G.; Erickson, Heidi; Kovacsovics, Tibor; O'Hare, Thomas; Heinrich, Michael C.; Druker, Brian J.

    2009-01-01

    Targeted therapy has vastly improved outcomes in certain types of cancer. Extension of this paradigm across a broad spectrum of malignancies will require an efficient method to determine the molecular vulnerabilities of cancerous cells. Improvements in sequencing technology will soon enable high-throughput sequencing of entire genomes of cancer patients; however, determining the relevance of identified sequence variants will require complementary functional analyses. Here, we report an RNAi-assisted protein target identification (RAPID) technology that individually assesses targeting of each member of the tyrosine kinase gene family. We demonstrate that RAPID screening of primary leukemia cells from 30 patients identifies targets that are critical to survival of the malignant cells from 10 of these individuals. We identify known, activating mutations in JAK2 and K-RAS, as well as patient-specific sensitivity to down-regulation of FLT1, CSF1R, PDGFR, ROR1, EPHA4/5, JAK1/3, LMTK3, LYN, FYN, PTK2B, and N-RAS. We also describe a previously undescribed, somatic, activating mutation in the thrombopoietin receptor that is sensitive to down-stream pharmacologic inhibition. Hence, the RAPID technique can quickly identify molecular vulnerabilities in malignant cells. Combination of this technique with whole-genome sequencing will represent an ideal tool for oncogenic target identification such that specific therapies can be matched with individual patients. PMID:19433805

  10. The confidence-accuracy relationship in eyewitness identification: effects of lineup instructions, foil similarity, and target-absent base rates.

    PubMed

    Brewer, Neil; Wells, Gary L

    2006-03-01

    Discriminating accurate from mistaken eyewitness identifications is a major issue facing criminal justice systems. This study examined whether eyewitness confidence assists such decisions under a variety of conditions using a confidence-accuracy (CA) calibration approach. Participants (N = 1,200) viewed a simulated crime and attempted 2 separate identifications from 8-person target-present or target-absent lineups. Confidence and accuracy were calibrated for choosers (but not nonchoosers) for both targets under all conditions. Lower overconfidence was associated with higher diagnosticity, lower target-absent base rates, and shorter identification latencies. Although researchers agree that courtroom expressions of confidence are uninformative, our findings indicate that confidence assessments obtained immediately after a positive identification can provide a useful guide for investigators about the likely accuracy of an identification.

  11. Identification of targets for rational pharmacological therapy in childhood craniopharyngioma.

    PubMed

    Gump, Jacob M; Donson, Andrew M; Birks, Diane K; Amani, Vladimir M; Rao, Karun K; Griesinger, Andrea M; Kleinschmidt-DeMasters, B K; Johnston, James M; Anderson, Richard C E; Rosenfeld, Amy; Handler, Michael; Gore, Lia; Foreman, Nicholas; Hankinson, Todd C

    2015-05-21

    Pediatric adamantinomatous craniopharyngioma (ACP) is a histologically benign but clinically aggressive brain tumor that arises from the sellar/suprasellar region. Despite a high survival rate with current surgical and radiation therapy (75-95 % at 10 years), ACP is associated with debilitating visual, endocrine, neurocognitive and psychological morbidity, resulting in excheptionally poor quality of life for survivors. Identification of an effective pharmacological therapy could drastically decrease morbidity and improve long term outcomes for children with ACP. Using mRNA microarray gene expression analysis of 15 ACP patient samples, we have found several pharmaceutical targets that are significantly and consistently overexpressed in our panel of ACP relative to other pediatric brain tumors, pituitary tumors, normal pituitary and normal brain tissue. Among the most highly expressed are several targets of the kinase inhibitor dasatinib - LCK, EPHA2 and SRC; EGFR pathway targets - AREG, EGFR and ERBB3; and other potentially actionable cancer targets - SHH, MMP9 and MMP12. We confirm by western blot that a subset of these targets is highly expressed in ACP primary tumor samples. We report here the first published transcriptome for ACP and the identification of targets for rational therapy. Experimental drugs targeting each of these gene products are currently being tested clinically and pre-clinically for the treatment of other tumor types. This study provides a rationale for further pre-clinical and clinical studies of novel pharmacological treatments for ACP. Development of mouse and cell culture models for ACP will further enable the translation of these targets from the lab to the clinic, potentially ushering in a new era in the treatment of ACP.

  12. Reduced-order model for underwater target identification using proper orthogonal decomposition

    NASA Astrophysics Data System (ADS)

    Ramesh, Sai Sudha; Lim, Kian Meng

    2017-03-01

    Research on underwater acoustics has seen major development over the past decade due to its widespread applications in domains such as underwater communication/navigation (SONAR), seismic exploration and oceanography. In particular, acoustic signatures from partially or fully buried targets can be used in the identification of buried mines for mine counter measures (MCM). Although there exist several techniques to identify target properties based on SONAR images and acoustic signatures, these methods first employ a feature extraction method to represent the dominant characteristics of a data set, followed by the use of an appropriate classifier based on neural networks or the relevance vector machine. The aim of the present study is to demonstrate the applications of proper orthogonal decomposition (POD) technique in capturing dominant features of a set of scattered pressure signals, and subsequent use of the POD modes and coefficients in the identification of partially buried underwater target parameters such as its location, size and material density. Several numerical examples are presented to demonstrate the performance of the system identification method based on POD. Although the present study is based on 2D acoustic model, the method can be easily extended to 3D models and thereby enables cost-effective representations of large-scale data.

  13. Prioritization of Biomarker Targets in Human Umbilical Cord Blood: Identification of Proteins in Infant Blood Serving as Validated Biomarkers in Adults

    PubMed Central

    Hansmeier, Nicole; Chao, Tzu-Chiao; Goldman, Lynn R.; Witter, Frank R.

    2012-01-01

    Background: Early diagnosis represents one of the best lines of defense in the fight against a wide array of human diseases. Umbilical cord blood (UCB) is one of the first easily available diagnostic biofluids and can inform about the health status of newborns. However, compared with adult blood, its diagnostic potential remains largely untapped. Objectives: Our goal was to accelerate biomarker research on UCB by exploring its detectable protein content and providing a priority list of potential biomarkers based on known proteins involved in disease pathways. Methods: We explored cord blood serum proteins by profiling a UCB pool of 12 neonates with different backgrounds using a combination of isoelectric focusing and liquid chromatography coupled with matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-MS/MS) and by comparing results with information contained in metabolic and disease databases available for adult blood. Results: A total of 1,210 UCB proteins were identified with a protein-level false discovery rate of ~ 5% as estimated by naïve target-decoy and MAYU approaches, signifying a 6-fold increase in the number of UCB proteins described to date. Identified proteins correspond to 138 different metabolic and disease pathways and provide a platform of mechanistically linked biomarker candidates for tracking disruptions in cellular processes. Moreover, among the identified proteins, 38 were found to be approved biomarkers for adult blood. Conclusions: The results of this study advance current knowledge of the human cord blood serum proteome. They showcase the potential of UCB as a diagnostic medium for assessing infant health by detection and identification of candidate biomarkers for known disease pathways using a global, nontargeted approach. These biomarkers may inform about mechanisms of exposure–disease relationships. Furthermore, biomarkers approved by the U.S. Food and Drug Administration for screening in adult blood were

  14. Identification of ground targets from airborne platforms

    NASA Astrophysics Data System (ADS)

    Doe, Josh; Boettcher, Evelyn; Miller, Brian

    2009-05-01

    The US Army RDECOM CERDEC Night Vision and Electronic Sensors Directorate (NVESD) sensor performance models predict the ability of soldiers to perform a specified military discrimination task using an EO/IR sensor system. Increasingly EO/IR systems are being used on manned and un-manned aircraft for surveillance and target acquisition tasks. In response to this emerging requirement, the NVESD Modeling and Simulation division has been tasked to compare target identification performance between ground-to-ground and air-to-ground platforms for both IR and visible spectra for a set of wheeled utility vehicles. To measure performance, several forced choice experiments were designed and administered and the results analyzed. This paper describes these experiments and reports the results as well as the NVTherm model calibration factors derived for the infrared imagery.

  15. Target identification using Zernike moments and neural networks

    NASA Astrophysics Data System (ADS)

    Azimi-Sadjadi, Mahmood R.; Jamshidi, Arta A.; Nevis, Andrew J.

    2001-10-01

    The development of an underwater target identification algorithm capable of identifying various types of underwater targets, such as mines, under different environmental conditions pose many technical problems. Some of the contributing factors are: targets have diverse sizes, shapes and reflectivity properties. Target emplacement environment is variable; targets may be proud or partially buried. Environmental properties vary significantly from one location to another. Bottom features such as sand, rocks, corals, and vegetation can conceal a target whether it is partially buried or proud. Competing clutter with responses that closely resemble those of the targets may lead to false positives. All the problems mentioned above contribute to overly difficult and challenging conditions that could lead to unreliable algorithm performance with existing methods. In this paper, we developed and tested a shape-dependent feature extraction scheme that provides features invariant to rotation, size scaling and translation; properties that are extremely useful for any target classification problem. The developed schemes were tested on an electro-optical imagery data set collected under different environmental conditions with variable background, range and target types. The electro-optic data set was collected using a Laser Line Scan (LLS) sensor by the Coastal Systems Station (CSS), located in Panama City, Florida. The performance of the developed scheme and its robustness to distortion, rotation, scaling and translation was also studied.

  16. Hierarchical relaxation methods for multispectral pixel classification as applied to target identification

    NASA Astrophysics Data System (ADS)

    Cohen, E. A., Jr.

    1985-02-01

    This report provides insights into the approaches toward image modeling as applied to target detection. The approach is that of examining the energy in prescribed wave-bands which emanate from a target and correlating the emissions. Typically, one might be looking at two or three infrared bands, possibly together with several visual bands. The target is segmented, using both first and second order modeling, into a set of interesting components and these components are correlated so as to enhance the classification process. A Markov-type model is used to provide an a priori assessment of the spatial relationships among critical parts of the target, and a stochastic model using the output of an initial probabilistic labeling is invoked. The tradeoff between this stochastic model and the Markov model is then optimized to yield a best labeling for identification purposes. In an identification of friend or foe (IFF) context, this methodology could be of interest, for it provides the ingredients for such a higher level of understanding.

  17. Uridine monophosphate kinase as potential target for tuberculosis: from target to lead identification.

    PubMed

    Arvind, Akanksha; Jain, Vaibhav; Saravanan, Parameswaran; Mohan, C Gopi

    2013-12-01

    Mycobacterium tuberculosis (Mtb) is a causative agent of tuberculosis (TB) disease, which has affected approximately 2 billion people worldwide. Due to the emergence of resistance towards the existing drugs, discovery of new anti-TB drugs is an important global healthcare challenge. To address this problem, there is an urgent need to identify new drug targets in Mtb. In the present study, the subtractive genomics approach has been employed for the identification of new drug targets against TB. Screening the Mtb proteome using the Database of Essential Genes (DEG) and human proteome resulted in the identification of 60 key proteins which have no eukaryotic counterparts. Critical analysis of these proteins using Kyoto Encyclopedia of Genes and Genomes (KEGG) metabolic pathways database revealed uridine monophosphate kinase (UMPK) enzyme as a potential drug target for developing novel anti-TB drugs. Homology model of Mtb-UMPK was constructed for the first time on the basis of the crystal structure of E. coli-UMPK, in order to understand its structure-function relationships, and which would in turn facilitate to perform structure-based inhibitor design. Furthermore, the structural similarity search was carried out using physiological inhibitor UTP of Mtb-UMPK to virtually screen ZINC database. Retrieved hits were further screened by implementing several filters like ADME and toxicity followed by molecular docking. Finally, on the basis of the Glide docking score and the mode of binding, 6 putative leads were identified as inhibitors of this enzyme which can potentially emerge as future drugs for the treatment of TB.

  18. Photoaffinity labeling in target- and binding-site identification

    PubMed Central

    Smith, Ewan; Collins, Ian

    2015-01-01

    Photoaffinity labeling (PAL) using a chemical probe to covalently bind its target in response to activation by light has become a frequently used tool in drug discovery for identifying new drug targets and molecular interactions, and for probing the location and structure of binding sites. Methods to identify the specific target proteins of hit molecules from phenotypic screens are highly valuable in early drug discovery. In this review, we summarize the principles of PAL including probe design and experimental techniques for in vitro and live cell investigations. We emphasize the need to optimize and validate probes and highlight examples of the successful application of PAL across multiple disease areas. PMID:25686004

  19. Identification of STAT target genes in adipocytes

    PubMed Central

    Zhao, Peng; Stephens, Jacqueline M.

    2013-01-01

    Adipocytes play important roles in lipid storage, energy homeostasis and whole body insulin sensitivity. Studies in the last two decades have identified the hormones and cytokines that activate specific STATs in adipocytes in vitro and in vivo. Five of the seven STAT family members are expressed in adipocyte (STATs 1, 3, 5A, 5B and 6). Many transcription factors, including STATs, have been shown to play an important role in adipose tissue development and function. This review will summarize the importance of adipocytes, indicate the cytokines and hormones that utilize the JAK-STAT signaling pathway in fat cells and focus on the identification of STAT target genes in mature adipocytes. To date, specific target genes have been identified for STATs, 1, 5A and 5B, but not for STATs 3 and 6. PMID:24058802

  20. Complementary Approaches to Existing Target Based Drug Discovery for Identifying Novel Drug Targets.

    PubMed

    Vasaikar, Suhas; Bhatia, Pooja; Bhatia, Partap G; Chu Yaiw, Koon

    2016-11-21

    In the past decade, it was observed that the relationship between the emerging New Molecular Entities and the quantum of R&D investment has not been favorable. There might be numerous reasons but few studies stress the introduction of target based drug discovery approach as one of the factors. Although a number of drugs have been developed with an emphasis on a single protein target, yet identification of valid target is complex. The approach focuses on an in vitro single target, which overlooks the complexity of cell and makes process of validation drug targets uncertain. Thus, it is imperative to search for alternatives rather than looking at success stories of target-based drug discovery. It would be beneficial if the drugs were developed to target multiple components. New approaches like reverse engineering and translational research need to take into account both system and target-based approach. This review evaluates the strengths and limitations of known drug discovery approaches and proposes alternative approaches for increasing efficiency against treatment.

  1. Potentials of single-cell biology in identification and validation of disease biomarkers.

    PubMed

    Niu, Furong; Wang, Diane C; Lu, Jiapei; Wu, Wei; Wang, Xiangdong

    2016-09-01

    Single-cell biology is considered a new approach to identify and validate disease-specific biomarkers. However, the concern raised by clinicians is how to apply single-cell measurements for clinical practice, translate the message of single-cell systems biology into clinical phenotype or explain alterations of single-cell gene sequencing and function in patient response to therapies. This study is to address the importance and necessity of single-cell gene sequencing in the identification and development of disease-specific biomarkers, the definition and significance of single-cell biology and single-cell systems biology in the understanding of single-cell full picture, the development and establishment of whole-cell models in the validation of targeted biological function and the figure and meaning of single-molecule imaging in single cell to trace intra-single-cell molecule expression, signal, interaction and location. We headline the important role of single-cell biology in the discovery and development of disease-specific biomarkers with a special emphasis on understanding single-cell biological functions, e.g. mechanical phenotypes, single-cell biology, heterogeneity and organization of genome function. We have reason to believe that such multi-dimensional, multi-layer, multi-crossing and stereoscopic single-cell biology definitely benefits the discovery and development of disease-specific biomarkers. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  2. Enhancing bioactive peptide release and identification using targeted enzymatic hydrolysis of milk proteins.

    PubMed

    Nongonierma, Alice B; FitzGerald, Richard J

    2018-06-01

    Milk proteins have been extensively studied for their ability to yield a range of bioactive peptides following enzymatic hydrolysis/digestion. However, many hurdles still exist regarding the widespread utilization of milk protein-derived bioactive peptides as health enhancing agents for humans. These mostly arise from the fact that most milk protein-derived bioactive peptides are not highly potent. In addition, they may be degraded during gastrointestinal digestion and/or have a low intestinal permeability. The targeted release of bioactive peptides during the enzymatic hydrolysis of milk proteins may allow the generation of particularly potent bioactive hydrolysates and peptides. Therefore, the development of milk protein hydrolysates capable of improving human health requires, in the first instance, optimized targeted release of specific bioactive peptides. The targeted hydrolysis of milk proteins has been aided by a range of in silico tools. These include peptide cutters and predictive modeling linking bioactivity to peptide structure [i.e., molecular docking, quantitative structure activity relationship (QSAR)], or hydrolysis parameters [design of experiments (DOE)]. Different targeted enzymatic release strategies employed during the generation of milk protein hydrolysates are reviewed herein and their limitations are outlined. In addition, specific examples are provided to demonstrate how in silico tools may help in the identification and discovery of potent milk protein-derived peptides. It is anticipated that the development of novel strategies employing a range of in silico tools may help in the generation of milk protein hydrolysates containing potent and bioavailable peptides, which in turn may be used to validate their health promoting effects in humans. Graphical abstract The targeted enzymatic hydrolysis of milk proteins may allow the generation of highly potent and bioavailable bioactive peptides.

  3. Proteomics meets blood banking: identification of protein targets for the improvement of platelet quality.

    PubMed

    Schubert, Peter; Devine, Dana V

    2010-01-03

    Proteomics has brought new perspectives to the fields of hematology and transfusion medicine in the last decade. The steady improvement of proteomic technology is propelling novel discoveries of molecular mechanisms by studying protein expression, post-translational modifications and protein interactions. This review article focuses on the application of proteomics to the identification of molecular mechanisms leading to the deterioration of blood platelets during storage - a critical aspect in the provision of platelet transfusion products. Several proteomic approaches have been employed to analyse changes in the platelet protein profile during storage and the obtained data now need to be translated into platelet biochemistry in order to connect the results to platelet function. Targeted biochemical applications then allow the identification of points for intervention in signal transduction pathways. Once validated and placed in a transfusion context, these data will provide further understanding of the underlying molecular mechanisms leading to platelet storage lesion. Future aspects of proteomics in blood banking will aim to make use of protein markers identified for platelet storage lesion development to monitor proteome changes when alterations such as the use of additive solutions or pathogen reduction strategies are put in place in order to improve platelet quality for patients. (c) 2009 Elsevier B.V. All rights reserved.

  4. Automatic identification of bird targets with radar via patterns produced by wing flapping.

    PubMed

    Zaugg, Serge; Saporta, Gilbert; van Loon, Emiel; Schmaljohann, Heiko; Liechti, Felix

    2008-09-06

    Bird identification with radar is important for bird migration research, environmental impact assessments (e.g. wind farms), aircraft security and radar meteorology. In a study on bird migration, radar signals from birds, insects and ground clutter were recorded. Signals from birds show a typical pattern due to wing flapping. The data were labelled by experts into the four classes BIRD, INSECT, CLUTTER and UFO (unidentifiable signals). We present a classification algorithm aimed at automatic recognition of bird targets. Variables related to signal intensity and wing flapping pattern were extracted (via continuous wavelet transform). We used support vector classifiers to build predictive models. We estimated classification performance via cross validation on four datasets. When data from the same dataset were used for training and testing the classifier, the classification performance was extremely to moderately high. When data from one dataset were used for training and the three remaining datasets were used as test sets, the performance was lower but still extremely to moderately high. This shows that the method generalizes well across different locations or times. Our method provides a substantial gain of time when birds must be identified in large collections of radar signals and it represents the first substantial step in developing a real time bird identification radar system. We provide some guidelines and ideas for future research.

  5. Comparative genomics identification of a novel set of temporally regulated hedgehog target genes in the retina.

    PubMed

    McNeill, Brian; Perez-Iratxeta, Carol; Mazerolle, Chantal; Furimsky, Marosh; Mishina, Yuji; Andrade-Navarro, Miguel A; Wallace, Valerie A

    2012-03-01

    The hedgehog (Hh) signaling pathway is involved in numerous developmental and adult processes with many links to cancer. In vertebrates, the activity of the Hh pathway is mediated primarily through three Gli transcription factors (Gli1, 2 and 3) that can serve as transcriptional activators or repressors. The identification of Gli target genes is essential for the understanding of the Hh-mediated processes. We used a comparative genomics approach using the mouse and human genomes to identify 390 genes that contained conserved Gli binding sites. RT-qPCR validation of 46 target genes in E14.5 and P0.5 retinal explants revealed that Hh pathway activation resulted in the modulation of 30 of these targets, 25 of which demonstrated a temporal regulation. Further validation revealed that the expression of Bok, FoxA1, Sox8 and Wnt7a was dependent upon Sonic Hh (Shh) signaling in the retina and their regulation is under positive and negative controls by Gli2 and Gli3, respectively. We also show using chromatin immunoprecipitation that Gli2 binds to the Sox8 promoter, suggesting that Sox8 is an Hh-dependent direct target of Gli2. Finally, we demonstrate that the Hh pathway also modulates the expression of Sox9 and Sox10, which together with Sox8 make up the SoxE group. Previously, it has been shown that Hh and SoxE group genes promote Müller glial cell development in the retina. Our data are consistent with the possibility for a role of SoxE group genes downstream of Hh signaling on Müller cell development. Crown Copyright © 2012. Published by Elsevier Inc. All rights reserved.

  6. Development and Validation of Targeted Next-Generation Sequencing Panels for Detection of Germline Variants in Inherited Diseases.

    PubMed

    Santani, Avni; Murrell, Jill; Funke, Birgit; Yu, Zhenming; Hegde, Madhuri; Mao, Rong; Ferreira-Gonzalez, Andrea; Voelkerding, Karl V; Weck, Karen E

    2017-06-01

    - The number of targeted next-generation sequencing (NGS) panels for genetic diseases offered by clinical laboratories is rapidly increasing. Before an NGS-based test is implemented in a clinical laboratory, appropriate validation studies are needed to determine the performance characteristics of the test. - To provide examples of assay design and validation of targeted NGS gene panels for the detection of germline variants associated with inherited disorders. - The approaches used by 2 clinical laboratories for the development and validation of targeted NGS gene panels are described. Important design and validation considerations are examined. - Clinical laboratories must validate performance specifications of each test prior to implementation. Test design specifications and validation data are provided, outlining important steps in validation of targeted NGS panels by clinical diagnostic laboratories.

  7. Sleep and eyewitness memory: Fewer false identifications after sleep when the target is absent from the lineup.

    PubMed

    Stepan, Michelle E; Dehnke, Taylor M; Fenn, Kimberly M

    2017-01-01

    Inaccurate eyewitness identifications are the leading cause of known false convictions in the United States. Moreover, improving eyewitness memory is difficult and often unsuccessful. Sleep consistently strengthens and protects memory from interference, particularly when a recall test is used. However, the effect of sleep on recognition memory is more equivocal. Eyewitness identification tests are often recognition based, thus leaving open the question of how sleep affects recognition performance in an eyewitness context. In the current study, we investigated the effect of sleep on eyewitness memory. Participants watched a video of a mock-crime and attempted to identify the perpetrator from a simultaneous lineup after a 12-hour retention interval that either spanned a waking day or night of sleep. In Experiment 1, we used a target-present lineup and, in Experiment 2, we used a target-absent lineup in order to investigate correct and false identifications, respectively. Sleep reduced false identifications in the target-absent lineup (Experiment 2) but had no effect on correct identifications in the target-present lineup (Experiment 1). These results are discussed with respect to memory strength and decision making strategies.

  8. Sleep and eyewitness memory: Fewer false identifications after sleep when the target is absent from the lineup

    PubMed Central

    Dehnke, Taylor M.; Fenn, Kimberly M.

    2017-01-01

    Inaccurate eyewitness identifications are the leading cause of known false convictions in the United States. Moreover, improving eyewitness memory is difficult and often unsuccessful. Sleep consistently strengthens and protects memory from interference, particularly when a recall test is used. However, the effect of sleep on recognition memory is more equivocal. Eyewitness identification tests are often recognition based, thus leaving open the question of how sleep affects recognition performance in an eyewitness context. In the current study, we investigated the effect of sleep on eyewitness memory. Participants watched a video of a mock-crime and attempted to identify the perpetrator from a simultaneous lineup after a 12-hour retention interval that either spanned a waking day or night of sleep. In Experiment 1, we used a target-present lineup and, in Experiment 2, we used a target-absent lineup in order to investigate correct and false identifications, respectively. Sleep reduced false identifications in the target-absent lineup (Experiment 2) but had no effect on correct identifications in the target-present lineup (Experiment 1). These results are discussed with respect to memory strength and decision making strategies. PMID:28877169

  9. Validation of photo-identification as a mark-recapture method in the spotted eagle ray Aetobatus narinari.

    PubMed

    González-Ramos, M S; Santos-Moreno, A; Rosas-Alquicira, E F; Fuentes-Mascorro, G

    2017-03-01

    The spotted eagle ray Aetobatus narinari is characterized by pigmentation patterns that are retained for up to 3·5 years. These pigmentations can be used to identify individuals through photo-identification. Only one study has validated this technique, but no study has estimated the percentage of correct identification of the rays using this technique. In order to carry out demographic research, a reliable photographic identification technique is needed. To achieve this validation for A. narinari, a double-mark system was established over 11 months and photographs of the dorsal surface of 191 rays were taken. Three body parts with distinctive natural patterns were analysed (dorsal surface of the cephalic region, dorsal surface of the pectoral fins and dorsal surface of the pelvic fins) in order to determine the body part that could be used to give the highest percentage of correct identification. The dorsal surface of the pectoral fins of A. narinari provides the most accurate photo-identification to distinguish individuals (88·2%). © 2016 The Fisheries Society of the British Isles.

  10. Identification of broad-spectrum antiviral compounds and assessment of the druggability of their target for efficacy against respiratory syncytial virus (RSV)

    PubMed Central

    Bonavia, Aurelio; Franti, Michael; Pusateri Keaney, Erin; Kuhen, Kelli; Seepersaud, Mohindra; Radetich, Branko; Shao, Jian; Honda, Ayako; Dewhurst, Janetta; Balabanis, Kara; Monroe, James; Wolff, Karen; Osborne, Colin; Lanieri, Leanne; Hoffmaster, Keith; Amin, Jakal; Markovits, Judit; Broome, Michelle; Skuba, Elizabeth; Cornella-Taracido, Ivan; Joberty, Gerard; Bouwmeester, Tewis; Hamann, Lawrence; Tallarico, John A.; Tommasi, Ruben; Compton, Teresa; Bushell, Simon M.

    2011-01-01

    The search for novel therapeutic interventions for viral disease is a challenging pursuit, hallmarked by the paucity of antiviral agents currently prescribed. Targeting of viral proteins has the inextricable challenge of rise of resistance. Safe and effective vaccines are not possible for many viral pathogens. New approaches are required to address the unmet medical need in this area. We undertook a cell-based high-throughput screen to identify leads for development of drugs to treat respiratory syncytial virus (RSV), a serious pediatric pathogen. We identified compounds that are potent (nanomolar) inhibitors of RSV in vitro in HEp-2 cells and in primary human bronchial epithelial cells and were shown to act postentry. Interestingly, two scaffolds exhibited broad-spectrum activity among multiple RNA viruses. Using the chemical matter as a probe, we identified the targets and identified a common cellular pathway: the de novo pyrimidine biosynthesis pathway. Both targets were validated in vitro and showed no significant cell cytotoxicity except for activity against proliferative B- and T-type lymphoid cells. Corollary to this finding was to understand the consequences of inhibition of the target to the host. An in vivo assessment for antiviral efficacy failed to demonstrate reduced viral load, but revealed microscopic changes and a trend toward reduced pyrimidine pools and findings in histopathology. We present here a discovery program that includes screen, target identification, validation, and druggability that can be broadly applied to identify and interrogate other host factors for antiviral effect starting from chemical matter of unknown target/mechanism of action. PMID:21502533

  11. Identification of broad-spectrum antiviral compounds and assessment of the druggability of their target for efficacy against respiratory syncytial virus (RSV).

    PubMed

    Bonavia, Aurelio; Franti, Michael; Pusateri Keaney, Erin; Kuhen, Kelli; Seepersaud, Mohindra; Radetich, Branko; Shao, Jian; Honda, Ayako; Dewhurst, Janetta; Balabanis, Kara; Monroe, James; Wolff, Karen; Osborne, Colin; Lanieri, Leanne; Hoffmaster, Keith; Amin, Jakal; Markovits, Judit; Broome, Michelle; Skuba, Elizabeth; Cornella-Taracido, Ivan; Joberty, Gerard; Bouwmeester, Tewis; Hamann, Lawrence; Tallarico, John A; Tommasi, Ruben; Compton, Teresa; Bushell, Simon M

    2011-04-26

    The search for novel therapeutic interventions for viral disease is a challenging pursuit, hallmarked by the paucity of antiviral agents currently prescribed. Targeting of viral proteins has the inextricable challenge of rise of resistance. Safe and effective vaccines are not possible for many viral pathogens. New approaches are required to address the unmet medical need in this area. We undertook a cell-based high-throughput screen to identify leads for development of drugs to treat respiratory syncytial virus (RSV), a serious pediatric pathogen. We identified compounds that are potent (nanomolar) inhibitors of RSV in vitro in HEp-2 cells and in primary human bronchial epithelial cells and were shown to act postentry. Interestingly, two scaffolds exhibited broad-spectrum activity among multiple RNA viruses. Using the chemical matter as a probe, we identified the targets and identified a common cellular pathway: the de novo pyrimidine biosynthesis pathway. Both targets were validated in vitro and showed no significant cell cytotoxicity except for activity against proliferative B- and T-type lymphoid cells. Corollary to this finding was to understand the consequences of inhibition of the target to the host. An in vivo assessment for antiviral efficacy failed to demonstrate reduced viral load, but revealed microscopic changes and a trend toward reduced pyrimidine pools and findings in histopathology. We present here a discovery program that includes screen, target identification, validation, and druggability that can be broadly applied to identify and interrogate other host factors for antiviral effect starting from chemical matter of unknown target/mechanism of action.

  12. The Development and Validation of a Life Experience Inventory for the Identification of Creative Electrical Engineers.

    ERIC Educational Resources Information Center

    Michael, William B.; Colson, Kenneth R.

    1979-01-01

    The construction and validation of the Life Experience Inventory (LEI) for the identification of creative electrical engineers are described. Using the number of patents held or pending as a criterion measure, the LEI was found to have high concurrent validity. (JKS)

  13. Leveraging 3D chemical similarity, target and phenotypic data in the identification of drug-protein and drug-adverse effect associations.

    PubMed

    Vilar, Santiago; Hripcsak, George

    2016-01-01

    Drug-target identification is crucial to discover novel applications for existing drugs and provide more insights about mechanisms of biological actions, such as adverse drug effects (ADEs). Computational methods along with the integration of current big data sources provide a useful framework for drug-target and drug-adverse effect discovery. In this article, we propose a method based on the integration of 3D chemical similarity, target and adverse effect data to generate a drug-target-adverse effect predictor along with a simple leveraging system to improve identification of drug-targets and drug-adverse effects. In the first step, we generated a system for multiple drug-target identification based on the application of 3D drug similarity into a large target dataset extracted from the ChEMBL. Next, we developed a target-adverse effect predictor combining targets from ChEMBL with phenotypic information provided by SIDER data source. Both modules were linked to generate a final predictor that establishes hypothesis about new drug-target-adverse effect candidates. Additionally, we showed that leveraging drug-target candidates with phenotypic data is very useful to improve the identification of drug-targets. The integration of phenotypic data into drug-target candidates yielded up to twofold precision improvement. In the opposite direction, leveraging drug-phenotype candidates with target data also yielded a significant enhancement in the performance. The modeling described in the current study is simple and efficient and has applications at large scale in drug repurposing and drug safety through the identification of mechanism of action of biological effects.

  14. A distributed computational search strategy for the identification of diagnostics targets: application to finding aptamer targets for methicillin-resistant staphylococci.

    PubMed

    Flanagan, Keith; Cockell, Simon; Harwood, Colin; Hallinan, Jennifer; Nakjang, Sirintra; Lawry, Beth; Wipat, Anil

    2014-06-30

    The rapid and cost-effective identification of bacterial species is crucial, especially for clinical diagnosis and treatment. Peptide aptamers have been shown to be valuable for use as a component of novel, direct detection methods. These small peptides have a number of advantages over antibodies, including greater specificity and longer shelf life. These properties facilitate their use as the detector components of biosensor devices. However, the identification of suitable aptamer targets for particular groups of organisms is challenging. We present a semi-automated processing pipeline for the identification of candidate aptamer targets from whole bacterial genome sequences. The pipeline can be configured to search for protein sequence fragments that uniquely identify a set of strains of interest. The system is also capable of identifying additional organisms that may be of interest due to their possession of protein fragments in common with the initial set. Through the use of Cloud computing technology and distributed databases, our system is capable of scaling with the rapidly growing genome repositories, and consequently of keeping the resulting data sets up-to-date. The system described is also more generically applicable to the discovery of specific targets for other diagnostic approaches such as DNA probes, PCR primers and antibodies.

  15. A distributed computational search strategy for the identification of diagnostics targets: Application to finding aptamer targets for methicillin-resistant staphylococci.

    PubMed

    Flanagan, Keith; Cockell, Simon; Harwood, Colin; Hallinan, Jennifer; Nakjang, Sirintra; Lawry, Beth; Wipat, Anil

    2014-06-01

    The rapid and cost-effective identification of bacterial species is crucial, especially for clinical diagnosis and treatment. Peptide aptamers have been shown to be valuable for use as a component of novel, direct detection methods. These small peptides have a number of advantages over antibodies, including greater specificity and longer shelf life. These properties facilitate their use as the detector components of biosensor devices. However, the identification of suitable aptamer targets for particular groups of organisms is challenging. We present a semi-automated processing pipeline for the identification of candidate aptamer targets from whole bacterial genome sequences. The pipeline can be configured to search for protein sequence fragments that uniquely identify a set of strains of interest. The system is also capable of identifying additional organisms that may be of interest due to their possession of protein fragments in common with the initial set. Through the use of Cloud computing technology and distributed databases, our system is capable of scaling with the rapidly growing genome repositories, and consequently of keeping the resulting data sets up-to-date. The system described is also more generically applicable to the discovery of specific targets for other diagnostic approaches such as DNA probes, PCR primers and antibodies.

  16. Feasibility of CRISPR-Cas9-Based In Vitro Drug Target Identification for Personalized Prostate Cancer Medicine

    DTIC Science & Technology

    2017-09-01

    AWARD NUMBER: W81XWH-16-1-0502 TITLE: Feasibility of CRISPR -Cas9-Based In Vitro Drug Target Identification for Personalized Prostate Cancer Medicine...CONTRACT NUMBER Feasibility of CRISPR -Cas9-Based In Vitro Drug Target Identification for Personalized Prostate Cancer Medicine 5b. GRANT NUMBER...Approved for Public Release; Distribution Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT This study tests the feasibility of using CRISPR -Cas9 to

  17. Identification of neuronal target genes for CCAAT/Enhancer Binding Proteins

    PubMed Central

    Kfoury, N.; Kapatos, G.

    2009-01-01

    CCAAT/Enhancer Binding Proteins (C/EBPs) play pivotal roles in development and plasticity of the nervous system. Identification of the physiological targets of C/EBPs (C/EBP target genes) should therefore provide insight into the underlying biology of these processes. We used unbiased genome-wide mapping to identify 115 C/EBPβ target genes in PC12 cells that include transcription factors, neurotransmitter receptors, ion channels, protein kinases and synaptic vesicle proteins. C/EBPβ binding sites were located primarily within introns, suggesting novel regulatory functions, and were associated with binding sites for other developmentally important transcription factors. Experiments using dominant negatives showed C/EBPβ to repress transcription of a subset of target genes. Target genes in rat brain were subsequently found to preferentially bind C/EBPα, β and δ. Analysis of the hippocampal transcriptome of C/EBPβ knockout mice revealed dysregulation of a high percentage of transcripts identified as C/EBP target genes. These results support the hypothesis that C/EBPs play non-redundant roles in the brain. PMID:19103292

  18. Review of validation and reporting of non-targeted fingerprinting approaches for food authentication.

    PubMed

    Riedl, Janet; Esslinger, Susanne; Fauhl-Hassek, Carsten

    2015-07-23

    Food fingerprinting approaches are expected to become a very potent tool in authentication processes aiming at a comprehensive characterization of complex food matrices. By non-targeted spectrometric or spectroscopic chemical analysis with a subsequent (multivariate) statistical evaluation of acquired data, food matrices can be investigated in terms of their geographical origin, species variety or possible adulterations. Although many successful research projects have already demonstrated the feasibility of non-targeted fingerprinting approaches, their uptake and implementation into routine analysis and food surveillance is still limited. In many proof-of-principle studies, the prediction ability of only one data set was explored, measured within a limited period of time using one instrument within one laboratory. Thorough validation strategies that guarantee reliability of the respective data basis and that allow conclusion on the applicability of the respective approaches for its fit-for-purpose have not yet been proposed. Within this review, critical steps of the fingerprinting workflow were explored to develop a generic scheme for multivariate model validation. As a result, a proposed scheme for "good practice" shall guide users through validation and reporting of non-targeted fingerprinting results. Furthermore, food fingerprinting studies were selected by a systematic search approach and reviewed with regard to (a) transparency of data processing and (b) validity of study results. Subsequently, the studies were inspected for measures of statistical model validation, analytical method validation and quality assurance measures. In this context, issues and recommendations were found that might be considered as an actual starting point for developing validation standards of non-targeted metabolomics approaches for food authentication in the future. Hence, this review intends to contribute to the harmonization and standardization of food fingerprinting, both

  19. Targeted nanodiamonds for identification of subcellular protein assemblies in mammalian cells

    PubMed Central

    Lake, Michael P.; Bouchard, Louis-S.

    2017-01-01

    Transmission electron microscopy (TEM) can be used to successfully determine the structures of proteins. However, such studies are typically done ex situ after extraction of the protein from the cellular environment. Here we describe an application for nanodiamonds as targeted intensity contrast labels in biological TEM, using the nuclear pore complex (NPC) as a model macroassembly. We demonstrate that delivery of antibody-conjugated nanodiamonds to live mammalian cells using maltotriose-conjugated polypropylenimine dendrimers results in efficient localization of nanodiamonds to the intended cellular target. We further identify signatures of nanodiamonds under TEM that allow for unambiguous identification of individual nanodiamonds from a resin-embedded, OsO4-stained environment. This is the first demonstration of nanodiamonds as labels for nanoscale TEM-based identification of subcellular protein assemblies. These results, combined with the unique fluorescence properties and biocompatibility of nanodiamonds, represent an important step toward the use of nanodiamonds as markers for correlated optical/electron bioimaging. PMID:28636640

  20. Comparison of conventional ultrasonography and ultrasonography-computed tomography fusion imaging for target identification using digital/real hybrid phantoms: a preliminary study.

    PubMed

    Soyama, Takeshi; Sakuhara, Yusuke; Kudo, Kohsuke; Abo, Daisuke; Wang, Jeff; Ito, Yoichi M; Hasegawa, Yu; Shirato, Hiroki

    2016-07-01

    This preliminary study compared ultrasonography-computed tomography (US-CT) fusion imaging and conventional ultrasonography (US) for accuracy and time required for target identification using a combination of real phantoms and sets of digitally modified computed tomography (CT) images (digital/real hybrid phantoms). In this randomized prospective study, 27 spheres visible on B-mode US were placed at depths of 3.5, 8.5, and 13.5 cm (nine spheres each). All 27 spheres were digitally erased from the CT images, and a radiopaque sphere was digitally placed at each of the 27 locations to create 27 different sets of CT images. Twenty clinicians were instructed to identify the sphere target using US alone and fusion imaging. The accuracy of target identification of the two methods was compared using McNemar's test. The mean time required for target identification and error distances were compared using paired t tests. At all three depths, target identification was more accurate and the mean time required for target identification was significantly less with US-CT fusion imaging than with US alone, and the mean error distances were also shorter with US-CT fusion imaging. US-CT fusion imaging was superior to US alone in terms of accurate and rapid identification of target lesions.

  1. Identification of microRNAs with regulatory potential using a matched microRNA-mRNA time-course data.

    PubMed

    Jayaswal, Vivek; Lutherborrow, Mark; Ma, David D F; Hwa Yang, Yee

    2009-05-01

    Over the past decade, a class of small RNA molecules called microRNAs (miRNAs) has been shown to regulate gene expression at the post-transcription stage. While early work focused on the identification of miRNAs using a combination of experimental and computational techniques, subsequent studies have focused on identification of miRNA-target mRNA pairs as each miRNA can have hundreds of mRNA targets. The experimental validation of some miRNAs as oncogenic has provided further motivation for research in this area. In this article we propose an odds-ratio (OR) statistic for identification of regulatory miRNAs. It is based on integrative analysis of matched miRNA and mRNA time-course microarray data. The OR-statistic was used for (i) identification of miRNAs with regulatory potential, (ii) identification of miRNA-target mRNA pairs and (iii) identification of time lags between changes in miRNA expression and those of its target mRNAs. We applied the OR-statistic to a cancer data set and identified a small set of miRNAs that were negatively correlated to mRNAs. A literature survey revealed that some of the miRNAs that were predicted to be regulatory, were indeed oncogenic or tumor suppressors. Finally, some of the predicted miRNA targets have been shown to be experimentally valid.

  2. Identification of HMX1 target genes: A predictive promoter model approach

    PubMed Central

    Boulling, Arnaud; Wicht, Linda

    2013-01-01

    Purpose A homozygous mutation in the H6 family homeobox 1 (HMX1) gene is responsible for a new oculoauricular defect leading to eye and auricular developmental abnormalities as well as early retinal degeneration (MIM 612109). However, the HMX1 pathway remains poorly understood, and in the first approach to better understand the pathway’s function, we sought to identify the target genes. Methods We developed a predictive promoter model (PPM) approach using a comparative transcriptomic analysis in the retina at P15 of a mouse model lacking functional Hmx1 (dmbo mouse) and its respective wild-type. This PPM was based on the hypothesis that HMX1 binding site (HMX1-BS) clusters should be more represented in promoters of HMX1 target genes. The most differentially expressed genes in the microarray experiment that contained HMX1-BS clusters were used to generate the PPM, which was then statistically validated. Finally, we developed two genome-wide target prediction methods: one that focused on conserving PPM features in human and mouse and one that was based on the co-occurrence of HMX1-BS pairs fitting the PPM, in human or in mouse, independently. Results The PPM construction revealed that sarcoglycan, gamma (35kDa dystrophin-associated glycoprotein) (Sgcg), teashirt zinc finger homeobox 2 (Tshz2), and solute carrier family 6 (neurotransmitter transporter, glycine) (Slc6a9) genes represented Hmx1 targets in the mouse retina at P15. Moreover, the genome-wide target prediction revealed that mouse genes belonging to the retinal axon guidance pathway were targeted by Hmx1. Expression of these three genes was experimentally validated using a quantitative reverse transcription PCR approach. The inhibitory activity of Hmx1 on Sgcg, as well as protein tyrosine phosphatase, receptor type, O (Ptpro) and Sema3f, two targets identified by the PPM, were validated with luciferase assay. Conclusions Gene expression analysis between wild-type and dmbo mice allowed us to develop a PPM

  3. An enhanced targeted identification strategy for the selective identification of flavonoid O-glycosides from Carthamus tinctorius by integrating offline two-dimensional liquid chromatography/linear ion-trap-Orbitrap mass spectrometry, high-resolution diagnostic product ions/neutral loss filtering and liquid chromatography-solid phase extraction-nuclear magnetic resonance.

    PubMed

    Yao, Chang-Liang; Yang, Wen-Zhi; Si, Wei; Shen, Yao; Zhang, Nai-Xia; Chen, Hua-Li; Pan, Hui-Qin; Yang, Min; Wu, Wan-Ying; Guo, De-An

    2017-03-31

    Targeted identification of potentially bioactive molecules from herbal medicines is often stymied by the insufficient chromatographic separation, ubiquitous matrix interference, and pervasive isomerism. An enhanced targeted identification strategy is presented and validated by the selective identification of flavonoid O-glycosides (FOGs) from Carthamus tinctorius. It consists of four steps: (i) enhanced separation and detection by offline two-dimensional liquid chromatography/LTQ-Orbitrap MS (offline 2D-LC/LTQ-Orbitrap MS) using collision-induced dissociation (CID) and high-energy C-trap dissociation (HCD); (ii) improved identification of the major aglycones by acid hydrolysis and LC-SPE-NMR; (iii) simplified spectral elucidation by high-resolution diagnostic product ions/neutral loss filtering; and (iv) more convincing structural identification by matching an in-house library. An offline 2D-LC system configuring an Acchrom XAmide column and a BEH Shield RP-18 UPLC ® column enabled much better separation of the easily co-eluting components. Combined use of CID and HCD could produce complementary fragmentation information. The intensity ratios of the aglycone ion species ([Y 0 -H] - /Y 0 - and [Y 0 -2H] - /Y 0 - ) in the HCD-MS 2 spectra were found diagnostic for discriminating the aglycone subtypes and characterizing the glycosylation patterns. Five aglycone structures (kaempferol, 6-hydroxykaempferol, 6-methoxykaempferol, carthamidin, and isocarthamidin) were identified based on the 1 H-NMR data recorded by LC-SPE-NMR. Of the 107 characterized flavonoids, 80 FOGs were first reported from C. tinctorius. Unknown aglycones, pentose, and novel acyl substituents were discovered. A new compound thereof was isolated and fully identified, which could partially validate the MS-oriented identification. This integral strategy can improve the potency, efficiency, and accuracy in the detection of new compounds from medicinal herbs and other natural sources. Copyright © 2017

  4. Emerging techniques for the discovery and validation of therapeutic targets for skeletal diseases.

    PubMed

    Cho, Christine H; Nuttall, Mark E

    2002-12-01

    Advances in genomics and proteomics have revolutionised the drug discovery process and target validation. Identification of novel therapeutic targets for chronic skeletal diseases is an extremely challenging process based on the difficulty of obtaining high-quality human diseased versus normal tissue samples. The quality of tissue and genomic information obtained from the sample is critical to identifying disease-related genes. Using a genomics-based approach, novel genes or genes with similar homology to existing genes can be identified from cDNA libraries generated from normal versus diseased tissue. High-quality cDNA libraries are prepared from uncontaminated homogeneous cell populations harvested from tissue sections of interest. Localised gene expression analysis and confirmation are obtained through in situ hybridisation or immunohistochemical studies. Cells overexpressing the recombinant protein are subsequently designed for primary cell-based high-throughput assays that are capable of screening large compound banks for potential hits. Afterwards, secondary functional assays are used to test promising compounds. The same overexpressing cells are used in the secondary assay to test protein activity and functionality as well as screen for small-molecule agonists or antagonists. Once a hit is generated, a structure-activity relationship of the compound is optimised for better oral bioavailability and pharmacokinetics allowing the compound to progress into development. Parallel efforts from proteomics, as well as genetics/transgenics, bioinformatics and combinatorial chemistry, and improvements in high-throughput automation technologies, allow the drug discovery process to meet the demands of the medicinal market. This review discusses and illustrates how different approaches are incorporated into the discovery and validation of novel targets and, consequently, the development of potentially therapeutic agents in the areas of osteoporosis and osteoarthritis

  5. Non-clinical studies required for new drug development - Part I: early in silico and in vitro studies, new target discovery and validation, proof of principles and robustness of animal studies.

    PubMed

    Andrade, E L; Bento, A F; Cavalli, J; Oliveira, S K; Freitas, C S; Marcon, R; Schwanke, R C; Siqueira, J M; Calixto, J B

    2016-10-24

    This review presents a historical overview of drug discovery and the non-clinical stages of the drug development process, from initial target identification and validation, through in silico assays and high throughput screening (HTS), identification of leader molecules and their optimization, the selection of a candidate substance for clinical development, and the use of animal models during the early studies of proof-of-concept (or principle). This report also discusses the relevance of validated and predictive animal models selection, as well as the correct use of animal tests concerning the experimental design, execution and interpretation, which affect the reproducibility, quality and reliability of non-clinical studies necessary to translate to and support clinical studies. Collectively, improving these aspects will certainly contribute to the robustness of both scientific publications and the translation of new substances to clinical development.

  6. Construction of a directed hammerhead ribozyme library: towards the identification of optimal target sites for antisense-mediated gene inhibition.

    PubMed Central

    Pierce, M L; Ruffner, D E

    1998-01-01

    Antisense-mediated gene inhibition uses short complementary DNA or RNA oligonucleotides to block expression of any mRNA of interest. A key parameter in the success or failure of an antisense therapy is the identification of a suitable target site on the chosen mRNA. Ultimately, the accessibility of the target to the antisense agent determines target suitability. Since accessibility is a function of many complex factors, it is currently beyond our ability to predict. Consequently, identification of the most effective target(s) requires examination of every site. Towards this goal, we describe a method to construct directed ribozyme libraries against any chosen mRNA. The library contains nearly equal amounts of ribozymes targeting every site on the chosen transcript and the library only contains ribozymes capable of binding to that transcript. Expression of the ribozyme library in cultured cells should allow identification of optimal target sites under natural conditions, subject to the complexities of a fully functional cell. Optimal target sites identified in this manner should be the most effective sites for therapeutic intervention. PMID:9801305

  7. RISC RNA sequencing for context-specific identification of in vivo microRNA targets.

    PubMed

    Matkovich, Scot J; Van Booven, Derek J; Eschenbacher, William H; Dorn, Gerald W

    2011-01-07

    MicroRNAs (miRs) are expanding our understanding of cardiac disease and have the potential to transform cardiovascular therapeutics. One miR can target hundreds of individual mRNAs, but existing methodologies are not sufficient to accurately and comprehensively identify these mRNA targets in vivo. To develop methods permitting identification of in vivo miR targets in an unbiased manner, using massively parallel sequencing of mouse cardiac transcriptomes in combination with sequencing of mRNA associated with mouse cardiac RNA-induced silencing complexes (RISCs). We optimized techniques for expression profiling small amounts of RNA without introducing amplification bias and applied this to anti-Argonaute 2 immunoprecipitated RISCs (RISC-Seq) from mouse hearts. By comparing RNA-sequencing results of cardiac RISC and transcriptome from the same individual hearts, we defined 1645 mRNAs consistently targeted to mouse cardiac RISCs. We used this approach in hearts overexpressing miRs from Myh6 promoter-driven precursors (programmed RISC-Seq) to identify 209 in vivo targets of miR-133a and 81 in vivo targets of miR-499. Consistent with the fact that miR-133a and miR-499 have widely differing "seed" sequences and belong to different miR families, only 6 targets were common to miR-133a- and miR-499-programmed hearts. RISC-sequencing is a highly sensitive method for general RISC profiling and individual miR target identification in biological context and is applicable to any tissue and any disease state.

  8. Approaches to Validate and Manipulate RNA Targets with Small Molecules in Cells.

    PubMed

    Childs-Disney, Jessica L; Disney, Matthew D

    2016-01-01

    RNA has become an increasingly important target for therapeutic interventions and for chemical probes that dissect and manipulate its cellular function. Emerging targets include human RNAs that have been shown to directly cause cancer, metabolic disorders, and genetic disease. In this review, we describe various routes to obtain bioactive compounds that target RNA, with a particular emphasis on the development of small molecules. We use these cases to describe approaches that are being developed for target validation, which include target-directed cleavage, classic pull-down experiments, and covalent cross-linking. Thus, tools are available to design small molecules to target RNA and to identify the cellular RNAs that are their targets.

  9. Validation of Laser-Induced Fluorescent Photogrammetric Targets on Membrane Structures

    NASA Technical Reports Server (NTRS)

    Jones, Thomas W.; Dorrington, Adrian A.; Shortis, Mark R.; Hendricks, Aron R.

    2004-01-01

    The need for static and dynamic characterization of a new generation of inflatable space structures requires the advancement of classical metrology techniques. A new photogrammetric-based method for non-contact ranging and surface profiling has been developed at NASA Langley Research Center (LaRC) to support modal analyses and structural validation of this class of space structures. This full field measurement method, known as Laser-Induced Fluorescence (LIF) photogrammetry, has previously yielded promising experimental results. However, data indicating the achievable measurement precision had not been published. This paper provides experimental results that indicate the LIF-photogrammetry measurement precision for three different target types used on a reflective membrane structure. The target types were: (1) non-contact targets generated using LIF, (2) surface attached retro-reflective targets, and (3) surface attached diffuse targets. Results from both static and dynamic investigations are included.

  10. Advances in the Study of Aptamer-Protein Target Identification Using the Chromatographic Approach.

    PubMed

    Drabik, Anna; Ner-Kluza, Joanna; Mielczarek, Przemyslaw; Civit, Laia; Mayer, Günter; Silberring, Jerzy

    2018-06-01

    Ever since the development of the process known as the systematic evolution of ligands by exponential enrichment (SELEX), aptamers have been widely used in a variety of studies, including the exploration of new diagnostic tools and the discovery of new treatment methods. Aptamers' ability to bind to proteins with high affinity and specificity, often compared to that of antibodies, enables the search for potential cancer biomarkers and helps us understand the mechanisms of carcinogenesis. The blind spot of those investigations is usually the difficulty in the selective extraction of targets attached to the aptamer. There are many studies describing the cell SELEX for the prime choice of aptamers toward living cancer cells or even whole tumors in the animal models. However, a dilemma arises when a large number of proteins are being identified as potential targets, which is often the case. In this article, we present a new analytical approach designed to selectively target proteins bound to aptamers. During studies, we have focused on the unambiguous identification of the molecular targets of aptamers characterized by high specificity to the prostate cancer cells. We have compared four assay approaches using electrophoretic and chromatographic methods for "fishing out" aptamer protein targets followed by mass spectrometry identification. We have established a new methodology, based on the fluorescent-tagged oligonucleotides commonly used for flow-cytometry experiments or as optic aptasensors, that allowed the detection of specific aptamer-protein interactions by mass spectrometry. The use of atto488-labeled aptamers for the tracking of the formation of specific aptamer-target complexes provides the possibility of studying putative protein counterparts without needing to apply enrichment techniques. Significantly, changes in the hydrophobic properties of atto488-labeled aptamer-protein complexes facilitate their separation by reverse-phase chromatography combined with

  11. miRTarBase update 2018: a resource for experimentally validated microRNA-target interactions.

    PubMed

    Chou, Chih-Hung; Shrestha, Sirjana; Yang, Chi-Dung; Chang, Nai-Wen; Lin, Yu-Ling; Liao, Kuang-Wen; Huang, Wei-Chi; Sun, Ting-Hsuan; Tu, Siang-Jyun; Lee, Wei-Hsiang; Chiew, Men-Yee; Tai, Chun-San; Wei, Ting-Yen; Tsai, Tzi-Ren; Huang, Hsin-Tzu; Wang, Chung-Yu; Wu, Hsin-Yi; Ho, Shu-Yi; Chen, Pin-Rong; Chuang, Cheng-Hsun; Hsieh, Pei-Jung; Wu, Yi-Shin; Chen, Wen-Liang; Li, Meng-Ju; Wu, Yu-Chun; Huang, Xin-Yi; Ng, Fung Ling; Buddhakosai, Waradee; Huang, Pei-Chun; Lan, Kuan-Chun; Huang, Chia-Yen; Weng, Shun-Long; Cheng, Yeong-Nan; Liang, Chao; Hsu, Wen-Lian; Huang, Hsien-Da

    2018-01-04

    MicroRNAs (miRNAs) are small non-coding RNAs of ∼ 22 nucleotides that are involved in negative regulation of mRNA at the post-transcriptional level. Previously, we developed miRTarBase which provides information about experimentally validated miRNA-target interactions (MTIs). Here, we describe an updated database containing 422 517 curated MTIs from 4076 miRNAs and 23 054 target genes collected from over 8500 articles. The number of MTIs curated by strong evidence has increased ∼1.4-fold since the last update in 2016. In this updated version, target sites validated by reporter assay that are available in the literature can be downloaded. The target site sequence can extract new features for analysis via a machine learning approach which can help to evaluate the performance of miRNA-target prediction tools. Furthermore, different ways of browsing enhance user browsing specific MTIs. With these improvements, miRTarBase serves as more comprehensively annotated, experimentally validated miRNA-target interactions databases in the field of miRNA related research. miRTarBase is available at http://miRTarBase.mbc.nctu.edu.tw/. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  12. Simultaneous analysis of coccidiostats and sulfonamides in non-target feed by HPLC-MS/MS and validation following the Commission Decision 2002/657/EC.

    PubMed

    Gavilán, Rosa Elvira; Nebot, Carolina; Patyra, Ewelina; Miranda, Jose Manuel; Franco, Carlos Manuel; Cepeda, Alberto

    2018-05-02

    Taking into consideration the maximum level for coccidiostats included in the European Regulation 574/2011 and the fact that the presence of residues of sulfonamides in non-target feed is forbidden, the aim of this article is to present an analytical method based on HPLC-MS/MS for the identification and quantification of sulfonamides and coccidiostats in non-target feeds. The method was validated following Decision 2002/657/EC and recovery, repeatability, and reproducibility were within the limits stablished in the Decision. For coccidiostats, the decision limit and detection capability were calculated for the different species taking into account the maximum level allowed in Regulation 574/2011. The applicability of the method was investigated in 50 feed samples collected from dairy farms, 50 obtained from feed mills, and 10 interlaboratory feed samples.

  13. Identification of Five Novel Variants in Chinese Oculocutaneous Albinism by Targeted Next-Generation Sequencing.

    PubMed

    Qiu, Biyuan; Ma, Tao; Peng, Chunyan; Zheng, Xiaoqin; Yang, Jiyun

    2018-04-01

    The diagnosis of oculocutaneous albinism (OCA) is established using clinical signs and symptoms. OCA is, however, a highly genetically heterogeneous disease with mutations identified in at least nineteen unique genes, many of which produce overlapping phenotypic traits. Thus, differentiating genetic OCA subtypes for diagnoses and genetic counseling is challenging, based on clinical presentation alone, and would benefit from a comprehensive molecular diagnostic. To develop and validate a more comprehensive, targeted, next-generation-sequencing-based diagnostic for the identification of OCA-causing variants. The genomic DNA samples from 28 OCA probands were analyzed by targeted next-generation sequencing (NGS), and the candidate variants were confirmed through Sanger sequencing. We observed mutations in the TYR, OCA2, and SLC45A2 genes in 25/28 (89%) patients with OCA. We identified 38 pathogenic variants among these three genes, including 5 novel variants: c.1970G>T (p.Gly657Val), c.1669A>C (p.Thr557Pro), c.2339-2A>C, and c.1349C>G (p.Thr450Arg) in OCA2; c.459_470delTTTTGCTGCCGA (p.Ala155_Phe158del) in SLC45A2. Our findings expand the mutational spectrum of OCA in the Chinese population, and the assay we developed should be broadly useful as a molecular diagnostic, and as an aid for genetic counseling for OCA patients.

  14. 3D high-content screening for the identification of compounds that target cells in dormant tumor spheroid regions.

    PubMed

    Wenzel, Carsten; Riefke, Björn; Gründemann, Stephan; Krebs, Alice; Christian, Sven; Prinz, Florian; Osterland, Marc; Golfier, Sven; Räse, Sebastian; Ansari, Nariman; Esner, Milan; Bickle, Marc; Pampaloni, Francesco; Mattheyer, Christian; Stelzer, Ernst H; Parczyk, Karsten; Prechtl, Stefan; Steigemann, Patrick

    2014-04-15

    Cancer cells in poorly vascularized tumor regions need to adapt to an unfavorable metabolic microenvironment. As distance from supplying blood vessels increases, oxygen and nutrient concentrations decrease and cancer cells react by stopping cell cycle progression and becoming dormant. As cytostatic drugs mainly target proliferating cells, cancer cell dormancy is considered as a major resistance mechanism to this class of anti-cancer drugs. Therefore, substances that target cancer cells in poorly vascularized tumor regions have the potential to enhance cytostatic-based chemotherapy of solid tumors. With three-dimensional growth conditions, multicellular tumor spheroids (MCTS) reproduce several parameters of the tumor microenvironment, including oxygen and nutrient gradients as well as the development of dormant tumor regions. We here report the setup of a 3D cell culture compatible high-content screening system and the identification of nine substances from two commercially available drug libraries that specifically target cells in inner MCTS core regions, while cells in outer MCTS regions or in 2D cell culture remain unaffected. We elucidated the mode of action of the identified compounds as inhibitors of the respiratory chain and show that induction of cell death in inner MCTS core regions critically depends on extracellular glucose concentrations. Finally, combinational treatment with cytostatics showed increased induction of cell death in MCTS. The data presented here shows for the first time a high-content based screening setup on 3D tumor spheroids for the identification of substances that specifically induce cell death in inner tumor spheroid core regions. This validates the approach to use 3D cell culture screening systems to identify substances that would not be detectable by 2D based screening in otherwise similar culture conditions. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  15. Biocomputational identification and validation of novel microRNAs predicted from bubaline whole genome shotgun sequences.

    PubMed

    Manku, H K; Dhanoa, J K; Kaur, S; Arora, J S; Mukhopadhyay, C S

    2017-10-01

    MicroRNAs (miRNAs) are small (19-25 base long), non-coding RNAs that regulate post-transcriptional gene expression by cleaving targeted mRNAs in several eukaryotes. The miRNAs play vital roles in multiple biological and metabolic processes, including developmental timing, signal transduction, cell maintenance and differentiation, diseases and cancers. Experimental identification of microRNAs is expensive and lab-intensive. Alternatively, computational approaches for predicting putative miRNAs from genomic or exomic sequences rely on features of miRNAs viz. secondary structures, sequence conservation, minimum free energy index (MFEI) etc. To date, not a single miRNA has been identified in bubaline (Bubalus bubalis), which is an economically important livestock. The present study aims at predicting the putative miRNAs of buffalo using comparative computational approach from buffalo whole genome shotgun sequencing data (INSDC: AWWX00000000.1). The sequences were blasted against the known mammalian miRNA. The obtained miRNAs were then passed through a series of filtration criteria to obtain the set of predicted (putative and novel) bubaline miRNA. Eight miRNAs were selected based on lowest E-value and validated by real time PCR (SYBR green chemistry) using RNU6 as endogenous control. The results from different trails of real time PCR shows that out of selected 8 miRNAs, only 2 (hsa-miR-1277-5p; bta-miR-2285b) are not expressed in bubaline PBMCs. The potential target genes based on their sequence complementarities were then predicted using miRanda. This work is the first report on prediction of bubaline miRNA from whole genome sequencing data followed by experimental validation. The finding could pave the way to future studies in economically important traits in buffalo. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. RISC RNA sequencing for context-specific identification of in vivo miR targets

    PubMed Central

    Matkovich, Scot J; Van Booven, Derek J; Eschenbacher, William H; Dorn, Gerald W

    2010-01-01

    Rationale MicroRNAs (miRs) are expanding our understanding of cardiac disease and have the potential to transform cardiovascular therapeutics. One miR can target hundreds of individual mRNAs, but existing methodologies are not sufficient to accurately and comprehensively identify these mRNA targets in vivo. Objective To develop methods permitting identification of in vivo miR targets in an unbiased manner, using massively parallel sequencing of mouse cardiac transcriptomes in combination with sequencing of mRNA associated with mouse cardiac RNA-induced silencing complexes (RISCs). Methods and Results We optimized techniques for expression profiling small amounts of RNA without introducing amplification bias, and applied this to anti-Argonaute 2 immunoprecipitated RISCs (RISC-Seq) from mouse hearts. By comparing RNA-sequencing results of cardiac RISC and transcriptome from the same individual hearts, we defined 1,645 mRNAs consistently targeted to mouse cardiac RISCs. We employed this approach in hearts overexpressing miRs from Myh6 promoter-driven precursors (programmed RISC-Seq) to identify 209 in vivo targets of miR-133a and 81 in vivo targets of miR-499. Consistent with the fact that miR-133a and miR-499 have widely differing ‘seed’ sequences and belong to different miR families, only 6 targets were common to miR-133a- and miR-499-programmed hearts. Conclusions RISC-sequencing is a highly sensitive method for general RISC profiling and individual miR target identification in biological context, and is applicable to any tissue and any disease state. Summary MicroRNAs (miRs) are key regulators of mRNA translation in health and disease. While bioinformatic predictions suggest that a single miR may target hundreds of mRNAs, the number of experimentally verified targets of miRs is low. To enable comprehensive, unbiased examination of miR targets, we have performed deep RNA sequencing of cardiac transcriptomes in parallel with cardiac RNA-induced silencing complex

  17. Combined effects of expectations and visual uncertainty upon detection and identification of a target in the fog.

    PubMed

    Quétard, Boris; Quinton, Jean-Charles; Colomb, Michèle; Pezzulo, Giovanni; Barca, Laura; Izaute, Marie; Appadoo, Owen Kevin; Mermillod, Martial

    2015-09-01

    Detecting a pedestrian while driving in the fog is one situation where the prior expectation about the target presence is integrated with the noisy visual input. We focus on how these sources of information influence the oculomotor behavior and are integrated within an underlying decision-making process. The participants had to judge whether high-/low-density fog scenes displayed on a computer screen contained a pedestrian or a deer by executing a mouse movement toward the response button (mouse-tracking). A variable road sign was added on the scene to manipulate expectations about target identity. We then analyzed the timing and amplitude of the deviation of mouse trajectories toward the incorrect response and, using an eye tracker, the detection time (before fixating the target) and the identification time (fixations on the target). Results revealed that expectation of the correct target results in earlier decisions with less deviation toward the alternative response, this effect being partially explained by the facilitation of target identification.

  18. Identification of lactoferricin B intracellular targets using an Escherichia coli proteome chip.

    PubMed

    Tu, Yu-Hsuan; Ho, Yu-Hsuan; Chuang, Ying-Chih; Chen, Po-Chung; Chen, Chien-Sheng

    2011-01-01

    Lactoferricin B (LfcinB) is a well-known antimicrobial peptide. Several studies have indicated that it can inhibit bacteria by affecting intracellular activities, but the intracellular targets of this antimicrobial peptide have not been identified. Therefore, we used E. coli proteome chips to identify the intracellular target proteins of LfcinB in a high-throughput manner. We probed LfcinB with E. coli proteome chips and further conducted normalization and Gene Ontology (GO) analyses. The results of the GO analyses showed that the identified proteins were associated with metabolic processes. Moreover, we validated the interactions between LfcinB and chip assay-identified proteins with fluorescence polarization (FP) assays. Sixteen proteins were identified, and an E. coli interaction database (EcID) analysis revealed that the majority of the proteins that interact with these 16 proteins affected the tricarboxylic acid (TCA) cycle. Knockout assays were conducted to further validate the FP assay results. These results showed that phosphoenolpyruvate carboxylase was a target of LfcinB, indicating that one of its mechanisms of action may be associated with pyruvate metabolism. Thus, we used pyruvate assays to conduct an in vivo validation of the relationship between LfcinB and pyruvate level in E. coli. These results showed that E. coli exposed to LfcinB had abnormal pyruvate amounts, indicating that LfcinB caused an accumulation of pyruvate. In conclusion, this study successfully revealed the intracellular targets of LfcinB using an E. coli proteome chip approach.

  19. Identification of Lactoferricin B Intracellular Targets Using an Escherichia coli Proteome Chip

    PubMed Central

    Chen, Po-Chung; Chen, Chien-Sheng

    2011-01-01

    Lactoferricin B (LfcinB) is a well-known antimicrobial peptide. Several studies have indicated that it can inhibit bacteria by affecting intracellular activities, but the intracellular targets of this antimicrobial peptide have not been identified. Therefore, we used E. coli proteome chips to identify the intracellular target proteins of LfcinB in a high-throughput manner. We probed LfcinB with E. coli proteome chips and further conducted normalization and Gene Ontology (GO) analyses. The results of the GO analyses showed that the identified proteins were associated with metabolic processes. Moreover, we validated the interactions between LfcinB and chip assay-identified proteins with fluorescence polarization (FP) assays. Sixteen proteins were identified, and an E. coli interaction database (EcID) analysis revealed that the majority of the proteins that interact with these 16 proteins affected the tricarboxylic acid (TCA) cycle. Knockout assays were conducted to further validate the FP assay results. These results showed that phosphoenolpyruvate carboxylase was a target of LfcinB, indicating that one of its mechanisms of action may be associated with pyruvate metabolism. Thus, we used pyruvate assays to conduct an in vivo validation of the relationship between LfcinB and pyruvate level in E. coli. These results showed that E. coli exposed to LfcinB had abnormal pyruvate amounts, indicating that LfcinB caused an accumulation of pyruvate. In conclusion, this study successfully revealed the intracellular targets of LfcinB using an E. coli proteome chip approach. PMID:22164243

  20. Approaches to the identification of new nonhormonal targets for male contraception.

    PubMed

    Kopf, Gregory S

    2008-10-01

    Fertility control is a global health issue with major personal and societal impact. Although, currently, there are several different options for contraception, the technologies behind these are antiquated, and the options for male-based contraception (i.e., withdrawal, condom and vasectomy) are inadequate. The genomic, proteomic and bioinformatic revolutions have provided new tools and new targets for contraceptive development, and the results of such approaches have identified gene products that play critical roles in male reproduction, thus expanding the array of potential targets for novel and innovative male-based contraceptives. This article will review the types of targets being considered in the development of nonhormonal male contraceptives and the technologies used to identify and validate these targets.

  1. PharmMapper server: a web server for potential drug target identification using pharmacophore mapping approach

    PubMed Central

    Liu, Xiaofeng; Ouyang, Sisheng; Yu, Biao; Liu, Yabo; Huang, Kai; Gong, Jiayu; Zheng, Siyuan; Li, Zhihua; Li, Honglin; Jiang, Hualiang

    2010-01-01

    In silico drug target identification, which includes many distinct algorithms for finding disease genes and proteins, is the first step in the drug discovery pipeline. When the 3D structures of the targets are available, the problem of target identification is usually converted to finding the best interaction mode between the potential target candidates and small molecule probes. Pharmacophore, which is the spatial arrangement of features essential for a molecule to interact with a specific target receptor, is an alternative method for achieving this goal apart from molecular docking method. PharmMapper server is a freely accessed web server designed to identify potential target candidates for the given small molecules (drugs, natural products or other newly discovered compounds with unidentified binding targets) using pharmacophore mapping approach. PharmMapper hosts a large, in-house repertoire of pharmacophore database (namely PharmTargetDB) annotated from all the targets information in TargetBank, BindingDB, DrugBank and potential drug target database, including over 7000 receptor-based pharmacophore models (covering over 1500 drug targets information). PharmMapper automatically finds the best mapping poses of the query molecule against all the pharmacophore models in PharmTargetDB and lists the top N best-fitted hits with appropriate target annotations, as well as respective molecule’s aligned poses are presented. Benefited from the highly efficient and robust triangle hashing mapping method, PharmMapper bears high throughput ability and only costs 1 h averagely to screen the whole PharmTargetDB. The protocol was successful in finding the proper targets among the top 300 pharmacophore candidates in the retrospective benchmarking test of tamoxifen. PharmMapper is available at http://59.78.96.61/pharmmapper. PMID:20430828

  2. Benchmark radar targets for the validation of computational electromagnetics programs

    NASA Technical Reports Server (NTRS)

    Woo, Alex C.; Wang, Helen T. G.; Schuh, Michael J.; Sanders, Michael L.

    1993-01-01

    Results are presented of a set of computational electromagnetics validation measurements referring to three-dimensional perfectly conducting smooth targets, performed for the Electromagnetic Code Consortium. Plots are presented for both the low- and high-frequency measurements of the NASA almond, an ogive, a double ogive, a cone-sphere, and a cone-sphere with a gap.

  3. Identification of STAT1 and STAT3 Specific Inhibitors Using Comparative Virtual Screening and Docking Validation

    PubMed Central

    Szelag, Malgorzata; Czerwoniec, Anna; Wesoly, Joanna; Bluyssen, Hans A. R.

    2015-01-01

    Signal transducers and activators of transcription (STATs) facilitate action of cytokines, growth factors and pathogens. STAT activation is mediated by a highly conserved SH2 domain, which interacts with phosphotyrosine motifs for specific STAT-receptor contacts and STAT dimerization. The active dimers induce gene transcription in the nucleus by binding to a specific DNA-response element in the promoter of target genes. Abnormal activation of STAT signaling pathways is implicated in many human diseases, like cancer, inflammation and auto-immunity. Searches for STAT-targeting compounds, exploring the phosphotyrosine (pTyr)-SH2 interaction site, yielded many small molecules for STAT3 but sparsely for other STATs. However, many of these inhibitors seem not STAT3-specific, thereby questioning the present modeling and selection strategies of SH2 domain-based STAT inhibitors. We generated new 3D structure models for all human (h)STATs and developed a comparative in silico docking strategy to obtain further insight into STAT-SH2 cross-binding specificity of a selection of previously identified STAT3 inhibitors. Indeed, by primarily targeting the highly conserved pTyr-SH2 binding pocket the majority of these compounds exhibited similar binding affinity and tendency scores for all STATs. By comparative screening of a natural product library we provided initial proof for the possibility to identify STAT1 as well as STAT3-specific inhibitors, introducing the ‘STAT-comparative binding affinity value’ and ‘ligand binding pose variation’ as selection criteria. In silico screening of a multi-million clean leads (CL) compound library for binding of all STATs, likewise identified potential specific inhibitors for STAT1 and STAT3 after docking validation. Based on comparative virtual screening and docking validation, we developed a novel STAT inhibitor screening tool that allows identification of specific STAT1 and STAT3 inhibitory compounds. This could increase our

  4. Signal-Noise Identification of Magnetotelluric Signals Using Fractal-Entropy and Clustering Algorithm for Targeted De-Noising

    NASA Astrophysics Data System (ADS)

    Li, Jin; Zhang, Xian; Gong, Jinzhe; Tang, Jingtian; Ren, Zhengyong; Li, Guang; Deng, Yanli; Cai, Jin

    A new technique is proposed for signal-noise identification and targeted de-noising of Magnetotelluric (MT) signals. This method is based on fractal-entropy and clustering algorithm, which automatically identifies signal sections corrupted by common interference (square, triangle and pulse waves), enabling targeted de-noising and preventing the loss of useful information in filtering. To implement the technique, four characteristic parameters — fractal box dimension (FBD), higuchi fractal dimension (HFD), fuzzy entropy (FuEn) and approximate entropy (ApEn) — are extracted from MT time-series. The fuzzy c-means (FCM) clustering technique is used to analyze the characteristic parameters and automatically distinguish signals with strong interference from the rest. The wavelet threshold (WT) de-noising method is used only to suppress the identified strong interference in selected signal sections. The technique is validated through signal samples with known interference, before being applied to a set of field measured MT/Audio Magnetotelluric (AMT) data. Compared with the conventional de-noising strategy that blindly applies the filter to the overall dataset, the proposed method can automatically identify and purposefully suppress the intermittent interference in the MT/AMT signal. The resulted apparent resistivity-phase curve is more continuous and smooth, and the slow-change trend in the low-frequency range is more precisely reserved. Moreover, the characteristic of the target-filtered MT/AMT signal is close to the essential characteristic of the natural field, and the result more accurately reflects the inherent electrical structure information of the measured site.

  5. Modeling and validation of photometric characteristics of space targets oriented to space-based observation.

    PubMed

    Wang, Hongyuan; Zhang, Wei; Dong, Aotuo

    2012-11-10

    A modeling and validation method of photometric characteristics of the space target was presented in order to track and identify different satellites effectively. The background radiation characteristics models of the target were built based on blackbody radiation theory. The geometry characteristics of the target were illustrated by the surface equations based on its body coordinate system. The material characteristics of the target surface were described by a bidirectional reflectance distribution function model, which considers the character of surface Gauss statistics and microscale self-shadow and is obtained by measurement and modeling in advance. The contributing surfaces of the target to observation system were determined by coordinate transformation according to the relative position of the space-based target, the background radiation sources, and the observation platform. Then a mathematical model on photometric characteristics of the space target was built by summing reflection components of all the surfaces. Photometric characteristics simulation of the space-based target was achieved according to its given geometrical dimensions, physical parameters, and orbital parameters. Experimental validation was made based on the scale model of the satellite. The calculated results fit well with the measured results, which indicates the modeling method of photometric characteristics of the space target is correct.

  6. Identification of the Transcriptional Targets of FOXP2, a Gene Linked to Speech and Language, in Developing Human Brain

    PubMed Central

    Spiteri, Elizabeth ; Konopka, Genevieve ; Coppola, Giovanni ; Bomar, Jamee ; Oldham, Michael ; Ou, Jing ; Vernes, Sonja C. ; Fisher, Simon E. ; Ren, Bing ; Geschwind, Daniel H. 

    2007-01-01

    Mutations in FOXP2, a member of the forkhead family of transcription factor genes, are the only known cause of developmental speech and language disorders in humans. To date, there are no known targets of human FOXP2 in the nervous system. The identification of FOXP2 targets in the developing human brain, therefore, provides a unique tool with which to explore the development of human language and speech. Here, we define FOXP2 targets in human basal ganglia (BG) and inferior frontal cortex (IFC) by use of chromatin immunoprecipitation followed by microarray analysis (ChIP-chip) and validate the functional regulation of targets in vitro. ChIP-chip identified 285 FOXP2 targets in fetal human brain; statistically significant overlap of targets in BG and IFC indicates a core set of 34 transcriptional targets of FOXP2. We identified targets specific to IFC or BG that were not observed in lung, suggesting important regional and tissue differences in FOXP2 activity. Many target genes are known to play critical roles in specific aspects of central nervous system patterning or development, such as neurite outgrowth, as well as plasticity. Subsets of the FOXP2 transcriptional targets are either under positive selection in humans or differentially expressed between human and chimpanzee brain. This is the first ChIP-chip study to use human brain tissue, making the FOXP2-target genes identified in these studies important to understanding the pathways regulating speech and language in the developing human brain. These data provide the first insight into the functional network of genes directly regulated by FOXP2 in human brain and by evolutionary comparisons, highlighting genes likely to be involved in the development of human higher-order cognitive processes. PMID:17999357

  7. TARGET's role in knowledge acquisition, engineering, validation, and documentation

    NASA Technical Reports Server (NTRS)

    Levi, Keith R.

    1994-01-01

    We investigate the use of the TARGET task analysis tool for use in the development of rule-based expert systems. We found TARGET to be very helpful in the knowledge acquisition process. It enabled us to perform knowledge acquisition with one knowledge engineer rather than two. In addition, it improved communication between the domain expert and knowledge engineer. We also found it to be useful for both the rule development and refinement phases of the knowledge engineering process. Using the network in these phases required us to develop guidelines that enabled us to easily translate the network into production rules. A significant requirement for TARGET remaining useful throughout the knowledge engineering process was the need to carefully maintain consistency between the network and the rule representations. Maintaining consistency not only benefited the knowledge engineering process, but also has significant payoffs in the areas of validation of the expert system and documentation of the knowledge in the system.

  8. Target identification, lead optimization and antitumor evaluation of some new 1,2,4-triazines as c-Met kinase inhibitors.

    PubMed

    El-Wakil, Marwa H; Ashour, Hayam M; Saudi, Manal N; Hassan, Ahmed M; Labouta, Ibrahim M

    2017-08-01

    In silico target fishing approach using PharmMapper server identified c-Met kinase as the selective target for our previously synthesized compound NCI 748494/1. This approach was validated by in vitro kinase assay which showed that NCI 748494/1 possessed promising inhibitory activity against c-Met kinase (IC 50 =31.70μM). Assessment of ADMET profiling, drug-likeness, drug score as well as docking simulation for the binding pose of that compound in the active site of c-Met kinase domain revealed that NCI 748494/1 could be considered as a promising drug lead. Based on target identification and validation, it was observed that there is structure similarity between NCI 748494/1 and the reported type II c-Met kinase inhibitor BMS-777607. Optimization of our lead NCI 748494/1 furnished newly synthesized 1,2,4-triazine derivatives based on well-established structure-activity relationships, whereas three compounds namely; 4d, 7a and 8c displayed excellent in vitro cytotoxicity against three c-Met addicted cancer cell lines; A549 (lung adenocarcinoma), HT-29 (colon cancer) and MKN-45 (gastric carcinoma); with IC 50 values in the range 0.01-1.86µM. In vitro c-Met kinase assay showed 8c to possess the highest c-Met kinase inhibition profile (IC 50 =4.31µM). Docking of the active compounds in c-Met kinase active site revealed strong binding interactions comparable to the lead NCI 748494/1 and BMS-777607, suggesting that c-Met inhibition is very likely to be the mechanism of the antitumor effect of these derivatives. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Systems biology-embedded target validation: improving efficacy in drug discovery.

    PubMed

    Vandamme, Drieke; Minke, Benedikt A; Fitzmaurice, William; Kholodenko, Boris N; Kolch, Walter

    2014-01-01

    The pharmaceutical industry is faced with a range of challenges with the ever-escalating costs of drug development and a drying out of drug pipelines. By harnessing advances in -omics technologies and moving away from the standard, reductionist model of drug discovery, there is significant potential to reduce costs and improve efficacy. Embedding systems biology approaches in drug discovery, which seek to investigate underlying molecular mechanisms of potential drug targets in a network context, will reduce attrition rates by earlier target validation and the introduction of novel targets into the currently stagnant market. Systems biology approaches also have the potential to assist in the design of multidrug treatments and repositioning of existing drugs, while stratifying patients to give a greater personalization of medical treatment. © 2013 Wiley Periodicals, Inc.

  10. In silico re-identification of properties of drug target proteins.

    PubMed

    Kim, Baeksoo; Jo, Jihoon; Han, Jonghyun; Park, Chungoo; Lee, Hyunju

    2017-05-31

    Computational approaches in the identification of drug targets are expected to reduce time and effort in drug development. Advances in genomics and proteomics provide the opportunity to uncover properties of druggable genomes. Although several studies have been conducted for distinguishing drug targets from non-drug targets, they mainly focus on the sequences and functional roles of proteins. Many other properties of proteins have not been fully investigated. Using the DrugBank (version 3.0) database containing nearly 6,816 drug entries including 760 FDA-approved drugs and 1822 of their targets and human UniProt/Swiss-Prot databases, we defined 1578 non-redundant drug target and 17,575 non-drug target proteins. To select these non-redundant protein datasets, we built four datasets (A, B, C, and D) by considering clustering of paralogous proteins. We first reassessed the widely used properties of drug target proteins. We confirmed and extended that drug target proteins (1) are likely to have more hydrophobic, less polar, less PEST sequences, and more signal peptide sequences higher and (2) are more involved in enzyme catalysis, oxidation and reduction in cellular respiration, and operational genes. In this study, we proposed new properties (essentiality, expression pattern, PTMs, and solvent accessibility) for effectively identifying drug target proteins. We found that (1) drug targetability and protein essentiality are decoupled, (2) druggability of proteins has high expression level and tissue specificity, and (3) functional post-translational modification residues are enriched in drug target proteins. In addition, to predict the drug targetability of proteins, we exploited two machine learning methods (Support Vector Machine and Random Forest). When we predicted drug targets by combining previously known protein properties and proposed new properties, an F-score of 0.8307 was obtained. When the newly proposed properties are integrated, the prediction performance

  11. Identification of distant drug off-targets by direct superposition of binding pocket surfaces.

    PubMed

    Schumann, Marcel; Armen, Roger S

    2013-01-01

    Correctly predicting off-targets for a given molecular structure, which would have the ability to bind a large range of ligands, is both particularly difficult and important if they share no significant sequence or fold similarity with the respective molecular target ("distant off-targets"). A novel approach for identification of off-targets by direct superposition of protein binding pocket surfaces is presented and applied to a set of well-studied and highly relevant drug targets, including representative kinases and nuclear hormone receptors. The entire Protein Data Bank is searched for similar binding pockets and convincing distant off-target candidates were identified that share no significant sequence or fold similarity with the respective target structure. These putative target off-target pairs are further supported by the existence of compounds that bind strongly to both with high topological similarity, and in some cases, literature examples of individual compounds that bind to both. Also, our results clearly show that it is possible for binding pockets to exhibit a striking surface similarity, while the respective off-target shares neither significant sequence nor significant fold similarity with the respective molecular target ("distant off-target").

  12. A Study of the Validity of Language Usage as an Indicator of Ethnic Identification.

    ERIC Educational Resources Information Center

    Patella, Victoria Morrow

    In determining whether Spanish usage is a valid indicator of ethnic identification, 669 Texas high school sophomores (3 Negroes, 70 Anglos, and 596 Mexican Americans) were interviewed. Mexican American respondents were then isolated on the basis of response to 4 questions, and their responses regarding family were grouped under 2 headings: (1)…

  13. Application of chemical biology in target identification and drug discovery.

    PubMed

    Zhu, Yue; Xiao, Ting; Lei, Saifei; Zhou, Fulai; Wang, Ming-Wei

    2015-09-01

    Drug discovery and development is vital to the well-being of mankind and sustainability of the pharmaceutical industry. Using chemical biology approaches to discover drug leads has become a widely accepted path partially because of the completion of the Human Genome Project. Chemical biology mainly solves biological problems through searching previously unknown targets for pharmacologically active small molecules or finding ligands for well-defined drug targets. It is a powerful tool to study how these small molecules interact with their respective targets, as well as their roles in signal transduction, molecular recognition and cell functions. There have been an increasing number of new therapeutic targets being identified and subsequently validated as a result of advances in functional genomics, which in turn led to the discovery of numerous active small molecules via a variety of high-throughput screening initiatives. In this review, we highlight some applications of chemical biology in the context of drug discovery.

  14. Identification of chemogenomic features from drug–target interaction networks using interpretable classifiers

    PubMed Central

    Tabei, Yasuo; Pauwels, Edouard; Stoven, Véronique; Takemoto, Kazuhiro; Yamanishi, Yoshihiro

    2012-01-01

    Motivation: Drug effects are mainly caused by the interactions between drug molecules and their target proteins including primary targets and off-targets. Identification of the molecular mechanisms behind overall drug–target interactions is crucial in the drug design process. Results: We develop a classifier-based approach to identify chemogenomic features (the underlying associations between drug chemical substructures and protein domains) that are involved in drug–target interaction networks. We propose a novel algorithm for extracting informative chemogenomic features by using L1 regularized classifiers over the tensor product space of possible drug–target pairs. It is shown that the proposed method can extract a very limited number of chemogenomic features without loosing the performance of predicting drug–target interactions and the extracted features are biologically meaningful. The extracted substructure–domain association network enables us to suggest ligand chemical fragments specific for each protein domain and ligand core substructures important for a wide range of protein families. Availability: Softwares are available at the supplemental website. Contact: yamanishi@bioreg.kyushu-u.ac.jp Supplementary Information: Datasets and all results are available at http://cbio.ensmp.fr/~yyamanishi/l1binary/ . PMID:22962471

  15. Computational Identification of MicroRNAs and Their Targets from Finger Millet (Eleusine coracana).

    PubMed

    Usha, S; Jyothi, M N; Suchithra, B; Dixit, Rekha; Rai, D V; Nagesh Babu, R

    2017-03-01

    MicroRNAs are endogenous small RNAs regulating intrinsic normal growth and development of plant. Discovering miRNAs, their targets and further inferring their functions had become routine process to comprehend the normal biological processes of miRNAs and their roles in plant development. In this study, we used homology-based analysis with available expressed sequence tag of finger millet (Eleusine coracana) to predict conserved miRNAs. Three potent miRNAs targeting 88 genes were identified. The newly identified miRNAs were found to be homologous with miR166 and miR1310. The targets recognized were transcription factors and enzymes, and GO analysis showed these miRNAs played varied roles in gene regulation. The identification of miRNAs and their targets is anticipated to hasten the pace of key epigenetic regulators in plant development.

  16. Identification of Distant Drug Off-Targets by Direct Superposition of Binding Pocket Surfaces

    PubMed Central

    Schumann, Marcel; Armen, Roger S.

    2013-01-01

    Correctly predicting off-targets for a given molecular structure, which would have the ability to bind a large range of ligands, is both particularly difficult and important if they share no significant sequence or fold similarity with the respective molecular target (“distant off-targets”). A novel approach for identification of off-targets by direct superposition of protein binding pocket surfaces is presented and applied to a set of well-studied and highly relevant drug targets, including representative kinases and nuclear hormone receptors. The entire Protein Data Bank is searched for similar binding pockets and convincing distant off-target candidates were identified that share no significant sequence or fold similarity with the respective target structure. These putative target off-target pairs are further supported by the existence of compounds that bind strongly to both with high topological similarity, and in some cases, literature examples of individual compounds that bind to both. Also, our results clearly show that it is possible for binding pockets to exhibit a striking surface similarity, while the respective off-target shares neither significant sequence nor significant fold similarity with the respective molecular target (“distant off-target”). PMID:24391782

  17. Drug Discovery for Neglected Diseases: Molecular Target-Based and Phenotypic Approaches

    PubMed Central

    2013-01-01

    Drug discovery for neglected tropical diseases is carried out using both target-based and phenotypic approaches. In this paper, target-based approaches are discussed, with a particular focus on human African trypanosomiasis. Target-based drug discovery can be successful, but careful selection of targets is required. There are still very few fully validated drug targets in neglected diseases, and there is a high attrition rate in target-based drug discovery for these diseases. Phenotypic screening is a powerful method in both neglected and non-neglected diseases and has been very successfully used. Identification of molecular targets from phenotypic approaches can be a way to identify potential new drug targets. PMID:24015767

  18. Development of a Photo-Cross-Linkable Diaminoquinazoline Inhibitor for Target Identification in Plasmodium falciparum.

    PubMed

    Lubin, Alexandra S; Rueda-Zubiaurre, Ainoa; Matthews, Holly; Baumann, Hella; Fisher, Fabio R; Morales-Sanfrutos, Julia; Hadavizadeh, Kate S; Nardella, Flore; Tate, Edward W; Baum, Jake; Scherf, Artur; Fuchter, Matthew J

    2018-04-13

    Diaminoquinazolines represent a privileged scaffold for antimalarial discovery, including use as putative Plasmodium histone lysine methyltransferase inhibitors. Despite this, robust evidence for their molecular targets is lacking. Here we report the design and development of a small-molecule photo-cross-linkable probe to investigate the targets of our diaminoquinazoline series. We demonstrate the effectiveness of our designed probe for photoaffinity labeling of Plasmodium lysates and identify similarities between the target profiles of the probe and the representative diaminoquinazoline BIX-01294. Initial pull-down proteomics experiments identified 104 proteins from different classes, many of which are essential, highlighting the suitability of the developed probe as a valuable tool for target identification in Plasmodium falciparum.

  19. PharmMapper 2017 update: a web server for potential drug target identification with a comprehensive target pharmacophore database.

    PubMed

    Wang, Xia; Shen, Yihang; Wang, Shiwei; Li, Shiliang; Zhang, Weilin; Liu, Xiaofeng; Lai, Luhua; Pei, Jianfeng; Li, Honglin

    2017-07-03

    The PharmMapper online tool is a web server for potential drug target identification by reversed pharmacophore matching the query compound against an in-house pharmacophore model database. The original version of PharmMapper includes more than 7000 target pharmacophores derived from complex crystal structures with corresponding protein target annotations. In this article, we present a new version of the PharmMapper web server, of which the backend pharmacophore database is six times larger than the earlier one, with a total of 23 236 proteins covering 16 159 druggable pharmacophore models and 51 431 ligandable pharmacophore models. The expanded target data cover 450 indications and 4800 molecular functions compared to 110 indications and 349 molecular functions in our last update. In addition, the new web server is united with the statistically meaningful ranking of the identified drug targets, which is achieved through the use of standard scores. It also features an improved user interface. The proposed web server is freely available at http://lilab.ecust.edu.cn/pharmmapper/. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  20. Rapid identification and validation of novel targeted approaches for Glioblastoma: A combined ex vivo-in vivo pharmaco-omic model.

    PubMed

    Daher, Ahmad; de Groot, John

    2018-01-01

    Tumor heterogeneity is a major factor in glioblastoma's poor response to therapy and seemingly inevitable recurrence. Only two glioblastoma drugs have received Food and Drug Administration approval since 1998, highlighting the urgent need for new therapies. Profiling "omics" analyses have helped characterize glioblastoma molecularly and have thus identified multiple molecular targets for precision medicine. These molecular targets have influenced clinical trial design; many "actionable" mutation-focused trials are underway, but because they have not yet led to therapeutic breakthroughs, new strategies for treating glioblastoma, especially those with a pharmacological functional component, remain in high demand. In that regard, high-throughput screening that allows for expedited preclinical drug testing and the use of GBM models that represent tumor heterogeneity more accurately than traditional cancer cell lines is necessary to maximize the successful translation of agents into the clinic. High-throughput screening has been successfully used in the testing, discovery, and validation of potential therapeutics in various cancer models, but it has not been extensively utilized in glioblastoma models. In this report, we describe the basic aspects of high-throughput screening and propose a modified high-throughput screening model in which ex vivo and in vivo drug testing is complemented by post-screening pharmacological, pan-omic analysis to expedite anti-glioma drugs' preclinical testing and develop predictive biomarker datasets that can aid in personalizing glioblastoma therapy and inform clinical trial design. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. A new efficient method of generating photoaffinity beads for drug target identification.

    PubMed

    Nishiya, Yoichi; Hamada, Tomoko; Abe, Masayuki; Takashima, Michio; Tsutsumi, Kyoko; Okawa, Katsuya

    2017-02-15

    Affinity purification is one of the most prevalent methods for the target identification of small molecules. Preparation of an appropriate chemical for immobilization, however, is a tedious and time-consuming process. A decade ago, a photoreaction method for generating affinity beads was reported, where compounds are mixed with agarose beads carrying a photoreactive group (aryldiazirine) and then irradiated with ultraviolet light under dry conditions to form covalent attachment. Although the method has proven useful for identifying drug targets, the beads suffer from inefficient ligand incorporation and tend to shrink and aggregate, which can cause nonspecific binding and low reproducibility. We therefore decided to craft affinity beads free from these shortcomings without compromising the ease of preparation. We herein report a modified method; first, a compound of interest is mixed with a crosslinker having an activated ester and a photoreactive moiety on each end. This mixture is then dried in a glass tube and irradiated with ultraviolet light. Finally, the conjugates are dissolved and reacted with agarose beads with a primary amine. This protocol enabled us to immobilize compounds more efficiently (approximately 500-fold per bead compared to the original method) and generated beads without physical deterioration. We herein demonstrated that the new FK506-immobilized beads specifically isolated more FKBP12 than the original beads, thereby proving our method to be applicable to target identification experiments. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Optimization and in Vivo Validation of Peptide Vectors Targeting the LDL Receptor.

    PubMed

    Jacquot, Guillaume; Lécorché, Pascaline; Malcor, Jean-Daniel; Laurencin, Mathieu; Smirnova, Maria; Varini, Karine; Malicet, Cédric; Gassiot, Fanny; Abouzid, Karima; Faucon, Aude; David, Marion; Gaudin, Nicolas; Masse, Maxime; Ferracci, Géraldine; Dive, Vincent; Cisternino, Salvatore; Khrestchatisky, Michel

    2016-12-05

    Active targeting and delivery to pathophysiological organs of interest is of paramount importance to increase specific accumulation of therapeutic drugs or imaging agents while avoiding systemic side effects. We recently developed a family of new peptide ligands of the human and rodent LDL receptor (LDLR), an attractive cell-surface receptor with high uptake activity and local enrichment in several normal or pathological tissues (Malcor et al., J. Med. Chem. 2012, 55 (5), 2227). Initial chemical optimization of the 15-mer, all natural amino acid compound 1/VH411 (DSGL[CMPRLRGC] c DPR) and structure-activity relationship (SAR) investigation led to the cyclic 8 amino acid analogue compound 22/VH445 ([cMPRLRGC] c ) which specifically binds hLDLR with a K D of 76 nM and has an in vitro blood half-life of ∼3 h. Further introduction of non-natural amino acids led to the identification of compound 60/VH4106 ([(d)-"Pen"M"Thz"RLRGC] c ), which showed the highest K D value of 9 nM. However, this latter analogue displayed the lowest in vitro blood half-life (∼1.9 h). In the present study, we designed a new set of peptide analogues, namely, VH4127 to VH4131, with further improved biological properties. Detailed analysis of the hLDLR-binding kinetics of previous and new analogues showed that the latter all displayed very high on-rates, in the 10 6 s -1. M -1 range, and off-rates varying from the low 10 -2 s -1 to the 10 -1 s -1 range. Furthermore, all these new analogues showed increased blood half-lives in vitro, reaching ∼7 and 10 h for VH4129 and VH4131, respectively. Interestingly, we demonstrate in cell-based assays using both VH445 and the most balanced optimized analogue VH4127 ([cM"Thz"RLRG"Pen"] c ), showing a K D of 18 nM and a blood half-life of ∼4.3 h, that its higher on-rate correlated with a significant increase in both the extent of cell-surface binding to hLDLR and the endocytosis potential. Finally, intravenous injection of tritium-radiolabeled 3 H

  3. CRISPR-Cas9-based target validation for p53-reactivating model compounds

    PubMed Central

    Wanzel, Michael; Vischedyk, Jonas B; Gittler, Miriam P; Gremke, Niklas; Seiz, Julia R; Hefter, Mirjam; Noack, Magdalena; Savai, Rajkumar; Mernberger, Marco; Charles, Joël P; Schneikert, Jean; Bretz, Anne Catherine; Nist, Andrea; Stiewe, Thorsten

    2015-01-01

    Inactivation of the p53 tumor suppressor by Mdm2 is one of the most frequent events in cancer, so compounds targeting the p53-Mdm2 interaction are promising for cancer therapy. Mechanisms conferring resistance to p53-reactivating compounds are largely unknown. Here we show using CRISPR-Cas9–based target validation in lung and colorectal cancer that the activity of nutlin, which blocks the p53-binding pocket of Mdm2, strictly depends on functional p53. In contrast, sensitivity to the drug RITA, which binds the Mdm2-interacting N terminus of p53, correlates with induction of DNA damage. Cells with primary or acquired RITA resistance display cross-resistance to DNA crosslinking compounds such as cisplatin and show increased DNA cross-link repair. Inhibition of FancD2 by RNA interference or pharmacological mTOR inhibitors restores RITA sensitivity. The therapeutic response to p53-reactivating compounds is therefore limited by compound-specific resistance mechanisms that can be resolved by CRISPR-Cas9-based target validation and should be considered when allocating patients to p53-reactivating treatments. PMID:26595461

  4. Neuronal Target Identification Requires AHA-1-Mediated Fine-Tuning of Wnt Signaling in C. elegans

    PubMed Central

    Zhang, Jingyan; Li, Xia; Jevince, Angela R.; Guan, Liying; Wang, Jiaming; Hall, David H.; Huang, Xun; Ding, Mei

    2013-01-01

    Electrical synaptic transmission through gap junctions is a vital mode of intercellular communication in the nervous system. The mechanism by which reciprocal target cells find each other during the formation of gap junctions, however, is poorly understood. Here we show that gap junctions are formed between BDU interneurons and PLM mechanoreceptors in C. elegans and the connectivity of BDU with PLM is influenced by Wnt signaling. We further identified two PAS-bHLH family transcription factors, AHA-1 and AHR-1, which function cell-autonomously within BDU and PLM to facilitate the target identification process. aha-1 and ahr-1 act genetically upstream of cam-1. CAM-1, a membrane-bound receptor tyrosine kinase, is present on both BDU and PLM cells and likely serves as a Wnt antagonist. By binding to a cis-regulatory element in the cam-1 promoter, AHA-1 enhances cam-1 transcription. Our study reveals a Wnt-dependent fine-tuning mechanism that is crucial for mutual target cell identification during the formation of gap junction connections. PMID:23825972

  5. A validated methodology for genetic identification of tuna species (genus Thunnus).

    PubMed

    Viñas, Jordi; Tudela, Sergi

    2009-10-27

    Tuna species of the genus Thunnus, such as the bluefin tunas, are some of the most important and yet most endangered trade fish in the world. Identification of these species in traded forms, however, may be difficult depending on the presentation of the products, which may hamper conservation efforts on trade control. In this paper, we validated a genetic methodology that can fully distinguish between the eight Thunnus species from any kind of processed tissue. After testing several genetic markers, a complete discrimination of the eight tuna species was achieved using Forensically Informative Nucleotide Sequencing based primarily on the sequence variability of the hypervariable genetic marker mitochondrial DNA control region (mtDNA CR), followed, in some specific cases, by a second validation by a nuclear marker rDNA first internal transcribed spacer (ITS1). This methodology was able to distinguish all tuna species, including those belonging to the subgenus Neothunnus that are very closely related, and in consequence can not be differentiated with other genetic markers of lower variability. This methodology also took into consideration the presence of introgression that has been reported in past studies between T. thynnus, T. orientalis and T. alalunga. Finally, we applied the methodology to cross-check the species identity of 26 processed tuna samples. Using the combination of two genetic markers, one mitochondrial and another nuclear, allows a full discrimination between all eight tuna species. Unexpectedly, the genetic marker traditionally used for DNA barcoding, cytochrome oxidase 1, could not differentiate all species, thus its use as a genetic marker for tuna species identification is questioned.

  6. Pharmacogenetics and target identification in diabetes.

    PubMed

    Pearson, Ewan R

    2018-02-24

    In diabetes, pharmacogenetics can be used both to identify patient subgroups who will have most benefit and/or least harm from a particularly treatment, and to gain insights into the molecular mechanisms of drug action and disease aetiology. There is increasing evidence that genetic variation alters response to diabetes treatments-both in terms of glycaemic response and side effects. This can be seen with dramatic impact on clinical care, in patients with genetic forms of diabetes such as Maturity Onset Diabetes of the Young caused by HNF1A mutations, and Neonatal diabetes due to activating mutations in ABCC8 or KCNJ11. Beyond monogenic diabetes, pharmacogenetic variants have yet to impact on clinical practice, yet the effect sizes (e.g. for metformin intolerance and OCT1 variants; or for metformin action and SLC2A2 variants) are potentially of clinical utility, especially if the genotype is already known at the point of prescribing. Over the next few years, increasing cohort sizes and linkage at scale to electronic medical records will provide considerable potential for stratification and novel target identification in diabetes. Copyright © 2018 Elsevier Ltd. All rights reserved.

  7. Gene silencing in Tribolium castaneum as a tool for the targeted identification of candidate RNAi targets in crop pests.

    PubMed

    Knorr, Eileen; Fishilevich, Elane; Tenbusch, Linda; Frey, Meghan L F; Rangasamy, Murugesan; Billion, Andre; Worden, Sarah E; Gandra, Premchand; Arora, Kanika; Lo, Wendy; Schulenberg, Greg; Valverde-Garcia, Pablo; Vilcinskas, Andreas; Narva, Kenneth E

    2018-02-01

    RNAi shows potential as an agricultural technology for insect control, yet, a relatively low number of robust lethal RNAi targets have been demonstrated to control insects of agricultural interest. In the current study, a selection of lethal RNAi target genes from the iBeetle (Tribolium castaneum) screen were used to demonstrate efficacy of orthologous targets in the economically important coleopteran pests Diabrotica virgifera virgifera and Meligethes aeneus. Transcript orthologs of 50 selected genes were analyzed in D. v. virgifera diet-based RNAi bioassays; 21 of these RNAi targets showed mortality and 36 showed growth inhibition. Low dose injection- and diet-based dsRNA assays in T. castaneum and D. v. virgifera, respectively, enabled the identification of the four highly potent RNAi target genes: Rop, dre4, ncm, and RpII140. Maize was genetically engineered to express dsRNA directed against these prioritized candidate target genes. T 0 plants expressing Rop, dre4, or RpII140 RNA hairpins showed protection from D. v. virgifera larval feeding damage. dsRNA targeting Rop, dre4, ncm, and RpII140 in M. aeneus also caused high levels of mortality both by injection and feeding. In summary, high throughput systems for model organisms can be successfully used to identify potent RNA targets for difficult-to-work with agricultural insect pests.

  8. Genetically Validated Drug Targets in Leishmania: Current Knowledge and Future Prospects.

    PubMed

    Jones, Nathaniel G; Catta-Preta, Carolina M C; Lima, Ana Paula C A; Mottram, Jeremy C

    2018-04-13

    There has been a very limited number of high-throughput screening campaigns carried out with Leishmania drug targets. In part, this is due to the small number of suitable target genes that have been shown by genetic or chemical methods to be essential for the parasite. In this perspective, we discuss the state of genetic target validation in the field of Leishmania research and review the 200 Leishmania genes and 36 Trypanosoma cruzi genes for which gene deletion attempts have been made since the first published case in 1990. We define a quality score for the different genetic deletion techniques that can be used to identify potential drug targets. We also discuss how the advances in genome-scale gene disruption techniques have been used to assist target-based and phenotypic-based drug development in other parasitic protozoa and why Leishmania has lacked a similar approach so far. The prospects for this scale of work are considered in the context of the application of CRISPR/Cas9 gene editing as a useful tool in Leishmania.

  9. A comparison of machine learning techniques for detection of drug target articles.

    PubMed

    Danger, Roxana; Segura-Bedmar, Isabel; Martínez, Paloma; Rosso, Paolo

    2010-12-01

    Important progress in treating diseases has been possible thanks to the identification of drug targets. Drug targets are the molecular structures whose abnormal activity, associated to a disease, can be modified by drugs, improving the health of patients. Pharmaceutical industry needs to give priority to their identification and validation in order to reduce the long and costly drug development times. In the last two decades, our knowledge about drugs, their mechanisms of action and drug targets has rapidly increased. Nevertheless, most of this knowledge is hidden in millions of medical articles and textbooks. Extracting knowledge from this large amount of unstructured information is a laborious job, even for human experts. Drug target articles identification, a crucial first step toward the automatic extraction of information from texts, constitutes the aim of this paper. A comparison of several machine learning techniques has been performed in order to obtain a satisfactory classifier for detecting drug target articles using semantic information from biomedical resources such as the Unified Medical Language System. The best result has been achieved by a Fuzzy Lattice Reasoning classifier, which reaches 98% of ROC area measure. Copyright © 2010 Elsevier Inc. All rights reserved.

  10. Specific detection and identification of [Actinobacillus] muris by PCR using primers targeting the 16S-23S rRNA internal transcribed spacer regions.

    PubMed

    Benga, Laurentiu; Benten, W Peter M; Engelhardt, Eva; Gougoula, Christina; Sager, Martin

    2013-08-01

    [Actinobacillus] muris represents along with [Pasteurella] pneumotropica the most prevalent Pasteurellaceae species isolated from the laboratory mouse. Despite the biological and economic importance of Pasteurellaceae in relation to experimental animals, no molecular based methods for the identification of [A.] muris are available. The aim of the present investigation was to develop a PCR method allowing detection and identification of [A.] muris. In this assay, a Pasteurellaceae common forward primer based on a conserved region of the 16S rRNA gene was used in conjunction with two different reverse primers specific for [A.] muris, targeting the 16S-23S internal transcribed spacer sequences. The specificity of the assay was tested against 78 reference and clinical isolates of Pasteurellaceae, including 37 strains of [A.] muris. In addition, eight other mice associated bacterial species which could pose a diagnostic problem were included. The assay showed 100% sensitivity and 97.95% specificity. Identification of the clinical isolates was validated by ITS profiling and when necessary by 16S rRNA sequencing. This multiplex PCR represents the first molecular tool able to detect [A.] muris and may become a reliable alternative to the present diagnostic methods. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Targeted Identification of Sialoglycoproteins in Hypoxic Endothelial Cells and Validation in Zebrafish Reveal Roles for Proteins in Angiogenesis

    PubMed Central

    Delcourt, Nicolas; Quevedo, Celia; Nonne, Christelle; Fons, Pierre; O'Brien, Donogh; Loyaux, Denis; Diez, Maria; Autelitano, François; Guillemot, Jean-Claude; Ferrara, Pascual; Muriana, Arantza; Callol, Carlos; Hérault, Jean-Pascal; Herbert, Jean-Marc; Favre, Gilles; Bono, Françoise

    2015-01-01

    The formation of new vessels in the tumor, termed angiogenesis, is essential for primary tumor growth and facilitates tumor invasion and metastasis. Hypoxia has been described as one trigger of angiogenesis. Indeed, hypoxia, which is characterized by areas of low oxygen levels, is a hallmark of solid tumors arising from an imbalance between oxygen delivery and consumption. Hypoxic conditions have profound effects on the different components of the tumoral environment. For example, hypoxia is able to activate endothelial cells, leading to angiogenesis but also thereby initiating a cascade of reactions involving neutrophils, smooth muscle cells, and fibroblasts. In addition, hypoxia directly regulates the expression of many genes for which the role and the importance in the tumoral environment remain to be completely elucidated. In this study, we used a method to selectively label sialoglycoproteins to identify new membrane and secreted proteins involved in the adaptative process of endothelial cells by mass spectrometry-based proteomics. We used an in vitro assay under hypoxic condition to observe an increase of protein expression or modifications of glycosylation. Then the function of the identified proteins was assessed in a vasculogenesis assay in vivo by using a morpholino strategy in zebrafish. First, our approach was validated by the identification of sialoglycoproteins such as CD105, neuropilin-1, and CLEC14A, which have already been described as playing key roles in angiogenesis. Second, we identified several new proteins regulated by hypoxia and demonstrated for the first time the pivotal role of GLUT-1, TMEM16F, and SDF4 in angiogenesis. PMID:25384978

  12. [Reliability and validity of the Chinese version on Alcohol Use Disorders Identification Test].

    PubMed

    Zhang, C; Yang, G P; Li, Z; Li, X N; Li, Y; Hu, J; Zhang, F Y; Zhang, X J

    2017-08-10

    Objective: To assess the reliability and validity of the Chinese version on Alcohol Use Disorders Identification Test (AUDIT) among medical students in China and to provide correct way of application on the recommended scales. Methods: An E-questionnaire was developed and sent to medical students in five different colleges. Students were all active volunteers to accept the testings. Cronbach's α and split-half reliability were calculated to evaluate the reliability of AUDIT while content, contract, discriminant and convergent validity were performed to measure the validity of the scales. Results: The overall Cronbach's α of AUDIT was 0.782 and the split-half reliability was 0.711. Data showed that the domain Cronbach's α and split-half reliability were 0.796 and 0.794 for hazardous alcohol use, 0.561 and 0.623 for dependence symptoms, and 0.647 and 0.640 for harmful alcohol use. Results also showed that the content validity index on the levels of items I-CVI) were from 0.83 to 1.00, the content validity index of scale level (S-CVI/UA) was 0.90, content validity index of average scale level (S-CVI/Ave) was 0.99 and the content validity ratios (CVR) were from 0.80 to 1.00. The simplified version of AUDIT supported a presupposed three-factor structure which could explain 61.175% of the total variance revealed through exploratory factor analysis. AUDIT semed to have good convergent and discriminant validity, with the success rate of calibration experiment as 100%. Conclusion: AUDIT showed good reliability and validity among medical students in China thus worth for promotion on its use.

  13. Identification of the Downstream Promoter Targets of Smad Tumor Suppressors in Human Breast Cancer Cells

    DTIC Science & Technology

    2004-10-01

    signaling mediator Smad2, Smad3 and Smad4 which form oligomeric complexes and migrate into nucleus to function as transcription factors to modulate... Smad3 and Smad4. 2. Identification of the downstream promoter targets of Smad3 or Smad4 in breast cancer cells. 3. Identify Smad4 regulated downstream...Development of a novel chromatin immunoprecipitation assay (CHIPS) using a TAP-TAG system to isolate in vivo binding targets of Smad3 and Smad4

  14. Raman fiber-optical method for colon cancer detection: Cross-validation and outlier identification approach

    NASA Astrophysics Data System (ADS)

    Petersen, D.; Naveed, P.; Ragheb, A.; Niedieker, D.; El-Mashtoly, S. F.; Brechmann, T.; Kötting, C.; Schmiegel, W. H.; Freier, E.; Pox, C.; Gerwert, K.

    2017-06-01

    Endoscopy plays a major role in early recognition of cancer which is not externally accessible and therewith in increasing the survival rate. Raman spectroscopic fiber-optical approaches can help to decrease the impact on the patient, increase objectivity in tissue characterization, reduce expenses and provide a significant time advantage in endoscopy. In gastroenterology an early recognition of malign and precursor lesions is relevant. Instantaneous and precise differentiation between adenomas as precursor lesions for cancer and hyperplastic polyps on the one hand and between high and low-risk alterations on the other hand is important. Raman fiber-optical measurements of colon biopsy samples taken during colonoscopy were carried out during a clinical study, and samples of adenocarcinoma (22), tubular adenomas (141), hyperplastic polyps (79) and normal tissue (101) from 151 patients were analyzed. This allows us to focus on the bioinformatic analysis and to set stage for Raman endoscopic measurements. Since spectral differences between normal and cancerous biopsy samples are small, special care has to be taken in data analysis. Using a leave-one-patient-out cross-validation scheme, three different outlier identification methods were investigated to decrease the influence of systematic errors, like a residual risk in misplacement of the sample and spectral dilution of marker bands (esp. cancerous tissue) and therewith optimize the experimental design. Furthermore other validations methods like leave-one-sample-out and leave-one-spectrum-out cross-validation schemes were compared with leave-one-patient-out cross-validation. High-risk lesions were differentiated from low-risk lesions with a sensitivity of 79%, specificity of 74% and an accuracy of 77%, cancer and normal tissue with a sensitivity of 79%, specificity of 83% and an accuracy of 81%. Additionally applied outlier identification enabled us to improve the recognition of neoplastic biopsy samples.

  15. A Validated Methodology for Genetic Identification of Tuna Species (Genus Thunnus)

    PubMed Central

    Viñas, Jordi; Tudela, Sergi

    2009-01-01

    Background Tuna species of the genus Thunnus, such as the bluefin tunas, are some of the most important and yet most endangered trade fish in the world. Identification of these species in traded forms, however, may be difficult depending on the presentation of the products, which may hamper conservation efforts on trade control. In this paper, we validated a genetic methodology that can fully distinguish between the eight Thunnus species from any kind of processed tissue. Methodology After testing several genetic markers, a complete discrimination of the eight tuna species was achieved using Forensically Informative Nucleotide Sequencing based primarily on the sequence variability of the hypervariable genetic marker mitochondrial DNA control region (mtDNA CR), followed, in some specific cases, by a second validation by a nuclear marker rDNA first internal transcribed spacer (ITS1). This methodology was able to distinguish all tuna species, including those belonging to the subgenus Neothunnus that are very closely related, and in consequence can not be differentiated with other genetic markers of lower variability. This methodology also took into consideration the presence of introgression that has been reported in past studies between T. thynnus, T. orientalis and T. alalunga. Finally, we applied the methodology to cross-check the species identity of 26 processed tuna samples. Conclusions Using the combination of two genetic markers, one mitochondrial and another nuclear, allows a full discrimination between all eight tuna species. Unexpectedly, the genetic marker traditionally used for DNA barcoding, cytochrome oxidase 1, could not differentiate all species, thus its use as a genetic marker for tuna species identification is questioned. PMID:19898615

  16. Identification of glycogen synthase kinase 3α as a therapeutic target in melanoma

    PubMed Central

    Madhunapantula, SubbaRao V.; Sharma, Arati; Gowda, Raghavendra; Robertson, Gavin P.

    2014-01-01

    Summary Deregulated expression or activity of kinases can lead to melanomas, but often the particular kinase isoform causing the effect is not well established, making identification and validation of different isoforms regulating disease development especially important. To accomplish this objective, an siRNA screen was undertaken that which identified glycogen synthase kinase 3α (GSK3α) as an important melanoma growth regulator. Melanocytes and melanoma cell lines representing various stages of melanoma tumor progression expressed both GSK3α and GSK3β, but analysis of tumors in patients with melanoma showed elevated expression of GSK3α in 72% of samples, which was not observed for GSK3β. Furthermore, 80% of tumors in patients with melanoma expressed elevated levels of catalytically active phosphorylated GSK3α (pGSK3αY279), but not phosphorylated GSK3β (pGSK3βY216). siRNA-mediated reduction in GSK3α protein levels reduced melanoma cell survival and proliferation, sensitized cells to apoptosis-inducing agents and decreased xenografted tumor development by up to 56%. Mechanistically, inhibiting GSK3α expression using siRNA or the pharmacological agent AR-A014418 arrested melanoma cells in the G0/G1 phase of the cell cycle and induced apoptotic death to retard tumorigenesis. Therefore, GSK3α is a key therapeutic target in melanoma. PMID:24034838

  17. Identification and validation of sRNAs in Edwardsiella tarda S08.

    PubMed

    Sun, Yuying; Zhang, Jiquan; Qin, Lei; Yan, Cui; Zhang, Xiaojun; Liu, Dandan

    2017-01-01

    Bacterial small non-coding RNAs (sRNAs) are known as novel regulators involved in virulence, stress responsibility, and so on. Recently, a lot of new researches have highlighted the critical roles of sRNAs in fine-tune gene regulation in both prokaryotes and eukaryotes. Edwardsiella tarda (E. tarda) is a gram-negative, intracellular pathogen that causes edwardsiellosis in fish. Thus far, no sRNA has been reported in E. tarda. The present study represents the first attempt to identify sRNAs in E. tarda S08. Ten sRNAs were validated by RNA sequencing and quantitative PCR (qPCR). ET_sRNA_1 and ET_sRNA_2 were homolous to tmRNA and GcvB, respectively. However, the other candidate sRNAs have not been reported till now. The cellular abundance of 10 validated sRNA was detected by qPCR at different growth phases to monitor their biosynthesis. Nine candidate sRNAs were expressed in the late-stage of exponential growth and stationary stages of growth (36~60 h). And the expression of the nine sRNAs was growth phase-dependent. But ET_sRNA_10 was almost expressed all the time and reached the highest peak at 48 h. Their targets were predicted by TargetRNA2 and each sRNA target contains some genes that directly or indirectly relate to virulence. These results preliminary showed that sRNAs probably play a regulatory role of virulence in E. tarda.

  18. Identification of apoptosis-related PLZF target genes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Bernardo, Maria Victoria; Yelo, Estefania; Gimeno, Lourdes

    2007-07-27

    The PLZF gene encodes a BTB/POZ-zinc finger-type transcription factor, involved in physiological development, proliferation, differentiation, and apoptosis. In this paper, we investigate proliferation, survival, and gene expression regulation in stable clones from the human haematopoietic K562, DG75, and Jurkat cell lines with inducible expression of PLZF. In Jurkat cells, but not in K562 and DG75 cells, PLZF induced growth suppression and apoptosis in a cell density-dependent manner. Deletion of the BTB/POZ domain of PLZF abrogated growth suppression and apoptosis. PLZF was expressed with a nuclear speckled pattern distinctively in the full-length PLZF-expressing Jurkat clones, suggesting that the nuclear speckled localizationmore » is required for PLZF-induced apoptosis. By microarray analysis, we identified that the apoptosis-inducer TP53INP1, ID1, and ID3 genes were upregulated, and the apoptosis-inhibitor TERT gene was downregulated. The identification of apoptosis-related PLZF target genes may have biological and clinical relevance in cancer typified by altered PLZF expression.« less

  19. EGFR, HER2 and VEGF pathways: validated targets for cancer treatment.

    PubMed

    Press, Michael F; Lenz, Heinz-Josef

    2007-01-01

    Targeted therapies are rationally designed to interfere with specific molecular events that are important in tumour growth, progression or survival. Several targeted therapies with anti-tumour activity in human cancer cell lines and xenograft models have now been shown to produce objective responses, delay disease progression and, in some cases, improve survival of patients with advanced malignancies. These targeted therapies include cetuximab, an anti-epidermal growth factor receptor (EGFR) monoclonal antibody; gefitinib and erlotinib, EGFR-specific tyrosine kinase inhibitors; trastuzumab, an anti-human EGFR type 2 (HER2)-related monoclonal antibody; lapatinib, a dual inhibitor of both EGFR- and HER2-associated tyrosine kinases; and bevacizumab, an anti-vascular endothelial growth factor (VEGF) monoclonal antibody. On the basis of preclinical and clinical evidence, EGFR, HER2 and VEGF represent validated targets for cancer therapy and remain the subject of intensive investigation. Both EGFR and HER2 are targets found on cancer cells, whereas VEGF is a target that acts in the tumour microenvironment. Clinical studies are focusing on how to best incorporate targeted therapy into current treatment regimens and other studies are exploring whether different strategies for inhibiting these targets will offer greater benefit. It is clear that optimal use of targeted therapy will depend on understanding how these drugs work mechanistically, and recognising that their activities may differ across patient populations, tumour types and disease stages, as well as when and how they are used in cancer treatment. The results achieved with targeted therapies to date are promising, although they illustrate the need for additional preclinical and clinical study.

  20. Network Understanding of Herb Medicine via Rapid Identification of Ingredient-Target Interactions

    NASA Astrophysics Data System (ADS)

    Zhang, Hai-Ping; Pan, Jian-Bo; Zhang, Chi; Ji, Nan; Wang, Hao; Ji, Zhi-Liang

    2014-01-01

    Today, herb medicines have become the major source for discovery of novel agents in countermining diseases. However, many of them are largely under-explored in pharmacology due to the limitation of current experimental approaches. Therefore, we proposed a computational framework in this study for network understanding of herb pharmacology via rapid identification of putative ingredient-target interactions in human structural proteome level. A marketing anti-cancer herb medicine in China, Yadanzi (Brucea javanica), was chosen for mechanistic study. Total 7,119 ingredient-target interactions were identified for thirteen Yadanzi active ingredients. Among them, about 29.5% were estimated to have better binding affinity than their corresponding marketing drug-target interactions. Further Bioinformatics analyses suggest that simultaneous manipulation of multiple proteins in the MAPK signaling pathway and the phosphorylation process of anti-apoptosis may largely answer for Yadanzi against non-small cell lung cancers. In summary, our strategy provides an efficient however economic solution for systematic understanding of herbs' power.

  1. Network understanding of herb medicine via rapid identification of ingredient-target interactions.

    PubMed

    Zhang, Hai-Ping; Pan, Jian-Bo; Zhang, Chi; Ji, Nan; Wang, Hao; Ji, Zhi-Liang

    2014-01-16

    Today, herb medicines have become the major source for discovery of novel agents in countermining diseases. However, many of them are largely under-explored in pharmacology due to the limitation of current experimental approaches. Therefore, we proposed a computational framework in this study for network understanding of herb pharmacology via rapid identification of putative ingredient-target interactions in human structural proteome level. A marketing anti-cancer herb medicine in China, Yadanzi (Brucea javanica), was chosen for mechanistic study. Total 7,119 ingredient-target interactions were identified for thirteen Yadanzi active ingredients. Among them, about 29.5% were estimated to have better binding affinity than their corresponding marketing drug-target interactions. Further Bioinformatics analyses suggest that simultaneous manipulation of multiple proteins in the MAPK signaling pathway and the phosphorylation process of anti-apoptosis may largely answer for Yadanzi against non-small cell lung cancers. In summary, our strategy provides an efficient however economic solution for systematic understanding of herbs' power.

  2. VALIDATION OF THE CORONAL THICK TARGET SOURCE MODEL

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fleishman, Gregory D.; Xu, Yan; Nita, Gelu N.

    2016-01-10

    We present detailed 3D modeling of a dense, coronal thick-target X-ray flare using the GX Simulator tool, photospheric magnetic measurements, and microwave imaging and spectroscopy data. The developed model offers a remarkable agreement between the synthesized and observed spectra and images in both X-ray and microwave domains, which validates the entire model. The flaring loop parameters are chosen to reproduce the emission measure, temperature, and the nonthermal electron distribution at low energies derived from the X-ray spectral fit, while the remaining parameters, unconstrained by the X-ray data, are selected such as to match the microwave images and total power spectra.more » The modeling suggests that the accelerated electrons are trapped in the coronal part of the flaring loop, but away from where the magnetic field is minimal, and, thus, demonstrates that the data are clearly inconsistent with electron magnetic trapping in the weak diffusion regime mediated by the Coulomb collisions. Thus, the modeling supports the interpretation of the coronal thick-target sources as sites of electron acceleration in flares and supplies us with a realistic 3D model with physical parameters of the acceleration region and flaring loop.« less

  3. Identification and characterization of potential druggable targets among hypothetical proteins of extensively drug resistant Mycobacterium tuberculosis (XDR KZN 605) through subtractive genomics approach.

    PubMed

    Uddin, Reaz; Siddiqui, Quratulain Nehal; Azam, Syed Sikander; Saima, Bibi; Wadood, Abdul

    2018-03-01

    Among the resistant isolates of tuberculosis (TB), the multidrug resistance tuberculosis (MDR-TB) and extensively drug resistant tuberculosis (XDR-TB) are the areas of growing concern for which the front-line antibiotics are no more effective. As a result, the search of new therapeutic targets against TB is an imperative need of time. On the other hand, the target identification is an a priori step in drug discovery based research. Furthermore, the availability of the complete proteomic data of extensively drug resistant Mycobacterium tuberculosis (XDR-MTB) made it possible to carry out in silico analysis for the discovery of new drug targets. In the current study, we aimed to prioritize the potential drug targets among the hypothetical proteins of XDR-TB via subtractive genomics approach. In the subtractive genomics, we stepwise reduced the complete proteome of XDR-MTB to only two hypothetical proteins and evidently proposed them as new therapeutic targets. The 3D structure of one of the two target proteins was predicted via homology modeling and later on, validated by various analysis tools. Our study suggested that the domains identified and the motif hits found in the sequences of the shortlisted drug targets are crucial for the survival of the XDR-MTB. To the best of our knowledge, the current study is the first attempt in which the complete proteomic data of XDR-MTB was subjected to the computational subtractive genomics approach and therefore, would provide an opportunity to identify the unique therapeutic targets against deadly XDR-MTB. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. The influence of camouflage, obstruction, familiarity and spatial ability on target identification from an unmanned ground vehicle.

    PubMed

    Fincannon, Thomas; Keebler, Joseph R; Jentsch, Florian; Curtis, Michael

    2013-01-01

    The purpose of this study was to examine the effects of environmental and cognitive factors on the identification of targets from an unmanned ground vehicle (UGV). This was accomplished by manipulating obstruction, camouflage and familiarity of objects in the environment, while also measuring spatial ability. The effects of these variables on target identification were studied by measuring performance of participants that observed pre-recorded video from a 1:35 scaled military operations in urban terrain facility. Analyses indicated that a combination of camouflage and obstruction caused the most detrimental effects on performance, and that there were differences in the recognition of familiar and unfamiliar targets. Further analysis indicated that these detrimental effects could only be overcome with a combination of target familiarity and spatial ability. The findings highlight the degree to which environmental factors hinder performance and the need for a multidimensional approach for improving performance under these conditions. Areas in need of future research are also discussed. Cognitive theory is applied to the problem of perception from UGVs. Results from an experimental study indicate that a combination of camouflage and obstruction caused the most detrimental effects on performance, with differences in the recognition of both familiar and unfamiliar targets. Familiarity and spatial ability interacted to predict the performance.

  5. Targeted identification of sialoglycoproteins in hypoxic endothelial cells and validation in zebrafish reveal roles for proteins in angiogenesis.

    PubMed

    Delcourt, Nicolas; Quevedo, Celia; Nonne, Christelle; Fons, Pierre; O'Brien, Donogh; Loyaux, Denis; Diez, Maria; Autelitano, François; Guillemot, Jean-Claude; Ferrara, Pascual; Muriana, Arantza; Callol, Carlos; Hérault, Jean-Pascal; Herbert, Jean-Marc; Favre, Gilles; Bono, Françoise

    2015-02-06

    The formation of new vessels in the tumor, termed angiogenesis, is essential for primary tumor growth and facilitates tumor invasion and metastasis. Hypoxia has been described as one trigger of angiogenesis. Indeed, hypoxia, which is characterized by areas of low oxygen levels, is a hallmark of solid tumors arising from an imbalance between oxygen delivery and consumption. Hypoxic conditions have profound effects on the different components of the tumoral environment. For example, hypoxia is able to activate endothelial cells, leading to angiogenesis but also thereby initiating a cascade of reactions involving neutrophils, smooth muscle cells, and fibroblasts. In addition, hypoxia directly regulates the expression of many genes for which the role and the importance in the tumoral environment remain to be completely elucidated. In this study, we used a method to selectively label sialoglycoproteins to identify new membrane and secreted proteins involved in the adaptative process of endothelial cells by mass spectrometry-based proteomics. We used an in vitro assay under hypoxic condition to observe an increase of protein expression or modifications of glycosylation. Then the function of the identified proteins was assessed in a vasculogenesis assay in vivo by using a morpholino strategy in zebrafish. First, our approach was validated by the identification of sialoglycoproteins such as CD105, neuropilin-1, and CLEC14A, which have already been described as playing key roles in angiogenesis. Second, we identified several new proteins regulated by hypoxia and demonstrated for the first time the pivotal role of GLUT-1, TMEM16F, and SDF4 in angiogenesis. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Estimation of Species Identification Error: Implications for Raptor Migration Counts and Trend Estimation

    Treesearch

    J.M. Hull; A.M. Fish; J.J. Keane; S.R. Mori; B.J Sacks; A.C. Hull

    2010-01-01

    One of the primary assumptions associated with many wildlife and population trend studies is that target species are correctly identified. This assumption may not always be valid, particularly for species similar in appearance to co-occurring species. We examined size overlap and identification error rates among Cooper's (Accipiter cooperii...

  7. Ground Target Modeling and Validation Conference (10th) Held in Houghton, Michigan, on 17-19 August 1999

    DTIC Science & Technology

    1999-08-01

    electrically small or only have a greater size in one dimension will not have a significant impact on the total RCS. At 1000 MHz, the components on the model ...7^/43- L"^y 16 % 6 ^Ly Cc>v y to-*^ r*r+r g,^\\oS^ Proceedings ? Tenth Annual Ground Target Modeling and Validation Conference August 1999...of the Tenth Annual Ground Target Modeling and Validation Conference (Unclassified) \\2. PERSONAL AUTHOR(S) William R Reynolds and Tracy T. Maki 13a

  8. Systems genetics for drug target discovery

    PubMed Central

    Penrod, Nadia M.; Cowper-Sal_lari, Richard; Moore, Jason H.

    2011-01-01

    The collection and analysis of genomic data has the potential to reveal novel druggable targets by providing insight into the genetic basis of disease. However, the number of drugs, targeting new molecular entities, approved by the US Food and Drug Administration (FDA) has not increased in the years since the collection of genomic data has become commonplace. The paucity of translatable results can be partly attributed to conventional analysis methods that test one gene at a time in an effort to identify disease-associated factors as candidate drug targets. By disengaging genetic factors from their position within the genetic regulatory system, much of the information stored within the genomic data set is lost. Here we discuss how genomic data is used to identify disease-associated genes or genomic regions, how disease-associated regions are validated as functional targets, and the role network analysis can play in bridging the gap between data generation and effective drug target identification. PMID:21862141

  9. Are multiple-trial experiments appropriate for eyewitness identification studies? Accuracy, choosing, and confidence across trials.

    PubMed

    Mansour, J K; Beaudry, J L; Lindsay, R C L

    2017-12-01

    Eyewitness identification experiments typically involve a single trial: A participant views an event and subsequently makes a lineup decision. As compared to this single-trial paradigm, multiple-trial designs are more efficient, but significantly reduce ecological validity and may affect the strategies that participants use to make lineup decisions. We examined the effects of a number of forensically relevant variables (i.e., memory strength, type of disguise, degree of disguise, and lineup type) on eyewitness accuracy, choosing, and confidence across 12 target-present and 12 target-absent lineup trials (N = 349; 8,376 lineup decisions). The rates of correct rejections and choosing (across both target-present and target-absent lineups) did not vary across the 24 trials, as reflected by main effects or interactions with trial number. Trial number had a significant but trivial quadratic effect on correct identifications (OR = 0.99) and interacted significantly, but again trivially, with disguise type (OR = 1.00). Trial number did not significantly influence participants' confidence in correct identifications, confidence in correct rejections, or confidence in target-absent selections. Thus, multiple-trial designs appear to have minimal effects on eyewitness accuracy, choosing, and confidence. Researchers should thus consider using multiple-trial designs for conducting eyewitness identification experiments.

  10. A statistical approach to selecting and confirming validation targets in -omics experiments

    PubMed Central

    2012-01-01

    Background Genomic technologies are, by their very nature, designed for hypothesis generation. In some cases, the hypotheses that are generated require that genome scientists confirm findings about specific genes or proteins. But one major advantage of high-throughput technology is that global genetic, genomic, transcriptomic, and proteomic behaviors can be observed. Manual confirmation of every statistically significant genomic result is prohibitively expensive. This has led researchers in genomics to adopt the strategy of confirming only a handful of the most statistically significant results, a small subset chosen for biological interest, or a small random subset. But there is no standard approach for selecting and quantitatively evaluating validation targets. Results Here we present a new statistical method and approach for statistically validating lists of significant results based on confirming only a small random sample. We apply our statistical method to show that the usual practice of confirming only the most statistically significant results does not statistically validate result lists. We analyze an extensively validated RNA-sequencing experiment to show that confirming a random subset can statistically validate entire lists of significant results. Finally, we analyze multiple publicly available microarray experiments to show that statistically validating random samples can both (i) provide evidence to confirm long gene lists and (ii) save thousands of dollars and hundreds of hours of labor over manual validation of each significant result. Conclusions For high-throughput -omics studies, statistical validation is a cost-effective and statistically valid approach to confirming lists of significant results. PMID:22738145

  11. Genome-wide identification of microRNA targets in the neglected disease pathogens of the genus Echinococcus.

    PubMed

    Macchiaroli, Natalia; Maldonado, Lucas L; Zarowiecki, Magdalena; Cucher, Marcela; Gismondi, María Inés; Kamenetzky, Laura; Rosenzvit, Mara Cecilia

    2017-06-01

    MicroRNAs (miRNAs), a class of small non-coding RNAs, are key regulators of gene expression at post-transcriptional level and play essential roles in biological processes such as development. MiRNAs silence target mRNAs by binding to complementary sequences in the 3'untranslated regions (3'UTRs). The parasitic helminths of the genus Echinococcus are the causative agents of echinococcosis, a zoonotic neglected disease. In previous work, we performed a comprehensive identification and characterization of Echinococcus miRNAs. However, current knowledge about their targets is limited. Since target prediction algorithms rely on complementarity between 3'UTRs and miRNA sequences, a major limitation is the lack of accurate sequence information of 3'UTR for most species including parasitic helminths. We performed RNA-seq and developed a pipeline that integrates the transcriptomic data with available genomic data of this parasite in order to identify 3'UTRs of Echinococcus canadensis. The high confidence set of 3'UTRs obtained allowed the prediction of miRNA targets in Echinococcus through a bioinformatic approach. We performed for the first time a comparative analysis of miRNA targets in Echinococcus and Taenia. We found that many evolutionarily conserved target sites in Echinococcus and Taenia may be functional and under selective pressure. Signaling pathways such as MAPK and Wnt were among the most represented pathways indicating miRNA roles in parasite growth and development. Genome-wide identification and characterization of miRNA target genes in Echinococcus provide valuable information to guide experimental studies in order to understand miRNA functions in the parasites biology. miRNAs involved in essential functions, especially those being absent in the host or showing sequence divergence with respect to host orthologs, might be considered as novel therapeutic targets for echinococcosis control. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Cross-cultural adaptation, reliability, and validation of the Korean version of the identification functional ankle instability (IdFAI).

    PubMed

    Ko, Jupil; Rosen, Adam B; Brown, Cathleen N

    2017-09-12

    To cross-culturally adapt the Identification Functional Ankle Instability for use with Korean-speaking participants. The English version of the IdFAI was cross-culturally adapted into Korean based on the guidelines. The psychometric properties in the Korean version of the IdFAI were measured for test-retest reliability, internal consistency, criterion-related validity, discriminative validity, and measurement error 181 native Korean-speakers. Intra-class correlation coefficients (ICC 2,1 ) between the English and Korean versions of the IdFAI for test-retest reliability was 0.98 (standard error of measurement = 1.41). The Cronbach's alpha coefficient was 0.89 for the Korean versions of IdFAI. The Korean versions of the IdFAI had a strong correlation with the SF-36 (r s  = -0.69, p < .001) and the Korean version of the Cumberland Ankle Instability Tool (r s  = -0.65, p < .001). The cutoff score of >10 was the optimal cutoff score to distinguish between the group memberships. The minimally detectable change of the Korean versions of the IdFAI score was 3.91. The Korean versions of the IdFAI have shown to be an excellent, reliable, and valid instrument. The Korean versions of the IdFAI can be utilized to assess the presence of Chronic Ankle Instability by researchers and clinicians working among Korean-speaking populations. Implications for rehabilitation The high recurrence rate of sprains may result into Chronic Ankle Instability (CAI). The Identification of Functional Ankle Instability Tool (IdFAI) has been validated and recommended to identify patients with Chronic Ankle Instability (CAI). The Korean version of the Identification of Functional Ankle Instability Tool (IdFAI) may be also recommend to researchers and clinicians for assessing the presence of Chronic Ankle Instability (CAI) in Korean-speaking population.

  13. Development and Validation of a Rotor-Gene Real-Time PCR Assay for Detection, Identification, and Quantification of Chlamydia trachomatis in a Single Reaction

    PubMed Central

    Jalal, Hamid; Stephen, Hannah; Curran, Martin D.; Burton, Janet; Bradley, Michelle; Carne, Christopher

    2006-01-01

    A multitarget real-time PCR (MRT-PCR) for detection of Chlamydia trachomatis DNA was developed and validated. There were three targets for amplification in a single reaction: the cryptic plasmid (CP), the major outer membrane protein (MOMP) gene, and an internal control. The assay had the following characteristics: (i) detection and confirmation of the presence of C. trachomatis DNA in a single reaction, (ii) detection of all genovars of C. trachomatis without any cross-reactivity with pathogenic bacteria or commensal organisms of the oropharynx and genital tract, (iii) a 95% probability of detection with three copies of MOMP and one copy of CP per reaction mixture, (iv) identification of the inhibition of amplification, (v) a quantitative dynamic range of 25 to 250,000 genome copies per reaction mixture, (vi) high intra- and interassay reproducibilities, and (vii) correct identification of all samples in the validation panel. There were 146 COBAS Amplicor PCR (Amplicor PCR)-positive samples and 122 Amplicor PCR-negative samples in the panel. MRT-PCR detected CP DNA alone in 6 (4%) Amplicor PCR-positive samples and both CP and MOMP DNAs in 140 (96%) of 146 Amplicor PCR-positive samples. The quantity of MOMP DNA in 95 (68%) of 140 samples was within the dynamic range of the assay. The median C. trachomatis load in these samples was 321 genome copies per reaction mixture (range, 26 to 40,137 genome copies per reaction mixture). Due to the inclusion of two different C. trachomatis-specific targets, the assay confirmed 259 (97%) of 268 results in a single reaction. This assay could be used in the qualitative format for the routine detection of C. trachomatis and in the quantitative format for study of the pathogenesis of C. trachomatis-associated diseases. The assay demonstrated the potential to eliminate the need for confirmatory testing in almost all samples, thus reducing the turnaround time and the workload. PMID:16390971

  14. Advanced forensic validation for human spermatozoa identification using SPERM HY-LITER™ Express with quantitative image analysis.

    PubMed

    Takamura, Ayari; Watanabe, Ken; Akutsu, Tomoko

    2017-07-01

    Identification of human semen is indispensable for the investigation of sexual assaults. Fluorescence staining methods using commercial kits, such as the series of SPERM HY-LITER™ kits, have been useful to detect human sperm via strong fluorescence. These kits have been examined from various forensic aspects. However, because of a lack of evaluation methods, these studies did not provide objective, or quantitative, descriptions of the results nor clear criteria for the decisions reached. In addition, the variety of validations was considerably limited. In this study, we conducted more advanced validations of SPERM HY-LITER™ Express using our established image analysis method. Use of this method enabled objective and specific identification of fluorescent sperm's spots and quantitative comparisons of the sperm detection performance under complex experimental conditions. For body fluid mixtures, we examined interference with the fluorescence staining from other body fluid components. Effects of sample decomposition were simulated in high humidity and high temperature conditions. Semen with quite low sperm concentrations, such as azoospermia and oligospermia samples, represented the most challenging cases in application of the kit. Finally, the tolerance of the kit against various acidic and basic environments was analyzed. The validations herein provide useful information for the practical applications of the SPERM HY-LITER™ Express kit, which were previously unobtainable. Moreover, the versatility of our image analysis method toward various complex cases was demonstrated.

  15. The verbal facilitation effect: re-reading person descriptions as a system variable to improve identification performance.

    PubMed

    Sporer, Siegfried L; Kaminski, Kristina S; Davids, Maike C; McQuiston, Dawn

    2016-11-01

    When witnesses report a crime, police usually ask for a description of the perpetrator. Several studies suggested that verbalising faces leads to a detriment in identification performance (verbal overshadowing effect [VOE]) but the effect has been difficult to replicate. Here, we sought to reverse the VOE by inducing context reinstatement as a system variable through re-reading one's own description before an identification task. Participants (N = 208) watched a video film and were then dismissed (control group), only described the perpetrator, or described and later re-read their own descriptions before identification in either target-present or target-absent lineups after a 2-day or a 5-week delay. Identification accuracy was significantly higher after re-reading (85.0%) than in the no description control group (62.5%) irrespective of target presence. Data were internally replicated using a second target and corroborated by several small meta-analyses. Identification accuracy was related to description quality. Moreover, there was a tendency towards a verbal facilitation effect (VFE) rather than a VOE. Receiver operating characteristic (ROC) curve analyses confirm that our findings are not due to a shift in response bias but truly reflect improvement of recognition performance. Differences in the ecological validity of study paradigms are discussed.

  16. Development, validation and implementation of a quadruplex real-time PCR assay for identification of potentially toxigenic corynebacteria.

    PubMed

    De Zoysa, Aruni; Efstratiou, Androulla; Mann, Ginder; Harrison, Timothy G; Fry, Norman K

    2016-12-01

    Toxigenic corynebacteria are uncommon in the UK; however, laboratory confirmation by the national reference laboratory can inform public health action according to national guidelines. Standard phenotypic tests for identification and toxin expression of isolates can take from ≥24 to ≥48 h from receipt. To decrease the time to result, a real-time PCR (qPCR) assay was developed for confirmation of both identification of Corynebacterium diphtheriae and Corynebacterium ulcerans/Corynebacterium pseudotuberculosis and detection of the diphtheria toxin gene. Target genes were the RNA polymerase β-subunit-encoding gene (rpoB) and A-subunit of the diphtheria toxin gene (tox). Green fluorescent protein DNA (gfp) was used as an internal process control. qPCR results were obtained within 3 to 4 h after receipt of isolate. The assay was validated according to published guidelines and demonstrated high diagnostic sensitivity (100 %), high specificity (98-100 %) and positive and negative predictive values of 91 to 100 % and 100 %, respectively, compared to both block-based PCR and the Elek test, together with a greatly reduced time from isolate receipt to reporting. Limitations of the qPCR assay were the inability to distinguish between C. ulcerans and C. pseudotuberculosis and that the presence of the toxin gene as demonstrated by qPCR may not always predict toxin expression. Thus, confirmation of expression of diphtheria toxin is always sought using the phenotypic Elek test. The new qPCR assay was formally introduced as the front-line test for putative toxigenic corynebacteria to inform public health action in England and Wales on 1 April 2014.

  17. Experiment research on infrared targets signature in mid and long IR spectral bands

    NASA Astrophysics Data System (ADS)

    Wang, Chensheng; Hong, Pu; Lei, Bo; Yue, Song; Zhang, Zhijie; Ren, Tingting

    2013-09-01

    Since the infrared imaging system has played a significant role in the military self-defense system and fire control system, the radiation signature of IR target becomes an important topic in IR imaging application technology. IR target signature can be applied in target identification, especially for small and dim targets, as well as the target IR thermal design. To research and analyze the targets IR signature systematically, a practical and experimental project is processed under different backgrounds and conditions. An infrared radiation acquisition system based on a MWIR cooled thermal imager and a LWIR cooled thermal imager is developed to capture the digital infrared images. Furthermore, some instruments are introduced to provide other parameters. According to the original image data and the related parameters in a certain scene, the IR signature of interested target scene can be calculated. Different background and targets are measured with this approach, and a comparison experiment analysis shall be presented in this paper as an example. This practical experiment has proved the validation of this research work, and it is useful in detection performance evaluation and further target identification research.

  18. Human genetics as a model for target validation: finding new therapies for diabetes.

    PubMed

    Thomsen, Soren K; Gloyn, Anna L

    2017-06-01

    Type 2 diabetes is a global epidemic with major effects on healthcare expenditure and quality of life. Currently available treatments are inadequate for the prevention of comorbidities, yet progress towards new therapies remains slow. A major barrier is the insufficiency of traditional preclinical models for predicting drug efficacy and safety. Human genetics offers a complementary model to assess causal mechanisms for target validation. Genetic perturbations are 'experiments of nature' that provide a uniquely relevant window into the long-term effects of modulating specific targets. Here, we show that genetic discoveries over the past decades have accurately predicted (now known) therapeutic mechanisms for type 2 diabetes. These findings highlight the potential for use of human genetic variation for prospective target validation, and establish a framework for future applications. Studies into rare, monogenic forms of diabetes have also provided proof-of-principle for precision medicine, and the applicability of this paradigm to complex disease is discussed. Finally, we highlight some of the limitations that are relevant to the use of genome-wide association studies (GWAS) in the search for new therapies for diabetes. A key outstanding challenge is the translation of GWAS signals into disease biology and we outline possible solutions for tackling this experimental bottleneck.

  19. Validity of Teacher Ratings in Selecting Influential Aggressive Adolescents for a Targeted Preventive Intervention

    PubMed Central

    Henry, David B.; Miller-Johnson, Shari; Simon, Thomas R.; Schoeny, Michael E.

    2009-01-01

    This study describes a method for using teacher nominations and ratings to identify socially influential, aggressive middle school students for participation in a targeted violence prevention intervention. The teacher nomination method is compared with peer nominations of aggression and influence to obtain validity evidence. Participants were urban, predominantly African American and Latino sixth-grade students who were involved in a pilot study for a large multi-site violence prevention project. Convergent validity was suggested by the high correlation of teacher ratings of peer influence and peer nominations of social influence. The teacher ratings of influence demonstrated acceptable sensitivity and specificity when predicting peer nominations of influence among the most aggressive children. Results are discussed m terms of the application of teacher nominations and ratings in large trials and full implementation of targeted prevention programs. PMID:16378226

  20. Nonlinear system identification technique validation

    NASA Astrophysics Data System (ADS)

    Rudko, M.; Bussgang, J. J.

    1982-01-01

    This final technical report describes the results obtained by SIGNATRON, Inc. of Lexington MA on Air Force Contract F30602-80-C-0104 for Rome Air Development Center. The objective of this effort is to develop a technique for identifying system response of nonlinear circuits by measurements of output response to known inputs. The report describes results of a study into the system identification technique based on the pencil-of-function method previously explored by Jain (1974) and Ewen (1979). The procedure identified roles of the linear response and is intended as a first step in nonlinear response and is intended as a first step in nonlinear circuit identification. There are serious implementation problems associated with the original approach such as loss of accuracy due to repeated integrations, lack of good measures of accuracy and computational iteration to identify the number of poles.

  1. Structure identification by Mass Spectrometry Non-Targeted Analysis using the US EPA’s CompTox Chemistry Dashboard

    EPA Science Inventory

    Identification of unknowns in mass spectrometry based non-targeted analyses (NTA) requires the integration of complementary pieces of data to arrive at a confident, consensus structure. Researchers use chemical reference databases, spectral matching, fragment prediction tools, r...

  2. Complex network theory for the identification and assessment of candidate protein targets.

    PubMed

    McGarry, Ken; McDonald, Sharon

    2018-06-01

    In this work we use complex network theory to provide a statistical model of the connectivity patterns of human proteins and their interaction partners. Our intention is to identify important proteins that may be predisposed to be potential candidates as drug targets for therapeutic interventions. Target proteins usually have more interaction partners than non-target proteins, but there are no hard-and-fast rules for defining the actual number of interactions. We devise a statistical measure for identifying hub proteins, we score our target proteins with gene ontology annotations. The important druggable protein targets are likely to have similar biological functions that can be assessed for their potential therapeutic value. Our system provides a statistical analysis of the local and distant neighborhood protein interactions of the potential targets using complex network measures. This approach builds a more accurate model of drug-to-target activity and therefore the likely impact on treating diseases. We integrate high quality protein interaction data from the HINT database and disease associated proteins from the DrugTarget database. Other sources include biological knowledge from Gene Ontology and drug information from DrugBank. The problem is a very challenging one since the data is highly imbalanced between target proteins and the more numerous nontargets. We use undersampling on the training data and build Random Forest classifier models which are used to identify previously unclassified target proteins. We validate and corroborate these findings from the available literature. Copyright © 2018 Elsevier Ltd. All rights reserved.

  3. Nuclear Security: Target Analysis-rev

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Singh, Surinder Paul; Gibbs, Philip W.; Bultz, Garl A.

    2014-03-01

    The objectives of this presentation are to understand target identification, including roll-up and protracted theft; evaluate target identification in the SNRI; recognize the target characteristics and consequence levels; and understand graded safeguards.

  4. [SIMBO: a Screening Instrument for Identification of Work-Related Disabilities--analyses of construct and prognostic validity].

    PubMed

    Streibelt, M; Gerwinn, H; Hansmeier, T; Thren, K; Müller-Fahrnow, W

    2007-10-01

    For a number of years, work-related interventions in medical rehabilitation (MBO) have been developed. Basically, these interventions concentrate on vocational problems of rehabilitees whose health disorders are strongly associated with contextual factors of the environment as well as personal factors. Previous studies showed a close relationship between the success of an intervention and identification of a specific demand. In fact there are several clinical concepts regarding specific demand. But there still is a lack of appropriate instruments for use in identification of occupational challenges. Therefore SIMBO (Screening Instrument for Identification of a Demand for Medical-Vocational Oriented Rehabilitation) has been developed recently. By using a scale for the intensity of work-related problems as well as a cut-off point, SIMBO is able to identify patients with and without a demand for work-related interventions. Analyses relative to construct validity and predictive validity were carried out on two different samples--a multi-clinic sample (patients with musculoskeletal disorders) and a sample from the German statutory pension insurance agency DRV Westfalen (successful applications for medical rehabilitation). In this context the cut-off level discussion is very important. By means of the multi-clinic sample--irrespective of cut-off definition--the SIMBO-decision and the clinical identification of MBO-demand were found to agree in 74-78% of the cases. This corresponds to a maximum adjusted correlation of r=0.59 (phi coefficient). Compared to the external ratings of vocational problems given by DRV staff in handling the applications, however, only little agreement is found (64%, r=0.25). In fact, SIMBO had in 77% (r=0.50) of the cases been able to correctly predict work-related problems to be expected. So the result obtained using this instrument is far better than prediction of these problems in the external ratings by DRV staff (54%, r=0,21). Also, return

  5. Experimental design and data analysis of Ago-RIP-Seq experiments for the identification of microRNA targets.

    PubMed

    Tichy, Diana; Pickl, Julia Maria Anna; Benner, Axel; Sültmann, Holger

    2017-03-31

    The identification of microRNA (miRNA) target genes is crucial for understanding miRNA function. Many methods for the genome-wide miRNA target identification have been developed in recent years; however, they have several limitations including the dependence on low-confident prediction programs and artificial miRNA manipulations. Ago-RNA immunoprecipitation combined with high-throughput sequencing (Ago-RIP-Seq) is a promising alternative. However, appropriate statistical data analysis algorithms taking into account the experimental design and the inherent noise of such experiments are largely lacking.Here, we investigate the experimental design for Ago-RIP-Seq and examine biostatistical methods to identify de novo miRNA target genes. Statistical approaches considered are either based on a negative binomial model fit to the read count data or applied to transformed data using a normal distribution-based generalized linear model. We compare them by a real data simulation study using plasmode data sets and evaluate the suitability of the approaches to detect true miRNA targets by sensitivity and false discovery rates. Our results suggest that simple approaches like linear regression models on (appropriately) transformed read count data are preferable. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  6. Proteomic Identification of Putative MicroRNA394 Target Genes in Arabidopsis thaliana Identifies Major Latex Protein Family Members Critical for Normal Development.

    PubMed

    Litholdo, Celso G; Parker, Benjamin L; Eamens, Andrew L; Larsen, Martin R; Cordwell, Stuart J; Waterhouse, Peter M

    2016-06-01

    Expression of the F-Box protein Leaf Curling Responsiveness (LCR) is regulated by microRNA, miR394, and alterations to this interplay in Arabidopsis thaliana produce defects in leaf polarity and shoot apical meristem organization. Although the miR394-LCR node has been documented in Arabidopsis, the identification of proteins targeted by LCR F-box itself has proven problematic. Here, a proteomic analysis of shoot apices from plants with altered LCR levels identified a member of the Latex Protein (MLP) family gene as a potential LCR F-box target. Bioinformatic and molecular analyses also suggested that other MLP family members are likely to be targets for this post-translational regulation. Direct interaction between LCR F-Box and MLP423 was validated. Additional MLP members had reduction in protein accumulation, in varying degrees, mediated by LCR F-Box. Transgenic Arabidopsis lines, in which MLP28 expression was reduced through an artificial miRNA technology, displayed severe developmental defects, including changes in leaf patterning and morphology, shoot apex defects, and eventual premature death. These phenotypic characteristics resemble those of Arabidopsis plants modified to over-express LCR Taken together, the results demonstrate that MLPs are driven to degradation by LCR, and indicate that MLP gene family is target of miR394-LCR regulatory node, representing potential targets for directly post-translational regulation mediated by LCR F-Box. In addition, MLP28 family member is associated with the LCR regulation that is critical for normal Arabidopsis development. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Proteomic Identification of Putative MicroRNA394 Target Genes in Arabidopsis thaliana Identifies Major Latex Protein Family Members Critical for Normal Development*

    PubMed Central

    Litholdo, Celso G.; Parker, Benjamin L.; Eamens, Andrew L.; Larsen, Martin R.; Cordwell, Stuart J.; Waterhouse, Peter M.

    2016-01-01

    Expression of the F-Box protein Leaf Curling Responsiveness (LCR) is regulated by microRNA, miR394, and alterations to this interplay in Arabidopsis thaliana produce defects in leaf polarity and shoot apical meristem organization. Although the miR394-LCR node has been documented in Arabidopsis, the identification of proteins targeted by LCR F-box itself has proven problematic. Here, a proteomic analysis of shoot apices from plants with altered LCR levels identified a member of the Latex Protein (MLP) family gene as a potential LCR F-box target. Bioinformatic and molecular analyses also suggested that other MLP family members are likely to be targets for this post-translational regulation. Direct interaction between LCR F-Box and MLP423 was validated. Additional MLP members had reduction in protein accumulation, in varying degrees, mediated by LCR F-Box. Transgenic Arabidopsis lines, in which MLP28 expression was reduced through an artificial miRNA technology, displayed severe developmental defects, including changes in leaf patterning and morphology, shoot apex defects, and eventual premature death. These phenotypic characteristics resemble those of Arabidopsis plants modified to over-express LCR. Taken together, the results demonstrate that MLPs are driven to degradation by LCR, and indicate that MLP gene family is target of miR394-LCR regulatory node, representing potential targets for directly post-translational regulation mediated by LCR F-Box. In addition, MLP28 family member is associated with the LCR regulation that is critical for normal Arabidopsis development. PMID:27067051

  8. Two-dimensional hidden semantic information model for target saliency detection and eyetracking identification

    NASA Astrophysics Data System (ADS)

    Wan, Weibing; Yuan, Lingfeng; Zhao, Qunfei; Fang, Tao

    2018-01-01

    Saliency detection has been applied to the target acquisition case. This paper proposes a two-dimensional hidden Markov model (2D-HMM) that exploits the hidden semantic information of an image to detect its salient regions. A spatial pyramid histogram of oriented gradient descriptors is used to extract features. After encoding the image by a learned dictionary, the 2D-Viterbi algorithm is applied to infer the saliency map. This model can predict fixation of the targets and further creates robust and effective depictions of the targets' change in posture and viewpoint. To validate the model with a human visual search mechanism, two eyetrack experiments are employed to train our model directly from eye movement data. The results show that our model achieves better performance than visual attention. Moreover, it indicates the plausibility of utilizing visual track data to identify targets.

  9. Identification and characterization of microRNAs and their target genes from Nile tilapia (Oreochromis niloticus).

    PubMed

    Huang, Yong; Ma, Xiu Ying; Yang, You Bing; Ren, Hong Tao; Sun, Xi Hong; Wang, Li Rui

    MicroRNAs (miRNAs) are a class of small single-stranded, endogenous 21-22 nt non-coding RNAs that regulate their target mRNA levels by causing either inactivation or degradation of the mRNAs. In recent years, miRNA genes have been identified from mammals, insects, worms, plants, and viruses. In this research, bioinformatics approaches were used to predict potential miRNAs and their targets in Nile tilapia from the expressed sequence tag (EST) and genomic survey sequence (GSS) database, respectively, based on the conservation of miRNAs in many animal species. A total of 19 potential miRNAs were detected following a range of strict filtering criteria. To test the validity of the bioinformatics method, seven predicted Nile tilapia miRNA genes were selected for further biological validation, and their mature miRNA transcripts were successfully detected by stem-loop RT-PCR experiments. Using these potential miRNAs, we found 56 potential targets in this species. Most of the target mRNAs appear to be involved in development, metabolism, signal transduction, transcription regulation and stress responses. Overall, our findings will provide an important foundation for further research on miRNAs function in the Nile tilapia.

  10. Large-Scale Chemical Similarity Networks for Target Profiling of Compounds Identified in Cell-Based Chemical Screens

    PubMed Central

    Lo, Yu-Chen; Senese, Silvia; Li, Chien-Ming; Hu, Qiyang; Huang, Yong; Damoiseaux, Robert; Torres, Jorge Z.

    2015-01-01

    Target identification is one of the most critical steps following cell-based phenotypic chemical screens aimed at identifying compounds with potential uses in cell biology and for developing novel disease therapies. Current in silico target identification methods, including chemical similarity database searches, are limited to single or sequential ligand analysis that have limited capabilities for accurate deconvolution of a large number of compounds with diverse chemical structures. Here, we present CSNAP (Chemical Similarity Network Analysis Pulldown), a new computational target identification method that utilizes chemical similarity networks for large-scale chemotype (consensus chemical pattern) recognition and drug target profiling. Our benchmark study showed that CSNAP can achieve an overall higher accuracy (>80%) of target prediction with respect to representative chemotypes in large (>200) compound sets, in comparison to the SEA approach (60–70%). Additionally, CSNAP is capable of integrating with biological knowledge-based databases (Uniprot, GO) and high-throughput biology platforms (proteomic, genetic, etc) for system-wise drug target validation. To demonstrate the utility of the CSNAP approach, we combined CSNAP's target prediction with experimental ligand evaluation to identify the major mitotic targets of hit compounds from a cell-based chemical screen and we highlight novel compounds targeting microtubules, an important cancer therapeutic target. The CSNAP method is freely available and can be accessed from the CSNAP web server (http://services.mbi.ucla.edu/CSNAP/). PMID:25826798

  11. Identification of Forensically Important Calliphoridae and Sarcophagidae Species Collected in Korea Using SNaPshot Multiplex System Targeting the Cytochrome c Oxidase Subunit I Gene

    PubMed Central

    Park, Ji Hye

    2018-01-01

    Estimation of postmortem interval (PMI) is paramount in modern forensic investigation. After the disappearance of the early postmortem phenomena conventionally used to estimate PMI, entomologic evidence provides important indicators for PMI estimation. The age of the oldest fly larvae or pupae can be estimated to pinpoint the time of oviposition, which is considered the minimum PMI (PMImin). The development rate of insects is usually temperature dependent and species specific. Therefore, species identification is mandatory for PMImin estimation using entomological evidence. The classical morphological identification method cannot be applied when specimens are damaged or have not yet matured. To overcome this limitation, some investigators employ molecular identification using mitochondrial cytochrome c oxidase subunit I (COI) nucleotide sequences. The molecular identification method commonly uses Sanger's nucleotide sequencing and molecular phylogeny, which are complex and time consuming and constitute another obstacle for forensic investigators. In this study, instead of using conventional Sanger's nucleotide sequencing, single-nucleotide polymorphisms (SNPs) in the COI gene region, which are unique between fly species, were selected and targeted for single-base extension (SBE) technology. These SNPs were genotyped using a SNaPshot® kit. Eleven Calliphoridae and seven Sarcophagidae species were covered. To validate this genotyping, fly DNA samples (103 adults, 84 larvae, and 4 pupae) previously confirmed by DNA barcoding were used. This method worked quickly with minimal DNA, providing a potential alternative to conventional DNA barcoding. Consisting of only a few simple electropherogram peaks, the results were more straightforward compared with those of the conventional DNA barcoding produced by Sanger's nucleotide sequencing. PMID:29682531

  12. The identification of new protein kinase inhibitors as targets in modern drug discovery.

    PubMed

    Akritopoulou-Zanze, Irini

    2006-07-01

    In recent years there has been great interest in developing protein kinase inhibitors as therapeutic agents for a variety of diseases. This article provides an overview on the history, development and validity of kinases as drug targets, as well as a description of kinase research, including its limitations, challenges and successes.

  13. Faster experimental validation of microRNA targets using cold fusion cloning and a dual firefly-Renilla luciferase reporter assay.

    PubMed

    Alvarez, M Lucrecia

    2014-01-01

    Different target prediction algorithms have been developed to provide a list of candidate target genes for a given animal microRNAs (miRNAs). However, these computational approaches provide both false-positive and false-negative predictions. Therefore, the target genes of a specific miRNA identified in silico should be experimentally validated. In this chapter, we describe a step-by-step protocol for the experimental validation of a direct miRNA target using a faster Dual Firefly-Renilla Luciferase Reporter Assay. We describe how to construct reporter plasmids using the simple, fast, and highly efficient cold fusion cloning technology, which does not require ligase, phosphatase, or restriction enzymes. In addition, we provide a protocol for co-transfection of reporter plasmids with either miRNA mimics or miRNA inhibitors in human embryonic kidney 293 (HEK293) cells, as well as a description on how to measure Firefly and Renilla luciferase activity using the Dual-Glo Luciferase Assay kit. As an example of the use of this technology, we will validate glucose-6-phosphate dehydrogenase (G6PD) as a direct target of miR-1207-5p.

  14. Validating An Analytic Completeness Model for Kepler Target Stars Based on Flux-level Transit Injection Experiments

    NASA Astrophysics Data System (ADS)

    Catanzarite, Joseph; Burke, Christopher J.; Li, Jie; Seader, Shawn; Haas, Michael R.; Batalha, Natalie; Henze, Christopher; Christiansen, Jessie; Kepler Project, NASA Advanced Supercomputing Division

    2016-06-01

    The Kepler Mission is developing an Analytic Completeness Model (ACM) to estimate detection completeness contours as a function of exoplanet radius and period for each target star. Accurate completeness contours are necessary for robust estimation of exoplanet occurrence rates.The main components of the ACM for a target star are: detection efficiency as a function of SNR, the window function (WF) and the one-sigma depth function (OSDF). (Ref. Burke et al. 2015). The WF captures the falloff in transit detection probability at long periods that is determined by the observation window (the duration over which the target star has been observed). The OSDF is the transit depth (in parts per million) that yields SNR of unity for the full transit train. It is a function of period, and accounts for the time-varying properties of the noise and for missing or deweighted data.We are performing flux-level transit injection (FLTI) experiments on selected Kepler target stars with the goal of refining and validating the ACM. “Flux-level” injection machinery inserts exoplanet transit signatures directly into the flux time series, as opposed to “pixel-level” injection, which inserts transit signatures into the individual pixels using the pixel response function. See Jie Li's poster: ID #2493668, "Flux-level transit injection experiments with the NASA Pleiades Supercomputer" for details, including performance statistics.Since FLTI is affordable for only a small subset of the Kepler targets, the ACM is designed to apply to most Kepler target stars. We validate this model using “deep” FLTI experiments, with ~500,000 injection realizations on each of a small number of targets and “shallow” FLTI experiments with ~2000 injection realizations on each of many targets. From the results of these experiments, we identify anomalous targets, model their behavior and refine the ACM accordingly.In this presentation, we discuss progress in validating and refining the ACM, and we

  15. A combined pre-clinical meta-analysis and randomized confirmatory trial approach to improve data validity for therapeutic target validation.

    PubMed

    Kleikers, Pamela W M; Hooijmans, Carlijn; Göb, Eva; Langhauser, Friederike; Rewell, Sarah S J; Radermacher, Kim; Ritskes-Hoitinga, Merel; Howells, David W; Kleinschnitz, Christoph; Schmidt, Harald H H W

    2015-08-27

    Biomedical research suffers from a dramatically poor translational success. For example, in ischemic stroke, a condition with a high medical need, over a thousand experimental drug targets were unsuccessful. Here, we adopt methods from clinical research for a late-stage pre-clinical meta-analysis (MA) and randomized confirmatory trial (pRCT) approach. A profound body of literature suggests NOX2 to be a major therapeutic target in stroke. Systematic review and MA of all available NOX2(-/y) studies revealed a positive publication bias and lack of statistical power to detect a relevant reduction in infarct size. A fully powered multi-center pRCT rejects NOX2 as a target to improve neurofunctional outcomes or achieve a translationally relevant infarct size reduction. Thus stringent statistical thresholds, reporting negative data and a MA-pRCT approach can ensure biomedical data validity and overcome risks of bias.

  16. Focus on flaviviruses: current and future drug targets.

    PubMed

    Geiss, Brian J; Stahla, Hillary; Hannah, Amanda M; Gari, Amanda M; Keenan, Susan M

    2009-05-01

    Infection by mosquito-borne flaviviruses (family Flaviviridae) is increasing in prevalence worldwide. The vast global, social and economic impact due to the morbidity and mortality associated with the diseases caused by these viruses necessitates therapeutic intervention. There is currently no effective clinical treatment for any flaviviral infection. Therefore, there is a great need for the identification of novel inhibitors to target the virus life cycle. In this article, we discuss structural and nonstructural viral proteins that are the focus of current target validation and drug discovery efforts. Both inhibition of essential enzymatic activities and disruption of necessary protein–protein interactions are considered. In addition, we address promising new targets for future research. As our molecular and biochemical understanding of the flavivirus life cycle increases, the number of targets for antiviral therapeutic discovery grows and the possibility for novel drug discovery continues to strengthen.

  17. Clinically Detectable Dental Identifiers Observed in Intra-oral Photographs and Extra-oral Radiographs, Validated for Human Identification Purposes.

    PubMed

    Angelakopoulos, Nikolaos; Franco, Ademir; Willems, Guy; Fieuws, Steffen; Thevissen, Patrick

    2017-07-01

    Screening the prevalence and pattern of dental identifiers contributes toward the process of human identification. This research investigated the uniqueness of clinical dental identifiers in photographs and radiographs. Panoramic and lateral cephalometric radiographs and five intra-oral photographs of 1727 subjects were used. In a target set, two observers examined different subjects. In a subset, both observers examined the same subjects (source set). The distance between source and target subjects was quantified for each identifier. The percentage of subjects in the target set being at least as close as the correct subject was assessed. The number of molars (34.6%), missing teeth (42%), and displaced teeth (59.9%) were the most unique identifiers in photographs and panoramic and lateral cephalometric radiographs, respectively. The pattern of rotated teeth (14.9%) was the most unique in photographs, while displaced teeth was in panoramic (37.6%) and lateral cephalometric (54.8%) radiographs. Morphological identifiers were the most unique, highlighting their importance for human identifications. © 2016 American Academy of Forensic Sciences.

  18. The accomplishments of lithium target and test facility validation activities in the IFMIF/EVEDA phase

    NASA Astrophysics Data System (ADS)

    Arbeiter, Frederik; Baluc, Nadine; Favuzza, Paolo; Gröschel, Friedrich; Heidinger, Roland; Ibarra, Angel; Knaster, Juan; Kanemura, Takuji; Kondo, Hiroo; Massaut, Vincent; Saverio Nitti, Francesco; Miccichè, Gioacchino; O'hira, Shigeru; Rapisarda, David; Sugimoto, Masayoshi; Wakai, Eiichi; Yokomine, Takehiko

    2018-01-01

    As part of the engineering validation and engineering design activities (EVEDA) phase for the international fusion materials irradiation facility IFMIF, major elements of a lithium target facility and the test facility were designed, prototyped and validated. For the lithium target facility, the EVEDA lithium test loop was built at JAEA and used to test the stability (waves and long term) of the lithium flow in the target, work out the startup procedures, and test lithium purification and analysis. It was confirmed by experiments in the Lifus 6 plant at ENEA that lithium corrosion on ferritic martensitic steels is acceptably low. Furthermore, complex remote handling procedures for the remote maintenance of the target in the test cell environment were successfully practiced. For the test facility, two variants of a high flux test module were prototyped and tested in helium loops, demonstrating their good capabilities of maintaining the material specimens at the desired temperature with a low temperature spread. Irradiation tests were performed for heated specimen capsules and irradiation instrumentation in the BR2 reactor at SCK-CEN. The small specimen test technique, essential for obtaining material test results with limited irradiation volume, was advanced by evaluating specimen shape and test technique influences.

  19. In silico Analysis of Toxins of Staphylococcus aureus for Validating Putative Drug Targets.

    PubMed

    Mohana, Ramadevi; Venugopal, Subhashree

    2017-01-01

    Toxins are one among the numerous virulence factors produced by the bacteria. These are powerful poisonous substances enabling the bacteria to encounter the defense mechanism of human body. The pathogenic system of Staphylococcus aureus is evolved with various exotoxins that cause detrimental effects on human immune system. Four toxins namely enterotoxin A, exfoliative toxin A, TSST-1 and γ-hemolysin were downloaded from Uniprot database and were analyzed to understand the nature of the toxins and for drug target validation. The results inferred that the toxins were found to interact with many protein partners and no homologous sequences for human proteome were found, and based on similarity search in Drugbank, the targets were identified as novel drug targets. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  20. Hazard identification and risk assessment for biologics targeting the immune system.

    PubMed

    Weir, Andrea B

    2008-01-01

    Biologic pharmaceuticals include a variety of products, such as monoclonal antibodies, fusion proteins and cytokines. Products in those classes include immunomodulatory biologics, which are intended to enhance or diminish the activity of the immune system. Immunomodulatory biologics have been approved by the U.S. FDA for a variety of indications, including cancer and inflammatory conditions. Prior to gaining approval for marketing, sponsoring companies for all types of products must demonstrate a product's safety in toxicology studies conducted in animals and show safety and efficacy in clinical trials conducted in patients. The overall goal of toxicology studies, which applies to immunomodulatory and other product types, is to identify the hazards that products pose to humans. Because biologics are generally highly selective for specific targets (receptors/epitopes), conducting toxicology studies in animal models with the target is essential. Such animals are referred to as pharmacologically relevant. Endpoints routinely included in toxicology studies, such as hematology, organ weight and histopathology, can be used to assess the effect of a product on the structure of the immune system. Additionally, specialized endpoints, such as immunophenotyping and immune function tests, can be used to define effects of immunomodulatory products on the immune system. Following hazard identification, risks posed to patients are assessed and managed. Risks can be managed through clinical trial design and risk communication, a practice that applies to immunomodulatory and other product types. Examples of risk management in clinical trial design include establishing a safe starting dose, defining the appropriate patient population and establishing appropriate patient monitoring. Risk communication starts during clinical trials and continues after product approval. A combination of hazard identification, risk assessment and risk management allows for drug development to proceed

  1. An integrated miRNA functional screening and target validation method for organ morphogenesis.

    PubMed

    Rebustini, Ivan T; Vlahos, Maryann; Packer, Trevor; Kukuruzinska, Maria A; Maas, Richard L

    2016-03-16

    The relative ease of identifying microRNAs and their increasing recognition as important regulators of organogenesis motivate the development of methods to efficiently assess microRNA function during organ morphogenesis. In this context, embryonic organ explants provide a reliable and reproducible system that recapitulates some of the important early morphogenetic processes during organ development. Here we present a method to target microRNA function in explanted mouse embryonic organs. Our method combines the use of peptide-based nanoparticles to transfect specific microRNA inhibitors or activators into embryonic organ explants, with a microRNA pulldown assay that allows direct identification of microRNA targets. This method provides effective assessment of microRNA function during organ morphogenesis, allows prioritization of multiple microRNAs in parallel for subsequent genetic approaches, and can be applied to a variety of embryonic organs.

  2. Pharmacologic Management of Duchenne Muscular Dystrophy: Target Identification and Preclinical Trials

    PubMed Central

    Kornegay, Joe N.; Spurney, Christopher F.; Nghiem, Peter P.; Brinkmeyer-Langford, Candice L.; Hoffman, Eric P.; Nagaraju, Kanneboyina

    2014-01-01

    Duchenne muscular dystrophy (DMD) is an X-linked human disorder in which absence of the protein dystrophin causes degeneration of skeletal and cardiac muscle. For the sake of treatment development, over and above definitive genetic and cell-based therapies, there is considerable interest in drugs that target downstream disease mechanisms. Drug candidates have typically been chosen based on the nature of pathologic lesions and presumed underlying mechanisms and then tested in animal models. Mammalian dystrophinopathies have been characterized in mice (mdx mouse) and dogs (golden retriever muscular dystrophy [GRMD]). Despite promising results in the mdx mouse, some therapies have not shown efficacy in DMD. Although the GRMD model offers a higher hurdle for translation, dogs have primarily been used to test genetic and cellular therapies where there is greater risk. Failed translation of animal studies to DMD raises questions about the propriety of methods and models used to identify drug targets and test efficacy of pharmacologic intervention. The mdx mouse and GRMD dog are genetically homologous to DMD but not necessarily analogous. Subcellular species differences are undoubtedly magnified at the whole-body level in clinical trials. This problem is compounded by disparate cultures in clinical trials and preclinical studies, pointing to a need for greater rigor and transparency in animal experiments. Molecular assays such as mRNA arrays and genome-wide association studies allow identification of genetic drug targets more closely tied to disease pathogenesis. Genes in which polymorphisms have been directly linked to DMD disease progression, as with osteopontin, are particularly attractive targets. PMID:24936034

  3. Identification of novel plant peroxisomal targeting signals by a combination of machine learning methods and in vivo subcellular targeting analyses.

    PubMed

    Lingner, Thomas; Kataya, Amr R; Antonicelli, Gerardo E; Benichou, Aline; Nilssen, Kjersti; Chen, Xiong-Yan; Siemsen, Tanja; Morgenstern, Burkhard; Meinicke, Peter; Reumann, Sigrun

    2011-04-01

    In the postgenomic era, accurate prediction tools are essential for identification of the proteomes of cell organelles. Prediction methods have been developed for peroxisome-targeted proteins in animals and fungi but are missing specifically for plants. For development of a predictor for plant proteins carrying peroxisome targeting signals type 1 (PTS1), we assembled more than 2500 homologous plant sequences, mainly from EST databases. We applied a discriminative machine learning approach to derive two different prediction methods, both of which showed high prediction accuracy and recognized specific targeting-enhancing patterns in the regions upstream of the PTS1 tripeptides. Upon application of these methods to the Arabidopsis thaliana genome, 392 gene models were predicted to be peroxisome targeted. These predictions were extensively tested in vivo, resulting in a high experimental verification rate of Arabidopsis proteins previously not known to be peroxisomal. The prediction methods were able to correctly infer novel PTS1 tripeptides, which even included novel residues. Twenty-three newly predicted PTS1 tripeptides were experimentally confirmed, and a high variability of the plant PTS1 motif was discovered. These prediction methods will be instrumental in identifying low-abundance and stress-inducible peroxisomal proteins and defining the entire peroxisomal proteome of Arabidopsis and agronomically important crop plants.

  4. Validation of the RAGE Hydrocode for Impacts into Volatile-Rich Targets

    NASA Astrophysics Data System (ADS)

    Plesko, C. S.; Asphaug, E.; Coker, R. F.; Wohletz, K. H.; Korycansky, D. G.; Gisler, G. R.

    2007-12-01

    In preparation for a detailed study of large-scale impacts into the Martian surface, we have validated the RAGE hydrocode (Gittings et al., in press, CSD) against a suite of experiments and statistical models. We present comparisons of hydrocode models to centimeter-scale gas gun impacts (Nakazawa et al. 2002), an underground nuclear test (Perret, 1971), and crater scaling laws (Holsapple 1993, O'Keefe and Ahrens 1993). We have also conducted model convergence and uncertainty analyses which will be presented. Results to date are encouraging for our current model goals, and indicate areas where the hydrocode may be extended in the future. This validation work is focused on questions related to the specific problem of large impacts into volatile-rich targets. The overall goal of this effort is to be able to realistically model large-scale Noachian, and possibly post- Noachian, impacts on Mars not so much to model the crater morphology as to understand the evolution of target volatiles in the post-impact regime, to explore how large craters might set the stage for post-impact hydro- geologic evolution both locally (in the crater subsurface) and globally, due to the redistribution of volatiles from the surface and subsurface into the atmosphere. This work is performed under the auspices of IGPP and the DOE at LANL under contracts W-7405-ENG-36 and DE-AC52-06NA25396. Effort by DK and EA is sponsored by NASA's Mars Fundamental Research Program.

  5. OCO-2 Observation and Validation Overview: Observations Data Modes and Target Observations, Taken During the First 15 Months of Operations

    NASA Astrophysics Data System (ADS)

    Osterman, G. B.; Fisher, B.; Wunch, D.; Eldering, A.; Wennberg, P. O.; Roehl, C. M.; Naylor, B. J.; Lee, R.; Pollock, R.; Gunson, M. R.

    2015-12-01

    The OCO-2 instrument was successfully launched on July 2, 2014 from Vandenberg Air Force Base in California. The instrument reached its observational orbit about three weeks later. The spacecraft is at the head of the A-train satellites and began collecting operational data on Sept 5, 2014. OCO-2 makes measurements in three modes: nadir, glint and target. Target observations are designed to provide large amounts of data in a small area near a ground validation site. The instruments of the Total Carbon Column Observing Network (TCCON) provide the ground validation data for the OCO-2 XCO2 observations and comparisons to TCCON form the basis of the OCO-2 validation plan. There are now 27 locations at which OCO-2 can perform target observations and CCON sites make up 23 of those possible target locations. For its first year in orbit, OCO-2 operated in nadir mode for 16 days and then in glint mode for 16 days. Each 16-day cycle spans 233 orbits. On July 1, 2015, OCO-2 changed to an observational mode of alternating nadir and glint measurements on an orbit-by-orbit basis. By December 2015, this operational mode may be modified such that orbits that measure only over ocean will always observed in glint mode. In this presentation we will provide information on the observations made by OCO-2 during its first 15 month in operations. We will show maps of the OCO-2 ground tracks and XCO2 data, calendars illustrating the observational schedule and statistics on the target observations taken. We will provide more information on what is involved in making target observations and how it affects the standard operational data acquisition patterns. Changes to the standard observational patterns of OCO-2 and to the list of locations for target observations will be discussed as well. We will provide an overview of some of the validation related analysis being done using nadir and glint mode OCO-2 data in addition to an overview on validation analyses that do not directly utilize TCCON

  6. Biophysics: for HTS hit validation, chemical lead optimization, and beyond.

    PubMed

    Genick, Christine C; Wright, S Kirk

    2017-09-01

    There are many challenges to the drug discovery process, including the complexity of the target, its interactions, and how these factors play a role in causing the disease. Traditionally, biophysics has been used for hit validation and chemical lead optimization. With its increased throughput and sensitivity, biophysics is now being applied earlier in this process to empower target characterization and hit finding. Areas covered: In this article, the authors provide an overview of how biophysics can be utilized to assess the quality of the reagents used in screening assays, to validate potential tool compounds, to test the integrity of screening assays, and to create follow-up strategies for compound characterization. They also briefly discuss the utilization of different biophysical methods in hit validation to help avoid the resource consuming pitfalls caused by the lack of hit overlap between biophysical methods. Expert opinion: The use of biophysics early on in the drug discovery process has proven crucial to identifying and characterizing targets of complex nature. It also has enabled the identification and classification of small molecules which interact in an allosteric or covalent manner with the target. By applying biophysics in this manner and at the early stages of this process, the chances of finding chemical leads with novel mechanisms of action are increased. In the future, focused screens with biophysics as a primary readout will become increasingly common.

  7. A single-laboratory validated method for the generation of DNA barcodes for the identification of fish for regulatory compliance.

    PubMed

    Handy, Sara M; Deeds, Jonathan R; Ivanova, Natalia V; Hebert, Paul D N; Hanner, Robert H; Ormos, Andrea; Weigt, Lee A; Moore, Michelle M; Yancy, Haile F

    2011-01-01

    The U.S. Food and Drug Administration is responsible for ensuring that the nation's food supply is safe and accurately labeled. This task is particularly challenging in the case of seafood where a large variety of species are marketed, most of this commodity is imported, and processed product is difficult to identify using traditional morphological methods. Reliable species identification is critical for both foodborne illness investigations and for prevention of deceptive practices, such as those where species are intentionally mislabeled to circumvent import restrictions or for resale as species of higher value. New methods that allow accurate and rapid species identifications are needed, but any new methods to be used for regulatory compliance must be both standardized and adequately validated. "DNA barcoding" is a process by which species discriminations are achieved through the use of short, standardized gene fragments. For animals, a fragment (655 base pairs starting near the 5' end) of the cytochrome c oxidase subunit 1 mitochondrial gene has been shown to provide reliable species level discrimination in most cases. We provide here a protocol with single-laboratory validation for the generation of DNA barcodes suitable for the identification of seafood products, specifically fish, in a manner that is suitable for FDA regulatory use.

  8. Targeted sequencing of clade-specific markers from skin microbiomes for forensic human identification.

    PubMed

    Schmedes, Sarah E; Woerner, August E; Novroski, Nicole M M; Wendt, Frank R; King, Jonathan L; Stephens, Kathryn M; Budowle, Bruce

    2018-01-01

    The human skin microbiome is comprised of diverse communities of bacterial, eukaryotic, and viral taxa and contributes millions of additional genes to the repertoire of human genes, affecting human metabolism and immune response. Numerous genetic and environmental factors influence the microbiome composition and as such contribute to individual-specific microbial signatures which may be exploited for forensic applications. Previous studies have demonstrated the potential to associate skin microbial profiles collected from touched items to their individual owner, mainly using unsupervised methods from samples collected over short time intervals. Those studies utilize either targeted 16S rRNA or shotgun metagenomic sequencing to characterize skin microbiomes; however, these approaches have limited species and strain resolution and susceptibility to stochastic effects, respectively. Clade-specific markers from the skin microbiome, using supervised learning, can predict individual identity using skin microbiomes from their respective donors with high accuracy. In this study the hidSkinPlex is presented, a novel targeted sequencing method using skin microbiome markers developed for human identification. The hidSkinPlex (comprised of 286 bacterial (and phage) family-, genus-, species-, and subspecies-level markers), initially was evaluated on three bacterial control samples represented in the panel (i.e., Propionibacterium acnes, Propionibacterium granulosum, and Rothia dentocariosa) to assess the performance of the multiplex. The hidSkinPlex was further evaluated for prediction purposes. The hidSkinPlex markers were used to attribute skin microbiomes collected from eight individuals from three body sites (i.e., foot (Fb), hand (Hp) and manubrium (Mb)) to their host donor. Supervised learning, specifically regularized multinomial logistic regression and 1-nearest-neighbor classification were used to classify skin microbiomes to their hosts with up to 92% (Fb), 96% (Mb

  9. Identification of inhibitors for putative malaria drug targets amongst novel antimalarial compounds

    PubMed Central

    Crowther, Gregory J.; Napuli, Alberto J.; Gilligan, James H.; Gagaring, Kerstin; Borboa, Rachel; Francek, Carolyn; Chen, Zhong; Dagostino, Eleanor F.; Stockmyer, Justin B.; Wang, Yu; Rodenbough, Philip P.; Castaneda, Lisa J.; Leibly, David J.; Bhandari, Janhavi; Gelb, Michael H.; Brinker, Achim; Engels, Ingo; Taylor, Jennifer; Chatterjee, Arnab K.; Fantauzzi, Pascal; Glynne, Richard J.; Van Voorhis, Wesley C.; Kuhen, Kelli L.

    2011-01-01

    The efficacy of most marketed antimalarial drugs has been compromised by evolution of parasite resistance, underscoring an urgent need to find new drugs with new mechanisms of action. We have taken a high-throughput approach toward identifying novel antimalarial chemical inhibitors of prioritized drug targets for P. falciparum, excluding targets which are inhibited by currently used drugs. A screen of commercially available libraries identified 5,655 low molecular weight compounds that inhibit growth of P. falciparum cultures with EC50 values below 1.25 μM. These compounds were then tested in 384- or 1536-well biochemical assays for activity against nine Plasmodium enzymes: adenylosuccinate synthetase (AdSS), choline kinase (CK), deoxyuridine triphosphate nucleotidohydrolase (dUTPase), glutamate dehydrogenase (GDH), guanylate kinase (GK), N-myristoyltransferase (NMT), orotidine 5′-monophosphate decarboxylase (OMPDC), farnesyl pyrophosphate synthase (FPPS) and S-adenosylhomocysteine hydrolase (SAHH). These enzymes were selected using TDRtargets.org, and are believed to have excellent potential as drug targets based on criteria such as their likely essentiality, druggability, and amenability to high-throughput biochemical screening. Six of these targets were inhibited by one or more of the antimalarial scaffolds and may have potential use in drug development, further target validation studies and exploration of P. falciparum biochemistry and biology. PMID:20813141

  10. Identification of inhibitors for putative malaria drug targets among novel antimalarial compounds.

    PubMed

    Crowther, Gregory J; Napuli, Alberto J; Gilligan, James H; Gagaring, Kerstin; Borboa, Rachel; Francek, Carolyn; Chen, Zhong; Dagostino, Eleanor F; Stockmyer, Justin B; Wang, Yu; Rodenbough, Philip P; Castaneda, Lisa J; Leibly, David J; Bhandari, Janhavi; Gelb, Michael H; Brinker, Achim; Engels, Ingo H; Taylor, Jennifer; Chatterjee, Arnab K; Fantauzzi, Pascal; Glynne, Richard J; Van Voorhis, Wesley C; Kuhen, Kelli L

    2011-01-01

    The efficacy of most marketed antimalarial drugs has been compromised by evolution of parasite resistance, underscoring an urgent need to find new drugs with new mechanisms of action. We have taken a high-throughput approach toward identifying novel antimalarial chemical inhibitors of prioritized drug targets for Plasmodium falciparum, excluding targets which are inhibited by currently used drugs. A screen of commercially available libraries identified 5655 low molecular weight compounds that inhibit growth of P. falciparum cultures with EC(50) values below 1.25μM. These compounds were then tested in 384- or 1536-well biochemical assays for activity against nine Plasmodium enzymes: adenylosuccinate synthetase (AdSS), choline kinase (CK), deoxyuridine triphosphate nucleotidohydrolase (dUTPase), glutamate dehydrogenase (GDH), guanylate kinase (GK), N-myristoyltransferase (NMT), orotidine 5'-monophosphate decarboxylase (OMPDC), farnesyl pyrophosphate synthase (FPPS) and S-adenosylhomocysteine hydrolase (SAHH). These enzymes were selected using TDRtargets.org, and are believed to have excellent potential as drug targets based on criteria such as their likely essentiality, druggability, and amenability to high-throughput biochemical screening. Six of these targets were inhibited by one or more of the antimalarial scaffolds and may have potential use in drug development, further target validation studies and exploration of P. falciparum biochemistry and biology. Copyright © 2010 Elsevier B.V. All rights reserved.

  11. Bioinformatic identification and expression analysis of banana microRNAs and their targets.

    PubMed

    Chai, Juan; Feng, Renjun; Shi, Hourui; Ren, Mengyun; Zhang, Yindong; Wang, Jingyi

    2015-01-01

    MicroRNAs (miRNAs) represent a class of endogenous non-coding small RNAs that play important roles in multiple biological processes by degrading targeted mRNAs or repressing mRNA translation. Thousands of miRNAs have been identified in many plant species, whereas only a limited number of miRNAs have been predicted in M. acuminata (A genome) and M. balbisiana (B genome). Here, previously known plant miRNAs were BLASTed against the Expressed Sequence Tag (EST) and Genomic Survey Sequence (GSS), a database of banana genes. A total of 32 potential miRNAs belonging to 13 miRNAs families were detected using a range of filtering criteria. 244 miRNA:target pairs were subsequently predicted, most of which encode transcription factors or enzymes that participate in the regulation of development, growth, metabolism, and other physiological processes. In order to validate the predicted miRNAs and the mutual relationship between miRNAs and their target genes, qRT-PCR was applied to detect the tissue-specific expression levels of 12 putative miRNAs and 6 target genes in roots, leaves, flowers, and fruits. This study provides some important information about banana pre-miRNAs, mature miRNAs, and miRNA target genes and these findings can be applied to future research of miRNA functions.

  12. Neuroinflammatory targets and treatments for epilepsy validated in experimental models.

    PubMed

    Aronica, Eleonora; Bauer, Sebastian; Bozzi, Yuri; Caleo, Matteo; Dingledine, Raymond; Gorter, Jan A; Henshall, David C; Kaufer, Daniela; Koh, Sookyong; Löscher, Wolfgang; Louboutin, Jean-Pierre; Mishto, Michele; Norwood, Braxton A; Palma, Eleonora; Poulter, Michael O; Terrone, Gaetano; Vezzani, Annamaria; Kaminski, Rafal M

    2017-07-01

    A large body of evidence that has accumulated over the past decade strongly supports the role of inflammation in the pathophysiology of human epilepsy. Specific inflammatory molecules and pathways have been identified that influence various pathologic outcomes in different experimental models of epilepsy. Most importantly, the same inflammatory pathways have also been found in surgically resected brain tissue from patients with treatment-resistant epilepsy. New antiseizure therapies may be derived from these novel potential targets. An essential and crucial question is whether targeting these molecules and pathways may result in anti-ictogenesis, antiepileptogenesis, and/or disease-modification effects. Therefore, preclinical testing in models mimicking relevant aspects of epileptogenesis is needed to guide integrated experimental and clinical trial designs. We discuss the most recent preclinical proof-of-concept studies validating a number of therapeutic approaches against inflammatory mechanisms in animal models that could represent novel avenues for drug development in epilepsy. Finally, we suggest future directions to accelerate preclinical to clinical translation of these recent discoveries. Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.

  13. In Situ Identification of Cyanobacteria with Horseradish Peroxidase-Labeled, rRNA-Targeted Oligonucleotide Probes

    PubMed Central

    Schönhuber, Wilhelm; Zarda, Boris; Eix, Stella; Rippka, Rosmarie; Herdman, Michael; Ludwig, Wolfgang; Amann, Rudolf

    1999-01-01

    Individual cyanobacterial cells are normally identified in environmental samples only on the basis of their pigmentation and morphology. However, these criteria are often insufficient for the differentiation of species. Here, a whole-cell hybridization technique is presented that uses horseradish peroxidase (HRP)-labeled, rRNA-targeted oligonucleotides for in situ identification of cyanobacteria. This indirect method, in which the probe-conferred enzyme has to be visualized in an additional step, was necessary since fluorescently monolabeled oligonucleotides were insufficient to overstain the autofluorescence of the target cells. Initially, a nonfluorescent detection assay was developed and successfully applied to cyanobacterial mats. Later, it was demonstrated that tyramide signal amplification (TSA) resulted in fluorescent signals far above the level of autofluorescence. Furthermore, TSA-based detection of HRP was more sensitive than that based on nonfluorescent substrates. Critical points of the assay, such as cell fixation and permeabilization, specificity, and sensitivity, were systematically investigated by using four oligonucleotides newly designed to target groups of cyanobacteria. PMID:10049892

  14. In Silico Prediction and Validation of Gfap as an miR-3099 Target in Mouse Brain.

    PubMed

    Abidin, Shahidee Zainal; Leong, Jia-Wen; Mahmoudi, Marzieh; Nordin, Norshariza; Abdullah, Syahril; Cheah, Pike-See; Ling, King-Hwa

    2017-08-01

    MicroRNAs are small non-coding RNAs that play crucial roles in the regulation of gene expression and protein synthesis during brain development. MiR-3099 is highly expressed throughout embryogenesis, especially in the developing central nervous system. Moreover, miR-3099 is also expressed at a higher level in differentiating neurons in vitro, suggesting that it is a potential regulator during neuronal cell development. This study aimed to predict the target genes of miR-3099 via in-silico analysis using four independent prediction algorithms (miRDB, miRanda, TargetScan, and DIANA-micro-T-CDS) with emphasis on target genes related to brain development and function. Based on the analysis, a total of 3,174 miR-3099 target genes were predicted. Those predicted by at least three algorithms (324 genes) were subjected to DAVID bioinformatics analysis to understand their overall functional themes and representation. The analysis revealed that nearly 70% of the target genes were expressed in the nervous system and a significant proportion were associated with transcriptional regulation and protein ubiquitination mechanisms. Comparison of in situ hybridization (ISH) expression patterns of miR-3099 in both published and in-house-generated ISH sections with the ISH sections of target genes from the Allen Brain Atlas identified 7 target genes (Dnmt3a, Gabpa, Gfap, Itga4, Lxn, Smad7, and Tbx18) having expression patterns complementary to miR-3099 in the developing and adult mouse brain samples. Of these, we validated Gfap as a direct downstream target of miR-3099 using the luciferase reporter gene system. In conclusion, we report the successful prediction and validation of Gfap as an miR-3099 target gene using a combination of bioinformatics resources with enrichment of annotations based on functional ontologies and a spatio-temporal expression dataset.

  15. Validation of automatic landmark identification for atlas-based segmentation for radiation treatment planning of the head-and-neck region

    NASA Astrophysics Data System (ADS)

    Leavens, Claudia; Vik, Torbjørn; Schulz, Heinrich; Allaire, Stéphane; Kim, John; Dawson, Laura; O'Sullivan, Brian; Breen, Stephen; Jaffray, David; Pekar, Vladimir

    2008-03-01

    Manual contouring of target volumes and organs at risk in radiation therapy is extremely time-consuming, in particular for treating the head-and-neck area, where a single patient treatment plan can take several hours to contour. As radiation treatment delivery moves towards adaptive treatment, the need for more efficient segmentation techniques will increase. We are developing a method for automatic model-based segmentation of the head and neck. This process can be broken down into three main steps: i) automatic landmark identification in the image dataset of interest, ii) automatic landmark-based initialization of deformable surface models to the patient image dataset, and iii) adaptation of the deformable models to the patient-specific anatomical boundaries of interest. In this paper, we focus on the validation of the first step of this method, quantifying the results of our automatic landmark identification method. We use an image atlas formed by applying thin-plate spline (TPS) interpolation to ten atlas datasets, using 27 manually identified landmarks in each atlas/training dataset. The principal variation modes returned by principal component analysis (PCA) of the landmark positions were used by an automatic registration algorithm, which sought the corresponding landmarks in the clinical dataset of interest using a controlled random search algorithm. Applying a run time of 60 seconds to the random search, a root mean square (rms) distance to the ground-truth landmark position of 9.5 +/- 0.6 mm was calculated for the identified landmarks. Automatic segmentation of the brain, mandible and brain stem, using the detected landmarks, is demonstrated.

  16. Identification of tissue-specific targeting peptide

    NASA Astrophysics Data System (ADS)

    Jung, Eunkyoung; Lee, Nam Kyung; Kang, Sang-Kee; Choi, Seung-Hoon; Kim, Daejin; Park, Kisoo; Choi, Kihang; Choi, Yun-Jaie; Jung, Dong Hyun

    2012-11-01

    Using phage display technique, we identified tissue-targeting peptide sets that recognize specific tissues (bone-marrow dendritic cell, kidney, liver, lung, spleen and visceral adipose tissue). In order to rapidly evaluate tissue-specific targeting peptides, we performed machine learning studies for predicting the tissue-specific targeting activity of peptides on the basis of peptide sequence information using four machine learning models and isolated the groups of peptides capable of mediating selective targeting to specific tissues. As a representative liver-specific targeting sequence, the peptide "DKNLQLH" was selected by the sequence similarity analysis. This peptide has a high degree of homology with protein ligands which can interact with corresponding membrane counterparts. We anticipate that our models will be applicable to the prediction of tissue-specific targeting peptides which can recognize the endothelial markers of target tissues.

  17. UXO detection and identification based on intrinsic target polarizabilities: A case history

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Gasperikova, E.; Smith, J.T.; Morrison, H.F.

    2008-07-15

    Electromagnetic induction data parameterized in time dependent object intrinsic polarizabilities allow discrimination of unexploded ordnance (UXO) from false targets (scrap metal). Data from a cart-mounted system designed for discrimination of UXO with 20 mm to 155 mm diameters are used. Discrimination of UXO from irregular scrap metal is based on the principal dipole polarizabilities of a target. A near-intact UXO displays a single major polarizability coincident with the long axis of the object and two equal smaller transverse polarizabilities, whereas metal scraps have distinct polarizability signatures that rarely mimic those of elongated symmetric bodies. Based on a training data setmore » of known targets, object identification was made by estimating the probability that an object is a single UXO. Our test survey took place on a military base where both 4.2-inch mortar shells and scrap metal were present. The results show that we detected and discriminated correctly all 4.2-inch mortars, and in that process we added 7%, and 17%, respectively, of dry holes (digging scrap) to the total number of excavations in two different survey modes. We also demonstrated a mode of operation that might be more cost effective than the current practice.« less

  18. Vitamin D Pathway Status and the Identification of Target Genes in the Mouse Mammary Gland

    DTIC Science & Technology

    2013-01-01

    12 Palmer HG et al. The vitamin D receptor is a Wnt effector that controls hair follicle differentiation and specifies tumor type in adult epidermis...AD_________________ Award Number: W81XWH-11-1-0152 TITLE: Vitamin D pathway status and the...December 2012 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER W81XWH-11-1-0152 Vitamin D pathway status and the identification of target genes in the

  19. Drug Target Protein-Protein Interaction Networks: A Systematic Perspective

    PubMed Central

    2017-01-01

    The identification and validation of drug targets are crucial in biomedical research and many studies have been conducted on analyzing drug target features for getting a better understanding on principles of their mechanisms. But most of them are based on either strong biological hypotheses or the chemical and physical properties of those targets separately. In this paper, we investigated three main ways to understand the functional biomolecules based on the topological features of drug targets. There are no significant differences between targets and common proteins in the protein-protein interactions network, indicating the drug targets are neither hub proteins which are dominant nor the bridge proteins. According to some special topological structures of the drug targets, there are significant differences between known targets and other proteins. Furthermore, the drug targets mainly belong to three typical communities based on their modularity. These topological features are helpful to understand how the drug targets work in the PPI network. Particularly, it is an alternative way to predict potential targets or extract nontargets to test a new drug target efficiently and economically. By this way, a drug target's homologue set containing 102 potential target proteins is predicted in the paper. PMID:28691014

  20. Gene expression complex networks: synthesis, identification, and analysis.

    PubMed

    Lopes, Fabrício M; Cesar, Roberto M; Costa, Luciano Da F

    2011-10-01

    Thanks to recent advances in molecular biology, allied to an ever increasing amount of experimental data, the functional state of thousands of genes can now be extracted simultaneously by using methods such as cDNA microarrays and RNA-Seq. Particularly important related investigations are the modeling and identification of gene regulatory networks from expression data sets. Such a knowledge is fundamental for many applications, such as disease treatment, therapeutic intervention strategies and drugs design, as well as for planning high-throughput new experiments. Methods have been developed for gene networks modeling and identification from expression profiles. However, an important open problem regards how to validate such approaches and its results. This work presents an objective approach for validation of gene network modeling and identification which comprises the following three main aspects: (1) Artificial Gene Networks (AGNs) model generation through theoretical models of complex networks, which is used to simulate temporal expression data; (2) a computational method for gene network identification from the simulated data, which is founded on a feature selection approach where a target gene is fixed and the expression profile is observed for all other genes in order to identify a relevant subset of predictors; and (3) validation of the identified AGN-based network through comparison with the original network. The proposed framework allows several types of AGNs to be generated and used in order to simulate temporal expression data. The results of the network identification method can then be compared to the original network in order to estimate its properties and accuracy. Some of the most important theoretical models of complex networks have been assessed: the uniformly-random Erdös-Rényi (ER), the small-world Watts-Strogatz (WS), the scale-free Barabási-Albert (BA), and geographical networks (GG). The experimental results indicate that the inference

  1. STARR: shortwave-targeted agile Raman robot for the detection and identification of emplaced explosives

    NASA Astrophysics Data System (ADS)

    Gomer, Nathaniel R.; Gardner, Charles W.

    2014-05-01

    In order to combat the threat of emplaced explosives (land mines, etc.), ChemImage Sensor Systems (CISS) has developed a multi-sensor, robot mounted sensor capable of identification and confirmation of potential threats. The system, known as STARR (Shortwave-infrared Targeted Agile Raman Robot), utilizes shortwave infrared spectroscopy for the identification of potential threats, combined with a visible short-range standoff Raman hyperspectral imaging (HSI) system for material confirmation. The entire system is mounted onto a Talon UGV (Unmanned Ground Vehicle), giving the sensor an increased area search rate and reducing the risk of injury to the operator. The Raman HSI system utilizes a fiber array spectral translator (FAST) for the acquisition of high quality Raman chemical images, allowing for increased sensitivity and improved specificity. An overview of the design and operation of the system will be presented, along with initial detection results of the fusion sensor.

  2. A novel algorithm for finding optimal driver nodes to target control complex networks and its applications for drug targets identification.

    PubMed

    Guo, Wei-Feng; Zhang, Shao-Wu; Shi, Qian-Qian; Zhang, Cheng-Ming; Zeng, Tao; Chen, Luonan

    2018-01-19

    The advances in target control of complex networks not only can offer new insights into the general control dynamics of complex systems, but also be useful for the practical application in systems biology, such as discovering new therapeutic targets for disease intervention. In many cases, e.g. drug target identification in biological networks, we usually require a target control on a subset of nodes (i.e., disease-associated genes) with minimum cost, and we further expect that more driver nodes consistent with a certain well-selected network nodes (i.e., prior-known drug-target genes). Therefore, motivated by this fact, we pose and address a new and practical problem called as target control problem with objectives-guided optimization (TCO): how could we control the interested variables (or targets) of a system with the optional driver nodes by minimizing the total quantity of drivers and meantime maximizing the quantity of constrained nodes among those drivers. Here, we design an efficient algorithm (TCOA) to find the optional driver nodes for controlling targets in complex networks. We apply our TCOA to several real-world networks, and the results support that our TCOA can identify more precise driver nodes than the existing control-fucus approaches. Furthermore, we have applied TCOA to two bimolecular expert-curate networks. Source code for our TCOA is freely available from http://sysbio.sibcb.ac.cn/cb/chenlab/software.htm or https://github.com/WilfongGuo/guoweifeng . In the previous theoretical research for the full control, there exists an observation and conclusion that the driver nodes tend to be low-degree nodes. However, for target control the biological networks, we find interestingly that the driver nodes tend to be high-degree nodes, which is more consistent with the biological experimental observations. Furthermore, our results supply the novel insights into how we can efficiently target control a complex system, and especially many evidences on the

  3. Large-scale identification of target proteins of a glycosyltransferase isozyme by Lectin-IGOT-LC/MS, an LC/MS-based glycoproteomic approach

    PubMed Central

    Sugahara, Daisuke; Kaji, Hiroyuki; Sugihara, Kazushi; Asano, Masahide; Narimatsu, Hisashi

    2012-01-01

    Model organisms containing deletion or mutation in a glycosyltransferase-gene exhibit various physiological abnormalities, suggesting that specific glycan motifs on certain proteins play important roles in vivo. Identification of the target proteins of glycosyltransferase isozymes is the key to understand the roles of glycans. Here, we demonstrated the proteome-scale identification of the target proteins specific for a glycosyltransferase isozyme, β1,4-galactosyltransferase-I (β4GalT-I). Although β4GalT-I is the most characterized glycosyltransferase, its distinctive contribution to β1,4-galactosylation has been hardly described so far. We identified a large number of candidates for the target proteins specific to β4GalT-I by comparative analysis of β4GalT-I-deleted and wild-type mice using the LC/MS-based technique with the isotope-coded glycosylation site-specific tagging (IGOT) of lectin-captured N-glycopeptides. Our approach to identify the target proteins in a proteome-scale offers common features and trends in the target proteins, which facilitate understanding of the mechanism that controls assembly of a particular glycan motif on specific proteins. PMID:23002422

  4. Validation approach for a fast and simple targeted screening method for 75 antibiotics in meat and aquaculture products using LC-MS/MS.

    PubMed

    Dubreil, Estelle; Gautier, Sophie; Fourmond, Marie-Pierre; Bessiral, Mélaine; Gaugain, Murielle; Verdon, Eric; Pessel, Dominique

    2017-04-01

    An approach is described to validate a fast and simple targeted screening method for antibiotic analysis in meat and aquaculture products by LC-MS/MS. The strategy of validation was applied for a panel of 75 antibiotics belonging to different families, i.e., penicillins, cephalosporins, sulfonamides, macrolides, quinolones and phenicols. The samples were extracted once with acetonitrile, concentrated by evaporation and injected into the LC-MS/MS system. The approach chosen for the validation was based on the Community Reference Laboratory (CRL) guidelines for the validation of screening qualitative methods. The aim of the validation was to prove sufficient sensitivity of the method to detect all the targeted antibiotics at the level of interest, generally the maximum residue limit (MRL). A robustness study was also performed to test the influence of different factors. The validation showed that the method is valid to detect and identify 73 antibiotics of the 75 antibiotics studied in meat and aquaculture products at the validation levels.

  5. Validation of the proteasome as a therapeutic target in Plasmodium using an epoxyketone inhibitor with parasite-specific toxicity

    PubMed Central

    Li, Hao; Ponder, Elizabeth L.; Verdoes, Martijn; Asbjornsdottir, Kristijana H.; Deu, Edgar; Edgington, Laura E.; Lee, Jeong Tae; Kirk, Christopher J.; Demo, Susan D.; Williamson, Kim C.; Bogyo, Matthew

    2012-01-01

    Summary The Plasmodium proteasome has been suggested to be a potential anti-malarial drug target, however toxicity of inhibitors has prevented validation of this enzyme in vivo. We report here a screen of a library of 670 analogs of the recently FDA approved inhibitor, carfilzomib, to identify compounds that selectively kill parasites. We identified one compound, PR3, that has significant parasite killing activity in vitro but dramatically reduced toxicity in host cells. We found that this parasite-specific toxicity is not due to selective targeting of the Plasmodium proteasome over the host proteasome, but instead is due to a lack of activity against one of the human proteasome subunits. Subsequently, we used PR3 to significantly reduce parasite load in P. berghei infected mice without host toxicity, thus validating the proteasome as a viable anti-malarial drug target. PMID:23142757

  6. Bioinformatic Identification and Expression Analysis of Banana MicroRNAs and Their Targets

    PubMed Central

    Shi, Hourui; Ren, Mengyun; Zhang, Yindong; Wang, Jingyi

    2015-01-01

    MicroRNAs (miRNAs) represent a class of endogenous non-coding small RNAs that play important roles in multiple biological processes by degrading targeted mRNAs or repressing mRNA translation. Thousands of miRNAs have been identified in many plant species, whereas only a limited number of miRNAs have been predicted in M. acuminata (A genome) and M. balbisiana (B genome). Here, previously known plant miRNAs were BLASTed against the Expressed Sequence Tag (EST) and Genomic Survey Sequence (GSS), a database of banana genes. A total of 32 potential miRNAs belonging to 13 miRNAs families were detected using a range of filtering criteria. 244 miRNA:target pairs were subsequently predicted, most of which encode transcription factors or enzymes that participate in the regulation of development, growth, metabolism, and other physiological processes. In order to validate the predicted miRNAs and the mutual relationship between miRNAs and their target genes, qRT-PCR was applied to detect the tissue-specific expression levels of 12 putative miRNAs and 6 target genes in roots, leaves, flowers, and fruits. This study provides some important information about banana pre-miRNAs, mature miRNAs, and miRNA target genes and these findings can be applied to future research of miRNA functions. PMID:25856313

  7. Activity-based proteome profiling of potential cellular targets of Orlistat--an FDA-approved drug with anti-tumor activities.

    PubMed

    Yang, Peng-Yu; Liu, Kai; Ngai, Mun Hong; Lear, Martin J; Wenk, Markus R; Yao, Shao Q

    2010-01-20

    Orlistat, or tetrahydrolipstatin (THL), is an FDA-approved antiobesity drug with potential antitumor activities. Cellular off-targets and potential side effects of Orlistat in cancer therapies, however, have not been extensively explored thus far. In this study, we report the total of synthesis of THL-like protein-reactive probes, in which extremely conservative modifications (i.e., an alkyne handle) were introduced in the parental THL structure to maintain the native biological properties of Orlistat, while providing the necessary functionality for target identification via the bio-orthogonal click chemistry. With these natural productlike, cell-permeable probes, we were able to demonstrate, for the first time, this chemical proteomic approach is suitable for the identification of previously unknown cellular targets of Orlistat. In addition to the expected fatty acid synthase (FAS), we identified a total of eight new targets, some of which were further validated by experiments including Western blotting, recombinant protein expression, and site-directed mutagenesis. Our findings have important implications in the consideration of Orlistat as a potential anticancer drug at its early stages of development for cancer therapy. Our strategy should be broadly useful for off-target identification against quite a number of existing drugs and/or candidates, which are also covalent modifiers of their biological targets.

  8. Tools for in silico target fishing.

    PubMed

    Cereto-Massagué, Adrià; Ojeda, María José; Valls, Cristina; Mulero, Miquel; Pujadas, Gerard; Garcia-Vallve, Santiago

    2015-01-01

    Computational target fishing methods are designed to identify the most probable target of a query molecule. This process may allow the prediction of the bioactivity of a compound, the identification of the mode of action of known drugs, the detection of drug polypharmacology, drug repositioning or the prediction of the adverse effects of a compound. The large amount of information regarding the bioactivity of thousands of small molecules now allows the development of these types of methods. In recent years, we have witnessed the emergence of many methods for in silico target fishing. Most of these methods are based on the similarity principle, i.e., that similar molecules might bind to the same targets and have similar bioactivities. However, the difficult validation of target fishing methods hinders comparisons of the performance of each method. In this review, we describe the different methods developed for target prediction, the bioactivity databases most frequently used by these methods, and the publicly available programs and servers that enable non-specialist users to obtain these types of predictions. It is expected that target prediction will have a large impact on drug development and on the functional food industry. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Global Identification of MicroRNAs and Their Targets in Barley under Salinity Stress

    PubMed Central

    Cui, Licao; Feng, Kewei; Liu, Fuyan; Du, Xianghong; Tong, Wei; Nie, Xiaojun; Ji, Wanquan; Weining, Song

    2015-01-01

    Salinity is a major limiting factor for agricultural production worldwide. A better understanding of the mechanisms of salinity stress response will aid efforts to improve plant salt tolerance. In this study, a combination of small RNA and mRNA degradome sequencing was used to identify salinity responsive-miRNAs and their targets in barley. A total of 152 miRNAs belonging to 126 families were identified, of which 44 were found to be salinity responsive with 30 up-regulated and 25 down-regulated respectively. The majority of the salinity-responsive miRNAs were up-regulated at the 8h time point, while down-regulated at the 3h and 27h time points. The targets of these miRNAs were further detected by degradome sequencing coupled with bioinformatics prediction. Finally, qRT-PCR was used to validate the identified miRNA and their targets. Our study systematically investigated the expression profile of miRNA and their targets in barley during salinity stress phase, which can contribute to understanding how miRNAs respond to salinity stress in barley and other cereal crops. PMID:26372557

  10. Identification of KasA as the cellular target of an anti-tubercular scaffold

    PubMed Central

    Abrahams, Katherine A.; Chung, Chun-wa; Ghidelli-Disse, Sonja; Rullas, Joaquín; Rebollo-López, María José; Gurcha, Sudagar S.; Cox, Jonathan A. G.; Mendoza, Alfonso; Jiménez-Navarro, Elena; Martínez-Martínez, María Santos; Neu, Margarete; Shillings, Anthony; Homes, Paul; Argyrou, Argyrides; Casanueva, Ruth; Loman, Nicholas J.; Moynihan, Patrick J.; Lelièvre, Joël; Selenski, Carolyn; Axtman, Matthew; Kremer, Laurent; Bantscheff, Marcus; Angulo-Barturen, Iñigo; Izquierdo, Mónica Cacho; Cammack, Nicholas C.; Drewes, Gerard; Ballell, Lluis; Barros, David; Besra, Gurdyal S.; Bates, Robert H.

    2016-01-01

    Phenotypic screens for bactericidal compounds are starting to yield promising hits against tuberculosis. In this regard, whole-genome sequencing of spontaneous resistant mutants generated against an indazole sulfonamide (GSK3011724A) identifies several specific single-nucleotide polymorphisms in the essential Mycobacterium tuberculosis β-ketoacyl synthase (kas) A gene. Here, this genomic-based target assignment is confirmed by biochemical assays, chemical proteomics and structural resolution of a KasA-GSK3011724A complex by X-ray crystallography. Finally, M. tuberculosis GSK3011724A-resistant mutants increase the in vitro minimum inhibitory concentration and the in vivo 99% effective dose in mice, establishing in vitro and in vivo target engagement. Surprisingly, the lack of target engagement of the related β-ketoacyl synthases (FabH and KasB) suggests a different mode of inhibition when compared with other Kas inhibitors of fatty acid biosynthesis in bacteria. These results clearly identify KasA as the biological target of GSK3011724A and validate this enzyme for further drug discovery efforts against tuberculosis. PMID:27581223

  11. Human target acquisition performance

    NASA Astrophysics Data System (ADS)

    Teaney, Brian P.; Du Bosq, Todd W.; Reynolds, Joseph P.; Thompson, Roger; Aghera, Sameer; Moyer, Steven K.; Flug, Eric; Espinola, Richard; Hixson, Jonathan

    2012-06-01

    The battlefield has shifted from armored vehicles to armed insurgents. Target acquisition (identification, recognition, and detection) range performance involving humans as targets is vital for modern warfare. The acquisition and neutralization of armed insurgents while at the same time minimizing fratricide and civilian casualties is a mounting concern. U.S. Army RDECOM CERDEC NVESD has conducted many experiments involving human targets for infrared and reflective band sensors. The target sets include human activities, hand-held objects, uniforms & armament, and other tactically relevant targets. This paper will define a set of standard task difficulty values for identification and recognition associated with human target acquisition performance.

  12. Modeling and performance of HF/OTH (High-Frequency/Over-the-Horizon) radar target identification systems

    NASA Astrophysics Data System (ADS)

    Strausberger, Donald J.

    Several Radar Target Identification (RTI) techniques have been developed at The Ohio State University in recent years. Using the ElectroScience Laboratory compact range a large database of coherent RCS measurement has been constructed for several types of targets (aircraft, ships, and ground vehicles) at a variety of polarizations, aspect angles, and frequency bands. This extensive database has been used to analyze the performance of several different classification algorithms through the use of computer simulations. In order to optimize classification performance, it was concluded that the radar frequency range should lie in the Rayleigh-resonance frequency range, where the wavelength is on the order of or larger than the target size. For aircraft and ships with general dimensions on the order of 10 meters to 100 meters it is apparent that the High Frequency (HF) band provides optimal classification performance. Since existing HF radars are currently being used for detection and tracking or aircraft and ships of these dimensions, it is natural to further investigate the possibility of using these existing radars as the measurement devices in a radar target classification system.

  13. Identification of downstream metastasis-associated target genes regulated by LSD1 in colon cancer cells.

    PubMed

    Chen, Jiang; Ding, Jie; Wang, Ziwei; Zhu, Jian; Wang, Xuejian; Du, Jiyi

    2017-03-21

    This study aims to identify downstream target genes regulated by lysine-specific demethylase 1 (LSD1) in colon cancer cells and investigate the molecular mechanisms of LSD1 influencing invasion and metastasis of colon cancer. We obtained the expression changes of downstream target genes regulated by small-interfering RNA-LSD1 and LSD1-overexpression via gene expression profiling in two human colon cancer cell lines. An Affymetrix Human Transcriptome Array 2.0 was used to identify differentially expressed genes (DEGs). We screened out LSD1-target gene associated with proliferation, metastasis, and invasion from DEGs via Gene Ontology and Pathway Studio. Subsequently, four key genes (CABYR, FOXF2, TLE4, and CDH1) were computationally predicted as metastasis-related LSD1-target genes. ChIp-PCR was applied after RT-PCR and Western blot validations to detect the occupancy of LSD1-target gene promoter-bound LSD1. A total of 3633 DEGs were significantly upregulated, and 4642 DEGs were downregulated in LSD1-silenced SW620 cells. A total of 4047 DEGs and 4240 DEGs were upregulated and downregulated in LSD1-overexpressed HT-29 cells, respectively. RT-PCR and Western blot validated the microarray analysis results. ChIP assay results demonstrated that LSD1 might be negative regulators for target genes CABYR and CDH1. The expression level of LSD1 is negatively correlated with mono- and dimethylation of histone H3 lysine4(H3K4) at LSD1- target gene promoter region. No significant mono-methylation and dimethylation of H3 lysine9 methylation was detected at the promoter region of CABYR and CDH1. LSD1- depletion contributed to the upregulation of CABYR and CDH1 through enhancing the dimethylation of H3K4 at the LSD1-target genes promoter. LSD1- overexpression mediated the downregulation of CABYR and CDH1expression through decreasing the mono- and dimethylation of H3K4 at LSD1-target gene promoter in colon cancer cells. CABYR and CDH1 might be potential LSD1-target genes in colon

  14. Identification of regulators of polyploidization presents therapeutic targets for treatment of AMKL.

    PubMed

    Wen, Qiang; Goldenson, Benjamin; Silver, Serena J; Schenone, Monica; Dancik, Vlado; Huang, Zan; Wang, Ling-Zhi; Lewis, Timothy A; An, W Frank; Li, Xiaoyu; Bray, Mark-Anthony; Thiollier, Clarisse; Diebold, Lauren; Gilles, Laure; Vokes, Martha S; Moore, Christopher B; Bliss-Moreau, Meghan; Verplank, Lynn; Tolliday, Nicola J; Mishra, Rama; Vemula, Sasidhar; Shi, Jianjian; Wei, Lei; Kapur, Reuben; Lopez, Cécile K; Gerby, Bastien; Ballerini, Paola; Pflumio, Francoise; Gilliland, D Gary; Goldberg, Liat; Birger, Yehudit; Izraeli, Shai; Gamis, Alan S; Smith, Franklin O; Woods, William G; Taub, Jeffrey; Scherer, Christina A; Bradner, James E; Goh, Boon-Cher; Mercher, Thomas; Carpenter, Anne E; Gould, Robert J; Clemons, Paul A; Carr, Steven A; Root, David E; Schreiber, Stuart L; Stern, Andrew M; Crispino, John D

    2012-08-03

    The mechanism by which cells decide to skip mitosis to become polyploid is largely undefined. Here we used a high-content image-based screen to identify small-molecule probes that induce polyploidization of megakaryocytic leukemia cells and serve as perturbagens to help understand this process. Our study implicates five networks of kinases that regulate the switch to polyploidy. Moreover, we find that dimethylfasudil (diMF, H-1152P) selectively increased polyploidization, mature cell-surface marker expression, and apoptosis of malignant megakaryocytes. An integrated target identification approach employing proteomic and shRNA screening revealed that a major target of diMF is Aurora kinase A (AURKA). We further find that MLN8237 (Alisertib), a selective inhibitor of AURKA, induced polyploidization and expression of mature megakaryocyte markers in acute megakaryocytic leukemia (AMKL) blasts and displayed potent anti-AMKL activity in vivo. Our findings provide a rationale to support clinical trials of MLN8237 and other inducers of polyploidization and differentiation in AMKL. Copyright © 2012 Elsevier Inc. All rights reserved.

  15. Development of a computer-assisted forensic radiographic identification method using the lateral cervical and lumbar spine.

    PubMed

    Derrick, Sharon M; Raxter, Michelle H; Hipp, John A; Goel, Priya; Chan, Elaine F; Love, Jennifer C; Wiersema, Jason M; Akella, N Shastry

    2015-01-01

    Medical examiners and coroners (ME/C) in the United States hold statutory responsibility to identify deceased individuals who fall under their jurisdiction. The computer-assisted decedent identification (CADI) project was designed to modify software used in diagnosis and treatment of spinal injuries into a mathematically validated tool for ME/C identification of fleshed decedents. CADI software analyzes the shapes of targeted vertebral bodies imaged in an array of standard radiographs and quantifies the likelihood that any two of the radiographs contain matching vertebral bodies. Six validation tests measured the repeatability, reliability, and sensitivity of the method, and the effects of age, sex, and number of radiographs in array composition. CADI returned a 92-100% success rate in identifying the true matching pair of vertebrae within arrays of five to 30 radiographs. Further development of CADI is expected to produce a novel identification method for use in ME/C offices that is reliable, timely, and cost-effective. © 2014 American Academy of Forensic Sciences.

  16. Identification of Host-Targeted Small Molecules That Restrict Intracellular Mycobacterium tuberculosis Growth

    PubMed Central

    Silvis, Melanie R.; Luo, Samantha S.; Sogi, Kimberly; Vokes, Martha; Bray, Mark-Anthony; Carpenter, Anne E.; Moore, Christopher B.; Siddiqi, Noman; Rubin, Eric J.; Hung, Deborah T.

    2014-01-01

    Mycobacterium tuberculosis remains a significant threat to global health. Macrophages are the host cell for M. tuberculosis infection, and although bacteria are able to replicate intracellularly under certain conditions, it is also clear that macrophages are capable of killing M. tuberculosis if appropriately activated. The outcome of infection is determined at least in part by the host-pathogen interaction within the macrophage; however, we lack a complete understanding of which host pathways are critical for bacterial survival and replication. To add to our understanding of the molecular processes involved in intracellular infection, we performed a chemical screen using a high-content microscopic assay to identify small molecules that restrict mycobacterial growth in macrophages by targeting host functions and pathways. The identified host-targeted inhibitors restrict bacterial growth exclusively in the context of macrophage infection and predominantly fall into five categories: G-protein coupled receptor modulators, ion channel inhibitors, membrane transport proteins, anti-inflammatories, and kinase modulators. We found that fluoxetine, a selective serotonin reuptake inhibitor, enhances secretion of pro-inflammatory cytokine TNF-α and induces autophagy in infected macrophages, and gefitinib, an inhibitor of the Epidermal Growth Factor Receptor (EGFR), also activates autophagy and restricts growth. We demonstrate that during infection signaling through EGFR activates a p38 MAPK signaling pathway that prevents macrophages from effectively responding to infection. Inhibition of this pathway using gefitinib during in vivo infection reduces growth of M. tuberculosis in the lungs of infected mice. Our results support the concept that screening for inhibitors using intracellular models results in the identification of tool compounds for probing pathways during in vivo infection and may also result in the identification of new anti-tuberculosis agents that work by

  17. Modeling and validation of spectral BRDF on material surface of space target

    NASA Astrophysics Data System (ADS)

    Hou, Qingyu; Zhi, Xiyang; Zhang, Huili; Zhang, Wei

    2014-11-01

    The modeling and the validation methods of the spectral BRDF on the material surface of space target were presented. First, the microscopic characteristics of the space targets' material surface were analyzed based on fiber-optic spectrometer using to measure the direction reflectivity of the typical materials surface. To determine the material surface of space target is isotropic, atomic force microscopy was used to measure the material surface structure of space target and obtain Gaussian distribution model of microscopic surface element height. Then, the spectral BRDF model based on that the characteristics of the material surface were isotropic and the surface micro-facet with the Gaussian distribution which we obtained was constructed. The model characterizes smooth and rough surface well for describing the material surface of the space target appropriately. Finally, a spectral BRDF measurement platform in a laboratory was set up, which contains tungsten halogen lamp lighting system, fiber optic spectrometer detection system and measuring mechanical systems with controlling the entire experimental measurement and collecting measurement data by computers automatically. Yellow thermal control material and solar cell were measured with the spectral BRDF, which showed the relationship between the reflection angle and BRDF values at three wavelengths in 380nm, 550nm, 780nm, and the difference between theoretical model values and the measured data was evaluated by relative RMS error. Data analysis shows that the relative RMS error is less than 6%, which verified the correctness of the spectral BRDF model.

  18. Regulation, overexpression, and target gene identification of Potato Homeobox 15 (POTH15) – a class-I KNOX gene in potato

    PubMed Central

    Mahajan, Ameya S.; Kondhare, Kirtikumar R.; Rajabhoj, Mohit P.; Kumar, Amit; Ghate, Tejashree; Ravindran, Nevedha; Habib, Farhat; Siddappa, Sundaresha; Banerjee, Anjan K.

    2016-01-01

    Potato Homeobox 15 (POTH15) is a KNOX-I (Knotted1-like homeobox) family gene in potato that is orthologous to Shoot Meristemless (STM) in Arabidopsis. Despite numerous reports on KNOX genes from different species, studies in potato are limited. Here, we describe photoperiodic regulation of POTH15, its overexpression phenotype, and identification of its potential targets in potato (Solanum tuberosum ssp. andigena). qRT-PCR analysis showed a higher abundance of POTH15 mRNA in shoot tips and stolons under tuber-inducing short-day conditions. POTH15 promoter activity was detected in apical and axillary meristems, stolon tips, tuber eyes, and meristems of tuber sprouts, indicating its role in meristem maintenance and leaf development. POTH15 overexpression altered multiple morphological traits including leaf and stem development, leaflet number, and number of nodes and branches. In particular, the rachis of the leaf was completely reduced and leaves appeared as a bouquet of leaflets. Comparative transcriptomic analysis of 35S::GUS and two POTH15 overexpression lines identified more than 6000 differentially expressed genes, including 2014 common genes between the two overexpression lines. Functional analysis of these genes revealed their involvement in responses to hormones, biotic/abiotic stresses, transcription regulation, and signal transduction. qRT-PCR of selected candidate target genes validated their differential expression in both overexpression lines. Out of 200 randomly chosen POTH15 targets, 173 were found to have at least one tandem TGAC core motif, characteristic of KNOX interaction, within 3.0kb in the upstream sequence of the transcription start site. Overall, this study provides insights to the role of POTH15 in controlling diverse developmental processes in potato. PMID:27217546

  19. MESSI: metabolic engineering target selection and best strain identification tool.

    PubMed

    Kang, Kang; Li, Jun; Lim, Boon Leong; Panagiotou, Gianni

    2015-01-01

    Metabolic engineering and synthetic biology are synergistically related fields for manipulating target pathways and designing microorganisms that can act as chemical factories. Saccharomyces cerevisiae's ideal bioprocessing traits make yeast a very attractive chemical factory for production of fuels, pharmaceuticals, nutraceuticals as well as a wide range of chemicals. However, future attempts of engineering S. cerevisiae's metabolism using synthetic biology need to move towards more integrative models that incorporate the high connectivity of metabolic pathways and regulatory processes and the interactions in genetic elements across those pathways and processes. To contribute in this direction, we have developed Metabolic Engineering target Selection and best Strain Identification tool (MESSI), a web server for predicting efficient chassis and regulatory components for yeast bio-based production. The server provides an integrative platform for users to analyse ready-to-use public high-throughput metabolomic data, which are transformed to metabolic pathway activities for identifying the most efficient S. cerevisiae strain for the production of a compound of interest. As input MESSI accepts metabolite KEGG IDs or pathway names. MESSI outputs a ranked list of S. cerevisiae strains based on aggregation algorithms. Furthermore, through a genome-wide association study of the metabolic pathway activities with the strains' natural variation, MESSI prioritizes genes and small variants as potential regulatory points and promising metabolic engineering targets. Users can choose various parameters in the whole process such as (i) weight and expectation of each metabolic pathway activity in the final ranking of the strains, (ii) Weighted AddScore Fuse or Weighted Borda Fuse aggregation algorithm, (iii) type of variants to be included, (iv) variant sets in different biological levels.Database URL: http://sbb.hku.hk/MESSI/. © The Author(s) 2015. Published by Oxford University

  20. 20180318 - Structure identification by Mass Spectrometry Non-Targeted Analysis using the US EPA’s CompTox Chemistry Dashboard (ACS Spring)

    EPA Science Inventory

    Identification of unknowns in mass spectrometry based non-targeted analyses (NTA) requires the integration of complementary pieces of data to arrive at a confident, consensus structure. Researchers use chemical reference databases, spectral matching, fragment prediction tools, r...

  1. Selection, application, and validation of a set of molecular descriptors for nuclear receptor ligands.

    PubMed

    Stewart, Eugene L; Brown, Peter J; Bentley, James A; Willson, Timothy M

    2004-08-01

    A methodology for the selection and validation of nuclear receptor ligand chemical descriptors is described. After descriptors for a targeted chemical space were selected, a virtual screening methodology utilizing this space was formulated for the identification of potential NR ligands from our corporate collection. Using simple descriptors and our virtual screening method, we are able to quickly identify potential NR ligands from a large collection of compounds. As validation of the virtual screening procedure, an 8, 000-membered NR targeted set and a 24, 000-membered diverse control set of compounds were selected from our in-house general screening collection and screened in parallel across a number of orphan NR FRET assays. For the two assays that provided at least one hit per set by the established minimum pEC(50) for activity, the results showed a 2-fold increase in the hit-rate of the targeted compound set over the diverse set.

  2. Visualization and Analysis of MiRNA-Targets Interactions Networks.

    PubMed

    León, Luis E; Calligaris, Sebastián D

    2017-01-01

    MicroRNAs are a class of small, noncoding RNA molecules of 21-25 nucleotides in length that regulate the gene expression by base-pairing with the target mRNAs, mainly leading to down-regulation or repression of the target genes. MicroRNAs are involved in diverse regulatory pathways in normal and pathological conditions. In this context, it is highly important to identify the targets of specific microRNA in order to understand the mechanism of its regulation and consequently its involvement in disease. However, the microRNA target identification is experimentally laborious and time-consuming. The in silico prediction of microRNA targets is an extremely useful approach because you can identify potential mRNA targets, reduce the number of possibilities and then, validate a few microRNA-mRNA interactions in an in vitro experimental model. In this chapter, we describe, in a simple way, bioinformatics guidelines to use miRWalk database and Cytoscape software for analyzing microRNA-mRNA interactions through their visualization as a network.

  3. Core Proteomic Analysis of Unique Metabolic Pathways of Salmonella enterica for the Identification of Potential Drug Targets.

    PubMed

    Uddin, Reaz; Sufian, Muhammad

    2016-01-01

    Infections caused by Salmonella enterica, a Gram-negative facultative anaerobic bacteria belonging to the family of Enterobacteriaceae, are major threats to the health of humans and animals. The recent availability of complete genome data of pathogenic strains of the S. enterica gives new avenues for the identification of drug targets and drug candidates. We have used the genomic and metabolic pathway data to identify pathways and proteins essential to the pathogen and absent from the host. We took the whole proteome sequence data of 42 strains of S. enterica and Homo sapiens along with KEGG-annotated metabolic pathway data, clustered proteins sequences using CD-HIT, identified essential genes using DEG database and discarded S. enterica homologs of human proteins in unique metabolic pathways (UMPs) and characterized hypothetical proteins with SVM-prot and InterProScan. Through this core proteomic analysis we have identified enzymes essential to the pathogen. The identification of 73 enzymes common in 42 strains of S. enterica is the real strength of the current study. We proposed all 73 unexplored enzymes as potential drug targets against the infections caused by the S. enterica. The study is comprehensive around S. enterica and simultaneously considered every possible pathogenic strain of S. enterica. This comprehensiveness turned the current study significant since, to the best of our knowledge it is the first subtractive core proteomic analysis of the unique metabolic pathways applied to any pathogen for the identification of drug targets. We applied extensive computational methods to shortlist few potential drug targets considering the druggability criteria e.g. Non-homologous to the human host, essential to the pathogen and playing significant role in essential metabolic pathways of the pathogen (i.e. S. enterica). In the current study, the subtractive proteomics through a novel approach was applied i.e. by considering only proteins of the unique metabolic

  4. Molecular Targets in Advanced Therapeutics of Cancers: The Role of Pharmacogenetics.

    PubMed

    Abubakar, Murtala B; Gan, Siew Hua

    2016-01-01

    The advent of advanced molecular targeted therapy has resulted in improved prognoses for patients with advanced malignancies. However, despite the significant success and specificity of this advocated targeted therapy, significant on- and off-target adverse effects and inter-individual variability in treatment responses have been reported. The interpatient variability in drug response has been suggested to be partly due to variations in patient genomes. Therefore, the identification of genetic biomarkers by conducting pharmacogenetics studies can help predict patient responses to targeted therapy and may serve as a basis for individualized treatment. In this review, both clinically established and potential molecular targets are highlighted. Overall, current literature suggests that individualization of targeted therapy is promising; however, integrating the clinical benefits of identified biomarkers into clinical practice for personalized medicine remains a major challenge, and further studies to validate these markers and identify novel therapeutic approaches are needed. © 2016 S. Karger AG, Basel.

  5. Hierarchical virtual screening for the discovery of new molecular scaffolds in antibacterial hit identification

    PubMed Central

    Ballester, Pedro J.; Mangold, Martina; Howard, Nigel I.; Robinson, Richard L. Marchese; Abell, Chris; Blumberger, Jochen; Mitchell, John B. O.

    2012-01-01

    One of the initial steps of modern drug discovery is the identification of small organic molecules able to inhibit a target macromolecule of therapeutic interest. A small proportion of these hits are further developed into lead compounds, which in turn may ultimately lead to a marketed drug. A commonly used screening protocol used for this task is high-throughput screening (HTS). However, the performance of HTS against antibacterial targets has generally been unsatisfactory, with high costs and low rates of hit identification. Here, we present a novel computational methodology that is able to identify a high proportion of structurally diverse inhibitors by searching unusually large molecular databases in a time-, cost- and resource-efficient manner. This virtual screening methodology was tested prospectively on two versions of an antibacterial target (type II dehydroquinase from Mycobacterium tuberculosis and Streptomyces coelicolor), for which HTS has not provided satisfactory results and consequently practically all known inhibitors are derivatives of the same core scaffold. Overall, our protocols identified 100 new inhibitors, with calculated Ki ranging from 4 to 250 μM (confirmed hit rates are 60% and 62% against each version of the target). Most importantly, over 50 new active molecular scaffolds were discovered that underscore the benefits that a wide application of prospectively validated in silico screening tools is likely to bring to antibacterial hit identification. PMID:22933186

  6. Hierarchical virtual screening for the discovery of new molecular scaffolds in antibacterial hit identification.

    PubMed

    Ballester, Pedro J; Mangold, Martina; Howard, Nigel I; Robinson, Richard L Marchese; Abell, Chris; Blumberger, Jochen; Mitchell, John B O

    2012-12-07

    One of the initial steps of modern drug discovery is the identification of small organic molecules able to inhibit a target macromolecule of therapeutic interest. A small proportion of these hits are further developed into lead compounds, which in turn may ultimately lead to a marketed drug. A commonly used screening protocol used for this task is high-throughput screening (HTS). However, the performance of HTS against antibacterial targets has generally been unsatisfactory, with high costs and low rates of hit identification. Here, we present a novel computational methodology that is able to identify a high proportion of structurally diverse inhibitors by searching unusually large molecular databases in a time-, cost- and resource-efficient manner. This virtual screening methodology was tested prospectively on two versions of an antibacterial target (type II dehydroquinase from Mycobacterium tuberculosis and Streptomyces coelicolor), for which HTS has not provided satisfactory results and consequently practically all known inhibitors are derivatives of the same core scaffold. Overall, our protocols identified 100 new inhibitors, with calculated K(i) ranging from 4 to 250 μM (confirmed hit rates are 60% and 62% against each version of the target). Most importantly, over 50 new active molecular scaffolds were discovered that underscore the benefits that a wide application of prospectively validated in silico screening tools is likely to bring to antibacterial hit identification.

  7. Automatic identification of variables in epidemiological datasets using logic regression.

    PubMed

    Lorenz, Matthias W; Abdi, Negin Ashtiani; Scheckenbach, Frank; Pflug, Anja; Bülbül, Alpaslan; Catapano, Alberico L; Agewall, Stefan; Ezhov, Marat; Bots, Michiel L; Kiechl, Stefan; Orth, Andreas

    2017-04-13

    For an individual participant data (IPD) meta-analysis, multiple datasets must be transformed in a consistent format, e.g. using uniform variable names. When large numbers of datasets have to be processed, this can be a time-consuming and error-prone task. Automated or semi-automated identification of variables can help to reduce the workload and improve the data quality. For semi-automation high sensitivity in the recognition of matching variables is particularly important, because it allows creating software which for a target variable presents a choice of source variables, from which a user can choose the matching one, with only low risk of having missed a correct source variable. For each variable in a set of target variables, a number of simple rules were manually created. With logic regression, an optimal Boolean combination of these rules was searched for every target variable, using a random subset of a large database of epidemiological and clinical cohort data (construction subset). In a second subset of this database (validation subset), this optimal combination rules were validated. In the construction sample, 41 target variables were allocated on average with a positive predictive value (PPV) of 34%, and a negative predictive value (NPV) of 95%. In the validation sample, PPV was 33%, whereas NPV remained at 94%. In the construction sample, PPV was 50% or less in 63% of all variables, in the validation sample in 71% of all variables. We demonstrated that the application of logic regression in a complex data management task in large epidemiological IPD meta-analyses is feasible. However, the performance of the algorithm is poor, which may require backup strategies.

  8. Genome-Scale Screening of Drug-Target Associations Relevant to Ki Using a Chemogenomics Approach

    PubMed Central

    Cao, Dong-Sheng; Liang, Yi-Zeng; Deng, Zhe; Hu, Qian-Nan; He, Min; Xu, Qing-Song; Zhou, Guang-Hua; Zhang, Liu-Xia; Deng, Zi-xin; Liu, Shao

    2013-01-01

    The identification of interactions between drugs and target proteins plays a key role in genomic drug discovery. In the present study, the quantitative binding affinities of drug-target pairs are differentiated as a measurement to define whether a drug interacts with a protein or not, and then a chemogenomics framework using an unbiased set of general integrated features and random forest (RF) is employed to construct a predictive model which can accurately classify drug-target pairs. The predictability of the model is further investigated and validated by several independent validation sets. The built model is used to predict drug-target associations, some of which were confirmed by comparing experimental data from public biological resources. A drug-target interaction network with high confidence drug-target pairs was also reconstructed. This network provides further insight for the action of drugs and targets. Finally, a web-based server called PreDPI-Ki was developed to predict drug-target interactions for drug discovery. In addition to providing a high-confidence list of drug-target associations for subsequent experimental investigation guidance, these results also contribute to the understanding of drug-target interactions. We can also see that quantitative information of drug-target associations could greatly promote the development of more accurate models. The PreDPI-Ki server is freely available via: http://sdd.whu.edu.cn/dpiki. PMID:23577055

  9. Identification and validation of reference genes for quantification of target gene expression with quantitative real-time PCR for tall fescue under four abiotic stresses.

    PubMed

    Yang, Zhimin; Chen, Yu; Hu, Baoyun; Tan, Zhiqun; Huang, Bingru

    2015-01-01

    Tall fescue (Festuca arundinacea Schreb.) is widely utilized as a major forage and turfgrass species in the temperate regions of the world and is a valuable plant material for studying molecular mechanisms of grass stress tolerance due to its superior drought and heat tolerance among cool-season species. Selection of suitable reference genes for quantification of target gene expression is important for the discovery of molecular mechanisms underlying improved growth traits and stress tolerance. The stability of nine potential reference genes (ACT, TUB, EF1a, GAPDH, SAND, CACS, F-box, PEPKR1 and TIP41) was evaluated using four programs, GeNorm, NormFinder, BestKeeper, and RefFinder. The combinations of SAND and TUB or TIP41 and TUB were most stably expressed in salt-treated roots or leaves. The combinations of GAPDH with TIP41 or TUB were stable in roots and leaves under drought stress. TIP41 and PEPKR1 exhibited stable expression in cold-treated roots, and the combination of F-box, TIP41 and TUB was also stable in cold-treated leaves. CACS and TUB were the two most stable reference genes in heat-stressed roots. TIP41 combined with TUB and ACT was stably expressed in heat-stressed leaves. Finally, quantitative real-time polymerase chain reaction (qRT-PCR) assays of the target gene FaWRKY1 using the identified most stable reference genes confirmed the reliability of selected reference genes. The selection of suitable reference genes in tall fescue will allow for more accurate identification of stress-tolerance genes and molecular mechanisms conferring stress tolerance in this stress-tolerant species.

  10. A Pathway to Freedom: An Evaluation of Screening Tools for the Identification of Trafficking Victims.

    PubMed

    Bespalova, Nadejda; Morgan, Juliet; Coverdale, John

    2016-02-01

    Because training residents and faculty to identify human trafficking victims is a major public health priority, the authors review existing assessment tools. PubMed and Google were searched using combinations of search terms including human, trafficking, sex, labor, screening, identification, and tool. Nine screening tools that met the inclusion criteria were found. They varied greatly in length, format, target demographic, supporting resources, and other parameters. Only two tools were designed specifically for healthcare providers. Only one tool was formally assessed to be valid and reliable in a pilot project in trafficking victim service organizations, although it has not been validated in the healthcare setting. This toolbox should facilitate the education of resident physicians and faculty in screening for trafficking victims, assist educators in assessing screening skills, and promote future research on the identification of trafficking victims.

  11. Lineup identification by children: effects of clothing bias.

    PubMed

    Freire, Alejo; Lee, Kang; Williamson, Karen S; Stuart, Sarah J E; Lindsay, R C L

    2004-06-01

    This study examined effects of clothing cues on children's identification accuracy from lineups. Four- to 14-year-olds (n = 228) saw 12 video clips of individuals, each wearing a distinctly colored shirt. After watching each clip children were presented with a target-present or target-absent photo lineup. Three clothing conditions were included. In 2 conditions all lineup members wore the same colored shirt; in the third, biased condition, the shirt color of only one individual matched that seen in the preceding clip (the target in target-present trials and the replacement in target-absent trials). Correct identifications of the target in target-present trials were most frequent in the biased condition, whereas in target-absent trials the biased condition led to more false identifications of the target replacement. Older children were more accurate than younger children, both in choosing the target from target-present lineups and rejecting target-absent lineups. These findings suggest that a simple clothing cue such as shirt color can have a significant impact on children's lineup identification accuracy.

  12. Lineup Identification by Children: Effects of Clothing Bias

    PubMed Central

    Freire, Alejo; Lee, Kang; Williamson, Karen S.; Stuart, Sarah J. E.; Lindsay, R. C. L.

    2008-01-01

    This study examined effects of clothing cues on children's identification accuracy from lineups. Four- to 14-year-olds (n = 228) saw 12 video clips of individuals, each wearing a distinctly colored shirt. After watching each clip children were presented with a target-present or target-absent photo lineup. Three clothing conditions were included. In 2 conditions all lineup members wore the same colored shirt; in the third, biased condition, the shirt color of only one individual matched that seen in the preceding clip (the target in target-present trials and the replacement in target-absent trials). Correct identifications of the target in target-present trials were most frequent in the biased condition, whereas in target-absent trials the biased condition led to more false identifications of the target replacement. Older children were more accurate than younger children, both in choosing the target from target-present lineups and rejecting target-absent lineups. These findings suggest that a simple clothing cue such as shirt color can have a significant impact on children's lineup identification accuracy. PMID:15264450

  13. Target identification by image analysis.

    PubMed

    Fetz, V; Prochnow, H; Brönstrup, M; Sasse, F

    2016-05-04

    Covering: 1997 to the end of 2015Each biologically active compound induces phenotypic changes in target cells that are characteristic for its mode of action. These phenotypic alterations can be directly observed under the microscope or made visible by labelling structural elements or selected proteins of the cells with dyes. A comparison of the cellular phenotype induced by a compound of interest with the phenotypes of reference compounds with known cellular targets allows predicting its mode of action. While this approach has been successfully applied to the characterization of natural products based on a visual inspection of images, recent studies used automated microscopy and analysis software to increase speed and to reduce subjective interpretation. In this review, we give a general outline of the workflow for manual and automated image analysis, and we highlight natural products whose bacterial and eucaryotic targets could be identified through such approaches.

  14. Drug target identification in protozoan parasites.

    PubMed

    Müller, Joachim; Hemphill, Andrew

    2016-08-01

    Despite the fact that diseases caused by protozoan parasites represent serious challenges for public health, animal production and welfare, only a limited panel of drugs has been marketed for clinical applications. Herein, the authors investigate two strategies, namely whole organism screening and target-based drug design. The present pharmacopoeia has resulted from whole organism screening, and the mode of action and targets of selected drugs are discussed. However, the more recent extensive genome sequencing efforts and the development of dry and wet lab genomics and proteomics that allow high-throughput screening of interactions between micromolecules and recombinant proteins has resulted in target-based drug design as the predominant focus in anti-parasitic drug development. Selected examples of target-based drug design studies are presented, and calcium-dependent protein kinases, important drug targets in apicomplexan parasites, are discussed in more detail. Despite the enormous efforts in target-based drug development, this approach has not yet generated market-ready antiprotozoal drugs. However, whole-organism screening approaches, comprising of both in vitro and in vivo investigations, should not be disregarded. The repurposing of already approved and marketed drugs could be a suitable strategy to avoid fastidious approval procedures, especially in the case of neglected or veterinary parasitoses.

  15. Drug target identification in protozoan parasites

    PubMed Central

    Müller, Joachim; Hemphill, Andrew

    2016-01-01

    Introduction Despite the fact that diseases caused by protozoan parasites represent serious challenges for public health, animal production and welfare, only a limited panel of drugs has been marketed for clinical applications. Areas covered Herein, the authors investigate two strategies, namely whole organism screening and target-based drug design. The present pharmacopoeia has resulted from whole organism screening, and the mode of action and targets of selected drugs are discussed. However, the more recent extensive genome sequencing efforts and the development of dry and wet lab genomics and proteomics that allow high-throughput screening of interactions between micromolecules and recombinant proteins has resulted in target-based drug design as the predominant focus in anti-parasitic drug development. Selected examples of target-based drug design studies are presented, and calcium-dependent protein kinases, important drug targets in apicomplexan parasites, are discussed in more detail. Expert opinion Despite the enormous efforts in target-based drug development, this approach has not yet generated market-ready antiprotozoal drugs. However, whole-organism screening approaches, comprising of both in vitro and in vivo investigations, should not be disregarded. The repurposing of already approved and marketed drugs could be a suitable strategy to avoid fastidious approval procedures, especially in the case of neglected or veterinary parasitoses. PMID:27238605

  16. RNAi phenotype profiling of kinases identifies potential therapeutic targets in Ewing's sarcoma.

    PubMed

    Arora, Shilpi; Gonzales, Irma M; Hagelstrom, R Tanner; Beaudry, Christian; Choudhary, Ashish; Sima, Chao; Tibes, Raoul; Mousses, Spyro; Azorsa, David O

    2010-08-18

    Ewing's sarcomas are aggressive musculoskeletal tumors occurring most frequently in the long and flat bones as a solitary lesion mostly during the teen-age years of life. With current treatments, significant number of patients relapse and survival is poor for those with metastatic disease. As part of novel target discovery in Ewing's sarcoma, we applied RNAi mediated phenotypic profiling to identify kinase targets involved in growth and survival of Ewing's sarcoma cells. Four Ewing's sarcoma cell lines TC-32, TC-71, SK-ES-1 and RD-ES were tested in high throughput-RNAi screens using a siRNA library targeting 572 kinases. Knockdown of 25 siRNAs reduced the growth of all four Ewing's sarcoma cell lines in replicate screens. Of these, 16 siRNA were specific and reduced proliferation of Ewing's sarcoma cells as compared to normal fibroblasts. Secondary validation and preliminary mechanistic studies highlighted the kinases STK10 and TNK2 as having important roles in growth and survival of Ewing's sarcoma cells. Furthermore, knockdown of STK10 and TNK2 by siRNA showed increased apoptosis. In summary, RNAi-based phenotypic profiling proved to be a powerful gene target discovery strategy, leading to successful identification and validation of STK10 and TNK2 as two novel potential therapeutic targets for Ewing's sarcoma.

  17. Targeting Thromboxane A2 Receptor for Anti-Metastasis Therapy of Breast Cancer

    DTIC Science & Technology

    2011-09-01

    of cell function by Rho GTPases." Drug News Perspect 14(7): 389-95. Erickson, J. W., R. A. Cerione, et al. (1997). "Identification of an actin...Focusing Tumor Microenvironment, Stem Cells and Metastasis 570 (MTOC) and Golgi apparatus to the front of the nucleus, oriented toward the direction of...define the function of TP in tumor cell motility and to validate TP as a target for anti-metastasis therapy of breast cancer. In the first aim, the

  18. Computational-experimental approach to drug-target interaction mapping: A case study on kinase inhibitors

    PubMed Central

    Ravikumar, Balaguru; Parri, Elina; Timonen, Sanna; Airola, Antti; Wennerberg, Krister

    2017-01-01

    Due to relatively high costs and labor required for experimental profiling of the full target space of chemical compounds, various machine learning models have been proposed as cost-effective means to advance this process in terms of predicting the most potent compound-target interactions for subsequent verification. However, most of the model predictions lack direct experimental validation in the laboratory, making their practical benefits for drug discovery or repurposing applications largely unknown. Here, we therefore introduce and carefully test a systematic computational-experimental framework for the prediction and pre-clinical verification of drug-target interactions using a well-established kernel-based regression algorithm as the prediction model. To evaluate its performance, we first predicted unmeasured binding affinities in a large-scale kinase inhibitor profiling study, and then experimentally tested 100 compound-kinase pairs. The relatively high correlation of 0.77 (p < 0.0001) between the predicted and measured bioactivities supports the potential of the model for filling the experimental gaps in existing compound-target interaction maps. Further, we subjected the model to a more challenging task of predicting target interactions for such a new candidate drug compound that lacks prior binding profile information. As a specific case study, we used tivozanib, an investigational VEGF receptor inhibitor with currently unknown off-target profile. Among 7 kinases with high predicted affinity, we experimentally validated 4 new off-targets of tivozanib, namely the Src-family kinases FRK and FYN A, the non-receptor tyrosine kinase ABL1, and the serine/threonine kinase SLK. Our sub-sequent experimental validation protocol effectively avoids any possible information leakage between the training and validation data, and therefore enables rigorous model validation for practical applications. These results demonstrate that the kernel-based modeling approach

  19. Identification of microRNA-mRNA modules using microarray data.

    PubMed

    Jayaswal, Vivek; Lutherborrow, Mark; Ma, David D F; Yang, Yee H

    2011-03-06

    MicroRNAs (miRNAs) are post-transcriptional regulators of mRNA expression and are involved in numerous cellular processes. Consequently, miRNAs are an important component of gene regulatory networks and an improved understanding of miRNAs will further our knowledge of these networks. There is a many-to-many relationship between miRNAs and mRNAs because a single miRNA targets multiple mRNAs and a single mRNA is targeted by multiple miRNAs. However, most of the current methods for the identification of regulatory miRNAs and their target mRNAs ignore this biological observation and focus on miRNA-mRNA pairs. We propose a two-step method for the identification of many-to-many relationships between miRNAs and mRNAs. In the first step, we obtain miRNA and mRNA clusters using a combination of miRNA-target mRNA prediction algorithms and microarray expression data. In the second step, we determine the associations between miRNA clusters and mRNA clusters based on changes in miRNA and mRNA expression profiles. We consider the miRNA-mRNA clusters with statistically significant associations to be potentially regulatory and, therefore, of biological interest. Our method reduces the interactions between several hundred miRNAs and several thousand mRNAs to a few miRNA-mRNA groups, thereby facilitating a more meaningful biological analysis and a more targeted experimental validation.

  20. Experimental and statistical post-validation of positive example EST sequences carrying peroxisome targeting signals type 1 (PTS1).

    PubMed

    Lingner, Thomas; Kataya, Amr R A; Reumann, Sigrun

    2012-02-01

    We recently developed the first algorithms specifically for plants to predict proteins carrying peroxisome targeting signals type 1 (PTS1) from genome sequences. As validated experimentally, the prediction methods are able to correctly predict unknown peroxisomal Arabidopsis proteins and to infer novel PTS1 tripeptides. The high prediction performance is primarily determined by the large number and sequence diversity of the underlying positive example sequences, which mainly derived from EST databases. However, a few constructs remained cytosolic in experimental validation studies, indicating sequencing errors in some ESTs. To identify erroneous sequences, we validated subcellular targeting of additional positive example sequences in the present study. Moreover, we analyzed the distribution of prediction scores separately for each orthologous group of PTS1 proteins, which generally resembled normal distributions with group-specific mean values. The cytosolic sequences commonly represented outliers of low prediction scores and were located at the very tail of a fitted normal distribution. Three statistical methods for identifying outliers were compared in terms of sensitivity and specificity." Their combined application allows elimination of erroneous ESTs from positive example data sets. This new post-validation method will further improve the prediction accuracy of both PTS1 and PTS2 protein prediction models for plants, fungi, and mammals.

  1. Experimental and statistical post-validation of positive example EST sequences carrying peroxisome targeting signals type 1 (PTS1)

    PubMed Central

    Lingner, Thomas; Kataya, Amr R. A.; Reumann, Sigrun

    2012-01-01

    We recently developed the first algorithms specifically for plants to predict proteins carrying peroxisome targeting signals type 1 (PTS1) from genome sequences.1 As validated experimentally, the prediction methods are able to correctly predict unknown peroxisomal Arabidopsis proteins and to infer novel PTS1 tripeptides. The high prediction performance is primarily determined by the large number and sequence diversity of the underlying positive example sequences, which mainly derived from EST databases. However, a few constructs remained cytosolic in experimental validation studies, indicating sequencing errors in some ESTs. To identify erroneous sequences, we validated subcellular targeting of additional positive example sequences in the present study. Moreover, we analyzed the distribution of prediction scores separately for each orthologous group of PTS1 proteins, which generally resembled normal distributions with group-specific mean values. The cytosolic sequences commonly represented outliers of low prediction scores and were located at the very tail of a fitted normal distribution. Three statistical methods for identifying outliers were compared in terms of sensitivity and specificity.” Their combined application allows elimination of erroneous ESTs from positive example data sets. This new post-validation method will further improve the prediction accuracy of both PTS1 and PTS2 protein prediction models for plants, fungi, and mammals. PMID:22415050

  2. In silico identification and characterization of conserved miRNAs and their target genes in sweet potato (Ipomoea batatas L.) Expressed Sequence Tags (ESTs)

    PubMed Central

    Dehury, Budheswar; Panda, Debashis; Sahu, Jagajjit; Sahu, Mousumi; Sarma, Kishore; Barooah, Madhumita; Sen, Priyabrata; Modi, Mahendra Kumar

    2013-01-01

    The endogenous small non-coding micro RNAs (miRNAs), which are typically ~21–24 nt nucleotides, play a crucial role in regulating the intrinsic normal growth of cells and development of the plants as well as in maintaining the integrity of genomes. These small non-coding RNAs function as the universal specificity factors in post-transcriptional gene silencing. Discovering miRNAs, identifying their targets, and further inferring miRNA functions is a routine process to understand normal biological processes of miRNAs and their roles in the development of plants. Comparative genomics based approach using expressed sequence tags (EST) and genome survey sequences (GSS) offer a cost-effective platform for identification and characterization of miRNAs and their target genes in plants. Despite the fact that sweet potato (Ipomoea batatas L.) is an important staple food source for poor small farmers throughout the world, the role of miRNA in various developmental processes remains largely unknown. In this paper, we report the computational identification of miRNAs and their target genes in sweet potato from their ESTs. Using comparative genomics-based approach, 8 potential miRNA candidates belonging to miR168, miR2911, and miR156 families were identified from 23 406 ESTs in sweet potato. A total of 42 target genes were predicted and their probable functions were illustrated. Most of the newly identified miRNAs target transcription factors as well as genes involved in plant growth and development, signal transduction, metabolism, defense, and stress response. The identification of miRNAs and their targets is expected to accelerate the pace of miRNA discovery, leading to an improved understanding of the role of miRNA in development and physiology of sweet potato, as well as stress response. PMID:24067297

  3. Real-Time Gas Identification by Analyzing the Transient Response of Capillary-Attached Conductive Gas Sensor

    PubMed Central

    Bahraminejad, Behzad; Basri, Shahnor; Isa, Maryam; Hambli, Zarida

    2010-01-01

    In this study, the ability of the Capillary-attached conductive gas sensor (CGS) in real-time gas identification was investigated. The structure of the prototype fabricated CGS is presented. Portions were selected from the beginning of the CGS transient response including the first 11 samples to the first 100 samples. Different feature extraction and classification methods were applied on the selected portions. Validation of methods was evaluated to study the ability of an early portion of the CGS transient response in target gas (TG) identification. Experimental results proved that applying extracted features from an early part of the CGS transient response along with a classifier can distinguish short-chain alcohols from each other perfectly. Decreasing time of exposition in the interaction between target gas and sensing element improved the reliability of the sensor. Classification rate was also improved and time of identification was decreased. Moreover, the results indicated the optimum interval of the early transient response of the CGS for selecting portions to achieve the best classification rates. PMID:22219666

  4. Regularities in eyewitness identification.

    PubMed

    Clark, Steven E; Howell, Ryan T; Davey, Sherrie L

    2008-06-01

    What do eyewitness identification experiments typically show? We address this question through a meta-analysis of 94 comparisons between target-present and target-absent lineups. The analyses showed that: (a) correct identifications and correct-nonidentifications were uncorrelated, (b) suspect identifications were more diagnostic with respect to the suspect's guilt or innocence than any other response, (c) nonidentifications were diagnostic of the suspect's innocence, (d) the diagnosticity of foil identifications depended on lineup composition, and (e) don't know responses were nondiagnostic with respect to guilt or innocence. Results of diagnosticity analyses for simultaneous and sequential lineups varied for full-sample versus direct-comparison analyses. Diagnosticity patterns also varied as a function of lineup composition. Theoretical, forensic, and legal implications are discussed.

  5. Zero-G experimental validation of a robotics-based inertia identification algorithm

    NASA Astrophysics Data System (ADS)

    Bruggemann, Jeremy J.; Ferrel, Ivann; Martinez, Gerardo; Xie, Pu; Ma, Ou

    2010-04-01

    The need to efficiently identify the changing inertial properties of on-orbit spacecraft is becoming more critical as satellite on-orbit services, such as refueling and repairing, become increasingly aggressive and complex. This need stems from the fact that a spacecraft's control system relies on the knowledge of the spacecraft's inertia parameters. However, the inertia parameters may change during flight for reasons such as fuel usage, payload deployment or retrieval, and docking/capturing operations. New Mexico State University's Dynamics, Controls, and Robotics Research Group has proposed a robotics-based method of identifying unknown spacecraft inertia properties1. Previous methods require firing known thrusts then measuring the thrust, and the velocity and acceleration changes. The new method utilizes the concept of momentum conservation, while employing a robotic device powered by renewable energy to excite the state of the satellite. Thus, it requires no fuel usage or force and acceleration measurements. The method has been well studied in theory and demonstrated by simulation. However its experimental validation is challenging because a 6- degree-of-freedom motion in a zero-gravity condition is required. This paper presents an on-going effort to test the inertia identification method onboard the NASA zero-G aircraft. The design and capability of the test unit will be discussed in addition to the flight data. This paper also introduces the design and development of an airbearing based test used to partially validate the method, in addition to the approach used to obtain reference value for the test system's inertia parameters that can be used for comparison with the algorithm results.

  6. Identification of agents effective against multiple toxins and viruses by host-oriented cell targeting.

    PubMed

    Zilbermintz, Leeor; Leonardi, William; Jeong, Sun-Young; Sjodt, Megan; McComb, Ryan; Ho, Chi-Lee C; Retterer, Cary; Gharaibeh, Dima; Zamani, Rouzbeh; Soloveva, Veronica; Bavari, Sina; Levitin, Anastasia; West, Joel; Bradley, Kenneth A; Clubb, Robert T; Cohen, Stanley N; Gupta, Vivek; Martchenko, Mikhail

    2015-08-27

    A longstanding and still-increasing threat to the effective treatment of infectious diseases is resistance to antimicrobial countermeasures. Potentially, the targeting of host proteins and pathways essential for the detrimental effects of pathogens offers an approach that may discover broad-spectrum anti-pathogen countermeasures and circumvent the effects of pathogen mutations leading to resistance. Here we report implementation of a strategy for discovering broad-spectrum host-oriented therapies against multiple pathogenic agents by multiplex screening of drugs for protection against the detrimental effects of multiple pathogens, identification of host cell pathways inhibited by the drug, and screening for effects of the agent on other pathogens exploiting the same pathway. We show that a clinically used antimalarial drug, Amodiaquine, discovered by this strategy, protects host cells against infection by multiple toxins and viruses by inhibiting host cathepsin B. Our results reveal the practicality of discovering broadly acting anti-pathogen countermeasures that target host proteins exploited by pathogens.

  7. Uncertainty quantification in operational modal analysis with stochastic subspace identification: Validation and applications

    NASA Astrophysics Data System (ADS)

    Reynders, Edwin; Maes, Kristof; Lombaert, Geert; De Roeck, Guido

    2016-01-01

    Identified modal characteristics are often used as a basis for the calibration and validation of dynamic structural models, for structural control, for structural health monitoring, etc. It is therefore important to know their accuracy. In this article, a method for estimating the (co)variance of modal characteristics that are identified with the stochastic subspace identification method is validated for two civil engineering structures. The first structure is a damaged prestressed concrete bridge for which acceleration and dynamic strain data were measured in 36 different setups. The second structure is a mid-rise building for which acceleration data were measured in 10 different setups. There is a good quantitative agreement between the predicted levels of uncertainty and the observed variability of the eigenfrequencies and damping ratios between the different setups. The method can therefore be used with confidence for quantifying the uncertainty of the identified modal characteristics, also when some or all of them are estimated from a single batch of vibration data. Furthermore, the method is seen to yield valuable insight in the variability of the estimation accuracy from mode to mode and from setup to setup: the more informative a setup is regarding an estimated modal characteristic, the smaller is the estimated variance.

  8. Identification of a pharmacological target for genioglossus reactivation throughout sleep.

    PubMed

    Grace, Kevin P; Hughes, Stuart W; Horner, Richard L

    2014-01-01

    Obstructive sleep apnea (OSA) is a significant public health problem caused by repeated episodes of upper airway closure that occur only during sleep. Attempts to treat OSA pharmacologically have been unsuccessful because there has not been identification of a target operating at cranial motor nuclei, blockade of which can reactivate pharyngeal muscle activity throughout sleep. Increasing potassium conductance is a common mechanism by which state-dependent neuromodulators reduce motoneuron excitability. Therefore, we aimed to determine if potassium channel blockade is an effective strategy to reactivate the pharyngeal musculature throughout sleep. In rats chronically instrumented for recording sleep-wake states and respiratory motor activities, we locally microperfused pharmacological agents into the hypoglossal motor pool to modulate potassium channels of three major classes: inwardly rectifying, two-pore domain, and voltage-gated. Microperfusion of the inwardly rectifying potassium channel blocker, barium, as well as the voltage-gated potassium channel blockers, tetraethylammonium and 4-aminopyridine, increased tonic and respiratory-related genioglossus activities throughout nonrapid eye movement (non-REM) and rapid eye movement (REM) sleep to 133-300% of levels present during baseline wakefulness. In contrast, microperfusion of methanandamide (TWIK-related acid-sensitive potassium [TASK] channel blocker/cannabinoid receptor agonist) activated genioglossus in wakefulness but not in sleep. These findings establish proof-of-principle that targeted blockade of certain potassium channels at the hypoglossal motor pool is an effective strategy for reversing upper airway hypotonia and causing sustained reactivation of genioglossus throughout nonrapid eye movement and rapid eye movement sleep. These findings identify an important new direction for translational approaches to the pharmacological treatment of obstructive sleep apnea.

  9. Large-Scale Off-Target Identification Using Fast and Accurate Dual Regularized One-Class Collaborative Filtering and Its Application to Drug Repurposing.

    PubMed

    Lim, Hansaim; Poleksic, Aleksandar; Yao, Yuan; Tong, Hanghang; He, Di; Zhuang, Luke; Meng, Patrick; Xie, Lei

    2016-10-01

    Target-based screening is one of the major approaches in drug discovery. Besides the intended target, unexpected drug off-target interactions often occur, and many of them have not been recognized and characterized. The off-target interactions can be responsible for either therapeutic or side effects. Thus, identifying the genome-wide off-targets of lead compounds or existing drugs will be critical for designing effective and safe drugs, and providing new opportunities for drug repurposing. Although many computational methods have been developed to predict drug-target interactions, they are either less accurate than the one that we are proposing here or computationally too intensive, thereby limiting their capability for large-scale off-target identification. In addition, the performances of most machine learning based algorithms have been mainly evaluated to predict off-target interactions in the same gene family for hundreds of chemicals. It is not clear how these algorithms perform in terms of detecting off-targets across gene families on a proteome scale. Here, we are presenting a fast and accurate off-target prediction method, REMAP, which is based on a dual regularized one-class collaborative filtering algorithm, to explore continuous chemical space, protein space, and their interactome on a large scale. When tested in a reliable, extensive, and cross-gene family benchmark, REMAP outperforms the state-of-the-art methods. Furthermore, REMAP is highly scalable. It can screen a dataset of 200 thousands chemicals against 20 thousands proteins within 2 hours. Using the reconstructed genome-wide target profile as the fingerprint of a chemical compound, we predicted that seven FDA-approved drugs can be repurposed as novel anti-cancer therapies. The anti-cancer activity of six of them is supported by experimental evidences. Thus, REMAP is a valuable addition to the existing in silico toolbox for drug target identification, drug repurposing, phenotypic screening, and

  10. Large-Scale Off-Target Identification Using Fast and Accurate Dual Regularized One-Class Collaborative Filtering and Its Application to Drug Repurposing

    PubMed Central

    Poleksic, Aleksandar; Yao, Yuan; Tong, Hanghang; Meng, Patrick; Xie, Lei

    2016-01-01

    Target-based screening is one of the major approaches in drug discovery. Besides the intended target, unexpected drug off-target interactions often occur, and many of them have not been recognized and characterized. The off-target interactions can be responsible for either therapeutic or side effects. Thus, identifying the genome-wide off-targets of lead compounds or existing drugs will be critical for designing effective and safe drugs, and providing new opportunities for drug repurposing. Although many computational methods have been developed to predict drug-target interactions, they are either less accurate than the one that we are proposing here or computationally too intensive, thereby limiting their capability for large-scale off-target identification. In addition, the performances of most machine learning based algorithms have been mainly evaluated to predict off-target interactions in the same gene family for hundreds of chemicals. It is not clear how these algorithms perform in terms of detecting off-targets across gene families on a proteome scale. Here, we are presenting a fast and accurate off-target prediction method, REMAP, which is based on a dual regularized one-class collaborative filtering algorithm, to explore continuous chemical space, protein space, and their interactome on a large scale. When tested in a reliable, extensive, and cross-gene family benchmark, REMAP outperforms the state-of-the-art methods. Furthermore, REMAP is highly scalable. It can screen a dataset of 200 thousands chemicals against 20 thousands proteins within 2 hours. Using the reconstructed genome-wide target profile as the fingerprint of a chemical compound, we predicted that seven FDA-approved drugs can be repurposed as novel anti-cancer therapies. The anti-cancer activity of six of them is supported by experimental evidences. Thus, REMAP is a valuable addition to the existing in silico toolbox for drug target identification, drug repurposing, phenotypic screening, and

  11. Systems-level modeling of mycobacterial metabolism for the identification of new (multi-)drug targets.

    PubMed

    Rienksma, Rienk A; Suarez-Diez, Maria; Spina, Lucie; Schaap, Peter J; Martins dos Santos, Vitor A P

    2014-12-01

    Systems-level metabolic network reconstructions and the derived constraint-based (CB) mathematical models are efficient tools to explore bacterial metabolism. Approximately one-fourth of the Mycobacterium tuberculosis (Mtb) genome contains genes that encode proteins directly involved in its metabolism. These represent potential drug targets that can be systematically probed with CB models through the prediction of genes essential (or the combination thereof) for the pathogen to grow. However, gene essentiality depends on the growth conditions and, so far, no in vitro model precisely mimics the host at the different stages of mycobacterial infection, limiting model predictions. These limitations can be circumvented by combining expression data from in vivo samples with a validated CB model, creating an accurate description of pathogen metabolism in the host. To this end, we present here a thoroughly curated and extended genome-scale CB metabolic model of Mtb quantitatively validated using 13C measurements. We describe some of the efforts made in integrating CB models and high-throughput data to generate condition specific models, and we will discuss challenges ahead. This knowledge and the framework herein presented will enable to identify potential new drug targets, and will foster the development of optimal therapeutic strategies. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

  12. Identification of the feedforward component in manual control with predictable target signals.

    PubMed

    Drop, Frank M; Pool, Daan M; Damveld, Herman J; van Paassen, Marinus M; Mulder, Max

    2013-12-01

    In the manual control of a dynamic system, the human controller (HC) often follows a visible and predictable reference path. Compared with a purely feedback control strategy, performance can be improved by making use of this knowledge of the reference. The operator could effectively introduce feedforward control in conjunction with a feedback path to compensate for errors, as hypothesized in literature. However, feedforward behavior has never been identified from experimental data, nor have the hypothesized models been validated. This paper investigates human control behavior in pursuit tracking of a predictable reference signal while being perturbed by a quasi-random multisine disturbance signal. An experiment was done in which the relative strength of the target and disturbance signals were systematically varied. The anticipated changes in control behavior were studied by means of an ARX model analysis and by fitting three parametric HC models: two different feedback models and a combined feedforward and feedback model. The ARX analysis shows that the experiment participants employed control action on both the error and the target signal. The control action on the target was similar to the inverse of the system dynamics. Model fits show that this behavior can be modeled best by the combined feedforward and feedback model.

  13. Identification of highly effective target genes for RNAi-mediated control of emerald ash borer, Agrilus planipennis.

    PubMed

    Rodrigues, Thais B; Duan, Jian J; Palli, Subba R; Rieske, Lynne K

    2018-03-22

    Recent study has shown that RNA interference (RNAi) is efficient in emerald ash borer (EAB), Agrilus planipennis, and that ingestion of double-stranded RNA (dsRNA) targeting specific genes causes gene silencing and mortality in neonates. Here, we report on the identification of highly effective target genes for RNAi-mediated control of EAB. We screened 13 candidate genes in neonate larvae and selected the most effective target genes for further investigation, including their effect on EAB adults and on a non-target organism, Tribolium castaneum. The two most efficient target genes selected, hsp (heat shock 70-kDa protein cognate 3) and shi (shibire), caused up to 90% mortality of larvae and adults. In EAB eggs, larvae, and adults, the hsp is expressed at higher levels when compared to that of shi. Ingestion of dsHSP and dsSHI caused mortality in both neonate larvae and adults. Administration of a mixture of both dsRNAs worked better than either dsRNA by itself. In contrast, injection of EAB.dsHSP and EAB.dsSHI did not cause mortality in T. castaneum. Thus, the two genes identified cause high mortality in the EAB with no apparent phenotype effects in a non-target organism, the red flour beetle, and could be used in RNAi-mediated control of this invasive pest.

  14. Fast identification of dermatophytes by MALDI-TOF/MS using direct transfer of fungal cells on ground steel target plates.

    PubMed

    da Cunha, Keith C; Riat, Arnaud; Normand, Anne-Cecile; Bosshard, Philipp P; de Almeida, Margarete T G; Piarroux, Renaud; Schrenzel, Jacques; Fontao, Lionel

    2018-05-15

    Dermatophytes cause human infections limited to keratinized tissues. We showed that the direct transfer method allows reliable identification of non-dermatophytes mould and yeast by MALDI-TOF/MS. We aimed at assessing whether the direct transfer method can be used for dermatophytes and whether an own mass spectra library would be superior to the Bruker library. We used the Bruker Biotyper to build a dermatophyte mass spectra library and assessed its performance by 1/ testing a panel of mass spectrum produced with strains genotypically identified and, 2/ comparing MALDI-TOF/MS identification to morphology-based methods. Identification of dermatophytes using the Bruker library is poor. Our library provided 97% concordance between ITS sequencing and MALDI-TOF/MS analysis with a panel of 1104 spectra corresponding to 276 strains. Direct transfer method using unpolished target plates allowed proper identification of 85% of dermatophytes clinical isolates most of which were common dermatophytes. A homemade dermatophyte MSP library is a prerequisite for accurate identification of species absent in the Bruker library but it also improves identification of species already listed in the database. The direct deposit method can be used to identify the most commonly found dermatophytes such as T. rubrum and T. interdigitale/mentagrophytes by MALDI-TOF/MS. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  15. Development and content validity of a screening instrument for gaming addiction in adolescents: the Gaming Addiction Identification Test (GAIT).

    PubMed

    Vadlin, Sofia; Åslund, Cecilia; Nilsson, Kent W

    2015-08-01

    This study describes the development of a screening tool for gaming addiction in adolescents - the Gaming Addiction Identification Test (GAIT). Its development was based on the research literature on gaming and addiction. An expert panel comprising professional raters (n = 7), experiential adolescent raters (n = 10), and parent raters (n = 10) estimated the content validity of each item (I-CVI) as well as of the whole scale (S-CVI/Ave), and participated in a cognitive interview about the GAIT scale. The mean scores for both I-CVI and S-CVI/Ave ranged between 0.97 and 0.99 compared with the lowest recommended I-CVI value of 0.78 and the S-CVI/Ave value of 0.90. There were no sex differences and no differences between expert groups regarding ratings in content validity. No differences in the overall evaluation of the scale emerged in the cognitive interviews. Our conclusions were that GAIT showed good content validity in capturing gaming addiction. The GAIT needs further investigation into its psychometric properties of construct validity (convergent and divergent validity) and criterion-related validity, as well as its reliability in both clinical settings and in community settings with adolescents. © 2015 Scandinavian Psychological Associations and John Wiley & Sons Ltd.

  16. Identification of GPC2 as an Oncoprotein and Candidate Immunotherapeutic Target in High-Risk Neuroblastoma.

    PubMed

    Bosse, Kristopher R; Raman, Pichai; Zhu, Zhongyu; Lane, Maria; Martinez, Daniel; Heitzeneder, Sabine; Rathi, Komal S; Kendsersky, Nathan M; Randall, Michael; Donovan, Laura; Morrissy, Sorana; Sussman, Robyn T; Zhelev, Doncho V; Feng, Yang; Wang, Yanping; Hwang, Jennifer; Lopez, Gonzalo; Harenza, Jo Lynne; Wei, Jun S; Pawel, Bruce; Bhatti, Tricia; Santi, Mariarita; Ganguly, Arupa; Khan, Javed; Marra, Marco A; Taylor, Michael D; Dimitrov, Dimiter S; Mackall, Crystal L; Maris, John M

    2017-09-11

    We developed an RNA-sequencing-based pipeline to discover differentially expressed cell-surface molecules in neuroblastoma that meet criteria for optimal immunotherapeutic target safety and efficacy. Here, we show that GPC2 is a strong candidate immunotherapeutic target in this childhood cancer. We demonstrate high GPC2 expression in neuroblastoma due to MYCN transcriptional activation and/or somatic gain of the GPC2 locus. We confirm GPC2 to be highly expressed on most neuroblastomas, but not detectable at appreciable levels in normal childhood tissues. In addition, we demonstrate that GPC2 is required for neuroblastoma proliferation. Finally, we develop a GPC2-directed antibody-drug conjugate that is potently cytotoxic to GPC2-expressing neuroblastoma cells. Collectively, these findings validate GPC2 as a non-mutated neuroblastoma oncoprotein and candidate immunotherapeutic target. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Predicting glycerophosphoinositol identities in lipidomic datasets using VaLID (Visualization and Phospholipid Identification)--an online bioinformatic search engine.

    PubMed

    McDowell, Graeme S V; Blanchard, Alexandre P; Taylor, Graeme P; Figeys, Daniel; Fai, Stephen; Bennett, Steffany A L

    2014-01-01

    The capacity to predict and visualize all theoretically possible glycerophospholipid molecular identities present in lipidomic datasets is currently limited. To address this issue, we expanded the search-engine and compositional databases of the online Visualization and Phospholipid Identification (VaLID) bioinformatic tool to include the glycerophosphoinositol superfamily. VaLID v1.0.0 originally allowed exact and average mass libraries of 736,584 individual species from eight phospholipid classes: glycerophosphates, glyceropyrophosphates, glycerophosphocholines, glycerophosphoethanolamines, glycerophosphoglycerols, glycerophosphoglycerophosphates, glycerophosphoserines, and cytidine 5'-diphosphate 1,2-diacyl-sn-glycerols to be searched for any mass to charge value (with adjustable tolerance levels) under a variety of mass spectrometry conditions. Here, we describe an update that now includes all possible glycerophosphoinositols, glycerophosphoinositol monophosphates, glycerophosphoinositol bisphosphates, and glycerophosphoinositol trisphosphates. This update expands the total number of lipid species represented in the VaLID v2.0.0 database to 1,473,168 phospholipids. Each phospholipid can be generated in skeletal representation. A subset of species curated by the Canadian Institutes of Health Research Training Program in Neurodegenerative Lipidomics (CTPNL) team is provided as an array of high-resolution structures. VaLID is freely available and responds to all users through the CTPNL resources web site.

  18. Development and Validation of a Novel Vancomycin Dosing Nomogram for Achieving High-Target Trough Levels at 2 Canadian Teaching Hospitals

    PubMed Central

    Thalakada, Rosanne; Legal, Michael; Lau, Tim T Y; Luey, Tiffany; Batterink, Josh; Ensom, Mary H H

    2012-01-01

    Background: Recent guidelines recommend a vancomycin trough (predose) level between 15 and 20 mg/L in the treatment of invasive gram-positive infections, but most initial dosing nomograms are designed to achieve lower targets (5–15 mg/L). Clinicians need guidance about appropriate initial dosing to achieve the higher target. Objective: To develop and validate a high-target vancomycin dosing nomogram to achieve trough levels of 15–20 mg/L. Methods: A retrospective study was conducted at 2 teaching hospitals, St Paul’s Hospital and Vancouver General Hospital in Vancouver, British Columbia. Patients who were treated with vancomycin between January 2008 and June 2010 and who had achieved a trough level of 14.5–20.5 mg/L were identified. Demographic and clinical data were collected. Multiple linear regression was used to develop a vancomycin dosing nomogram for each hospital site. An integrated nomogram was constructed by merging the data from the 2 hospitals. A unique set of patients at each institution was used for validating their respective nomograms and a pooled group of patients for validating the integrated nomogram. Predictive success was evaluated, and a nomogram was deemed significantly different from another nomogram if p < 0.05 via “χ2 testing. Results: Data from 78 patients at one hospital and 91 patients at the other were used in developing the respective institutional nomograms. For each hospital’s data set, both age and initial serum creatinine were significantly associated with the predicted dosing interval (p < 0.001). Validation in a total of 105 test patients showed that the integrated nomogram had a predictive success rate of 56%. Conclusions: A novel vancomycin dosing nomogram was developed and validated at 2 Canadian teaching hospitals. This integrated nomogram is a tool that clinicians can use in selecting appropriate initial vancomycin regimens on the basis of age and serum creatinine, to achieve high-target levels of 15–20 mg

  19. Emotional System for Military Target Identification

    DTIC Science & Technology

    2009-10-01

    algorithm [23], and used it to solve a facial recognition problem. In other works [24,25], we explored the potential of using emotional neural...other application areas, such as security ( facial recognition ) and medical (blood cell identification), can be also efficiently used in military...Application of an emotional neural network to facial recognition . Neural Computing and Applications, 18(4), 309-320. [25] Khashman, A. (2009). Blood cell

  20. Identification and characterization of DNAzymes targeting DNA methyltransferase I for suppressing bladder cancer proliferation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wang, Xiangbo; Zhang, Lu; Ding, Nianhua

    2015-05-29

    Epigenetic inactivation of genes plays a critical role in many important human diseases, especially in cancer. A core mechanism for epigenetic inactivation of the genes is methylation of CpG islands in genome DNA, which is catalyzed by DNA methyltransferases (DNMTs). The inhibition of DNMTs may lead to demethylation and expression of the silenced tumor suppressor genes. Although DNMT inhibitors are currently being developed as potential anticancer agents, only limited success is achieved due to substantial toxicity. Here, we utilized a multiplex selection system to generate efficient RNA-cleaving DNAzymes targeting DNMT1. The lead molecule from the selection was shown to possessmore » efficient kinetic profiles and high efficiency in inhibiting the enzyme activity. Transfection of the DNAzyme caused significant down-regulation of DNMT1 expression and reactivation of p16 gene, resulting in reduced cell proliferation of bladder cancers. This study provides an alternative for targeting DNMTs for potential cancer therapy. - Highlights: • Identified DNMT1-targeted DNAzymes by multiplex selection system. • Biochemically characterized a lead DNAzyme with high kinetic efficiency. • Validated DNMT1-targeted DNAzyme in its enzymatic and cellular activities.« less

  1. Development and validation of an extended database for yeast identification by MALDI-TOF MS in Argentina.

    PubMed

    Taverna, Constanza Giselle; Mazza, Mariana; Bueno, Nadia Soledad; Alvarez, Christian; Amigot, Susana; Andreani, Mariana; Azula, Natalia; Barrios, Rubén; Fernández, Norma; Fox, Barbara; Guelfand, Liliana; Maldonado, Ivana; Murisengo, Omar Alejandro; Relloso, Silvia; Vivot, Matias; Davel, Graciela

    2018-05-11

    Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) has revolutionized the identification of microorganisms in clinical laboratories because it is rapid, relatively simple to use, accurate, and can be used for a wide number of microorganisms. Several studies have demonstrated the utility of this technique in the identification of yeasts; however, its performance is usually improved by the extension of the database. Here we developed an in-house database of 143 strains belonging to 42 yeast species in the MALDI Biotyper platform, and we validated the extended database with 388 regional strains and 15 reference strains belonging to 55 yeast species. We also performed an intra- and interlaboratory study to assess reproducibility and analyzed the use of the cutoff values of 1.700 and 2.000 to correctly identify at species level. The creation of an in-house database that extended the manufacturer's database was successful in view of no incorrect identification was introduced. The best performance was observed by using the extended database and a cutoff value of 1.700 with a sensitivity of .94 and specificity of .96. A reproducibility study showed utility to detect deviations and could be used for external quality control. The extended database was able to differentiate closely related species and it has potential in distinguishing the molecular genotypes of Cryptococcus neoformans and Cryptococcus gattii.

  2. An Assessment of Database-Validated microRNA Target Genes in Normal Colonic Mucosa: Implications for Pathway Analysis.

    PubMed

    Slattery, Martha L; Herrick, Jennifer S; Stevens, John R; Wolff, Roger K; Mullany, Lila E

    2017-01-01

    Determination of functional pathways regulated by microRNAs (miRNAs), while an essential step in developing therapeutics, is challenging. Some miRNAs have been studied extensively; others have limited information. In this study, we focus on 254 miRNAs previously identified as being associated with colorectal cancer and their database-identified validated target genes. We use RNA-Seq data to evaluate messenger RNA (mRNA) expression for 157 subjects who also had miRNA expression data. In the replication phase of the study, we replicated associations between 254 miRNAs associated with colorectal cancer and mRNA expression of database-identified target genes in normal colonic mucosa. In the discovery phase of the study, we evaluated expression of 18 miR-NAs (those with 20 or fewer database-identified target genes along with miR-21-5p, miR-215-5p, and miR-124-3p which have more than 500 database-identified target genes) with expression of 17 434 mRNAs to identify new targets in colon tissue. Seed region matches between miRNA and newly identified targeted mRNA were used to help determine direct miRNA-mRNA associations. From the replication of the 121 miRNAs that had at least 1 database-identified target gene using mRNA expression methods, 97.9% were expressed in normal colonic mucosa. Of the 8622 target miRNA-mRNA associations identified in the database, 2658 (30.2%) were associated with gene expression in normal colonic mucosa after adjusting for multiple comparisons. Of the 133 miRNAs with database-identified target genes by non-mRNA expression methods, 97.2% were expressed in normal colonic mucosa. After adjustment for multiple comparisons, 2416 miRNA-mRNA associations remained significant (19.8%). Results from the discovery phase based on detailed examination of 18 miRNAs identified more than 80 000 miRNA-mRNA associations that had not previously linked to the miRNA. Of these miRNA-mRNA associations, 15.6% and 14.8% had seed matches for CRCh38 and CRCh37

  3. F-18 High Alpha Research Vehicle (HARV) parameter identification flight test maneuvers for optimal input design validation and lateral control effectiveness

    NASA Technical Reports Server (NTRS)

    Morelli, Eugene A.

    1995-01-01

    Flight test maneuvers are specified for the F-18 High Alpha Research Vehicle (HARV). The maneuvers were designed for open loop parameter identification purposes, specifically for optimal input design validation at 5 degrees angle of attack, identification of individual strake effectiveness at 40 and 50 degrees angle of attack, and study of lateral dynamics and lateral control effectiveness at 40 and 50 degrees angle of attack. Each maneuver is to be realized by applying square wave inputs to specific control effectors using the On-Board Excitation System (OBES). Maneuver descriptions and complete specifications of the time/amplitude points define each input are included, along with plots of the input time histories.

  4. Identification of anti-cancer targets of eco-friendly waste Punica granatum peel by dual reverse virtual screening and binding analysis.

    PubMed

    Usha, Talambedu; Goyal, Arvind Kumar; Lubna, Syed; Prashanth, Hp; Mohan, T Madhan; Pande, Veena; Middha, Sushil Kumar

    2014-01-01

    Punica granatum (family: Lythraceae) is mainly found in Iran, which is considered to be its primary centre of origin. Studies on pomegranate peel have revealed antioxidant, anti-inflammatory, anti- angiogenesis activities, with prevention of premature aging and reducing inflammation. In addition to this it is also useful in treating various diseases like diabetes, maintaining blood pressure and treatment of neoplasms such as prostate and breast cancer. In this study we identified anti-cancer targets of active compounds like corilagin (tannins), quercetin (flavonoids) and pseudopelletierine (alkaloids) present in pomegranate peel by employing dual reverse screening and binding analysis. The potent targets of the pomegranate peel were annotated by the PharmMapper and ReverseScreen 3D, then compared with targets identified from different Bioassay databases (NPACT and HIT's). Docking was then further employed using AutoDock pyrx and validated through discovery studio for studying molecular interactions. A number of potent anti-cancerous targets were attained from the PharmMapper server according to their fit score and from ReverseScreen 3D server according to decreasing 3D scores. The identified targets now need to be further validated through in vitro and in vivo studies.

  5. Electro-Optic Identification Research Program

    DTIC Science & Technology

    2002-04-01

    Electro - optic identification (EOID) sensors provide photographic quality images that can be used to identify mine-like contacts provided by long...tasks such as validating existing electro - optic models, development of performance metrics, and development of computer aided identification and

  6. Transcriptome-wide identification of Rauvolfia serpentina microRNAs and prediction of their potential targets.

    PubMed

    Prakash, Pravin; Rajakani, Raja; Gupta, Vikrant

    2016-04-01

    MicroRNAs (miRNAs) are small non-coding RNAs of ∼ 19-24 nucleotides (nt) in length and considered as potent regulators of gene expression at transcriptional and post-transcriptional levels. Here we report the identification and characterization of 15 conserved miRNAs belonging to 13 families from Rauvolfia serpentina through in silico analysis of available nucleotide dataset. The identified mature R. serpentina miRNAs (rse-miRNAs) ranged between 20 and 22nt in length, and the average minimal folding free energy index (MFEI) value of rse-miRNA precursor sequences was found to be -0.815 kcal/mol. Using the identified rse-miRNAs as query, their potential targets were predicted in R. serpentina and other plant species. Gene Ontology (GO) annotation showed that predicted targets of rse-miRNAs include transcription factors as well as genes involved in diverse biological processes such as primary and secondary metabolism, stress response, disease resistance, growth, and development. Few rse-miRNAs were predicted to target genes of pharmaceutically important secondary metabolic pathways such as alkaloids and anthocyanin biosynthesis. Phylogenetic analysis showed the evolutionary relationship of rse-miRNAs and their precursor sequences to homologous pre-miRNA sequences from other plant species. The findings under present study besides giving first hand information about R. serpentina miRNAs and their targets, also contributes towards the better understanding of miRNA-mediated gene regulatory processes in plants. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Creation and Validation of Sintered PTFE BRDF Targets & Standards

    PubMed Central

    Durell, Christopher; Scharpf, Dan; McKee, Greg; L’Heureux, Michelle; Georgiev, Georgi; Obein, Gael; Cooksey, Catherine

    2016-01-01

    Sintered polytetrafluoroethylene (PTFE) is an extremely stable, near-perfect Lambertian reflecting diffuser and calibration standard material that has been used by national labs, space, aerospace and commercial sectors for over two decades. New uncertainty targets of 2 % on-orbit absolute validation in the Earth Observing Systems community have challenged the industry to improve is characterization and knowledge of almost every aspect of radiometric performance (space and ground). Assuming “near perfect” reflectance for angular dependent measurements is no longer going to suffice for many program needs. The total hemispherical spectral reflectance provides a good mark of general performance; but, without the angular characterization of bidirectional reflectance distribution function (BRDF) measurements, critical data is missing from many applications and uncertainty budgets. Therefore, traceable BRDF measurement capability is needed to characterize sintered PTFE’s angular response and provide a full uncertainty profile to users. This paper presents preliminary comparison measurements of the BRDF of sintered PTFE from several laboratories to better quantify the BRDF of sintered PTFE, assess the BRDF measurement comparability between laboratories, and improve estimates of measurement uncertainties under laboratory conditions. PMID:26900206

  8. Creation and Validation of Sintered PTFE BRDF Targets & Standards.

    PubMed

    Durell, Christopher; Scharpf, Dan; McKee, Greg; L'Heureux, Michelle; Georgiev, Georgi; Obein, Gael; Cooksey, Catherine

    2015-09-21

    Sintered polytetrafluoroethylene (PTFE) is an extremely stable, near-perfect Lambertian reflecting diffuser and calibration standard material that has been used by national labs, space, aerospace and commercial sectors for over two decades. New uncertainty targets of 2 % on-orbit absolute validation in the Earth Observing Systems community have challenged the industry to improve is characterization and knowledge of almost every aspect of radiometric performance (space and ground). Assuming "near perfect" reflectance for angular dependent measurements is no longer going to suffice for many program needs. The total hemispherical spectral reflectance provides a good mark of general performance; but, without the angular characterization of bidirectional reflectance distribution function (BRDF) measurements, critical data is missing from many applications and uncertainty budgets. Therefore, traceable BRDF measurement capability is needed to characterize sintered PTFE's angular response and provide a full uncertainty profile to users. This paper presents preliminary comparison measurements of the BRDF of sintered PTFE from several laboratories to better quantify the BRDF of sintered PTFE, assess the BRDF measurement comparability between laboratories, and improve estimates of measurement uncertainties under laboratory conditions.

  9. In silico prediction of novel therapeutic targets using gene-disease association data.

    PubMed

    Ferrero, Enrico; Dunham, Ian; Sanseau, Philippe

    2017-08-29

    Target identification and validation is a pressing challenge in the pharmaceutical industry, with many of the programmes that fail for efficacy reasons showing poor association between the drug target and the disease. Computational prediction of successful targets could have a considerable impact on attrition rates in the drug discovery pipeline by significantly reducing the initial search space. Here, we explore whether gene-disease association data from the Open Targets platform is sufficient to predict therapeutic targets that are actively being pursued by pharmaceutical companies or are already on the market. To test our hypothesis, we train four different classifiers (a random forest, a support vector machine, a neural network and a gradient boosting machine) on partially labelled data and evaluate their performance using nested cross-validation and testing on an independent set. We then select the best performing model and use it to make predictions on more than 15,000 genes. Finally, we validate our predictions by mining the scientific literature for proposed therapeutic targets. We observe that the data types with the best predictive power are animal models showing a disease-relevant phenotype, differential expression in diseased tissue and genetic association with the disease under investigation. On a test set, the neural network classifier achieves over 71% accuracy with an AUC of 0.76 when predicting therapeutic targets in a semi-supervised learning setting. We use this model to gain insights into current and failed programmes and to predict 1431 novel targets, of which a highly significant proportion has been independently proposed in the literature. Our in silico approach shows that data linking genes and diseases is sufficient to predict novel therapeutic targets effectively and confirms that this type of evidence is essential for formulating or strengthening hypotheses in the target discovery process. Ultimately, more rapid and automated target

  10. Genomic identification of potential targets unique to Candida albicans for the discovery of antifungal agents.

    PubMed

    Tripathi, Himanshu; Luqman, Suaib; Meena, Abha; Khan, Feroz

    2014-01-01

    Despite of modern antifungal therapy, the mortality rates of invasive infection with human fungal pathogen Candida albicans are up to 40%. Studies suggest that drug resistance in the three most common species of human fungal pathogens viz., C. albicans, Aspergillus fumigatus (causing mortality rate up to 90%) and Cryptococcus neoformans (causing mortality rate up to 70%) is due to mutations in the target enzymes or high expression of drug transporter genes. Drug resistance in human fungal pathogens has led to an imperative need for the identification of new targets unique to fungal pathogens. In the present study, we have used a comparative genomics approach to find out potential target proteins unique to C. albicans, an opportunistic fungus responsible for severe infection in immune-compromised human. Interestingly, many target proteins of existing antifungal agents showed orthologs in human cells. To identify unique proteins, we have compared proteome of C. albicans [SC5314] i.e., 14,633 total proteins retrieved from the RefSeq database of NCBI, USA with proteome of human and non-pathogenic yeast Saccharomyces cerevisiae. Results showed that 4,568 proteins were identified unique to C. albicans as compared to those of human and later when these unique proteins were compared with S. cerevisiae proteome, finally 2,161 proteins were identified as unique proteins and after removing repeats total 1,618 unique proteins (42 functionally known, 1,566 hypothetical and 10 unknown) were selected as potential antifungal drug targets unique to C. albicans.

  11. System Identification Methods for Aircraft Flight Control Development and Validation

    DOT National Transportation Integrated Search

    1995-10-01

    System-identification methods compose a mathematical model, or series of models, : from measurements of inputs and outputs of dynamic systems. This paper : discusses the use of frequency-domain system-identification methods for the : development and ...

  12. Computer aided manual validation of mass spectrometry-based proteomic data.

    PubMed

    Curran, Timothy G; Bryson, Bryan D; Reigelhaupt, Michael; Johnson, Hannah; White, Forest M

    2013-06-15

    Advances in mass spectrometry-based proteomic technologies have increased the speed of analysis and the depth provided by a single analysis. Computational tools to evaluate the accuracy of peptide identifications from these high-throughput analyses have not kept pace with technological advances; currently the most common quality evaluation methods are based on statistical analysis of the likelihood of false positive identifications in large-scale data sets. While helpful, these calculations do not consider the accuracy of each identification, thus creating a precarious situation for biologists relying on the data to inform experimental design. Manual validation is the gold standard approach to confirm accuracy of database identifications, but is extremely time-intensive. To palliate the increasing time required to manually validate large proteomic datasets, we provide computer aided manual validation software (CAMV) to expedite the process. Relevant spectra are collected, catalogued, and pre-labeled, allowing users to efficiently judge the quality of each identification and summarize applicable quantitative information. CAMV significantly reduces the burden associated with manual validation and will hopefully encourage broader adoption of manual validation in mass spectrometry-based proteomics. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Linking rare and common disease: mapping clinical disease-phenotypes to ontologies in therapeutic target validation.

    PubMed

    Sarntivijai, Sirarat; Vasant, Drashtti; Jupp, Simon; Saunders, Gary; Bento, A Patrícia; Gonzalez, Daniel; Betts, Joanna; Hasan, Samiul; Koscielny, Gautier; Dunham, Ian; Parkinson, Helen; Malone, James

    2016-01-01

    The Centre for Therapeutic Target Validation (CTTV - https://www.targetvalidation.org/) was established to generate therapeutic target evidence from genome-scale experiments and analyses. CTTV aims to support the validity of therapeutic targets by integrating existing and newly-generated data. Data integration has been achieved in some resources by mapping metadata such as disease and phenotypes to the Experimental Factor Ontology (EFO). Additionally, the relationship between ontology descriptions of rare and common diseases and their phenotypes can offer insights into shared biological mechanisms and potential drug targets. Ontologies are not ideal for representing the sometimes associated type relationship required. This work addresses two challenges; annotation of diverse big data, and representation of complex, sometimes associated relationships between concepts. Semantic mapping uses a combination of custom scripting, our annotation tool 'Zooma', and expert curation. Disease-phenotype associations were generated using literature mining on Europe PubMed Central abstracts, which were manually verified by experts for validity. Representation of the disease-phenotype association was achieved by the Ontology of Biomedical AssociatioN (OBAN), a generic association representation model. OBAN represents associations between a subject and object i.e., disease and its associated phenotypes and the source of evidence for that association. The indirect disease-to-disease associations are exposed through shared phenotypes. This was applied to the use case of linking rare to common diseases at the CTTV. EFO yields an average of over 80% of mapping coverage in all data sources. A 42% precision is obtained from the manual verification of the text-mined disease-phenotype associations. This results in 1452 and 2810 disease-phenotype pairs for IBD and autoimmune disease and contributes towards 11,338 rare diseases associations (merged with existing published work [Am J Hum Genet

  14. Panorama: A Targeted Proteomics Knowledge Base

    PubMed Central

    2015-01-01

    Panorama is a web application for storing, sharing, analyzing, and reusing targeted assays created and refined with Skyline,1 an increasingly popular Windows client software tool for targeted proteomics experiments. Panorama allows laboratories to store and organize curated results contained in Skyline documents with fine-grained permissions, which facilitates distributed collaboration and secure sharing of published and unpublished data via a web-browser interface. It is fully integrated with the Skyline workflow and supports publishing a document directly to a Panorama server from the Skyline user interface. Panorama captures the complete Skyline document information content in a relational database schema. Curated results published to Panorama can be aggregated and exported as chromatogram libraries. These libraries can be used in Skyline to pick optimal targets in new experiments and to validate peak identification of target peptides. Panorama is open-source and freely available. It is distributed as part of LabKey Server,2 an open source biomedical research data management system. Laboratories and organizations can set up Panorama locally by downloading and installing the software on their own servers. They can also request freely hosted projects on https://panoramaweb.org, a Panorama server maintained by the Department of Genome Sciences at the University of Washington. PMID:25102069

  15. Does methodology matter in eyewitness identification research? The effect of live versus video exposure on eyewitness identification accuracy.

    PubMed

    Pozzulo, Joanna D; Crescini, Charmagne; Panton, Tasha

    2008-01-01

    The present study examined the effect of mode of target exposure (live versus video) on eyewitness identification accuracy. Adult participants (N=104) were exposed to a staged crime that they witnessed either live or on videotape. Participants were then asked to rate their stress and arousal levels prior to being presented with either a target-present or -absent simultaneous lineup. Across target-present and -absent lineups, mode of target exposure did not have a significant effect on identification accuracy. However, mode of target exposure was found to have a significant effect on stress and arousal levels. Participants who witnessed the crime live had higher levels of stress and arousal than those who were exposed to the videotaped crime. A higher level of arousal was significantly related to poorer identification accuracy for those in the video condition. For participants in the live condition however, stress and arousal had no effect on eyewitness identification accuracy. Implications of these findings in regards to the generalizability of laboratory-based research on eyewitness testimony to real-life crime are discussed.

  16. Concealed identification symbols and nondestructive determination of the identification symbols

    DOEpatents

    Nance, Thomas A.; Gibbs, Kenneth M.

    2014-09-16

    The concealing of one or more identification symbols into a target object and the subsequent determination or reading of such symbols through non-destructive testing is described. The symbols can be concealed in a manner so that they are not visible to the human eye and/or cannot be readily revealed to the human eye without damage or destruction of the target object. The identification symbols can be determined after concealment by e.g., the compilation of multiple X-ray images. As such, the present invention can also provide e.g., a deterrent to theft and the recovery of lost or stolen objects.

  17. Identification of plant glutaredoxin targets.

    PubMed

    Rouhier, Nicolas; Villarejo, Arsenio; Srivastava, Manoj; Gelhaye, Eric; Keech, Olivier; Droux, Michel; Finkemeier, Iris; Samuelsson, Göran; Dietz, Karl Josef; Jacquot, Jean-Pierre; Wingsle, Gunnar

    2005-01-01

    Glutaredoxins (Grxs) are small ubiquitous proteins of the thioredoxin (Trx) family, which catalyze dithiol-disulfide exchange reactions or reduce protein-mixed glutathione disulfides. In plants, several Trx-interacting proteins have been isolated from different compartments, whereas very few Grx-interacting proteins are known. We describe here the determination of Grx target proteins using a mutated poplar Grx, various tissular and subcellular plant extracts, and liquid chromatography coupled to tandem mass spectrometry detection. We have identified 94 putative targets, involved in many processes, including oxidative stress response [peroxiredoxins (Prxs), ascorbate peroxidase, catalase], nitrogen, sulfur, and carbon metabolisms (methionine synthase, alanine aminotransferase, phosphoglycerate kinase), translation (elongation factors E and Tu), or protein folding (heat shock protein 70). Some of these proteins were previously found to interact with Trx or to be glutathiolated in other organisms, but others could be more specific partners of Grx. To substantiate further these data, Grx was shown to support catalysis of the stroma beta-type carbonic anhydrase and Prx IIF of Arabidopsis thaliana, but not of poplar 2-Cys Prx. Overall, these data suggest that the interaction could occur randomly either with exposed cysteinyl disulfide bonds formed within or between target proteins or with mixed disulfides between a protein thiol and glutathione.

  18. MRM validation of targeted nonglycosylated peptides from N-glycoprotein biomarkers using direct trypsin digestion of undepleted human plasma.

    PubMed

    Lee, Ju Yeon; Kim, Jin Young; Cheon, Mi Hee; Park, Gun Wook; Ahn, Yeong Hee; Moon, Myeong Hee; Yoo, Jong Shin

    2014-02-26

    A rapid, simple, and reproducible MRM-based validation method for serological glycoprotein biomarkers in clinical use was developed by targeting the nonglycosylated tryptic peptides adjacent to N-glycosylation sites. Since changes in protein glycosylation are known to be associated with a variety of diseases, glycoproteins have been major targets in biomarker discovery. We previously found that nonglycosylated tryptic peptides adjacent to N-glycosylation sites differed in concentration between normal and hepatocellular carcinoma (HCC) plasma due to differences in steric hindrance of the glycan moiety in N-glycoproteins to tryptic digestion (Lee et al., 2011). To increase the feasibility and applicability of clinical validation of biomarker candidates (nonglycosylated tryptic peptides), we developed a method to effectively monitor nonglycosylated tryptic peptides from a large number of plasma samples and to reduce the total analysis time with maximizing the effect of steric hindrance by the glycans during digestion of glycoproteins. The AUC values of targeted nonglycosylated tryptic peptides were excellent (0.955 for GQYCYELDEK, 0.880 for FEDGVLDPDYPR and 0.907 for TEDTIFLR), indicating that these could be effective biomarkers for hepatocellular carcinoma. This method provides the necessary throughput required to validate glycoprotein biomarkers, as well as quantitative accuracy for human plasma analysis, and should be amenable to clinical use. Difficulties in verifying and validating putative protein biomarkers are often caused by complex sample preparation procedures required to determine their concentrations in a large number of plasma samples. To solve the difficulties, we developed MRM-based protein biomarker assays that greatly reduce complex, time-consuming, and less reproducible sample pretreatment steps in plasma for clinical implementation. First, we used undepleted human plasma samples without any enrichment procedures. Using nanoLC/MS/MS, we targeted

  19. Chemical proteomics for target discovery of head-to-tail cyclized mini-proteins

    NASA Astrophysics Data System (ADS)

    Hellinger, Roland; Thell, Kathrin; Vasileva, Mina; Muhammad, Taj; Gunasekera, Sunithi; Kümmel, Daniel; Göransson, Ulf; Becker, Christian W.; Gruber, Christian W.

    2017-10-01

    Target deconvolution is one of the most challenging tasks in drug discovery, but a key step in drug development. In contrast to small molecules, there is a lack of validated and robust methodologies for target elucidation of peptides. In particular, it is difficult to apply these methods to cyclic and cysteine-stabilized peptides since they exhibit reduced amenability to chemical modification and affinity capture; however, such ribosomal synthesized and post-translationally modified peptide natural products are rich sources of promising drug candidates. For example, plant-derived circular peptides called cyclotides have recently attracted much attention due to their immunosuppressive effects and oral activity in the treatment of multiple sclerosis in mice, but their molecular target has hitherto not been reported. In this study a chemical proteomics approach using photo-affinity crosslinking was developed to determine a target of the circular peptide [T20K]kalata B1. Using this prototypic nature-derived peptide enabled the identification of a possible modulation of 14-3-3 proteins. This biochemical interaction was validated via competition pull down assays as well as a cellular reporter assay indicating an effect on 14-3-3-dependent transcriptional activity. As proof of concept, the presented approach may be applicable for target elucidation of various cyclic peptides and mini-proteins, in particular cyclotides, which represent a promising class of molecules in drug discovery and development.

  20. Active imaging system performance model for target acquisition

    NASA Astrophysics Data System (ADS)

    Espinola, Richard L.; Teaney, Brian; Nguyen, Quang; Jacobs, Eddie L.; Halford, Carl E.; Tofsted, David H.

    2007-04-01

    The U.S. Army RDECOM CERDEC Night Vision & Electronic Sensors Directorate has developed a laser-range-gated imaging system performance model for the detection, recognition, and identification of vehicle targets. The model is based on the established US Army RDECOM CERDEC NVESD sensor performance models of the human system response through an imaging system. The Java-based model, called NVLRG, accounts for the effect of active illumination, atmospheric attenuation, and turbulence effects relevant to LRG imagers, such as speckle and scintillation, and for the critical sensor and display components. This model can be used to assess the performance of recently proposed active SWIR systems through various trade studies. This paper will describe the NVLRG model in detail, discuss the validation of recent model components, present initial trade study results, and outline plans to validate and calibrate the end-to-end model with field data through human perception testing.

  1. A Statistics-based Platform for Quantitative N-terminome Analysis and Identification of Protease Cleavage Products*

    PubMed Central

    auf dem Keller, Ulrich; Prudova, Anna; Gioia, Magda; Butler, Georgina S.; Overall, Christopher M.

    2010-01-01

    Terminal amine isotopic labeling of substrates (TAILS), our recently introduced platform for quantitative N-terminome analysis, enables wide dynamic range identification of original mature protein N-termini and protease cleavage products. Modifying TAILS by use of isobaric tag for relative and absolute quantification (iTRAQ)-like labels for quantification together with a robust statistical classifier derived from experimental protease cleavage data, we report reliable and statistically valid identification of proteolytic events in complex biological systems in MS2 mode. The statistical classifier is supported by a novel parameter evaluating ion intensity-dependent quantification confidences of single peptide quantifications, the quantification confidence factor (QCF). Furthermore, the isoform assignment score (IAS) is introduced, a new scoring system for the evaluation of single peptide-to-protein assignments based on high confidence protein identifications in the same sample prior to negative selection enrichment of N-terminal peptides. By these approaches, we identified and validated, in addition to known substrates, low abundance novel bioactive MMP-2 targets including the plasminogen receptor S100A10 (p11) and the proinflammatory cytokine proEMAP/p43 that were previously undescribed. PMID:20305283

  2. Predictive validity of the identification of seniors at risk screening tool in a German emergency department setting.

    PubMed

    Singler, Katrin; Heppner, Hans Jürgen; Skutetzky, Andreas; Sieber, Cornel; Christ, Michael; Thiem, Ulrich

    2014-01-01

    The identification of patients at high risk for adverse outcomes [death, unplanned readmission to emergency department (ED)/hospital, functional decline] plays an important role in emergency medicine. The Identification of Seniors at Risk (ISAR) instrument is one of the most commonly used and best-validated screening tools. As to the authors' knowledge so far there are no data on any screening tool for the identification of older patients at risk for a negative outcome in Germany. To evaluate the validity of the ISAR screening tool in a German ED. This was a prospective single-center observational cohort study in an ED of an urban university-affiliated hospital. Participants were 520 patients aged ≥75 years consecutively admitted to the ED. The German version of the ISAR screening tool was administered directly after triage of the patients. Follow-up telephone interviews to assess outcome variables were conducted 28 and 180 days after the index visit in the ED. The primary end point was death from any cause or hospitalization or recurrent ED visit or change of residency into a long-term care facility on day 28 after the index ED visit. The mean age ± SD was 82.8 ± 5.0 years. According to ISAR, 425 patients (81.7%) scored ≥2 points, and 315 patients (60.5%) scored ≥3 points. The combined primary end point was observed in 250 of 520 patients (48.1%) on day 28 and in 260 patients (50.0%) on day 180. Using a continuous ISAR score the area under the curve on day 28 was 0.621 (95% confidence interval, CI 0.573-0.669) and 0.661 (95% CI 0.615-0.708) on day 180, respectively. The German version of the ISAR screening tool acceptably identified elderly patients in the ED with an increased risk of a negative outcome. Using the cutoff ≥3 points instead of ≥2 points yielded better overall results.

  3. Early Identification of Children at Risk for Academic Difficulties Using Standardized Assessment: Stability and Predictive Validity of Preschool Math and Language Scores

    ERIC Educational Resources Information Center

    Frans, Niek; Post, Wendy J.; Huisman, Mark; Oenema-Mostert, Ineke C. E.; Keegstra, Anne L.; Minnaert, Alexander E. M. G.

    2017-01-01

    Despite the claim by several researchers that variability in performance may complicate the identification of "at-risk" children, variability in the academic performance of young children remains an undervalued area of research. The goal of this study is to examine the predictive validity for future scores and the score stability of two…

  4. Nuclear Security: Target Analysis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Singh, Surinder Paul; Gibbs, Philip W.; Bultz, Garl A.

    2014-03-01

    This objectives of this session were to understand the basic steps of target identification; describe the SNRI targets in detail; characterize specific targets with more detail; prioritize targets based on guidance documents; understand the graded safeguards concept; identify roll up and understand why it is a concern; and recognize the category for different materials.

  5. Clinical Validation of Targeted Next Generation Sequencing for Colon and Lung Cancers

    PubMed Central

    D’Haene, Nicky; Le Mercier, Marie; De Nève, Nancy; Blanchard, Oriane; Delaunoy, Mélanie; El Housni, Hakim; Dessars, Barbara; Heimann, Pierre; Remmelink, Myriam; Demetter, Pieter; Tejpar, Sabine; Salmon, Isabelle

    2015-01-01

    Objective Recently, Next Generation Sequencing (NGS) has begun to supplant other technologies for gene mutation testing that is now required for targeted therapies. However, transfer of NGS technology to clinical daily practice requires validation. Methods We validated the Ion Torrent AmpliSeq Colon and Lung cancer panel interrogating 1850 hotspots in 22 genes using the Ion Torrent Personal Genome Machine. First, we used commercial reference standards that carry mutations at defined allelic frequency (AF). Then, 51 colorectal adenocarcinomas (CRC) and 39 non small cell lung carcinomas (NSCLC) were retrospectively analyzed. Results Sensitivity and accuracy for detecting variants at an AF >4% was 100% for commercial reference standards. Among the 90 cases, 89 (98.9%) were successfully sequenced. Among the 86 samples for which NGS and the reference test were both informative, 83 showed concordant results between NGS and the reference test; i.e. KRAS and BRAF for CRC and EGFR for NSCLC, with the 3 discordant cases each characterized by an AF <10%. Conclusions Overall, the AmpliSeq colon/lung cancer panel was specific and sensitive for mutation analysis of gene panels and can be incorporated into clinical daily practice. PMID:26366557

  6. Identification and Validation of Small-Gatekeeper Kinases as Drug Targets in Giardia lamblia

    PubMed Central

    Hennessey, Kelly M.; Smith, Tess R.; Xu, Jennifer W.; Alas, Germain C. M.; Ojo, Kayode K.; Merritt, Ethan A.

    2016-01-01

    Giardiasis is widely acknowledged to be a neglected disease in need of new therapeutics to address toxicity and resistance issues associated with the limited available treatment options. We examined seven protein kinases in the Giardia lamblia genome that are predicted to share an unusual structural feature in their active site. This feature, an expanded active site pocket resulting from an atypically small gatekeeper residue, confers sensitivity to “bumped” kinase inhibitors (BKIs), a class of compounds that has previously shown good pharmacological properties and minimal toxicity. An initial phenotypic screen for biological activity using a subset of an in-house BKI library found that 5 of the 36 compounds tested reduced trophozoite growth by at least 50% at a concentration of 5 μM. The cellular localization and the relative expression levels of the seven protein kinases of interest were determined after endogenously tagging the kinases. Essentiality of these kinases for parasite growth and infectivity were evaluated genetically using morpholino knockdown of protein expression to establish those that could be attractive targets for drug design. Two of the kinases were critical for trophozoite growth and attachment. Therefore, recombinant enzymes were expressed, purified and screened against a BKI library of >400 compounds in thermal stability assays in order to identify high affinity compounds. Compounds with substantial thermal stabilization effects on recombinant protein were shown to have good inhibition of cell growth in wild-type G. lamblia and metronidazole-resistant strains of G. lamblia. Our data suggest that BKIs are a promising starting point for the development of new anti-giardiasis therapeutics that do not overlap in mechanism with current drugs. PMID:27806042

  7. Exploring the Process of Energy Generation in Pathophysiology by Targeted Metabolomics: Performance of a Simple and Quantitative Method.

    PubMed

    Riera-Borrull, Marta; Rodríguez-Gallego, Esther; Hernández-Aguilera, Anna; Luciano, Fedra; Ras, Rosa; Cuyàs, Elisabet; Camps, Jordi; Segura-Carretero, Antonio; Menendez, Javier A; Joven, Jorge; Fernández-Arroyo, Salvador

    2016-01-01

    Abnormalities in mitochondrial metabolism and regulation of energy balance contribute to human diseases. The consequences of high fat and other nutrient intake, and the resulting acquired mitochondrial dysfunction, are essential to fully understand common disorders, including obesity, cancer, and atherosclerosis. To simultaneously and noninvasively measure and quantify indirect markers of mitochondrial function, we have developed a method based on gas chromatography coupled to quadrupole-time of flight mass spectrometry and an electron ionization interface, and validated the system using plasma from patients with peripheral artery disease, human cancer cells, and mouse tissues. This approach was used to increase sensibility in the measurement of a wide dynamic range and chemical diversity of multiple intermediate metabolites used in energy metabolism. We demonstrate that our targeted metabolomics method allows for quick and accurate identification and quantification of molecules, including the measurement of small yet significant biological changes in experimental samples. The apparently low process variability required for its performance in plasma, cell lysates, and tissues allowed a rapid identification of correlations between interconnected pathways. Our results suggest that delineating the process of energy generation by targeted metabolomics can be a valid surrogate for predicting mitochondrial dysfunction in biological samples. Importantly, when used in plasma, targeted metabolomics should be viewed as a robust and noninvasive source of biomarkers in specific pathophysiological scenarios.

  8. Exploring the Process of Energy Generation in Pathophysiology by Targeted Metabolomics: Performance of a Simple and Quantitative Method

    NASA Astrophysics Data System (ADS)

    Riera-Borrull, Marta; Rodríguez-Gallego, Esther; Hernández-Aguilera, Anna; Luciano, Fedra; Ras, Rosa; Cuyàs, Elisabet; Camps, Jordi; Segura-Carretero, Antonio; Menendez, Javier A.; Joven, Jorge; Fernández-Arroyo, Salvador

    2016-01-01

    Abnormalities in mitochondrial metabolism and regulation of energy balance contribute to human diseases. The consequences of high fat and other nutrient intake, and the resulting acquired mitochondrial dysfunction, are essential to fully understand common disorders, including obesity, cancer, and atherosclerosis. To simultaneously and noninvasively measure and quantify indirect markers of mitochondrial function, we have developed a method based on gas chromatography coupled to quadrupole-time of flight mass spectrometry and an electron ionization interface, and validated the system using plasma from patients with peripheral artery disease, human cancer cells, and mouse tissues. This approach was used to increase sensibility in the measurement of a wide dynamic range and chemical diversity of multiple intermediate metabolites used in energy metabolism. We demonstrate that our targeted metabolomics method allows for quick and accurate identification and quantification of molecules, including the measurement of small yet significant biological changes in experimental samples. The apparently low process variability required for its performance in plasma, cell lysates, and tissues allowed a rapid identification of correlations between interconnected pathways. Our results suggest that delineating the process of energy generation by targeted metabolomics can be a valid surrogate for predicting mitochondrial dysfunction in biological samples. Importantly, when used in plasma, targeted metabolomics should be viewed as a robust and noninvasive source of biomarkers in specific pathophysiological scenarios.

  9. Culture-Independent Identification of Periodontitis-Associated Porphyromonas and Tannerella Populations by Targeted Molecular Analysis

    PubMed Central

    de Lillo, A.; Booth, V.; Kyriacou, L.; Weightman, A. J.; Wade, W. G.

    2004-01-01

    Periodontitis is the commonest bacterial disease of humans and is the major cause of adult tooth loss. About half of the oral microflora is unculturable; and 16S rRNA PCR, cloning, and sequencing techniques have demonstrated the high level of species richness of the oral microflora. In the present study, a PCR primer set specific for the genera Porphyromonas and Tannerella was designed and used to analyze the bacterial populations in subgingival plaque samples from inflamed shallow and deep sites in subjects with periodontitis and shallow sites in age- and sex-matched controls. A total of 308 clones were sequenced and found to belong to one of six Porphyromonas or Tannerella species or phylotypes, one of which, Porphyromonas P3, was novel. Tannerella forsythensis was found in significantly higher proportions in patients than in controls. Porphyromonas catoniae and Tannerella phylotype BU063 appeared to be associated with shallow sites. Targeted culture-independent molecular ecology studies have a valuable role to play in the identification of bacterial targets for further investigations of the pathogenesis of bacterial infections. PMID:15583276

  10. Mining, identification and function analysis of microRNAs and target genes in peanut (Arachis hypogaea L.).

    PubMed

    Zhang, Tingting; Hu, Shuhao; Yan, Caixia; Li, Chunjuan; Zhao, Xiaobo; Wan, Shubo; Shan, Shihua

    2017-02-01

    In the present investigation, a total of 60 conserved peanut (Arachis hypogaea L.) microRNA (miRNA) sequences, belonging to 16 families, were identified using bioinformatics methods. There were 392 target gene sequences, identified from 58 miRNAs with Target-align software and BLASTx analyses. Gene Ontology (GO) functional analysis suggested that these target genes were involved in mediating peanut growth and development, signal transduction and stress resistance. There were 55 miRNA sequences, verified employing a poly (A) tailing test, with a success rate of up to 91.67%. Twenty peanut target gene sequences were randomly selected, and the 5' rapid amplification of the cDNA ends (5'-RACE) method were used to validate the cleavage sites of these target genes. Of these, 14 (70%) peanut miRNA targets were verified by means of gel electrophoresis, cloning and sequencing. Furthermore, functional analysis and homologous sequence retrieval were conducted for target gene sequences, and 26 target genes were chosen as the objects for stress resistance experimental study. Real-time fluorescence quantitative PCR (qRT-PCR) technology was applied to measure the expression level of resistance-associated miRNAs and their target genes in peanut exposed to Aspergillus flavus (A. flavus) infection and drought stress, respectively. In consequence, 5 groups of miRNAs & targets were found accorded with the mode of miRNA negatively controlling the expression of target genes. This study, preliminarily determined the biological functions of some resistance-associated miRNAs and their target genes in peanut. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  11. Identification of Novel Pax8 Targets in FRTL-5 Thyroid Cells by Gene Silencing and Expression Microarray Analysis

    PubMed Central

    Di Palma, Tina; Conti, Anna; de Cristofaro, Tiziana; Scala, Serena; Nitsch, Lucio; Zannini, Mariastella

    2011-01-01

    Background The differentiation program of thyroid follicular cells (TFCs), by far the most abundant cell population of the thyroid gland, relies on the interplay between sequence-specific transcription factors and transcriptional coregulators with the basal transcriptional machinery of the cell. However, the molecular mechanisms leading to the fully differentiated thyrocyte are still the object of intense study. The transcription factor Pax8, a member of the Paired-box gene family, has been demonstrated to be a critical regulator required for proper development and differentiation of thyroid follicular cells. Despite being Pax8 well-characterized with respect to its role in regulating genes involved in thyroid differentiation, genomics approaches aiming at the identification of additional Pax8 targets are lacking and the biological pathways controlled by this transcription factor are largely unknown. Methodology/Principal Findings To identify unique downstream targets of Pax8, we investigated the genome-wide effect of Pax8 silencing comparing the transcriptome of silenced versus normal differentiated FRTL-5 thyroid cells. In total, 2815 genes were found modulated 72 h after Pax8 RNAi, induced or repressed. Genes previously reported to be regulated by Pax8 in FRTL-5 cells were confirmed. In addition, novel targets genes involved in functional processes such as DNA replication, anion transport, kinase activity, apoptosis and cellular processes were newly identified. Transcriptome analysis highlighted that Pax8 is a key molecule for thyroid morphogenesis and differentiation. Conclusions/Significance This is the first large-scale study aimed at the identification of new genes regulated by Pax8, a master regulator of thyroid development and differentiation. The biological pathways and target genes controlled by Pax8 will have considerable importance to understand thyroid disease progression as well as to set up novel therapeutic strategies. PMID:21966443

  12. Site selection and directional models of deserts used for ERBE validation targets

    NASA Technical Reports Server (NTRS)

    Staylor, W. F.

    1986-01-01

    Broadband shortwave and longwave radiance measurements obtained from the Nimbus 7 Earth Radiation Budget scanner were used to develop reflectance and emittance models for the Sahara, Gibson, and Saudi Deserts. These deserts will serve as in-flight validation targets for the Earth Radiation Budget Experiment being flown on the Earth Radiation Budget Satellite and two National Oceanic and Atmospheric Administration polar satellites. The directional reflectance model derived for the deserts was a function of the sum and product of the cosines of the solar and viewing zenith angles, and thus reciprocity existed between these zenith angles. The emittance model was related by a power law of the cosine of the viewing zenith angle.

  13. Identification of a novel LMF1 nonsense mutation responsible for severe hypertriglyceridemia by targeted next-generation sequencing.

    PubMed

    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.

  14. Meat Species Identification using Loop-mediated Isothermal Amplification Assay Targeting Species-specific Mitochondrial DNA.

    PubMed

    Cho, Ae-Ri; Dong, Hee-Jin; Cho, Seongbeom

    2014-01-01

    Meat source fraud and adulteration scandals have led to consumer demands for accurate meat identification methods. Nucleotide amplification assays have been proposed as an alternative method to protein-based assays for meat identification. In this study, we designed Loop-mediated isothermal amplification (LAMP) assays targeting species-specific mitochondrial DNA to identify and discriminate eight meat species; cattle, pig, horse, goat, sheep, chicken, duck, and turkey. The LAMP primer sets were designed and the target genes were discriminated according to their unique annealing temperature generated by annealing curve analysis. Their unique annealing temperatures were found to be 85.56±0.07℃ for cattle, 84.96±0.08℃ for pig, and 85.99±0.05℃ for horse in the BSE-LAMP set (Bos taurus, Sus scrofa domesticus and Equus caballus); 84.91±0.11℃ for goat and 83.90±0.11℃ for sheep in the CO-LAMP set (Capra hircus and Ovis aries); and 86.31±0.23℃ for chicken, 88.66±0.12℃ for duck, and 84.49±0.08℃ for turkey in the GAM-LAMP set (Gallus gallus, Anas platyrhynchos and Meleagris gallopavo). No cross-reactivity was observed in each set. The limits of detection (LODs) of the LAMP assays in raw and cooked meat were determined from 10 pg/μL to 100 fg/μL levels, and LODs in raw and cooked meat admixtures were determined from 0.01% to 0.0001% levels. The assays were performed within 30 min and showed greater sensitivity than that of the PCR assays. These novel LAMP assays provide a simple, rapid, accurate, and sensitive technology for discrimination of eight meat species.

  15. Identification of Novel Molecular Targets for Endometrial Cancer Using a Drill-Down LC-MS/MS Approach with iTRAQ

    PubMed Central

    Voisin, Sébastien N.; Krakovska, Olga; Matta, Ajay; DeSouza, Leroi V.; Romaschin, Alexander D.; Colgan, Terence J.; Siu, K. W. Michael

    2011-01-01

    Background The number of patients with endometrial carcinoma (EmCa) with advanced stage or high histological grade is increasing and prognosis has not improved for over the last decade. There is an urgent need for the discovery of novel molecular targets for diagnosis, prognosis and treatment of EmCa, which will have the potential to improve the clinical strategy and outcome of this disease. Methodology and Results We used a “drill-down” proteomics approach to facilitate the identification of novel molecular targets for diagnosis, prognosis and/or therapeutic intervention for EmCa. Based on peptide ions identified and their retention times in the first LC-MS/MS analysis, an exclusion list was generated for subsequent iterations. A total of 1529 proteins have been identified below the Proteinpilot® 5% error threshold from the seven sets of iTRAQ experiments performed. On average, the second iteration added 78% new peptides to those identified after the first run, while the third iteration added 36% additional peptides. Of the 1529 proteins identified, only 40 satisfied our criteria for significant differential expression in EmCa in comparison to normal proliferative tissues. These proteins included metabolic enzymes (pyruvate kinase M2 and lactate dehydrogenase A); calcium binding proteins (S100A6, calcyphosine and calumenin), and proteins involved in regulating inflammation, proliferation and invasion (annexin A1, interleukin enhancer-binding factor 3, alpha-1-antitrypsin, macrophage capping protein and cathepsin B). Network analyses revealed regulation of these molecular targets by c-myc, Her2/neu and TNF alpha, suggesting intervention with these pathways may be a promising strategy for the development of novel molecular targeted therapies for EmCa. Conclusions Our analyses revealed the significance of drill-down proteomics approach in combination with iTRAQ to overcome some of the limitations of current proteomics strategies. This study led to the

  16. System identification methods for aircraft flight control development and validation

    NASA Technical Reports Server (NTRS)

    Tischler, Mark B.

    1995-01-01

    System-identification methods compose a mathematical model, or series of models, from measurements of inputs and outputs of dynamic systems. The extracted models allow the characterization of the response of the overall aircraft or component subsystem behavior, such as actuators and on-board signal processing algorithms. This paper discusses the use of frequency-domain system-identification methods for the development and integration of aircraft flight-control systems. The extraction and analysis of models of varying complexity from nonparametric frequency-responses to transfer-functions and high-order state-space representations is illustrated using the Comprehensive Identification from FrEquency Responses (CIFER) system-identification facility. Results are presented for test data of numerous flight and simulation programs at the Ames Research Center including rotorcraft, fixed-wing aircraft, advanced short takeoff and vertical landing (ASTOVL), vertical/short takeoff and landing (V/STOL), tiltrotor aircraft, and rotor experiments in the wind tunnel. Excellent system characterization and dynamic response prediction is achieved for this wide class of systems. Examples illustrate the role of system-identification technology in providing an integrated flow of dynamic response data around the entire life-cycle of aircraft development from initial specifications, through simulation and bench testing, and into flight-test optimization.

  17. VIRmiRNA: a comprehensive resource for experimentally validated viral miRNAs and their targets.

    PubMed

    Qureshi, Abid; Thakur, Nishant; Monga, Isha; Thakur, Anamika; Kumar, Manoj

    2014-01-01

    Viral microRNAs (miRNAs) regulate gene expression of viral and/or host genes to benefit the virus. Hence, miRNAs play a key role in host-virus interactions and pathogenesis of viral diseases. Lately, miRNAs have also shown potential as important targets for the development of novel antiviral therapeutics. Although several miRNA and their target repositories are available for human and other organisms in literature, but a dedicated resource on viral miRNAs and their targets are lacking. Therefore, we have developed a comprehensive viral miRNA resource harboring information of 9133 entries in three subdatabases. This includes 1308 experimentally validated miRNA sequences with their isomiRs encoded by 44 viruses in viral miRNA ' VIRMIRNA: ' and 7283 of their target genes in ' VIRMIRTAR': . Additionally, there is information of 542 antiviral miRNAs encoded by the host against 24 viruses in antiviral miRNA ' AVIRMIR': . The web interface was developed using Linux-Apache-MySQL-PHP (LAMP) software bundle. User-friendly browse, search, advanced search and useful analysis tools are also provided on the web interface. VIRmiRNA is the first specialized resource of experimentally proven virus-encoded miRNAs and their associated targets. This database would enhance the understanding of viral/host gene regulation and may also prove beneficial in the development of antiviral therapeutics. Database URL: http://crdd.osdd.net/servers/virmirna. © The Author(s) 2014. Published by Oxford University Press.

  18. Customizable PCR-microplate array for differential identification of multiple pathogens

    PubMed Central

    Woubit, Abdela; Yehualaeshet, Teshome; Roberts, Sherrelle; Graham, Martha; Kim, Moonil; Samuel, Temesgen

    2014-01-01

    Customizable PCR-microplate arrays were developed for the rapid identification of Francisella tularensis subsp. tularensis, Salmonella Typhi, Shigella dysenteriae, Yersinia pestis, Vibrio cholerae Escherichia coli O157:H7, Salmonella Typhimurium, Salmonella Saintpaul, Francisella tularensis subsp. novicida, Vibrio parahaemolyticus, and Yersinia pseudotuberculosis. Previously, we identified highly specific primers targeting each of the pathogens above. Here, we report the development of customizable PCR-microplate arrays for simultaneous identification of the pathogens using the primers. A mixed aliquot of genomic DNA from 38 different strains was used to validate three PCR-microplate array formats. Identical PCR conditions were used to run all the samples on the three formats. Results show specific amplifications on all the three custom plates. In a preliminary test to evaluate the sensitivity of these assays in laboratory-inoculated samples, detection limits as low as 9 cfu/g/ml S. Typhimurium were obtained from beef hot dog, and 78 cfu/ml from milk. Such microplate arrays could serve as valuable tools for initial identification or secondary confirmation of these pathogens. PMID:24215700

  19. Customizable PCR-microplate array for differential identification of multiple pathogens.

    PubMed

    Woubit, Abdela; Yehualaeshet, Teshome; Roberts, Sherrelle; Graham, Martha; Kim, Moonil; Samuel, Temesgen

    2013-11-01

    Customizable PCR-microplate arrays were developed for the rapid identification of Salmonella Typhimurium, Salmonella Saintpaul, Salmonella Typhi, Shigella dysenteriae, Escherichia coli O157:H7, Francisella tularensis subsp. tularensis, Francisella tularensis subsp. novicida, Vibrio cholerae, Vibrio parahaemolyticus, Yersinia pestis, and Yersinia pseudotuberculosis. Previously, we identified highly specific primers targeting each of these pathogens. Here, we report the development of customizable PCR-microplate arrays for simultaneous identification of the pathogens using the primers identified. A mixed aliquot of genomic DNA from 38 strains was used to validate three PCR-microplate array formats. Identical PCR conditions were used to run all the samples on the three formats. Specific amplifications were obtained on all three custom plates. In preliminary tests performed to evaluate the sensitivity of these assays in samples inoculated in the laboratory with Salmonella Typhimurium, amplifications were obtained from 1 g of beef hot dog inoculated at as low as 9 CFU/ml or from milk inoculated at as low as 78 CFU/ml. Such microplate arrays could be valuable tools for initial identification or secondary confirmation of contamination by these pathogens.

  20. Discovery of novel drugs for promising targets.

    PubMed

    Martell, Robert E; Brooks, David G; Wang, Yan; Wilcoxen, Keith

    2013-09-01

    Once a promising drug target is identified, the steps to actually discover and optimize a drug are diverse and challenging. The goal of this study was to provide a road map to navigate drug discovery. Review general steps for drug discovery and provide illustrating references. A number of approaches are available to enhance and accelerate target identification and validation. Consideration of a variety of potential mechanisms of action of potential drugs can guide discovery efforts. The hit to lead stage may involve techniques such as high-throughput screening, fragment-based screening, and structure-based design, with informatics playing an ever-increasing role. Biologically relevant screening models are discussed, including cell lines, 3-dimensional culture, and in vivo screening. The process of enabling human studies for an investigational drug is also discussed. Drug discovery is a complex process that has significantly evolved in recent years. © 2013 Elsevier HS Journals, Inc. All rights reserved.

  1. Evaluating the effect of database inflation in proteogenomic search on sensitive and reliable peptide identification.

    PubMed

    Li, Honglan; Joh, Yoon Sung; Kim, Hyunwoo; Paek, Eunok; Lee, Sang-Won; Hwang, Kyu-Baek

    2016-12-22

    Proteogenomics is a promising approach for various tasks ranging from gene annotation to cancer research. Databases for proteogenomic searches are often constructed by adding peptide sequences inferred from genomic or transcriptomic evidence to reference protein sequences. Such inflation of databases has potential of identifying novel peptides. However, it also raises concerns on sensitive and reliable peptide identification. Spurious peptides included in target databases may result in underestimated false discovery rate (FDR). On the other hand, inflation of decoy databases could decrease the sensitivity of peptide identification due to the increased number of high-scoring random hits. Although several studies have addressed these issues, widely applicable guidelines for sensitive and reliable proteogenomic search have hardly been available. To systematically evaluate the effect of database inflation in proteogenomic searches, we constructed a variety of real and simulated proteogenomic databases for yeast and human tandem mass spectrometry (MS/MS) data, respectively. Against these databases, we tested two popular database search tools with various approaches to search result validation: the target-decoy search strategy (with and without a refined scoring-metric) and a mixture model-based method. The effect of separate filtering of known and novel peptides was also examined. The results from real and simulated proteogenomic searches confirmed that separate filtering increases the sensitivity and reliability in proteogenomic search. However, no one method consistently identified the largest (or the smallest) number of novel peptides from real proteogenomic searches. We propose to use a set of search result validation methods with separate filtering, for sensitive and reliable identification of peptides in proteogenomic search.

  2. False discovery rates in spectral identification.

    PubMed

    Jeong, Kyowon; Kim, Sangtae; Bandeira, Nuno

    2012-01-01

    Automated database search engines are one of the fundamental engines of high-throughput proteomics enabling daily identifications of hundreds of thousands of peptides and proteins from tandem mass (MS/MS) spectrometry data. Nevertheless, this automation also makes it humanly impossible to manually validate the vast lists of resulting identifications from such high-throughput searches. This challenge is usually addressed by using a Target-Decoy Approach (TDA) to impose an empirical False Discovery Rate (FDR) at a pre-determined threshold x% with the expectation that at most x% of the returned identifications would be false positives. But despite the fundamental importance of FDR estimates in ensuring the utility of large lists of identifications, there is surprisingly little consensus on exactly how TDA should be applied to minimize the chances of biased FDR estimates. In fact, since less rigorous TDA/FDR estimates tend to result in more identifications (at higher 'true' FDR), there is often little incentive to enforce strict TDA/FDR procedures in studies where the major metric of success is the size of the list of identifications and there are no follow up studies imposing hard cost constraints on the number of reported false positives. Here we address the problem of the accuracy of TDA estimates of empirical FDR. Using MS/MS spectra from samples where we were able to define a factual FDR estimator of 'true' FDR we evaluate several popular variants of the TDA procedure in a variety of database search contexts. We show that the fraction of false identifications can sometimes be over 10× higher than reported and may be unavoidably high for certain types of searches. In addition, we further report that the two-pass search strategy seems the most promising database search strategy. While unavoidably constrained by the particulars of any specific evaluation dataset, our observations support a series of recommendations towards maximizing the number of resulting

  3. TipMT: Identification of PCR-based taxon-specific markers.

    PubMed

    Rodrigues-Luiz, Gabriela F; Cardoso, Mariana S; Valdivia, Hugo O; Ayala, Edward V; Gontijo, Célia M F; Rodrigues, Thiago de S; Fujiwara, Ricardo T; Lopes, Robson S; Bartholomeu, Daniella C

    2017-02-11

    Molecular genetic markers are one of the most informative and widely used genome features in clinical and environmental diagnostic studies. A polymerase chain reaction (PCR)-based molecular marker is very attractive because it is suitable to high throughput automation and confers high specificity. However, the design of taxon-specific primers may be difficult and time consuming due to the need to identify appropriate genomic regions for annealing primers and to evaluate primer specificity. Here, we report the development of a Tool for Identification of Primers for Multiple Taxa (TipMT), which is a web application to search and design primers for genotyping based on genomic data. The tool identifies and targets single sequence repeats (SSR) or orthologous/taxa-specific genes for genotyping using Multiplex PCR. This pipeline was applied to the genomes of four species of Leishmania (L. amazonensis, L. braziliensis, L. infantum and L. major) and validated by PCR using artificial genomic DNA mixtures of the Leishmania species as templates. This experimental validation demonstrates the reliability of TipMT because amplification profiles showed discrimination of genomic DNA samples from Leishmania species. The TipMT web tool allows for large-scale identification and design of taxon-specific primers and is freely available to the scientific community at http://200.131.37.155/tipMT/ .

  4. Modeling human target acquisition in ground-to-air weapon systems

    NASA Technical Reports Server (NTRS)

    Phatak, A. V.; Mohr, R. L.; Vikmanis, M.; Wei, K. C.

    1982-01-01

    The problems associated with formulating and validating mathematical models for describing and predicting human target acquisition response are considered. In particular, the extension of the human observer model to include the acquisition phase as well as the tracking segment is presented. Relationship of the Observer model structure to the more complex Standard Optimal Control model formulation and to the simpler Transfer Function/Noise representation is discussed. Problems pertinent to structural identifiability and the form of the parameterization are elucidated. A systematic approach toward the identification of the observer acquisition model parameters from ensemble tracking error data is presented.

  5. 5 CFR 2100.4 - Requirements for identification.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... must provide their full name and as much information as possible in order that a proper search for..., military or annuitant identification card, or any official document as acceptable identification validation...

  6. Identification of Target Complaints by Computer Interview: Evaluation of the Computerized Assessment System for Psychotherapy Evaluation and Research.

    ERIC Educational Resources Information Center

    Farrell, Albert D.; And Others

    1987-01-01

    Evaluated computer interview to standardize collection of target complaints. Adult outpatients (N=103) completed computer interview, unstructured intake interview, Symptoms Checklist-90, and Minnesota Multiphasic Personality Inventory. Results provided support for the computer interview in regard to reliability and validity though there was low…

  7. Manufacturing validation of biologically functional T cells targeted to CD19 antigen for autologous adoptive cell therapy.

    PubMed

    Hollyman, Daniel; Stefanski, Jolanta; Przybylowski, Mark; Bartido, Shirley; Borquez-Ojeda, Oriana; Taylor, Clare; Yeh, Raymond; Capacio, Vanessa; Olszewska, Malgorzata; Hosey, James; Sadelain, Michel; Brentjens, Renier J; Rivière, Isabelle

    2009-01-01

    On the basis of promising preclinical data demonstrating the eradication of systemic B-cell malignancies by CD19-targeted T lymphocytes in vivo in severe combined immunodeficient-beige mouse models, we are launching phase I clinical trials in patients with chronic lymphocytic leukemia (CLL) and acute lymphoblastic leukemia. We present here the validation of the bioprocess which we developed for the production and expansion of clinical grade autologous T cells derived from patients with CLL. We demonstrate that T cells genetically modified with a replication-defective gammaretroviral vector derived from the Moloney murine leukemia virus encoding a chimeric antigen receptor (CAR) targeted to CD19 (1928z) can be expanded with Dynabeads CD3/CD28. This bioprocess allows us to generate clinical doses of 1928z+ T cells in approximately 2 to 3 weeks in a large-scale semiclosed culture system using the Wave Bioreactor. These 1928z+ T cells remain biologically functional not only in vitro but also in severe combined immunodeficient-beige mice bearing disseminated tumors. The validation requirements in terms of T-cell expansion, T-cell transduction with the 1928z CAR, biologic activity, quality control testing, and release criteria were met for all 4 validation runs using apheresis products from patients with CLL. Additionally, after expansion of the T cells, the diversity of the skewed Vbeta T-cell receptor repertoire was significantly restored. This validated process will be used in phase I clinical trials in patients with chemorefractory CLL and in patients with relapsed acute lymphoblastic leukemia. It can also be adapted for other clinical trials involving the expansion and transduction of patient or donor T cells using any CAR or T-cell receptor.

  8. Food for Thought ... Mechanistic Validation

    PubMed Central

    Hartung, Thomas; Hoffmann, Sebastian; Stephens, Martin

    2013-01-01

    Summary Validation of new approaches in regulatory toxicology is commonly defined as the independent assessment of the reproducibility and relevance (the scientific basis and predictive capacity) of a test for a particular purpose. In large ring trials, the emphasis to date has been mainly on reproducibility and predictive capacity (comparison to the traditional test) with less attention given to the scientific or mechanistic basis. Assessing predictive capacity is difficult for novel approaches (which are based on mechanism), such as pathways of toxicity or the complex networks within the organism (systems toxicology). This is highly relevant for implementing Toxicology for the 21st Century, either by high-throughput testing in the ToxCast/ Tox21 project or omics-based testing in the Human Toxome Project. This article explores the mostly neglected assessment of a test's scientific basis, which moves mechanism and causality to the foreground when validating/qualifying tests. Such mechanistic validation faces the problem of establishing causality in complex systems. However, pragmatic adaptations of the Bradford Hill criteria, as well as bioinformatic tools, are emerging. As critical infrastructures of the organism are perturbed by a toxic mechanism we argue that by focusing on the target of toxicity and its vulnerability, in addition to the way it is perturbed, we can anchor the identification of the mechanism and its verification. PMID:23665802

  9. Nematode Species Identification-Current Status, Challenges and Future Perspectives for Cyathostomins.

    PubMed

    Bredtmann, Christina M; Krücken, Jürgen; Murugaiyan, Jayaseelan; Kuzmina, Tetiana; von Samson-Himmelstjerna, Georg

    2017-01-01

    Human and animal health is globally affected by a variety of parasitic helminths. The impact of co-infections and development of anthelmintic resistance requires improved diagnostic tools, especially for parasitic nematodes e.g., to identify resistant species or attribute pathological effects to individual species or particular species combinations. In horses, co-infection with cyathostomins is rather a rule than an exception with typically 5 to 15 species (out of more than 40 described) per individual host. In cyathostomins, reliable morphological species differentiation is currently limited to adults and requires highly specialized expertize while precise morphological identification of eggs and early stage larvae is impossible. The situation is further complicated by a questionable validity of some cyathostomins while others might actually represent cryptic species complexes. Several molecular methods using different target sequences were established to overcome these limitations. For adult worms, PCR followed by sequencing of mitochondrial genes or external or internal ribosomal RNA spacers is suitable to genetically confirm morphological identifications. The most commonly used method to differentiate eggs or larvae is the reverse-line-blot hybridization assay. However, both methods suffer from the fact that target sequences are not available for many species or even that GenBank® entries are unreliable regarding the cyathostomin species. Recent advances in proteomic tools for identification of metazoans including insects and nematodes of the genus Trichinella will be evaluated for suitability to diagnose cyathostomins. Future research should focus on the comparative analysis of morphological, molecular and proteomic data from the same cyathostomin specimen to optimize tools for species-specific identification.

  10. Cy5 maleimide labelling for sensitive detection of free thiols in native protein extracts: identification of seed proteins targeted by barley thioredoxin h isoforms.

    PubMed Central

    Maeda, Kenji; Finnie, Christine; Svensson, Birte

    2004-01-01

    Barley thioredoxin h isoforms HvTrxh1 and HvTrxh2 differ in temporal and spatial distribution and in kinetic properties. Target proteins of HvTrxh1 and HvTrxh2 were identified in mature seeds and in seeds after 72 h of germination. Improvement of the established method for identification of thioredoxin-targeted proteins based on two-dimensional electrophoresis and fluorescence labelling of thiol groups was achieved by application of a highly sensitive Cy5 maleimide dye and large-format two-dimensional gels, resulting in a 10-fold increase in the observed number of labelled protein spots. The technique also provided information about accessible thiol groups in the proteins identified in the barley seed proteome. In total, 16 different putative target proteins were identified from 26 spots using tryptic in-gel digestion, matrix-assisted laser-desorption ionization-time-of-flight MS and database search. HvTrxh1 and HvTrxh2 were shown to have similar target specificity. Barley alpha-amylase/subtilisin inhibitor, previously demonstrated to be reduced by both HvTrxh1 and HvTrxh2, was among the identified target proteins, confirming the suitability of the method. Several alpha-amylase/trypsin inhibitors, some of which are already known as target proteins of thioredoxin h, and cyclophilin known as a target protein of m-type thioredoxin were also identified. Lipid transfer protein, embryospecific protein, three chitinase isoenzymes, a single-domain glyoxalase-like protein and superoxide dismutase were novel identifications of putative target proteins, suggesting new physiological roles of thioredoxin h in barley seeds. PMID:14636158

  11. Rapid Identification of Chemoresistance Mechanisms Using Yeast DNA Mismatch Repair Mutants

    PubMed Central

    Ojini, Irene; Gammie, Alison

    2015-01-01

    Resistance to cancer therapy is a major obstacle in the long-term treatment of cancer. A greater understanding of drug resistance mechanisms will ultimately lead to the development of effective therapeutic strategies to prevent resistance from occurring. Here, we exploit the mutator phenotype of mismatch repair defective yeast cells combined with whole genome sequencing to identify drug resistance mutations in key pathways involved in the development of chemoresistance. The utility of this approach was demonstrated via the identification of the known CAN1 and TOP1 resistance targets for two compounds, canavanine and camptothecin, respectively. We have also experimentally validated the plasma membrane transporter HNM1 as the primary drug resistance target of mechlorethamine. Furthermore, the sequencing of mitoxantrone-resistant strains identified inactivating mutations within IPT1, a gene encoding inositolphosphotransferase, an enzyme involved in sphingolipid biosynthesis. In the case of bactobolin, a promising anticancer drug, the endocytosis pathway was identified as the drug resistance target responsible for conferring resistance. Finally, we show that that rapamycin, an mTOR inhibitor previously shown to alter the fitness of the ipt1 mutant, can effectively prevent the formation of mitoxantrone resistance. The rapid and robust nature of these techniques, using Saccharomyces cerevisiae as a model organism, should accelerate the identification of drug resistance targets and guide the development of novel therapeutic combination strategies to prevent the development of chemoresistance in various cancers. PMID:26199284

  12. eap Gene as novel target for specific identification of Staphylococcus aureus.

    PubMed

    Hussain, Muzaffar; von Eiff, Christof; Sinha, Bhanu; Joost, Insa; Herrmann, Mathias; Peters, Georg; Becker, Karsten

    2008-02-01

    The cell surface-associated extracellular adherence protein (Eap) mediates adherence of Staphylococcus aureus to host extracellular matrix components and inhibits inflammation, wound healing, and angiogenesis. A well-characterized collection of S. aureus and non-S. aureus staphylococcal isolates (n = 813) was tested for the presence of the Eap-encoding gene (eap) by PCR to investigate the use of the eap gene as a specific diagnostic tool for identification of S. aureus. Whereas all 597 S. aureus isolates were eap positive, this gene was not detectable in 216 non-S. aureus staphylococcal isolates comprising 47 different species and subspecies of coagulase-negative staphylococci and non-S. aureus coagulase-positive or coagulase-variable staphylococci. Furthermore, non-S. aureus isolates did not express Eap homologs, as verified on the transcriptional and protein levels. Based on these data, the sensitivity and specificity of the newly developed PCR targeting the eap gene were both 100%. Thus, the unique occurrence of Eap in S. aureus offers a promising tool particularly suitable for molecular diagnostics of this pathogen.

  13. Structure identification for Non-Targeted Analytical Chemistry using the US EPA’s CompTox Chemistry Dashboard (ACS 2017 Fall meeting 3 of 3)

    EPA Science Inventory

    Identification of unknowns in non-targeted analyses (NTA) requires the integration of complementary data types to generate a confident consensus structure. Researchers use a variety of data and tools (e.g., chemical reference databases, spectral matching, fragment prediction too...

  14. Direct Quantification of Methane Emissions Across the Supply Chain: Identification of Mitigation Targets

    NASA Astrophysics Data System (ADS)

    Darzi, M.; Johnson, D.; Heltzel, R.; Clark, N.

    2017-12-01

    Researchers at West Virginia University's Center for Alternative Fuels, Engines, and Emissions have recently participated in a variety of studies targeted at direction quantification of methane emissions from across the natural gas supply chain. These studies included assessing methane emissions from heavy-duty vehicles and their fuel stations, active unconventional well sites - during both development and production, natural gas compression and storage facilities, natural gas engines - both large and small, two- and four-stroke, and low-throughput equipment associated with coal bed methane wells. Engine emissions were sampled using conventional instruments such as Fourier transform infrared spectrometers and heated flame ionization detection analyzers. However, to accurately quantify a wide range of other sources beyond the tailpipe (both leaks and losses), a full flow sampling system was developed, which included an integrated cavity-enhanced absorption spectrometer. Through these direct quantification efforts and analysis major sources of methane emissions were identified. Technological solutions and best practices exist or could be developed to reduce methane emissions by focusing on the "lowest-hanging fruit." For example, engine crankcases from across the supply chain should employ vent mitigation systems to reduce methane and other emissions. An overview of the direct quantification system and various campaign measurements results will be presented along with the identification of other targets for additional mitigation.

  15. A targeted mass spectrometry-based approach for the identification and characterization of proteins containing α-aminoadipic and γ-glutamic semialdehyde residues

    PubMed Central

    Chavez, Juan D.; Bisson, William H.

    2011-01-01

    The site-specific identification of α-aminoadipic semialdehyde (AAS) and γ-glutamic semialdehyde (GGS) residues in proteins is reported. Semialdehydic protein modifications result from the metal-catalyzed oxidation of Lys or Arg and Pro residues, respectively. Most of the analytical methods for the analysis of protein carbonylation measure change to the global level of carbonylation and fail to provide details regarding protein identity, site, and chemical nature of the carbonylation. In this work, we used a targeted approach, which combines chemical labeling, enrichment, and tandem mass spectrometric analysis, for the site-specific identification of AAS and GGS sites in proteins. The approach is applied to in vitro oxidized glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and an untreated biological sample, namely cardiac mitochondrial proteins. The analysis of GAPDH resulted in the site-specific identification of two AAA and four GGS residues. Computational evaluation of the identified AAS and GGS sites in GAPDH indicated that these sites are located in flexible regions, show high solvent accessibility values, and are in proximity with possible metal ion binding sites. The targeted proteomic analysis of semialdehydic modifications in cardiac mitochondria yielded nine AAS modification sites which were unambiguously assigned to distinct lysine residues in the following proteins: ATP/ATP translocase isoforms 1 and 2, ubiquinol cytochrome-c reductase core protein 2, and ATP synthase α-subunit. PMID:20957471

  16. Identification of Reduced-Order Thermal Therapy Models Using Thermal MR Images: Theory and Validation

    PubMed Central

    2013-01-01

    In this paper, we develop and validate a method to identify computationally efficient site- and patient-specific models of ultrasound thermal therapies from MR thermal images. The models of the specific absorption rate of the transduced energy and the temperature response of the therapy target are identified in the reduced basis of proper orthogonal decomposition of thermal images, acquired in response to a mild thermal test excitation. The method permits dynamic reidentification of the treatment models during the therapy by recursively utilizing newly acquired images. Such adaptation is particularly important during high-temperature therapies, which are known to substantially and rapidly change tissue properties and blood perfusion. The developed theory was validated for the case of focused ultrasound heating of a tissue phantom. The experimental and computational results indicate that the developed approach produces accurate low-dimensional treatment models despite temporal and spatial noises in MR images and slow image acquisition rate. PMID:22531754

  17. Identification of reduced-order thermal therapy models using thermal MR images: theory and validation.

    PubMed

    Niu, Ran; Skliar, Mikhail

    2012-07-01

    In this paper, we develop and validate a method to identify computationally efficient site- and patient-specific models of ultrasound thermal therapies from MR thermal images. The models of the specific absorption rate of the transduced energy and the temperature response of the therapy target are identified in the reduced basis of proper orthogonal decomposition of thermal images, acquired in response to a mild thermal test excitation. The method permits dynamic reidentification of the treatment models during the therapy by recursively utilizing newly acquired images. Such adaptation is particularly important during high-temperature therapies, which are known to substantially and rapidly change tissue properties and blood perfusion. The developed theory was validated for the case of focused ultrasound heating of a tissue phantom. The experimental and computational results indicate that the developed approach produces accurate low-dimensional treatment models despite temporal and spatial noises in MR images and slow image acquisition rate.

  18. Identification and validation of novel prognostic markers in Renal Cell Carcinoma.

    PubMed

    Rabjerg, Maj

    2017-10-01

    insights since we could detect currents compatible with KCa3.1 and KCa1.1 in the cell membrane of primary and commercial ccRCC cell lines. Nonetheless, we were not able to show any significant inhibition of cell growth by the selective inhibitors of KCa3.1 and KCa1.1, TRAM-34, RA-2 and Paxilline. In study III our aim was to investigate the prognostic role of 19 genes selected on the basis of an earlier study done by the group. We used Taqman® Low Density Array to perform a quantitative real-time PCR analysis. By selecting an optimal cut-point and correct for overestimation of the p-value, we could identify three genes with impact on prognosis of ccRCC in both univariate and multivariate analysis. High expression of the genes SPP1 and CSNK2A1 (encoding Osteopontin and CK2α respectively) correlated with poor prognosis while high expression of DEFB1 (encoding β-Defensin) correlated with better prognosis. Study IV focused on validating the results obtained in Paper III by investigating the protein expression of CK2α (Protein kinase 2, alpha subunit) in the different subtypes of RCC and oncocytoma. Furthermore, we investigated whether protein expression of CK2α in ccRCC correlated with prognosis. Here we could show, that a positive nuclear staining was a marker of poor prognosis in high-stage ccRCC. Moreover, enzyme activity analysis revealed a higher activity of the protein kinase in tumor tissue of ccRCC than in normal renal cortex. Novel insights were provided in a proliferation study where we investigated the selective inhibitors of CK2α, CX-4945 and E9. CX-4945 was able to inhibit ccRCC cell growth by nearly 50%. All together the studies presented in this thesis add additional information to the ongoing research within identification of novel prognostic markers in ccRCC. We have discovered four new molecular markers, which reliably can predict prognosis at the time of diagnosis. Additionally, we identified CK2α as a novel therapeutic target of ccRCC. The studies

  19. The effect on cadaver blood DNA identification by the use of targeted and whole body post-mortem computed tomography angiography.

    PubMed

    Rutty, Guy N; Barber, Jade; Amoroso, Jasmin; Morgan, Bruno; Graham, Eleanor A M

    2013-12-01

    Post-mortem computed tomography angiography (PMCTA) involves the injection of contrast agents. This could have both a dilution effect on biological fluid samples and could affect subsequent post-contrast analytical laboratory processes. We undertook a small sample study of 10 targeted and 10 whole body PMCTA cases to consider whether or not these two methods of PMCTA could affect post-PMCTA cadaver blood based DNA identification. We used standard methodology to examine DNA from blood samples obtained before and after the PMCTA procedure. We illustrate that neither of these PMCTA methods had an effect on the alleles called following short tandem repeat based DNA profiling, and therefore the ability to undertake post-PMCTA blood based DNA identification.

  20. The single lineup paradigm: A new way to manipulate target presence in eyewitness identification experiments.

    PubMed

    Oriet, Chris; Fitzgerald, Ryan J

    2018-02-01

    The suspect in eyewitness lineups may be guilty or innocent. These possibilities are traditionally simulated in eyewitness identification studies using a dual-lineup paradigm: All witnesses observe the same perpetrator and then receive one of two lineups. In this paradigm, the suspect's guilt is manipulated by including the perpetrator in one lineup and an innocent suspect in the other. The lineup is then filled with people matched to either the suspect (resulting in different fillers in perpetrator-present and perpetrator-absent lineups) or to the perpetrator (resulting in the same fillers in each lineup). An inescapable feature of the dual-lineup paradigm is that the perpetrator-present and perpetrator-absent lineups differ not only in the suspect's guilt, but also in their composition. Here, we describe a single-lineup paradigm: Subjects observe one of two perpetrators and then all subjects receive the same lineup containing one of the perpetrators. This alternative paradigm allows manipulation of the suspect's guilt without changing the lineup's composition. In three experiments, we applied the single-lineup paradigm to explore suspect-filler similarity and consistently found that increasing similarity reduced perpetrator identifications but did little to prevent innocent suspect misidentifications. Conversely, when fillers were matched to the perpetrator using a dual-lineup paradigm, increasing similarity reduced identification of perpetrators and innocent suspects. This finding suggests that the effect of filler similarity may depend on the person to whom the fillers are matched. We suggest that the single-lineup paradigm is a more ecologically valid and better controlled approach to creating suspect-matched lineups in laboratory investigations of eyewitness memory than existing procedures. (PsycINFO Database Record (c) 2018 APA, all rights reserved).

  1. Computational identification of structural factors affecting the mutagenic potential of aromatic amines: study design and experimental validation.

    PubMed

    Slavov, Svetoslav H; Stoyanova-Slavova, Iva; Mattes, William; Beger, Richard D; Brüschweiler, Beat J

    2018-07-01

    A grid-based, alignment-independent 3D-SDAR (three-dimensional spectral data-activity relationship) approach based on simulated 13 C and 15 N NMR chemical shifts augmented with through-space interatomic distances was used to model the mutagenicity of 554 primary and 419 secondary aromatic amines. A robust modeling strategy supported by extensive validation including randomized training/hold-out test set pairs, validation sets, "blind" external test sets as well as experimental validation was applied to avoid over-parameterization and build Organization for Economic Cooperation and Development (OECD 2004) compliant models. Based on an experimental validation set of 23 chemicals tested in a two-strain Salmonella typhimurium Ames assay, 3D-SDAR was able to achieve performance comparable to 5-strain (Ames) predictions by Lhasa Limited's Derek and Sarah Nexus for the same set. Furthermore, mapping of the most frequently occurring bins on the primary and secondary aromatic amine structures allowed the identification of molecular features that were associated either positively or negatively with mutagenicity. Prominent structural features found to enhance the mutagenic potential included: nitrobenzene moieties, conjugated π-systems, nitrothiophene groups, and aromatic hydroxylamine moieties. 3D-SDAR was also able to capture "true" negative contributions that are particularly difficult to detect through alternative methods. These include sulphonamide, acetamide, and other functional groups, which not only lack contributions to the overall mutagenic potential, but are known to actively lower it, if present in the chemical structures of what otherwise would be potential mutagens.

  2. Dana-Farber Cancer Institute: Identification of Therapeutic Targets Across Cancer Types | Office of Cancer Genomics

    Cancer.gov

    The Dana Farber Cancer Institute CTD2 Center focuses on the use of high-throughput genetic and bioinformatic approaches to identify and credential oncogenes and co-dependencies in cancers. This Center aims to provide the cancer research community with information that will facilitate the prioritization of targets based on both genomic and functional evidence, inform the most appropriate genetic context for downstream mechanistic and validation studies, and enable the translation of this information into therapeutics and diagnostics.

  3. Identification of alsterpaullone as a novel small molecule inhibitor to target group 3 medulloblastoma.

    PubMed

    Faria, Claudia C; Agnihotri, Sameer; Mack, Stephen C; Golbourn, Brian J; Diaz, Roberto J; Olsen, Samantha; Bryant, Melissa; Bebenek, Matthew; Wang, Xin; Bertrand, Kelsey C; Kushida, Michelle; Head, Renee; Clark, Ian; Dirks, Peter; Smith, Christian A; Taylor, Michael D; Rutka, James T

    2015-08-28

    Advances in the molecular biology of medulloblastoma revealed four genetically and clinically distinct subgroups. Group 3 medulloblastomas are characterized by frequent amplifications of the oncogene MYC, a high incidence of metastasis, and poor prognosis despite aggressive therapy. We investigated several potential small molecule inhibitors to target Group 3 medulloblastomas based on gene expression data using an in silico drug screen. The Connectivity Map (C-MAP) analysis identified piperlongumine as the top candidate drug for non-WNT medulloblastomas and the cyclin-dependent kinase (CDK) inhibitor alsterpaullone as the compound predicted to have specific antitumor activity against Group 3 medulloblastomas. To validate our findings we used these inhibitors against established Group 3 medulloblastoma cell lines. The C-MAP predicted drugs reduced cell proliferation in vitro and increased survival in Group 3 medulloblastoma xenografts. Alsterpaullone had the highest efficacy in Group 3 medulloblastoma cells. Genomic profiling of Group 3 medulloblastoma cells treated with alsterpaullone confirmed inhibition of cell cycle-related genes, and down-regulation of MYC. Our results demonstrate the preclinical efficacy of using a targeted therapy approach for Group 3 medulloblastomas. Specifically, we provide rationale for advancing alsterpaullone as a targeted therapy in Group 3 medulloblastoma.

  4. Identification of Hepatitis C Virus Inhibitors Targeting Different Aspects of Infection Using a Cell-Based Assay

    PubMed Central

    Yu, Xuemei; Sainz, Bruno; Petukhov, Pavel A.

    2012-01-01

    With 2 to 3% of the worldwide population chronically infected, hepatitis C virus (HCV) infection continues to be a major health care burden. Unfortunately, current interferon-based treatment options are not effective in all patients and are associated with significant side effects. Consequently, there is an ongoing need to identify and develop new anti-HCV therapies. Toward this goal, we previously developed a cell-based HCV infection assay for antiviral compound screening based on a low-multiplicity-of-infection approach that uniquely allows for the identification of antiviral compounds that target cell culture-derived HCV (HCVcc) at any step of the viral infection cycle. Using this assay, here we report the screening of the NCI Diversity Set II library, containing 1,974 synthesized chemical compounds, and the identification of compounds with specific anti-HCV activity. In combination with toxicity counterscreening, we identified 30 hits from the compound library, 13 of which showed reproducible and dose-dependent inhibition of HCV with mean therapeutic indices (50% cytotoxic concentration [CC50]/50% effective concentration [EC50]) of greater than 6. Using HCV pseudotype and replicon systems of multiple HCV genotypes, as well as infectious HCVcc-based assembly and secretion analysis, we determined that different compounds within this group of candidate inhibitors target different steps of viral infection. The compounds identified not only will serve as biological probes to study and further dissect the biology of viral infection but also should facilitate the development of new anti-HCV therapeutic treatments. PMID:22948883

  5. Identification of active sources inside cavities using the equivalent source method-based free-field recovery technique

    NASA Astrophysics Data System (ADS)

    Bi, Chuan-Xing; Hu, Ding-Yu; Zhang, Yong-Bin; Jing, Wen-Qian

    2015-06-01

    In previous studies, an equivalent source method (ESM)-based technique for recovering the free sound field in a noisy environment has been successfully applied to exterior problems. In order to evaluate its performance when applied to a more general noisy environment, that technique is used to identify active sources inside cavities where the sound field is composed of the field radiated by active sources and that reflected by walls. A patch approach with two semi-closed surfaces covering the target active sources is presented to perform the measurements, and the field that would be radiated by these target active sources into free space is extracted from the mixed field by using the proposed technique, which will be further used as the input of nearfield acoustic holography for source identification. Simulation and experimental results validate the effectiveness of the proposed technique for source identification in cavities, and show the feasibility of performing the measurements with a double layer planar array.

  6. Computational identification of conserved microRNAs and their targets from expression sequence tags of blueberry (Vaccinium corybosum)

    PubMed Central

    Li, Xuyan; Hou, Yanming; Zhang, Li; Zhang, Wenhao; Quan, Chen; Cui, Yuhai; Bian, Shaomin

    2014-01-01

    MicroRNAs (miRNAs) are a class of endogenous, approximately 21nt in length, non-coding RNA, which mediate the expression of target genes primarily at post-transcriptional levels. miRNAs play critical roles in almost all plant cellular and metabolic processes. Although numerous miRNAs have been identified in the plant kingdom, the miRNAs in blueberry, which is an economically important small fruit crop, still remain totally unknown. In this study, we reported a computational identification of miRNAs and their targets in blueberry. By conducting an EST-based comparative genomics approach, 9 potential vco-miRNAs were discovered from 22,402 blueberry ESTs according to a series of filtering criteria, designated as vco-miR156–5p, vco-miR156–3p, vco-miR1436, vco-miR1522, vco-miR4495, vco-miR5120, vco-miR5658, vco-miR5783, and vco-miR5986. Based on sequence complementarity between miRNA and its target transcript, 34 target ESTs from blueberry and 70 targets from other species were identified for the vco-miRNAs. The targets were found to be involved in transcription, RNA splicing and binding, DNA duplication, signal transduction, transport and trafficking, stress response, as well as synthesis and metabolic process. These findings will greatly contribute to future research in regard to functions and regulatory mechanisms of blueberry miRNAs. PMID:25763692

  7. Computational identification of conserved microRNAs and their targets from expression sequence tags of blueberry (Vaccinium corybosum).

    PubMed

    Li, Xuyan; Hou, Yanming; Zhang, Li; Zhang, Wenhao; Quan, Chen; Cui, Yuhai; Bian, Shaomin

    2014-01-01

    MicroRNAs (miRNAs) are a class of endogenous, approximately 21nt in length, non-coding RNA, which mediate the expression of target genes primarily at post-transcriptional levels. miRNAs play critical roles in almost all plant cellular and metabolic processes. Although numerous miRNAs have been identified in the plant kingdom, the miRNAs in blueberry, which is an economically important small fruit crop, still remain totally unknown. In this study, we reported a computational identification of miRNAs and their targets in blueberry. By conducting an EST-based comparative genomics approach, 9 potential vco-miRNAs were discovered from 22,402 blueberry ESTs according to a series of filtering criteria, designated as vco-miR156-5p, vco-miR156-3p, vco-miR1436, vco-miR1522, vco-miR4495, vco-miR5120, vco-miR5658, vco-miR5783, and vco-miR5986. Based on sequence complementarity between miRNA and its target transcript, 34 target ESTs from blueberry and 70 targets from other species were identified for the vco-miRNAs. The targets were found to be involved in transcription, RNA splicing and binding, DNA duplication, signal transduction, transport and trafficking, stress response, as well as synthesis and metabolic process. These findings will greatly contribute to future research in regard to functions and regulatory mechanisms of blueberry miRNAs.

  8. Target validation: linking target and chemical properties to desired product profile.

    PubMed

    Wyatt, Paul G; Gilbert, Ian H; Read, Kevin D; Fairlamb, Alan H

    2011-01-01

    The discovery of drugs is a lengthy, high-risk and expensive business taking at least 12 years and is estimated to cost upwards of US$800 million for each drug to be successfully approved for clinical use. Much of this cost is driven by the late phase clinical trials and therefore the ability to terminate early those projects destined to fail is paramount to prevent unwanted costs and wasted effort. Although neglected diseases drug discovery is driven more by unmet medical need rather than financial considerations, the need to minimise wasted money and resources is even more vital in this under-funded area. To ensure any drug discovery project is addressing the requirements of the patients and health care providers and delivering a benefit over existing therapies, the ideal attributes of a novel drug needs to be pre-defined by a set of criteria called a target product profile. Using a target product profile the drug discovery process, clinical study design, and compound characteristics can be defined all the way back through to the suitability or druggability of the intended biochemical target. Assessment and prioritisation of the most promising targets for entry into screening programmes is crucial for maximising chances of success.

  9. Bioinformatic identification and experimental validation of miRNAs from foxtail millet (Setaria italica).

    PubMed

    Han, Jun; Xie, Hao; Sun, Qingpeng; Wang, Jun; Lu, Min; Wang, Weixiang; Guo, Erhu; Pan, Jinbao

    2014-08-10

    MiRNAs are a novel group of non-coding small RNAs that negatively regulate gene expression. Many miRNAs have been identified and investigated extensively in plant species with sequenced genomes. However, few miRNAs have been identified in foxtail millet (Setaria italica), which is an ancient cereal crop of great importance for dry land agriculture. In this study, 271 foxtail millet miRNAs belonging to 44 families were identified using a bioinformatics approach. Twenty-three pairs of sense/antisense miRNAs belonging to 13 families, and 18 miRNA clusters containing members of 8 families were discovered in foxtail millet. We identified 432 potential targets for 38 miRNA families, most of which were predicted to be involved in plant development, signal transduction, metabolic pathways, disease resistance, and environmental stress responses. Gene ontology (GO) analysis revealed that 101, 56, and 23 target genes were involved in molecular functions, biological processes, and cellular components, respectively. We investigated the expression patterns of 43 selected miRNAs using qRT-PCR analysis. All of the miRNAs were expressed ubiquitously with many exhibiting different expression levels in different tissues. We validated five predicted targets of four miRNAs using the RNA ligase mediated rapid amplification of cDNA end (5'-RLM-RACE) method. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. The topoisomerase II-Hsp90 complex: a new chemotherapeutic target?

    PubMed

    Barker, Catherine R; Hamlett, Jane; Pennington, Stephen R; Burrows, Francis; Lundgren, Karen; Lough, Rachel; Watson, Alastair J M; Jenkins, John R

    2006-06-01

    The modulation of DNA topology by topoisomerase II plays a crucial role during chromosome condensation and segregation in mitosis and has thus become a highly attractive target for chemotherapeutic drugs. However, these drugs are highly toxic, and so new approaches are required. One such strategy is to target topoisomerase II-interacting proteins. Here we report the identification of potential topoisomerase II-associated proteins using immunoprecipitation, followed by 1-D and 2-D gel electrophoresis and MALDI-TOF mass spectrometry. A total of 23 proteins were identified and, of these, 17 were further validated as topoisomerase IIalpha-associated proteins by coimmunoprecipitation and Western blot. Six of the interacting proteins were cellular chaperones, including 3 members of the heat shock protein-90 (Hsp90) family, and so the effect of Hsp90 modulation on the antitumor activity of topoisomerase II drugs was tested using the sulforhodamine B assay, clonogenic assays and a xenograft model. The Hsp90 inhibitors geldanamycin, 17-AAG (17-allylamino-17-demethoxygeldanamycin) and radicicol significantly enhanced the activity of the topoisomerase II poisons etoposide and mitoxantrone in vitro and in vivo. Thus, our method of identifying topoisomerase II-interacting proteins appears to be effective, and at least 1 novel topoisomerase IIalpha-associated protein, Hsp90, may represent a valid drug target in the context of topoisomerase II-directed chemotherapy.

  11. Fruitful research: drug target discovery for neurodegenerative diseases in Drosophila.

    PubMed

    Konsolaki, Mary

    2013-12-01

    Although vertebrate model systems have obvious advantages in the study of human disease, invertebrate organisms have contributed enormously to this field as well. The conservation of genome structure and physiology among organisms poses unexpected peculiarities, and the redundancy in certain gene families or the presence of polymorphisms that can slightly alter gene expression can, in certain instances, bring invertebrate systems, such as Drosophila, closer to humans than mice and vice versa. This necessitates the analysis of disease pathways in multiple model organisms. The author highlights findings from Drosophila models of neurodegenerative diseases that have occurred in the past few years. She also highlights and discusses various molecular, genetic and genomic tools used in flies, as well as methods for generating disease models. Finally, the author describes Drosophila models of Alzheimer's, Parkinson's tri-nucleotide repeat diseases, and Fragile X syndrome and summarizes insights in disease mechanisms that have been discovered directly in fly models. Full genome genetic screens in Drosophila can lead to the rapid identification of drug target candidates that can be subsequently validated in a vertebrate system. In addition, the Drosophila models of neurodegeneration may often show disease phenotypes that are absent in equivalent mouse models. The author believes that the extensive contribution of Drosophila to both new disease drug target discovery, in addition to target validation, makes them indispensible to drug discovery and development.

  12. Identification of the TFII-I family target genes in the vertebrate genome.

    PubMed

    Chimge, Nyam-Osor; Makeyev, Aleksandr V; Ruddle, Frank H; Bayarsaihan, Dashzeveg

    2008-07-01

    GTF2I and GTF2IRD1 encode members of the TFII-I transcription factor family and are prime candidates in the Williams syndrome, a complex neurodevelopmental disorder. Our previous expression microarray studies implicated TFII-I proteins in the regulation of a number of genes critical in various aspects of cell physiology. Here, we combined bioinformatics and microarray results to identify TFII-I downstream targets in the vertebrate genome. These results were validated by chromatin immunoprecipitation and siRNA analysis. The collected evidence revealed the complexity of TFII-I-mediated processes that involve distinct regulatory networks. Altogether, these results lead to a better understanding of specific molecular events, some of which may be responsible for the Williams syndrome phenotype.

  13. Indel-seq: a fast-forward genetics approach for identification of trait-associated putative candidate genomic regions and its application in pigeonpea (Cajanus cajan).

    PubMed

    Singh, Vikas K; Khan, Aamir W; Saxena, Rachit K; Sinha, Pallavi; Kale, Sandip M; Parupalli, Swathi; Kumar, Vinay; Chitikineni, Annapurna; Vechalapu, Suryanarayana; Sameer Kumar, Chanda Venkata; Sharma, Mamta; Ghanta, Anuradha; Yamini, Kalinati Narasimhan; Muniswamy, Sonnappa; Varshney, Rajeev K

    2017-07-01

    Identification of candidate genomic regions associated with target traits using conventional mapping methods is challenging and time-consuming. In recent years, a number of single nucleotide polymorphism (SNP)-based mapping approaches have been developed and used for identification of candidate/putative genomic regions. However, in the majority of these studies, insertion-deletion (Indel) were largely ignored. For efficient use of Indels in mapping target traits, we propose Indel-seq approach, which is a combination of whole-genome resequencing (WGRS) and bulked segregant analysis (BSA) and relies on the Indel frequencies in extreme bulks. Deployment of Indel-seq approach for identification of candidate genomic regions associated with fusarium wilt (FW) and sterility mosaic disease (SMD) resistance in pigeonpea has identified 16 Indels affecting 26 putative candidate genes. Of these 26 affected putative candidate genes, 24 genes showed effect in the upstream/downstream of the genic region and two genes showed effect in the genes. Validation of these 16 candidate Indels in other FW- and SMD-resistant and FW- and SMD-susceptible genotypes revealed a significant association of five Indels (three for FW and two for SMD resistance). Comparative analysis of Indel-seq with other genetic mapping approaches highlighted the importance of the approach in identification of significant genomic regions associated with target traits. Therefore, the Indel-seq approach can be used for quick and precise identification of candidate genomic regions for any target traits in any crop species. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  14. Manufacturing validation of biologically functional T cells targeted to CD19 antigen for autologous adoptive cell therapy

    PubMed Central

    Hollyman, Daniel; Stefanski, Jolanta; Przybylowski, Mark; Bartido, Shirley; Borquez-Ojeda, Oriana; Taylor, Clare; Yeh, Raymond; Capacio, Vanessa; Olszewska, Malgorzata; Hosey, James; Sadelain, Michel; Brentjens, Renier J.; Rivière, Isabelle

    2009-01-01

    Summary Based on promising pre-clinical data demonstrating the eradication of systemic B cell malignancies by CD19-targeted T lymphocytes in vivo in SCID beige mouse models, we are launching Phase 1 clinical trials in patients with chronic lymphocytic leukemia (CLL) and acute lymphoblastic leukemia (ALL). We present here the validation of the bioprocess we developed for the production and expansion of clinical grade autologous T cells derived from patients with CLL. We demonstrate that T cells genetically modified with a replication-defective gammaretroviral vector derived from the Moloney murine leukemia virus encoding a chimeric antigen receptor (CAR) targeted to CD19 (1928z) can be expanded with Dynabeads® CD3/CD28. This bioprocess allows us to generate clinical doses of 1928z+ T cells in approximately 2 to 3 weeks in a large-scale semi-closed culture system using the Wave bioreactor. These 1928z+ T cells remain biologically functional not only in vitro but also in SCID beige mice bearing disseminated tumors. The validation requirements in terms of T cell expansion, T cell transduction with the 1928z CAR, biological activity, quality control testing and release criteria were met for all four validation runs using apheresis products from patients with CLL. Additionally, following expansion of the T cells, the diversity of the skewed Vβ T cell receptor repertoire was significantly restored. This validated process will be used in phase I clinical trials in patients with chemo-refractory CLL and in patients with relapsed ALL. It can also be adapted for other clinical trials involving the expansion and transduction of patient or donor T cells using any chimeric antigen receptor or T cell receptor. PMID:19238016

  15. Summary of tracking and identification methods

    NASA Astrophysics Data System (ADS)

    Blasch, Erik; Yang, Chun; Kadar, Ivan

    2014-06-01

    Over the last two decades, many solutions have arisen to combine target tracking estimation with classification methods. Target tracking includes developments from linear to non-linear and Gaussian to non-Gaussian processing. Pattern recognition includes detection, classification, recognition, and identification methods. Integrating tracking and pattern recognition has resulted in numerous approaches and this paper seeks to organize the various approaches. We discuss the terminology so as to have a common framework for various standards such as the NATO STANAG 4162 - Identification Data Combining Process. In a use case, we provide a comparative example highlighting that location information (as an example) with additional mission objectives from geographical, human, social, cultural, and behavioral modeling is needed to determine identification as classification alone does not allow determining identification or intent.

  16. Predicting Glycerophosphoinositol Identities in Lipidomic Datasets Using VaLID (Visualization and Phospholipid Identification)—An Online Bioinformatic Search Engine

    PubMed Central

    McDowell, Graeme S. V.; Taylor, Graeme P.; Fai, Stephen; Bennett, Steffany A. L.

    2014-01-01

    The capacity to predict and visualize all theoretically possible glycerophospholipid molecular identities present in lipidomic datasets is currently limited. To address this issue, we expanded the search-engine and compositional databases of the online Visualization and Phospholipid Identification (VaLID) bioinformatic tool to include the glycerophosphoinositol superfamily. VaLID v1.0.0 originally allowed exact and average mass libraries of 736,584 individual species from eight phospholipid classes: glycerophosphates, glyceropyrophosphates, glycerophosphocholines, glycerophosphoethanolamines, glycerophosphoglycerols, glycerophosphoglycerophosphates, glycerophosphoserines, and cytidine 5′-diphosphate 1,2-diacyl-sn-glycerols to be searched for any mass to charge value (with adjustable tolerance levels) under a variety of mass spectrometry conditions. Here, we describe an update that now includes all possible glycerophosphoinositols, glycerophosphoinositol monophosphates, glycerophosphoinositol bisphosphates, and glycerophosphoinositol trisphosphates. This update expands the total number of lipid species represented in the VaLID v2.0.0 database to 1,473,168 phospholipids. Each phospholipid can be generated in skeletal representation. A subset of species curated by the Canadian Institutes of Health Research Training Program in Neurodegenerative Lipidomics (CTPNL) team is provided as an array of high-resolution structures. VaLID is freely available and responds to all users through the CTPNL resources web site. PMID:24701584

  17. Effects of postidentification feedback on eyewitness identification and nonidentification confidence.

    PubMed

    Semmler, Carolyn; Brewer, Neil; Wells, Gary L

    2004-04-01

    Two experiments investigated new dimensions of the effect of confirming feedback on eyewitness identification confidence using target-absent and target-present lineups and (previously unused) unbiased witness instructions (i.e., "offender not present" option highlighted). In Experiment 1, participants viewed a crime video and were later asked to try to identify the thief from an 8-person target-absent photo array. Feedback inflated witness confidence for both mistaken identifications and correct lineup rejections. With target-present lineups in Experiment 2, feedback inflated confidence for correct and mistaken identifications and lineup rejections. Although feedback had no influence on the confidence-accuracy correlation, it produced clear overconfidence. Confidence inflation varied with the confidence measure reference point (i.e., retrospective vs. current confidence) and identification response latency.

  18. Genetic Validation of Aminoacyl-tRNA Synthetases as Drug Targets in Trypanosoma brucei

    PubMed Central

    Kalidas, Savitha; Cestari, Igor; Monnerat, Severine; Li, Qiong; Regmi, Sandesh; Hasle, Nicholas; Labaied, Mehdi; Parsons, Marilyn; Stuart, Kenneth

    2014-01-01

    Human African trypanosomiasis (HAT) is an important public health threat in sub-Saharan Africa. Current drugs are unsatisfactory, and new drugs are being sought. Few validated enzyme targets are available to support drug discovery efforts, so our goal was to obtain essentiality data on genes with proven utility as drug targets. Aminoacyl-tRNA synthetases (aaRSs) are known drug targets for bacterial and fungal pathogens and are required for protein synthesis. Here we survey the essentiality of eight Trypanosoma brucei aaRSs by RNA interference (RNAi) gene expression knockdown, covering an enzyme from each major aaRS class: valyl-tRNA synthetase (ValRS) (class Ia), tryptophanyl-tRNA synthetase (TrpRS-1) (class Ib), arginyl-tRNA synthetase (ArgRS) (class Ic), glutamyl-tRNA synthetase (GluRS) (class 1c), threonyl-tRNA synthetase (ThrRS) (class IIa), asparaginyl-tRNA synthetase (AsnRS) (class IIb), and phenylalanyl-tRNA synthetase (α and β) (PheRS) (class IIc). Knockdown of mRNA encoding these enzymes in T. brucei mammalian stage parasites showed that all were essential for parasite growth and survival in vitro. The reduced expression resulted in growth, morphological, cell cycle, and DNA content abnormalities. ThrRS was characterized in greater detail, showing that the purified recombinant enzyme displayed ThrRS activity and that the protein localized to both the cytosol and mitochondrion. Borrelidin, a known inhibitor of ThrRS, was an inhibitor of T. brucei ThrRS and showed antitrypanosomal activity. The data show that aaRSs are essential for T. brucei survival and are likely to be excellent targets for drug discovery efforts. PMID:24562907

  19. Targeted Mass Spectrometric Approach for Biomarker Discovery and Validation with Nonglycosylated Tryptic Peptides from N-linked Glycoproteins in Human Plasma*

    PubMed Central

    Lee, Ju Yeon; Kim, Jin Young; Park, Gun Wook; Cheon, Mi Hee; Kwon, Kyung-Hoon; Ahn, Yeong Hee; Moon, Myeong Hee; Lee, Hyoung–Joo; Paik, Young Ki; Yoo, Jong Shin

    2011-01-01

    A simple mass spectrometric approach for the discovery and validation of biomarkers in human plasma was developed by targeting nonglycosylated tryptic peptides adjacent to glycosylation sites in an N-linked glycoprotein, one of the most important biomarkers for early detection, prognoses, and disease therapies. The discovery and validation of novel biomarkers requires complex sample pretreatment steps, such as depletion of highly abundant proteins, enrichment of desired proteins, or the development of new antibodies. The current study exploited the steric hindrance of glycan units in N-linked glycoproteins, which significantly affects the efficiency of proteolytic digestion if an enzymatically active amino acid is adjacent to the N-linked glycosylation site. Proteolytic digestion then results in quantitatively different peptide products in accordance with the degree of glycosylation. The effect of glycan steric hindrance on tryptic digestion was first demonstrated using alpha-1-acid glycoprotein (AGP) as a model compound versus deglycosylated alpha-1-acid glycoprotein. Second, nonglycosylated tryptic peptide biomarkers, which generally show much higher sensitivity in mass spectrometric analyses than their glycosylated counterparts, were quantified in human hepatocellular carcinoma plasma using a label-free method with no need for N-linked glycoprotein enrichment. Finally, the method was validated using a multiple reaction monitoring analysis, demonstrating that the newly discovered nonglycosylated tryptic peptide targets were present at different levels in normal and hepatocellular carcinoma plasmas. The area under the receiver operating characteristic curve generated through analyses of nonglycosylated tryptic peptide from vitronectin precursor protein was 0.978, the highest observed in a group of patients with hepatocellular carcinoma. This work provides a targeted means of discovering and validating nonglycosylated tryptic peptides as biomarkers in human plasma

  20. Flanking sequence determination and specific PCR identification of transgenic wheat B102-1-2.

    PubMed

    Cao, Jijuan; Xu, Junyi; Zhao, Tongtong; Cao, Dongmei; Huang, Xin; Zhang, Piqiao; Luan, Fengxia

    2014-01-01

    The exogenous fragment sequence and flanking sequence between the exogenous fragment and recombinant chromosome of transgenic wheat B102-1-2 were successfully acquired using genome walking technology. The newly acquired exogenous fragment encoded the full-length sequence of transformed genes with transformed plasmid and corresponding functional genes including ubi, vector pBANF-bar, vector pUbiGUSPlus, vector HSP, reporter vector pUbiGUSPlus, promoter ubiquitin, and coli DH1. A specific polymerase chain reaction (PCR) identification method for transgenic wheat B102-1-2 was established on the basis of designed primers according to flanking sequence. This established specific PCR strategy was validated by using transgenic wheat, transgenic corn, transgenic soybean, transgenic rice, and non-transgenic wheat. A specifically amplified target band was observed only in transgenic wheat B102-1-2. Therefore, this method is characterized by high specificity, high reproducibility, rapid identification, and excellent accuracy for the identification of transgenic wheat B102-1-2.

  1. Identification and functional validation of HPV-mediated hypermethylation in head and neck squamous cell carcinoma

    PubMed Central

    2013-01-01

    Background Human papillomavirus-positive (HPV+) head and neck squamous cell carcinoma (HNSCC) represents a distinct clinical and epidemiological condition compared with HPV-negative (HPV-) HNSCC. To test the possible involvement of epigenetic modulation by HPV in HNSCC, we conducted a genome-wide DNA-methylation analysis. Methods Using laser-capture microdissection of 42 formalin-fixed paraffin wax-embedded (FFPE) HNSCCs, we generated DNA-methylation profiles of 18 HPV+ and 14 HPV- samples, using Infinium 450 k BeadArray technology. Methylation data were validated in two sets of independent HPV+/HPV- HNSCC samples (fresh-frozen samples and cell lines) using two independent methods (Infinium 450 k and whole-genome methylated DNA immunoprecipitation sequencing (MeDIP-seq)). For the functional analysis, an HPV- HNSCC cell line was transduced with lentiviral constructs containing the two HPV oncogenes (E6 and E7), and effects on methylation were assayed using the Infinium 450 k technology. Results and discussion Unsupervised clustering over the methylation variable positions (MVPs) with greatest variation showed that samples segregated in accordance with HPV status, but also that HPV+ tumors are heterogeneous. MVPs were significantly enriched at transcriptional start sites, leading to the identification of a candidate CpG island methylator phenotype in a sub-group of the HPV+ tumors. Supervised analysis identified a strong preponderance (87%) of MVPs towards hypermethylation in HPV+ HNSCC. Meta-analysis of our HNSCC and publicly available methylation data in cervical and lung cancers confirmed the observed DNA-methylation signature to be HPV-specific and tissue-independent. Grouping of MVPs into functionally more significant differentially methylated regions identified 43 hypermethylated promoter DMRs, including for three cadherins of the Polycomb group target genes. Integration with independent expression data showed strong negative correlation, especially for the

  2. Identification of microRNAs and their targets in Finger millet by high throughput sequencing.

    PubMed

    Usha, S; Jyothi, M N; Sharadamma, N; Dixit, Rekha; Devaraj, V R; Nagesh Babu, R

    2015-12-15

    MicroRNAs are short non-coding RNAs which play an important role in regulating gene expression by mRNA cleavage or by translational repression. The majority of identified miRNAs were evolutionarily conserved; however, others expressed in a species-specific manner. Finger millet is an important cereal crop; nonetheless, no practical information is available on microRNAs to date. In this study, we have identified 95 conserved microRNAs belonging to 39 families and 3 novel microRNAs by high throughput sequencing. For the identified conserved and novel miRNAs a total of 507 targets were predicted. 11 miRNAs were validated and tissue specificity was determined by stem loop RT-qPCR, Northern blot. GO analyses revealed targets of miRNA were involved in wide range of regulatory functions. This study implies large number of known and novel miRNAs found in Finger millet which may play important role in growth and development. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. In Vitro Evaluation of Molecular Tumor Targets in Nuclear Medicine: Immunohistochemistry Is One Option, but Under Which Conditions?

    PubMed

    Reubi, Jean Claude

    2017-12-01

    The identification of new molecular targets for diagnostic and therapeutic applications using in vitro methods is an important challenge in nuclear medicine. One such method is immunohistochemistry, increasingly popular because it is easy to perform. This review presents the case for conducting receptor immunohistochemistry to evaluate potential molecular targets in human tumor tissue sections. The focus is on the immunohistochemistry of G-protein-coupled receptors, one of the largest families of cell surface proteins, representing a major class of drug targets and thus playing an important role in nuclear medicine. This review identifies common pitfalls and challenges and provides guidelines on performing such immunohistochemical studies. An appropriate validation of the target is a prerequisite for developing robust and informative new molecular probes. © 2017 by the Society of Nuclear Medicine and Molecular Imaging.

  4. The neural substrates of action identification.

    PubMed

    Marsh, Abigail A; Kozak, Megan N; Wegner, Daniel M; Reid, Marguerite E; Yu, Henry H; Blair, R J R

    2010-12-01

    Mentalization is the process by which an observer views a target as possessing higher cognitive faculties such as goals, intentions and desires. Mentalization can be assessed using action identification paradigms, in which observers choose mentalistic (goals-focused) or mechanistic (action-focused) descriptions of targets' actions. Neural structures that play key roles in inferring goals and intentions from others' observed or imagined actions include temporo-parietal junction, ventral premotor cortex and extrastriate body area. We hypothesized that these regions play a role in action identification as well. Data collected using functional magnetic resonance imaging (fMRI) confirmed our predictions that activity in ventral premotor cortex and middle temporal gyrus near the extrastriate body area varies both as a function of the valence of the target and the extent to which actions are identified as goal-directed. In addition, the inferior parietal lobule is preferentially engaged when participants identify the actions of mentalized targets. Functional connectivity analyses suggest support from other regions, including the medial prefrontal cortex and amygdala, during mentalization. We found correlations between action identification and Autism Quotient scores, suggesting that understanding the neural correlates of action identification may enhance our understanding of the underpinnings of essential social cognitive processes.

  5. Messenger RNA biomarker signatures for forensic body fluid identification revealed by targeted RNA sequencing.

    PubMed

    Hanson, E; Ingold, S; Haas, C; Ballantyne, J

    2018-05-01

    The recovery of a DNA profile from the perpetrator or victim in criminal investigations can provide valuable 'source level' information for investigators. However, a DNA profile does not reveal the circumstances by which biological material was transferred. Some contextual information can be obtained by a determination of the tissue or fluid source of origin of the biological material as it is potentially indicative of some behavioral activity on behalf of the individual that resulted in its transfer from the body. Here, we sought to improve upon established RNA based methods for body fluid identification by developing a targeted multiplexed next generation mRNA sequencing assay comprising a panel of approximately equal sized gene amplicons. The multiplexed biomarker panel includes several highly specific gene targets with the necessary specificity to definitively identify most forensically relevant biological fluids and tissues (blood, semen, saliva, vaginal secretions, menstrual blood and skin). In developing the biomarker panel we evaluated 66 gene targets, with a progressive iteration of testing target combinations that exhibited optimal sensitivity and specificity using a training set of forensically relevant body fluid samples. The current assay comprises 33 targets: 6 blood, 6 semen, 6 saliva, 4 vaginal secretions, 5 menstrual blood and 6 skin markers. We demonstrate the sensitivity and specificity of the assay and the ability to identify body fluids in single source and admixed stains. A 16 sample blind test was carried out by one lab with samples provided by the other participating lab. The blinded lab correctly identified the body fluids present in 15 of the samples with the major component identified in the 16th. Various classification methods are being investigated to permit inference of the body fluid/tissue in dried physiological stains. These include the percentage of reads in a sample that are due to each of the 6 tissues/body fluids tested and

  6. Target selection for a hypervelocity asteroid intercept vehicle flight validation mission

    NASA Astrophysics Data System (ADS)

    Wagner, Sam; Wie, Bong; Barbee, Brent W.

    2015-02-01

    Asteroids and comets have collided with the Earth in the past and will do so again in the future. Throughout Earth's history these collisions have played a significant role in shaping Earth's biological and geological histories. The planetary defense community has been examining a variety of options for mitigating the impact threat of asteroids and comets that approach or cross Earth's orbit, known as near-Earth objects (NEOs). This paper discusses the preliminary study results of selecting small (100-m class) NEO targets and mission analysis and design trade-offs for validating the effectiveness of a Hypervelocity Asteroid Intercept Vehicle (HAIV) concept, currently being investigated for a NIAC (NASA Advanced Innovative Concepts) Phase 2 study. In particular this paper will focus on the mission analysis and design for single spacecraft direct impact trajectories, as well as several mission types that enable a secondary rendezvous spacecraft to observe the HAIV impact and evaluate it's effectiveness.

  7. Identification of Aspergillus fumigatus and Related Species by Nested PCR Targeting Ribosomal DNA Internal Transcribed Spacer Regions

    PubMed Central

    Zhao, Jun; Kong, Fanrong; Li, Ruoyu; Wang, Xiaohong; Wan, Zhe; Wang, Duanli

    2001-01-01

    Aspergillus fumigatus is the most common species that causes invasive aspergillosis. In order to identify A. fumigatus, partial ribosomal DNA (rDNA) from two to six strains of five different Aspergillus species was sequenced. By comparing sequence data from GenBank, we designed specific primer pairs targeting rDNA internal transcribed spacer (ITS) regions of A. fumigatus. A nested PCR method for identification of other A. fumigatus-related species was established by using the primers. To evaluate the specificities and sensitivities of those primers, 24 isolates of A. fumigatus and variants, 8 isolates of Aspergillus nidulans, 7 isolates of Aspergillus flavus and variants, 8 isolates of Aspergillus terreus, 9 isolates of Aspergillus niger, 1 isolate each of Aspergillus parasiticus, Aspergillus penicilloides, Aspergillus versicolor, Aspergillus wangduanlii, Aspergillus qizutongii, Aspergillus beijingensis, and Exophiala dermatitidis, 4 isolates of Candida, 4 isolates of bacteria, and human DNA were used. The nested PCR method specifically identified the A. fumigatus isolates and closely related species and showed a high degree of sensitivity. Additionally, four A. fumigatus strains that were recently isolated from our clinic were correctly identified by this method. Our results demonstrate that these primers are useful for the identification of A. fumigatus and closely related species in culture and suggest further studies for the identification of Aspergillus fumigatus species in clinical specimens. PMID:11376067

  8. Molecular Validation of PACE4 as a Target in Prostate Cancer12

    PubMed Central

    D'Anjou, François; Routhier, Sophie; Perreault, Jean-Pierre; Latil, Alain; Bonnel, David; Fournier, Isabelle; Salzet, Michel; Day, Robert

    2011-01-01

    Prostate cancer remains the single most prevalent cancer in men. Standard therapies are still limited and include androgen ablation that initially causes tumor regression. However, tumor cells eventually relapse and develop into a hormone-refractory prostate cancer. One of the current challenges in this disease is to define new therapeutic targets, which have been virtually unchanged in the past 30 years. Recent studies have suggested that the family of enzymes known as the proprotein convertases (PCs) is involved in various types of cancers and their progression. The present study examined PC expression in prostate cancer and validates one PC, namely PACE4, as a target. The evidence includes the observed high expression of PACE4 in all different clinical stages of human prostate tumor tissues. Gene silencing studies targeting PACE4 in the DU145 prostate cancer cell line produced cells (cell line 4-2) with slower proliferation rates, reduced clonogenic activity, and inability to grow as xenografts in nude mice. Gene expression and proteomic profiling of the 4-2 cell line reveals an increased expression of known cancer-related genes (e.g., GJA1, CD44, IGFBP6) that are downregulated in prostate cancer. Similarly, cancer genes whose expression is decreased in the 4-2 cell line were upregulated in prostate cancer (e.g., MUC1, IL6). The direct role of PACE4 in prostate cancer is most likely through the upregulated processing of growth factors or through the aberrant processing of growth factors leading to sustained cancer progression, suggesting that PACE4 holds a central role in prostate cancer. PMID:21633671

  9. Crowd Sourcing a New Paradigm for Interactome Driven Drug Target Identification in Mycobacterium tuberculosis

    PubMed Central

    Rohira, Harsha; Bhat, Ashwini G.; Passi, Anurag; Mukherjee, Keya; Choudhary, Kumari Sonal; Kumar, Vikas; Arora, Anshula; Munusamy, Prabhakaran; Subramanian, Ahalyaa; Venkatachalam, Aparna; S, Gayathri; Raj, Sweety; Chitra, Vijaya; Verma, Kaveri; Zaheer, Salman; J, Balaganesh; Gurusamy, Malarvizhi; Razeeth, Mohammed; Raja, Ilamathi; Thandapani, Madhumohan; Mevada, Vishal; Soni, Raviraj; Rana, Shruti; Ramanna, Girish Muthagadhalli; Raghavan, Swetha; Subramanya, Sunil N.; Kholia, Trupti; Patel, Rajesh; Bhavnani, Varsha; Chiranjeevi, Lakavath; Sengupta, Soumi; Singh, Pankaj Kumar; Atray, Naresh; Gandhi, Swati; Avasthi, Tiruvayipati Suma; Nisthar, Shefin; Anurag, Meenakshi; Sharma, Pratibha; Hasija, Yasha; Dash, Debasis; Sharma, Arun; Scaria, Vinod; Thomas, Zakir; Chandra, Nagasuma; Brahmachari, Samir K.; Bhardwaj, Anshu

    2012-01-01

    A decade since the availability of Mycobacterium tuberculosis (Mtb) genome sequence, no promising drug has seen the light of the day. This not only indicates the challenges in discovering new drugs but also suggests a gap in our current understanding of Mtb biology. We attempt to bridge this gap by carrying out extensive re-annotation and constructing a systems level protein interaction map of Mtb with an objective of finding novel drug target candidates. Towards this, we synergized crowd sourcing and social networking methods through an initiative ‘Connect to Decode’ (C2D) to generate the first and largest manually curated interactome of Mtb termed ‘interactome pathway’ (IPW), encompassing a total of 1434 proteins connected through 2575 functional relationships. Interactions leading to gene regulation, signal transduction, metabolism, structural complex formation have been catalogued. In the process, we have functionally annotated 87% of the Mtb genome in context of gene products. We further combine IPW with STRING based network to report central proteins, which may be assessed as potential drug targets for development of drugs with least possible side effects. The fact that five of the 17 predicted drug targets are already experimentally validated either genetically or biochemically lends credence to our unique approach. PMID:22808064

  10. Drug Target Mining and Analysis of the Chinese Tree Shrew for Pharmacological Testing

    PubMed Central

    Liu, Jie; Lee, Wen-hui; Zhang, Yun

    2014-01-01

    The discovery of new drugs requires the development of improved animal models for drug testing. The Chinese tree shrew is considered to be a realistic candidate model. To assess the potential of the Chinese tree shrew for pharmacological testing, we performed drug target prediction and analysis on genomic and transcriptomic scales. Using our pipeline, 3,482 proteins were predicted to be drug targets. Of these predicted targets, 446 and 1,049 proteins with the highest rank and total scores, respectively, included homologs of targets for cancer chemotherapy, depression, age-related decline and cardiovascular disease. Based on comparative analyses, more than half of drug target proteins identified from the tree shrew genome were shown to be higher similarity to human targets than in the mouse. Target validation also demonstrated that the constitutive expression of the proteinase-activated receptors of tree shrew platelets is similar to that of human platelets but differs from that of mouse platelets. We developed an effective pipeline and search strategy for drug target prediction and the evaluation of model-based target identification for drug testing. This work provides useful information for future studies of the Chinese tree shrew as a source of novel targets for drug discovery research. PMID:25105297

  11. Identification and validation of Asteraceae miRNAs by the expressed sequence tag analysis.

    PubMed

    Monavar Feshani, Aboozar; Mohammadi, Saeed; Frazier, Taylor P; Abbasi, Abbas; Abedini, Raha; Karimi Farsad, Laleh; Ehya, Farveh; Salekdeh, Ghasem Hosseini; Mardi, Mohsen

    2012-02-10

    MicroRNAs (miRNAs) are small non-coding RNA molecules that play a vital role in the regulation of gene expression. Despite their identification in hundreds of plant species, few miRNAs have been identified in the Asteraceae, a large family that comprises approximately one tenth of all flowering plants. In this study, we used the expressed sequence tag (EST) analysis to identify potential conserved miRNAs and their putative target genes in the Asteraceae. We applied quantitative Real-Time PCR (qRT-PCR) to confirm the expression of eight potential miRNAs in Carthamus tinctorius and Helianthus annuus. We also performed qRT-PCR analysis to investigate the differential expression pattern of five newly identified miRNAs during five different cotyledon growth stages in safflower. Using these methods, we successfully identified and characterized 151 potentially conserved miRNAs, belonging to 26 miRNA families, in 11 genus of Asteraceae. EST analysis predicted that the newly identified conserved Asteraceae miRNAs target 130 total protein-coding ESTs in sunflower and safflower, as well as 433 additional target genes in other plant species. We experimentally confirmed the existence of seven predicted miRNAs, (miR156, miR159, miR160, miR162, miR166, miR396, and miR398) in safflower and sunflower seedlings. We also observed that five out of eight miRNAs are differentially expressed during cotyledon development. Our results indicate that miRNAs may be involved in the regulation of gene expression during seed germination and the formation of the cotyledons in the Asteraceae. The findings of this study might ultimately help in the understanding of miRNA-mediated gene regulation in important crop species. Copyright © 2011 Elsevier B.V. All rights reserved.

  12. Effects of video compression on target acquisition performance

    NASA Astrophysics Data System (ADS)

    Espinola, Richard L.; Cha, Jae; Preece, Bradley

    2008-04-01

    The bandwidth requirements of modern target acquisition systems continue to increase with larger sensor formats and multi-spectral capabilities. To obviate this problem, still and moving imagery can be compressed, often resulting in greater than 100 fold decrease in required bandwidth. Compression, however, is generally not error-free and the generated artifacts can adversely affect task performance. The U.S. Army RDECOM CERDEC Night Vision and Electronic Sensors Directorate recently performed an assessment of various compression techniques on static imagery for tank identification. In this paper, we expand this initial assessment by studying and quantifying the effect of various video compression algorithms and their impact on tank identification performance. We perform a series of controlled human perception tests using three dynamic simulated scenarios: target moving/sensor static, target static/sensor static, sensor tracking the target. Results of this study will quantify the effect of video compression on target identification and provide a framework to evaluate video compression on future sensor systems.

  13. A computable phenotype for asthma case identification in adult and pediatric patients: External validation in the Chicago Area Patient-Outcomes Research Network (CAPriCORN).

    PubMed

    Afshar, Majid; Press, Valerie G; Robison, Rachel G; Kho, Abel N; Bandi, Sindhura; Biswas, Ashvini; Avila, Pedro C; Kumar, Harsha Vardhan Madan; Yu, Byung; Naureckas, Edward T; Nyenhuis, Sharmilee M; Codispoti, Christopher D

    2017-10-13

    Comprehensive, rapid, and accurate identification of patients with asthma for clinical care and engagement in research efforts is needed. The original development and validation of a computable phenotype for asthma case identification occurred at a single institution in Chicago and demonstrated excellent test characteristics. However, its application in a diverse payer mix, across different health systems and multiple electronic health record vendors, and in both children and adults was not examined. The objective of this study is to externally validate the computable phenotype across diverse Chicago institutions to accurately identify pediatric and adult patients with asthma. A cohort of 900 asthma and control patients was identified from the electronic health record between January 1, 2012 and November 30, 2014. Two physicians at each site independently reviewed the patient chart to annotate cases. The inter-observer reliability between the physician reviewers had a κ-coefficient of 0.95 (95% CI 0.93-0.97). The accuracy, sensitivity, specificity, negative predictive value, and positive predictive value of the computable phenotype were all above 94% in the full cohort. The excellent positive and negative predictive values in this multi-center external validation study establish a useful tool to identify asthma cases in in the electronic health record for research and care. This computable phenotype could be used in large-scale comparative-effectiveness trials.

  14. Targeting mammalian organelles with internalizing phage (iPhage) libraries

    PubMed Central

    Rangel, Roberto; Dobroff, Andrey S.; Guzman-Rojas, Liliana; Salmeron, Carolina C.; Gelovani, Juri G.; Sidman, Richard L.; Pasqualini, Renata; Arap, Wadih

    2015-01-01

    Techniques largely used for protein interaction studies and discovery of intracellular receptors, such as affinity capture complex purification and yeast two-hybrid, may produce inaccurate datasets due to protein insolubility, transient or weak protein interactions, or irrelevant intracellular context. A versatile tool to overcome these limitations as well as to potentially create vaccines and engineer peptides and antibodies as targeted diagnostic and therapeutic agents, is the phage display technique. We have recently developed a new technology for screening internalizing phage (iPhage) vectors and libraries utilizing a ligand/receptor-independent mechanism to penetrate eukaryotic cells. iPhage particles provide a unique discovery platform for combinatorial intracellular targeting of organelle ligands along with their corresponding receptors and to fingerprint functional protein domains in living cells. Here we explain the design, cloning, construction, and production of iPhage-based vectors and libraries, along with basic ligand-receptor identification and validation methodologies for organelle receptors. An iPhage library screening can be performed in ~8 weeks. PMID:24030441

  15. Molecular basis for the action of a dietary flavonoid revealed by the comprehensive identification of apigenin human targets

    PubMed Central

    Arango, Daniel; Morohashi, Kengo; Yilmaz, Alper; Kuramochi, Kouji; Parihar, Arti; Brahimaj, Bledi; Grotewold, Erich; Doseff, Andrea I.

    2013-01-01

    Flavonoids constitute the largest class of dietary phytochemicals, adding essential health value to our diet, and are emerging as key nutraceuticals. Cellular targets for dietary phytochemicals remain largely unknown, posing significant challenges for the regulation of dietary supplements and the understanding of how nutraceuticals provide health value. Here, we describe the identification of human cellular targets of apigenin, a flavonoid abundantly present in fruits and vegetables, using an innovative high-throughput approach that combines phage display with second generation sequencing. The 160 identified high-confidence candidate apigenin targets are significantly enriched in three main functional categories: GTPase activation, membrane transport, and mRNA metabolism/alternative splicing. This last category includes the heterogeneous nuclear ribonucleoprotein A2 (hnRNPA2), a factor involved in splicing regulation, mRNA stability, and mRNA transport. Apigenin binds to the C-terminal glycine-rich domain of hnRNPA2, preventing hnRNPA2 from forming homodimers, and therefore, it perturbs the alternative splicing of several human hnRNPA2 targets. Our results provide a framework to understand how dietary phytochemicals exert their actions by binding to many functionally diverse cellular targets. In turn, some of them may modulate the activity of a large number of downstream genes, which is exemplified here by the effects of apigenin on the alternative splicing activity of hnRNPA2. Hence, in contrast to small-molecule pharmaceuticals designed for defined target specificity, dietary phytochemicals affect a large number of cellular targets with varied affinities that, combined, result in their recognized health benefits. PMID:23697369

  16. Finite strain transient creep of D16T alloy: identification and validation employing heterogeneous tests

    NASA Astrophysics Data System (ADS)

    Shutov, A. V.; Larichkin, A. Yu

    2017-10-01

    A cyclic creep damage model, previously proposed by the authors, is modified for a better description of the transient creep of D16T alloy observed in the finite strain range under rapidly changing stresses. The new model encompasses the concept of kinematic hardening, which allows us to account for the creep-induced anisotropy. The model kinematics is based on the nested multiplicative split of the deformation gradient, proposed by Lion. The damage evolution is accounted for by the classical Kachanov-Rabotnov approach. The material parameters are identified using experimental data on cyclic torsion of thick-walled samples with different holding times between load reversals. For the validation of the proposed material model, an additional experiment is analyzed. Although this additional test is not involved in the identification procedure, the proposed cyclic creep damage model describes it accurately.

  17. Therapeutic Targeting of Eosinophil Adhesion and Accumulation in Allergic Conjunctivitis

    PubMed Central

    Baiula, Monica; Bedini, Andrea; Carbonari, Gioia; Dattoli, Samantha Deianira; Spampinato, Santi

    2012-01-01

    Considerable evidence indicates that eosinophils are important effectors of ocular allergy. Increased worldwide prevalence of allergic eye pathologies has stimulated the identification of novel drug targets, including eosinophils and adhesion molecules. Accumulation of eosinophils in the eye is a key event in the onset and maintenance of allergic inflammation and is mediated by different adhesion molecules. Antihistamines with multiple mechanisms of action can be effective during the early and late phases of allergic conjunctivitis by blocking the interaction between β1 integrins and vascular cell adhesion molecule (VCAM)-1. Small molecule antagonists that target key elements in the process of eosinophil recruitment have been identified and reinforce the validity of α4β1 integrin as a therapeutic target. Glucocorticoids are among the most effective drugs for ocular allergy, but their use is limited by adverse effects. Novel dissociated glucocorticoids can prevent eosinophil accumulation and induce apoptosis of eosinophils, making them promising candidates for ophthalmic drugs. This article reviews recent understanding of the role of adhesion molecules in eosinophil recruitment in the inflamed conjunctiva along with effective treatments for allergic conjunctivitis. PMID:23271999

  18. Precise Geolocation Of Persistent Scatterers Aided And Validated By Lidar DSM

    NASA Astrophysics Data System (ADS)

    Chang, Ling; Dheenathayalan, Prabu; Hanessen, Ramon

    2013-12-01

    Persistent Scatterers (PS) interferometry results in the de- formation history of time-coherent scatterers. Although several applications focus on smooth, spatially correlated signals, we aim for the detection, identification and analysis of single anomalies. These targets can be indicative of, e.g., strain in structures, potentially leading to the failure of such structures. For the identification and analysis it is of the greatest importance to know the exact position of the effective scattering center, to avoid an improper interpretation of the driving mechanism. Here we present an approach to optimize the geolocation of important scatterers, when necessary aided by an a priori Lidar-derived DSM (AHN-1 data) with 15cm and 5m resolution in vertical and horizontal directions, respectively. The DSM is also used to validate the geocoding. We implement our approach on a near-collapse event of a shopping mall in Heerlen, the Netherlands, to generate the precise geolocation of local PS points.

  19. Identification of Thiotetronic Acid Antibiotic Biosynthetic Pathways by Target-directed Genome Mining.

    PubMed

    Tang, Xiaoyu; Li, Jie; Millán-Aguiñaga, Natalie; Zhang, Jia Jia; O'Neill, Ellis C; Ugalde, Juan A; Jensen, Paul R; Mantovani, Simone M; Moore, Bradley S

    2015-12-18

    Recent genome sequencing efforts have led to the rapid accumulation of uncharacterized or "orphaned" secondary metabolic biosynthesis gene clusters (BGCs) in public databases. This increase in DNA-sequenced big data has given rise to significant challenges in the applied field of natural product genome mining, including (i) how to prioritize the characterization of orphan BGCs and (ii) how to rapidly connect genes to biosynthesized small molecules. Here, we show that by correlating putative antibiotic resistance genes that encode target-modified proteins with orphan BGCs, we predict the biological function of pathway specific small molecules before they have been revealed in a process we call target-directed genome mining. By querying the pan-genome of 86 Salinispora bacterial genomes for duplicated house-keeping genes colocalized with natural product BGCs, we prioritized an orphan polyketide synthase-nonribosomal peptide synthetase hybrid BGC (tlm) with a putative fatty acid synthase resistance gene. We employed a new synthetic double-stranded DNA-mediated cloning strategy based on transformation-associated recombination to efficiently capture tlm and the related ttm BGCs directly from genomic DNA and to heterologously express them in Streptomyces hosts. We show the production of a group of unusual thiotetronic acid natural products, including the well-known fatty acid synthase inhibitor thiolactomycin that was first described over 30 years ago, yet never at the genetic level in regards to biosynthesis and autoresistance. This finding not only validates the target-directed genome mining strategy for the discovery of antibiotic producing gene clusters without a priori knowledge of the molecule synthesized but also paves the way for the investigation of novel enzymology involved in thiotetronic acid natural product biosynthesis.

  20. Fast and Confident: Postdicting Eyewitness Identification Accuracy in a Field Study

    ERIC Educational Resources Information Center

    Sauerland, Melanie; Sporer, Siegfried L.

    2009-01-01

    The combined postdictive value of postdecision confidence, decision time, and Remember-Know-Familiar (RKF) judgments as markers of identification accuracy was evaluated with 10 targets and 720 participants. In a pedestrian area, passers-by were asked for directions. Identifications were made from target-absent or target-present lineups. Fast…

  1. Interventions targeted at primary care practitioners to improve the identification and referral of patients with co-morbid obesity: a realist review protocol.

    PubMed

    Blane, David N; Macdonald, Sara; Morrison, David; O'Donnell, Catherine A

    2015-05-01

    Obesity is one of the most significant public health challenges in the developed world. Recent policy has suggested that more can be done in primary care to support adults with obesity. In particular, general practitioners (GPs) and practice nurses (PNs) could improve the identification and referral of adults with obesity to appropriate weight management services. Previous interventions targeted at primary care practitioners in this area have had mixed results, suggesting a more complex interplay between patients, practitioners, and systems. The objectives of this review are (i) to identify the underlying 'programme theory' of interventions targeted at primary care practitioners to improve the identification and referral of adults with obesity and (ii) to explore how and why GPs and PNs identify and refer individuals with obesity, particularly in the context of weight-related co-morbidity. This protocol will explain the rationale for using a realist review approach and outline the key steps in this process. Realist review is a theory-led approach to knowledge synthesis that provides an explanatory analysis aimed at discerning what works, for whom, in what circumstances, how, and why. In this review, scoping interviews with key stakeholders involved in the planning and delivery of adult weight management services in Scotland helped to inform the identification of formal theories - from psychology, sociology, and implementation science - that will be tested as the review progresses. A comprehensive search strategy is described, including scope for iterative searching. Data analysis is outlined in three stages (describing context-mechanism-outcome configurations, exploring patterns in these configurations, and developing and testing middle-range theories, informed by the formal theories previously identified), culminating in the production of explanatory programme theory that considers individual, interpersonal, and institutional/systems-level components. This is the

  2. Evaluation of matrix-assisted laser desorption/ionization time of flight mass spectrometry for the identification of ceratopogonid and culicid larvae.

    PubMed

    Steinmann, I C; Pflüger, V; Schaffner, F; Mathis, A; Kaufmann, C

    2013-03-01

    Matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) was evaluated for the rapid identification of ceratopogonid larvae. Optimal sample preparation as evaluated with laboratory-reared biting midges Culicoides nubeculosus was the homogenization of gut-less larvae in 10% formic acid, and analysis of 0.2 mg/ml crude protein homogenate mixed with SA matrix at a ratio of 1:1.5. Using 5 larvae each of 4 ceratopogonid species (C. nubeculosus, C. obsoletus, C. decor, and Dasyhelea sp.) and of 2 culicid species (Aedes aegypti, Ae. japonicus), biomarker mass sets between 27 and 33 masses were determined. In a validation study, 67 larvae belonging to the target species were correctly identified by automated database-based identification (91%) or manual full comparison (9%). Four specimens of non-target species did not yield identification. As anticipated for holometabolous insects, the biomarker mass sets of adults cannot be used for the identification of larvae, and vice versa, because they share only very few similar masses as shown for C. nubeculosus, C. obsoletus, and Ae. japonicus. Thus, protein profiling by MALDI-TOF as a quick, inexpensive and accurate alternative tool is applicable to identify insect larvae of vector species collected in the field.

  3. Analytic validation and real-time clinical application of an amplicon-based targeted gene panel for advanced cancer

    PubMed Central

    Wing, Michele R.; Reeser, Julie W.; Smith, Amy M.; Reeder, Matthew; Martin, Dorrelyn; Jewell, Benjamin M.; Datta, Jharna; Miya, Jharna; Monk, J. Paul; Mortazavi, Amir; Otterson, Gregory A.; Goldberg, Richard M.; VanDeusen, Jeffrey B.; Cole, Sharon; Dittmar, Kristin; Jaiswal, Sunny; Kinzie, Matthew; Waikhom, Suraj; Freud, Aharon G.; Zhou, Xiao-Ping; Chen, Wei; Bhatt, Darshna; Roychowdhury, Sameek

    2017-01-01

    Multiplex somatic testing has emerged as a strategy to test patients with advanced cancer. We demonstrate our analytic validation approach for a gene hotspot panel and real-time prospective clinical application for any cancer type. The TruSight Tumor 26 assay amplifies 85 somatic hotspot regions across 26 genes. Using cell line and tumor mixes, we observed that 100% of the 14,715 targeted bases had at least 1000x raw coverage. We determined the sensitivity (100%, 95% CI: 96-100%), positive predictive value (100%, 95% CI: 96-100%), reproducibility (100% concordance), and limit of detection (3% variant allele frequency at 1000x read depth) of this assay to detect single nucleotide variants and small insertions and deletions. Next, we applied the assay prospectively in a clinical tumor sequencing study to evaluate 174 patients with metastatic or advanced cancer, including frozen tumors, formalin-fixed tumors, and enriched peripheral blood mononuclear cells in hematologic cancers. We reported one or more somatic mutations in 89 (53%) of the sequenced tumors (167 passing quality filters). Forty-three of these patients (26%) had mutations that would enable eligibility for targeted therapies. This study demonstrates the validity and feasibility of applying TruSight Tumor 26 for pan-cancer testing using multiple specimen types. PMID:29100271

  4. The use of alcohol use disorders identification test (AUDIT) in detecting alcohol use disorder and risk drinking in the general population: validation of AUDIT using schedules for clinical assessment in neuropsychiatry.

    PubMed

    Lundin, Andreas; Hallgren, Mats; Balliu, Natalja; Forsell, Yvonne

    2015-01-01

    The alcohol use disorders identification test (AUDIT) and AUDIT-Consumption (AUDIT-C) are commonly used in population surveys but there are few validations studies in the general population. Validity should be estimated in samples close to the targeted population and setting. This study aims to validate AUDIT and AUDIT-C in a general population sample (PART) in Stockholm, Sweden. We used a general population subsample age 20 to 64 that answered a postal questionnaire including AUDIT who later participated in a psychiatric interview (n = 1,093). Interviews using Schedules for Clinical Assessment in Neuropsychiatry was used as criterion standard. Diagnoses were set according to the fourth version of the Diagnostic and Statistical Manual of Mental Disorders (DSM-IV). Agreement between the diagnostic test and criterion standard was measured with area under the receiver operator characteristics curve (AUC). A total of 1,086 (450 men and 636 women) of the interview participants completed AUDIT. There were 96 individuals with DSM-IV-alcohol dependence, 36 DSM-IV-Alcohol Abuse, and 153 Risk drinkers. AUCs were for DSM-IV-alcohol use disorder 0.90 (AUDIT-C 0.85); DSM-IV-dependence 0.94 (AUDIT-C 0.89); risk drinking 0.80 (AUDIT-C 0.80); and any criterion 0.87 (AUDIT-C 0.84). In this general population sample, AUDIT and AUDIT-C performed outstanding or excellent in identifying dependency, risk drinking, alcohol use disorder, any disorder, or risk drinking. Copyright © 2015 by the Research Society on Alcoholism.

  5. Human L-DOPA decarboxylase mRNA is a target of miR-145: A prediction to validation workflow.

    PubMed

    Papadopoulos, Emmanuel I; Fragoulis, Emmanuel G; Scorilas, Andreas

    2015-01-10

    l-DOPA decarboxylase (DDC) is a multiply-regulated gene which encodes the enzyme that catalyzes the biosynthesis of dopamine in humans. MicroRNAs comprise a novel class of endogenously transcribed small RNAs that can post-transcriptionally regulate the expression of various genes. Given that the mechanism of microRNA target recognition remains elusive, several genes, including DDC, have not yet been identified as microRNA targets. Nevertheless, a number of specifically designed bioinformatic algorithms provide candidate miRNAs for almost every gene, but still their results exhibit moderate accuracy and should be experimentally validated. Motivated by the above, we herein sought to discover a microRNA that regulates DDC expression. By using the current algorithms according to bibliographic recommendations we found that miR-145 could be predicted with high specificity as a candidate regulatory microRNA for DDC expression. Thus, a validation experiment followed by firstly transfecting an appropriate cell culture system with a synthetic miR-145 sequence and sequentially assessing the mRNA and protein levels of DDC via real-time PCR and Western blotting, respectively. Our analysis revealed that miR-145 had no significant impact on protein levels of DDC but managed to dramatically downregulate its mRNA expression. Overall, the experimental and bioinformatic analysis conducted herein indicate that miR-145 has the ability to regulate DDC mRNA expression and potentially this occurs by recognizing its mRNA as a target. Copyright © 2014. Published by Elsevier B.V.

  6. Targeting specific facial variation for different identification tasks.

    PubMed

    Aeria, Gillian; Claes, Peter; Vandermeulen, Dirk; Clement, John Gerald

    2010-09-10

    A conceptual framework that allows faces to be studied and compared objectively with biological validity is presented. The framework is a logical extension of modern morphometrics and statistical shape analysis techniques. Three dimensional (3D) facial scans were collected from 255 healthy young adults. One scan depicted a smiling facial expression and another scan depicted a neutral expression. These facial scans were modelled in a Principal Component Analysis (PCA) space where Euclidean (ED) and Mahalanobis (MD) distances were used to form similarity measures. Within this PCA space, property pathways were calculated that expressed the direction of change in facial expression. Decomposition of distances into property-independent (D1) and dependent components (D2) along these pathways enabled the comparison of two faces in terms of the extent of a smiling expression. The performance of all distances was tested and compared in dual types of experiments: Classification tasks and a Recognition task. In the Classification tasks, individual facial scans were assigned to one or more population groups of smiling or neutral scans. The property-dependent (D2) component of both Euclidean and Mahalanobis distances performed best in the Classification task, by correctly assigning 99.8% of scans to the right population group. The recognition task tested if a scan of an individual depicting a smiling/neutral expression could be positively identified when shown a scan of the same person depicting a neutral/smiling expression. ED1 and MD1 performed best, and correctly identified 97.8% and 94.8% of individual scans respectively as belonging to the same person despite differences in facial expression. It was concluded that decomposed components are superior to straightforward distances in achieving positive identifications and presents a novel method for quantifying facial similarity. Additionally, although the undecomposed Mahalanobis distance often used in practice outperformed

  7. Biased lineup instructions and face identification from video images.

    PubMed

    Thompson, W Burt; Johnson, Jaime

    2008-01-01

    Previous eyewitness memory research has shown that biased lineup instructions reduce identification accuracy, primarily by increasing false-positive identifications in target-absent lineups. Because some attempts at identification do not rely on a witness's memory of the perpetrator but instead involve matching photos to images on surveillance video, the authors investigated the effects of biased instructions on identification accuracy in a matching task. In Experiment 1, biased instructions did not affect the overall accuracy of participants who used video images as an identification aid, but nearly all correct decisions occurred with target-present photo spreads. Both biased and unbiased instructions resulted in high false-positive rates. In Experiment 2, which focused on video-photo matching accuracy with target-absent photo spreads, unbiased instructions led to more correct responses (i.e., fewer false positives). These findings suggest that investigators should not relax precautions against biased instructions when people attempt to match photos to an unfamiliar person recorded on video.

  8. In situ identification of nocardioform actinomycetes in activated sludge using fluorescent rRNA-targeted oligonucleotide probes.

    PubMed

    Schuppler, M; Wagner, M; Schön, G; Göbel, U B

    1998-01-01

    Hitherto, few environmental samples have been investigated by a 'full cycle rRNA analysis'. Here the results of in situ hybridization experiments with specific rRNA-targeted oligonucleotide probes developed on the basis of new sequences derived from a previously described comparative 16S rRNA analysis of nocardioform actinomycetes in activated sludge are reported. Application of the specific probes enabled identification and discrimination of the distinct populations of nocardioform actinomycetes in activated sludge. One of the specific probes (DLP) detected rod-shaped bacteria which were found in 13 of the 16 investigated sludge samples from various wastewater treatment plants, suggesting their importance in the wastewater treatment process. Another probe (GLP2) hybridized with typically branched filaments of nocardioforms mainly found in samples from enhanced biological phosphorus removal plants, suggesting that these bacteria are involved in sludge foaming. The combination of in situ hybridization with fluorescently labelled rRNA-targeted oligonucleotide probes and confocal laser scanning microscopy improved the detection of nocardioform actinomycetes, which often showed only weak signals inside the activated-sludge flocs.

  9. Identification of Molecular Targets for 4,5-Dichloro-2-n-octyl-4-isothiazolin-3-one (DCOIT) in Teleosts: New Insight into Mechanism of Toxicity.

    PubMed

    Chen, Lianguo; Au, Doris W T; Hu, Chenyan; Peterson, Drew R; Zhou, Bingsheng; Qian, Pei-Yuan

    2017-02-07

    Environmental pollutants are capable of concomitantly inducing diverse toxic effects. However, it is largely unknown which effects are directly induced and which effects are secondary, thus calling for definitive identification of the initiating molecular event for a pollutant to elucidate the mechanism of toxicity. In the present study, affinity pull-down assays were used to identify target proteins for 4,5-dichloro-2-n-octyl-4-isothiazolin-3-one (DCOIT), a costal pollutant of emerging concern, in various tissues (e.g., brain, liver, plasma, and gonad) from marine medaka (Oryzias melastigma) and zebrafish (Danio rerio). Pull-down results showed that, in male and female brains from medaka and zebrafish, DCOIT had a consistently high affinity for G protein alpha subunits (Gα), suggesting the targeted effects of DCOIT on signaling transduction from G protein-coupled receptors (GPCRs) and an extrapolatable mode of action in teleost brains. Validation using recombinant proteins and molecular docking analysis confirmed that binding of DCOIT to Gα protein competitively inhibited its activation by substrate. Considering the involvement of GPCRs in the regulation of myriad biological processes, including the hypothalamus-pituitary-gonadal-liver axis, binding of DCOIT to upstream Gα proteins in the brain may provide a plausible explanation for the diversity of toxic effects resulting from DCOIT challenge, especially abnormal hormonal production through the mitogen-activated protein kinase pathway. A new mechanism of action based on GPCR signaling is thus hypothesized for endocrine disrupting chemicals and warrants further research to clearly elucidate the link between GPCR signaling and endocrine disruption.

  10. Validating presupposed versus focused text information.

    PubMed

    Singer, Murray; Solar, Kevin G; Spear, Jackie

    2017-04-01

    There is extensive evidence that readers continually validate discourse accuracy and congruence, but that they may also overlook conspicuous text contradictions. Validation may be thwarted when the inaccurate ideas are embedded sentence presuppositions. In four experiments, we examined readers' validation of presupposed ("given") versus new text information. Throughout, a critical concept, such as a truck versus a bus, was introduced early in a narrative. Later, a character stated or thought something about the truck, which therefore matched or mismatched its antecedent. Furthermore, truck was presented as either given or new information. Mismatch target reading times uniformly exceeded the matching ones by similar magnitudes for given and new concepts. We obtained this outcome using different grammatical constructions and with different antecedent-target distances. In Experiment 4, we examined only given critical ideas, but varied both their matching and the main verb's factivity (e.g., factive know vs. nonfactive think). The Match × Factivity interaction closely resembled that previously observed for new target information (Singer, 2006). Thus, readers can successfully validate given target information. Although contemporary theories tend to emphasize either deficient or successful validation, both types of theory can accommodate the discourse and reader variables that may regulate validation.

  11. Model Identification in Time-Series Analysis: Some Empirical Results.

    ERIC Educational Resources Information Center

    Padia, William L.

    Model identification of time-series data is essential to valid statistical tests of intervention effects. Model identification is, at best, inexact in the social and behavioral sciences where one is often confronted with small numbers of observations. These problems are discussed, and the results of independent identifications of 130 social and…

  12. Identification and Expression Analyses of miRNAs from Two Contrasting Flower Color Cultivars of Canna by Deep Sequencing.

    PubMed

    Roy, Sribash; Tripathi, Abhinandan Mani; Yadav, Amrita; Mishra, Parneeta; Nautiyal, Chandra Shekhar

    2016-01-01

    miRNAs are endogenous small RNA (sRNA) that play critical roles in plant development processes. Canna is an ornamental plant belonging to family Cannaceae. Here, we report for the first time the identification and differential expression of miRNAs in two contrasting flower color cultivars of Canna, Tropical sunrise and Red president. A total of 313 known miRNAs belonging to 78 miRNA families were identified from both the cultivars. Thirty one miRNAs (17 miRNA families) were specific to Tropical sunrise and 43 miRNAs (10 miRNA families) were specific to Red president. Thirty two and 18 putative new miRNAs were identified from Tropical sunrise and Red president, respectively. One hundred and nine miRNAs were differentially expressed in the two cultivars targeting 1343 genes. Among these, 16 miRNAs families targeting 60 genes were involved in flower development related traits and five miRNA families targeting five genes were involved in phenyl propanoid and pigment metabolic processes. We further validated the expression analysis of a few miRNA and their target genes by qRT-PCR. Transcription factors were the major miRNA targets identified. Target validation of a few randomly selected miRNAs by RLM-RACE was performed but was successful with only miR162. These findings will help in understanding flower development processes, particularly the color development in Canna.

  13. Identification of Small Molecule Translesion Synthesis Inhibitors That Target the Rev1-CT/RIR Protein-Protein Interaction.

    PubMed

    Sail, Vibhavari; Rizzo, Alessandro A; Chatterjee, Nimrat; Dash, Radha C; Ozen, Zuleyha; Walker, Graham C; Korzhnev, Dmitry M; Hadden, M Kyle

    2017-07-21

    Translesion synthesis (TLS) is an important mechanism through which proliferating cells tolerate DNA damage during replication. The mutagenic Rev1/Polζ-dependent branch of TLS helps cancer cells survive first-line genotoxic chemotherapy and introduces mutations that can contribute to the acquired resistance so often observed with standard anticancer regimens. As such, inhibition of Rev1/Polζ-dependent TLS has recently emerged as a strategy to enhance the efficacy of first-line chemotherapy and reduce the acquisition of chemoresistance by decreasing tumor mutation rate. The TLS DNA polymerase Rev1 serves as an integral scaffolding protein that mediates the assembly of the active multiprotein TLS complexes. Protein-protein interactions (PPIs) between the C-terminal domain of Rev1 (Rev1-CT) and the Rev1-interacting region (RIR) of other TLS DNA polymerases play an essential role in regulating TLS activity. To probe whether disrupting the Rev1-CT/RIR PPI is a valid approach for developing a new class of targeted anticancer agents, we designed a fluorescence polarization-based assay that was utilized in a pilot screen for small molecule inhibitors of this PPI. Two small molecule scaffolds that disrupt this interaction were identified, and secondary validation assays confirmed that compound 5 binds to Rev1-CT at the RIR interface. Finally, survival and mutagenesis assays in mouse embryonic fibroblasts and human fibrosarcoma HT1080 cells treated with cisplatin and ultraviolet light indicate that these compounds inhibit mutagenic Rev1/Polζ-dependent TLS in cells, validating the Rev1-CT/RIR PPI for future anticancer drug discovery and identifying the first small molecule inhibitors of TLS that target Rev1-CT.

  14. Improved prediction of drug-target interactions using regularized least squares integrating with kernel fusion technique.

    PubMed

    Hao, Ming; Wang, Yanli; Bryant, Stephen H

    2016-02-25

    Identification of drug-target interactions (DTI) is a central task in drug discovery processes. In this work, a simple but effective regularized least squares integrating with nonlinear kernel fusion (RLS-KF) algorithm is proposed to perform DTI predictions. Using benchmark DTI datasets, our proposed algorithm achieves the state-of-the-art results with area under precision-recall curve (AUPR) of 0.915, 0.925, 0.853 and 0.909 for enzymes, ion channels (IC), G protein-coupled receptors (GPCR) and nuclear receptors (NR) based on 10 fold cross-validation. The performance can further be improved by using a recalculated kernel matrix, especially for the small set of nuclear receptors with AUPR of 0.945. Importantly, most of the top ranked interaction predictions can be validated by experimental data reported in the literature, bioassay results in the PubChem BioAssay database, as well as other previous studies. Our analysis suggests that the proposed RLS-KF is helpful for studying DTI, drug repositioning as well as polypharmacology, and may help to accelerate drug discovery by identifying novel drug targets. Published by Elsevier B.V.

  15. Targeting host factors to treat West Nile and dengue viral infections.

    PubMed

    Krishnan, Manoj N; Garcia-Blanco, Mariano A

    2014-02-10

    West Nile (WNV) and Dengue (DENV) viruses are major arboviral human pathogens belonging to the genus Flavivirus. At the current time, there are no approved prophylactics (e.g., vaccines) or specific therapeutics available to prevent or treat human infections by these pathogens. Due to their minimal genome, these viruses require many host molecules for their replication and this offers a therapeutic avenue wherein host factors can be exploited as treatment targets. Since several host factors appear to be shared by many flaviviruses the strategy may result in pan-flaviviral inhibitors and may also attenuate the rapid emergence of drug resistant mutant viruses. The scope of this strategy is greatly enhanced by the recent en masse identification of host factors impacting on WNV and DENV infection. Excellent proof-of-principle experimental demonstrations for host-targeted control of infection and infection-induced pathogenesis have been reported for both WNV and DENV. These include exploiting not only those host factors supporting infection, but also targeting host processes contributing to pathogenesis and innate immune responses. While these early studies validated the host-targeting approach, extensive future investigations spanning a range of aspects are needed for a successful deployment in humans.

  16. Targeting Host Factors to Treat West Nile and Dengue Viral Infections

    PubMed Central

    Krishnan, Manoj N.; Garcia-Blanco, Mariano A.

    2014-01-01

    West Nile (WNV) and Dengue (DENV) viruses are major arboviral human pathogens belonging to the genus Flavivirus. At the current time, there are no approved prophylactics (e.g., vaccines) or specific therapeutics available to prevent or treat human infections by these pathogens. Due to their minimal genome, these viruses require many host molecules for their replication and this offers a therapeutic avenue wherein host factors can be exploited as treatment targets. Since several host factors appear to be shared by many flaviviruses the strategy may result in pan-flaviviral inhibitors and may also attenuate the rapid emergence of drug resistant mutant viruses. The scope of this strategy is greatly enhanced by the recent en masse identification of host factors impacting on WNV and DENV infection. Excellent proof-of-principle experimental demonstrations for host-targeted control of infection and infection-induced pathogenesis have been reported for both WNV and DENV. These include exploiting not only those host factors supporting infection, but also targeting host processes contributing to pathogenesis and innate immune responses. While these early studies validated the host-targeting approach, extensive future investigations spanning a range of aspects are needed for a successful deployment in humans. PMID:24517970

  17. Multi-look fusion identification: a paradigm shift from quality to quantity in data samples

    NASA Astrophysics Data System (ADS)

    Wong, S.

    2009-05-01

    A multi-look identification method known as score-level fusion is found to be capable of achieving very high identification accuracy, even when low quality target signatures are used. Analysis using measured ground vehicle radar signatures has shown that a 97% correct identification rate can be achieved using this multi-look fusion method; in contrast, only a 37% accuracy rate is obtained when single target signature input is used. The results suggest that quantity can be used to replace quality of the target data in improving identification accuracy. With the advent of sensor technology, a large amount of target signatures of marginal quality can be captured routinely. This quantity over quality approach allows maximum exploitation of the available data to improve the target identification performance and this could have the potential of being developed into a disruptive technology.

  18. Targeted proteomics identifies liquid-biopsy signatures for extracapsular prostate cancer

    PubMed Central

    Kim, Yunee; Jeon, Jouhyun; Mejia, Salvador; Yao, Cindy Q; Ignatchenko, Vladimir; Nyalwidhe, Julius O; Gramolini, Anthony O; Lance, Raymond S; Troyer, Dean A; Drake, Richard R; Boutros, Paul C; Semmes, O. John; Kislinger, Thomas

    2016-01-01

    Biomarkers are rapidly gaining importance in personalized medicine. Although numerous molecular signatures have been developed over the past decade, there is a lack of overlap and many biomarkers fail to validate in independent patient cohorts and hence are not useful for clinical application. For these reasons, identification of novel and robust biomarkers remains a formidable challenge. We combine targeted proteomics with computational biology to discover robust proteomic signatures for prostate cancer. Quantitative proteomics conducted in expressed prostatic secretions from men with extraprostatic and organ-confined prostate cancers identified 133 differentially expressed proteins. Using synthetic peptides, we evaluate them by targeted proteomics in a 74-patient cohort of expressed prostatic secretions in urine. We quantify a panel of 34 candidates in an independent 207-patient cohort. We apply machine-learning approaches to develop clinical predictive models for prostate cancer diagnosis and prognosis. Our results demonstrate that computationally guided proteomics can discover highly accurate non-invasive biomarkers. PMID:27350604

  19. Validation of DNA-based identification software by computation of pedigree likelihood ratios.

    PubMed

    Slooten, K

    2011-08-01

    Disaster victim identification (DVI) can be aided by DNA-evidence, by comparing the DNA-profiles of unidentified individuals with those of surviving relatives. The DNA-evidence is used optimally when such a comparison is done by calculating the appropriate likelihood ratios. Though conceptually simple, the calculations can be quite involved, especially with large pedigrees, precise mutation models etc. In this article we describe a series of test cases designed to check if software designed to calculate such likelihood ratios computes them correctly. The cases include both simple and more complicated pedigrees, among which inbred ones. We show how to calculate the likelihood ratio numerically and algebraically, including a general mutation model and possibility of allelic dropout. In Appendix A we show how to derive such algebraic expressions mathematically. We have set up these cases to validate new software, called Bonaparte, which performs pedigree likelihood ratio calculations in a DVI context. Bonaparte has been developed by SNN Nijmegen (The Netherlands) for the Netherlands Forensic Institute (NFI). It is available free of charge for non-commercial purposes (see www.dnadvi.nl for details). Commercial licenses can also be obtained. The software uses Bayesian networks and the junction tree algorithm to perform its calculations. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

  20. Identification accuracy of children versus adults: a meta-analysis.

    PubMed

    Pozzulo, J D; Lindsay, R C

    1998-10-01

    Identification accuracy of children and adults was examined in a meta-analysis. Preschoolers (M = 4 years) were less likely than adults to make correct identifications. Children over the age of 5 did not differ significantly from adults with regard to correct identification rate. Children of all ages examined were less likely than adults to correctly reject a target-absent lineup. Even adolescents (M = 12-13 years) did not reach an adult rate of correct rejection. Compared to simultaneous lineup presentation, sequential lineups increased the child-adult gap for correct rejections. Providing child witnesses with identification practice or training did not increase their correct rejection rates. Suggestions for children's inability to correctly reject target-absent lineups are discussed. Future directions for identification research are presented.

  1. GIS prospectivity mapping and 3D modeling validation for potential uranium deposit targets in Shangnan district, China

    NASA Astrophysics Data System (ADS)

    Xie, Jiayu; Wang, Gongwen; Sha, Yazhou; Liu, Jiajun; Wen, Botao; Nie, Ming; Zhang, Shuai

    2017-04-01

    Integrating multi-source geoscience information (such as geology, geophysics, geochemistry, and remote sensing) using GIS mapping is one of the key topics and frontiers in quantitative geosciences for mineral exploration. GIS prospective mapping and three-dimensional (3D) modeling can be used not only to extract exploration criteria and delineate metallogenetic targets but also to provide important information for the quantitative assessment of mineral resources. This paper uses the Shangnan district of Shaanxi province (China) as a case study area. GIS mapping and potential granite-hydrothermal uranium targeting were conducted in the study area combining weights of evidence (WofE) and concentration-area (C-A) fractal methods with multi-source geoscience information. 3D deposit-scale modeling using GOCAD software was performed to validate the shapes and features of the potential targets at the subsurface. The research results show that: (1) the known deposits have potential zones at depth, and the 3D geological models can delineate surface or subsurface ore-forming features, which can be used to analyze the uncertainty of the shape and feature of prospectivity mapping at the subsurface; (2) single geochemistry anomalies or remote sensing anomalies at the surface require combining the depth exploration criteria of geophysics to identify potential targets; and (3) the single or sparse exploration criteria zone with few mineralization spots at the surface has high uncertainty in terms of the exploration target.

  2. The identification of cellular targets of 17β estradiol using a lytic (T7) cDNA phage display approach.

    PubMed

    Van Dorst, Bieke; Mehta, Jaytry; Rouah-Martin, Elsa; De Coen, Wim; Blust, Ronny; Robbens, Johan

    2011-02-01

    To unravel the mechanism of action of chemical compounds, it is crucial to know their cellular targets. A novel in vitro tool that can be used as a fast, simple and cost effective alternative is cDNA phage display. This tool is used in our study to select cellular targets of 17β estradiol (E2). It was possible to select two potential cellular targets of E2 out of the T7 Select™ Human Breast cDNA phage library. The selected cellular targets, autophagy/beclin-1 regulator 1 (beclin 1) and ATP synthase F(0) subunit 6 (ATP6) have so far been unknown as binding proteins of E2. To confirm the E2 binding properties of these selected proteins, surface plasmon resonance (SPR) was used. With SPR the K(d) values were determined to be 0.178±0.031 and 0.401±0.142 nM for the ATP6 phage and beclin 1 phage, respectively. These K(d) values in the low nM range verify that the selected cellular proteins are indeed binding proteins for E2. The selection and identification of these two potential cellular targets of E2, can enhance our current understanding of its mechanism of action. This illustrates the potential of lytic (T7) cDNA phage display in toxicology, to provide important information about cellular targets of chemical compounds. Copyright © 2010 Elsevier Ltd. All rights reserved.

  3. Systematic Identification of MCU Modulators by Orthogonal Interspecies Chemical Screening.

    PubMed

    Arduino, Daniela M; Wettmarshausen, Jennifer; Vais, Horia; Navas-Navarro, Paloma; Cheng, Yiming; Leimpek, Anja; Ma, Zhongming; Delrio-Lorenzo, Alba; Giordano, Andrea; Garcia-Perez, Cecilia; Médard, Guillaume; Kuster, Bernhard; García-Sancho, Javier; Mokranjac, Dejana; Foskett, J Kevin; Alonso, M Teresa; Perocchi, Fabiana

    2017-08-17

    The mitochondrial calcium uniporter complex is essential for calcium (Ca 2+ ) uptake into mitochondria of all mammalian tissues, where it regulates bioenergetics, cell death, and Ca 2+ signal transduction. Despite its involvement in several human diseases, we currently lack pharmacological agents for targeting uniporter activity. Here we introduce a high-throughput assay that selects for human MCU-specific small-molecule modulators in primary drug screens. Using isolated yeast mitochondria, reconstituted with human MCU, its essential regulator EMRE, and aequorin, and exploiting a D-lactate- and mannitol/sucrose-based bioenergetic shunt that greatly minimizes false-positive hits, we identify mitoxantrone out of more than 600 clinically approved drugs as a direct selective inhibitor of human MCU. We validate mitoxantrone in orthogonal mammalian cell-based assays, demonstrating that our screening approach is an effective and robust tool for MCU-specific drug discovery and, more generally, for the identification of compounds that target mitochondrial functions. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. CCTop: An Intuitive, Flexible and Reliable CRISPR/Cas9 Target Prediction Tool

    PubMed Central

    del Sol Keyer, Maria; Wittbrodt, Joachim; Mateo, Juan L.

    2015-01-01

    Engineering of the CRISPR/Cas9 system has opened a plethora of new opportunities for site-directed mutagenesis and targeted genome modification. Fundamental to this is a stretch of twenty nucleotides at the 5’ end of a guide RNA that provides specificity to the bound Cas9 endonuclease. Since a sequence of twenty nucleotides can occur multiple times in a given genome and some mismatches seem to be accepted by the CRISPR/Cas9 complex, an efficient and reliable in silico selection and evaluation of the targeting site is key prerequisite for the experimental success. Here we present the CRISPR/Cas9 target online predictor (CCTop, http://crispr.cos.uni-heidelberg.de) to overcome limitations of already available tools. CCTop provides an intuitive user interface with reasonable default parameters that can easily be tuned by the user. From a given query sequence, CCTop identifies and ranks all candidate sgRNA target sites according to their off-target quality and displays full documentation. CCTop was experimentally validated for gene inactivation, non-homologous end-joining as well as homology directed repair. Thus, CCTop provides the bench biologist with a tool for the rapid and efficient identification of high quality target sites. PMID:25909470

  5. Electro-Optic Identification (EOID) Research Program

    DTIC Science & Technology

    2002-09-30

    The goal of this research is to provide computer-assisted identification of underwater mines in electro - optic imagery. Identification algorithms will...greatly reduce the time and risk to reacquire mine-like-objects for positive classification and identification. The objectives are to collect electro ... optic data under a wide range of operating and environmental conditions and develop precise algorithms that can provide accurate target recognition on this data for all possible conditions.

  6. Short communication: Validation of 4 candidate causative trait variants in 2 cattle breeds using targeted sequence imputation.

    PubMed

    Pausch, Hubert; Wurmser, Christine; Reinhardt, Friedrich; Emmerling, Reiner; Fries, Ruedi

    2015-06-01

    Most association studies for pinpointing trait-associated variants are performed within breed. The availability of sequence data from key ancestors of several cattle breeds now enables immediate assessment of the frequency of trait-associated variants in populations different from the mapping population and their imputation into large validation populations. The objective of this study was to validate the effects of 4 putatively causative variants on milk production traits, male fertility, and stature in German Fleckvieh and Holstein-Friesian animals using targeted sequence imputation. We used whole-genome sequence data of 456 animals to impute 4 missense mutations in DGAT1, GHR, PRLR, and PROP1 into 10,363 Fleckvieh and 8,812 Holstein animals. The accuracy of the imputed genotypes exceeded 95% for all variants. Association testing with imputed variants revealed consistent antagonistic effects of the DGAT1 p.A232K and GHR p.F279Y variants on milk yield and protein and fat contents, respectively, in both breeds. The allele frequency of both polymorphisms has changed considerably in the past 20 yr, indicating that they were targets of recent selection for milk production traits. The PRLR p.S18N variant was associated with yield traits in Fleckvieh but not in Holstein, suggesting that it may be in linkage disequilibrium with a mutation affecting yield traits rather than being causal. The reported effects of the PROP1 p.H173R variant on milk production, male fertility, and stature could not be confirmed. Our results demonstrate that population-wide imputation of candidate causal variants from sequence data is feasible, enabling their rapid validation in large independent populations. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  7. Identification and Validation of Novel Small Molecule Disruptors of HuR-mRNA Interaction

    PubMed Central

    Wu, Xiaoqing; Lan, Lan; Wilson, David Michael; Marquez, Rebecca T.; Tsao, Wei-chung; Gao, Philip; Roy, Anuradha; Turner, Benjamin Andrew; McDonald, Peter; Tunge, Jon A; Rogers, Steven A; Dixon, Dan A.; Aubé, Jeffrey; Xu, Liang

    2015-01-01

    HuR, an RNA binding protein, binds to adenine- and uridine-rich elements (ARE) in the 3′-untranslated region (UTR) of target mRNAs, regulating their stability and translation. HuR is highly abundant in many types of cancer, and it promotes tumorigenesis by interacting with cancer-associated mRNAs, which encode proteins that are implicated in different tumor processes including cell proliferation, cell survival, angiogenesis, invasion, and metastasis. Drugs that disrupt the stabilizing effect of HuR upon mRNA targets could have dramatic effects on inhibiting cancer growth and persistence. In order to identify small molecules that directly disrupt the HuR–ARE interaction, we established a fluorescence polarization (FP) assay optimized for high throughput screening (HTS) using HuR protein and an ARE oligo from Musashi RNA-binding protein 1 (Msi1) mRNA, a HuR target. Following the performance of an HTS of ~6000 compounds, we discovered a cluster of potential disruptors, which were then validated by AlphaLISA (Amplified Luminescent Proximity Homogeneous Assay), surface plasmon resonance (SPR), ribonucleoprotein immunoprecipitation (RNP IP) assay, and luciferase reporter functional studies. These compounds disrupted HuR–ARE interactions at the nanomolar level and blocked HuR function by competitive binding to HuR. These results support future studies toward chemical probes for a HuR function study and possibly a novel therapy for HuR-overexpressing cancers. PMID:25750985

  8. Dana-Farber Cancer Institute: Identification of Therapeutic Targets in KRAS Driven Lung Cancer | Office of Cancer Genomics

    Cancer.gov

    The CTD2 Center at Dana Farber Cancer Institute focuses on the use of high-throughput genetic and bioinformatic approaches to identify and credential oncogenes and co-dependencies in cancers. This Center aims to provide the cancer research community with information that will facilitate the prioritization of targets based on both genomic and functional evidence, inform the most appropriate genetic context for downstream mechanistic and validation studies, and enable the translation of this information into therapeutics and diagnostics.

  9. Effects of Resolution, Range, and Image Contrast on Target Acquisition Performance.

    PubMed

    Hollands, Justin G; Terhaar, Phil; Pavlovic, Nada J

    2018-05-01

    We sought to determine the joint influence of resolution, target range, and image contrast on the detection and identification of targets in simulated naturalistic scenes. Resolution requirements for target acquisition have been developed based on threshold values obtained using imaging systems, when target range was fixed, and image characteristics were determined by the system. Subsequent work has examined the influence of factors like target range and image contrast on target acquisition. We varied the resolution and contrast of static images in two experiments. Participants (soldiers) decided whether a human target was located in the scene (detection task) or whether a target was friendly or hostile (identification task). Target range was also varied (50-400 m). In Experiment 1, 30 participants saw color images with a single target exemplar. In Experiment 2, another 30 participants saw monochrome images containing different target exemplars. The effects of target range and image contrast were qualitatively different above and below 6 pixels per meter of target for both tasks in both experiments. Target detection and identification performance were a joint function of image resolution, range, and contrast for both color and monochrome images. The beneficial effects of increasing resolution for target acquisition performance are greater for closer (larger) targets.

  10. One-week 96-well soft agar growth assay for cancer target validation.

    PubMed

    Ke, Ning; Albers, Aaron; Claassen, Gisela; Yu, De-hua; Chatterton, Jon E; Hu, Xiuyuan; Meyhack, Bernd; Wong-Staal, Flossie; Li, Qi-Xiang

    2004-05-01

    Soft agar growth, used to measure cell anchorage-independent proliferation potential, is one of the most important and most commonly used assays to detect cell transformation. However, the traditional soft agar assay is time-consuming, labor-intensive, and plagued with inconsistencies due to individual subjectivity. It does not, therefore, meet the increasing demands of today's oncology drug target screening or validation processes. This report describes an alternative 96-well soft agar growth assay that can function as a replacement for the traditional method and overcomes the aforementioned limitations. It offers the following advantages: a shortened assay duration (1 week instead of 4 weeks) that makes transient transfection or treatment possible; plate reader quantification of soft agar growth (measuring cloning efficiency and colony size); and a significant reduction in required labor. Higher throughput also makes it possible to process large numbers of samples and treatments simultaneously and in a much more efficient manner, while saving precious workspace and overall cost.

  11. Detection and identification of 700 drugs by multi-target screening with a 3200 Q TRAP LC-MS/MS system and library searching.

    PubMed

    Dresen, S; Ferreirós, N; Gnann, H; Zimmermann, R; Weinmann, W

    2010-04-01

    The multi-target screening method described in this work allows the simultaneous detection and identification of 700 drugs and metabolites in biological fluids using a hybrid triple-quadrupole linear ion trap mass spectrometer in a single analytical run. After standardization of the method, the retention times of 700 compounds were determined and transitions for each compound were selected by a "scheduled" survey MRM scan, followed by an information-dependent acquisition using the sensitive enhanced product ion scan of a Q TRAP hybrid instrument. The identification of the compounds in the samples analyzed was accomplished by searching the tandem mass spectrometry (MS/MS) spectra against the library we developed, which contains electrospray ionization-MS/MS spectra of over 1,250 compounds. The multi-target screening method together with the library was included in a software program for routine screening and quantitation to achieve automated acquisition and library searching. With the help of this software application, the time for evaluation and interpretation of the results could be drastically reduced. This new multi-target screening method has been successfully applied for the analysis of postmortem and traffic offense samples as well as proficiency testing, and complements screening with immunoassays, gas chromatography-mass spectrometry, and liquid chromatography-diode-array detection. Other possible applications are analysis in clinical toxicology (for intoxication cases), in psychiatry (antidepressants and other psychoactive drugs), and in forensic toxicology (drugs and driving, workplace drug testing, oral fluid analysis, drug-facilitated sexual assault).

  12. Identification of target genes of synovial sarcoma-associated fusion oncoprotein using human pluripotent stem cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hayakawa, Kazuo; Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto; Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya

    2013-03-22

    Highlights: ► We tried to identify targets of synovial sarcoma (SS)-associated SYT–SSX fusion gene. ► We established pluripotent stem cell (PSC) lines with inducible SYT–SSX gene. ► SYT–SSX responsive genes were identified by the induction of SYT–SSX in PSC. ► SS-related genes were selected from database by in silico analyses. ► 51 genes were finally identified among SS-related genes as targets of SYT–SSX in PSC. -- Abstract: Synovial sarcoma (SS) is a malignant soft tissue tumor harboring chromosomal translocation t(X; 18)(p11.2; q11.2), which produces SS-specific fusion gene, SYT–SSX. Although precise function of SYT–SSX remains to be investigated, accumulating evidences suggestmore » its role in gene regulation via epigenetic mechanisms, and the product of SYT–SSX target genes may serve as biomarkers of SS. Lack of knowledge about the cell-of-origin of SS, however, has placed obstacle in the way of target identification. Here we report a novel approach to identify SYT–SSX2 target genes using human pluripotent stem cells (hPSCs) containing a doxycycline-inducible SYT–SSX2 gene. SYT–SSX2 was efficiently induced both at mRNA and protein levels within three hours after doxycycline administration, while no morphological change of hPSCs was observed until 24 h. Serial microarray analyses identified genes of which the expression level changed more than twofold within 24 h. Surprisingly, the majority (297/312, 95.2%) were up-regulated genes and a result inconsistent with the current concept of SYT–SSX as a transcriptional repressor. Comparing these genes with SS-related genes which were selected by a series of in silico analyses, 49 and 2 genes were finally identified as candidates of up- and down-regulated target of SYT–SSX, respectively. Association of these genes with SYT–SSX in SS cells was confirmed by knockdown experiments. Expression profiles of SS-related genes in hPSCs and human mesenchymal stem cells (hMSCs) were

  13. Blind sequential lineup administration reduces both false identifications and confidence in those false identifications.

    PubMed

    Charman, Steve D; Quiroz, Vanessa

    2016-10-01

    One of the most recommended procedures proposed by eyewitness experts is the use of double-blind lineups, in which the administrator does not know the identity of the suspect in the lineup. But despite the near universality of this recommendation, there is surprisingly little empirical research to support the claim that nonblind administration inflates false identifications. What little research has been conducted has shown conflicting findings with regard to the conditions under which nonblind administration affects false identifications, as well as its effects on witness confidence. The current study attempts to elucidate this effect. Student-participants (n = 312) were randomly assigned to play the role of either a lineup administrator (who were either told the identity of the suspect in the lineup or not) or a mock crime witness. Following unbiased instructions, administrators presented either a target-present or target-absent sequential lineup to the witness while being surreptitiously videorecorded. Nonblind administration significantly inflated false, but not correct, identifications, and significantly inflated witness confidence in those false identifications. Video recordings indicated that nonblind administrators were significantly more likely than blind administrators to smile (a) while the witness was viewing a photograph of the suspect, and (b) after a suspect identification. Results provide stronger support for the use of blind lineup administration by broadening the conditions under which nonblind administration is shown to inflate false identifications. Possible reconciliations for conflicting findings in the literature are discussed. (PsycINFO Database Record (c) 2016 APA, all rights reserved).

  14. Microfluidic droplet enrichment for targeted sequencing

    PubMed Central

    Eastburn, Dennis J.; Huang, Yong; Pellegrino, Maurizio; Sciambi, Adam; Ptáček, Louis J.; Abate, Adam R.

    2015-01-01

    Targeted sequence enrichment enables better identification of genetic variation by providing increased sequencing coverage for genomic regions of interest. Here, we report the development of a new target enrichment technology that is highly differentiated from other approaches currently in use. Our method, MESA (Microfluidic droplet Enrichment for Sequence Analysis), isolates genomic DNA fragments in microfluidic droplets and performs TaqMan PCR reactions to identify droplets containing a desired target sequence. The TaqMan positive droplets are subsequently recovered via dielectrophoretic sorting, and the TaqMan amplicons are removed enzymatically prior to sequencing. We demonstrated the utility of this approach by generating an average 31.6-fold sequence enrichment across 250 kb of targeted genomic DNA from five unique genomic loci. Significantly, this enrichment enabled a more comprehensive identification of genetic polymorphisms within the targeted loci. MESA requires low amounts of input DNA, minimal prior locus sequence information and enriches the target region without PCR bias or artifacts. These features make it well suited for the study of genetic variation in a number of research and diagnostic applications. PMID:25873629

  15. Development and Validation of a Decision Tool for Early Identification of Adult Patients with Severe and Complex Eating Disorder Psychopathology in Need of Highly Specialized Care.

    PubMed

    Dingemans, Alexandra E; Goorden, Maartje; Lötters, Freek J B; Bouwmans, Clazien; Danner, Unna N; van Elburg, Annemarie A; van Furth, Eric F; Hakkaart-van Roijen, Leona

    2017-09-01

    Patients with complex and severe eating disorders often receive a number of ineffective or/and insufficient treatments. Direct referral of these patients to highly specialized tertiary treatment facilities in an earlier stage of the disorder is likely to be more (cost)-effective. The aim of the study was to develop a decision tool that aids clinicians in early identification of these patients. After identification of criteria that were indicative of severity and complexity of eating disorder psychopathology by means of a systematic review of literature and consultation of a focus group, a Delphi method was applied to obtain consensus from experts on the list of relevant criteria. Finally, the decision tool was validated in clinical practice, and cut-off criteria were established. The tool demonstrated good feasibility and validity to identify patients for highly specialized tertiary care. The final decision tool consisted of five criteria that can easily be implemented in clinical practice. Copyright © 2017 John Wiley & Sons, Ltd and Eating Disorders Association. Copyright © 2017 John Wiley & Sons, Ltd and Eating Disorders Association.

  16. Validation of a personalized dosimetric evaluation tool (Oedipe) for targeted radiotherapy based on the Monte Carlo MCNPX code

    NASA Astrophysics Data System (ADS)

    Chiavassa, S.; Aubineau-Lanièce, I.; Bitar, A.; Lisbona, A.; Barbet, J.; Franck, D.; Jourdain, J. R.; Bardiès, M.

    2006-02-01

    Dosimetric studies are necessary for all patients treated with targeted radiotherapy. In order to attain the precision required, we have developed Oedipe, a dosimetric tool based on the MCNPX Monte Carlo code. The anatomy of each patient is considered in the form of a voxel-based geometry created using computed tomography (CT) images or magnetic resonance imaging (MRI). Oedipe enables dosimetry studies to be carried out at the voxel scale. Validation of the results obtained by comparison with existing methods is complex because there are multiple sources of variation: calculation methods (different Monte Carlo codes, point kernel), patient representations (model or specific) and geometry definitions (mathematical or voxel-based). In this paper, we validate Oedipe by taking each of these parameters into account independently. Monte Carlo methodology requires long calculation times, particularly in the case of voxel-based geometries, and this is one of the limits of personalized dosimetric methods. However, our results show that the use of voxel-based geometry as opposed to a mathematically defined geometry decreases the calculation time two-fold, due to an optimization of the MCNPX2.5e code. It is therefore possible to envisage the use of Oedipe for personalized dosimetry in the clinical context of targeted radiotherapy.

  17. Ships and Maritime Targets Observation Campaigns Using Available C- and X-Band SAR Satellite

    NASA Astrophysics Data System (ADS)

    Velotto, Domenico; Bentes, Carlos; Lehner, Susanne

    2015-04-01

    Obviously, radar resolution and swath width are two very important factors when it comes to synthetic aperture radar (SAR) maritime targets detections. The dilemma of using single polarization SAR imagery with higher resolution and coverage or quad- (or dual- polarimetric) imagery with its richness of information, is still unsolved when it comes to this application.In the framework of ESA project MARISS and EU project DOLPHIN, in situ campaigns aimed at solving this dilemma have been carried out. Single and multi- polarimetric SAR data acquired by TerraSAR-X, RADARSAT-2 and COSMO-SkyMed have been acquired with close time gaps and partial coverage overlap. In this way several moving and non-moving maritime targets have been imaged with different polarization, geometry and working frequency. Available ground truth reports provided by Automatic Identification System (AIS) data, nautical chart and wind farm location are used to validate the different types of maritime targets.

  18. Ada Compiler Validation Summary Report: Harris Corporation, Harris Ada Compiler, Version 4.0, Harris HCX-9 (Host) and (Target), 880603W1.09059

    DTIC Science & Technology

    1988-06-06

    TYPE Of REPORT & PERIOD COVERED Ada Compiler Validation Summary Report : Harris 6 June 1988 to 6 June 1988 Corporation, Harris Ada Compiler, Version...4.0, Harris 1 PERFORINGDRG REPORT NUMBER HCX-9 (Host) and (Target), 880603W1.09059 7. AUTHOR(s) S. CONTRACT OR 6RANT NUMBER(s) Wright-Patterson AFB...88-03-02-HAR Ada COMPILER VALIDATION SUMMARY REPORT : Certificate Number: 880603WI.09059 A Harris Corporation AccessionFor Harris Ada Compiler, Version

  19. [Thalidomide teratogenicity and its direct target identification].

    PubMed

    Ito, Takumi; Ando, Hideki; Handa, Hiroshi

    2015-01-01

    Half a century ago, thalidomide was developed as a sedative drug and was wildly used over 40 countries. However the drug has serious birth defects such as amelia and phocomelia. Now thalidomide is regarded as a clinically effective drug and used for the treatment of multiple myeloma under strict controls. The direct target of thalidomide had been a long-standing question. We identified cereblon as a primary direct target protein for thalidomide teratogenicity using new affinity bead technology in 2010. In this review, we introduce an overview of thalidomide teratogenicity, a story about how we identified cereblon, and recent advances in cereblon studies.

  20. Targeting protein translation, RNA splicing, and degradation by morpholino-based conjugates in Plasmodium falciparum.

    PubMed

    Garg, Aprajita; Wesolowski, Donna; Alonso, Dulce; Deitsch, Kirk W; Ben Mamoun, Choukri; Altman, Sidney

    2015-09-22

    Identification and genetic validation of new targets from available genome sequences are critical steps toward the development of new potent and selective antimalarials. However, no methods are currently available for large-scale functional analysis of the Plasmodium falciparum genome. Here we present evidence for successful use of morpholino oligomers (MO) to mediate degradation of target mRNAs or to inhibit RNA splicing or translation of several genes of P. falciparum involved in chloroquine transport, apicoplast biogenesis, and phospholipid biosynthesis. Consistent with their role in the parasite life cycle, down-regulation of these essential genes resulted in inhibition of parasite development. We show that a MO conjugate that targets the chloroquine-resistant transporter PfCRT is effective against chloroquine-sensitive and -resistant parasites, causes enlarged digestive vacuoles, and renders chloroquine-resistant strains more sensitive to chloroquine. Similarly, we show that a MO conjugate that targets the PfDXR involved in apicoplast biogenesis inhibits parasite growth and that this defect can be rescued by addition of isopentenyl pyrophosphate. MO-based gene regulation is a viable alternative approach to functional analysis of the P. falciparum genome.

  1. Accurate identification of motor unit discharge patterns from high-density surface EMG and validation with a novel signal-based performance metric

    NASA Astrophysics Data System (ADS)

    Holobar, A.; Minetto, M. A.; Farina, D.

    2014-02-01

    Objective. A signal-based metric for assessment of accuracy of motor unit (MU) identification from high-density surface electromyograms (EMG) is introduced. This metric, so-called pulse-to-noise-ratio (PNR), is computationally efficient, does not require any additional experimental costs and can be applied to every MU that is identified by the previously developed convolution kernel compensation technique. Approach. The analytical derivation of the newly introduced metric is provided, along with its extensive experimental validation on both synthetic and experimental surface EMG signals with signal-to-noise ratios ranging from 0 to 20 dB and muscle contraction forces from 5% to 70% of the maximum voluntary contraction. Main results. In all the experimental and simulated signals, the newly introduced metric correlated significantly with both sensitivity and false alarm rate in identification of MU discharges. Practically all the MUs with PNR > 30 dB exhibited sensitivity >90% and false alarm rates <2%. Therefore, a threshold of 30 dB in PNR can be used as a simple method for selecting only reliably decomposed units. Significance. The newly introduced metric is considered a robust and reliable indicator of accuracy of MU identification. The study also shows that high-density surface EMG can be reliably decomposed at contraction forces as high as 70% of the maximum.

  2. Neuromorphic Optical Signal Processing and Image Understanding for Automated Target Recognition

    DTIC Science & Technology

    1989-12-01

    34 Stochastic Learning Machine " Neuromorphic Target Identification * Cognitive Networks 3. Conclusions ..... ................ .. 12 4. Publications...16 5. References ...... ................... . 17 6. Appendices ....... .................. 18 I. Optoelectronic Neural Networks and...Learning Machines. II. Stochastic Optical Learning Machine. III. Learning Network for Extrapolation AccesFon For and Radar Target Identification

  3. Functional validation of the oncogenic cooperativity and targeting potential of tuberous sclerosis mutation in medulloblastoma using a MYC-amplified model cell line.

    PubMed

    Henderson, Jacob J; Wagner, Jacob P; Hofmann, Nicolle E; Eide, Christopher A; Cho, Yoon-Jae; Druker, Brian J; Davare, Monika A

    2017-10-01

    Medulloblastoma is the most common malignant brain tumor of childhood. To identify targetable vulnerabilities, we employed inhibitor screening that revealed mTOR inhibitor hypersensitivity in the MYC-overexpressing medulloblastoma cell line, D341. Concomitant exome sequencing unveiled an uncharacterized missense mutation, TSC2 A415V , in these cells. We biochemically demonstrate that the TSC2 A415V mutation is functionally deleterious, leading to shortened half-life and proteasome-mediated protein degradation. These data suggest that MYC cooperates with activated kinase pathways, enabling pharmacologic intervention in these treatment refractory tumors. We propose that identification of activated kinase pathways may allow for tailoring targeted therapy to improve survival and treatment-related morbidity in medulloblastoma. © 2017 Wiley Periodicals, Inc.

  4. Examination of soldier target recognition with direct view optics

    NASA Astrophysics Data System (ADS)

    Long, Frederick H.; Larkin, Gabriella; Bisordi, Danielle; Dorsey, Shauna; Marianucci, Damien; Goss, Lashawnta; Bastawros, Michael; Misiuda, Paul; Rodgers, Glenn; Mazz, John P.

    2017-10-01

    Target recognition and identification is a problem of great military and scientific importance. To examine the correlation between target recognition and optical magnification, ten U.S. Army soldiers were tasked with identifying letters on targets at 800 and 1300 meters away. Letters were used since they are a standard method for measuring visual acuity. The letters were approximately 90 cm high, which is the size of a well-known rifle. Four direct view optics with angular magnifications of 1.5x, 4x, 6x, and 9x were used. The goal of this approach was to measure actual probabilities for correct target identification. Previous scientific literature suggests that target recognition can be modeled as a linear response problem in angular frequency space using the established values for the contrast sensitivity function for a healthy human eye and the experimentally measured modulation transfer function of the optic. At the 9x magnification, the soldiers could identify the letters with almost no errors (i.e., 97% probability of correct identification). At lower magnification, errors in letter identification were more frequent. The identification errors were not random but occurred most frequently with a few pairs of letters (e.g., O and Q), which is consistent with the literature for letter recognition. In addition, in the small subject sample of ten soldiers, there was considerable variation in the observer recognition capability at 1.5x and a range of 800 meters. This can be directly attributed to the variation in the observer visual acuity.

  5. Design and Validation of CRISPR/Cas9 Systems for Targeted Gene Modification in Induced Pluripotent Stem Cells.

    PubMed

    Lee, Ciaran M; Zhu, Haibao; Davis, Timothy H; Deshmukh, Harshahardhan; Bao, Gang

    2017-01-01

    The CRISPR/Cas9 system is a powerful tool for precision genome editing. The ability to accurately modify genomic DNA in situ with single nucleotide precision opens up new possibilities for not only basic research but also biotechnology applications and clinical translation. In this chapter, we outline the procedures for design, screening, and validation of CRISPR/Cas9 systems for targeted modification of coding sequences in the human genome and how to perform genome editing in induced pluripotent stem cells with high efficiency and specificity.

  6. Near-Field Chipless Radio-Frequency Identification (RFID) Sensing and Identification System with Switching Reading.

    PubMed

    Paredes, Ferran; Herrojo, Cristian; Mata-Contreras, Javier; Moras, Miquel; Núñez, Alba; Ramon, Eloi; Martín, Ferran

    2018-04-09

    A chipless radio-frequency identification (chipless-RFID) and sensing system, where tags are read by proximity (near-field) through a switch, is presented. The tags consist of a set of identical resonant elements (split-ring resonators or SRRs), printed or etched at predefined and equidistant positions, forming a linear chain, each SRR providing a bit of information. The logic state ('1' or '0') associated with each resonator depends on whether it is present or not in the predefined position. The reader is an array of power splitters used to feed a set of SRR-loaded transmission lines (in equal number to the number of resonant elements, or bits, of the tag). The feeding (interrogation) signal is a harmonic (single-tone) signal tuned to a frequency in the vicinity of the fundamental resonance of the SRRs. The set of SRR-loaded lines must be designed so that the corresponding SRRs are in perfect alignment with the SRRs of the tag, provided the tag is positioned on top of the reader. Thus, in a reading operation, as long as the tag is very close to the reader, the SRRs of the tag modify (decrease) the transmission coefficient of the corresponding reader line (through electromagnetic coupling between both SRRs), and the amplitude of the output signal is severely reduced. Therefore, the identification (ID) code of the tag is contained in the amplitudes of the output signals of the SRR-loaded lines, which can be inferred sequentially by means of a switching system. Unlike previous chipless-RFID systems based on near-field and sequential bit reading, the tags in the proposed system can be merely positioned on top of the reader, conveniently aligned, without the need to mechanically place them across the reader. Since tag reading is only possible if the tag is very close to the reader, this system can be also used as a proximity sensor with applications such as target identification. The proposed chipless-RFID and sensing approach is validated by reading a designed 4-bit

  7. Near-Field Chipless Radio-Frequency Identification (RFID) Sensing and Identification System with Switching Reading

    PubMed Central

    Mata-Contreras, Javier; Moras, Miquel; Ramon, Eloi; Martín, Ferran

    2018-01-01

    A chipless radio-frequency identification (chipless-RFID) and sensing system, where tags are read by proximity (near-field) through a switch, is presented. The tags consist of a set of identical resonant elements (split-ring resonators or SRRs), printed or etched at predefined and equidistant positions, forming a linear chain, each SRR providing a bit of information. The logic state (‘1’ or ‘0’) associated with each resonator depends on whether it is present or not in the predefined position. The reader is an array of power splitters used to feed a set of SRR-loaded transmission lines (in equal number to the number of resonant elements, or bits, of the tag). The feeding (interrogation) signal is a harmonic (single-tone) signal tuned to a frequency in the vicinity of the fundamental resonance of the SRRs. The set of SRR-loaded lines must be designed so that the corresponding SRRs are in perfect alignment with the SRRs of the tag, provided the tag is positioned on top of the reader. Thus, in a reading operation, as long as the tag is very close to the reader, the SRRs of the tag modify (decrease) the transmission coefficient of the corresponding reader line (through electromagnetic coupling between both SRRs), and the amplitude of the output signal is severely reduced. Therefore, the identification (ID) code of the tag is contained in the amplitudes of the output signals of the SRR-loaded lines, which can be inferred sequentially by means of a switching system. Unlike previous chipless-RFID systems based on near-field and sequential bit reading, the tags in the proposed system can be merely positioned on top of the reader, conveniently aligned, without the need to mechanically place them across the reader. Since tag reading is only possible if the tag is very close to the reader, this system can be also used as a proximity sensor with applications such as target identification. The proposed chipless-RFID and sensing approach is validated by reading a designed

  8. Improved Targeting Through Collaborative Decision-Making and Brain Computer Interfaces

    NASA Technical Reports Server (NTRS)

    Stoica, Adrian; Barrero, David F.; McDonald-Maier, Klaus

    2013-01-01

    This paper reports a first step toward a brain-computer interface (BCI) for collaborative targeting. Specifically, we explore, from a broad perspective, how the collaboration of a group of people can increase the performance on a simple target identification task. To this end, we requested a group of people to identify the location and color of a sequence of targets appearing on the screen and measured the time and accuracy of the response. The individual results are compared to a collective identification result determined by simple majority voting, with random choice in case of drawn. The results are promising, as the identification becomes significantly more reliable even with this simple voting and a small number of people (either odd or even number) involved in the decision. In addition, the paper briefly analyzes the role of brain-computer interfaces in collaborative targeting, extending the targeting task by using a BCI instead of a mechanical response.

  9. Modal identification experiment

    NASA Technical Reports Server (NTRS)

    Kvaternik, Raymond G.

    1992-01-01

    The Modal Identification Experiment (MIE) is a proposed on-orbit experiment being developed by NASA's Office of Aeronautics and Space Technology wherein a series of vibration measurements would be made on various configurations of Space Station Freedom (SSF) during its on-orbit assembly phase. The experiment is to be conducted in conjunction with station reboost operations and consists of measuring the dynamic responses of the spacecraft produced by station-based attitude control system and reboost thrusters, recording and transmitting the data, and processing the data on the ground to identify the natural frequencies, damping factors, and shapes of significant vibratory modes. The experiment would likely be a part of the Space Station on-orbit verification. Basic research objectives of MIE are to evaluate and improve methods for analytically modeling large space structures, to develop techniques for performing in-space modal testing, and to validate candidate techniques for in-space modal identification. From an engineering point of view, MIE will provide the first opportunity to obtain vibration data for the fully-assembled structure because SSF is too large and too flexible to be tested as a single unit on the ground. Such full-system data is essential for validating the analytical model of SSF which would be used in any engineering efforts associated with structural or control system changes that might be made to the station as missions evolve over time. Extensive analytical simulations of on-orbit tests, as well exploratory laboratory simulations using small-scale models, have been conducted in-house and under contract to develop a measurement plan and evaluate its potential performance. In particular, performance trade and parametric studies conducted as part of these simulations were used to resolve issues related to the number and location of the measurements, the type of excitation, data acquisition and data processing, effects of noise and nonlinearities

  10. Simulation of target interpretation based on infrared image features and psychology principle

    NASA Astrophysics Data System (ADS)

    Lin, Wei; Chen, Yu-hua; Gao, Hong-sheng; Wang, Zhan-feng; Wang, Ji-jun; Su, Rong-hua; Huang, Yan-ping

    2009-07-01

    It's an important and complicated process in target interpretation that target features extraction and identification, which effect psychosensorial quantity of interpretation person to target infrared image directly, and decide target viability finally. Using statistical decision theory and psychology principle, designing four psychophysical experiment, the interpretation model of the infrared target is established. The model can get target detection probability by calculating four features similarity degree between target region and background region, which were plotted out on the infrared image. With the verification of a great deal target interpretation in practice, the model can simulate target interpretation and detection process effectively, get the result of target interpretation impersonality, which can provide technique support for target extraction, identification and decision-making.

  11. From benchmarking HITS-CLIP peak detection programs to a new method for identification of miRNA-binding sites from Ago2-CLIP data

    PubMed Central

    Bottini, Silvia; Hamouda-Tekaya, Nedra; Tanasa, Bogdan; Zaragosi, Laure-Emmanuelle; Grandjean, Valerie; Repetto, Emanuela

    2017-01-01

    Abstract Experimental evidence indicates that about 60% of miRNA-binding activity does not follow the canonical rule about the seed matching between miRNA and target mRNAs, but rather a non-canonical miRNA targeting activity outside the seed or with a seed-like motifs. Here, we propose a new unbiased method to identify canonical and non-canonical miRNA-binding sites from peaks identified by Ago2 Cross-Linked ImmunoPrecipitation associated to high-throughput sequencing (CLIP-seq). Since the quality of peaks is of pivotal importance for the final output of the proposed method, we provide a comprehensive benchmarking of four peak detection programs, namely CIMS, PIPE-CLIP, Piranha and Pyicoclip, on four publicly available Ago2-HITS-CLIP datasets and one unpublished in-house Ago2-dataset in stem cells. We measured the sensitivity, the specificity and the position accuracy toward miRNA binding sites identification, and the agreement with TargetScan. Secondly, we developed a new pipeline, called miRBShunter, to identify canonical and non-canonical miRNA-binding sites based on de novo motif identification from Ago2 peaks and prediction of miRNA::RNA heteroduplexes. miRBShunter was tested and experimentally validated on the in-house Ago2-dataset and on an Ago2-PAR-CLIP dataset in human stem cells. Overall, we provide guidelines to choose a suitable peak detection program and a new method for miRNA-target identification. PMID:28108660

  12. Targeting Conserved Genes in Penicillium Species.

    PubMed

    Peterson, Stephen W

    2017-01-01

    Polymerase chain reaction amplification of conserved genes and sequence analysis provides a very powerful tool for the identification of toxigenic as well as non-toxigenic Penicillium species. Sequences are obtained by amplification of the gene fragment, sequencing via capillary electrophoresis of dideoxynucleotide-labeled fragments or NGS. The sequences are compared to a database of validated isolates. Identification of species indicates the potential of the fungus to make particular mycotoxins.

  13. Simple SNP-based minimal marker genotyping for Humulus lupulus L. identification and variety validation.

    PubMed

    Henning, John A; Coggins, Jamie; Peterson, Matthew

    2015-10-06

    in the study to be genetically differentiated from each other. Next, we compared genetic matrices calculated from the minimal marker sets [(Table 2; 6-, 7-, 8-, 10- and 12-marker set matrices] and that of a matrix calculated from a set of markers with no missing data across all 116 samples (1006 SNP markers). The minimum number of markers required to meet both specifications was a set of 7-markers (Table 3). These seven SNPs were then aligned with a genome assembly, and DNA sequence both upstream and downstream were used to identify primer sequences that can be used to develop seven amplicons for high resolution melting curve PCR detection or other SNP-based PCR detection methods. This study identifies a set of 7 SNP markers that may prove useful for the identification and validation of hop varieties and accessions. Variety validation of unknown samples assumes that the variety under question has been included a priori in a discovery panel. These results are based upon in silica studies and markers need to be validated using different SNP marker technology upon a differential set of hop genotypes. The marker sequence data and suggested primer sets provide potential means to fingerprint hop varieties in most genetic laboratories utilizing SNP-marker technology.

  14. Prediction of microRNA target genes using an efficient genetic algorithm-based decision tree.

    PubMed

    Rabiee-Ghahfarrokhi, Behzad; Rafiei, Fariba; Niknafs, Ali Akbar; Zamani, Behzad

    2015-01-01

    MicroRNAs (miRNAs) are small, non-coding RNA molecules that regulate gene expression in almost all plants and animals. They play an important role in key processes, such as proliferation, apoptosis, and pathogen-host interactions. Nevertheless, the mechanisms by which miRNAs act are not fully understood. The first step toward unraveling the function of a particular miRNA is the identification of its direct targets. This step has shown to be quite challenging in animals primarily because of incomplete complementarities between miRNA and target mRNAs. In recent years, the use of machine-learning techniques has greatly increased the prediction of miRNA targets, avoiding the need for costly and time-consuming experiments to achieve miRNA targets experimentally. Among the most important machine-learning algorithms are decision trees, which classify data based on extracted rules. In the present work, we used a genetic algorithm in combination with C4.5 decision tree for prediction of miRNA targets. We applied our proposed method to a validated human datasets. We nearly achieved 93.9% accuracy of classification, which could be related to the selection of best rules.

  15. Prediction of microRNA target genes using an efficient genetic algorithm-based decision tree

    PubMed Central

    Rabiee-Ghahfarrokhi, Behzad; Rafiei, Fariba; Niknafs, Ali Akbar; Zamani, Behzad

    2015-01-01

    MicroRNAs (miRNAs) are small, non-coding RNA molecules that regulate gene expression in almost all plants and animals. They play an important role in key processes, such as proliferation, apoptosis, and pathogen–host interactions. Nevertheless, the mechanisms by which miRNAs act are not fully understood. The first step toward unraveling the function of a particular miRNA is the identification of its direct targets. This step has shown to be quite challenging in animals primarily because of incomplete complementarities between miRNA and target mRNAs. In recent years, the use of machine-learning techniques has greatly increased the prediction of miRNA targets, avoiding the need for costly and time-consuming experiments to achieve miRNA targets experimentally. Among the most important machine-learning algorithms are decision trees, which classify data based on extracted rules. In the present work, we used a genetic algorithm in combination with C4.5 decision tree for prediction of miRNA targets. We applied our proposed method to a validated human datasets. We nearly achieved 93.9% accuracy of classification, which could be related to the selection of best rules. PMID:26649272

  16. Assessment of Dengue virus helicase and methyltransferase as targets for fragment-based drug discovery.

    PubMed

    Coutard, Bruno; Decroly, Etienne; Li, Changqing; Sharff, Andrew; Lescar, Julien; Bricogne, Gérard; Barral, Karine

    2014-06-01

    Seasonal and pandemic flaviviruses continue to be leading global health concerns. With the view to help drug discovery against Dengue virus (DENV), a fragment-based experimental approach was applied to identify small molecule ligands targeting two main components of the flavivirus replication complex: the NS3 helicase (Hel) and the NS5 mRNA methyltransferase (MTase) domains. A library of 500 drug-like fragments was first screened by thermal-shift assay (TSA) leading to the identification of 36 and 32 fragment hits binding Hel and MTase from DENV, respectively. In a second stage, we set up a fragment-based X-ray crystallographic screening (FBS-X) in order to provide both validated fragment hits and structural binding information. No fragment hit was confirmed for DENV Hel. In contrast, a total of seven fragments were identified as DENV MTase binders and structures of MTase-fragment hit complexes were solved at resolution at least 2.0Å or better. All fragment hits identified contain either a five- or six-membered aromatic ring or both, and three novel binding sites were located on the MTase. To further characterize the fragment hits identified by TSA and FBS-X, we performed enzymatic assays to assess their inhibition effect on the N7- and 2'-O-MTase enzymatic activities: five of these fragment hits inhibit at least one of the two activities with IC50 ranging from 180μM to 9mM. This work validates the FBS-X strategy for identifying new anti-flaviviral hits targeting MTase, while Hel might not be an amenable target for fragment-based drug discovery (FBDD). This approach proved to be a fast and efficient screening method for FBDD target validation and discovery of starting hits for the development of higher affinity molecules that bind to novel allosteric sites. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Discerning cultural identification from a thinly sliced behavioral sample.

    PubMed

    Hamamura, Takeshi; Li, Liman Man Wai

    2012-12-01

    This research examined whether individual differences in cultural identification can be discerned at zero acquaintance. This issue was examined in Hong Kong, where the idiosyncrasy of cultural identification is a salient social-psychological issue. The participants were able to perceive accurately the targets' identification with Western culture from a video clip and from a still image. Findings also indicated that a stereotype of Western cultural identity (i.e., extraversion and particular hairstyle) facilitated these perceptions. Specifically, (a) the participants with a stronger stereotype were more accurate in perceiving Western cultural identification, (b) the targets who were experimentally manipulated to appear extraverted were rated as more strongly identifying with Western culture, and (c) the participants relatively unfamiliar with these stereotypes did not correctly perceive Western cultural identification. Implications of these findings on research on multiculturalism are also discussed.

  18. Targeted quantitative analysis of Streptococcus pyogenes virulence factors by multiple reaction monitoring.

    PubMed

    Lange, Vinzenz; Malmström, Johan A; Didion, John; King, Nichole L; Johansson, Björn P; Schäfer, Juliane; Rameseder, Jonathan; Wong, Chee-Hong; Deutsch, Eric W; Brusniak, Mi-Youn; Bühlmann, Peter; Björck, Lars; Domon, Bruno; Aebersold, Ruedi

    2008-08-01

    In many studies, particularly in the field of systems biology, it is essential that identical protein sets are precisely quantified in multiple samples such as those representing differentially perturbed cell states. The high degree of reproducibility required for such experiments has not been achieved by classical mass spectrometry-based proteomics methods. In this study we describe the implementation of a targeted quantitative approach by which predetermined protein sets are first identified and subsequently quantified at high sensitivity reliably in multiple samples. This approach consists of three steps. First, the proteome is extensively mapped out by multidimensional fractionation and tandem mass spectrometry, and the data generated are assembled in the PeptideAtlas database. Second, based on this proteome map, peptides uniquely identifying the proteins of interest, proteotypic peptides, are selected, and multiple reaction monitoring (MRM) transitions are established and validated by MS2 spectrum acquisition. This process of peptide selection, transition selection, and validation is supported by a suite of software tools, TIQAM (Targeted Identification for Quantitative Analysis by MRM), described in this study. Third, the selected target protein set is quantified in multiple samples by MRM. Applying this approach we were able to reliably quantify low abundance virulence factors from cultures of the human pathogen Streptococcus pyogenes exposed to increasing amounts of plasma. The resulting quantitative protein patterns enabled us to clearly define the subset of virulence proteins that is regulated upon plasma exposure.

  19. Validation of ICD-9-CM coding algorithm for improved identification of hypoglycemia visits.

    PubMed

    Ginde, Adit A; Blanc, Phillip G; Lieberman, Rebecca M; Camargo, Carlos A

    2008-04-01

    Accurate identification of hypoglycemia cases by International Classification of Diseases, Ninth Revision, Clinical Modification (ICD-9-CM) codes will help to describe epidemiology, monitor trends, and propose interventions for this important complication in patients with diabetes. Prior hypoglycemia studies utilized incomplete search strategies and may be methodologically flawed. We sought to validate a new ICD-9-CM coding algorithm for accurate identification of hypoglycemia visits. This was a multicenter, retrospective cohort study using a structured medical record review at three academic emergency departments from July 1, 2005 to June 30, 2006. We prospectively derived a coding algorithm to identify hypoglycemia visits using ICD-9-CM codes (250.3, 250.8, 251.0, 251.1, 251.2, 270.3, 775.0, 775.6, and 962.3). We confirmed hypoglycemia cases by chart review identified by candidate ICD-9-CM codes during the study period. The case definition for hypoglycemia was documented blood glucose 3.9 mmol/l or emergency physician charted diagnosis of hypoglycemia. We evaluated individual components and calculated the positive predictive value. We reviewed 636 charts identified by the candidate ICD-9-CM codes and confirmed 436 (64%) cases of hypoglycemia by chart review. Diabetes with other specified manifestations (250.8), often excluded in prior hypoglycemia analyses, identified 83% of hypoglycemia visits, and unspecified hypoglycemia (251.2) identified 13% of hypoglycemia visits. The absence of any predetermined co-diagnosis codes improved the positive predictive value of code 250.8 from 62% to 92%, while excluding only 10 (2%) true hypoglycemia visits. Although prior analyses included only the first-listed ICD-9 code, more than one-quarter of identified hypoglycemia visits were outside this primary diagnosis field. Overall, the proposed algorithm had 89% positive predictive value (95% confidence interval, 86-92) for detecting hypoglycemia visits. The proposed algorithm

  20. SuperTarget goes quantitative: update on drug–target interactions

    PubMed Central

    Hecker, Nikolai; Ahmed, Jessica; von Eichborn, Joachim; Dunkel, Mathias; Macha, Karel; Eckert, Andreas; Gilson, Michael K.; Bourne, Philip E.; Preissner, Robert

    2012-01-01

    There are at least two good reasons for the on-going interest in drug–target interactions: first, drug-effects can only be fully understood by considering a complex network of interactions to multiple targets (so-called off-target effects) including metabolic and signaling pathways; second, it is crucial to consider drug-target-pathway relations for the identification of novel targets for drug development. To address this on-going need, we have developed a web-based data warehouse named SuperTarget, which integrates drug-related information associated with medical indications, adverse drug effects, drug metabolism, pathways and Gene Ontology (GO) terms for target proteins. At present, the updated database contains >6000 target proteins, which are annotated with >330 000 relations to 196 000 compounds (including approved drugs); the vast majority of interactions include binding affinities and pointers to the respective literature sources. The user interface provides tools for drug screening and target similarity inclusion. A query interface enables the user to pose complex queries, for example, to find drugs that target a certain pathway, interacting drugs that are metabolized by the same cytochrome P450 or drugs that target proteins within a certain affinity range. SuperTarget is available at http://bioinformatics.charite.de/supertarget. PMID:22067455

  1. Using the QCM Biosensor-Based T7 Phage Display Combined with Bioinformatics Analysis for Target Identification of Bioactive Small Molecule.

    PubMed

    Takakusagi, Yoichi; Takakusagi, Kaori; Sugawara, Fumio; Sakaguchi, Kengo

    2018-01-01

    Identification of target proteins that directly bind to bioactive small molecule is of great interest in terms of clarifying the mode of action of the small molecule as well as elucidating the biological phenomena at the molecular level. Of the experimental technologies available, T7 phage display allows comprehensive screening of small molecule-recognizing amino acid sequence from the peptide libraries displayed on the T7 phage capsid. Here, we describe the T7 phage display strategy that is combined with quartz-crystal microbalance (QCM) biosensor for affinity selection platform and bioinformatics analysis for small molecule-recognizing short peptides. This method dramatically enhances efficacy and throughput of the screening for small molecule-recognizing amino acid sequences without repeated rounds of selection. Subsequent execution of bioinformatics programs allows combinatorial and comprehensive target protein discovery of small molecules with its binding site, regardless of protein sample insolubility, instability, or inaccessibility of the fixed small molecules to internally located binding site on larger target proteins when conventional proteomics approaches are used.

  2. Mapping the pathways of resistance to targeted therapies

    PubMed Central

    Wood, Kris C.

    2015-01-01

    Resistance substantially limits the depth and duration of clinical responses to targeted anticancer therapies. Through the use of complementary experimental approaches, investigators have revealed that cancer cells can achieve resistance through adaptation or selection driven by specific genetic, epigenetic, or microenvironmental alterations. Ultimately, these diverse alterations often lead to the activation of signaling pathways that, when co-opted, enable cancer cells to survive drug treatments. Recently developed methods enable the direct and scalable identification of the signaling pathways capable of driving resistance in specific contexts. Using these methods, novel pathways of resistance to clinically approved drugs have been identified and validated. By combining systematic resistance pathway mapping methods with studies revealing biomarkers of specific resistance pathways and pharmacological approaches to block these pathways, it may be possible to rationally construct drug combinations that yield more penetrant and lasting responses in patients. PMID:26392071

  3. Targeted next-generation sequencing at copy-number breakpoints for personalized analysis of rearranged ends in solid tumors.

    PubMed

    Kim, Hyun-Kyoung; Park, Won Cheol; Lee, Kwang Man; Hwang, Hai-Li; Park, Seong-Yeol; Sorn, Sungbin; Chandra, Vishal; Kim, Kwang Gi; Yoon, Woong-Bae; Bae, Joon Seol; Shin, Hyoung Doo; Shin, Jong-Yeon; Seoh, Ju-Young; Kim, Jong-Il; Hong, Kyeong-Man

    2014-01-01

    The concept of the utilization of rearranged ends for development of personalized biomarkers has attracted much attention owing to its clinical applicability. Although targeted next-generation sequencing (NGS) for recurrent rearrangements has been successful in hematologic malignancies, its application to solid tumors is problematic due to the paucity of recurrent translocations. However, copy-number breakpoints (CNBs), which are abundant in solid tumors, can be utilized for identification of rearranged ends. As a proof of concept, we performed targeted next-generation sequencing at copy-number breakpoints (TNGS-CNB) in nine colon cancer cases including seven primary cancers and two cell lines, COLO205 and SW620. For deduction of CNBs, we developed a novel competitive single-nucleotide polymorphism (cSNP) microarray method entailing CNB-region refinement by competitor DNA. Using TNGS-CNB, 19 specific rearrangements out of 91 CNBs (20.9%) were identified, and two polymerase chain reaction (PCR)-amplifiable rearrangements were obtained in six cases (66.7%). And significantly, TNGS-CNB, with its high positive identification rate (82.6%) of PCR-amplifiable rearrangements at candidate sites (19/23), just from filtering of aligned sequences, requires little effort for validation. Our results indicate that TNGS-CNB, with its utility for identification of rearrangements in solid tumors, can be successfully applied in the clinical laboratory for cancer-relapse and therapy-response monitoring.

  4. Fast Open-World Person Re-Identification.

    PubMed

    Zhu, Xiatian; Wu, Botong; Huang, Dongcheng; Zheng, Wei-Shi

    2018-05-01

    Existing person re-identification (re-id) methods typically assume that: 1) any probe person is guaranteed to appear in the gallery target population during deployment (i.e., closed-world) and 2) the probe set contains only a limited number of people (i.e., small search scale). Both assumptions are artificial and breached in real-world applications, since the probe population in target people search can be extremely vast in practice due to the ambiguity of probe search space boundary. Therefore, it is unrealistic that any probe person is assumed as one target people, and a large-scale search in person images is inherently demanded. In this paper, we introduce a new person re-id search setting, called large scale open-world (LSOW) re-id, characterized by huge size probe images and open person population in search thus more close to practical deployments. Under LSOW, the under-studied problem of person re-id efficiency is essential in addition to that of commonly studied re-id accuracy. We, therefore, develop a novel fast person re-id method, called Cross-view Identity Correlation and vErification (X-ICE) hashing, for joint learning of cross-view identity representation binarisation and discrimination in a unified manner. Extensive comparative experiments on three large-scale benchmarks have been conducted to validate the superiority and advantages of the proposed X-ICE method over a wide range of the state-of-the-art hashing models, person re-id methods, and their combinations.

  5. Identification of microRNAs in Caragana intermedia by high-throughput sequencing and expression analysis of 12 microRNAs and their targets under salt stress.

    PubMed

    Zhu, Jianfeng; Li, Wanfeng; Yang, Wenhua; Qi, Liwang; Han, Suying

    2013-09-01

    142 miRNAs were identified and 38 miRNA targets were predicted, 4 of which were validated, in C. intermedia . The expression of 12 miRNAs in salt-stressed leaves was assessed by qRT-PCR. MicroRNAs (miRNAs) are endogenous small RNAs that play important roles in various biological and metabolic processes in plants. Caragana intermedia is an important ecological and economic tree species prominent in the desert environment of west and northwest China. To date, no investigation into C. intermedia miRNAs has been reported. In this study, high-throughput sequencing of small RNAs and analysis of transcriptome data were performed to identify both conserved and novel miRNAs, and also their target mRNA genes in C. intermedia. Based on sequence similarity and hairpin structure prediction, 132 putative conserved miRNAs (12 of which were confirmed to form hairpin precursors) belonging to 31 known miRNA families were identified. Ten novel miRNAs (including the miRNA* sequences of three novel miRNAs) were also discovered. Furthermore, 36 potential target genes of 17 known miRNA families and 2 potential target genes of 1 novel miRNA were predicted; 4 of these were validated by 5' RACE. The expression of 12 miRNAs was validated in different tissues, and these and five target mRNAs were assessed by qRT-PCR after salt treatment. The expression levels of seven miRNAs (cin-miR157a, cin-miR159a, cin-miR165a, cin-miR167b, cin-miR172b, cin-miR390a and cin-miR396a) were upregulated, while cin-miR398a expression was downregulated after salt treatment. The targets of cin-miR157a, cin-miR165a, cin-miR172b and cin-miR396a were downregulated and showed an approximately negative correlation with their corresponding miRNAs under salt treatment. These results would help further understanding of miRNA regulation in response to abiotic stress in C. intermedia.

  6. A pan-European ring trial to validate an International Standard for detection of Vibrio cholerae, Vibrio parahaemolyticus and Vibrio vulnificus in seafoods.

    PubMed

    Hartnell, R E; Stockley, L; Keay, W; Rosec, J-P; Hervio-Heath, D; Van den Berg, H; Leoni, F; Ottaviani, D; Henigman, U; Denayer, S; Serbruyns, B; Georgsson, F; Krumova-Valcheva, G; Gyurova, E; Blanco, C; Copin, S; Strauch, E; Wieczorek, K; Lopatek, M; Britova, A; Hardouin, G; Lombard, B; In't Veld, P; Leclercq, A; Baker-Austin, C

    2018-02-10

    Globally, vibrios represent an important and well-established group of bacterial foodborne pathogens. The European Commission (EC) mandated the Comite de European Normalisation (CEN) to undertake work to provide validation data for 15 methods in microbiology to support EC legislation. As part of this mandated work programme, merging of ISO/TS 21872-1:2007, which specifies a horizontal method for the detection of V. parahaemolyticus and V. cholerae, and ISO/TS 21872-2:2007, a similar horizontal method for the detection of potentially pathogenic vibrios other than V. cholerae and V. parahaemolyticus was proposed. Both parts of ISO/TS 21872 utilized classical culture-based isolation techniques coupled with biochemical confirmation steps. The work also considered simplification of the biochemical confirmation steps. In addition, because of advances in molecular based methods for identification of human pathogenic Vibrio spp. classical and real-time PCR options were also included within the scope of the validation. These considerations formed the basis of a multi-laboratory validation study with the aim of improving the precision of this ISO technical specification and providing a single ISO standard method to enable detection of these important foodborne Vibrio spp.. To achieve this aim, an international validation study involving 13 laboratories from 9 countries in Europe was conducted in 2013. The results of this validation have enabled integration of the two existing technical specifications targeting the detection of the major foodborne Vibrio spp., simplification of the suite of recommended biochemical identification tests and the introduction of molecular procedures that provide both species level identification and discrimination of putatively pathogenic strains of V. parahaemolyticus by the determination of the presence of theromostable direct and direct related haemolysins. The method performance characteristics generated in this have been included in revised

  7. Measurement and validation of benchmark-quality thick-target tungsten X-ray spectra below 150 kVp.

    PubMed

    Mercier, J R; Kopp, D T; McDavid, W D; Dove, S B; Lancaster, J L; Tucker, D M

    2000-11-01

    Pulse-height distributions of two constant potential X-ray tubes with fixed anode tungsten targets were measured and unfolded. The measurements employed quantitative alignment of the beam, the use of two different semiconductor detectors (high-purity germanium and cadmium-zinc-telluride), two different ion chamber systems with beam-specific calibration factors, and various filter and tube potential combinations. Monte Carlo response matrices were generated for each detector for unfolding the pulse-height distributions into spectra incident on the detectors. These response matrices were validated for the low error bars assigned to the data. A significant aspect of the validation of spectra, and a detailed characterization of the X-ray tubes, involved measuring filtered and unfiltered beams at multiple tube potentials (30-150 kVp). Full corrections to ion chamber readings were employed to convert normalized fluence spectra into absolute fluence spectra. The characterization of fixed anode pitting and its dominance over exit window plating and/or detector dead layer was determined. An Appendix of tabulated benchmark spectra with assigned error ranges was developed for future reference.

  8. Simultaneous identification and DNA barcoding of six Eimeria species infecting turkeys using PCR primers targeting the mitochondrial cytochrome c oxidase subunit I (mtCOI) locus.

    PubMed

    Hafeez, Mian A; Shivaramaiah, Srichaitanya; Dorsey, Kristi Moore; Ogedengbe, Mosun E; El-Sherry, Shiem; Whale, Julia; Cobean, Julie; Barta, John R

    2015-05-01

    Species-specific PCR primers targeting the mitochondrial cytochrome c oxidase subunit I (mtCOI) locus were generated that allow for the specific identification of the most common Eimeria species infecting turkeys (i.e., Eimeria adenoeides, Eimeria meleagrimitis, Eimeria gallopavonis, Eimeria meleagridis, Eimeria dispersa, and Eimeria innocua). PCR reaction chemistries were optimized with respect to divalent cation (MgCl2) and dNTP concentrations, as well as PCR cycling conditions (particularly anneal temperature for primers). Genomic DNA samples from single oocyst-derived lines of six Eimeria species were tested to establish specificity and sensitivity of these newly designed primer pairs. A mixed 60-ng total DNA sample containing 10 ng of each of the six Eimeria species was used as DNA template to demonstrate specific amplification of the correct product using each of the species-specific primer pairs. Ten nanograms of each of the five non-target Eimeria species was pooled to provide a non-target, control DNA sample suitable to test the specificity of each primer pair. The amplifications of the COI region with species-specific primer pairs from pooled samples yielded products of expected sizes (209 to 1,012 bp) and no amplification of non-target Eimeria sp. DNA was detected using the non-target, control DNA samples. These primer pairs specific for Eimeria spp. of turkeys did not amplify any of the seven Eimeria species infecting chickens. The newly developed PCR primers can be used as a diagnostic tool capable of specifically identifying six turkey Eimeria species; additionally, sequencing of the PCR amplification products yields sequence-based genotyping data suitable for identification and molecular phylogenetics.

  9. Validation of the kidney disease quality of life-short form: a cross-sectional study of a dialysis-targeted health measure in Singapore.

    PubMed

    Joshi, Veena D; Mooppil, Nandakumar; Lim, Jeremy Fy

    2010-12-20

    In Singapore, the prevalence of end-stage renal disease (ESRD) and the number of people on dialysis is increasing. The impact of ESRD on patient quality of life has been recognized as an important outcome measure. The Kidney Disease Quality Of Life-Short Form (KDQOL-SF™) has been validated and is widely used as a measure of quality of life in dialysis patients in many countries, but not in Singapore. We aimed to determine the reliability and validity of the KDQOL-SF™ for haemodialysis patients in Singapore. From December 2006 through January 2007, this cross-sectional study gathered data on patients ≥21 years old, who were undergoing haemodialysis at National Kidney Foundation in Singapore. We used exploratory factor analysis to determine construct validity of the eight KDQOL-SF™ sub-scales, Cronbach's alpha coefficient to determine internal consistency reliability, correlation of the overall health rating with kidney disease-targeted scales to confirm validity, and correlation of the eight sub-scales with age, income and education to determine convergent and divergent validity. Of 1980 haemodialysis patients, 1180 (59%) completed the KDQOL-SF™. Full information was available for 980 participants, with a mean age of 56 years. The sample was representative of the total dialysis population in Singapore, except Indian ethnicity that was over-represented. The instrument designers' proposed eight sub-scales were confirmed, which together accounted for 68.4% of the variance. All sub-scales had a Cronbach's α above the recommended minimum value of 0.7 to indicate good reliability (range: 0.72 to 0.95), except for Social function (0.66). Correlation of items within subscales was higher than correlation of items outside subscales in 90% of the cases. The overall health rating positively correlated with kidney disease-targeted scales, confirming validity. General health subscales were found to have significant associations with age, income and education

  10. A forward model-based validation of cardiovascular system identification

    NASA Technical Reports Server (NTRS)

    Mukkamala, R.; Cohen, R. J.

    2001-01-01

    We present a theoretical evaluation of a cardiovascular system identification method that we previously developed for the analysis of beat-to-beat fluctuations in noninvasively measured heart rate, arterial blood pressure, and instantaneous lung volume. The method provides a dynamical characterization of the important autonomic and mechanical mechanisms responsible for coupling the fluctuations (inverse modeling). To carry out the evaluation, we developed a computational model of the cardiovascular system capable of generating realistic beat-to-beat variability (forward modeling). We applied the method to data generated from the forward model and compared the resulting estimated dynamics with the actual dynamics of the forward model, which were either precisely known or easily determined. We found that the estimated dynamics corresponded to the actual dynamics and that this correspondence was robust to forward model uncertainty. We also demonstrated the sensitivity of the method in detecting small changes in parameters characterizing autonomic function in the forward model. These results provide confidence in the performance of the cardiovascular system identification method when applied to experimental data.

  11. Identification of (antioxidative) plants in herbal pharmaceutical preparations and dietary supplements.

    PubMed

    Deconinck, Eric; Custers, Deborah; De Beer, Jacques Omer

    2015-01-01

    The standard procedures for the identification, authentication, and quality control of medicinal plants and herbs are nowadays limited to pure herbal products. No guidelines or procedures, describing the detection or identification of a targeted plant or herb in pharmaceutical preparations or dietary supplements, can be found. In these products the targeted plant is often present together with other components of herbal or synthetic origin. This chapter describes a strategy for the fast development of a chromatographic fingerprint approach that allows the identification of a targeted plant in herbal preparations and dietary supplements. The strategy consists of a standard chromatographic gradient that is tested for the targeted plant with different extraction solvents and different mobile phases. From the results obtained, the optimal fingerprint is selected. Subsequently the samples are analyzed according to the selected methodological parameters, and the obtained fingerprints can be compared with the one obtained for the pure herbal product or a standard preparation. Calculation of the dissimilarity between these fingerprints will result in a probability of presence of the targeted plant. Optionally mass spectrometry can be used to improve specificity, to confirm identification, or to identify molecules with a potential medicinal or antioxidant activity.

  12. Identification of cutting force coefficients in machining process considering cutter vibration

    NASA Astrophysics Data System (ADS)

    Yao, Qi; Luo, Ming; Zhang, Dinghua; Wu, Baohai

    2018-03-01

    Among current cutting force models, cutting force coefficients still are the foundation of predicting calculation combined with consideration of geometry engagement variation, equipment characteristics, material properties and so on. Attached with unimpeachable significance, the traditional and some novel identification methods of cutting force coefficient are still faced with trouble, including repeated onerous work, over ideal measuring condition, variation of value due to material divergence, interference from measuring units. To utilize the large amount of data from real manufacturing section, enlarge data sources and enrich cutting data base for former prediction task, a novel identification method is proposed by considering stiffness properties of the cutter-holder-spindle system in this paper. According to previously proposed studies, the direct result of cutter vibration is the form of dynamic undeformed chip thickness. This fluctuation is considered in two stages of this investigation. Firstly, a cutting force model combined with cutter vibration is established in detailed way. Then, on the foundation of modeling, a novel identification method is developed, in which the dynamic undeformed chip thickness could be obtained by using collected data. In a carefully designed experiment procedure, the reliability of model is validated by comparing predicted and measured results. Under different cutting condition and cutter stiffness, data is collected for the justification of identification method. The results showed divergence in calculated coefficients is acceptable confirming the possibility of accomplishing targets by applying this new method. In discussion, the potential directions of improvement are proposed.

  13. Species and hybrid identification of sturgeon caviar: a new molecular approach to detect illegal trade.

    PubMed

    Boscari, E; Barmintseva, A; Pujolar, J M; Doukakis, P; Mugue, N; Congiu, L

    2014-05-01

    Overexploitation of wild populations due to the high economic value of caviar has driven sturgeons to near extinction. The high prices commanded by caviar on world markets have made it a magnet for illegal and fraudulent caviar trade, often involving low-value farmed caviar being sold as top-quality caviar. We present a new molecular approach for the identification of pure sturgeon species and hybrids that are among the most commercialized species in Europe and North America. Our test is based on the discovery of species-specific single nucleotide polymorphisms (SNPs) in the ribosomal protein S7, supplemented with the Vimentin gene and the mitochondrial D-loop. Test validations performed in 702 specimens of target and nontarget sturgeon species demonstrated a 100% identification success for Acipenser naccarii, A. fulvescens, A. stellatus, A. sinensis and A. transmontanus. In addition to species identification, our approach allows the identification of Bester and AL hybrids, two of the most economically important hybrids in the world, with 80% and 100% success, respectively. Moreover, the approach has the potential to identify many other existing sturgeon hybrids. The development of a standardized sturgeon identification tool will directly benefit trade law enforcement, providing the tools to monitor and regulate the legal trade of caviar and protect sturgeon stocks from illicit producers and traders, hence contributing to safeguarding this group of heavily threatened species. © 2013 John Wiley & Sons Ltd.

  14. Approach and Instrument Placement Validation

    NASA Technical Reports Server (NTRS)

    Ator, Danielle

    2005-01-01

    The Mars Exploration Rovers (MER) from the 2003 flight mission represents the state of the art technology for target approach and instrument placement on Mars. It currently takes 3 sols (Martian days) for the rover to place an instrument on a designated rock target that is about 10 to 20 m away. The objective of this project is to provide an experimentally validated single-sol instrument placement capability to future Mars missions. After completing numerous test runs on the Rocky8 rover under various test conditions, it has been observed that lighting conditions, shadow effects, target features and the initial target distance have an effect on the performance and reliability of the tracking software. Additional software validation testing will be conducted in the months to come.

  15. Clinical Validation of Copy Number Variant Detection from Targeted Next-Generation Sequencing Panels.

    PubMed

    Kerkhof, Jennifer; Schenkel, Laila C; Reilly, Jack; McRobbie, Sheri; Aref-Eshghi, Erfan; Stuart, Alan; Rupar, C Anthony; Adams, Paul; Hegele, Robert A; Lin, Hanxin; Rodenhiser, David; Knoll, Joan; Ainsworth, Peter J; Sadikovic, Bekim

    2017-11-01

    Next-generation sequencing (NGS) technology has rapidly replaced Sanger sequencing in the assessment of sequence variations in clinical genetics laboratories. One major limitation of current NGS approaches is the ability to detect copy number variations (CNVs) approximately >50 bp. Because these represent a major mutational burden in many genetic disorders, parallel CNV assessment using alternate supplemental methods, along with the NGS analysis, is normally required, resulting in increased labor, costs, and turnaround times. The objective of this study was to clinically validate a novel CNV detection algorithm using targeted clinical NGS gene panel data. We have applied this approach in a retrospective cohort of 391 samples and a prospective cohort of 2375 samples and found a 100% sensitivity (95% CI, 89%-100%) for 37 unique events and a high degree of specificity to detect CNVs across nine distinct targeted NGS gene panels. This NGS CNV pipeline enables stand-alone first-tier assessment for CNV and sequence variants in a clinical laboratory setting, dispensing with the need for parallel CNV analysis using classic techniques, such as microarray, long-range PCR, or multiplex ligation-dependent probe amplification. This NGS CNV pipeline can also be applied to the assessment of complex genomic regions, including pseudogenic DNA sequences, such as the PMS2CL gene, and to mitochondrial genome heteroplasmy detection. Copyright © 2017 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

  16. Nuclease Target Site Selection for Maximizing On-target Activity and Minimizing Off-target Effects in Genome Editing

    PubMed Central

    Lee, Ciaran M; Cradick, Thomas J; Fine, Eli J; Bao, Gang

    2016-01-01

    The rapid advancement in targeted genome editing using engineered nucleases such as ZFNs, TALENs, and CRISPR/Cas9 systems has resulted in a suite of powerful methods that allows researchers to target any genomic locus of interest. A complementary set of design tools has been developed to aid researchers with nuclease design, target site selection, and experimental validation. Here, we review the various tools available for target selection in designing engineered nucleases, and for quantifying nuclease activity and specificity, including web-based search tools and experimental methods. We also elucidate challenges in target selection, especially in predicting off-target effects, and discuss future directions in precision genome editing and its applications. PMID:26750397

  17. A network model of genomic hormone interactions underlying dementia and its translational validation through serendipitous off-target effect

    PubMed Central

    2013-01-01

    Background While the majority of studies have focused on the association between sex hormones and dementia, emerging evidence supports the role of other hormone signals in increasing dementia risk. However, due to the lack of an integrated view on mechanistic interactions of hormone signaling pathways associated with dementia, molecular mechanisms through which hormones contribute to the increased risk of dementia has remained unclear and capacity of translating hormone signals to potential therapeutic and diagnostic applications in relation to dementia has been undervalued. Methods Using an integrative knowledge- and data-driven approach, a global hormone interaction network in the context of dementia was constructed, which was further filtered down to a model of convergent hormone signaling pathways. This model was evaluated for its biological and clinical relevance through pathway recovery test, evidence-based analysis, and biomarker-guided analysis. Translational validation of the model was performed using the proposed novel mechanism discovery approach based on ‘serendipitous off-target effects’. Results Our results reveal the existence of a well-connected hormone interaction network underlying dementia. Seven hormone signaling pathways converge at the core of the hormone interaction network, which are shown to be mechanistically linked to the risk of dementia. Amongst these pathways, estrogen signaling pathway takes the major part in the model and insulin signaling pathway is analyzed for its association to learning and memory functions. Validation of the model through serendipitous off-target effects suggests that hormone signaling pathways substantially contribute to the pathogenesis of dementia. Conclusions The integrated network model of hormone interactions underlying dementia may serve as an initial translational platform for identifying potential therapeutic targets and candidate biomarkers for dementia-spectrum disorders such as Alzheimer

  18. A database de-identification framework to enable direct queries on medical data for secondary use.

    PubMed

    Erdal, B S; Liu, J; Ding, J; Chen, J; Marsh, C B; Kamal, J; Clymer, B D

    2012-01-01

    To qualify the use of patient clinical records as non-human-subject for research purpose, electronic medical record data must be de-identified so there is minimum risk to protected health information exposure. This study demonstrated a robust framework for structured data de-identification that can be applied to any relational data source that needs to be de-identified. Using a real world clinical data warehouse, a pilot implementation of limited subject areas were used to demonstrate and evaluate this new de-identification process. Query results and performances are compared between source and target system to validate data accuracy and usability. The combination of hashing, pseudonyms, and session dependent randomizer provides a rigorous de-identification framework to guard against 1) source identifier exposure; 2) internal data analyst manually linking to source identifiers; and 3) identifier cross-link among different researchers or multiple query sessions by the same researcher. In addition, a query rejection option is provided to refuse queries resulting in less than preset numbers of subjects and total records to prevent users from accidental subject identification due to low volume of data. This framework does not prevent subject re-identification based on prior knowledge and sequence of events. Also, it does not deal with medical free text de-identification, although text de-identification using natural language processing can be included due its modular design. We demonstrated a framework resulting in HIPAA Compliant databases that can be directly queried by researchers. This technique can be augmented to facilitate inter-institutional research data sharing through existing middleware such as caGrid.

  19. Species-specific identification of Dekkera/Brettanomyces yeasts by fluorescently labeled DNA probes targeting the 26S rRNA.

    PubMed

    Röder, Christoph; König, Helmut; Fröhlich, Jürgen

    2007-09-01

    Sequencing of the complete 26S rRNA genes of all Dekkera/Brettanomyces species colonizing different beverages revealed the potential for a specific primer and probe design to support diagnostic PCR approaches and FISH. By analysis of the complete 26S rRNA genes of all five currently known Dekkera/Brettanomyces species (Dekkera bruxellensis, D. anomala, Brettanomyces custersianus, B. nanus and B. naardenensis), several regions with high nucleotide sequence variability yet distinct from the D1/D2 domains were identified. FISH species-specific probes targeting the 26S rRNA gene's most variable regions were designed. Accessibility of probe targets for hybridization was facilitated by the construction of partially complementary 'side'-labeled probes, based on secondary structure models of the rRNA sequences. The specificity and routine applicability of the FISH-based method for yeast identification were tested by analyzing different wine isolates. Investigation of the prevalence of Dekkera/Brettanomyces yeasts in the German viticultural regions Wonnegau, Nierstein and Bingen (Rhinehesse, Rhineland-Palatinate) resulted in the isolation of 37 D. bruxellensis strains from 291 wine samples.

  20. Validation of CoaBC as a Bactericidal Target in the Coenzyme A Pathway of Mycobacterium tuberculosis.

    PubMed

    Evans, Joanna C; Trujillo, Carolina; Wang, Zhe; Eoh, Hyungjin; Ehrt, Sabine; Schnappinger, Dirk; Boshoff, Helena I M; Rhee, Kyu Y; Barry, Clifton E; Mizrahi, Valerie

    2016-12-09

    Mycobacterium tuberculosis relies on its own ability to biosynthesize coenzyme A to meet the needs of the myriad enzymatic reactions that depend on this cofactor for activity. As such, the essential pantothenate and coenzyme A biosynthesis pathways have attracted attention as targets for tuberculosis drug development. To identify the optimal step for coenzyme A pathway disruption in M. tuberculosis, we constructed and characterized a panel of conditional knockdown mutants in coenzyme A pathway genes. Here, we report that silencing of coaBC was bactericidal in vitro, whereas silencing of panB, panC, or coaE was bacteriostatic over the same time course. Silencing of coaBC was likewise bactericidal in vivo, whether initiated at infection or during either the acute or chronic stages of infection, confirming that CoaBC is required for M. tuberculosis to grow and persist in mice and arguing against significant CoaBC bypass via transport and assimilation of host-derived pantetheine in this animal model. These results provide convincing genetic validation of CoaBC as a new bactericidal drug target.

  1. Open-source chemogenomic data-driven algorithms for predicting drug-target interactions.

    PubMed

    Hao, Ming; Bryant, Stephen H; Wang, Yanli

    2018-02-06

    While novel technologies such as high-throughput screening have advanced together with significant investment by pharmaceutical companies during the past decades, the success rate for drug development has not yet been improved prompting researchers looking for new strategies of drug discovery. Drug repositioning is a potential approach to solve this dilemma. However, experimental identification and validation of potential drug targets encoded by the human genome is both costly and time-consuming. Therefore, effective computational approaches have been proposed to facilitate drug repositioning, which have proved to be successful in drug discovery. Doubtlessly, the availability of open-accessible data from basic chemical biology research and the success of human genome sequencing are crucial to develop effective in silico drug repositioning methods allowing the identification of potential targets for existing drugs. In this work, we review several chemogenomic data-driven computational algorithms with source codes publicly accessible for predicting drug-target interactions (DTIs). We organize these algorithms by model properties and model evolutionary relationships. We re-implemented five representative algorithms in R programming language, and compared these algorithms by means of mean percentile ranking, a new recall-based evaluation metric in the DTI prediction research field. We anticipate that this review will be objective and helpful to researchers who would like to further improve existing algorithms or need to choose appropriate algorithms to infer potential DTIs in the projects. The source codes for DTI predictions are available at: https://github.com/minghao2016/chemogenomicAlg4DTIpred. Published by Oxford University Press 2018. This work is written by US Government employees and is in the public domain in the US.

  2. Efforts Aimed To Reduce Attrition in Antimalarial Drug Discovery: A Systematic Evaluation of the Current Antimalarial Targets Portfolio.

    PubMed

    Chaparro, María Jesús; Calderón, Félix; Castañeda, Pablo; Fernández-Alvaro, Elena; Gabarró, Raquel; Gamo, Francisco Javier; Gómez-Lorenzo, María G; Martín, Julio; Fernández, Esther

    2018-04-13

    Malaria remains a major global health problem. In 2015 alone, more than 200 million cases of malaria were reported, and more than 400,000 deaths occurred. Since 2010, emerging resistance to current front-line ACTs (artemisinin combination therapies) has been detected in endemic countries. Therefore, there is an urgency for new therapies based on novel modes of action, able to relieve symptoms as fast as the artemisinins and/or block malaria transmission. During the past few years, the antimalarial community has focused their efforts on phenotypic screening as a pragmatic approach to identify new hits. Optimization efforts on several chemical series have been successful, and clinical candidates have been identified. In addition, recent advances in genetics and proteomics have led to the target deconvolution of phenotypic clinical candidates. New mechanisms of action will also be critical to overcome resistance and reduce attrition. Therefore, a complementary strategy focused on identifying well-validated targets to start hit identification programs is essential to reinforce the clinical pipeline. Leveraging published data, we have assessed the status quo of the current antimalarial target portfolio with a focus on the blood stage clinical disease. From an extensive list of reported Plasmodium targets, we have defined triage criteria. These criteria consider genetic, pharmacological, and chemical validation, as well as tractability/doability, and safety implications. These criteria have provided a quantitative score that has led us to prioritize those targets with the highest probability to deliver successful and differentiated new drugs.

  3. Time domain nonlinear SMA damper force identification approach and its numerical validation

    NASA Astrophysics Data System (ADS)

    Xin, Lulu; Xu, Bin; He, Jia

    2012-04-01

    Most of the currently available vibration-based identification approaches for structural damage detection are based on eigenvalues and/or eigenvectors extracted from vibration measurements and, strictly speaking, are only suitable for linear system. However, the initiation and development of damage in engineering structures under severe dynamic loadings are typical nonlinear procedure. Studies on the identification of restoring force which is a direct indicator of the extent of the nonlinearity have received increasing attention in recent years. In this study, a date-based time domain identification approach for general nonlinear system was developed. The applied excitation and the corresponding response time series of the structure were used for identification by means of standard least-square techniques and a power series polynomial model (PSPM) which was utilized to model the nonlinear restoring force (NRF). The feasibility and robustness of the proposed approach was verified by a 2 degree-of-freedoms (DOFs) lumped mass numerical model equipped with a shape memory ally (SMA) damper mimicking nonlinear behavior. The results show that the proposed data-based time domain method is capable of identifying the NRF in engineering structures without any assumptions on the mass distribution and the topology of the structure, and provides a promising way for damage detection in the presence of structural nonlinearities.

  4. Systematic Identification, Characterization and Target Gene Analysis of microRNAs Involved in Osteoarthritis Subchondral Bone Pathogenesis.

    PubMed

    Prasadam, Indira; Batra, Jyotsna; Perry, Samuel; Gu, Wenyi; Crawford, Ross; Xiao, Yin

    2016-07-01

    This study aimed to identify the microRNAs associated with sclerotic status of subchondral bone in the pathogenesis of osteoarthritis (OA). Total RNA was extracted from non-sclerotic and sclerotic OA subchondral bone from patients undergoing knee replacement surgeries. miRCURY™ LNA miRNA chip and qRT-PCR were used to profile and validate differential microRNA expression. In addition, we further confirmed profiles of altered miRNAs in an OA rat meniscectomy animal model and their putative targets of the miRNAs were predicted using ingenuity (IPA) software. Finally, five short-listed miRNAs were reactivated by transient in vitro overexpression (miRNA mimics) in subchondral bone osteoblasts and their phenotypes were assessed. Functional screening identified 30 differentiated miRNAs in sclerotic subchondral bone compared to non-sclerotic bone of OA patients. Data integration resulted in confirmation of the eight miRNAs, with aberrant expression in independent human OA bone sample set. In silico analysis (IPA) identified 732 mRNA transcripts as putative targets of the eight altered miRNAs, of which twenty genes were validated to be differentially expressed in sclerotic compared to non-sclerotic bone samples. Out of eight dysregulated miRNA's, five of them showed consistent time-dependent downregulation in a rat OA model. Furthermore, synthetic miR-199a-3p, miR-199a-5p, miR-590-5p, and miR-211-5p mimics rescued the abnormal osteoarthritic subchondral bone osteoblast gene expression and mineralization. We have identified four novel miRNAs that play important roles in subchondral bone pathogenesis in OA. Additional studies are required to develop these miRNAs into therapeutic modalities for OA.

  5. Validation of a New Web Application for Identification of Fungi by Use of Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry

    PubMed Central

    Becker, P.; Gabriel, F.; Cassagne, C.; Accoceberry, I.; Gari-Toussaint, M.; Hasseine, L.; De Geyter, D.; Pierard, D.; Surmont, I.; Djenad, F.; Donnadieu, J. L.; Piarroux, M.; Hendrickx, M.; Piarroux, R.

    2017-01-01

    ABSTRACT Matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometry has emerged as a reliable technique to identify molds involved in human diseases, including dermatophytes, provided that exhaustive reference databases are available. This study assessed an online identification application based on original algorithms and an extensive in-house reference database comprising 11,851 spectra (938 fungal species and 246 fungal genera). Validation criteria were established using an initial panel of 422 molds, including dermatophytes, previously identified via DNA sequencing (126 species). The application was further assessed using a separate panel of 501 cultured clinical isolates (88 mold taxa including dermatophytes) derived from five hospital laboratories. A total of 438 (87.35%) isolates were correctly identified at the species level, while 26 (5.22%) were assigned to the correct genus but the wrong species and 37 (7.43%) were not identified, since the defined threshold of 20 was not reached. The use of the Bruker Daltonics database included in the MALDI Biotyper software resulted in a much higher rate of unidentified isolates (39.76 and 74.30% using the score thresholds 1.7 and 2.0, respectively). Moreover, the identification delay of the online application remained compatible with real-time online queries (0.15 s per spectrum), and the application was faster than identifications using the MALDI Biotyper software. This is the first study to assess an online identification system based on MALDI-TOF spectrum analysis. We have successfully applied this approach to identify molds, including dermatophytes, for which diversity is insufficiently represented in commercial databases. This free-access application is available to medical mycologists to improve fungal identification. PMID:28637907

  6. Validation of a New Web Application for Identification of Fungi by Use of Matrix-Assisted Laser Desorption Ionization-Time of Flight Mass Spectrometry.

    PubMed

    Normand, A C; Becker, P; Gabriel, F; Cassagne, C; Accoceberry, I; Gari-Toussaint, M; Hasseine, L; De Geyter, D; Pierard, D; Surmont, I; Djenad, F; Donnadieu, J L; Piarroux, M; Ranque, S; Hendrickx, M; Piarroux, R

    2017-09-01

    Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry has emerged as a reliable technique to identify molds involved in human diseases, including dermatophytes, provided that exhaustive reference databases are available. This study assessed an online identification application based on original algorithms and an extensive in-house reference database comprising 11,851 spectra (938 fungal species and 246 fungal genera). Validation criteria were established using an initial panel of 422 molds, including dermatophytes, previously identified via DNA sequencing (126 species). The application was further assessed using a separate panel of 501 cultured clinical isolates (88 mold taxa including dermatophytes) derived from five hospital laboratories. A total of 438 (87.35%) isolates were correctly identified at the species level, while 26 (5.22%) were assigned to the correct genus but the wrong species and 37 (7.43%) were not identified, since the defined threshold of 20 was not reached. The use of the Bruker Daltonics database included in the MALDI Biotyper software resulted in a much higher rate of unidentified isolates (39.76 and 74.30% using the score thresholds 1.7 and 2.0, respectively). Moreover, the identification delay of the online application remained compatible with real-time online queries (0.15 s per spectrum), and the application was faster than identifications using the MALDI Biotyper software. This is the first study to assess an online identification system based on MALDI-TOF spectrum analysis. We have successfully applied this approach to identify molds, including dermatophytes, for which diversity is insufficiently represented in commercial databases. This free-access application is available to medical mycologists to improve fungal identification. Copyright © 2017 American Society for Microbiology.

  7. A practical guide to epidemiological practice and standards in the identification and validation of diagnostic markers using a bladder cancer example.

    PubMed

    Behrens, T; Bonberg, N; Casjens, S; Pesch, B; Brüning, T

    2014-01-01

    Technical advances to analyze biological markers have generated a plethora of promising new marker candidates for early detection of cancer. However, in subsequent analyses only few could be successfully validated as being predictive, clinically useful, or effective. This failure is partially due to rapid publication of results that were detected in early stages of biomarker research. Methodological considerations are a major concern when carrying out molecular epidemiological studies of diagnostic markers to avoid errors that increase the potential for bias. Although guidelines for conducting studies and reporting of results have been published to improve the quality of marker studies, their planning and execution still need to be improved. We will discuss different sources of bias in study design, handling of specimens, and statistical analysis to illustrate possible pitfalls associated with marker research, and present legal, ethical, and technical considerations associated with storage and handling of specimens. This article presents a guide to epidemiological standards in marker research using bladder cancer as an example. Because of the possibility to detect early cancer stages due to leakage of molecular markers from the target organ or exfoliation of tumor cells into the urine, bladder cancer is particularly useful to study diagnostic markers. To improve the overall quality of marker research, future developments should focus on networks of studies and tissue banks according to uniform legal, ethical, methodological, and technical standards. This article is part of a Special Issue entitled: Computational Proteomics in the Post-Identification Era. Guest Editors: Martin Eisenacher and Christian Stephan. © 2013.

  8. Group-level self-definition and self-investment: a hierarchical (multicomponent) model of in-group identification.

    PubMed

    Leach, Colin Wayne; van Zomeren, Martijn; Zebel, Sven; Vliek, Michael L W; Pennekamp, Sjoerd F; Doosje, Bertjan; Ouwerkerk, Jaap W; Spears, Russell

    2008-07-01

    Recent research shows individuals' identification with in-groups to be psychologically important and socially consequential. However, there is little agreement about how identification should be conceptualized or measured. On the basis of previous work, the authors identified 5 specific components of in-group identification and offered a hierarchical 2-dimensional model within which these components are organized. Studies 1 and 2 used confirmatory factor analysis to validate the proposed model of self-definition (individual self-stereotyping, in-group homogeneity) and self-investment (solidarity, satisfaction, and centrality) dimensions, across 3 different group identities. Studies 3 and 4 demonstrated the construct validity of the 5 components by examining their (concurrent) correlations with established measures of in-group identification. Studies 5-7 demonstrated the predictive and discriminant validity of the 5 components by examining their (prospective) prediction of individuals' orientation to, and emotions about, real intergroup relations. Together, these studies illustrate the conceptual and empirical value of a hierarchical multicomponent model of in-group identification.

  9. Application of multi-target phytotherapeutic concept in malaria drug discovery: a systems biology approach in biomarker identification.

    PubMed

    Tarkang, Protus Arrey; Appiah-Opong, Regina; Ofori, Michael F; Ayong, Lawrence S; Nyarko, Alexander K

    2016-01-01

    There is an urgent need for new anti-malaria drugs with broad therapeutic potential and novel mode of action, for effective treatment and to overcome emerging drug resistance. Plant-derived anti-malarials remain a significant source of bioactive molecules in this regard. The multicomponent formulation forms the basis of phytotherapy. Mechanistic reasons for the poly-pharmacological effects of plants constitute increased bioavailability, interference with cellular transport processes, activation of pro-drugs/deactivation of active compounds to inactive metabolites and action of synergistic partners at different points of the same signaling cascade. These effects are known as the multi-target concept. However, due to the intrinsic complexity of natural products-based drug discovery, there is need to rethink the approaches toward understanding their therapeutic effect. This review discusses the multi-target phytotherapeutic concept and its application in biomarker identification using the modified reverse pharmacology - systems biology approach. Considerations include the generation of a product library, high throughput screening (HTS) techniques for efficacy and interaction assessment, High Performance Liquid Chromatography (HPLC)-based anti-malarial profiling and animal pharmacology. This approach is an integrated interdisciplinary implementation of tailored technology platforms coupled to miniaturized biological assays, to track and characterize the multi-target bioactive components of botanicals as well as identify potential biomarkers. While preserving biodiversity, this will serve as a primary step towards the development of standardized phytomedicines, as well as facilitate lead discovery for chemical prioritization and downstream clinical development.

  10. Targeting of PHOX2B expression allows the identification of drugs effective in counteracting neuroblastoma cell growth

    PubMed Central

    Zanni, Eleonora Di; Bianchi, Giovanna; Ravazzolo, Roberto; Raffaghello, Lizzia; Ceccherini, Isabella; Bachetti, Tiziana

    2017-01-01

    The pathogenic role of the PHOX2B gene in neuroblastoma is indicated by heterozygous mutations in neuroblastoma patients and by gene overexpression in both neuroblastoma cell lines and tumor samples. PHOX2B encodes a transcription factor which is crucial for the correct development and differentiation of sympathetic neurons. PHOX2B overexpression is considered a prognostic marker for neuroblastoma and it is also used by clinicians to monitor minimal residual disease. Furthermore, it has been observed that neuronal differentiation in neuroblastoma is dependent on down-regulation of PHOX2B expression, which confirms that PHOX2B expression may be considered a target in neuroblastoma. Here, PHOX2B promoter or 3′ untranslated region were used as molecular targets in an in vitro high-throughput approach that led to the identification of molecules able to decrease PHOX2B expression at transcriptional and likely even at post-transcriptional levels. Further functional investigations carried out on PHOX2B mRNA levels and biological consequences, such as neuroblastoma cell apoptosis and growth, showed that chloroquine and mycophenolate mofetil are most promising agents for neuroblastoma therapy based on down-regulation of PHOX2B expression. Finally, a strong correlation between the effect of drugs in terms of down-regulation of PHOX2B expression and of biological consequences in neuroblastoma cells confirms the role of PHOX2B as a potential molecular target in neuroblastoma. PMID:29069774

  11. Predicting essential genes for identifying potential drug targets in Aspergillus fumigatus.

    PubMed

    Lu, Yao; Deng, Jingyuan; Rhodes, Judith C; Lu, Hui; Lu, Long Jason

    2014-06-01

    Aspergillus fumigatus (Af) is a ubiquitous and opportunistic pathogen capable of causing acute, invasive pulmonary disease in susceptible hosts. Despite current therapeutic options, mortality associated with invasive Af infections remains unacceptably high, increasing 357% since 1980. Therefore, there is an urgent need for the development of novel therapeutic strategies, including more efficacious drugs acting on new targets. Thus, as noted in a recent review, "the identification of essential genes in fungi represents a crucial step in the development of new antifungal drugs". Expanding the target space by rapidly identifying new essential genes has thus been described as "the most important task of genomics-based target validation". In previous research, we were the first to show that essential gene annotation can be reliably transferred between distantly related four Prokaryotic species. In this study, we extend our machine learning approach to the much more complex Eukaryotic fungal species. A compendium of essential genes is predicted in Af by transferring known essential gene annotations from another filamentous fungus Neurospora crassa. This approach predicts essential genes by integrating diverse types of intrinsic and context-dependent genomic features encoded in microbial genomes. The predicted essential datasets contained 1674 genes. We validated our results by comparing our predictions with known essential genes in Af, comparing our predictions with those predicted by homology mapping, and conducting conditional expressed alleles. We applied several layers of filters and selected a set of potential drug targets from the predicted essential genes. Finally, we have conducted wet lab knockout experiments to verify our predictions, which further validates the accuracy and wide applicability of the machine learning approach. The approach presented here significantly extended our ability to predict essential genes beyond orthologs and made it possible to

  12. High-throughput identification and dendritic cell-based functional validation of MHC class I-restricted Mycobacterium tuberculosis epitopes

    PubMed Central

    Nair, Smita K.; Tomaras, Georgia D.; Sales, Ana Paula; Boczkowski, David; Chan, Cliburn; Plonk, Kelly; Cai, Yongting; Dannull, Jens; Kepler, Thomas B.; Pruitt, Scott K.; Weinhold, Kent J.

    2014-01-01

    Emergence of drug-resistant strains of the pathogen Mycobacterium tuberculosis (Mtb) and the ineffectiveness of BCG in curtailing Mtb infection makes vaccine development for tuberculosis an important objective. Identifying immunogenic CD8+ T cell peptide epitopes is necessary for peptide-based vaccine strategies. We present a three-tiered strategy for identifying and validating immunogenic peptides: first, identify peptides that form stable complexes with class I MHC molecules; second, determine whether cytotoxic T lymphocytes (CTLs) raised against the whole protein antigen recognize and lyse target cells pulsed with peptides that passed step 1; third, determine whether peptides that passed step 2, when administered in vivo as a vaccine in HLA-A2 transgenic mice, elicit CTLs that lyse target cells expressing the whole protein antigen. Our innovative approach uses dendritic cells transfected with Mtb antigen-encoding mRNA to drive antigen expression. Using this strategy, we have identified five novel peptide epitopes from the Mtb proteins Apa, Mtb8.4 and Mtb19. PMID:24755960

  13. A targeted genotyping approach enhances identification of variants in taste receptor and appetite/reward genes of potential functional importance for obesity-related porcine traits.

    PubMed

    Cirera, S; Clop, A; Jacobsen, M J; Guerin, M; Lesnik, P; Jørgensen, C B; Fredholm, M; Karlskov-Mortensen, P

    2018-04-01

    Taste receptors (TASRs) and appetite and reward (AR) mechanisms influence eating behaviour, which in turn affects food intake and risk of obesity. In a previous study, we used next generation sequencing to identify potentially functional mutations in TASR and AR genes and found indications for genetic associations between identified variants and growth and fat deposition in a subgroup of animals (n = 38) from the UNIK resource pig population. This population was created for studying obesity and obesity-related diseases. In the present study we validated results from our previous study by investigating genetic associations between 24 selected single nucleotide variants in TASR and AR gene variants and 35 phenotypes describing obesity and metabolism in the entire UNIK population (n = 564). Fifteen variants showed significant association with specific obesity-related phenotypes after Bonferroni correction. Six of the 15 genes, namely SIM1, FOS, TAS2R4, TAS2R9, MCHR2 and LEPR, showed good correlation between known biological function and associated phenotype. We verified a genetic association between potentially functional variants in TASR/AR genes and growth/obesity and conclude that the combination of identification of potentially functional variants by next generation sequencing followed by targeted genotyping and association studies is a powerful and cost-effective approach for increasing the power of genetic association studies. © 2018 Stichting International Foundation for Animal Genetics.

  14. Genomic identification of direct target genes of LEAFY

    PubMed Central

    William, Dilusha A.; Su, Yanhui; Smith, Michael R.; Lu, Meina; Baldwin, Don A.; Wagner, Doris

    2004-01-01

    The switch from vegetative to reproductive development in plants necessitates a switch in the developmental program of the descendents of the stem cells in the shoot apical meristem. Genetic and molecular investigations have demonstrated that the plant-specific transcription factor and meristem identity regulator LEAFY (LFY) controls this developmental transition by inducing expression of a second transcription factor, APETALA1, and by regulating the expression of additional, as yet unknown, genes. Here we show that the additional LFY targets include the APETALA1-related factor, CAULI-FLOWER, as well as three transcription factors and two putative signal transduction pathway components. These genes are up-regulated by LFY even when protein synthesis is inhibited and, hence, appear to be direct targets of LFY. Supporting this conclusion, cis-regulatory regions upstream of these genes are bound by LFY in vivo. The newly identified LFY targets likely initiate the transcriptional changes that are required for the switch from vegetative to reproductive development in Arabidopsis. PMID:14736918

  15. 21 CFR 886.1880 - Fusion and stereoscopic target.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Fusion and stereoscopic target. 886.1880 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1880 Fusion and stereoscopic target. (a) Identification. A fusion and stereoscopic target is a device intended for use as a viewing object...

  16. 21 CFR 886.1880 - Fusion and stereoscopic target.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Fusion and stereoscopic target. 886.1880 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1880 Fusion and stereoscopic target. (a) Identification. A fusion and stereoscopic target is a device intended for use as a viewing object...

  17. A Novel mRNA Level Subtraction Method for Quick Identification of Target-Orientated Uniquely Expressed Genes Between Peanut Immature Pod and Leaf

    PubMed Central

    2010-01-01

    Subtraction technique has been broadly applied for target gene discovery. However, most current protocols apply relative differential subtraction and result in great amount clone mixtures of unique and differentially expressed genes. This makes it more difficult to identify unique or target-orientated expressed genes. In this study, we developed a novel method for subtraction at mRNA level by integrating magnetic particle technology into driver preparation and tester–driver hybridization to facilitate uniquely expressed gene discovery between peanut immature pod and leaf through a single round subtraction. The resulting target clones were further validated through polymerase chain reaction screening using peanut immature pod and leaf cDNA libraries as templates. This study has resulted in identifying several genes expressed uniquely in immature peanut pod. These target genes can be used for future peanut functional genome and genetic engineering research. PMID:21406066

  18. High-throughput identification of off-targets for the mechanistic study of severe adverse drug reactions induced by analgesics

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pan, Jian-Bo; Ji, Nan; Pan, Wen

    2014-01-01

    Drugs may induce adverse drug reactions (ADRs) when they unexpectedly bind to proteins other than their therapeutic targets. Identification of these undesired protein binding partners, called off-targets, can facilitate toxicity assessment in the early stages of drug development. In this study, a computational framework was introduced for the exploration of idiosyncratic mechanisms underlying analgesic-induced severe adverse drug reactions (SADRs). The putative analgesic-target interactions were predicted by performing reverse docking of analgesics or their active metabolites against human/mammal protein structures in a high-throughput manner. Subsequently, bioinformatics analyses were undertaken to identify ADR-associated proteins (ADRAPs) and pathways. Using the pathways and ADRAPsmore » that this analysis identified, the mechanisms of SADRs such as cardiac disorders were explored. For instance, 53 putative ADRAPs and 24 pathways were linked with cardiac disorders, of which 10 ADRAPs were confirmed by previous experiments. Moreover, it was inferred that pathways such as base excision repair, glycolysis/glyconeogenesis, ErbB signaling, calcium signaling, and phosphatidyl inositol signaling likely play pivotal roles in drug-induced cardiac disorders. In conclusion, our framework offers an opportunity to globally understand SADRs at the molecular level, which has been difficult to realize through experiments. It also provides some valuable clues for drug repurposing. - Highlights: • A novel computational framework was developed for mechanistic study of SADRs. • Off-targets of drugs were identified in large scale and in a high-throughput manner. • SADRs like cardiac disorders were systematically explored in molecular networks. • A number of ADR-associated proteins were identified.« less

  19. Prediction of miRNA targets.

    PubMed

    Oulas, Anastasis; Karathanasis, Nestoras; Louloupi, Annita; Pavlopoulos, Georgios A; Poirazi, Panayiota; Kalantidis, Kriton; Iliopoulos, Ioannis

    2015-01-01

    Computational methods for miRNA target prediction are currently undergoing extensive review and evaluation. There is still a great need for improvement of these tools and bioinformatics approaches are looking towards high-throughput experiments in order to validate predictions. The combination of large-scale techniques with computational tools will not only provide greater credence to computational predictions but also lead to the better understanding of specific biological questions. Current miRNA target prediction tools utilize probabilistic learning algorithms, machine learning methods and even empirical biologically defined rules in order to build models based on experimentally verified miRNA targets. Large-scale protein downregulation assays and next-generation sequencing (NGS) are now being used to validate methodologies and compare the performance of existing tools. Tools that exhibit greater correlation between computational predictions and protein downregulation or RNA downregulation are considered the state of the art. Moreover, efficiency in prediction of miRNA targets that are concurrently verified experimentally provides additional validity to computational predictions and further highlights the competitive advantage of specific tools and their efficacy in extracting biologically significant results. In this review paper, we discuss the computational methods for miRNA target prediction and provide a detailed comparison of methodologies and features utilized by each specific tool. Moreover, we provide an overview of current state-of-the-art high-throughput methods used in miRNA target prediction.

  20. Construct Validation of the Fairy Tale Test--Standardization Data.

    ERIC Educational Resources Information Center

    Coulacoglou, Carina

    2002-01-01

    Studied the construct validity of the Fairy Tale Test (C. Coulacoglu, 1993), a personality projective test for children, in a sample of 800 Greek children aged 8, 10, and 12. Factor analysis led to identification of eight primary factors, and correlations with other measures provide construct validity evidence. (SLD)