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PubMed1995

Independent contributions of the orienting of attention, fixation offset and bilateral stimulation on human saccadic latencies.

In a series of experiments we examined the effects of the endogenous orienting of visual attention on human saccade latency. Three separate manipulations were performed: the orienting of visual attention, the prior offset of fixation (gap paradigm) and the bilateral presentation of saccade targets. Each of these manipulations was shown to make an independent contribution to saccade latency. In experiments 1 and 2 subjects were instructed to orient their attention covertly to a location by a verbal pre-cue; targets could appear in the attended hemifield (valid) or in the non-attended hemifield (invalid) together with a no-instruction (neutral) condition. Saccades were made under fixation gap and overlap conditions, to either single target or two bilaterally presented targets which appeared at equal and opposite eccentricities in both hemifields. The results showed a large increase (cost) of saccade latency to invalid targets and a small non-significant decrease (benefit) of saccade latency to valid targets. The cost associated with invalid targets replicates the "meridan crossing effect" shown in manual reaction time experiments and is consistent with the hemifield inhibition and premotor models of attentional orienting. The use of a "gap" procedure produced a generalised facilitation of saccade latency, which was not modified by the prior orienting of visual attention. The magnitude of the gap effect was similar for saccades made to attended and non-attended stimulis. This suggests that the gap effect may be due to ocular motor disengagement, or a warning signal effect, rather than to the prior disengagement of visual attention. When two targets were presented simultaneously, one in each hemifield, saccade latency was slowed compared with the single target condition. The magnitude of this slowing was unaffected by the prior orienting of visual attention or by the fixation condition. The slowing was examined in more detail in experiment 3, by presenting targets with brief offset delays. The latency increase was maximal if the two targets were presented simultaneously and decreased if the distractor appeared at short intervals (20-80 ms) before or after the saccade target onset. If the non-attended stimulus was presented at greater intervals (160, 240 ms) before the saccade target, then a facilitation effect was observed. This demonstrates that the onset of a distractor in the non-attended hemifield can have both an inhibitory and a facilitatory effect on a saccade production.

Attention↗
PubMed2005

1,4-bis(3-aminopropyl)piperazine libraries: from the discovery of classical chloroquine-like antimalarials to the identification of new targets.

The purpose of this review is to provide an update on our work based on the 1,4-bis(3-aminopropyl)piperazine skeleton and how it allowed our group to validate a new target. After a brief introduction where we will relate the way this substructure was introduced in our 4-aminoquinolinyl derivatives, we will present first the different libraries synthesized around this moiety: (1) libraries of sulfonamides, amides and amines derived from 4-aminoquinolines and, (2) libraries where the 4-aminoquinoline nucleus is replaced. High throughput evaluation of biological activity and physicochemical parameters will be presented. The evaluation of the anti-malarial activity of the compounds will be discussed in the light of a chloroquine-like mechanism (accumulation in the acidic food vacuole and inhibition of beta-hematin formation). In a second part we will present active 1,4-bis(3-aminopropyl)piperazine as tools for identification and/or validation of new antimalarial targets. Fluorescence assays on some derivatives show that they are surprisingly localized outside the food vacuole, suggesting the existence of other target(s). Secondly, we will present a library of 1,4-bis(3-aminopropyl)piperazine as inhibitors of the cytosolic aminopeptidase Pfa-M1, a new potential target for antimalarials.

Antimalarials↗
PubMed1994

Adult age differences in shifting focused attention.

Young and older adults performed a choice response task in which 1 of 2 target letters was presented visually at 1 of 4 display locations. In 2 experiments, the validity of a target location cue and the presence of nontarget characters (distractors) were varied. With target-only displays and 40% cue validity (Experiment 1), the estimated time to shift attention between display locations was essentially 0 ms for both age groups. With 70% cue validity, Experiment 2 demonstrated significant increases in the attention shift time as a function of both increased age and the presence of distractors (asterisks). The results suggest that age-related changes in the shifting of focused attention are minimal except when the processing of nontarget information is required.

Adolescent↗
PubMed2004

Focus groups in psychological assessment: enhancing content validity by consulting members of the target population.

A review of articles in Psychological Assessment reveals that many researchers develop instruments without the benefit of consultation with members of the target population. To the extent that researchers do consult the target population, most fail to bring consultation in early enough to inform the identification and specification of key constructs. Moreover, this consultation typically takes the form of one-to-one interviews. The authors' goal in this article was to elaborate on the importance of population consultation as part of content validation and to critically evaluate the potential of one method of consultation, focus groups, to inform multiple stages of instrument development. The authors suggest that this method holds promise for enhancing the content validity of instruments and, ultimately, the validity of research findings.

Combat Disorders↗
PubMedDate not supplied

Considerations in compound database preparation--"hidden" impact on virtual screening results.

Structure-based virtual screening (SBVS) utilizing docking algorithms has become an essential tool in the drug discovery process, and significant progress has been made in successfully applying the technique to a wide range of receptor targets. In silico validation of virtual screening protocols before application to a receptor target using a corporate or commercially available compound collection is key to establishing a successful process. Ultimately, retrieval of a set of active compounds from a database of inactives is required, and the metric of enrichment (E) is habitually used to discern the quality of separation of the two. Numerous reports have addressed the performance of docking algorithms with regard to the quality of binding mode prediction and the issue of postprocessing "hit lists" of docked ligands. However, the impact of ligand database preprocessing has yet to be examined in the context of virtual screening and prioritization of compounds for biological evaluation. We provide an insight into the implications of cheminformatic preprocessing of a validation database of compounds where multiple protonated, tautomeric, stereochemical, and conformational states have been enumerated. Several commonly used methods for the generation of ligand conformations and conformational ensembles are examined, paired with an exhaustive rigid-body algorithm for the docking of different "multimeric" compound representations to the ligand binding site of the human estrogen receptor alpha. Chemgauss, a shapegaussian scoring function with intrinsic chemical knowledge, was combined with PLP as a consensus-scoring scheme to rank output from the docking protocol and enrichment rates calculated for each screen. The overheads of CPU consumption and the effect on relative database size (disk requirement) for each of the protocols employed are considered. Assessment of these parameters indicates that SBVS enrichments are highly dependent on the initial cheminformatic treatment(s) used in database construction. The interplay of SMILES representations, stereochemical information, protonation state enumeration, and ligand conformation ensembles are critical in achieving optimum enrichment rates in such screening.

Computer Simulation↗
PubMed2005

In vivo efficacy of HSV-TK transcriptionally targeted to the tumour vasculature is augmented by combination with cytotoxic chemotherapy.

BACKGROUND: Retroviral vectors are suitable for targeting endothelial cells in the tumour neovasculature because of their intrinsic selectivity for proliferating cells. Previously, we inserted regulatory elements of the endothelial-specific prepro-endothelin-1 (ppET1) promoter in retroviral vectors to generate high-titre, replication-defective recombinant retroviruses that restricted gene expression to the vascular compartment of tumours. METHODS: A retroviral vector was generated in which expression of herpes simplex virus thymidine kinase (HSV-TK) was transcriptionally restricted to endothelial cells, under the control of a hybrid ppET-1 LTR. Xenograft tumour models were used to determine the efficacy of targeting HSV-TK to the tumour vasculature. Subsequently, vascular-targeted gene therapy was combined with chemotherapeutic agents. RESULTS: Breast or colorectal xenograft tumour growth was reduced and survival was increased in response to ganciclovir treatment. Treatment resulted in widespread vascular disruption and tumour cell apoptosis. In colorectal tumours, combination with irinotecan, a cytotoxic drug used to treat colorectal cancer, significantly increased survival compared to drug alone. No beneficial effect on survival was observed when combined with cisplatin, a cytotoxic drug not in clinical use for this tumour type. On the basis of their relative efficacies in vitro against tumour and endothelial cells, co-operativity with irinotecan likely derives from additionally targeting the peripheral tumour cells that survive the anti-vascular treatment. CONCLUSIONS: We show that the ppET1-targeted vector is efficacious for therapeutic gene expression in vivo, validating a strategy targeted to tumour vasculature, and demonstrate that vascular targeting combined with appropriate chemotherapy is more effective than either therapy alone.

Animals↗
PubMed2006

Antithrombotic therapy in cardiac surgery.

PURPOSE: To review the perioperative management of antithrombotic therapy in cardiac surgery, including the management of cardiopulmonary bypass (CPB) and off-pump surgery. METHODS: A review of the relevant English literature over the period 1975-2005 was undertaken, in addition to a review of international practices in antithrombotic therapy in cardiac surgery. PRINCIPAL FINDINGS: Cardiopulmonary bypass is required in most procedures and makes anticoagulation mandatory. Anticoagulation is, usually, achieved with unfractionnated heparin (UFH). Unfractionated heparin is monitored by point-of-care (POC) testing, such as the activated clotting time or the determination of heparin concentration. The target values of both tests remain empirical, with no clearly validated thresholds. The target value needs to be adjusted according to the POC test, given significant variations between devices and activators. After CABG, the need for antiplatelet therapy is well demonstrated, in order to limit the risk of postoperative death or ischemic events, and improve venous graft patency. Immediately after valvular surgery, antithrombotic therapy should take into account the specific risk carried by each patient and by each prosthetic device. The risk of venous thromboembolism, though poorly defined, is also present in the postoperative period and may require additional attention. Given the frequent exposure to UFH, occurrence of heparin-induced thrombocytopenia is not infrequent in these patients and requires careful individual management. CONCLUSIONS: Antithrombotic therapy is an essential component of cardiac surgery. Yet, with the exception of antiplatelet agents in CABG patients, antithrombotic therapy is often based on the clinical experience of medical teams more than on an evidence-based assessment of the literature.

Anticoagulants↗
PubMed2003

Knockouts model the 100 best-selling drugs--will they model the next 100?

The biopharmaceutical industry is currently faced with a tremendous number of potential drug targets identified through the sequencing of the human genome. The challenge ahead is to delineate those targets with the greatest value for therapeutic intervention. Here, we critically evaluate mouse-knockout technology for target discovery and validation. A retrospective evaluation of the knockout phenotypes for the targets of the 100 best-selling drugs indicates that these phenotypes correlate well with known drug efficacy, illuminating a productive path forward for discovering future drug targets. Prospective mining of the druggable genome is being catalysed by large-scale mouse knockout programs combined with phenotypic screens focused on identifying targets that modulate mammalian physiology in a therapeutically relevant manner.

Animals↗
PubMed2004

Covert orienting to exogenous and endogenous cues in children with spina bifida.

Children with spina bifida meningomyelocele and hydrocephalus (SBM) have congenital dysmorphology of the midbrain and thinning of the posterior cortex, brain regions associated with the control of covert orienting. We studied cued covert orienting in 92 children with SBM, and 40 age-matched typically developing controls. Cues were of three types: exogenous (luminance change in a peripheral box either valid or invalid for upcoming target location), endogenous arrow (a central arrow either valid or invalid for upcoming target location), or endogenous word (a central word either valid or invalid for upcoming target location). Compared to controls, children with SBM showed slowed covert orienting to both exogenous and endogenous cues and a higher cost of attentional disengagement (e.g., a greater cue-validity effect) for exogenous although not for endogenous cues. Covert orienting deficits were associated with midbrain dysmorphology in the form of beaking of the tectum, and with right posterior brain volume loss.

Adolescent↗
PubMed2005

Intrathecal gene transfer by adeno-associated virus for pain.

Chronic pain is among the most prevalent medical problems, affecting more than half of patients with advanced cancer and many with other common diseases. Current analgesics often fail to provide satisfactory symptom relief and frequently cause severe side effects. Intrathecal (IT) gene transfer is an attractive method for pain research in rodent models, because it allows targeting of a wide variety of secretable peptides and proteins to the spinal cord, an important neural center for the processing of nociceptive signals. The potential of IT gene transfer for improving opioid therapy and for validating new analgesic targets, such as cytokines involved in spinal glial activation, is discussed. The IT space has been notoriously resistant to efficient gene transfer, limiting therapeutic gene expression to less than 2 weeks with most vector systems. Recent progress with adeno-associated virus (AAV) technology allowed efficient long-term gene expression, facilitating studies reflective of the chronic nature of many pain states. AAV is one of the most advanced gene therapy vectors currently undergoing clinical trials for a variety of disorders. In patients, AAV vectors could be administered intrathecally by a lumbar puncture, a safe procedure routinely performed at the bedside. AAV vectors may therefore become an important tool for translational studies to validate newly identified therapeutic targets in clinical pain states.

Analgesics, Opioid↗
PubMed2002

Recognizing the un-real McCoy: priming and the modularity of face recognition.

Fodor (1983) has proposed that face perception is carried out by an informationally encapsulated module, whose operation is unaffected by context or expectancies. We tested the modularity hypothesis by examining whether discriminations between normal and distorted versions of famous faces can be primed, either by the name of an associated person (semantic context) or by a valid cue as to the identity of the target face (expectancy). A preliminary experiment showed that, in the absence of priming, discriminations between normal and distorted versions of a face were unaffected by whether the target faces were familiar or not, confirming that these judgments tap perceptual, not postperceptual (semantic), coding processes. In Experiment 1, accuracy was significantly higher when target face pairs were preceded by related name primes, as compared with unrelated ones. In Experiment 2, reaction times were significantly faster for targets preceded by a valid identity cue than for targets preceded by an invalid one. Neither effect could be explained as a speed-accuracy tradeoff. These results fail to support Fodor's conjecture that face processing is encapsulated.

Adult↗
PubMed2025

Lit-OTAR framework for extracting biological evidences from literature.

SUMMARY: The lit-OTAR framework, developed through a collaboration between Europe PMC and Open Targets, leverages deep learning to revolutionize drug discovery by extracting evidence from scientific literature for drug target identification and validation. This novel framework combines named entity recognition for identifying gene/protein (target), disease, organism, and chemical/drug within scientific texts, and entity normalization to map these entities to databases like Ensembl, Experimental Factor Ontology, and ChEMBL. Continuously operational, it has processed over 39 million abstracts and 4.5 million full-text articles and preprints to date, identifying more than 48.5 million unique associations that significantly help accelerate the drug discovery process and scientific research >29.9 m distinct target-disease, 11.8 m distinct target-drug, and 8.3 m distinct disease-drug relationships. AVAILABILITY AND IMPLEMENTATION: The results are accessible through Europe PMC's SciLite web app (https://europepmc.org/) and its annotations API (https://europepmc.org/annotationsapi), as well as via the Open Targets Platform (https://platform.opentargets.org/). The daily pipeline is available at https://github.com/ML4LitS/otar-maintenance, and the Open Targets ETL processes are available at https://github.com/opentargets.

Drug Discovery↗
PubMed2006

Microplate assay for inhibitors of the transpeptidase activity of PBP1b of Escherichia coli.

The transpeptidase (TP) activity of penicillin-binding proteins (PBPs), target of the beta-lactam antibiotics, is a well-validated antibacterial drug target. The TP activity of PBP1b converts un-cross-linked peptidoglycan to the cross-linked form. Directly measuring TP activity is difficult because cross-linked and un-cross-linked peptidoglycan have very similar chromatographic properties. The authors report a microdilution plate method to directly measure the TP enzyme activity, uncoupled from the transglycosylase (TG), for detection of TP inhibitors. Escherichia coli membranes were incubated with 100 mM ampicillin, followed by removal of unbound ampicillin. The substrate for the TP, un-cross-linked peptidoglycan, was prepared by incubating these membranes with peptidoglycan sugar precursors, 1 of which was radiolabeled. Subsequently, solubilized PBP1b was added and TP activity assayed. The cross-linked peptidoglycan formed was monitored by addition of wheat germ agglutinin scintillation proximity assay beads plus N-laurylsarcosine, which selectively captures cross-linked peptidoglycan. The PBP1bcatalyzed activity was inhibited by penicillin G but not by cephalexin or cephradine, which have higher affinity for PBP1a. Moenomycin, a TG inhibitor, also inhibited TP activity. Because this is a true enzyme assay, it has the potential to detect novel, non-beta-lactam TP inhibitors and could lead to the discovery of new antibacterial agents.

Bacterial Proteins↗
PubMed2005

Progress in the discovery of polo-like kinase inhibitors.

Polo-like kinases (PLKs) are key enzymes that control mitotic entry of proliferating cells and regulate many aspects of mitosis necessary for successful cytokinesis. Of the four known human PLKs, PLK1 is the best characterized and is overexpressed in many tumour types with aberrant elevation frequently constituting a prognostic indicator of poor disease outcome. Despite the fact that PLK1 has been regarded as a validated mitotic cancer target for a number of years, very few reports of small-molecule PLK inhibitors have appeared to date. In order to provide a starting point for the discovery and development of selective PLK inhibitors, we have characterized a number of known generic kinase inhibitors with hitherto unknown activity against PLK1, as well as discovering novel inhibitors through structure-guided design. Previously, the only characterized biochemical PLK1 inhibitor was scytonemin, a symmetric indolic marine natural product that is a micromolar non-specific ATP competitor. In addition to the progress in the development of ATP-competitive small-molecule PLK inhibitors, recent reports on the use of antisense oligonucleotides (ASONs) and small interfering RNAs (siRNAs) directed against PLK1 have shown selective antiproliferative effects on cancer cells both in vitro and in vivo, producing phenotypes consistent with known PLK functions, and confirming that targeting PLKs with conventional small-molecule agents may be a valid and effective anticancer strategy. Here we present a progress update on the approaches taken so far in developing PLK inhibitors.

Amino Acid Sequence↗
PubMed2006

A loss-of-function RNA interference screen for molecular targets in cancer.

The pursuit of novel therapeutic agents in cancer relies on the identification and validation of molecular targets. Hallmarks of cancer include self-sufficiency in growth signals and evasion from apoptosis; genes that regulate these processes may be optimal for therapeutic attack. Here we describe a loss-of-function screen for genes required for the proliferation and survival of cancer cells using an RNA interference library. We used a doxycycline-inducible retroviral vector for the expression of small hairpin RNAs (shRNAs) to construct a library targeting 2,500 human genes. We used retroviral pools from this library to infect cell lines representing two distinct molecular subgroups of diffuse large B-cell lymphoma (DLBCL), termed activated B-cell-like DLBCL and germinal centre B-cell-like DLBCL. Each vector was engineered to contain a unique 60-base-pair 'bar code', allowing the abundance of an individual shRNA vector within a population of transduced cells to be measured using microarrays of the bar-code sequences. We observed that a subset of shRNA vectors was depleted from the transduced cells after three weeks in culture only if shRNA expression was induced. In activated B-cell-like DLBCL cells, but not germinal centre B-cell-like DLBCL cells, shRNAs targeting the NF-kappaB pathway were depleted, in keeping with the essential role of this pathway in the survival of activated B-cell-like DLBCL. This screen uncovered CARD11 as a key upstream signalling component responsible for the constitutive IkappaB kinase activity in activated B-cell-like DLBCL. The methodology that we describe can be used to establish a functional taxonomy of cancer and help reveal new classes of therapeutic targets distinct from known oncogenes.

Adaptor Proteins, Signal Transducing↗
PubMed2003

A transient tobacco expression system coupled to MALDI-TOF-MS allows validation of the impact of differential targeting on structure and activity of a recombinant therapeutic glycoprotein produced in plants.

Tobacco-based transient expression was employed to elucidate the impact of differential targeting to subcellular compartments on activity and quality of gastric lipase as a model for the production of recombinant glycoproteins in plants. Overall N-linked glycan structures of recombinant lipase were analyzed and for the first time sugar structures of its four individual N-glycosylation sites were determined in situ by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) on a trypsin digest without isolation or deglycosylation of the peptides. Three glycosylation sites contain both complex-type N-glycans and high-mannose-type structures, the fourth is exclusively linked to high-mannose glycans. Although the overall pattern of glycan structures is influenced by the targeting, our results show that the type of glycans found linked to a given Asn residue is largely influenced by the physico-chemical environment of the site. The transient tobacco system combined with MALDI-TOF-MS appears to be a useful tool for the evaluation of glycoprotein production in plants.

Animals↗
PubMed2001

Geometry of metal-ligand interactions in proteins.

The geometry of metal-ligand interactions in proteins is examined and compared with information for small-molecule complexes from the Cambridge Structural Database (CSD). The paper deals with the metals Ca, Mg, Mn, Fe, Cu, Zn and with metal-donor atom distances, coordination numbers and extent of distortion from ideal geometry (octahedral, tetrahedral etc.). It assesses the agreement between geometry found in all metalloprotein structures in the Protein Data Bank (PDB) determined at resolution < or = 1.6 A with that predicted from the CSD for ligands which are analogues of amino-acid side chains in proteins [Harding (1999), Acta Cryst. D55, 1432-1443; Harding (2000), Acta Cryst. D56, 857-867]. The agreement is reasonably good for these structures but poorer for many determined at lower resolution (examined to 2.8 A resolution). For metal-donor distances, the predictions from the CSD, with minor adjustments, provide good targets either for validation or for restraints in refinement of structures where only poorer resolution data is available. These target distances are tabulated and the use of restraints is recommended. Validation of angles or the use in refinement of restraints on angles at the metal atom is more difficult because of the inherent flexibility of these angles. A much simplified set of parameters for angle restraints with quite large standard deviations is provided. (Despite the flexibility of the angles, acceptable and preferred coordination numbers and shapes are well established and a summary table is provided.) An unusual and perhaps biochemically important feature of Zn coordination with carboxylate seen in the CSD examples is also clearly present in metalloprotein structures. With metals like Ca, carboxylate coordination is monodentate or bidentate (two M-O bonds of nearly equal length). In Zn carboxylates a continuous range between monodentate and bidentate coordination is found, with one Zn-O bond of normal length and another of any length between this and a van der Waals contact.

Amino Acids↗