Discovery of Novel Allosteric Modulators of Metabotropic Glutamate Receptor Subtype 5 Reveals Chemical and Functional Diversity and In Vivo Activity in Rat Behavioral Models of Anxiolytic and Antipsychotic Activity

Modulators of metabotropic glutamate receptor subtype 5 (mGluR5) may provide novel treatments for multiple central nervous system (CNS) disorders, including anxiety and schizophrenia. Although compounds have been developed to better understand the physiological roles of mGluR5 and potential usefulness for the treatment of these disorders, there are limitations in the tools available, including poor selectivity, low potency, and limited solubility. To address these issues, we developed an innovative assay that allows simultaneous screening for mGluR5 agonists, antagonists, and potentiators. We identified multiple scaffolds that possess diverse modes of activity at mGluR5, including both positive and negative allosteric modulators (PAMs and NAMs, respectively). 3-Fluoro-5-(3-(pyridine-2-yl)-1,2,4-oxadiazol-5-yl) benzonitrile (VU0285683) was developed as a novel selective mGluR5 NAM with high affinity for the 2-methyl-6-(phenyl-ethynyl)-pyridine (MPEP) binding site. VU0285683 had anxiolytic-like activity in two rodent models for anxiety but did not potentiate phen-cyclidine-induced hyperlocomotor activity. (4-Hydroxypiperidin-1-yl)(4-phenylethynyl) phenyl) methanone (VU0092273) was identified as a novel mGluR5 PAM that also binds to the MPEP site. VU0092273 was chemically optimized to an orally active analog, N-cyclobutyl-6-((3-fluorophenyl) ethynyl) nicotinamide hydrochloride (VU0360172), which is selective for mGluR5. This novel mGluR5 PAM produced a dose-dependent reversal of amphetamine-induced hyperlocomotion, a rodent model predictive of antipsychotic activity. Discovery of structurally and functionally diverse allosteric modulators of mGluR5 that demonstrate in vivo efficacy in rodent models of anxiety and antipsychotic activity provide further support for the tremendous diversity of chemical scaffolds and modes of efficacy of mGluR5 ligands. In addition, these studies provide strong support for the hypothesis that multiple structurally distinct mGluR5 modulators have robust activity in animal models that predict efficacy in the treatment of CNS disorders.

Positive allosteric modulators of the metabotropic glutamate receptor subtype 4 (mGluR4). Part II: Challenges in hit-to-lead

This Letter describes the synthesis and SAR of two mGluR4 positive allosteric modulator leads, 6 and 7. VU001171 (6) represents the most potent (EC50 = 650 nM), efficacious (141% Glu Max) and largest fold shift (36-fold) of any mGluR4 PAM reported to date. However, this work highlights the challenges in hit-to-lead for mGluR4 PAMs, with multiple confirmed HTS hits displaying little or no tractable SAR. (C) 2008 Elsevier Ltd. All rights reserved.

Synthesis and SAR of novel, 4-(phenylsulfamoyl)phenylacetamide mGlu(4) positive allosteric modulators (PAMs) identified by functional high-throughput screening (HTS)

Herein we disclose the synthesis and SAR of a series of 4-(phenylsulfamoyl)phenylacetamide compounds as mGlu(4) positive allosteric modulators (PAMs) that were identified via a functional HTS. An iterative parallel approach to these compounds culminated in the discovery of VU0364439 (11) which represents the most potent (19.8 nM) mGlu(4) PAM reported to date. (C) 2010 Elsevier Ltd. All rights reserved.

Optimization of an ether series of mGluR5 positive allosteric modulators: Molecular determinants of MPEP site interaction crossover

We report the optimization of a series of non-MPEP site metabotropic glutamate receptor 5 (mGlu5) pos. allosteric modulators (PAMs) based on a simple acyclic ether series. Modifications led to a gain of MPEP site interaction through incorporation of a chiral amide in conjunction with a nicotinamide core. A highly potent PAM, 8v (VU0404251), was shown to be efficacious in a rodent model of psychosis. These studies suggest that potent PAMs within topol. similar chemotypes can be developed to preferentially interact or not interact with the MPEP allosteric binding site.

Discovery and SAR of a novel series of non-MPEP site mGlu(5) PAMs based on an aryl glycine sulfonammide scaffold

Herein we report the discovery and SAR of a novel series of non-MPEP site metabotropic glutamate receptor 5 (mGlu(5)) positive allosteric modulators (PAMs) based on an aryl glycine sulfonamide scaffold. This series represents a rare non-MPEP site mGlu(5) PAM chemotype.

Using Non-Monetary Deprivation Indicators to Analyze Poverty and Social Exclusion: Lessons from Europe?

Non-monetary indicators of deprivation are now widely used in studying poverty in Europe. While measuring financial resources remains central, having reliable information about material deprivation adds to the ability to capture poverty and social exclusion. Non-monetary indicators can help improve the identification of those experiencing poverty and understand how it comes about. They are most productively used when multidimensionality is explicitly taken into account, both in framing the question and in empirical application. While serious methodological and measurement issues remain to be addressed, material deprivation indicators allow for new insights in making poverty comparisons across countries and analyzing changes over time. (C) 2010 by the Association for Public Policy Analysis and Management.

Polyamine system in developing rat eye and an animal model of retinopathy of prematurity

BACKGROUND: In experimental models of retinopathy of prematurity (ROP), a vasoproliferative disorder of the retina, retinal lesions are usually assessed by morphological examination. However, studies suggest that the polyamine system may be useful in monitoring proliferation processes. For this reason, polyamine concentrations in rat erythrocytes (RBC) and the regulation of polyamine system in rat eyes under the conditions relevant to ROP were investigated. METHODS: Newborn Wistar rats were reared in room air (control) or exposed first to hyperoxia (60% or 80% oxygen, 2 weeks) and then to normoxia (relative hypoxia, 1 or 2 weeks). Blood was collected from orbital vessels at 2 weeks of age and before death. Polyamine system-related enzyme activities were measured in retina and lens with radioassays. Polyamines were quantified by fluorometry after extraction, dansylation and HPLC separation. RESULTS: Oxygen (80% only) significantly decreased RBC polyamine concentrations, which then markedly increased after rats were transferred for a week to normal air, suggesting retardation of growth processes and compensatory stimulation, respectively. However, polyamine system changes in the rat eye were not so pronounced. Enzyme activities and polyamine concentrations tended to be lower in retina after hyperoxia and were only slightly higher, with the exception of ornithine decarboxylase, after a subsequent 1 week of normoxia. In litters subjected to normoxia for longer periods no changes were found. CONCLUSION: The transient and short-lived alteration in polyamine metabolism, especially in the eye, suggests that exposure of newborn rats to high oxygen supplementation followed by normoxia does not necessarily result in marked retinopathy.

Experimental Evaluations of Elementary Science Programs: A Best-Evidence Synthesis

This article presents a systematic review of research on the achievement outcomes of all types of approaches to teaching science in elementary schools. Study inclusion criteria included use of randomized or matched control groups, a study duration of at least 4 weeks, and use of achievement measures independent of the experimental treatment. A total of 23 studies met these criteria. Among studies evaluating inquiry-based teaching approaches, programs that used science kits did not show positive outcomes on science achievement measures (weighted ES=+0.02 in 7 studies), but inquiry-based programs that emphasized professional development but not kits did show positive outcomes (weighted ES=+0.36 in 10 studies). Technological approaches integrating video and computer resources with teaching and cooperative learning showed positive outcomes in a few small, matched studies (ES=+0.42 in 6 studies). The review concludes that science teaching methods focused on enhancing teachers’ classroom instruction throughout the year, such as cooperative learning and science-reading integration, as well as approaches that give teachers technology tools to enhance instruction, have significant potential to improve science learning.

Educational gaming in the health sciences: systematic review

Educational gaming in the health sciences: systematic review. Aim. This paper is a report of a review to investigate the use of games to support classroom learning in the health sciences. Background. One aim of education in the health sciences is to enable learners to develop professional competence. Students have a range of learning styles and innovative teaching strategies assist in creating a dynamic learning environment. New attitudes towards experiential learning methods have contributed to the expansion of gaming as a strategy. Data sources. A search for studies published between January 1980 and June 2008 was undertaken, using appropriate search terms. The databases searched were: British Education Index, British Nursing Index, The Cochrane Library, CINAHLPlus, Medline, PubMed, ERIC, PsychInfo and Australian Education Index. Methods. All publications and theses identified through the search were assessed for relevance. Sixteen papers reporting empirical studies or reviews that involved comparison of gaming with didactic methods were included. Results. The limited research available indicates that, while both traditional didactic methods and gaming have been successful in increasing student knowledge, neither method is clearly more helpful to students. The use of games generally enhances student enjoyment and may improve long-term retention of information. Conclusion. While the use of games can be viewed as a viable teaching strategy, care should be exercised in the use of specific games that have not been assessed objectively. Further research on the use of gaming is needed to enable educators to gaming techniques appropriately for the benefit of students and, ultimately, patients.

Measurement of the Electrogenicity of Bile Salt/H+ Antiport in Escherichia coli

The transmembrane proton gradient (ΔpH) is the primary source of energy exploited by secondary active substrate/H+ antiporters to drive the electroneutral transport of substrates across the Escherichia coli (E. coli) inner membrane. Such electroneutral transport results in no net movement of charges across the membrane. The charge on the transported substrate and the stoichiometry of the exchange reaction, however, can result in an electrogenic reaction which is driven by both the ΔpH and the electrical (∆Ψ) components of the proton electrochemical gradient, resulting in a net movement of electrical charges across the membrane. We have shown that the major facilitator superfamily transporter MdtM - a multidrug efflux protein from E. coli that functions physiologically in protection of bacterial cells against bile salts - imparts bile salt resistance to the bacterial cell by coupling the exchange of external protons (H+) to the efflux of bile salts from the cell interior via an electrogenic antiport reaction (Paul et al., 2014). This protocol describes, using fluorometry, how to detect electrogenic antiport activity of MdtM in inverted membrane vesicles of an antiporter-deficient strain of E. coli TO114 cells by measuring transmembrane ∆Ψ. The method exploits changes that occur in the intensity of the fluorescence signal (quenching and dequenching) of the probe Oxonol V in response to changes in membrane potential due to the MdtM-catalysed sodium cholate/H+ exchange reaction. The protocol can be adapted to detect activity of any secondary active antiporter that couples the electrogenic translocation of H+ across a biological membrane to that of its counter-substrate, and may be used to unmask otherwise camouflaged transport activities and physiological roles.

Measurement of proton-driven antiport in Escherichia coli

Secondary active transport of substrates across the inner membrane is vital to the bacterial cell. Of the secondary active transporter families, the ubiquitous major facilitator superfamily (MFS) is the largest and most functionally diverse (Reddy et al., 2012). Recently, it was reported that the MFS multidrug efflux protein MdtM from Escherichia coli (E. coli) functions physiologically in protection of bacterial cells against bile salts (Paul et al., 2014). The MdtM transporter imparts bile salt resistance to the bacterial cell by coupling the exchange of external protons (H+) to the efflux of bile salts from the cell interior via an antiport reaction. This protocol describes, using fluorometry, how to detect the bile salt/H+ antiport activity of MdtM in inverted membrane vesicles of an antiporter-deficient strain of E. coli TO114 cells by measuring transmembrane ∆pH. This method exploits the changes that occur in the intensity of the fluorescence signal (quenching and dequenching) of the pH-sensitive dye acridine orange in response to changes in [H+] in the vesicular lumen. Due to low levels of endogenous transporter expression that would normally make the contribution of individual transporters such as MdtM to proton-driven antiport difficult to detect, the method typically necessitates that the transporter of interest be overexpressed from a multicopy plasmid. Although the first section of the protocol described here is very specific to the overexpression of MdtM from the pBAD/Myc-His A expression vector, the protocol describing the subsequent measurement of bile salt efflux by MdtM can be readily adapted for measurement of antiport of other substrates by any other antiporter that exchanges protons for countersubstrate.

Characteristics, complications, and gaps in evidence-based interventions in rheumatic heart disease: the Global Rheumatic Heart Disease Registry (the REMEDY study)

AIMS: Rheumatic heart disease (RHD) accounts for over a million premature deaths annually; however, there is little contemporary information on presentation, complications, and treatment.

METHODS AND RESULTS: This prospective registry enrolled 3343 patients (median age 28 years, 66.2% female) presenting with RHD at 25 hospitals in 12 African countries, India, and Yemen between January 2010 and November 2012. The majority (63.9%) had moderate-to-severe multivalvular disease complicated by congestive heart failure (33.4%), pulmonary hypertension (28.8%), atrial fibrillation (AF) (21.8%), stroke (7.1%), infective endocarditis (4%), and major bleeding (2.7%). One-quarter of adults and 5.3% of children had decreased left ventricular (LV) systolic function; 23% of adults and 14.1% of children had dilated LVs. Fifty-five percent (n = 1761) of patients were on secondary antibiotic prophylaxis. Oral anti-coagulants were prescribed in 69.5% (n = 946) of patients with mechanical valves (n = 501), AF (n = 397), and high-risk mitral stenosis in sinus rhythm (n = 48). However, only 28.3% (n = 269) had a therapeutic international normalized ratio. Among 1825 women of childbearing age (12-51 years), only 3.6% (n = 65) were on contraception. The utilization of valvuloplasty and valve surgery was higher in upper-middle compared with lower-income countries.

CONCLUSION: Rheumatic heart disease patients were young, predominantly female, and had high prevalence of major cardiovascular complications. There is suboptimal utilization of secondary antibiotic prophylaxis, oral anti-coagulation, and contraception, and variations in the use of percutaneous and surgical interventions by country income level.