Corrigendum: Crystal structure of the GLP-1 receptor bound to a peptide agonist.

This corrects the article DOI: 10.1038/nature22800.

Crystal structure of the GLP-1 receptor bound to a peptide agonist.

Glucagon-like peptide 1 (GLP-1) regulates glucose homeostasis through the control of insulin release from the pancreas. GLP-1 peptide agonists are efficacious drugs for the treatment of diabetes. To gain insight into the molecular mechanism of action of GLP-1 peptides, here we report the crystal structure of the full-length GLP-1 receptor bound to a truncated peptide agonist. The peptide agonist retains an α-helical conformation as it sits deep within the receptor-binding pocket. The arrangement of the transmembrane helices reveals hallmarks of an active conformation similar to that observed in class A receptors. Guided by this structural information, we design peptide agonists with potent in vivo activity in a mouse model of diabetes.

Intracellular allosteric antagonism of the CCR9 receptor.

Chemokines and their G-protein-coupled receptors play a diverse role in immune defence by controlling the migration, activation and survival of immune cells. They are also involved in viral entry, tumour growth and metastasis and hence are important drug targets in a wide range of diseases. Despite very significant efforts by the pharmaceutical industry to develop drugs, with over 50 small-molecule drugs directed at the family entering clinical development, only two compounds have reached the market: maraviroc (CCR5) for HIV infection and plerixafor (CXCR4) for stem-cell mobilization. The high failure rate may in part be due to limited understanding of the mechanism of action of chemokine antagonists and an inability to optimize compounds in the absence of structural information. CC chemokine receptor type 9 (CCR9) activation by CCL25 plays a key role in leukocyte recruitment to the gut and represents a therapeutic target in inflammatory bowel disease. The selective CCR9 antagonist vercirnon progressed to phase 3 clinical trials in Crohn's disease but efficacy was limited, with the need for very high doses to block receptor activation. Here we report the crystal structure of the CCR9 receptor in complex with vercirnon at 2.8 Å resolution. Remarkably, vercirnon binds to the intracellular side of the receptor, exerting allosteric antagonism and preventing G-protein coupling. This binding site explains the need for relatively lipophilic ligands and describes another example of an allosteric site on G-protein-coupled receptors that can be targeted for drug design, not only at CCR9, but potentially extending to other chemokine receptors.

Effects of Bacopa monnieri (CDRI 08®) in a population of males exhibiting inattention and hyperactivity aged 6 to 14 years: A randomized, double-blind, placebo-controlled trial.

The current study investigated the efficacy of extract of Bacopa monnieri (BM; CDRI 08®) in reducing levels of inattention and hyperactivity in young children. BM has demonstrated improvements in cognitive outcomes in adults, yet little research is available on its effects in younger populations. A 14-week randomized, double-blind, placebo-controlled clinical trial, with placebo run-in and run-out phases, investigated the effects of BM on behavioural, cognitive, mood, and sleep effects in male children aged 6 to 14 years against placebo. One-hundred and twelve participants were recruited into the trial, with 93 datasets available for analysis. No significant behavioural differences were noted between treatment groups. Cognitive outcomes indicated decreased error-making in children taking CDRI 08® (p = .04) and increased speed of reaction time in those taking placebo (p = .04) at study end. Improvements in cognitive flexibility (p = .01), executive functioning (p = .04), interpersonal problems (p = .02), and sleep routine (p = .04) were noted in those consuming CDRI 08® over placebo. CDRI 08® did not improve behavioural outcomes, but may have cognitive, mood and sleep benefits in children aged 6 to 14 years. Further study is required to support the findings presented here.

Structure of class B GPCR corticotropin-releasing factor receptor 1.

Structural analysis of class B G-protein-coupled receptors (GPCRs), cell-surface proteins that respond to peptide hormones, has been restricted to the amino-terminal extracellular domain, thus providing little understanding of the membrane-spanning signal transduction domain. The corticotropin-releasing factor receptor type 1 is a class B receptor which mediates the response to stress and has been considered a drug target for depression and anxiety. Here we report the crystal structure of the transmembrane domain of the human corticotropin-releasing factor receptor type 1 in complex with the small-molecule antagonist CP-376395. The structure provides detailed insight into the architecture of class B receptors. Atomic details of the interactions of the receptor with the non-peptide ligand that binds deep within the receptor are described. This structure provides a model for all class B GPCRs and may aid in the design of new small-molecule drugs for diseases of brain and metabolism.

The Cryo-EM structure of the CorA channel from Methanocaldococcus jannaschii in low magnesium conditions.

CorA channels are responsible for the uptake of essential magnesium ions by bacteria. X-ray crystal structures have been resolved for two full-length CorA channels, each in a non-conducting state with magnesium ions bound to the protein: These structures reveal a homo-pentameric quaternary structure with approximate 5-fold rotational symmetry about a central pore axis. We report the structure of the detergent solubilized Methanocaldococcus jannaschii CorA channel determined by Cryo-Electron Microscopy and Single Particle Averaging, supported by Small Angle X-ray Scattering and X-ray crystallography. This structure also shows a pentameric channel but with a highly asymmetric domain structure. The asymmetry of the domains includes differential separations between the trans-membrane segments, which reflects mechanical coupling of the cytoplasmic domain to the trans-membrane domain. This structure therefore reveals an important aspect of the gating mechanism of CorA channels by providing an indication of how the absence of magnesium ions leads to major structural changes.

A Molecular Pharmacologist's Guide to G Protein-Coupled Receptor Crystallography.

G protein-coupled receptor (GPCR) structural biology has progressed dramatically in the last decade. There are now over 120 GPCR crystal structures deposited in the Protein Data Bank of 32 different receptors from families scattered across the phylogenetic tree, including class B, C, and Frizzled GPCRs. These structures have been obtained in combination with a wide variety of ligands and captured in a range of conformational states. This surge in structural knowledge has enlightened research into the molecular recognition of biologically active molecules, the mechanisms of receptor activation, the dynamics of functional selectivity, and fueled structure-based drug design efforts for GPCRs. Here we summarize the innovations in both protein engineering/molecular biology and crystallography techniques that have led to these advances in GPCR structural biology and discuss how they may influence the resulting structural models. We also provide a brief molecular pharmacologist's guide to GPCR X-ray crystallography, outlining some key aspects in the process of structure determination, with the goal to encourage noncrystallographers to interrogate structures at the molecular level. Finally, we show how chemogenomics approaches can be used to marry the wealth of existing receptor pharmacology data with the expanding repertoire of structures, providing a deeper understanding of the mechanistic details of GPCR function.

A randomized controlled trial investigating the effects of PCSO-524, a patented oil extract of the New Zealand green lipped mussel (Perna canaliculus), on the behaviour, mood, cognition and neurophysiology of children and adolescents (aged 6-14 years) experiencing clinical and sub-clinical levels of hyperactivity and inattention: study protocol ACTRN12610000978066.

BACKGROUND: The prevalence rate of attention-deficit/hyperactivity disorder (ADHD) within Western cultures is between 5% and 12%, and is the most common psychiatric illness among school-aged children, with an estimated 50% of these children retaining ADHD symptoms for the rest of their lives. Children with ADHD have lower blood levels of long-chain Poly Unsaturated Fatty Acids (LC PUFAs) compared with children without ADHD, and following PUFA supplementation, have shown improvements in ADHD-related symptoms. One highly promising marine based LC PUFA preparation is the Omega-3-rich Lyprinol/Omega XL which is a natural formulation containing standardised lipid extract of the New Zealand green lipped mussel (Perna canaliculus) known as PCSO-524® which contains a unique combination of free fatty acids, sterol esters, polar lipids and carotenoids. It is this unique combination of marine lipids that may assist in correcting the decreased levels of LC PUFA levels in children with symptoms of ADHD. The compound is a mixture belonging to a lipid group called sterol esters (SE). The fatty acids in the SE fraction are mainly myristic acid, palmitic acid, palmitoleic acid, stearic acid, oleic acid, linoleic acid, eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA). Lyprinol/Omega XL has previously been shown to contain a potent group of Omega-3 lipids that block the 5 - lipoxygenase metabolic pathway responsible for inflammation in the body. METHODS: A randomized double blind placebo controlled trial will be utilized to assess the effects of 14 weeks administration of Lyprinol/Omega XL versus placebo in 150 children aged 6 to 14 years with high levels of hyperactivity and inattention. Additionally, a range of cognitive, mood and central electrophysiological measures will be undertaken during the 14 week supplementation trial. The primary outcome measure, the Conners' Parent Rating Scales will be completed initially at baseline, then in weeks 4, 8, 10, 14 and then again at 4 weeks post-administration (week 18). The results will contribute to our understanding of the efficacy of marine based Omega-3 s with high anti-inflammatory actions on inattention and hyperactivity in children aged 6 to 14 years.

An acute, double-blind, placebo-controlled crossover study of 320 mg and 640 mg doses of a special extract of Bacopa monnieri (CDRI 08) on sustained cognitive performance.

Standardized extracts of the traditional Ayurvedic medicine Bacopa monnieri (BM) (Brahmi) have been recently shown to have cognitive enhancing effects in chronic administration studies. Pre-clinical work has also identified a number of acute anxiolytic, nootropic, and cardiovascular effects of BM. There has, however, been little research on the acute effects of BM on cognitive function. The current study aimed to assess the acute effects of a specific extract of BM (KeenMind®-CDRI 08) in a double-blind, placebo-controlled study in normal healthy participants who completed a cognitively demanding series of tests. Twenty-four healthy volunteers completed six repetitions of the Cognitive Demand Battery (CDB) after consuming a placebo, 320 mg BM or 640 mg of BM in a cross-over design and provided cardiovascular and mood assessments before and after treatment. Change from baseline scores indicated that the 320 mg dose of BM improved performance at the first, second, and fourth repetition post-dosing on the CDB, and the treatments had no effect upon cardiovascular activity or in attenuating task-induced ratings of stress and fatigue. It was concluded that assessment of an earlier pharmacological window and use of less memory-specific cognitive tests together with more temporally sensitive measures of brain activity may improve our understanding of the acute neurocognitive properties of BM.

Digital technology for addressing cognitive impairment in recent-onset psychosis: A perspective.

Cognitive impairments in psychosis negatively impact functional recovery and quality of life. Existing interventions for improving cognitive impairment in recent-onset psychosis show inconsistent treatment efficacy, small effects, suboptimal engagement and limited generalizability to daily life functioning. In this perspective we explore how digital technology has the potential to address these limitations in order to improve cognitive and functional outcomes in recent-onset psychosis. Computer programs can be used for standardized, automated delivery of cognitive remediation training. Virtual reality provides the opportunity for learning and practicing cognitive skills in real-world scenarios within a virtual environment. Smartphone apps could be used for notification reminders for everyday tasks to compensate for cognitive difficulties. Internet-based technologies can offer psychoeducation and training materials for enhancing cognitive skills. Early findings indicate some forms of digital interventions for cognitive enhancement can be effective, with well-established evidence for human-supported computer-based cognitive remediation in recent-onset psychosis. Emerging evidence regarding virtual reality is favorable for improving social cognition. Overall, blending digital interventions with human support improves engagement and effectiveness. Despite the potential of digital interventions for enhancing cognition in recent-onset psychosis, few studies have been conducted to date. Implementation challenges affecting application of digital technologies for cognitive impairment in recent-onset psychosis are sustained engagement, clinical integration, and lack of quality in the commercial marketplace. Future opportunities lie in including motivational frameworks and behavioral change interventions, increasing service engagement in young people and lived experience involvement in digital intervention development.

Characterization of a CorA Mg2+ transport channel from Methanococcus jannaschii using a Thermofluor-based stability assay.

The Thermofluor assay has been a valuable asset in structural genomics, providing a high-throughput method for assessing the crystallizability of proteins. The technique has been well characterized for soluble proteins but has been less extensively described for membrane proteins. Here we show the successful application of a Thermofluor-based stability assay to an ion channel, CorA from Methanococcus jannaschii. Optimization of the concentration of free detergent within the assay was important, as excessive concentrations mask the fluorescence change associated with thermal unfolding of the protein. CorA was shown to be stabilized by low pH, but relatively insensitive to salt concentration. Divalent metal cations were also capable of stabilizing the protein, in the order Co2+>Ni2+>Mn2+>Mg2+>Ca2+. Finally, removal of the oligohistidine tag was also shown to improve the thermal stability of CorA. Conclusions are drawn from this detailed study about the general applicability of this technique to other membrane proteins.

Systematic Overview of Bacopa monnieri (L.) Wettst. Dominant Poly-Herbal Formulas in Children and Adolescents.

Background: The Ayurvedic medicinal system employs a holistic approach to health, utilising the synergistic properties of organic resources. Research into the Ayurvedic herb Bacopa monnieri (L.) Wettst. (B.monnieri) has reported improvements in cognitive outcomes in child and adult populations. The aim of current review is to systematically assess and critically summarize clinical trials investigating B.monnieri-dominant poly-herbal formulas and their effects on the cognition, memory, learning, and behaviour in children and adolescents. Methods: Key word searches were performed using PubMed, Scopus, Cochrane Library, DHARA, and CINAHL for publications meeting inclusion criteria up to November 2017. There were no restrictions in study design. Effect sizes were calculated for all significant findings to allow for direct comparisons, and each study was evaluated on design quality. Cognitive and behavioural outcomes were grouped into validated constructs for cross-study comparison. Results: Nine trials met inclusion criteria. Five studies reported sufficient data for effect size analysis with most improvements reported in behavioural outcomes. True cognitive abilities and behavioural constructs were reviewed in six studies, with visual perception, impulsivity, and attention demonstrating the greatest improvements. The veracity of the evidence for the formulations reviewed is weakened by inconsistent statistical design and under-reporting of safety and tolerability data (44%). Conclusions: The current review extends research supporting B.monnieri as a cognitive enhancer and provides modest evidence for the use of B.monnieri in poly-herbal preparations for improving cognitive and behavioural outcomes in child and adolescent populations. Greater emphasis on statistical vigour and the reporting of tolerability data are essential for future trials to adequately document poly-herbal treatment efficacy.

A Randomized, Double-Blind Study Assessing Changes in Cognitive Function in Indian School Children Receiving a Combination of Bacopa monnieri and Micronutrient Supplementation vs. Placebo.

Several studies have indicated a chronic cognitive enhancing effect of Bacopa monnieri across different ages and cognitive impairment associated with vitamin and mineral deficiencies in children. Therefore, we investigated the effects of 4-month supplementation with a combination of B. monnieri extract and multiple micronutrients on cognitive functions in Indian school children aged 7-12 years. This was a randomized, double-blind, parallel design, single-center study in which 300 children were randomized to receive a beverage either fortified with B. monnieri and multiple micronutrients ("fortified") or a non-fortified isocaloric equivalent ("control") twice-daily for 4 months. Cognitive function was assessed by the Cambridge Neuropsychological Automated Test Battery (CANTAB) administered at baseline, Day 60 and Day 121. The primary endpoint was change in short-term memory (working memory) from baseline in subjects receiving "fortified" vs. "control" beverages after 4 months. Secondary endpoints included sustained attention, episodic memory, and executive function. The "fortified" beverage did not significantly improve short-term memory or any of the secondary outcomes tested relative to the "control" beverage. However, the spatial working memory "strategy" score showed significant improvement on Day 60 (difference between groups in change from baseline: -0.55; p < 0.05), but not on Day 121 due to the active intervention. Study products were well-tolerated. Reasons for these unexpected findings are discussed.

Reduced inattention and hyperactivity and improved cognition after marine oil extract (PCSO-524®) supplementation in children and adolescents with clinical and subclinical symptoms of attention-deficit hyperactivity disorder (ADHD): a randomised, double-blind, placebo-controlled trial.

INTRODUCTION: This study investigated the effects of a marine oil extract (PCSO-524®) on inattention, hyperactivity, mood and cognition in children and adolescents. PCSO-524® is a standardised lipid extract of the New Zealand green-lipped mussel and is an inflammatory modulator that inhibits the 5'-lipoxygenase and cyclooxygenase pathways and decreases concentrations of the pro-inflammatory arachidonic acid (AA). METHODS: PCSO-524® or a matched placebo was administered for 14 weeks to 144 participants (123 males/21 females; mean age 8.7 years) with high hyperactivity and inattention in a randomised, double-blind, placebo-controlled study. The primary outcome was the Conners Parent Rating Scale assessing parental reports of behavioural problems. Secondary outcomes assessed changes in cognition and mood. RESULTS: The results of the present study did not support the hypothesis that PCSO-524® improves parental reports of hyperactivity, inattention and impulsivity in children ages 6 to 14 years over placebo. Repeated measures ANOVA on post hoc subsample analysis indicated significant improvements in hyperactivity (p = 0.04), attention (p = 0.02), learning (p = 0.05) and probability of ADHD (p = 0.04) with a medium to large average effect size (d = 0.65) in those children who did not meet criteria for combined hyperactivity and inattention. Furthermore, significant improvements in the PCSO-524® group were indicated in a whole sample repeated measures ANCOVA on recognition memory between baseline and week 8 over placebo (p = 0.02, d = 0.56); this difference was not sustained at week 14. CONCLUSIONS: The results presented indicate that PCSO-524® may be beneficial in reducing levels of hyperactivity and inattention in a population of children with clinical and subclinical symptoms of ADHD.

A systematic review of the Ayurvedic medicinal herb Bacopa monnieri in child and adolescent populations.

OBJECTIVES: Clinicians utilise critical research to advance their knowledge when prescribing standard and alternative therapies for developmental disorders. Recent research has reported that the traditional Ayurvedic medicine Bacopa monnieri may improve cognitive outcomes in adult populations; however, few studies have investigated its benefits in younger cohorts. The aim of the current review is to systematically assess and critically summarize clinical trial outcomes and safety of Bacopa and its effects on the cognition and behaviour in children and adolescents. METHOD: PubMed, Scopus, Cochrane Library, Google and CINAHL were searched up to August 2015 for trials investigating Bacopa monnieri in child and adolescent populations. There were no restrictions in study design. Cognitive and behavioural outcomes were grouped into validated constructs and effect sizes were calculated for all significant data to allow for direct comparisons. RESULTS: Five studies met inclusion criteria for this review. The results demonstrated significant consistent improvements in the language behaviour cognitive domain and in a number of the memory sub-domains. Significant improvements were also seen in hyperactivity and attention-deficit domains. Overall outcome data demonstrated small to medium effect sizes (mean d=0.42). Safety and tolerability data was well reported for 80% of studies with only 2.3% of all participants reporting mild side-effects. CONCLUSION: This review highlights the safe use of Bacopa monnieri in child and adolescent populations for improving elements of cognition as well as behaviour and attention-deficit domains. However, there is a significant need for replicated study designs and stringent statistical analysis to validate these outcomes.

Conformational thermostabilisation of corticotropin releasing factor receptor 1.

Recent technical advances have greatly facilitated G-protein coupled receptors crystallography as evidenced by the number of successful x-ray structures that have been reported recently. These technical advances include novel detergents, specialised crystallography techniques as well as protein engineering solutions such as fusions and conformational thermostabilisation. Using conformational thermostabilisation, it is possible to generate variants of GPCRs that exhibit significantly increased stability in detergent micelles whilst preferentially occupying a single conformation. In this paper we describe for the first time the application of this technique to a member of a class B GPCR, the corticotropin releasing factor receptor 1 (CRF1R). Mutational screening in the presence of the inverse agonist, CP-376395, resulted in the identification of a construct with twelve point mutations that exhibited significantly increased thermal stability in a range of detergents. We further describe the subsequent construct engineering steps that eventually yielded a crystallisation-ready construct which recently led to the solution of the first x-ray structure of a class B receptor. Finally, we have used molecular dynamic simulation to provide structural insight into CRF1R instability as well as the stabilising effects of the mutants, which may be extended to other class B receptors considering the high degree of structural conservation.

A Randomized Controlled Trial Investigating the Effects of a Special Extract of Bacopa monnieri (CDRI 08) on Hyperactivity and Inattention in Male Children and Adolescents: BACHI Study Protocol (ANZCTRN12612000827831).

UNLABELLED: Clinical diagnoses of Attention Deficit Hyperactivity Disorder (ADHD) and the use of prescription medications for its treatment have increased in recent years. Current treatments may involve the administration of amphetamine-type substances, a treatment path many parents are apprehensive to take. Therefore, alternative pharmacological treatments are required. Few nutritional or pharmacological alternatives that reduce ADHD associated symptoms (hyperactivity and inattention) have been subjected to rigorous clinical trials. Bacopa monnieri is a perennial creeping herb. CDRI 08 is a special extract of Bacopa monnieri which has been subjected to hundreds of scientific studies and has been shown in human randomized controlled trials (RCTs) to improve memory, attention, and mood. It is hypothesised that chronic administration of CDRI 08 will improve attention, concentration and behaviour in children with high levels of hyperactivity and/or inattention. This paper reports the protocol for the first 16-week, randomized, placebo-controlled, double-blind, parallel groups trial examining the efficacy and safety of CDRI 08 in male children aged 6-14 years with high levels of inattention and hyperactivity. The primary outcome variable will be the level of hyperactivity and inattention measured by the Conners' Parent Rating Scale (CPRS). Secondary outcome variables include cognition, mood, sleep, and EEG. TRIAL REGISTRATION: Australia and New Zealand Clinical Trials Register (ANZCTR): ACTRN12612000827831.

The cognitive-enhancing effects of Bacopa monnieri: a systematic review of randomized, controlled human clinical trials.

OBJECTIVES: Traditional knowledge suggests that Bacopa monnieri enhances cognitive performance. Such traditional beliefs have now been scientifically tested through a handful of randomized, controlled human clinical trials. The current systematic review aimed to examine the scientific evidence as to whether Bacopa can enhance cognitive performance in humans. DESIGN: A systematic review of randomized controlled trials is presented. Multiple databases were systematically searched by multiple authors. Relevant trials were objectively assessed for methodological quality. SUBJECTS: The subjects studied were adult humans without dementia or significant cognitive impairment. INTERVENTIONS: B. monnieri, including Bacopa extracts, were administered over long-term supplementation periods. OUTCOME MEASURES: Any validated cognitive test, whether a primary or secondary outcome. RESULTS: Six (6) studies met the final inclusion criteria and were included in review. Trials were all conducted over 12 weeks. Across trials, three different Bacopa extracts were used at dosages of 300-450 mg extract per day. All reviewed trials examined the effects of Bacopa on memory, while other cognitive domains were less well studied. There were no cognitive tests in the areas of auditory perceptual abilities or idea production and only a paucity of research in the domains of reasoning, number facility, and language behavior. Across studies, Bacopa improved performance on 9 of 17 tests in the domain of memory free recall. There was little evidence of enhancement in any other cognitive domains. CONCLUSIONS: There is some evidence to suggest that Bacopa improves memory free recall with evidence for enhancement in other cognitive abilities currently lacking perhaps due to inconsistent measures employed by studies across these cognitive domains. Research into the nootropic effects of Bacopa is in its infancy, with research still yet to investigate the effects of Bacopa across all human cognitive abilities. Similarly, future research should examine the nootropic effects of Bacopa at varied dosages and across different extracts.

Complementary medicines (herbal and nutritional products) in the treatment of Attention Deficit Hyperactivity Disorder (ADHD): a systematic review of the evidence.

OVERVIEW: Complementary and Alternative Medicines (CAMs) are frequently given to children and adolescents for reputed benefits in the treatment of hyperkinetic and concentration disorders such as Attention Deficit Hyperactivity Disorder (ADHD). In such vulnerable populations high quality evidence is required to support such claims. AIMS: The aim of the paper is to assess the current evidence of herbal and nutritional interventions for ADHD using a systematic search of clinical trials meeting an acceptable standard of evidence. METHODS: PubMed, PsycINFO, Cochrane Library and CINAHL were searched up to May 26th, 2011 for randomised, controlled clinical trials using CAM products as interventions to treat ADHD. A quality analysis using a purpose-designed scale, and an estimation of effect sizes (Cohen's d) where data were available, were also calculated. RESULTS: The review revealed that 16 studies met inclusion criteria, with predominant evidentiary support found for zinc, iron, Pinus marinus (French maritime pine bark), and a Chinese herbal formula (Ningdong); and mixed (mainly inconclusive) evidence for omega-3, and l-acetyl carnitine. Current data suggest that Ginkgo biloba (ginkgo), and Hypercium perforatum (St. John's wort) are ineffective in treating ADHD. CONCLUSION: The research suggests only some CAMs may be beneficial in ADHD, thus clinicians need to be aware of the current evidence. Promising candidates for future research include Bacopa monniera (brahmi) and Piper methysticum (kava), providing potential efficacy in improving attentional and hyperkinetic disorders via a combination of cognitive enhancing and sedative effects.