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The type 5 metabotropic glutamate receptor, mGlu 5 , has been proposed as a potential therapeutic target for the treatment of several neurodegenerative diseases. In preclinical neurodegenerative disease models, novel allosteric modulators have been shown to improve cognitive performance and reduce disease-related pathology. A common pathological hallmark of neurodegenerative diseases is a chronic neuroinflammatory response, involving glial cells such as astrocytes and microglia. Since mGlu 5 is expressed in astrocytes, targeting this receptor could provide a potential mechanism by which neuroinflammatory processes in neurodegenerative disease may be modulated. This review will discuss current evidence that highlights the potential of mGlu 5 allosteric modulators to treat neurodegenerative diseases, including Alzheimer’s disease, Huntington’s disease, Parkinson’s disease, and amyotrophic lateral sclerosis. Furthermore, this review will explore the role of mGlu 5 in neuroinflammatory responses, and the potential for this G protein-coupled receptor to modulate neuroinflammation.

Adenosine A 2A receptor structure Adenosine receptors are G protein-coupled receptors that are found in the heart and the brain, and adenosine is the endogenous ligand for this class of transmembrane receptor. Lebon et al . present two X-ray crystal structures of a thermostabilized human adenosine A 2A receptor bound to its endogenous agonist adenosine and the synthetic agonist NECA. Comparison of the agonist-bound structures of A 2A receptor with the agonist-bound structures of β-adrenoceptors suggests that the contraction of the ligand binding pocket caused by the inward motion of several helices may be a common feature in the activation of all G protein-coupled receptors. Adenosine receptors and β-adrenoceptors are G-protein-coupled receptors (GPCRs) that activate intracellular G proteins on binding the agonists adenosine 1 or noradrenaline 2 , respectively. GPCRs have similar structures consisting of seven transmembrane helices that contain well-conserved sequence motifs, indicating that they are probably activated by a common mechanism 3 , 4 . Recent structures of β-adrenoceptors highlight residues in transmembrane region 5 that initially bind specifically to agonists rather than to antagonists, indicating that these residues have an important role in agonist-induced activation of receptors 5 , 6 , 7 . Here we present two crystal structures of the thermostabilized human adenosine A 2A receptor (A 2A R-GL31) bound to its endogenous agonist adenosine and the synthetic agonist NECA. The structures represent an intermediate conformation between the inactive and active states, because they share all the features of GPCRs that are thought to be in a fully activated state, except that the cytoplasmic end of transmembrane helix 6 partially occludes the G-protein-binding site. The adenine substituent of the agonists binds in a similar fashion to the chemically related region of the inverse agonist ZM241385 (ref. 8 ). Both agonists contain a ribose group, not found in ZM241385, which extends deep into the ligand-binding pocket where it makes polar interactions with conserved residues in H7 (Ser 277 7.42 and His 278 7.43 ; superscripts refer to Ballesteros–Weinstein numbering 9 ) and non-polar interactions with residues in H3. In contrast, the inverse agonist ZM241385 does not interact with any of these residues and comparison with the agonist-bound structures indicates that ZM241385 sterically prevents the conformational change in H5 and therefore it acts as an inverse agonist. Comparison of the agonist-bound structures of A 2A R with the agonist-bound structures of β-adrenoceptors indicates that the contraction of the ligand-binding pocket caused by the inward motion of helices 3, 5 and 7 may be a common feature in the activation of all GPCRs.

Metabotropic glutamate receptors are class C G-protein-coupled receptors which respond to the neurotransmitter glutamate. Structural studies have been restricted to the amino-terminal extracellular domain, providing little understanding of the membrane-spanning signal transduction domain. Metabotropic glutamate receptor 5 is of considerable interest as a drug target in the treatment of fragile X syndrome, autism, depression, anxiety, addiction and movement disorders. Here we report the crystal structure of the transmembrane domain of the human receptor in complex with the negative allosteric modulator, mavoglurant. The structure provides detailed insight into the architecture of the transmembrane domain of class C receptors including the precise location of the allosteric binding site within the transmembrane domain and key micro-switches which regulate receptor signalling. This structure also provides a model for all class C G-protein-coupled receptors and may aid in the design of new small-molecule drugs for the treatment of brain disorders. An X-ray structure is presented for metabotropic glutamate receptor 5, a class C G-protein-coupled glutamate receptor linked to fragile X syndrome and neurological disorders; this study provides insights into the protein’s mechanism of action. Structure of a class C GPCR This manuscript presents the X-ray crystal structure of the transmembrane portion of the metabotropic glutamate receptor 5 (mGlu5), a class C G-protein-coupled receptor (GPCR), in the presence of mavoglurant, a negative allosteric modulator. Negative allosteric modulators of mGlu5 are in clinical trials for the treatment of fragile X syndrome, depression, anxiety, migraine and dyskinesias; positive allosteric modulators of mGlu5 might be useful for the treatment of schizophrenia and cognitive disorders. In this structure, mavoglurant is found in a pocket that is about 8 Å from the extracellular receptor surface.

The crystal structure of the CCR9 chemokine receptor in complex with vercirnon at 2.8 Å resolution. Small-molecule chemokine receptor antagonists Chemokine receptors are a family of G-protein-coupled receptors that regulate the migration of immune cells; their function has been implicated in a range of diseases. Two groups reporting in this issue of Nature describe crystal structures of two different chemokine receptors bound to small-molecule inhibitors. Tracy Handel and colleagues describe the structure of CCR2—a promising drug target for autoimmune, inflammatory and metabolic diseases as well as cancer—bound to orthosteric (BMS-681) and allosteric (CCR2-RA-[ R ]) antagonists. Fiona Marshall and colleagues describe the structure of CCR9—involved in immune cell recruitment to the gut and a promising drug target in inflammatory bowel disease—in complex with the selective CCR9 antagonist vercirnon. Both CCR2 and CCR9 structures reveal an allosteric pocket on the cytoplasmic face of the receptor. This allosteric pocket appears to be highly druggable, and homologous pockets may be present on other chemokine receptors. Chemokines and their G-protein-coupled receptors play a diverse role in immune defence by controlling the migration, activation and survival of immune cells 1 . They are also involved in viral entry, tumour growth and metastasis and hence are important drug targets in a wide range of diseases 2 , 3 . 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 4 . 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 5 . 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 6 . 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 6 . 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 7 that can be targeted for drug design, not only at CCR9, but potentially extending to other chemokine receptors.

Adenosine receptors and β-adrenoceptors are G-protein-coupled receptors (GPCRs) that activate intracellular G proteins on binding the agonists adenosine or noradrenaline, respectively. GPCRs have similar structures consisting of seven transmembrane helices that contain well-conserved sequence motifs, indicating that they are probably activated by a common mechanism. Recent structures of β-adrenoceptors highlight residues in transmembrane region 5 that initially bind specifically to agonists rather than to antagonists, indicating that these residues have an important role in agonist-induced activation of receptors. Here we present two crystal structures of the thermostabilized human adenosine A^sub 2A^ receptor (A^sub 2A^R-GL31) bound to its endogenous agonist adenosine and the synthetic agonist NECA. The structures represent an intermediate conformation between the inactive and active states, because they share all the features of GPCRs that are thought to be in a fully activated state, except that the cytoplasmic end of transmembrane helix 6 partially occludes the G-protein-binding site. The adenine substituent of the agonists binds in a similar fashion to the chemically related region of the inverse agonist ZM241385 (ref. 8). Both agonists contain a ribose group, not found in ZM241385, which extends deep into the ligand-binding pocket where it makes polar interactions with conserved residues in H7 (Ser 277^sup 7.42^ and His 278^sup 7.43^; superscripts refer to Ballesteros-Weinstein numbering) and non-polar interactions with residues in H3. In contrast, the inverse agonist ZM241385 does not interact with any of these residues and comparison with the agonist-bound structures indicates that ZM241385 sterically prevents the conformational change in H5 and therefore it acts as an inverse agonist. Comparison of the agonist-bound structures of A^sub 2A^R with the agonist-bound structures of β-adrenoceptors indicates that the contraction of the ligand-binding pocket caused by the inward motion of helices 3, 5 and 7 may be a common feature in the activation of all GPCRs. [PUBLICATION ABSTRACT]

Adenosine A.sub.2A receptor structure Adenosine receptors are G protein-coupled receptors that are found in the heart and the brain, and adenosine is the endogenous ligand for this class of transmembrane receptor. Lebon et al. present two X-ray crystal structures of a thermostabilized human adenosine A.sub.2A receptor bound to its endogenous agonist adenosine and the synthetic agonist NECA. Comparison of the agonist-bound structures of A.sub.2A receptor with the agonist-bound structures of [beta]-adrenoceptors suggests that the contraction of the ligand binding pocket caused by the inward motion of several helices may be a common feature in the activation of all G protein-coupled receptors. Adenosine receptors and [beta]-adrenoceptors are G-protein-coupled receptors (GPCRs) that activate intracellular G proteins on binding the agonists adenosine.sup.1 or noradrenaline.sup.2, respectively. GPCRs have similar structures consisting of seven transmembrane helices that contain well-conserved sequence motifs, indicating that they are probably activated by a common mechanism.sup.3,4. Recent structures of [beta]-adrenoceptors highlight residues in transmembrane region 5 that initially bind specifically to agonists rather than to antagonists, indicating that these residues have an important role in agonist-induced activation of receptors.sup.5,6,7. Here we present two crystal structures of the thermostabilized human adenosine A.sub.2A receptor (A.sub.2AR-GL31) bound to its endogenous agonist adenosine and the synthetic agonist NECA. The structures represent an intermediate conformation between the inactive and active states, because they share all the features of GPCRs that are thought to be in a fully activated state, except that the cytoplasmic end of transmembrane helix 6 partially occludes the G-protein-binding site. The adenine substituent of the agonists binds in a similar fashion to the chemically related region of the inverse agonist ZM241385 (ref. 8). Both agonists contain a ribose group, not found in ZM241385, which extends deep into the ligand-binding pocket where it makes polar interactions with conserved residues in H7 (Ser 277.sup.7.42 and His 278.sup.7.43; superscripts refer to Ballesteros-Weinstein numbering.sup.9) and non-polar interactions with residues in H3. In contrast, the inverse agonist ZM241385 does not interact with any of these residues and comparison with the agonist-bound structures indicates that ZM241385 sterically prevents the conformational change in H5 and therefore it acts as an inverse agonist. Comparison of the agonist-bound structures of A.sub.2AR with the agonist-bound structures of [beta]-adrenoceptors indicates that the contraction of the ligand-binding pocket caused by the inward motion of helices 3, 5 and 7 may be a common feature in the activation of all GPCRs.

The ghrelin receptor (GRLN-R) was cloned in 1996 after the discovery that a series of synthetic growth hormone-releasing compounds (the growth hormone secretagogues) acted through a receptor distinct from the growth hormone-releasing hormone receptor. In 1999 the endogenous ligand of the receptor, ghrelin, was discovered. As well as stimulating growth hormone release, ghrelin has been shown to be involved in many other processes such as appetite stimulation and the regulation of energy homeostasis, making the ghrelin/GRLN-R system an attractive pharmaceutical target for the treatment of disorders such as growth hormone deficiency, cachexia and obesity. The GRLN-R displays a high level of ligand-independent (constitutive) activity and has been suggested to couple to Gaq/11, Gai/o, Gas and Ga12/13 G protein pathways, although little is known about the signalling of ghrelin and the growth hormone secretagogues in all but the Gaq/11 pathway. Two of the growth hormone secretagogues, GHRP-6 and L-692,429, have been described as ‘ago-allosteric modulators’ of the GRLN-R as, when co-administered with ghrelin, GHRP-6 and L-692,429 were reported to act both as co-agonists (increasing the efficacy of the ghrelin response) and as negative or positive (respectively) regulators of the potency of ghrelin. This study sought to investigate the pharmacology of the GRLN-R through the Gai/o pathway. [35S]GTPyS binding assays were used to measure activation of the Gai/o pathway, demonstrating that GHRP-6, L-692,585 (a commercially available analogue of L-692,429) and a third growth hormone secretagogues, MK-677, acted with higher efficacy than ghrelin. At least in the system tested, upon co-administration with ghrelin each of the growth hormone secretagogues acted in a simple competitive fashion with ghrelin. Radioligand binding experiments showed that the dissociation kinetics of [His[125I]]- ghrelin from the GRLN-R were not altered by co-administration of the growth hormone secretagogues. Fitting data to a modified operational model of allosterism demonstrated that GHRP-6, L-692,585 and MK-677 were not ago-allosteric modulators of the GRLN-R but simple orthosteric agonists. In order to further examine the receptor-specific effects of ghrelin and the growth hormone secretagogues, a Flp-In™ T-REx™ HEK293 cell line expressing the GRLN-R was constructed. [35S]GTPyS binding assays confirmed that the GRLN-R was constitutively active through both the Gaq/11 and Gai/o pathways, demonstrated by an increase in [35S]GTPyS loading upon receptor expression which could be reduced by the administration of the GRLN-R inverse agonist SPA. Upon expression of the GRLN-R a considerable level of cell detachment was observed with the remaining cells appearing rounded compared to parental HEK293 cells, an affect that appeared to be mediated by the constitutive activity of the receptor. In contrast to the [35S]GTPyS assays used to measure activation of Gai/o, [35S]GTPyS assays with a Gaq-immunoprecipitation step demonstrated that the growth hormone secretagogues acted with equal efficacies to that of ghrelin, demonstrating functional selectivity at the GRLN-R. Intact cell assays were also used to measure Gaq/11 and Gai/o responses, however, a Gai/o-mediated response could not be measured in cAMP accumulation assays, suggesting that the GRLN-R could signal via activation of Gao but not Gai1-3. Although the activation of the Gas pathway by the GRLN-R remains controversial, in this study ghrelin and the growth secretagogues could evoke a Gas-mediated cAMP response (although L-692,585 acted with a lower efficacy than ghrelin). Finally, two naturally occurring missense mutations of the GRLN-R (A204E in the second extracellular loop and I134T in the third transmembrane helix) were analysed to investigate whether these mutations led to retention of the receptor within the endoplasmic reticulum and to investigate whether the mutations affected the ability of the GRLN-R to signal to the growth hormone secretagogues. The A204E mutation caused partial retention of the GRLN-R within the endoplasmic reticulum, whilst receptor that was transported to the plasma membrane did not display any measurable constitutive activity. In contrast, the I134T mutation did not alter receptor localisation, nor did it have any effect on the constitutive or ligand-induced activation of the GRLN-R, however, it appeared to lower the efficacy of the inverse agonist SPA.

Patient-reported outcomes (PROs) are used in clinical trials to provide valuable evidence on the impact of disease and treatment on patients’ symptoms, function and quality of life. High-quality PRO data from trials can inform shared decision-making, regulatory and economic analyses and health policy. Recent evidence suggests the PRO content of past trial protocols was often incomplete or unclear, leading to research waste. To address this issue, international, consensus-based, PRO-specific guidelines were developed: the Standard Protocol Items: Recommendations for Interventional Trials (SPIRIT)-PRO Extension. The SPIRIT-PRO Extension is a 16-item checklist which aims to improve the content and quality of aspects of clinical trial protocols relating to PRO data collection to minimise research waste, and ultimately better inform patient-centred care. This SPIRIT-PRO explanation and elaboration (E&E) paper provides information to promote understanding and facilitate uptake of the recommended checklist items, including a comprehensive protocol template. For each SPIRIT-PRO item, we provide a detailed description, one or more examples from existing trial protocols and supporting empirical evidence of the item’s importance. We recommend this paper and protocol template be used alongside the SPIRIT 2013 and SPIRIT-PRO Extension paper to optimise the transparent development and review of trial protocols with PROs.