We found that 3-cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide (CDPPB) is a potent and selective positive allosteric modulator of the metabotropic glutamate receptor subtype 5 (mGluR5). In Chinese hamster ovary cells expressing human mGluR5, CDPPB potentiated threshold responses to glutamate in fluorometric Ca 2ϩ assays more than 7-fold with an EC 50 value of approximately 27 nM. At 1 M, CDPPB shifted mGluR5 agonist concentration response curves to glutamate, quisqualate, and (R,S)-3,5-dihydroxyphenylglycine 3-to 9-fold to the left. At higher concentrations, CDPPB exhibited agonist-like activity on cells expressing mGluR5. No other activity was observed on any other mGluR or cell type at concentrations up to 10 M. CDPPB had no effect on [ 3 H]quisqualate binding to mGluR5 but did compete for binding of [ 3 H]methoxyPEPy, an analog of the selective mGluR5 negative allosteric modulator MPEP. CDPPB was found to be brain penetrant and reversed amphetamine-induced locomotor activity and amphetamineinduced deficits in prepulse inhibition in rats, two models sensitive to antipsychotic drug treatment. These results demonstrate that positive allosteric modulation of mGluR5 produces behavioral effects, suggesting that such modulation serves as a viable approach to increasing mGluR5 activity in vivo. These effects are consistent with the hypothesis that allosteric potentiation of mGluR5 may provide a novel approach for development of antipsychotic agents.Glutamate, the predominant excitatory neurotransmitter in the mammalian central nervous system (CNS), exerts its effects through two classes of receptors. The first class of receptor, ionotropic glutamate receptors, are postsynaptic, multimeric ligand-gated ion channels classified into three groups named for group-selective agonists: N-methyl-D-aspartate (NMDA), ␣-amino-3-hydroxy-5-methyl-4-isoxazopropionic acid, and kainate receptors. The NMDA receptor (NMDAR) is known to play an important role in processes related to schizophrenia. NMDAR antagonists, such as phencyclidine and ketamine, induce positive, negative, and cognitive symptoms reminiscent of schizophrenia in human volunteers and worsen existing symptoms in schizophrenic patients. This observation has led to the hypothesis that changes in CNS circuits induced by NMDAR hypofunction may play a key role in the development and/or in the underlying symptoms of schizophrenia (Olney et al., 1999). Therefore, development of compounds that selectively increase NMDAR function could be used to test this hypothesis.The metabotropic glutamate receptors (mGluRs), the second class of glutamate receptor, are members of family C of the G-protein-coupled receptors and are characterized by a large extracellular agonist binding domain on the aminoterminal end of the receptor that is distinct from the seventransmembrane domain characteristic of all G-protein-couArticle, publication date, and citation information can be found at

Despite increased understanding of the biological basis for sleep control in the brain, few novel mechanisms for the treatment of insomnia have been identified in recent years. One notable exception is inhibition of the excitatory neuropeptides orexins A and B by design of orexin receptor antagonists. Herein, we describe how efforts to understand the origin of poor oral pharmacokinetics in a leading HTS-derived diazepane orexin receptor antagonist led to the identification of compound 10 with a 7-methyl substitution on the diazepane core. Though 10 displayed good potency, improved pharmacokinetics, and excellent in vivo efficacy, it formed reactive metabolites in microsomal incubations. A mechanistic hypothesis coupled with an in vitro assay to assess bioactivation led to replacement of the fluoroquinazoline ring of 10 with a chlorobenzoxazole to provide 3 (MK-4305), a potent dual orexin receptor antagonist that is currently being tested in phase III clinical trials for the treatment of primary insomnia.

We found that N- {4-chloro-2-[(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)methyl]phenyl}-2-hydroxybenzamide (CPPHA), is a potent and selective positive allosteric modulator of the metabotropic glutamate receptor subtype 5 (mGluR5). CPPHA alone had no agonist activity and acted as a selective positive allosteric modulator of human and rat mGluR5. CPPHA potentiated threshold responses to glutamate in fluorometric Ca 2ϩ assays 7-to 8-fold with EC 50 values in the 400 to 800 nM range, and at 10 M shifted mGluR5 agonist concentration-response curves to glutamate, quisqualate, and (R,S)-3,5-dihydroxyphenylglycine (DHPG) 4-to 7-fold to the left. The only effect of CPPHA on other mGluRs was weak inhibition of mGluR4 and 8. Neither CPPHA nor the previously described 3,3Ј-difluorobenzaldazine (DFB) affected [ 3 H]quisqualate binding to mGluR5, but although DFB partially competed for [ 3 H]3-methoxy-5-(2-pyridinylethynyl)pyridine binding, CPPHA had no effect on the binding of this 2-methyl-6-(phenylethynyl)-pyridine analog to mGluR5. Although the binding sites for the two classes of allosteric modulators seem to be different, these different allosteric sites can modulate functionally and mechanistically similar allosteric effects. In electrophysiological studies of brain slice preparations, it had been previously shown that activation of mGluR5 receptors by agonists increased N-methyl-D-aspartate (NMDA) receptor currents in the CA1 region of hippocampal slices. We found that CPPHA (10 M) potentiated NMDA receptor currents in hippocampal slices induced by threshold levels of DHPG, whereas having no effect on these currents by itself. Similarly, 10 M CPPHA also potentiated mGluR5-mediated DHPG-induced depolarization of rat subthalamic nucleus neurons. These results demonstrate that allosteric potentiation of mGluR5 increases the effect of threshold agonist concentrations in native systems.Metabotropic glutamate receptors (mGluRs) are G proteincoupled receptors (GPCRs) that bind glutamate to modulate neurotransmitter release or postsynaptic excitatory neurotransmission, and hence they modulate the strength of synaptic transmission. The mGluRs are members of GPCR family C and possess a large extracellular agonist binding domain in the amino-terminal portion of the receptor. This agonist binding domain distinguishes family C from the other GPCR families in which the agonist binding sites are associated with the seven-strand transmembrane spanning region or with the extracellular loops that connect the strands of this region. Thus, in the mGluRs, interaction of the agonist with the transmembrane domains is thought to be indirect (O'Hara et al., 1993; for reviews, see Conn and Pin, 1997;Bockaert and Pin, 1999).Article, publication date, and citation information can be found at

We have identified a family of highly selective allosteric modulators of the group I metabotropic glutamate receptor subtype 5 (mGluR5). This family of closely related analogs exerts a spectrum of effects, ranging from positive to negative allosteric modulation, and includes compounds that do not themselves modulate mGluR5 agonist activity but rather prevent other family members from exerting their modulatory effects. 3,3Ј-Difluorobenzaldazine (DFB) has no agonist activity, but it acts as a selective positive allosteric modulator of human and rat mGluR5. DFB potentiates threshold responses to glutamate, quisqualate, and 3,5-dihydroxyphenylglycine in fluorometric Ca 2ϩ assays 3-to 6-fold, with EC 50 values in the 2 to 5 M range, and at 10 to 100 M, it shifts mGluR5 agonist concentration-response curves approximately 2-fold to the left. The analog 3,3Ј-dimethoxybenzaldazine (DMeOB) acts as a negative modulator of mGluR5 agonist activity, with an IC 50 of 3 M in fluorometric Ca 2ϩ assays, whereas the analog 3,3Ј-dichlorobenzaldazine (DCB) does not exert any apparent modulatory effect on mGluR5 activity. However, DCB seems to act as an allosteric ligand with neutral cooperativity, preventing the positive allosteric modulation of mGluRs by DFB as well as the negative modulatory effect of DMeOB. None of these analogs affects binding of [ 3 H]quisqualate to the orthosteric (glutamate) site, but they do inhibit [ 3 H]3-methoxy-5-(2-pyridinylethynyl)pyridine binding to the site for 2-methyl-6-(phenylethynyl)-pyridine, a previously identified negative allosteric modulator. With the use of these compounds, we provide evidence that allosteric sites on GPCRs can respond to closely related ligands with a range of pharmacological activities from positive to negative modulation as well as to neutral competition of this modulation.Metabotropic glutamate receptors (mGluRs) are G proteincoupled receptors that bind glutamate to exert a modulatory influence on neuronal excitability and synaptic transmission in the central nervous system. The eight known members of the mGluR subfamily have been divided into three groups on the basis of their sequence identity, pharmacology, and preferred signal transduction mechanism. Group I mGluRs (mGluRs 1 and 5) are primarily localized postsynaptically where they modulate ion channel activity and neuronal excitability. The group I mGluRs are coupled to G ␣q and its associated effectors, such as phospholipase C. Groups II (mGluRs 2 and 3) and III (mGluRs 4, 6, 7, and 8) are primarily located presynaptically and regulate the release of neurotransmitters, including glutamate. Group II and III mGluRs are coupled to G ␣i and its associated effectors, such as adenylate cyclase. These latter two groups are distinguished from each other by their pharmacology; selective agonists and antagonists have been identified for each group (Conn and Pin, 1997). All mGluR subtypes possess a large (ϳ560 amino acids) extracellular amino-terminal domain that contains the glutamate agonist binding site. Thus, in...

This report describes the discovery of the first centrally active allosteric modulators of the metabotropic glutamate receptor subtype 5 (mGluR5). Appropriately substituted N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamides (e.g., 8) have been identified as a novel class of potent positive allosteric modulators of mGluR5 that potentiate the response to glutamate. An iterative analogue library synthesis approach provided potentiators with excellent potency and selectivity for mGluR5 (vs mGluRs 1-4, 7, 8). Compound 8q demonstrated in vivo proof of concept in an animal behavior model where known antipsychotics are active, supporting the development of new antipsychotics based on the NMDA hypofunction model for schizophrenia.