A Novel Class of Succinimide-Derived Negative Allosteric Modulators of Metabotropic Glutamate Receptor Subtype 1 Provides Insight into a Disconnect in Activity between the Rat and Human Receptors

Recent progress in the discovery of mGlu₁ allosteric modulators has suggested the modulation of mGlu₁ could offer possible treatment for a number of central nervous system disorders; however, the available chemotypes are inadequate to fully investigate the therapeutic potential of mGlu₁ modulation. To address this issue, we used a fluorescence-based high-throughput screening assay to screen an allosteric modulator-biased library of compounds to generate structurally diverse mGlu₁ negative allosteric modulator hits for chemical optimization. Herein, we describe the discovery and characterization of a novel mGlu₁ chemotype. This series of succinimide negative allosteric modulators, exemplified by VU0410425, exhibited potent inhibitory activity at rat mGlu₁ but was, surprisingly, inactive at human mGlu₁. VU0410425 and a set of chemically diverse mGlu₁ negative allosteric modulators previously reported in the literature were utilized to examine this species disconnect between rat and human mGlu₁ activity. Mutation of the key transmembrane domain residue 757 and functional screening of VU0410425 and the literature compounds suggests that amino acid 757 plays a role in the activity of these compounds, but the contribution of the residue is scaffold specific, ranging from critical to minor. The operational model of allosterism was used to estimate the binding affinities of each compound to compare to functional data. This novel series of mGlu₁ negative allosteric modulators provides valuable insight into the pharmacology underlying the disconnect between rat and human mGlu₁ activity, an issue that must be understood to progress the therapeutic potential of allosteric modulators of mGlu₁

Identification of Metabotropic Glutamate Receptor Subtype 5 Potentiators Using Virtual High-Throughput Screening

Selective potentiators of glutamate response at metabotropic glutamate receptor subtype 5 (mGluR5) have exciting potential for the development of novel treatment strategies for schizophrenia. A total of 1,382 compounds with positive allosteric modulation (PAM) of the mGluR5 glutamate response were identified through high-throughput screening (HTS) of a diverse library of 144,475 substances utilizing a functional assay measuring receptor-induced intracellular release of calcium. Primary hits were tested for concentration-dependent activity, and potency data (EC₅₀ values) were used for training artificial neural network (ANN) quantitative structure−activity relationship (QSAR) models that predict biological potency from the chemical structure. While all models were trained to predict EC₅₀, the quality of the models was assessed by using both continuous measures and binary classification. Numerical descriptors of chemical structure were used as input for the machine learning procedure and optimized in an iterative protocol. The ANN models achieved theoretical enrichment ratios of up to 38 for an independent data set not used in training the model. A database of ∼450,000 commercially available drug-like compounds was targeted in a virtual screen. A set of 824 compounds was obtained for testing based on the highest predicted potency values. Biological testing found 28.2% (232/824) of these compounds with various activities at mGluR5 including 177 pure potentiators and 55 partial agonists. These results represent an enrichment factor of 23 for pure potentiation of the mGluR5 glutamate response and 30 for overall mGluR5 modulation activity when compared with those of the original mGluR5 experimental screening data (0.94% hit rate). The active compounds identified contained 72% close derivatives of previously identified PAMs as well as 28% nontrivial derivatives of known active compounds

Design and Synthesis of mGlu₂ NAMs with Improved Potency and CNS Penetration Based on a Truncated Picolinamide Core

Herein, we detail the optimization of the mGlu₂ negative allosteric modulator (NAM), VU6001192, by a reductionist approach to afford a novel, simplified mGlu₂ NAM scaffold. This new chemotype not only affords potent and selective mGlu₂ inhibition, as exemplified by VU6001966 (mGlu₂ IC₅₀ = 78 nM, mGlu₃ IC₅₀ > 30 μM), but also excellent central nervous system (CNS) penetration (<i>K</i>_p = 1.9, <i>K</i>_p,uu = 0.78), a feature devoid in all previously disclosed mGlu₂ NAMs (<i>K</i>_ps ≈ 0.3, <i>K</i>_p,uus ≈ 0.1). Moreover, this series, based on overall properties, represents an exciting lead series for potential mGlu₂ PET tracer development

Design of 4‑Oxo-1-aryl-1,4-dihydroquinoline-3-carboxamides as Selective Negative Allosteric Modulators of Metabotropic Glutamate Receptor Subtype 2

Both orthosteric and allosteric antagonists of the group II metabotropic glutamate receptors (mGlus) have been used to establish a link between mGlu_2/3 inhibition and a variety of CNS diseases and disorders. Though these tools typically have good selectivity for mGlu_2/3 versus the remaining six members of the mGlu family, compounds that are selective for only one of the individual group II mGlus have proved elusive. Herein we report on the discovery of a potent and highly selective mGlu₂ negative allosteric modulator <b>58</b> (VU6001192) from a series of 4-oxo-1-aryl-1,4-dihydroquinoline-3-carboxamides. The concept for the design of this series centered on morphing a quinoline series recently disclosed in the patent literature into a chemotype previously used for the preparation of muscarinic acetylcholine receptor subtype 1 positive allosteric modulators. Compound <b>58</b> exhibits a favorable profile and will be a useful tool for understanding the biological implications of selective inhibition of mGlu₂ in the CNS

Substituted 1‑Phenyl-3-(pyridin-2-yl)urea Negative Allosteric Modulators of mGlu₅: Discovery of a New Tool Compound VU0463841 with Activity in Rat Models of Cocaine Addiction

Cocaine is a powerful and highly addictive stimulant that disrupts the normal reward circuitry in the central nervous system (CNS), producing euphoric effects. Cocaine use can lead to acute and life threatening emergencies, and abuse is associated with increased risk for contracting infectious diseases. Though certain types of behavioral therapy have proven effective for treatment of cocaine addiction, relapse remains high, and there are currently no approved medications for the treatment of cocaine abuse. Evidence has continued to accumulate that indicates a critical role for the metabotropic glutamate receptor subtype 5 (mGlu₅) in the modulation of neural circuitry associated with the addictive properties of cocaine. While the small molecule mGlu₅ negative allosteric modulator (NAM) field is relatively advanced, investigation into the potential of small molecule mGlu₅ NAMs for the treatment of cocaine addiction remains an area of high interest. Herein we describe the discovery and characterization of a potent and selective compound <b>29</b> (VU0463841) with good CNS exposure in rats. The utility of <b>29</b> (VU0463841) was demonstrated by its ability to attenuate drug seeking behaviors in relevant rat models of cocaine addiction

Chemical Modulation of Mutant mGlu₁ Receptors Derived from Deleterious <i>GRM1</i> Mutations Found in Schizophrenics

Schizophrenia is a complex and highly heterogeneous psychiatric disorder whose precise etiology remains elusive. While genome-wide association studies (GWAS) have identified risk genes, they have failed to determine if rare coding single nucleotide polymorphisms (nsSNPs) contribute in schizophrenia. Recently, two independent studies identified 12 rare, deleterious nsSNPS in the <i>GRM1</i> gene, which encodes the metabotropic glutamate receptor subtype 1 (mGlu₁), in schizophrenic patients. Here, we generated stable cell lines expressing the mGlu₁ mutant receptors and assessed their pharmacology. Using both the endogenous agonist glutamate and the synthetic agonist DHPG, we found that several of the mutant mGlu₁ receptors displayed a loss of function that was not due to a loss in plasma membrane expression. Due to a lack of mGlu₁ positive allosteric modulators (PAM) tool compounds active at human mGlu₁, we optimized a known mGlu₄ PAM/mGlu₁ NAM chemotype into a series of potent and selective mGlu₁ PAMs by virtue of a double “molecular switch”. Employing mGlu₁ PAMs from multiple chemotypes, we demonstrate that the mutant receptors can be potentiated by small molecules and in some cases efficacy restored to that comparable to wild type mGlu₁ receptors, suggesting deficits in patients with schizophrenia due to these mutations may be amenable to intervention with an mGlu₁ PAM. However, in wild type animals, mGlu₁ negative allosteric modulators (NAMs) are efficacious in classic models predictive of antipsychotic activity, whereas we show that mGlu₁ PAMs have no effect to slight potentiation in these models. These data further highlight the heterogeneity of schizophrenia and the critical role of patient selection strategies in psychiatric clinical trials to match genotype with therapeutic mechanism

Discovery of a Selective and CNS Penetrant Negative Allosteric Modulator of Metabotropic Glutamate Receptor Subtype 3 with Antidepressant and Anxiolytic Activity in Rodents

Previous preclinical work has demonstrated the therapeutic potential of antagonists of the group II metabotropic glutamate receptors (mGlus). Still, compounds that are selective for the individual group II mGlus (mGlu₂ and mGlu₃) have been scarce. There remains a need for such compounds with the balance of properties suitable for convenient use in a wide array of rodent behavioral studies. We describe here the discovery of a selective mGlu₃ NAM <b>106</b> (VU0650786) suitable for in vivo work. Compound <b>106</b> is a member of a series of 5-aryl-6,7-dihydropyrazolo­[1,5-<i>a</i>]­pyrazine-4­(5<i>H</i>)-one compounds originally identified as a mGlu₅ positive allosteric modulator (PAM) chemotype. Its suitability for use in rodent behavioral models has been established by extensive in vivo PK studies, and the behavioral experiments presented here with compound <b>106</b> represent the first examples in which an mGlu₃ NAM has demonstrated efficacy in models where prior efficacy had previously been noted with nonselective group II antagonists

Discovery of VU6005649, a CNS Penetrant mGlu_7/8 Receptor PAM Derived from a Series of Pyrazolo[1,5‑<i>a</i>]pyrimidines

Herein, we report the structure–activity relationships within a series of mGlu₇ PAMs based on a pyrazolo­[1,5-<i>a</i>]­pyrimidine core with excellent CNS penetration (<i>K</i>_ps > 1 and <i>K</i>_p,uus > 1). Analogues in this series proved to display a range of Group III mGlu receptor selectivity, but VU6005649 emerged as the first dual mGlu_7/8 PAM, filling a void in the Group III mGlu receptor PAM toolbox and demonstrating <i>in vivo</i> efficacy in a mouse contextual fear conditioning model

VU6010608, a Novel mGlu₇ NAM from a Series of <i>N</i>‑(2-(1<i>H</i>‑1,2,4-Triazol-1-yl)-5-(trifluoromethoxy)phenyl)benzamides

Herein, we report the structure–activity relationships within a series of mGlu₇ NAMs based on an <i>N</i>-(2-(1<i>H</i>-1,2,4-triazol-1-yl)-5-(trifluoromethoxy)­phenyl)­benzamide core with excellent CNS penetration (<i>K</i>_p 1.9–5.8 and <i>K</i>_p,uu 0.4–1.4). Analogues in this series displayed steep SAR. Of these, VU6010608 (<b>11a</b>) emerged with robust efficacy in blocking high frequency stimulated long-term potentiation in electrophysiology studies

Discovery of <i>N</i>‑(5-Fluoropyridin-2-yl)-6-methyl-4-(pyrimidin-5-yloxy)picolinamide (VU0424238): A Novel Negative Allosteric Modulator of Metabotropic Glutamate Receptor Subtype 5 Selected for Clinical Evaluation

Preclinical evidence in support of the potential utility of mGlu₅ NAMs for the treatment of a variety of psychiatric and neurodegenerative disorders is extensive, and multiple such molecules have entered clinical trials. Despite some promising results from clinical studies, no small molecule mGlu₅ NAM has yet to reach market. Here we present the discovery and evaluation of <i>N</i>-(5-fluoropyridin-2-yl)-6-methyl-4-(pyrimidin-5-yloxy)­picolinamide (<b>27</b>, VU0424238), a compound selected for clinical evaluation. Compound <b>27</b> is more than 900-fold selective for mGlu₅ versus the other mGlu receptors, and binding studies established a <i>K</i>_i value of 4.4 nM at a known allosteric binding site. Compound <b>27</b> had a clearance of 19.3 and 15.5 mL/min/kg in rats and cynomolgus monkeys, respectively. Imaging studies using a known mGlu₅ PET ligand demonstrated 50% receptor occupancy at an oral dose of 0.8 mg/kg in rats and an intravenous dose of 0.06 mg/kg in baboons