Discovery of 4-(5-Membered)Heteroarylether-6-methylpicolinamide Negative Allosteric Modulators of Metabotropic Glutamate Receptor Subtype 5.

This Letter details our efforts to develop novel, non-acetylene-containing metabotropic glutamate receptor subtype 5 (mGlu) negative allosteric modulators (NAMs) with improved pharmacological properties. This endeavor involved replacing the ether-linked pyrimidine moiety, a metabolic liability, with various 5-membered heterocycles. From this exercise, we identified , a highly brain penetrant and selective mGlu NAM which displayed moderate potency against both human and rat mGlu.

Discovery of VU6024578/BI02982816: An mGlu Positive Allosteric Modulator with Efficacy in Preclinical Antipsychotic and Cognition Models.

Herein, we report progress toward a metabotropic glutamate receptor subtype 1 (mGlu) positive allosteric modulator (PAM) clinical candidate and the discovery of VU6024578/BI02982816. From a weak high-throughput screening hit (VU0538160, EC > 10 μM, 71% Glu), optimization efforts improved functional potency over 185-fold to deliver the selective (inactive on mGlu) and CNS penetrant (rat K = 0.99, K = 0.

Discovery of VU0467319: an M Positive Allosteric Modulator Candidate That Advanced into Clinical Trials.

Herein we detail the of VU0467319 (VU319), an M Positive Allosteric Modulator (PAM) clinical candidate that successfully completed a Phase I Single Ascending Dose (SAD) clinical trial. VU319 () is a moderately potent M PAM (M PAM EC = 492 nM ± 2.9 nM, 71.

The cannabinoid CB receptor positive allosteric modulator EC21a exhibits complicated pharmacology .

Schizophrenia is a complex disease involving the dysregulation of numerous brain circuits and patients exhibit positive symptoms (hallucinations, delusions), negative symptoms (anhedonia), and cognitive impairments. We have shown that the antipsychotic efficacy of positive allosteric modulators (PAMs) of both the M muscarinic receptor and metabotropic glutamate receptor 1 (mGlu) involve the retrograde activation of the presynaptic cannabinoid type-2 (CB) receptor, indicating that CB activation or potentiation could result in a novel therapeutic strategy for schizophrenia. We used two complementary assays, receptor-mediated phosphoinositide hydrolysis and GIRK channel activation, to characterize a CB PAM scaffold, represented by the compound EC21a, to explore its potential as a starting point to optimize therapeutics for schizophrenia.

Elucidating the molecular logic of a metabotropic glutamate receptor heterodimer.

Metabotropic glutamate (mGlu) receptor protomers can heterodimerize, leading to different pharmacology compared to their homodimeric counterparts. Here, we use complemented donor-acceptor resonance energy transfer (CODA-RET) technology that distinguishes signaling from defined mGlu heterodimers or homodimers, together with targeted mutagenesis of receptor protomers and computational docking, to elucidate the mechanism of activation and differential pharmacology in mGlu heteromers. We demonstrate that positive allosteric modulators (PAMs) that bind an upper allosteric pocket in the mGlu transmembrane domain are active at both mGlu homomers and mGlu heteromers, while those that bind a lower allosteric pocket within the same domain are efficacious in homomers but not heteromers.

Discovery of VU6016235: A Highly Selective, Orally Bioavailable, and Structurally Distinct Tricyclic M Muscarinic Acetylcholine Receptor Positive Allosteric Modulator (PAM).

Herein, we report structure-activity relationship (SAR) studies to develop novel tricyclic M PAM scaffolds with improved pharmacological properties. This endeavor involved a "tie-back" strategy to replace a 5-amino-2,4-dimethylthieno[2,3-]pyrimidine-6-carboxamide core, which led to the discovery of two novel tricyclic cores. While both tricyclic cores displayed low nanomolar potency against both human and rat M and were highly brain-penetrant, the 2,4-dimethylpyrido[4',3':4,5]thieno[2,3-]pyrimidine tricycle core provided lead compound, , with an overall superior pharmacological and drug metabolism and pharmacokinetics (DMPK) profile, as well as efficacy in a preclinical antipsychotic animal model.

Discovery of VU6007496: Challenges in the Development of an M Positive Allosteric Modulator Backup Candidate.

Herein we report progress toward a backup clinical candidate to the M positive allosteric modulator (PAM) VU319/ACP-319. Scaffold-hopping from the pyrrolo[2,3-]pyridine-based M PAM VU6007477 to isomeric pyrrolo[3,2-]pyridine and thieno[3,2-]pyridine congeners identified several backup contenders. Ultimately, VU6007496, a pyrrolo[3,2-]pyridine, advanced into late stage profiling, only to be plagued with unanticipated, species-specific metabolism and active/toxic metabolites which were identified in our phenotypic seizure liability screen, preventing further development.

Discovery of VU6008677: A Structurally Distinct Tricyclic M Positive Allosteric Modulator with Improved CYP450 Profile.

This Letter details our efforts to develop novel tricyclic muscarinic acetylcholine receptor subtype 4 (M) positive allosteric modulator (PAM) scaffolds with improved pharmacological properties. This endeavor involved a "tie-back" strategy to replace the 3-amino-5-chloro-4,6-dimethylthieno[2,3-]pyridine-2-carboxamide core, which led to the discovery of two novel tricyclic cores: an 8-chloro-9-methylpyrido[3',2':4,5]thieno[3,2-]pyrimidin-4-amine core and 8-chloro-7,9-dimethylpyrido[3',2':4,5]furo[3,2-]pyrimidin-4-amine core. Both tricyclic cores displayed low nanomolar potency against human M and greatly reduced cytochrome P450 inhibition when compared with parent compound .

Development of VU6036864: A Triazolopyridine-Based High-Quality Antagonist Tool Compound of the M Muscarinic Acetylcholine Receptor.

While the muscarinic acetylcholine receptor mAChR subtype 5 (M) has been studied over decades, recent findings suggest that more in-depth research is required to elucidate a thorough understanding of its physiological function related to neurological and psychiatric disorders. Our efforts to identify potent, selective, and pharmaceutically favorable next-generation M antagonist tool compounds have led to the discovery of a novel triazolopyridine-based series. In particular, () showed exquisite potency (human M IC = 20 nM), good subtype selectivity (>500 fold selectivity against human M), desirable brain exposure ( = 0.

Potentiation of the muscarinic acetylcholine receptor 1 modulates neurophysiological features in a mouse model of Rett syndrome.

Rett syndrome (RTT) is a neurodevelopmental disorder primarily caused by mutations in the X chromosome-linked gene Methyl-CpG Binding Protein 2 (MECP2). Restoring MeCP2 expression after disease onset in a mouse model of RTT reverses phenotypes, providing hope for development of treatments for RTT. Translatable biomarkers of improvement and treatment responses have the potential to accelerate both preclinical and clinical evaluation of targeted therapies in RTT.

MeCP2 gene therapy ameliorates disease phenotype in mouse model for Pitt Hopkins syndrome.

The neurodevelopmental disorder Pitt Hopkins syndrome (PTHS) causes clinical symptoms similar to Rett syndrome (RTT) patients. However, RTT is caused by MECP2 mutations whereas mutations in the TCF4 gene lead to PTHS. The mechanistic commonalities underling these two disorders are unknown, but their shared symptomology suggest that convergent pathway-level disruption likely exists.

Novel pharmacological targets for GABAergic dysfunction in ADHD.

Attention deficit/hyperactivity disorder (ADHD) is a neurodevelopment disorder that affects approximately 5% of the population. The disorder is characterized by impulsivity, hyperactivity, and deficits in attention and cognition, although symptoms vary across patients due to the heterogenous and polygenic nature of the disorder. Stimulant medications are the standard of care treatment for ADHD patients, and their effectiveness has led to the dopaminergic hypothesis of ADHD in which deficits in dopaminergic signaling, especially in cortical brain regions, mechanistically underly ADHD pathophysiology.

Therapeutic Potential for Metabotropic Glutamate Receptor 7 Modulators in Cognitive Disorders.

Metabotropic glutamate receptor 7 (mGlu) is the most highly conserved and abundantly expressed mGlu receptor in the human brain. The presynaptic localization of mGlu, coupled with its low affinity for its endogenous agonist, glutamate, are features that contribute to the receptor's role in modulating neuronal excitation and inhibition patterns, including long-term potentiation, in various brain regions. These characteristics suggest that mGlu modulation may serve as a novel therapeutic strategy in disorders of cognitive dysfunction, including neurodevelopmental disorders that cause impairments in learning, memory, and attention.

Activation of Metabotropic Glutamate Receptor 3 Modulates Thalamo-accumbal Transmission and Rescues Schizophrenia-Like Physiological and Behavioral Deficits.

Background: Polymorphisms in the gene encoding for metabotropic glutamate receptor 3 (mGlu) are associated with an increased likelihood of schizophrenia diagnosis and can predict improvements in negative symptoms following treatment with antipsychotics. However, the mechanisms by which mGlu can regulate brain circuits involved in schizophrenia pathophysiology are not clear.

Methods: We employed selective pharmacological tools and a variety of approaches including whole-cell patch-clamp electrophysiology, slice optogenetics, and fiber photometry to investigate the effects of mGlu activation on phencyclidine (PCP)-induced impairments in thalamo-accumbal transmission and sociability deficits.

Development of a Selective and High Affinity Radioligand, [H]VU6013720, for the M Muscarinic Receptor.

M muscarinic receptors are highly expressed in the striatum and cortex, brain regions that are involved in diseases such as Parkinson's disease, schizophrenia, and dystonia. Despite potential therapeutic advantages of specifically targeting the M receptor, it has been historically challenging to develop highly selective ligands, resulting in undesired off-target activity at other members of the muscarinic receptor family. Recently, we have reported first-in-class, potent, and selective M receptor antagonists.

Differential Activity of Orthosteric Agonists and Allosteric Modulators at Metabotropic Glutamate Receptor 7.

Metabotropic glutamate receptor 7 (mGlu) is a G protein coupled receptor that has demonstrated promise as a therapeutic target across a number of neurologic and psychiatric diseases. Compounds that modulate the activity of mGlu, such as positive and negative allosteric modulators, may represent new therapeutic strategies to modulate receptor activity. The endogenous neurotransmitter associated with the mGlu receptor family, glutamate, exhibits low efficacy and potency in activating mGlu, and surrogate agonists, such as the compound L-(+)-2-Amino-4-phosphonobutyric acid (L-AP4), are often used for receptor activation and compound profiling.

GRM7 gene mutations and consequences for neurodevelopment.

The metabotropic glutamate receptor 7 (mGlu), encoded by the GRM7 gene in humans, is a presynaptic, G protein-coupled glutamate receptor that is essential for modulating neurotransmission. Mutations in or reduced expression of GRM7 have been identified in different genetic neurodevelopmental disorders (NDDs), and rare biallelic missense variants have been proposed to underlie a subset of NDDs. Clinical GRM7 variants have been associated with a range of symptoms consistent with neurodevelopmental molecular features, including hypomyelination, brain atrophy and defects in axon outgrowth.

Persistent challenges in the development of an mGlu PAM in vivo tool compound: The discovery of VU6046980.

Herein, we report on the further chemical optimization of the first reported mGlu positive allosteric modulator (PAM), VU6027459. Replacement of the quinoline core by a cinnoline scaffold increased mGlu PAM potency by ∼ 10-fold, and concomitant introduction of a chiral tricyclic motif led to potent mGlu PAMs with enantioselective mGlu receptor selectivity profiles. Of these, VU6046980 emerged as a putative in vivo tool compound with excellent CNS penetration (K = 4.

Discovery of a potent M antagonist with improved clearance profile. Part 2: Pyrrolidine amide-based antagonists.

This Letter describes our ongoing effort to improve the clearance of selective M antagonists. Herein, we report the replacement of the previously disclosed piperidine amide (4, disclosed in Part 1) with a pyrrolidine amide core. Several compounds within this series provided good potency, subtype selectivity, and low to moderate clearance profiles.

Discovery of a potent M antagonist with improved clearance profile. Part 1: Piperidine amide-based antagonists.

The lack of potent and selective tool compounds with pharmaceutically favorable properties limits the in vivo understanding of muscarinic acetylcholine receptor subtype 5 (M) biology. Previously, we presented a highly potent and selective M antagonist VU6019650 with a suboptimal clearance profile as our second-generation tool compound. Herein, we disclose our ongoing efforts to generate next-generation M antagonists with improved clearance profiles.

Differential activity of mGlu allosteric modulators provides evidence for mGlu heterodimers at hippocampal Schaffer collateral-CA1 synapses.

Glutamate acts at eight metabotropic glutamate (mGlu) receptor subtypes expressed in a partially overlapping fashion in distinct brain circuits. Recent evidence indicates that specific mGlu receptor protomers can heterodimerize and that these heterodimers can exhibit different pharmacology when compared to their homodimeric counterparts. Group III mGlu agonist-induced suppression of evoked excitatory potentials and induction of long-term potentiation at Schaffer collateral-CA1 (SC-CA1) synapses in the rodent hippocampus can be blocked by the selective mGlu negative allosteric modulator (NAM), ADX71743.

Development and profiling of mGlu NAMs with a range of saturable inhibition of agonist responses in vitro.

We describe here a series of metabotropic glutamate receptor 7 (mGlu) negative allosteric modulators (NAMs) with a saturable range of activity in inhibiting responses to an orthosteric agonist in two distinct in vitro pharmacological assays. The range of inhibition among compounds in this scaffold provides highly structurally related ligands with differential degrees of receptor blockade that can be used to understand inhibitory efficacy profiles in native tissue or in vivo.

Optimized Administration of the M PAM VU0467154 Demonstrates Broad Efficacy, but Limited Effective Concentrations in Mice.

Hypofunction of cholinergic circuits and diminished cholinergic tone have been associated with the neurodevelopmental disorder Rett syndrome (RTT). Specifically, deletion of in cholinergic neurons evokes the same social and cognitive phenotypes in mice seen with global knockout, and decreased choline acetyltransferase activity and vesamicol binding have been reported in RTT autopsy samples. Further, we recently identified significant decreases in muscarinic acetylcholine receptor subtype 4 (M) expression in both the motor cortex and cerebellum of RTT patient autopsies and established proof of concept that an acute dose of the positive allosteric modulator (PAM) VU0467154 (VU154) rescued phenotypes in mice.

Clinical and Preclinical Evidence for M Muscarinic Acetylcholine Receptor Potentiation as a Therapeutic Approach for Rett Syndrome.

Rett syndrome (RTT) is a neurodevelopmental disorder that is characterized by developmental regression, loss of communicative ability, stereotyped hand wringing, cognitive impairment, and central apneas, among many other symptoms. RTT is caused by loss-of-function mutations in a methyl-reader known as methyl-CpG-binding protein 2 (MeCP2), a protein that links epigenetic changes on DNA to larger chromatin structure. Historically, target identification for RTT has relied heavily on Mecp2 knockout mice; however, we recently adopted the alternative approach of performing transcriptional profiling in autopsy samples from RTT patients.