Rescue of Fmr1KO phenotypes with mGluR5 inhibitors: MRZ-8456 versus AFQ-056.

Metabotropic glutamate receptor 5 (mGluR5) is a drug target for central nervous system disorders such as fragile X syndrome that involve excessive glutamate-induced excitation. We tested the efficacy of a novel negative allosteric modulator of mGluR5 developed by Merz Pharmaceuticals, MRZ-8456, in comparison to MPEP and AFQ-056 (Novartis, a.k.a. mavoglurant) in both in vivo and in vitro assays in a mouse model of fragile X syndrome, Fmr1KO mice. The in vivo assays included susceptibility to audiogenic-induced seizures and pharmacokinetic measurements of drug availability. The in vitro assays included dose response assessments of biomarker expression and dendritic spine length and density in cultured primary neurons. Both MRZ-8456 and AFQ-056 attenuated wild running and audiogenic-induced seizures in Fmr1KO mice with similar pharmacokinetic profiles. Both drugs significantly reduced dendritic expression of amyloid-beta protein precursor (APP) and rescued the ratio of mature to immature dendritic spines. These findings demonstrate that MRZ-8456, a drug being developed for the treatment of motor complications of L-DOPA in Parkinson's disease and which completed a phase I clinical trial, is effective in attenuating both well-established (seizures and dendritic spine maturity) and exploratory biomarker (APP expression) phenotypes in a mouse model of fragile X syndrome.

Assessment of the pulmonary CYP1A1 metabolism of mavoglurant (AFQ056) in rat.

1. AFQ056 phenotyping results indicate that CYP1A1 is responsible for the formation of the oxidative metabolite, M3. In line with the predominant assumption that CYP1A1 is mainly expressed in extrahepatic tissues, only traces of M3 were detected in hepatic systems. The aim of this study was to investigate the pulmonary CYP1A1 mediated metabolism of AFQ056 in rat. 2. Western blot analysis confirmed that CYP1A1 is expressed in rat lung albeit at low levels. M3 formation was clearly observed in recombinant rat CYP1A1, lung microsomes and lung tissue slices and was strongly inhibited by ketoconazole in the incubations. As CYP3A4 and CYP2C9 metabolites were only observed at trace levels, we concluded that the reduced M3 formation was due to CYP1A1 inhibition. 3. AFQ056 lung clearance (CLlung) as estimated from in vitro data was predicted to be negligible (<1% pulmonary blood flow). This was confirmed by in vivo experiments where intravenous and intra-arterial dosing to rats failed to show significant pulmonary extraction. 4. While rat lung may make a contribution to the formation of M3, it is unlikely to be the only organ involved in this process and further experiments are required to investigate the potential metabolic elimination routes for AFQ056.

Mavoglurant (AFQ056) in combination with increased levodopa dosages in Parkinson's disease patients.

Long-term use of levodopa (L-dopa) in patients with Parkinson's disease is associated with development of dyskinesia. This study explored whether Parkinson's disease patients with L-dopa-induced dyskinesia experience improved OFF-time from higher L-dopa doses without worsening of dyskinesias when receiving adjunctive mavoglurant. Patients with moderate-to-severe L-dopa-induced dyskinesia were randomized to receive mavoglurant or placebo. Mavoglurant (AFQ056) was up-titrated over two weeks from 25 mg twice daily (bid) to 100 mg bid (L-dopa kept stable), followed by three weeks during which the daily L-dopa dosage was increased by up to 300 mg/day. A sample size of 30 was initially planned; however, the study was terminated prematurely due to enrollment challenges. OFF-time showed greater improvements in the mavoglurant group (n = 7) compared with the placebo group (n = 7); difference at week 5 was -2.77 h (90% confidence interval -5.44, -0.09 h; p = 0.09). ON-time without troublesome dyskinesia increased more from baseline to week 5 in the mavoglurant group (4.38 h) versus the placebo group (0.63 h). Clinician-rated measures were conflicting. The Modified Abnormal Involuntary Movement Scale scores showed a slight improvement with mavoglurant compared with placebo, while the Unified Dyskinesia Rating Scale parts III and IV worsened slightly with mavoglurant compared with placebo. Due to the low patient numbers and conflicting clinician-rated outcomes data, our findings are not conclusive. However, our results suggest that mavoglurant combined with higher doses of L-dopa may be effective in treating patients with Parkinson's disease experiencing L-dopa-related motor fluctuations and dyskinesias.

A randomized, placebo-controlled trial of AFQ056 for the treatment of chorea in Huntington's disease.

BACKGROUND: This study investigated the hypothesis that AFQ056 (mavoglurant), a selective metabotropic glutamate receptor 5 antagonist, reduces chorea in Huntington's disease (HD). METHODS: This 32-day randomized, double-blind, parallel-group, proof-of-concept study investigated AFQ056 (25-150 mg [incremental doses], twice-daily) versus placebo in patients with HD. Primary efficacy assessments were the chorea-sum score and orientation index (nondominant hand) from the quantitative motor (Q-Motor) grasping task at day 28. Key secondary efficacy assessments included finger-tapping in the Unified Huntington's Disease Rating Scale-Total Motor Score and Q-Motor measures. Safety and tolerability were assessed. RESULTS: Overall, 42 patients were randomized. At day 28, no improvement was observed on the primary efficacy assessments (P > 0.10) with AFQ056 versus placebo. The Q-Motor speeded-tapping interonset interval variability was reduced with AFQ056 versus placebo for the nondominant hand (P = 0.01). The incidence of adverse events was 66.7% with AFQ056 and 57.1% with placebo. CONCLUSIONS: AFQ056 did not reduce choreatic movements in HD, but was well tolerated. The clinical relevance of the Q-Motor findings (speeded-tapping) are unknown and may warrant further investigation.

Development of allosteric modulators of GPCRs for treatment of CNS disorders.

The discovery of allosteric modulators of G protein-coupled receptors (GPCRs) provides a promising new strategy with potential for developing novel treatments for a variety of central nervous system (CNS) disorders. Traditional drug discovery efforts targeting GPCRs have focused on developing ligands for orthosteric sites which bind endogenous ligands. Allosteric modulators target a site separate from the orthosteric site to modulate receptor function. These allosteric agents can either potentiate (positive allosteric modulator, PAM) or inhibit (negative allosteric modulator, NAM) the receptor response and often provide much greater subtype selectivity than orthosteric ligands for the same receptors. Experimental evidence has revealed more nuanced pharmacological modes of action of allosteric modulators, with some PAMs showing allosteric agonism in combination with positive allosteric modulation in response to endogenous ligand (ago-potentiators) as well as "bitopic" ligands that interact with both the allosteric and orthosteric sites. Drugs targeting the allosteric site allow for increased drug selectivity and potentially decreased adverse side effects. Promising evidence has demonstrated potential utility of a number of allosteric modulators of GPCRs in multiple CNS disorders, including neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease, as well as psychiatric or neurobehavioral diseases such as anxiety, schizophrenia, and addiction.

AFQ056 in Parkinson patients with levodopa-induced dyskinesia: 13-week, randomized, dose-finding study.

AFQ056 is a novel, selective metabotropic glutamate receptor 5 antagonist. This was a 13-week, double-blind, placebo-controlled study. Patients with Parkinson's disease and moderate-to-severe levodopa (l-dopa)-induced dyskinesia who were receiving stable l-dopa/anti-parkinsonian treatment and were not currently receiving amantadine were randomized to receive either AFQ056 (at doses of 20, 50, 100, 150, or 200 mg daily) or placebo (1:1:1:1:2:3 ratio) for 12 weeks. The primary outcome was the modified Abnormal Involuntary Movements Scale. Secondary outcomes included the 26-item Parkinson's Disease Dyskinesia Scale, the Patient's/Clinician's Global Impression of Change, and the Unified Parkinson's Disease Rating Scale parts III (motor evaluation) and IV (severity of motor complications). Safety was assessed. In total, 98 of 133 (73.7%) AFQ056-treated patients and 47 of 64 (73.4%) patients in the placebo group completed the study. Baseline characteristics were comparable. Patients randomized to AFQ056 200 mg daily administered in 2 doses demonstrated significant improvements at Week 12 on the modified Abnormal Involuntary Movements Scale compared with placebo (difference, -2.8; 95% confidence interval [CI], -5.2, -0.4; P = 0.007). Based on final actual doses, there was a dose-response relationship on the modified Abnormal Involuntary Movements Scale, with 200 mg daily demonstrating the most robust effect (difference, -3.6; 95% CI, -7.0, -0.3; P = 0.012). Improvements in dyskinesia were supported by change on Unified Parkinson's Disease Rating Scale part IV item 32 (50 mg daily: difference, -0.7; 95% CI, -1.1, -0.2; P = 0.003; 200 mg daily: difference, -0.5; 95% CI, -0.8, -0.1; P = 0.005). No significant changes were observed on the 26-item Parkinson's Disease Dyskinesia Scale, the Unified Parkinson's Disease Rating Scale part IV item 33 or items 32 and 33, or the Patient's/Clinician's Global Impression of Change. Unified Parkinson's Disease Rating Scale part III scores were not significantly changed, indicating no worsening of motor symptoms. The most common adverse events (with incidence greater with AFQ056 than with placebo) were dizziness, hallucination, fatigue, nasopharyngitis, diarrhea, and insomnia. AFQ056 demonstrated anti-dyskinetic efficacy in this population without worsening underlying motor symptoms. These results will guide dose selection for future clinical trials.

Scaffold hopping approach towards various AFQ-056 analogs as potent metabotropic glutamate receptor 5 negative allosteric modulators.

The metabotropic glutamate receptor subtype 5 has evolved into a promising target for the treatment of various diseases of the central nervous system, such as Fragile X and L-DOPA induced dyskinesia. One of the most advanced clinical compound is Novartis' AFQ-056 (Mavoglurant), which served us as a template for a scaffold hopping approach, generating a structurally diverse set of potent analogs. Both the limited aqueous solubility and the relatively poor metabolic stability of AFQ-056 were improved with hexahydrocyclopenta[c]pyrrole derivative 54a, which proved to be a valuable candidate for further development.

Targeted Treatments for Fragile X Syndrome.

The histories of targeted treatment trials in fragile X syndrome (FXS) are reviewed in animal studies and human trials. Advances in understanding the neurobiology of FXS have identified a number of pathways that are dysregulated in the absence of FMRP and are therefore pathways that can be targeted with new medication. The utilization of quantitative outcome measures to assess efficacy in multiple studies has improved the quality of more recent trials. Current treatment trials including the use of cannabidiol (CBD) topically and metformin orally have positive preliminary data, and both of these medications are available clinically. The use of the phosphodiesterase inhibitor (PDE4D), BPN1440, which raised the level of cAMP that is low in FXS has very promising results for improving cognition in adult males who underwent a controlled trial. There are many more targeted treatments that will undergo trials in FXS, so the future looks bright for new treatments.

Comparison of Rat and Human Pulmonary Metabolism Using Precision-cut Lung Slices (PCLS).

BACKGROUND: Although the liver is the primary organ of drug metabolism, the lungs also contain drug-metabolizing enzymes and may, therefore, contribute to the elimination of drugs. In this investigation, the Precision-cut Lung Slice (PCLS) technique was standardized with the aims of characterizing and comparing rat and human pulmonary drug metabolizing activity. METHOD: Due to the limited availability of human lung tissue, standardization of the PCLS method was performed with rat lung tissue. Pulmonary enzymatic activity was found to vary significantly with rat age and rat strain. The Dynamic Organ Culture (DOC) system was superior to well-plates for tissue incubations, while oxygen supply appeared to have a limited impact within the 4h incubation period used here. RESULTS: The metabolism of a range of phase I and phase II probe substrates was assessed in rat and human lung preparations. Cytochrome P450 (CYP) activity was relatively low in both species, whereas phase II activity appeared to be more significant. CONCLUSION: PCLS is a promising tool for the investigation of pulmonary drug metabolism. The data indicates that pulmonary CYP activity is relatively low and that there are significant differences in enzyme activity between rat and human lung.

Mavoglurant Augmentation in OCD Patients Resistant to Selective Serotonin Reuptake Inhibitors: A Proof-of-Concept, Randomized, Placebo-Controlled, Phase 2 Study.

INTRODUCTION: To determine if mavoglurant (modified release) as an augmentation therapy to selective serotonin reuptake inhibitors (SSRIs) could have beneficial effects reducing Yale-Brown Obsessive Compulsive Scale (Y-BOCS) total score in patients with obsessive-compulsive disorder (OCD) resistant to SSRI treatment. METHODS: This was a multicenter, randomized, double-blind, placebo-controlled, parallel-group, phase 2 study. Patients remained on their SSRI treatment and mavoglurant or placebo was added on. Non-smoking men and women aged 18-65 years primarily diagnosed with OCD according to Diagnostic and Statistical Manual of Mental Disorders (4th ed., text rev.; DSM-IV-TR) criteria were randomized (1:1) to mavoglurant or placebo groups. After 50 patients were randomized, an interim analysis was conducted to determine whether the study should be continued. The primary outcome measure was absolute change in Y-BOCS from baseline at week 17. Safety was assessed by recording adverse events (AEs) and serious adverse events (SAEs). RESULTS: Interim analysis led to a decision to terminate the study. In total 38 (76.0%) participants completed 17 weeks of treatment and 37 (74.0%) completed the study. There was no significant difference in least squares (LS) mean change from baseline at week 17 in Y-BOCS total score for mavoglurant compared with placebo groups [-6.9 (1.75) vs. -8.0 (1.78), respectively; LS mean difference 1.1; 95% CI -3.9, 6.2; p = 0.671]. The incidence of AEs was higher in the mavoglurant compared with the placebo group (80.8% vs. 70.8%, respectively). CONCLUSION: This study of mavoglurant in OCD was terminated because of the lack of efficacy at interim analysis. The study did not support the use of an antagonist of mGluR5 receptors for OCD treatment. TRIAL REGISTRATION: The study was registered with ClinicalTrials.gov: NCT01813019. FUNDING: This study was sponsored by Novartis Pharma AG, Basel, Switzerland.

Mavoglurant in adolescents with fragile X syndrome: analysis of Clinical Global Impression-Improvement source data from a double-blind therapeutic study followed by an open-label, long-term extension study.

BACKGROUND: A phase II randomized, placebo-controlled, double-blind study and subsequent open-label extension study evaluated the efficacy, safety, and tolerability of mavoglurant (AFQ056), a selective metabotropic glutamate receptor subtype-5 antagonist, in treating behavioral symptoms in adolescent patients with fragile X syndrome (FXS). A novel method was applied to analyze changes in symptom domains in patients with FXS using the narratives associated with the clinician-rated Clinical Global Impression-Improvement (CGI-I) scale. METHODS: In the core study, patients were randomized to receive mavoglurant (25, 50, or 100 mg BID) or placebo over 12 weeks. In the extension, patients received 100 mg BID mavoglurant (or the highest tolerated dose) for up to 32 months. Global improvement, as a measure of treatment response, was assessed using the CGI-I scale. Investigators assigning CGI-I scores of 1 (very much improved), 2 (much improved), 6 (much worse), or 7 (very much worse) were provided a standard narrative template to collect further information about the changes observed in patients. Investigator feedback was coded and clustered into categories of improvement or worsening to identify potential areas of improvement with mavoglurant. Treatment effect in each category was characterized using the Cochran-Mantel-Haenszel test. RESULTS: A total of 134 and 103 patients had reached 2 weeks or more of core and extension study treatment, respectively, by the pre-assigned cutoff date for investigator feedback. In the core study, 34 CGI-I scores of 1 or 2 were reported in 28 patients; one patient scored 6. Analysis of the CGI-I narratives did not indicate greater treatment response in patients receiving mavoglurant compared with placebo in any specific improvement domain. There were 54 CGI-I scores of 1 or 2 in 47 patients in the extension study. The most frequently reported categories of improvement were behavior and mood (79.3 and 76.6 % in core and extension studies, respectively), engagement (75.9 and 78.7 %), and communication (69.0 and 61.7 %). CONCLUSIONS: A method was established to capture and categorize FXS symptoms using CGI-I narratives. Although this method did not show benefit of drug over placebo, narratives from investigators were mostly based on parental report and thus do not represent a completely objective alternative assessment. TRIAL REGISTRATION: The studies described are registered at ClinicalTrials.gov with clinical trial identifier numbers NCT01357239 and NCT01433354.

Progress toward therapeutic potential for AFQ056 in Fragile X syndrome.

Fragile X syndrome (FXS) is the most common form of inherited intellectual disability and the leading single-gene cause of autism. It is caused by the lack of production of the Fragile X mental retardation protein (FMRP), resulting in cognitive deficits, hyperactivity, and autistic behaviors. Breakthrough advances in potential therapy for FXS followed the discovery that aberrant group 1 metabotropic glutamate receptor (mGluR) signaling is an important constituent of the pathophysiology of the syndrome. Research has indicated that upon neuronal stimulation, FMRP acts downstream of group 1 mGluRs (mGluRs1/5) to inhibit protein synthesis, long-term depression, and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor internalization. To offset the deficits caused by the lack of FMRP, many pharmaceutical companies have designed medicinal drugs to target the unrestrained stimulation of mGluR5 signaling in FXS. Indeed, promising results from animal and clinical studies suggest that mGluR5 antagonists such as AFQ056 can successfully correct many of the deficits in FXS. In this review, we cover the animal studies performed to date that test the role of AFQ056 as a selective mGluR5 antagonist to alleviate the phenotypes of FXS.

Neurophysiological signals as potential translatable biomarkers for modulation of metabotropic glutamate 5 receptors.

The Group I metabotropic glutamate receptor subtype 5 (mGluR5) is widely distributed in the brain with dense expression in the cerebral cortex, hippocampus, and basal ganglia. These receptors have been implicated in psychiatric and neurological disorders such as schizophrenia, Fragile X syndrome, addiction, anxiety/depression, Parkinson's disease and neuropathic pain. The present study evaluated the effects of the mGluR5 negative allosteric modulators (NAMs) 4-difluoromethoxy-3-(pyridine-2-ylethynyl)phenyl)5H-pyrrolo[3,4-b]pyridine-6(7H)-yl methanone (GRN-529) and methyl (3aR,4S,7aR)-4-hydroxy-4-[(3-methylphenyl)ethynyl]octahydro-1H-indole-1-carboxylate (AFQ056) on polysomnographic (PSG) and quantitative electroencephalographic (qEEG) measures in freely moving rats. Furthermore, the anxiolytic profile of GRN-529 was characterized in anesthetized rats by measuring stimulation-induced hippocampal theta oscillation. The present findings demonstrate that inhibition of mGluR5 via its allosteric site profoundly modulates high-level neuronal network activities as indicated by changes in sleep-wake activity and power distribution of qEEG. Both GRN-529 and AFQ056 reduced the total time spent in rapid-eye movement with AFQ056 producing a significant increase in wakefulness at the highest dose tested. Additionally, qEEG revealed significant compound-induced increases in delta power concomitant with more subtle decreases in theta and alpha band power. Receptor occupancy (RO) studies revealed that GRN-529 and AFQ056 at all doses resulted in over 45% mGluR5 occupancy. Furthermore, GRN-529 dose-dependently decreased elicited hippocampal theta frequency, consistent with previous findings using clinically active anxiolytic compounds. The described changes in neurophysiological signals identified in freely moving rats may be considered suitable translational biomarkers for the clinical evaluation of mGluR5 NAMs.