TY - JOUR

T1 - Detailed in vitro pharmacological characterization of clinically tested negative allosteric modulators of the metabotropic glutamate receptor 5 (mGlu5)

AU - Arsova, Angela

AU - Møller, Thor C

AU - Vedel, Line

AU - Hansen, Jakob L

AU - Foster, Simon R

AU - Gregory, Karen J

AU - Bräuner-Osborne, Hans

N1 - The American Society for Pharmacology and Experimental Therapeutics.

PY - 2020

Y1 - 2020

N2 - Negative allosteric modulation of the metabotropic glutamate 5 (mGlu5) receptor has emerged as a potential strategy for the treatment of neurological disorders. Despite the success in preclinical studies, many mGlu5 negative allosteric modulators (NAMs) that have reached clinical trials failed due to lack of efficacy. In this study, we provide a detailed in vitro pharmacological characterization of nine clinically and preclinically tested NAMs. We evaluated inhibition of L-glutamate-induced signaling with Ca2+ mobilization, inositol monophosphate (IP1) accumulation, extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation, and real-time receptor internalization assays on rat mGlu5 expressed in HEK293A cells. Moreover, we determined association (kon) and dissociation (koff) rates, as well as NAM affinity with [3H]methoxy-PEPy binding experiments. kon and koff values varied greatly between the nine NAMs (34- and 139-fold, respectively) resulting in long receptor residence times (>400 min) for basimglurant and mavoglurant, medium residence times (10-30 min) for AZD2066, remeglurant and (RS)-remeglurant, and low residence time (<10 min) for dipraglurant, F169521, F1699611 and STX107. We found that all NAMs inhibited L-glutamate induced mGlu5 receptor internalization, generally with a similar potency to IP1 accumulation and ERK1/2 phosphorylation while Ca2+ mobilization was less potently inhibited. Operational model of allosterism analyses revealed that dipraglurant and (RS)-remeglurant were biased towards (affinity) receptor internalization and away (cooperativity) from the ERK1/2 phosphorylation pathway, respectively. Our study is the first to measure binding kinetics and negative allosteric modulation of mGlu5 receptor internalization, and adds significant new knowledge about the molecular pharmacology of a diverse range of clinically relevant NAMs. SIGNIFICANCE STATEMENT: The metabotropic glutamate 5 (mGlu5) receptor is important in many brain functions and implicated in several neurological pathologies. Negative allosteric modulators (NAMs) have shown promising results in preclinical models but have so far failed in human clinical trials. Here we provide the most comprehensive and comparative molecular pharmacological study to date of nine preclinical/clinical tested negative allosteric modulators (NAMs) at the mGlu5 receptor, which is also the first study to measure binding kinetics and negative allosteric modulation of mGlu5 receptor internalization.

AB - Negative allosteric modulation of the metabotropic glutamate 5 (mGlu5) receptor has emerged as a potential strategy for the treatment of neurological disorders. Despite the success in preclinical studies, many mGlu5 negative allosteric modulators (NAMs) that have reached clinical trials failed due to lack of efficacy. In this study, we provide a detailed in vitro pharmacological characterization of nine clinically and preclinically tested NAMs. We evaluated inhibition of L-glutamate-induced signaling with Ca2+ mobilization, inositol monophosphate (IP1) accumulation, extracellular signal-regulated kinase 1/2 (ERK1/2) phosphorylation, and real-time receptor internalization assays on rat mGlu5 expressed in HEK293A cells. Moreover, we determined association (kon) and dissociation (koff) rates, as well as NAM affinity with [3H]methoxy-PEPy binding experiments. kon and koff values varied greatly between the nine NAMs (34- and 139-fold, respectively) resulting in long receptor residence times (>400 min) for basimglurant and mavoglurant, medium residence times (10-30 min) for AZD2066, remeglurant and (RS)-remeglurant, and low residence time (<10 min) for dipraglurant, F169521, F1699611 and STX107. We found that all NAMs inhibited L-glutamate induced mGlu5 receptor internalization, generally with a similar potency to IP1 accumulation and ERK1/2 phosphorylation while Ca2+ mobilization was less potently inhibited. Operational model of allosterism analyses revealed that dipraglurant and (RS)-remeglurant were biased towards (affinity) receptor internalization and away (cooperativity) from the ERK1/2 phosphorylation pathway, respectively. Our study is the first to measure binding kinetics and negative allosteric modulation of mGlu5 receptor internalization, and adds significant new knowledge about the molecular pharmacology of a diverse range of clinically relevant NAMs. SIGNIFICANCE STATEMENT: The metabotropic glutamate 5 (mGlu5) receptor is important in many brain functions and implicated in several neurological pathologies. Negative allosteric modulators (NAMs) have shown promising results in preclinical models but have so far failed in human clinical trials. Here we provide the most comprehensive and comparative molecular pharmacological study to date of nine preclinical/clinical tested negative allosteric modulators (NAMs) at the mGlu5 receptor, which is also the first study to measure binding kinetics and negative allosteric modulation of mGlu5 receptor internalization.

U2 - 10.1124/mol.119.119032

DO - 10.1124/mol.119.119032

M3 - Journal article

C2 - 32358164

VL - 98

SP - 49

EP - 60

JO - Molecular Pharmacology

JF - Molecular Pharmacology

SN - 0026-895X

IS - 1

ER -