Ashwin Muraleetharan, Yuyang Wang, Matthew C Rowe, Ashleigh Gould, Karen J Gregory, Shane D Hellyer
{"title":"人类代谢性谷氨酸受体1变构调节的严格表征揭示了探针和测定依赖的药理学。","authors":"Ashwin Muraleetharan, Yuyang Wang, Matthew C Rowe, Ashleigh Gould, Karen J Gregory, Shane D Hellyer","doi":"10.1124/molpharm.122.000664","DOIUrl":null,"url":null,"abstract":"<p><p>Allosteric modulation of metabotropic glutamate receptor subtype 1 (mGlu<sub>1</sub>) represents a viable therapeutic target for treating numerous central nervous system disorders. Although multiple chemically distinct mGlu<sub>1</sub> positive (PAMs) and negative (NAMs) allosteric modulators have been identified, drug discovery paradigms have not included rigorous pharmacological analysis. In the present study, we hypothesized that existing mGlu<sub>1</sub> allosteric modulators possess unappreciated probe-dependent or biased pharmacology. Using human embryonic kidney 293 (HEK293A) cells stably expressing human mGlu<sub>1,</sub> we screened mGlu<sub>1</sub> PAMs and NAMs from divergent chemical scaffolds for modulation of different mGlu<sub>1</sub> orthosteric agonists in intracellular calcium (iCa<sup>2+</sup>) mobilization and inositol monophosphate (IP<sub>1</sub>) accumulation assays. Operational models of agonism and allosterism were used to derive estimates for important pharmacological parameters such as affinity, efficacy, and cooperativity. Modulation of glutamate and quisqualate-mediated iCa<sup>2+</sup> mobilization revealed probe dependence at the level of affinity and cooperativity for both mGlu<sub>1</sub> PAMs and NAMs. We also identified the previously described mGlu<sub>5</sub> selective NAM PF-06462894 as an mGlu<sub>1</sub> NAM with a different pharmacological profile from other NAMs. Differential profiles were also observed when comparing ligand pharmacology between iCa<sup>2+</sup> mobilization and IP<sub>1</sub> accumulation. The PAMs Ro67-4853 and CPPHA displayed apparent negative cooperativity for modulation of quisqualate affinity, and the NAMs CPCCOEt and PF-06462894 had a marked reduction in cooperativity with quisqualate in IP<sub>1</sub> accumulation and upon extended incubation in iCa<sup>2+</sup> mobilization assays. These data highlight the importance of rigorous assessment of mGlu<sub>1</sub> modulator pharmacology to inform future drug discovery programs for mGlu<sub>1</sub> allosteric modulators. SIGNIFICANCE STATEMENT: Metabotropic glutamate receptor subtype 1 (mGlu<sub>1</sub>) positive and negative allosteric modulators have therapeutic potential in multiple central nervous system disorders. We show that chemically distinct modulators display differential pharmacology with different orthosteric ligands and across divergent signaling pathways at human mGlu<sub>1</sub>. Such complexities in allosteric ligand pharmacology should be considered in future mGlu<sub>1</sub> allosteric drug discovery programs.</p>","PeriodicalId":18767,"journal":{"name":"Molecular Pharmacology","volume":null,"pages":null},"PeriodicalIF":3.2000,"publicationDate":"2023-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"1","resultStr":"{\"title\":\"Rigorous Characterization of Allosteric Modulation of the Human Metabotropic Glutamate Receptor 1 Reveals Probe- and Assay-Dependent Pharmacology.\",\"authors\":\"Ashwin Muraleetharan, Yuyang Wang, Matthew C Rowe, Ashleigh Gould, Karen J Gregory, Shane D Hellyer\",\"doi\":\"10.1124/molpharm.122.000664\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>Allosteric modulation of metabotropic glutamate receptor subtype 1 (mGlu<sub>1</sub>) represents a viable therapeutic target for treating numerous central nervous system disorders. Although multiple chemically distinct mGlu<sub>1</sub> positive (PAMs) and negative (NAMs) allosteric modulators have been identified, drug discovery paradigms have not included rigorous pharmacological analysis. In the present study, we hypothesized that existing mGlu<sub>1</sub> allosteric modulators possess unappreciated probe-dependent or biased pharmacology. Using human embryonic kidney 293 (HEK293A) cells stably expressing human mGlu<sub>1,</sub> we screened mGlu<sub>1</sub> PAMs and NAMs from divergent chemical scaffolds for modulation of different mGlu<sub>1</sub> orthosteric agonists in intracellular calcium (iCa<sup>2+</sup>) mobilization and inositol monophosphate (IP<sub>1</sub>) accumulation assays. Operational models of agonism and allosterism were used to derive estimates for important pharmacological parameters such as affinity, efficacy, and cooperativity. Modulation of glutamate and quisqualate-mediated iCa<sup>2+</sup> mobilization revealed probe dependence at the level of affinity and cooperativity for both mGlu<sub>1</sub> PAMs and NAMs. We also identified the previously described mGlu<sub>5</sub> selective NAM PF-06462894 as an mGlu<sub>1</sub> NAM with a different pharmacological profile from other NAMs. Differential profiles were also observed when comparing ligand pharmacology between iCa<sup>2+</sup> mobilization and IP<sub>1</sub> accumulation. The PAMs Ro67-4853 and CPPHA displayed apparent negative cooperativity for modulation of quisqualate affinity, and the NAMs CPCCOEt and PF-06462894 had a marked reduction in cooperativity with quisqualate in IP<sub>1</sub> accumulation and upon extended incubation in iCa<sup>2+</sup> mobilization assays. These data highlight the importance of rigorous assessment of mGlu<sub>1</sub> modulator pharmacology to inform future drug discovery programs for mGlu<sub>1</sub> allosteric modulators. SIGNIFICANCE STATEMENT: Metabotropic glutamate receptor subtype 1 (mGlu<sub>1</sub>) positive and negative allosteric modulators have therapeutic potential in multiple central nervous system disorders. We show that chemically distinct modulators display differential pharmacology with different orthosteric ligands and across divergent signaling pathways at human mGlu<sub>1</sub>. 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Rigorous Characterization of Allosteric Modulation of the Human Metabotropic Glutamate Receptor 1 Reveals Probe- and Assay-Dependent Pharmacology.
Allosteric modulation of metabotropic glutamate receptor subtype 1 (mGlu1) represents a viable therapeutic target for treating numerous central nervous system disorders. Although multiple chemically distinct mGlu1 positive (PAMs) and negative (NAMs) allosteric modulators have been identified, drug discovery paradigms have not included rigorous pharmacological analysis. In the present study, we hypothesized that existing mGlu1 allosteric modulators possess unappreciated probe-dependent or biased pharmacology. Using human embryonic kidney 293 (HEK293A) cells stably expressing human mGlu1, we screened mGlu1 PAMs and NAMs from divergent chemical scaffolds for modulation of different mGlu1 orthosteric agonists in intracellular calcium (iCa2+) mobilization and inositol monophosphate (IP1) accumulation assays. Operational models of agonism and allosterism were used to derive estimates for important pharmacological parameters such as affinity, efficacy, and cooperativity. Modulation of glutamate and quisqualate-mediated iCa2+ mobilization revealed probe dependence at the level of affinity and cooperativity for both mGlu1 PAMs and NAMs. We also identified the previously described mGlu5 selective NAM PF-06462894 as an mGlu1 NAM with a different pharmacological profile from other NAMs. Differential profiles were also observed when comparing ligand pharmacology between iCa2+ mobilization and IP1 accumulation. The PAMs Ro67-4853 and CPPHA displayed apparent negative cooperativity for modulation of quisqualate affinity, and the NAMs CPCCOEt and PF-06462894 had a marked reduction in cooperativity with quisqualate in IP1 accumulation and upon extended incubation in iCa2+ mobilization assays. These data highlight the importance of rigorous assessment of mGlu1 modulator pharmacology to inform future drug discovery programs for mGlu1 allosteric modulators. SIGNIFICANCE STATEMENT: Metabotropic glutamate receptor subtype 1 (mGlu1) positive and negative allosteric modulators have therapeutic potential in multiple central nervous system disorders. We show that chemically distinct modulators display differential pharmacology with different orthosteric ligands and across divergent signaling pathways at human mGlu1. Such complexities in allosteric ligand pharmacology should be considered in future mGlu1 allosteric drug discovery programs.
期刊介绍:
Molecular Pharmacology publishes findings derived from the application of innovative structural biology, biochemistry, biophysics, physiology, genetics, and molecular biology to basic pharmacological problems that provide mechanistic insights that are broadly important for the fields of pharmacology and toxicology. Relevant topics include:
Molecular Signaling / Mechanism of Drug Action
Chemical Biology / Drug Discovery
Structure of Drug-Receptor Complex
Systems Analysis of Drug Action
Drug Transport / Metabolism