Identification of global and ligand-specific calcium sensing receptor activation mechanisms S

Andrew Keller, Irina Kufareva, Tracy M Josephs, Jiayin Diao, Vyvyan Tuong-Vy Mai, Arthur D. Conigrave, Arthur Christopoulos, Karen J Gregory, Katie Leach

Research output: Contribution to journalArticleResearchpeer-review

Abstract

Calcium sensing receptor (CaSR) positive allosteric modulators (PAMs) are therapeutically important. However, few are approved for clinical use, in part due to complexities in assessing allostery at a receptor where the endogenous agonist (extracellular calcium) is present in all biologic fluids. Such complexity impedes efforts to quantify and optimize allosteric drug parameters (affinity, cooperativity, and efficacy) that dictate PAM structure-activity relationships (SARs). Furthermore, an underappreciation of the structural mechanisms underlying CaSR activation hinders predictions of how PAM SAR relates to in vitro and in vivo activity. Herein, we combined site-directed mutagenesis and calcium mobilization assays with analytical pharmacology to compare modes of PAM binding, positive modulation, and agonism. We demonstrate that 3-(2-chlorophenyl)-N-((1R)-1-(3-methoxyphenyl)ethyl)-1-propanamine (NPS R568) binds to a 7 transmembrane domain (7TM) cavity common to class C G protein-coupled receptors and used by (aR)-(2)-a-methyl-N-[3-[3-[trifluoromethylphenyl]propyl]-1-napthalenemethanamine (cinacalcet) and 1-benzothiazol-2-yl-1-(2,4-dimethylphenyl)-ethanol (AC265347); however, there are subtle distinctions in the contribution of select residues to the binding and transmission of cooperativity by PAMs. Furthermore, we reveal some common activation mechanisms used by different CaSR activators, but also demonstrate some differential contributions of residues within the 7TM bundle and extracellular loops to the efficacy of the PAM-agonist, AC265347, versus cooperativity. Finally, we show that PAMS potentiate the affinity of divalent cations. Our results support the existence of both global and ligand-specific CaSR activation mechanisms and reveal that allosteric agonism is mediated in part via distinct mechanisms to positive modulation.

Original languageEnglish
Pages (from-to)619-630
Number of pages12
JournalMolecular Pharmacology
Volume93
Issue number6
DOIs
Publication statusPublished - 1 Jun 2018

Cite this

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title = "Identification of global and ligand-specific calcium sensing receptor activation mechanisms S",
abstract = "Calcium sensing receptor (CaSR) positive allosteric modulators (PAMs) are therapeutically important. However, few are approved for clinical use, in part due to complexities in assessing allostery at a receptor where the endogenous agonist (extracellular calcium) is present in all biologic fluids. Such complexity impedes efforts to quantify and optimize allosteric drug parameters (affinity, cooperativity, and efficacy) that dictate PAM structure-activity relationships (SARs). Furthermore, an underappreciation of the structural mechanisms underlying CaSR activation hinders predictions of how PAM SAR relates to in vitro and in vivo activity. Herein, we combined site-directed mutagenesis and calcium mobilization assays with analytical pharmacology to compare modes of PAM binding, positive modulation, and agonism. We demonstrate that 3-(2-chlorophenyl)-N-((1R)-1-(3-methoxyphenyl)ethyl)-1-propanamine (NPS R568) binds to a 7 transmembrane domain (7TM) cavity common to class C G protein-coupled receptors and used by (aR)-(2)-a-methyl-N-[3-[3-[trifluoromethylphenyl]propyl]-1-napthalenemethanamine (cinacalcet) and 1-benzothiazol-2-yl-1-(2,4-dimethylphenyl)-ethanol (AC265347); however, there are subtle distinctions in the contribution of select residues to the binding and transmission of cooperativity by PAMs. Furthermore, we reveal some common activation mechanisms used by different CaSR activators, but also demonstrate some differential contributions of residues within the 7TM bundle and extracellular loops to the efficacy of the PAM-agonist, AC265347, versus cooperativity. Finally, we show that PAMS potentiate the affinity of divalent cations. Our results support the existence of both global and ligand-specific CaSR activation mechanisms and reveal that allosteric agonism is mediated in part via distinct mechanisms to positive modulation.",
author = "Andrew Keller and Irina Kufareva and Josephs, {Tracy M} and Jiayin Diao and Mai, {Vyvyan Tuong-Vy} and Conigrave, {Arthur D.} and Arthur Christopoulos and Gregory, {Karen J} and Katie Leach",
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Identification of global and ligand-specific calcium sensing receptor activation mechanisms S. / Keller, Andrew; Kufareva, Irina; Josephs, Tracy M; Diao, Jiayin; Mai, Vyvyan Tuong-Vy; Conigrave, Arthur D.; Christopoulos, Arthur; Gregory, Karen J; Leach, Katie.

In: Molecular Pharmacology, Vol. 93, No. 6, 01.06.2018, p. 619-630.

Research output: Contribution to journalArticleResearchpeer-review

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AU - Kufareva, Irina

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AU - Diao, Jiayin

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AU - Leach, Katie

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