Identification of a series of CCK-2 receptor nonpeptide agonists: Sensitivity to stereochemistry and a receptor point mutation

Kopin, Alan S.; McBride, Edward W.; Chen, Ci; Freidinger, Roger; Chen, Duan; Zhao, Chun‐Mei; Beinborn, Martin

doi:10.1073/pnas.0831223100

Cited by 17 publications

(20 citation statements)

References 28 publications

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“…As originally demonstrated for the CCK-2R L325E mutant, constitutively active receptors provide a magnifying glass of ligand function (Beinborn et al, 1998b) and therefore have considerable utility in discovering agonist lead compounds Kopin, 2003). Once identified, these hits can then be further optimized into clinically useful drugs.…”

Section: Ligand Function At Constitutively Active Receptor Mutants 757mentioning

confidence: 99%

“…CCK-2R-modulated processes include gastric acid secretion and mucosal growth, as well as the perception of pain. Given the longstanding interest in the development of drugs that target the CCK-2R, a broad range of nonpeptide ligands, including agonists, antagonists, and inverse agonists, have been identified (Beinborn et al, 1998b;de Tullio et al, 2000;Kopin et al, 2003). Using selected compounds, we have previously demonstrated that a prototype constitutively active mutant of the CCK-2R (L325E) shows a systematic increase in ligand affinities and efficacies versus respective values at the wild-type receptor (Beinborn et al, 1998b).…”

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confidence: 99%

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Ligand Function at Constitutively Active Receptor Mutants Is Affected by Two Distinct Yet Interacting Mechanisms

Beinborn

Ren²,

Bläker³

et al. 2004

Molecular Pharmacology

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It has been proposed that mutations that induce constitutive activity in G-protein-coupled receptors (GPCRs) concomitantly enhance the ability of partial agonists to trigger second-messenger signaling. Using the cholecystokinin type 2 receptor (CCK-2R) as a model system, we have explored whether this association applies to a diverse set of activating mutations. Consistent with established principles, constitutively active CCK-2Rs resulting from amino acid substitutions within the third intracellular loop each systematically increased partial agonist activities versus corresponding wild-type values. In contrast, activating mutations within transmembrane domain segments near the extracellular loops led to an increase in efficacy of only a subset of compounds but decreased or did not change the function of others. When transmembrane domain amino acid substitutions were introduced in combination with intracellular amplifying mutations, observed changes in ligand activity were defined by the product of two discernible factors 1) systematic amplification caused by an equilibrium shift from the inactive to the active receptor conformation and 2) ligand-specific alterations in signaling, which probably result from mutation-induced changes in the putative binding pocket. These findings illustrate functional heterogeneity among GPCR mutants with ligand-independent signaling. A subgroup of activating mutations facilitates receptor isomerization to the active state and in parallel perturbs ligand receptor interactions. These mutants do not adhere to the previously proposed "hallmark criteria" of constitutive activity.

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Section: Ligand Function At Constitutively Active Receptor Mutants 757mentioning

confidence: 99%

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confidence: 99%

Ligand Function at Constitutively Active Receptor Mutants Is Affected by Two Distinct Yet Interacting Mechanisms

Beinborn

Ren²,

Bläker³

et al. 2004

Molecular Pharmacology

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“…Mutation of nonconserved amino acids from transmembrane helices led to the identification of a single amino acid in the canine sequence (Leu355, TM VI) that is responsible for this reversal of specificity (38). In general, studies with so-called nonpeptide antagonists of CCK2R revealed that although the amino acid sequence homology of CCK2R in the different species is near 90%, the efficacy of the nonpeptide molecules to stimulate phospholipase C varied from 0 to 60% of the CCK-induced maximal response according to the species and the compound tested (L365,260, L740,093, YM022, PD135158, PD136,450, PD134,308) (39, 47, 48, 227,232,255,256). Incorporation of nonconservative amino acids in the human CCK2R sequence enabled identification of amino acids in transmembrane helices that account for these variations (47, 48).…”

Section: A Localization Of Ligand Binding Sitesmentioning

confidence: 99%

Cholecystokinin and Gastrin Receptors

Dufresne

Seva²,

Fourmy³

2006

Physiological Reviews

422

418

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Cholecystokinin and gastrin receptors (CCK1R and CCK2R) are G protein-coupled receptors that have been the subject of intensive research in the last 10 years with corresponding advances in the understanding of their functioning and physiology. In this review, we first describe general properties of the receptors, such as the different signaling pathways used to exert short- and long-term effects and the structural data that explain their binding properties, activation, and regulation. We then focus on peripheral cholecystokinin receptors by describing their tissue distribution and physiological actions. Finally, pathophysiological peripheral actions of cholecystokinin receptors and their relevance in clinical disorders are reviewed.

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“…(Table 3). Given some rare precedents for mutation-induced conversion of antagonists to agonists (Strader et al, 1989;Blaker et al, 1998;Kopin et al, 2003), we also examined SNAP-7941 activity on the R210H and P377S variants. Experiments revealed that this compound did not induce signaling at either of the mutants (not shown).…”

Section: Resultsmentioning

confidence: 99%

Two Naturally Occurring Mutations in the Type 1 Melanin-Concentrating Hormone Receptor Abolish Agonist-Induced Signaling

Goldstein¹,

Schroeder²,

Fortin³

et al. 2010

The Journal of Pharmacology and Experimental Therapeutics

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The melanin-concentrating hormone (MCH) receptor type 1 (MCHR1) is a seven-transmembrane domain protein that modulates orexigenic activity of MCH, the corresponding endogenous peptide agonist. MCH antagonists are being explored as a potential treatment for obesity. In the current study, we examined the pharmacological impact of 11 naturally occurring mutations in the human MCHR1. Wild-type and mutant receptors were transiently expressed in human embryonic kidney 293 cells. MCHR1-mediated, G␣ i -dependent signaling was monitored by using luciferase reporter gene assays. Two mutants, R210H and P377S, failed to respond to MCH. Five other variants showed significant alterations in MCH efficacy, ranging from 44 to 142% of the wild-type value. At each of the MCH-responsive mutants, agonist potency and inhibition by (S)-methyl 3-((3-(4-(3-acetamidophenyl) piperidin-1-yl)propyl)carbamoyl)-4-(3,4-difluorophenyl)-6-(methoxymethyl)-2-oxo-1,2,3,4-tetrahydropyrimidine-5-carboxylate (SNAP-7941), an established MCHR1 small-molecule antagonist, were similar to wild type. To explore the basis for inactivity of the R210H and P377S mutants, we examined expression levels of these receptors. Assessment by enzyme-linked immunosorbent assay revealed that cell surface expression of both nonfunctional receptors was comparable with wild type. Overnight treatment with SNAP-7941, followed by washout of antagonist, enhanced MCH induced signaling by the wild-type receptor and restored MCH responsiveness of the P377S but not the R210H variant. It is of note that the two loss-of-function mutants were identified in markedly underweight individuals, raising the possibility that a lean phenotype may be linked to deficient MCHR1 signaling. Formal association studies with larger cohorts are needed to explore the extent to which signaling-deficient MCHR1 variants influence the maintenance of body weight.

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Identification of a series of CCK-2 receptor nonpeptide agonists: Sensitivity to stereochemistry and a receptor point mutation

Cited by 17 publications

References 28 publications

Ligand Function at Constitutively Active Receptor Mutants Is Affected by Two Distinct Yet Interacting Mechanisms

Ligand Function at Constitutively Active Receptor Mutants Is Affected by Two Distinct Yet Interacting Mechanisms

Cholecystokinin and Gastrin Receptors

Two Naturally Occurring Mutations in the Type 1 Melanin-Concentrating Hormone Receptor Abolish Agonist-Induced Signaling

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