The Experts below are selected from a list of 297 Experts worldwide ranked by ideXlab platform
Eric L Barker - One of the best experts on this subject based on the ideXlab platform.
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comparative molecular field analysis using selectivity fields reveals residues in the third transmembrane helix of the serotonin transporter associated with substrate and Antagonist recognition
Journal of Pharmacology and Experimental Therapeutics, 2008Co-Authors: Crystal C Walline, David E Nichols, Ivy F Carroll, Eric L BarkerAbstract:The human serotonin transporter (hSERT) regulates the spatial and temporal actions of serotonin (5-HT) neurotransmission by removing 5-HT from the synapse. Previous studies have identified residues in the third transmembrane helix (TMH) that may be important for substrate translocation or Antagonist recognition. We identified hSERT residues in TMH III that are divergent from Drosophila SERT and used species-scanning mutagenesis to generate reciprocal mutants. Transport inhibition assays suggest that the Potency of substituted amphetamines was decreased for the hSERT mutants A169D, I172M, and S174M. In addition, there was a loss of Potency for several antidepressants and 3-phenyltropane analogs for the I172M mutant. These results suggest that residues in TMH III may contribute to Antagonist recognition. We carried out comparative molecular field analyses using selectivity fields to directly visualize the mutation-induced effects of Antagonist Potency for antidepressants, 3-phenyltropane analogs, and amphetamines. The hSERT I172M selectivity field analysis for the 3-phenyltropane analogs revealed that electrostatic interactions resulted in decreased Potency. The amphetamine and antidepressant selectivity field analyses reveal the observed decreases in potencies for the hSERT I172M mutant are due to a change in tertiary structure of the hSERT protein and are not due to disruption of a direct binding site. Finally, the hSERT mutant A169D displayed altered kinetics for sodium binding, indicating that this residue may lie near the putative sodium binding site. A SERT homology model developed from the Aquifex aeolicus leucine transporter structure provides a structural context for further interpreting the results of the TMH III mutations.
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high affinity recognition of serotonin transporter Antagonists defined by species scanning mutagenesis an aromatic residue in transmembrane domain i dictates species selective recognition of citalopram and mazindol
Journal of Biological Chemistry, 1998Co-Authors: Eric L Barker, Melody A Perlman, Erika M Adkins, Zdenek B Pristupa, William J. Houlihan, Hyman B Niznik, Randy D BlakelyAbstract:Abstract Human and Drosophila melanogasterserotonin (5-HT) transporters (SERTs) exhibit similar 5-HT transport kinetics and can be distinguished pharmacologically by many, but not all, biogenic amine transporter Antagonists. By using human andDrosophila SERT chimeras, major determinants of potencies of two transporter Antagonists, mazindol and citalopram, were tracked to the amino-terminal domains encompassing transmembrane domains I and II. Species-scanning mutagenesis, whereby amino acid substitutions are made switching residues from one species to another, was employed on the eight amino acids that differ between human andDrosophila SERTs in this region, and Antagonist potencies were reassessed in 5-HT uptake assays. A single mutation in transmembrane domain I of human SERT, Y95F, shifted both citalopram and mazindol to Drosophila SERT-like potencies. Strikingly, these Potency changes were in opposite directions suggesting Tyr95 contributes both positive and negative determinants of Antagonist Potency. To gain insight into how the Y95F mutant might influence mazindol Potency, we determined how structural variants of mazindol responded to the mutation. Our studies demonstrate the importance of the hydroxyl group on the heterocyclic nucleus of mazindol for maintaining species-selective recognition of mazindol and suggest that transmembrane domain I participates in the formation of Antagonist-binding sites for amine transporters.
T J Opgenorth - One of the best experts on this subject based on the ideXlab platform.
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endothelin receptors in human smooth muscle cells Antagonist Potency differs on agonist evoked responses
American Journal of Physiology-cell Physiology, 1994Co-Authors: J R Wuwong, W J Chiou, Zeijing Huang, M J Vidal, T J OpgenorthAbstract:The present study characterized endothelin (ET) receptors in human pericardium smooth muscle cells (HPSMC) and examined the Potency of Antagonists on ET-evoked signal transduction and DNA synthesis. HPSMC contain both ETA and ETB receptors. ET-1 binding was completely inhibited by a nonselective Antagonist (Ro-46-2005) or a combination of ETA-selective and ETB-selective ligands (BQ-123 and ET-3). The molecular masses for ETA and ETB receptors were 69 and 42 kDa, respectively. ET-1, but not ET-3, stimulated phosphatidylinositol hydrolysis and arachidonic acid release in a dose- and time-dependent manner, reaching a plateau within 20-40 min. These immediate effects of ET-1 on signal transduction were completely inhibited by 1 microM, BQ-123, ET-1, but not ET-3, stimulated DNA synthesis in a dose-dependent manner, and the effect became prominent after 24 h. BQ-123 (1 microM) or Ro-46-2005 (10 microM) did not completely inhibit this mitogenic effect of ET-1. The reduced Potency of BQ-123 and Ro-46-2005 on the delayed effect of ET-1 was not the result of ligand degradation or a difference in receptor internalization; rather, the decrease in Potency was due to the fact that Antagonist binding was more reversible than ET-1 binding.
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endothelin receptors in human smooth muscle cells Antagonist Potency differs on agonist evoked responses
American Journal of Physiology-cell Physiology, 1994Co-Authors: J R Wuwong, W J Chiou, Zeijing Huang, M J Vidal, T J OpgenorthAbstract:The present study characterized endothelin (ET) receptors in human pericardium smooth muscle cells (HPSMC) and examined the Potency of Antagonists on ET-evoked signal transduction and DNA synthesis...
Helmut R Maecke - One of the best experts on this subject based on the ideXlab platform.
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Evaluation of Three Different Families of Bombesin Receptor RadioAntagonists for Targeted Imaging and Therapy of Gastrin Releasing Peptide Receptor (GRP-R) Positive Tumors
2015Co-Authors: Rosalba Mansi, Xuejuan Wang, Renzo Cescato, Jean Claude Reubi, Keelara Abiraj, Maria Luisa Tamma, Eleni Gourni, Sandra Berndt, Helmut R MaeckeAbstract:Two new classes of radiolabeled GRP receptor Antagonists are studied and compared with the well-established statine-based receptor Antagonist DOTA-4-amino-1-carboxymethylpiperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2 (RM2, 1; DOTA:1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid; Sta:(3S,4S)-4-amino-3-hydroxy-6-methylheptanoic acid). The bombesin-based pseudopeptide DOTA-4-amino-1-carboxymethylpiperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Leuψ(CHOH-CH2)-(CH2)2-CH3 (RM7, 2), and the methyl ester DOTA-4-amino-1-carboxymethylpiperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-OCH3 (ARBA05, 3) analogues are labeled with 111In and evaluated in vitro in PC-3 cell line and in vivo in PC-3 tumor-bearing nude mice. Antagonist Potency was assessed by immunofluorescence-based receptor internalization and Ca2+ mobilization assays. The conjugates showed good binding affinity, the IC50 value of 2 (3.2 ± 1.8 nM) being 2 and 10 times lower than 1 and 3. Compared to 111In-1, 111In-2 showed higher uptake in target tissues such as pancreas (1.5 ± 0.5%IA/g and 39.8 ± 9.3%IA/g at 4 h, respectively), whereas the compounds had similar tumor uptake (11.5 ± 2.4%IA/g and 11.8 ± 3.9%IA/g at 4h, respectively). The displacement of the radioligand in vivo was different in different receptor positive organs and depended on the displacing peptide
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Evaluation of three different families of bombesin receptor radioAntagonists for targeted imaging and therapy of gastrin releasing peptide receptor (GRP-R) positive tumors.
Journal of medicinal chemistry, 2014Co-Authors: Rosalba Mansi, Xuejuan Wang, Marialuisa Tamma, Renzo Cescato, Jean Claude Reubi, Keelara Abiraj, Eleni Gourni, Sandra Berndt, Helmut R MaeckeAbstract:Two new classes of radiolabeled GRP receptor Antagonists are studied and compared with the well-established statine-based receptor Antagonist DOTA-4-amino-1-carboxymethylpiperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2 (RM2, 1; DOTA:1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid; Sta:(3S,4S)-4-amino-3-hydroxy-6-methylheptanoic acid). The bombesin-based pseudopeptide DOTA-4-amino-1-carboxymethylpiperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Leuψ(CHOH-CH2)-(CH2)2-CH3 (RM7, 2), and the methyl ester DOTA-4-amino-1-carboxymethylpiperidine-d-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-OCH3 (ARBA05, 3) analogues are labeled with (111)In and evaluated in vitro in PC-3 cell line and in vivo in PC-3 tumor-bearing nude mice. Antagonist Potency was assessed by immunofluorescence-based receptor internalization and Ca(2+) mobilization assays. The conjugates showed good binding affinity, the IC50 value of 2 (3.2 ± 1.8 nM) being 2 and 10 times lower than 1 and 3. Compared to (111)In-1, (111)In-2 showed higher uptake in target tissues such as pancreas (1.5 ± 0.5%IA/g and 39.8 ± 9.3%IA/g at 4 h, respectively), whereas the compounds had similar tumor uptake (11.5 ± 2.4%IA/g and 11.8 ± 3.9%IA/g at 4h, respectively). The displacement of the radioligand in vivo was different in different receptor positive organs and depended on the displacing peptide.
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evaluation of a 1 4 7 10 tetraazacyclododecane 1 4 7 10 tetraacetic acid conjugated bombesin based radioAntagonist for the labeling with single photon emission computed tomography positron emission tomography and therapeutic radionuclides
Clinical Cancer Research, 2009Co-Authors: Rosalba Mansi, Xuejuan Wang, Flavio Forrer, Stefan Kneifel, Marialuisa Tamma, Beatrice Waser, Renzo Cescato, Jean Claude Reubi, Helmut R MaeckeAbstract:Purpose: G protein–coupled receptor agonists are being used as radiolabeled vectors for in vivo localization and therapy of tumors. Recently, somatostatin-based Antagonists were shown to be superior to agonists. Here, we compare the new [ 111 In/ 68 Ga]-labeled bombesin-based Antagonist RM1 with the agonist [ 111 In]-AMBA for targeting the gastrin-releasing peptide receptor (GRPR). Experimental Design: IC 50 , K d values, and Antagonist Potency were determined using PC-3 and HEK-GRPR cells. Biodistribution and imaging studies were done in nude mice transplanted with the PC-3 tumor. The Antagonist Potency was assessed by evaluating the effects on calcium release and on receptor internalization monitored by immunofluorescence microscopy. Results: The IC 50 value of [ nat In]-RM1 was 14 ± 3.4 nmol/L. [ nat/111 In]-RM1 was found to bind to the GRPR with a K d of 8.5 ± 2.7 nmol/L compared with a K d of 0.6 ± 0.3 nmol/L of [ 111 In]-AMBA. A higher maximum number of binding site value was observed for [ 111 In]-RM1 (2.4 ± 0.2 nmol/L) compared with [ 111 In]-AMBA (0.7 ± 0.1 nmol/L). [ nat Lu]-AMBA is a potent agonist in the immunofluorescence-based internalization assay, whereas [ nat In]-RM1 is inactive alone but efficiently antagonizes the bombesin effect. These data are confirmed by the calcium release assay. The pharmacokinetics showed a superiority of the radioAntagonist with regard to the high tumor uptake (13.4 ± 0.8% IA/g versus 3.69 ± 0.75% IA/g at 4 hours after injection. as well as to all tumor-to-normal tissue ratios. Conclusion: Despite their relatively low GRPR affinity, the Antagonists [ 111 In/ 68 Ga]-RM1 showed superior targeting properties compared with [ 111 In]-AMBA. As found for somatostatin receptor–targeting radiopeptides, GRP-based radioAntagonists seem to be superior to radioagonists for in vivo imaging and potentially also for targeted radiotherapy of GRPR-positive tumors. (Clin Cancer Res 2009;15(16):5240–9)
J R Wuwong - One of the best experts on this subject based on the ideXlab platform.
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endothelin receptors in human smooth muscle cells Antagonist Potency differs on agonist evoked responses
American Journal of Physiology-cell Physiology, 1994Co-Authors: J R Wuwong, W J Chiou, Zeijing Huang, M J Vidal, T J OpgenorthAbstract:The present study characterized endothelin (ET) receptors in human pericardium smooth muscle cells (HPSMC) and examined the Potency of Antagonists on ET-evoked signal transduction and DNA synthesis. HPSMC contain both ETA and ETB receptors. ET-1 binding was completely inhibited by a nonselective Antagonist (Ro-46-2005) or a combination of ETA-selective and ETB-selective ligands (BQ-123 and ET-3). The molecular masses for ETA and ETB receptors were 69 and 42 kDa, respectively. ET-1, but not ET-3, stimulated phosphatidylinositol hydrolysis and arachidonic acid release in a dose- and time-dependent manner, reaching a plateau within 20-40 min. These immediate effects of ET-1 on signal transduction were completely inhibited by 1 microM, BQ-123, ET-1, but not ET-3, stimulated DNA synthesis in a dose-dependent manner, and the effect became prominent after 24 h. BQ-123 (1 microM) or Ro-46-2005 (10 microM) did not completely inhibit this mitogenic effect of ET-1. The reduced Potency of BQ-123 and Ro-46-2005 on the delayed effect of ET-1 was not the result of ligand degradation or a difference in receptor internalization; rather, the decrease in Potency was due to the fact that Antagonist binding was more reversible than ET-1 binding.
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endothelin receptors in human smooth muscle cells Antagonist Potency differs on agonist evoked responses
American Journal of Physiology-cell Physiology, 1994Co-Authors: J R Wuwong, W J Chiou, Zeijing Huang, M J Vidal, T J OpgenorthAbstract:The present study characterized endothelin (ET) receptors in human pericardium smooth muscle cells (HPSMC) and examined the Potency of Antagonists on ET-evoked signal transduction and DNA synthesis...
Randy D Blakely - One of the best experts on this subject based on the ideXlab platform.
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high affinity recognition of serotonin transporter Antagonists defined by species scanning mutagenesis an aromatic residue in transmembrane domain i dictates species selective recognition of citalopram and mazindol
Journal of Biological Chemistry, 1998Co-Authors: Eric L Barker, Melody A Perlman, Erika M Adkins, Zdenek B Pristupa, William J. Houlihan, Hyman B Niznik, Randy D BlakelyAbstract:Abstract Human and Drosophila melanogasterserotonin (5-HT) transporters (SERTs) exhibit similar 5-HT transport kinetics and can be distinguished pharmacologically by many, but not all, biogenic amine transporter Antagonists. By using human andDrosophila SERT chimeras, major determinants of potencies of two transporter Antagonists, mazindol and citalopram, were tracked to the amino-terminal domains encompassing transmembrane domains I and II. Species-scanning mutagenesis, whereby amino acid substitutions are made switching residues from one species to another, was employed on the eight amino acids that differ between human andDrosophila SERTs in this region, and Antagonist potencies were reassessed in 5-HT uptake assays. A single mutation in transmembrane domain I of human SERT, Y95F, shifted both citalopram and mazindol to Drosophila SERT-like potencies. Strikingly, these Potency changes were in opposite directions suggesting Tyr95 contributes both positive and negative determinants of Antagonist Potency. To gain insight into how the Y95F mutant might influence mazindol Potency, we determined how structural variants of mazindol responded to the mutation. Our studies demonstrate the importance of the hydroxyl group on the heterocyclic nucleus of mazindol for maintaining species-selective recognition of mazindol and suggest that transmembrane domain I participates in the formation of Antagonist-binding sites for amine transporters.