title: Characterization of the interaction of the novel antihypertensive etamicastat with human dopamine-β-hydroxylase: comparison with nepicastat.
abstract: The interaction of etamicastat, a novel peripherally acting dopamine-β-hydroxylase (DBH) inhibitor, with the enzyme was studied using a classical kinetic approach and the pharmacodynamics effect of the compound upon administration to rats was also evaluated. SK-N-SH cell homogenates convert tyramine into octopamine with a Km value of 9 mM, and a Vmax of 1747 nmol/mg protein/h. The K(m) value for ascorbate was 3 mM. The inhibition of DBH by etamicastat and nepicastat, a known centrally acting DBH inhibitor, with IC50 values of 107 and 40 nM, respectively, was fully reversed by dilution. Non-linear fitting of the velocities, determined at various concentrations of substrate (tyramine) and co-substrate (ascorbic acid), and of etamicastat and nepicastat, indicated that the inhibition of DBH by both compounds follows a mixed-model inhibition mechanism, approaching competitive behavior with regards to the substrate tyramine, with K(i) values of 34 and 11 nM, respectively. Relatively to ascorbate, both compounds followed a mixed-model inhibition mechanism, approaching uncompetitive behavior. Oral administration of both compounds (at 30 mg/kg) inhibited adrenal DBH activity over time and significantly decreased noradrenaline levels in the heart. Nepicastat also decreased noradrenaline levels in the parietal cortex, but not etamicastat. Both compounds significantly decreased systolic and diastolic blood pressure in spontaneously hypertensive rats. In conclusion, etamicastat and nepicastat behave as multisubstrate DBH inhibitors, binding reversibly and preferentially to the reduced form of the enzyme, and simultaneously at the substrate and oxygen binding sites. Etamicastat, in contrast to nepicastat, offers the advantage of peripheral selectivity without central effects.
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Maria João
authors: MJ
authors: Bonifácio
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Filipa
authors: F
authors: Sousa
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Marco
authors: M
authors: Neves
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Nuno
authors: N
authors: Palma
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Bruno
authors: B
authors: Igreja
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Nuno Miguel
authors: NM
authors: Pires
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal.
authors: Lyndon C
authors: LC
authors: Wright
authors: Department of Research & Development, BIAL, 4745-457 São Mamede do Coronado, Portugal; Department of Pharmacology & Therapeutics, Faculty of Medicine, University Porto, Porto, Portugal; MedInUP - Center for Drug Discovery and Innovative Medicines, University of Porto, Porto, Portugal. Electronic address: psoares.silva@bial.com.
authors: Patrício
authors: P
authors: Soares-da-Silva
mesh_terms: D000311:Adrenal Glands / Q000187:drug effects / Q000201:enzymology; D000818:Animals; D000959:Antihypertensive Agents / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D001578:Benzopyrans / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D002460:Cell Line; D004299:Dopamine beta-Hydroxylase / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007093:Imidazoles / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D011918:Rats, Inbred SHR; D017208:Rats, Wistar; D013871:Thiones / Q000378:metabolism*
chemical_list: D000959:Antihypertensive Agents; D001578:Benzopyrans; D004791:Enzyme Inhibitors; D007093:Imidazoles; D013871:Thiones; C509002:etamicastat; D004299:Dopamine beta-Hydroxylase; C109487:nepicastat
keywords: Dopamine beta hydroxylase; Enzyme kinetics; Etamicastat; Inhibition mechanism; Nepicastat; Noradrenaline