RESUMO
We report here the first orally active, selenium-based antihypertensive agent, and we demonstrate its restricted CNS permeability using inductively coupled plasma/mass spectroscopy (ICP/MS) and operant behavioral analysis. The biochemistry and pharmacology of selenium are subjects of intense current interest. As a consequence of the redox chemistry of the selenium moiety, phenylaminoalkyl selenides possess the remarkable characteristic of propagating a cycle of turnover-dependent local depletion of reduced ascorbate when processed by the key enzyme of catecholamine metabolism, dopamine-beta-monooxygenase. ICP/MS analysis was used to determine the pharmacokinetic parameters for selenide compounds after i.v. administration to anesthetized rats. Analysis of the data using a two-compartment pharmacokinetic model established very rapid initial clearance and a short beta-elimination half-life from blood. We developed an oxidative procedure for digestion and processing of tissue samples in order to obtain ICP/MS data on the tissue distributions of Se-containing metabolites after the administration of selenide compounds. The results establish that aromatic ring hydroxylation of the selenides results in a marked reduction in brain levels of Se-containing metabolites. The comparative effects of selenide compounds on locomotor activity and operant behavior were then investigated, and the results fully corroborate the ICP/MS analytical results. The novel compound, 4-hydroxy-alpha-methyl-phenyl-2-aminoethyl selenide, exhibits both restricted CNS permeability and oral antihypertensive activity in spontaneously hypertensive rats. This compound is the first orally active selenium-based antihypertensive agent ever reported, and it possesses properties that are highly desirable in pharmacological agents being developed for treatment of chronic diseases such as hypertension.
Assuntos
Anti-Hipertensivos/farmacocinética , Encéfalo/metabolismo , Compostos Organosselênicos/farmacocinética , Administração Oral , Animais , Anti-Hipertensivos/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Masculino , Camundongos , Compostos Organosselênicos/farmacologia , Permeabilidade , Ratos , Ratos Sprague-Dawley , Ratos Wistar , Relação Estrutura-AtividadeRESUMO
Carboxyl-terminal amidation, a required post-translational modification for the bioactivation of many neuropeptides, entails sequential enzymatic action by peptidylglycine monooxygenase (PAM, EC 1.14.17.3) and peptidylamidoglycolate lyase (PGL, EC 4.3.2.5). The monooxygenase, PAM, first catalyzes conversion of a glycine-extended pro-peptide to the corresponding alpha-hydroxyglycine derivative, and the lyase, PGL, then catalyzes breakdown of this alpha-hydroxyglycine derivative to the amidated peptide plus glyoxylate. We now introduce the first potent inhibitors for peptidylamidoglycolate lyase. These inhibitors, which can be viewed as pyruvate-extended N-acetyl amino acids, constitute a novel class of compounds. They were designed to resemble likely transient species along the reaction pathway of PGL catalysis. A general synthetic procedure for preparation of pyruvate-extended N-acetyl amino acids or peptides is described. Since these compounds possess the 2,4-dioxo-carboxylate moiety, their solution tautomerization was investigated using both NMR and high performance liquid chromatography analyses. The results establish that freshly prepared solutions of N-Ac-Phe-pyruvate consist predominantly of the enol tautomer, which then slowly tautomerizes to the diketo form when left standing for several days in an aqueous medium; upon acidification, formation of the hydrate tautomer occurs. Kinetic experiments established that these novel compounds are highly potent, pure competitive inhibitors of PGL. Kinetic experiments with the ascorbate-dependent copper monooxygenases, PAM and dopamine-beta-monooxygenase, established that these compounds also bind competitively with respect to ascorbate; however, pyruvate-extended N-acyl-amino acid derivatives possessing hydrophobic side chains are much more potent inhibitors of PGL than of PAM. Selective targeting of N-Ac-Phe-pyruvate so as to inhibit the lyase, but not the monooxygenase, domain was demonstrated with the bifunctional amidating enzyme of Xenopus laevis. The availability of potent inhibitors of PGL should facilitate studies regarding the possible biological role of alpha-hydroxyglycine-extended peptides.
Assuntos
Amidas/metabolismo , Amidina-Liases , Liases/metabolismo , Oxigenases de Função Mista/metabolismo , Complexos Multienzimáticos , Peptídeos/metabolismo , Ácido Pirúvico/metabolismo , Animais , Bovinos , Xenopus laevisRESUMO
Dopamine beta-monoxygenase (DBM, E.C. 1.14.17.1) is an attractive target point for possible modulation of adrenergic activity, and a variety of DBM-targeted pseudosubstrates and inhibitors have been developed in this laboratory and other laboratories. We now demonstrate the efficacy of a DBM-targeted mechanism-based inactivator, as well as enzymatic processing of two alternate DBM substrates, within functional adrenal chromaffin cells. When cultured adrenal medullary chromaffin cells were incubated with the mechanism-based inactivator 1-(4'-hydroxyphenyl)-1-(aminomethyl)-ethene (HOPAME), vesicular DBM activity was markedly decreased. Similarly, the alternate substrates 4'-hydroxyphenyl-2-aminoethyl sulfide and 4'-hydroxyphenyl-2-aminopropyl selenide each undergo uptake and DBM-catalyzed oxygenation within these cells. The simultaneous action of both the mechanism-based inactivator and an alternate substrate within functional chromaffin cells was also demonstrated. These results provide support for a direct mechanistic link between the enzymological properties of DBM-targeted adrenergic agents and their in-vivo pharmacological activities.