RESUMO
The aim of the study was to investigate the hepatoprotective properties of the conjugate of the xymedon drug substance with succinic acid (1a). The study presents an in vitro comparative evaluation of the cytotoxicity and cytoprotective properties of 1a and succinic acid on a cell line of normal human hepatocytes Chang Liver, and in vivo investigation of the ability of 1a to restore liver from the toxic damage caused by CCl4 in Wistar rats. It was shown that the cytotoxicity of 1a was 19.9 ± 0.8 mmol/L, and that of succinic acid was 14.1 ± 0.2 mmol/L. Against the background of d-galactosamine exposure, the cytoprotective effect of 1a was found to be superior to that of succinic acid. It was shown that 1a caused a significant reduction in necrotic and steatosis changes in the liver and restoration of biochemical markers of cytolysis, as well as bilirubin metabolism and synthetic liver function.
Assuntos
Hepatócitos/patologia , Fígado/metabolismo , Pirimidinas , Ácido Succínico , Animais , Linhagem Celular , Citoproteção/efeitos dos fármacos , Humanos , Masculino , Pirimidinas/química , Pirimidinas/farmacologia , Ratos , Ratos Wistar , Ácido Succínico/química , Ácido Succínico/farmacologiaRESUMO
INTRODUCTION: The aim of this study was to evaluate the effects of adenosine 5'-triphosphate (ATP) and adenosine on the contractility of mammalian skeletal muscle under hypothermic conditions. METHODS: Contractions of isolated rat soleus muscle were induced by either electrical stimulation (ES) or carbachol at physiological temperatures (37°C) and hypothermic conditions (30-14°C) and recorded in the presence of ATP, adenosine, suramin, and 8-(p-sulfophenyl)-theophylline (8-SPT). RESULTS: At 37°C, incubation of the muscles with ATP inhibited ES-induced contractions; the inhibitory effect of ATP disappeared at 14°C. Adenosine inhibited ES-induced contractions at all temperature levels; 8-SPT fully prevented the action of adenosine. ATP and adenosine did not significantly affect carbachol-induced contractions at 37°C, while at lower temperatures ATP potentiated them. Suramin fully prevented effects of ATP. CONCLUSIONS: ATP is involved in both pre- and postsynaptic regulation of rat soleus muscle contractility, and these processes are significantly more pronounced at low temperatures. Muscle Nerve 55: 417-423, 2017.
Assuntos
Trifosfato de Adenosina/farmacologia , Adenosina/farmacologia , Contração Muscular/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/fisiologia , Temperatura , Análise de Variância , Animais , Carbacol/farmacologia , Agonistas Colinérgicos/farmacologia , Estimulação Elétrica , Hipotermia/induzido quimicamente , Masculino , Antagonistas Nicotínicos/farmacologia , Antagonistas de Receptores Purinérgicos P1/farmacologia , Antagonistas do Receptor Purinérgico P2/farmacologia , Ratos , Ratos Wistar , Suramina/farmacologia , Teofilina/análogos & derivados , Teofilina/farmacologia , Tubocurarina/farmacologiaRESUMO
Inhibition of human AChE (acetylcholinesterase) and BChE (butyrylcholinesterase) by an alkylammonium derivative of 6-methyluracil, C-547, a potential drug for the treatment of MG (myasthenia gravis) was studied. Kinetic analysis of AChE inhibition showed that C-547 is a slow-binding inhibitor of type B, i.e. after formation of the initial enzyme·inhibitor complex (Ki=140 pM), an induced-fit step allows establishment of the final complex (Ki*=22 pM). The estimated koff is low, 0.05 min(-1) On the other hand, reversible inhibition of human BChE is a fast-binding process of mixed-type (Ki=1.77 µM; Ki'=3.17 µM). The crystal structure of mouse AChE complexed with C-547 was solved at 3.13 Å resolution. The complex is stabilized by cation-π, stacking and hydrogen-bonding interactions. Molecular dynamics simulations of the binding/dissociation processes of C-547 and C-35 (a non-charged analogue) to mouse and human AChEs were performed. Molecular modelling on mouse and human AChE showed that the slow step results from an enzyme conformational change that allows C-547 to cross the bottleneck in the active-site gorge, followed by formation of tight complex, as observed in the crystal structure. In contrast, the related non-charged compound C-35 is not a slow-binding inhibitor. It does not cross the bottleneck because it is not sensitive to the electrostatic driving force to reach the bottom of the gorge. Thus C-547 is one of the most potent and selective reversible inhibitors of AChE with a long residence time, τ=20 min, longer than for other reversible inhibitors used in the treatment of MG. This makes C-547 a promising drug for the treatment of this disease.
Assuntos
Acetilcolinesterase/química , Inibidores da Colinesterase/química , Simulação de Acoplamento Molecular , Miastenia Gravis , Compostos de Amônio Quaternário/química , Uracila/análogos & derivados , Animais , Células CHO , Inibidores da Colinesterase/uso terapêutico , Cricetinae , Cricetulus , Humanos , Camundongos , Miastenia Gravis/tratamento farmacológico , Miastenia Gravis/enzimologia , Compostos de Amônio Quaternário/uso terapêutico , Uracila/química , Uracila/uso terapêuticoRESUMO
We report a novel class of carbamate-type ChE inhibitors, structural analogs of pyridostigmine. A small library of congeneric pyridoxine-based compounds was designed, synthesized and evaluated for AChE and BChE enzymes inhibition in vitro. The most active compounds have potent enzyme inhibiting activity with IC50 values in the range of 0.46-2.1µM (for AChE) and 0.59-8.1µM (for BChE), with moderate selectivity for AChE comparable with that of pyridostigmine and neostigmine. Acute toxicity studies using mice models demonstrated excellent safety profile of the obtained compounds with LD50 in the range of 22-326mg/kg, while pyridostigmine and neostigmine are much more toxic (LD50 3.3 and 0.51mg/kg, respectively). The obtained results pave the way to design of novel potent and safe cholinesterase inhibitors for symptomatic treatment of neuromuscular disorders.
Assuntos
Acetilcolinesterase/metabolismo , Butirilcolinesterase/metabolismo , Inibidores da Colinesterase/química , Brometo de Piridostigmina/análogos & derivados , Piridoxina/química , Acetilcolinesterase/química , Animais , Butirilcolinesterase/química , Inibidores da Colinesterase/metabolismo , Inibidores da Colinesterase/toxicidade , Dose Letal Mediana , Camundongos , Ligação Proteica , Brometo de Piridostigmina/metabolismo , Brometo de Piridostigmina/toxicidade , Relação Estrutura-Atividade , Testes de Toxicidade AgudaRESUMO
Acetylcholinesterase (AChE) is an enzyme that hydrolyses the neurotransmitter acetylcholine, thereby limiting spillover and duration of action. This study demonstrates the existence of an endogenous mechanism for the regulation of synaptic AChE activity. At the rat extensor digitorum longus neuromuscular junction, activation of N-methyl-d-aspartate (NMDA) receptors by combined application of glutamate and glycine led to enhancement of nitric oxide (NO) production, resulting in partial AChE inhibition. Partial AChE inhibition was measured using increases in miniature endplate current amplitude. AChE inhibition by paraoxon, inactivation of NO synthase by N(x)-nitro-L-arginine methyl ester, and NMDA receptor blockade by DL-2-amino-5-phosphopentanoic acid prevented the increase in miniature endplate current amplitude caused by amino acids. High-frequency (10 Hz) motor nerve stimulation in a glycine-containing bathing solution also resulted in an increase in the amplitude of miniature endplate currents recorded during the interstimulus intervals. Pretreatment with an NO synthase inhibitor and NMDA receptor blockade fully eliminated this effect. This suggests that endogenous glutamate, released into the synaptic cleft as a co-mediator of acetylcholine, is capable of triggering the NMDA receptor/NO synthase-mediated pathway that modulates synaptic AChE activity. Therefore, in addition to well-established modes of synaptic plasticity (e.g. changes in the effectiveness of neurotransmitter release and/or the sensitivity of the postsynaptic membrane), another mechanism exists based on the prompt regulation of AChE activity.
Assuntos
Acetilcolinesterase/metabolismo , Junção Neuromuscular/metabolismo , Óxido Nítrico/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Inibidores da Colinesterase/farmacologia , Ácido Glutâmico/metabolismo , Masculino , Potenciais Pós-Sinápticos em Miniatura , NG-Nitroarginina Metil Éster/farmacologia , Junção Neuromuscular/fisiologia , Plasticidade Neuronal , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Paraoxon/farmacologia , Ratos , Ratos Wistar , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Valina/análogos & derivados , Valina/farmacologiaRESUMO
The creation of mitochondria-targeted vector systems is a new tool for the treatment of socially significant diseases. Phosphonium groups provide targeted delivery of drugs through biological barriers to organelles. For this purpose, a new class of alkyl(diethylAmino)(Phenyl) Phosphonium halides (APPs) containing one, two, or three diethylamino groups was obtained by the reaction of alkyl iodides (bromides) with (diethylamino)(phenyl)phosphines under mild conditions (20 °C) and high yields (93-98%). The structure of APP was established by NMR and XRD. A high in vitro cytotoxicity of APPs against M-HeLa, HuTu 80, PC3, DU-145, PANC-1, and MCF-7 lines was found. The selectivity index is in the range of 0.06-4.0 µM (SI 17-277) for the most active APPs. The effect of APPs on cancer cells is characterized by hyperproduction of ROS and depolarization of the mitochondrial membrane. APPs induce apoptosis, proceeding along the mitochondrial pathway. Incorporation of APPs into lipid systems (liposomes and solid lipid nanoparticles) improves cytotoxicity toward tumor cells and decrease toxicity against normal cell lines. The IC50s of lipid systems are lower than for the reference drug DOX, with a high SI (30-56) toward MCF-7 and DU-145. APPs exhibit high selective activity against Gram-positive bacteria S. aureus 209P and B. segeus 8035, including methicillin-resistant S. aureus (MRSA-1, MRSA-2), comparable to the activity of the fluoroquinolone antibiotic norfloxacin. A moderate in vivo toxicity in CD-1 mice was established for the lead APP.
RESUMO
Organophosphorus (OP) compounds represent a serious health hazard worldwide. The dominant mechanism of their action results from covalent inhibition of acetylcholinesterase (AChE). Standard therapy of acute OP poisoning is partially effective. However, prophylactic administration of reversible or pseudo-irreversible AChE inhibitors before OP exposure increases the efficiency of standard therapy. The purpose of the study was to test the duration of the protective effect of a slow-binding reversible AChE inhibitor (C547) in a mouse model against acute exposure to paraoxon (POX). It was shown that the rate of inhibition of AChE by POX in vitro after pre-inhibition with C547 was several times lower than without C547. Ex vivo pre-incubation of mouse diaphragm with C547 significantly prevented the POX-induced muscle weakness. Then it was shown that pre-treatment of mice with C547 at the dose of 0.01 mg/kg significantly increased survival after poisoning by 2xLD50 POX. The duration of the pre-treatment was effective up to 96 h, whereas currently used drug for pre-exposure treatment, pyridostigmine at a dose of 0.15 mg/kg was effective less than 24 h. Thus, long-lasting slow-binding reversible AChE inhibitors can be considered as new potential drugs to increase the duration of pre-exposure treatment of OP poisoning.
Assuntos
Compostos de Benzilamônio/administração & dosagem , Brometos/administração & dosagem , Inibidores da Colinesterase/administração & dosagem , Intoxicação por Organofosfatos/prevenção & controle , Compostos Organofosforados/toxicidade , Paraoxon/toxicidade , Brometo de Piridostigmina/administração & dosagem , Animais , Compostos de Benzilamônio/farmacologia , Brometos/farmacologia , Inibidores da Colinesterase/farmacologia , Preparações de Ação Retardada , Modelos Animais de Doenças , Camundongos , Brometo de Piridostigmina/farmacologia , Fatores de TempoRESUMO
We compared the effects of the novel acetylcholinesterase (AChE) inhibitor C-547 on action potential configuration and sinus rhythm in the isolated right atrium preparation of rat with those of armin and neostigmine. Both armin (10(-7), 10(-6), and 10(-5) M) and neostigmine (10(-7), 10(-6), and 5 x 10(-6) M) produced a marked decrease in action potential duration and slowing of sinus rate. These effects were abolished by atropine and are attributable to the accumulation of acetylcholine in the myocardium. The novel selective AChE inhibitor C-547 (10(-9) to 10(-7) M), an alkylammonium derivative of 6-methyluracil, had no such effects. The inhibition constant of C-547 on cardiac AChE is 40-fold higher than that on extensor digitorum longus muscle AChE. These results suggest that C-547 might be employed to treat diseases such as myasthenia gravis or Alzheimer disease, without having unwanted effects on the heart.
Assuntos
Inibidores da Colinesterase/farmacologia , Coração/efeitos dos fármacos , Compostos de Amônio Quaternário/farmacologia , Uracila/análogos & derivados , Potenciais de Ação/efeitos dos fármacos , Animais , Armina/farmacologia , Função Atrial/efeitos dos fármacos , Atropina/farmacologia , Relação Dose-Resposta a Droga , Eletrofisiologia , Coração/fisiologia , Técnicas In Vitro , Neostigmina/farmacologia , Ratos , Nó Sinoatrial/efeitos dos fármacos , Nó Sinoatrial/fisiologia , Uracila/farmacologiaRESUMO
In order to obtain a non-toxic amphiphilic calixresorcinarene capable to form nanoconjugates for drug encapsulation, tetraundecylcalixresorcinarene functionalized by methoxy poly(ethylene glycol) chains has been synthesized. The macrocycle obtained is characterized by low hemotoxicity. In aqueous solution it forms nanoassociates that are able to encapsulate organic substrates of different hydrophobicity, including drugs (doxorubicin, naproxen, ibuprofen, quercetin). The micelles of the macrocycle slowed down the release of the hydrophilic substrates in vitro. In physiological sodium chloride solution and phosphate-buffered saline, the micelles of the macrocycle acquire thermoresponsive properties and exhibit a temperature-controlled release of doxorubicin in vitro. The combination of the low toxicity and the encapsulation properties of the obtained calixresorcinarene-mPEG conjugate shows promising potential for the use as a supramolecular drug-delivery system.
RESUMO
Aggregation properties of mono (mono-CS) and dicationic (di-CS) surfactants, namely quaternised derivatives of 1,4-diazabicyclo[2.2.2]octane (DABCO), have been evaluated in water and in nutrient broths of different pH, i.e. in Hottinger broth (ÑÐ=7.2) and Sabouraud dextrose broth (ÑÐ=5.6). Aggregation capacity of surfactants was shown to be responsible for the solubilization properties of a complex composed of a hydrophobic probe (Sudan I) and a selected drug (quercetin), contributing to the antimicrobial activity of this surfactant system. The effect of N-methyl-d-glucamine (NmDg) additive on the antimicrobial activity of mono-CS, and its aggregation and solubilization parameters, has also been evaluated. A substantial decrease in critical micelle concentration (CMC) of cationic surfactants in nutrient broths (up to 60 times) has been reported. Twofold dilution of monocationic surfactant by NmDg slightly changed the CMC of surfactant; however, it provided a remarkable increase in solubilization capacity (â¼by 4 times) and decrease in its toxicity. The data anticipate the potential use of DABCO quaternized derivatives as innovative non-toxic delivery systems for hydrophobic drugs.
Assuntos
Anti-Infecciosos/química , Interações Hidrofóbicas e Hidrofílicas , Preparações Farmacêuticas/química , Piperazinas/farmacologia , Anisotropia , Anti-Infecciosos/farmacologia , Candida albicans/efeitos dos fármacos , Cátions , Meios de Cultura , Portadores de Fármacos , Fluorescência , Hemólise/efeitos dos fármacos , Humanos , Hidrodinâmica , Micelas , Tamanho da Partícula , Piperazinas/química , Quercetina/química , Soluções , Staphylococcus aureus/efeitos dos fármacos , Tensão Superficial , Tensoativos/química , TemperaturaRESUMO
Novel 6-methyluracil derivatives with ω-(substituted benzylethylamino)alkyl chains at the nitrogen atoms of the pyrimidine ring were designed and synthesized. The numbers of methylene groups in the alkyl chains were varied along with the electron-withdrawing substituents on the benzyl rings. The compounds are mixed-type reversible inhibitors of cholinesterases, and some of them show remarkable selectivity for human acetylcholinesterase (hAChE), with inhibitory potency in the nanomolar range, more than 10,000-fold higher than that for human butyrylcholinesterase (hBuChE). Molecular modeling studies indicate that these compounds are bifunctional AChE inhibitors, spanning the enzyme active site gorge and binding to its peripheral anionic site (PAS). In vivo experiments show that the 6-methyluracil derivatives are able to penetrate the blood-brain barrier (BBB), inhibiting brain-tissue AChE. The most potent AChE inhibitor, 3 d (1,3-bis[5-(o-nitrobenzylethylamino)pentyl]-6-methyluracil), was found to improve working memory in scopolamine and transgenic APP/PS1 murine models of Alzheimer's disease, and to significantly decrease the number and area of ß-amyloid peptide plaques in the brain.
Assuntos
Acetilcolinesterase/metabolismo , Doença de Alzheimer/tratamento farmacológico , Inibidores da Colinesterase/farmacologia , Uracila/análogos & derivados , Doença de Alzheimer/enzimologia , Animais , Comportamento Animal/efeitos dos fármacos , Encéfalo/enzimologia , Inibidores da Colinesterase/síntese química , Inibidores da Colinesterase/química , Cristalografia por Raios X , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Humanos , Camundongos , Camundongos Transgênicos , Modelos Moleculares , Estrutura Molecular , Relação Estrutura-Atividade , Uracila/síntese química , Uracila/química , Uracila/farmacologiaRESUMO
3-Phenyl(methyl)-5-alkyl-1-(pyridin-3-yl)imidazo[1,5-a]quinoxalin-4-ones (2a-f) and their N-alkyl-pyridinium salts (3a-o), including 1,n-bis{3-(3-phenylimidazo[1,5-a]quinoxalin-4(5H)-on-1-yl)pyridinium}alkane dibromides (4a-d, 5, 6) have been synthesized. It has been established that the antimicrobial properties of imidazo[1,5-a]quinoxaline derivatives are connected with the presence of various alkyl substituents in the position 1 of the pyridine ring and in the position 5 of the imidazo[1,5-a]quinoxaline system. Chlorides and iodides are more active towards bacteria than fungi. Compounds 3d, 3e, 3m and 3n showed an effective bacteriostatic activity. Compound showed not only well defined bacteriostatic activities but also good fungistatic activities, with the MIC values comparable with the reference drugs. Toxicity of more effective (imidazo[1,5-a]quinoxalin-4-on-1-yl)-1-pyridinium halides was examined in mice.
Assuntos
Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Imidazóis/química , Imidazóis/farmacologia , Compostos de Piridínio/química , Animais , Anti-Infecciosos/síntese química , Anti-Infecciosos/toxicidade , Bactérias/efeitos dos fármacos , Desenho de Fármacos , Feminino , Fungos/efeitos dos fármacos , Imidazóis/síntese química , Imidazóis/toxicidade , Masculino , Camundongos , Testes de Sensibilidade MicrobianaRESUMO
Reactions of pyrimidinophanes with two 6-methylthiocytosine and one 5(6)-alkyluracil moieties bridged with each other by polymethylene spacers with methyl or nonyl p-toluenesulfonate, p-toluenesulfonic acid, methanesulfonate and trifluorosulfonate afforded amphiphilic macrocyclic bis-p-toluene-, methane- and trifluorosulfonates. Despite the presence of several reaction centers in the initial pyrimidinophane molecules, protonation and methylation occurred only at the N(1) atom (with quaternization) of the 6-methylthiocytosine moieties. The bacteriostatic and fungistatic activity of the products was estimated. Macrocyclic tosylates exhibit a remarkable selectivity towards Staphylococcus aureus, with MIC values comparable with a reference drug. Bacteriostatic activity of the amphiphilic pyrimidinophanes depends on the size of the macrocycles, and the highest activity corresponds to definite lengths of polymethylene bridges. Besides, the antimicrobial activity of the screened pyrimidine derivatives depends on their topology. While macrocyclic tosylates are more active against bacteria than against fungi, acyclic tosylate with the same structural fragments shows a dramatical decrease of MIC towards mold and yeast with respect to the corresponding macrocycle. It is found that macrocyclic and acyclic tosylates in high dilutions decrease the extracellular lipase activity.
Assuntos
Antibacterianos/farmacologia , Citosina/química , Pirimidinas/farmacologia , Uracila/química , Animais , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
BACKGROUND AND PURPOSE: The rat respiratory muscle diaphragm has markedly lower sensitivity than the locomotor muscle extensor digitorum longus (EDL) to the new acetylcholinesterase (AChE) inhibitors, alkylammonium derivatives of 6-methyluracil (ADEMS). This study evaluated several possible reasons for differing sensitivity between the diaphragm and limb muscles and between the muscles and the brain. EXPERIMENTAL APPROACH: Increased amplitude and prolonged decay time of miniature endplate currents were used to assess anti-cholinesterase activity in muscles. In hippocampal slices, induction of synchronous network activity was used to follow cholinesterase inhibition. The inhibitor sensitivities of purified AChE from the EDL and brain were also estimated. KEY RESULTS: The intermuscular difference in sensitivity to ADEMS is partly explained caused by a higher level of mRNA and activity of 1,3-bis[5(diethyl-o-nitrobenzylammonium)pentyl]-6-methyluracildibromide (C-547)-resistant BuChE in the diaphragm. Moreover, diaphragm AChE was more than 20 times less sensitive to C-547 than that from the EDL. Sensitivity of the EDL to C-547 dramatically decreased after treadmill exercises that increased the amount of PRiMA AChE(G4), but not ColQ AChE(A12) molecular forms. The A12 form present in muscles appeared more sensitive to C-547. The main form of AChE in brain, PRiMA AChE(G4), was apparently less sensitive because brain cholinesterase activity was almost three orders of magnitude more resistant to C-547 than that of the EDL. CONCLUSIONS AND IMPLICATIONS: Our findings suggest that ADEMS compounds could be used for the selective inhibition of AChEs and as potential therapeutic tools.
Assuntos
Inibidores da Colinesterase/farmacologia , Diafragma/efeitos dos fármacos , Hipocampo/efeitos dos fármacos , Músculo Esquelético/efeitos dos fármacos , Compostos de Amônio Quaternário/farmacologia , Uracila/análogos & derivados , Acetilcolinesterase/metabolismo , Animais , Butirilcolinesterase/metabolismo , Diafragma/enzimologia , Feminino , Hipocampo/enzimologia , Masculino , Potenciais Pós-Sinápticos em Miniatura/efeitos dos fármacos , Músculo Esquelético/enzimologia , Condicionamento Físico Animal/fisiologia , Distribuição Aleatória , Ratos , Ratos Wistar , Uracila/farmacologiaRESUMO
Agent No. 547 (1,3-bis[omega-(diethyl-ortho-nitrobenzylammonio)-pentyl]-6-methyluracil dibromide), a newly synthesized inhibitor of mammalian-specific acetyltcholinesterase (EC 3.1.1.7) was investigated for genotoxicity using the DNA-repair test, Ames test and in vivo micronucleus test with mouse peripheral blood erythrocytes. Agent No. 547 did not cause significant changes in growth of repair-deficient Escherichia coli tester strains. The compound was non-mutagenic in Salmonella typhimurium strains TA98 and TA100 with and without rat microsomal activation mixture. However, we observed a marked increase in number of His(+) revertants for both tester strains in preincubation assays. The results obtained in the micronucleus test indicate that agent No. 547 possesses significant clastogenic activity. At the high dose tested (0.5 mg/kg), the compound induced a seven-fold increase in the number of micronuclei over the spontaneous background 48 h after treatment. The results suggest that further work should be promoted to identify the metabolic pathways involved in genotoxicity of agent No. 547 in mammalian cells and to evaluate the real risk of its exposure.