RESUMO
Francisella tularensis is a highly infectious intracellular bacterium causing tularemia disease and is regarded as a potential biological weapon. The development of a vaccine, effective treatment, or prophylactic substances targeted against tularemia is in the forefront of interest and could help to prevent or mitigate possible malevolent acts by bioterrorism utilizing F. tularensis. The viability of F. tularensis, and thus of a tularemia disease outbreak, might potentially be suppressed by simple commonly available natural substances. Epigallocatechin gallate (EGCG) is contained in green tea and its antimicrobial effect has been described. Here, we show that EGCG can suppress F. tularensis growth and is able to reduce the bacterium's ability to replicate inside mouse bone marrow-derived macrophages (BMMs) without side effects on BMMs' own viability. We suggest one (but not the only) mechanism of EGCG action. We demonstrate that EGCG can block the main functions of HU protein, the important regulator of F. tularensis virulence, leading to overall attenuation of F. tularensis viability. EGCG can delay death of mice infected by F. tularensis and can be used as a prophylactic agent against tularemia disease. Postponing death by up to 2 days can provide sufficient opportunity to administer another treatment agent.
Assuntos
Catequina , Francisella tularensis , Tularemia , Animais , Camundongos , Tularemia/microbiologia , Proteínas de Ligação a DNA/metabolismo , Catequina/uso terapêuticoRESUMO
Organophosphorus compounds (OPs) involving life-threatening nerve agents (NA) have been known for several decades. Despite a clear mechanism of their lethality caused by the irreversible inhibition of acetylcholinesterase (AChE) and manifested via overstimulation of peripheral nicotinic and muscarinic acetylcholine (ACh) receptors, the mechanism for central neurotoxicity responsible for acute or delayed symptoms of the poisoning has not been thoroughly uncovered. One of the reasons is the lack of a suitable model. In our study, we have chosen the SH-SY5Y model in both the differentiated and undifferentiated state to study the effects of NAs (GB, VX and A234). The activity of expressed AChE in cell lysate assessed by Ellman's method showed 7.3-times higher activity in differentiated SH-SY5Y cells in contrast to undifferentiated cells, and with no involvement of BuChE as proved by ethopropazine (20 µM). The activity of AChE was found to be, in comparison to untreated cells, 16-, 9.3-, and 1.9-times lower upon A234, VX, and GB (100 µM) administration respectively. The cytotoxic effect of given OPs expressed as the IC50 values for differentiated and undifferentiated SH-SY5Y, respectively, was found 12 mM and 5.7 mM (A234), 4.8 mM and 1.1 mM (VX) and 2.6 mM and 3.8 mM (GB). In summary, although our results confirm higher AChE expression in the differentiated SH-SY5Y cell model, the such higher expression does not lead to a more pronounced NA cytotoxic effect. On the contrary, higher expression of AChE may attenuate NA-induced cytotoxicity by scavenging the NA. Such finding highlights a protective role for cholinesterases by scavenging Novichoks (A-agents). Second, we confirmed the mechanism of cytotoxicity of NAs, including A-agents, can be ascribed rather to the non-specific effects of OPs than to AChE-mediated effects.
Assuntos
Antineoplásicos , Agentes Neurotóxicos , Neuroblastoma , Síndromes Neurotóxicas , Humanos , Acetilcolinesterase/metabolismo , Linhagem Celular Tumoral , Síndromes Neurotóxicas/etiologiaRESUMO
Cucurbit[n]urils are macrocyclic compounds capable of forming host-guest complexes with different molecules. In this study, we focused on cucurbit[7]uril (CB[7]) safety and pharmacokinetics. We investigated CB[7] cytotocixity in human renal cells ACHN using the xCELLigence system. We also determined maximum tolerated doses (MTD) and no observed adverse effect levels (NOAEL) after intramuscular (i.m.), intraperitoneal (i.p.), and intragastric (i.g.) administration in mice using clinical observation, blood biochemistry, and histopathology. At NOAELs, we studied its pharmacokinetics in plasma and kidneys. Finally, we performed a 7 day repeated-dose toxicity study at 50% of NOAEL after i.p. administration, assaying CB[7] concentration in plasma, brain, kidney, and liver; we also assessed the liver and kidney histopathology. In vitro, CB[7] did not show toxicity up to 0.94 mg/mL. MTDs in vivo were set at 300, 350, and 600 mg/kg, and NOAEL were established at 150, 100, and 300 mg/kg after i.m., i.p., and i.g. administration, respectively. Parenteral administration produced tissue damage mainly to the kidney, while i.g. administration caused only minor liver damage. Parenteral CB[7] administration led to fast elimination from blood, accompanied with kidney accumulation; absorption from the gastrointestinal tract was minimal. Short repeated i.p. administration was well tolerated. After initial CB[7] accumulation in blood and kidney, the concentrations stabilised and decreased during the experiment. Approximately 3.6% of animals showed signs of nephrotoxicity. Although CB[7] appears to be a promising molecule, nephrotoxicity may be the most critical drawback of its parenteral use, because the kidney represents the main organ of its elimination.
Assuntos
Compostos Heterocíclicos com 2 Anéis , Imidazolidinas , Compostos Macrocíclicos , Animais , Compostos Heterocíclicos com 2 Anéis/toxicidade , Imidazolidinas/toxicidade , Compostos Macrocíclicos/toxicidade , Dose Máxima Tolerável , CamundongosRESUMO
A series of tacrine - benzothiazole hybrids incorporate inhibitors of acetylcholinesterase (AChE), amyloid ß (Aß) aggregation and mitochondrial enzyme ABAD, whose interaction with Aß leads to mitochondrial dysfunction, into a single molecule. In vitro, several of 25 final compounds exerted excellent anti-AChE properties and interesting capabilities to block Aß aggregation. The best derivative of the series could be considered 10w that was found to be highly potent and selective towards AChE with the IC50 value in nanomolar range. Moreover, the same drug candidate exerted absolutely the best results of the series against ABAD, decreasing its activity by 23% at 100 µM concentration. Regarding the cytotoxicity profile of highlighted compound, it roughly matched that of its parent compound - 6-chlorotacrine. Finally, 10w was forwarded for in vivo scopolamine-induced amnesia experiment consisting of Morris Water Maze test, where it demonstrated mild procognitive effect. Taking into account all in vitro and in vivo data, highlighted derivative 10w could be considered as the lead structure worthy of further investigation.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Benzotiazóis/farmacologia , Colinérgicos/farmacologia , Inibidores Enzimáticos/farmacologia , Fármacos Neuroprotetores/farmacologia , Tacrina/farmacologia , 3-Hidroxiacil-CoA Desidrogenases/antagonistas & inibidores , 3-Hidroxiacil-CoA Desidrogenases/metabolismo , Acetilcolinesterase/metabolismo , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/antagonistas & inibidores , Peptídeos beta-Amiloides/metabolismo , Benzotiazóis/química , Colinérgicos/síntese química , Colinérgicos/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Estrutura Molecular , Fármacos Neuroprotetores/síntese química , Fármacos Neuroprotetores/química , Agregados Proteicos/efeitos dos fármacos , Relação Estrutura-Atividade , Tacrina/químicaRESUMO
Sulfur mustard [bis(2-chloroethyl) sulfide; SM] is a highly poisonous chemical warfare agent. The mechanism of its cytotoxicity affects several pathways, which cause cell damage or death. The main organ affected in case of exposure to both aerosol and vapor is lungs. The present study focuses on time- and concentration-dependent changes in human lung fibroblasts NHLF and lung epithelial cell line A-549. The cells were treated with SM at the concentrations of 5, 10 and 100 µM and signs of stress response were evaluated during 1-72 h post-treatment. Parameters for testing included cell viability and morphology, loss of transmembrane mitochondrial potential, apoptosis, oxidative stress, changes in the cell cycle, and ATM kinase activation. The cytotoxic effect of SM resulted in a time-dependent decrease in viability of A-459 associated with apoptosis more markedly than in NHLF. We did not observe any generation of reactive oxygen species by SM. SM at concentrations of 5 and 10 µM induced the S-phase cell cycle arrest at both cell lines. On the other hand, 100 µM caused nonspecific cell cycle arrest. ATM kinase was activated transiently. The results indicate that NHLF cells are less prone to toxic damage by SM in case of cell viability, apoptosis and loss of transmembrane mitochondrial potential. The analysis provides a time-related cytotoxic profile of A-549 and NHLF cells for further investigation into the prevention of SM toxic effects and their potential treatment.
Assuntos
Substâncias para a Guerra Química/toxicidade , Células Epiteliais/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Gás de Mostarda/toxicidade , Estresse Fisiológico/efeitos dos fármacos , Células A549 , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Humanos , Pulmão/citologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Espécies Reativas de Oxigênio , Fatores de TempoRESUMO
The development of acetylcholinesterase reactivators, i.e., antidotes against organophosphorus poisoning, is an important goal of defense research. The aim of this study was to compare cytotoxicity and chemical structure of five currently available oximes (pralidoxime, trimedoxime, obidoxime, methoxime, and asoxime) together with four perspective oximes from K-series (K027, K074, K075, and K203). The cytotoxicity of tested substances was measured using two methods - colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay and impedance based real-time cytotoxicity assay - in three different cell lines (HepG2, ACHN, and NHLF). Toxicity was subsequently expressed as toxicological index IC50. The tested compounds showed different cytotoxicity ranging from 0.92 to 40.06 mM. In HepG2 cells, K027 was the least and asoxime was the most toxic reactivator. In ACHN and NHLF cell lines, trimedoxime was the compound with the lowest adverse effects, whereas the highest toxicity was found in methoxime-treated cells. The results show that at least five structural features affect the reactivators' toxicity such as the number of oxime groups in the molecule, their position on pyridinium ring, the length of carbon linker, and the oxygen substitution or insertion of the double bond into the connection chain. Newly synthetized oximes with IC50 ≥ 1 mM evaluated in this three cell lines model might appear suitable for further testing.
Assuntos
Reativadores da Colinesterase/química , Reativadores da Colinesterase/toxicidade , Oximas/química , Oximas/toxicidade , Alternativas aos Testes com Animais , Sobrevivência Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Fibroblastos/efeitos dos fármacos , Células Hep G2 , Humanos , Concentração Inibidora 50 , Dose Letal Mediana , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
Autophagy inhibition through small-molecule inhibitors is one of the approaches to increase the efficiency of radiotherapy in oncological patients. A new inhibitor-Lys05-with the potential to accumulate within lysosomes and to block autophagy was discovered a few years ago. Several studies have addressed its chemosensitizing effects but nothing is known about its impact in the context of ionizing radiation (IR). To describe its role in radiosensitization, we employed radioresistant human non-small cell lung carcinoma cells (H1299, p53-negative). Combined treatment of H1299 cells by Lys05 together with IR decreased cell survival in the clonogenic assay and real-time monitoring of cell growth more than either Lys05 or IR alone. Immunodetection of LC3 and p62/SQSTM1 indicated that autophagy was inhibited, which correlated with increased SQSTM1 and decreased BNIP3 gene expression determined by qRT-PCR. Fluorescence microscopy and flow cytometry uncovered an accumulation of lysosomes. Similarly, transmission electron microscopy demonstrated the accumulation of autophagosomes confirming the ability of Lys05 to potentiate autophagy inhibition in H1299 cells. We report here for the first time that Lys05 could be utilized in combination with IR as a promising future strategy in the eradication of lung cancer cells.
Assuntos
Neoplasias Pulmonares/metabolismo , Radiação Ionizante , Apoptose/efeitos da radiação , Western Blotting , Linhagem Celular Tumoral , Citometria de Fluxo , Humanos , Microscopia Eletrônica de Transmissão , Microscopia de FluorescênciaRESUMO
2-chloroethyl ethyl sulfide (CEES) is a vesicant agent, commonly referred to half mustard due to its ability to form monofunctional adducts with DNA. In this study, we evaluated the chemoprotective potential of 13 compounds and their mixtures with sodium 2-mercaptoethanesulfonate (MESNA) against CEES-induced geno- and cytotoxicity in human lung cell line A-549. MESNA, L-glutathione (GSH), thiourea, sodium thiosulfate, hexamethylenetetramine, 4-acetamidophenol, asoxime dichloride (HI-6), N-acetyl-L-cysteine (NAC), sodium pyruvate, myo-inositol, 3-aminobenzamide (3-AB), nicotinamide, and Nω-nitro-L-arginine methyl ester hydrochloride and combinations of these compounds with MESNA were applied 30 min before CEES. DNA alkylation was measured using modified comet assay 1 and 24 h after the exposure. Cell viability was determined using MTT assay at 24 and 72 h. The mono-therapeutical approach identified MESNA and GSH to provide significant chemoprotection. NAC and 3-AB supported DNA damage repair, while cell viability remained unaffected. Mixtures of GSH or NAC with MESNA showed protective synergism against DNA damage. Other compounds or their combinations with MESNA failed due to the potentiation of CEES-induced cytotoxicity. The chemoprotection against CEES remains limited; however, the combination of substances can provide protective synergy and may represent a promising strategy in the treatment of accidental exposure to monoalkylating agents.
RESUMO
Alzheimer's disease (AD) is a progressive neurodegenerative disorder and currently there is no efficient treatment. The classic drug-design strategy based on the "one-molecule-one-target" paradigm was found to be ineffective in the case of multifactorial diseases like AD. A novel multi-target-directed ligand strategy based on the assumption that a single compound consisting of two or more distinct pharmacophores is able to hit multiple targets has been proposed as promising. Herein, we investigated 7-methoxytacrine - memantine heterodimers developed with respect to the multi-target-directed ligand theory. The spectroscopic, microscopic and cell culture methods were used for systematic investigation of the interference of the heterodimers with ß-secretase (BACE1) activity, Aß peptide amyloid fibrillization (amyloid theory) and interaction with M1 subtype of muscarinic (mAChRs), nicotinic (nAChRs) acetylcholine receptors (cholinergic theory) and N-methyl-d-aspartate receptors (NMDA) (glutamatergic theory). The drug-like properties of selected compounds have been evaluated from the point of view of blood-brain barrier penetration and cell proliferation. We have confirmed the multipotent effect of novel series of compounds. They inhibited effectively Aß peptide amyloid fibrillization and affected the BACE1 activity. Moreover, they have AChE inhibitory potency but they could not potentiate cholinergic transmission via direct interaction with cholinergic receptors. All compounds were reported to act as an antagonist of both M1 muscarinic and muscle-type nicotinic receptors. We have found that 7-methoxytacrine - memantine heterodimers are able to hit multiple targets associated with Alzheimer's disease and thus, have a potential clinical impact for slowing or blocking the neurodegenerative process related to this disease.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Amantadina/farmacologia , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Peptídeos beta-Amiloides/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Tacrina/análogos & derivados , Doença de Alzheimer/metabolismo , Amantadina/análogos & derivados , Secretases da Proteína Precursora do Amiloide/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Células CHO , Colinesterases/metabolismo , Cricetulus , Dimerização , Inibidores Enzimáticos/química , Células HEK293 , Humanos , Terapia de Alvo Molecular , Receptor Muscarínico M1/antagonistas & inibidores , Receptor Muscarínico M1/metabolismo , Receptores Colinérgicos/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Tacrina/química , Tacrina/farmacologia , XenopusRESUMO
A novel series of 6-chlorotacrine-scutellarin hybrids was designed, synthesized and the biological activity as potential anti-Alzheimer's agents was assessed. Their inhibitory activity towards human acetylcholinesterase (hAChE) and human butyrylcholinesterase (hBChE), antioxidant activity, ability to cross the blood-brain barrier (BBB) and hepatotoxic profile were evaluated in vitro. Among these compounds, hybrid K1383, bearing two methylene tether between two basic scaffolds, was found to be very potent hAChE inhibitor (IC50 = 1.63 nM). Unfortunately, none of the hybrids displayed any antioxidant activity (EC50 ≥ 500 µM). Preliminary data also suggests a comparable hepatotoxic profile with 6-Cl-THA (established on a HepG2 cell line). Kinetic studies performed on hAChE with the most active compound in the study, K1383, pointed out to a mixed, non-competitive enzyme inhibition. These findings were further corroborated by docking studies.
Assuntos
Apigenina/química , Inibidores da Colinesterase/química , Inibidores da Colinesterase/farmacologia , Glucuronatos/química , Tacrina/análogos & derivados , Acetilcolinesterase/metabolismo , Doença de Alzheimer/enzimologia , Barreira Hematoencefálica/metabolismo , Butirilcolinesterase/metabolismo , Inibidores da Colinesterase/síntese química , Desenho de Fármacos , Ativação Enzimática/efeitos dos fármacos , Humanos , Simulação de Acoplamento Molecular , Relação Estrutura-Atividade , Tacrina/químicaRESUMO
Tacrine (THA), the first clinically effective acetylcholinesterase (AChE) inhibitor and the first approved drug for the treatment of Alzheimer's disease (AD), was withdrawn from the market due to its side effects, particularly its hepatotoxicity. Nowadays, THA serves as a valuable scaffold for the design of novel agents potentially applicable for AD treatment. One such compound, namely 7-methoxytacrine (7-MEOTA), exhibits an intriguing profile, having suppressed hepatotoxicity and concomitantly retaining AChE inhibition properties. Another interesting class of AChE inhibitors represents Huprines, designed by merging two fragments of the known AChE inhibitors-THA and (-)-huperzine A. Several members of this compound family are more potent human AChE inhibitors than the parent compounds. The most promising are so-called huprines X and Y. Here, we report the design, synthesis, biological evaluation, and in silico studies of 2-methoxyhuprine that amalgamates structural features of 7-MEOTA and huprine Y in one molecule.
Assuntos
Aminoquinolinas/química , Aminoquinolinas/farmacologia , Descoberta de Drogas , Acetilcolinesterase , Doença de Alzheimer/tratamento farmacológico , Aminoquinolinas/síntese química , Sítios de Ligação , Barreira Hematoencefálica/metabolismo , Butirilcolinesterase , Domínio Catalítico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Inibidores da Colinesterase/química , Inibidores da Colinesterase/farmacologia , Desenho de Fármacos , Ativação Enzimática/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/química , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Humanos , Hidrólise , Concentração Inibidora 50 , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Permeabilidade , Ligação Proteica , Relação Estrutura-Atividade , Tacrina/análogos & derivados , Tacrina/química , Tacrina/farmacologiaRESUMO
OBJECTIVE: The genotoxic vesicant sulphur mustard [bis-2-(chloroethyl)sulphide] is a chemical warfare agent which is easily available due to its relatively simple synthesis. Thus, sulphur mustard is a potential agent for mass contamination. In this study, we focused on sulphur mustard toxicity and decontamination in a rat model using commercially available detergent mixtures for dermal decontamination. METHODS: Male Wistar rats were percutaneously treated with sulphur mustard and subjected to wet decontamination 2 min postexposure. Commercially produced detergents Neodekont™, Argos™, Dermogel™ and FloraFree™ were tested for their decontamination efficacy against an exposed group and their protective ratios determined. RESULTS AND CONCLUSION: The results showed that all tested detergent solutions produced an increase in the median lethal dose [LD(50) = 9.83 (5.87-13.63) mg·kg(-1)] in comparison to controls, which led to increased survival of experimental animals. In general, all tested detergents provided modest decontamination efficacy (PR = 2.0-5.7). The highest protective ratio (5.7) was consistently achieved with Argos™. Accordingly, Argos™ should be considered in further investigation of mass casualty decontamination.
Assuntos
Substâncias para a Guerra Química/toxicidade , Descontaminação/métodos , Detergentes/farmacologia , Gás de Mostarda/toxicidade , Administração Cutânea , Animais , Caquexia/induzido quimicamente , Diarreia/induzido quimicamente , Hematúria/induzido quimicamente , Dose Letal Mediana , Masculino , Ratos , Ratos Wistar , Pele/efeitos dos fármacos , Pele/patologia , Espirro/efeitos dos fármacosRESUMO
Oxime HI-6 is an efficient reactivator of the acetylcholinesterase inhibited by organophosphorous nerve agents. In this study we have estimated cytotoxicity of HI-6 by the colony forming assay and genotoxicity by the comet assay on human and rodent cell lines. IC50 of HI-6 assessed by the colony forming capacity was 3.59 mM for HeLa cells and 5.18 mM for a mouse cell line L929. Small difference in cytotoxicity was found among other cell lines tested: IC50 was 1.61 mM for human A549 cells, 1.14 mM for UROtse line, 1.96 mM and 1.71 mM for Chinese hamster cells AA8 and UV-20, respectively. The A549 cell viability measured with the MTT test was 5 times decreased comparing 2 and 24 hours of HI-6 oxime treatment. The 5 mM HI-6 concentration reduced the viability within 2 hours to 95% only, however, it induced a significant number of DNA breaks in mouse cells L929, and also in human UROtse and HepG2 cells. 1-ß-D-arabinofuranosylcytosine (10(-4) M) and hydroxyurea (10(-2) M), supplemented to the cultivation medium, did not cause any significant accumulation of DNA breaks during treatment, which indicated that the nucleotide excision repair was not acting on the induced DNA damage.
Assuntos
Linhagem Celular/efeitos dos fármacos , Linhagem Celular/fisiologia , Dano ao DNA/fisiologia , Oximas/toxicidade , Compostos de Piridínio/toxicidade , Animais , Antídotos/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Reativadores da Colinesterase/toxicidade , Ensaio de Unidades Formadoras de Colônias , Cricetinae , Citotoxinas/toxicidade , Humanos , Dose Letal Mediana , Camundongos , Mutagênicos/toxicidade , Ratos , Especificidade da EspécieRESUMO
Oxime-based acetylcholinesterase reactivators (briefly oximes) regenerate organophosphate-inactivated acetylcholinesterase and restore its function. Poor blood-brain-barrier passage and fast elimination from blood limit their actual use in treatment of patients exposed to organophosphates. Previous in vitro results implicated further testing of cucurbit[7]uril as a delivery vehicle for bisquaternary oximes. The present paper focuses on cell toxicity, in vivo safety and influence of cucurbit[7]uril on oxime pharmacokinetics and pharmacodynamics. Neither the K027 nor the complex caused any cell toxicity, changes in blood biochemistry or hepato- or nephrotoxicity in tested concentrations. The encapsulation of K027 increased and accelerated the blood-brain-barrier penetration. The peripheral oxime exposure also increased, supporting the suggestion that cucurbit[7]uril protects the circulating oxime from rapid renal clearance. Contrary to the comparable in vitro reactivation power of K027 and the encapsulated K027, we failed to confirm this in vivo. In theory, this might result from the non-specific binding of molecules to the cucurbit[7]uril or the interaction of K027 with cucurbit[7]uril being too strong for acetylcholinesterase reactivation. Precise explanation requires additional in silico, in vitro and also in vivo experiments.
Assuntos
Acetilcolinesterase/sangue , Acetilcolinesterase/metabolismo , Encéfalo/efeitos dos fármacos , Hidrocarbonetos Aromáticos com Pontes/farmacocinética , Reativadores da Colinesterase/farmacocinética , Eritrócitos/efeitos dos fármacos , Imidazóis/farmacocinética , Oximas/farmacocinética , Compostos de Piridínio/farmacocinética , Células A549 , Animais , Encéfalo/enzimologia , Hidrocarbonetos Aromáticos com Pontes/administração & dosagem , Hidrocarbonetos Aromáticos com Pontes/toxicidade , Sobrevivência Celular/efeitos dos fármacos , Reativadores da Colinesterase/administração & dosagem , Reativadores da Colinesterase/toxicidade , Relação Dose-Resposta a Droga , Eritrócitos/enzimologia , Feminino , Proteínas Ligadas por GPI/sangue , Proteínas Ligadas por GPI/metabolismo , Células Hep G2 , Humanos , Imidazóis/administração & dosagem , Imidazóis/toxicidade , Injeções Intramusculares , Masculino , Dose Máxima Tolerável , Camundongos Endogâmicos ICR , Oximas/administração & dosagem , Oximas/toxicidade , Compostos de Piridínio/administração & dosagem , Compostos de Piridínio/toxicidade , Medição de Risco , Distribuição TecidualRESUMO
BACKGROUND: Based on the prevalence studies, the number of people suffering from dementia will almost double every 20 years, to 65.7 million in 2030 and 115.4 million in 2050, assuming no changes in mortality, effective preventative measures, definitive diagnostic guidelines or curative treatment. From the abovementioned epidemiological data, it is obvious that dementia constitutes a major public health problem not only at present, but unfortunately also in the future. OBJECTIVES AND METHODS: Several N-alkylated tacrine (THA) derivatives have already been synthesized by Pomponi et al., in 1997. However, these compounds were tested for their anti-AChE activity using enzyme isolated from Electrophorus electricus. For this reason, we have decided to extend the previously reported series of THA derivatives and consequently test them in the battery of experiments, the results of which have served to more relevant evaluation of these compounds from the perspective of Alzeimer´s disease compared to that published by Pomponi. RESULTS AND CONCLUSION: In summary, all compounds of interest effectively inhibited ChEs in vitro. One of the most promising derivatives 8 bearing an N-octyl chain showed 2.5-fold higher AChE inhibitory activity in relation to tacrine. With respect to blood-brain barrier (BBB) penetration, it can be claimed that synthesized analogues are presumably able to cross the BBB. From the point of view of hepatotoxicity, selected Nalkylated tacrine derivatives exerted worse results compared to tacrine. However, in vitro results are only illustrative, therefore, only in vivo experiments could determine the real value of selected N-alkylated THA derivatives.
Assuntos
Doença de Alzheimer , Inibidores da Colinesterase/farmacologia , Descoberta de Drogas , Tacrina/análogos & derivados , Tacrina/farmacologia , Células Hep G2 , HumanosRESUMO
A combination of tacrine and tryptophan led to the development of a new family of heterodimers as multi-target agents with potential to treat Alzheimer's disease. Based on the in vitro biological profile, compound S-K1035 was found to be the most potent inhibitor of human acetylcholinesterase (hAChE) and human butyrylcholinesterase (hBChE), demonstrating balanced IC50 values of 6.3 and 9.1â¯nM, respectively. For all the tacrine-tryptophan heterodimers, favorable inhibitory effect on hAChE as well as on hBChE was coined to the optimal spacer length ranging from five to eight carbon atoms between these two pharmacophores. S-K1035 also showed good ability to inhibit Aß42 self-aggregation (58.6⯱â¯5.1% at 50⯵M) as well as hAChE-induced Aß40 aggregation (48.3⯱â¯6.3% at 100⯵M). The X-ray crystallographic analysis of TcAChE in complex with S-K1035 pinpointed the utility of the hybridization strategy applied and the structures determined with the two K1035 enantiomers in complex with hBChE could explain the higher inhibition potency of S-K1035. Other in vitro evaluations predicted the ability of S-K1035 to cross blood-brain barrier and to exert a moderate inhibition potency against neuronal nitric oxide synthase. Based on the initial promising biochemical data and a safer in vivo toxicity compared to tacrine, S-K1035 was administered to scopolamine-treated rats being able to dose-dependently revert amnesia.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Inibidores da Colinesterase/farmacologia , Fármacos Neuroprotetores/farmacologia , Tacrina/farmacologia , Triptofano/farmacologia , Acetilcolinesterase/metabolismo , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/antagonistas & inibidores , Peptídeos beta-Amiloides/metabolismo , Animais , Butirilcolinesterase/metabolismo , Inibidores da Colinesterase/síntese química , Inibidores da Colinesterase/química , Relação Dose-Resposta a Droga , Humanos , Ligantes , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Estrutura Molecular , Fármacos Neuroprotetores/síntese química , Fármacos Neuroprotetores/química , Agregados Proteicos/efeitos dos fármacos , Ratos , Ratos Wistar , Relação Estrutura-Atividade , Tacrina/química , Triptofano/químicaRESUMO
Inter-strand crosslinks (ICL) in the DNA are regarded to be the main toxic lesions induced by sulphur mustard (SM). We have followed the induction of ICL in the DNA of different organs of Wistar rats and Balb/c or NMRI mice by the percutaneous application of SM using the modified (reverse) comet assay. Significant amounts of ICL were found in Balb/C lymphocytes, in bone marrow and liver cells after the dose of 80â¯mg/kg. A dose-dependent amount of ICL was induced in rats, with efficient induction in lymphocytes and spleen cells already after 5â¯mg SM/kg, indicating a higher susceptibility of rats to the DNA-damaging effect of SM compared with mice. A significant induction of ICL in other tested tissues (liver, bone marrow, colon epithelium) was seen at the dose of 20â¯mg/kg. The induced ICL were removed from the DNA during 48â¯h except for rats at the dose of 80â¯mg/kg. In fact, we observed that ICL are almost completely repaired in tissues of rats receiving high lethal doses. Results suggest that the unhooking of ICL, which we followed with the comet assay, may lead to the formation of another toxic DNA lesion during the repair process.
Assuntos
Substâncias para a Guerra Química/toxicidade , Reagentes de Ligações Cruzadas/toxicidade , Dano ao DNA , Reparo do DNA , Gás de Mostarda/toxicidade , Animais , Colo/efeitos dos fármacos , Colo/metabolismo , Colo/patologia , Ensaio Cometa , Reagentes de Ligações Cruzadas/administração & dosagem , Adutos de DNA , Feminino , Humanos , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Linfócitos/efeitos dos fármacos , Linfócitos/metabolismo , Linfócitos/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Gás de Mostarda/administração & dosagem , Ratos , Ratos Wistar , Baço/efeitos dos fármacos , Baço/metabolismo , Baço/patologiaRESUMO
Bacterial biofilms pose a serious medical problem due to their significant resistance to antimicrobials, and staphylococci are recognized as the most frequent cause of biofilm-associated infections. The hop plant (Humulus lupulus L.) contains substances that have been determined to act as anti-infective agents against bacteria, mainly in planktonic form. Therefore, we decided to investigate the antibiofilm properties of H. lupulus L.-derived compounds (humulone, lupulone and xanthohumol) against a selected group of Staphylococcus spp., including methicillin-susceptible and resistant strains. All tested hop compounds were shown to possess antimicrobial properties against all tested staphylococci, both planktonic and biofilm-dwelling, with no significant difference between resistant and susceptible strains. All compounds lowered the number of bacterial cells released from the biofilm, with the strongest effect seen for lupulone, followed by xanthohumol. Moreover, lupulone and xanthohumol were not only able to penetrate the biofilm and reduce the number of bacteria within it, but their higher concentrations (â¼60 µg/mL for xanthohumol and â¼125 µg/mL for lupulone) reduced the number of surviving bacterial cells to zero.
Assuntos
Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Cicloexenos/farmacologia , Flavonoides/farmacologia , Propiofenonas/farmacologia , Staphylococcus/efeitos dos fármacos , Staphylococcus/crescimento & desenvolvimento , Terpenos/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Humulus/química , Testes de Sensibilidade Microbiana , Extratos Vegetais/farmacologia , Staphylococcus/genéticaRESUMO
BACKGROUND: In the last decade, the concept of uncharged reactivators potentially able to penetrate the CNS has been introduced as an alternative to the classic charged oxime reactivators. However, this concept brings with it several associated drawbacks such as higher lipophilicity, difficulty in administration, lower affinity to cholinesterases, and higher toxicity risk. OBJECTIVE: In this study, we compare data obtained for a set of five classic charged reactivators and a set of three recently published uncharged oximes supplemented by two novel ones. METHODS: This time, we used only in silico prediction and in vitro approaches. RESULTS: Our data showed that tested uncharged oximes have low affinity for cholinesterases, do not possess high reactivation potency, and certainly represent a greater toxicity risk due to higher lipophilicity. We assume that balanced physicochemical properties will be required for the successful treatment of OP poisoning. Nevertheless, the compound meeting such criteria and pinpointed in silico (K1280) failed in this particular case. CONCLUSION: From the presented data, it seems that the concept of uncharged reactivators will have to be modified, at least to improve the bioavailability and to satisfy requirements for in vivo administration.
Assuntos
Antídotos/farmacologia , Reativadores da Colinesterase/farmacologia , Oximas/farmacologia , Animais , Antídotos/síntese química , Antídotos/toxicidade , Barreira Hematoencefálica/efeitos dos fármacos , Butirilcolinesterase/metabolismo , Inibidores da Colinesterase/síntese química , Inibidores da Colinesterase/farmacologia , Inibidores da Colinesterase/toxicidade , Reativadores da Colinesterase/síntese química , Reativadores da Colinesterase/toxicidade , Simulação por Computador , Camundongos , Intoxicação por Organofosfatos/tratamento farmacológico , Organofosfatos/toxicidade , Oximas/síntese química , Oximas/toxicidade , Paraoxon/toxicidade , Ratos , Sarina/toxicidadeRESUMO
The purpose of this study was to evaluate the efficacy of potential candidate molecules or their combinations against strong alkylation agent sulfur mustard (SM) on the human lung alveolar epithelial cell line A-549. Candidate molecules were chosen on the basis of their previously observed protective effects in vitro. The tested compounds, including antioxidants, sulfhydryl or other sulfur-containing molecules, nitrogen-containing molecules, PARP inhibitors and a NO synthase inhibitor, were applicated 30min before SM treatment. The efficiency of candidate molecules to protect cells against DNA damage and cell death induced by SM was determined using single-cell gel electrophoresis (comet assay) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction by viable cells. The damage of DNA was assessed 1 and 24h after dose 50µM SM. Cell survival was assessed 24 and 72h after the exposure. To achieve maximal cytoprotection, combinations of selected compounds with sodium 2-mercaptoethane sulphonate (MESNA) were tested. We found significant protective effects by several drugs used individually and also in combination with MESNA. High protection was achieved by sodium thiosulphate, which was further potentiated when combined with MESNA. Most of the selected compounds or mixture provided only moderate genoptotection without having any effect towards cell viability.