RESUMO
BACKGROUND: Considering the interesting role in the peptidoglycan biosynthesis pathway, the enzyme UDP-N-acetylglucosamine enolpyruvyl transferase is an attractive target to develop new antibacterial agents. It catalyzes the first key step of this pathway and its inhibition leads to bacterial cell death. Fosfomycin is known as the natural inhibitor of MurA. OBJECTIVE: The study aimed to introduce new inhibitors of MurA by virtual screening of different chemical compounds libraries, and test the best scored "virtual hits" against three pathogenic bacteria: Escherichia coli, Bacillus subtilis and Staphylococcus aureus. METHODS: A virtual screening of the structural analogues of fosfomycin downloaded from the Pub- Chem database was performed. Moreover, French National Chemical Library and ZINC database were also utilized to identify new structures different from fosfomycin. FlexX was the software used for this study. The antibacterial testing was divided into two methods: disk diffusion and broth dilution. RESULTS: A set of virtual hits was found to have better energy score than that of fosfomycin, seven of them were tested in vitro. In addition, the disk diffusion method explored four compounds that exhibited antibacterial activity: CID-21680357 (fosfomycin analogue), AB-00005001, ZINC04658565, and ZINC901335. The testing was continued by broth dilution method for both compounds CID-21680357 and ZINC901335 to determine their minimum inhibitory concentrations, and ZINC901335 had the best value with 457µg/ml against Staphylococcus aureus. CONCLUSION: Four compounds were found and proven in silico and in vitro to have antibacterial activity, namely CID-21680357, AB-00005001, ZINC04658565, and ZINC901335.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antibacterianos/química , Inibidores Enzimáticos/química , Fosfomicina/análogos & derivados , Simulação de Acoplamento Molecular/métodos , Alquil e Aril Transferases/metabolismo , Antibacterianos/farmacologia , Testes de Sensibilidade a Antimicrobianos por Disco-Difusão/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Inibidores Enzimáticos/farmacologia , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Fosfomicina/farmacologia , Testes de Sensibilidade Microbiana/métodosRESUMO
BACKGROUND: Streptococcus mutans and Streptococcus sanguinis are Gram-positive bacteria that cause dental caries. MurA enzyme acts as a catalyst in the formation of peptidoglycan in bacterial cell walls, making it ideal as an antibacterial target. Basil (Ocimum americanum) is an edible plant that is diverse and has been used as a herbal medicine for a long time. It has been reported that basil has a pharmacological effect as well as antibacterial activity. The purpose of this study was to identify antibacterial compounds in O. americanum and analyze their inhibition activity on MurA enzyme. METHODS: Fresh leaves from O. americanum were extracted with n-hexane and purified by a combination of column chromatography on normal and reverse phases together with in vitro bioactivity assay against S. mutans ATCC 25175 and S. sanguinis ATCC 10556, respectively, while in silico molecular docking simulation of lauric acid (1) was conducted using PyRx 0.8. RESULTS: The structure determination of antibacterial compound by spectroscopic methods resulted in an active compound lauric acid (1). The in vitro evaluation of antibacterial activity in compound 1 showed Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) values of 78.13 and 156.3 ppm and 1250 and 2500 ppm against S. sanguinis and S. mutans, respectively. Further analysis and in silico evaluation determined lauric acid (1) as MurA Enzyme inhibitor. Lauric acid (1) showed a binding affinity of -5.2 Kcal/mol, which was higher than fosfomycin. CONCLUSION: Lauric acid showed the potential as a new natural antibacterial agent through MurA inhibition in bacterial cell wall biosynthesis.
Assuntos
Antibacterianos/farmacologia , Cárie Dentária/tratamento farmacológico , Ácidos Láuricos/farmacologia , Ocimum basilicum/química , Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/metabolismo , Antibacterianos/isolamento & purificação , Antibacterianos/uso terapêutico , Cárie Dentária/microbiologia , Humanos , Ácidos Láuricos/isolamento & purificação , Ácidos Láuricos/uso terapêutico , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Folhas de Planta/química , Streptococcus mutans/efeitos dos fármacos , Streptococcus mutans/enzimologia , Streptococcus sanguis/efeitos dos fármacos , Streptococcus sanguis/enzimologiaRESUMO
Coenzyme Q10 (CoQ, ubiquinone) is a redox-active lipid endogenously synthesized by the cells. The final stage of CoQ biosynthesis is performed at the mitochondrial level by the 'complex Q', where coq2 is responsible for the prenylation of the benzoquinone ring of the molecule. We report that the competitive coq2 inhibitor 4-nitrobenzoate (4-NB) decreased the cellular CoQ content and caused severe impairment of mitochondrial function in the T67 human glioma cell line. In parallel with the reduction in CoQ biosynthesis, the cholesterol level increased, leading to significant perturbation of the plasma membrane physicochemical properties. We show that 4-NB treatment did not significantly affect the cell viability, because of an adaptive metabolic rewiring toward glycolysis. Hypoxia-inducible factor 1α (HIF-1α) stabilization was detected in 4-NB-treated cells, possibly due to the contribution of both reduction in intracellular oxygen tension and ROS overproduction. Exogenous CoQ supplementation partially recovered cholesterol content, HIF-1α degradation, and ROS production, whereas only weakly improved the bioenergetic impairment induced by the CoQ depletion. Our data provide new insights on the effect of CoQ depletion and contribute to shed light on the pathogenic mechanisms of ubiquinone deficiency syndrome.
Assuntos
Metabolismo Energético , Glicólise , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Ubiquinona/análogos & derivados , Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/metabolismo , Ataxia/metabolismo , Linhagem Celular Tumoral , Colesterol/metabolismo , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Doenças Mitocondriais/metabolismo , Debilidade Muscular/metabolismo , Nitrobenzoatos/farmacologia , Estabilidade Proteica/efeitos dos fármacos , Ubiquinona/antagonistas & inibidores , Ubiquinona/biossíntese , Ubiquinona/deficiência , Ubiquinona/metabolismoRESUMO
BACKGROUND: Streptococcus sanguinis is Gram-positive bacteria that contribute to caries. Many antibacterial agents are resistant against bacteria so that the discovery of new antibacterial agents is a crucial issue. Mechanism of antibacterial agents by disrupting cell wall bacteria is a promising target to be developed. One of the enzymes contributing to the cell wall is MurA enzyme. MurA is an enzyme catalyzing the first step of peptidoglycan biosynthesis in the cell wall formation. Inhibiting MurA is an effective and efficient way to kill the bacteria. Source of bioactive compounds including the antibacterial agent can be found in natural product such as herbal plant. Piper betle L. was reported to contain active antibacterial compounds. However, there is no more information on the antibacterial activity and molecular mechanism of P. betle's compound against S. sanguinis. PURPOSE: The study aims to identify antibacterial constituents of P. betle L. and evaluate their activities through two different methods including in vitro and in silico analysis. MATERIALS AND METHODS: The antibacterial agent was purified by bioactivity-guided isolation with combination chromatography methods and the chemical structure was determined by spectroscopic methods. The in vitro antibacterial activity was evaluated by disc diffusion and dilution methods while the in silico study of a compound binds on the MurA was determined using PyRx program. RESULTS: The antibacterial compound identified as allylpyrocatechol showed inhibitory activity against S. sanguinis with an inhibition zone of 11.85 mm at 1%, together with MIC and MBC values of 39.1 and 78.1 µg/mL, respectively. Prediction for molecular inhibition mechanism of allylpyrocatechols against the MurA presented two allylpyrocatechol derivatives showing binding activity of -5.4, stronger than fosfomycin as a reference with the binding activity of -4.6. CONCLUSION: Two allylpyrocatechol derivatives were predicted to have a good potency as a novel natural antibacterial agent against S. sanguinis through blocking MurA activity that causes disruption of bacterial cell wall.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antibacterianos/farmacologia , Catecóis/farmacologia , Inibidores Enzimáticos/farmacologia , Extratos Vegetais/farmacologia , Streptococcus sanguis/efeitos dos fármacos , Alquil e Aril Transferases/metabolismo , Antibacterianos/química , Antibacterianos/isolamento & purificação , Catecóis/química , Catecóis/isolamento & purificação , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Inibidores Enzimáticos/isolamento & purificação , Testes de Sensibilidade Microbiana , Estrutura Molecular , Piper betle/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Folhas de Planta/química , Streptococcus sanguis/enzimologia , Relação Estrutura-AtividadeRESUMO
BACKGROUND: A significant number of antibiotics are known to inhibit peptidoglycan synthesis in the cross-linking stage, while the drug fosfomycin is the only one known to inhibit MurA. Escalated antibiotic resistance has had an impact on the efficacy of fosfomycin, thus demanding the discovery of suitable substitutes with improved potential for MurA inhibition. The aim of this work is to isolate antibacterial compounds from Sarang Semut (Myrmecodia pendans) and to evaluate their antibacterial activity against pathogenic oral bacteria of Enterococcus faecalis ATCC 29212 and inhibitory activity against MurA enzyme. METHODS: The antibacterial compounds from Sarang Semut were isolated by a bioactivity-guided separation method with various solvents and combination of column chromatography on normal and reverse phases. The compounds with concentrations of 1000 and 5000 ppm were assessed against E. faecalis ATCC 29212 by agar well diffusion method, with chlorhexidine and fosfomycin being used as positive controls. RESULTS: Two antibacterial compounds isolated from Sarang Semut were identified as two new flavonoids derivates of 1 (10 mg) and 2 (4 mg). Both compounds were tested for antibacterial activities against E. faecalis. MIC values of compounds 1 and 2 were 8.15 and 8.05 mm at 1000 ppm and 8.62 and 8.55 mm at 5000 ppm, respectively. MBC values were 156 and 625 ppm for 1 and 625 and 2500 ppm for 2, respectively. In an inhibitory murA enzyme activity assay, compounds 1 and 2 were shown to inhibit the enzyme activity by IC50 values of 21.7 and 151.3 ppm. CONCLUSION: The study demonstrated that ethyl acetate fraction of Sarang Semut contained antibacterial flavonoids as active constituents that showed activity against E. faecalis. These results showed the plant's potential in herbal medicine and the development of new antibacterial agent for pathogenic dental caries.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antibacterianos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Enterococcus faecalis/efeitos dos fármacos , Flavonoides/farmacologia , Rubiaceae , Antibacterianos/química , Enterococcus faecalis/enzimologia , Enterococcus faecalis/crescimento & desenvolvimento , Flavonoides/química , Boca/microbiologiaRESUMO
Fusobacterium nucleatum plays a key role in several diseases such as periodontitis, gingivitis, appendicitis, and inflammatory bowel disease (IBD). The development of antibiotic resistance by this bacterium demands novel therapeutic intervention. Our recent study has reported UDP-N-acetylglucosamine 1-carboxyvinyltransferase (MurA) as one of the potential target proteins in F. nucleatum. In this study, we proposed two novel MurA inhibitors through in silico screening and evaluated their mode of inhibition by in vitro experiments. It was found that MurA structural arrangement (inside-out α/ß barrel) was stabilized by L/FXXXG(A) motif-based interactions. The protein was maintained in an open or substrate-free conformation due to repulsive forces between two parallelly arranged positively charged residues of domain I and II. In this conformation, we identified six best compounds that held key interactions with the substrate-binding pocket via a structure-based virtual screening of natural and chemical compound libraries. However, among these, only orientin and quercetin-3-O-d-glucuronide (Q3G) showed better interaction capability through consistent H-bond occupancy and lowest binding free energy during molecular dynamic simulations. In vitro inhibition studies evidenced the mixed and uncompetitive mode of inhibition by orientin and Q3G, respectively, with purified MurA protein. This explains the binding of orientin in both open and closed (substrate-bound) conformations of MurA, and Q3G binding in only closed conformation. Therefore, the Q3G binding mode was predicted on a MurA-substrate complex, which highlighted its constant H-bond with Cys118, a phosphoenolpyruvate (PEP) interacting residue. This suggests that Q3G may interrupt the PEP binding, thereby inhibiting the MurA activity. Thus, the current study discusses the structure of MurA and demonstrates the inhibitory action of two novel compounds.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Biologia Computacional/métodos , Inibidores Enzimáticos/farmacologia , Fusobacterium nucleatum/enzimologia , Alquil e Aril Transferases/química , Proteínas de Bactérias/antagonistas & inibidores , Simulação por Computador , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/química , Flavonoides/química , Flavonoides/farmacologia , Glucosídeos/química , Glucosídeos/farmacologia , Ligação de Hidrogênio , Modelos Moleculares , Simulação de Dinâmica Molecular , Conformação Proteica , Quercetina/análogos & derivados , Quercetina/química , Quercetina/farmacologia , Relação Estrutura-AtividadeRESUMO
Menaquinone (MK) biosynthesis pathway is a potential target for evaluating antimicrobials in gram-positive bacteria. Here, 1,4-dihydroxy-2-naphthoate prenyltransferase (MenA) was targeted to reduce methicillin-resistant Staphylococcus aureus (MRSA) growth. MenA inhibiting, long chain-based compounds were designed, synthesized and evaluated against MRSA and menaquinone utilizing bacteria in aerobic conditions. The results showed that these bacteria were susceptible to most of the compounds. Menaquinone (MK-4) supplementation rescued MRSA growth, suggesting these compounds inhibit MK biosynthesis. 3a and 7c exhibited promising inhibitory activities with MICs ranging 1-8 µg/mL against MRSA strains. The compounds did not facilitate small colony variant formation. These compounds also inhibited the biofilm growth by MRSA at high concentration. Compounds 3a, 6b and 7c displayed a promising extracellular bactericidal activity against MRSA at concentrations equal to and four-fold less than their respective MICs. We also observed cytokines released from THP-1 macrophages treated with compounds 3a, 6b and 7c and found decreases in TNF-α and IL-6 release and increase in IL-1ß. These data provide evidence that MenA inhibitors act as TNF-α and IL-6 inhibitors, raising the potential for development and application of these compounds as potential immunomodulatory agents.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antibacterianos/farmacologia , Biofilmes/crescimento & desenvolvimento , Citocinas/metabolismo , Inibidores Enzimáticos/farmacologia , Fatores Imunológicos/farmacologia , Staphylococcus aureus Resistente à Meticilina/crescimento & desenvolvimento , Antibacterianos/síntese química , Bactérias Aeróbias/efeitos dos fármacos , Bactérias Aeróbias/crescimento & desenvolvimento , Biofilmes/efeitos dos fármacos , Inibidores Enzimáticos/síntese química , Humanos , Fatores Imunológicos/síntese química , Macrófagos/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Viabilidade Microbiana/efeitos dos fármacos , Células THP-1 , Vitamina K 2/metabolismoRESUMO
The bacterial cell wall has long been a celebrated target for antibacterial drug discovery due to its critical nature in bacteria and absence in mammalian systems. At the heart of the cell wall biosynthetic pathway lies undecaprenyl phosphate (Und-P), the lipid-linked carrier upon which the bacterial cell wall is built. This study exploits recent insights into the link between late-stage wall teichoic acid inhibition and Und-P production, in Gram-positive organisms, to develop a cell-based small-molecule screening platform that enriches for inhibitors of undecaprenyl diphosphate synthase (UppS). Screening a chemical collection of 142,000 small molecules resulted in the identification of 6 new inhibitors of UppS. To date, inhibitors of UppS have generally shown off-target effects on membrane potential due to their physical-chemical characteristics. We demonstrate that MAC-0547630, one of the six inhibitors identified, exhibits selective, nanomolar inhibition against UppS without off-target effects on membrane potential. Such characteristics make it a unique chemical probe for exploring the inhibition of UppS in bacterial cell systems.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Proteínas de Bactérias/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/farmacologia , Alquil e Aril Transferases/genética , Alquil e Aril Transferases/metabolismo , Antibacterianos/química , Bactérias/enzimologia , Bactérias/genética , Bactérias/crescimento & desenvolvimento , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Parede Celular/efeitos dos fármacos , Parede Celular/enzimologia , Parede Celular/genética , Avaliação Pré-Clínica de Medicamentos , Testes de Sensibilidade Microbiana , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-AtividadeRESUMO
Expanding antibiotic use in clinical practice and emergence of bacterial resistance are fueling research efforts for the development of novel antibacterials. Underexploited or completely novel mechanistic approaches and biological targets are of especial interest. Undecaprenyl pyrophosphate synthase (UppS) is an essential enzyme in the biosynthesis of the bacterial cell wall. Although UppS is a validated target, no selective inhibitors occur in materia medica. Nevertheless, several native substrate analogues have been reported and used in enzyme kinetics studies or as pharmacological probes. The majority of small-molecule UppS inhibitors belong to the well-known class of bisphosphonates that are primarily used for treatment of bone resorption disorders. The most potent compound of this class has an IC50 of 0.59 µM. Inherently, the selectivity and suitability of such compounds for antimicrobial drug design can be questioned. Therefore, highthroughput and virtual screenings for non-bisphosphonate inhibitors were performed, and nanomolar inhibitors of UppS were identified, some with antimicrobial activities towards clinically relevant strains. The reported scaffolds belong to tetramic and tetronic acids with IC50 in the 100-nM range, and to dihydropyridines with IC50 down to 40 nM, all with antibacterial activity. Aryl-diketo acids are also potent inhibitors with MRSA antimicrobial activity, with the allosteric inhibitor methylisoxazole-4-carboxamide (IC50, 50 nM) active on several pathogenic Streptococcus pneumoniae strains. Clomiphene is a well-known oestrogen receptor modulator, and it has been reported to inhibit UppS. Although conclusions on the structure activity relationships cannot be drawn from all these data, these compound series represent an important contribution to the field of antibiotics.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Antibacterianos/farmacologia , Inibidores Enzimáticos/farmacologia , Isoxazóis/farmacologia , Streptococcus pneumoniae/efeitos dos fármacos , Alquil e Aril Transferases/metabolismo , Antibacterianos/química , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Isoxazóis/química , Testes de Sensibilidade Microbiana , Streptococcus pneumoniae/enzimologia , Relação Estrutura-AtividadeRESUMO
A second isoprene unit biosynthetic pathway, via 2-C-methyl-D-erythritol 4-phosphate (MEP), was discovered in the 1990s. We screened and isolated the cyclic dipeptide, maculosin, which is a probable novel MEP pathway inhibitor, from the culture broth of Bacillus subtilis strain KN07. To identify the target enzyme of maculosin, we applied an avidin-biotin complex method using biotinylated maculosin and the lysates of seven Escherichia coli strains, each overexpressing one enzyme of the MEP pathway, and performed quartz crystal microbalance (QCM) experiments using maculosin and each enzyme. The results indicate that IspG, the sixth enzyme on the MEP pathway, was bound to maculosin.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Alquil e Aril Transferases/metabolismo , Inibidores Enzimáticos/farmacologia , Eritritol/análogos & derivados , Fosfatos Açúcares/metabolismo , Avidina/metabolismo , Biotina/metabolismo , Biotinilação , Avaliação Pré-Clínica de Medicamentos , Eritritol/metabolismo , Escherichia coli K12/metabolismo , Peptídeos Cíclicos/farmacologia , Piperazinas/farmacologia , Staphylococcus aureus/metabolismoRESUMO
Uridinediphospho-N-acetylglucosamine enolpyruvyl transferase (MurA, E.C. 2.5.1.7) is an essential bacterial enzyme that catalyzes the first step of the cell wall biosynthetic pathway, which involves the transfer of an enolpyruvyl group from phosphoenolpyruvate to uridinediphospho-Nacetylglucosamine. In this study, novel inhibitors of Haemophilus influenzae MurA (Hi MurA) were identified using high-throughput screening of a chemical library from the Korea Chemical Bank. The identified compounds contain a quinoline moiety and have much lower effective inhibitory concentrations (IC(50)) than fosfomycin, a wellknown inhibitor of MurA. These inhibitors appear to covalently modify the sulfhydryl group of the active site cysteine (C117), since the C117D mutant Hi MurA was not inhibited by these compounds and excess dithiothreitol abolished their inhibitory activities. The increased mass value of Hi MurA after treatment with the identified inhibitor further confirmed that the active-site cysteine residue of Hi MurA is covalently modified by the inhibitor.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Inibidores Enzimáticos/isolamento & purificação , Inibidores Enzimáticos/farmacologia , Haemophilus influenzae/enzimologia , Cisteína/metabolismo , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/química , Ensaios de Triagem em Larga Escala , Concentração Inibidora 50 , Coreia (Geográfico) , Bibliotecas de Moléculas Pequenas , Reagentes de Sulfidrila/metabolismoRESUMO
Statin drugs inhibit 3-hydroxy-3-methylglutaryl CoA reductase, which reduces the synthesis of both cholesterol and isoprenoids (geranylgeranyl pyrophosphate and farnesyl pyrophosphate), with the latter being lipid molecules responsible for the posttranslational modification of small GTP-binding proteins such as Rho. Effects of statins, independent of lowering blood cholesterol levels, are thought to occur by inhibition of Rho/Rho kinase. The Rho kinase inhibitor Y-27632 has been reported to increase both erythrocyte deformability and low O2 tension-induced ATP release. Here, we tested the hypothesis that by inhibiting Rho/Rho kinase, simvastatin would increase both erythrocyte deformability and low O2 tension-induced ATP release. Male Sprague-Dawley rats were divided into two groups, control or simvastatin treated [simvastatin-supplemented chow (0.02%)], for 4 wk. Simvastatin treatment increased rat erythrocyte deformability compared with controls (n = 6, P < 0.05). However, erythrocytes of simvastatin-treated rats (n = 9, P < 0.05) exhibited impaired low O2 tension-induced ATP release. Similarly, the geranylgeranyl transferase inhibitor GGTI-2133 (10 µM) also increased deformability and impaired low O2 tension-induced ATP release in healthy human erythrocytes (P < 0.05). Interestingly, ATP release in response to mastoparan 7 (n = 7, P < 0.05), which directly activates Gi, and isoproterenol (n = 5, P < 0.05), which signals through Gs, was not altered by incubation with GGTI-2133. These results suggest that although statins increase erythrocyte deformability, likely by inhibiting geranylgeranylation, the finding that both statins and a geranylgeranyl transferase inhibitor attenuated low O2 tension-induced ATP release demonstrates that factors in addition to erythrocyte deformability are critical for ATP release in response to this physiological stimulus.
Assuntos
Trifosfato de Adenosina/metabolismo , Alquil e Aril Transferases/antagonistas & inibidores , Deformação Eritrocítica/efeitos dos fármacos , Imidazóis/farmacologia , Leucina/análogos & derivados , Naftalenos/farmacologia , Oxigênio/metabolismo , Sinvastatina/farmacologia , Agonistas Adrenérgicos beta/farmacologia , Adulto , Alquil e Aril Transferases/metabolismo , Animais , Anticolesterolemiantes/farmacologia , Anticolesterolemiantes/uso terapêutico , Colesterol/sangue , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Isoproterenol/farmacologia , Leucina/farmacologia , Masculino , Pessoa de Meia-Idade , Óxido Nítrico Sintase Tipo III/metabolismo , Pressão Parcial , Peptídeos/farmacologia , Ratos , Ratos Sprague-Dawley , Sinvastatina/uso terapêutico , Venenos de Vespas/farmacologia , Adulto Jovem , Quinases Associadas a rho/metabolismoRESUMO
Giant cell tumor (GCT) is the most common nonmalignant primary bone tumor reported in Hong Kong. It usually affects young adults between the ages of 20 and 40. This tumor is well known for its potential to recur following treatment. To date no effective adjuvant therapy exists for GCT. Our project aimed to study the effects of pamidronate (PAM), farnesyl transferase inhibitor (FTI-277), geranylgeranyl transferase inhibitor (GGTI-298), and their combinations on GCT stromal cells (SC). Individual treatment with PAM, FTI-277, and GGTI-298, inhibited the cell viability and proliferation of GCT SC in a dose-dependent way. Combination of FTI-277 with GGTI-298 caused synergistic effects in reducing cell viability, and its combination index was 0.49, indicating a strong synergism. Moreover, the combination of FTI-277 with GGTI-298 synergistically enhanced cell apoptosis and activated caspase-3/7, -8, and -9 activities. PAM induced cell-cycle arrest at the S-phase. The combination of PAM with GGTI-298 significantly increased OPG/RANKL mRNA ratio and activated caspase-3/7 activity. Our findings support that the combination of bisphosphonates with GGTIs or FTIs with GGTIs may be used as potential adjuvants in the treatment of GCT of bone.
Assuntos
Neoplasias Ósseas , Difosfonatos/farmacologia , Inibidores Enzimáticos/farmacologia , Tumor de Células Gigantes do Osso , Osteoprotegerina/genética , Ligante RANK/genética , Alquil e Aril Transferases/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Benzamidas/farmacologia , Conservadores da Densidade Óssea/farmacologia , Neoplasias Ósseas/tratamento farmacológico , Neoplasias Ósseas/patologia , Neoplasias Ósseas/fisiopatologia , Caspases/metabolismo , Divisão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Farnesiltranstransferase/antagonistas & inibidores , Expressão Gênica/efeitos dos fármacos , Tumor de Células Gigantes do Osso/tratamento farmacológico , Tumor de Células Gigantes do Osso/patologia , Tumor de Células Gigantes do Osso/fisiopatologia , Humanos , Metionina/análogos & derivados , Metionina/farmacologia , Pamidronato , Prenilação/efeitos dos fármacos , RNA Mensageiro/metabolismo , Fase S/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
The semisynthesis of xanthanolide derivatives is reported from xanthinin and 4-epi-isoxanthanol, two sesquiterpene lactones isolated from the crude chloroformic extract of the leaves of Xanthium macrocarpum DC. (Asteraceae) by liquid/liquid chromatography. In vitro evaluation of their protein farnesyltransferase (PFTase) inhibitory activity has been investigated. In contrast to other biological activities of xanthanolides, PFTase inhibition is not associated with the presence of the potentially toxic alpha-methylene-gamma-lactone function.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Clorofórmio/química , Inibidores Enzimáticos , Lactonas , Extratos Vegetais/química , Folhas de Planta/química , Sesquiterpenos/síntese química , Sesquiterpenos/isolamento & purificação , Sesquiterpenos/farmacologia , Xanthium/química , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/patologia , Ensaios Enzimáticos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Células HCT116 , Humanos , Lactonas/síntese química , Lactonas/química , Lactonas/farmacologia , Sesquiterpenos/química , Solventes/químicaRESUMO
The endocannabinoid system regulates cell proliferation in human breast cancer cells. Recently, we described that a metabolically stable anandamide analog, 2-methyl-2'-F-anandamide, by activation of CB1 receptors significantly inhibited cell proliferation of human breast cancer cell lines. In this study, we observed that the activation of the CB1 receptor, in two human mammary carcinoma cell lines, MDA-MB-231 and MCF7, caused the inhibition of 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase activity due to a reduction of HMG-CoA reductase transcript levels. The decrease of HMG-CoA reductase activity induced the inhibition of the prenylation of proteins, in particular of the farnesylation of Ras oncogenic protein involved in cell proliferation of these cell lines. We suggest that the inhibitory effect of anandamide analog on tumor cell proliferation could be related to the inhibition of Ras farnesylation.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Ácidos Araquidônicos/farmacologia , Neoplasias da Mama/patologia , Carcinoma/patologia , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Alcamidas Poli-Insaturadas/farmacologia , Antineoplásicos/farmacologia , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Moduladores de Receptores de Canabinoides/farmacologia , Carcinoma/genética , Carcinoma/metabolismo , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Endocanabinoides , Inibidores Enzimáticos/farmacologia , Feminino , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Hidroximetilglutaril-CoA Redutases/genética , Hidroximetilglutaril-CoA Redutases/metabolismo , Proteínas ras/genética , Proteínas ras/metabolismoRESUMO
Eighteen new meroterpene derivatives, dichrostachines A-R (1-18), have been isolated from the root and stem barks of Dichrostachys cinerea, and their structures determined by spectroscopic means and molecular modeling. From a biosynthetic standpoint these compounds arise from a Diels-Alder reaction between a labdane diene of the raimonol type and a flavonoid B-ring-derived quinone. The hypothesis was tested by the partial synthesis of similar compounds by simply mixing methyl communate and a synthetic flavonoid quinone. The hemisynthetic compounds were shown by NMR to have configurations different from those of the natural products, thus allowing a refinement of the biosynthesis hypothesis. Most of the compounds were assayed for their ability to inhibit the enzyme protein farnesyl transferase. The most active compounds exhibited IC50 and cytotoxicity values in the 1 microM range.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Fabaceae/química , Plantas Medicinais/química , Terpenos/isolamento & purificação , Terpenos/farmacologia , República Democrática do Congo , Estrutura Molecular , Ressonância Magnética Nuclear Biomolecular , Casca de Planta/química , Terpenos/químicaRESUMO
Geranylgeranylation is critical to the function of several proteins including Rho, Rap1, Rac, Cdc42, and G-protein gamma subunits. Geranylgeranyltransferase type I (GGTase-I) inhibitors (GGTIs) have therapeutic potential to treat inflammation, multiple sclerosis, atherosclerosis, and many other diseases. Following our standard workflow, we have developed and rigorously validated quantitative structure-activity relationship (QSAR) models for 48 GGTIs using variable selection k nearest neighbor (kNN), automated lazy learning (ALL), and partial least squares (PLS) methods. The QSAR models were employed for virtual screening of 9.5 million commercially available chemicals, yielding 47 diverse computational hits. Seven of these compounds with novel scaffolds and high predicted GGTase-I inhibitory activities were tested in vitro, and all were found to be bona fide and selective micromolar inhibitors. Notably, these novel hits could not be identified using traditional similarity search. These data demonstrate that rigorously developed QSAR models can serve as reliable virtual screening tools, leading to the discovery of structurally novel bioactive compounds.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Avaliação Pré-Clínica de Medicamentos/métodos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Relação Quantitativa Estrutura-Atividade , Algoritmos , Animais , Linhagem Celular , Técnicas de Química Combinatória , Reprodutibilidade dos TestesRESUMO
Potent inhibitors of MenA (1,4-dihydroxy-2-naphtoate prenyltrasferase) in Mycobacterium tuberculosis are identified, and are also effective in inhibiting growth of Mycobacterium tuberculosis at low concentrations. The MenA inhibitors possess common chemical structural features of (alkylamino)oalkoxyphenyl)(phenyl)methanones. Significantly, the MenA inhibitors can be synthesized in a few steps with high overall yields. The representative MenA inhibitors are highly effective in killing nonreplicating Mycobacterium tuberculosis that is evaluated by using the Wayne low oxygen model. In addition, a series of drug resistant Mycobacterium spp. are sensitive to the MenA inhibitors. The results are expected to be of significance in terms of discovering new lead compounds that can be developed into new drugs to combat unmet diseases caused by Mycobacterium tuberculosis.
Assuntos
Alquil e Aril Transferases/antagonistas & inibidores , Proteínas de Bactérias/antagonistas & inibidores , Dimetilaliltranstransferase/antagonistas & inibidores , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/enzimologia , Tuberculose/tratamento farmacológico , Tuberculose/microbiologia , Alquil e Aril Transferases/química , Proteínas de Bactérias/química , Benzofenonas/farmacologia , Catálise/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Dimetilaliltranstransferase/química , Farmacorresistência Bacteriana , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Isoniazida/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Estrutura Molecular , Mycobacterium/efeitos dos fármacos , Mycobacterium tuberculosis/metabolismo , Rifampina/farmacologia , Relação Estrutura-Atividade , Vitamina K 2/metabolismoRESUMO
Distinct clinicopathologic acute lymphoblastic leukemia (ALL) entities have been identified, resulting in the adoption of risk-oriented treatment approaches. In Philadelphia chromosome-positive ALL, the optimal treatment requires the addition of BCR-ABL tyrosine kinase inhibitors, as imatinib. Despite advances, the outcome remains poor, and novel agents are desperately required. The emergence of resistance to the Bcr-Abl inhibitor imatinib mesylate in patients with Philadelphia chromosome-positive (Ph+) ALL has prompted the development of second-generation compounds active against mutant forms, including dasatinib and nilotinib.
Assuntos
Antineoplásicos/uso terapêutico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Inibidores de Proteínas Quinases/uso terapêutico , Alquil e Aril Transferases/antagonistas & inibidores , Benzamidas , Benzenossulfonatos/uso terapêutico , Dasatinibe , Resistencia a Medicamentos Antineoplásicos , Proteínas de Fusão bcr-abl/genética , Humanos , Mesilato de Imatinib , Indóis/uso terapêutico , Niacinamida/análogos & derivados , Compostos de Fenilureia , Piperazinas/uso terapêutico , Piridinas/uso terapêutico , Pirimidinas/uso terapêutico , Pirróis/uso terapêutico , Sorafenibe , Sunitinibe , Tiazóis/uso terapêuticoRESUMO
The methanolic extract of the leaves of Liriodendron tulipifera was found to show inhibitory activity towards farnesyl protein transferase (FPTase). Bioassay-guided fractionation of the methanolic extract resulted in the isolation of lipiferolide, an inhibitor of FPTase. This compound inhibited the FPTase activity in a dose-dependent manner, and showed cell growth inhibitory activity against several tumor cells.