RESUMO
Rheumatoid arthritis (RA) is an autoimmune disease involving T and B lymphocytes. Autoantibodies contribute to joint deterioration and worsening symptoms. Adenosine deaminase (ADA), an enzyme in purine metabolism, influences adenosine levels and joint inflammation. Inhibiting ADA could impact RA progression. Intracellular ATP breakdown generates adenosine, which increases in hypoxic and inflammatory conditions. Lymphocytes with ADA play a role in RA. Inhibiting lymphocytic ADA activity has an immune-regulatory effect. Synovial fluid levels of ADA are closely associated with the disease's systemic activity, making it a useful parameter for evaluating joint inflammation. Flavonoids, such as quercetin (QUE), are natural substances that can inhibit ADA activity. QUE demonstrates immune-regulatory effects and restores T-cell homeostasis, making it a promising candidate for RA therapy. In this review, we will explore the impact of QUE in suppressing ADA and reducing produced the inflammation in RA, including preclinical investigations and clinical trials.
Assuntos
Inibidores de Adenosina Desaminase , Artrite Reumatoide , Quercetina , Humanos , Adenosina , Adenosina Desaminase/metabolismo , Artrite Reumatoide/tratamento farmacológico , Inflamação/tratamento farmacológico , Quercetina/farmacologia , Inibidores de Adenosina Desaminase/farmacologiaRESUMO
Adenosine deaminase (ADA) is a Zn2+-containing enzyme that catalyzes the irreversible deamination of adenosine to inosine or deoxyadenosine to deoxyinosine. In addition to this enzymatic function, ADA mediates cell-to-cell interactions involved in lymphocyte co-stimulation or endothelial activation. ADA is implicated in cardiovascular pathologies such as atherosclerosis and certain types of cancers, including lymphoma and leukemia. To date, only two drugs (pentostatin and cladribine) have been approved for the treatment of hairy cell leukemia. In search of natural ADA inhibitors, we demonstrated the binding of selected phenolic compounds to the active site of ADA using molecular docking and molecular dynamics simulation. Our results show that phenolic compounds (chlorogenic acid, quercetin, and hyperoside) stabilized the ADA complex by forming persistent interactions with the catalytically essential Zn2+ ion. Furthermore, MM-GBSA ligand binding affinity calculations revealed that hyperoside had a comparable binding energy score (ΔG = - 46.56 ± 8.26 kcal/mol) to that of the cocrystal ligand in the ADA crystal structure (PDB ID: 1O5R) (ΔG = - 51.97 ± 4.70 kcal/mol). Similarly, chlorogenic acid exhibited a binding energy score (ΔG = - 18.76 ± 4.60 kcal/mol) comparable to those of the two approved ADA inhibitor drugs pentostatin (ΔG = - 14.54 ± 2.25 kcal/mol) and cladribine (ΔG = - 25.52 ± 4.10 kcal/mol) while quercetin was found to have modest binding affinity (ΔG = - 8.85 ± 7.32 kcal/mol). This study provides insights into the possible inhibitory potential of these phenolic compounds against ADA.
Assuntos
Inibidores de Adenosina Desaminase , Pentostatina , Inibidores de Adenosina Desaminase/farmacologia , Inibidores de Adenosina Desaminase/química , Simulação de Acoplamento Molecular , Quercetina/farmacologia , Cladribina , Ligantes , Ácido Clorogênico , Simulação de Dinâmica MolecularRESUMO
Altered RNA editing has been linked to several neurodevelopmental disorders, including autism spectrum disorder (ASD) and intellectual disability, in addition to depression, schizophrenia, some cancers, viral infections and autoimmune disorders. The human ADAR2 is a potential therapeutic target for managing these various disorders due to its crucial role in adenosine to inosine editing. This study applied consensus scoring to rank potential ADAR2 inhibitors after performing molecular docking with AutoDock Vina and Glide (Maestro), using a library of 35,161 compounds obtained from traditional Chinese medicine. A total of 47 compounds were predicted to be good binders of the human ADAR2 and had insignificant toxicity concerns. Molecular dynamics (MD) simulations, including the molecular mechanics Poisson-Boltzmann surface area (MM/PBSA) procedure, also emphasized the binding of the shortlisted compounds. The potential compounds had plausible binding free energies ranging from -81.304 to -1068.26 kJ/mol from the MM/PBSA calculations. ZINC000085511995, a naphthoquinone had more negative binding free energy (-1068.26 kJ/mol) than inositol hexakisphosphate (IHP) [-873.873 kJ/mol], an agonist and a strong binder of ADAR2. The potential displacement of IHP by ZINC000085511995 in the IHP binding site of ADAR2 could be explored for possible deactivation of ADAR2. Bayesian-based biological activity prediction corroborates the neuropharmacological, antineoplastic and antiviral activity of the potential lead compounds. All the potential lead compounds, except ZINC000014612330 and ZINC000013462928, were predicted to be inhibitors of various deaminases. The potential lead compounds also had probability of activity (Pa) > 0.442 and probability of inactivity (Pi) < 0.116 values for treating acute neurologic disorders, except for ZINC000085996580 and ZINC000013462928. Pursuing these compounds for their anti-ADAR2 activities holds a promising future, especially against neurological disorders, some cancers and viral infections caused by RNA viruses. Molecular interaction, hydrogen bond and per-residue decomposition analyses predicted Arg400, Arg401, Lys519, Trp687, Glu689, and Lys690 as hot-spot residues in the ADAR2 IHP binding site. Most of the top compounds were observed to have naphthoquinone, indole, furanocoumarin or benzofuran moieties. Serotonin and tryptophan, which are beneficial in digestive regulation, improving sleep cycle and mood, are indole derivatives. These chemical series may have the potential to treat neurological disorders, prion diseases, some cancers, specific viral infections, metabolic disorders and eating disorders through the disruption of ADAR2 pathways. A total of nine potential lead compounds were shortlisted as plausible modulators of ADAR2.
Assuntos
Inibidores de Adenosina Desaminase , Doenças Transmissíveis , Neoplasias , Humanos , Teorema de Bayes , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Inibidores de Adenosina Desaminase/farmacologiaRESUMO
Protein-ligand (GOLD) docking of the NCI compounds into the ligand-binding site of Plasmodium falciparum adenosine deaminase (PfADA) identified three most active azo compounds containing 4-[(4-hydroxy-2-oxo-1H-quinolin-3-yl) moiety. These compounds showed IC50 of 3.7-15.4 µM against PfADA, as well as inhibited the growth of P. falciparum strains 3D7 (chloroquine (CQ)-sensitive) and K1 (CQ-resistant) with IC50 of 1.8-3.1 and 1.7-3.6 µM, respectively. The identified compounds have structures similar to the backbone structure (4-N-(7-chloroquinolin-4-yl)) in CQ, and NSC45545 could mimic CQ by inhibiting the bioformation of hemozoin in parasitic food vacuole. The amount of in situ hemozoin in the ring-stage parasite was determined using a combination of synchrotron transmission Fourier transform infrared microspectroscopy and Principal Component Analysis. Stretching of the C-O bond of hemozoin propionate group measured at 1220-1210 cm-1 in untreated intraerythrocytic P. falciparum strains 3D7 and K1 was disappeared following treatment with 1.85 and 1.74 µM NSC45545, similar to those treated with 0.02 and 0.13 µM CQ, respectively. These findings indicate a novel dual function of 4-[(4-hydroxy-2-oxo-1H-quinolin-3-yl) azo compounds in inhibiting both PfADA and in situ hemozoin biocrystallization. These lead compounds hold promise for further development of new antimalarial therapeutics that could delay the onset of parasitic drug resistance.
Assuntos
Inibidores de Adenosina Desaminase , Antimaláricos , Compostos Azo , Plasmodium falciparum , Adenosina Desaminase , Antimaláricos/farmacologia , Compostos Azo/farmacologia , Biomineralização , Cloroquina/farmacologia , Resistência a Medicamentos , Ligantes , Plasmodium falciparum/efeitos dos fármacos , Inibidores de Adenosina Desaminase/farmacologiaRESUMO
Inhibitors of the enzyme adenosine monophosphate deaminase (AMPD) show interesting levels of herbicidal activity. An enzyme mechanism-based approach has been used to design new inhibitors of AMPD starting from nebularine (6) and resulting in the synthesis of 2-deoxy isonebularine (16). This compound is a potent inhibitor of the related enzyme adenosine deaminase (ADA; IC50 16 nM), binding over 5000 times more strongly than nebularine. It is proposed that the herbicidal activity of compound 16 is due to 5Ì-phosphorylation in planta to give an inhibitor of AMPD. Subsequently, an enzyme structure-based approach was used to design new non-ribosyl AMPD inhibitors. The initial lead structure was discovered by in silico screening of a virtual library against plant AMPD. In a second step, binding to AMPD was further optimised via more detailed molecular modeling leading to 2-(benzyloxy)-5-(imidazo[2,1-f][1,2,4]triazin-7-yl)benzoic acid (36) (IC50 300 nM). This compound does not inhibit ADA and shows excellent selectivity for plant over human AMPD.
Assuntos
AMP Desaminase/antagonistas & inibidores , Inibidores de Adenosina Desaminase/farmacologia , Adenosina Desaminase/metabolismo , Desenho de Fármacos , AMP Desaminase/metabolismo , Inibidores de Adenosina Desaminase/síntese química , Inibidores de Adenosina Desaminase/química , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
Adenosine deaminase (ADA) is an enzyme present in purine metabolic pathway. Its inhibitors are considered to be potent drug lead compounds against inflammatory and malignant diseases. This study aimed to test ADA inhibitory activity of some Streptomyces secondary metabolites by using computational and in vitro methods. The in silico screening of the inhibitory properties has been carried out using pharmacophore modeling, docking, and molecular dynamics studies. The in vitro validation of the selected antibiotics has been carried out by enzyme kinetics and fluorescent spectroscopic studies. The results indicated that novobiocin, an aminocoumarin antibiotic from Streptomyces niveus, has significant inhibition on ADA activity. Hence, the antibiotic can be used as a lead compound for the development of potential ADA inhibitors.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Adenosina Desaminase/metabolismo , Antibacterianos/farmacologia , Reposicionamento de Medicamentos , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Streptomyces/química , Inibidores de Adenosina Desaminase/química , Aminoglicosídeos/química , Aminoglicosídeos/farmacologia , Domínio Catalítico , Avaliação Pré-Clínica de Medicamentos , Ensaios Enzimáticos , Humanos , Análise dos Mínimos Quadrados , Ligantes , Novobiocina/química , Novobiocina/farmacologia , Relação Quantitativa Estrutura-Atividade , Espectrometria de FluorescênciaRESUMO
BACKGROUND: Measurement of adenosine deaminase (ADA) can provide information about cell-mediated immunity. This report's objective was to study the enzymatic activity of total ADA (tADA) and its isoenzymes ADA1 and ADA2 in canine, equine, porcine, and bovine serum and saliva and their changes in different inflammatory situations in each species. Besides, an automated method for ADA2 measurement was developed and validated. RESULTS: tADA was present in serum and saliva of healthy animals of the four species. Erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) concentration of 0.47 mM was needed for ADA1 inhibition in canine and porcine samples (serum and saliva) and bovine saliva, whereas for equine saliva 0.94 mM was needed. ADA2 activity was not detected in bovine serum and was very low or absent in equine serum and bovine saliva. An automated procedure to measure ADA2 consisting of adding EHNA to a commercial reagent for tADA measurement provided repetitive (coefficients of variation < 8.8% in serum and < 10% in saliva) and accurate (linearity of serial sample dilutions with R2 > 0.90) results, being equivalent to a manual incubation of the sample with EHNA at a similar concentration. Salivary tADA, as well as ADA1 and ADA2, were higher in dogs with leishmaniosis, horses with acute abdominal disease and pigs with lameness than in healthy animals. tADA and isoenzymes in saliva showed a positive significant correlation with serum ferritin in dogs (r = 0.602, P < 0.01; r = 0.555, P < 0.05; and r = 0.632, P < 0.01; respectively for tADA, ADA1 and ADA2) and serum C-reactive protein in pigs (r = 0.700, P < 0.01, for both tADA and ADA1; r = 0.770, P < 0.001, for ADA2), whereas salivary ADA2 significantly correlated with serum amyloid A in horses (r = 0.649, P < 0.01). In cows, salivary tADA and ADA1 significantly increased after calving, correlating with total white blood cell count (r = 0.487, P < 0.05, for both tADA and ADA1). CONCLUSIONS: The activity of total ADA and its different isoenzymes, can be measured in serum and saliva of dogs, horses, pigs and cows by a simple and fast procedure described in this report. When measured in saliva, these analytes correlated with other biomarkers of inflammation and it could potentially be used as a biomarkers of inflammation and immune activation in the species of this study.
Assuntos
Adenosina Desaminase/metabolismo , Bovinos/metabolismo , Cães/metabolismo , Cavalos/metabolismo , Inflamação/veterinária , Saliva/metabolismo , Suínos/metabolismo , Adenina/análogos & derivados , Adenosina Desaminase/sangue , Inibidores de Adenosina Desaminase , Animais , Automação , Biomarcadores/sangue , Biomarcadores/metabolismo , Bovinos/sangue , Ensaios Enzimáticos Clínicos/métodos , Ensaios Enzimáticos Clínicos/veterinária , Cães/sangue , Cavalos/sangue , Inflamação/sangue , Inflamação/enzimologia , Isoenzimas/sangue , Isoenzimas/metabolismo , Suínos/sangueRESUMO
This study was designed to investigate the effects and underlying mechanisms of Astaxanthin (AST) on high-fructose-induced hyperuricemia (HUA) from the perspectives of the uric acid (UA) synthesis and excretion in rat models. Following six weeks of a 10% fructose diet, the level of serum UA effectively decreased in the AST groups as compared to the model group. The enzymatic activities of xanthine oxidase (XOD) and adenosine deaminase (ADA) were significantly inhibited, and the mRNA expression levels of XOD and ADA significantly decreased after the AST administration. These results suggested that the AST reduced UA synthesis by inhibiting the mRNA expressions and enzyme activities of XOD and ADA, thereby contributing to HUA improvement. On the hand, the relative expressions of the mRNA and protein of kidney reabsorption transport proteins (GLUT9 and URAT1) were significantly down-regulated by AST, while that of the kidney secretion proteins (OAT1, OAT3 and ABCG2) were significantly up-regulated by AST. These results indicated that the AST promoted UA excretion by regulating the urate transport proteins, and thus alleviated HUA. This study suggested that the AST could serve as an effective alternative to traditional medicinal drugs for the prevention of fructose-induced HUA.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Adenosina Desaminase/metabolismo , Hiperuricemia/prevenção & controle , Proteínas de Membrana Transportadoras/efeitos dos fármacos , Ácido Úrico/sangue , Xantina Oxidase/antagonistas & inibidores , Adenosina Desaminase/genética , Animais , Biomarcadores/sangue , Biomarcadores/urina , Modelos Animais de Doenças , Frutose , Hiperuricemia/induzido quimicamente , Hiperuricemia/enzimologia , Rim/efeitos dos fármacos , Rim/metabolismo , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Ratos Sprague-Dawley , Reabsorção Renal/efeitos dos fármacos , Ácido Úrico/urina , Xantina Oxidase/genética , Xantina Oxidase/metabolismo , Xantofilas/farmacologiaRESUMO
Cordycepin has recently received increased attention owing to its extensive pharmacological activity. Adenosine deaminase (ADA) is widely distributed in mammalian blood and tissues; as a result, cordycepin is quickly metabolized upon entering into the body and converted into the inactive metabolite 3'-deoxyinosine, thus limiting its activity when administered alone. We herein present a novel ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) method for screening ADA inhibitors against the metabolism of cordycepin. Cordycepin and 3'-deoxyinosine were chosen as substrate and product, respectively. A proper separation was achieved for all analytes within 3 min. 3'-Deoxyinosine was quantified in the presence or absence of potential ADA inhibitors to evaluate ADA activity. The assay can simultaneously determine substrate and product, with the endogenous substance and ADA inhibitors added not interfering in its activity. After optimizing the enzymatic incubation and UHPLC-MS/MS conditions, Km and Vmax values for ADA deamination of cordycepin were 95.18 ± 7.85 µm and 363.90 ± 12.16 µmol/min/unit, respectively. Oleanolic acid and ursolic acid from Ligustri Lucidi Fructus were chosen as ADA inhibitors with half maximal inhibitory concentration values of 21.82 ± 0.39 and 18.41 ± 0.14 µm, respectively. A non-competitive inhibition model was constructed and this assay can be used to screen other potential ADA inhibitors quickly and accurately.
Assuntos
Inibidores de Adenosina Desaminase , Cromatografia Líquida de Alta Pressão/métodos , Desoxiadenosinas , Ligustrum/química , Extratos Vegetais , Inibidores de Adenosina Desaminase/análise , Inibidores de Adenosina Desaminase/química , Inibidores de Adenosina Desaminase/isolamento & purificação , Desoxiadenosinas/análise , Desoxiadenosinas/metabolismo , Descoberta de Drogas , Extratos Vegetais/análise , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Espectrometria de Massas em Tandem/métodos , Triterpenos , Ácido UrsólicoRESUMO
Adenosine deaminase (ADA) is an enzyme of purine metabolism that irreversibly converts adenosine to inosine or 2'deoxyadenosine to 2'deoxyinosine. ADA is active both inside the cell and on the cell surface where it was found to interact with membrane proteins, such as CD26 and adenosine receptors, forming ecto-ADA (eADA). In addition to adenosine uptake, the activity of eADA is an essential mechanism that terminates adenosine signaling. This is particularly important in cardiovascular system, where adenosine protects against endothelial dysfunction, vascular inflammation, or thrombosis. Besides enzymatic function, ADA protein mediates cell-to-cell interactions involved in lymphocyte co-stimulation or endothelial activation. Furthermore, alteration in ADA activity was demonstrated in many cardiovascular pathologies such as atherosclerosis, myocardial ischemia-reperfusion injury, hypertension, thrombosis, or diabetes. Modulation of ADA activity could be an important therapeutic target. This work provides a systematic review of ADA activity and anchoring inhibitors as well as summarizes the perspectives of their therapeutic use in cardiovascular pathologies associated with increased activity of ADA.
Assuntos
Inibidores de Adenosina Desaminase/uso terapêutico , Animais , Descoberta de Drogas/métodos , Proteínas de Choque Térmico/metabolismo , Humanos , Simulação de Dinâmica Molecular , Ligação Proteica , Água/químicaRESUMO
Neuroinflammation is one of the driving forces of progressive neurodegeneration in Parkinson's disease (PD). The metabolomics approach has been proved highly useful in identifying potential therapeutic targets. Here, to identify inflammation-relevant treatment targets for PD, mass spectrometry-based untargeted metabolomics was applied to characterize metabolic changes in the striatum of mice with double-hit PD induced by lipopolysaccharide plus 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Seven days after the final MPTP administration, metabolites from the purine metabolism pathway, including adenosine, 1-methyladenosine, adenine, inosine, hypoxanthine, xanthine, xanthosine, and guanosine, were found to be significantly dysregulated. The metabolite-protein interaction network and changes in the concentration ratio of these metabolites indicated that adenosine and adenosine deaminase (ADA; EC 3.5.4.4) were the most promising therapeutic targets and adenosine augmentation might be a rational approach to slow PD progression. These findings were then verified in a subacute MPTP-induced PD mouse model treated with ADA inhibition alone or in conjunction with antagonism of adenosine A2A receptors (A2A R). Behavioral, biochemical, and immunohistochemical analysis demonstrated that ADA inhibition significantly ameliorated the MPTP-mediated motor disabilities, dopamine depletion, and dopaminergic cell death. Significantly enhanced neuroprotective effects were further observed when the ADA inhibitor was utilized in conjunction with an A2A R antagonist. Together, our study indicated for the first time that ADA inhibitors protected against neurodegeneration induced by the neurotoxin MPTP, and ADA inhibitors in combination with A2A R antagonists showed additive antiparkinsonian effects.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Adenosina Desaminase/metabolismo , Antiparkinsonianos/farmacologia , Transtornos Parkinsonianos/metabolismo , Animais , Modelos Animais de Doenças , Masculino , Metabolômica , Camundongos , Camundongos Endogâmicos C57BL , Antagonistas de Receptores Purinérgicos P1/farmacologiaRESUMO
Plasmodium falciparum (Pf), a malarial pathogen, can only synthesize purine nucleotides employing a salvage pathway because it lacks de novo biosynthesis. Adenosine deaminase (ADA), one of the three purine salvage enzymes, catalyzes the irreversible hydrolytic deamination of adenosine to inosine, which is further converted to GMP and AMP for DNA/RNA production. In addition to adenosine conversion, Plasmodium ADA also catalyzes the conversion of 5'-methylthioadenosine, derived from polyamine biosynthesis, into 5'-methylthioinosine whereas the human enzyme is not capable of this function. Here we report the crystal structure of a surface engineered PfADA at a resolution of 2.48â¯Å, together with results on kinetic studies of PfADA wild-type and active site variants. The structure reveals a novel inosine binding pocket linked to a distinctive PfADA substructure (residues 172-179) derived from a non-conserved gating helix loop (172-188) in Plasmodium spp. and other ADA enzymes. Variants of PfADA and human (h) ADA active site amino acids were generated in order to study their role in catalysis, including PfADA- Phe136, -Thr174, -Asp176, and -Leu179, and hADA-Met155, equivalent to PfADA-Asp176. PfADA-Leu179His showed no effect on kinetic parameters. However, kinetic results of PfADA-Asp176Met/Ala mutants and hADA-Met155Asp/Ala showed that the mutation reduced adenosine and 5'-methylthioadenosine substrate affinity in PfADA and kcat in hADA, thereby reducing catalytic efficiency of the enzyme. Phe136Leu mutant showed increased Km (>10-fold) for both substrates whereas Thr174Ile/Ala only affected 5'-methylthioadenosine binding affinity. Together, the structure with the novel inosine binding pocket and the kinetic data provide insights for rational design of inhibitors against PfADA.
Assuntos
Adenosina Desaminase/química , Plasmodium falciparum/enzimologia , Proteínas de Protozoários/química , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Inibidores de Adenosina Desaminase/química , Inibidores de Adenosina Desaminase/farmacologia , Sequência de Aminoácidos , Substituição de Aminoácidos , Domínio Catalítico , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Inosina/metabolismo , Cinética , Modelos Moleculares , Mutagênese Sítio-Dirigida , Plasmodium falciparum/genética , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Homologia de Sequência de Aminoácidos , Especificidade por SubstratoRESUMO
Adenosine deaminase (ADA), which catalyzes the irreversible deamination of adenosine to inosine, is related to various human diseases such as tuberculous peritonitis and leukemia. Therefore, the method used to detect ADA activity and screen the effectiveness of various inhibitor candidates has important implications for the diagnosis treatment for various human diseases. A simple and rapid assay method for ADA, based on the enzymatic formation of a luminescent lanthanide complex, is proposed in this study. Inosine, an enzymatic product of ADA with stronger sensitization efficiency for Tb3+ than adenosine, produced a strong luminescence by forming an inosine-Tb3+ complex, and it enabled the direct monitoring of ADA activity in real-time. By introducing only Tb3+ to adenosine and ADA in the buffer, the enhancement of luminescence enabled the detection of a low concentration of ADA (detection limit 1.6 U/L). Moreover, this method could accurately determine the inhibition efficiency (IC50) of the known ADA inhibitor, erhythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), and the inhibition of ADA could be confirmed by the naked eye. Considering its simplicity, this assay could be extended to the high-throughput screening of various ADA inhibitor candidates.
Assuntos
Adenosina Desaminase/isolamento & purificação , Técnicas Biossensoriais , Complexos de Coordenação/química , Inosina/química , Adenosina/química , Adenosina Desaminase/química , Inibidores de Adenosina Desaminase/química , Humanos , Elementos da Série dos Lantanídeos/químicaRESUMO
The activity of a cell-surface ecto-adenosine deaminase (eADA) is markedly increased in the endothelial activation and vascular inflammation leading to decreased adenosine concentration and alterations in adenosine signalling. Depending on the specific pathway activated, extracellular purines mediate host cell response or regulate growth and cytotoxicity on tumour cells. The aim of this study was to test the effects of adenosine deaminase inhibition by 2'deoxycoformycin (dCF) on the breast cancer development. dCF treatment decreased a tumour growth and a final tumour mass in female BALB/c mice injected orthotopically with 4T1 cancer cells. dCF also counteracted cancer-induced endothelial dysfunction in orthotopic and intravenous 4T1 mouse breast cancer models. In turn, this low dCF dose had a minor effect on immune stimulation exerted by 4T1 cell implantation. In vitro studies revealed that dCF suppressed migration and invasion of 4T1 cells via A2a and A3 adenosine receptor activation as well as 4T1 cell adhesion and transmigration through the endothelial cell layer via A2a receptor stimulation. Similar effects of dCF were observed in human breast cancer cells. Moreover, dCF improved a barrier function of endothelial cells decreasing its permeability. This study highlights beneficial effects of adenosine deaminase inhibition on breast cancer development. The inhibition of adenosine deaminase activity by dCF reduced tumour size that was closely related to the decreased aggressiveness of tumour cells by adenosine receptor-dependent mechanisms and endothelial protection.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Progressão da Doença , Neoplasias Mamárias Animais/metabolismo , Neoplasias Mamárias Animais/patologia , Receptores Purinérgicos P1/metabolismo , Adenosina Desaminase/metabolismo , Animais , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Modelos Animais de Doenças , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/patologia , Espaço Extracelular/metabolismo , Feminino , Humanos , Neoplasias Mamárias Animais/sangue , Neoplasias Mamárias Animais/irrigação sanguínea , Camundongos Endogâmicos BALB C , Invasividade Neoplásica , Nucleotídeos/sangue , Pentostatina/farmacologia , Fenótipo , Migração Transendotelial e Transepitelial/efeitos dos fármacosRESUMO
Adenosine deaminase (ADA) is one of the major enzymes involved in purin metabolism, it has a significant role in cell growth and differentiation. Over-activity of ADA has been noticed in some pathology, like malignancy and inflammation and makes it an attractive target for the development of drugs for such diseases. In the present study, ADA inhibitory activity of morin, a bioactive flavonoid, was assessed through computational and biophysical methods. The enzyme kinetics data showed that morin is a competitive inhibitor of ADA. Binding energy calculated from ITC analysis was -7.11 kcal/mol. Interaction of morin with ADA was also studied using fluorescence quenching method. Molecular docking studies revealed the structural details of the interaction. Molecular dynamics study in explicit solvent was also conducted to assess the structural stability of protein ligand complex.
Assuntos
Inibidores de Adenosina Desaminase/química , Adenosina Desaminase/química , Flavonoides/química , Adenosina Desaminase/efeitos dos fármacos , Adenosina Desaminase/metabolismo , Inibidores de Adenosina Desaminase/uso terapêutico , Sítios de Ligação/efeitos dos fármacos , Flavonoides/uso terapêutico , Humanos , Inflamação/tratamento farmacológico , Cinética , Ligantes , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Complexos Multiproteicos/químicaRESUMO
Breast cancer is a heterogeneous and multifactorial disease with variable disease progression risk, and treatment response. Urtica dioica is a traditional herb used as an adjuvant therapeutic agent in cancer. In the present study, we have evaluated the effects of the aqueous extract of Urtica dioica on Adenosine deaminase (ADA) and Ornithine decarboxylase (ODC1) gene expression in MCF-7, MDA-MB-231, two breast cancer cell lines being estrogen receptor positive and estrogen receptor negative, respectively. Cell lines were cultured in suitable media. After 24 h, different concentrations of the extract were added and after 72 h, ADA and ODC1 gene expression as well as BCL2 and BAX apoptotic genes were assessed by Taqman real time PCR assay. Cells viability was assessed by MTT assay, and apoptosis was also evaluated at cellular level. The intra and extracellular levels of ODC1 and ADA enzymes were evaluated by ELISA. Results showed differential expression of ADA and ODC1 genes in cancer cell lines. In MCF-7 cell line, the expression level of ADA was upregulated in a dose-dependent manner but its expression did not change in MDA-MB cell line. ODC1 expression was increased in both examined cell lines. Also, increased level of the apoptotic BAX/BCL-2 ratio was detected in MCF-7 cells. These results demonstrated that Urtica dioica induces apoptosis in breast cancer cells by influencing ODC1 and ADA genes expression, and estrogen receptors. The different responses observed with these cell lines could be due to the interaction of Urtica dioica as a phytoestrogen with the estrogen receptor.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Inibidores da Ornitina Descarboxilase/farmacologia , Urtica dioica/química , Adenosina/metabolismo , Adenosina Desaminase/metabolismo , Inibidores de Adenosina Desaminase/química , Antineoplásicos Fitogênicos/química , Neoplasias da Mama/metabolismo , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Células MCF-7 , Ornitina Descarboxilase/metabolismo , Inibidores da Ornitina Descarboxilase/química , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Poliaminas/metabolismoRESUMO
Elevated circulating uric acid has been postulated to play an important pathophysiological role in estrogen-progestin combined oral contraceptive (COC)-induced hypertension and endothelial dysfunction. We hypothesized that disruption of glucoregulation and liver triglyceride (TG) accumulation induced by COC use would be abated by valproic acid (VPA) treatment through suppression of adenosine deaminase (ADA) and xanthine oxidase (XO) activities. Female Wistar rats aged 9-10 weeks were treated with a combination of estrogen-progestin COC steroids (1.0 µg ethinylestradiol and 5.0 µg levonorgestrel; p.o.) with or without VPA (100.0 mg/kg; p.o.) daily for 6 weeks. The result shows that the disrupted glucoregulation and associated elevated hepatic ADA activity, plasma and hepatic XO activity, uric acid (UA), TG/HDL-cholesterol, total cholesterol, and malondialdehyde induced by COC treatment were attenuated by VPA treatment. However, VPA did not have any effect on plasma aldosterone, corticosterone, ADA, circulating and hepatic free fatty acid. Our results demonstrate that suppression of plasma and hepatic XO activities, along with hepatic ADA activity and UA by VPA treatment, protects against disrupted glucoregulation and increased liver TG by COC independent of elevated corticosteroids. The findings imply that VPA would provide protection against the development of cardiometabolic disorder via inhibition of the ADA/XO/UA-mediated pathway.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Anticoncepcionais Orais Combinados/efeitos adversos , Hipertensão/tratamento farmacológico , Ácido Valproico/farmacologia , Xantina Oxidase/antagonistas & inibidores , Adenosina Desaminase/sangue , Adenosina Desaminase/metabolismo , Inibidores de Adenosina Desaminase/uso terapêutico , Aldosterona/sangue , Animais , Anticoncepcionais Orais Combinados/administração & dosagem , Corticosterona/sangue , Modelos Animais de Doenças , Estrogênios/administração & dosagem , Estrogênios/efeitos adversos , Feminino , Glucose/metabolismo , Humanos , Hipertensão/sangue , Hipertensão/induzido quimicamente , Hipertensão/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Progestinas/administração & dosagem , Progestinas/efeitos adversos , Ratos , Ratos Wistar , Triglicerídeos/metabolismo , Ácido Úrico/sangue , Ácido Valproico/uso terapêutico , Xantina Oxidase/metabolismoRESUMO
Adenosine deaminase (ADA) is an enzyme widely distributed from bacteria to humans. ADA is known as a potential therapeutic target for the treatment of lymphoproliferative disorders and cancer. Endophytes are endosymbionts, often bacteria or fungi, which live within plant tissues and internal organs or intercellular space. Endophytes have a broad variety of bioactive metabolites that are used for the identification of novel natural compounds. Here, 54 morphologically distinct endophyte strains were isolated from six plants such as Peganum harmala Linn., Rheum officinale Baill., Gentiana macrophylla Pall., Radix stephaniae tetrandrae, Myrrha, and Equisetum hyemale Linn. The isolated strains were used for the search of ADA inhibitors that resulted in the identification of the strain with the highest inhibition activity, Aspergillus niger sp. Four compounds were isolated from this strain using three-step chromatography procedure, and compound 2 was determined as the compound with the highest inhibition activity of ADA. Based on the results of 1H and 13C NMR spectroscopies, compound 2 was identified as 3-(4-nitrophenyl)-5-phenyl isoxazole. We showed that compound 2 was a new uncompetitive inhibitor of ADA with high cytotoxic effect on HepG2 and SMCC-7721 cells (the IC50 values were 0.347 and 0.380 mM, respectively). These results suggest that endophyte strains serve as promising sources for the identification of ADA inhibitors, and compound 2 could be an effective drug in the cancer treatment.
Assuntos
Inibidores de Adenosina Desaminase/química , Aspergillus niger/química , Endófitos/química , Plantas/microbiologia , Adenosina Desaminase/química , Adenosina Desaminase/metabolismo , Inibidores de Adenosina Desaminase/metabolismo , Aspergillus niger/genética , Aspergillus niger/isolamento & purificação , Aspergillus niger/metabolismo , Linhagem Celular , Endófitos/genética , Endófitos/isolamento & purificação , Endófitos/metabolismo , Humanos , Espectroscopia de Ressonância Magnética , Estrutura MolecularRESUMO
Adenosine deaminase (ADA), able to catalyze the irreversible deamination of adenosine into inosine, can be found in almost all tissues and plays an important role in several diseases. In this work, we developed a label-free fluorescence method for the detection of adenosine deaminase activity and inhibition. In the presence of ADA, ATP has been shown to be hydrolyzed. The ATP aptamer was shown to form a G-quadruplex/thioflavin T (ThT) complex with ThT and exhibited an obvious fluorescence signal. However, the ATP aptamer could bind with ATP and exhibited a low fluorescence signal because of the absence of ADA. This assay showed high sensitivity to ADA with a detection limit of 1 U/L based on an SNR of 3 and got a good linear relationship within the range of 1â»100 U/L with R² = 0.9909. The LOD is lower than ADA cutoff value (4 U/L) in the clinical requirement and more sensitive than most of the reported methods. This technique exhibited high selectivity for ADA against hoGG I, UDG, RNase H and λexo. Moreover, this strategy was successfully applied for assaying the inhibition of ADA using erythro-9-(2-hydroxy-3-nonyl) adenine (EHNA) and, as such, demonstrated great potential for the future use in the diagnosis of ADA-relevant diseases, particularly in advanced drug development.
Assuntos
Inibidores de Adenosina Desaminase/farmacologia , Adenosina Desaminase/análise , Adenosina Desaminase/metabolismo , Ensaios Enzimáticos/métodos , Técnicas Biossensoriais , Fluorescência , Humanos , Limite de Detecção , Fatores de TempoRESUMO
A series of terminal nonyl chain and nucleobase modified analogues of (+)-EHNA (III) were synthesized and evaluated for their ability to inhibit adenosine deaminase (ADA). The constrained carbon analogues of (+)-EHNA, 7a-7h, 10a-c, 12, 13, 14 and 17a-c appeared very potent with Ki values in the low nanomolar range. Thio-analogues of (+)-EHNA 24a-e wherein 5'C of nonyl chain replaced by sulfur atom found to be less potent compared to (+)-EHNA. Docking of the representative compounds into the active site of ADA was performed to understand structure-activity relationships. Compounds 7a (Ki: 1.1nM) 7b (Ki: 5.2nM) and 26a (Ki: 5.9nM) showed suitable balance of potency, microsomal stability and demonstrated better pharmacokinetic properties as compared to (+)-EHNA and therefore may have therapeutic potential for various inflammatory diseases, hypertension and cancer.