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1.
Toxicol Lett ; 314: 164-171, 2019 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-31330168

RESUMO

In routine regulatory toxicology studies, the anatomic pathology endpoints are frequently the most significant element of the study data. They may profoundly influence subsequent clinical development and use of a test article, with implications for both human safety and for the fate of key commercial assets. Unfortunately (in common with other perceptual medical specialties), anatomic pathology data are also among the most subjective endpoints in regulatory toxicology studies - a challenge magnified by the fact that not only the diagnostic data but the anatomic pathologist's interpretation of it in their narrative report represent raw data within a regulated study (United States Federal Register, 1987). A strategy for minimizing and managing the risk of misdiagnosis/misinterpretation of pathology data is critical for any preclinical toxicology development program and is a collaborative approach between study directors, study monitors and toxicologists and toxicologic pathologists. The article provides a basic understanding of the sources of error and limitations of anatomic pathology evaluation, a starting point for troubleshooting and a basis for a sound management strategy. It describes common reasons for unexpected or inconsistent pathology findings and sets out to provide a framework for toxicologists to approach commissioning and critically evaluating their pathology data, and for identifying situations where additional third-party advice and review may be justified.


Assuntos
Confiabilidade dos Dados , Avaliação Pré-Clínica de Medicamentos/métodos , Microscopia , Patologia/métodos , Toxicologia/métodos , Animais , Comportamento Cooperativo , Humanos , Comunicação Interdisciplinar , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Medição de Risco , Fluxo de Trabalho
2.
Nat Commun ; 10(1): 2620, 2019 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-31197165

RESUMO

Conventional drug screens and treatments often ignore the underlying complexity of brain network dysfunctions, resulting in suboptimal outcomes. Here we ask whether we can correct abnormal functional connectivity of the entire brain by identifying and combining multiple neuromodulators that perturb connectivity in complementary ways. Our approach avoids the combinatorial complexity of screening all drug combinations. We develop a high-speed platform capable of imaging more than 15000 neurons in 50ms to map the entire brain functional connectivity in large numbers of vertebrates under many conditions. Screening a panel of drugs in a zebrafish model of human Dravet syndrome, we show that even drugs with related mechanisms of action can modulate functional connectivity in significantly different ways. By clustering connectivity fingerprints, we algorithmically select small subsets of complementary drugs and rapidly identify combinations that are significantly more effective at correcting abnormal networks and reducing spontaneous seizures than monotherapies, while minimizing behavioral side effects. Even at low concentrations, our polytherapy performs superior to individual drugs even at highest tolerated concentrations.


Assuntos
Epilepsias Mioclônicas/tratamento farmacológico , Modelos Biológicos , Rede Nervosa/efeitos dos fármacos , Fenômenos Fisiológicos do Sistema Nervoso/efeitos dos fármacos , Neurotransmissores/farmacologia , Algoritmos , Animais , Animais Geneticamente Modificados , Comportamento Animal/efeitos dos fármacos , Encéfalo/citologia , Encéfalo/diagnóstico por imagem , Encéfalo/efeitos dos fármacos , Encéfalo/fisiologia , Mapeamento Encefálico/métodos , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos/métodos , Sinergismo Farmacológico , Quimioterapia Combinada/métodos , Epilepsias Mioclônicas/genética , Epilepsias Mioclônicas/patologia , Ensaios de Triagem em Larga Escala/métodos , Humanos , Microscopia Confocal/métodos , Rede Nervosa/diagnóstico por imagem , Rede Nervosa/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Neurotransmissores/uso terapêutico , Peixe-Zebra
3.
Nat Commun ; 10(1): 2621, 2019 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-31197168

RESUMO

The high selectivity of the human blood-brain barrier (BBB) restricts delivery of many pharmaceuticals and therapeutic antibodies to the central nervous system. Here, we describe an in vitro microfluidic organ-on-a-chip BBB model lined by induced pluripotent stem cell-derived human brain microvascular endothelium interfaced with primary human brain astrocytes and pericytes that recapitulates the high level of barrier function of the in vivo human BBB for at least one week in culture. The endothelium expresses high levels of tight junction proteins and functional efflux pumps, and it displays selective transcytosis of peptides and antibodies previously observed in vivo. Increased barrier functionality was accomplished using a developmentally-inspired induction protocol that includes a period of differentiation under hypoxic conditions. This enhanced BBB Chip may therefore represent a new in vitro tool for development and validation of delivery systems that transport drugs and therapeutic antibodies across the human BBB.


Assuntos
Barreira Hematoencefálica/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Células Endoteliais/metabolismo , Microfluídica/instrumentação , Anticorpos/farmacologia , Astrócitos , Barreira Hematoencefálica/citologia , Avaliação Pré-Clínica de Medicamentos/instrumentação , Avaliação Pré-Clínica de Medicamentos/métodos , Endotélio Vascular/citologia , Humanos , Dispositivos Lab-On-A-Chip , Microfluídica/métodos , Microvasos/citologia , Pericitos , Permeabilidade , Células-Tronco Pluripotentes , Cultura Primária de Células/instrumentação , Cultura Primária de Células/métodos
4.
AAPS PharmSciTech ; 20(5): 210, 2019 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-31161269

RESUMO

Dorzolamide HCl (DRZ) ophthalmic drop is one of the most common glaucoma medications which rapidly eliminates after instillation leading to short residence time of the drug on cornea. The purpose of the present study is to develop a pH-triggered in situ gel system for ophthalmic delivery of DRZ for treatment of ocular hypertension. In this study, a 32 full factorial design was used for preparation of in situ gel formulations using different levels of Carbopol® and hydroxyl propyl methyl cellulose (HPMC). Rheological behavior, in vitro drug release, ex vivo corneal permeability, and IOP-lowering activity were investigated. DRZ solution (2% w/v) containing of 0.1% (w/v) Carbopol® and 0.1% (w/v) HPMC was selected as the optimal formulation considering its free flow under non-physiological conditions (initial pH and 25 ± 2°C) and transition to appropriate gel form under physiological circumstance (pH 7.4 and 34°C). This in situ gel presented the mucoadhesive property. Ex vivo corneal permeability of this combined solution was similar to those of DRZ solution. The developed formulation compared to the marketed drop (Biosopt®) and DRZ 2% solution had a better performance in intraocular pressure activity. The efficiency and long duration of IOP reduction could be due to the prolonged residence time of the in situ gel. The presence of Carbopol® as a pH triggered and mucoadhesive polymer causes to attach to the ocular mucosal surface for a long term.


Assuntos
Resinas Acrílicas/farmacocinética , Anti-Hipertensivos/farmacocinética , Portadores de Fármacos/farmacocinética , Derivados da Hipromelose/farmacocinética , Sulfonamidas/farmacocinética , Tiofenos/farmacocinética , Resinas Acrílicas/administração & dosagem , Resinas Acrílicas/síntese química , Administração Oftálmica , Animais , Anti-Hipertensivos/administração & dosagem , Anti-Hipertensivos/síntese química , Córnea/efeitos dos fármacos , Córnea/metabolismo , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/síntese química , Avaliação Pré-Clínica de Medicamentos/métodos , Liberação Controlada de Fármacos , Géis , Glaucoma/tratamento farmacológico , Glaucoma/metabolismo , Concentração de Íons de Hidrogênio , Derivados da Hipromelose/administração & dosagem , Derivados da Hipromelose/síntese química , Pressão Intraocular/efeitos dos fármacos , Pressão Intraocular/fisiologia , Masculino , Soluções Oftálmicas/administração & dosagem , Soluções Oftálmicas/síntese química , Soluções Oftálmicas/farmacocinética , Coelhos , Sulfonamidas/administração & dosagem , Sulfonamidas/síntese química , Suínos , Tiofenos/administração & dosagem , Tiofenos/síntese química
5.
Planta Med ; 85(9-10): 766-773, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31167297

RESUMO

Osteoporosis is a clinical condition characterized by low bone strength that leads to an increased risk of fracture. Strategies for the treatment of osteoporosis involve inhibition of bone resorption by osteoclasts and an increase of bone formation by osteoblasts. Here, we identified the extract derived from the stem part of Edgeworthia papyrifera that enhanced differentiation of MC3T3-E1 cells to osteoblast-like cells and inhibited osteoclast differentiation of RAW 264.7 cells in vitro. In support of our observation, rutin and daphnoretin, which were previously reported to inhibit osteoclast differentiation, were identified in E. papyrifera extract. In an animal model of osteoporosis, the ovariectomy-induced increases in bone resorption biomarkers such as pyridinoline and tartrate-resistant acid phosphatase were significantly reduced by E. papyrifera extract administration at 25.6 and 48.1%, respectively. Furthermore, the ovariectomy-induced bone loss in animal models of osteoporosis was significantly prevented by the administration of E. papyrifera in our study. Taking these observations into account, we suggest that E. papyrifera is an interesting candidate for further exploration as an anti-osteoporotic agent.


Assuntos
Osteoblastos/efeitos dos fármacos , Osteoclastos/efeitos dos fármacos , Osteoporose/tratamento farmacológico , Extratos Vegetais/farmacologia , Thymelaeaceae/química , Fosfatase Alcalina/metabolismo , Aminoácidos/urina , Animais , Biomarcadores/sangue , Biomarcadores/urina , Reabsorção Óssea/tratamento farmacológico , Diferenciação Celular/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Feminino , Camundongos , Camundongos Endogâmicos , Modelos Animais , Osteoporose/etiologia , Extratos Vegetais/análise , Células RAW 264.7 , Ratos Sprague-Dawley
6.
Nat Commun ; 10(1): 2082, 2019 05 07.
Artigo em Inglês | MEDLINE | ID: mdl-31064985

RESUMO

Single-cell resolution technologies warrant computational methods that capture cell heterogeneity while allowing efficient comparisons of populations. Here, we summarize cell populations by adding features' dispersion and covariances to population averages, in the context of image-based profiling. We find that data fusion is critical for these metrics to improve results over the prior alternatives, providing at least ~20% better performance in predicting a compound's mechanism of action (MoA) and a gene's pathway.


Assuntos
Biologia Computacional/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Análise de Célula Única/métodos , Células Cultivadas/efeitos dos fármacos , Análise de Dados , Conjuntos de Dados como Assunto , Descoberta de Drogas/métodos
7.
Anal Bioanal Chem ; 411(15): 3257-3268, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31089788

RESUMO

It has been demonstrated that triterpenes in Alismatis rhizoma (Zexie in Chinese, ZX) contributed to the lipid-lowering effect on high-fat diet-induced hyperlipidemia. Alisol B 23-acetate, one of the abundant triterpenes in ZX, was used as the marker of quality control for ZX in Chinese Pharmacopoeia, while it could not reflect the lipid-lowering effect because other triterpenes in ZX also had prominent medicinal efficacy. To identify the significantly bioactive triterpenes in ZX, a multiple reaction monitoring (MRM)-based characteristic chemical profile (CCP)-support vector machine (SVM) model was used to explore the relationship between triterpenes and lipid-lowering effect of ZX. Firstly, the content of 87 targeted triterpenes was quantified by the MRM-based CCP using UHPLC-QTRAP-MS/MS. Secondly, the lipid-lowering effect of 30 ZX samples was assessed by 3T3-L1 preadipocytes. Thirdly, 9 of the 87 triterpenes possessing high mean impact value were identified to have significant lipid-lowering effect via the particle swarm-optimized SVM model. The new SVM model constructed by the 9 triterpenes showed good prediction performance and the overall prediction accuracy reached 81.94%. Finally, the real activity of these triterpenes was partly confirmed and was consistent with the prediction of SVM. These results showed that the method for discovery of triterpenes with prominent lipid-lowering activity in ZX was reliable. The proposed method is expected to provide an efficient and rapid approach for screening of active component and drug discovery in traditional herbs. Graphical abstract.


Assuntos
Alismataceae/química , Hipolipemiantes/química , Hipolipemiantes/farmacologia , Rizoma/química , Máquina de Vetores de Suporte , Triterpenos/química , Triterpenos/farmacologia , Células 3T3-L1 , Adipogenia/efeitos dos fármacos , Animais , Cromatografia Líquida de Alta Pressão , Descoberta de Drogas/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Hiperlipidemias/tratamento farmacológico , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipídeos/análise , Camundongos , Espectrometria de Massas em Tandem
8.
BMC Bioinformatics ; 20(Suppl 7): 198, 2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-31074386

RESUMO

BACKGROUND: For treating a complex disease such as cancer, some effective means are needed to control biological networks that underlies the disease. The one-target one-drug paradigm has been the dominating drug discovery approach in the past decades. Compared to single target-based drugs, combination drug targets may overcome many limitations of single drug target and achieve a more effective and safer control of the disease. Most of existing combination drug targets are developed based on clinical experience or text-and-trial strategy, which cannot provide theoretical guidelines for designing and screening effective drug combinations. Therefore, systematic identification of multiple drug targets and optimal intervention strategy needs to be developed. RESULTS: We developed a strategy to screen the synergistic combinations of two drug targets in disease networks based on the classification of single drug targets. The method tried to identify the sensitivity of single intervention and then the combination of multiple interventions that can restore the disease network to a desired normal state. In our strategy of screening drug target combinations, we first classified all drug targets into sensitive and insensitive single drug targets. Then, we identified the synergistic and antagonistic of drug target combinations, including the combinations of sensitive drug targets, the combinations of insensitive drug target and the combination of sensitive and insensitive targets. Finally, we applied our strategy to Arachidonic Acid (AA) metabolic network and found 18 pairs of synergistic drug target combinations, five of which have been proven to be viable by biological or medical experiments. CONCLUSIONS: Different from traditional methods for judging drug synergy and antagonism, we propose the framework of how to enhance the efficiency by perturbing two sensitive targets in a combinatorial way, how to decrease the drug dose and therefore its side effect and cost by perturbing combinatorially a main sensitive target and an auxiliary insensitive target, and how to perturb two insensitive targets to realize the transition from a disease state to a healthy one which cannot be realized by perturbing each insensitive target alone. Although the idea is mainly applied to an AA metabolic network, the strategy holds for more general molecular networks such as combinatorial regulation in gene regulatory networks.


Assuntos
Algoritmos , Combinação de Medicamentos , Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos/métodos , Redes e Vias Metabólicas , Neoplasias/tratamento farmacológico , Sistemas de Liberação de Medicamentos , Humanos
9.
Enzyme Microb Technol ; 127: 65-69, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31088619

RESUMO

Matrix metalloproteinases (MMPs) are zinc-dependent proteases involved in the degradation of extracellular matrix proteins. As one of the isoforms, MMP-1 breaks down collagen, and its activity is known to be important in wound healing. Its timely and adequate level of expression is pivotal because MMP-1 is also involved in the damage or aging of skins as well as in certain types of cancers. Thus, both assaying the MMP-1 activity and developing its inhibitors are of great importance. We here developed an in-house assay system that gave us the high degree of freedom in screening peptide inhibitors of MMP-1. The assay system utilized a circularly permutated fusion of ß-lactamase and its inhibitory protein through an MMP-1-sensitive linker so that the activity of MMP-1 could be translated into that of ß-lactamase. As a proof of concept, we applied the developed assay system to initial screens of MMP-1 inhibitors and successfully identified one lead peptide that inhibited the collagenase activity of the enzyme.


Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Metaloproteinase 1 da Matriz/análise , Inibidores de Metaloproteinases de Matriz/isolamento & purificação , Inibidores de Metaloproteinases de Matriz/farmacologia , Peptídeos/isolamento & purificação , Peptídeos/farmacologia
10.
Nat Commun ; 10(1): 2265, 2019 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-31118422

RESUMO

Hepatitis delta virus (HDV) depends on the helper function of hepatitis B virus (HBV), which provides the envelope proteins for progeny virus secretion. Current infection-competent cell culture models do not support assembly and secretion of HDV. By stably transducing HepG2 cells with genes encoding the NTCP-receptor and the HBV envelope proteins we produce a cell line (HepNB2.7) that allows continuous secretion of infectious progeny HDV following primary infection. Evaluation of antiviral drugs shows that the entry inhibitor Myrcludex B (IC50: 1.4 nM) and interferon-α (IC50: 28 IU/ml, but max. 60-80% inhibition) interfere with primary infection. Lonafarnib inhibits virus secretion (IC50: 36 nM) but leads to a substantial intracellular accumulation of large hepatitis delta antigen and replicative intermediates, accompanied by the induction of innate immune responses. This work provides a cell line that supports the complete HDV replication cycle and presents a convenient tool for antiviral drug evaluation.


Assuntos
Antivirais/farmacologia , Vírus Delta da Hepatite/efeitos dos fármacos , Proteínas do Envelope Viral/metabolismo , Replicação Viral/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Células Hep G2 , Vírus da Hepatite B/metabolismo , Vírus Delta da Hepatite/fisiologia , Antígenos da Hepatite delta/metabolismo , Humanos , Imunidade Inata/efeitos dos fármacos , Concentração Inibidora 50
11.
Planta Med ; 85(9-10): 719-728, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31137047

RESUMO

Abnormal lipid metabolism, such as increased fatty acid uptake and esterification, is associated with nonalcoholic fatty liver disease (NAFLD). The aqueous extract of the aerial part of Angelica tenuissima Nakai (ATX) inhibited high-fat diet-induced hepatic steatosis in mice as well as oleic acid-induced neutral lipid accumulation in HepG2 cells. ATX decreased the mRNA and protein levels of CD36 and diglyceride acyltransferase 2 (DGAT2), the maturation of sterol regulatory element-binding proteins (SREBP), and the expression of the lipogenic target genes fasn and scd1. The ATX components, Z-ligustilide and n-butylidenephthalide, inhibited the expression of FATP5 and DGAT2 and thus oleic acid-induced lipid accumulation in HepG2 cells. These results suggest that ATX and its active components Z-ligustilide and n-butylidenephthalide inhibit fatty acid uptake and esterification in mice and have potential as therapeutics for NAFLD.


Assuntos
4-Butirolactona/análogos & derivados , Angelica/química , Metabolismo dos Lipídeos/efeitos dos fármacos , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Anidridos Ftálicos/farmacologia , 4-Butirolactona/isolamento & purificação , 4-Butirolactona/farmacologia , Animais , Dieta Hiperlipídica/efeitos adversos , Avaliação Pré-Clínica de Medicamentos/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Humanos , Lipogênese/efeitos dos fármacos , Masculino , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Ácido Oleico/farmacologia , Anidridos Ftálicos/isolamento & purificação , Componentes Aéreos da Planta/química , Extratos Vegetais/análise , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Proteína de Ligação a Elemento Regulador de Esterol 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo
12.
Environ Sci Pollut Res Int ; 26(21): 21871-21881, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31134551

RESUMO

Utilization of non-host plants semiochemicals to mediate insect behavior offers a promising opportunity for novel management of insect pests in field crops and fruits. Therefore, there is still a substantial opportunity for the development of natural prophylactic as an eco-friendly approach in the novel pest management programs. Sophora alopecuroides extract has been used as a natural pesticide in the control of agricultural and household pests, but the low persistence effect and rapid biodegradability limit its use on a wider scale in pest management programs. In this study, an emulsifiable concentrate formulation containing S. alopecuroides extract (SAE-EC) was developed with a simple procedure and evaluated for its ovicidal, antifeedant, and repellent effects against Diaphorina citri under laboratory and semi-field conditions. Our results indicated that SAE-EC at 15, 30, and 50 mg/mL concentrations provide complete protection against psyllids for a period of 96 h after application both under laboratory and semi-field conditions, while the aqueous methanolic extract of S. alopecuroides loses its persistence 48 h after application. Furthermore, the emulsifiable concentrate at 20 and 30 mg/mL concentrations, only 15.97% and 31.97% of eggs were able to hatch, and at similar concentrations, 72.86% and 85.5% of honeydew secretion were reduced as compared to the control. SAE-EC at 30 mg/mL concentration has not shown any phytotoxic symptoms on Murraya paniculata seedlings. Fourier transform infrared (FTIR) study revealed the presence of alkaloids in emulsifiable concentrate after 3 months of its preparation placed under ambient temperature. Furthermore, the particle size and polydispersity index (PDI) of the emulsifiable concentrate were also confirmed by dynamic light scattering (DLS). Our finding indicated that emulsifiable concentrate formulation prolongs the persistence of S. alopecuroides extract and enhances its efficacy both under laboratory and semi-field conditions. It has been concluded that the emulsifiable concentrate formulation containing S. alopecuroides extract might be developed as an eco-friendly novel prophylactic against citrus psyllid.


Assuntos
Hemípteros/efeitos dos fármacos , Repelentes de Insetos/farmacologia , Extratos Vegetais/farmacologia , Sophora/química , Alcaloides/análise , Animais , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos/métodos , Difusão Dinâmica da Luz , Emulsões/química , Emulsões/farmacologia , Hemípteros/fisiologia , Controle de Insetos/métodos , Repelentes de Insetos/administração & dosagem , Repelentes de Insetos/química , Murraya/efeitos dos fármacos , Extratos Vegetais/administração & dosagem , Extratos Vegetais/química , Plântula/efeitos dos fármacos , Espectroscopia de Infravermelho com Transformada de Fourier , Zigoto/efeitos dos fármacos
13.
Anal Chim Acta ; 1063: 110-116, 2019 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-30967174

RESUMO

BACKGROUND: Hyphenation of liquid chromatography (LC) with high-resolution mass spectrometry (HRMS) offers the potential to develop broad-spectrum screening procedures from low volumes of biological matrices. In parallel, dried blood spot (DBS) has become a valuable tool in the bioanalysis landscape to overcome conventional blood collection issues. Herein, we demonstrated the applicability of DBS as micro-sampling procedure for broad-spectrum toxicological screening. METHODS: A method was developed on a HRMS system in data dependant acquisition (DDA) mode using an extensive inclusion list to promote collection of relevant data. 104 real toxicology cases were analysed, and the results were cross-validated with one published and one commercial screening procedures. Quantitative MRM analyses were also performed on identified substances on a triple quadrupole instrument as a complementary confirmation procedure. RESULTS: The method showed limits of identification (LOIs) in appropriateness with therapeutic ranges for all the classes of interest. Applying the three screening approaches on 104 real cases, 271 identifications were performed including 14 and 6 classes of prescribed and illicit drugs, respectively. Among the detected substances, 23% were only detected by the proposed method. Based on confirmatory analyses, we demonstrated that the use of blood micro-samples did not impair the sensitivity allowing more identifications in the low concentration ranges. CONCLUSION: A LC-HRMS assay was successfully developed for toxicological screening of blood microsamples demonstrating a high identification power at low concentration ranges. The validation procedure and the analysis of real cases demonstrated the potential of this assay by supplementing screening approaches of reference.


Assuntos
Teste em Amostras de Sangue Seco , Avaliação Pré-Clínica de Medicamentos/instrumentação , Avaliação Pré-Clínica de Medicamentos/métodos , Drogas Ilícitas/sangue , Cromatografia Líquida , Humanos , Espectrometria de Massas em Tandem
14.
Phytomedicine ; 58: 152748, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31005722

RESUMO

BACKGROUND: Several species of Salvia are used as medicinal plants around the world. Biological activities of isolated compounds have been described, being diterpenes frequently responsible for the effects. PURPOSE: Isolation of diterpenes from Salvia uliginosa Benth. and evaluation of the antichemotactic and leishmanicidal activities of the isolated compounds. STUDY DESIGN: To isolate diterpenes from S. uliginosa and evaluate their antichemotactic and leishmanicidal activities in vitro. METHODS: The exudate of S. uliginosa was obtained by rapidly dipping the aerial parts in dichloromethane. The compounds were isolated by repeated column chromatography over silica gel. The effects on L. amazonensis growth, survival, DNA degradation, ROS generation, as well as the antichemotactic activity and cytotoxicity of the compounds towards human erythrocytes and macrophages were evaluated. RESULTS: A novel icetexane diterpene, isoicetexone (IsoICT) along with the known diterpenes icetexone (ICT), and 7-acetoxy-6,7-dihydroicetexone were isolated from the dichloromethane surface exudate of S. uliginosa. The structures were elucidated using NMR and MS experiments, and by comparison with previously reported data. IsoICT and ICT at low concentrations caused completely inhibition of neutrophils migration in vitro. In addition, IsoICT and ICT showed high leishmanicidal activity against L. amazonensis, induced ROS production in parasites and presented low cytotoxicity against macrophages and human erythrocytes, and moderate to high selectivity index. CONCLUSION: These data indicated that IsoICT and ICT exhibit potent antichemotactic and leishmanicidal effects. Further studies are needed in order to evaluate the in vivo activities as well as the toxicity of the compounds.


Assuntos
Antiprotozoários/química , Quimiotaxia/efeitos dos fármacos , Diterpenos/química , Salvia/química , Antiprotozoários/farmacologia , Células Cultivadas , Diterpenos/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Eritrócitos/efeitos dos fármacos , Humanos , Leishmania/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Estrutura Molecular , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Espécies Reativas de Oxigênio/metabolismo
15.
Methods Mol Biol ; 1971: 265-277, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30980309

RESUMO

The study of in vitro infections is essential to evaluate distinct aspects of Leishmania biology and also invaluable for more meaningful in vitro screening of promising chemical entities. Macrophage-like cells lines from different origins are amenable to Leishmania infection. Cell lines due to their stability and standardization potential are highly valued for their capacity to support reproducible infections and consistent data. In fact, these cells have been a mainstay of leishmaniasis research for more than 40 years. In this context, the human monocytic THP-1 cell line is commonly used as it can be differentiated with phorbol-12myristate-13-acetate (PMA) into macrophages that are susceptible to Leishmania infection. In this section, we will describe generalities concerning the use of cell lines for in vitro Leishmania infection using THP-1 derived macrophages and Leishmania infantum axenic amastigotes expressing luciferase associated to preclinical drug screening as example.


Assuntos
Antiprotozoários/farmacologia , Leishmania infantum/crescimento & desenvolvimento , Leishmaniose Visceral/tratamento farmacológico , Macrófagos/parasitologia , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Leishmaniose Visceral/metabolismo , Leishmaniose Visceral/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Células THP-1 , Acetato de Tetradecanoilforbol/farmacologia
16.
Eur J Pharm Sci ; 135: 103-112, 2019 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-31034983

RESUMO

Tuberculosis, caused by Mycobacterium tuberculosis has been one of the primal afflictions to human, and owing to the current scenario of drug resistance, newer drugs, and alternate targets are required to mitigate the disease. FtsZ is a GTP hydrolyzing protein, conserved in prokaryotes that plays a central role in Z-ring formation during cell division cytokinesis stage. This study employs the use of pharmacophore models derived from two different datasets based on Mtb-FtsZ GTPase inhibition and whole cell antibacterial activity, to virtually screen and shortlist novel compounds from In-house small molecule library as Mtb-FtsZ inhibitors and evaluate their in-vitro and ex-vivo activity. The results revealed Piperine (IC50 = 21.2 ±â€¯0.7 µM), 4-Bromo di-methoxy coumarin (IC50 = 13.0 ±â€¯1.6 µM) and Di-ethyl amino methyl coumarin (IC50 = 19.4 ±â€¯1.1) as potent Mtb-FtsZ GTPase inhibitors which showed considerable antibacterial activity (84.0 ±â€¯2.6 µM, 56.0 ±â€¯4.3 µM and 108 ±â€¯7.1 µM respectively) against M. smegmatis. They appear to be bacteriostatic, as well as treatment with these compounds led to a 3× increase in cell length of M. smegmatis. Further these molecules also altered the FtsZ gene expression by 3-fold when compared to untreated. In addition compound Aloin, an Aloe exudate showed potent Mtb-FtsZ inhibition (IC50 = 16.7 ±â€¯0.4 µM) but exhibited poor anti-mycobacterial activity (>500 µM).


Assuntos
Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Proteínas do Citoesqueleto/metabolismo , Mycobacterium smegmatis/efeitos dos fármacos , Alcaloides/farmacologia , Proteínas de Bactérias/genética , Benzodioxóis/farmacologia , Divisão Celular/efeitos dos fármacos , Cumarínicos/farmacologia , Citocinese , Proteínas do Citoesqueleto/genética , Bases de Dados de Compostos Químicos , Avaliação Pré-Clínica de Medicamentos/métodos , GTP Fosfo-Hidrolases/antagonistas & inibidores , Humanos , Simulação de Acoplamento Molecular , Mycobacterium smegmatis/citologia , Mycobacterium tuberculosis/metabolismo , Piperidinas/farmacologia , Alcamidas Poli-Insaturadas/farmacologia , Relação Estrutura-Atividade
17.
Eur J Pharm Biopharm ; 139: 186-196, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30951820

RESUMO

The preclinical evaluation of nasally administered drug candidates requires screening studies based on in vitro models of the nasal mucosa. The aim of this study was to evaluate the morpho-functional characteristics of the 3D MucilAir™ nasal model with a pharmacological focus on [ATP]-binding cassette (ABC) efflux transporters. We initially performed a phenotypic characterization of the MucilAir™ model and assessed its barrier properties by immunofluorescence (IF), protein mass spectrometry and examination of histological sections. We then focused on the functional expression of the ABC transporters P-glycoprotein (P-gp), multidrug resistance associated protein (MRP)1, MRP2 and breast cancer resistance protein (BCRP) in bidirectional transport experiments. The MucilAir™ model comprises a tight, polarized, pseudo-stratified nasal epithelium composed of fully differentiated ciliated, goblet and basal cells. These ABC transporters were all expressed by the cell membranes. P-gp and BCRP were both functional and capable of actively effluxing substrates. The MucilAir™ model could consequently represent a potent tool for evaluating the interaction of nasally administered drugs with ABC transporters.


Assuntos
Mucosa Nasal/metabolismo , Técnicas de Cultura de Tecidos/métodos , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Administração Intranasal , Células CACO-2 , Técnicas de Cultura de Células , Avaliação Pré-Clínica de Medicamentos/métodos , Corantes Fluorescentes/administração & dosagem , Corantes Fluorescentes/farmacocinética , Voluntários Saudáveis , Humanos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Mucosa Nasal/citologia , Proteínas de Neoplasias/metabolismo , Permeabilidade
18.
Carbohydr Polym ; 212: 252-259, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30832855

RESUMO

Niclosamide, previously used as an anthelmintic drug is currently being repurposed for its anticancer activity. Niclosamide is a brick like biopharmaceutical classification system (BCS) class II drug with poor aqueous solubility and dissolution consequently leading to low bioavailability. By considering the physicochemical properties and geometry of niclosamide, inclusion complex with cyclodextrin was prepared by freeze drying method and characterized using FT-IR, DSC, PXRD, and 1HNMR. In silico molecular modeling study was performed to study the possible interactions between niclosamide and cyclodextrin. The anticancer activity of niclosamide formulation was evaluated through in vitro cell cytotoxicity study using various cancer cell lines. The potential of niclosamide complex for improvement of the bioavailability was evaluated in male BALB/c mice. In vitro cytotoxicity studies indicated significantly higher cytotoxicity at lower concentrations and the pharmacokinetic studies showed significant improvement in Cmax and Tmax of niclosamide from cyclodextrin complex in comparison to pure niclosamide alone.


Assuntos
Antineoplásicos/síntese química , Ciclodextrinas/síntese química , Composição de Medicamentos/métodos , Reposicionamento de Medicamentos/métodos , Niclosamida/síntese química , Animais , Anticestoides/síntese química , Anticestoides/metabolismo , Antineoplásicos/metabolismo , Ciclodextrinas/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Células HCT116 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Niclosamida/metabolismo
19.
Carbohydr Polym ; 212: 59-66, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-30832881

RESUMO

A novel bilayer film of chitosan and konjac glucomannan were prepared by the two-step casting technique. Blend films were also prepared to investigate the interactions between the two polymers in the interfacial region of the bilayer structure. Scanning electron microscopy, Fourier transform infrared spectroscopy, and X-ray diffraction analysis showed that, unlike in the blends, the physicochemical properties of each biopolymer were preserved in the bilayer film. Differential scanning calorimetry and thermogravimetric analysis also indicated a good thermostability and miscibility for both polymers, probably due to strong hydrogen bonds between their polymer chains. Biological, mechanical and water vapor transmission tests showed a high biocompatibility, low cytotoxicity, and suitable mechanical and barrier properties of the bilayer films for wound dressing applications.


Assuntos
Bandagens , Quitosana/síntese química , Mananas/síntese química , Plantas , Candida albicans/efeitos dos fármacos , Candida albicans/fisiologia , Quitosana/farmacologia , Cromatografia em Gel/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Mananas/farmacologia , Resistência à Tração/efeitos dos fármacos , Resistência à Tração/fisiologia , Difração de Raios X/métodos
20.
AAPS PharmSciTech ; 20(4): 155, 2019 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-30924008

RESUMO

In this paper, a novel formulation of dual-release dry suspension of mosapride citrate (DRDS-MC) was designed which can be quickly released in the stomach while having sustained-release effect. Co-grinding mixture of mosapride citrate (MC) together with L-HPC as hydrophilic excipient was prepared in order to improve the solubility of MC. The co-grinding mixture was characterized by solubility studies, DSC, X-RD, SEM, FTIR, and size distribution before the preparation of the DRDS-MC. Then, the co-grinding mixture was used to prepare DRDS-MC via wet granulation method. The evaluation of DRDS-MC was focused on physicochemical properties, intestinal absorption, and pharmacokinetics. The results of DSC, X-RD, SEM, FTIR, and size distribution indicated that MC resides in co-grinding mixture with no crystalline changes, hydrogen bonds made L-HPC greatly improving the solubility of MC. Then, the dissolution of DRDS-MC reached 70% in pH 1.2 within 2 h, and the 12-h dissolution of MC in pH 6.8 was nearly 80%. The sedimentation volume after 3 h was 0.94 and redispersibility was good. The linear regression equation between in vitro release of DRDS-MC and intestinal absorption fraction in rats was: Y = 29.215 + 47.535*X (r = 0.952). At last, pharmacokinetic studies in beagle dogs demonstrated that DRDS-MC has prolonged effect compared with commercial formulation Gasmotin as a reference. All results indicated that the DRDS-MC could be quickly released in the stomach while having sustained-release effect.


Assuntos
Benzamidas/síntese química , Benzamidas/farmacocinética , Absorção Gastrointestinal/efeitos dos fármacos , Fármacos Gastrointestinais/síntese química , Fármacos Gastrointestinais/farmacocinética , Morfolinas/síntese química , Morfolinas/farmacocinética , Animais , Estudos Cross-Over , Preparações de Ação Retardada/síntese química , Preparações de Ação Retardada/farmacocinética , Cães , Avaliação Pré-Clínica de Medicamentos/métodos , Liberação Controlada de Fármacos/efeitos dos fármacos , Liberação Controlada de Fármacos/fisiologia , Excipientes/síntese química , Excipientes/farmacocinética , Absorção Gastrointestinal/fisiologia , Masculino , Distribuição Aleatória , Ratos , Solubilidade , Suspensões
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