Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 363
Filtrar
1.
Science ; 373(6554): 541-547, 2021 07 30.
Artigo em Inglês | MEDLINE | ID: mdl-34326236

RESUMO

Repurposing drugs as treatments for COVID-19, the disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has drawn much attention. Beginning with sigma receptor ligands and expanding to other drugs from screening in the field, we became concerned that phospholipidosis was a shared mechanism underlying the antiviral activity of many repurposed drugs. For all of the 23 cationic amphiphilic drugs we tested, including hydroxychloroquine, azithromycin, amiodarone, and four others already in clinical trials, phospholipidosis was monotonically correlated with antiviral efficacy. Conversely, drugs active against the same targets that did not induce phospholipidosis were not antiviral. Phospholipidosis depends on the physicochemical properties of drugs and does not reflect specific target-based activities-rather, it may be considered a toxic confound in early drug discovery. Early detection of phospholipidosis could eliminate these artifacts, enabling a focus on molecules with therapeutic potential.


Assuntos
Antivirais/farmacologia , COVID-19/tratamento farmacológico , Reposicionamento de Medicamentos , Lipidoses/induzido quimicamente , Fosfolipídeos/metabolismo , SARS-CoV-2/efeitos dos fármacos , Células A549 , Animais , Antivirais/química , Antivirais/uso terapêutico , Antivirais/toxicidade , COVID-19/virologia , Cátions , Chlorocebus aethiops , Relação Dose-Resposta a Droga , Feminino , Humanos , Camundongos , Testes de Sensibilidade Microbiana , SARS-CoV-2/fisiologia , Tensoativos/química , Tensoativos/farmacologia , Tensoativos/toxicidade , Células Vero , Replicação Viral/efeitos dos fármacos
2.
Anal Chem ; 92(16): 11223-11231, 2020 08 18.
Artigo em Inglês | MEDLINE | ID: mdl-32664717

RESUMO

Lipid droplets (LDs) are organelles that play a major role in regulating the storage of neutral lipids. Dysregulation of LDs is associated with metabolic disorders, such as fatty liver diseases, obesity, diabetes, and atherosclerosis. We have developed LD-selective small-molecule fluorescence probes (probes 3 and 4) that are available for both one- and two-photon microscopy, employing live or fixed cells. We found that probes 3 and 4 sensitively detect the increased LDs in response to oleic acid or endoplasmic reticulum stress, both in cells and tissues of the liver. The narrow absorption and emission bands of probes 3 and 4 allow multicolor imaging for the study of the role of LDs in pathophysiology and LD-associated signaling by the coapplication of the probes for different organelles or antibodies against specific proteins. In addition, we show here, for the first time, that two-photon microscopy imaging using our LD-selective probes with LysoTracker provides a novel method for screening drugs to potentially induce steatosis and/or phospholipidosis.


Assuntos
Fígado Gorduroso/diagnóstico por imagem , Corantes Fluorescentes/química , Gotículas Lipídicas/metabolismo , Lipidoses/diagnóstico por imagem , Animais , Benzofuranos/síntese química , Benzofuranos/química , Benzofuranos/efeitos da radiação , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Fígado Gorduroso/induzido quimicamente , Corantes Fluorescentes/síntese química , Corantes Fluorescentes/efeitos da radiação , Células HeLa , Humanos , Lipidoses/induzido quimicamente , Camundongos , Microscopia de Fluorescência , Fótons
3.
Bioorg Med Chem Lett ; 30(9): 126933, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32044185

RESUMO

In this paper, we present the phospholipidosis-inducing potential (PLIP) of forty fragment-sized diamines derived from N-benzyl-4-(methylamino)piperidine and discuss the relationship between their PLIP and the physicochemical properties. Our results demonstrate that the previously reported methods are not suitable for predicting the PLIP of fragment-sized diamines; the second basic pKa can distinguish PLIP-positive diamines from PLIP-negative diamines more accurately than ClogP or most basic pKa. To the best of our knowledge, this is the first report describing the relationship between PLIP and second basic pKa.


Assuntos
Diaminas/farmacologia , Lipidoses/induzido quimicamente , Diaminas/efeitos adversos , Diaminas/química , Concentração de Íons de Hidrogênio , Estrutura Molecular , Relação Estrutura-Atividade
4.
Anal Bioanal Chem ; 411(30): 8023-8032, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31776643

RESUMO

Within drug development and pre-clinical trials, a common, significant and poorly understood event is the development of drug-induced lipidosis in tissues and cells. In this manuscript, we describe a mass spectrometry imaging strategy, involving repeated analysis of tissue sections by DESI MS, in positive and negative polarities, using MS and MS/MS modes. We present results of the detected distributions of the administered drug, drug metabolites, lipid molecules and a putative marker of lipidosis, di-docosahexaenoyl (22:6)-bis(monoacylglycerol) phosphate (di-22:6-BMP). A range of strategies have previously been reported for detection, isolation and identification of this compound, which is an isomer of di-docosahexaenoic (22:6 n-3) phosphatidylglycerol (di-22:6 PG), a commonly found lipid that acts as a surfactant in lung tissues. We show that MS imaging using MS/MS can be used to differentiate these compounds of identical mass, based upon the different distributions of abundant fragment ions. Registration of images of these fragments, and detected drugs and metabolites, is presented as a new method for studying drug-induced lipidosis in tissues. Graphical abstract.


Assuntos
Biomarcadores/metabolismo , Lipidoses/induzido quimicamente , Pulmão/diagnóstico por imagem , Espectrometria de Massas/métodos , Amiodarona/efeitos adversos , Animais , Antiarrítmicos/efeitos adversos , Masculino , Ratos Wistar , Roedores
5.
J Theor Biol ; 479: 37-47, 2019 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-31310757

RESUMO

Phospholipidosis is characterized by the presence of excessive accumulation of phospholipids in different tissue types (lungs, liver, eyes, kidneys etc.) caused by cationic amphiphilic drugs. Electron microscopy analysis has revealed the presence of lamellar inclusion bodies as the hallmark of phospholipidosis. Some phospholipidosis causing compounds can cause tissue specific inflammatory/retrogressive changes. Reliable and accurate in silico methods could facilitate early screening of phospholipidosis inducing compounds which can subsequently speed up the pharmaceutical drug discovery pipelines. In the present work, stacking ensembles are implemented for combining a number of different base learners to develop predictive models (a total of 256 trained machine learning models were tested) for phospholipidosis inducing compounds using a wide range of molecular descriptors (ChemMine, JOELib, Open babel and RDK descriptors) and structural alerts as input features. The best model consisting of stacked ensemble of machine learning algorithms with random forest as the second level learner outperformed other base and ensemble learners. JOELib descriptors along with structural alerts performed better than the other types of descriptor sets. The best ensemble model achieved an overall accuracy of 88.23%, sensitivity of 86.27%, specificity of 90.20%, mcc of 0.765, auc of 0.896 with 88.21 g-means. To assess the robustness and stability of the best ensemble model, it is further evaluated using stratified 10×10 fold cross validation and holdout testing sets (repeated 10 times) achieving 84.83% mean accuracy with 0.708 mean mcc and 88.46% mean accuracy with 0.771 mean mcc respectively. A comparison of different meta classifiers (Generalized linear regression, Gradient boosting machines, Random forest and Deep learning neural networks) in stacking ensemble revealed that random forest is the better choice for combining multiple classification models.


Assuntos
Lipidoses/diagnóstico , Modelos Estatísticos , Fosfolipídeos/metabolismo , Área Sob a Curva , Descoberta de Drogas , Humanos , Lipidoses/induzido quimicamente , Lipidoses/etiologia , Aprendizado de Máquina/normas , Sensibilidade e Especificidade
6.
Toxicol Pathol ; 47(1): 26-34, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30373479

RESUMO

Cationic amphiphilic drugs (CADs) can induce phospholipidosis (PLD) in organs/tissues. Several ophthalmic pharmaceuticals containing CADs are marketed and used in children. To investigate the effect of PLD on the developing cornea, chloroquine and amiodarone, which are representative CADs, were applied topically to the eyes of juvenile rabbits, and the effects in juvenile rabbits were compared with those in young adult rabbits. Diffuse corneal cloudiness was observed in chloroquine- and amiodarone-treated eyes. Histopathologically, vacuolation was observed in the corneal epithelium and keratocytes. On ultrastructural examination, these vacuoles contained multilamellar inclusion bodies, which are a characteristic of PLD. The size of the vacuoles in the corneal epithelium was reduced in juveniles compared with young adults. Cytoplasmic lamellar bodies and exocytosis in the corneal endothelium were observed in young adult rabbits but not in juvenile rabbits. This study revealed that topical application of chloroquine or amiodarone induces corneal PLD in juvenile and young adult rabbits. Corneal endothelial changes occurred only in young adult rabbits, but ophthalmological changes were similar between juveniles and young adults. The results of the study suggest that the effects of corneal PLD were similar among age groups based on risk assessment.


Assuntos
Envelhecimento/metabolismo , Amiodarona/toxicidade , Cloroquina/toxicidade , Córnea/efeitos dos fármacos , Lipidoses/induzido quimicamente , Fosfolipídeos/metabolismo , Administração Oftálmica , Envelhecimento/patologia , Animais , Córnea/metabolismo , Córnea/ultraestrutura , Modelos Animais de Doenças , Feminino , Corpos de Inclusão/metabolismo , Instilação de Medicamentos , Lipidoses/metabolismo , Lipidoses/patologia , Masculino , Coelhos
7.
Am J Pathol ; 188(9): 1993-2003, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29981744

RESUMO

Drug-induced phospholipidosis is a lysosomal storage disorder characterized by the excess accumulation of tissue phospholipids. Although azithromycin can be used to induce phospholipidosis, no experimental studies evaluating the relationship between drug accumulation and phospholipid localization have been performed. In this study, azithromycin was orally administered to rats for 7 days, and the relationship between drug and phospholipid accumulation was performed using imaging mass microscopy. The administration of azithromycin induced tubular epithelial vacuolation in the inner stripe of the outer medulla of the kidney, consistent with the lamellar bodies that are typical manifestations of drug-induced phospholipidosis. Azithromycin and phospholipid tissue levels were extensively elevated in the kidneys of azithromycin-treated rats. Imaging mass microscopy revealed that both azithromycin and its metabolites were found in the kidneys of azithromycin-treated rats but not in control animals. The vacuolated areas of the kidneys were primarily found in the inner stripe of the outer medulla, consistent with the areas of high azithromycin concentration. Azithromycin was colocalized with several phospholipids-phosphatidylinositol (18:0/20:4), phosphatidylethanolamine (18:0/20:4 and 16:0/20:4), and possibly didocosahexaenoyl (C22:6)-bis(monoacylglycerol) phosphate, a putative biomarker of drug-induced phospholipidosis. In summary, we found correlations between regions of kidney damage and the accumulation of azithromycin, its metabolites, and phospholipids using imaging mass microscopy. Such analyses may help reveal the mechanism and identify putative biomarkers of drug-induced phospholipidosis.


Assuntos
Azitromicina/toxicidade , Nefropatias/patologia , Lipidoses/patologia , Espectrometria de Massas/métodos , Microscopia Eletrônica de Transmissão/métodos , Fosfolipídeos/metabolismo , Animais , Antibacterianos/toxicidade , Processamento de Imagem Assistida por Computador , Nefropatias/induzido quimicamente , Nefropatias/complicações , Nefropatias/metabolismo , Lipidoses/induzido quimicamente , Lipidoses/complicações , Lipidoses/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley
8.
BMC Nephrol ; 19(1): 53, 2018 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-29510679

RESUMO

BACKGROUND: It is well-recognized that injection of iodinated radiographic contrast media (CM) sometimes causes acute renal injury via multiple mechanisms, such as vasoconstriction, toxicity on glomerular endothelium and tubular epithelium and so forth. CASE PRESENTATION: A 51-year-old man developed acute renal injury with proteinuria after CM administration. To our surprise, in his renal biopsy sample the myelin figure like structure was observed in glomerular endothelium and mesangial cells by transmission electron microscopy. However the patient didn't has any clinic clues of Fabry disease and other lysosomal storage disorders. Moreover in vitro cultured glomerular endothelial and mesangial cells we found CM triggers lipid aggregation along with the increased CD36 and decreased ABCA1 abundance. Thus this patient was administrated statin to correct the aberrant lipid trafficking, 2 months later at his next visit we found his renal function partially recovered with reduced proteinuria. CONCLUSIONS: Besides the well-known underlying mechanisms, CM may cause renal impairment by triggering the dysregulated transportation of lipid. Furthermore statin is suggested to be a very promising medicine to decrease side effects of CM.


Assuntos
Injúria Renal Aguda/induzido quimicamente , Meios de Contraste/efeitos adversos , Glomérulos Renais/efeitos dos fármacos , Lipidoses/induzido quimicamente , Células Mesangiais/efeitos dos fármacos , Injúria Renal Aguda/patologia , Humanos , Glomérulos Renais/patologia , Glomérulos Renais/ultraestrutura , Lipidoses/patologia , Masculino , Células Mesangiais/patologia , Células Mesangiais/ultraestrutura , Pessoa de Meia-Idade
9.
Toxicol Lett ; 284: 184-194, 2018 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-29248575

RESUMO

Phospholipidosis is a metabolic disorder characterized by intracellular accumulation of phospholipids. It can be caused by short-term or chronic exposure to cationic amphiphilic drugs (CADs). These compounds bind to phospholipids, leading to inhibition of their degradation and consequently to their accumulation in lysosomes. Drug-induced phospholipidosis (DIPL) is frequently at the basis of discontinuation of drug development and post-market drug withdrawal. Therefore, reliable human-relevant in vitro models must be developed to speed up the identification of compounds that are potential inducers of phospholipidosis. Here, hepatic cells derived from human skin (hSKP-HPC) were evaluated as an in vitro model for DIPL. These cells were exposed over time to amiodarone, a CAD known to induce phospholipidosis in humans. Transmission electron microscopy revealed the formation of the typical lamellar inclusions in the cell cytoplasm. Increase of phospholipids was already detected after 24 h exposure to amiodarone, whereas a significant increase of neutral lipid vesicles could be observed after 72 h. At the transcriptional level, the modulation of genes involved in DIPL was detected. These results provide a valuable indication of the applicability of hSKP-HPC for the quick assessment of drug-induced phospholipidosis in vitro, early in the drug development process.


Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Hepatócitos/efeitos dos fármacos , Lipidoses/induzido quimicamente , Fosfolipídeos/metabolismo , Pele/citologia , Células-Tronco/citologia , Amiodarona/toxicidade , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Células Hep G2 , Hepatócitos/ultraestrutura , Humanos , Lipidoses/genética , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Masculino , Fosfolipídeos/genética
10.
Mol Pharm ; 14(12): 4362-4373, 2017 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-29099189

RESUMO

Drug induced phospholipidosis (PLD) may be observed in the preclinical phase of drug development and pose strategic questions. As lysosomes have a central role in pathogenesis of PLD, assessment of lysosomal concentrations is important for understanding the pharmacokinetic basis of PLD manifestation and forecast of potential clinical appearance. Herein we present a systematic approach to provide insight into tissue-specific PLD by evaluation of unbound intracellular and lysosomal (reflecting acidic organelles) concentrations of two structurally related diprotic amines, GRT1 and GRT2. Their intratissue distribution was assessed using brain and lung slice assays. GRT1 induced PLD both in vitro and in vivo. GRT1 showed a high intracellular accumulation that was more pronounced in the lung, but did not cause cerebral PLD due to its effective efflux at the blood-brain barrier. Compared to GRT1, GRT2 revealed higher interstitial fluid concentrations in lung and brain, but more than 30-fold lower lysosomal trapping capacity. No signs of PLD were seen with GRT2. The different profile of GRT2 relative to GRT1 is due to a structural change resulting in a reduced basicity of one amino group. Hence, by distinct chemical modifications, undesired lysosomal trapping can be separated from desired drug delivery into different organs. In summary, assessment of intracellular unbound concentrations was instrumental in delineating the intercompound and intertissue differences in PLD induction in vivo and could be applied for identification of potential lysosomotropic compounds in drug development.


Assuntos
Diaminas/farmacologia , Lipidoses/induzido quimicamente , Modelos Biológicos , Animais , Encéfalo/metabolismo , Química Farmacêutica , Líquido Extracelular/metabolismo , Feminino , Células Hep G2 , Humanos , Pulmão/metabolismo , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Masculino , Modelos Animais , Modelos Químicos , Fosfolipídeos/metabolismo , Ratos , Ratos Sprague-Dawley , Ratos Wistar , Distribuição Tecidual
11.
Mol Pharm ; 14(12): 4346-4352, 2017 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-29077420

RESUMO

The drug-induced accumulation of phospholipids in lysosomes of various tissues is predominantly observed in regular repeat dose studies, often after prolonged exposure, and further investigated in mechanistic studies prior to candidate nomination. The finding can cause delays in the discovery process inflicting high costs to the affected projects. This article presents an in vitro imaging-based method for early detection of phospholipidosis liability and a hybrid approach for early detection and risk mitigation of phospolipidosis utilizing the in vitro readout with in silico model prediction. A set of reference compounds with phospolipidosis annotation was used as an external validation set yielding accuracies between 77.6% and 85.3% for various in vitro and in silico models, respectively. By means of a small set of chemically diverse known drugs with in vivo phospholipidosis annotation, the advantages of combining different prediction methods to reach an overall improved phospholipidosis prediction will be discussed.


Assuntos
Descoberta de Drogas/métodos , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/prevenção & controle , Lipidoses/induzido quimicamente , Lisossomos/metabolismo , Fosfolipídeos/metabolismo , Animais , Linhagem Celular Tumoral , Biologia Computacional/métodos , Simulação por Computador , Descoberta de Drogas/economia , Avaliação Pré-Clínica de Medicamentos/métodos , Técnicas In Vitro , Aprendizado de Máquina , Microscopia de Fluorescência
12.
J Toxicol Sci ; 42(5): 589-596, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28904294

RESUMO

It is important to consider susceptibility to drug-induced toxicity between animals and humans. Chimeric mice with a humanized liver are expected to predict hepatotoxicity in humans. Drug-induced phospholipidosis (DIPL), in which phospholipids accumulate, is a known entity. In this study, we examined whether chimeric mice can reveal species differences in DIPL. Changes in various phosphatidylcholine (PhC) molecules were investigated in the liver of chimeric mice after administering amiodarone, which induces phospholipidosis. Liquid chromatography-tandem mass spectrometry revealed that levels of PhCs tended to increase in the liver after administration of amiodarone. The liver of chimeric mice consists of human hepatocytes and residual mouse hepatocytes. We used imaging mass spectrometry (IMS) to evaluate the increase of PhCs in human and mouse hepatocytes after administration of amiodarone. IMS visualizes localization of endogenous and exogenous molecules in tissues. The IMS analysis suggested that the localized levels of several PhCs tended to be higher in the human hepatocytes than those in mouse hepatocytes, and PhC levels changed in response to amiodarone. Chimeric mice with a humanized liver will be useful to evaluate species differences in DIPL between mice and humans.


Assuntos
Amiodarona/toxicidade , Hepatócitos/metabolismo , Hepatócitos/transplante , Lipidoses/induzido quimicamente , Lipidoses/metabolismo , Fígado/metabolismo , Fosfatidilcolinas/metabolismo , Fosfolipídeos/metabolismo , Quimeras de Transplante , Animais , Cromatografia Líquida , Humanos , Camundongos , Especificidade da Espécie , Espectrometria de Massas em Tandem
13.
J Toxicol Sci ; 42(5): 641-650, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28904299

RESUMO

The utility of HepaRG cells as an in vitro cell-based assay system for predicting drug-induced phospholipidosis (PLD) was investigated. In experiment 1, 10 PLD-positive compounds and 11 PLD-negative compounds were selected. HepaRG cells were treated with each compound for 48 hr. In experiment 2, loratadine and desloratadine, a major metabolite of loratadine, were used to assess metabolic activation for PLD. HepaRG cells were treated with loratadine and desloratadine in the presence or absence of 500 µM 1-aminobenzotriazole (ABT), a broad CYP inhibitor, for 48 hr. After treatment with compounds in experiments 1 and 2, the relative fluorescence intensity (RFI) was measured using LYSO-ID Red dye to assess the PLD induction. In experiment 1, our cell-based assay system using HepaRG cells exhibited 100% sensitivity and 100% specificity for predicting drug-induced PLD. In experiment 2, loratadine increased the RFI in the PLD assay. However, the increase in the RFI was not observed in co-treatment with loratadine and ABT. In addition, desloratadine increased the RFI in the presence and absence of ABT. These results suggested that metabolic activation of loratadine may contribute to PLD in HepaRG cells. We newly demonstrated that HepaRG cells have a high ability for predicting drug-induced PLD. In addition, we newly showed that HepaRG cells may predict drug-induced PLD mediated by metabolic activation of loratadine. Thus, a cell-based assay system using HepaRG cells is a useful model for predicting drug-induced PLD.


Assuntos
Bioensaio/métodos , Lipidoses/induzido quimicamente , Lipidoses/metabolismo , Fosfolipídeos/metabolismo , Amicacina/toxicidade , Amiodarona/toxicidade , Amitriptilina/toxicidade , Clorpromazina/toxicidade , Feminino , Células Hep G2 , Humanos , Imipramina/efeitos adversos , Loratadina/análogos & derivados , Loratadina/toxicidade , Valor Preditivo dos Testes , Triazóis/toxicidade
14.
Int J Toxicol ; 36(5): 386-394, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28820006

RESUMO

Immunohistochemical staining for the lysosome-associated membrane protein 2 (LAMP-2) has been proposed previously as an alternative to electron microscopy to identify hepatic phospholipidosis. This study used LAMP-2 immunohistochemistry (IHC) to diagnose phospholipidosis in rats exhibiting renal tubular injury. Rats were administered toreforant, a histamine H4 receptor antagonist by oral gavage at a dose of 3, 10, or 100 mg/kg/d for 6 months. Hematoxylin and eosin staining revealed renal tubular epithelial cell vacuolation, hypertrophy, degeneration, and luminal dilation in the 100 mg/kg/d group animals. Renal tubular injury was confirmed using kidney injury marker 1 (KIM-1) IHC. The involvement of phosopholipidosis in the renal injury was investigated by LAMP-2. Adipophilin IHC was included to differentiate phospholipidosis from lipidosis. Increased LAMP-2 staining was observed in the 100 mg/kg/d group animals when compared to vehicle group animals. Lysosome-associated membrane protein-2 staining was most prominent in the outer stripe of the outer medulla where KIM-1 staining was also most prominent. By contrast, adipophilin staining was not increased. Phospholipidosis was also confirmed by electron microscopy. These data support the use of LAMP-2 IHC as a diagnostic tool and suggest an association between phospholipidosis and the renal tubular injury caused by toreforant.


Assuntos
Antagonistas dos Receptores Histamínicos/toxicidade , Nefropatias/diagnóstico , Rim/efeitos dos fármacos , Lipidoses/diagnóstico , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Fosfolipídeos/metabolismo , Receptores Histamínicos H4/antagonistas & inibidores , Injúria Renal Aguda , Animais , Moléculas de Adesão Celular/metabolismo , Feminino , Imuno-Histoquímica , Rim/metabolismo , Rim/patologia , Rim/ultraestrutura , Nefropatias/induzido quimicamente , Nefropatias/metabolismo , Nefropatias/patologia , Lipidoses/induzido quimicamente , Lipidoses/metabolismo , Lipidoses/patologia , Proteína 2 de Membrana Associada ao Lisossomo/análise , Masculino , Microscopia Eletrônica de Transmissão , Perilipina-2/análise , Perilipina-2/metabolismo , Ratos Sprague-Dawley
16.
Arch Toxicol ; 91(12): 3885-3895, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28551711

RESUMO

Recent reports have noted that a number of compounds that block the human Ether-à-go-go related gene (hERG) ion channel also induce phospholipidosis (PLD). To explore a hypothesis explaining why most PLD inducers are also hERG inhibitors, a modeling approach was undertaken with data sets comprised of 4096 compounds assayed for hERG inhibition and 5490 compounds assayed for PLD induction. To eliminate the chemical domain effect, a filtered data set of 567 compounds tested in quantitative high-throughput screening (qHTS) format for both hERG inhibition and PLD induction was constructed. Partial least squares (PLS) modeling followed by 3D-SDAR mapping of the most frequently occurring bins and projection on to the chemical structure suggested that both adverse effects are driven by similar structural features, namely two aromatic rings and an amino group forming a three-center toxicophore. Non-parametric U-tests performed on the original 3D-SDAR bins indicated that the distance between the two aromatic rings is the main factor determining the differences in activity; at distances of up to about 5.5 Å, a phospholipidotic compound would also inhibit hERG, while at longer distances, a sharp reduction of the PLD-inducing potential leaves only a well-pronounced hERG blocking effect. The hERG activity itself diminishes after the distance between the centroids of the two aromatic rings exceeds 12.5 Å. Further comparison of the two toxicophores revealed that the almost identical aromatic rings to amino group distances play no significant role in distinguishing between PLD and hERG activity. The hypothesis that the PLD toxicophore appears to be a subset of the hERG toxicophore explains why about 80% of all phospholipidotic chemicals (the remaining 20% are thought to act via a different mechanism) also inhibit the hERG ion channel. These models were further validated in large-scale qHTS assays testing 1085 chemicals for their PLD-inducing potential and 1570 compounds for hERG inhibition. After removal of the modeling and experimental inconclusive compounds, the area under the receiver-operating characteristic (ROC) curve was 0.92 for the PLD model and 0.87 for the hERG model. Due to the exceptional ability of these models to recognize safe compounds (negative predictive values of 0.99 for PLD and 0.94 for hERG were achieved), their use in regulatory settings might be particularly useful.


Assuntos
Canal de Potássio ERG1/antagonistas & inibidores , Lipidoses/induzido quimicamente , Bloqueadores dos Canais de Potássio/química , Bloqueadores dos Canais de Potássio/farmacologia , Relação Quantitativa Estrutura-Atividade , Algoritmos , Humanos , Modelos Moleculares , Fosfolipídeos/metabolismo , Bloqueadores dos Canais de Potássio/efeitos adversos , Reprodutibilidade dos Testes
17.
Chem Res Toxicol ; 30(5): 1145-1156, 2017 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-28398727

RESUMO

In vitro safety assessment in early drug discovery represents an important step to detect potential safety-related liabilities. It reduces late stage attrition and allows candidate optimization. In this study, we report on the use of the LogDBPL assay (a recently published assay for the determination of drug distribution coefficients between an aqueous phase and porcine brain polar lipids extract) for phospholipidosis (PLD) risk evaluation. The LogDBPL parameter was first compared to the effective permeability in the parallel artificial membrane permeability assay (PAMPA), previously reported as correlating with PLD risk. Subsequently, the LogDBPL for a set of 234 drugs with known PLD effect was measured, representing the largest data set of LogDBPL data published so far, and the correlation with phospholipid accumulation was further investigated. In addition, a comparison with other in silico methods based on physicochemical parameters is reported. Results showed that LogDBPL is an efficient descriptor to assess PLD risk, especially when corrected using the pKa value of compounds, being superior to the distribution coefficient in octanol, LogDOCT, and the effective permeability in the PAMPA assay. A multivariate statistical analysis approach was finally used to better define the intrinsic features of LogDBPL, whose effect proved to be highly similar to that of volume of distribution in silico when used to predict brain distribution and PLD.


Assuntos
Encéfalo/metabolismo , Lipidoses/induzido quimicamente , Fosfolipídeos/metabolismo , Animais , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Preparações Farmacêuticas/classificação , Análise de Componente Principal , Suínos
18.
Artigo em Inglês | MEDLINE | ID: mdl-28285021

RESUMO

Cationic amphiphilic drugs (CADs) can induce the hyperaccumulation of phospholipids in cells and tissues. This side effect, which is known as drug-induced phospholipidosis, is sometimes problematic in the development and clinical use of CADs. It is known that CADs generally interact with phospholipids via both hydrophobic and acid-base interactions, and CADs with the larger affinity to phospholipid exhibit the larger induction risk. To develop a chromatographic assay system to predict the phospholipidosis-inducing potential with considering the acid-base interaction between CAD and phosphate group of phospholipid, hydrophilic interaction chromatographic (HILIC) methods were tested in this study. First, a PC HILIC column with phosphocholine groups on a packed material was used. The acid-base or other hydrophilic interactions to the stationary phase differed among basic drugs, and retention to the PC HILIC column did not accurately reflect the induction potential of phospholipidosis. As an alternative HILIC approach, the elution of CADs with the phosphate buffer from an amide column was tested. The elution effect, which is expressed as ratio of retention factors between different phosphate content in the mobile phase, closely correlated with the induction potential. Using the elution effect and retention factor to a reversed-phase HPLC column, the phospholipidosis-inducing drugs were clearly discriminated from the non-inducers. These results suggest that the proposed chromatographic approach can screen phospholipidosis-inducing drugs.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Lipidoses/induzido quimicamente , Preparações Farmacêuticas/metabolismo , Fosfatos/metabolismo , Fosfolipídeos/metabolismo , Humanos , Interações Hidrofóbicas e Hidrofílicas , Lipidoses/metabolismo , Preparações Farmacêuticas/química
19.
J Appl Toxicol ; 37(8): 943-953, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28138993

RESUMO

Lipid profiling has emerged as an effective approach to not only screen disease and drug toxicity biomarkers but also understand their underlying mechanisms of action. Tamoxifen, a widely used antiestrogenic agent for adjuvant therapy against estrogen-positive breast cancer, possesses side effects such as hepatic steatosis and phospholipidosis (PLD). In the present study, we administered tamoxifen to Sprague-Dawley rats and used lipidomics to reveal tamoxifen-induced alteration of the hepatic lipid profile and its association with the plasma lipid profile. Treatment with tamoxifen for 28 days caused hepatic PLD in rats. We compared the plasma and liver lipid profiles in treated vs. untreated rats using a multivariate analysis to determine differences between the two groups. In total, 25 plasma and 45 liver lipids were identified and altered in the tamoxifen-treated group. Of these lipids, arachidonic acid (AA)-containing phosphatidylcholines (PCs), such as PC (17:0/20:4) and PC (18:1/20:4), were commonly reduced in both plasma and liver. Conversely, tamoxifen increased other phosphoglycerolipids in the liver, such as phosphatidylethanolamine (18:1/18:1) and phosphatidylinositol (18:0/18:2). We also examined alteration of AA-containing PCs and some phosphoglycerolipids in the pre-PLD stage and found that these lipid alterations were initiated before pathological alteration in the liver. In addition, changes in plasma and liver levels of AA-containing PCs were linearly associated. Moreover, levels of free AA and mRNA levels of AA-synthesizing enzymes, such as fatty acid desaturase 1 and 2, were decreased by tamoxifen treatment. Therefore, our study demonstrated that AA-containing PCs might have potential utility as novel and predictive biomarkers for tamoxifen-induced PLD. Copyright © 2017 John Wiley & Sons, Ltd.


Assuntos
Ácido Araquidônico/sangue , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Lipidoses/metabolismo , Fígado/metabolismo , Fosfatidilcolinas/sangue , Tamoxifeno/toxicidade , Animais , Ácido Araquidônico/metabolismo , Biomarcadores/sangue , Biomarcadores/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Relação Dose-Resposta a Droga , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipidoses/induzido quimicamente , Fígado/efeitos dos fármacos , Masculino , Fosfatidilcolinas/metabolismo , Ratos Sprague-Dawley
20.
Toxicol Sci ; 156(1): 39-53, 2017 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-28013220

RESUMO

Cationic amphiphilic drugs (CADs) are small molecules that can induce phospholipidosis (PLD), causing the intracellular accumulation of phospholipid in the lamellar bodies. Nanotechnology based drug delivery systems have been used widely, while it is unknown if drug-induced PLD (DIP) can be potentiated through drug retention by indigestible nanocarriers. Due to the high drug loading capacity of graphene, we investigated if PEGylated graphene oxide (PEG-GO) loaded with CAD could potentiate DIP. Tamoxifen induced the accumulation of NBD-PE, a fluorescence labeled phospholipid in human hepatoma HepG2 cells, while PEG-GO loaded with tamoxifen (PEG-GO/tamoxifen) further potentiated PLD. PEG-GO/tamoxifen induced more gene expression of PLD marker than tamoxifen alone. PEG-GO enhanced DIP was also observed for other CAD, indicating that nanocarrier potentiated DIP could be universal. More lamellar bodies were observed in PEG-GO/tamoxifen treated cells than tamoxifen alone by transmission electron microscopy. When compared with tamoxifen alone, PEG-GO/tamoxifen showed a delayed but potent PLD. In addition, the retarded PLD recovery by PEG-GO/tamoxifen indicated that the reversibility of DIP was interfered. Confocal microscopy revealed the increased number of lysosomes, greater expression of lysosomal associated membrane protein 2 (LAMP2) (a PLD marker), and an increase in the co-localization between lysosome/LAMP2 and NBD-PE by PEG-GO/tamoxifen rather than tamoxifen alone. Finally, we found that PEG-GO or/and tamoxifen-induced PLD seemed to have no correlation with autophagy. This research suggests pharmaceutical companies and regulatory agencies that if nanoparticles are used as the vectors for drug delivery, the adverse drug effects may be further potentiated probably through the long-term accumulation of nanocarriers.


Assuntos
Portadores de Fármacos/administração & dosagem , Grafite/efeitos adversos , Lipidoses/induzido quimicamente , Lisossomos/efeitos dos fármacos , Nanoestruturas/efeitos adversos , Fosfolipídeos/metabolismo , Polietilenoglicóis/efeitos adversos , Absorção Fisiológica , Animais , Portadores de Fármacos/metabolismo , Portadores de Fármacos/farmacologia , Antagonistas de Estrogênios/administração & dosagem , Antagonistas de Estrogênios/metabolismo , Antagonistas de Estrogênios/farmacologia , Corantes Fluorescentes/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Grafite/química , Grafite/metabolismo , Células Hep G2 , Humanos , Lipidoses/metabolismo , Lisossomos/metabolismo , Lisossomos/patologia , Lisossomos/ultraestrutura , Camundongos , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Nanoestruturas/química , Nanoestruturas/ultraestrutura , Fosfatidiletanolaminas/metabolismo , Polietilenoglicóis/química , Polietilenoglicóis/metabolismo , Células RAW 264.7 , Propriedades de Superfície , Tamoxifeno/administração & dosagem , Tamoxifeno/metabolismo , Tamoxifeno/farmacologia , Testes de Toxicidade Aguda
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...