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1.
Biophys Chem ; 258: 106317, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31918025

RESUMO

The antimicrobial activity of amphotericin B (AmB) depends on its interaction with ergosterol-containing cell membranes of fungus. Cholesterol is a sterol in mammalian cell membrane, and its structure is very similar to ergosterol, which caused to the toxic of amphotericin B to mammalian or human cell membranes. Even so, it is still the gold standard for the treatment of fungal infections. The mechanism of its toxicity to mammalian cell membrane has become a hot topic. The toxicity mechanism of amphotericin B on the cell membrane is also related to the phospholipids on the membrane. The effects of saturated and unsaturated fat chains on the interaction of amphotericin B with phospholipid monolayers containing cholesterol or ergosterol were studied at the molecular level using an air-water interface monolayer model. Both atomic force microscope and Brewster angle microscope were used to observe the surface morphology of the monolayer. The analysis of limiting molecular area suggested that the interaction between AmB and the two kinds of sterol is significantly different on the unsaturated lipid monolayer. According to the elastic modulus, the AmB molecules can increase the compressibility or viscoelasticity of the phospholipid/sterol monolayer. However, this impact of AmB on the DOPC/sterol monolayer containing ergosterol was stronger than that containing cholesterol at 25 ~ 50 mN/m. While this impact of AmB on the DPPC/sterol monolayer containing cholesterol was stronger than that containing ergosterol at 32 ~ 56 mN/m. The excess Gibbs free energy of the monolayer showed that, in the presence of saturated fat chain, amphotericin B could make the molecules of the DPPC/cholesterol monolayer and the DPPC/ergosterol monolayer arrange more closely and make intermolecular interaction stronger. There was no significant difference between DPPC/cholesterol monolayer and DPPC/ergosterol monolayer. However, in the presence of unsaturated chain, the effects of amphotericin B on the DOPC/cholesterol monolayer and the DOPC/ergosterol monolayer were significantly different. Amphotericin B made the molecular arrangement of DOPC/ergosterol monolayer more loosed, and the intermolecular force weakened at 5-35 mN/m. AFM images reflect that AmB can perforate the phospholipid-ergosterol monolayer, which was no significant correlation with saturation of the lipid monolayer. But the areas of dark areas shaped holes on the DPPC/ergosterol monolayer were larger than that on the DOPC/ergosterol monolayer. The adsorption of amphotericin B on lipid/sterol monolayer suggests that the orientation of amphotericin B may be different when it is inserted into the monolayer of phospholipid-sterol in the presence of saturated or unsaturated chains. The results are helpful to understand the complex mechanism of toxicity of amphotericin B to cell membrane.


Assuntos
Anfotericina B/química , Antibacterianos/química , Colesterol/química , Ergosterol/química , Fosfolipídeos/química , Ar , Humanos , Tamanho da Partícula , Pressão , Propriedades de Superfície , Água/química
2.
Med Chem ; 16(1): 128-139, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-30501601

RESUMO

BACKGROUND: It is assumed that the unfavorable selective toxicity of an antifungal drug Amphotericin B (AmB) can be improved upon chemical modification of the antibiotic molecule. OBJECTIVE: The aim of this study was verification of the hypothesis that introduction of bulky substituents at the amino sugar moiety of the antibiotic may result in diminishment of mammalian in vitro toxicity of thus prepared AmB derivatives. METHODS: Twenty-eight derivatives of AmB were obtained upon chemical modification of the amino group of mycosamine residue. This set comprised 10 N-succinimidyl-, 4 N-benzyl-, 5 Nthioureidyl- and 9 N-aminoacyl derivatives. Parameters characterizing biological in vitro activity of novel compounds were determined. RESULTS: All the novel compounds demonstrated lower than AmB antifungal in vitro activity but most of them exhibited negligible cytotoxicity against human erythrocytes and three mammalian cell lines. In consequence, the selective toxicity of majority of novel antifungals, reflected by the selective toxicity index (STI = EH50/IC50) was improved in comparison with that of AmB, especially in the case of 5 compounds. The novel AmB derivatives with the highest STI, induced substantial potassium efflux from Candida albicans cells at concentrations slightly lower than IC50s but did not trigger potassium release from human erythrocytes at concentrations lower than 100 µg/mL. CONCLUSION: Some of the novel AmB derivatives can be considered promising antifungal drug candidates.


Assuntos
Anfotericina B/farmacologia , Antifúngicos/farmacologia , Hexosaminas/farmacologia , Anfotericina B/síntese química , Anfotericina B/química , Antifúngicos/síntese química , Antifúngicos/química , Aspergillus/efeitos dos fármacos , Candida/efeitos dos fármacos , Cryptococcus/efeitos dos fármacos , Relação Dose-Resposta a Droga , Fusarium/efeitos dos fármacos , Hexosaminas/química , Testes de Sensibilidade Microbiana , Estrutura Molecular , Mucor/efeitos dos fármacos , Rhizopus/efeitos dos fármacos , Relação Estrutura-Atividade
3.
Int J Nanomedicine ; 14: 7593-7607, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31802863

RESUMO

Background: Amphotericin B (Amp) and Betulinic acid (BA) as antileishmanial agents have negligible water solubility and high toxicity. To solve these problems, for the first time, chitosan nanoparticles and Anionic Linear Globular Dendrimer (D) were synthesized for the treatment of Leishmania major (L. major). Method: Chitosan and dendrimer nanoparticles were synthesized, and Amp and BA were loaded into the nanoparticles. The particles were then characterized using various methods and their efficacy was evaluated in vitro and in vivo environments (parasite burden was confirmed using pathological studies and real-time PCR methods). Result: The results of docking showed that Amp and BA can be loaded into chitosan and dendrimer nanoparticles. The results of physically drug loading efficiency for AK (Amphotericin B-chitosan), BK (Betulinic acid-chitosan), AD (Amphotericin B-Dendrimer) and BD (Betulinic acid- Dendrimer) were 90, 93, 84 and 96 percent, respectively. The characterization results indicated that the drugs were loaded into nanoparticles physically. Moreover, the increased solubility rate for AD=478, BD=790, AK=80 and BK=300 folds. Furthermore, the results of the drug delivery system showed the slow controlled drug release pattern with cellular uptake of more than 90%. The treatment results showed a 100 percent decrease of toxicity for the all nanodrugs was observed in vivo and in vitro environments. Moreover, AK10 and BK20 mg/kg reduced parasite burden by 83 percent (P<0.001), while AD50 and BD40 mg/kg reduced it to a lesser extent compared to glucantime. Conclusion: All the synthesized nanodrugs were completely succeeded by 100% to recovery the L. major induced pathological effects in the infected footpad. Also, the results of present study were confirmed with real-time PCR and the results showed that AK and BK were succeeded in a large extent to the treatment of L. major infection (P<0.001), therefore AK and BK could be considered as proper alternatives of choices drugs.


Assuntos
Anfotericina B/farmacologia , Quitosana/química , Dendrímeros/química , Leishmania major/efeitos dos fármacos , Leishmania major/genética , Nanopartículas/química , Reação em Cadeia da Polimerase em Tempo Real/métodos , Triterpenos/química , Anfotericina B/química , Animais , Antiprotozoários/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Sistemas de Liberação de Medicamentos , Liberação Controlada de Fármacos , Simulação de Acoplamento Molecular , Nanopartículas/uso terapêutico , Nanopartículas/toxicidade , Parasitos/efeitos dos fármacos , Parasitos/genética , Solubilidade , Termodinâmica
4.
Int J Nanomedicine ; 14: 6073-6101, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31686803

RESUMO

Background: Amphotericin B (AmB) as a liposomal formulation of AmBisome is the first line of treatment for the disease, visceral leishmaniasis, caused by the parasite Leishmania donovani. However, nephrotoxicity is very common due to poor water solubility and aggregation of AmB. This study aimed to develop a water-soluble covalent conjugate of gold nanoparticle (GNP) with AmB for improved antileishmanial efficacy and reduced cytotoxicity. Methods: Citrate-reduced GNPs (~39 nm) were functionalized with lipoic acid (LA), and the product GNP-LA (GL ~46 nm) was covalently conjugated with AmB using carboxyl-to-amine coupling chemistry to produce GNP-LA-AmB (GL-AmB ~48 nm). The nanoparticles were characterized by dynamic light scattering, transmission electron microscopy (TEM), and spectroscopic (ultraviolet-visible and infrared) methods. Experiments on AmB uptake of macrophages, ergosterol depletion of drug-treated parasites, cytokine ELISA, fluorescence anisotropy, flow cytometry, and gene expression studies established efficacy of GL-AmB over standard AmB. Results: Infrared spectroscopy confirmed the presence of a covalent amide bond in the conjugate. TEM images showed uniform size with smooth surfaces of GL-AmB nanoparticles. Efficiency of AmB conjugation was ~78%. Incubation in serum for 72 h showed <7% AmB release, indicating high stability of conjugate GL-AmB. GL-AmB with AmB equivalents showed ~5-fold enhanced antileishmanial activity compared with AmB against parasite-infected macrophages ex vivo. Macrophages treated with GL-AmB showed increased immunostimulatory Th1 (IL-12 and interferon-γ) response compared with standard AmB. In parallel, AmB uptake was ~5.5 and ~3.7-fold higher for GL-AmB-treated (P<0.001) macrophages within 1 and 2 h of treatment, respectively. The ergosterol content in GL-AmB-treated parasites was ~2-fold reduced compared with AmB-treated parasites. Moreover, GL-AmB was significantly less cytotoxic and hemolytic than AmB (P<0.01). Conclusion: GNP-based delivery of AmB can be a better, cheaper, and safer alternative than available AmB formulations.


Assuntos
Anfotericina B/síntese química , Antiprotozoários/farmacologia , Ouro/química , Nanopartículas Metálicas/química , Anfotericina B/química , Anfotericina B/farmacologia , Animais , Antiprotozoários/química , Candida albicans/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular , Citocinas/metabolismo , Difusão Dinâmica da Luz , Ergosterol/metabolismo , Hemólise/efeitos dos fármacos , Humanos , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/crescimento & desenvolvimento , Estágios do Ciclo de Vida/efeitos dos fármacos , Peroxidação de Lipídeos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Nanopartículas Metálicas/ultraestrutura , Camundongos , Carbonilação Proteica/efeitos dos fármacos , Compostos de Sulfidrila/metabolismo , Ácido Tióctico/química , Resultado do Tratamento
5.
Molecules ; 24(18)2019 Sep 09.
Artigo em Inglês | MEDLINE | ID: mdl-31505853

RESUMO

Complement (C) activation can underlie the infusion reactions to liposomes and other nanoparticle-based medicines, a hypersensitivity syndrome that can be partially reproduced in animal models. However, the sensitivities and manifestations substantially differ in different species, and C activation may not be the only cause of pathophysiological changes. In order to map the species variation of C-dependent and -independent pseudoallergy (CARPA/CIPA), here we used known C activators and C activator liposomes to compare their acute hemodynamic, hematological, and biochemical effects in rats. These C activators were cobra venom factor (CVF), zymosan, AmBisome (at 2 doses), its amphotericin B-free vehicle (AmBisombo), and a PEGylated cholesterol-containing liposome (PEG-2000-chol), all having different powers to activate C in rat blood. The pathophysiological endpoints measured were blood pressure, leukocyte and platelet counts, and plasma thromboxane B2, while C activation was assessed by C3 consumption using the Pan-Specific C3 assay. The results showed strong linear correlation between C activation and systemic hypotension, pointing to a causal role of C activation in the hemodynamic changes. The observed thrombocytopenia and leukopenia followed by leukocytosis also correlated with C3 conversion in case of C activators, but not necessarily with C activation by liposomes. These findings are consistent with the double hit hypothesis of hypersensitivity reactions (HSRs), inasmuch as strong C activation can fully account for all symptoms of HSRs, but in case of no-, or weak C activators, the pathophysiological response, if any, is likely to involve other activation pathways.


Assuntos
Ativação do Complemento/efeitos dos fármacos , Síndrome de Hipersensibilidade a Medicamentos/tratamento farmacológico , Leucocitose/sangue , Lipossomos/farmacologia , Anfotericina B/química , Anfotericina B/farmacologia , Animais , Colesterol/química , Convertases de Complemento C3-C5/química , Convertases de Complemento C3-C5/farmacologia , Proteínas do Sistema Complemento/química , Proteínas do Sistema Complemento/metabolismo , Síndrome de Hipersensibilidade a Medicamentos/etiologia , Síndrome de Hipersensibilidade a Medicamentos/patologia , Venenos Elapídicos/química , Venenos Elapídicos/farmacologia , Humanos , Hipotensão/sangue , Hipotensão/induzido quimicamente , Leucocitose/induzido quimicamente , Leucopenia/sangue , Leucopenia/induzido quimicamente , Lipossomos/química , Nanopartículas/química , Polietilenoglicóis/química , Ratos , Trombocitopenia/sangue , Trombocitopenia/induzido quimicamente , Zimosan/química , Zimosan/farmacologia
6.
Curr Drug Deliv ; 16(7): 645-653, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31362675

RESUMO

BACKGROUND: Amphotericin B (AmB) is important for the treatment of systemic fungal infections. Nowadays, only intravenous administration (IV) of AmB has been available due to its low aqueous solubility. Two forms of AmB are available. The first is Fungizone®, a mixture of AmB and sodium deoxcycholate that produces severe nephrotoxicity. The second are lipid-based formulations that reduce nephrotoxicity, but they are costly and require higher dose than Fungizone®. Thus, a cheaper delivery system with reduced AmB toxicity is required. OBJECTIVE: To develop and characterize AmB loaded-nanostructured lipid carriers (AmB-loaded NLCs) for IV administration to reduce AmB toxicity. METHODS: AmB-loaded NLCs with different solid lipids were prepared by the high-pressure homogenization technique. Their physicochemical properties and the drug release profile were examined. The molecular structure of AmB, antifungal and hemolysis activities of developed AmB-loaded NLCs were also evaluated. RESULTS: AmB-loaded NLCs ~110 to ~140 nm in diameter were successfully produced with a zeta potential of ~-19 mV and entrapment efficiency of ~75%. In vitro release showed fast release characteristics. AmB-loaded NLCs could reduce the AmB molecular aggregation as evident from the absorbance ratio of the first to the fourth peak showing a partial aggregation of AmB. This result suggested that AmB-loaded NLCs could offer less nephrotoxicity compared to Fungizone®. In vitro antifungal activity of AmB-loaded NLCs showed a minimum inhibitory concentration of 0.25 µgmL-1. CONCLUSION: AmB-loaded NLCs present high potential carriers for effective IV treatment with prolonged circulation time and reduced toxicity.


Assuntos
Anfotericina B , Antifúngicos , Portadores de Fármacos , Nanoestruturas , Administração Intravenosa , Anfotericina B/administração & dosagem , Anfotericina B/química , Anfotericina B/toxicidade , Animais , Antifúngicos/administração & dosagem , Antifúngicos/química , Antifúngicos/toxicidade , Candida albicans/efeitos dos fármacos , Candida albicans/crescimento & desenvolvimento , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/química , Portadores de Fármacos/toxicidade , Liberação Controlada de Fármacos , Eritrócitos/efeitos dos fármacos , Hemólise/efeitos dos fármacos , Lipídeos/administração & dosagem , Lipídeos/química , Lipídeos/toxicidade , Nanoestruturas/administração & dosagem , Nanoestruturas/química , Nanoestruturas/toxicidade , Ovinos
7.
Int J Pharm ; 569: 118603, 2019 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-31401296

RESUMO

Conventional quantitation of drug content in the liposome formulation involves the breakdown of bulk liposomes, which ignores details on the distribution of the active pharmaceutical ingredient (API) and excipients in liposomes of different sizes. The objective of this study is to develop an analytical method which can separate the liposomes into different sizes and obtain information of the drug and excipient distribution in the different sized liposomes. We developed an asymmetric flow field-flow fractionation (AF4) method for size-based separation of AmBisome, an amphotericin B liposomal formulation, and a high-performance liquid chromatography ultraviolet-visible and charged aerosol detection (HPLC-UV-CAD) method for simultaneous quantitation of the API (Amphotericin B) and the lipid excipients [1,2-Distearoyl-sn-glycero-3-phosphoglycerol (DSPG), hydrogenated soy phosphatidylcholine (HSPC), and cholesterol]. The measured drug content in the bulk liposome formulation was consistent with the drug product labeling. Liposomes were separated using AF4 into eleven size fractions and the liposomes particles sizes of each fraction were measured with nanoparticle tracking analysis. The drug to total lipid ratios in fractionated liposomes increased from 0.1 to 0.45 when the liposome size increased from 75 nm to 124 nm, while the lipid composition remained constant throughout the fractioned size range (cholesterol:DSPG, 0.7 and HSPC:DSPG, 0.3). These study results suggest that, for liposomal formulations of Amphotericin B in liposomes, the drug to lipid ratio increases with the size of the liposomes. This new analytical method provided a more in-depth characterization of liposomes, i.e., determining drug and excipient distributions in different sizes of liposomes, in a more efficient manner with more specific size-based composition information.


Assuntos
Anfotericina B/química , Excipientes/química , Colesterol/química , Nanopartículas/química , Tamanho da Partícula , Fosfatidilcolinas/química , Fosfatidilgliceróis/química
8.
Curr Pharm Des ; 25(14): 1616-1622, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31298163

RESUMO

Leishmaniasis is one of the most neglected diseases in the world. Its most severe clinical form, called visceral, if left untreated, can be fatal. Conventional therapy is based on the use of pentavalent antimonials and includes amphotericin B (AmB) as a second-choice drug. The micellar formulation of AmB, although effective, is associated with acute and chronic toxicity. Commercially-available lipid formulations emerged to overcome such drawbacks, but their high cost limits their widespread use. Drug delivery systems such as nanoemulsions (NE) have proven ability to solubilize hydrophobic compounds, improve absorption and bioavailability, increase efficacy and reduce toxicity of encapsulated drugs. NE become even more attractive because they are inexpensive and easy to prepare. The aim of this work was to incorporate AmB in NE prepared by sonicating a mixture of surfactants, Kolliphor® HS15 (KHS15) and Brij® 52, and an oil, isopropyl myristate. NE exhibited neutral pH, conductivity values consistent with oil in water systems, spherical structures with negative Zeta potential value, monomodal size distribution and average diameter of drug-containing droplets ranging from 33 to 132 nm. AmB did not modify the thermal behavior of the system, likely due to its dispersion in the internal phase. Statistically similar antileishmanial activity of AmB-loaded NE to that of AmB micellar formulation suggests further exploring them in terms of toxicity and effectiveness against amastigotes, with the aim of offering an alternative to treat visceral leishmaniasis.


Assuntos
Anfotericina B/química , Antiprotozoários/química , Emulsões/química , Leishmania infantum/efeitos dos fármacos , Micelas , Nanopartículas/química
9.
mBio ; 10(3)2019 05 28.
Artigo em Inglês | MEDLINE | ID: mdl-31138748

RESUMO

Cryptococcus neoformans is an encapsulated yeast responsible for approximately a quarter of a million deaths worldwide annually despite therapy, and upwards of 11% of HIV/AIDS-related deaths, rivaling the impact of tuberculosis and malaria. However, the most effective antifungal agent, amphotericin B, requires intravenous delivery and has significant renal and hematopoietic toxicity, making it difficult to utilize, especially in resource-limited settings. The present studies describe a new nanoparticle crystal encapsulated formulation of amphotericin B known as encochleated amphotericin B (CAmB) that seeks to provide an oral formulation that is low in toxicity and cost. Using a 3-day delayed model of murine cryptococcal meningoencephalitis and a large inoculum of a highly virulent strain of serotype A C. neoformans, CAmB, in combination with flucytosine, was found to have efficacy equivalent to parental amphotericin B deoxycholate with flucytosine and superior to oral fluconazole without untoward toxicity. Transport of fluorescent CAmB particles to brain as well as significant brain levels of amphotericin drug was demonstrated in treated mice, and immunological profiles were similar to those of mice treated with conventional amphotericin B. Additional toxicity studies using a standardized rat model showed negligible toxicity after a 28-day treatment schedule. These studies thus offer the potential for an efficacious oral formulation of a known fungicidal drug against intrathecal cryptococcal disease.IMPORTANCE Cryptococcus neoformans is a significant global fungal pathogen that kills an estimated quarter of a million HIV-infected individuals yearly and has poor outcomes despite therapy. The most effective therapy, amphotericin B, is highly effective in killing the fungus but is available only in highly toxic, intravenous formulations that are unavailable in most of the developing world, where cryptococcal disease in most prevalent. For example, in Ethiopia, reliance on the orally available antifungal fluconazole results in high mortality, even when initiated as preemptive therapy at the time of HIV diagnosis. Thus, alternative agents could result in significant saving of lives. Toward this end, the present work describes the development of a new formulation of amphotericin B (CAmB) that encapsulates the drug as a crystal lipid nanoparticle that facilitates oral absorption and prevents toxicity. Successful oral absorption of the drug was demonstrated in a mouse model that, in combination with the antifungal flucytosine, provided efficacy equal to a parental preparation of amphotericin B plus flucytosine. These studies demonstrate the potential for CAmB in combination with flucytosine to provide an effective oral formulation of a well-known, potent fungicidal drug combination.


Assuntos
Anfotericina B/uso terapêutico , Antifúngicos/uso terapêutico , Criptococose/tratamento farmacológico , Meningoencefalite/tratamento farmacológico , Administração Oral , Anfotericina B/química , Animais , Antifúngicos/química , Cryptococcus neoformans/efeitos dos fármacos , Ácido Desoxicólico/uso terapêutico , Modelos Animais de Doenças , Combinação de Medicamentos , Composição de Medicamentos , Quimioterapia Combinada , Feminino , Flucitosina/uso terapêutico , Lipídeos/química , Masculino , Meningoencefalite/microbiologia , Camundongos , Nanopartículas/química , Ratos , Ratos Sprague-Dawley
10.
Int J Pharm ; 565: 447-457, 2019 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-31071418

RESUMO

Identifying the critical process parameters (CPPs) of a complex drug product manufacture and the associated impact on critical quality attributes (CQAs) is essential to the development and quality control of both new and generic drugs. AmBisome, a liposomal amphotericin B (AMB) macrolide antibiotic widely adopted as an important antifungal drug product, was used as a model complex drug product in the current study. This study investigated how multi-step production approaches and related manufacturing conditions may affect essential physico-chemical and toxicological properties of the final drug product. A key challenge in the manufacture and analysis of liposomal AMB was the drug substance's propensity to aggregate, with associated poor solubility in water and organic solvents. This study identified three key CPPs in a four step manufacturing process: (i) proper acidification during formation of the drug-lipid complexes (Step 1), (ii) liposome heat curing following liposomal particle sizing (Step 3), and (iii) flash-freezing at the initial stages of the lyophilization cycle (Step 4). Over-acidification led to rapid degradation of the drug, whereas under-acidification hampered full solubilization and formation of the soluble drug-lipid complexes. Extended heat treatment of the formed liposomes at 65 °C, just above the lipid phase transition temperature, brought dramatic changes in the aggregated state and/or packing of the drug in the liposomal bilayer, as followed by the complex changes in the UV/Vis spectra. Such thermal conditioning resulted in a five- to ten-fold reduction in the in-vitro toxicity of the drug product, bringing it close to the values for AmBisome used as control and measured by the RBC assay. Finally, flash-freezing conditions during lyophilization was critical to prevent aggregation and maintaining the 80-120 nm liposome size when reconstituted. Our research found that changes in the amphotericin's UV/Vis spectra were a sensitive CQA measure and provided a set of quantitative parameters for a facile non-destructive process monitoring in-situ, as well as for comparison of the quality of final formulations.


Assuntos
Anfotericina B/química , Antibacterianos/química , Antifúngicos/química , Anfotericina B/toxicidade , Animais , Antibacterianos/toxicidade , Antifúngicos/toxicidade , Composição de Medicamentos , Eritrócitos/efeitos dos fármacos , Congelamento , Temperatura Alta , Tamanho da Partícula , Ratos
11.
Biochemistry ; 58(17): 2282-2291, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30973009

RESUMO

The clinically important antibiotic amphotericin B (AmB) is a membrane-active natural product that targets membrane sterol. The antimicrobial activity of AmB is generally attributed to its membrane permeabilization, which occurs when a pore is formed across a lipid bilayer. In this study, the molecular orientation of AmB was investigated using solid-state nuclear magnetic resonance (NMR) to better understand the mechanism of antifungal activity. The methyl ester of AmB (AME) labeled with NMR isotopes, d3-AME, and its fluorinated and/or 13C-labeled derivatives were prepared. All of the AmB derivatives showed similar membrane-disrupting activities and ultraviolet spectra in phospholipid liposomes, suggesting that their molecular assemblies in membranes closely mimic those of AmB. Solid-state 2H NMR measurements of d3-AME in a hydrated membrane showed that the mobility of AME molecules depends on concentration and temperature. At a 1:5:45 AME:Erg:dimyristoylphosphatidylcholine ratio, AME became sufficiently mobilized to observe the motional averaging of quadrupole coupling. On the basis of the rotational averaging effect of 19F chemical shift anisotropy, 2H quadrupolar splitting, and 13C-19F dipolar coupling of 14ß-F-AMEs, we deduced that the molecular axis of AME is predominantly parallel to the normal of a lipid bilayer. This result supports the barrel-stave model as a molecular assembly of AmB in membranes.


Assuntos
Anfotericina B/análogos & derivados , Antifúngicos/química , Ergosterol/química , Bicamadas Lipídicas/química , Fosfolipídeos/química , Anfotericina B/química , Anfotericina B/metabolismo , Anfotericina B/farmacologia , Antifúngicos/metabolismo , Antifúngicos/farmacologia , Membrana Celular/química , Membrana Celular/efeitos dos fármacos , Ergosterol/metabolismo , Fungos/citologia , Fungos/efeitos dos fármacos , Fungos/metabolismo , Marcação por Isótopo , Bicamadas Lipídicas/metabolismo , Lipossomos/química , Lipossomos/metabolismo , Espectroscopia de Ressonância Magnética/métodos , Modelos Químicos , Modelos Moleculares , Estrutura Molecular , Fosfolipídeos/metabolismo , Esteróis/química , Esteróis/metabolismo
12.
Am J Health Syst Pharm ; 76(11): 810-813, 2019 May 17.
Artigo em Inglês | MEDLINE | ID: mdl-30994894

RESUMO

PURPOSE: We describe the use of liposomal amphotericin B and amphotericin B deoxycholate in a critically ill patient with pulmonary blastomycosis receiving both venovenous extracorporeal membrane oxygenation (ECMO) and continuous renal replacement therapy (CRRT). SUMMARY: A 50-year-old African American man presented for dyspnea and cough and was noted to have blastomycosis on bronchoscopy. He developed respiratory failure and acute kidney injury, requiring mechanical ventilation, ECMO, and CRRT. After 4 days of liposomal amphotericin, the transmembrane pressure gradient on the membrane oxygenator increased dramatically without visualization of a clot, requiring a circuit exchange. A trough amphotericin B level taken the day before the exchange was undetectable for amphotericin B. After the circuit exchange, the patient was switched to amphotericin B deoxycholate. A subsequent trough level was 3.8 µg/mL. The patient improved and was able to be decannulated. However, he did require tracheostomy and long-term hemodialysis. CONCLUSION: In our case we believe that liposomal amphotericin B was significantly removed by ECMO and was responsible for the failure of the ECMO circuit. We would suggest amphotericin B deoxycholate be used in such patients preferentially and that serum levels of the drug be assessed when possible.


Assuntos
Anfotericina B/farmacologia , Blastomicose/terapia , Ácido Desoxicólico/farmacologia , Oxigenação por Membrana Extracorpórea/instrumentação , Lesão Renal Aguda/etiologia , Lesão Renal Aguda/terapia , Anfotericina B/química , Anfotericina B/uso terapêutico , Área Sob a Curva , Blastomicose/sangue , Blastomicose/complicações , Terapia Combinada/métodos , Estado Terminal/terapia , Ácido Desoxicólico/uso terapêutico , Combinação de Medicamentos , Substituição de Medicamentos , Falha de Equipamento , Humanos , Masculino , Pessoa de Meia-Idade , Oxigenadores de Membrana/efeitos adversos , Insuficiência Respiratória/etiologia , Insuficiência Respiratória/terapia , Resultado do Tratamento
13.
Bull Exp Biol Med ; 166(6): 735-738, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31020586

RESUMO

Comparative analysis of the effects of chemically transformed polyene antibiotics pimaricin, nystatin, lucensomycin, amphotericin B, and levorin on biological objects in vivo and in vitro revealed the greatest biological activity of original amphotericin B and levorin with its derivatives. The study also examined the effects of alkyl derivatives of amphotericin B and levorin modified in certain parts of the lactone ring on the lipid and biological membranes. It is established that methylated levorin possesses larger biological activity than the original antibiotic. Examination of the effects of alkyl derivatives of levorin and amphotericin B on cell cultures C6 (rat glioma) and HeLa (human cervical carcinoma) in vitro revealed the antitumor action of methylated levorin and original amphotericin B.


Assuntos
Anfotericina B/farmacologia , Antibacterianos/farmacologia , Antineoplásicos/farmacologia , Candicidina/farmacologia , Alquilação , Anfotericina B/química , Animais , Antibacterianos/química , Antineoplásicos/química , Candicidina/química , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células HeLa , Humanos , Lucensomycin/química , Lucensomycin/farmacologia , Natamicina/química , Natamicina/farmacologia , Neuroglia , Nistatina/química , Nistatina/farmacologia , Ratos , Relação Estrutura-Atividade
14.
AAPS PharmSciTech ; 20(3): 136, 2019 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-30838459

RESUMO

Surface-modified nanostructured lipid carriers (NLC) represent a promising mode of drug delivery used to enhance retention of drugs at absorption site. Formulated chitosan-coated amphotericin-B-loaded NLC (ChiAmp NLC) had a size of 394.4 ± 6.4 nm, encapsulation and loading efficiencies of 86.0 ± 3% and 11.0 ± 0.1% respectively. Amphotericin-B release from NLCs was biphasic with no changes in physical properties upon exposure to simulated gastrointestinal conditions. Antifungal properties of Amphotericin-B and ChiAmpB NLC were comparable but ChiAmpB NLC was twice less toxic to red blood cells and ten times safer on HT-29 cell lines. In vitro mucoadhesion data were observed ex vivo, where ChiAmpB NLC resulted in higher retention within the small intestine compared to the uncoated formulation. The data strongly offers the possibility of orally administering a non-toxic, yet effective Amphotericin-B nanoformulation for the treatment of systemic fungal infections.


Assuntos
Anfotericina B/administração & dosagem , Anfotericina B/farmacologia , Antifúngicos/administração & dosagem , Antifúngicos/farmacologia , Quitosana/administração & dosagem , Quitosana/farmacologia , Portadores de Fármacos/química , Lipídeos/química , Nanoestruturas/química , Administração Oral , Anfotericina B/química , Animais , Antifúngicos/química , Candida albicans/efeitos dos fármacos , Quitosana/química , Eritrócitos/efeitos dos fármacos , Células HT29 , Humanos , Mucosa Intestinal/efeitos dos fármacos , Jejuno/efeitos dos fármacos , Masculino , Testes de Sensibilidade Microbiana , Ratos Sprague-Dawley
15.
Bioconjug Chem ; 30(3): 966-973, 2019 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-30793876

RESUMO

Due to the advanced fluorescence property of N-carbon quantum dots (N-CQDs), a new method to detect pathogenic fungi by newly synthesized cornstalk N-CQDs modified with water-soluble amphotericin B (N-CQDs@AmpB) was developed. Specifically, N-CQDs with blue fluorescence were initially synthesized according to a previous report and modified with amphotericin B on their surfaces. Subsequently, the as-prepared N-CQDs@AmpB was used to detect Candida albicans, exhibiting a linear range of 2.60 × 105 to 1.99 × 108 cfu/mL and a detection limit of 1124 cfu/mL. Compared with other common methods, the method largely shortened the detection time and enabled the process to be performed with minimal interference from complex samples such as beef sausage. The high cost of water-soluble amphotericin B may hamper the large-scale application of the new detection method using N-CQDs@AmpB. Thus, alcohol-soluble amphotericin B was used in subsequent experiments, confirming its potential to broaden avenues for the detection of fungi.


Assuntos
Anfotericina B/química , Antifúngicos/química , Candida albicans/isolamento & purificação , Carbono/química , Fluorometria/métodos , Pontos Quânticos/química , Animais , Bovinos , Limite de Detecção , Produtos da Carne/microbiologia , Microscopia Eletrônica de Transmissão , Espectroscopia de Infravermelho com Transformada de Fourier
16.
AAPS PharmSciTech ; 20(3): 122, 2019 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-30805739

RESUMO

Cutaneous leishmaniasis is a neglected parasitic disease. Treatment is preferably performed with pentavalent antimony associated or not with amphotericin B (AmB). This study aimed to develop an emulgel with different chemical enhancers of cutaneous release. Initially, AmB emulsions were obtained with the chemical promoters, oleic acid and geraniol and without promoter, then for the evaluation of the formulations, a preliminary stability study was carried out where the formulations were submitted to centrifugation, before and after the freeze-thaw cycle and analyzed appearance, color, pH, spreadability, viscosity, conductivity, droplet size, assay, in vitro release study, in vitro antileishmania activity in Leishmania major promastigotes, and macrophage toxicity in the MTT test. The emulsions were yellowish, with no signs of instability after the centrifugation test. The pH range corresponded to that of the skin, which is 4.6 to 5.8, before and after the freeze-thaw cycle, the formulations had good spreadability and did not present significant viscosity differences before and after the freeze-thaw cycle, presenting a non-Newtonian characteristic. AmB content was within the kinetic model of zero order release, the formulation of 3% AmB and 5% oleic acid (formulation 1) was chosen to proceed with the antileishmania activity test and showed potential activity against the in vitro parasite with significant reduction of cytotoxicity on murine macrophages, indicating that the formulation is promising for the treatment of cutaneous leishmaniasis.


Assuntos
Anfotericina B/administração & dosagem , Antiprotozoários/administração & dosagem , Leishmaniose Cutânea/tratamento farmacológico , Anfotericina B/química , Anfotericina B/uso terapêutico , Animais , Antiprotozoários/química , Antiprotozoários/uso terapêutico , Composição de Medicamentos , Liberação Controlada de Fármacos , Estabilidade de Medicamentos , Emulsões , Concentração de Íons de Hidrogênio , Camundongos , Viscosidade
17.
Drug Dev Ind Pharm ; 45(4): 560-567, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30632399

RESUMO

Amphotericin B (AmB) is one of the most effective systemic antifungal agents, but its use is circumscribed by the dose-limiting toxicity of the conventional micellar dispersion formulation, Fungizone®. Significantly lesser toxicity is obtained when AmB incorporated into the aqueous dispersion of lipid nanoparticles. The aim of this study was to develop and characterize AmB loaded solid lipid nanoparticles (SLNs) and nanostructured lipid carriers (NLCs). NLC differed from SLN by the presence of liquid lipid, glyceryl tri(caprylate/caprate) in the lipid matrix. Various surfactants i.e. tween 20, cremophor RH40, poloxamer 407 (P407) and Myrj 52 were used to stabilize SLN and NLC. The effect of phospholipid incorporated in those lipid dispersions was also determined. Among surfactants tested, only P407 could stabilize AmB lipid dispersions. There was no chemical reaction occurred between AmB and other components that confirmed by Fourier transform infrared spectroscopy (FT-IR) spectra. The differential scanning calorimetry (DSC), hot-stage microscopy (HSM), powder X-ray diffractometry (PXRD) data showed that AmB was molecularly dispersed or in amorphous form in the lipid matrix. The proton nuclear magnetic resonance (1H-NMR) results showed that in the presence of phospholipid oil clusters within the lipid matrix are formed. These results indicate that SLN and NLC stabilized by P407 and/or phospholipid as the colloidal carrier for AmB were successfully developed.


Assuntos
Anfotericina B/administração & dosagem , Antifúngicos/administração & dosagem , Portadores de Fármacos/química , Composição de Medicamentos/métodos , Anfotericina B/química , Antifúngicos/química , Química Farmacêutica/métodos , Estabilidade de Medicamentos , Excipientes/química , Lipídeos/química , Nanopartículas/química , Tamanho da Partícula , Tensoativos/química
18.
Eur J Pharm Biopharm ; 134: 107-116, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30481559

RESUMO

AmBisome® is a liposomal formulation of amphotericin B (Amp B), a complex parenteral antifungal product with no US FDA approved generic version available to date. For generic Amp B liposomal product development, examination of the drug release profile is important for product quality control and analytical comparability evaluation with the reference listed drug. Yet, there is no standardized in vitro drug release (IVR) assay currently available for Amp B liposomes. In this study, we describe the development of a USP-4 apparatus-based IVR assay capable of discriminating liposomal Amp B formulations based on the drug release profile. The goal of the IVR assay development was to identify release media compositions and assay temperatures capable of facilitating 70-100% of drug release from AmBisome® in 24 h without Amp B precipitation or disruption of liposome structure. We found that an addition of 5% w/v of γ-cyclodextrin to the release media of 5% sucrose, 10 mM HEPES, and 0.01% NaN3 (pH = 7.4) prevented Amp B precipitation and facilitated drug release. Increased IVR assay temperature led to increased drug release rate, and 55 °C was selected as the highest temperature that induced drug release close to our target without causing product precipitation. The developed IVR assay was used to discriminate between drug release rates from AmBisome® and micellar Amp B products like Fungizone® and Fungcosome. The IVR assay was also capable of discriminating between Amp B liposomes with the same composition as AmBisome® but prepared by either extrusion or homogenization processes, both of which resulted in measurable liposomal particle size heterogeneity and Amp B concentration differences. Finally, the USP-4 IVR assay was used to compare Amp B release profiles between AmBisome® and two generic products approved in India, Amphonex® (Bharat Serums and Vaccines Ltd.) (f2 = 66.3) and Phosome® (Cipla Ltd.) (f2 = 55.4). Taken together, the developed USP-4 IVR assay can be a useful tool for drug release profile characterization in generic liposomal Amp B formulation development.


Assuntos
Anfotericina B/química , Antifúngicos/química , Química Farmacêutica/instrumentação , Desenvolvimento de Medicamentos/instrumentação , Liberação Controlada de Fármacos , Química Farmacêutica/métodos , Desenvolvimento de Medicamentos/métodos , Tamanho da Partícula
19.
Drug Deliv Transl Res ; 9(1): 76-84, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30484256

RESUMO

Cutaneous leishmaniasis (CL) is an infectious, parasitic disease caused by the protozoan Leishmania. Amphotericin B (AMB) is a macrolide polyene antibiotic presenting potent antifungal and antileishmanial activity, but due to poor water solubility at physiological pH, side effects, and toxicity, its therapeutic efficiency is limited. In the present study, poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) loaded with AMB were generated to reduce drug toxicity and facilitate localized delivery over a prolonged time. AMB NPs were characterized for particle size, zeta potential, polydispersity index, and degree of aggregation. In vitro assessments demonstrated its sustained activity against Leishmania major promastigotes and parasite-infected macrophages. A single intralesional administration to infected BALB/c mice revealed that AMB NPs were more effective than AMB deoxycholate in terms of reducing lesion area. Taken together, these findings suggest that AMB NPs improve AMB delivery and can be used for local treatment of CL.


Assuntos
Anfotericina B/administração & dosagem , Antiprotozoários/administração & dosagem , Leishmania major/efeitos dos fármacos , Leishmaniose Cutânea/tratamento farmacológico , Poliésteres/química , Ácido Poliglicólico/química , Administração Tópica , Anfotericina B/química , Anfotericina B/farmacologia , Animais , Antiprotozoários/química , Antiprotozoários/farmacologia , Modelos Animais de Doenças , Portadores de Fármacos/química , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Nanopartículas , Tamanho da Partícula , Células THP-1
20.
J Microbiol Biotechnol ; 29(1): 171-177, 2019 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-30415525

RESUMO

Parasitic infections have remained a significant burden on human and animal health. In part, this is due to lack of clinically-approved, novel antimicrobials and a lack of interest by the pharmaceutical industry. An alternative approach is to modify existing clinically-approved drugs for efficient delivery formulations to ensure minimum inhibitory concentration is achieved at the target site. Nanotechnology offers the potential to enhance the therapeutic efficacy of drugs through modification of nanoparticles with ligands. Amphotericin B, nystatin, and fluconazole are clinically available drugs in the treatment of amoebal and fungal infections. These drugs were conjugated with gold nanoparticles. To characterize these gold-conjugated drug, atomic force microscopy, ultraviolet-visible spectrophotometry and Fourier transform infrared spectroscopy were performed. These drugs and their gold nanoconjugates were examined for antimicrobial activity against the protist pathogen, Acanthamoeba castellanii of the T4 genotype. Moreover, host cell cytotoxicity assays were accomplished. Cytotoxicity of these drugs and drug-conjugated gold nanoparticles was also determined by lactate dehydrogenase assay. Gold nanoparticles conjugation resulted in enhanced bioactivity of all three drugs with amphotericin B producing the most significant effects against Acanthamoeba castellanii (p < 0.05). In contrast, bare gold nanoparticles did not exhibit antimicrobial potency. Furthermore, amoebae treated with drugs-conjugated gold nanoparticles showed reduced cytotoxicity against HeLa cells. In this report, we demonstrated the use of nanotechnology to modify existing clinically-approved drugs and enhance their efficacy against pathogenic amoebae. Given the lack of development of novel drugs, this is a viable approach in the treatment of neglected diseases.


Assuntos
Acanthamoeba castellanii/efeitos dos fármacos , Amebicidas/química , Amebicidas/farmacologia , Ouro/química , Nanopartículas Metálicas/química , Acanthamoeba castellanii/crescimento & desenvolvimento , Anfotericina B/química , Anfotericina B/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Fluconazol/química , Fluconazol/farmacologia , Células HeLa , Humanos , Nanomedicina , Nistatina/química , Nistatina/farmacologia
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