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1.
Sci Rep ; 10(1): 12243, 2020 07 22.
Artigo em Inglês | MEDLINE | ID: mdl-32699361

RESUMO

The development of an effective oral therapeutics is an immediate need for the control and elimination of visceral leishmaniasis (VL). We exemplify the preparation and optimization of 2-hydroxypropyl-ß-cyclodextrin (HPCD) modified solid lipid nanoparticles (SLNs) based oral combinational cargo system of Amphotericin B (AmB) and Paromomycin (PM) against murine VL. The emulsion solvent evaporation method was employed to prepare HPCD modified dual drug-loaded solid lipid nanoparticles (m-DDSLNs). The optimized formulations have a mean particle size of 141 ± 3.2 nm, a polydispersity index of 0.248 ± 0.11 and entrapment efficiency for AmB and PM was found to be 96% and 90% respectively. The morphology of m-DDSLNs was confirmed by scanning electron microscopy and transmission electron microscopy. The developed formulations revealed a sustained drug release profile upto 57% (AmB) and 21.5% (PM) within 72 h and were stable at both 4 °C and 25 °C during short term stability studies performed for 2 months. Confocal laser scanning microscopy confirmed complete cellular internalization of SLNs within 24 h of incubation. In vitro cytotoxicity study against J774A.1 macrophage cells confirmed the safety and biocompatibility of the developed formulations. Further, m-DDSLNs did not induce any hepatic/renal toxicities in Swiss albino mice. The in vitro simulated study was performed to check the stability in simulated gastric fluids and simulated intestinal fluids and the release was found almost negligible. The in vitro anti-leishmanial activity of m-DDSLNs (1 µg/ml) has shown a maximum percentage of inhibition (96.22%) on intra-cellular amastigote growth of L. donovani. m-DDSLNs (20 mg/kg × 5 days, p.o.) has significantly (P < 0.01) reduced the liver parasite burden as compared to miltefosine (3 mg/kg × 5 days, p.o.) in L. donovani-infected BALB/c mice. This work suggests that the superiority of as-prepared m-DDSLNs as a promising approach towards the oral delivery of anti-leishmanial drugs.


Assuntos
Anfotericina B/química , Anfotericina B/farmacologia , Leishmania donovani/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Nanopartículas/química , Paromomicina/química , Paromomicina/farmacologia , Animais , Antiprotozoários/química , Antiprotozoários/farmacologia , Linhagem Celular , Emulsões/química , Lipídeos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica de Transmissão/métodos , Tamanho da Partícula , Fosforilcolina/análogos & derivados , Fosforilcolina/química , Fosforilcolina/farmacologia
2.
Sci Rep ; 10(1): 6420, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32286337

RESUMO

The opportunistic pathogen, Acanthamoeba castellanii is the causative agent for the sight threatening infection Acanthamoeba keratitis (AK). It is commonly associated with contact lens wearers, and prevalence is increasing at an alarming rate due to an inadequate preventive strategy to protect the lens from this protist. This problem is compounded by the lack of an effective acanthamoebocide, particularly with cysticidal activity in the contact lens solutions. We have used cytotoxicity assays and a variety of biophysical approaches to show that two molecules with tails made of alkyl carbon, alkylphosphocholines (APCs) and quaternary ammonium compounds (QACs) had significant chain-length dependent efficacy against A. castellanii trophozoites, the latter producing death via permeabilization, and DNA complexing. QACs were more effective than APCs and had activity against cysts. Conversely, the QAC with 12 alkyl carbon chain, was non toxic, its presence increased A. castellanii trophozoites biomass and delayed encystation by 96 h. Interestingly, it was unable to induce excystation and increased trophozoite sensitivity to APC16. These results present a mono- and multi-inhibitor management strategy effective against trophozoites and cysts that may be useful for formulating into contact lense cleaning solutions and reducing AK incidence.


Assuntos
Acanthamoeba castellanii/efeitos dos fármacos , Antiprotozoários/química , Antiprotozoários/farmacologia , Carbono/química , Acanthamoeba castellanii/citologia , Morte Celular/efeitos dos fármacos , Linhagem Celular , Citoplasma/metabolismo , DNA/metabolismo , Concentração Inibidora 50 , Fosforilcolina/química , Fosforilcolina/farmacologia , Compostos de Amônio Quaternário/química , Compostos de Amônio Quaternário/farmacologia
3.
Parasit Vectors ; 13(1): 96, 2020 Feb 22.
Artigo em Inglês | MEDLINE | ID: mdl-32087758

RESUMO

BACKGROUND: Since the introduction of miltefosine (MIL) as first-line therapy in the kala-azar elimination programme in the Indian subcontinent, treatment failure rates have been increasing. Since parasite infectivity and virulence may become altered upon treatment relapse, this laboratory study assessed the phenotypic effects of repeated in vitro and in vivo MIL exposure. METHODS: Syngeneic Leishmania donovani lines either or not exposed to MIL were compared for drug susceptibility, rate of promastigote multiplication and metacyclogenesis, macrophage infectivity and behaviour in the sand fly vector, Lutzomyia longipalpis. RESULTS: Promastigotes of both in vitro and in vivo MIL-selected strains displayed a slightly reduced drug susceptibility that was associated with a reduced MIL-accumulation linked to a lower copy number (disomic state) of chromosome 13 harboring the miltefosine transporter (LdMT) gene. In vitro selected promastigotes showed a lower rate of metacyclogenesis whereas the in vivo derived promastigotes displayed a moderately increased growth rate. Repeated MIL exposure did neither influence the parasite load nor metacyclogenesis in the sand fly vector. CONCLUSIONS: Recurrent in vitro and in vivo MIL exposure evokes a number of very subtle phenotypic and genotypic changes which could make promastigotes less susceptible to MIL without attaining full resistance. These changes did not significantly impact on infection in the sand fly vector.


Assuntos
Antiprotozoários/farmacologia , Insetos Vetores/parasitologia , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/fisiologia , Fosforilcolina/análogos & derivados , Psychodidae/parasitologia , Aclimatação , Animais , Resistência a Medicamentos , Humanos , Leishmania donovani/patogenicidade , Leishmaniose Visceral/parasitologia , Leishmaniose Visceral/transmissão , Testes de Sensibilidade Parasitária , Fenótipo , Fosforilcolina/farmacologia , Virulência
4.
Biochem Biophys Res Commun ; 524(4): 1018-1024, 2020 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-32063363

RESUMO

Pten deletion in the hematopoietic stem cells (HSC) causes a myeloproliferative disorder, which may subsequently develop into a T-cell acute lymphoblastic leukemia (T-ALL). ß-catenin expression was dramatically increased in the c-KitmidCD3+Lin- leukemia stem cells (LSC) and was critical for T-ALL development. Therefore, the inactivation of ß-catenin in LSC may have a potential to eliminate the LSC. In this study, we investigated the mechanism of enhancement of the ß-catenin expression and subsequently used a drug to inactivate ß-catenin expression in T-ALL. Western blot (WB) analysis revealed an increased level of ß-catenin in the leukemic cells, but not in the pre-leukemic cells. Furthermore, the WB analysis of the thymic cells from different stages of leukemia development showed that increased expression of ß-catenin was not via the pS9-GSK3ß signaling, but was dependent on the pT308-Akt activation. Miltefosine (Hexadecylphosphocholine) is the first oral anti-Leishmania drug, which is a phospholipid agent and has been shown to inhibit the PI3K/Akt activity. Treatment of the PtenΔ/Δ leukemic mice with Miltefosine for different durations demonstrated that the pT308-Akt and the ß-catenin expressions were inhibited in the leukemia blast cells. Miltefosine treatment also suppressed the TGFß1/Smad3 signaling pathway. Analysis of TGFß1 in the sorted subpopulations of the blast cells showed that TGFß1 was secreted by the CD3+CD4- subpopulation and may exert effects on the subpopulations of both CD3+CD4+ and CD3+CD4- leukemia blast cells. When a TGFßR1 inhibitor, SB431542 was injected into the PtenΔ/Δ leukemic mice, the Smad3 and ß-catenin expressions were down-regulated. On the basis of the results, we conclude that Miltefosine can suppress leukemia by degrading ß-catenin through repression of the pT308-Akt and TGFß1/Smad3 signaling pathways. This study demonstrates a possibility to inhibit Pten loss-associated leukemia genesis via targeting Akt and Smad3.


Assuntos
Antineoplásicos/uso terapêutico , Fosforilcolina/análogos & derivados , Leucemia-Linfoma Linfoblástico de Células T Precursoras/tratamento farmacológico , Proteólise/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , beta Catenina/metabolismo , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Deleção de Genes , Humanos , Camundongos , PTEN Fosfo-Hidrolase/genética , Fosforilcolina/farmacologia , Fosforilcolina/uso terapêutico , Leucemia-Linfoma Linfoblástico de Células T Precursoras/genética , Leucemia-Linfoma Linfoblástico de Células T Precursoras/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/metabolismo
5.
Artigo em Inglês | MEDLINE | ID: mdl-32058031

RESUMO

Hepatocellular carcinoma (HCC) is an aggressive and widespread cancer. Patients with liver cirrhosis of different aetiologies are at a risk to develop HCC. It is important to know that in approximately 20% of cases primary liver tumors arise in a non-cirrhotic liver. Lipid metabolism is variable in patients with chronic liver diseases, and lipid metabolites involved therein do play a role in the development of HCC. Of note, lipid composition of carcinogenic tissues differs from non-affected liver tissues. High cholesterol and low ceramide levels in the tumors protect the cells from oxidative stress and apoptosis, and do also promote cell proliferation. So far, detailed characterization of the mechanisms by which lipids enable the development of HCC has received little attention. Evaluation of the complex roles of lipids in HCC is needed to better understand the pathophysiology of HCC, the later being of paramount importance for the development of urgently needed therapeutic interventions. Disturbed hepatic lipid homeostasis has systemic consequences and lipid species may emerge as promising biomarkers for early diagnosis of HCC. The challenge is to distinguish lipids specifically related to HCC from changes simply related to the underlying liver disease. This review article discusses aberrant lipid metabolism in patients with HCC.


Assuntos
Antineoplásicos/uso terapêutico , Carcinoma Hepatocelular/metabolismo , Metabolismo dos Lipídeos/fisiologia , Lipídeos/sangue , Neoplasias Hepáticas/metabolismo , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Biomarcadores Tumorais/sangue , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/sangue , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Diagnóstico Diferencial , Modelos Animais de Doenças , Progressão da Doença , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipídeos/farmacologia , Lipídeos/uso terapêutico , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática/sangue , Cirrose Hepática/diagnóstico , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Neoplasias Hepáticas/sangue , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/tratamento farmacológico , Estresse Oxidativo/efeitos dos fármacos , Éteres Fosfolipídicos/farmacologia , Éteres Fosfolipídicos/uso terapêutico , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Fosforilcolina/uso terapêutico , Plasmalogênios/sangue , Plasmalogênios/metabolismo , Plasmalogênios/farmacologia , Plasmalogênios/uso terapêutico , Índice de Gravidade de Doença
6.
PLoS One ; 15(1): e0223463, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31914134

RESUMO

Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) DNA cytosine deaminase 3B (A3B) is a DNA editing enzyme which induces genomic DNA mutations in multiple myeloma and in various other cancers. APOBEC family proteins are highly homologous so it is especially difficult to investigate the biology of specifically A3B in cancer cells. To easily and comprehensively investigate A3B function in myeloma cells, we used CRISPR/Cas9 to generate A3B reporter cells that contain 3×FLAG tag and IRES-EGFP sequences integrated at the end of the A3B gene. These reporter cells stably express 3xFLAG tagged A3B and the reporter EGFP and this expression is enhanced by known stimuli, such as PMA. Conversely, shRNA knockdown of A3B decreased EGFP fluorescence and 3xFLAG tagged A3B protein levels. We screened a series of anticancer treatments using these cell lines and identified that most conventional therapies, such as antimetabolites or radiation, exacerbated endogenous A3B expression, but recent molecular targeted therapeutics, including bortezomib, lenalidomide and elotuzumab, did not. Furthermore, chemical inhibition of ATM, ATR and DNA-PK suppressed EGFP expression upon treatment with antimetabolites. These results suggest that DNA damage triggers A3B expression through ATM, ATR and DNA-PK signaling.


Assuntos
Citidina Desaminase/genética , Dano ao DNA/genética , Antígenos de Histocompatibilidade Menor/genética , Mieloma Múltiplo/genética , Anticorpos Monoclonais Humanizados/farmacologia , Bortezomib/farmacologia , Sistemas CRISPR-Cas/genética , Linhagem Celular Tumoral , Núcleo Celular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Humanos , Lenalidomida/farmacologia , Mieloma Múltiplo/tratamento farmacológico , Mieloma Múltiplo/patologia , Mieloma Múltiplo/radioterapia , Mutação/genética , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Ácidos Polimetacrílicos/farmacologia , RNA Interferente Pequeno/genética , Radiação , Transdução de Sinais/efeitos dos fármacos
7.
Chem Biol Interact ; 315: 108850, 2020 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-31634447

RESUMO

1,2,3-triazolium salts are poorly understood regarding their antileishmanial activity. Hence, as an effort to identify novel chemical scaffolds as antileishmanial agents, a series of 1,2,3-triazolium salts (TS) and corresponding 1,2,3-triazole (T) precursors including new epoxide derivatives were synthesized and assayed against Leishmania amazonensis promastigote and intracellular amastigote forms. Among them, the compound TS-6 exhibited promising activity on promastigotes (IC50 = 3.61 µM) and intracellular amastigotes (IC50 = 7.61 µM) of L. amazonensis, superior to miltefosine (IC50 > 10.0 µM), used as reference drug. In addition, TS-6 showed negligible cytotoxicity on murine peritoneal macrophages with a SI of about 10. Studies on the mode of action of TS-6 indicate mitochondrial dysfunction through an increase in 'total' and mitochondrial-ROS as well as depolarization of mitochondrial membrane potential of L. amazonensis promastigotes. In silico physicochemical studies indicate that the TS-6 could potentially be used as an oral drug.


Assuntos
Antiprotozoários/farmacologia , Leishmania mexicana/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Triazóis/farmacologia , Animais , Leishmania mexicana/metabolismo , Leishmaniose Cutânea/tratamento farmacológico , Leishmaniose Cutânea/parasitologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/metabolismo , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia
8.
Ann Biomed Eng ; 48(3): 913-926, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30815762

RESUMO

Vascular tissue engineering of the middle layer of natural arteries requires contractile smooth muscle cells (SMC) which can be differentiated from adipose-derived mesenchymal stem cells (ASC) by treatment with transforming growth factor-ß, sphingosylphosphorylcholine and bone morphogenetic protein-4 (TSB). Since mechanical stimulation may support or replace TSB-driven differentiation, we investigated its effect plus TSB-treatment on SMC orientation and contractile protein expression. Tubular fibrin scaffolds with incorporated ASC or pre-differentiated SMC were exposed to pulsatile perfusion for 10 days with or without TSB. Statically incubated scaffolds served as controls. Pulsatile incubation resulted in collagen-I expression and orientation of either cell type circumferentially around the lumen as shown by alpha smooth muscle actin (αSMA), calponin and smoothelin staining as early, intermediate and late marker proteins. Semi-quantitative Westernblot analyses revealed strongly increased αSMA and calponin expression by either pulsatile (12.48-fold; p < 0.01 and 38.15-fold; p = 0.07) or static incubation plus TSB pre-treatment (8.91-fold; p < 0.05 and 37.69-fold; p < 0.05). In contrast, contractility and smoothelin expression required both mechanical and TSB stimulation since it was 2.57-fold increased (p < 0.05) only by combining pulsatile perfusion and TSB. Moreover, pre-differentiation of ASC prior to pulsatile perfusion was not necessary since it could not further increase the expression level of any marker.


Assuntos
Células-Tronco Mesenquimais/citologia , Miócitos de Músculo Liso/citologia , Túnica Média , Adipogenia , Adulto , Idoso , Reatores Biológicos , Proteína Morfogenética Óssea 4/farmacologia , Diferenciação Celular , Colágeno Tipo I , Feminino , Fibrina , Humanos , Masculino , Células-Tronco Mesenquimais/efeitos dos fármacos , Pessoa de Meia-Idade , Miócitos de Músculo Liso/efeitos dos fármacos , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Estimulação Física , Pressão , Esfingosina/análogos & derivados , Esfingosina/farmacologia , Estresse Mecânico , Engenharia Tecidual , Tecidos Suporte , Fator de Crescimento Transformador beta/farmacologia
9.
Nat Commun ; 10(1): 5627, 2019 12 09.
Artigo em Inglês | MEDLINE | ID: mdl-31819054

RESUMO

Current genome-wide screens allow system-wide study of drug resistance but detecting small nucleotide variants (SNVs) is challenging. Here, we use chemical mutagenesis, drug selection and next generation sequencing to characterize miltefosine and paromomycin resistant clones of the parasite Leishmania. We highlight several genes involved in drug resistance by sequencing the genomes of 41 resistant clones and by concentrating on recurrent SNVs. We associate genes linked to lipid metabolism or to ribosome/translation functions with miltefosine or paromomycin resistance, respectively. We prove by allelic replacement and CRISPR-Cas9 gene-editing that the essential protein kinase CDPK1 is crucial for paromomycin resistance. We have linked CDPK1 in translation by functional interactome analysis, and provide evidence that CDPK1 contributes to antimonial resistance in the parasite. This screen is powerful in exploring networks of drug resistance in an organism with diploid to mosaic aneuploid genome, hence widening the scope of its applicability.


Assuntos
Resistência a Medicamentos/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Leishmania/genética , Mutagênese , Mutação/genética , Paromomicina/farmacologia , Fosforilcolina/análogos & derivados , Fosforilação/efeitos dos fármacos , Fosforilcolina/farmacologia
10.
SAR QSAR Environ Res ; 30(12): 919-933, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31702401

RESUMO

Folates are essential biomolecules required to carry out many crucial processes in leishmania parasite. Dihydrofolate reductase-thymidylate synthase (DHFR-TS) and pteridine reductase 1 (PTR1) involved in folate biosynthesis in leishmania have been established as suitable targets for development of chemotherapy against leishmaniasis. In the present study, various computational tools such as homology modelling, pharmacophore modelling, docking, molecular dynamics and molecular mechanics have been employed to design dual DHFR-TS and PTR1 inhibitors. Two designed molecules, i.e. 2-(4-((4-nitrobenzyl)oxy)phenyl)-1H-benzo[d]imidazole and 2-(4-((2,4-dichlorobenzyl)oxy)phenyl)-1H-benzo[d]oxazolemolecules were synthesized. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) assay was performed to evaluate in vitro activity of molecules against promastigote form of Leishmania donovani using Miltefosine as standard. 2-(4-((4-nitrobenzyl)oxy)phenyl)-1H-benzo[d]imidazole and 2-(4-((2,4-dichlorobenzyl)oxy)phenyl)-1H-benzo[d]oxazolemolecules were found to be moderately active with showed IC50 = 68 ± 2.8 µM and 57 ± 4.2 µM, respectively.


Assuntos
Antiprotozoários/farmacologia , Leishmania donovani/efeitos dos fármacos , Complexos Multienzimáticos/química , Oxirredutases/química , Proteínas de Protozoários/química , Tetra-Hidrofolato Desidrogenase/química , Timidilato Sintase/química , Antiprotozoários/síntese química , Antiprotozoários/química , Benzimidazóis/síntese química , Benzimidazóis/química , Benzimidazóis/farmacologia , Benzoxazóis/síntese química , Benzoxazóis/química , Benzoxazóis/farmacologia , Descoberta de Drogas , Concentração Inibidora 50 , Leishmania donovani/metabolismo , Modelos Moleculares , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Relação Estrutura-Atividade
11.
J Pharm Pharmacol ; 71(12): 1871-1878, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31595517

RESUMO

OBJECTIVE: This work describes the isolation of anti-Leishmania amazonensis metabolites from Saururus cernuus (Saururaceae). Additionally, ultrastructural changes in promastigotes were evidenced by electron microscopy. METHODS: The MeOH extract from the leaves of S. cernuus was subjected to bioactivity-guided fractionation. Anti-L. amazonensis activity of purified compounds was performed in vitro against promastigote and amastigote forms. KEY FINDINGS: Bioactivity-guided fractionation of the MeOH extract from the leaves of S. cernuus afforded two related tetrahydrofuran dineolignans: threo,threo-manassantin A (1) and threo,erythro-manassantin A (2). Compounds 1 and 2 displayed activity against promastigotes (EC50 of 35.4 ± 7.7 and 17.6 ± 4.2 µm, respectively) and amastigotes (EC50 of 20.4 ± 1.9 and 16.0 ± 1.1 µm, respectively), superior to that determined for the positive control miltefosine (EC50 of 28.7 ± 3.5 µm). Reduced cytotoxicity for host cells was observed for both compounds. Additionally, ultrastructural changes in promastigotes leading to an alteration of structural morphology were observed, as evidenced by electron microscopy. Furthermore, these compounds altered the morphology and physiology of the plasmatic membrane of L. amazonensis. CONCLUSIONS: The obtained results indicated that dineolignans 1 and 2 could be considered as a scaffold for the design of novel and selective drug candidates for the treatment of leishmaniasis.


Assuntos
Antiprotozoários/farmacologia , Leishmania/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Lignanas/farmacologia , Saururaceae/química , Animais , Antiprotozoários/química , Antiprotozoários/isolamento & purificação , Furanos/química , Furanos/isolamento & purificação , Furanos/farmacologia , Leishmaniose/parasitologia , Lignanas/química , Lignanas/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C , Microscopia Eletrônica , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Folhas de Planta
12.
J Pharm Pharmacol ; 71(12): 1854-1863, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31595530

RESUMO

OBJECTIVES: The in vitro antileishmanial effect of analogues of resveratrol (AR) present in the N-aryl imines and N-aryl hydrazones series was investigated. In addition, possible parasite targets were evaluated. METHODS: Antipromastigote activity of Leishmania amazonensis, L. braziliensis and L. infantum, as well as the cytotoxicity on macrophages was determined by MTT assay and L. braziliensis-infected macrophages effect by Giemsa stain. After staining, effects on the parasite targets were analysed by flow cytometry or by fluorescence microscopy. KEY-FINDINGS: Among the tested compounds, the derivative AR26 showed the best effect against promastigotes of all Leishmania species (IC50  < 3.0 µg/ml), being more active than miltefosine, the control drug. AR26 was also effective against amastigotes of L. braziliensis (IC50  = 15.9 µg/ml), with low toxicity to mammalian cells. The evaluation of mechanism of action of AR26 on L. braziliensis promastigotes indicates mitochondrial potential depolarization, plasma membrane permeabilization, interference in the progression of the cell cycle and accumulation of autophagic vacuoles. In addition, any increase of the reactive oxygen species levels was detected in the treated L. braziliensis-macrophages. CONCLUSIONS: Data indicate that the antileishmanial activity of AR26 is related to multitarget action, and the resveratrol analogues could be used in future studies as antileishmanial agent.


Assuntos
Antiprotozoários/farmacologia , Leishmania/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Resveratrol/farmacologia , Animais , Antiprotozoários/administração & dosagem , Antiprotozoários/química , Modelos Animais de Doenças , Feminino , Hidrazonas/administração & dosagem , Hidrazonas/química , Hidrazonas/farmacologia , Iminas/administração & dosagem , Iminas/química , Iminas/farmacologia , Concentração Inibidora 50 , Leishmaniose/parasitologia , Macrófagos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Resveratrol/administração & dosagem , Resveratrol/análogos & derivados
13.
Biomacromolecules ; 20(11): 4135-4142, 2019 11 11.
Artigo em Inglês | MEDLINE | ID: mdl-31609601

RESUMO

Under pathological conditions, the joint is not well lubricated, which inevitably leads to osteoarthritis. Currently, in clinics injection of hyaluronic acid (HA) as an intra-articular viscosupplement is one of the main methods for alleviation of osteoarthritis. However, the viscosity of HA reduces dramatically under high shear rate due to the shear-thinning effect. Therefore, it is crucial to enhance the lubrication property of HA in order to treat osteoarthritis effectively. In this study, we successfully grafted 2-methacryloyloxyethyl phosphorylcholine (MPC), which is a zwitterionic biomaterial with excellent hydration lubrication, onto the HA with two different molecular weights (HAMPC) to enhance lubrication. The lubrication test performed using an atomic force microscope showed that, compared with HA, the friction coefficient of HAMPC was greatly reduced under various conditions. The in vitro test demonstrated that HAMPC was biocompatible and could upregulate cartilage anabolic genes while simultaneously downregulating cartilage catabolic proteases and pain-related genes. Importantly, high molecular weight HAMPC exhibited improved the capability to regulate these genes compared with low molecular weight HAMPC. In conclusion, the high molecular weight HAMPC developed herein, with enhanced lubrication and anti-inflammation, may be a promising polymer for the treatment of osteoarthritis.


Assuntos
Ácido Hialurônico/farmacologia , Articulações/efeitos dos fármacos , Metacrilatos/farmacologia , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Animais , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/química , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/ultraestrutura , Fricção/efeitos dos fármacos , Humanos , Ácido Hialurônico/síntese química , Ácido Hialurônico/química , Inflamação/tratamento farmacológico , Articulações/ultraestrutura , Lubrificantes/síntese química , Lubrificantes/química , Lubrificantes/farmacologia , Metacrilatos/síntese química , Metacrilatos/química , Camundongos , Microscopia de Força Atômica , Osteoartrite/tratamento farmacológico , Fosforilcolina/síntese química , Fosforilcolina/química , Polímeros/síntese química , Polímeros/química , Polímeros/farmacologia , Viscosidade/efeitos dos fármacos
14.
Langmuir ; 35(40): 13189-13195, 2019 10 08.
Artigo em Inglês | MEDLINE | ID: mdl-31547644

RESUMO

In clinics it is extremely important for implanted devices to achieve the property of enhanced lubrication and bacterial resistance; however, such a strategy has rarely been reported in previous literature. In the present study, a surface functionalization method, motivated by articular cartilage-inspired superlubrication and mussel-inspired adhesion, was proposed to modify titanium alloy (Ti6Al4V) using the copolymer (DMA-MPC) synthesized via free radical copolymerization. The copolymer-coated Ti6Al4V (Ti6Al4V@DMA-MPC) was evaluated by X-ray photoelectron spectroscopy, water contact angle, and Raman spectra to confirm that the DMA-MPC copolymer was successfully coated onto the Ti6Al4V substrate. In addition, the tribological test, with the polystyrene microsphere and Ti6Al4V or Ti6Al4V@DMA-MPC as the tribopair, indicated that the friction coefficient was greatly reduced for Ti6Al4V@DMA-MPC. Furthermore, the bacterial resistance test showed that bacterial attachment was significantly inhibited for Ti6Al4V@DMA-MPC for the three types of bacteria tested. The enhanced lubrication and bacterial resistance of Ti6Al4V@DMA-MPC was due to the tenacious hydration shell formed surrounding the zwitterionic charges in the phosphorylcholine group of the DMA-MPC copolymer. In summary, a bioinspired surface functionalization strategy is developed in this study, which can act as a universal and promising method to achieve enhanced lubrication and bacterial resistance for biomedical implants.


Assuntos
Antibacterianos/farmacologia , Biofilmes/efeitos dos fármacos , Materiais Revestidos Biocompatíveis/química , Lubrificantes/farmacologia , Metacrilatos/farmacologia , Fosforilcolina/análogos & derivados , Titânio/química , Animais , Antibacterianos/química , Antibacterianos/toxicidade , Linhagem Celular , Materiais Revestidos Biocompatíveis/toxicidade , Escherichia coli/efeitos dos fármacos , Escherichia coli/fisiologia , Lubrificantes/química , Lubrificantes/toxicidade , Lubrificação , Metacrilatos/química , Metacrilatos/toxicidade , Camundongos , Fosforilcolina/química , Fosforilcolina/farmacologia , Fosforilcolina/toxicidade , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/fisiologia , Streptococcus mutans/efeitos dos fármacos , Streptococcus mutans/fisiologia , Titânio/toxicidade
15.
Artigo em Inglês | MEDLINE | ID: mdl-31525614

RESUMO

OBJECTIVES: To gain insight into the propagation of miltefosine (MIL) resistance in visceral leishmaniasis, this laboratory study explored development of resistant parasites with a defective miltefosine transporter (MT) in sand flies. METHODS: Infectivity, colonization of stomodeal valve and metacyclogenesis of a MIL-resistant (MIL-R) Leishmania infantum LEM3323 line with a defective MT were assessed in the natural sand fly vectors Phlebotomus perniciosus and Lutzomyia longipalpis. Given our recent description of partial drug dependency of the MT-deficient line, the impact of MIL pre-exposure on sand fly infectivity was explored as well. RESULTS: A significant reduction in sand fly infection, stomodeal valve colonization and differentiation into metacyclics (determined by a lower flagellum/cell body length ratio) was observed in both vectors for MIL-R as compared to the isogenic parent MIL-susceptible line. Re-introduction of the wildtype MT gene into MIL-R tended to partially rescue the capacity to infect sand flies. Pre-exposure to MIL did not alter infectivity of the MIL-R line. CONCLUSION: The MIL resistant L. infantum LEM3323 line is significantly hampered in its development and transmissibility potential in two sand fly vector species. Additional studies are warranted to evaluate whether this applies to other visceral Leishmania parasites with acquired MIL-resistance.


Assuntos
Antiprotozoários/farmacologia , Insetos Vetores/parasitologia , Leishmania infantum/crescimento & desenvolvimento , Phlebotomus/parasitologia , Fosforilcolina/análogos & derivados , Psychodidae/parasitologia , Análise de Variância , Animais , Resistência a Medicamentos , Feminino , Concentração Inibidora 50 , Leishmania infantum/efeitos dos fármacos , Leishmaniose Visceral/tratamento farmacológico , Leishmaniose Visceral/parasitologia , Leishmaniose Visceral/transmissão , Camundongos , Camundongos Endogâmicos BALB C , Fosforilcolina/farmacologia , Coelhos
16.
Eur J Med Chem ; 183: 111676, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31542713

RESUMO

Leishmaniasis, a major health problem worldwide, has a limited arsenal of drugs for its control. The appearance of resistance to first- and second-line anti-leishmanial drugs confirms the need to develop new and less toxic drugs that overcome spontaneous resistance. In the present study, we report the design and synthesis of a novel library of 38 flavonol-like compounds and their evaluation in a panel of assays encompassing parasite killing, pharmacokinetics, genomics and ADME-Toxicity resulting in the progression of a compound in the drug discovery value chain. Compound 19, 2-(benzo[b]thiophen-3-yl)-3-hydroxy-6-methoxy-4H-chromen-4-one, exhibited a broad-spectrum activity against Leishmania spp. (EC50 1.9 µM for Leishmania infantum, 3.4 µM for L. donovani, 6.7 µM for L. major), Trypanosoma cruzi (EC50 7.5 µM) and T. brucei (EC50 0.8 µM). Focusing on anti-Leishmania activity, compound 19 challenge in vitro did not select for resistance markers in L. donovani, while a Cos-Seq screening for dominant resistance genes identified a gene locus on chromosome 36 that became ineffective at concentrations beyond EC50. Thus, compound 19 is a promising scaffold to tackle drug resistance in Leishmania infection. In vivo pharmacokinetic studies indicated that compound 19 has a long half-life (intravenous (IV): 63.2 h; per os (PO): 46.9 h) with an acceptable ADME-Toxicity profile. When tested in Leishmania infected hamsters, no toxicity and limited efficacy were observed. Low solubility and degradation were investigated spectroscopically as possible causes for the sub-optimal pharmacokinetic properties. Compound 19 resulted a specific compound based on the screening against a protein set, following the intrinsic fluorescence changes.


Assuntos
Antiprotozoários , Flavonóis , Leishmania/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Fosforilcolina/análogos & derivados , Tiofenos , Animais , Antiprotozoários/síntese química , Antiprotozoários/química , Antiprotozoários/farmacologia , Cricetinae , Avaliação Pré-Clínica de Medicamentos , Resistência a Medicamentos/efeitos dos fármacos , Flavonóis/síntese química , Flavonóis/química , Flavonóis/farmacologia , Genômica , Humanos , Fosforilcolina/química , Fosforilcolina/farmacologia , Tiofenos/síntese química , Tiofenos/química , Tiofenos/farmacologia
17.
Molecules ; 24(19)2019 Sep 22.
Artigo em Inglês | MEDLINE | ID: mdl-31546686

RESUMO

A novel stereoisomer of eushearilide, 23-demethyleushearilide, was synthesized, and the structure-activity relationships of this compound along with known eushearilide stereoisomers were investigated in order to design novel lead compounds for the treatment of fungal infections. It was discovered that all of these congeners, together with the natural product, exhibited a wide range of antimicrobial activity against not only fungi but also against bacteria, including methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE).


Assuntos
Macrolídeos/síntese química , Macrolídeos/farmacologia , Fosforilcolina/análogos & derivados , Anti-Infecciosos/síntese química , Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Macrolídeos/química , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Fosforilcolina/síntese química , Fosforilcolina/química , Fosforilcolina/farmacologia , Estereoisomerismo , Resistência a Vancomicina , Enterococos Resistentes à Vancomicina/efeitos dos fármacos
18.
Eur J Med Chem ; 183: 111660, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31514064

RESUMO

This mini-review focuses on leishmanicidal drugs that were sourced from small molecules previously approved for other diseases. The mechanisms of action of these molecules are herein explored, to probe the origins of their inter-species growth inhibitory activities. It is shown how the transversal action of the azoles - fluconazole, posaconazole and itraconazole - in both fungi and Leishmania is due to the occurrence of the same target, lanosterol 14-α-demethylase, in these two groups of species. In turn, the drugs miltefosine and amphotericin B are presented as truly multi-target agents, acting on small molecules, proteins, genes and even organelles. Steps towards future leishmanicidal drug candidates based on the multi-target strategy and on drug repurposing are also briefly presented.


Assuntos
Antifúngicos , Itraconazol , Leishmaniose/tratamento farmacológico , Anfotericina B/farmacologia , Antifúngicos/química , Antifúngicos/farmacologia , Reposicionamento de Medicamentos , Farmacorresistência Fúngica/efeitos dos fármacos , Fluconazol/farmacologia , Humanos , Itraconazol/química , Itraconazol/farmacologia , Leishmaniose/enzimologia , Terapia de Alvo Molecular , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Esterol 14-Desmetilase/metabolismo , Triazóis/farmacologia , Voriconazol/farmacologia
19.
Exp Parasitol ; 206: 107730, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31494215

RESUMO

Phospholipids are the main component of membranes and are responsible for cell integrity. Alkylphospholipid analogues (APs) were first designed as antitumoral agents and were later tested against different cell types. Trypanosoma cruzi, the Chagas disease etiological agent, is sensitive to APs (edelfosine, miltefosine and ilmofosine) in vitro. We investigated the effect of synthetic ring substituted AP against epimastigotes, amastigotes and trypomastigotes. TCAN26, could inhibit the in vitro growth of epimastigotes and amastigotes with the 50% inhibitory concentrations (IC50) in the nanomolar range. Trypomastigotes lysis was also induced with 24-h treatment and a LC50 of 2.3 µM. Ultrastructural analysis by electron microscopy demonstrated that TCAN26 mainly affected the parasite's membranes leading to mitochondrial and Golgi cisternae swelling, membrane blebs, and autophagic figures in the different parasite developmental stages. While the Golgi of the parasites was significantly affected, the Golgi complex of the host cells remained normal suggesting a specific mechanism of action. In summary, our results suggest that TCAN 26 is a potent and selective inhibitor of T. cruzi growth probably due to disturbances of phospholipid biosynthesis.


Assuntos
Adamantano/farmacologia , Fosforilcolina/análogos & derivados , Fosforilcolina/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Adamantano/química , Animais , Antiprotozoários/química , Antiprotozoários/farmacologia , Autofagia/efeitos dos fármacos , Membrana Celular/efeitos dos fármacos , Doença de Chagas/tratamento farmacológico , Doença de Chagas/parasitologia , Relação Dose-Resposta a Droga , Citometria de Fluxo , Complexo de Golgi/efeitos dos fármacos , Concentração Inibidora 50 , Dose Letal Mediana , Macrófagos Peritoneais/efeitos dos fármacos , Camundongos , Microscopia Eletrônica de Varredura , Microscopia Eletrônica de Transmissão , Microscopia de Fluorescência , Microscopia Imunoeletrônica , Mitocôndrias/efeitos dos fármacos , Fosforilcolina/química , Tripanossomicidas/química , Trypanosoma cruzi/crescimento & desenvolvimento , Trypanosoma cruzi/ultraestrutura
20.
Mater Sci Eng C Mater Biol Appl ; 104: 109916, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31499946

RESUMO

Composite resins (CRs) are widely used as dental restorative materials for caries treatment. They cause problems of secondary caries since Streptococcus mutans stays in the dental plaque, which the surface exists and produces acidic compounds during metabolism. The dental plaque depositions are induced by the protein adsorption on the surface. Therefore, suppression of protein adsorption on the surface of the CRs is important for inhibiting the formation of plaque and secondary caries. In this study we developed a surface treatment to provide an antibiofouling nature to the CRs by chemical reaction with 2-methacryloyloxyethyl phosphorylcholine (MPC) polymers in the oral cavity during dental treatment. To carry out the photochemical reaction on the remaining polymerizable groups of CRs, we synthesized the MPC polymer with a polymerizable group in the side chain. The MPC polymer could bind on the surfaces of the CRs chemically under dental treatment procedures. The treated surface showed significant resistance to oral protein adsorption and bacterial adhesion even when the surface was brushed with a toothbrush. Thus, we concluded that the photochemical reaction of the MPC polymer with the CRs in the oral cavity was good for making an antibiofouling surface and preventing secondary caries.


Assuntos
Resinas Acrílicas/farmacologia , Resinas Compostas/farmacologia , Placa Dentária/prevenção & controle , Metacrilatos/farmacologia , Fosforilcolina/análogos & derivados , Polímeros/farmacologia , Poliuretanos/farmacologia , Incrustação Biológica , Mucinas/metabolismo , Fósforo/análise , Fosforilcolina/farmacologia , Espectroscopia Fotoeletrônica , Polímeros/síntese química , Polímeros/química , Espectroscopia de Prótons por Ressonância Magnética , Silício/análise , Espectroscopia de Infravermelho com Transformada de Fourier , Streptococcus mutans/efeitos dos fármacos , Propriedades de Superfície
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