RESUMO
Decoquinate (DQ) is an antimicrobial agent commonly used as a feed additive for birds for human consumption. Its use as an additive is well established, but DQ has the potential for therapy as an antimicrobial drug for veterinary treatment and its optimized derivatives and/or formulations, mainly nanoformulations, have antimicrobial activity against pathogens that infect humans. However, DQ has a high partition coefficient and low solubility in aqueous fluids, and these biopharmaceutical properties have limited its use in humans. In this review, we highlight the antimicrobial activity and pharmacokinetic properties of DQ and highlight the solutions currently under investigation to overcome these drawbacks. A literature search was conducted focusing on the use of decoquinate against various infectious diseases in humans and animals. The search was conducted in several databases, including scientific and patent databases. Pharmaceutical nanotechnology and medicinal chemistry are the tools of choice to achieve human applications, and most of these applications have been able to improve the biopharmaceutical properties and pharmacokinetic profile of DQ. Based on the results presented here, DQ prototypes could be tested in clinical trials for human application in the coming years.
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Leishmaniasis are endemic diseases caused by different species of intracellular parasites of the genus Leishmania. Due to the high toxicity and drug resistance of current antileishmanial drugs, it is necessary to identify new and more effective drugs. Previously, we investigated the immunomodulatory and anti-Trypanosoma cruzi action of BA5, a derivative of betulinic acid. In the present study, we investigated the in vitro activity of BA5 against different species of Leishmania and their action mechanism. BA5 exhibited low cytotoxicity against macrophages and inhibited the proliferation of promastigote forms of Leishmania amazonensis (IC50 = 4.5 ± 1.1 µM), Leishmania major (IC50 = 3.0 ± 0.8 µM), Leishmania braziliensis (IC50 = 0.9 ± 1.1 µM) and Leishmania infantum (IC50 = 0.15 ± 0.05 µM). Incubation with BA5 reduced the percentage of Leishmania amazonensis-infected macrophages and the number of intracellular parasites (IC50 = 4.1 ± 0.7 µM). To understand the mechanism of action underlying BA5 antileishmanial activity (incubation at IC50/2, IC50 or 2xIC50 values of the drug), we investigated ultrastructural changes by scanning electron microscopy and evaluated cell cycle, membrane mitochondrial potential, and cell death against promastigote forms of Leishmania amazonensis by flow cytometry. Promastigotes incubated with BA5 presented membrane blebbing, flagella damage, increased size, and body deformation. Flow cytometry analysis showed that parasite death is mainly caused by apoptosis-like death, arrested cell cycle in G0/G1 phase and did not alter the membrane mitochondrial potential of Leishmania amazonensis. Surprisingly, the combination of BA5 and amphotericin B, an assay used to determine the degree of drug interaction, revealed synergistic effects (CI = 0.15 ± 0.09) on promastigotes forms of Leishmania amazonensis. In conclusion, BA5 compound is an effective and selective antileishmanial agent.
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Chagas disease is caused by Trypanosoma cruzi infection and remains a relevant cause of chronic heart failure in Latin America. The pharmacological arsenal for Chagas disease is limited, and the available anti-T. cruzi drugs are not effective when administered during the chronic phase. Cardiomyocytes derived from human-induced pluripotent stem cells (hiPSC-CMs) have the potential to accelerate the process of drug discovery for Chagas disease, through predictive preclinical assays in target human cells. Here, we aimed to establish a novel high-content screening- (HCS-) based method using hiPSC-CMs to simultaneously evaluate anti-T. cruzi activity and cardiotoxicity of chemical compounds. To provide proof-of-concept data, the reference drug benznidazole and three compounds with known anti-T. cruzi activity (a betulinic acid derivative named BA5 and two thiazolidinone compounds named GT5A and GT5B) were evaluated in the assay. hiPSC-CMs were infected with T. cruzi and incubated for 48 h with serial dilutions of the compounds for determination of EC50 and CC50 values. Automated multiparametric analyses were performed using an automated high-content imaging system. Sublethal toxicity measurements were evaluated through morphological measurements related to the integrity of the cytoskeleton by phalloidin staining, nuclear score by Hoechst 33342 staining, mitochondria score following MitoTracker staining, and quantification of NT-pro-BNP, a peptide released upon mechanical myocardial stress. The compounds showed EC50 values for anti-T. cruzi activity similar to those previously described for other cell types, and GT5B showed a pronounced trypanocidal activity in hiPSC-CMs. Sublethal changes in cytoskeletal and nucleus scores correlated with NT-pro-BNP levels in the culture supernatant. Mitochondrial score changes were associated with increased cytotoxicity. The assay was feasible and allowed rapid assessment of anti-T. cruzi action of the compounds, in addition to cardiotoxicity parameters. The utilization of hiPSC-CMs in the drug development workflow for Chagas disease may help in the identification of novel compounds.
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The need for new immunomodulatory drugs is due to the side effects associated with the prolonged use of the currently used immunomodulatory drugs. In this context, the present work aimed to investigate the immunomodulatory effect of an ethanolic concentrated extract from Physalis angulata. The cytotoxicity of samples was determined using peritoneal macrophages though the Alamar Blue assay. The immunomodulatory activity of the ethanolic extract from P. angulata on activated macrophages was determined by measurement of nitrite and cytokine production. The immunosuppressive effects of the ethanolic extract from P. angulata was evaluated on lymphocyte proliferation and cytokine production. The effects of the extract on cell cycle progression and cell death on lymphocytes were evaluated by flow cytometry. Lastly, the ethanolic extract from P. angulata was tested in vivo in toxicological tests and in models of peritonitis and delayed-type hypersensitivity response. The ethanolic extract from P. angulata decreased nitrite, interleukin-6, interleukin-12, and TNF-α production by activated macrophages without affecting the cell viability. In addition, the ethanolic extract from P. angulata inhibited lymphoproliferation and the secretion of interleukin-2, interleukin-6, and IFN-γ, and increased interleukin-4 secretion by activated splenocytes. Flow cytometry analysis in lymphocyte cultures showed that treatment with the ethanolic extract from P. angulata induces cell cycle arrest in the G1 phase followed by cell death by apoptosis. Moreover, mice treated with the extract from P. angulata at 100 or 200 mg/kg did not show signs of toxicity or alterations in serum components. Finally, the ethanolic extract from P. angulata significantly reduced neutrophil migration and reduced paw edema in bovine serum albumin-induced the delayed-type hypersensitivity response model. Our results demonstrate the potential of the ethanolic extract of P. angulata as an alternative for the treatment of immune-inflammatory diseases.
Assuntos
Physalis , Animais , Etanol , Macrófagos , Macrófagos Peritoneais , Camundongos , Extratos Vegetais/farmacologiaRESUMO
Leishmaniasis are group of neglected diseases with worldwide distribution that affect about 12 million people. The current treatment is limited and may cause severe adverse effects, and thus, the search for new drugs more effective and less toxic is relevant. We have previously investigated the immunomodulatory effects of LASSBio-1386, an N-acylhydrazone derivative. Here we investigated the in vitro and in vivo activity of LASSBio-1386 against L. amazonensis. LASSBio-1386 inhibited the proliferation of promastigotes of L. amazonensis (EC50 = 2.4 ± 0.48 µM), while presenting low cytotoxicity to macrophages (CC50 = 74.1 ± 2.9 µM). In vitro incubation with LASSBio-1386 reduced the percentage of Leishmania-infected macrophages and the number of intracellular parasites (EC50 = 9.42 ± 0.64 µM). Also, in vivo treatment of BALB/c mice infected with L. amazonensis resulted in a decrease of lesion size, parasitic load and caused histopathological alterations, when compared to vehicle-treated control. Moreover, LASSBio-1386 caused ultrastructural changes, arrested cell cycle in G0/G1 phase and did not alter the membrane mitochondrial potential of L. amazonensis. Aiming to its possible molecular interactions, we performed docking and molecular dynamics studies on Leishmania phosphodiesterase B1 (PDB code: 2R8Q) and LASSBio-1386. The computational analyses suggest that LASSBio-1386 acts against Leishmania through the modulation of leishmanial PDE activity. In conclusion, our results indicate that LASSBio-1386 is a promising candidate for the development of new leishmaniasis treatment.
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Asthma is a chronic, complex and heterogeneous inflammatory illness, characterized by obstruction of the lower airways. About 334 million people worldwide suffer from asthma, and these estimates, as well as the severity of the disease, have increased in the last decades. Glucocorticoids are currently the most widely used drugs in the treatment and control of asthma symptoms, but their prolonged use can cause serious adverse effects. N-acylhydrazone derivatives have been tested in pre-clinical studies in models of inflammatory diseases. Here we tested SintMed65 (N'-[(1E)-3-(4-nitrophenylhydrazono)]-(2E)-propan-2-ylidene-3,5-dinitrobenzohydrazide), a compound belonging to a novel class of immunosuppressive drugs, in a mouse model of allergic airway inflammation. BALB/c mice were sensitized previously and challenged with ovalbumin for five consecutive days and SintMed65 treatment was performed orally 1â¯h prior to challenge with ovalbumin. Administration of SintMed65, as well as the reference drug dexamethasone, reduced cellularity and the number of eosinophils in the bronchoalveolar fluid (BALF). SintMed65 also reduced the production of Th2 cytokines IL-4, IL-5 and IL-13 in the BALF, and IL-4, IL-10 and CCL8 gene expression in lung, compared to vehicle-treated mice. Importantly, a reduction in the number of leukocytes and in the mucus production in lungs of SintMed65-treated mice was found, compared to the vehicle-treated group. In contrast, IgE production was not significantly altered after treatment with SintMed65. Our results demonstrate that compound SintMed65 possesses anti-inflammatory characteristics, suggesting its therapeutic potential for the treatment of allergic diseases.
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Antiasmáticos/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Asma/tratamento farmacológico , Hidrazonas/uso terapêutico , Alérgenos , Animais , Asma/imunologia , Asma/patologia , Líquido da Lavagem Broncoalveolar/imunologia , Citocinas/imunologia , Modelos Animais de Doenças , Contagem de Leucócitos , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Masculino , Camundongos Endogâmicos BALB C , Muco/imunologia , OvalbuminaRESUMO
Chronic Chagas disease cardiomyopathy (CCC) is a major cause of heart disease in Latin America and treatment for this condition is unsatisfactory. Here we investigated the effects of BA5, an amide semi-synthetic derivative betulinic acid, in a model of CCC. Mice chronically infected with T. cruzi were treated orally with BA5 (10 or 1 mg/Kg), three times per week, for 2 months. BA5 treatment decreased inflammation and fibrosis in heart sections but did not improve exercise capacity or ameliorate cardiac electric disturbances in infected mice. Serum concentrations of TNF-α, IFN-γ, and IL-1ß, as well as cardiac gene expression of pro-inflammatory mediators, were reduced after BA5 treatment. In contrast, a significant increase in the anti-inflammatory cytokine IL-10 concentration was observed in BA5-treated mice in both tested doses compared to vehicle-treated mice. Moreover, polarization to anti-inflammatory/M2 macrophage phenotype was evidenced by a decrease in the expression of NOS2 and proinflammatory cytokines and the increase in M2 markers, such as Arg1 and CHI3 in mice treated with BA5. In conclusion, BA5 had a potent anti-inflammatory activity on a model of parasite-driven heart disease related to IL-10 production and a switch from M1 to M2 subset of macrophages.
Assuntos
Anti-Inflamatórios/farmacologia , Cardiomiopatia Chagásica/tratamento farmacológico , Interleucina-10/imunologia , Macrófagos/imunologia , Triterpenos/farmacologia , Trypanosoma cruzi/imunologia , Animais , Cardiomiopatia Chagásica/imunologia , Cardiomiopatia Chagásica/patologia , Doença Crônica , Modelos Animais de Doenças , Fibrose , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Macrófagos/patologia , Camundongos , Triterpenos Pentacíclicos , Ácido BetulínicoRESUMO
The objective of this work was to obtain and evaluate anti-inflammatory in vitro, in vivo and in silico potential of novel indole-N-acylhydrazone derivatives. In total, 10 new compounds (3a-j) were synthesized in satisfactory yields, through a condensation reaction in a single synthesis step. In the lymphoproliferation assay, using mice splenocytes, 3a and 3b showed inhibition of lymphocyte proliferation of 62.7% (±3.5) and 50.7% (±2), respectively, while dexamethasone presented an inhibition of 74.6% (±2.4). Moreover, compound 3b induced higher Th2 cytokines production in mice splenocytes cultures. The results for COX inhibition assays showed that compound 3b is a selective COX-2 inhibitor, but with less potency when compared to celecoxib, and compound 3a not presented selectivity towards COX-2. The molecular docking results suggest compounds 3a and 3b interact with the active site of COX-2 in similar conformations, but not with the active site of COX-1, and this may be the main reason to the COX-2 selectivity of compound 3b. In vivo carrageenan-induced paw edema assays were adopted for the confirmation of the anti-inflammatory activity. Compound 3b showed better results in suppressing edema at all tested concentrations and was able to induce an edema inhibition of 100% after 5â¯h of carrageenan injection at the 30â¯mgâ¯kg-1 dosage, corroborating with the COX inhibition and lymphoproliferation results. I addition to our experimental results, in silico analysis suggest that compounds 3a and 3b present a well-balanced profile between pharmacodynamics and pharmacokinetics. Thus, our preliminary results revealed the potentiality of a new COX-2 selective derivative in the modulation of the inflammatory process.
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Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/metabolismo , Inibidores de Ciclo-Oxigenase/química , Inibidores de Ciclo-Oxigenase/farmacologia , Hidrazonas/química , Hidrazonas/farmacologia , Acilação , Animais , Anti-Inflamatórios não Esteroides/síntese química , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacologia , Anti-Inflamatórios não Esteroides/uso terapêutico , Carragenina , Linhagem Celular , Inibidores de Ciclo-Oxigenase 2/síntese química , Inibidores de Ciclo-Oxigenase 2/química , Inibidores de Ciclo-Oxigenase 2/farmacologia , Inibidores de Ciclo-Oxigenase/síntese química , Inibidores de Ciclo-Oxigenase/uso terapêutico , Edema/induzido quimicamente , Edema/tratamento farmacológico , Edema/enzimologia , Feminino , Humanos , Hidrazonas/síntese química , Hidrazonas/uso terapêutico , Indóis/síntese química , Indóis/química , Indóis/farmacologia , Indóis/uso terapêutico , Camundongos Endogâmicos BALB C , Simulação de Acoplamento MolecularRESUMO
Immunosuppressive drugs are widely used for the treatment of immune-mediated diseases and inflammation, but the toxicity and side effects of the available immunosuppressors make the search of new agents of great relevance. Here, we evaluated the immunomodulatory activity of an N-acylhydrazone derivative, (E)-N'-(3,4-dimethoxybenzylidene)-4-methoxybenzohydrazide (LASSBio-1386), a phosphodiesterase-4 (PDE-4) inhibitor. LASSBio-1386 inhibited lymphocyte activation in a concentration-dependent fashion, decreasing lymphoproliferation and IFN-γ and IL-2 production stimulated by anti-CD3/CD28 mAbs or concanavalin A (Con A) and inducing cell-cycle arrest in the G0/G1 phase. These effects were not blocked by RU486, a glucocorticoid receptor (GR) antagonist, indicating an effect independent of glucocorticoid receptor activation. Combination index-isobologram analysis indicates a synergistic effect between LASSBio-1386 and dexamethasone in lymphoproliferation inhibition. LASSBio-1386 presented immunomodulatory action in macrophage cultures, as observed by a significant and concentration-dependent decrease in NO and TNF-α production, an effect achieved by reducing IĸB expression and NF-κB activation. In the mouse model of endotoxic shock, LASSBio-1386 at 50 and 100â¯mg/kg protected 50 and 85% of mice against LPS-induced lethality, respectively. In agreement to its in vitro action, treatment with 100â¯mg/kg of LASSBio-1386 reduced TNF-α and IL-1ß serum levels, while increased IL-6 and IL-10. Finally, LASSBio-1386 reduced the paw edema in a BSA-induced delayed-type hypersensitivity model. These findings demonstrate the immunomodulatory and immunosuppressant effects of LASSBio-1386 and indicate this molecule is a promising pharmacologic agent for immune-mediated diseases.
Assuntos
Hidrazonas/farmacologia , Hipersensibilidade Tardia/tratamento farmacológico , Imunossupressores/farmacologia , Lipopolissacarídeos/toxicidade , Inibidores da Fosfodiesterase 4/farmacologia , Choque/tratamento farmacológico , Animais , Benzamidas , Citocinas/genética , Citocinas/metabolismo , Dexametasona/farmacologia , Antagonistas de Hormônios/farmacologia , Hidrazonas/química , Macrófagos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mifepristona/farmacologia , Estrutura Molecular , NF-kappa B/genética , NF-kappa B/metabolismo , Óxido Nítrico/metabolismo , Células RAW 264.7RESUMO
Betulinic acid (BA) is a naturally occurring triterpenoid with several biological properties already described, including immunomodulatory activity. Here we investigated the immunomodulatory activity of eight semi-synthetic amide derivatives of betulinic acid. Screening of derivatives BA1-BA8 led to the identification of compounds with superior immunomodulatory activity than BA on activated macrophages and lymphocytes. BA5, the most potent derivative, inhibited nitric oxide and TNFα production in a concentration-dependent manner, and decreased NF-κB activation in Raw 264.7 cells. Additionally, BA5 inhibited the proliferation of activated lymphocytes and the secretion of IL-2, IL-4 IL-6, IL-10, IL-17A and IFNÉ£, in a concentration-dependent manner. Flow cytometry analysis in lymphocyte cultures showed that treatment with BA5 induces cell cycle arrest in pre-G1 phase followed by cell death by apoptosis. Moreover, BA5 also inhibited the activity of calcineurin, an enzyme that plays a critical role in the progression of cell cycle and T lymphocyte activation. BA5 has a synergistic inhibitory effect with dexamethasone on lymphoproliferation, showing a promising profile for drug combination. Finally, we observed immunosuppressive effects of BA5 in vivo in mouse models of lethal endotoxemia and delayed type hypersensitivity. Our results reinforce the potential use of betulinic acid and its derivatives in the search for potent immunomodulatory drugs.