RESUMO
The design of immunogens that elicit broadly reactive neutralizing antibodies (bnAbs) has been a major obstacle to HIV-1 vaccine development. One approach to assess potential immunogens is to use mice expressing precursors of human bnAbs as vaccination models. The bnAbs of the VRC01-class derive from the IGHV1-2 immunoglobulin heavy chain and neutralize a wide spectrum of HIV-1 strains via targeting the CD4 binding site of the envelope glycoprotein gp120. We now describe a mouse vaccination model that allows a germline human IGHV1-2(∗)02 segment to undergo normal V(D)J recombination and, thereby, leads to the generation of peripheral B cells that express a highly diverse repertoire of VRC01-related receptors. When sequentially immunized with modified gp120 glycoproteins designed to engage VRC01 germline and intermediate antibodies, IGHV1-2(∗)02-rearranging mice, which also express a VRC01-antibody precursor light chain, can support the affinity maturation of VRC01 precursor antibodies into HIV-neutralizing antibody lineages.
Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , HIV-1/imunologia , Imunização , Cadeias Pesadas de Imunoglobulinas/imunologia , Células Precursoras de Linfócitos B/imunologia , Animais , Anticorpos Monoclonais/genética , Linfócitos B/imunologia , Anticorpos Amplamente Neutralizantes , Linhagem Celular , Modelos Animais de Doenças , Regulação da Expressão Gênica/imunologia , Anticorpos Anti-HIV , Cadeias Pesadas de Imunoglobulinas/química , Cadeias Pesadas de Imunoglobulinas/genética , Concentração Inibidora 50 , Camundongos , Deleção de Sequência , Linfócitos T/imunologiaRESUMO
The B.1.1.529/Omicron variant of SARS-CoV-2 was only recently detected in southern Africa, but its subsequent spread has been extensive, both regionally and globally1. It is expected to become dominant in the coming weeks2, probably due to enhanced transmissibility. A striking feature of this variant is the large number of spike mutations3 that pose a threat to the efficacy of current COVID-19 vaccines and antibody therapies4. This concern is amplified by the findings of our study. Here we found that B.1.1.529 is markedly resistant to neutralization by serum not only from patients who recovered from COVID-19, but also from individuals who were vaccinated with one of the four widely used COVID-19 vaccines. Even serum from individuals who were vaccinated and received a booster dose of mRNA-based vaccines exhibited substantially diminished neutralizing activity against B.1.1.529. By evaluating a panel of monoclonal antibodies against all known epitope clusters on the spike protein, we noted that the activity of 17 out of the 19 antibodies tested were either abolished or impaired, including ones that are currently authorized or approved for use in patients. Moreover, we also identified four new spike mutations (S371L, N440K, G446S and Q493R) that confer greater antibody resistance on B.1.1.529. The Omicron variant presents a serious threat to many existing COVID-19 vaccines and therapies, compelling the development of new interventions that anticipate the evolutionary trajectory of SARS-CoV-2.
Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/virologia , Evasão da Resposta Imune/imunologia , SARS-CoV-2/imunologia , Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , COVID-19/sangue , COVID-19/imunologia , Vacinas contra COVID-19/administração & dosagem , Vacinas contra COVID-19/imunologia , Linhagem Celular , Convalescença , Evolução Molecular , Humanos , Soros Imunes/imunologia , Concentração Inibidora 50 , Modelos Moleculares , Mutação , Testes de Neutralização , SARS-CoV-2/química , SARS-CoV-2/classificação , SARS-CoV-2/genética , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologiaRESUMO
The COVID-19 pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has posed a worldwide threat in the past 3 years. Although it has been widely and intensively investigated, the mechanism underlying the coronavirus-host interaction requires further elucidation, which may contribute to the development of new antiviral strategies. Here, we demonstrated that the host cAMP-responsive element-binding protein (CREB1) interacts with the non-structural protein 13 (nsp13) of SARS-CoV-2, a conserved helicase for coronavirus replication, both in cells and in lung tissues subjected to SARS-CoV-2 infection. The ATPase and helicase activity of viral nsp13 were shown to be potentiated by CREB1 association, as well as by Protein kinase A (PKA)-mediated CREB1 activation. SARS-CoV-2 replication is significantly suppressed by PKA Cα, cAMP-activated protein kinase catalytic subunit alpha (PRKACA), and CREB1 knockdown or inhibition. Consistently, the CREB1 inhibitor 666-15 has shown significant antiviral effects against both the WIV04 strain and the Omicron strain of the SARS-CoV-2. Our findings indicate that the PKA-CREB1 signaling axis may serve as a novel therapeutic target against coronavirus infection. IMPORTANCE: In this study, we provide solid evidence that host transcription factor cAMP-responsive element-binding protein (CREB1) interacts directly with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) helicase non-structural protein 13 (nsp13) and potentiate its ATPase and helicase activity. And by live SARS-CoV-2 virus infection, the inhibition of CREB1 dramatically impairs SARS-CoV-2 replication in vivo. Notably, the IC50 of CREB1 inhibitor 666-15 is comparable to that of remdesivir. These results may extend to all highly pathogenic coronaviruses due to the conserved nsp13 sequences in the virus.
Assuntos
RNA-Polimerase RNA-Dependente de Coronavírus , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico , Proteínas Quinases Dependentes de AMP Cíclico , Interações entre Hospedeiro e Microrganismos , SARS-CoV-2 , Proteínas não Estruturais Virais , Replicação Viral , Humanos , Adenosina Trifosfatases/metabolismo , Antivirais/farmacologia , RNA-Polimerase RNA-Dependente de Coronavírus/metabolismo , COVID-19/virologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/antagonistas & inibidores , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/deficiência , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , DNA Helicases/metabolismo , Concentração Inibidora 50 , RNA Helicases/metabolismo , SARS-CoV-2/classificação , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/enzimologia , SARS-CoV-2/crescimento & desenvolvimento , Transdução de Sinais/efeitos dos fármacos , Proteínas não Estruturais Virais/metabolismo , Replicação Viral/efeitos dos fármacos , Feminino , Animais , CamundongosRESUMO
Pseudorabies virus (PRV) has become a "new life-threatening zoonosis" since the human-originated PRV strain was first isolated in 2020. To identify novel anti-PRV agents, we screened a total of 107 ß-carboline derivatives and found 20 compounds displaying antiviral activity against PRV. Among them, 14 compounds showed better antiviral activity than acyclovir. We found that compound 45 exhibited the strongest anti-PRV activity with an IC50 value of less than 40 nM. Our in vivo studies showed that treatment with 45 significantly reduced the viral loads and protected mice challenged with PRV. To clarify the mode of action of 45, we conducted a time of addition assay, an adsorption assay, and an entry assay. Our results indicated that 45 neither had a virucidal effect nor affected viral adsorption while significantly inhibiting PRV entry. Using the FITC-dextran uptake assay, we determined that 45 inhibits macropinocytosis. The actin-dependent plasma membrane protrusion, which is important for macropinocytosis, was also suppressed by 45. Furthermore, the kinase DYRK1A (dual-specificity tyrosine phosphorylation-regulated kinase 1A) was predicted to be a potential target for 45. The binding of 45 to DYRK1A was confirmed by drug affinity responsive target stability and cellular thermal shift assay. Further analysis revealed that knockdown of DYRK1A by siRNA suppressed PRV macropinocytosis and the tumor necrosis factor alpha-TNF-induced formation of protrusions. These results suggested that 45 could restrain PRV macropinocytosis by targeting DYRK1A. Together, these findings reveal a unique mechanism through which ß-carboline derivatives restrain PRV infection, pointing to their potential value in the development of anti-PRV agents.
Assuntos
Antivirais , Carbolinas , Herpesvirus Suídeo 1 , Animais , Humanos , Camundongos , Aciclovir/farmacologia , Aciclovir/toxicidade , Antivirais/química , Antivirais/farmacologia , Antivirais/uso terapêutico , Carbolinas/química , Carbolinas/farmacologia , Carbolinas/uso terapêutico , Técnicas de Silenciamento de Genes , Herpesvirus Suídeo 1/efeitos dos fármacos , Concentração Inibidora 50 , Pinocitose/efeitos dos fármacos , Proteínas Tirosina Quinases/antagonistas & inibidores , Pseudorraiva/tratamento farmacológico , Pseudorraiva/prevenção & controle , Pseudorraiva/virologia , Internalização do Vírus/efeitos dos fármacos , Células HeLa , Modelos Químicos , Quinases DyrkRESUMO
Testing Plasmodium vivax antimicrobial sensitivity is limited to ex vivo schizont maturation assays, which preclude determining the IC50s of delayed action antimalarials such as doxycycline. Using Plasmodium cynomolgi as a model for P. vivax, we determined the physiologically significant delayed death effect induced by doxycycline [IC50(96 h), 1,401 ± 607 nM]. As expected, IC50(96 h) to chloroquine (20.4 nM), piperaquine (12.6 µM), and tafenoquine (1,424 nM) were not affected by extended exposure.
Assuntos
Aminoquinolinas , Antimaláricos , Doxiciclina , Piperazinas , Plasmodium cynomolgi , Plasmodium vivax , Doxiciclina/farmacologia , Antimaláricos/farmacologia , Aminoquinolinas/farmacologia , Plasmodium vivax/efeitos dos fármacos , Plasmodium cynomolgi/efeitos dos fármacos , Cloroquina/farmacologia , Animais , Malária Vivax/tratamento farmacológico , Malária Vivax/parasitologia , Quinolinas/farmacologia , Concentração Inibidora 50 , Humanos , Testes de Sensibilidade ParasitáriaRESUMO
To date, there is no licensed vaccine for Middle East respiratory syndrome coronavirus (MERS-CoV). Therefore, MERS-CoV is one of the diseases targeted by the Coalition for Epidemic Preparedness Innovations (CEPI) vaccine development programs and has been classified as a priority disease by the World Health Organization (WHO). An important measure of vaccine immunogenicity and antibody functionality is the detection of virus-neutralizing antibodies. We have developed and optimized a microneutralization assay (MNA) using authentic MERS-CoV and standardized automatic counting of virus foci. Compared to our standard virus neutralization assay, the MNA showed improved sensitivity when analyzing 30 human sera with good correlation of results (Spearman's correlation coefficient r = 0.8917, p value < 0.0001). It is important to use standardized materials, such as the WHO international standard (IS) for anti-MERS-CoV immunoglobulin G, to compare the results from clinical trials worldwide. Therefore, in addition to the neutralizing titers (NT50 = 1384, NT80 = 384), we determined the IC50 and IC80 of WHO IS in our MNA to be 0.67 IU/ml and 2.6 IU/ml, respectively. Overall, the established MNA is well suited to reliably quantify vaccine-induced neutralizing antibodies with high sensitivity.
Assuntos
Anticorpos Neutralizantes , Anticorpos Antivirais , Coronavírus da Síndrome Respiratória do Oriente Médio , Testes de Neutralização , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Humanos , Testes de Neutralização/métodos , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/diagnóstico , Animais , Concentração Inibidora 50 , Sensibilidade e EspecificidadeRESUMO
New affordable drugs are needed for the treatment of infection with the protozoan parasite Trypanosoma cruzi responsible for the Chagas disease (CD). Only two old drugs are currently available, nifurtimox and benznidazole (Bz) but they exhibit unwanted side effects and display a weak activity in the late chronic phase of the disease. In this context, we evaluated the activity of a series of aryl-pyrazolone derivatives against T cruzi, using both bloodstream trypomastigote and intracellular amastigote forms of the parasite. The test compounds originate from a series of anticancer agents targeting the immune checkpoint ligand PD-L1 and bear an analogy with known anti-trypanosomal pyrazolones. A first group of 6 phenyl-pyrazolones was tested, revealing the activity of a single pyridyl-pyrazolone derivative. Then a second group of 8 compounds with a common pyridyl-pyrazolone core was evaluated. The in vitro testing process led to the identification of two non-cytotoxic and highly potent molecules against the intracellular form of T. cruzi, with an activity comparable to Bz. Moreover, one compound revealed an activity largely superior to that of Bz against bloodstream trypomastigotes, while being non-cytotoxic (selectivity index >1000). Unfortunately, the compound showed little activity in vivo, most likely due to its very limited plasma stability. However, the study opens novel perspectives for the design of new anti-trypanosomal products and the mechanism of action of the compounds is discussed.
Assuntos
Doença de Chagas , Pirazolonas , Tripanossomicidas , Trypanosoma cruzi , Trypanosoma cruzi/efeitos dos fármacos , Pirazolonas/farmacologia , Pirazolonas/química , Tripanossomicidas/farmacologia , Tripanossomicidas/química , Animais , Camundongos , Doença de Chagas/tratamento farmacológico , Doença de Chagas/parasitologia , Piridinas/farmacologia , Piridinas/química , Concentração Inibidora 50 , Nitroimidazóis/farmacologia , Nitroimidazóis/químicaRESUMO
Trypanosoma cruzi (T. cruzi) causes Chagas, which is a neglected tropical disease (NTD). WHO estimates that 6 to 7 million people are infected worldwide. Current treatment is done with benznidazole (BZN), which is very toxic and effective only in the acute phase of the disease. In this work, we designed, synthesized, and characterized thirteen new phenoxyhydrazine-thiazole compounds and applied molecular docking and in vitro methods to investigate cell cytotoxicity, trypanocide activity, nitric oxide (NO) production, cell death, and immunomodulation. We observed a higher predicted affinity of the compounds for the squalene synthase and 14-alpha demethylase enzymes of T. cruzi. Moreover, the compounds displayed a higher predicted affinity for human TLR2 and TLR4, were mildly toxic in vitro for most mammalian cell types tested, and LIZ531 (IC50 2.8 µM) was highly toxic for epimastigotes, LIZ311 (IC50 8.6 µM) for trypomastigotes, and LIZ331 (IC50 1.9 µM) for amastigotes. We observed that LIZ311 (IC50 2.5 µM), LIZ431 (IC50 4.1 µM) and LIZ531 (IC50 5 µM) induced 200 µg/mL of NO and JM14 induced NO production in three different concentrations tested. The compound LIZ331 induced the production of TNF and IL-6. LIZ311 induced the secretion of TNF, IFNγ, IL-2, IL-4, IL-10, and IL-17, cell death by apoptosis, decreased acidic compartment formation, and induced changes in the mitochondrial membrane potential. Taken together, LIZ311 is a promising anti-T. cruzi compound is not toxic to mammalian cells and has increased antiparasitic activity and immunomodulatory properties.
Assuntos
Doença de Chagas , Simulação de Acoplamento Molecular , Óxido Nítrico , Tiazóis , Tripanossomicidas , Trypanosoma cruzi , Trypanosoma cruzi/efeitos dos fármacos , Tiazóis/farmacologia , Tiazóis/química , Doença de Chagas/tratamento farmacológico , Doença de Chagas/imunologia , Humanos , Animais , Camundongos , Óxido Nítrico/metabolismo , Óxido Nítrico/biossíntese , Tripanossomicidas/farmacologia , Tripanossomicidas/química , Concentração Inibidora 50 , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Hidrazinas/farmacologia , Hidrazinas/química , Citocinas/metabolismo , Camundongos Endogâmicos BALB CRESUMO
OBJECTIVES: Malaria is a significant global health challenge, particularly in Africa, Asia, and Latin America, necessitating immediate investigation into innovative and efficacious treatments. This work involves the development of pyrazole substituted 1,3,5-triazine derivatives as antimalarial agent. METHODS: In this study, ten compounds 7(a-j) were synthesized by using nucleophilic substitution reaction, screened for in silico study and their antimalarial activity were evaluated against 3D7 (chloroquine-sensitive) strain of P. falciparum. KEY FINDING: The present work involves the development of hybrid trimethoxy pyrazole 1,3,5-triazine derivatives 7 (a-j). Through in silico analysis, four compounds were identified with favorable binding energy and dock scores. The primary focus of the docking investigations was on the examination of hydrogen bonding and the associated interactions with certain amino acid residues, including Arg A122, Ser A108, Ser A111, Ile A164, Asp A54, and Cys A15. The IC50 values of the four compounds were measured in vitro to assess their antimalarial activity against the chloroquine sensitive 3D7 strain of P. falciparum. The IC50 values varied from 25.02 to 54.82 µg/mL. CONCLUSION: Among the ten derivatives, compound 7J has considerable potential as an antimalarial agent, making it a viable contender for further refinement in the realm of pharmaceutical exploration, with the aim of mitigating the global malaria load.
Assuntos
Antimaláricos , Concentração Inibidora 50 , Simulação de Acoplamento Molecular , Plasmodium falciparum , Pirazóis , Triazinas , Antimaláricos/farmacologia , Antimaláricos/síntese química , Antimaláricos/química , Pirazóis/farmacologia , Pirazóis/química , Pirazóis/síntese química , Triazinas/farmacologia , Triazinas/química , Triazinas/síntese química , Plasmodium falciparum/efeitos dos fármacos , Simulação por Computador , Desenho de Fármacos , Relação Estrutura-Atividade , Humanos , Cloroquina/farmacologia , Cloroquina/química , Ligação de HidrogênioRESUMO
Giardiasis is a prevalent parasitic diarrheal disease caused by Giardia lamblia, affecting people worldwide. Recently, the availability of several drugs for its treatment has highlighted issues such as multidrug resistance, limited effectiveness and undesirable side effects. Therefore, it is necessary to develop alternative new drugs and treatment strategies that can enhance therapeutic outcomes and effectively treat giardiasis. Natural compounds show promise in the search for more potent anti-giardial agents. Our investigation focused on the effect of Andrographolide (ADG), an active compound of the Andrographis paniculata plant, on Giardia lamblia, assessing trophozoite growth, morphological changes, cell cycle arrest, DNA damage and inhibition of gene expression associated with pathogenic factors. ADG demonstrated anti-Giardia activity almost equivalent to the reference drug metronidazole, with an IC50 value of 4.99 µM after 24 h of incubation. In cytotoxicity assessments and morphological examinations, it showed significant alterations in trophozoite shape and size and effectively hindered the adhesion of trophozoites. It also caused excessive ROS generation, DNA damage, cell cycle arrest and inhibited the gene expression related to pathogenesis. Our findings have revealed the anti-giardial efficacy of ADG, suggesting its potential as an agent against Giardia infections. This could offer a natural and low-risk treatment option for giardiasis, reducing the risk of side effects and drug resistance.
Assuntos
Antiprotozoários , Pontos de Checagem do Ciclo Celular , Dano ao DNA , Diterpenos , Giardia lamblia , Concentração Inibidora 50 , Espécies Reativas de Oxigênio , Trofozoítos , Diterpenos/farmacologia , Giardia lamblia/efeitos dos fármacos , Giardia lamblia/crescimento & desenvolvimento , Giardia lamblia/genética , Trofozoítos/efeitos dos fármacos , Trofozoítos/crescimento & desenvolvimento , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Dano ao DNA/efeitos dos fármacos , Antiprotozoários/farmacologia , Humanos , Animais , Expressão Gênica/efeitos dos fármacos , Metronidazol/farmacologiaRESUMO
Sheep haemonchosis is a disease that causes serious losses in livestock production, particularly with the increase of cases of anthelmintic resistance around the world. This justifies the urgent need of alternative solutions. The aim of this study was to determine the chemical profile, in vitro, and, in vivo, anthelmintic properties of Thymus capitatus essential oil. To evaluate the, in vitro, anthelmintic activity of the T. capitatus EO on Haemonchus contortus, two tests were used: egg hatch assay (EHA) and adult worm motility (AWM) assay. The nematicidal effect of this oil was evaluated, in vivo, in mice infected artificially with Heligmosomoides polygyrus using faecal egg count reduction (FECR) and total worm count reduction (TWCR). Chromatographic characterization of T.capitatus composition using gas chromatography coupled to mass spectrometry (GC-MS) demonstrated the presence of carvacrol (81.16%), as the major constituents. The IC50 values obtained was 1.9 mg/mL in the EHT. In the AWM assay; T. capitatus essential oil achieved 70.8% inhibition at 1 mg/mL after 8 h incubation. The in vivo, evaluation on H. polygyrus revealed a significant nematicidal effect 7 days post-treatment by inducing 49.5% FECR and 64.5% TWCR, using the highest dose (1600 mg/kg). The results of present study, demonstrate that T.capitatus EO possess a significant anthelmintic properties. Furthermore, it could be an alternative source of anthelmintic agents against gastrointestinal infections caused by H. contortus.
Assuntos
Anti-Helmínticos , Fezes , Flores , Cromatografia Gasosa-Espectrometria de Massas , Hemoncose , Haemonchus , Nematospiroides dubius , Óleos Voláteis , Contagem de Ovos de Parasitas , Infecções por Strongylida , Thymus (Planta) , Animais , Haemonchus/efeitos dos fármacos , Óleos Voláteis/farmacologia , Óleos Voláteis/química , Óleos Voláteis/isolamento & purificação , Camundongos , Nematospiroides dubius/efeitos dos fármacos , Thymus (Planta)/química , Hemoncose/veterinária , Hemoncose/tratamento farmacológico , Hemoncose/parasitologia , Infecções por Strongylida/tratamento farmacológico , Infecções por Strongylida/veterinária , Infecções por Strongylida/parasitologia , Anti-Helmínticos/farmacologia , Anti-Helmínticos/isolamento & purificação , Anti-Helmínticos/uso terapêutico , Anti-Helmínticos/química , Fezes/parasitologia , Contagem de Ovos de Parasitas/veterinária , Flores/química , Feminino , Ovinos , Concentração Inibidora 50 , Monoterpenos/farmacologia , Monoterpenos/isolamento & purificação , Monoterpenos/química , Masculino , Doenças dos Ovinos/parasitologia , Doenças dos Ovinos/tratamento farmacológico , CimenosRESUMO
A new tetramic acid glycoside, aurantoside L (1), was isolated from the sponge Siliquariaspongia japonica collected at Tsushima Is., Nagasaki Prefecture, Japan. The structure of aurantoside L (1) composed of a tetramic acid bearing a chlorinated polyene system and a trisaccharide part was elucidated using spectral analysis. Aurantoside L (1) showed anti-parasitic activity against L. amazonensis with an IC50 value of 0.74 µM.
Assuntos
Glicosídeos , Leishmania , Poríferos , Poríferos/química , Animais , Glicosídeos/farmacologia , Glicosídeos/química , Glicosídeos/isolamento & purificação , Leishmania/efeitos dos fármacos , Antiprotozoários/farmacologia , Antiprotozoários/química , Antiprotozoários/isolamento & purificação , Pirrolidinonas/farmacologia , Pirrolidinonas/química , Pirrolidinonas/isolamento & purificação , Japão , Concentração Inibidora 50RESUMO
Octocoral of the genus Clavularia is a kind of marine invertebrate possessing abundant cytotoxic secondary metabolites, such as prostanoids and dolabellanes. In our continuous natural product study of C. spp., two previously undescribed prostanoids [clavulone I-15-one (1) and 12-O-deacetylclavulone I (2)] and eleven known analogs (3-13) were identified. The structures of these new compounds were elucidated based on analysis of their 1D and 2D NMR, HRESIMS, and IR data. Additionally, all tested prostanoids (1 and 3-13) showed potent cytotoxic activities against the human oral cancer cell line (Ca9-22). The major compound 3 showed cytotoxic activity against the Ca9-22 cells with the IC50 value of 2.11 ± 0.03 µg/mL, which echoes the cytotoxic effect of the coral extract. In addition, in silico tools were used to predict the possible effects of isolated compounds on human tumor cell lines and nitric oxide production, as well as the pharmacological potentials.
Assuntos
Antozoários , Antineoplásicos , Prostaglandinas , Humanos , Antozoários/química , Animais , Linhagem Celular Tumoral , Prostaglandinas/farmacologia , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Óxido Nítrico/metabolismo , Concentração Inibidora 50 , Organismos Aquáticos , Estrutura MolecularRESUMO
Two new meroterpenoids, aspergienynes O and P (1 and 2), one new natural compound, aspergienyne Q (3), and a new α-pyrone derivative named 3-(4-methoxy-2-oxo-2H-pyran-6-yl)butanoic acid (4) were isolated from the mangrove endophytic fungal strain Aspergillus sp. GXNU-Y85, along with five known compounds (5-9). The absolute configurations of those new isolates were confirmed through extensive analysis using spectroscopic data (HRESIMS, NMR, and ECD). The pharmacological study of the anti-proliferation activity indicated that isolates 5 and 9 displayed moderate inhibitory effects against HeLa and A549 cells, with the IC50 values ranging from 16.6 to 45.4 µM.
Assuntos
Aspergillus , Pironas , Terpenos , Aspergillus/química , Humanos , Pironas/farmacologia , Pironas/química , Pironas/isolamento & purificação , Terpenos/farmacologia , Terpenos/química , Terpenos/isolamento & purificação , Células A549 , Células HeLa , Antineoplásicos/farmacologia , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Estrutura Molecular , Endófitos/química , Concentração Inibidora 50 , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Espectroscopia de Ressonância MagnéticaRESUMO
The development of a versatile platform for the synthesis of 1,2-difunctionalized bicyclo[1.1.1]pentanes to potentially mimic ortho/meta-substituted arenes is described. The syntheses of useful building blocks bearing alcohol, amine, and carboxylic acid functional handles have been achieved from a simple common intermediate. Several ortho- and meta-substituted benzene analogs, as well as simple molecular matched pairs, have also been prepared using this platform. The results of in-depth ADME (absorption, distribution, metabolism, and excretion) investigations of these systems are presented, as well as computational studies which validate the ortho- or meta-character of these bioisosteres.
Assuntos
Hidrocarbonetos Aromáticos/química , Pentanos/química , Bioensaio , Cristalografia por Raios X , Hepatócitos/metabolismo , Humanos , Concentração Inibidora 50 , Pentanos/síntese química , EstereoisomerismoRESUMO
Plasmodium malaria parasites are obligate intracellular protozoans that use a unique form of locomotion, termed gliding motility, to move through host tissues and invade cells. The process is substrate dependent and powered by an actomyosin motor that drives the posterior translocation of extracellular adhesins which, in turn, propel the parasite forward. Gliding motility is essential for tissue translocation in the sporozoite and ookinete stages; however, the short-lived erythrocyte-invading merozoite stage has never been observed to undergo gliding movement. Here we show Plasmodium merozoites possess the ability to undergo gliding motility in vitro and that this mechanism is likely an important precursor step for successful parasite invasion. We demonstrate that two human infective species, Plasmodium falciparum and Plasmodium knowlesi, have distinct merozoite motility profiles which may reflect distinct invasion strategies. Additionally, we develop and validate a higher throughput assay to evaluate the effects of genetic and pharmacological perturbations on both the molecular motor and the complex signaling cascade that regulates motility in merozoites. The discovery of merozoite motility provides a model to study the glideosome and adds a dimension for work aiming to develop treatments targeting the blood stage invasion pathways.
Assuntos
Eritrócitos/parasitologia , Merozoítos/fisiologia , Plasmodium falciparum/genética , Plasmodium/metabolismo , Proteínas de Protozoários/metabolismo , Esporozoítos/fisiologia , Citoesqueleto de Actina/metabolismo , Actomiosina/química , Animais , Eritrócitos/citologia , Células Endoteliais da Veia Umbilical Humana , Humanos , Concentração Inibidora 50 , Locomoção , Proteínas de Membrana/metabolismo , Transdução de SinaisRESUMO
BACKGROUND: Conventional microscopic counting is a widely utilised method for evaluating the trypanocidal effects of drugs on intracellular amastigotes. This is a low-cost approach, but it is time-consuming and reliant on the expertise of the microscopist. So, there is a pressing need for developing technologies to enhance the efficiency of low-cost anti-Trypanosoma cruzi drug screening. OBJECTIVES: In our laboratory, we aimed to expedite the screening of anti-T. cruzi drugs by implementing a fluorescent method that correlates emitted fluorescence from green fluorescent protein (GFP)-expressing T. cruzi (Tc-GFP) with cellular viability. METHODS: Epimastigotes (Y strain) were transfected with the pROCKGFPNeo plasmid, resulting in robust and sustained GFP expression across epimastigotes, trypomastigotes, and intracellular amastigotes. Tc-GFP epimastigotes and intracellular amastigotes were exposed to a serial dilution of benznidazole (Bz). Cell viability was assessed through a combination of microscopic counting, MTT, and fluorimetry. FINDINGS: The fluorescence data indicated an underestimation of the activity of Bz against epimastigotes (IC50 75 µM x 14 µM). Conversely, for intracellular GFP-amastigotes, both fluorimetry and microscopy yielded identical IC50 values. Factors influencing the fluorimetry approach are discussed. MAIN CONCLUSIONS: Our proposed fluorometric assessment is effective and can serve as a viable substitute for the time-consuming microscopic counting of intracellular amastigotes.
Assuntos
Proteínas de Fluorescência Verde , Tripanossomicidas , Trypanosoma cruzi , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/genética , Proteínas de Fluorescência Verde/genética , Tripanossomicidas/farmacologia , Nitroimidazóis/farmacologia , Testes de Sensibilidade Parasitária , Animais , Concentração Inibidora 50 , Avaliação Pré-Clínica de Medicamentos , Sobrevivência Celular/efeitos dos fármacosRESUMO
Toxoplasmosis poses a global health threat, ranging from asymptomatic cases to severe, potentially fatal manifestations, especially in immunocompromised individuals and congenital transmission. Prior research suggests that oregano essential oil (OEO) exhibits diverse biological effects, including antiparasitic activity against Toxoplasma gondii. Given concerns about current treatments, exploring new compounds is important. This study was to assess the toxicity of OEO on BeWo cells and T. gondii tachyzoites, as well as to evaluate its effectiveness in in vitro infection models and determine its direct action on free tachyzoites. OEO toxicity on BeWo cells and T. gondii tachyzoites was assessed by MTT and trypan blue methods, determining cytotoxic concentration (CC50), inhibitory concentration (IC50), and selectivity index (SI). Infection and proliferation indices were analyzed. Direct assessments of the parasite included reactive oxygen species (ROS) levels, mitochondrial membrane potential, necrosis, and apoptosis, as well as electron microscopy. Oregano oil exhibited low cytotoxicity on BeWo cells (CC50: 114.8 µg/mL ± 0.01) and reduced parasite viability (IC50 12.5 ± 0.06 µg/mL), demonstrating 9.18 times greater selectivity for parasites than BeWo cells. OEO treatment significantly decreased intracellular proliferation in infected cells by 84% after 24 h with 50 µg/mL. Mechanistic investigations revealed increased ROS levels, mitochondrial depolarization, and lipid droplet formation, linked to autophagy induction and plasma membrane permeabilization. These alterations, observed through electron microscopy, suggested a necrotic process confirmed by propidium iodide labeling. OEO treatment demonstrated anti-T. gondii action through cellular and metabolic change while maintaining low toxicity to trophoblastic cells.
Assuntos
Autofagia , Óleos Voláteis , Origanum , Espécies Reativas de Oxigênio , Toxoplasma , Óleos Voláteis/farmacologia , Óleos Voláteis/química , Toxoplasma/efeitos dos fármacos , Toxoplasma/crescimento & desenvolvimento , Origanum/química , Humanos , Autofagia/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Linhagem Celular , Antiprotozoários/farmacologia , Concentração Inibidora 50 , Necrose/tratamento farmacológico , Sobrevivência Celular/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Potencial da Membrana Mitocondrial/efeitos dos fármacosRESUMO
In pursuit of potential agents to treat Chagas disease and leishmaniasis, we report the design, synthesis, and identification novel naphthoquinone hydrazide-based molecular hybrids. The compounds were subjected to in vitro trypanocide and leishmanicidal activities. N'-(1,4-Dioxo-1,4-dihydronaphthalen-2-yl)-3,5-dimethoxybenzohydrazide (13) showed the best performance against Trypanosoma cruzi (IC50 1.83 µM) and Leishmania amazonensis (IC50 9.65 µM). 4-Bromo-N'-(1,4-dioxo-1,4-dihydronaphthalen-2-yl)benzohydrazide (16) exhibited leishmanicidal activity (IC50 12.16 µM). Regarding trypanocide activity, compound 13 was low cytotoxic to LLC-MK2 cells (SI = 95.28). Furthermore, through molecular modeling studies, the cysteine proteases cruzain, rhodesain and CPB2.8 were identified as the potential biological targets.
Assuntos
Desenho de Fármacos , Hidrazinas , Leishmania , Naftoquinonas , Tripanossomicidas , Trypanosoma cruzi , Naftoquinonas/farmacologia , Naftoquinonas/química , Naftoquinonas/síntese química , Trypanosoma cruzi/efeitos dos fármacos , Tripanossomicidas/farmacologia , Tripanossomicidas/síntese química , Tripanossomicidas/química , Leishmania/efeitos dos fármacos , Hidrazinas/química , Hidrazinas/farmacologia , Animais , Antiprotozoários/farmacologia , Antiprotozoários/síntese química , Antiprotozoários/química , Testes de Sensibilidade Parasitária , Concentração Inibidora 50 , Relação Estrutura-Atividade , Cisteína EndopeptidasesRESUMO
Leishmaniasis is a disease of public health relevance that demands new therapeutic alternatives due to the toxicity of conventional treatments. In this study, 27 plants of interest to the Unified Health System (SUS) were evaluated for cytotoxicity in macrophages, leishmanicidal activity and production of nitric oxide (NO). None of the species demonstrated cytotoxicity to macrophages (CC50 >100 µg/mL). Extracts from Chenopodium ambrosioides, Equisetum arvense, Maytenus ilicifolia showed greater efficacy in inducing the death of Leishmania amazonensis amastigotes with IC50 of 68.4, 82.3, 75.7 µg/mL, respectively. The species Cynara scolymus, Punica granatum and Passiflora alata were the most effective in inducing an increase in the indirect concentration of NO (41.31, 29.30 and 28.86 µM, respectively) in cultures of macrophages infected with L. amazonensis. Furthermore, Punica granatum was also the most effective species in inducing an increase in NO in macrophages infected by Leishmania chagasi (19.90 µM). The results obtained so far support the continuation of studies, with the possibility of developing safer and more effective treatments for leishmaniasis, using natural products. The identification of plants that stimulate the production of NO in macrophages infected by Leishmania opens doors for more detailed investigations of the mechanism of action of these natural products.