RESUMEN
Glioblastoma multiforme (GBM) is account for 70% of all primary malignancies of the central nervous system. The median survival of human patients after treatment is around 15 months. There are several biological targets which have been reported that can be pursued using ligands with varied structures to treat this disease. In our group, we have developed several ligands that target a wide range of proteins involved in anticancer effects, such as histone deacetylase (HDACs), G protein-coupled estrogen receptor 1 (GPER), estrogen receptor-beta (ERß) and NADPH oxidase (NOX), that were screened on bidimensional (2D) and tridimensional (3D) GBM stem cells like (GSC). Our results show that some HDAC inhibitors show antiproliferative properties at 21-32 µM. These results suggest that in this 3D culture, HDACs could be the most relevant targets that are modulated to induce the antiproliferative effects that require in the future further experimental studies.
Asunto(s)
Neoplasias Encefálicas , Glioblastoma , Neoplasias Encefálicas/tratamiento farmacológico , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Proliferación Celular , Glioblastoma/patología , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas , Humanos , LigandosRESUMEN
BACKGROUND: Some reports have demonstrated the role of the G Protein-coupled Estrogen Receptor (GPER) in growth and proliferation of breast cancer cells. OBJECTIVE: In an effort to develop new therapeutic strategies against breast cancer, we employed an in silico study to explore the binding modes of tetrahydroquinoline 2 and 4 to be compared with the reported ligands G1 and G1PABA. METHODS: This study aimed to design and filter ligands by in silico studies determining their Lipinski's rule, toxicity and binding properties with GPER to achieve experimental assays as anti-proliferative compounds of breast cancer cell lines. RESULTS: In silico studies suggest as promissory two tetrahydroquinoline 2 and 4 which contain a carboxyl group instead of the acetyl group (as is needed for G1 synthesis), which add low (2) and high hindrance (4) chemical moieties to explore the polar, hydrophobic and hindrance effects. Docking and molecular dynamics simulations of the target compounds were performed with GPER to explore their binding mode and free energy values. In addition, the target small molecules were synthesized and assayed in vitro using breast cancer cells (MCF-7 and MDA-MB-231). Experimental assays showed that compound 2 decreased cell proliferation, showing IC50 values of 50µM and 25µM after 72h of treatment of MCF-7 and MDA-MB-231 cell lines, respectively. Importantly, compound 2 showed a similar inhibitory effect on proliferation as G1 compound in MDA-MB-231 cells, suggesting that both ligands reach the GPER-binding site in a similar way, as was demonstrated through in silico studies. CONCLUSION: A concentration-dependent inhibition of cell proliferation occurred with compound 2 in the two cell lines regardless of GPER.
Asunto(s)
Antineoplásicos/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Quinolinas/farmacología , Receptores de Estrógenos/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Antineoplásicos/síntesis química , Antineoplásicos/química , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Modelos Moleculares , Estructura Molecular , Quinolinas/síntesis química , Quinolinas/química , Receptores de Estrógenos/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Relación Estructura-Actividad , Termodinámica , Células Tumorales CultivadasRESUMEN
Since the neuraminidase (NA) enzyme of the influenza A virus plays a key role in the process of release of new viral particles from a host cell, it is often a target for new drug design. The emergence of NA mutations, such as H275Y, has led to great resistance against neuraminidase inhibitors, including oseltamivir and zanamivir. Hence, we herein designed a set of derivatives by modifying the amine and/or carboxylic groups of oseltamivir. After being screened for their physicochemical (Lipinski's rule) and toxicological properties, the remaining compounds were submitted to molecular and theoretical studies. The docking simulations provided insights into NA recognition patterns, demonstrating that oseltamivir modified at the carboxylic moiety and coupled with anilines had higher affinity and a better binding pose for NA than the derivatives modified at the amine group. Based on these theoretical studies, the new oseltamivir derivatives may have higher affinity to mutant variants and possibly to other viral subtypes. Accordingly, two compounds were selected for synthesis, which together with their respective intermediates were evaluated for their cytotoxicity and antiviral activities. Their biological activity was then tested in cells infected with the A/Puerto Rico/916/34 (H1N1) influenza virus, and virus yield reduction assays were performed. Additionally, by measuring neuraminidase activity with the neuraminidase assay kit it was found that the compounds produced inhibitory activity on this enzyme. Finally, the infected cells were analysed with atomic force microscopy (AFM), observing morphological changes strongly suggesting that these compounds interfered with cellular release of viral particles.
Asunto(s)
Antivirales/farmacología , Diseño de Fármacos , Inhibidores Enzimáticos/farmacología , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Neuraminidasa/antagonistas & inhibidores , Oseltamivir/farmacología , Animales , Antivirales/química , Chlorocebus aethiops , Simulación por Computador , Perros , Farmacorresistencia Viral , Células HeLa , Humanos , Técnicas In Vitro , Gripe Humana/tratamiento farmacológico , Gripe Humana/virología , Células de Riñón Canino Madin Darby , Microscopía de Fuerza Atómica , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Infecciones por Orthomyxoviridae/virología , Oseltamivir/química , Células Vero , Proteínas Virales/antagonistas & inhibidoresRESUMEN
BACKGROUND: Annona cherimola Miller (Annonaceae) is a medicinal plant frequently recommended in Mexican traditional medicine for the treatment of gastrointestinal disorders such as diarrhea and dysentery. OBJECTIVE: This work was undertaken to obtain information that support the traditional use of A. cherimola, on pharmacological basis using in vitro and computational experiments. MATERIAL AND METHODS: Bioassay-guided fractionation of the ethanol extract of the leaves of A. cherimola afforded five phenolic compounds: caffeic acid, quercetin, kaempferol, nicotinflorin, and rutin. RESULTS: The in vitro antiprotozoal assay showed that kaempferol was the most potent antiamoebic and antigiardial compound with IC50 values of 7.9 µg/mL for Entamoeba histolytica and 8.7 µg/mL for Giardia lamblia. Computational molecular docking study showed that kaempferol interacted in a region different than metronidazole in the enzyme pyruvate: ferredoxin oxidoreductase (PFOR). CONCLUSION: Considering that PFOR is a target of metronidazole; kaempferol may be a lead compound for the development of novel antiprotozoal agent. Also, these findings give support to the use of A. cherimola in the traditional medicine from México for the treatment of diarrhea and dysentery. SUMMARY: Bioassay-guided fractionation of the ethanol extract of the leaves of Annona cherimola afforded five phenolic compounds: caffeic acid, quercetin, kaempferol, nicotinflorin and rutin. The in vitro antiprotozoal assay showed that kaempferol was the most potent antiamoebic and antigiardial compound with IC50 values of 7.9 µg/mL for Entamoeba histolytica and 8.7 µg/mL for Giardia lamblia. Computational molecular docking study showed that kaempferol interacted in a region different that metronidazole in the enzyme pyruvate: ferredoxin oxidoreductase. Abbreviations used: PFOR:Pyruvate:ferredoxin oxidoreductase, G: lamblia: Giardia lamblia, E: histolytica: Entamoeba histolytica.
RESUMEN
The G-protein coupled receptors (GPCRs) represent the largest superfamily of membrane proteins in charge to pass the cell signaling after binding with their cognate ligands to the cell interior. In breast cancer, a GPCR named GPER1 plays a key role in the process of growth and the proliferation of cancer cells. In a previous study, theoretical methods were applied to construct a model of GPER1, which later was submitted to molecular dynamics (MD) simulations to perform a docking calculation. Based on this preceding work, it is known that GPER1 is sensitive to structural differences in its binding site. However, due to the nature of that past study, conformational changes linked to the ligand binding were not observed. Therefore, in this study, in order to explore the conformational changes coupled to the agonist/antagonist binding, MD simulations of about 0.25µs were performed for the free and bound states, summarizing 0.75µs of MD simulation in total. For the bound states, one agonist (G-1) and antagonist (G-15) were chosen since is widely known that these two molecules cause an impact on GPER1 mobility. Based on the conformational ensemble generated through MD simulations, we found that despite G-1 and G-15 being stabilized by similar map of residues, the structural differences between both ligands impact the hydrogen bond pattern not only at the GPER1 binding site but also along the seven-helix bundle, causing significant differences in the conformational mobility along the extracellular and cytoplasmic domain, and to a lesser degree in the curvatures of helix 2, helix 3 and helix 7 between the free and bound states, which is in agreement with reported literature, and might be linked to microscopic characteristics of the activated-inactivated transition. Furthermore, binding free energy calculations using the MM/GBSA method for the bound states, followed by an alanine scanning analysis allowed us to identify some important residues for the complex stabilization.
Asunto(s)
Receptores de Estrógenos , Receptores Acoplados a Proteínas G , Benzodioxoles/metabolismo , Sitios de Unión , Ciclopentanos/metabolismo , Humanos , Membrana Dobles de Lípidos/metabolismo , Simulación de Dinámica Molecular , Conformación Proteica , Quinolinas/metabolismo , Receptores de Estrógenos/antagonistas & inhibidores , Receptores de Estrógenos/química , Receptores de Estrógenos/metabolismo , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/química , Receptores Acoplados a Proteínas G/metabolismoRESUMEN
We have provided evidence that the stimulatory effects of (-)-epicatechin ((-)-EPI) on endothelial cell nitric oxide (NO) production may involve the participation of a cell-surface receptor. Thus far, such entity(ies) has not been fully elucidated. The G protein-coupled estrogen receptor (GPER) is a cell-surface receptor that has been linked to protective effects on the cardiovascular system and activation of intracellular signaling pathways (including NO production) similar to those reported with (-)-EPI. In bovine coronary artery endothelial cells (BCAEC) by the use of confocal imaging, we evidence the presence of GPER at the cell-surface and on F-actin filaments. Using in silico studies we document the favorable binding mode between (-)-EPI and GPER. Such binding is comparable to that of the GPER agonist, G1. By the use of selective blockers, we demonstrate that the activation of ERK 1/2 and CaMKII by (-)-EPI is dependent on the GPER/c-SRC/EGFR axis mimicking those effects noted with G1. We also evidence by the use of siRNA the role that GPER has on mediating ERK1/2 activation by (-)-EPI. GPER appears to be coupled to a non Gαi/o or Gαs, protein subtype. To extrapolate our findings to an ex vivo model, we employed phenylephrine pre-contracted aortic rings evidencing that (-)-EPI can mediate vasodilation through GPER activation. In conclusion, we provide evidence that suggests the GPER as a potential mediator of (-)-EPI effects and highlights the important role that GPER may have on cardiovascular system protection.