RESUMEN
The signaling of sphingosine kinase 1 (SphK1) and sphingosine-1-phosphate (S1P) regulates various diseases, including multiple sclerosis, atherosclerosis, rheumatoid arthritis, inflammation-related ailments, diabetes, and cancer. SphK1 is considered an attractive potential drug target and is extensively explored in cancer and other inflammatory diseases. In this study, we have investigated the inhibitory potential and binding affinity of SphK1 with cholic acid (CA), syringic acid (SA), and mangiferin (MF) using a combination of docking and molecular dynamics (MD) simulation studies followed by experimental measurements of binding affinity and enzyme inhibition assays. We observed these compounds bind to SphK1 with a significantly high affinity and eventually inhibit its kinase activity with IC50 values of 28.23 µM, 33.35 µM, and 57.2 µM for CA, SA, and MF, respectively. Further, the docking and 100 ns MD simulation studies showed that CA, SA, and MF bind with the active site residues of SphK1 with favorable energy and strong non-covalent interactions that might be accountable for inhibiting its kinase activity. Our finding indicates that CA, SA, and MF may be implicated in designing novel anti-cancer therapeutics with an improved affinity and lesser side effects by targeting SphK1.
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Neoplasias , Humanos , Ácido Cólico , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismoRESUMEN
Bacopa monnieri has been used for centuries in Ayurvedic medicine, alone or in combination with other herbs, as a memory and learning enhancer, sedative, and anti-epileptic. This review aimed to highlight the health benefits of B. monnieri extracts (BME), focusing on anti-cancer and neurodegenerative diseases. We examined the clinical studies on phytochemistry and pharmacological application of BME. We further highlighted the mechanism of action of these extracts in varying types of cancer and their therapeutic implications. In addition, we investigated the underlying molecular mechanism in therapeutic interventions, toxicities, safety concerns and synergistic potential in cognition and neuroprotection. Overall, this review provides deeper insights into the therapeutic implications of Brahmi as a lead formulation for treating neurological disorders and exerting cognitive-enhancing effects.
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Sphingosine kinase 1 (SphK1) and sphingosine-1-phosphate (S1P) signaling regulates numerous diseases such as cancer, diabetes, and inflammation-related ailments, rheumatoid arthritis, atherosclerosis, and multiple sclerosis. The importance of SphK1 in chemo-resistance has been extensively explored in breast, lung, colon, and hepatocellular carcinomas. SphK1 is considered an attractive drug target for the development of anticancer therapy. New drug molecules targeting the S1P signaling are required owing to its pleiotropic nature and association with multiple downstream targets. Here, we have investigated the binding affinity and SphK1 inhibitory potential of cinchonine and colcemid using a combined molecular docking and simulation studies followed by experimental analysis. These compounds bind to SphK1 with a significantly high affinity and subsequently inhibit kinase activity (IC50 7-9 µM). Further, MD simulation studies revealed that both cinchonine and colcemid bind to the residues at the active site pocket of SphK1 with several non-covalent interactions, which may be responsible for inhibiting its kinase activity. Besides, the binding of cinchonine and colcemid causes substantial conformational changes in the structure of SphK1. Taken together, cinchonine and colcemid may be implicated in designing potential drug molecules with improved affinity and specificity for SphK1 targeting anticancer therapy.Communicated by Ramaswamy H. Sarma.
Asunto(s)
Fosfotransferasas (Aceptor de Grupo Alcohol) , Alcaloides de Cinchona , Demecolcina , Simulación del Acoplamiento Molecular , Fosfotransferasas (Aceptor de Grupo Alcohol)/químicaRESUMEN
Sphingosine kinase 1 (SphK1) has emerged as an attractive drug target for different diseases. Recently, discovered SphK1 inhibitors have been recommended in cancer therapeutics; however, selectivity and potency are great challenges. In this study, a novel series of benzimidazoles was synthesized and evaluated as SphK1 inhibitors. Our design strategy is twofold: It aimed first to study the effect of replacing the 5-position of the benzimidazole ring with a polar carboxylic acid group on the SphK1-inhibitory activity and cytotoxicity. Our second aim was to optimize the structures of the benzimidazoles through the elongation of the chain. The enzyme inhibition potentials against all the synthesized compounds toward SphK1 were evaluated, and the results revealed that most of the studied compounds inhibited SphK1 effectively. The binding affinity of the benzimidazole derivatives toward SphK1 was measured by fluorescence binding and molecular docking. Compounds 33, 37, 39, 41, 42, 43, and 45 showed an appreciable binding affinity. Therefore, the SphK1-inhibitory potentials of compounds 33, 37, 39, 41, 42, 43, and 45 were studied and IC50 values were determined, to reveal high potency. The study showed that these compounds inhibited SphK1 with effective IC50 values. Among the studied compounds, compound 41 was the most effective one with the lowest IC50 value and a high cytotoxicity on a wide spectrum of cell lines. Molecular docking revealed that most of these compounds fit well into the ATP-binding site of SphK1 and form hydrogen bond interactions with catalytically important residues. Overall, the findings suggest the therapeutic potential of benzimidazoles in the clinical management of SphK1-associated diseases.
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Antineoplásicos/farmacología , Bencimidazoles/farmacología , Neoplasias/tratamiento farmacológico , Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Adenosina Trifosfato/metabolismo , Antineoplásicos/síntesis química , Antineoplásicos/química , Bencimidazoles/síntesis química , Bencimidazoles/química , Sitios de Unión , Línea Celular Tumoral , Humanos , Simulación del Acoplamiento Molecular , Neoplasias/enzimología , Neoplasias/patología , Relación Estructura-ActividadRESUMEN
The sphingosine kinase-1/sphingosine-1-phosphate pathway is linked with the cancer progression and survival of the chemotherapy-challenged cells. Sphingosine kinase-1 (SphK1) has emerged as an attractive drug target, but their inhibitors from natural sources are limited. In this study, we have chosen harmaline, one of the ß-carboline alkaloids, and report its mechanism of binding to SphK1 and subsequent inhibition. Molecular docking combined with fluorescence binding studies revealed that harmaline binds to the substrate-binding pocket of SphK1 with an appreciable binding affinity and significantly inhibits the kinase activity of SphK1 with an IC50 value in the micromolar range. The cytotoxic effect of harmaline on non-small-cell lung cancer cells by MTT assay was found to be higher for H1299 compared to A549. Harmaline induces apoptosis in non-small-cell lung carcinoma cells (H1299 and A549), possibly via the intrinsic pathway. Our findings suggest that harmaline could be implicated as a scaffold for designing potent anticancer molecules with SphK1 inhibitory potential.
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Sphingosine kinase 1 (SphK1) plays a significant role in various cellular processes, including cell proliferation, apoptosis, and angiogenesis. SphK1 is considered as an attractive target for drug development owing to its connection with several diseases, including cancer. In the current work, the urea-induced unfolding of SphK1 was performed at pH 8.0 and 25 °C using CD and fluorescence spectroscopy. SphK1 follows a biphasic unfolding transition (N â I â D) with an intermediate (I) state populated around 4.0 M urea concentration. The circular dichroism ([θ]222) and fluorescence emission spectra (λmax) of SphK1 with increasing concentrations of urea were analyzed to calculate Gibbs free energy (ΔG0) for both the transitions (N â I and I â D). A significant overlap of both the transitions obtained by two spectroscopic properties ([θ]222 and λmax) was observed, indicating that both N â I and I â D transition follow two-step equilibrium unfolding pattern. Also, we performed 100 ns molecular dynamics (MD) simulations to get atomistic insights into the structural changes in SphK1 with increasing urea concentrations. Our results showed a consistent pattern of the SphK1 unfolding with increasing urea concentrations. Together, spectroscopic and MD simulation findings provide deep insights into the unfolding mechanism and conformational features of SphK1.
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Fosfotransferasas (Aceptor de Grupo Alcohol)/química , Desnaturalización Proteica , Urea/química , Algoritmos , Humanos , Enlace de Hidrógeno , Modelos Moleculares , Conformación Proteica , Desnaturalización Proteica/efectos de los fármacos , Desplegamiento Proteico , Solventes , Análisis Espectral , Relación Estructura-Actividad , Urea/farmacologíaRESUMEN
Sphingosine kinase 1 (SphK1) is one of the well-studied drug targets for cancer and inflammatory diseases. Recently discovered small-molecule inhibitors of SphK1 have been recommended in cancer therapeutics; however, selectivity and potency of first-generation inhibitors are great challenge. In search of effective SphK1 inhibitors, a set of small molecules have been designed and synthesized bearing urea, sulfonylurea, sulfonamide, and sulfonyltriurea groups. The binding affinity of these inhibitors was measured by fluorescence-binding assay and isothermal titration calorimetry. Compounds 1, 5, 6, and 7 showed an admirable binding affinity to the SphK1 in the sub-micromolar range and significantly inhibited SphK1 activity with admirable IC50 values. Molecular docking studies revealed that these compounds fit well into the sphingosine binding pocket of SphK1 and formed significant number of hydrogen bonds and van der Waals interactions. These molecules may be exploited as potent and selective inhibitors of SphK1 that could be implicated in cancer therapeutics after the required in vivo validation.
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Sphingosine kinase 1 (SphK1) catalyzes the conversion of sphingosine to sphingosine-1-phosphate that acts as a bioactive signalling molecule, and regulates various cellular processes including lymphocyte trafficking, angiogenesis and response to apoptotic stimuli. Abnormal expression of SphK1 has been observed in a wide range of cancers highlighting their role in tumour growth and metastasis. This enzyme also plays a critical role in metabolic and inflammatory diseases, including pulmonary fibrosis, diabetic neuropathy and Alzheimer's disease. In the present study, we have investigated the structural and conformational changes in SphK1 at varying pH using various spectroscopic techniques. Consistent results were observed with the function of SphK1 at corresponding pH values. SphK1 maintains its secondary and tertiary structure in the pH range of 7.5-10.0. However, protein aggregation was observed in the acidic pH range (4.0-6.5). At pHâ¯2.0, the SphK1 exists in the molten-globule state. Kinase assay also shows that SphK1 activity was optimal in the pH range of 7.5-8.5. To complement in vitro results, we have performed 100â¯ns molecular dynamics simulation to examine the effect of pH on the structural stability of SphK1 at molecular level. SphK1 maintains its native conformation in the alkaline pH range with some residual fluctuations detected at acidic pH. A considerable correlation was noticed between spectroscopic, enzymatic activity and MD simulation studies. pH dependent structural changes can be further implicated to understand its association with disease condition, and cellular homeostasis with respect to protein function under variable pH conditions.
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Fosfotransferasas (Aceptor de Grupo Alcohol)/química , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Dicroismo Circular , Estabilidad de Enzimas , Humanos , Enlace de Hidrógeno , Concentración de Iones de Hidrógeno , Simulación de Dinámica Molecular , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Análisis de Componente Principal , Agregado de Proteínas , Conformación Proteica , Estructura Secundaria de Proteína , Estructura Terciaria de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Espectrometría de Fluorescencia , Espectrofotometría , Esfingolípidos/metabolismoRESUMEN
Sphingosine kinase 1 (SphK1) has recently gained attention as a potential drug target for its association with cancer and other inflammatory diseases. Here, we have investigated the binding affinity of dietary phytochemicals viz., ursolic acid, capsaicin, DL-α tocopherol acetate, quercetin, vanillin, citral, limonin and simvastatin with the SphK1. Docking studies revealed that all these compounds bind to the SphK1 with varying affinities. Fluorescence binding and isothermal titration calorimetric measurements suggested that quercetin and capsaicin bind to SphK1 with an excellent affinity, and significantly inhibits its activity with an admirable IC50 values. The binding mechanism of quercetin was assessed by docking and molecular dynamics simulation studies for 100 ns in detail. We found that quercetin acts as a lipid substrate competitive inhibitor, and it interacts with important residues of active-site pocket through hydrogen bonds and other non-covalent interactions. Quercetin forms a stable complex with SphK1 without inducing any significant conformational changes in the protein structure. In conclusion, we infer that quercetin and capsaicin provide a chemical scaffold to develop potent and selective inhibitors of SphK1 after required modifications for the clinical management of cancer.
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Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/fisiología , Fitoquímicos/metabolismo , Humanos , Lisofosfolípidos/metabolismo , Simulación de Dinámica Molecular , Neoplasias , Fitoquímicos/uso terapéutico , Unión Proteica/fisiología , Quercetina/farmacología , Esfingosina/metabolismoRESUMEN
Pyruvate dehydrogenase kinase 3 (PDK3) is a mitochondrial protein, has recently been considered as a potential pharmacological target for varying types of cancer. Here, we report the binding mechanism of quercetin to the PDK3 by using molecular docking, simulation, fluorescence spectroscopy and isothermal titration calorimetric assays. Molecular docking along with simulation provided an in-depth analysis of protein-ligand interactions. We have observed that quercetin interacts to the important residues of active site cavity of PDK3 and shows a well-ordered conformational fitting. The stability of quercetin-PDK3 complex is maintained by several non-covalent interactions throughout the simulation. To complement in silico findings with the experiments, we have successfully expressed and purified human PDK3. Both fluorescence and isothermal titration calorimetric experiments showed excellent binding affinity of quercetin to the PDK3. Kinase inhibition assay further revealed a significant inhibitory potential of quercetin to the PDK3 with the IC50 values in µM range. Quercetin is non-toxic to HEK293, and significantly inhibits the proliferation of cancer (HepG2 and A549) cell lines. All these observations clearly indicate that quercetin may be further evaluated as promising therapeutic molecule for PDK3 with required modifications and in vivo validation.
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Antineoplásicos/farmacología , Inhibidores Enzimáticos/farmacología , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/antagonistas & inhibidores , Quercetina/farmacología , Antineoplásicos/metabolismo , Proliferación Celular/efectos de los fármacos , Inhibidores Enzimáticos/metabolismo , Células HEK293 , Células Hep G2 , Humanos , Concentración 50 Inhibidora , Simulación del Acoplamiento Molecular , Estructura Secundaria de Proteína/efectos de los fármacos , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/química , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/metabolismo , Quercetina/metabolismoRESUMEN
Selective carbonic anhydrase (CA) inhibitors have gained a lot of importance owing to the implication of specific isoforms of CA in certain diseases like glaucoma, leukemia, cystic fibrosis, and epilepsy. A novel class of sulfonylurea derivatives was synthesized from corresponding sulfonyl chlorides and amines. Compounds with different pendant moieties in the sulfonylurea derivatives show significant interactions with human carbonic anhydrase II (CAII). In vitro evaluation of the sulfonylurea derivatives revealed that three compounds possess admirable inhibitory activity against CAII. Compounds containing methyl (G2), isopropyl (G4) and o-tosyl (G5) groups displayed IC50 (109-137⯵m) for CAII. Fluorescence binding and cytotoxicity studies revealed that these compounds are showing good binding affinity (18-34⯵M) to CAII and non- toxic to human cells. Further, molecular docking studies of G2, G4 and G5 with CAII showed that these compounds fit nicely in the active site of CAII. Molecular dynamics simulation studies of these compounds complexed with CAII showed that essential interactions were maintained up to 50â¯ns of simulation. These results indicate the promising nature of the sulfonylurea scaffold towards CAII inhibition and opens scope of hit to-lead optimization for discovery of effective drugs against CAII-associated disorders.