Novel insight on IRE1 in the regulation of chondrocyte dedifferentiation through ER stress independent pathway.

Inositol-requiring enzyme-1 (IRE1) is the master regulator of the unfolded protein response pathway, associated with the endoplasmic reticulum (ER) in sensing and regulating cell stress. The activity of IRE1 is highly explored and well-characterized in cancer and other cells. However, the IRE1 molecular mechanism in chondrocytes is poorly understood. The present study explored the effect of IRE1 on chondrocytes regarding its chondrogenic gene expression and its correlation with different cellular pathways and cell behavior. Chondrocytes transfected with the cDNA of IRE1 reduced the expression of type II collagen, disrupting chondrocyte differentiation as confirmed by western blotting and immunofluorescence. Upon siRNA treatment, the influence of IRE1 on chondrocyte differentiation is restored by reviving the normal expression of type II collagen. Different molecular pathways were explored to investigate the role of IRE1 in causing chondrocyte dedifferentiation. However, we found no significant correlation, as IRE1 induces dedifferentiation through independent pathways. In response to various endoplasmic reticulum (ER) agonists (2-deoxy-D-glucose), and ER stress antagonists (tauroursodeoxycholic acid and salubrinal), IRE1 overexpression did not affect GRP78/94, as implicated in the pathogenesis of ER stress. Moreover, when IRE1 overexpression was correlated with the inflammation pathway, nuclear factor-kappa B (NFκB), IRE1 substantially increased the expression of p50 while decreasing the expression of nuclear factor kappa light polypeptide alpha (IκBα). These results suggest that IRE1 induces dedifferentiation in chondrocytes by modulating inflammatory pathways that cause dedifferentiation by disrupting type II collagen expression.

Recent Development on Sensing Strategies for Small Molecules Detections.

Sensors play a critical role in the detection and monitoring of various substances present in our environment, providing us with valuable information about the world around us. Within the field of sensor development, one area that holds particular importance is the detection of small molecules. Small molecules encompass a wide range of organic or inorganic compounds with low molecular weight, typically below 900 Daltons including gases, volatile organic compounds, solvents, pesticides, drugs, biomarkers, toxins, and pollutants. The accurate and efficient detection of these small molecules has attracted significant interest from the scientific community due to its relevance in diverse fields such as environmental pollutants monitoring, medical diagnostics, industrial optimization, healthcare remedies, food safety, ecosystems, and aquatic and terrestrial life preservation. To meet the demand for precise and efficient monitoring of small molecules, this summary aims to provide an overview of recent advancements in sensing and quantification strategies for various organic small molecules including Hydrazine, Glucose, Morpholine, Ethanol amine, Nitrosamine, Oxygen, Nitro-aromatics, Phospholipids, Carbohydrates, Antibiotics, Pesticides, Drugs, Adenosine Triphosphate, Aromatic Amine, Glutathione, Hydrogen Peroxide, Acetone, Methyl Parathion, and Thiophenol. The focus is on understanding the receptor sensing mechanism, along with the electrical, optical, and electrochemical response. Additionally, the variations in UV-visible spectral properties of the ligands upon treatment with the receptor, fluorescence and absorption titration analysis for limit of detection (LOD) determination, and bioimaging analysis are discussed wherever applicable. It is anticipated that the information gathered from this literature survey will be helpful for the perusal of innovation regarding sensing strategies.

Design and synthesis of thiadiazole-oxadiazole-acetamide derivatives: Elastase inhibition, cytotoxicity, kinetic mechanism, and computational studies.

Considering the biological significance of 1,3,4-thiadiazole/oxadiazole heterocyclic scaffolds, a novel series of 1,3,4-thiadiazole-1,3,4-oxadiazole-acetamide derivatives (7a-j) was designed and synthesized using molecular hybridization. The inhibitory effects of the target compounds on elastase were evaluated, and all of these molecules were found to be potent inhibitors compared to the standard reference oleanolic acid. Compound 7f exhibited the excellent inhibitory activity (IC50 = 0.06 ± 0.02 µM), which is 214-fold more active than oleanolic acid (IC50 = 12.84 ± 0.45 µM). Kinetic analysis was also performed on the most potent compound (7f) to determine the mode of binding with the target enzyme, and it was discovered that 7f inhibits the enzyme in a competitive manner. Furthermore, the MTT assay method was used to assess their toxicity on the viability of B16F10 melanoma cell lines, and all compounds did not display any toxic effect on the cells even at high concentrations. The molecular docking studies of all compounds also justified with their good docking score and among them, compound 7f had a good conformational state with hydrogen bond interactions within the receptor binding pocket, which is consistent with the experimental inhibition studies.

E3 Ubiquitin Ligase Constitutive Photomorphogenic 1 Regulates Differentiation and Inflammation via MAPK Signaling Pathway in Rabbit Articular Chondrocytes.

Constitutive photomorphogenic 1 (COP1), is an E3 ubiquitin ligase that plays a role in the regulation of various cellular processes including cell growth, differentiation, and survival in mammals. In certain conditions such as overexpression or loss of function, COP1 acts either as an oncogenic protein or as a tumor suppressor by targeting specific proteins for ubiquitination-mediated degradation. However, the precise role of COP1 has not been well studied in primary articular chondrocytes. In this study, we investigated the role of COP1 in chondrocyte differentiation. Western blotting and reverse transcription-polymerase chain reaction analysis demonstrated that COP1 overexpression reduced type II collagen expression, promoted cyclooxygenase 2 (COX-2) expression, and reduced sulfated proteoglycan synthesis, as detected by Alcian blue staining. Upon siRNA treatment, revived type II collagen, sulfated proteoglycan production, and decreased COX-2 expression. Phosphorylation of p38 kinase and ERK-1/-2 signaling pathways was regulated by COP1 upon cDNA and siRNA transfection in chondrocytes. The inhibition of the p38 kinase and ERK-1/-2 signaling pathways with SB203580 and PD98059 ameliorated the expression of type II collagen and COX-2 in transfected chondrocytes, thus suggesting that COP1 regulates differentiation and inflammation in rabbit articular chondrocytes via the p38 kinase and ERK-1/-2 signaling pathway.

Synthesis and discovery of potential tyrosinase inhibitor of new coumarin-based thiophenyl-pyrazolylthiazole nuclei: In vitro evaluation, cytotoxicity, kinetic, and computational studies.

A well-known key enzyme in melanogenesis and hyperpigmentation is tyrosinase. The present study introduces a novel series of thiophenyl-pyrazolylthiazole-coumarin hybrids (6a-6h) as tyrosinase inhibitors. The in-vitro tyrosinase inhibition results indicated that all compounds have strong tyrosinase inhibitory activity, particularly compound 6g (IC50  = 0.043 ± 0.006 µM), was identified as the most active compound compared to the positive control (kojic acid, IC50  = 18.521 ± 1.162 µM). Lineweaver-Burk plots were employed to analyze the kinetic mechanism, and compound 6g formed an enzyme-inhibitor complex by inhibiting tyrosinase non-competitively. Furthermore, all compounds demonstrated excellent antioxidant activity against DPPH. MTT assay was used to screen the cytotoxicity of all compounds on B16F10 melanoma cells, and they had no toxic effect on the cells. The binding affinity of compounds with tyrosinase was also investigated using molecular docking, and the ligands displayed good binding energy values. These molecules could be a promising lead for skin pigmentation and associated diseases as nontoxic pharmacological scaffolds.

Synthesis, kinetic studies and in-silico investigations of novel quinolinyl-iminothiazolines as alkaline phosphatase inhibitors.

Deposition of hydroxyapatite (HA) or alkaline phosphate crystals on soft tissues causes the pathological calcification diseases comprising of end-stage osteoarthritis (OA), ankylosing spondylitis (AS), medial artery calcification and tumour calcification. The pathological calcification is symbolised by increased concentration of tissue non-specific alkaline phosphatase (TNAP). An efficient therapeutic strategy to eradicate these diseases is required, and for this the alkaline phosphatase inhibitors can play a potential role. In this context a series of novel quinolinyl iminothiazolines was synthesised and evaluated for alkaline phosphatase inhibition potential. All the compounds were subjected to DFT studies where N-benzamide quinolinyl iminothiazoline (6g), N-dichlorobenzamide quinolinyl iminothiazoline (6i) and N-nitrobenzamide quinolinyl iminothiazoline (6j) were found as the most reactive compounds. Then during the in-vitro testing, the compound N-benzamide quinolinyl iminothiazoline (6g) exhibited the maximum alkaline phosphatase inhibitory effect (IC50 = 0.337 ± 0.015 µM) as compared to other analogues and standard KH2PO4 (IC50 = 5.245 ± 0.477 µM). The results were supported by the molecular docking studies, molecular dynamics simulations and kinetic analysis which also revealed the inhibitory potential of compound N-benzamide quinolinyl iminothiazoline (6g) against alkaline phosphatase. This compound can be act as lead molecule for the synthesis of more effective inhibitors and can be suggested to test at the molecular level.

Synthesis, carbonic anhydrase inhibition, anticancer activity, and molecular docking studies of 1,3,4-oxadiazole derivatives.

In this work, we have synthesized various organic compounds possessing 1,3,4-oxadiazole as a core structure and the structure of the newly synthesized target compounds has been revealed using different analytical approaches such as FT-IR, LCMS, and NMR (proton and carbon), respectively. The in vitro carbonic anhydrase potentials of these synthesized 17 different analogues were investigated. The result suggests that compound 7g, a 3-pyridine substituted analogue with an IC50 of 0.1 µM, was found to have the most potent carbonic inhibitory activity (11-fold more active) than the positive control (acetazolamide) with an IC50 of 1.1 ± 0.1 µM. Besides, among the series 7(a-q) approved in the identification of four potent carbonic anhydrase inhibitors with the IC50 standards varies from 0.1 to 1.0 ± 0.1 µM. Additionally, the non-competitive behaviour for potent compound 7g was analysed using the Lineweaver-Burk plot from the kinetic study. Furthermore, the anticancer activity of all the synthesized compounds screened against B16F10 melanoma cells using the MTT assay method. Additionally, the molecular docking studies revealed that 7g inhibitor shows good binding energy as well as good binding interaction pattern along with enzyme.

Synthesis, anticancer evaluation, and molecular docking studies of thiazolyl-pyrazoline derivatives.

The molecular hybridization of thiazole and pyrazoline heterocyclic structures with diverse activities appears to be an interesting strategy for developing new anticancer compounds. This study presents the synthesis of eleven new thiazolyl-pyrazoline derivatives (7a-k) and the evaluation of their in-vitro anti-proliferative activities against human lung carcinoma (A549) and human melanoma cancer (A375) cell lines through MTT assay. In comparison to the positive reference drug erlotinib (IC50 = 34.16 µM in A549 and IC50 = 25.85 µM in A375), four compounds (7e, 7h, 7j, and 7k) were identified as the most active against both cell lines (especially compound 7k with IC50 = 20.28 µM in A549 and 16.08 µM in A375). Additionally, these potent compounds were selected to be investigated for their anti-metastasis and anti-inflammatory properties via inhibition of the expression of matrix metalloproteinase 2, 9 (MMP-2, 9) and cyclooxygenase 2 (COX-2). In A549 cells, upon exposure to compounds 7e and 7j, COX-2 expression is decreased, whereas compounds 7e, 7j, and 7k reduced COX-2 expression in A375 cell lines. Molecular docking studies were carried out to show the possible interactions of synthesized compounds with the predicted active site of the COX-2 protein. The results revealed that compounds 7e and 7j can bind well to the active site of COX-2 protein. Collectively, compounds 7e, 7j, and 7k are all promising candidates for further research towards the development of novel anticancer agents.

Potent Alkaline Phosphatase Inhibitors, Pyrazolo-Oxothiazolidines: Synthesis, Biological Evaluation, Molecular Docking, and Kinetic Studies.

To develop new alkaline phosphatase inhibitors (ALP), a series of pyrazolo-oxothiazolidine derivatives were synthesized and biologically assessed, and the results showed that all of the synthesized compounds significantly inhibited ALP. Specifically, compound 7g displayed the strongest inhibitory activity (IC50 = 0.045 ± 0.004 µM), which is 116-fold more active than monopotassium phosphate (IC50 = 5.242 ± 0.472 µM) as a standard reference. The most potent compound among the series (7g) was checked for its mode of binding with the enzyme and shown as non-competitively binding with the target enzyme. The antioxidant activity of these compounds was examined to investigate the radical scavenging effect. Moreover, the MTT assay method was performed to evaluate their toxic effects on the viability of MG-63 human osteosarcoma cells, and all compounds have no toxic effect on the cells at 4 µM. Computational research was also conducted to examine the binding affinity of the ligands with alkaline phosphatase, and the results revealed that all compounds showed good binding energy values within the active site of the target. Therefore, these novel pyrazolo-oxothiazolidine derivatives might be employed as promising pharmacophores for potent and selective alkaline phosphatase inhibitors.

Design, Synthesis, Kinetic Analysis and Pharmacophore-Directed Discovery of 3-Ethylaniline Hybrid Imino-Thiazolidinone as Potential Inhibitor of Carbonic Anhydrase II: An Emerging Biological Target for Treatment of Cancer.

Carbonic anhydrases (CA), having Zn2+ metal atoms, are responsible for the catalysis of CO2 and water to bicarbonate and protons. Any abnormality in the functioning of these enzymes may lead to morbidities such as glaucoma and different types of cancers including brain, renal and pancreatic carcinomas. To cope with the lack of presence of a promising therapeutic agent against these cancers, searching for an efficient and suitable carbonic anhydrase inhibitor is crucial. In the current study, ten novel 3-ethylaniline hybrid imino-thiazolidinones were synthesized and characterized by FTIR, NMR (1H, 13C), and mass spectrometry. Synthesis was carried out by diethyl but-2-ynedioate cyclization and different acyl thiourea substitutions of 3-ethyl amine. The CA (II) enzyme inhibition profile for all synthesized derivatives was determined. It was observed that compound 6e demonstrated highest inhibition of CA-II with an IC50 value of 1.545 ± 0.016 µM. In order to explore the pharmacophoric properties and develop structure activity relationship, in silico screening was performed. In silico investigations included density functional theory (DFT) studies, pharmacophore-guided model development, molecular docking, molecular dynamic (MD) simulations, and prediction of drug likeness scores. DFT investigations provided insight into the electronic characteristics of compounds, while molecular docking determined the binding orientation of derivatives within the CA-II active site. Compounds 6a, 6e, and 6g had a reactive profile and generated stable protein-ligand interactions with respective docking scores of -6.12, -6.99, and -6.76 kcal/mol. MD simulations were used to evaluate the stability of the top-ranked complex. In addition, pharmacophore-guided modeling demonstrated that compound 6e produced the best pharmacophore model (HHAAARR) compared to standard brinzolamide. In vitro and in silico investigations anticipated that compound 6e would be an inhibitor of carbonic anhydrase II with high efficacy. Compound 6e may serve as a potential lead for future synthesis that can be investigated at the molecular level, and additional in vivo studies are strongly encouraged.

The Inhibitory Effect of Polyphenon 60 from Green Tea on Melanin and Tyrosinase in Zebrafish and A375 Human Melanoma Cells.

Polyphenon 60 (PP60) from green tea has long been used as an antioxidant, anticancer, antimicrobial, and antimutagenic. Aim of the Study. To investigate tyrosinase inhibition-related kinetic mechanism and antimelanogenesis potential of PP60. Materials and Methods. The effect of PP60 on melanin and tyrosinase was evaluated in A375 melanoma cells and zebrafish embryos. The melanoma cells were treated with 20, 40, and 60 µg/mL of PP60, and tyrosinase expression was induced by using L-DOPA. The western blot method was used for the evaluation of tyrosinase expression. Cell lysates were prepared from treated and untreated cells for cellular tyrosinase and melanin quantification. Furthermore, zebrafish embryos were treated with 20, 40, and 60 µg/mL of PP60 and reference drug kojic acid for determination of depigmentation and melanin quantification. In vitro assays were also performed to examine the impact of PP60 on mushroom tyrosinase activity. To determine cytotoxicity, MTT was used against melanoma cell line A375. Results. PP60 showed good tyrosinase inhibitory activity with an IC50 value of 0.697 ± 0.021 µg/mL as compared to kojic acid a reference drug with an IC50 value of 2.486 ± 0.085 µg/mL. Kinetic analysis revealed its mixed type of inhibition against mushroom tyrosinase. In addition, western blot analysis showed that at 60 µg/mL dose of PP60 significantly reduced L-DOPA-induced tyrosinase expression in melanoma cells. PP60 significantly inhibits the cellular tyrosinase (p < 0.05) and reduces the melanin (p < 0.05) contents of melanoma cells. Furthermore, PP60 was found to be very potent in significantly reducing the zebrafish embryos' pigmentation (p < 0.05) and melanin (p < 0.05) content at the dose of 60 µg/mL. Conclusions. Our results demonstrate that PP60 has a strong potency to reduce pigmentation. It may be useful for the cosmetic industries to develop skin whitening agents with minimal toxic effects.

Inhibitors of EYA3 Protein in Ewing Sarcoma.

OBJECTIVE: Among sarcomas, Ewing sarcoma (EWS) is characterized as a highly malignant type of bone tumor caused by the fusion of EWS RNA Binding Protein-1 (EWSR1)/ Friend leukemia integration 1 (FLI1) genes. The product of fusion gene gives rise to EWSR1/FLI1 which activates the activity of Eyes absent homolog 3 (EYA3) which causes tumor growth and angiogenesis. EYA3 is now considered as a therapeutic drug target for EWS . The study was designed to gather potential inhibitors for the EYA3 target using medicinal compounds. METHODS: In this study, we have obtained a list of medicinal compounds from the NuBBE database and downloaded their structural information. Then insilico screening analysis of >2,000 medicinal compounds was performed with PyRX virtual drug screening software to discover potential inhibitors for the treatment of EWS. RESULTS: Our investigation revealed that Sorbifolin and 1,7-Dihydroxy-3-methylanthracene-9.10-dione show interactive affinity for EYA3 active residues. Moreover, these compounds have adequate toxicity, can induce cytotoxicity in EWS cells, and are capable of regulating the expression of genes activated by EWSR1/FLI1. CONCLUSION: Our study concluded that Sorbifolin and 1,7-Dihydroxy-3-methylanthracene-9.10-dione are promising drug candidates for the treatment of EWS and should be further subjected to invitro testing.

Identification of Novel Natural Product Inhibitors against Matrix Metalloproteinase 9 Using Quantum Mechanical Fragment Molecular Orbital-Based Virtual Screening Methods.

Matrix metalloproteinases (MMPs) are calcium-dependent zinc-containing endopeptidases involved in multiple cellular processes. Among the MMP isoforms, MMP-9 regulates cancer invasion, rheumatoid arthritis, and osteoarthritis by degrading extracellular matrix proteins present in the tumor microenvironment and cartilage and promoting angiogenesis. Here, we identified two potent natural product inhibitors of the non-catalytic hemopexin domain of MMP-9 using a novel quantum mechanical fragment molecular orbital (FMO)-based virtual screening workflow. The workflow integrates qualitative pharmacophore modeling, quantitative binding affinity prediction, and a raw material search of natural product inhibitors with the BMDMS-NP library. In binding affinity prediction, we made a scoring function with the FMO method and applied the function to two protein targets (acetylcholinesterase and fibroblast growth factor 1 receptor) from DUD-E benchmark sets. In the two targets, the FMO method outperformed the Glide docking score and MM/PBSA methods. By applying this workflow to MMP-9, we proposed two potent natural product inhibitors (laetanine 9 and genkwanin 10) that interact with hotspot residues of the hemopexin domain of MMP-9. Laetanine 9 and genkwanin 10 bind to MMP-9 with a dissociation constant (KD) of 21.6 and 0.614 µM, respectively. Overall, we present laetanine 9 and genkwanin 10 for MMP-9 and demonstrate that the novel FMO-based workflow with a quantum mechanical approach is promising to discover potent natural product inhibitors of MMP-9, satisfying the pharmacophore model and good binding affinity.

Sulforaphane induces cell differentiation, melanogenesis and also inhibit the proliferation of melanoma cells.

Sulforaphane (SFN) is an organosulfur compound extracted from cruciferous vegetables and has biological effects. The effect of SFN has been studied in different types of cancers, as this compound incites various cytotoxic mechanisms to stunt cancer proliferation. However, the role of SFN activity in melanoma is yet to be known. The current study has been devised to elucidate the effects induced by SFN treatment in the B16F10 melanoma cell line and zebrafish model. Cells were treated with SFN reduced cell proliferation and increased tyrosinase production. Moreover, microscopic and immunofluorescence analysis confirmed the elongated appearance of melanoma cells due to cytoskeletal reorganization induced by SFN. Western blotting showed that SFN regulates the protein expression of Microphthalmia-associated transcription factor (MITF), Protein kinase C beta 1 (PKCß1), and tyrosinase. The relationship between melanin biosynthesis and changes in the actin cytoskeleton encouraged by SFN on melanoma was determined by treating it with Cytochalasin D (CD) and Jasplakinolide (JAS). Co-treatment of SFN with CD increased more tyrosinase expression than SFN alone whereas with JAS, slightly reduced the expression. Immature zebrafish were pretreated with phenylthiourea (PTU) and then exposed to different SFN concentrations yielded the same results by upregulating the melanin levels despite the presence of melanin inhibitor (PTU). These study results show that SFN induces the biosynthesis of melanin in the B16F10 melanoma cell line, which occurs through changes in actin.

Identification of medicinal compounds as potential inhibitors for mutated isocitrate dehydrogenases against chondrosarcoma.

Chondrosarcoma is the third most common cartilaginous bone tumour that is insusceptible to radio- and chemotherapy and it is inclined to metastasis. These resistant qualities are facilitated by mutant variants of isocitrate dehydrogenases (IDH) 1-2 enzyme. These mutant enzymes promote oncogenesis of chondrocytes by changing their epigenetic wardrobe leading to tumour formation. Presently, there are lack of drugs available to be exploited as a remedy for this disease. On the other hand, majority of chemotherapeutic drugs induce cytotoxicity in the cancer cells at the cost of harming surrounding healthy cells, jeopardizing human life. The current study is focused on screening various medicinal compounds against IDH1 and IDH2 combined with insilico gene expression, cancer cells cytotoxicity and ADMET (absorption, distribution, metabolism, excretion and toxicity) studies to elucidate the molecular mechanism against chondrosarcoma and also to uncover pharmacokinetic profile of these compounds. Screening of 5000+ compounds filtered two efficacious compounds (Artocarpetin and 5-Galloylquinic acid) capable of establishing hydrogen bond connections with both IDH variants. Other studies showed that these compounds downregulate ITGAV, CARPIN1, CCL5 and COG5 and TNFRSF10B gene that reduces chondrogenesis and inflammation, Artocarpetin and 5-galloylquinic acid are TP53 expression enhancer and inhibit MM9 expression that promote immunomodulation and apoptosis in these cancers. These compounds are both active against CHSA8926 and CHSA011 cell line of chondrosarcoma. However, the ADME profile of 5-galloylquinic acid is slightly unsatisfactory based on druglikness and bioavailability score criteria as compared to artocarpetin. Both of these compounds are class-5 chemicals and require high doses to elicit adverse response. Our results suggest that artocarpetin and 5-galloylquinic acid are efficacious drug candidates and could be further exploited to validate these findings in vitro.

Novel 1,3,4-oxadiazole compounds inhibit the tyrosinase and melanin level: Synthesis, in-vitro, and in-silico studies.

In this research work, we have designed and synthesized some biologically useful of 1,3,4-Oxadiazoles. The structural interpretation of the synthesized compounds has been validated by using FT-IR, LC-MS, HRMS, 1H NMR and 13C NMR techniques. Moreover, the in-vitro mushroom tyrosinase inhibitory potential of the target compounds was assessed. The in-vitro study reveals that, all compounds demonstrate an excellent tyrosinase inhibitory activity. Especially, 2-(5-(2-methoxyphenyl)-1,3,4-oxadiazol-2-ylthio)-N-phenylacetamide (IC50 = 0.003 ± 0.00 µM) confirms much more significant potent inhibition activity compared with standard drug kojic acid (IC50 = 16.83 ± 1.16 µM). Subsequently, the most potent five oxadiazole compounds were screened for cytotoxicity study against B16F10 melanoma cells using an MTT assay method. The survival rate for the most potent compound was more pleasant than other compounds. Furthermore, the western blot results proved that the most potent compound considerably decreased the expression level of tyrosinase at 50 µM (P < 0.05). The molecular docking investigation exposed that the utmost potent compound displayed the significant interactions pattern within the active region of the tyrosinase enzyme and which might be responsible for the decent inhibitory activity towards the enzyme. A molecular dynamic simulation experiment was presented to recognize the residual backbone stability of protein structure.

Targeting FGL2, a molecular drug target for glioblastoma, with natural compounds through virtual screening method.

Background: Fibroleukin-2 protein (FGL2) causes redevelopment of brain tumors. Inhibition of these proteins has shown to improve glioblastoma prognosis and treatment efficacy. Aim: The current study gathered recently exploited natural compounds that suppress glioblastoma proliferation in vitro, tested against FGL2 protein. Method: Twenty-five compounds were explored through a virtual screening platform. Results: Three natural compounds (betanine, hesperetin and ovatodiolide) hit the active site of FGL2. Furthermore, the influence of these compounds was also assessed using in silico gene expression, and ADMET tools showed downregulation of some genes, which caused rapid tumor development while possessing a moderate acute toxicity and pharmacokinetic profile. Conclusion: Our study presents three compounds that are good candidates for evaluation in FGL2 mutated glioblastoma animal models.

Novel 1,2,4-triazole analogues as mushroom tyrosinase inhibitors: synthesis, kinetic mechanism, cytotoxicity and computational studies.

We have created a novel series of mushroom tyrosinase inhibitors with 1,2,4-triazole as fundamental skeleton. The target compound 1,2,4-triazol-3-ylthio)-N-phenyl acetamide derivatives 9(a-l) were synthesized by the reaction of 4- and 5-substituted 1,2,4-triazole-3-thiol derivatives 6(a-c) with 2-chloro-N-sub/un-substituted phenyl acetamide derivatives 8(a-d) under basic condition. By using the analytical techniques for instance, FTIR, LC-MS, 1H NMR and 13C NMR, the structural verification was evaluated. The novel series of the target compounds 9(a-l) has been scanned for biological activity (mushroom tyrosinase inhibition potential) which demonstrates adequate results. Interestingly, compound 9k (IC50 = 0.0048 ± 0.0016 µM) exhibits 3500 times more activity compared with standard drug kojic acid (IC50 = 16.8320 ± 1.1600 µM) against mushroom tyrosinase inhibitor. Furthermore, the cytotoxicity experiment was carried out for the highly effective target compounds (9d, 9i, 9j and 9k) by using MTT assay method for A375 human melanoma cells to define the nontoxic performance of the most effective compounds ranging from 1 to 25 µM. Furthermore, the molecular docking study delivers the thought concerning the interface of the ligand with an enzyme. Also, the dynamic simulation was accomplished for compound 9k to govern the plausible binding model.

Inhibitory effects on melanogenesis by thymoquinone are mediated through the ß­catenin pathway in B16F10 mouse melanoma cells.

Thymoquinone (TQ) is a component found in the seeds of Nigella sativa, an annual plant growing on the Mediterranean coast, and is known for its anticancer and anti­inflammatory effects. However, to date, at least to the best of our knowledge, limited studies are available examining the molecular mechanisms through which TQ inhibits melanogenesis. Accordingly, this study aimed to treat B16F10 mouse melanoma cells with TQ to investigate its apparent effects and its molecular regulatory mechanisms. Treatment of the B16F10 cells with 10, 15 and 20 µM of TQ for 48 h resulted in a dose­dependent decrease in the expression of microphthalmia­associated transcription factor (MITF), tyrosinase expression and tyrosinase activity, and these treatments simultaneously led to a decrease in the protein expression and transcription of ß­catenin, a Wnt signaling pathway protein. Pre­treatment of the cells with the proteasome inhibitor, MG132, to confirm the inhibition of melanogenesis through the ß­catenin pathway by TQ treatment resulted in an increase in the expression of ß­catenin that was initially reduced by TQ, and the expression and activity of MITF and tyrosinase also increased. Pre­treatment with LiCl, which is known to inactivate glycogen synthase kinase 3ß (GSK3ß) by inducing the phosphorylation of the Ser­9 site, resulted in an increased phospho­GSK3ß expression accompanied by ß­catenin that was initially reduced by TQ, and the recovery of the expression and activity of tyrosinase was also confirmed. The transfection of S37A cDNA into B16F10 cells that overexpress ß­catenin resulted in the recovery of ß­catenin expression that was initially reduced by TQ, and this treatment also recovered the expression and activity of tyrosinase. When zebrafish eggs were treated with 1, 2.5 and 5 µM of TQ at 10 h following fertilization, their melanin content decreased in a dose­dependent manner. On the whole, these findings demonstrated that the inhibition of melanogenesis in B16F10 mouse melanoma cells by TQ treatment resulted from the inhibition of the ß­catenin pathway and confirmed that TQ treatment inhibited melanogenesis in zebrafish.

Gallotannin attenuates 2­deoxy­D­glucose­induced dedifferentiation and endoplasmic reticulum stress through inhibition of inositol­requiring enzyme 1 downstream p38 kinase pathway in chondrocytes.

Gallotannin (GT) is a class of polyphenols with antioxidant, anticancer, and antiviral activities. 2­Deoxy­D­glucose (2DG), a glucose­derived molecule, can inhibit glucose metabolism and induce endoplasmic reticulum (ER) stress. GT in primary­cultured chondrocytes enhances expression of type II collagen, an indicator of differentiation, and cyclooxygenase­2 (COX­2), which mediates inflammatory reactions. In contrast, 2DG reduces type II collagen and COX­2 expression while driving ER­stress­induced unglycosylation. In the present study, it was investigated whether GT could attenuate 2DG­induced dedifferentiation and ER­stress. Following treatment with GT and 2DG, chondrocytes were assessed using western blotting, RT­PCR, immunofluorescence, and alcian blue staining. GT restored type II collagen expression that was reduced by 2DG, inhibited ER­stress­induced COX­2 unglycosylation, and induced COX­2 expression. The expression of a glucose­regulated protein, GRP78, which is an indicator of reduced ER­stress, was decreased. To link the GT signaling pathway with pathways that inhibit 2DG­induced dedifferentiation and ER­stress, inhibitors were treated in chondrocytes. The results revealed that, among the different signaling pathways triggered by ER­stress, the p38 kinase pathway was involved in the inositol­requiring enzyme 1 (IRE1) downstream signaling pathway. Following inhibition of the IRE1 pathway, type II collagen expression was increased and COX­2 expression was decreased. In addition, after examining the splicing of X­box binding protein 1 (XBP­1) which is dependent on IRE1 activation induced by ER­stress, it was revealed that GT inhibited the increase of XBP­1s after splicing due to 2DG­induced ER stress. GT in chondrocytes inhibited 2DG­induced dedifferentiation and ER­stress­induced COX­2 unglycosylation while regulating differentiation and inflammation via the ER­stress­induced p38 kinase pathway downstream from the IRE1 pathway.