Anticancer Effects of New Ceramides Isolated from the Red Sea Red Algae Hypnea musciformis in a Model of Ehrlich Ascites Carcinoma: LC-HRMS Analysis Profile and Molecular Modeling.

Different classes of phytochemicals were previously isolated from the Red Sea algae Hypnea musciformis as sterols, ketosteroids, fatty acids, and terpenoids. Herein, we report the isolation of three fatty acids-docosanoic acid 4, hexadecenoic acid 5, and alpha hydroxy octadecanoic acid 6-as well as three ceramides-A (1), B (2), and C (3)-with 9-methyl-sphinga-4,8-dienes and phytosphingosine bases. Additionally, different phytochemicals were determined using the liquid chromatography coupled with electrospray ionization high-resolution mass spectrometry (LC-ESI-HRMS) technique. Ceramides A (1) and B (2) exhibited promising in vitro cytotoxic activity against the human breast adenocarcinoma (MCF-7) cell line when compared with doxorubicin as a positive control. Further in vivo study and biochemical estimation in a mouse model of Ehrlich ascites carcinoma (EAC) revealed that both ceramides A (1) and B (2) at doses of 1 and 2 mg/kg, respectively, significantly decreased the tumor size in mice inoculated with EAC cells. The higher dose (2 mg/kg) of ceramide B (2) particularly expressed the most pronounced decrease in serum levels of vascular endothelial growth factor -B (VEGF-B) and tumor necrosis factor-α (TNF-α) markers, as well as the expression levels of the growth factor midkine in tumor tissue relative to the EAC control group. The highest expression of apoptotic factors, p53, Bax, and caspase 3 was observed in the same group that received 2 mg/kg of ceramide B (2). Molecular docking simulations suggested that ceramides A (1) and B (2) could bind in the deep grove between the H2 helix and the Ser240-P250 loop of p53, preventing its interaction with MDM2 and leading to its accumulation. In conclusion, this study reports the cytotoxic, apoptotic, and antiangiogenic effects of ceramides isolated from the Red Sea algae Hypnea musciformis in an experimental model of EAC.

New Antiproliferative Triflavanone from Thymelaea hirsuta-Isolation, Structure Elucidation and Molecular Docking Studies.

In this study isolates from Thymelaea hirsuta, a wild plant from the Sinai Peninsula of Egypt, were identified and their selective cytotoxicity levels were evaluated. Phytochemical examination of the ethyl acetate (EtOAc) fraction of the methanolic (MeOH) extract of the plant led to the isolation of a new triflavanone compound (1), in addition to the isolation of nine previously reported compounds. These included five dicoumarinyl ethers found in Thymelaea: daphnoretin methyl ether (2), rutamontine (3), neodaphnoretin (4), acetyldaphnoretin (5), and edgeworthin (6); two flavonoids: genkwanin (7) and trans-tiliroside (8); p-hydroxy benzoic acid (9) and ß sitosterol glucoside (10). Eight of the isolated compounds were tested for in vitro cytotoxicity against Vero and HepG2 cell lines using a sulforhodamine-B (SRB) assay. Compounds 1, 2 and 5 exhibited remarkable cytotoxic activities against HepG2 cells, with IC50 values of 8.6, 12.3 and 9.4 µM, respectively, yet these compounds exhibited non-toxic activities against the Vero cells. Additionally, compound 1 further exhibited promising cytotoxic activity against both MCF-7 and HCT-116 cells, with IC50 values of 4.26 and 9.6 µM, respectively. Compound 1 significantly stimulated apoptotic breast cancer cell death, resulting in a 14.97-fold increase and arresting 40.57% of the cell population at the Pre-G1 stage of the cell cycle. Finally, its apoptosis-inducing activity was further validated through activation of BAX and caspase-9, and inhibition of BCL2 levels. In silico molecular docking experiments revealed a good binding mode profile of the isolates towards Ras activation/pathway mitogen-activated protein kinase (Ras/MAPK); a common molecular pathway in the development and progression of liver tumors.

Chemical Constituent Profiling of Phyllostachys heterocycla var. Pubescens with Selective Cytotoxic Polar Fraction through EGFR Inhibition in HepG2 Cells.

Different extracts of the Bamboo shoot skin Phyllostachys heterocycla var. pubescens were screened against panel of cancer cell lines and normal one. The cell viability results exhibited that the ethyl acetate extract showed the least vitality percentage of 2.14% of HepG2 cells. Accordingly, it was subjected to chromatographic separation, which resulted in the isolation of a new natural product; 7-hydroxy, 5-methoxy, methyl cinnamate (1), together with four known compounds. The structures of the pure isolated compounds were deduced based on different spectroscopic data. The new compound (1) was screened against the HepG2 and MCF-7 cells and showed IC50 values of 7.43 and 10.65 µM, respectively. It induced apoptotic cell death in HepG2 with total apoptotic cell death of 58.6% (12.44-fold) compared to 4.71% in control by arresting cell cycle progression at the G1 phase. Finally, compound 1 was validated as EGFR tyrosine kinase inhibitor in both enzymatic levels (IC50 = 98.65 nM compared to Erlotinib (IC50 = 78.65 nM). Finally, in silico studies of compound 1 through the molecular docking indicated its high binding affinity towards EGFR protein and the ADME pharmacokinetics indicated it as a drug-like.

Thalassosterol, a New Cytotoxic Aromatase Inhibitor Ergosterol Derivative from the Red Sea Seagrass Thalassodendron ciliatum.

Thalassodendron ciliatum (Forssk.) Den Hartog is a seagrass belonging to the plant family Cymodoceaceae with ubiquitous phytoconstituents and important pharmacological potential, including antioxidant, antiviral, and cytotoxic activities. In this work, a new ergosterol derivative named thalassosterol (1) was isolated from the methanolic extract of T. ciliatum growing in the Red Sea, along with two known first-reported sterols, namely ergosterol (2) and stigmasterol (3), using different chromatographic techniques. The structure of the new compound was established based on 1D and 2D NMR spectroscopy and high-resolution mass spectrometry (HR-MS) and by comparison with the literature data. The new ergosterol derivative showed significant in vitro antiproliferative potential against the human cervical cancer cell line (HeLa) and human breast cancer (MCF-7) cell lines, with IC50 values of 8.12 and 14.24 µM, respectively. In addition, docking studies on the new sterol 1 explained the possible binding interactions with an aromatase enzyme; this inhibition is beneficial in both cervical and breast cancer therapy. A metabolic analysis of the crude extract of T. ciliatum using liquid chromatography combined with high-resolution electrospray ionization mass spectrometry (LC-ESI-HR-MS) revealed the presence of an array of phenolic compounds, sterols and ceramides, as well as di- and triglycerides.

New Cytotoxic Natural Products from the Red Sea Sponge Stylissa carteri.

Bioactivity-guided isolation supported by LC-HRESIMS metabolic profiling led to the isolation of two new compounds, a ceramide, stylissamide A (1), and a cerebroside, stylissoside A (2), from the methanol extract of the Red Sea sponge Stylissa carteri. Structure elucidation was achieved using spectroscopic techniques, including 1D and 2D NMR and HRMS. The bioactive extract's metabolomic profiling showed the existence of various secondary metabolites, mainly oleanane-type saponins, phenolic diterpenes, and lupane triterpenes. The in vitro cytotoxic activity of the isolated compounds was tested against two human cancer cell lines, MCF-7 and HepG2. Both compounds, 1 and 2, displayed strong cytotoxicity against the MCF-7 cell line, with IC50 values at 21.1 ± 0.17 µM and 27.5 ± 0.18 µM, respectively. They likewise showed a promising activity against HepG2 with IC50 at 36.8 ± 0.16 µM for 1 and IC50 30.5 ± 0.23 µM for 2 compared to the standard drug cisplatin. Molecular docking experiments showed that 1 and 2 displayed high affinity to the SET protein and to inhibitor 2 of protein phosphatase 2A (I2PP2A), which could be a possible mechanism for their cytotoxic activity. This paper spreads light on the role of these metabolites in holding fouling organisms away from the outer surface of the sponge, and the potential use of these defensive molecules in the production of novel anticancer agents.

Cytotoxic, Apoptosis-Inducing Activities, and Molecular Docking of a New Sterol from Bamboo Shoot Skin Phyllostachys heterocycla var. pubescens.

Phytochemical screening of nonpolar fractions from the methanol extract of the Bamboo shoot skin Phyllostachys heterocycla var. pubescens resulted in the isolation of a new sterol-glucoside-fatty acid derivative (6'-O-octadeca-8'',11''-dienoyl)-sitosterol-3-O-ß-d-glucoside (1), together with six known compounds. The chemical structures of the pure isolated compounds were deduced based on different spectral data. The isolated compounds were assessed to determine their cytotoxic activity, and the results were confirmed by determining their apoptotic activity. Compound 1 was more cytotoxic against the MCF-7 cells (IC50 = 25.8 µM) compared to Fluorouracil (5-FU) (26.98 µM), and it significantly stimulated apoptotic breast cancer cell death with 32.6-fold (16.63% compared to 0.51 for the control) at pre-G1 and G2/M-phase cell cycle arrest and blocked the progression of MCF-7 cells. Additionally, RT-PCR results further confirmed the apoptotic activity of compound 1 by the upregulation of proapoptotic genes (P53; Bax; and caspases 3, 8, and 9) and downregulation of the antiapoptotic genes (BCL2). Finally, the identified compounds, especially 1, were found to have high binding affinity towards both tyrosine-specific protein kinase (TPK) and vascular endothelial growth factor receptor (VEGFR-2) through the molecular docking studies that highlight its mode of action.

Chemical profiling, cytotoxic activities through apoptosis induction in MCF-7 cells and molecular docking of Phyllostachys heterocycla bark nonpolar extract.

The chemical constituents of the nonpolar fractions of the bamboo shoot skin Phyllostachys heterocycla were extensively studied. The phytochemical study was divided into two parts: the first deals with isolation of the chemical constituents using different chromatographic techniques that resulted in isolation of four compounds. The chemical structures of the pure isolated compounds were elucidated using different spectroscopic data. The second part deals with identification of the rest of the constituents using the GC technique. Additionally, both crude extract and the pure isolated compounds were investigated for cytotoxic activity. One of the isolated compounds; namely glyceryl 1-monopalmitate showed highly promising effect against the MCF-7 cells with (IC50 = 19.78 µM) compared to 5-FU (26.98 µM), and it remarkably stimulated apoptotic breast cancer cell death with 31.6-fold (16.13% compared to 0.51 for the control) at pre-G1 and G2/M-phase cell cycle arrest and blocked the progression of MCF-7 cells. Moreover, the identified compounds especially 1 were found to have high binding affinity towards both TPK and VEGFR-2 through the molecular docking studies which highlight its mode of action. HighlightsChemical profiling of Phyllostachys heterocycla bark nonpolar extract was fully identified.Glyceryl 1-monopalmitate showed highly promising effect against the MCF-7 cells with (IC50 = 19.78 µM) compared to 5-FU (26.98 µM).Glyceryl 1-monopalmitate significantly stimulated apoptotic breast cancer cell death with 31.6-fold by arresting cell cycle at G2/M and preG1 phases.Molecular docking simulation showed good binding affinities towards TPK and VEGFR-2 proteins.Communicated by Ramaswamy H. Sarma.

Isolates From Thymelaea Hirsuta Inhibit Progression Of Hepatocellular Carcinoma In Vitro And In Vivo.

Herein, we investigated effect of Thymelaea hirsuta isolates on hepatocellular carcinoma. Methanolic extract of T. hirsuta led to isolation of two new compounds [6` hydroxyDaphnoretin (9) and Mithnin (15)], seven compounds reported for the first time from genus Thymelaea [Dotriacontanol (1), and 3-ketopentatriacontanoic (2), Docosylcoumarate (5), Docosylcaffeate (6), Daphnodorin B (11), 3`` -epi-dihydrodaphnodorin B (12) and Wikstaiwanone B (14)], and six known compounds. Eight compounds (5, 6, 9, 10, 11, 12, 14, and 15) showed significant anti-proliferative activity on HepG2 cells. These compounds caused significant reduction (p < 0.05) in serum levels of AST, ALT, ALP, total bilirubin, GGT, and AFP, a significant increase in Bax and p53 expression, and a significant decrease in Bcl2 gene in liver as compared to the HCC group. These results indicate that T. hirsuta isolates inhibited HCC progression, possibly through induction of apoptosis and therefore they could be used as a beneficial source for treating HCC.

Anti-inflammatory effect of methoxyflavonoids from Chiliadenus montanus (Jasonia Montana) growing in Egypt.

Chiliadenus montanus is a medicinal plant that grows in Sinai Peninsula in Egypt. Phytochemical investigation of C. montanus methanolic extract led to the isolation of five methoxy flavonoids; Chrysosplenol-D (1), 5,7,4'-trihydroxy- 3,3'-dimethoxy flavone (2), 5,7-dihydroxy -3,3',4'-trimethoxyflavone (3), Bonanzin (4), 3,5,6,7,4'-pentamethoxy flavone (5), a sesquiterpene, Cryptomeridiol (6) and stigmast-5,22-dien-3-O-ß-D-glucopyranoside (7). The anti-inflammatory activity of compounds 2 and 5 was assessed in vitro on CaCo2 cells stimulated by lipopolysaccharide (LPS). Both compounds downregulated LPS-induced expression of inflammatory cytokines; tumor necrosis factor alpha (TNFα), interleukin 1ß (IL1ß), nuclear factor kappa B (NFκB), cyclooxygenase 1 (Cox1), cyclooxygenase 2 (Cox2), and 5-lipoxygenase (5Lox). In vivo, both compounds significantly decreased paw edema thickness in rats relative to carrageenan, showing better anti-inflammatory activity than celecoxib (36.98%) after 1 h (46.60% and 48.11%, respectively). An in silico study was performed, where both compounds were docked into the active site of the crystal structure of the human Cox2 enzyme.

Chemical Composition of the Red Sea Green Algae Ulva lactuca: Isolation and In Silico Studies of New Anti-COVID-19 Ceramides.

Coronavirus disease 2019 (COVID-19) is the disease caused by the virus SARS-CoV-2 responsible for the ongoing pandemic which has claimed the lives of millions of people. This has prompted the scientific research community to act to find treatments against the SARS-CoV-2 virus that include safe antiviral medicinal compounds. The edible green algae U. lactuca. is known to exhibit diverse biological activities such as anti-influenza virus, anti-Japanese encephalitis virus, immunomodulatory, anticoagulant, antioxidant and antibacterial activities. Herein, four new ceramides in addition to two known ones were isolated from Ulva lactuca. The isolated ceramides, including Cer-1, Cer-2, Cer-3, Cer-4, Cer-5 and Cer-6 showed promising antiviral activity against SARS-CoV-2 when investigated using in silico approaches by preventing its attachment to human cells and/or inhibiting its viral replication. Cer-4 and Cer-5 were the most effective in inhibiting the human angiotensin converting enzyme (hACE)-spike protein complex which is essential for the virus to enter the human host. In addition to this, Cer-4 also showed an inhibition of the SARS-CoV-2 protease (Mpro) that is responsible for its viral replication and transcription. In this study, we also used liquid chromatography coupled to electrospray ionization high-resolution mass spectroscopy (LC-ESI-HRMS) to identify several metabolites of U. lactuca, including metabolites such as fatty acids, their glyceride derivatives, terpenoids, sterols and oxysterols from the organic extract. Some of these metabolites also possessed promising antiviral activity, as previously reported.

Antioxidant and Anti-Inflammatory Activity of Cynanchum acutum L. Isolated Flavonoids Using Experimentally Induced Type 2 Diabetes Mellitus: Biological and In Silico Investigation for NF-κB Pathway/miR-146a Expression Modulation.

Cynanchum acutum L. is a climbing vine that belongs to the family Apocynaceae. Using different chromatographic techniques, seven compounds were isolated from the methanolic extract of the plant. The isolated compounds include six flavonoid compounds identified as rutin (1), quercetin-3-O-neohesperidoside (2), quercetin-3-O-ß-galactoside (3), isoquercitrin (4), quercetin (5), and kaempferol 3-O-ß-glucoside (6), in addition to a coumarin, scopoletin (7). The structures of the compounds were elucidated based on 1D NMR spectroscopy and high-resolution mass spectrometry (HR-MS), and by comparison with data reported in the literature. The first five compounds were selected for in vivo investigation of their anti-inflammatory and antioxidant properties in a rat model of type 2 diabetes. All tested compounds significantly reduced oxidative stress and increased erythrocyte lysate levels of antioxidant enzymes, along with the amelioration of the serum levels of inflammatory markers. Upregulation of miR-146a expression and downregulation of nuclear factor kappa B (NF-κB) expression were detected in the liver and adipose tissue of rats treated with the isolated flavonoids. Results from the biological investigation and those from the validated molecular modeling approach on two biological targets of the NF-κB pathway managed to highlight the superior anti-inflammatory activity of quercetin-3-O-galactoside (3) and quercetin (5), as compared to other bioactive metabolites.

VEGFR-Mediated Cytotoxic Activity of Pulicaria undulata Isolated Metabolites: A Biological Evaluation and In Silico Study.

Natural products play a remarkable role not only in the synthesis, design, and discovery of new drugs but also as the most prominent source of drugs and bioactive substances. Adding to the search for new sources of safe innovative antitumor drugs, here we reported a phytochemical study on Pulicaria undulata which revealed promising antiangiogenic agents. Six compounds were isolated and identified as xanthoxyline (1), stigmasterol (2), oleanolic acid (3), salvigenin (4), rhamnetin (5) and dihydroquercetin-4'-methyl ether (6) using nuclear magnetic resonance (NMR) spectroscopic techniques. Compound 3 and 4 are first reported in Pulicaria genus. Both the extract and isolated compounds were evaluated for in vitro antiproliferative activity against breast cancer cell line (MCF-7). In vivo antiproliferative activity against Ehrlich's ascites carcinoma (EAC) were also assessed. The P. undulata extract and isolates showed significant reduction in tumor weight, decreased both serum vascular endothelial growth factor B (VEGF-B) levels and vascular endothelial growth factor receptor 2 (VEGFR-2) expression significantly compared to the control EAC group, suggesting an antiangiogenic activity through the inhibition of VEGF signaling. Besides, they displayed reduction in CD34 expression, confirming their antiangiogenic effect. Moreover, the potential affinity of isolated compounds to human estrogen nuclear receptor-alpha (hER-α), the most recognized modulator of VEGFR-2 expression, was virtually estimated through molecular modeling studies. The most promising activity profiles were assigned to the investigated flavonoids, compounds 4-6, as well as the alkyl-phenylketone, compound 1. Additionally, these four top active compounds showed respective high to intermediate docking scores while possessing preferential binding with hER-α critical pocket residues. Based on the provided data, these isolated compounds illustrated promising inhibitors of VEGF-stimulated angiogenesis, which could be a possible mechanism for their anticancer activity.

Ergosterol Peroxide from the Egyptian Red Lingzhi or Reishi Mushroom, Ganoderma resinaceum (Agaricomycetes), Showed Preferred Inhibition of MCF-7 over MDA-MB-231 Breast Cancer Cell Lines.

Ergosterol peroxide and ganoderic acid AMI were isolated for the first time from the mycelium of the Egyptian Ganoderma resinaceum mushroom. The structure of these two metabolites was established by detailed analysis of 1D and 2D NMR. The isolated compounds were tested for their antitumor in vitro activities in MCF-7 and MDA-MB-231 breast cancer cell lines. Ergosterol peroxide showed preferred inhibition of MCF-7 (ER +ve) cell lines relative to MDA-MB-231 (ER -ve) cell lines with an IC50 of 1.18 µM and 12.82 µM respectively. Our data suggest that ergosterol peroxide targets estrogen receptors.

Selective kappa opioid antagonists for treatment of addiction, are we there yet?

Kappa opioid receptor (KOP) is a G-protein coupled receptor mainly expressed in the cerebral cortex and hypothalamus. It is implicated in nociception, diuresis, emotion, cognition, and immune system functions. KOP agonists possess a strong analgesic effect accompanied by a feeling of dysphoria. On the other hand, antagonists of this receptor were found to block depression, anxiety, and drug-seeking behaviors in animal models. Recently, great interest has been given to the development of selective KOP antagonists as an addiction treatment that does not cause dependence itself or show high relapse rates like the currently used agents. This review provides a comprehensive survey of the KOP antagonists developed for this purpose together with their in vivo studies and clinical trials. In addition, a future perspective and recommendations for the work needed to develop clinically relevant KOP antagonists are presented.

IN VITRO ANTI-PROTOZOAL ACTIVITY OF PROPOLIS EXTRACT AND CYSTEINE PROTEASES INHIBITOR (PHENYL VINYL SULFONE) ON BLASTOCYSTIS SPECIES.

Blastocystis is one of the commonest enteric protozoan parasites worldwide. Despite its controversial clinical significance, frequent association with symptoms has necessitated treatment of cases with persistent symptoms. For long time, metronidzole (MTZ) was considered as a basic drug for blastocystosis treatment, however reports of treatment failure as well as its well-known side effects has promoted the search for more safe and effective alternatives. In vitro antiprotozoal activity of ethanolic extract of Egyptian propolis and a cysteine protease inhibitor, phenyl vinyl sulfone (PVS) on Blastocystis spp. was assessed through challenging with graded concentrations of propolis extract (125, 250, 500 & 1000pg/ml) and PVS (100, 200 and 300 ptg/ml) compared to MTZ (10, 50 and 100 pg/ml) and viable parasites were counted after 24, 48 and 72 hr. of incubation. Molecular subtyling of Blastocystis spp. was done using subtype specific sequence-tagged site (STS) primers. Propolis extract inhibited the growth of Blastocystis spp. in both of the detected subtypes (STI and ST3), which was especially observed in cultures exposed to 500 & 1000 µg/ml through all incuba- tion periods with the later concentration producing comparable results to MTZ. While PVS showed significant parasite count reduction on ST3 isolates, especially with the highest concentration, however the effect on STl isolate was nonsignificant. These findings highlight the potential antiprotozoal activity of propolis extract as a potent natural alternative for MTZ in treatment of blastocystosis.