LC-HRMS Profiling and Cytotoxic Potential of Actinomycetes Associated with the Red Sea Soft Coral Sarcophyton glaucum: In vitro and In silico Studies.

In the current study, the actinomycetes associated with the red sea-derived soft coral Sarcophyton glaucum were investigated in terms of biological and chemical diversity. Four different media, M1, ISP2, Marine Agar (MA), and Actinomycete isolation agar (AIA) were used for the isolation of three strains of actinomycetes that were identified as Streptomyces sp. UR 25, Micromonospora sp. UR32 and Saccharomonospora sp. UR 19. LC-HRMS analysis was used to investigate the chemical diversity of the isolated actinobacteria. The LC-HRMS data were statistically processed using MetaboAnalyst 5.0 viz to differentiate the extract groups and determine the optimal growth culturing conditions. Multivariate data statistical analysis revealed that the Micromonospora sp. extract cultured on (MA) medium is the most distinctive extract in terms of chemical composition. While, the Streptomyces sp. UR 25 extracts are differ significantly from Micromonospora sp. UR32 and Saccharomonospora sp. UR 19. Biological investigation using in vitro cytotoxic assay for actinobacteria extracts revealed the prominent potentiality of the Streptomyces sp. UR 25 cultured on oligotrophic medium against human hepatoma (HepG2), human breast adenocarcinoma (MCF-7) and human colon adenocarcinoma (CACO2) cell lines (IC50 =3.3, 4.2 and 6.8 µg/mL, respectively). SwissTarget Prediction speculated that among the identified compounds, 16-deethyl, indanomycin (8) could have reasonable affinity on HDM2 active site. In this respect, molecular docking study was performed for compound (8) to reveal a substantial affinity on HDM2 active site. In addition, molecular dynamics simulations were carried out at 200 ns for the most active compound (8) compared to the co-crystallized inhibitor DIZ giving deeper information regarding their thermodynamic and dynamic properties as well.

Promising Cytotoxic butenolides from the Soybean endophytic fungus Aspergillus terreus: a combined molecular docking and in-vitro studies.

AIM: This study aimed to use one strain many compounds approach (OSMAC) to investigate the cytotoxic potential of Aspergillus terreus associated with soybean versus several cancer cell lines, by means of in-silico and in vitro approaches. METHODS AND RESULTS: Fermentation of the isolated strain was done on five media. The derived extracts were investigated for their inhibitory activities against three human cancer cell lines; mammary gland breast cancer (MCF-7), colorectal adenocarcinoma (Caco-2), and hepatocellular carcinoma (HepG2) using MTT Assay. The fungal mycelia fermented in Modified Potato Dextrose Broth (MPDB) was the most cytotoxic extract against HepG2, MCF-7, and Caco-2 cell lines with IC50 4.2 ± 0.13, 5.9 ± 0.013 and 7.3 ± 0.004 µg mL-1, respectively. MPDB extract was scaled up resulting in the isolation of six metabolites; three fatty acids (1, 2, and 4), one sterol (3) and two butenolides (5 and 6) by column chromatography. The isolated compounds (1-6) were screened through a molecular docking approach for their binding aptitude to various active sites. butyrolactone-I (5) revealed a significant interaction within the CDK2 active site, while aspulvinone E (6) showed promising binding affinity to FLT3 and EGFR active sites that was confirmed by in vitro CDK2, FLT3 and EGFR inhibitory activity. Finally, the in vitro cytotoxic activities of butyrolactone-I (5) and aspulvinone E (6) revealed the antiproliferative activity of butyrolactone-I (5), against HepG2 cell line (IC50 = 17.85 ± 0.32 µM). CONCLUSION: Molecular docking analysis and in vitro assays suggested the CDK2/A2 inhibitory potential of butyrolactone-I (5) in addition to the promising interaction abilities of aspulvinone E (6) with EGFR and FLT3 active sites as a possible mechanism of their biological activities.

Induction of Antibacterial Metabolites by Co-Cultivation of Two Red-Sea-Sponge-Associated Actinomycetes Micromonospora sp. UR56 and Actinokinespora sp. EG49.

Liquid chromatography coupled with high resolution mass spectrometry (LC-HRESMS)-assisted metabolomic profiling of two sponge-associated actinomycetes, Micromonospora sp. UR56 and Actinokineospora sp. EG49, revealed that the co-culture of these two actinomycetes induced the accumulation of metabolites that were not traced in their axenic cultures. Dereplication suggested that phenazine-derived compounds were the main induced metabolites. Hence, following large-scale co-fermentation, the major induced metabolites were isolated and structurally characterized as the already known dimethyl phenazine-1,6-dicarboxylate (1), phenazine-1,6-dicarboxylic acid mono methyl ester (phencomycin; 2), phenazine-1-carboxylic acid (tubermycin; 3), N-(2-hydroxyphenyl)-acetamide (9), and p-anisamide (10). Subsequently, the antibacterial, antibiofilm, and cytotoxic properties of these metabolites (1-3, 9, and 10) were determined in vitro. All the tested compounds except 9 showed high to moderate antibacterial and antibiofilm activities, whereas their cytotoxic effects were modest. Testing against Staphylococcus DNA gyrase-B and pyruvate kinase as possible molecular targets together with binding mode studies showed that compounds 1-3 could exert their bacterial inhibitory activities through the inhibition of both enzymes. Moreover, their structural differences, particularly the substitution at C-1 and C-6, played a crucial role in the determination of their inhibitory spectra and potency. In conclusion, the present study highlighted that microbial co-cultivation is an efficient tool for the discovery of new antimicrobial candidates and indicated phenazines as potential lead compounds for further development as antibiotic scaffold.

Bioactive Brominated Oxindole Alkaloids from the Red Sea Sponge Callyspongia siphonella.

In the present study, LC-HRESIMS-assisted dereplication along with bioactivity-guided isolation led to targeting two brominated oxindole alkaloids (compounds 1 and 2) which probably play a key role in the previously reported antibacterial, antibiofilm, and cytotoxicity of Callyspongia siphonella crude extracts. Both metabolites showed potent antibacterial activity against Gram-positive bacteria, Staphylococcus aureus (minimum inhibitory concentration (MIC) = 8 and 4 µg/mL) and Bacillus subtilis (MIC = 16 and 4 µg/mL), respectively. Furthermore, they displayed moderate biofilm inhibitory activity in Pseudomonas aeruginosa (49.32% and 41.76% inhibition, respectively), and moderate in vitro antitrypanosomal activity (13.47 and 10.27 µM, respectively). In addition, they revealed a strong cytotoxic effect toward different human cancer cell lines, supposedly through induction of necrosis. This study sheds light on the possible role of these metabolites (compounds 1 and 2) in keeping fouling organisms away from the sponge outer surface, and the possible applications of these defensive molecules in the development of new anti-infective agents.

Aberration of Nrf2­Bach1 pathway in colorectal carcinoma; role in carcinogenesis and tumor progression.

Nrf2 and Bach1 are important transcriptional factors that protect against reactive oxygen species (ROS). Although aberration of these molecules was associated with malignant transformation and progression, their aberration pattern in colorectal carcinoma (CRC) is not yet fully studied. In this study, Nrf2 and Bach1 were immunohistochemically examined in 93 formalin-fixed paraffin-embedded blocks of colonic tumors (65 carcinoma with their corresponding surgical margins and 28 adenomas). Nrf2 expression was gradually increased in the apparently normal mucosa (57 ±â€¯41)-adenoma (90 ±â€¯36)-carcinoma (198 ±â€¯78) direction and only showed significant higher mean of expression in CRC with brisk inflammatory peritumoral response. The mean of Bach1 expression was highest in apparently normal colonic mucosa (267 ±â€¯16), lowest in adenoma (53 ±â€¯31) and high in carcinoma tissues (194 ±â€¯93). Significant higher mean of expression was detected in carcinoma with: LN metastasis (p = 0.04), lymphovascular invasion (p = 0.024); perineural invasion (p = 0.03) and advanced pathological stage (p < 0.001). Significant higher mean of expression of Nrf2 and Bach1 was detected in adenoma specimens with high grade dysplasia (p = 0.016 and p = 0.024) respectively. In conclusion, Nfr2 and Bach1 expression are altered in CRC but in different way. Nrf2 is gradually increasing from normal mucosa to adenoma and was highest in carcinoma but was not associated with features of tumor invasiveness. Bach1 was highest in normal mucosa; less in adenoma then increased in carcinoma and was associated with features of tumor invasiveness and metastasis. This may indicate a possible role of Nrf2 in CRC carcinogenesis and a role of Bach1 in CRC progression.

Does loss of hormonal receptors influence the pathophysiological characteristics of the HER-2 breast cancer phenotype?

BACKGROUND: Some breast carcinomas (BC) of the HER-2 type respond poorly to endocrine therapy, indicating that hormonal receptor (HR) status possibly impacts the biological criteria of this tumor class. The aim of this study was to compare the clinicopathological characteristics of HR-positive and HR-negative tumors occurring in HER-2 and non-HER-2 BC. METHODS: Tissue microarray sections from 336 primary invasive BC specimens were stained immunohistochemically with antibodies against HER-2, ER, and PR. Proliferation was assessed using Ki67 and the P53 status was identified. RESULTS: The HER-2 phenotype was identified in 42/336 (12.5%) specimens, while 293/336 (87.5%) were of the non-HER-2 phenotype. In the non-HER-2 group, 103/293 (35%) were HR-negative tumors. HR negativity was significantly associated with higher tumor grades (P < 0.0001), higher proliferation rates (P < 0.0001), presence of necrosis (P < 0.0001), and with a higher P53 expression (P < 0.0001). There were no differences in patient age, tumor size, LN status, or presence of vascular invasion (VI) between the HR-negative and HR-positive groups. In the HER-2 group, 16/42 (38%) had HR-negative tumors. No significant difference in clinicopathological characteristics, except for tumor grade, was detected between the HR-positive and HR-negative tumors in this group. CONCLUSION: Loss of HR does not influence the biological features of HER-2 BC. This finding may indicate that some tumors will 'biologically' move from being HER-2-positive/HR-positive tumors to behaving more like HER-2-positive/HR-negative tumors even when ER are present on the cell surface. Further studies are needed to explore this hypothesis and to identify the subset of tumors that will benefit from endocrine therapy.

Epigenetic Modifiers Induce Bioactive Phenolic Metabolites in the Marine-Derived Fungus Penicillium brevicompactum.

Fungi usually contain gene clusters that are silent or cryptic under normal laboratory culture conditions. These cryptic genes could be expressed for a wide variety of bioactive compounds. One of the recent approaches to induce production of such cryptic fungal metabolites is to use histone deacetylases (HDACs) inhibitors. In the present study, the cultures of the marine-derived fungus Penicillium brevicompactum treated with nicotinamide and sodium butyrate were found to produce a lot of phenolic compounds. Nicotinamide treatment resulted in the isolation and identification of nine compounds 1⁻9. Sodium butyrate also enhanced the productivity of anthranilic acid (10) and ergosterol peroxide (11). The antioxidant as well as the antiproliferative activities of each metabolite were determined. Syringic acid (4), sinapic acid (5), and acetosyringone (6) exhibited potent in vitro free radical scavenging, (IC50 20 to 30 µg/mL) and antiproliferative activities (IC50 1.14 to 1.71 µM) against HepG2 cancer cell line. Furthermore, a pharmacophore model of the active compounds was generated to build up a structure-activity relationship.

Pattern of HER-2 Gene Amplification and Protein Expression in Benign, Borderline, and Malignant Ovarian Serous and Mucinous Neoplasms.

Amplification of HER-2 gene and overexpression of HER-2 receptor play a significant role in the progression of a number of malignancies such as breast cancer. Trastuzumab (anti-HER-2 therapeutic agent) has been used successfully in treatment of breast cancer. The aim of this study was to assess the pattern of HER-2 gene amplification and of HER-2 receptor expression in a spectrum of serous and mucinous ovarian tumors to determine whether HER-2 is altered in these neoplasms similar to that occurring in breast cancer. Formalin-fixed paraffin-embedded microarray tissue sections from 212 specimens were stained with HER-2 antibody using immunohistochemistry and with anti-HER-2 DNA probe using chromogenic in situ hybridization. Specimens consisted of 65 benign tumors (50 serous and 15 mucinous), 26 borderline (13 serous and 13 mucinous), 73 malignant tumors (53 serous carcinoma and 20 mucinous carcinoma), 18 metastatic deposits (13 serous and 5 mucinous), in addition to 30 normal tissues (16 ovarian surface and 14 normal fallopian tube). HER-2 protein-positive expression was not detected in the normal or the benign tissues. Borderline neoplasms showed positive staining, but no overexpression. HER-2 overexpression was seen only in 4 carcinoma specimens: 1/53 (1.8%) primary serous carcinomas and 3/20 (15%) primary mucinous carcinomas. HER-2 gene amplification was seen in 4 specimens: 2 primary mucinous carcinomas and 2 malignant deposits of these 2 mucinous carcinomas. In conclusion, alteration of HER-2 was not detected in ovarian serous neoplasms; however, in mucinous carcinoma, HER-2 amplification and overexpression occur.

Bioactive Formylated Flavonoids from Eugenia rigida: Isolation, Synthesis, and X-ray Crystallography.

Two new flavonoids, rac-6-formyl-5,7-dihydroxyflavanone (1) and 2',6'-dihydroxy-4'-methoxy-3'-methylchalcone (2), together with five known derivatives, rac-8-formyl-5,7-dihydroxyflavanone (3), 4',6'-dihydroxy-2'-methoxy-3'-methyldihydrochalcone (4), rac-7-hydroxy-5-methoxy-6-methylflavanone (5), 3'-formyl-2',4',6'-trihydroxy-5'-methyldihydrochalcone (6), and 3'-formyl-2',4',6'-trihydroxydihydrochalcone (7), were isolated from the leaves of Eugenia rigida. The individual (S)- and (R)-enantiomers of 1 and 3, together with the corresponding formylated flavones 8 (6-formyl-5,7-dihydroxyflavone) and 9 (8-formyl-5,7-dihydroxyflavone), as well as 2',4',6'-trihydroxychalcone (10), 3'-formyl-2',4',6'-trihydroxychalcone (11), and the corresponding 3'-formyl-2',4',6'-trihydroxydihydrochalcone (7) and 2',4',6'-trihydroxydihydrochalcone (12), were synthesized. The structures of the isolated and synthetic compounds were established via NMR, HRESIMS, and electronic circular dichroism data. In addition, the structures of 3, 5, and 8 were confirmed by single-crystal X-ray diffraction crystallography. The isolated and synthetic flavonoids were evaluated for their antimicrobial and cytotoxic activities against a panel of microorganisms and solid tumor cell lines.

Prognostic significance of lymphatic invasion in lymph node-positive breast carcinoma: findings from a large case series with long-term follow-up using immunohistochemical endothelial marker.

The poor prognostic significance of lymphatic invasion (LI) in breast carcinoma (BC) as a whole and in lymph node (LN)-negative patients in particular has been recognized in several studies; however, its prognostic role in LN-positive patients is still questionable. Aim of the current study was to assess prognostic role of LI in LN-positive BC specimens. Sections from non-selected 557 LN-positive BC specimens were stained with antibody to podoplanin/D2-40. LI was identified and correlated with clinicopathological features and patients' outcome. Twenty-year overall survival (OS), disease-free interval (DFI), and development of distant metastasis (DM) or recurrence were known for all patients. LI was detected in 262/557 (47%) of specimens ranging from 1 to 350 lesion per tumor section. Its presence was associated with higher grade tumors (P<0.0001), negative hormonal receptors (P<0.0001), high HER-2 expression (P=0.006), and with increased number of positive LNs (P=0.019). In the whole LN-positive BC, presence of LI was a poor prognostic factor for OS, DFI, and development of DM both in univariate and in multivariate analysis. In further stratification of patients, LI was associated with poorer prognosis in patients with single positive LN and not in patients with >1 positive LN. In T1N1 stage, LI was highly associated with poor OS (P=0.002), DFI (P<0.0001), and DM (P<0.0001). In T2N1 patients, LI was associated only with poorer DFI (P=0.037) but not with death or DM. In the two former patient groups, LI lost significance in multivariate analysis. In conclusion, LI is a poor prognostic factor in LN-positive BC particularly for patients having single positive LN. LI therefore would add further prognostic significance when considered in treatment in those patients. We recommend incorporation of LI in breast carcinoma staging and in prognostic indices.

Metabolomic profiling of Medicago sativa-derived fungal endophytes and evaluation of their biological activities.

This study aimed to discover the potential of Medicago sativa-derived fungal endophytes as a prospective source of bioactive metabolites. In the present study, three different strains of fungal endophyte Aspergillus terreus were isolated from leaves L, roots T and stems St of Medicago sativa to explore their biological and chemical diversity. These isolated fungi were exposed to different fermentation conditions by adding various chemical elicitors to their solid fermentation media. According to LC-HRESIMS-based metabolomics and multivariate analysis, each chemical treatment had a different effect on the chemical profiles of the fungi. Orthogonal Projections to Latent Structures Discriminant Analysis (OPLS-DA) proposed several compounds with anticancer action against MCF-7 (a human breast cancer cell line) and MDA-MB-231 (a human epithelial breast cancer cell line).

New insights into the anti-inflammatory and anti-melanoma mechanisms of action of azelaic acid and other Fusarium solani metabolites via in vitro and in silico studies.

Metabolites exploration of the ethyl acetate extract of Fusarium solani culture broth that was isolated from Euphorbia tirucalli root afforded five compounds; 4-hydroxybenzaldehyde (1), 4-hydroxybenzoic acid (2), tyrosol (3), azelaic acid (4), malic acid (5), and fusaric acid (6). Fungal extract as well as its metabolites were evaluated for their anti-inflammatory and anti-hyperpigmentation potential via in vitro cyclooxygenases and tyrosinase inhibition assays, respectively. Azelaic acid (4) exhibited powerful and selective COX-2 inhibition followed by fusaric acid (6) with IC50 values (2.21 ± 0.06 and 4.81 ± 0.14 µM, respectively). As well, azelaic acid (4) had the most impressive tyrosinase inhibitory effect with IC50 value of 8.75 ± 0.18 µM compared to kojic acid (IC50 = 9.27 ± 0.19 µM). Exclusive computational studies of azelaic acid and fusaric acid with COX-2 were in good accord with the in vitro results. Interestingly, this is the first time to investigate and report the potential of compounds 3-6 to inhibit cyclooxygenase enzymes. One of the most invasive forms of skin cancer is melanoma, a molecular docking study using a set of enzymes related to melanoma suggested pirin to be therapeutic target for azelaic acid and fusaric acid as a plausible mechanism for their anti-melanoma activity.

Cytotoxicity and modulation of cancer-related signaling by (Z)- and (E)-3,4,3',5'-tetramethoxystilbene isolated from Eugenia rigida.

Bioassay-guided fractionation of the leaves of Eugenia rigida yielded three stilbenes, (Z)-3,4,3',5'-tetramethoxystilbene (1), (E)-3,4,3',5'-tetramethoxystilbene (2), and (E)-3,5,4'-trimethoxystilbene (3). Their structures were determined using 1D- and 2D-NMR spectroscopy and HRESIMS. The sterically hindered Z-stereoisomer 1, a new natural product, was prepared by time-dependent photoisomerization of the E-isomer (2) under UV irradiation at λ254 nm, while 2,3,5,7-tetramethoxyphenanthrene (5) was identified at λ365 nm by UHPLC/APCI-MS and NMR spectroscopy. Compounds 1-3 were tested against a panel of luciferase reporter gene assays that assess the activity of many cancer-related signaling pathways, and the Z-isomer (1) was found to be more potent than the E-isomer (2) in inhibiting the activation of Stat3, Smad3/4, myc, Ets, Notch, and Wnt signaling, with IC50 values between 40 and 80 µM. However, both compounds showed similar inhibition against Ap-1 and NF-κB signaling. In addition, 1 demonstrated cytotoxic activity toward human leukemia cells, solid tumor cells of epidermal, breast, and cervical carcinomas, and skin melanoma, with IC50 values between 3.6 and 4.3 µM, while 2 was weakly active against leukemia, cervical carcinoma, and skin melanoma cells. Interestingly, 2 showed antioxidant activity by inhibition of ROS generation to 50% at 33.3 µM in PMA-induced HL-60 cells, while 1 was inactive at 100 µM (vs Trolox 1.4 µM).

Elicitation for activation of the actinomycete genome's cryptic secondary metabolite gene clusters.

This review summarizes the recent advances in the elicitation approaches used to activate the actinomycete genome's cryptic secondary metabolite gene clusters and shows the diversity of natural products obtained by various elicitation methods up to June 2022, such as co-cultivation of actinomycetes with actinomycetes, other non-actinomycete bacteria, fungi, cell-derived components, and/or algae. Chemical elicitation and molecular elicitation as transcription factor decoys, engineering regulatory genes, the promoter replacement strategy, global regulatory genes, and reporter-guided mutant selection were also reported. For researchers interested in this field, this review serves as a valuable resource for the latest studies and references.

Objective assessment of blood and lymphatic vessel invasion and association with macrophage infiltration in cutaneous melanoma.

The aims of this study were to investigate the role of vascular invasion (blood and lymphatic), vessel density and the presence of tumour-associated macrophages as prognostic markers in 202 cutaneous melanoma patients. Sections of primary melanoma were stained with lymphatic-specific antibody D2-40 to assess lymphatic vessel invasion and density in intratumoural and peritumoural areas; an antibody against endothelial marker CD34 was used to determine blood vessel invasion and density, and an antibody against CD68 was used to determine macrophage counts. Immunohistochemically determined vascular invasion (combined blood and lymphatic) was compared with that determined using haematoxylin and eosin (H&E) staining. The use of immunohistochemistry increased detection of vascular invasion from 8-30% of patients, and histological exam of H&E-stained tissue was associated with a false positive rate of 64%. Lymphatic vessel invasion occurred at a much higher frequency than blood vessel invasion (27 and 4% of patients, respectively). Although immunohistochemically detected vessel invasion was significantly associated with histological markers of adverse prognosis, such as increased Breslow thickness, ulceration and mitotic rate (all P<0.001), no associations with relapse-free or overall survival were observed. High macrophage counts were significantly associated with markers of aggressive disease, such as Breslow thickness, ulceration and mitotic rate (P<0.001, P<0.001, P=0.005, respectively), and lymphatic vessel invasion and high microvessel density (P=0.002 and P=0.003, respectively). These results suggest that vascular invasion is more accurately detected using immunohistochemistry and occurs predominantly via lymphatic vessels. The association of vessel characteristics with histological characteristics of the primary melanoma provides evidence for their biological importance in melanoma, but that they were not associated with clinical outcome attests to the value of existing histological prognostic biomarkers. We note that a high macrophage count may be associated with neovascularisation and primary tumour growth, and may also promote invasion through lymphatic vessels.

Objective assessment of lymphatic and blood vascular invasion in lymph node-negative breast carcinoma: findings from a large case series with long-term follow-up.

In a previous study on a small series of breast cancers, we developed objective methods for the assessment of vascular invasion (VI), using immunohistochemical staining. We found that VI was predominantly lymphovascular invasion (LVI), with minimal contribution of blood vascular invasion (BVI). The aims of the current study were: (a) to assess the frequency, extent and prognostic role of LVI and BVI in a large, well-characterized series of LN-negative breast cancers; and (b) to assess the ability of VI to stratify early breast cancer into different prognostic groups. Paraffin-embedded sections from 1005 lymph-node (LN)-negative primary invasive breast cancers were stained for CD34, CD31 and podoplanin/D240 to detect BVI and LVI. VI lesions were assessed and the results were correlated with clinicopathological criteria and survival. VI was detected in 218 (22%); 211/218 (97%) were LVI, while BVI was detected in 7/218 (3%). The frequency of LVIs/section ranged from 1 to 79, with no significant difference between the frequency of LVI and outcome. The presence of LVI was significantly associated with adverse disease-free interval (DFI) and poor overall survival (OS) in both univariate and multivariate analyses. The results from the study indicated that VI in early stage breast cancer is predominantly LVI and that its objective assessment is a powerful independent prognostic factor. Efforts to detect early metastatic activity, such as diligent pathological examination of sentinel LN biopsies would be complimented by the objective evaluation of VI status of the primary tumour. VI status should be included routinely in breast cancer staging systems.

Simultaneous determination of saponins and a flavonoid from aerial parts of Zygophyllum coccineum L.

Triterpenoid saponins are a class of glycosides with a wide range of bioactivities, which make them interesting research candidates. Zygophyllum coccineum is an Egyptian desert plant rich in triterpenoid saponins. Reviewing the relevant literature, no data concerning the HPLC or ultra-performance LC (UPLC) analysis of Zygophyllum content were found. This paper presents two methods, HPLC-UV and UPLC-UV-evaporative light scattering detector (ELSD)/MS, for the simultaneous determination of 10 compounds in the alcohol extract of Z. coccineum. The HPLC method uses a C18 column and water-acetonitrile (both containing 0.1% trifluoroacetic acid) gradient system. The separation was achieved within 32 min. The developed UPLC method simultaneously detects and quantifies the 10 compounds using an Acquity UPLC BEH Shield RP18 column and reagent alcohol-acetonitrile (80/20, v/v) and water (both containing 0.5% formic acid) gradient system within 14 min with UV, ELS, and MS detectors. The methods were used to analyze another species, Z. simplex, and results revealed a great variation between the secondary metabolite pattern of both species.

Antitrypanosomal activity of new semi-synthetic bergenin derivatives.

Bergenin and 11-O-(4'-O-methyl galloyl)-bergenin, previously isolated from Crassula capitella extract, were used as starting materials for the synthesis of eight derivatives; four derivatives 2a-2d were synthesized from bergenin through the formation of ester derivatives and four alkyl derivatives 4a-4d were synthesized from 11-O-(4'-O-methyl galloyl)-bergenin. The structures of the synthesized analogues were confirmed upon 1 H and 13 C NMR spectroscopic elucidation. Antileishmanial and antitrypanosomal activities of the synthesized derivatives were evaluated, compounds 11-O-(3',5' di-benzyl, 4'-O-methyl galloyl)-8,10-di-O-benzyl-bergenin (4c) and 11-O-(3',5'di-4-chlorobenzyl,4'-O-methyl galloyl)-8,10di-O-4-chlorobenzyl bergenin (4d) showed potent antitrypanosomal activity with IC50 values of 0.52 and 0.5 µM, respectively and IC90  values of 0.66 µM against T. brucei compared with IC50 and IC90 values of 21.7 and 50.3 µM for the positive control difluoromethylornithine.

Digalloyl Glycoside: A Potential Inhibitor of Trypanosomal PFK from Euphorbia abyssinica J.F. Gmel.

Human African trypanosomiasis is an endemic infectious disease caused by Trypanosoma brucei via the bite of tsetse-fly. Most of the drugs used for the treatment, e.g., Suramin, have shown several problems, including the high level of toxicity. Accordingly, the discovery of anti-trypanosomal drugs from natural sources has become an urgent requirement. In our previous study on the anti-trypanosomal potential of Euphorbia species, Euphorbia abyssinica displayed significant anti-trypanosomal activity. Therefore, a phytochemical investigation of the methanolic extract of E. abyssinica was carried out. Twelve compounds, including two triterpenes (1, 2); one sterol-glucoside (4); three ellagic acid derivatives (3, 9, 11); three gallic acid derivatives (5, 6, 10); and three flavonoids (7, 8, 12), were isolated. The structures of isolated compounds were determined through different spectroscopic techniques. Compound (10) was obtained for the first time from genus Euphorbia while all other compounds except compound (4), were firstly reported in E. abyssinica. Consequently, an in silico study was used to estimate the anti-trypanosomal activity of the isolated compounds. Several compounds displayed interesting activity where 1,6-di-O-galloyl-d-glucose (10) appeared as the most potent inhibitor of trypanosomal phosphofructokinase (PFK). Moreover, molecular dynamics (MD) simulations and ADMET calculations were performed for 1,6-di-O-galloyl-d-glucose. In conclusion, 1,6-di-O-galloyl-d-glucose revealed high binding free energy as well as desirable molecular dynamics and pharmacokinetic properties; therefore, it could be suggested for further in vitro and in vivo studies for trypanosomiasis.

Characterization of Promising Cytotoxic Metabolites from Tabebuia guayacan Hemsl.: Computational Prediction and In Vitro Testing.

Genus Tabebuia is famous for its traditional uses and valuable phytoconstituents. Our previous investigation of Tabebuia species noted the promising anticancer activity of T. guayacan Hemsl. leaves extract, however, the mechanism underlying the observed anticancer activity is still unexplored. The current research was designed to explore the phytochemical content as well as to address the phytoconstituent(s) responsible for the recorded anticancer activity. Accordingly, sixteen compounds were isolated, and their structures were elucidated using different spectroscopic techniques. The drug-likeness of the isolated compounds, as well as their binding affinity with four anticancer drug target receptors: CDK-2/6, topoisomerase-1, and VEGFR-2, were evaluated. Additionally, the most promising compounds were in vitro evaluated for inhibitory activities against CDK-2/6 and VEGFR-2 enzymes using kinase assays method. Corosolic acid (3) and luteolin-7-O-ß-glucoside (16) were the most active inhibitors against CDK-2 (-13.44 kcal/mol) and topoisomerase 1 (-13.83 kcal/mol), respectively. Meanwhile, quercetin 3-O-ß-xyloside (10) scored the highest binding free energies against both CDK-6 (-16.23 kcal/mol) as well as against VEGFR-2 protein targets (-10.39 kcal/mol). Molecular dynamic simulation indicated that quercetin 3-O-ß-xyloside (10) exhibited the least fluctuations and deviations from the starting binding pose with RMSD (2.6 Å). Interestingly, in vitro testing results confirmed the potent activity of 10 (IC50 = 0.154 µg/mL) compared to IC50 = 0.159 µg/mL of the reference drug ribociclib. These findings suggest the three noted compounds (3, 10, and 16) for further in vivo anticancer studies.