Berberis lyceum and Fumaria indica: in vitro cytotoxicity, antioxidant activity, and in silico screening of their selected phytochemicals as novel hepatitis C virus nonstructural protein 5A inhibitors.

Berberis lyceum and Fumaria indica are two Pakistani indigenous herbal medicines used to treat liver infections, including hepatitis C virus (HCV). This study aimed to evaluate the cytotoxicity, and antioxidant activity of these plant extracts and computationally screen their selected phytoconstituents as HCV NS5A inhibitors. The viability of HepG2 cells was assessed 24 h and 48 h post-treatment using colorimetric and dye exclusion methods. Antioxidant properties were examined by the 2,2-diphenyl-1-picrylhydrazyl (DPPH), reducing power, and total antioxidant capacity assays. Seventeen known phytochemicals identified from each plant were docked into the active binding site of HCV NS5A protein. The top hit ligands were analyzed for their druglikeness properties and the indices of absorption, distribution, metabolism, elimination, and toxicity (ADMET). The results showed that both plant extracts were non-toxic (CC50 > 200 µg/ml). The IC50 values of DPPH-radical scavenging activity were 51.02 ± 0.94 and 62.91 ± 1.85 µg/ml for B. lyceum and F. indica, respectively. They also exhibited reducing power and total antioxidant capacity.The phytochemicals were identified as potent HCV NS5A inhibitors with good druglikeness and ADMET properties. Six of the docked phytochemicals exhibited higher binding scores (-17.9 to -19.2 kcal/mol) with HCV NS5A protein than the standard drug, daclatasvir (-17.2 kcal/mol). Molecular dynamics (MD) simulation confirmed the stability of two compounds, berbamine and paprafumine at 100 ns with active site of HCV NS5A protein. The identified compounds through molecular docking and MD simulation could have potential as HCV NS5A inhibitor after further validation.

In vitro phytochemical and anticancer activity of Misopates orontium L. and Dicliptera bupleuroides Nees.

In the present study phytochemical analysis and anticancer activity of Misopates orontium L. and Dicliptera bupleuroides Nees was carried out. Methanolic extracts of M. orontium and D. bupleuroides were selected for phytochemical analysis. The present analysis showed the presence of phytochemical such as carbohydrates, proteins, tannins, glycosides, alkaloids, saponins, phenols and flavonoids in M. orontium and D. bupleuroides. Anticancer assays including MTT, Alamar Blue (AB), Neutral Red (NR) and lactate dehydrogenase (LDH) were employed on whole herb methanolic extract and all other fractions of both plants to calculate the % age of cell viability and cell cytotoxicity. The percentage of cell viability was highly significant in all anticancer assays for all fractions. Therefore, ethyl acetate and aqueous fractions showed the excellent profile in evaluation of cytotoxicity in each assay. All above findings indicated that the whole herb of both selected plants have strong anticancer activity.

Discovery of Novel HCV NS5B polymerase inhibitor, 2-(3,4-dimethyl-5,5-dioxidobenzo[e]pyrazolo[4,3-c][1,2]thiazin-2(4H)-yl)-N-(2-fluorobenzyl)acetamide via molecular docking and experimental approach.

Hepatitis C Virus (HCV) is a viral infection posing a severe global threat that left untreated progresses to end-stage liver disease, including cirrhosis and hepatocellular carcinoma (HCC). Moreover, no prophylactic approach exists so far enabling its prevention. The NS5B polymerase holds special significance as the target of intervention against HCV infection. The current study kindles benzothiazine derivatives against HCV NS5B polymerase through in silico and experimental approaches. Following docking, the compound 2-(3,4-dimethyl-5,5-dioxidobenzo[e]pyrazolo[4,3-c][1,2]thiazin-2(4H)-yl)-N-(2-fluorobenzyl)acetamide was revealed to form effective binding interaction in the proposed site of HCV NS5B with a score of -10 kcal/mol and subsequently was deciphered through molecular dynamics (MD) simulation study which indicated interaction of residues TYR_382, VAL_381 and HIS_467 through hydrophobic interaction and two residues such as GLU_202 and LYS_209 contributed in the formation of water bridges. The subsequent in silico pharmacological analysis revealed its safe drug profile. The cytotoxicity activity of compound 6c indicated to be non-toxic in HepG2 cells at concentration ranges from 0.001-1.0 µmol/L with >80% cell viability and diminished expression of the HCV NS5B to 98% at the dose of 1.0 µmol/L and 90% at 0.5µmol/L. Thus the hit compound 6c might be a potent NS5B polymerase inhibitor required to be validated further through in vivo and preclinical studies.

GC-MS analysis, anticancer and anti-inflammatory activities of Saussurea hypoleuca spreng. Root.

Study has been premeditated to appraise the anticancer and anti-inflammatory activities of a native medicinal plant Saussurea hypoleuca Spreng root. Anticancer assays including MTT, Alamar Blue (AB), Neutral Red (NR) & LDH were employed on root methanolic extract (RME) and all fractions to calculate % age of cell viability and cell cytotoxicity. All fractions of plant root were tested for in vitro as well as in vivo anti-inflammatory assays by reported methods. GC-MS analysis of n-hexane: chloroform fractions in column chromatography has shown isopropyl myristate, hexadecanoic acid, 11-octadecenoic acid, Di-n-octyl phthalate, dioctyl ether, decanedioic acid, 1H-3a,7-Methanoazulene, 3,4-hexanedione and Tetracosapentaene. Percentage of cell viability in anticancer assays was significantly high in all fractions. However, whole results were momentous with ethyl acetate and aqueous fractions owning to excellent profile in evaluating cytotoxicity in each assay. COX-2 inhibition was calculated which was high in RME (68.69%), ethyl acetate (56.52%), aqueous (55.21%) and chloroform fraction (53.47%). Carrageenan and formalin models were developed on rats to investigate in vivo anti-inflammatory activity. RME (56.19%, 71.09%, 66.4%, 67.99%) and ethyl acetate (51.36%, 64.97%, 55.63% & 61.01%) produced significant % age inhibition in dose dependent manner at 200 and 400 mg/kg doses respectively. All above findings direct that plant root holds strong anticancer and anti-inflammatory activities.

Perception of breast cancer risk factors: Dysregulation of TGF-ß/miRNA axis in Pakistani females.

Breast cancer poses a serious health risk for women throughout the world. Among the Asian population, Pakistani women have the highest risk of developing breast cancer. One out of nine women is diagnosed with breast cancer in Pakistan. The etiology and the risk factor leading to breast cancer are largely unknown. In the current study the risk factors that are most pertinent to the Pakistani population, the etiology, molecular mechanisms of tumor progression, and therapeutic targets of breast cancer are studied. A correlative, cross-sectional, descriptive, and questionnaire-based study was designed to predict the risk factors in breast cancer patients. Invasive Ductal Carcinoma (90%) and grade-II tumor (73.2%) formation are more common in our patient's data set. Clinical parameters such as mean age of 47.5 years (SD ± 11.17), disturbed menstrual cycle (> 2), cousin marriages (repeated), and lactation period (< 0.5 Y) along with stress, dietary and environmental factors have an essential role in the development of breast cancer. In addition to this in silico analysis was performed to screen the miRNA regulating the TGF-beta pathway using TargetScanHuman, and correlation was depicted through Mindjet Manager. The information thus obtained was observed in breast cancer clinical samples both in peripheral blood mononuclear cells, and biopsy through quantitative real-time PCR. There was a significant dysregulation (**P>0.001) of the TGF-ß1 signaling pathway and the miRNAs (miR-29a, miR-140, and miR-148a) in patients' biopsy in grade and stage specifically, correlated with expression in blood samples. miRNAs (miR-29a and miR-140, miR-148a) can be an effective diagnostic and prognostic marker as they regulate SMAD4 and SMAD2 expression respectively in breast cancer blood and biopsy samples. Therefore, proactive therapeutic strategies can be devised considering negatively regulated cascade genes and amalgamated miRNAs to control breast cancer better.

Tectona grandis leaf extract ameliorates hepatic fibrosis: Modulation of TGF- ß /Smad signaling pathway and upregulating MMP3/TIMP1 ratio.

ETHNOBOTANICAL RELEVANCE: Tectona grandis L.f (or syn: Jatus grandis (L.f.) Kuntze Revis), from family Lamiaceae, also known as Teak, is widely recognized in ayurvedic system of medicine and confer curative potential against inflammation, liver disorders, biliousness, diabetes, bronchitis, leprosy and dysentery. Its leaves are rich source of edible food colorant and reported nontoxic for liver and various organs. AIM OF STUDY: Hepatic injury progression to liver cirrhosis and cancer is a serious health issue across the world. Currently, anti-fibrotic therapeutic options are limited and expensive with no FDA approved direct anti-hepato-fibrotic drug validated in clinic. Thus, the aim of this study was to understand ameliorative effect of Tectona grandis L.f, leaves in early liver fibrosis. METHOD AND RESULTS: C57BL/6 mice suffering from CCl4 induced liver injury, were orally administered at three different doses (50, 100 & 200 mg/kg) of Tectona grandis L.f, leaf extract, thrice a week, up to 4 and 8 weeks. Anti-fibrotic effect was evaluated through animal body/liver weight measurements, serological tests (AST, ALT, GSH, MDA and LDH assays), tissue hydroxyproline content, and histochemical analysis (H&E, Masson trichrome, Sirius red and αSMA localization). Moreover, transcriptional and post-transcriptional expression of fibrosis associated biomarkers and TGF-ß/Smad cascade were analyzed. It was observed that 100 mg/kg dose optimally downregulated TGF-ß1/Smad2 with upregulation of Smad7 and regulated αSMA, Col 1, PDGF, TIMP1 and MMP3 expression, post 8 weeks of treatment. In addition, MMP3/TIMP1 ratio was upregulated to 0.7, 2.5 and 1.7 fold at 50 mg/kg, 100 mg/kg & 200 mg/kg treatments respectively, in comparison to untreated liver fibrosis models. The extract contains gallic acid, caffeic acid, sinapinic acid and myricetin when analyzed through high performance liquid chromatography. CONCLUSION: Tectona grandis L.f, leaves have potential to ameliorate liver fibrosis induced by CCl4 in mice via modulation of TGF-ß1/Smad pathway and upregulated MMP3/TIMP1 ratio.