Assessment cytotoxic assay of Rhizophora plants mangrove using brine shrimp (Artemia salina L) model

Rhizophoraceae is the main family of mangroves as a source of bioactive compounds originating from the coast. Ethnophamacologically Rhizophoraceae has been used in various traditional medicine. Natural sources as anticancer from the Rhizophoraceae family are interesting to know. This study aimed to determine the cytotoxic bioactivity of methanolic extracts of roots, bark, leaves, and fruit/hypocotyl from five species of Rhizophoraceae (Bruguieria cylindrica, B. gymnorrhiza, Ceriops decandra, Rhizophora apiculata, and R. mucronata) from the Langsa mangrove forest, Aceh. The method used in this study was the Brine Shrimp Lethality Test (BSLT) bioassay using Artemia salina Leach at extract concentrations of 1, 10, 100, 500, and 1000 μg/ml. Samples were extracted using the maceration method and methanol as the solvent. The cytotoxic activity of 20 Rhizophoraceae methanol extracts showed that 12 extracts were toxic with an LC50 range of 31.5 - 934.9 μg/ml (based on LC50 ≤ 1000 μg/ml). The two extracts of which the closest to highly toxic (based on LC50 ≤ 30 μg/ml) were C. decandra bark showed LC50 of 31.5 μg/ml, and R. mucronata bark showed LC50 31.8 μg/ml. This shows that Rhizophoraceae extract has potential as a natural anticancer agent. In the five rhizophoraceae species, C. decandra was the most active compared to other species. In the four plant parts, the bark was the most toxic.

Antioxidant and Cytotoxicity Activities of the Fermentation Extract of the Endophytic Fungi from the Marine Biota of Colt Coral

Indonesia has the largest and most biodiverse coral reef in the world. Colt coral has not been studied and explored especially endophytic fungi associated with the coral. Endophytic fungi are highly potential for the production of antioxidant and anticancer compounds. This research aimed to study the antioxidant and cytotoxic activities of fermentation extract from endophytic fungi from colt coral. Filtrate and mycelium extracts were obtained from static and shake fermentations of isolate SKF 15. Antioxidant and cytotoxic assays were conducted by free radical scavenger 1,1-diphenyl-2-picrylhydrazyl (DPPH) and AlamarBlue methods, respectively. The result showed that FD extract provided the highest antioxidant activity with inhibition of 49.36% at 200 ppm of DPPH. Variation of fermentation time (3-21 days) demonstrated the highest activity with inhibition of 66.97% for antioxidant assay (7 days) and 81.13% for cytotoxic assay (3 days). FTIR analysis presented the existence of hydroxyl groups O-H (3452.58 cm-1), C=C groups (1668.43 cm-1); C-O hydroxyl group (1230.58 cm -1), and C-H sp3 groups (2941.44 cm-1). Based on LC-MS analysis, FD extract has a mass of m/z 305.63, [M+H]+, predicted as dihydroquercetin (C15H24O7). Keywords: Antioxidant assay, cytotoxic assay, endophytic fungi, colt coral, DPPH method, AlamarBlue method

Combined inhibition of PD-1/PD-L1, Lag-3, and Tim-3 axes augments antitumor immunity in gastric cancer–T cell coculture models

Background Immunotherapy targeting PD-1 provides a limited survival benefit in patients with unresectable advanced or recurrent gastric cancer (GC). Beside PD-L1, the expression of inhibitory ligands such as CEACAM-1 and LSECtin on GC cells account for this limitation. Here we assessed their expression and immune suppressive effect in GC patients. Methods Using multiplexed immunohistochemistry staining, we evaluated the distribution of different inhibitory ligands, including PD-L1, CEACAM-1, LSECtin, and MHC class II, in 365 GC patients. We analyzed their correlations and overall survival (OS) based on the expression of each inhibitory ligand and the independent prognostic factors that affect OS. Subsequently, we evaluated the additive effect of anti-PD-1 mAb or anti-PD-L1 mAb with/without anti-Lag-3 mAb with/without anti-Tim-3 mAb in cytotoxic assay using tumor-antigen specific CTL clones against GC cell lines. Results Co-expression of the inhibitory ligands for PD-1, Tim-3, and Lag-3 was observed in the largest proportion (34.7%). CEACAM-1, LSECtin, and MHC class II expression showed significant correlation with PD-L1 expression and OS. Multivariable analysis demonstrated that CEACAM-1 low is an independent prognostic factor. Furthermore, combining dual and triple ICIs yielded additive effect on cytotoxicity of CTL clones against each immune inhibitory ligand positive GC cell lines. Conclusions Our findings suggested that the expression of inhibitory ligands for Tim-3 and Lag-3 on GC cells serve as potential biomarkers to predict the response to anti-PD-1 therapy and the combinatorial immunotherapy with ICIs targeting for PD-1, Tim-3, and Lag-3 has a therapeutic potential for GC patients.

PHYTOCHEMICAL COMPOSITION AND ANTIINFLAMMATORY ACTIVITY OF NELUMBO NUCIFERA GAERTN

Objective: Inflammation is a process of injuries caused by physical, chemical, and biological factors. Nitric oxide (NO) plays an important role in theregulation of various pathological and pathophysiological processes. Overproduction of NO induces tissue damage associated with acute and chronicinflammations. This study was conducted to determine the phytochemical composition and the NO inhibitory properties of Nelumbo nucifera extractsin lipopolysaccharide(LPS)-stimulated macrophage cell line.Methods: The dried leaf, stalk, and flower materials of roseum plenum and album plenum (AP) were extracted with 95% ethanol solvents. Thephytochemical compounds of the extraction were analysed by gas chromatography-mass spectrometry. The cytotoxic assay of extracts againstmacrophage cells was conducted using resazurin. The NO was determined using LPS-induced RAW264.7 cells to measure inhibitory activity of extracton the production of NO.Results: The extracts from Lotus, which exhibited the non-cytotoxic to the RAW264.7 cells. The AP-stalk extracts were capable to reduce the NO levelin LPS-activated RAW264.7 cells. GC-MS analysis of AP-stalk extraction revealed pharmacologically active compounds.Conclusion: The results conduct that the AP-stalk extract effectively inhibited the NO production and may be useful in preventing inflammatorydiseases mediated by excessive production of NO. Bio-active phytoconstituents from AP stalk extract could potentially be used for anti-inflammation.These data also suggest that AP-stalk extract may serve as a good indicator of the pharmacological activities of medicinal plants.

Am Synthesis, Docking Study and In vitro Anticancer Evaluation of Some New Flurbiprofen Derivatives Against MCF-7 and WRL-68 Cell Lines

A new series of flurbiprofen derivatives containing thiosemicarbazide moiety (3-7) was synthesized from flurbiprofen as parent nucleus by esterification, hydrazide formation, and heating with different aryl isothiocyanate substituents, respectively. Flurbiprofen was also treated with thiosemicarbazide in the presence POCl3 as a catalyst, to produce 1,3,4 -thiadiazole -2-amine (8). Treatment of (8) with different aryl isothiocyanates produced thiourea derivatives (9-12). Also, the reaction of (8) with different benzoyl chloride substituents produced benzamide compounds (13-15). Eventually , treatment of (8) with ethyl acetoacetate(EAA) produced [1,3,4]thiadiazolo[3,2-a]pyrimidin-7-one (16) .The new compounds were characterized by spectroscopic techniques :FTIR, 1HNMR, and CHNS analysis. A molecular docking study for the synthesized compounds (3-16), against the Vascular Endothelial Growth Factor receptor (VEGFR-2) was applied and it indicated that compounds 4,7,13, and 15,exhibited the optimum binding energy of -6.77, -6.12,-6.68, and -6.43 kcal/mol, respectively. Target compounds were also assessed for their in vitro anticancer effects in a cell-line study. All of the compounds tested showed the most plausible anticancer activity, compared to a positive control(Sorafenib), using in vitro MTT cytotoxic assay ,against human breast tumor (MCF-7), and normal WRL-68 cell line. The in vitro results revealed that compounds 4,5,10,11,13, and 15 exhibited the highest inhibitory activity at their IC50 concentrations, against MCF-7 cell lines, as follows (122.7,113.9,95,7. 109.1,40.32 and 112.29µg/mL, respectively. While their cytotoxic effect against normal WRL-68 cell line at their IC50 concentrations, as follow 210.2, 181.3 ,151.7,278.7,80.28, and 236 µg/mL, respectively, therefore, such compounds were considered more selective toward MCF-7 than normal WRL-68,and their selectivity index (SI): 1.71,1.59,1.59 ,2.55 ,1.99 , and 2.10,respectively . Among the synthesized compounds, the compound 15 was chosen to screen its effect in vitro through multi-parameter cytotoxic assay against MCF-7 breast cancer implemented in High Content Screening (ArrayScan XTI, Thermo Scientific),which could be taken in consideration as a starting point for the development of new anticancer drugs

Synthesis, Cytotoxicity and Docking Simulation of Novel Annulated Dihydroisoquinoline Heterocycles

Objective: Coupling of ethyl 2-(6,7-dimethoxy-3,4-dihydroisoquinolin-1-yl)acetate 2 with diazotized anilines in ethanol in the presence of sodium acetate yielded 2-(2-arylhydrazono)-2-(6,7- dimethoxy-3,4-dihydroisoquinolin-1-yl)acetate (4a-f). Methods: Treatment of 2 with α-bromoketones 6a-f in dry benzene at reflux gave the corresponding isoquinolinium bromides 7a-f. Refluxing of each of the salts 7a-f in dry benzene and in the presence of triethylamine yielded 2-arylpyrrolo-[2,1-a]isoquinoline structures 8a-f, that converted to ethyl (E)-8,9- dimethoxy-3-(phenyldiazen-yl)-2-(aryl)-5,6-dihydropyrrolo[2,1-a]isoquinoline-1-carboxylate (9a-f) upon treatment with diazotized anilines 3 in ethanol in the presence of sodium acetate. Results and Conclusion: Cytotoxic assay was performed for in vitro antitumor screening against caucasian breast adenocarcinoma (MCF7), hepatocellular carcinoma (HepG2) and colorectal carcinoma (HCT-116) cell lines. The results were compared with the standard anticancer drug (doxorubicin). Molecular docking using MOE 2014.09 software was carried out for the most potent compound 4d, which showed the highest binding affinity towards the four tested proteins and thus initiated apoptosis of cancer cells.

Cisplatin Nanoparticles for Folate Targeted Delivery in Ovarian Cancer: Study in SKOV-3 Cancer Cells

Tumor targeting of drug to specific site via use of folate is one the choicest method for enhancing specificity of Cisplatin. Chitosan was conjugated with folate via amino acylation method and Folate-chitosan-Cisplatin-nanoparticles (FA-CS-Cis-NP's) were prepared using ionotropic gelation method. Thus prepared nanoparticles were evaluated for various parameters such as particles size, % encapsulation efficiency, % drug release, cytotoxic assay, cellular uptake. The average particle size of nanoparticles was found to be 265 nm with encapsulation efficiency of 89.43%. The cumulative release of drug from dosage form was found to be 82.31% for 240 min. The higher level of intra-cellular accumulation of Cisplatin was found to be in case of FA-CS-Cis-NP's than conventional drug delivery when cultured with folate. The results indicate that targeting of FA-CSCis-NP's is possible with folate targeting and are proved as potential folate receptor positive tumor cell targeting drug delivery system.