Induction of apoptosis by a synergistic lignand composition from Cedrus deodarain human cancer cells

AP9-cd, a synergistic lignan mixture from Cedrus deodara (Pinaceae) consisting of (−)-wikstromal, (−)-matairesinol and dibenzyl butyrolactol, depicted cytotoxic effects against numerous human cancer cell lines reported previously.The aim of this study was to investigate the mechanism of cell death in human cancer cells. The viability, morphological and ultrastructural changes in Molt-4 cells were investigated. Using the trypan blue exclusion assay, we demonstrated that AP9-cd significantly reduced the viability of Molt-4 cells in a time- and dose-dependent manner. Apoptotic assays using light microscopy revealed that this agent induced Molt-4 cell apoptosis at varied concentrations. The treatment causes a loss in cell viability by activating the apoptotic process as identified by light and electron microscopy. The morphological changes of intracellular organelles in Molt-4 cells treated with 30 μg/ml of AP9-cd revealed the disruption of mitochondrial cristae. Other features included the vacuolization, chromatin condensation and formation of micronuclei. Surface ultrastructural studies of four different tumor cell lines (Molt-4, HL-60, PC-3 and A-549) treated with AP9-cd depicted loss of surface projections, condensation and formation of apoptotic bodies. AP9-cd treatment to transgenic fruit fly, Drosophila, carrying human adenomatous polpyposis coli (hAPC) gene enhanced eye phenotypes and therefore may inhibit Wnt/Wg pathway which is important in the aetiology of a number of human cancers

Dynamics of essential oil biosynthesis in relation to inflorescence and glandular ontogeny inSalvia sclarea

Changes in the essential oil concentration, composition and glandular morphology of Salvia sclarea L. were studied at different stages of inflorescence maturity. The chemical composition of oil was determined by GC–MS, NMR and the peak enrichment technique. The oil yield at bud stage on a fresh basis was minimum (0.08%), peaked at full bloom stage (0.18%) and was followed by sharp decline on maturation (0.07%). The main components of the oil were linalool (36.6–41.9%) and linalyl acetate (13.2–19.2%). The maximum percentage composition of various constituents was coincident with full bloom stage. β -Humulene (6.4–8.9%), α -cadinene (t–1.5%), β -caryophyllene (t–1.4%), β - caryophyllene oxide (0.4–1.4%) and sclareol (0.3–1.8%) present in the oil showed gradual increase in percentage over the different stages of maturity, with no significant turnover losses at maturation stage. Turnover of essential oil (61.1% loss) and monoterpenes (23.6% loss) occurred late in development at full maturity. Scanning electron microscopy (SEM) was used to follow the changes in the oil secretory glands over different temporal phases of maturation. The decline in oil concentration and monoterpene constituents compared very well with the observed deterioration and lyses of secretory glandular system. An abrupt fall in oil concentration apparently appears due to differential evaporation of the more volatile constituents, rather than as a dynamic balance between biosynthetic and catabolic processes

Boeravinone B, A Novel Dual Inhibitor of NorA Bacterial Efflux Pump of Staphylococcus aureus and Human P-Glycoprotein, Reduces the Biofilm Formation and Intracellular Invasion of Bacteria

This study elucidated the role of boeravinone B, a NorA multidrug efflux pump inhibitor, in biofilm inhibition. The effects of boeravinone B plus ciprofloxacin, a NorA substrate, were evaluated in NorA-overexpressing, wild-type, and knocked-out Staphylococcus aureus (SA-1199B, SA-1199, and SA-K1758, respectively). The mechanism of action was confirmed using the ethidium bromide accumulation and efflux assay. The role of boeravinone B as a human P-glycoprotein (P-gp) inhibitor was examined in the LS-180 (colon cancer) cell line. Moreover, its role in the inhibition of biofilm formation and intracellular invasion of S. aureus in macrophages was studied. Boeravinone B reduced the minimum inhibitory concentration (MIC) of ciprofloxacin against S. aureus and its methicillin-resistant strains; the effect was stronger in SA-1199B. Furthermore, time–kill kinetics revealed that boeravinone B plus ciprofloxacin, at subinhibitory concentration (0.25 × MIC), is as equipotent as that at the MIC level. This combination also had a reduced mutation prevention concentration. Boeravinone B reduced the efflux of ethidium bromide and increased the accumulation, thus strengthening the role as a NorA inhibitor. Biofilm formation was reduced by four–eightfold of the minimal biofilm inhibitory concentration of ciprofloxacin, effectively preventing bacterial entry into macrophages. Boeravinone B effectively inhibited P-gp with half maximal inhibitory concentration (IC50) of 64.85 μM. The study concluded that boeravinone B not only inhibits the NorA-mediated efflux of fluoroquinolones but also considerably inhibits the biofilm formation of S. aureus. Its P-gp inhibition activity demonstrates its potential as a bioavailability and bioefficacy enhancer

Synthesis and cytotoxity of novel benzopyran derivatives

1619-1629In-vitro cytotoxic activity of benzopyrones and benzopyran derivatives has been evaluated against a panel of several human cancer cell lines. Out of 16 compounds prepared and screened, four compounds have shown significant cytotoxicity against human breast, liver, colon and prostrate cancer cell lines. Apoptosis rather than necrosis has been established as the mode of mechanism for the cytotoxicity of the studied molecules using Light Microscopic and Scanning Electron Microscopic techniques

Anticancer activity of an essential oil from Cymbopogon flexuosus

The essential oil froma lemon grass variety of Cymbopogon flexuosuswas studied for its in vitro cytotoxicity against twelve human cancer cell lines. The in vivo anticancer activity of the oil was also studied using both solid and ascitic Ehrlich and Sarcoma-180 tumor models in mice. In addition, the morphological changes in tumor cells were studied to ascertain the mechanism of cell death. The in vitro cytotoxicity studies showed dose-dependent effects against various human cancer cell lines. The IC50 values of oil ranged from 4.2 to 79�g/ml depending upon the cell line. In 502713 (colon) and IMR-32 (neuroblastoma) cell lines, the oil showed highest cytotoxicity with IC50 value of 4.2 and 4.7�g/ml, respectively. Intraperitoneal administration of the oil significantly inhibited both ascitic and solid forms of Ehrlich and Sarcoma-180 tumors in a dose-dependent manner. The tumor growth inhibition at 200 mg/kg (i.p.) of the oil observed with both ascitic and solid tumor forms of Ehrlich Ascites carcinoma was 97.34 and 57.83 respectively. In case of Sarcoma-180, the growth inhibition at similar dose of oil was 94.07 and 36.97% in ascitic and solid forms respectively. Morphological studies of the oil treated HL-60 cells revealed loss of surface projections, chromatin condensation and apoptosis. The mitochondria showed apparent loss of cristae in the cells undergoing apoptosis. The morphological studies of Sarcoma-180 solid tumor cells from animals treated with the oil revealed condensation and fragmentation of nuclei typical of apoptosis. Morphological studies of ascites cells from animals treated with the oil too revealed the changes typical of apoptosis. Our results indicate that the oil has a promising anticancer activity and causes loss in tumor cell viability by activating the apoptotic process as identified by electron microscopy

6-Nitro-2-(3-hydroxypropyl)-1H-benz[de]isoquinoline-1,3-dione, a potent antitumor agent, induces cell cycle arrest and apoptosis

<p>Abstract</p> <p>Background</p> <p>Anticancer activities of several substituted naphthalimides (1H-benz[de]isoquinoline-1,3-diones) are well documented. Some of them have undergone Phase I-II clinical trials. Presently a series of ten N-(hydroxyalkyl) naphthalimides (compounds <b>1a-j) </b>were evaluated as antitumor agents.</p> <p>Methods</p> <p>Compounds <b>1a-j </b>were initially screened in MOLT-4, HL-60 and U-937 human tumor cell lines and results were compared with established clinical drugs. Cytotoxicities of compounds <b>1d </b>and <b>1i </b>were further evaluated in a battery of human tumor cell lines and in normal human peripheral blood mononuclear cells. Cell cycle analysis of compound <b>1i </b>treated MOLT-4 cells was studied by flow cytometry. Its apoptosis inducing effect was carried out in MOLT-4 and HL-60 cells by flow cytometry using annexin V-FITC/PI double staining method. The activities of caspase-3 and caspase-6 in MOLT-4 cells following incubation with compound <b>1i </b>were measured at different time intervals. Morphology of the MOLT-4 cells after treatment with <b>1i </b>was examined under light microscope and transmission electron microscope. ³H-Thymidine and ³H-uridine incorporation in S-180 cells in vitro following treatment with 8 μM concentration of compounds <b>1d </b>and <b>1i </b>were studied.</p> <p>Results</p> <p>6-Nitro-2-(3-hydroxypropyl)-1H-benz[de]isoquinoline-1,3-dione (compound <b>1i</b>), has exhibited maximum activity as it induced significant cytotoxicity in 8 out of 13 cell lines employed. Interestingly it did not show any cytotoxicity against human PBMC (IC₅₀value 273 μM). Cell cycle analysis of compound <b>1i </b>treated MOLT-4 cells demonstrated rise in sub-G₁fraction and concomitant accumulation of cells in S and G₂/M phases, indicating up-regulation of apoptosis along with mitotic arrest and/or delay in exit of daughter cells from mitotic cycle respectively. Its apoptosis inducing effect was confirmed in flow cytometric study in MOLT-4 and the action was mediated by activation of both caspase 3 and 6. Light and transmission electron microscopic studies corroborated its apoptosis inducing efficacy at a concentration of 10 μM in MOLT-4 cells. Its apoptosis induction was also observed in HL-60 cells to an extent much greater than well known apoptosis inducing agents as camptothecin and cis-platin at 10 μM concentration each. It significantly inhibited DNA and RNA synthesis in S-180.</p> <p>Conclusions</p> <p>In essence, compound <b>1i </b>showed potential as an antitumor agent.</p

Potentiation of the antitumor effect of 11-keto-β-boswellic acid by its 3-α-hexanoyloxy derivative

We recently discovered that a propionyloxy derivative of 11-keto-β-boswellic acid (PKBA) showed better anticancer potential than other boswellic acids including AKBA, encompassing the importance of acyl group at the 3-α-hydroxy position of KBA. In continuation of our previous work, other higher derivatives (with increasing alkoxy chain length at 3-α-hydroxy position) including butyryloxy (BKBA) and hexanoyloxy(HKBA) derivatives of KBA were synthesized. The respective IC50 values of BKBA and HKBA in HL-60 cells were found to be 7.7 and 4.5 μg/ml. IC50 value of HKBA was comparatively lower than that of BKBA, and further lower than that of the previously reported derivative (PKBA, IC50 8.7 μg/ml). In order to compare the anticancer potential of HKBA with PKBA, detailed in vitro pro-apoptotic and in vivo anticancer studies were carried out. The induction of apoptosis by HKBA was measured using various parameters including fluorescence and scanning electron microscopy, DNA fragmentation and Annexin V-FITC binding. The extent of DNA damage was measured using neutral comet assay. HKBA was further evaluated for its effect on DNA cell cycle and mitochondria where it was found to arrest cells in G2/M phase and also induced loss of mitochondrial membrane potential. These events were associated with increased expression of cytosolic cytochrome c and cleavage of PARP. Target based studies showed that HKBA inhibited the enzymatic activity of topoisomerases I and II at low doses than that of PKBA. In vivo studies also revealed a low dose inhibitory effect of HKBA on ascitic and solid murine tumor models

Dual role of Par-4 in abrogation of EMT and switching on mesenchymal to epithelial transition (MET) in metastatic pancreatic cancer cells

Epithelial-mesenchymal transition (EMT) is a critical event that occurs during the invasion and metastatic spread of cancer cells. Here, we conceive a dual mechanism of Par-4-mediated inhibition of EMT and induction of MET in metastatic pancreatic cancer cells. First, we demonstrate that 1,1′-β-D-glucopyranosyl-3,3′-bis(5-bromoindolyl)-octyl methane (NGD16), an N-glycosylated derivative of medicinally important phytochemical 3,3′-diindolylmethane (DIM) abrogates EMT by inducing pro-apoptotic protein Par-4. Induction of Par-4 (by NGD16 or ectopic overexpression) strongly impedes invasion with inhibition of major mesenchymal markers viz. Vimentin and Twist-1 epithelial marker- E-cadherin. Further, NGD16 triggers MET phenotypes in pancreatic cancer cells by augmenting ALK2/Smad4 signaling in a Par-4-dependent manner. Conversely, siRNA-mediated silencing of endogenous Par-4 unveil reversal of MET with diminished E-cadherin expression and invasive phenotypes. Additionally, we demonstrate that intact Smad4 is essential for Par-4-mediated maintenance of E-cadherin level in MET induced cells. Notably, we imply that Par-4 induction regulates E-cadherin levels in the pancreatic cancer cells via modulating Twist-1 promoter activity. Finally, in vivo studies with syngenic mouse metastatic pancreatic cancer model reveal that NGD16 strongly suppresses metastatic burden, ascites formation, and prolongs the overall survival of animals effectively