Chitinase inhibits growth of human breast and colorectal cancer cells in vitro and Ehrlich ascites carcinoma cells in vivo

Trichosanthes dioica seed extract was loaded on a QA-cellulose column and the unbound fraction with the chitinase activity was run on SDS-PAGE. Multiple bands were observed and were separated by a Sephadex G-50 column. The combination of the 6 and 33 kDa masses supported the degradation of chitinase as purified earlier. Only the 33 kDa fraction contained sugar and showed chitinase activity. The chitinase was also isolated by using a chitin column. At 200 µg/ml protein concentration, the chitinase inhibited 49.1 %, 48.8 % and 38.12 % of Ehrlich ascites carcinoma, HCT-116 and MCF-7 cells growth, respectively, in a dose-dependent manner. Exactly, 46 % and 82 % EAC cell growth inhibition were observed after treating the EAC cells bearing Swiss albino mice with the chitinase at the doses of 1.0 and 2.0 mg/Kg/day respectively. EAC, HCT-116 and MCF-7 cells growth inhibitions were due to the induction of apoptosis. ROS was accumulated in HCT-116 and MCF-7 cells. After treatment of HCT-116 cells, the expression level of p53 and TNFα genes increased and PARP gene decreased. On the other hand, elevated expression was observed for PARP, MAPK, NFκB, FAS, FADD, and Caspase-8 genes in MCF-7 cells. The induction of apoptosis in HCT-116 was further confirmed by caspase protein expression. The chitinase causes ‘S’ cell cycle arrest in MCF-7 and HCT-116 cells. T. dioica seed chitinase inhibited EAC, HCT-116 and MCF-7 cells by inducing apoptosis in vitro and EAC in vivo in mice. These promising results indicated that T. dioica seed chitinase can be an anticancer agent.     

Antibacterial Activity of Kaempferia rotunda Rhizome Lectin and Its Induction of Apoptosis in Ehrlich Ascites Carcinoma Cells

The tuberous rhizome Kaempferia rotunda Linn. has been used as food and traditional medicinal plant, and the purified K. rotunda lectin (KRL) showed antiproliferative activity against Ehrlich ascites carcinoma cells [1]. In the present study, KRL showed agglutination activity against Escherichia coli and Staphylococcus aureus, with partial inhibition of their growth. MTT assay was used to investigate the effect of KRL on EAC cells in vitro in RPMI-1640 medium, and it was found that lectin inhibited 6.2–50.5 % cell growth at the range of 7.5–120 μg/ml protein concentration. The cell cycle arrest at G₀/G₁ phase of EAC cells was also determined by flow cytometry after treatment with lectin. The apoptotic cell morphological changes of the treated EAC cells were confirmed by fluorescence and optical microscope. In the presence of caspase-3 inhibitor, the cell growth inhibition of the lectin was reduced significantly. RT-PCR was used to evaluate the expression of apoptosis-related genes, bcl-2, bcl-X, and bax. Bax gene expression was intensively increased with the despaired of bcl-X gene expression and significant decrease of bcl-2 gene expression in the cells treated with KRL. Thus, lectin induced apoptotic cell death in Ehrlich ascites carcinoma cells.     

Biosynthesis of Silver Chloride Nanoparticles by Rhizospheric Bacteria and Their Antibacterial Activity against Phytopathogenic Bacterium Ralstonia solanacearum

Ralstonia solanacearum is the most destructive pathogen, causing bacterial wilt disease of eggplant. The present study aimed to develop green synthesis and characterization of silver chloride nanoparticles (AgCl-NPs) by using a native bacterial strain and subsequent evaluation of their antibacterial activity against R. solanacearum. Here, a total of 10 bacterial strains were selected for the biosynthesis of AgCl-NPs. Among them, the highest yield occurred in the synthesis of AgCl-NPs using a cell-free aqueous filtrate of strain IMA13. Ultrastructural observation revealed that the AgCl-NPs were spherical and oval with smooth surfaces and 5–35 nm sizes. XRD analysis studies revealed that these particles contained face-centered cubic crystallites of metallic Ag and AgCl. Moreover, FTIR analysis showed the presence of capping proteins, carbohydrates, lipids, and lipopeptide compounds and crystalline structure of AgCl-NPs. On the basis of phylogenetic analysis using a combination of six gene sequences (16S, gyrA, rpoB, purH, polC, and groEL), we identified strain IMA13 as Bacillus mojavensis. Three kinds of lipopeptide compounds, namely, bacillomycin D, iturin, and fengycin, forming cell-free supernatant produced by strain IAM13, were identified by MALDI-TOF mass spectrometry. Biogenic AgCl-NPs showed substantial antibacterial activity against R. solanacearum at a concentration of 20 µg/mL⁻¹. Motility assays showed that the AgCl-NPs significantly inhibited the swarming and swimming motility (61.4 and 55.8%) against R. solanacearum. Moreover, SEM and TEM analysis showed that direct interaction of AgCl-NPs with bacterial cells caused rupture of cell wall and cytoplasmic membranes, as well as leakage of nucleic acid materials, which ultimately resulted in the death of R. solanacearum. Overall, these findings will help in developing a promising nanopesticide against phytopathogen plant disease management.     

Synthesis of bis[benzyl‐N′‐hydrazinecarbodithioato‐κ2 N′,S]nickel(II) complex as a novel lead molecule for cancer treatment

A novel bis[benzyl‐N′‐hydrazinecarbodithioato‐κ² N′,S]nickel(II) complex was synthesized and characterized by means of various physical, chemical, and spectroscopic techniques. The X‐ray single crystal diffraction analysis indicated two independent close comparable bis‐chelated square planar complexes of trans‐configuration, where S‐benzyl dithiocarbazate (SBDTC) ligand is coordinated via N,S‐donor set. The complex is able to inhibit Ehrlich ascites carcinoma (EAC) cell proliferation by 51.81% and 75.75%, with 0.3 and 50 mg kg^?1 (dose adjusted) dose, respectively, administered intraperitoneally for five successive days in mice model. Apoptotic cell morphological changes were examined using optical and fluorescence microscopy techniques. Expression pattern of apoptosis regulatory genes in EAC cells treated with the synthesized nickel(II) complex for five consecutive days showed an increased expression of P⁵³, Bax, Cas‐8, Cas‐9, Cas‐3, Cyt‐c, and TNF‐α proapoptotic genes and decreased expression of antiapoptotic Bcl‐2 gene. The Ni(II) complex and Bleomycin (standard drug) were used in molecular docking coupled with molecular dynamics simulation studies with the aim to support the experimental results and to investigate the apoptotic effect towards the targeting apoptotic genes. Both experimental and computational studies reveal that the nickel(II) complex inhibits EAC cells growth successfully, suggesting a potential compound for cancer treatment.     

In vitro and In vivo Antiproliferative Effect of a Combination of Ultraviolet‐A and Alkoxy Furocoumarins Isolated from Umbelliferae Medicinal Plants,in Melanoma Cells

We examined the effects of six furocoumarins with alkoxy groups at the C‐5 or C‐8 position isolated from Umbelliferae medicinal plants on cell proliferation, and their mechanisms of action against B16F10 melanoma cells or in melanin‐possessing hairless mice implanted with B16F10 cells, under UVA irradiation. Three furocoumarins with an alkoxy group at C‐5, isoimperatorin (1), oxypeucedanin (2) and oxypeucedanin hydrate (3), showed antiproliferative activity and caused G₂/M arrest at concentrations of 0.1–10.0 μm . Furthermore, three furocoumarins with an alkoxy group at C‐8, imperatorin (4), heraclenin (5) and heraclenol (6), inhibited the proliferation of melanoma cells and cell cycle at G₂/M at concentrations of 0.1–1.0 μm . UVA plus 1, 2, 3, 4 and 6 reduced tumor growth and final tumor weight in B16F10‐bearing mice at a dose of 0.3, 0.5 or 1.0 mg kg^?1 (intraperitoneal injection). UVA plus 1, 3 and 6 increased Chk1 phosphorylation and reduced cdc2 (Thr 161) phosphorylation in melanoma cells. We suggest that the antitumor actions of UVA plus furocoumarins with an alkoxy group at C‐5 or C‐8 were due to G₂/M arrest of the cell cycle by an increase in phosphor‐Chk1 and decrease in phospho‐cdc2.     

Expression of GD3 synthase modifies ganglioside profile and increases migration of MCF-7 breast cancer cells

B- and c-series of gangliosides are over-expressed in neuro-ectoderm-related cancers, including breast cancer. It has been shown that G_D3 ganglioside is over-expressed in about 50% of invasive ductal breast carcinoma and the G_D3 synthase (GD3S) gene displays higher expression among estrogen receptor (ER) negative breast tumors. We previously showed that GD3S expression in MDA-MB-231 breast cancer cells induces the expression of G_D2 and increased cell proliferation and migration via a G_D2-dependent activation of c-Met receptor. Here, we show that in ER-positive MCF-7 breast cancer cells, GD3S expression resulted in an increase of G_D1b, which was associated with a decrease of G_M1a and G_M2. Meanwhile, GD3S expressing MCF-7 cells exhibited an increased migration without any modification of proliferation rate. Therefore, GD3S expression can result in different modifications of both ganglioside profiles and cell phenotypes depending on breast cell types.     

The Bioflavonoid Galangin Suppresses the Growth of Ehrlich Ascites Carcinoma in Swiss Albino Mice: A Molecular Insight

Bioflavonoids are plant compounds touted for their potential to treat or prevent several diseases including cancer caused by various stress conditions. Galangin (4H-1-Benzopyran-4-one, 3, 5, 7-trihydroxy-2-phenyl-), a flavonoid, is a polyphenolic compound found primarily in medicinal herb, Alpinia galanga. This study aims to demonstrate the galangin as a pharmacological lead compound using in vitro, in vivo, and in silico model targeting specific cancer condition and proteins. The proliferation of MCF-7 and Ehrlich ascites carcinoma (EAC) cells was significantly inhibited with an IC₅₀ of 34.11 and 22.29???g/ml, respectively. In an animal model system, galangin has inhibited the tumor growth by 73.51%?±?4.742 in EAC-induced Swiss Albino mice with no evidences of mortality as compared to standard drug, 5-fluorouracil. The effectiveness of galangin is proven in an animal system suggesting its pharmacokinetics behavior in an animal model which is also complemented by outcome of in silico analysis with more than 88?% of human intestinal absorption and significant Caco-2 cell, MDCK cell, and skin permeability as predicted by in silico methods. Galangin was docked against 19 different proteins involved in tumorogenesis and apoptosis; the energetic analysis indicates that it exhibits higher predicted binding free energy of ?12.7?kcal/mol with Bcl-xL protein.     

Pogostemon cablin Triggered ROS-Induced DNA Damage to Arrest Cell Cycle Progression and Induce Apoptosis on Human Hepatocellular Carcinoma In Vitro and In Vivo

The purpose of the study was to elucidate the anti-hepatoma effects and mechanisms of Pogostemon cablin essential oils (PPa extract) in vitro and in vivo. PPa extract exhibited an inhibitory effect on hepatocellular carcinoma (HCC) cells and was less cytotoxic to normal cells, especially normal liver cells, than it was to HCC cells, exerting a good selective index. Additionally, PPa extract inhibited HCC cell growth by blocking the cell cycle at the G₀/G₁ phase via p53 dependent or independent pathway to down regulated cell cycle regulators. Moreover, PPa extract induced the FAS-FASL-caspase-8 system to activate the extrinsic apoptosis pathway, and it increased the bax/bcl-2 ratio and reduced ΔΨm to activate the intrinsic apoptosis pathway that might be due to lots of reactive oxygen species (ROS) production which was induced by PPa extract. In addition, PPa extract presented to the potential to act synergistically with sorafenib to effectively inhibit HCC cell proliferation through the Akt/mTOR pathway and reduce regrowth of HCC cells. In an animal model, PPa extract suppressed HCC tumor growth and prolonged lifespan by reducing the VEGF/VEGFR axis and inducing tumor cell apoptosis in vivo. Ultimately, PPa extract demonstrated nearly no or low system-wide, physiological, or pathological toxicity in vivo. In conclusion, PPa extract effectively inhibited HCC cell growth through inducing cell cycle arrest and activating apoptosis in vitro and in vivo. Furthermore, PPa extract exhibits less toxicity toward normal cells and organs than it does toward HCC cells, which might lead to fewer side effects in clinical applications. PPa extract may be developed into a clinical drug to suppress tumor growth or functional food to prevent HCC initiation or chemoprotection of HCC recurrence.     

Engineered Bacteriorhodopsin May Induce Lung Cancer Cell Cycle Arrest and Suppress Their Proliferation and Migration

Highly expressible bacteriorhodopsin (HEBR) is a light-triggered protein (optogenetic protein) that has seven transmembrane regions with retinal bound as their chromophore to sense light. HEBR has controllable photochemical properties and regulates activity on proton pumping. In this study, we generated HEBR protein and incubated with lung cancer cell lines (A549 and H1299) to evaluate if there was a growth-inhibitory effect with or without light illumination. The data revealed that the HEBR protein suppressed cell proliferation and induced the G₀/G₁ cell cycle arrest without light illumination. Moreover, the migration abilities of A549 and H1299 cells were reduced by ~17% and ~31% after incubation with HEBR (40 μg/mL) for 4 h. The Snail-1 gene expression level of the A549 cells was significantly downregulated by ~50% after the treatment of HEBR. In addition, HEBR significantly inhibited the gene expression of Sox-2 and Oct-4 in H1299 cells. These results suggested that the HEBR protein may inhibit cell proliferation and cell cycle progression of lung cancer cells, reduce their migration activity, and suppress some stemness-related genes. These findings also suggested the potential of HEBR protein to regulate the growth and migration of tumor cells, which may offer the possibility for an anticancer drug.     

Polyphenol Oxidase as a Promising Alternative Therapeutic Agent for Cancer Therapy

Cancers have always been the most difficult to fight, the treatment of cancer is still not considered. Thus, exploring new anticancer drugs is still imminent. Traditional Chinese medicine has played an important role in the treatment of cancer. Polyphenol oxidase (PPO) extracted from Edible mushroom has many related reports on its characteristics, but its role in cancer treatment is still unclear. This study aims to investigate the effects of PPO extracted from Edible mushroom on the proliferation, migration, invasion, and apoptosis of cancer cells in vitro and explore the therapeutic effects of PPO on tumors in vivo. A cell counting kit-8 (CCK8) assay was used to detect the effect of PPO on the proliferation of cancer cells. The effect of PPO on cancer cell migration ability was detected by scratch test. The effect of PPO on the invasion ability of cancer cells was detected by a transwell assay. The effect of PPO on the apoptosis of cancer cells was detected by flow cytometry. Female BALB/c mice (18–25 g, 6–8 weeks) were used for in vivo experiments. The experiments were divided into control group, model group, low-dose group (25 mg/kg), and high-dose group (50 mg/kg). In vitro, PPO extracted from Edible mushroom significantly inhibited the proliferation, migration, and invasion capability of breast cancer cell 4T1, lung cancer cell A549, and prostate cancer cell C4-2, and significantly promoted the apoptosis of 4T1, A549, and C4-2. In vivo experiments showed PPO inhibitory effect on tumor growth. Collectively, the edible fungus extract PPO could play an effective role in treating various cancers, and it may potentially be a promising agent for treating cancers.     

pIL-12 delivered by polymer based nanovector for anti-tumor genetherapy

Finding more effective and safe non-viral vectors to transfer genes into cancer cells has become the key of immune gene therapy for cancer. Herein a triblock compound MPEG₂₀₀₀–PDLLA₄₀₀₀–MPEG₂₀₀₀ modified by cationic liposome DOTAP was used as a non-viral vector DOTAP/MPEG₂₀₀₀–PDLLA₄₀₀₀–MPEG₂₀₀₀ (DMPM) to effectively transfer interleukin (IL)-12 plasmid (pIL-12) into tumor tissue. IL-12 produced by transfected tumor cells successfully inducing lymphocyte proliferation and promoting interferon-γ (IFN-γ) secretion, which resulted in tumor cells death. The ability of DMPM to transfer pIL-12 and the immune effect induced by IL-12 in cells had been explored. The anti-tumor effect, mechanism and safety of pIL-12/DMPM in mice cancer model were investigated in this study. Our results showed that the pIL-12 transferred by DMPM was highly expressed both in CT26 cells and B16-F10 cells. IL-12 expressed in the culture supernatant of transfected tumor cells stimulated lymphocyte proliferation and promoted IFN-γ secretion. The experimental result confirmed that pIL-12/DMPM therapy significantly reduced tumor growth in mice model. We designed the nanocomposite DMPM to deliver pIL-12 for cancer treatment and explored its therapeutic efficacy and the underlying anti-tumor mechanism. Our study suggested pIL-12 loaded by DMPM complex would be an effective strategy for cancer treatment.     

Enhancement of antitumor effect using dendritic cells activated with natural killer cells in the presence of Toll-like receptor agonist

Dendritic cells (DCs) play a role in natural killer (NK) cell activation, while NK cells are also able to activate and mature DCs. Toll-like receptors (TLRs) on the surface of DCs and NK cells induce the maturation and activation of these cells when engaged with their cognate ligand. We investigated to generate potent DCs by maturation with NK cells in the presence of TLR agonist in vitro and tested the efficacy of these DC vaccinations in mouse colon cancer model. The optimal ratios of DCs versus NK cells were 1:1 to 1:2. Immature DCs were mature with NK cells in the presence of lipopolysaccharide, which is TLR4 agonist, and further addition of IL-2 induced phenotypically and functionally mature bone marrow-derived DCs. These potent DCs exhibited not only high expression of several costimulatory molecules and high production of IL-12p40 and IL-12p70, but also high allogeneic T cells stimulatory capacity, and the induction of the high activities to generate tumor-specific CTLs. Consistently, vaccination with these DCs efficiently inhibited CT-26 tumor growth in mouse colon cancer model when compared to other vaccination strategies. Interestingly, combination therapy of these DC-based vaccines and with low-dose cyclophosphamide showed dramatic inhibition effects of tumor growth. These results suggest that the DCs maturated with NK cells in the presence of TLR agonist are potent inducer of antitumor immune responses in mouse model and may provide a new source of DC-based vaccines for the development of immunotherapy against colon cancer.     

Cytotoxicity and anticancer activity of natural rubber latex particles for cancer cells

To broaden the knowledge of cytotoxicity of natural rubber latex (NRL) nanoparticles we for the first time examined the latex biocompatibility in vitro against a panel of cancer cells (A549, A2780, and MDA-MB-231). Owing to fractionation of NRL nanoparticles by ultra-centrifuge, the effect of the non-rubber constituents (intermediate of 5.8 wt% and sediment of 0.2 wt%) on the cytotoxicity was clarified. For intermediate constituent, the half maximal inhibitory concentration (IC₅₀) values at 24 h was 1.05 mg/mL for A549 cells, which was one order of magnitude higher in toxicity as compared to that for A2780 (0.24 mg/mL) and MDA-MB-231 (0.36 mg/mL) cells. In addition, profound studies including cell cycle arrest abilities and apoptosis induction profiles against cancer cells were discussed in detail. It was found that the constituents exhibit some significant effect on the cell cycle arrest and trigger apoptosis for A2780 cells. This effective apoptosis induction profiles was more prominent in MDA-MB-231 cells incubated with NRL nanoparticles and sediment loading conditions. The percentage of apoptotic cells was ca. 6–8% of the total cells.     

Anti-Cancer Properties of Ginkgolic Acids in Human Nasopharyngeal Carcinoma CNE-2Z Cells via Inhibition of Heat Shock Protein 90

Ginkgo biloba L. has been used in traditional Chinese medicine (TCM) for thousands of years. However, the anti-cancer properties of ginkgolic acids (GAS) isolated from G. biloba have not been investigated in human nasopharyngeal carcinoma cells. In this study, GAS exhibited an inhibitory effect on the ATPase activity of heat shock protein 90 (Hsp90) and anti-proliferative activities against four human cancer cell lines, with IC₅₀ values ranging from 14.91 to 23.81 μg·mL⁻¹. In vivo experiments confirmed that GAS inhibited tumor growth in CNE-2Z cell-xenografted nude mice with low hepatotoxicity. We further demonstrated that GAS suppressed migration and invasion and induced the apoptosis of CNE-2Z cells by inducing the degradation of Hsp90 client proteins (MMP-2, MMP-9, Her-2, c-Raf, Akt, and Bcl-2). Together, GAS are new Hsp90 inhibitors by binding to Hsp90 (hydrogen bond and hydrophobic interaction). Thus, GAS from G. biloba might represent promising Hsp90 inhibitors for the development of anti-nasopharyngeal carcinoma agents.     

Antibacterial,antifungal and in vivo anticancer activities of chitin-binding lectins from Tomato (Solanum lycopersicum) fruits

A mixture of chitin-binding lectins from Tomato (Solanum lycopersicum) fruits, designated as Tomato chitin-binding lectins (TCLs), was isolated through affinity chromatography using an acetylated chitin column. Molecular weights of TCLs were determined to be 30 to 115 KDa which possessed mild toxicity with an LC₅₀ value of 521 µg/ml examined by the brine shrimp nauplii toxicity assay. Strong antibacterial activity of TCLs was found against Escherichia coli, Staphylococcus aureus and Shigella boydii at a concentration of 500 µg/ml by using disc diffusion method. Minimum inhibitory concentrations (MIC) of TCLs against Staphylococcus aureus and Escherichia coli were found to be 200 µg/ml and 140 µg/ml, respectively whereas minimum bactericidal concentrations (MBC) against the same bacterial species were 840 and 600 µg/ml, respectively. TCLs also exerted antibiofilm activity (53.32% at 250 μg/ml) against Escherichia coli. Strong antifungal activity of TCLs against Aspergillus niger was found at 600 µg/ml whereas the lectin mixture agglutinated A. niger spores at 200 µg/ml. TCLs exhibited 19.63% and 59.91% anti-proliferative activity against Ehrlich ascites carcinoma (EAC) cells in vivo in Swiss albino mice when intraperitonealy injected at doses 1.0 mg/kg/day and 2.0 mg/kg/day, respectively for five consecutive days. Morphological changes of apoptosis in EAC cells under fluorescence microscope and alteration of the expression of apoptosis-related genes (Fas, Caspase 8 and Caspase 3) had also been observed. MTT assay showed 27.61%, 38.74% and 49.23% of in vitro anticancer activity of the tomato lectins at concentrations of 37.5, 75 and 150 µg/ml, respectively.     

Flow Cytometry Analysis of <Emphasis Type="Italic">G</Emphasis><Subscript>0</Subscript>/<Emphasis Type="Italic">G</Emphasis><Subscript>1</Subscript> Diploid Cell Fraction in Ovarian Cancer Tissue

The proportion of diploid cells in the G₀/G₁ cell cycle phases was estimated by flow cytometry in 60 samples of stage III serous ovarian cancer tissue. The tumor tissue shows considerable heterogeneity with regard to the content of this tissue fraction, which ranged from 27 to 95% with a median of 73%. Statistically significant differences in the size of this fraction were identified by comparing tumor subgroups sensitive and resistant to first-line platinum-taxane chemotherapy. Predictive significance of the G₀/G₁ fraction was concluded and quantitative evaluation of this fraction is recommended for clinical use.     

Bioinformatics and Functional Analyses Implicate Potential Roles for EOGT and L-fringe in Pancreatic Cancers

Notch signaling receptors, ligands, and their downstream target genes are dysregulated in pancreatic ductal adenocarcinoma (PDAC), suggesting a role of Notch signaling in pancreatic tumor development and progression. However, dysregulation of Notch signaling by post-translational modification of Notch receptors remains poorly understood. Here, we analyzed the Notch-modifying glycosyltransferase involved in the regulation of the ligand-dependent Notch signaling pathway. Bioinformatic analysis revealed that the expression of epidermal growth factor (EGF) domain-specific O-linked N-acetylglucosamine (EOGT) and Lunatic fringe (LFNG) positively correlates with a subset of Notch signaling genes in PDAC. The lack of EOGT or LFNG expression inhibited the proliferation and migration of Panc-1 cells, as observed by the inhibition of Notch activation. EOGT expression is significantly increased in the basal subtype, and low expression of both EOGT and LFNG predicts better overall survival in PDAC patients. These results imply potential roles for EOGT- and LFNG-dependent Notch signaling in PDAC.