DEVELOPMENT OF A THERAPEUTIC MODEL OF EARLY LIVER CANCER USING CROCIN-COATED MAGNETITE NANOPARTICLES

Hepatocellular carcinoma is one of the most common health problems that is difficult to treat. As a result of the side effects frequently experienced with conventional cancer treatments, there has been a growing interest to develop controlled drug delivery system that can reduce the mortality rate of liver cancer patients and un-harm healthy tissues. Magnetite nanoparticles are potentially important in hepatocellular carcinoma treatment, since they can be used as delivery system. Pure and coated magnetite nanoparticles were synthesized via modified co-precipitation method in air at low temperature. Various reaction parameters and coating materials have been investigated and characterized. Among these parameters and coating materials, 1.0 % of dextran was selected as an optimum coating for nanoparticles using a slow feeding rate for the Fe2+/Fe3+ reactants, maintaining the stirring and soaking temperatures at 60ºC. After that dextran-coated magnetite nanoparticles were bound to crocin, a pharmacologically active component of saffron, via cross-linker. Crocin alone has shown anti-cancer activity in different in vitro and in vivo settings by several studies. The aim of this study was to synthesize dextran-coated magnetite nanoparticles containing crocin with a higher therapeutic index for hepatocellular carcinoma treatment. The nanoparticles with crocin were tested in vitro and in vivo for their anti-cancer effects as compared to free crocin. HepG2 cells treated with crocin-dextran-coated magnetite nanoparticles showed a decrease in cell proliferation compared to control (non-treated cells) or to those treated with free crocin or dextran-coated nanoparticles. The anti-cancer activity of crocin-dextran-coated nanoparticles was also evaluated in Balb/c mice. These mice were injected with carcinogenic agent, diethylnitrosamine. Histological examination revealed several precancerous changes. The immunohistochemical analysis using antibodies indication of cell proliferation (Ki-67), apoptosis (M30-Cytodeath and Bcl-2), inflammation (cyclooxygenase-2) and angiogenesis (vascular endothelial growth factor), indicated that magnetite nanoparticles conjugated with dextran plus crocin does indeed improve its anti-tumorigenic activity over free crocin. These results provide the basis for designing new modalities for treatment of liver cancer which could hopefully reduce its high mortality rate

Immobilized Ag NPs on chitosan-biguanidine coated magnetic nanoparticles for synthesis of propargylamines and treatment of human lung cancer

The magnetically isolable nanobiocomposites have significant impact as the modified new generation catalysts in recent days. This has persuaded us to design and synthesis of a novel Ag NPs decorated biguanidine-chitosan (Bigua-CS) dual biomolecular functionalized core-shell type magnetic nanocomposite (Ag/Bigua-CS@Fe3O4). Bigua-CS could be introducing polysaccharide materials as potential coating agent to immobilizing and stabilizing metal nanoparticles. The material was characterized using several advanced techniques like fourier transformed infrared spectroscopy (FT-IR), inductively coupled plasma (ICP), field emission scanning electron microscopy (FE-SEM), energy dispersive X-ray spectroscopy (EDX), atomic mapping, high resolution transmission electron microscopy (HR-TEM), vibrating sample magnetometer (VSM) and X-ray diffraction (XRD). Towards the chemical applications of the material, we headed the multicomponent synthesis of diverse propargylamines by A(3) coupling in water, which ended up with excellent yields. Due to strong paramagnetism, the catalyst was easily isolable and reused in 9cycles without any leaching and considerable change in reactivity. In addition, the catalyst was engaged in biological assays like study of anti-oxidant properties by DPPH mediated free radical scavenging test using BHT as a reference molecule. Thereafter, on having a significant IC50 value in radical scavenging assay, we extended the bio-application of the catalyst in anticancer study of adenocarcinoma cells of human lungs. The three different cancer cell lines, PC-14, LC-2/ad and HLC-1 were used in this regard. The best result was achieved in the case of PC-14 cell line with strong IC50 values

Intravenous administration of manuka honey inhibits tumor growth and improves host survival when used in combination with chemotherapy in a melanoma mouse model.

Manuka honey has been recognized for its anti-bacterial and wound-healing activity but its potential antitumor effect is poorly studied despite the fact that it contains many antioxidant compounds. In this study, we investigated the antiproliferative activity of manuka honey on three different cancer cell lines, murine melanoma (B16.F1) and colorectal carcinoma (CT26) as well as human breast cancer (MCF-7) cells in vitro. The data demonstrate that manuka honey has potent anti-proliferative effect on all three cancer cell lines in a time- and dose-dependent manner, being effective at concentrations as low as 0.6% (w/v). This effect is mediated via the activation of a caspase 9-dependent apoptotic pathway, leading to the induction of caspase 3, reduced Bcl-2 expression, DNA fragmentation and cell death. Combination treatment of cancer cells with manuka and paclitaxel in vitro, however, revealed no evidence of a synergistic action on cancer cell proliferation. Furthermore, we utilized an in vivo syngeneic mouse melanoma model to assess the potential effect of intravenously-administered manuka honey, alone or in combination with paclitaxel, on the growth of established tumors. Our findings indicate that systemic administration of manuka honey was not associated with any alterations in haematological or clinical chemistry values in serum of treated mice, demonstrating its safety profile. Treatment with manuka honey alone resulted in about 33% inhibition of tumor growth, which correlated with histologically observable increase in tumor apoptosis. Although better control of tumor growth was observed in animals treated with paclitaxel alone or in combination with manuka honey (61% inhibition), a dramatic improvement in host survival was seen in the co-treatment group. This highlights a potentially novel role for manuka honey in alleviating chemotherapy-induced toxicity

Clinical chemistry parameters are unaltered in mice following intravenous injection with manuka honey.

<p>Mice were treated as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0055993#pone-0055993-g004" target="_blank">Figure 4</a> legend, following which blood was collected and analyzed for the indicated parameters. In each graph, the values for individual mice in a group are shown, together with the mean ± SEM. The shaded box in each graph represents the normal range for that particular parameter. The results are representative of three independent experiments.</p

Systemic administration of manuka honey is not associated with any alterations in hematological values.

<p>Mice were injected with saline or manuka (50% w/v) 2 times per week for a total of 3 weeks, following which blood was collected and analyzed for the indicated parameters. In each graph, the values for individual mice in a group are shown, together with the mean ± SEM. The shaded box in each graph represents the normal range for that particular parameter. The results are representative of three independent experiments.</p

Co-treatment with manuka and taxol results in additive effect.

<p>B16.F1 cells were seeded at 1×10³ cells per well in a 96-well plate and incubated with the indicated concentrations of manuka, alone or in combination with taxol (10 ng/ml), for 72 hrs. Cell viability was determined using CellTiter-Glo luminescent assay. Results are expressed as percentage viability in treated cell cultures compared to untreated cells and are representative of 3 independent experiments. Asterisks denote statistically significant differences between corresponding cell cultures treated with each manuka concentration in absence or presence of taxol (*, <i>p</i><0.05).</p

Effect of systemic administration of manuka on tumor growth and host survival.

<p>(<b><i>A</i></b>) Animals with established tumors were treated i.v. with either manuka honey (50% w/v), taxol (10 mg/Kg), manuka+taxol, or saline as control. All treatments were given twice per week until the end of observation period. Each data point represents the mean ± SEM of 19–20 mice per group, pooled from 2 individual experiments. Asterisks denote statistically significant differences between each experimental group and the saline control group; also shown is a comparison between manuka alone and manuka+taxol groups (**, <i>p</i><0.01; ***, <i>p</i><0.001). (<b><i>B</i></b>) Co-treatment with taxol and manuka leads to a significant enhancement in host survival. Experimental animals were followed for survival for up to day 25 post treatment. Each data point represents the mean ± SEM of 19–20 mice per group, pooled from 2 individual experiments. Asterisks denote statistically significant differences between experimental and saline control groups; also shown is a comparison between taxol alone and manuka+taxol groups (**, <i>p</i><0.01; *, <i>p</i><0.05).</p

Manuka induces caspase-mediated apoptosis in cancer cells.

<p>B16.F1 melanoma cells were treated with manuka (5% w/v), taxol (10 or 50 ng/ml) or medium as control. After 24 hrs of culture, enzymatic activity of caspase 3/7 (graph <b><i>A</i></b>) and caspase 8 (graph <b><i>B</i></b>) were determined using specific kits and following manufacturer's recommendation. The data is presented as fold increase in caspase activity after normalization to the number of viable cells per culture. <b><i>C</i></b>. Western blot analysis of caspase-9 activation B16.F1 cells treated with manuka or taxol. Whole cell extracts were prepared after a 24-hr treatment with manuka (5% w/v) or taxol (10 ng/ml). Protein extracts were resolved on 10% SDS-PAGE and immunoblotted with caspase-9-specific ployclonal antibody capable of detecting both full length and cleaved forms of caspase-9. The cell extracts were also probed with an antibody against β-actin as a control for protein loading.</p