Phytochemicals in Cancer Prevention and Therapy

Despite advances in modern medicine, cancer is still the major cause of mortality in both developing and developed countries. Search for safer and more effective chemoprevention and treatment strategy is a need for the improvement of patient care in the field. Prevention may be more effective and less costly because cancer is largely a preventable disease which could be attributed to a greater extent to lifestyle. Dietary phytochemicals have been used for the treatment of cancer throughout history due to their safety, low toxicity, and general availability. Population based studies suggest that a reduced risk of cancer is associated with high consumption of vegetables and fruits. Promising phytochemicals not only disrupt aberrant signaling pathways leading to cancer but also synergize with chemotherapy and radiotherapy. Thus, the cancer chemoprevention and therapeutic potential of naturally occurring phytochemicals are of great interest. In this special issue we have collected many interesting original research articles and reviews that provide solid evidence to support the application of phytochemicals or dietary agents in prevention and treatment of cancer

Plumbagin inhibits invasion and migration of breast and gastric cancer cells by downregulating the expression of chemokine receptor CXCR4

<p>Abstract</p> <p>Background</p> <p>Increasing evidence indicates that the interaction between the CXC chemokine receptor-4 (CXCR4) and its ligand CXCL12 is critical in the process of metastasis that accounts for more than 90% of cancer-related deaths. Thus, novel agents that can downregulate the CXCR4/CXCL12 axis have therapeutic potential in inhibiting cancer metastasis.</p> <p>Methods</p> <p>In this report, we investigated the potential of an agent, plumbagin (5-hydroxy-2-methyl-1, 4-naphthoquinone), for its ability to modulate CXCR4 expression and function in various tumor cells using Western blot analysis, DNA binding assay, transient transfection, real time PCR analysis, chromatin immunoprecipitation, and cellular migration and invasion assays.</p> <p>Results</p> <p>We found that plumbagin downregulated the expression of CXCR4 in breast cancer cells irrespective of their HER2 status. The decrease in CXCR4 expression induced by plumbagin was not cell type-specific as the inhibition also occurred in gastric, lung, renal, oral, and hepatocellular tumor cell lines. Neither proteasome inhibition nor lysosomal stabilization had any effect on plumbagin-induced decrease in CXCR4 expression. Detailed study of the underlying molecular mechanism(s) revealed that the regulation of the downregulation of CXCR4 was at the transcriptional level, as indicated by downregulation of mRNA expression, inhibition of NF-κB activation, and suppression of chromatin immunoprecipitation activity. In addition, using a virtual, predictive, functional proteomics-based tumor pathway platform, we tested the hypothesis that NF-κB inhibition by plumbagin causes the decrease in CXCR4 and other metastatic genes. Suppression of CXCR4 expression by plumbagin was found to correlate with the inhibition of CXCL12-induced migration and invasion of both breast and gastric cancer cells.</p> <p>Conclusions</p> <p>Overall, our results indicate, for the first time, that plumbagin is a novel blocker of CXCR4 expression and thus has the potential to suppress metastasis of cancer.</p

Emodin Suppresses Migration and Invasion through the Modulation of CXCR4 Expression in an Orthotopic Model of Human Hepatocellular Carcinoma

10.1371/journal.pone.0057015PLoS ONE83

γ-Tocotrienol is a novel inhibitor of constitutive and inducible STAT3 signalling pathway in human hepatocellular carcinoma: potential role as an antiproliferative, pro-apoptotic and chemosensitizing agent

10.1111/j.1476-5381.2010.01187.xBritish Journal of Pharmacology1632283-298BJPC

Emodin suppresses migration and invasion of HepG2 cells.

<p>A, Wound-healing assay was performed for evaluating the inhibitory effect of emodin on HepG2 cell migration. Confluent monolayers of HepG2 cells were scarred, and repair was monitored microscopically after 12 h of pre-treatment with emodin (50 µM) before being exposed to 100ng/mL CXCL12 for 24 h. Width of wound was measured at time zero and 24 h of incubation with and without emodin in the absence or presence of CXCL12. The representative photographs showed the same area at time zero and after 24 h of incubation. B, HepG2 (2×10⁵ cells) were seeded in the top-chamber of the Matrigel. After pre-incubation with or without emodin (50 µM) for 12 h, transwell chambers were then placed into the wells of a 24-well plate, in which we had added either the basal medium only or basal medium containing 100 ng/mL CXCL12 for 24 h. After incubation, they were assessed for cell invasion as described in Materials and Methods. Columns represent percentage of invaded cells; bars, S.E. *indicates p value <0.05. Representative results of two independent experiments are shown.</p

Emodin suppresses CXCR4 mRNA level in HCC cells.

<p>A and B, Emodin suppresses CXCR4, neither by proteasomal nor by lysosomal degradation. HepG2 cells (1×10⁶) were treated with indicated concentrations of ALLN or chloroquine for 1 h at 37°C, followed by treatment of 50 µM emodin for 6 h. Whole-cell extracts were prepared and analyzed by Western blot analysis using antibodies against CXCR4. The same blots were stripped and reprobed with β-actin antibody to show equal protein loading. Representative results of two independent experiments are shown. C, Emodin suppresses the expression of CXCR4 mRNA in HepG2 cells. HepG2 cells were treated with 50µM emodin for the indicated time intervals, after which cells were harvested after treatment and total RNA samples were extracted. 1µg portions of the respective RNA extracts then proceed for Reverse Transcription to generate corresponding cDNA. Real time PCR was performed to measure the relative quantities of CXCR4 mRNA using targeted TaqMan probes, with GAPDH as endogenous control for measurement of equal loading of RNA samples. Results were analyzed using Sequence Detection Software version 1.3 provided by Applied Biosystems. Relative gene expression was obtained after normalization with endogenous GAPDH and determination of the difference in threshold cycle (Ct) between treated and untreated cells using 2-ΔΔCt method.</p

Emodin inhibits TNF-α inducible NF-κB activation in HepG2 cells.

<p>A, HepG2 cells were incubated with 50 µM emodin for the indicated time points and then stimulated with TNF-α (1 nM) for 24 h. The nuclear extracts were assayed for NF-κB activation by TransAM p65 transcription factor assay kit. Bars indicate standard error. * indicates p value <0.05. B, HepG2 cells were transiently transfected with an NF-κB luciferase plasmid and co-transfected with β-galactosidase and then treated with 50 µM emodin for the indicated time points and then stimulated with TNF-α (1 nM) for 24 h. Cell supernatants were thereafter collected and assayed for luciferase activity as described in Materials and Methods. Representative results of two independent experiments are shown. Results are expressed as fold activity over the activity of the vector control. Bars indicate standard error. * indicates p value <0.05; ** indicates p value <0.001. C, Emodin inhibits binding of NF-κB to the CXCR4 promoter. Hep3B cells were treated with 50 µM emodin for indicated time intervals and the proteins were crosslinked with DNA by formaldehyde and then subjected to ChIP assay using an anti-p65 antibody with the CXCR4 primer. Reaction products were resolved by electrophoresis.</p

Emodin suppresses lung metastasis and CXCR4 expression in orthotopic mice model.

<p>Spontaneous metastatic model of human liver cancer cells orthotopically implanted in nude mice. Cubes of HCCLM3_Luc tumor were implanted orthotopically into the liver of female Balb/c nude mice. Mice were euthanized when the detectable liver tumor signal was >1×10¹¹ p/s. A, At the time of necropsy, bioluminescent imaging was employed to monitor signals from lung metastasis. The lung biopsy was imaged to detect metastasis in the lungs. The color bar depicts the photon flux (p/s) emitted. # = mouse ID; D = day at necropsy. Lung and liver signals are in p/s unit. B, Comparison of the lung metastasis signals detected at time of necropsy for the untreated and emodin-treated groups. C, Tumor tissue extracts were prepared as described in Methods. 30 µg of protein was taken and analyzed by Western blot analysis with antibodies against CXCR4. The same blots were stripped and reprobed with β-actin antibody to show equal protein loading. D, Immunohistochemical analysis of representative section of tumor tissues stained with anti-CXCR4 antibody demonstrate strong cytoplasmic staining for CXCR4 in the control section and weak cytoplasmic staining in the emodin treated section E, Emodin decreases CXCR4 levels in tumor tissues. Tumor tissue homogenate was prepared as described in Methods. 150 µg of the tissue supernatant was taken for ELISA assay. Assay was performed according to manufacturer’s instructions using ELISA kit to determine the levels of CXCR4 in the control and treated group. Data is represented as Mean ± SE. * indicates p value <0.05.</p