Molecular Mechanisms Involved In Induction Of Cell Growth Arrest And Cell Death In Human Colon Cancer Cells By Tangutorine A Beta Carboline

Gallotannin (GT) is a polyphenol that is active against colon cancer. GT has bee n shown to induce cell cycle arrest in the human colon cancer cell line HCT116 a nd apoptosis in the isogenic cell line HCT116 p21-/-. The mechanisms underlying these anti-carcinogenic effects are not well defined. The aim of this study is to elucidate the molecular mechanisms of action of GT, to determine whether GT c auses DNA damage, to determine the role of p21 in antagonizing apoptosis and cau sing senescence in HCT116 human colon cancer cells and to determine the molecula r mechanisms involved in the antitumorigenic effects of GT in vivo. This work shows that GT causes DNA damage and decreases the survival of HCT116 cells. GT downregulated Mdm2 and upregulated p53 expression levels in wild type HCT116 cells. Conversely, GT upregulated Mdm2 and downregulated p53 expression l evels in HCT p21-/- cells. It also downregulated E2F-1 protein expression levels in wild type HCT116 cells and upregulated it in HCT p21-/- cells. GT induced tr ansient upregulation of p21 at 20 microg/ml followed by remarkable reduction at 40 microg/ml in wild type HCT116 cells. GT-induced apoptosis in HCT116 p21-/- ce lls correlated to decreased p53 and Puma. Transfection of HCT116 p21-/- cells wi th p21 plasmid rescued the cells from apoptosis and restored wild type phenotype upon GT treatment. Moreover, GT induced senescence in wild type HCT116 cells bu t not in HCT116 p21-/- cells. Previous work in this laboratory has shown that GT decreases and stabilizes the growth of human colon cancer xenografts in NOD/SCID mice. In an attempt to inve stigate the mechanism of in vivo growth inhibition, tumor tissues were stained f or Ki-67, VEGFA, MMP-2, and MMP-9. GT decreased the expression of Ki-67, a proli feration marker, VEGFA, an angiogenesis marker, and MMP-2, a metastasis marker i n the NOD/SCID xenograft model of colon cancer. GT had no effect on the expressi on levels of MMP-9 protein. In conclusion, the presence of p21gene modulates the response of HCT116 cells to GT. When p21 is present, senescence is induced while in p21 absence higher prop ortion of cells die by apoptosis. The senescence inducing property of GT in HCT1 16 cells supports its development for use in anti cancer therapy.

Cancer cells are remarkably resilient to therapies aimed at their elimination. The exploration of pathways that sustain cancer cells and that allow cancer cells to become resistant has revealed new avenues for chemotherapeutic development, as well as rational approaches to combination therapies based on existing treatment options. Several signaling pathways, such as Wnt, phosphoinositide 3-kinase (PI3K), and Ras-Raf-MEK, constitute integrated networks that work together to maintain cellular homeostasis under basal conditions and to drive cell-mass accumulation and cell cycle progression in the presence of appropriate mitogenic stimuli. During cancer development, these pathways are corrupted in malignant cells to maintain viability and proliferative activity under environmentally stressful conditions such as limited growth factors, oxygen, and nutrients that drive normal cells into quiescence or death. Importantly, dysfunction within any one of these pathways results in compensatory responses from the other networks. Thus, biological research is gradually shifting toward more general approaches that target entire pathways rather than isolated components and integrate those pathways into biological networks.

The endoplasmic reticulum (ER) stress pathway is a prominent regulator of cancer cell death and as a result, ER stress inducers are being exploited pharmacologically. Cytotoxic ER stress is typically regulated by the transcription factor, C/EBP homologous protein 10 (CHOP10). The products of CHOP10 transcription include three pro-apoptotic proteins; tribbles-related protein 3 (TRB3), death receptor-5 (DR5), and ER oxidoreductase 1[alpha] (ERO1[alpha]). Our previous findings showed that cell death induced by 15deoxy, [delta]12,14 prostamide J2 (15d PMJ2) occurred in an ER stress-dependent manner. However, the signaling pathway that regulates15d PMJ2-mediated ER stress cell death has not been identified. Therefore, the goal of this study is to determine the role of CHOP10 and its transcriptional products in cytotoxic ER stress that is induced by 15d PMJ2 in colon cancer cells. Our results show that 15d PMJ2 exhibited a preferential cytotoxicity toward the human colon cancer cell line, HCT116, compared to the non-tumorigenic colon cell line, FHC. The induction of cell death in HCT116 cells was accompanied by a significant increase in CHOP10 protein expression. In addition, pharmacological blockade of ER stress prevented 15d PMJ2 induced cell death indicating that cell death is reliant upon ER stress. To determine the role of CHOP10 in this process, a genetic deletion of CHOP10 (CHOP10-KO) was performed using CRISPR/Cas9. 15d PMJ2 induced death in WT but not in CHOP10-KO cells. Interestingly, TRB3 synthesis appears to require 15d PMJ2-induced CHOP10 expression. Therefore, the activity of protein kinase B (PKB)/Akt, a known TRB3 target, was investigated. CHOP10 stimulation by 15d PMJ2 inactivated Akt in WT but not in CHOP10-KO cells. TRB3 transfected CHOP10-KO cells restored 15d PMJ2-induced death, suggesting the importance of TRB3 in CHOP10-initiated inhibition of Akt and the control of the ER stress-mediated cell death by 15d PMJ2. Moreover, PD-L1, which is a T cell inhibitory ligand, is downstream of Akt, thus cell surface PD-L1 expression was examined. 15d PMJ2 decreased PD-L1 expression in WT cells and it was comparable to CHOP10-KO cells. The activity of NF-kB, which is a modulator of PD-L1 expression, was investigated. A decrease in PD-L1 expression was mediated by an inhibition of NF-kB activity by 15d PMJ2, and NF-kB transfection mitigated 15d PMJ2-reduced PD-L1 expression. In 15d PMJ2 treated cells, a decrease in PD-L1 expression reduced apoptosis of T cells, indicating that 15d PMJ2 enhanced anti-tumor immunity via PD-L1 down-regulation. Thus, 15d PMJ2 is a potential anti-tumor agent that possesses the dual actions of tumor destruction and anti-tumor immune cell stimulation.

This important volume commences with an overview of the modes of action of defensive secondary metabolites, followed by detailed surveys of chemical defense in marine ecosystems, the biochemistry of induced defense, plant-microbe interactions and medical applications. A chapter is also included covering biotechnological aspects of producing valuable secondary metabolites in plant cell and organ cultures. This is a comprehensive and fully updated new edition, edited by Professor Michael Wink and including contributions from many internationally acknowledged experts in the field.

This volume focuses on reclamation, management, and utilization of salt-affected soils, their sustainable use, and evaluation of plants inhabiting naturally occurring saline habitats. It is of interest to scientists and students as well as agricultural institutions and farmers to increase the awareness of salinity problems. The volume is supported by UNESCO Doha, Qatar, and has an international authorship.

Desertification (land degradation in arid, semi-arid and dry sub-humid areas resulting mainly from adverse human impacts) is the main environmental problem of dry lands, which occupy more than 40 per cent of the total global land area. The phenomenon threatens about 3.6 billion hectares and currently affects the livelihood of about 900 million people. Thl! world is now losing annually about 1.5 million hectares of total irrigated lands (240 million hectares) due mostly to salinization, mainly in drylands. Salt affected soils are widely distributed throughout the arid and semi-arid regions, and particularly severe in China (7 million ha), India (20 million ha), Pakistan (3.2 million ha), USA (5.2 million ha), as well as Near East, southern Europe and elsewhere. Demands on production have increased the pressure on existing productive land and moved the limits of production onto increasingly marginal lands. Wise land-use practices have yet to be developed for such conditions. The Executive Director of UNEP reported to the Governing Council in February 1992 concerning the "Status of Desertification and Implementation of the United Nations Plan of Action to Comtat Desertification (PACD)". The Report concludes that major efforts to implement the PACD had gJne into supporting measures rather than concrete corrective field operations. Little evidence of progrl!ss was found in irrigated croplands, rainfed croplands or rangelands. It was recommended that every piece of land should be used in keeping with its ecological characteristics, natural capabilities and constraints.

"The book is designed for use by advanced students, researchers and professionals in plant biochemistry, physiology, molecular biology, genetics, pharmacology, medicine, pharmacy and agriculture working in the academic and industrial sectors, including the pesticide and pharmaceutical industries."--Jacket.