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The Legume Inflammation Feeding Experiment is, to our knowledge, the first randomized crossover feeding trial testing the effects of a legume-enriched, low-glycemic index (GI) diet among men characterized for colorectal adenomas and insulin resistance (IR) status. This study was designed to test the effects of a legume-enriched diet compared with a healthy American (HA) diet under weight-stable conditions. The primary objective was to assess effects on C-reactive protein (CRP) and C-peptide levels. The secondary objective was to assess changes by IR status or history of adenomas. A total of 64 men who completed a colonoscopy within the previous 2 y consumed 2 diets in random order each for 4 wk separated by a washout period. The diets were a legume-enriched (250 g/d), low-GI (GI 38) diet and a high-GI (GI 69) HA diet. We measured fasting glucose, insulin, C-peptide, CRP, and soluble tumor necrosis factor-alpha receptors I and II (sTNFRI/II) at the beginning and end of the diet periods. Participants who consumed both the legume and HA diets had favorably improved CRP (-20.2 and -18.3%) and sTNFRI (-3.7 and -4.4%) concentrations, respectively. The sTNFRII concentrations declined marginally during the legume diet period (-3.8%; P = 0.060) and significantly during the HA diet period (-5.1%; P < 0.001). Fasting glucose increased significantly during both the legume (+1.8%) and HA (-2.2%) diet periods. Only the changes in glucose differed between the diet periods. Serum C-peptide and plasma insulin levels did not change in participants consuming either diet. Healthful dietary changes can improve biomarkers of IR and inflammation. [ABSTRACT FROM AUTHOR]

The present study was undertaken to test the hypothesis that the toxicity and carcinogenicity of vanadium might arise from elevation of reactive oxygen species leading to activation of the transcription factor activator protein-1 (AP-1). The AP-1 transactivation response has been implicated as causal in transformation responses to phorbol esters and growth factors. To investigate the possible activity of vanadium in the activation of AP-1, we treated mouse epidermal Jb6 P+ cells stably transfected with an AP-1 luciferase reporter plasmid with various concentrations of vanadate. This resulted in concentration-dependent transactivation of AP-1. Superoxide dismutase (SOD) and catalase inhibited AP-1 activation induced by vanadate, indicating the involvement of superoxide anion radical (O2-·),hydroxyl radical (·OH) and/or H2O2 in the mechanism of vanadate-induced AP-1 activation. However, sodium formate, a specific ·OH scavenger, did not alter vanadate-induced AP-1 activation, suggesting a minimal role for the ·OH radical. NADPH enhanced AP-1 activation by increasing vanadate-mediated generation of O2-·. N-acetylcysteine, a thiol-containing antioxidant, decreased activation, further showing that vanadate-induced AP-1 activation involved redox reactions. Calphostin C, a specific inhibitor of protein kinase C (PKC), inhibited activation of AP-1, demonstrating that PKC is involved in the cell signal cascades leading to vanadate-induced AP-1 activation. Electron spin resonance (ESR) measurements show that Hb6 P+ cells are able to reduce vanadate to generate vanadium(IV) in the presence of NADPH. Molecular oxygen was consumed during the vanadate reduction process to generate O2-· as measured by ESR spin trapping using 5,5-dimethyl-L-pyrroline N-oxide as the spin trapping agent. SOD inhibited the ESR spin adduct signal, further demonstrating the generation of O2-· in the cellular reduction of vanadate. These results provide support for a model in which vanadium, like other classes of tumor promoters, transactivates AP-1-dependent gene expression. In the case of vanadium, AP-1 transactivation is dependent on the generation of O2-· and H2O2, but not ·OH. [ABSTRACT FROM PUBLISHER]

We have reported the tumor suppressive effects of manganese-containing superoxide dismutase (MnSOD) in human breast cancer cells. In order to understand the molecular mechanism of this anti-tumor effect, we asked whether tumor suppressor gene(s), especially the ones inhibiting tumor invasion and motility, are involved in MnSOD-induced tumor suppression. Maspin is one of the serpin family of protease inhibitors that has been shown to function as a tumor-suppressor in human breast epithelium. In the present study, we demonstrated that maspin expression was up-regulated in human breast cancer MCF-7 cells that overexpress a normal MnSOD gene. The induced maspin transcripts were detected by RT-PCR and Northern blot and identified by sequencing. Maspin gene expression was induced in parallel with the level of exogenous MnSOD protein, which was induced by transfection with varied amounts of cDNA. In order to analyze cell invasion ability, which may be related to the induced maspin gene expression. MnSOD stable transfectants were tested using a matrigel invasion chamber. The invasion ability was reduced to 24% and 36% in the cloned (MCF + SOD) and pooled MnSOD-transfectants (MCF + SODp) respectively, compared with the wild-type MCF-7 cell line. In conclusion, these results suggest that overexpression of a normal MnSOD cDNA in human breast cancer cells up-regulates the gene expression of the protease inhibitor, maspin, which may play a role in the inhibitory function of MnSOD on tumor invasion. [ABSTRACT FROM PUBLISHER]

Programmed cell death 4 (Pdcd4), a tumor invasion suppressor, is frequently downregulated in colorectal cancer and other cancers. In this study, we find that loss of Pdcd4 increases the activity of mammalian target of rapamycin complex 2 (mTORC2) and thereby upregulates Snail expression. Examining the components of mTORC2 showed that Pdcd4 knockdown increased the protein but not mRNA level of stress-activated-protein kinase interacting protein 1 (Sin1), which resulted from enhanced Sin1 translation. To understand how Pdcd4 regulates Sin1 translation, the SIN1 5' untranslated region (5'UTR) was fused with luciferase reporter and named as 5'Sin1-Luc. Pdcd4 knockdown/knockout significantly increased the translation of 5'Sin1-Luc but not the control luciferase without the SIN1 5'UTR, suggesting that Sin1 5'UTR is necessary for Pdcd4 to inhibit Sin1 translation. Ectopic expression of wild-type Pdcd4 and Pdcd4(157-469), a deletion mutant that binds to translation initiation factor 4A (eIF4A), sufficiently inhibited Sin1 translation, and thus suppressed mTORC2 kinase activity and invasion in colon tumor cells. By contrast, Pdcd4(157-469)(D253A,D418A), a mutant that does not bind to eIF4A, failed to inhibit Sin1 translation, and consequently failed to repress mTORC2 activity and invasion. In addition, directly inhibiting eIF4A with silvestrol significantly suppressed Sin1 translation and attenuated invasion. These results indicate that Pdcd4-inhibited Sin1 translation is through suppressing eIF4A, and functionally important for suppression of mTORC2 activity and invasion. Moreover, in colorectal cancer tissues, the Sin1 protein but not mRNA was significantly upregulated while Pdcd4 protein was downregulated, suggesting that loss of Pdcd4 might correlate with Sin1 protein level but not mRNA level in colorectal cancer patients. Taken together, our work reveals a novel mechanism by which Pdcd4 inhibits Sin1 translation to attenuatemTORC2 activity and thereby suppresses invasion.

There is a strong belief that garlic has medicinal properties and may even reduce the risk of developing certain cancers including those of the gastrointestinal tract. The chemopreventive effects of garlic may be attributed to the anti-inflammatory properties of the sulfur-containing constituents of garlic, which includes diallyl disulfide (DADS). Here, we demonstrate that DADS prevented colorectal tumorigenesis in a mouse model of colitis-induced colorectal cancer. Supplementation with 85 ppm of DADS (60 mg daily human equivalent dose) in the diet of FVB/N mice treated with chemical carcinogen azoxymethane (AOM) and colonic irritant dextran sodium sulfate (DSS) resulted in the reduction in tumor incidence, tumor number, and tumor burden by 21.54%, 47.3%, and 66.4%, respectively. Further analysis revealed that mice fed the DADS-supplemented diet resolved the initial DSS-induced inflammation faster than those on the control diet, preventing prolonged inflammation and cellular transformation. Subsequent mechanistic studies in vitro suggest that DADS chemopreventive effects are mediated through NF-κB signaling. When SW480 colorectal cancer cells were treated with DADS, NF-κB nuclear localization and activity were diminished. Interestingly, NF-κB suppression was found to be dependent on DADS inhibition of GSK-3β, a positive regulator of NF-κB. Inhibition of GSK-3β and loss of nuclear NF-κB activity were also observed in vivo in AOM/DSS-treated mice fed a diet supplemented with 85 ppm DADS. Our results indicate that DADS can prevent tumorigenesis by suppressing inflammation, a process largely involving GSK-3β inhibition and consequential reduction in NF-κB nuclear localization. Cancer Prev Res; 9(7); 607-15. ©2016 AACR., (©2016 American Association for Cancer Research.)

The translation inhibitor and tumor suppressor Pdcd4 was reported to be lost in various tumors and put forward as prognostic marker in tumorigenesis. Decreased Pdcd4 protein stability due to PI3K-mTOR-p70S6K1 dependent phosphorylation of Pdcd4 followed by β-TrCP1-mediated ubiquitination, and proteasomal destruction of the protein was characterized as a major mechanism contributing to the loss of Pdcd4 expression in tumors. In an attempt to identify stabilizers of Pdcd4, we used a luciferase-based high-throughput compatible cellular assay to monitor phosphorylation-dependent proteasomal degradation of Pdcd4 in response to mitogen stimulation. Following a screen of approximately 2000 compounds, we identified 1,2-bis(4-chlorophenyl)disulfide as a novel Pdcd4 stabilizer. To determine an initial structure-activity relationship, we used 3 additional compounds, synthesized according to previous reports, and 2 commercially available compounds for further testing, in which either the linker between the aryls was modified (compounds 2-4) or the chlorine residues were replaced by groups with different electronic properties (compounds 5 and 6). We observed that those compounds with alterations in the sulfide linker completely lost the Pdcd4 stabilizing potential. In contrast, modifications in the chlorine residues showed only minor effects on the Pdcd4 stabilizing activity. A reporter with a mutated phospho-degron verified the specificity of the compounds for stabilizing the Pdcd4 reporter. Interestingly, the active diaryl disulfides inhibited proliferation and viability at concentrations where they stabilized Pdcd4, suggesting that Pdcd4 stabilization might contribute to the anti-proliferative properties. Finally, computational modelling indicated that the flexibility of the disulfide linker might be necessary to exert the biological functions of the compounds, as the inactive compound appeared to be energetically more restricted.

Colorectal cancer, a leading cause of cancer death, has been linked to inflammation and obesity. Berberine, an isoquinoline alkaloid, possesses anti-inflammatory, anti-diabetes and anti-tumor properties. In the azoxymethane initiated and dextran sulfate sodium (AOM/DSS) promoted colorectal carcinogenesis mouse model, berberine treated mice showed a 60% reduction in tumor number (P = 0.009), a 48% reduction in tumors <2 mm, (P = 0.05); 94% reduction in tumors 2-4 mm, (P = 0.001), and 100% reduction in tumors >4 mm (P = 0.02) compared to vehicle treated mice. Berberine also decreased AOM/DSS induced Ki-67 and COX-2 expression. In vitro analysis showed that in addition to its anti-proliferation activity, berberine also induced apoptosis in colorectal cancer cell lines. Berberine activated AMP-activated protein kinase (AMPK), a major regulator of metabolic pathways, and inhibited mammalian target of rapamycin (mTOR), a downstream target of AMPK. Furthermore, 4E-binding protein-1 and p70 ribosomal S6 kinases, downstream targets of mTOR, were down regulated by berberine treatment. Berberine did not affect Liver kinase B1 (LKB1) activity or the mitogen-activated protein kinase pathway. Berberine inhibited Nuclear Factor kappa-B (NF-κB) activity, reduced the expression of cyclin D1 and survivin, induced phosphorylation of p53 and increased caspase-3 cleavage in vitro. Berberine inhibition of mTOR activity and p53 phosphorylation was found to be AMPK dependent, while inhibition NF-κB was AMPK independent. In vivo, berberine also activated AMPK, inhibited mTOR and p65 phosphorylation and activated caspase-3 cleavage. Our data suggests that berberine suppresses colon epithelial proliferation and tumorigenesis via AMPK dependent inhibition of mTOR activity and AMPK independent inhibition of NF-κB., (© 2014 Wiley Periodicals, Inc.)

An important characteristic of cancer is that the disease can overcome the surveillance of the immune system. A possible explanation for this resistance arises from the ability of tumor cells to block the tumoricidal activity of host immune cells such as natural killer (NK) cells by inducing the localized accumulation of regulatory T (Treg) cells. Evidence exists that components in commonly consumed foods including vitamins A, D, and E, water-soluble constituents of mushrooms, polyphenolics in fruits and vegetables, and n-3 fatty acids in fish oil can modulate NK cell activities, Treg cell properties, and the interactions between those two cell types. Thus, it is extremely important for cancer prevention to understand the involvement of dietary components with the early stage dynamics of interactions among these immune cells. This review addresses the potential significance of diet in supporting the function of NK cells, Treg cells, and the balance between those two cell types, which ultimately results in decreased cancer risk., (Published 2015. This article is a U.S. Government work and is in the public domain in the USA.)

Cryptotanshinone (CPT) is a natural compound extracted from herbal medicine that has been previously shown to possess antitumor properties in various types of human cancer cells. In the present study, we examined the potential role of CPT in the treatment of colorectal cancer. Using SW480, HCT116, and LOVO colorectal cancer cell lines, the effects of CPT on cell viability, apoptosis, and tumorigenicity were evaluated. The results showed that CPT significantly inhibited the growth and viability of SW480, HCT116, and LOVO cell lines by inducing apoptosis and prevented anchorage dependent growth on agar. In addition, CPT inhibited the activation of Signal transducer and activator of transcription 3 (Stat3) pathways in colorectal cancer cells. Stat3 is a transcription factor that mediates the expression of various genes associated with many cellular processes, such as inflammation and cell growth, and has been shown to promote several cancer types, including colorectal cancer. These findings indicate that CPT may be a potential candidate for the treatment and prevention of colorectal cancer in part by inhibiting the activation of Stat3.

Scope: Aim of the study was to identify and monitor metabolite markers of dry bean consumption in parallel human and mouse studies that each had shown chemopreventive effects of dry bean consumption on colorectal neoplasia risk., Methods and Results: Using LC/mass spectroscopy ± ESI and GC/mass spectroscopy, serum metabolites of dry beans were measured in 46 men before and after a 4-week dry bean enriched diet (250 g/day) and 12 mice that received a standardized diet containing either 0 or 10% navy bean ethanol extract for 6 weeks; we also investigated fecal metabolites in the mice. The serum metabolites identified in these controlled feeding studies were then investigated in 212 polyp-free participants from the Polyp Prevention Trial who self-reported either increased (≥+31 g/day from baseline), high dry bean intake of ≥42 g/day in year 3 or low, unchanged dry bean consumption of <8 g/day; serum was analyzed from baseline and year 3. Serum pipecolic acid and S-methyl cysteine were elevated after dry bean consumption in human and mouse studies and reflected dry bean consumption in the Polyp Prevention Trial., Conclusion: Serum levels of pipecolic acid and S-methyl cysteine are useful biomarkers of dry bean consumption., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Valosin-containing protein (VCP or p97), a member of the AAA family (ATPases associated with diverse cellular activities), plays a key role in many important cellular activities. A genetic deficiency of VCP can cause inclusion body myopathy associated with Paget's disease of bone and frontotemporal dementia (IBMPFD). Previous studies showed that the VCP N domain is essential for the regulation of nuclear entry of VCP. Here we report that IBMPFD mutations, which are mainly located in the N domain, suppress the nuclear entry of VCP. Moreover, the peptide sequence G780AGPSQ in the C-terminal region regulates the retention of VCP in the nucleus. A mutant lacking this sequence can increase the nuclear distribution of IBMPFD VCP, suggesting that this sequence is a potential molecular target for correcting the deficient nucleocytoplasmic shuttling of IBMPFD VCP proteins., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Sporadic and non-hereditary mutations account for the majority of colorectal cancers (CRC). After the loss of adenomatous polyposis coli (APC) function and activation of the β-catenin/LEF signaling pathway, activating mutations in Kras are major drivers of sporadic CRC. Preventing the outgrowth of cells that develop sporadic mutations will decrease CRC. Resveratrol, a naturally occurring polyphenolic compound has anti-inflammatory, anti-oxidant and anti-cancer activities. We used a genetically engineered mouse model for sporadic CRC where the APC locus is knocked out and Kras is activated specifically in the distal colon to determine the effects of resveratrol on preventing and treating CRC. Feeding mice a diet supplemented with 150 or 300 ppm resveratrol (105 and 210mg daily human equivalent dose, respectively) before tumors were visible by colonoscopy resulted in a 60% inhibition of tumor production. In the 40% of mice that did develop tumors Kras expression was lost in the tumors. In a therapeutic assay where tumors were allowed to develop prior to treatment, feeding tumor bearing mice with resveratrol resulted in a complete remission in 33% of the mice and a 97% decrease in tumor size in the remaining mice. Analysis of miRNA expression in non-tumoral and tumoral colonic tissue of resveratrol treated mice showed an increased expression of miR-96, a miRNA previously shown to regulate Kras translation. These data indicate that resveratrol can prevent the formation and growth of colorectal tumors by downregulating Kras expression., (Published by Oxford University Press 2014.)

A cell-based high-throughput screen that assessed the cellular stability of a tumor suppressor protein PDCD4 (Programmed cell death 4) was used to identify a new guanidine-containing marine alkaloid mirabilin K (3), as well as the known compounds mirabilin G (1) and netamine M (2). The structures of these tricyclic guanidine alkaloids were established from extensive spectroscopic analyses. Compounds 1 and 2 inhibited cellular degradation of PDCD4 with EC50 values of 1.8 μg/mL and 2.8 μg/mL, respectively. Mirabilin G (1) and netamine M (2) are the first marine natural products reported to stabilize PDCD4 under tumor promoting conditions.

Background: The small molecule NSC676914A was previously identified as an NF-κB inhibitor in TPA-stimulated HEK293 cells (Mol Can Ther 8:571-581, 2009). We hypothesized that this effect would also be seen in ovarian cancer cells, and serve as its mechanism of cytotoxicity. OVCAR3 and HEK293 cell lines stably containing a NF-κB luciferase reporter gene were generated., Methods: Levels of NF-κB activity were assessed by luciferase reporter assays, after stimulation with LPA, LPS, TPA, and TNFα, in the presence or absence of a known NF-κB inhibitor or NSC676914A, and cytotoxicity was measured., Results: NSC676914A was toxic to both OVCAR3 and HEK293 cells. We also investigated the cytotoxicity of NSC676914A on a panel of lymphoma cell lines with well characterized mutations previously shown to determine sensitivity or resistance to NF-κB inhibition. The compound did not show predicted patterns of effects on NF-κB activity in either lymphoma, ovarian or HEK293 cell lines. In HEK293 cells, the small molecule inhibited NF-κB when cells were stimulated, while in OVCAR3 cells it only partially inhibited NF-κB. Interestingly, we observed rescue of cell death with ROS inhibition., Conclusions: The current study suggests that the effect of NSC676914A on NF-κB depends on cell type and the manner in which the pathway is stimulated. Furthermore, as it is similarly toxic to lymphoma, OVCAR3 and HEK293 cells, NSC676914A shows promising NF-κB-independent anti-cancer activity in ovarian tumor cells.

PDCD4 is a tumor suppressor induced by apoptotic stimuli that regulates both translation and transcription. Previously, we showed that overexpression of PDCD4 leads to decreased anchorage-independent growth in glioblastoma (GBM)-derived cell lines and decreased tumor growth in a GBM xenograft model. In inflammatory cells, PDCD4 stimulates tumor necrosis factor-induced activation of the transcription factor NF-κB, an oncogenic driver in many cancer sites. However, the effect of PDCD4 on NF-κB transcriptional activity in most cancers including GBM is still unknown. We studied the effect of PDCD4 on NF-κB-dependent transcriptional activity in GBM by stably overexpressing PDCD4 in U251 and LN229 cells. Stable PDCD4 expression inhibits NF-κB transcriptional activation measured by a luciferase reporter. The molecular mechanism by which PDCD4 inhibits NF-κB transcriptional activation does not involve inhibited expression of NF-κB p65 or p50 proteins. PDCD4 does not inhibit pathways upstream of NF-κB including the activation of IKKα and IKKβ kinases or degradation of IκBα, events needed for nuclear transport of p65 and p50. PDCD4 overexpression does inhibit localization of p65 but not p50 in the nucleus. PDCD4 protein interacts preferentially with p65 protein as shown by co-immunoprecipitation and confocal imaging. PDCD4 overexpression inhibits the mRNA expression of two NF-κB target genes in a p65-dependent manner. These results suggest that PDCD4 can significantly inhibit NF-κB activity in GBM cells by a mechanism that involves direct or indirect protein-protein interaction independent of the expected mRNA-selective translational inhibition. These findings offer novel opportunities for NF-κB-targeted interventions to prevent or treat cancer., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Sulfiredoxin (Srx), the exclusive enzyme that reduces the hyperoxidized inactive form of peroxiredoxins (Prxs), has been found highly expressed in several types of human skin cancer. To determine whether Srx contributed to skin tumorigenesis in vivo, Srx null mice were generated on an FVB background. Mouse skin tumorigenesis was induced by a 7,12-dimethylbenz[α]anthracene/12-O-tetradecanoylphorbol-13-acetate (DMBA/TPA) protocol. We found that the number, volume and size of papillomas in Srx(-/-) mice were significantly fewer compared with either wild-type (Wt) or heterozygous (Het) siblings. Histopathological analysis revealed more apoptotic cells in tumors from Srx(-/-) mice. Mechanistic studies in cell culture revealed that Srx was stimulated by TPA in a redox-independent manner. This effect was mediated transcriptionally through the activation of mitogen-activated protein kinase and Jun-N-terminal kinase. We also demonstrated that Srx was capable of reducing hyperoxidized Prxs to facilitate cell survival under oxidative stress conditions. These findings suggested that loss of Srx protected mice, at least partially, from DMBA/TPA-induced skin tumorigenesis. Therefore, Srx has an oncogenic role in skin tumorigenesis and targeting Srx may provide novel strategies for skin cancer prevention or treatment.

Deregulation of the phosphatidylinositol 3-kinase (PI3K)-Akt-mammalian target of rapamycin (mTOR)-70kDa ribosomal protein S6 kinase 1 (p70(S6K)) pathway is commonly observed in many tumors. This pathway controls proliferation, survival, and translation, and its overactivation is associated with poor prognosis for tumor-associated survival. Current efforts focus on the development of novel inhibitors of this pathway. In a cell-based high-throughput screening assay of 15,272 pure natural compounds, we identified pomiferin triacetate as a potent stabilizer of the tumor suppressor programmed cell death 4 (Pdcd4). Mechanistically, pomiferin triacetate appeared as a general inhibitor of the PI3K-Akt-mTOR-p70(S6K) cascade. Interference with this pathway occurred downstream of Akt but upstream of p70(S6K). Specifically, mTOR kinase emerged as the molecular target of pomiferin triacetate, with similar activities against mTOR complexes 1 and 2. In an in vitro mTOR kinase assay pomiferin triacetate dose-dependently inhibited mTOR with an IC50 of 6.2 μM. Molecular docking studies supported the interaction of the inhibitor with the catalytic site of mTOR. Importantly, pomiferin triacetate appeared to be highly selective for mTOR compared to a panel of 17 lipid and 50 protein kinases tested. As a consequence of the mTOR inhibition, pomiferin triacetate efficiently attenuated translation. In summary, pomiferin triacetate emerged as a novel and highly specific mTOR inhibitor with strong translation inhibitory effects. Thus, it might be an interesting lead structure for the development of mTOR- and translation-targeted anti-tumor therapies., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Analysis of the Polyp Prevention Trial showed an association between an isorhamnetin-rich diet and a reduced risk of advanced adenoma recurrence; however, the mechanism behind the chemoprotective effects of isorhamnetin remains unclear. Here, we show that isorhamnetin prevents colorectal tumorigenesis of FVB/N mice treated with the chemical carcinogen azoxymethane and subsequently exposed to colonic irritant dextran sodium sulfate (DSS). Dietary isorhamnetin decreased mortality, tumor number, and tumor burden by 62%, 35%, and 59%, respectively. MRI, histopathology, and immunohistochemical analysis revealed that dietary isorhamnetin resolved the DSS-induced inflammatory response faster than the control diet. Isorhamnetin inhibited AOM/DSS-induced oncogenic c-Src activation and β-catenin nuclear translocation, while promoting the expression of C-terminal Src kinase (CSK), a negative regulator of Src family of tyrosine kinases. Similarly, in HT-29 colon cancer cells, isorhamnetin inhibited oncogenic Src activity and β-catenin nuclear translocation by inducing expression of csk, as verified by RNA interference knockdown of csk. Our observations suggest the chemoprotective effects of isorhamnetin in colon cancer are linked to its anti-inflammatory activities and its inhibition of oncogenic Src activity and consequential loss of nuclear β-catenin, activities that are dependent on CSK expression.

Objectives: Screening colonoscopy to monitor for early colitis-associated colon cancer (CAC) is difficult due to the aberrant mucosal patterns associated with long-standing colitis. The aim of this study was to develop a rapid fluorescent detection method for use during colonoscopy for improving the detection of CAC utilising a topically applied enzymatically activatable probe (gGlu-HMRG) which fluoresces in the presence of γ-glutamyltranspeptidase (GGT), an enzyme associated with cancer., Methods: Expression of GGT in colon cell lines was examined with fluorescence microscopy and flow cytometry. A mouse model (azoxymethane/dextran sulphate sodium) of CAC was used and mice were examined with white light and fluorescence colonoscopy before and after topical gGlu-HMRG administration., Results: Expression of GGT, although variable, was higher in human colon cancer cells than normal human colon cells. Using fluorescence colonoscopy in mice, gGlu-HMRG fluorescent lesions were detected 5 min after topical administration and fluorescence persisted for at least 30 min. Fluorescence guided biopsy revealed all fluorescent lesions that contained cancer or dysplasia (n=16), whereas three out of 12 non-fluorescent lesions contained low grade dysplasia and others did not contain neoplastic histology. Microscopic inflammatory infiltration also had variable fluorescence but in general was much lower (∼10-fold) in signal than cancer. Repeat fluorescence endoscopy allowed individual tumours to be monitored., Conclusion: These results suggest that gGlu-HMRG can improve endoscopic detection of CAC with a higher target to background ratio than conventional white light colonoscopy. This could be of benefit to patients with long-standing colitis who must undergo repeated screening colonoscopies.

Recent studies have suggested that changes in serum phosphate levels influence pathological states associated with aging such as cancer, bone metabolism, and cardiovascular function, even in individuals with normal renal function. The causes are only beginning to be elucidated but are likely a combination of endocrine, paracrine, autocrine, and cell autonomous effects. We have used an integrated quantitative biology approach, combining transcriptomics and proteomics to define a multi-phase, extracellular phosphate-induced, signaling network in pre-osteoblasts as well as primary human and mouse mesenchymal stromal cells. We identified a rapid mitogenic response stimulated by elevated phosphate that results in the induction of immediate early genes including c-fos. The mechanism of activation requires FGF receptor signaling followed by stimulation of N-Ras and activation of AP-1 and serum response elements. A distinct long-term response also requires FGF receptor signaling and results in N-Ras activation and expression of genes and secretion of proteins involved in matrix regulation, calcification, and angiogenesis. The late response is synergistically enhanced by addition of FGF23 peptide. The intermediate phase results in increased oxidative phosphorylation and ATP production and is necessary for the late response providing a functional link between the phases. Collectively, the results define elevated phosphate, as a mitogen and define specific mechanisms by which phosphate stimulates proliferation and matrix regulation. Our approach provides a comprehensive understanding of the cellular response to elevated extracellular phosphate, functionally connecting temporally coordinated signaling, transcriptional, and metabolic events with changes in long-term cell behavior., (Copyright © 2012 Wiley Periodicals, Inc.)

Sulfiredoxin (Srx) is the enzyme that reduces the hyperoxidized inactive form of peroxiredoxins. To study the function of Srx in carcinogenesis in vivo, we tested whether loss of Srx protects mice from cancer development. Srx null mice were generated and colon carcinogenesis was induced by an azoxymethane (AOM) and dextran sulfate sodium (DSS) protocol. Compared with either wild-type (Wt) or heterozygotes, Srx(-/-) mice had significantly reduced rates in both tumor multiplicity and volume. Mechanistic studies reveal that loss of Srx did not alter tumor cell proliferation; however, increased apoptosis and decreased inflammatory cell infiltration were obvious in tumors from Srx null mice compared with those from Wt control. In addition to the AOM/DSS model, examination of Srx expression in human reveals a tissue-specific expression pattern. Srx expression was also demonstrated in tumors from colorectal cancer patients and the levels of expression were associated with patients' clinic stages. These data provide the first in vivo evidence that loss of Srx renders mice resistant to AOM/DSS-induced colon carcinogenesis, suggesting that Srx has a critical oncogenic role in cancer development, and Srx may be used as a marker for human colon cancer pathogenicity.

Background: Prenatal exposure to ethanol (EtOH) elicits a range of neuro-developmental abnormalities, microcephaly to behavioral deficits. Impaired protein synthesis has been connected to pathogenesis of EtOH-induced brain damage and abnormal neuron development. However, mechanisms underlying these impairments of protein synthesis are not known. In this study, we illustrate the effects of EtOH on programmed cell death protein 4 (PDCD4), a tumor and translation repressor., Methods: Primary cortical neurons (PCNs) were treated with 2.5 and 4 mg/ml EtOH for different time points (4 to 24 hours), and PDCD4 expression was detected by Western blotting. Protein synthesis was determined using [(35) S] methionine incorporation assay. Methyl cap pull-down assay was performed to establish the effect of EtOH on association of eukaryotic initiation factor 4A (eIF4A) with capped mRNA. Luciferase assay was performed to determine the in vivo translation. A 2-day acute 5-dose binge model with EtOH (4 g/kg body wt, 25% v/v) was performed in Sprague-Dawley rats at 12-hour intervals and analyzed for PDCD4, eIF4A, and eIF4A-methyl cap association., Results: EtOH increased PDCD4 expression in a time- and dose-dependent manner in PCNs, which inhibited the association of eIF4A with methyl cap. EtOH and ectopic PDCD4 expression suppressed in vivo translation in PCNs and RNAi targeting of PDCD4 blocked the inhibitory effect of EtOH on protein synthesis. In utero exposure of pregnant rats to EtOH resulted in a significant increase in PDCD4 in fetal cerebral cortex along with the inhibition of methyl cap-associated eIF4A, compared with isocaloric controls. Increased PDCD4 also occurred in pooled fractions of remaining brain regions., Conclusions: Our data, for the first time, illustrate that PDCD4 mediates inhibitory effects of EtOH on protein synthesis in PCNs and developing brain., (Copyright © 2012 by the Research Society on Alcoholism.)

Oral consumption of freeze-dried black raspberries attenuated neoplastic changes in colorectal tissue markers of apoptosis, cell proliferation, and angiogenesis in colorectal cancer (CRC) patients. To determine whether plasma concentrations of interleukin (IL)-1β, IL-2, IL-6, IL-8, IL-10, IL-12p70, granulocyte macrophage colony stimulating factor (GM-CSF), interferon-γ, and tumor necrosis factor-α (TNF-α) were associated with berry treatment and changes in colorectal tissue markers of apoptosis, cell proliferation, and angiogenesis, plasma and biopsy samples of adenocarcinoma and adjacent normal-appearing colorectal tissue were collected before and during berry treatment from 24 CRC patients who had not received prior therapy and drank a slurry of black raspberry powder (20 g in 100 ml drinking water) 3 times a day for 1 to 9 wk. Plasma concentrations of GM-CSF (+0.12 ± 0.04 pg/mL; P = 0.01) and IL-8 (-1.61 ± 0.71 pg/mL; P = 0.04) changed in patients receiving berries for more than 10 days. These changes were correlated with beneficial changes in markers of proliferation (r(ΔGM-CSF, ΔKi67 carcinoma - normal) = -0.51) and apoptosis (r(ΔIL-8, ΔTUNEL carcinoma - normal) = -0.52) observed in colorectal tissue taken within the same week. Plasma concentrations of GM-CSF and IL-8 may serve as noninvasive indicators to monitor tissue response to berry-based interventions for CRC.

Cancer stem cells often have phenotypic and functional characteristics similar to normal stem cells including the properties of self-renewal and differentiation. Recent findings suggest that uncontrolled self-renewal may explain cancer relapses and may represent a critical target for cancer prevention. It is conceivable that the loss of regulatory molecules resulting from inappropriate consumption of specific foods and their constituents may foster the aberrant self-renewal of cancer stem cells. In fact, increasing evidence points to the network delivering signals for self-renewal from extracellular compartments to the nucleus including changes in stem cell environments, inducible expression of microRNAs, hyperplastic nuclear chromatin structures, and the on/off of differentiation process as possible sites of action for bioactive food components. Diverse dietary constituents such as vitamins A and D, genistein, (-)-epigallocatechin-3-gallate (EGCG), sulforaphane, curcumin, piperine, theanine and choline have been shown to modify self-renewal properties of cancer stem cells. The ability of these bioactive food components to influence the balance between proliferative and quiescent cells by regulating critical feedback molecules in the network including dickkopf 1 (DKK-1), secreted frizzled-related protein 2 (sFRP2), B cell-specific Moloney murine leukemia virus integration site 1 (Bmi-1) and cyclin-dependent kinase 6 (CDK6) may account for their biological response. Overall, the response to food components does not appear to be tissue or organ specific, suggesting there may be common cellular mechanisms. Unquestionably, additional studies are needed to clarify the physiological role of these dietary components in preventing the resistance of tumor cells to traditional drugs and cancer recurrence., (Published by Elsevier Inc.)

The transcription factor AP-1 (activator protein-1) regulates a number of genes that drive tumor promotion and progression. While basal levels of AP-1 activity are important for normal cell proliferation and cell survival, overactivated AP-1-dependent gene expression stimulates inflammation, angiogenesis, invasion, and other events that propel carcinogenesis. We seek to discover genes targeted by carcinogenesis inhibitors that do not also inhibit cell proliferation or survival. Transgenic TAM67 (dominant-negative c-Jun) inhibits mouse skin tumorigenesis and tumor progression without inhibiting cell proliferation or induced hyperproliferation. Expression profiling of wild-type and K14-TAM67 mouse epidermis has revealed a number of functionally significant genes that are induced by tumor promoters in wild-type mice but not in those expressing the AP-1 blocker. The current study now identifies Wnt5a signaling as a new target of TAM67 when it inhibits DMBA/TPA-induced carcinogenesis. Wnt5a is required to maintain the tumor phenotype in tumorigenic mouse JB6 cells and Ras-transformed human squamous carcinoma HaCaT-II4 cells, as Wnt5a knockdown suppresses anchorage-independent and tumor xenograft growth. The oncogenic Wnt5a-mediated pathway signals through activation of the protein kinase PKCα and oncogenic transcription factor STAT3 phosphorylation and not through the canonical Wnt/β-catenin pathway. Similar to Wnt5a knockdown, inhibitors of PKCα blocked STAT3 activation in both mouse JB6 and human HaCaT-II4 tumor cells. Moreover, expression of STAT3-regulated genes FAS, MMP3, IRF1, and cyclin D1 was suppressed with Wnt5a knockdown. Treatment of mouse Wnt5a knockdown cells with a PKCα-specific activator rescued phosphorylation of STAT3. Thus, Wnt5a signaling is required for maintaining the tumor phenotype in squamous carcinoma cells, Wnt5a targeting by the AP-1 blockade contributes to inhibition of skin carcinogenesis, and the signaling pathway traverses PKCα and STAT3 activation. Coordinate overactivation of Wnt5a expression and STAT3 signaling is observed in human skin and colon cancers as well as glioblastoma.

Loss of the tumor suppressor Pdcd4 was reported for various tumor entities and proposed as a prognostic marker in tumorigenesis. We previously characterized decreased Pdcd4 protein stability in response to mitogenic stimuli, which resulted from p70(S6K1)-dependent protein phosphorylation, β-TrCP1-mediated ubiquitination, and proteasomal destruction. Following high-throughput screening of natural product extract libraries using a luciferase-based reporter assay to monitor phosphorylation-dependent proteasomal degradation of the tumor suppressor Pdcd4, we succeeded in showing that a crude extract from Eriophyllum lanatum stabilized Pdcd4 from TPA-induced degradation. Erioflorin was identified as the active component and inhibited not only degradation of the Pdcd4-luciferase-based reporter but also of endogenous Pdcd4 at low micromolar concentrations. Mechanistically, erioflorin interfered with the interaction between the E3-ubiquitin ligase β-TrCP1 and Pdcd4 in cell culture and in in vitro binding assays, consequently decreasing ubiquitination and degradation of Pdcd4. Interestingly, while erioflorin stabilized additional β-TrCP-targets (such as IκBα and β-catenin), it did not prevent the degradation of targets of other E3-ubiquitin ligases such as p21 (a Skp2-target) and HIF-1α (a pVHL-target), implying selectivity for β-TrCP. Moreover, erioflorin inhibited the tumor-associated activity of known Pdcd4- and IκBα-regulated αtranscription factors, that is, AP-1 and NF-κB, altered cell cycle progression and suppressed proliferation of various cancer cell lines. Our studies succeeded in identifying erioflorin as a novel Pdcd4 stabilizer that inhibits the interaction of Pdcd4 with the E3-ubiquitin ligase β-TrCP1. Inhibition of E3-ligase/target-protein interactions may offer the possibility to target degradation of specific proteins only as compared to general proteasome inhibition.

We have accomplished a parallel screen of cycloaddition partners for o-quinols utilizing a plate-based microwave system. Microwave irradiation improves the efficiency of retro-Diels-Alder/Diels-Alder cascades of o-quinol dimers which generally proceed in a diastereoselective fashion. Computational studies indicate that asynchronous transition states are favored in Diels-Alder cycloadditions of o-quinols. Subsequent biological evaluation of a collection of cycloadducts has identified an inhibitor of activator protein-1 (AP-1), an oncogenic transcription factor.

The tumor suppressor programmed cell death 4 (Pdcd4) is lost in various tumor tissues. Loss of Pdcd4 has been associated with increased tumorigenic potential and tumor progression. While various mechanisms of Pdcd4 regulation have been described, the effect of an inflammatory tumor microenvironment on Pdcd4 protein expression has not been characterized so far. In the present study, we aimed to elucidate the molecular mechanisms of Pdcd4 protein regulation in tumor cells under inflammatory conditions. 12-O-tetradecanoylphorbol 13-acetate-induced differentiation of human U937 monocytes increased the expression and secretion of inflammatory cytokines such as tumor necrosis factor α, interleukin (IL)-6 and IL-8. Exposure to conditioned medium (CM) of these activated macrophages markedly decreased Pdcd4 protein expression in various tumor cells. Similarly, indirect coculture with such activated U937 monocyte-derived macrophages resulted in the loss of Pdcd4 protein in tumor cells. Decreased Pdcd4 protein levels were attributable to enhanced proteasomal degradation, diminishing Pdcd4 protein half-life. Proteasomal degradation required activation of phosphatidylinositol-3-kinase (PI3K)-mammalian target of rapamycin (mTOR) signaling. Since macrophage-CM sufficed to induce Pdcd4 degradation, Pdcd4 downregulation was determined to be an indirect unidirectional effect of the macrophages on the tumor cells. Pdcd4 protein expression was also attenuated in vivo in mouse colon tissue in response to dextran sodium sulfate-induced colitis. In summary, we characterized PI3K-mTOR-dependent proteasome-mediated Pdcd4 degradation in tumor cells in the inflammatory tumor microenvironment. Consequently, stabilization of Pdcd4 protein could provide a promising novel avenue for therapeutics targeting inflammation-associated tumors.

Our current natural product program utilizes new actinomycetes originating from unexplored and underexplored ecological niches, employing cytotoxicity against a selected panel of cancer cell lines as the preliminary screen to identify hit strains for natural product dereplication, followed by mechanism-based assays of the purified natural products to discover potential anticancer drug leads. Three new linear polyketides, actinopolysporins A (1), B (2), and C (3), along with the known antineoplastic antibiotic tubercidin (4), were isolated from the halophilic actinomycete Actinopolyspora erythraea YIM 90600, and the structures of the new compounds were elucidated on the basis of spectroscopic data interpretation. All four compounds were assayed for their ability to stabilize the tumor suppressor programmed cell death protein 4 (Pdcd4), which is known to antagonize critical events in oncogenic pathways. Only 4 significantly inhibited proteasomal degradation of a model Pdcd4-luciferase fusion protein, with an IC50 of 0.88±0.09 μM, unveiling a novel biological activity for this well-studied natural product.

Grassypeptolides F (1) and G (2), bis-thiazoline-containing cyclic depsipeptides with a rare β-amino acid, extensive N-methylation, and a large number of d-amino acids, are reported from an extract of the Palauan cyanobacterium Lyngbya majuscula. Both 1 and 2 were found to have moderate inhibitory activity against the transcription factor AP-1 (IC₅₀ = 5.2 and 6.0 μM, respectively).

A high throughput screen for inhibitors of the oncogenic transcription factor activator protein-1 (AP-1) was applied to the NCI repository of natural product extracts. The liphophilic extract of the plant Nothospondias staudtii (Simaroubaceae) displayed significant AP-1 inhibition. Bioassay-guided fractionation of the extract lead to a new quassinoid named nothospondin (1), and the known compound glaucarubinone (2). The structure of 1 was elucidated by spectroscopic methods. Compounds 1 and 2 showed potent, dose-dependent AP-1 inhibition at noncytotoxic concentrations., (Published by Elsevier Ltd.)

MicroRNAs (miRNAs) are small, noncoding RNAs that play a critical role in developmental and physiological processes and are implicated in the pathogenesis of several human diseases, including cancer. They function by regulating target gene expression post-transcriptionally. In this study, we examined the role of oncogenic mir-21 in the pathogenesis of glioblastoma, the most aggressive form of primary brain tumor. We have previously reported that mir-21 is expressed at higher levels in primary glioblastoma-tissue and glioblastoma-derived cell lines than in normal brain tissue. We demonstrate that downregulation of mir-21 in glioblastoma-derived cell lines results in increased expression of its target, programmed cell death 4 (Pdcd4), a known tumor-suppressor gene. In addition, our data indicate that either downregulation of mir-21 or overexpression of its target, Pdcd4, in glioblastoma-derived cell lines leads to decreased proliferation, increased apoptosis, and decreased colony formation in soft agar. Using a glioblastoma xenograft model in immune-deficient nude mice, we observe that glioblastoma-derived cell lines in which mir-21 levels are downregulated or Pdcd4 is over-expressed exhibit decreased tumor formation and growth. Significantly, tumors grow when the glioblastoma-derived cell lines are transfected with anti-mir-21 and siRNA to Pdcd4, confirming that the tumor growth is specifically regulated by Pdcd4. These critical in vivo findings demonstrate an important functional linkage between mir-21 and Pdcd4 and further elucidate the molecular mechanisms by which the known high level of mir-21 expression in glioblastoma can attribute to tumorigenesis--namely, inhibition of Pdcd4 and its tumor-suppressive functions.

A high-throughput cell-based reporter assay designed to identify small-molecule stabilizers of the tumor suppressor Pdcd4 was used to screen extracts in the NCI Natural Products Repository. Bioassay-guided fractionation of an extract from a Papua New Guinea collection of the tropical tree Cryptocarya sp. provided a series of new 5,6-dihydro-α-pyrone-containing 1,3-polyols (1-8), named cryptocaryols A-H. Their structures were assigned from a combination of NMR, MS, and CD studies in conjunction with NMR database comparisons. Compounds 1-8 were found to rescue Pdcd4 from TPA-induced degradation with EC50 concentrations that ranged from 1.3 to 4.9 μM.

Oxidative stress is known to cause tumorigenesis through induction of DNA and lipid damage. It also promotes cancer progression through a largely unknown mechanism. Sulfiredoxin (Srx) is a novel oxidative stress-induced antioxidant protein whose function in tumorigenesis and cancer progression has not been well studied. We report that Srx is highly expressed in human lung cancer. Knockdown of Srx reduces anchorage-independent colony formation, cell migration, and invasion of human lung cancer cells. Srx preferentially interacts with Peroxiredoxin (Prx) IV relative to other Prxs due to its intrinsic higher binding affinity. Knockdown of Prx IV recapitulates the phenotypic changes of depleting Srx. Disruption or enhancement of the Srx-Prx IV axis leads respectively to reduction or acceleration of tumor growth and metastasis formation in vivo. Through identification and validation of the downstream mediators we unraveled the Srx-mediated signaling network that traverses AP-1-activating and other phosphokinase signaling cascades. Our work reveals that the Srx-Prx IV axis is critical for lung cancer maintenance and metastasis, suggesting that targeting the Srx-Prx IV axis may provide unique effective strategies for cancer prevention and treatment.

An activator protein-1 (AP-1) based bioassay-guided phytochemical investigation on Podocarpus latifolius led to the isolation of three new sempervirol-type diterpenes, cycloinumakiol (1), inumakal (2), and inumakoic acid (3), along with three known norditerpenes (4-6). Compounds 4 and 6 were responsible for the observed bioactivity.

Chemopreventive dietary compounds, such as flavonols, may inhibit colorectal carcinogenesis partly by altering cytokine expression and attenuating inflammation. Single nucleotide polymorphisms (SNPs) in the promoter regions of genes encoding cytokines may influence flavonol-induced changes in cytokine expression and consequently cancer risk. Using logistic regression, we estimated odds ratios (OR) and 95% confidence intervals (CI) for the association between SNPs of interleukin (IL)-1β, 6, 8, and 10 alone or combined with flavonol intake or serum IL concentration changes, and adenoma recurrence in 808 participants from the intervention arm of the Polyp Prevention Trial, a 4-year intervention study evaluating the effectiveness of a low-fat, high-fiber, high-fruit and vegetable diet on adenoma recurrence. Overall, SNPs in genes encoding IL-1β, 6, 8, and 10 were not associated with their corresponding serum concentrations or adenoma recurrence. However, individuals homozygous for IL-10 -592 C (OR=2.23, 95% CI: 1.07-4.66, P(interaction)=0.03) orIL-10 -819 C (OR=2.18, 95% CI: 1.05-4.51, P(interaction)=0.05) had an elevated risk of high-risk adenoma recurrence when their serum IL-10 concentrations increased during the trial. In addition, IL-6 -174 GG in combination with above median flavonol intake (OR=0.14, 95% CI: 0.03-0.66) or with decreased IL-6 concentrations (OR=0.14, 95% CI: 0.03-0.65) reduced the risk of advanced adenoma recurrence, although the interaction term was not statistically significant. In conclusion, our results suggest that IL SNPs, in combination with a flavonol-rich diet or decreased serum IL, may lower the risk of adenoma recurrence.

Background: Serum cytokine concentrations may reflect inflammatory processes occurring during the development of colorectal neoplasms. Flavonols, bioactive compounds found in plant-based foods and beverages, may inhibit colorectal neoplasms partly by attenuating inflammation., Methods: Using logistic regression, we estimated odds ratios (ORs) and 95% confidence intervals (CIs) to investigate the association between serum concentrations of interleukin (IL) β, 2, 8, 10, 12p70, granulocyte macrophage colony stimulating factor, interferon-γ, and tumour necrosis factor-α, measured over time, flavonol intake, estimated from a flavonol database used in conjunction with a food frequency questionnaire, and adenoma recurrence in 872 participants from the intervention arm of the Polyp Prevention Trial., Results: Decreased IL-2 concentration during the trial increased the risk of any adenoma recurrence (4th vs 1st quartile, OR=1.68, 95% CI=1.13-2.49), whereas decreased IL-1β or IL-10 reduced the risk of advanced adenoma recurrence (OR=0.37, 95% CI=0.15-0.94; OR=0.39, 95% CI=0.15-0.98, respectively). Individuals with flavonol intake above the median (29.7 mg per day) and decreased cytokine concentrations had the lowest risk of advanced adenoma recurrence., Conclusion: Overall, no consistent associations were observed between serum cytokine profile and colorectal adenoma recurrence; however, decreased cytokine concentrations during high flavonol consumption may indicate prevention of colorectal neoplasms.

Clinical studies have shown that fiber consumption facilitates weight loss and improves lipid profiles; however, the beneficial effects of high fermentable fiber low glycemic index (GI) diets under conditions of weight maintenance are unclear. In the Legume Inflammation Feeding Experiment, a randomized controlled cross-over feeding study, 64 middle-aged men who had undergone colonoscopies within the previous 2 years received both a healthy American (HA) diet (no legume consumption, fiber consumption = 9 g/1,000 kcal, and GI = 69) and a legume enriched (1.5 servings/1,000 kcal), high fiber (21 g/1,000 kcal), low GI (GI = 38) diet (LG) in random order. Diets were isocaloric and controlled for macronutrients including saturated fat; they were consumed each for 4 weeks with a 2-4 week break separating dietary treatments. Compared to the HA diet, the LG diet led to greater declines in both fasting serum total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) (P < 0.001 and P < 0.01, respectively). Insulin-resistant (IR) subjects had greater reductions in high density lipoprotein cholesterol (HDL-C; P < 0.01), and triglycerides (TAG)/HDL-C (P = 0.02) after the LG diet, compared to the HA diet. Insulin-sensitive (IS) subjects had greater reductions in TC (P < 0.001), LDL-C (P < 0.01), TC/HDL-C (P < 0.01), and LDL-C/HDL-C (P = 0.02) after the LG diet, compared to the HA diet. In conclusion, a high legume, high fiber, low GI diet improves serum lipid profiles in men, compared to a healthy American diet. However, IR individuals do not achieve the full benefits of the same diet on cardiovascular disease (CVD) lipid risk factors.

Lung cancer is the leading cause of cancer mortality in the United States and worldwide. The identification of key regulatory and molecular mechanisms involved in lung tumorigenesis is therefore critical to increase our understanding of this disease and could ultimately lead to targeted therapies to improve prevention and treatment. Induction of members of the activator protein-1 (AP-1) transcription factor family has been described in human non-small cell lung carcinoma. Activation of AP-1 can either stimulate or repress transcription of multiple gene targets, ultimately leading to increased cell proliferation and inhibition of apoptosis. In the present study, we show induction of AP-1 in carcinogen-induced mouse lung tumors compared with surrounding normal lung tissue. We then used a transgenic mouse model directing conditional expression of the dominant-negative c-jun mutant TAM67 in lung epithelial cells to determine the effect of AP-1 inhibition on mouse lung tumorigenesis. Consistent with low AP-1 activity in normal lung tissue, TAM67 expression had no observed effects in adult mouse lung. TAM67 decreased tumor number and overall lung tumor burden in chemically induced mouse lung tumor models. The most significant inhibitory effect was observed on carcinoma burden compared with lower-grade lesions. Our results support the concept that AP-1 is a key regulator of mouse lung tumorigenesis, and identify AP-1-dependent transcription as a potential target to prevent lung tumor progression.

Ample evidence has shown key roles of inflammation in tumor promotion and carcinogenesis, and tumor-associated macrophages are known to promote tumor growth and dissemination. Programmed cell death 4 (Pdcd4) is a novel tumor suppressor, and although various studies have revealed that the functions and expression mechanisms of Pdcd4 in tumor promotion, those in regard to inflammation remain unclear. In the present study, we examined whether inflammatory stimuli regulate Pdcd4 expression. 12-O-tetradecanoylphorbol 13-acetate (TPA) suppressed expression of pdcd4 mRNA in human monocytic cell lines (U937, THP-1). Similarly, the bacterial endotoxin lipopolysaccharide (LPS) downregulated pdcd4 level in mouse RAW264.7 and peritoneal macrophages. Furthermore, conditioned medium from LPS-stimulated RAW264.7 macrophages suppressed pdcd4 mRNA in RAW264.7 macrophages, and findings obtained with recombinant tumor necrosis factor-alpha (TNF-alpha) and TNF-alpha-specific siRNA suggested that TNF-alpha partly mediates LPS-triggered Pdcd4 downregulation via an autocrine mechanism. Specific inhibitors of phosphoinositide-3-kinase (PI3K) and c-jun N-terminus kinase (JNK) restored LPS-abolished pdcd4 mRNA. Consistently, in MCF7 mammary carcinoma cells, conditioned medium from TPA-differentiated/activated U937 cells suppressed pdcd4 mRNA. Additionally, knockdown of pdcd4 in RAW264.7 macrophages using siRNA significantly enhanced LPS-induced TNF-alpha protein production, and interferon-gamma, CC chemokine ligand (Ccl) 1, Ccl20, and interleukin-10 mRNA expression. These results suggest that Pdcd4 suppresses the induction of these inflammatory mediators. Taken together, loss of Pdcd4 in macrophages may be a critical step in establishing the inflammatory environment while that in tumor cells contributes to tumor progression., (2010 Wiley-Liss, Inc.)

Metastasis to bone, liver and lungs is the primary cause of death in breast cancer patients. Our studies have revealed that the novel tumor suppressor Pdcd4 inhibits breast cancer cell migration and invasion in vitro. Loss of Pdcd4 in human nonmetastatic breast cancer cells increased the expression of lysyl oxidase (LOX) mRNA. LOX is a hypoxia-inducible amine oxidase, the activity of which enhances breast cancer cell invasion in vitro and in vivo. Specific inhibition of LOX activity by beta-aminopropionitrile or small interfering RNA decreased the invasiveness of T47D and MCF7 breast cancer cells attenuated for Pdcd4 function. Most significantly, loss of Pdcd4 augments hypoxia induction of LOX as well. Conversely, overexpression of Pdcd4 significantly reversed the hypoxia induction of LOX expression in T47D cells attenuated for Pdcd4. However, Pdcd4 did not affect hypoxia-inducible factor-1 (HIF-1) protein expression or HIF-1-responsive element-luciferase activity in response to hypoxia, suggesting that Pdcd4 regulation of LOX occurs through an HIF-independent mechanism. Nevertheless, the loss of Pdcd4 early in cancer progression may have an important role in the increased sensitivity of cancer cells to hypoxia through increased LOX activity and concomitant enhanced invasiveness.

Serum interleukin-6 (IL-6), a proinflammatory cytokine, is considered an indicator of inflammation and may be an indicator of colorectal carcinogenesis given that inflammation can promote carcinogenesis. Flavonols, which can be found in fruits and vegetables, may inhibit colorectal carcinogenesis partly by inhibiting inflammation. We estimated odds ratios and 95% confidence intervals (95% CI) to determine whether serum IL-6 was associated with colorectal adenoma recurrence and flavonol intake and thus may serve as a risk indicator and as a response indicator to dietary flavonols. Serum IL-6 concentrations at baseline, year 1, and year 3 were measured in 872 participants from the intervention arm of the Polyp Prevention Trial, a 4-year trial that examined the effectiveness of a low-fat, high-fiber, high-fruit and vegetable diet on adenoma recurrence. Intake of flavonols, especially of isorhamnetin, kaempferol, and quercetin, was inversely associated with serum IL-6 concentrations (highest versus lowest flavonol intake quartile, 1.80 versus 2.20 pg/mL) and high-risk (OR, 0.51; 95% CI, 0.26-0.98) and advanced adenoma recurrence (OR, 0.17; 95% CI, 0.06-0.50). A decrease in IL-6 concentration during the trial was inversely associated with high-risk (OR, 0.44; 95% CI, 0.23-0.84) and advanced adenoma recurrence (OR, 0.47; 95% CI, 0.19-1.18). Individuals with above median flavonol intake and equal or below median IL-6 change after baseline had the lowest risk of recurrence of high-risk and advanced adenoma. Our results suggest that serum IL-6 may serve as a risk indicator and as a response indicator to dietary flavonols for colorectal cancer prevention., (2010 AACR.)

Signal transducer and activator of transcription 2 (STAT2) is an essential transcription factor in the type I IFN (IFN-alpha/beta) signal transduction pathway and known for its role in mediating antiviral immunity and cell growth inhibition. Unlike other members of the STAT family, IFNs are the only cytokines known to date that can activate STAT2. Given the inflammatory and antiproliferative dual nature of IFNs, we hypothesized that STAT2 prevents inflammation-induced colorectal and skin carcinogenesis by altering the inflammatory immune response. Contrary to our hypothesis, deletion of STAT2 inhibited azoxymethane/dextran sodium sulfate-induced colorectal carcinogenesis as measured by prolonged survival, lower adenoma incidence, smaller polyps, and less chronic inflammation. STAT2 deficiency also inhibited 7,12-dimethylbenz(a)anthracene/12-O-tetradecanoylphorbol-13-acetate-induced skin carcinogenesis as indicated by reduced papilloma multiplicity. A potential mechanism by which STAT2 promotes carcinogenesis is through activation of proinflammatory mediators. Deletion of STAT2 decreased azoxymethane/dextran sodium sulfate-induced expression and release of proinflammatory mediators, such as interleukin-6 and CCL2, and decreased interleukin-6 release from skin carcinoma cells, which then decreased STAT3 activation. Our findings identify STAT2 as a novel contributor to colorectal and skin carcinogenesis that may act to increase the gene expression and secretion of proinflammatory mediators, which in turn activate the oncogenic STAT3 signaling pathway., ((c) 2010 AACR.)