Role of natural products in breast cancer related symptomology: Targeting chronic inflammation.

Breast cancer is the most common cancer in women worldwide. There have been many advancements in the treatment of breast cancer leading to an increased population of patients living with this disease. Accumulating evidence suggests that cancer diagnosis and aftermath experienced stress could not only affect the quality of life of cancer patients, but it could also influence their disease outcome. The magnitude of stress experienced by breast cancer patients is often compared to the post-traumatic stress disorder-like symptoms suggested to be mediated by the chronic inflammation including NF-κB, AKt, p53 and other inflammatory pathways. Here, we describe the symptomology of PTSD-like symptoms in breast cancer patients and argue that they may in fact be caused by or maintained through aspects of chronic inflammation mediated by the pro-inflammatory markers. Evidence exists that natural products that might attenuate or lessen the effects of chronic inflammation abound in the diet. We summarize some possible agents that might abate the genesis of symptoms experienced by breast cancer patients while mitigating the effect of inflammation.

The Highly Pure Neem Leaf Extract, SCNE, Inhibits Tumorigenesis in Oral Squamous Cell Carcinoma via Disruption of Pro-tumor Inflammatory Cytokines and Cell Signaling.

Oral squamous cell carcinoma (OSCC) is a deadly disease that comprises 60% of all head and neck squamous cell cancers. The leaves of the Neem tree (Azadirachta indica) have been used in traditional Ayurvedic medicine for centuries to treat numerous oral maladies and are known to have significant anti-inflammatory properties. We hypothesize that a highly pure super critical CO2 Neem leaf extract (SCNE) prevents initiation and progression of OSCC via downregulation of intra-tumor pro-inflammatory pathways, which promote tumorigenesis. Hence, we investigated the anticancer effects of SCNE using in vitro and in vivo platforms. OSCC cell lines (SCC4, Cal27, and HSC3) were treated with SCNE while inflammation, proliferation, and migration were analyzed over time. SCNE treatment significantly inhibited OSCC cell proliferation and migration and reduced MMP activity in vitro, suggesting its potential to inhibit tumor growth and metastasis. The preventive effects of SCNE in ectopic xenograft and 4NQO-1 (4-Nitroquinoline-1-oxide) carcinogen-induced mouse models of OSCC were also evaluated. Indeed, xenografted nude mice showed significant reduction of OSCC tumor volumes. Likewise, SCNE significantly reduced the incidence of tongue dysplasia in the 4NQO-1 OSCC initiation model. In both OSCC animal models, SCNE significantly depressed circulating pro-cancer inflammatory cytokines (host and tumor-secreted) including NFkB, COX2, IL-1, IL-6, TNFα, and IFNγ. In addition, we demonstrate that SCNE downregulates STAT3 and AKT expression and activity in vitro. We also demonstrate that the primary active component, nimbolide (NIM), has significant anticancer activity in established OSCC xenografts. Lastly, we show that SCNE induces an M1 phenotype in tumor associated macrophages (TAMS) in vivo. Taken together, these data strongly support SCNE as means of preventing OSCC via downregulation of pro-cancer inflammatory cascades and NIM as a potential new therapy for existing OSCC.

Disruption of Epigenetic Silencing in Human Colon Cancer Cells Lines Utilizing a Novel Supercritical CO2 Extract of Neem Leaf (Azadirachta indica).

BACKGROUND/AIM: Aerial parts and seeds of the neem tree (Azadirachta indica) have long been used in traditional medicine such as Ayurveda for health-related purposes. Our interest in neem bioactives lies in their potential use as standalone anticancer agents, or as adjuvants to standard therapy. The aim of the present study was to explore a supercritical CO2 extract (SCNE) of neem leaf and a prominent liminoid in neem leaf, nimbolide, for epigenetic activity. MATERIALS AND METHODS: Human colorectal cancer cell lines (HCT116 and HT29) were cultured for 48 h in the presence of neem extract or nimbolide and evaluated for growth inhibition and evidence of suppression of histone deacetylation and DNA methylation. RESULTS: Both SCNE and nimbolide suppressed the proliferation of colon cancer cells by inducing epigenetic modifications. CONCLUSION: Neem leaf contains bioactive constituents which modify epigenetic activity.

A supercritical CO2 extract of neem leaf (A. indica) and its bioactive liminoid, nimbolide, suppresses colon cancer in preclinical models by modulating pro-inflammatory pathways.

Colorectal cancer (CRC) is the third most commonly diagnosed cancer and the second leading cause of cancer death in men and women in the United States. Anti-inflammatory blockade has been proven to be a promising avenue of colorectal cancer prevention. However, NSAIDs while effective in curbing CRC risk are too toxic for long-term use in cancer prevention. The Neem tree (Azadirachta indica) is rich in liminoid terpenoids, collectively known as azadiractoids and has been shown to have anti-inflammatory effects. To explore a role of neem in CRC, human colon cancer cell lines HCT116 and HT29 cells were treated with purified Super Critical Neem Extract (SCNE) or the neem liminoid, nimbolide. SCNE treatment resulted in a dose dependent inhibition of CRC cell proliferation and an increase in apoptosis. Treatment with SCNE and nimbolide decreased the expression of transcriptional factors, STAT3 and NF-κB which plays a major role in gene regulation of multiple cellular processes. Protein expression of COX1, IL-6, and TNF-α were decreased on treatment with SCNE in CRC cells. Western blots and Zymogram assays results revealed anti-invasive effect by decreased expression of MMP2 and MMP9 proteins in CRC cells. Overall, these data confirm a potential anti-cancer effect of SCNE, reducing cell proliferation, inflammation, migration, and invasion in human colon cancer cells. Confirming these indications, we found that treatment of mice bearing HT29 and HCT116 xenografted tumors exhibited striking inhibition of colon tumor growth. Clearly we must explore the effect of neem in preclinical animal models for anti-cancer therapy.

Therapeutic effects of the euglenoid ichthyotoxin, euglenophycin, in colon cancer.

Colorectal cancer (CRC) remains one of the most commonly diagnosed cancers and the 3rd leading cause of cancer-related mortality. The emergence of drug resistance poses a major challenge in CRC care or treatment. This can be addressed by determining cancer mechanisms, discovery of druggable targets, and development of new drugs. In search for novel agents, aquatic microorganisms offer a vastly untapped pharmacological source that can be developed for cancer therapeutics. In this study, we characterized the anti-colorectal cancer potential of euglenophycin, a microalgal toxin from Euglena sanguinea. The toxin (49.1-114.6 µM) demonstrated cytotoxic, anti-proliferative, anti-clonogenic, and anti-migration effects against HCT116, HT29, and SW620 CRC cells. We identified G1 cell cycle arrest and cell type - dependent modulation of autophagy as mechanisms of growth inhibition. We validated euglenophycin's anti-tumorigenic activity in vivo using CRL:Nu(NCr)Foxn1nu athymic nude mouse CRC xenograft models. Intraperitoneal toxin administration (100 mg/kg; 5 days) decreased HCT116 and HT29 xenograft tumor volumes (n=10 each). Tumor inhibition was associated with reduced expression of autophagy negative regulator mechanistic target of rapamycin (mTOR) and decreased trend of serum pro-inflammatory cytokines. Together, these results provide compelling evidence that euglenophycin can be a promising anti-colorectal cancer agent targeting multiple cancer-promoting processes. Furthermore, this study supports expanding natural products drug discovery to freshwater niches as prospective sources of anti-cancer compounds.

Reduction in promotor methylation utilizing EGCG (epigallocatechin-3-gallate) restores RXRα expression in human colon cancer cells.

Silencing of regulatory genes through hypermethylation of CpG islands is an important mechanism in tumorigenesis. In colon cancer, RXRα, an important dimerization partner with other nuclear transcription factors, is silenced through this mechanism. We previously found that colon tumors in ApcMin/+ mice had diminished levels of RXRα protein and expression levels of this gene were restored by treatment with a green tea intervention, due to reduced promoter methylation of RXRα. We hypothesized that CIMP+ cell lines, which epigenetically silence key regulatory genes would also evidence silencing of RXRα and EGCG treatment would restore its expression. We indeed found EGCG to restore RXRα activity levels in the human cell lines, in a dose dependent manner and reduced RXRα promoter methylation. EGCG induced methylation changes in several other colon cancer related genes but did not cause a decrease in global methylation. Numerous epidemiological reports have shown the benefits of green tea consumption in reducing colon cancer risk but to date no studies have shown that the risk reduction may be related to the epigenetic restoration by tea polyphenols. Our results show that EGCG modulates the reversal of gene silencing involved in colon carcinogenesis providing a possible avenue for colon cancer prevention and treatment.

Natural Agents Used in Chemoprevention of Aerodigestive and GI Cancers.

Aerodigestive cancers are on an increasing level in both occurrence and mortality. A major cause in many of these cancers is disruption of the inflammatory pathway, leading to increased cell proliferation, and epigenetic silencing of normal regulatory genes. Here we review the research on several natural products: silibinin, silymarin, quercetin, neem & nimbolide, gingerol, epigallatecatechin-3- gallate, curcumin, genistein and resveratrol conducted on aerodigestive cancers. These types of cancers are primarily those from oral cavity, esophagus/windpipe, stomach, small and large intestine, colon/rectum and bile/pancreas tissues. We report on the utilization in vivo and in vitro systems to research these dose effects on the inflammatory and epigenetic pathway components within the aerodigestive cancer. To follow up on the basic research we will discuss remaining research questions and future directions involving these natural products as putative stand alone or in combination with clinical agents.

Chemoprevention in gastrointestinal physiology and disease. Targeting the progression of cancer with natural products: a focus on gastrointestinal cancer.

The last decade has witnessed remarkable progress in the utilization of natural products for the prevention and treatment of human cancer. Many agents now in the pipeline for clinical trial testing have evolved from our understanding of how human nutritional patterns account for widespread differences in cancer risk. In this review, we have focused on many of these promising agents arguing that they may provide a new strategy for cancer control: natural products once thought to be only preventive in their mode of action now are being explored for efficacy in tandem with cancer therapeutics. Natural products may reduce off-target toxicity of therapeutics while making cancers more amenable to therapy. On the horizon is the use of certain natural products, in their own right, as mitigants of late-stage cancer, a new frontier for small-molecule natural product drug discovery.

Future directions in the prevention of prostate cancer.

The high global incidence of prostate cancer has led to a focus on chemoprevention strategies to reduce the public health impact of the disease. Early studies indicating that selenium and vitamin E might protect against prostate cancer encouraged large-scale studies that produced mixed clinical results. Next-generation prostate cancer prevention trials validated the impact of 5α-reductase inhibitors in hormone-responsive prostate cancer, and these results were confirmed in follow-up studies. Other interventions on the horizon, involving both dietary and pharmacological agents, hold some promise but require further investigation to validate their efficacy. In this Review, we discuss the clinical and preclinical evidence for dietary and pharmacological prevention of prostate cancer and give an overview of future opportunities for chemoprevention.

Green tea polyphenol epigallocatechin 3-gallate, contributes to the degradation of DNMT3A and HDAC3 in HCT 116 human colon cancer cells.

BACKGROUND: Colon cancer is still the second leading cause of cancer deaths in the United States. Epigenetic gene silencing involving DNA methyltransferases (DNMTs) and histone deacetylases (HDACs) plays an important role in the progression of colon cancer. MATERIALS AND METHODS: In the present study we found that the sensitivity of colon cancer cells to methylation plays a role in its response to alternative therapy involving the green tea polyphenol, epigallocatechin 3-gallate. HDAC and DNMT protein expression were reduced when methylation-sensitive HCT 116 human colon cancer cells was treated with EGCG, but was relatively stable in the HT-29 cell line. This decrease in expression may be partially explained by our finding that DNMT3A and HDAC3 are degraded in the methylation-sensitive colon cancer cells in part by inhibiting their association with the E3 ubiquitin ligase, UHRF1. CONCLUSION: These findings provide a rationale for the development of a targeted therapy for methylation-sensitive colon cancer that can include EGCG in combination with other DNMT and HDAC inhibitors.

Vitamin D receptor and retinoid X receptor α status and vitamin D insufficiency in models of murine colitis.

The anti-inflammatory actions of vitamin D have long been recognized and its importance in modulating colon cancer and colitis development is becoming apparent. The vitamin D receptor (VDR) is downregulated in human ulcerative colitis and colitis-associated cancer (CAC); however, its status in murine models of colitis has yet to be explored. Snail and Snail2, zinc-finger transcription factors regulated by inflammatory pathways and able to transcriptionally silence VDR, are upregulated in human Ulcerative Colitis and are associated with localized VDR silencing. To signal, VDR must heterodimerize with retinoid X receptor α (RXRα). If either VDR or RXRα are compromised, vitamin D cannot regulate inflammatory pathways. RXRα is downregulated in human colorectal cancer, yet its expression in human and murine colitis has yet to be investigated. To explore the importance of vitamin D and VDR in murine colitis, we used acute and chronic azoxymethane/dextran sulfate sodium models of murine colitis. VDR was downregulated early in the onset of colitis, whereas RXRα downregulation only occurred as colitis became chronic and developed into CAC. Receptor downregulation was associated with an early increase in the expression of the inflammatory markers, Snail and Snail2. The acute colitis model induced in combination with a vitamin D-deficient diet resulted in increased morbidity, receptor downregulation, inflammatory marker expression, and Snail and Snail2 upregulation. These experiments show the importance of vitamin D and VDR in modulating murine colitis development.

Resveratrol induces premature senescence in lung cancer cells via ROS-mediated DNA damage.

Resveratrol (RV) is a natural component of red wine and grapes that has been shown to be a potential chemopreventive and anticancer agent. However, the molecular mechanisms underlying RV's anticancer and chemopreventive effects are incompletely understood. Here we show that RV treatment inhibits the clonogenic growth of non-small cell lung cancer (NSCLC) cells in a dose-dependent manner. Interestingly, the tumor-suppressive effect of low dose RV was not associated with any significant changes in the expression of cleaved PARP and activated caspase-3, suggesting that low dose RV treatment may suppress tumor cell growth via an apoptosis-independent mechanism. Subsequent studies reveal that low dose RV treatment induces a significant increase in senescence-associated ß-galactosidase (SA-ß-gal) staining and elevated expression of p53 and p21 in NSCLC cells. Furthermore, we show that RV-induced suppression of lung cancer cell growth is associated with a decrease in the expression of EF1A. These results suggest that RV may exert its anticancer and chemopreventive effects through the induction of premature senescence. Mechanistically, RV-induced premature senescence correlates with increased DNA double strand breaks (DSBs) and reactive oxygen species (ROS) production in lung cancer cells. Inhibition of ROS production by N-acetylcysteine (NAC) attenuates RV-induced DNA DSBs and premature senescence. Furthermore, we show that RV treatment markedly induces NAPDH oxidase-5 (Nox5) expression in both A549 and H460 cells, suggesting that RV may increase ROS generation in lung cancer cells through upregulating Nox5 expression. Together, these findings demonstrate that low dose RV treatment inhibits lung cancer cell growth via a previously unappreciated mechanism, namely the induction of premature senescence through ROS-mediated DNA damage.

Racial disparities in advanced-stage colorectal cancer survival.

PURPOSE: African-Americans (AA) have a higher incidence of and lower survival from colorectal cancer (CRC) compared with European Americans (EA). In the present study, statewide, population-based data from South Carolina Central Cancer Registry are used to investigate the relationship between race and age on advanced-stage CRC survival. METHODS: The study population was comprised of 3,865 advanced pathologically documented colon and rectal adenocarcinoma cases diagnosed between 01 January 1996 and 31 December 2006: 2,673 (69 %) EA and 1,192 (31 %) AA. Kaplan-Meier methods were used to generate median survival time and corresponding 95 % confidence intervals (CI) by race, age, and gender. Factors associated with survival were evaluated by fitting Cox proportional hazards regression models to generate hazard ratios (HR) and 95 % CI. RESULTS: We observed a significant interaction between race and age on CRC survival (p = 0.04). Among younger patients (<50 years), AA race was associated with a 1.34 times (95 % CI 1.06-1.71) higher risk of death compared with EA. Among older patients, we observed a modest increase in risk of death among AA men compared with EA [HR 1.16 (95 % CI 1.01-1.32)] but no difference by race between women [HR 0.94 (95 % CI 0.82-1.08)]. Moreover, we observed that the disparity in survival has worsened over the past 15 years. CONCLUSIONS: Future studies that integrate clinical, molecular, and treatment-related data are needed for advancing understanding of the racial disparity in CRC survival, especially for those <50 years old.

Epigenetic mechanisms underlying diet-sourced compounds in the prevention and treatment of gastrointestinal cancer.

The development of colon cancer, the third most diagnosed cancer and third leading cause of cancer deaths in the United States, can be influenced by genetic predispositions and environmental exposures. As 80% of colon cancer cases are sporadic in nature, much interest lies in determining risk factors that may foster its development, as well as identifying compounds that could inhibit colon cancer development or halt progression. A major risk factor for sporadic colon cancer is a high fat, Western diet which has been linked to a cancer-prone, pro-inflammatory state. Cultures which place an emphasis on fresh fruits and vegetables demonstrate lower colon cancer incidences. Diet not only has the potential to encourage colon cancer development, but recent evidence demonstrates that certain dietary natural products can halt colon cancer development and progression via epigenetic regulation. Epigenetic dysregulation may contribute to inflammation-driven diseases, such as cancer, and can lead to the inappropriate silencing of genes necessary to inhibit cancer development. Natural compounds have shown the ability to reverse epigenetic dysregulation in in vitro and in vivo models. As current allopathic medicines aimed at reversing epigenetic silencing are accompanied with the risk of toxicity and side effects, much interest lies in being able to harness the disease preventing properties in natural products. Here, we discuss the epidemiology of colon cancer, describe the need for natural approaches to inhibit disease development and highlight natural products which have been shown to inhibit gastrointestinal cancer initiation and progression in vitro or in vivo through epigenetic modulation.

A limited role of p53 on the ability of a Hexane fraction of American ginseng to suppress mouse colitis.

Ulcerative colitis (UC) is debilitating and carries a high colon cancer risk. Apoptosis of inflammatory cells is a key mechanism regulating UC. We have recently shown that American ginseng (AG), and to a greater extent, a Hexane fraction of AG (HAG) can cause apoptosis and suppress mouse colitis through a p53-mediated mechanism. Here, we tested the hypothesis that HAG suppresses colitis through a p53 mechanism. We found only a limited impact of p53 in the ability of HAG to induce inflammatory cell apoptosis and suppress mouse colitis in vitro and in vivo. Finally, we asked whether HAG could cause cell cycle arrest of HCT116 colon cancer cells in vitro. Interestingly, HAG caused a G1 arrest of such cells independent of p53 status. Findings are significant because HAG suppresses colitis and associated colon cancer, and mutation in p53 is observed in most colitis-driven colon cancers. Therefore, HAG might be very effective in targeting the inflammatory cells and cancer cells since it induces apoptosis of inflammatory cells and cell cycle arrest in both p53-/- and WT p53 colon cancer cells.

Homeostatic imbalance and colon cancer: the dynamic epigenetic interplay of inflammation, environmental toxins, and chemopreventive plant compounds.

The advent of modern medicine has allowed for significant advances within the fields of emergency care, surgery, and infectious disease control. Health threats that were historically responsible for immeasurable tolls on human life are now all but eradicated within certain populations, specifically those that enjoy higher degrees of socio-economic status and access to healthcare. However, modernization and its resulting lifestyle trends have ushered in a new era of chronic illness; one in which an unprecedented number of people are estimated to contract cancer and other inflammatory diseases. Here, we explore the idea that homeostasis has been redefined within just a few generations, and that diseases such as colorectal cancer are the result of fluctuating physiological and molecular imbalances. Phytochemical-deprived, pro-inflammatory diets combined with low-dose exposures to environmental toxins, including bisphenol-A (BPA) and other endocrine disruptors, are now linked to increasing incidences of cancer in westernized societies and developing countries. There is recent evidence that disease determinants are likely set in utero and further perpetuated into adulthood dependent upon the innate and environmentally acquired phenotype unique to each individual. In order to address a disease as multi-factorial, case-specific, and remarkably adaptive as cancer, research must focus on its root causes in order to elucidate the molecular mechanisms by which they can be prevented or counteracted via plant-derived compounds such as epigallocatechin-3-gallate (EGCG) and resveratrol. The significant role of epigenetics in the regulation of these complex processes is emphasized here to form a comprehensive view of the dynamic interactions that influence modern-day carcinogenesis, and how sensibly restoring homeostatic balance may be the key to the cancer riddle.

A hexane fraction of American ginseng suppresses mouse colitis and associated colon cancer: anti-inflammatory and proapoptotic mechanisms.

Ulcerative colitis is a chronic inflammatory condition associated with a high colon cancer risk. We have previously reported that American ginseng extract significantly reduced the inflammatory parameters of chemically induced colitis. The aim of this study was to further delineate the components of American ginseng that suppress colitis and prevent colon cancer. Among five different fractions of American ginseng (butanol, hexane, ethylacetate, dichloromethane, and water), a hexane fraction has particularly potent antioxidant and proapoptotic properties. The effects of this fraction were shown in a mouse macrophage cell line (ANA-1 cells), in a human lymphoblastoid cell line (TK6), and in an ex vivo model (CD4(+)/CD25(-) primary effector T cells). A key in vivo finding was that compared with the whole American ginseng extract, the hexane fraction of American ginseng was more potent in treating colitis in a dextran sodium sulfate (DSS) mouse model, as well as suppressing azoxymethane/DSS-induced colon cancer. Furthermore, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) labeling of inflammatory cells within the colonic mesenteric lymph nodes was elevated in mice consuming DSS + the hexane fraction of American ginseng. Results are consistent with our in vitro data and with the hypothesis that the hexane fraction of American ginseng has anti-inflammatory properties and drives inflammatory cell apoptosis in vivo, providing a mechanism by which this fraction protects from colitis in this DSS mouse model. This study moves us closer to understanding the molecular components of American ginseng that suppress colitis and prevent colon cancer associated with colitis.

Nutraceutical use in late-stage cancer.

Access to a wealth of information on the internet has led many cancer patients to use complementary methods as an adjunct to traditional therapy for cancer, with, and more often, without informing their primary caregiver. Of the common complementary modalities, the use of dietary supplements appears to be highly prevalent in patients in active treatment for cancer, and later in cancer survivors. Emerging research suggests that some plant-based agents may, indeed, impact late-stage cancer, influencing molecular processes corrupted by tumor cells to evade detection, expand clonally, and invade surrounding tissues. The intent of this article is to review some of the current science underpinning the use of nutraceuticals in the latter stages of cancer.

Aberrant crypt foci: the case for inclusion as a biomarker for colon cancer.

Aberrant crypt foci (ACF) are one of the earliest histopathological manifestations of colon cancer. In this review, we critically present the molecular, cellular, histopathological, and chemopreventive evidence that ACF are relevant biomarkers for colon cancer. The laboratory and clinical evidence are highly suggestive that ACF are in the pathway leading to colon cancer, but not all ACF will do so. The possible fate and outcome of ACF in the progression toward colon cancer may be dependent on a number of features that define their predictive power for the prevention or progression of cancer.

Epigenetic modulation of the retinoid X receptor alpha by green tea in the azoxymethane-Apc Min/+ mouse model of intestinal cancer.

We investigated the possible mechanisms of inhibition of colorectal carcinogenesis by green tea (GT) in azoxymethane-treated (AOM) Apc(Min/+) mice. Mice received water or a 0.6% (w/v) solution of GT as the only source of beverage. GT treatment commenced at the 8th week of age and lasted for 8 wk. The treatment caused a statistically significant reduction in the number of newly formed tumors (28%, P < 0.05). Immunohistochemical analysis showed that GT decreased the levels of beta-catenin and its downstream target cyclin D1. To probe a mechanism, we further investigated the expression of retinoic X receptor alpha (RXR alpha) in AOM/Apc(Min/+) tumors. Our results show that RXR alpha is selectively downregulated in AOM/Apc(Min/+) mouse intestinal tumors. In contrast, other retinoic receptors including retinoic acid receptor alpha (RAR alpha), RAR beta, RXR beta, and RXR gamma were all expressed in Apc(Min/+) adenomas. Furthermore, our results show that RXR alpha downregulation is an early event in colorectal carcinogenesis and is independent of beta-catenin expression. GT significantly increased the protein levels of RXR alpha. In addition, RT-PCR analysis showed that GT induced a similar increase in the levels of RXR alpha mRNA. Genomic bisulfite treatment of colonic DNA followed by pyrosequencing of 24 CpG sites in the promoter region of RXR alpha gene showed a significant decrease in CpG methylation with GT treatment. The results suggest that a low concentration of GT is sufficient to desilence RXR alpha and inhibit intestinal tumorigenesis in the Apc(Min/+) mouse.