EDIL3 as an Angiogenic Target of Immune Exclusion Following Checkpoint Blockade.

Immune checkpoint blockade (ICB) has become the standard of care for several solid tumors. Multiple combinatorial approaches have been studied to improve therapeutic efficacy. The combination of antiangiogenic agents and ICB has demonstrated efficacy in several cancers. To improve the mechanistic understanding of synergies with these treatment modalities, we performed screens of sera from long-term responding patients treated with ipilimumab and bevacizumab. We discovered a high-titer antibody response against EGF-like repeats and discoidin I-like domains protein 3 (EDIL3) that correlated with favorable clinical outcomes. EDIL3 is an extracellular protein, previously identified as a marker of poor prognosis in various malignancies. Our Tumor Immune Dysfunction and Exclusion analysis predicted that EDIL3 was associated with immune exclusion signatures for cytotoxic immune cell infiltration and nonresponse to ICB. Cancer-associated fibroblasts (CAF) were predicted as the source of EDIL3 in immune exclusion-related cells. Furthermore, The Cancer Genome Atlas Skin Cutaneous Melanoma (TCGA-SKCM) and CheckMate 064 data analyses correlated high levels of EDIL3 with increased pan-fibroblast TGFß response, enrichment of angiogenic signatures, and induction of epithelial-to-mesenchymal transition. Our in vitro studies validated EDIL3 overexpression and TGFß regulation in patient-derived CAFs. In pretreatment serum samples from patients, circulating levels of EDIL3 were associated with circulating levels of VEGF, and like VEGF, EDIL3 increased the angiogenic abilities of patient-derived tumor endothelial cells (TEC). Mechanistically, three-dimensional microfluidic cultures and two-dimensional transmigration assays with TEC endorsed EDIL3-mediated disruption of the lymphocyte function-associated antigen-1 (LFA-1)-ICAM-1 interaction as a possible means of T-cell exclusion. We propose EDIL3 as a potential target for improving the transendothelial migration of immune cells and efficacy of ICB therapy.

Capacitively coupled nonthermal plasma synthesis of aluminum nanocrystals for enhanced yield and size control.

Uniform-size, non-native oxide-passivated metallic aluminum nanoparticles (Al NPs) have desirable properties for fuel applications, battery components, plasmonics, and hydrogen catalysis. Nonthermal plasma-assisted synthesis of Al NPs was previously achieved with an inductively coupled plasma (ICP) reactor, but the low production rate and limited tunability of particle size were key barriers to the applications of this material. This work focuses on the application of capacitively coupled plasma (CCP) to achieve improved control over Al NP size and a ten-fold increase in yield. In contrast with many other materials, where NP size is controlled via the gas residence time in the reactor, the Al NP size appeared to depend on the power input to the CCP system. The results indicate that the CCP reactor assembly, with a hydrogen-rich argon/hydrogen plasma, was able to produce Al NPs with diameters that were tunable between 8 and 21 nm at a rate up ∼ 100 mg h-1. X-ray diffraction indicates that a hydrogen-rich environment results in crystalline metal Al particles. The improved synthesis control of the CCP system compared to the ICP system is interpreted in terms of the CCP's lower plasma density, as determined by double Langmuir probe measurements, leading to reduced NP heating in the CCP that is more amenable to NP nucleation and growth.

Impact of federalization for health financing and workforce in Nepal.

The adoption of its 2015 constitution has converted Nepal to a federal government while simultaneously resulted in significant reforms of the health system in Nepal in terms of both structure and commitment. In this commentary, we review evidence ranging from health financing to health workforce development to show that the impact of federalization on Nepal's health system and its efforts to achieve equitable and affordable universal health care have been mixed. On the one hand, careful efforts of the federal government to support subnational governments during the transition appears to have avoided serious disruption, subnational governments have successfully taken on the financial burden of the health system, and increase subnational control has allowed more flexible adaptation to changing needs than might have otherwise been possible. On the other hand, financing resource and ability disparities across subnational governments contributes to significant disparities in workforce development, and subnational authorities appear to have underestimated significant health issues (e.g. NCDs) in their budgets. We then provide three recommendations to improve the success of the Nepalese system: (1) to assess whether the services covered by health financing and insurance schemes like the National Health Insurance Program adequately address the needs of the rising burden of NCDs in Nepal, (2) to set clear minimum requirements on key metrics for subnational health systems, and (3) to extend grant programs to address resource disparities.

Attenuation of NAD[P]H:quinone oxidoreductase 1 aggravates prostate cancer and tumor cell plasticity through enhanced TGFß signaling.

NAD[P]H:quinone oxidoreductase 1 (NQO1) regulates cell fate decisions in response to stress. Oxidative stress supports cancer maintenance and progression. Previously we showed that knockdown of NQO1 (NQO1low) prostate cancer cells upregulate pro-inflammatory cytokines and survival under hormone-deprived conditions. Here, we tested the ability of NQO1low cells to form tumors. We found NQO1low cells form aggressive tumors compared with NQO1high cells. Biopsy specimens and circulating tumor cells showed biochemical recurrent prostate cancer was associated with low NQO1. NQO1 silencing was sufficient to induce SMAD-mediated TGFß signaling and mesenchymal markers. TGFß treatment decreased NQO1 levels and induced molecular changes similar to NQO1 knockdown cells. Functionally, NQO1 depletion increased migration and sensitivity to oxidative stress. Collectively, this work reveals a possible new gatekeeper role for NQO1 in counteracting cellular plasticity in prostate cancer cells. Further, combining NQO1 with TGFß signaling molecules may serve as a better signature to predict biochemical recurrence.

Potential for enriching next-generation health-promoting gut bacteria through prebiotics and other dietary components.

The human intestinal commensal microbiota and associated metabolic products have long been regarded as contributors to host health. As the identity and activities of the various members of this community have become clearer, newly identified health-associated bacteria, such as Faecalibacterium prausnitzii, Akkermansia muciniphila, Ruminococcus bromii and Roseburia species, have emerged. Notably, the abundance of many of these bacteria is inversely correlated to several disease states. While technological and regulatory hurdles may limit the use of strains from these taxa as probiotics, it should be possible to utilize prebiotics and other dietary components to selectively enhance their growth in situ. Dietary components of potential relevance include well-established prebiotics, such as galacto-oligosaccharides, fructo-oligosaccharides and inulin, while other putative prebiotics, such as other oligosaccharides, polyphenols, resistant starch, algae and seaweed as well as host gut metabolites such as lactate and acetate, may also be applied with the aim of selectively and/or differentially affecting the beneficial bacterial community within the gastrointestinal environment. The present review provides an overview of the dietary components that could be applied in this manner.

Genoprotective Effects of Essential Oil Compounds Against Oxidative and Methylated DNA Damage in Human Colon Cancer Cells.

Essential oils (EO) are widely used in foods as flavoring and preservative agents. Many of the biological activities of EO have been attributed to major essential oil compounds (EOC) but their direct interaction with colonic epithelial cells and their genotoxic and genoprotective effects are not well established. In this study, the cytotoxicity and genotoxicity of EOC including nerolidol, thymol, geraniol, methylisoeugenol, eugenol, linalool, and a commercial blend (Agolin) were determined. Furthermore, the genoprotective effects of EOC against oxidative and methylating damage were assessed using the comet assay in HT-29 colorectal adenocarcinoma cells. The majority of EOC were cytotoxic to HT-29 cells at or above 250 ppm after 24 hr exposure. At noncytotoxic doses, none of the EOC was genotoxic in the comet assay. Genoprotection against oxidative DNA damage was observed for nerolidol (at 62.5 ppm), thymol (at 12.5 ppm), geraniol, and methylisoeugenol (both at 125 ppm), as well as linalool and Agolin (both at 250 ppm). Thymol was the most protective compound against oxidative DNA damage and geraniol (at 125 ppm) also protected cells against methylating DNA damage. This study highlights the potential of EOC such as thymol to protect the colonic epithelium against oxidative DNA damage and geraniol against methylating DNA damage. Further in vivo studies are needed to confirm these findings for safety and efficacy to exploit their potential pharmaceutical or nutraceutical uses for colonic health.

Herbal remedies for urinary stones used in India and China: A review.

ETHANOPHARMACOLOGICAL RELEVANCE: The process of formation or appearance of a urinary stone anywhere in the renal tract is known as urolithiasis. It is a longstanding health problem, known to exist since early age of civilization. Records about symptoms, signs and treatment strategies of urinary stones diseases are found in the several ancient texts of traditional medicines such as Ayurveda, Traditional Chinese Medicine (TCM), Siddha and Unani. In Ayurveda, urolithiasis has been considered as one of the eight most troublesome diseases. Ayurvedic management and cure of urinary stone involves herbal formulas, alkaline liquids and surgical procedures. Whereas, TCM recommends polyherbal drugs, acupuncture and mexibustion for treatment of the urinary stones. Among these therapies, herbal remedies are in practice till today for the treatment and cure urinary stone diseases. MATERIALS AND METHODS: A comprehensive review of the scientific literature about pathophysiology of urinary stones and antiurolithiatic plants was undertaken using the following bibliographic databases: MEDLINE/PubMed, Scopus, Web of Knowledge and Google Scholar. The search was conducted from publications from all years until Dec., 2015 by combination of the search terms and Boolean operators; 'urinary stone' OR 'kidney stone' AND 'plant' OR 'medicine' OR 'antiurolithiatic plants'. Outputs were restricted to those completed studies only published in English. In this review, literatures about plants which are used as diuretic and/or in treatment urinary tract infections have not also been considered. The Plant List and Royal Botanical Garden, Kew databases were used to authenticate botanical names of plants. Books and monographs published in English were used to collect information about historical records of antiurolithiatic plants. RESULTS: Recent pharmacological interventions accredited ancient antiurolithiatic claims to several plants and their formulations. The majority of antiurolithiatic plants were found to either dissolve the stones or inhibit the process of urinary stone formation. Plants such as Phyllanthus niruri L. and Elymus repens (L.) Gould, as well as herbal products including 'Wu-Ling-San' formula, 'Cystone' and 'Herbmed' have been proved their utility as promising antiurolithiatic medicines in the different phases of clinical trials. In addition, some of the isolated phytochemicals such as berberine, lupeol, khelin, visnagin, 7-hydroxy-2',4',5'-trimethoxyisoflavone and 7-hydroxy-4'-methoxyisoflavone were reported to have potent antiurolithiatic activity. CONCLUSION: In ancient medicinal texts, antiurolithiatic potential has been ascribed to several plants and their formulations. Present scientific studies provide scientific evidences for few of these claims however, they are insufficient to establish many of these plants and herbal formulations as therapeutic remedies for the treatment and management of urinary stones. Conversely, findings of pre-clinical and clinical studies about some plants and herbal formulations are promising, which underlines the utility of herbal remedies as alternative medicines for the treatment and management of urinary stones in the future.

Chronic inflammatory mediators enhance prostate cancer development and progression.

Chronic inflammation is postulated to influence prostate cancer progression. Preclinical studies have claimed that inflammatory mediators are involved in prostate cancer development and therefore suggested these as attractive targets for intervention. However, among the many pro-inflammatory mediators, there is no consensus regarding the identity of the primary one(s). In clinical studies, chronic inflammation has been found in prostate tumor specimens, and tissues resected for treatment of benign prostatic hyperplasia (BPH). Although collective evidence from molecular, experimental and clinical data suggests that inflammation can contribute or promote prostate carcinogenesis, an etiologic link has not yet been established. Moreover, the role of chronic inflammation in the onset of castration resistant and metastatic disease is unclear. Therefore it is important to open a dialog regarding recent findings on how chronic inflammatory mediators contribute to prostate cancer progression, and their usefulness to prevent disease progression. In this commentary, we assess the current literature with respect to chronic inflammation as a potential initiator and promoter of prostate carcinogenesis and discuss the prospects for its potential clinical applications.

NQO1 suppresses NF-κB-p300 interaction to regulate inflammatory mediators associated with prostate tumorigenesis.

NADPH reductase NAD(P)H: quinone oxidoreductase 1 (NQO1) is needed to maintain a cellular pool of antioxidants, and this enzyme may contribute to tumorigenesis on the basis of studies in NQO1-deficient mice. In this work, we sought deeper insights into how NQO1 contributes to prostate carcinogenesis, a setting in which oxidative stress and inflammation are established contributors to disease development and progression. In the TRAMP mouse model of prostate cancer, NQO1 was highly expressed in tumor cells. NQO1 silencing in prostate cancer cells increased levels of nuclear IKKα and NF-κB while decreasing the levels of p53, leading to interactions between NF-κB and p300 that reinforce survival signaling. Gene expression analysis revealed upregulation of a set of immune-associated transcripts associated with inflammation and tumorigenesis in cells in which NQO1 was attenuated, with IL8 confirmed functionally in cell culture as one key NQO1-supported cytokine. Notably, NQO1-silenced prostate cancer cells were more resistant to androgen deprivation. Furthermore, NQO1 inhibition increased migration, including under conditions of androgen deprivation. These results reveal a molecular link between NQO1 expression and proinflammatory cytokine signaling in prostate cancer. Furthermore, our results suggest that altering redox homeostasis through NQO1 inhibition might promote androgen-independent cell survival via opposing effects on NF-κB and p53 function.

Biotransformation of caffeoyl quinic acids from green coffee extracts by Lactobacillus johnsonii NCC 533.

The potential of Lactobacillus johnsonii NCC 533 to metabolize chlorogenic acids from green coffee extract was investigated. Two enzymes, an esterase and a hydroxycinnamate decarboxylase (HCD), were involved in this biotransformation. The complete hydrolysis of 5-caffeoylquinic acid (5-CQA) into caffeic acid (CA) by L. johnsonii esterase occurred during the first 16 h of reaction time. No dihydrocaffeic acid was identified in the reaction mixture. The decarboxylation of CA into 4-vinylcatechol (4-VC) started only when the maximum concentration of CA was reached (10 µmol/ml). CA was completely transformed into 4-VC after 48 h of incubation. No 4-vinylphenol or other derivatives could be identified in the reaction media. In this study we demonstrate the capability of L. johnsonii to transform chlorogenic acids from green coffee extract into 4-VC in two steps one pot reaction. Thus, the enzymatic potential of certain lactobacilli might be explored to generate flavor compounds from plant polyphenols.

Dietary resveratrol prevents development of high-grade prostatic intraepithelial neoplastic lesions: involvement of SIRT1/S6K axis.

SIRT1 (mammalian ortholog of the yeast silent information regulator 2) is a NAD-dependent histone deacetylase belonging to the multigene family of sirtuins. Anecdotal and epidemiologic observations provide evidence for beneficial effects of the calorie restriction mimetic resveratrol (RES), a SIRT1 activator in preventing cardiovascular diseases and cancer. Although SIRT1 possesses both tumorigenic and antitumorigenic potential, the molecular mechanisms underlying SIRT1-mediated tumor progression or inhibition are poorly understood. In this study, we investigated the role of SIRT1 in multiple human prostate cancer cell lines and prostate-specific PTEN knockout mouse model using resveratrol. Androgen-independent prostate cancer cell lines (C42B, PC3, and DU145) express higher levels of SIRT1 than androgen-responsive (LNCaP) and nontumorigenic prostate cells (RWPE-1). Resveratrol enhanced this expression without any significant effect on SIRT1 enzymatic activity. Inhibition of SIRT1 expression using shRNA enhanced cell proliferation and inhibited autophagy by repressing phosphorylation of S6K and 4E-BP1. These biologic correlates were reversed in the presence of resveratrol. Analysis of prostates from dietary intervention with resveratrol showed a significant reduction in prostate weight and reduction in the incidence of high-grade prostatic intraepithelial neoplastic (HGPIN) lesions by approximately 54% with no significant change in body weight. Consistent with the in vitro findings, resveratrol intervention in the PTEN knockout mouse model was associated with reduction in the prostatic levels of mTOR complex 1 (mTORC1) activity and increased expression of SIRT1. These data suggest that SIRT1/S6K-mediated inhibition of autophagy drives prostate tumorigenesis. Therefore, modulation of SIRT1/S6K signaling represents an effective strategy for prostate cancer prevention.

Anti-tumor activity of the novel hexahydrocannabinol analog LYR-8 in Human colorectal tumor xenograft is mediated through the inhibition of Akt and hypoxia-inducible factor-1α activation.

Cannabinoid compounds have been shown to exert anti-tumor effects by affecting angiogenesis, invasion, and metastasis. In the present study, we examined the action mechanism by which LYR-8, a novel hexahydrocannabinol analog, exerts anti-angiogenic and anti-tumor activity in human cancer xenografts. In the xenografted tumor tissues, LYR-8 significantly reduced the expression of hypoxia-inducible factor-1 alpha (HIF-1α), a transcription factor responsible for induction of angiogenesis-promoting factors, and its target genes, vascular endothelial growth factor (VEGF) and cyclooxygenase-2 (COX-2). In HT-29 human colon cancer cells treated with a hypoxia-inducing agent (CoCl(2)), LYR-8 dose-dependently suppressed the induction of HIF-1α and subsequently its targets, VEGF and COX-2. In addition, highly elevated prostaglandin E(2) (PGE(2)) concentrations in CoCl(2)-treated HT-29 cells were also significantly suppressed by LYR-8. However, LYR-8 alone in the absence of CoCl(2) did not alter the basal expression of VEGF and COX-2, or PGE(2) production. Furthermore, LYR-8 effectively suppressed Akt signaling, which corresponded to the suppression of CoCl(2)-induced HIF-1α accumulation. Taken together, LYR-8 exerts anti-tumor effects through the inhibition of Akt and HIF-1α activation, and subsequently suppressing factors regulating tumor microenvironment, such as VEGF and COX-2. These results indicate a novel function of cannabinoid-like compound LYR-8 as an anti-tumor agent with a HIF-1α inhibitory activity.

Antioxidants for prostate cancer chemoprevention: challenges and opportunities.

Extensive research has led to the firm conclusion that antioxidants protect cells from damage caused by oxidative stress and its associated pathological conditions including inflammation. It has also been established that inflammation is a precursor in neoplastic transformation of the prostate. Although, a vast body of experimental and clinical evidence shows efficacy of antioxidants as preventive strategies for prostate cancer, there is a lack of consistent agreement in outcomes especially from recent large-scale randomized clinical trials. Despite these concerns, our understanding of the preventive mechanisms as well as clinical efficacy and safety data indicate that novel antioxidant therapeutics still hold great promise for prostate cancer chemoprevention. We propose that for effective use of antioxidants for prostate cancer prevention, further high impact translational research is needed with special attention on selecting those patients who will benefit from such intervention. Therefore, it is important to validate predictive biomarkers from successful trials and combine this with knowledge of preclinical characterization of antioxidants (and combinations) that will eventually facilitate the development of 'personalized prostate cancer chemoprevention'. In this review, we briefly describe some common and emerging antioxidants that have shown benefits in preclinical and clinical settings. Above all, we focus on summarizing the progress we made thus far in prostate cancer chemoprevention using antioxidants, the heightened interest and challenges in the future.

The anti-angiogenic and anti-tumor activity of synthetic phenylpropenone derivatives is mediated through the inhibition of receptor tyrosine kinases.

Abnormal angiogenesis plays a critical role in the pathogenesis of various diseases such as cancer and chronic inflammation. A variety of pro-angiogenic factors, including vascular endothelial growth factor (VEGF), exert their action through endothelial receptor tyrosine kinases (RTKs). The synthetic phenylpropenone derivatives, used in this study were the following: 1,3-diphenyl-propenone (DPhP), 3-phenyl-1-thiophen-2-yl-propenone (PhT2P), 3-phenyl-1-thiophen-3-yl-propenone (PhT3P) and 1-furan-2-yl-3-phenyl-propenone (FPhP). These derivatives were screened for their inhibitory effect on VEGF-induced angiogenesis in vitro using HUVECs and in vivo using chick chorioallantoic membrane (CAM). The order of anti-angiogenic activity was DPhP>FPhP>PhT3P>PhT2P. The most effective compound DPhP, also known as chalcone, showed weak VEGF receptor tyrosine kinase activity compared with the specific inhibitor, SU4312 (3-[[4-(dimethylamino)phenyl]methylene]-1,3-dihydro-2H-indol-2-one). However, DPhP also inhibited several other receptor tyrosine kinases including Tie-2, epithermal growth factor (EGF) receptor, EphB2, fibroblast growth factor (FGF) receptor 3 and insulin-like growth factor-1 (IGF-1) receptor, as revealed by a receptor tyrosine kinase array assay. In addition, the down-stream signaling, including ERK phosphorylation and NF-κB activation, after receptor activation was significantly inhibited by DPhP. Furthermore, in the HT29 human colon cancer cell-inoculated CAM assay, the tumor growth and tumor-induced angiogenesis was significantly inhibited by DPhP (10µg/ml). These results suggest that the simple flavonoid, DPhP (chalcone), has valuable potential as an antiangiogenic and anti-cancer agent, and its action is mediated through the inhibition of multi-target RTKs including VEGF receptor 2.

Role of nonsteroidal anti-inflammatory drug-activated gene-1 in docetaxel-induced cell death of human colorectal cancer cells with different p53 status.

Nonsteroidal anti-inflammatory drug-activated gene-1 (NAG-1) expression is upregulated not only by NSAIDs such as sulindac sulfide, but also by several antitumorigenic dietary compounds, suggesting that NAG-1 is a specific target for the development of effective anticancer agents. Despite being a downstream target of p53, NAG-1 induction is both p53-dependent and p53-independent. It is not clear whether NAG-1 induction is the responsible factor in cancer cell apoptosis with mutated p53. In this study, we report that NAG-1 induction alone cannot determine apoptotic cell fate in colon cancer cells. Although docetaxel induced an increase in NAG-1 and apoptosis in both HCT-116 (wild-type p53) and HT-29 (mutant p53) colon cancer cells, NAG-1 knockdown with siRNA prevented docetaxel-induced cell death in only HCT-116 cells. Docetaxel decreased Bcl-2 in HCT-116 cells, which have functionally active p53, according to luciferase reporter gene analyses, and docetaxel-induced cell death and changes in Bcl-2 and NAG-1 expression were blocked by PFT-α, a p53 inhibitor. In HT-29 cells with functionally inactive p53, the docetaxel-induced Bcl-xL decrease, NAG-1 increase, and cell death were not blocked by PFT-α. On the other hand, sulindac sulfide at concentrations that significantly induced NAG-1 did not decrease cell viability comparable to docetaxel, and it did not affect the level of p53, Bax, Bcl-2, and Bcl-xL in either cell line. The present study demonstrates that p53-dependent NAG-1 induction is linked to cell death and that NAG-1 induction without accompanying alteration of antiapoptosis protein Bcl-2 family members may not lead to cancer cell death.

NRF2 blockade suppresses colon tumor angiogenesis by inhibiting hypoxia-induced activation of HIF-1α.

Transcription factor NRF2 is an important modifier of cellular responses to oxidative stress. Although its cytoprotective effects are firmly established, recent evidence suggesting important roles in cancer pathobiology has yet to be mechanistically developed. In the current study, we investigated the role of NRF2 in colon tumor angiogenesis. Stable RNAi-mediated knockdown of NRF2 in human colon cancer cells suppressed tumor growth in mouse xenograft settings with a concomitant reduction in blood vessel formation and VEGF expression. Similar antiangiogenic effects of NRF2 knockdown were documented in chick chorioallantoic membrane assays and endothelial tube formation assays. Notably, NRF2-inhibited cancer cells failed to accumulate HIF-1α protein under hypoxic conditions, limiting expression of VEGF and other HIF-1α target genes. In these cells, HIF-1α was hydroxylated but pharmacological inhibition of PHD domain-containing prolyl hydroxylases was sufficient to restore hypoxia-induced accumulation of HIF-1α. Mechanistic investigations demonstrated that reduced mitochondrial O(2) consumption in NRF2-inhibited cells was probably responsible for HIF-1α degradation during hypoxia; cellular O(2) consumption and ATP production were lower in NRF2 knockdown cells than in control cells. Our findings offer novel insights into how cellular responses to O(2) and oxidative stress are integrated in cancer cells, and they highlight NRF2 as a candidate molecular target to control tumor angiogenesis by imposing a blockade to HIF-1α signaling.

Novel hexahydrocannabinol analogs as potential anti-cancer agents inhibit cell proliferation and tumor angiogenesis.

Both natural and synthetic cannabinoids have been shown to suppress the growth of tumor cells in culture and in animal models by affecting key signaling pathways including angiogenesis, a pivotal step in tumor growth, invasion, and metastasis. In our search for cannabinoid-like anticancer agents devoid of psychoactive side effects, we synthesized and evaluated the anti-angiogenic effects of a novel series of hexahydrocannabinol analogs. Among these, two analogs LYR-7 [(9S)-3,6,6,9-tetramethyl-6a,7,8,9,10,10a-hexahydro-6H-benzo[c]chromen-1-ol] and LYR-8 [(1-((9S)-1-hydroxy-6,6,9-trimethyl-6a,7,8,9,10,10a-hexahydro-6H-benzo[c]chromen-2-yl)ethanone)] were selected based on their anti-angiogenic activity and lack of binding affinity for cannabinoid receptors. Both LYR-7 and LYR-8 inhibited VEGF-induced proliferation, migration, and capillary-like tube formation of HUVECs in a concentration-dependent manner. The inhibitory effect of the compounds on cell proliferation was more selective in endothelial cells than in breast cancer cells (MCF-7 and tamoxifen-resistant MCF-7). We also noted effective inhibition of VEGF-induced new blood vessel formation by the compounds in the in vivo chick chorioallantoic membrane (CAM) assay. Furthermore, both LYR analogs potently inhibited VEGF production and NF-κB transcriptional activity in cancer cells. Additionally, LYR-7 or LYR-8 strongly inhibited breast cancer cell-induced angiogenesis and tumor growth. Together, these results suggest that novel synthetic hexahydrocannabinol analogs, LYR-7 and LYR-8, inhibit tumor growth by targeting VEGF-mediated angiogenesis signaling in endothelial cells and suppressing VEGF production and cancer cell growth.

HMC05, Herbal Formula, Inhibits TNF-α-Induced Inflammatory Response in Human Umbilical Vein Endothelial Cells.

Vascular inflammation has been implicated in the progression of cardiovascular diseases such as atherosclerosis. In the present study, we found that HMC05, an extract from eight different herbal mixtures, dose-dependently inhibited tumor necrosis factor-α (TNF-α)-induced adhesion of monocytes to endothelial cells. Such inhibitory effect of HMC05 correlated with suppressed expression of monocyte chemoattractant protein-1, CC chemokine receptor 2, vascular cell adhesion molecule-1 and intercellular cell adhesion molecule-1. In addition, HMC05 significantly inhibited production of reactive oxygen species (ROS) and nuclear factor (NF)-κB activation by TNF-α. Those inhibitory effects of HMC05 (1-10 µg mL(-1)) on the TNF-α-induced inflammatory event was similar to those of berberine (1-10 µM), which is a major component of HMC05 and one of herbal compounds known to have vasorelaxing and lipid-lowering activities. However, berberine significantly reduced the viability of HUVECs in a time- and concentration-dependent manner. In contrast, HMC05 (1-10 µg ml(-1)) did not affect the cell viability for up to 48 h treatment. In conclusion, we propose that HMC05 may be a safe and potent herbal formula against vascular inflammation, and its action may be attributable to the inhibition of ROS- and NF-κB-dependent expression of adhesion molecules and chemokines.

Induction of p53-independent apoptosis by a novel synthetic hexahydrocannabinol analog is mediated via Sp1-dependent NSAID-activated gene-1 in colon cancer cells.

Nonsteroidal anti-inflammatory drug (NSAID)-activated gene-1 (NAG-1) has received greater attention as a novel molecular target for anti-cancer therapeutics in recent years. We identified a novel synthetic hexahydrocannabinol analog, LYR-8 [(1-((9S)-1-hydroxy-6,6,9-trimethyl-6a,7,8,9,10,10a-hexahydro-6H-benzo[c]chromen-2-yl)ethanone)], as a potent NAG-1 and apoptosis inducer in a panel of human cancer cells. LYR-8 did not possess any affinity for cannabinoid receptor CB(1) or CB(2), which eliminates the concern about potential psychoactive side effects. LYR-8 dramatically induced NAG-1 expression and apoptosis in HCT116 (wild-type p53) and HT29 (mutant p53) colon cancer cells. The NAG-1 expression by LYR-8 was not blocked by pifithrin-alpha, a specific p53 inhibitor, which was different from doxorubicin that induced p53-dependent NAG-1 transcriptional activity. The induction of NAG-1 promoter activity by LYR-8 was strongly correlated with increased Sp1 activation as noted in various luc-promoter activities. Furthermore, pretreatment with the specific Sp1 inhibitor mithramycin A completely reversed the LYR-8-induced NAG-1 expression in both HCT116 and HT29 cells. Knockdown of NAG-1 using siRNA significantly reversed LYR-8-induced cell death in both wild-type and mutant p53-expressing colon cancer cells. Furthermore, sensitization with NAG-1 inducer sulindac sulfide synergized LYR-8-induced cell death in both colon cancer cells. These results suggest that induction of NAG-1 via Sp1 activation is a promising therapeutic approach in cancer treatment, and that a novel compound like LYR-8 could be a potent chemotherapeutic agent for colon cancers including p53-mutated cancer.

Grifola frondosa water extract alleviates intestinal inflammation by suppressing TNF-alpha production and its signaling.

TNF-alpha is a major cytokine involved in inflammatory bowel disease (IBD). In this study, water extract of Grifola frondosa (GFW) was evaluated for its protective effects against colon inflammation through the modulation of TNF-alpha action. In coculture of HT-29 human colon cancer cells with U937 human monocytic cells, TNF-alpha-induced monocyte adhesion to HT-29 cells was significantly suppressed by GFW (10, 50, 100 micg/ml). The reduced adhesion by GFW correlated with the suppressed expression of MCP-1 and IL-8, the major IBD-associated chemokines. In addition, treatment with GFW significantly suppressed TNF-alpha-induced reactive oxygen species production and NF-kappaB transcriptional activity in HT-29 cells. In differentiated U937 monocytic cells, LPS-induced TNF-alpha production, which is known to be mediated through NF-kappaB activation, was significantly suppressed by GFW. In an in vivo rat model of IBD, oral administration of GFW for 5 days (1 g/kg per day) significantly inhibited the trinitrobenzene sulfonic acid (TNBS)-induced weight loss, colon ulceration, myeloperoxidase activity, and TNF-alpha expression in the colon tissue. Moreover, the effect of GFW was similar to that of intra-peritoneal injection of 5-aminosalicylic acid (5-ASA), an active metabolite of sulfasalazine, commonly used drug for the treatment of IBD. The results suggest that GFW ameliorates colon inflammation by suppressing production of TNF-alpha as well as its signaling through NF-kappaB leading to the expression of inflammatory chemokines, MCP-1 and IL-8. Taken together, the results strongly suggest GFW is a valuable medicinal food for IBD treatment, and thus may be used as an alternative medicine for IBD.