The significance of abdominal para-aortic lymph node metastasis in patients with lower thoracic esophageal cancer.

Para-aortic lymph node (PALN) recurrence is often seen in patients with lower thoracic esophageal cancer treated by esophagectomy with extended lymph node dissection. However, the clinicopathological characteristics of patients with PALN metastasis and the significance of PALN dissection are unknown. A total of 283 patients with lower thoracic esophageal cancer underwent esophagectomy with lymphadenectomy at our hospital between April 1984 and March 2007. Among these 283 patients, 60 patients were enrolled in this retrospective study according to following criteria: (i) clinical T2 to T4 tumor, (ii) no clinical PALN metastasis, and (iii) received PALN dissection. PALN dissection was indicated by a tumor depth of at least T2 and no severe complications. The clinicopathological data, recurrence pattern, and overall survival were compared between patients with PALN and without PALN metastasis. The mean length of surgery was 587 min and the mean blood loss was 1383 mL. The morbidity was 33.3% and mortality was 5% in this series. Sixteen patients (26.7%) had PALN metastasis; these showed significantly more lymph node metastases (15.8 ± 13.2 vs. 3.0 ± 3.2, P < 0.0001) and significantly worse survival rates (53.3% vs. 79.9% at 1 year, 6.7% vs. 62.0% at 3 years, P < 0.0001) than patients without PALN metastasis. The incidence of lymph node recurrence (P < 0.0001) and hematogenous recurrence (P= 0.0487) was also higher in patients with PALN metastasis than in patients without PALN metastasis. Among the 16 patients with PALN metastasis, a univariate analysis revealed total number of metastatic nodes < 8 (P= 0.0325) to be a significant prognostic factor. A multivariate logistic regression analysis of the regional lymph nodes identified the invasion of the lower mediastinal nodes (hazard ratio = 6.120) and retroperitoneal nodes (hazard ratio = 15.167) to be significantly correlated with PALN metastasis. PALN metastasis is suggested to be related to the systemic spread of lymphatic metastasis even in lower thoracic esophageal cancer. PALN dissection for pathological PALN(+) patients should not be performed. It remains to be determined in future prospective studies whether patients without pathological PALN metastasis, but showing PALN micrometastasis, could achieve improved survival with PALN dissection.

Prevention of gastroduodenal content reflux and delayed gastric emptying after esophagectomy: gastric tube reconstruction with duodenal diversion plus Roux-en-Y anastomosis.

Reflux of gastroduodenal contents and delayed gastric emptying are the most common and serious problems after esophagectomy with gastric reconstruction. However, attempts to reduce the above symptoms, surgically as well as non-surgically, had no or limited effect. To address this issue, we performed retrosternal gastric reconstruction with duodenal diversion plus Roux-en-Y anastomosis (RY) in eight patients with thoracic esophageal cancer and compared the outcomes with control patients who underwent standard reconstruction. The procedure is simple, safe, and not associated with any postoperative complications. The pancreatic amylase concentrations in the gastric juice samples on postoperative day 2 were slightly lower in the non-RY group than in the RY group (1884 ± 2152 vs. 25,790 ± 23,542IU/mL, respectively, P= 0.07). Postoperative endoscopic examination showed neither reflux esophagitis nor residual gastric content in the RY group. Quality of life assessed by the Dysfunction After Upper Gastrointestinal Surgery-32 questionnaire postoperatively was significantly better in the RY group than in the non-RY group for 'decreased physical activity,''symptoms of reflux,''nausea and vomiting,' and 'pain.' The results of this pilot study suggest that gastric reconstruction with duodenal diversion plus RY seems effective in improving both the reflux and delayed gastric emptying. The benefits of this procedure need to be further assessed in a large-scale, randomized controlled trial.

Correlation between pretherapeutic d-dimer levels and response to neoadjuvant chemotherapy in patients with advanced esophageal cancer.

Neoadjuvant chemotherapy may improve survival of responders in esophageal cancer patients but is useless and harmful in non-responders. Thus, it is important to predict the effect of the chemotherapy, and that any predictor must be applicable clinically. The aim of this study is to examine the correlation between pretherapeutic hypercoagulopathy as determined by plasma d-dimer levels and response to chemotherapy. In 71 patients with esophageal cancer who underwent neoadjuvant chemotherapy (cisplatin, adriamycin and 5-fluorouracil) followed by surgery, plasma d-dimer levels were measured before chemotherapy and the clinical and pathological responses to chemotherapy were assessed at 4 weeks after therapy (after surgery). Pretherapeutic plasma d-dimer level was significantly lower in clinical responders (complete response/partial response [CR/PR]; 0.62 +/- 1.10 microg/mL, mean +/- SD) than in non-responders (no change/progressive disease [NC/PD]; 1.15 +/- 1.08 microg/mL, P = 0.0491), and in pathological responders (Grade 1b-3; 0.62 +/- 1.11 microg/mL) and non-responders (Grade 0-1a; 1.15 +/- 1.05 microg/mL, P = 0.0107). The optimal cut-off level of the plasma d-dimer levels for predicting clinical and pathological responses was 0.6 microg/mL. Then, sensitivity and specificity for the prediction of CR/PR were 68% and 73%, and those for Grade 1b-3 were 91% and 69%, respectively. Our results suggested that pretherapeutic plasma d-dimer level correlated significantly with clinical and pathological responses to chemotherapy. Pretherapeutic plasma d-dimer level can be used as a predictor for chemosensitivity.

The toxic conformation of the 42-residue amyloid beta peptide and its relevance to oxidative stress in Alzheimer's disease.

Senile plaques in the brain of patients with Alzheimer's disease mainly consist of aggregates of amyloid beta peptides (Abeta42, Abeta40). Abeta42 is more neurotoxic than Abeta40. This review describes recent findings from a structural analysis of Abeta42 aggregates and discusses their relevance to neurotoxicity through the formation of radicals.

Cellular thiol status-dependent inhibition of tumor cell growth via modulation of p27(kip1) translocation and retinoblastoma protein phosphorylation by 1'-acetoxychavicol acetate.

1'-Acetoxychavicol acetate (ACA) has been shown to inhibit tumor cell growth, but there is limited information on its effects on cell signaling and the cell cycle control pathway. In this study, we sought to determine how ACA alters cell cycle and its related control factors in its growth inhibitory effect in Ehrlich ascites tumor cells (EATC). ACA caused an accumulation of cells in the G1 phase and an inhibition of DNA synthesis, which were reversed by supplementation with N-acetylcysteine (NAC) or glutathione ethyl ester (GEE). Furthermore, ACA decreased hyperphosphorylated Rb levels and increased hypophosphorylated Rb levels. NAC and GEE also abolished the decease in Rb phosphorylation by ACA. As Rb phosphorylation is regulated by G1 cyclin dependent kinase and CDK inhibitor p27(kip1), which is an important regulator of the mammalian cell cycle, we estimated the amount of p27(kip1) levels by western blotting. Treatment with ACA had virtually no effect on the amount of p27(kip1) levels, but caused a decrease in phosphorylated p27(kip1) and an increase in unphosphorylated p27(kip1) as well as an increase in the nuclear localization of p27(kip1). These events were abolished in the presence of NAC or GEE. These results suggest that in EATC, cell growth inhibition elicited by ACA involves decreases in Rb and p27(kip1) phosphorylation and an increase in nuclear localization of p27(kip1), and these events are dependent on the cellular thiol status.

FDG-PET predicts treatment efficacy and surgical outcome of pre-operative chemoradiation therapy for resectable and borderline resectable pancreatic cancer.

BACKGROUND: The efficacy of neoadjuvant chemoradiotherapy (NACRT) for resectable and borderline resectable pancreatic cancer is important for predicting outcomes after radical surgery, but few clinical indicators predict outcome before resection. This study examined the utility of FDG-PET in predicting the efficacy of NACRT and outcome after radical surgery. METHODS: Eighty-three pancreatic cancer patients who underwent FDG-PET before and after NACRT and had positive standard uptake values (SUVs) before NACRT were enrolled in this study. Peri-operative clinical factors, including FDG-PET findings, were examined to predict the efficacy of NACRT and outcome after surgery. RESULTS: Evans grade I, IIA, IIB, III, and IV was determined in 11, 31, 27, 11, and 3 patients, respectively. The maximum SUVs after NACRT (post SUV-max) and tumor size were significantly decreased compared to pretreatment values (p < 0.001 and p = 0.007, respectively). The post SUV-max and regression index were significantly related to grade III/IV (p = 0.04 and p < 0.001, respectively), but only the regression index predicted NACRT efficacy (p = 0.002). The AUC of the regression index for the detection of grade III/IV was 0.822, and 13 of 14 grade III/IV patients were picked up using 50% as the threshold (p < 0.001). Patients with a regression index >50% had a significantly better prognosis after radical resection than patients with <50% (p = 0.032). Regression index as well as pathological lymph node status and resectability status were independent prognostic factors in multivariate analysis (exp 2.086, p = 0.043). CONCLUSION: The regression index is potentially a good indicator of the efficacy of NACRT and outcome after radical resection for pancreatic cancer.

Suppression of tumor promoter-induced oxidative stress and inflammatory responses in mouse skin by a superoxide generation inhibitor 1'-acetoxychavicol acetate.

Double applications of phorbol esters trigger excessive reactive oxygen species (ROS) production in mouse skin. Previously reported data suggest that the two applications induce distinguishable biochemical events, namely, priming and activation. The former is characterized as a recruitment of inflammatory cells, such as neutrophils, by chemotactic factors to inflammatory regions and edema formation. The latter is responsible for ROS generation. Thus, inhibitory effects of 1'-acetoxychavicol acetate (ACA), previously reported to be a superoxide generation inhibitor in vitro, on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced oxidative stress and inflammatory responses in mouse skin model were examined using a double application of ACA. We demonstrated that two pretreatments and pretreatment with ACA (810 nmol) in the activation phase suppressed double TPA application-induced H2O2 formation in mouse skin. ACA exhibited no inhibitory effects on edema formation and the enhancement of myeloperoxidase activity during the first TPA treatment, whereas the anti-inflammatory agent genistein administered at the same dose inhibited both biomarkers. No inhibitory potential of ACA for TPA-induced H2O2 formation in the priming phase was confirmed. On the other hand, in the in vitro study, ACA inhibited ROS generation in differentiated HL-60 cells more strongly than did 1'-hydroxychavicol, which showed no inhibition by pretreatment in the activation phase. In addition, allopurinol did not inhibit double TPA application-induced H2O2 formation in mouse skin. These findings suggest that the NADPH oxidase system of neutrophils rather than the epithelial xanthine oxidase system is dominant for the O2--generating potential in double TPA-treated mouse skin. ACA significantly inhibited mouse epidermis thiobarbituric acid-reacting substance formation, known as an overall oxidative damage biomarker. Moreover, histological studies demonstrated that ACA inhibited double TPA treatment-induced morphological changes reflecting inflammatory response, such as edema formation, leukocyte infiltration, hyperplasia, and cell proliferation. Furthermore, pretreatment with ACA but not 1'-hydroxychavicol in the activation phase inhibits double TPA application-induced increases in both number of leukocytes and proliferating cell nuclear antigen index. These results suggested that ROS from leukocytes including O2- plays an important role for continuous and excessive production of chemotactic factors, leading to chronic inflammation and hyperplasia, which are inhibitable by ACA. Thus, we concluded that O2- generation inhibitors are agents that effectively inhibit oxidative stress and inflammatory responses in mouse skin.

Nitric oxide synthase is induced in tumor promoter-sensitive, but not tumor promoter-resistant, JB6 mouse epidermal cells cocultured with interferon-gamma-stimulated RAW 264.7 cells: the role of tumor necrosis factor-alpha.

Expression of inducible nitric oxide synthase (iNOS) has been reported to be involved in certain organs of potential tumorigenesis, including the stomach and colon. The mechanisms for iNOS expression in epithelial cells, however, are not fully understood. In the present study, we investigated the role of macrophages in epithelial iNOS expression by coculturing a stimulated murine macrophage-like cell line, RAW 264.7, with either tumor promoter-sensitive (P+) or promoter-resistant (P-) JB6 murine epidermal cells. After monoculture, treatment of RAW 264.7 cells with IFN-gamma for 24 h generated a large amount of nitrite (NO2-), as reported previously, whereas no increase in NO2- concentration was observed in the IFN-gamma-treated P+ or P-subclones. Interestingly, when IFN-gamma-treated RAW 264.7 cells were cocultured with P+ but not P- cells, we observed a marked increase in NO2- concentration (30.8+/-3.6 microM), which significantly exceeded (P < 0.01) the sum of the concentrations (20.0+/-2.3 microM) added from each cell line monoculture. Western blotting analysis revealed that, after coculture, iNOS protein was up-regulated 55-fold more than the control in JB6 P+ but not in P- cells. IFN-gamma-treated RAW 264.7 cells secreted proinflammatory cytokines, including tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta. The addition of IFN-gamma-treated RAW 264.7 cell-conditioned media to P+ subclones led to a significant enhancement of NO2- formation that was diminished by the TNF-alpha-specific but not IL-1beta-specific antibody. When combined with IFN-gamma, the recombinant TNF-alpha (1-100 ng/ml) enhanced NO2- formation in JB6 P+ cells, whereas IL-1beta (1-100 ng/ml) did not. These results led us to conclude that IFN-gamma-treated RAW 264.7 cells release TNF-alpha to induce iNOS expression in promoter-sensitive JB6 cells. Thus, we propose the hypothesis that macrophages stimulate neoplastic cells with TNF-alpha via a paracrine loop to induce epithelial iNOS protein expression.

Redox regulation of glutathione S-transferase induction by benzyl isothiocyanate: correlation of enzyme induction with the formation of reactive oxygen intermediates.

Here we report the molecular mechanism underlying the induction of glutathione S-transferase (GST) in rat liver epithelial RL34 cells treated with a cancer chemopreventive isothiocyanate compound, benzylisothiocyanate (BITC). BITC was found to significantly induce GST activity in RL34 cells. Northern and Western blot analyses demonstrated that BITC specifically enhanced the production of the class pi GST isozyme (GSTP1). Our studies demonstrated for the first time that the addition of BITC to the cells resulted in an immediate increase in the reactive oxygen intermediates (ROIs) detected by a fluorescence probe, 2',7'-dichlorofluorescin diacetate. The level of the ROIs in the cells treated with BITC (10 microM) was approximately 50-fold higher than those in the control cells. Furthermore, glutathione depletion by diethyl maleate significantly enhanced BITC-induced ROI production and accelerated the BITC-induced elevation of the GST activity, whereas pretreatment of the cells with glutathione inhibited both the ROI production and GST induction. The structure-activity relationship of the isothiocyanates also indicated that the ROI-producing activities closely correlated with their GST-inducing potencies. Moreover, the GSTP1 enhancer I-containing region was found to be essential for induction of the GSTP1 gene by intracellular ROI inducers such as BITC and diethyl maleate. These data suggest the involvement of the redox regulation on the induction of GSTP1 by BITC.

Inhibitory effect of citrus nobiletin on phorbol ester-induced skin inflammation, oxidative stress, and tumor promotion in mice.

The intake of citrus fruits has been suggested as a way to prevent the development of some types of human cancer. Nitric oxide (NO) is closely associated with the processes of epithelial carcinogenesis. We attempted a search for NO generation inhibitors in Citrus unshiu. The active constituent was traced by an activity-guiding separation. NO and superoxide (O2-) generation was induced by a combination of lipopolysaccharide and IFN-gamma in mouse macrophage RAW 264.7 cells, and by 12-O-tetradecanoylphorbol-13-acetate (TPA) in differentiated human promyelocyte HL-60, respectively. Expression of inducible NO synthase and cyclooxygenase 2 proteins were detected by Western blotting. The in vivo anti-inflammatory and antitumor promoting activities were evaluated by topical TPA application to ICR mouse skin with measurement of edema formation, epidermal thickness, leukocyte infiltration, hydrogen peroxide production, and the rate of proliferating cell nuclear antigen-stained cells. As a result, nobiletin, a polymethoxyflavonoid, was identified as an inhibitor of both NO and O2- generation. Nobiletin significantly inhibited two distinct stages of skin inflammation induced by double TPA application [first stage priming (leukocyte infiltration) and second stage activation (oxidative insult by leukocytes)] by decreasing the inflammatory parameters. It also suppressed the expression of cyclooxygenase-2 and inducible NO synthase proteins and prostaglandin E2 release. Nobiletin inhibited dimethylbenz[a]anthracene (0.19 micromol)/TPA (1.6 nmol)-induced skin tumor formation at doses of 160 and 320 nmol by reducing the number of tumors per mouse by 61.2% (P < 0.001) and 75.7% (P < 0.001), respectively. The present study suggests that nobiletin is a functionally novel and possible chemopreventive agent in inflammation-associated tumorigenesis.

Citrus auraptene exerts dose-dependent chemopreventive activity in rat large bowel tumorigenesis: the inhibition correlates with suppression of cell proliferation and lipid peroxidation and with induction of phase II drug-metabolizing enzymes.

In our previous short-term experiment, Citrus auraptene inhibited the development of azoxymethane (AOM)-induced aberrant crypt foci, which are precursor lesions for colorectal carcinoma. In the present study, the possible inhibitory effect of dietary administration of auraptene was investigated using an animal colon carcinogenesis model with a colon carcinogen AOM. Male F344 rats were given s.c. injections of AOM (15 mg/kg body weight) once a week for 3 weeks to induce colon neoplasms. They also received diets containing 100 or 500 ppm auraptene for 4 weeks in groups of "initiation" feeding, starting 1 week before the first dosing of AOM. The diets containing auraptene were also given to rats for 38 weeks in groups of "postinitiation" feeding. At the termination of the study (38 weeks), dietary administration of auraptene caused dose-dependent inhibition in AOM-induced large bowel carcinogenesis. Auraptene feeding during the initiation phase reduced the incidence of colon adenocarcinoma by 49% at 100 ppm (P = 0.099) and 65% at 500 ppm (P = 0.0075). Auraptene administration during the postinitiation phase inhibited the incidence of colon adenocarcinoma by 58% at 100 ppm (P = 0.021) and 65% at 500 ppm (P = 0.0075). Also, the multiplicity of colon carcinoma was significantly reduced by initiation feeding at a dose level of 500 ppm (P < 0.01) and postinitiation feeding at a level of 100 and 500 ppm (P < 0.05 and P < 0.01, respectively). Feeding of auraptene suppressed the expression of cell proliferation biomarkers (ornithine decarboxylase activity and polyamine content) in the colonic mucosa and reduced the production of aldehydic lipid peroxidation [malondialdehyde and 4-hydroxy-2(E)-nonenal]. In addition, auraptene increased the activities of Phase II drug-metabolizing enzymes (glutathione S-transferase and quinone reductase) in the liver and colon. These findings suggest that the inhibitory effects of auraptene on AOM-induced colon tumorigenesis at the initiation level might be associated, in part, with increased activity of Phase II enzymes, and those at the postinitiation stage might be related to suppression of cell proliferation and lipid peroxidation in the colonic mucosa.

Expression of N-acetylglucosaminyltransferase V in colorectal cancer correlates with metastasis and poor prognosis.

N-Acetylglucosaminyltransferase V (GnT-V) is an enzyme that catalyzes beta 1-6 branching of N-acetylglucosamine on asparagine-linked oligosaccharides of cell proteins. Metastatic potential of various cancer cells has been shown to correlate with increase of GnT-V activity and concomitant beta 1-6 branching of N-acetylglucosamine. However, protein expression of GnT-V in human cancer tissue and its clinical significance have not yet been demonstrated. To clarify the possible relationship between metastasis and GnT-V in human colorectal cancer, protein expression of GnT-V was studied using surgically resected specimens. We established a monoclonal antibody against GnT-V and performed immunohistochemical analysis of 103 human colorectal cancer cases. Of 103 cases, 26 cases (25.2 %) showed specific expression of GnT-V in colorectal cancer tissues. The expression of GnT-V was significantly correlated with distant metastasis (P < 0.05, chi2 test). Overall 5-year survival rate was 52.8% for GnT-V-positive patients and 81.7% for GnT-V-negative patients (P < 0.01, Log-rank test). We showed direct evidence for the relationship between GnT-V and metastasis in human colorectal cancer. Screening of GnT-V expression in colorectal cancer may provide useful information for prognosis of postoperative patients.

A catechol antioxidant protocatechuic acid potentiates inflammatory leukocyte-derived oxidative stress in mouse skin via a tyrosinase bioactivation pathway.

The modifying effects of topical application of a catechol antioxidant protocatechuic acid (PA) on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced inflammatory responses in mouse skin were investigated. Treatment with a high dose (20,000 nmol) of PA, based on time of application, modifies inflammatory responses in the skin of the B6C3F(1) mouse, a resistant strain to inflammatory response induction by TPA, but shows much higher tyrosinase expression than that of an albino mouse. The responsibility of a large amount of PA-induced leukocyte infiltration to an inflamed region in a B6C3F(1) mouse is more sensitive than that of an ICR albino mouse. When ICR mice were treated with TPA (1.6 nmol) twice weekly for 5 weeks to induce chronic inflammatory responses, pretreatment with 1600 nmol PA 30 min prior to each TPA treatment significantly enhanced the inflammatory responses including edema formation, leukocyte infiltration, and the level of thiobarbituric acid-reacting substances. The dose-dependency was closely parallel to the results of a tumor promotion study of PA previously reported. Further, the treatment of PA alone resulted in tyrosinase-dependent contact hypersensitivity in ICR mouse skin. In addition, the in vitro study of cytotoxicity demonstrated that bioactivation by tyrosinase but not myeloperoxidase of PA significantly enhanced cytotoxicity and intracellular glutathione consumption. We conclude that the tyrosinase-derived reactive quinone intermediate(s) of PA, which binds nucleophilic residues of proteins including sulfhydryl group and conjugates of which are recognized as haptens, was partially involved in alteration of the cellular immune functions including oxygen radical-generating leukocytes migration to inflamed regions.

Down-regulation of focal adhesion kinase, pp125FAK, in endothelial cell retraction during tumor cell invasion.

Although endothelial cell retraction is required before tumor cell invasion, its molecular mechanism still remains obscure. We previously demonstrated that conditioned medium (CM) derived from a human pancreatic cancer cell line, PSN-1, induced endothelial cell retraction and facilitated tumor cell invasion. To investigate the molecular change of events in the transduction of extracellular signals during endothelial cell retraction, we examined the effect of the CM derived from PSN-1 cells on the tyrosine phosphorylation in endothelial cells. Immunoblot analyses revealed that the PSN-1 CM decreased tyrosine phosphorylation of a 120-130 kD protein, and induced the concomitant down-regulation of focal adhesion kinase, pp125FAK, during endothelial cell retraction in time- and dose-dependent fashions. These changes preceded endothelial cell retraction and were reversible after removal of the CM. Further quantitative densitometric analyses demonstrated that the extent of decrease in tyrosine phosphorylated 120-130 kD protein during the endothelial cell retraction was likely to be proportional to that of the down-regulation of pp125FAK. A tyrosine phosphorylated 120-130 kD protein immunoprecipitated by anti-phosphotyrosine antibody immunoreacted with anti-pp125FAK antibody. These results suggested that decreased amount of a tyrosine phosphorylated 120-130 kD protein probably due to the down-regulation of pp125FAK might be associated with the signal transduction pathway in the endothelial cells during their retraction. Furthermore, these findings were also observed in the CM from another four human cancer cell lines, suggesting the down-regulation of pp125FAK in endothelial cells during tumor cell invasion.

Phosphodiesterase type III inhibitor, cilostazol, inhibits colon cancer cell motility.

Metastasis of cancer cells is initiated by the cellular migration into extracellular matrix and surrounding vessels. We previously showed that elevation of cAMP levels in cancer cells suppressed trans-cellular migration in vitro. Drugs that can elevate cAMP levels in cancer cells effectively may be applied to prevent metastasis in cancer patients. Cilostazol, an oral anti-platelet drug, is a specific cAMP phosphodiesterase type III inhibitor and has been clinically used to treat thrombosis patients. In chemotaxis assay, cellular migration of human colon cancer cells, DLD- 1, was induced by 10 microg/ml of soluble fibronectin or 10% of fetal bovine serum (FBS). Treatment with cilostazol (50 microM) suppressed 92.3% or 84.6% of the migration in control cells, respectively. When DLD-1 cells were stimulated by soluble fibronectin in phagokinetic assay, migration assessed by the area of gold particle phagocytosis track was induced and cilostazol also decreased 67.3% of the cellular migration in control cells. Furthermore, in the trans-cellular migration assay, cilostazol suppressed cancer cell invasion induced by FBS. Thus, cilostazol can suppress colon cancer cell motility and might be effective as an anti-metastasis drug for cancer patients.

Inhibitory effects of ursolic and oleanolic acid on skin tumor promotion by 12-O-tetradecanoylphorbol-13-acetate.

Ursolic acid (UA) and oleanolic acid (OA), which had been isolated from Glechoma hederacea as inhibitors of Epstein-Barr virus (EBV) activation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), were tested against inhibitory effect on tumor promotion by TPA in vivo. They inhibited effectively the tumor promotion in mouse skin and the activities were comparable to that of a known inhibitor of tumor promotion, retinoic acid (RA). Interestingly, UA was more effective on a single application before initial TPA-treatment than on a continuous application before each TPA-treatment, while OA and RA were ineffective in the same treatment. These data suggest that the role of UA for inhibitory action on tumor promotion differs slightly from those of RA and OA.

Inhibitory effect of pheophorbide a, a chlorophyll-related compound, on skin tumor promotion in ICR mouse.

Anti-tumor-promoting activity of pheophorbide a (PPBa) a chlorophyll-related compound, was examined in a two-stage carcinogenesis experiment in ICR mouse skin by 7,12-dimethylbenz[a] anthracene (DMBA, 0.19 mumol) and 12-O-tetradecanoylphorbol-13-acetate (TPA, 1.6 nmol). Topical application of PPBa (160 nmol) markedly reduced the average number of tumors per mouse and the ratio of tumor-bearing mice (inhibitory ratio: IR = 56%, P < 0.01 and 31%, P < 0.005, respectively). PPBa exhibited potent anti-inflammatory activity in ICR mouse ears and moderate inhibitory activity toward TPA-induced superoxide (O2-) generation in differentiated HL-60 cells. While CuPPBa, a synthetic copper complex of PPBa, exhibited higher anti-inflammatory activity than that of indomethacin, it showed little antioxidative effect against formation of lipid hydroperoxides (LOOHs) and malondialdehyde (MDA), suggesting that the antioxidative effect of PPBa might not be important for anti-inflammatory activity. These results imply that the active mechanism of PPBa for anti-tumor promotion might be partly involved in inhibition of TPA-induced inflammatory responses by suppressing leukocyte activation.

Screening for in vitro anti-tumor promoting activities of edible plants from Thailand.

A total of 112 species of edible plants (122 samples) from Thailand were randomly collected, and their methanol extracts were screened for in vitro anti-tumor promoting activity using the inhibition test of Epstein-Barr virus (EBV) activation in Raji cells induced by 12-O-hexadecanoylphorbol-13-acetate (HPA, 40 ng/ml). It was found that 60% of these extracts inhibited EBV activation by 30% or more at a concentration of 200 mg/ml. Significantly, the ratio is markedly higher than that (26%) previously observed in common edible plants in Japan. Thus, physiological potentiality of edible Thai plants has been implied in terms of cancer chemoprevention.

Epstein-Barr virus-activating principle in the ether extracts of soils collected from under plants which contain active diterpene esters.

Soil samples were collected from the ground under the plants of Euphorbiaceae and Thymelaeaceae known to possess Epstein-Barr virus-activating diterpene esters. In a test system, the ether extracts of such soil samples at a concentration of 20 micrograms/ml induced Epstein-Barr virus early antigen in approximately 5-25% of the non-producer Raji cells. These findings suggest a possible interaction between plant-derived diterpene esters and the human system, and provide a new aspect in considering the cause of Epstein-Barr virus-associated diseases, particularly nasopharyngeal carcinoma.

Screening of edible plants against possible anti-tumor promoting activity.

One hundred twenty-one species of edible plants (133 test-parts) were screened against possible anti-tumor promoting activity by an in vitro short-term assay system of inhibition of Epstein-Barr virus (EBV) activation induced by a phorbol-ester promoter, 12-O-hexadecanoylphorbol-13-acetate (HPA). The methanol-extracts (ME) of 14 species of the edible plants strongly inhibited the activation, 7 moderately and 12 weakly inhibited it. On partition of the randomly selected inactive ME (26 species) with ethyl acetate and water, 13 and 2 species were active, more or less, in the ethyl acetate and water soluble part, respectively. Thus, this result suggested that anti-tumor promoters occur in a wide variety of edible plants. The anti-tumor promoting activity in the crude extracts may be enhanced or reduced with co-occurring factors acting additively, synergistically or antagonistically.