Four New Xanthones from Cratoxylum cochinchinense and Their In Vitro Antiproliferative Effects.

The study of the chemical constituents of branches and twigs of Cratoxylum cochinchinense collected in Singapore led to the isolation and structural elucidation of four new xanthones, named cratoxanthone A (1), B (2), C (3), and D (4), together with six known xanthones (5-10) and one known dihydroanthracenone (11). Eight xanthones (including 1 and 2) and 11 were tested for their antiproliferative activity in three human carcinoma cell lines (lung adenocarcinoma A549, colorectal carcinoma Colo205, and epidermoid carcinoma KB) and a human acute lymphoblastic leukemia B cell line (NALM-6), and the mitochondrial membrane potential was determined in KB cells. New xanthones 1 and 2 attenuated NALM-6 cell proliferation with IC50 values of 17.78 and 8.27 µM, respectively. Furthermore, KB cells treated with these compounds had significantly decreased mitochondrial membrane potentials. Notably, the proliferation of A549 cells was specifically inhibited by 11, but not the xanthones.

Three new acridone alkaloids from Glycosmis lanceolata (Blume) D.Dietr. and their cytotoxic effects on tumour cell lines.

We separated and structurally elucidated three new acridone alkaloids (glycomontamine A (1), B (2), and C (3)), together with three known compounds (glycofoline, kokusaginine and dictamnine) from the acetone extract of Glycosmis lanceolata (Blume) D.Dietr. branches collected in Thailand. The compounds were assayed for cell viability using human lung adenocarcinoma cell line A549, breast adenocarcinoma cell line T47D, cervix epithelioid carcinoma cell line Hela, acute lymphoid leukaemia B cell line NALM-6, and human dermal fibroblasts. The viability of Hela cells treated with compound 1 (IC50 17.6 µM) and T47D cells treated with compound 2 (IC50 17.4 µM) decreased dose-dependently. Both compounds also showed cytotoxicity against NALM-6 cells (IC50 16.5 and 9.3 µM). Additionally, compound 1 decreased the mitochondrial membrane potential of Hela cells, whereas compound 2 did not change the mitochondrial membrane potential in T47D cells.

Lansiumamide B and SB-204900 isolated from Clausena lansium inhibit histamine and TNF-α release from RBL-2H3 cells.

AIMS AND OBJECTIVE: Mast cells play a central role in allergic and chronic inflammation. Extracts from Clausena lansium (Lour.) Skeels (Rutaceae) possess many pharmacological effects including anti-inflammatory, anti-oxidant, anti-cancer, and anti-trichomonal activities. In addition, the leaves and fruit are used in Chinese folk medicine. We have isolated and identified four known cinnamamides from this plant: lansiumamide C, lansamide I, lansiumamide B, and SB-204900. However, the biological activities of these compounds are not yet understood. The purpose of this paper is to clarify the pharmacological effects of these compounds on mast cells. METHODS: We measured inflammatory molecules in A23187-stimulated rat basophilic leukemia cells (RBL-2H3) treated with these compounds using HPLC, ELISA, and immunoblotting methods. In addition, some signaling molecules were investigated by immunoblotting. RESULTS: Lansamide I, lansiumamide B, and SB-204900 significantly decreased histamine release. Furthermore, lansiumamide B- and SB-204900-treated cells also reduced the protein and/or mRNA levels of TNF-α. SB-204900 markedly suppressed the phosphorylation of p38 MAPK. CONCLUSION: Our findings suggest that lansiumamide B and SB-204900 attenuate mast-cell-induced inflammation.

Three phlorotannins from Sargassum carpophyllum are effective against the secretion of allergic mediators from antigen-stimulated rat basophilic leukemia cells.

Sargassum carpophyllum (Sargassaceae) is a brown seaweed that contains phlorotannins, which are phloroglucinol polymers with reported anti-inflammatory activities. The phlorotannins 2-[2-(3,5-dihydroxyphenoxy)-3,5-dihydroxyphenoxy]-1,3,5-benzenetriol (1), 2,2'-[[2-(3,5-dihydroxyphenoxy)-5-hydroxy-1,3-phenylene]bis(oxy)]bis(1,3,5-benzenetriol) (2), and 2-[2-[4-[2-(3,5-dihydroxyphenoxy)-3,5-dihydroxyphenoxy]-3,5-dihydroxyphenoxy]-3,5-dihydroxyphenoxy]-1,3,5-benzenetriol (3) were isolated from S. carpophyllum. Here, we evaluated the anti-allergic activities of these compounds and comprehensively explored their effects on intracellular protein levels. Immunoglobulin E-sensitized rat basophilic leukemia cells pretreated with any of these three compounds exhibited reduced ß-hexosaminidase, prostaglandin D2, and tumor necrosis factor-α secretion compared with dinitrophenyl-human serum albumin (DNP-HSA)-stimulated cells. Reduction of ß-hexosaminidase release was dose-dependent but the half-maximal inhibitory concentrations of the compounds were similar (36-51 µM). Proteomics analysis revealed that the three compounds up-regulated 25 proteins and down-regulated 33 proteins compared with DNP-HSA stimulation alone, and slightly suppressed proteasome 5 expression linked to the regulation of IκB. These results demonstrate that these phlorotannins are potentially useful for preventing immediate hypersensitivity. S. carpophyllum may be a functional food.

Four new isoflavones from Derris scandens and their in vitro antiproliferative effects.

Four new compounds (derriscandenon D (1), E (2), F (3), G (4)) and six known isoflavones (warangalone (5), millewanin E (6), rhynedlin A (7), 6,8-diprenylgenistein (8), isolupalbigenin (9), isoscandinone (10)) were isolated from the acetone extract of the branches of Derris scandens. These compounds were assayed for cell viability using the human lung carcinoma cell line A549, colorectal carcinoma cell line Colo205, epidermoid carcinoma cell line KB, the human acute lymphoblastic leukaemia cell line NALM-6, and human dermal fibroblasts. Compounds 2 and 3 significantly decreased the viability of KB cells, with IC50 values of 2.7 and 12.9 µM, respectively. In addition, compounds 2 and 3 reduced the mitochondrial membrane potential in KB cells. Compounds 2 and 3 strongly down-regulated the cell viability of cell lines KB and NALM-6, achieving IC50 values of 2.7 and 0.9 µM, respectively, compared with the positive control staurosporine at 1.25 and 0.01 µM, respectively.

Evaluation of flavoglaucin, its derivatives and pyranonigrins produced by molds used in fermented foods for inhibiting tumor promotion.

Flavoglaucin, its derivatives, and pyranonigrins, which are antioxidants produced by the molds used in fermented foods, were examined for their inhibition of tumor promotion by the Epstein-Barr virus early antigen activation test. Flavoglaucin and its derivatives exhibited high activity. Flavoglaucin and such a derivative as isodihydroauroglaucin inhibited mouse skin tumor promotion in a two-stage carcinogenesis test and appear to be antitumor promoters.

Cytotoxic activity of dimeric acridone alkaloids derived from Citrus plants towards human leukaemia HL-60 cells.

OBJECTIVES: Acridone alkaloids from Citrus and their derivatives show various kinds of biological activity. However, the anticancer activities of dimeric acridone alkaloids with unique structures and the molecular mechanism of these effects are poorly understood. METHODS: We investigated the cytotoxicity effects of dimeric acridone alkaloids isolated from Marsh grapefruit on human myeloid leukaemia HL-60 cells. KEY FINDINGS: Of the six dimeric acridone alkaloids tested, citbismine-E, the most potent, dose- and time-dependently decreased HL-60 cell viability by inducing apoptosis. The treatment of HL-60 cells with citbismine-E yielded a significant increase in levels of intracellular reactive oxygen species (ROS). Citbismine-E lowered the mitochondrial membrane potential and increased the activities of caspase-9 and -3. In addition, citbismine-E-induced apoptosis, decrease in mitochondrial membrane potential and caspase activation were significantly alleviated by pretreatment of the cells with antioxidant N-acetylcysteine (NAC). Citbismine-E induced intrinsic caspase-dependent apoptosis through ROS-mediated c-Jun N-terminal kinase activation. Citbismine-E-induced production of oxidative stress biomarkers, malondialdehyde and 8-hydroxy-2'-deoxyguanosine was also attenuated by pretreatment with NAC. CONCLUSIONS: Citbismine-E is a powerful cytotoxic agent against HL-60 cells that acts by inducing mitochondrial dysfunction-mediated apoptosis through ROS-dependent JNK activation. Citbismine-E also induced oxidative stress damage via ROS-mediated lipid peroxidation and DNA damage in HL-60 cells.

Three isoflavones from Derris scandens (Roxb.) Benth and their cancer chemopreventive activity and in vitro antiproliferative effects.

Three undescribed isoflavones, derriscandenon A, B, and C, together with seven known isoflavones were isolated and structurally characterized during a study of the chemical constituents in the leaves of Derris scandens (Roxb.) Benth (Leguminosae, Fabaceae) collected in Bangladesh. The inhibitory activity of the compounds against activation of Epstein-Barr virus antigen (EBV-EA) by 12-O-tetradecanoylphorbo-13-acetate (TPA) was measured to identify possible chemopreventive agents. Mild inhibitory effects (IC50 278-290 mol ratio/32 pmol TPA) against EBV-EA induction compared with curcumin (IC50 341 mol ratio/32 pmol TPA) were observed for four known compounds (lupalbigenin, isopalbigenin, glyurallin, and isangustone A). Next, we focused on antitumor effects and investigated cell viability, cell proliferation, and mitochondria membrane potential by using an MTT assay, a live cell monitoring system, and fluorescence staining. Of the seven isoflavones tested for cell viability, a dose-dependent decrease in cell viability was observed for four isoflavones (derriscandenon B and C, derrubone, and glyurallin) in KB cells and two compounds (derriscandenon B and isochandaisone) in NALM6-MSH+ cells. In addition, the proliferation of KB cells was significantly inhibited by these four compounds at a concentration of 5 µM. The mitochondria membrane potentials of KB cells treated with derriscandenon C, derrubone, and glyurallin at the IC50 concentration were decreased by about 55%, whereas undescribed compound derriscandenon B had no effect. Our results show that some of the compounds isolated from D. scandens may be suitable as seed compounds for cancer prevention and therapy.

Gamma-lactone carbazoles from Clausena anisata.

The study of chemical constituents of the stems of Clausena anisata collected in Thailand led to the isolation and identification of eight known and two new carbazole alkaloids named furanoclausamines A (1) and B (2). Clausamine E (3) was found to exhibit cytotoxicity against the human leukemia cell line HL-60.

Antioxidants produced by Eurotium herbariorum of filamentous fungi used for the manufacture of karebushi, dried bonito (Katsuobushi).

Extracts prepared by culturing ten filamentous fungi from Aspergillus and Eurotium species isolated from dried bonito (katsuobushi) were examined for 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capacity. The extracts prepared by culturing E. herbariorum NE-1 and NE-4, which are used in the molding process for the manufacture of karebushi (a kind of katsuobushi), were shown to have higher activity than the others. Five antioxidants were isolated from the extracts and identified as isodihydroauroglaucin (IDAG), auroglaucin (AG), dihydroauroglaucin (DAG), tetrahydroauroglaucin (TAG), and flavoglaucin (FG) by (1)H-NMR, (13)C-NMR, and EI-MS analyses. Compared with alpha-tocopherol, the isolated antioxidants exhibited high antioxidative activity for the radical scavenging capacity of DPPH and superoxide, but low activity for inhibiting the autoxidation of docosahexaenoic acid (DHA). The isolated antioxidants were produced by the Eurotium species, but not by the Aspergillus species. DAG and TAG exhibited higher radical scavenging capacity than the other antioxidants and were abundantly contained in the extracts of E. herbariorum NE-1 and NE-4.

Effect of natsudaidain isolated from Citrus plants on TNF-alpha and cyclooxygenase-2 expression in RBL-2H3 cells.

OBJECTIVES: Flavonoids inhibit the activity of chemical mediators released from mast cells. Our aim was to investigate the effects of natsudaidain, a polymethoxyflavone isolated from Citrus plants, on mast cells. METHODS: We investigated the inhibitory effects of natsudaidain, which is a polymethoxyflavone isolated from Citrus plants, on histamine release, tumour necrosis factor-alpha production and cyclooxygenase-2 expression in Ca ionophore-stimulated rat basophilic leukemia cells (A23187-stimulated RBL-2H3 cells) by spectrofluorometric, ELISA and immunoblotting methods. KEY FINDINGS: The percent of histamine release from A23187-stimulated RBL-2H3 cells pretreated with natsudaidain at 5, 25 and 50 microM was not changed as compared with non-treated A23187-stimulated cells. At 100 and 200 microM, natsudaidain pretreatment resulted in slightly reduced histamine release (% histamine release, 89.8+/-3.5% and 71.5+/-5.6% at 100 and 200 microM). Thus, natsudaidain hardly affects histamine release from RBL-2H3 cells, except at high concentrations. On the other hand, natsudaidain dose-dependently inhibited tumour necrosis factor-alpha protein and mRNA levels in A23187-stimulated RBL-2H3 cells; a concentration of 6.8 microM was required for a 50% reduction. In addition, all concentrations of this compound that we tested also inhibited cyclooxygenase-2 protein expression. The mRNA levels of cyclooxygenase-2 in A23187-stimulated RBL-2H3 cells treated with natsudaidain were also markedly decreased. The phosphorylated-p38 MAPK protein levels in A23187-stimulated RBL-2H3 cells treated with natsudaidain were lower than in the non-treated cells. CONCLUSIONS: These findings suggest that natsudaidain inhibits tumour necrosis factor-alpha and cyclooxygenase-2 production by suppressing p38 MAPK phosphorylation but not p65 NFkappaB phosphorylation, and that natsudaidain might alleviate inflammatory diseases.

Acrofolione A and B, acetophenone dimers from Acronychia pendunculata, induce an apoptotic effect on human NALM-6 pre-B cell leukaemia cells.

OBJECTIVES: We investigated the apoptotic activities of acrofolione A (1) and B (2) isolated from Acronychia pedunculata against a human pre-B cell leukaemia cell line (NALM-6) to explore the apoptosis-related signalling molecules targeted by 1 and 2. METHODS: The apoptosis effects of 1 and 2 in NALM-6 cells were investigated by TUNEL staining, annexin V, mitochondria membrane potential and caspase 3/7 activity. We carried out a protein array to explore the signalling molecules involved in apoptosis comprehensively. KEY FINDINGS: Acrofolione A (1) suppressed the growth of NALM-6, K562 and HPB-ALL cells (IC50 16.7 ± 1.9, 17.9 ± 0.3 and 10.1 ± 0.2 µm, respectively) more effectively than acrofolione B (2). Both compounds time-dependently increased the number of NALM-6 cells with abnormal nuclei, and increased the number of annexin V-positive cells and decreased the mitochondrial membrane potential of NALM-6 cells. Acrofolione A (1) markedly elevated caspase 3/7 activity and increased the number of TUNEL-positive cells. Cells treated with either compound showed enhanced expression of cleaved PARP and cleaved caspase 3 and 7, and reduced survivin protein levels. CONCLUSIONS: Acrofolione A (1) and B (2) may be useful in the treatment of various types of leukaemia.

Antioxidative pyranonigrins in rice mold starters and their suppressive effect on the expression of blood adhesion molecules.

Antioxidants having a 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging capacity in rice mold starters, which are used for the preparation of various Japanese fermented foods, and their effectiveness against the expression of blood adhesion molecules were examined. An antioxidant was isolated from the rice mold starters used for shochu and identified as pyranonigrin-S (PG-S) by (1)H-NMR, (13)C-NMR, and FAB-MS analyses. It was a derivative of pyranonigrin-A (PG-A), which has been isolated as an antioxidant from the rice mold starters. Pyranonigrins PG-A and PG-S were found to exist in spores on rice mold starters which had been prepared by Aspergillus awamori, A. kawachii, and A. saitoi. PG-S exhibited a higher level of DPPH radical scavenging activity than PG-A. PG-A was found to have a significant suppressive effect on the expression of vascular cell adhesion molecule-1 (VCAM-1) in human umbilical vein endothelial cells (HUVECs) induced by tumor necrosis factor-alpha (TNF-alpha) (P<0.05).

Induction of apoptosis in human leukaemia HL-60 cells by furanone-coumarins from Murraya siamensis.

To identify potential anti-tumour agents, we screened five furanone-coumarins isolated from Murraya siamensis Craib (Rutaceae) for their ability to inhibit the growth of human leukaemia HL-60 cells. Among the furanone-coumarins tested, murrayacoumarin B (compound 2) showed significant cytotoxicity against HL-60 cells. Fluorescence microscopy with Hoechst 33342 staining revealed that the percentage of apoptotic cells with fragmented nuclei and condensed chromatin increased in a time-dependent manner after treatment with murrayacoumarin B. Interestingly, this furanone-coumarin induced the loss of the mitochondrial membrane potential. In addition, treatment with murrayacoumarin B stimulated the activities of caspase-9 and caspase-3, and caspase-9 and caspase-3 inhibitors suppressed the apoptosis induced by murrayacoumarin B. These results suggest that murrayacoumarin B induced apoptosis in HL-60 cells through activation of the caspase9/caspase-3 pathway triggered by mitochondrial dysfunction.

Anti-cell proliferation effect of naphthoquinone dimers isolated from Plumbago zeylanica.

Study of the chemical constituents of the roots of Plumbago zeylanica L. collected in Taiwan led to the isolation and identification of a new naphthoquinone dimer, plumzeylanone (1), along with eight known compounds (2-9). Nine naphthoquinones isolated from this plant were assayed for cell growth inhibition activity using NALM-6 (human B cell precursor leukaemia), A549 (human lung adenocarcinoma), Colo205 (human colorectal adenocarcinoma) and KB (human epidermoid carcinoma). Plumzeylanone (1), a novel plumbagin dimer, suppressed cell proliferation in only NALM-6 cells (IC50 3.98 µM). However, maritinone (9) showed strong inhibition of cell growth in all cell lines tested (0.12 < IC50 < 9.06 µM). This compound appeared to affect the cell cycle.

Synthetic cinnamylphenol derivatives as cancer chemopreventive agents.

Several substituted cinnamylphenol (1,3-diphenylpropene) derivatives were synthesized and tested for their inhibitory activities against in vitro Epstein-Barr virus early antigen activation induced by 12-O-tetradecanoylphorbol-13-acetate in Raji cells. The prenylated cinnamylphenols were found to show remarkably potent activity. Furthermore, prenylated cinnamylphenols (19 and 25) exhibited a marked inhibitory effect on mouse skin tumor promotion in an in vivo two-stage carcinogenesis test. These results indicate that some prenylated cinnamylphenols might be valuable as potential cancer chemopreventive agents (anti-tumor promoters).

Apoptosis inducing activity of 4-substituted coumarins from Calophyllum brasiliense in human leukaemia HL-60 cells.

With the objective of identifying anti-tumour-promoting agents, we carried out a primary screening of ten 4-substituted coumarins isolated from Calophyllum brasiliense Camb. (Guttiferae), to determine the ability of these compounds to inhibit proliferation of the human leukaemia cell line HL-60. Among the 4-substituted coumarins isolated, calophyllolide (2) and mammea B/BB (3) showed significant cytotoxicity against HL-60 cells. Fluorescence microscopy with Hoechst 33342 staining revealed that the percentage of apoptotic cells with fragmented nuclei and condensed chromatin increased in a time-dependent manner after treatment with calophyllolide (2) or mammea B/BB (3). In addition, the activity of caspase-9 and caspase-3 was also enhanced in a time-dependent manner upon treatment with the 4-substituted coumarins 2 and 3. Caspase-9 and caspase-3 inhibitors suppressed apoptosis induced by 4-substituted coumarins 2 and 3. These results suggest that calophyllolide (2) and mammea B/ BB (3) induced apoptosis in HL-60 cells through activation of the caspase-9/caspase-3 pathway, which is triggered by mitochondrial dysfunction.

Acetophenones Isolated from Acronychia pedunculata and their Anti-proliferative Activities.

Study of the chemical constituents of Acronychia pedunculata (L.) Miq. (Rutaceae) stems collected in Taiwan led to the isolation and identification of eight known and three new acetophenones, named acrophenone A (1), B (2), and C (3). Of them, acrovestone (5), acropyrone (6) and acrovestenol (7), which are dimer compounds, strikingly inhibited the proliferation of human leukemia cell lines.

Acetophenones from Acronychia pedunculata and their Cancer Chemopreventive Activity.

From the roots of Acronychia pedunculata (L.) Miq. (Rutaceae) collected in Taiwan, six known and three new acetophenones have been isolated. The new compounds were named acrophenones D (1), E (2), and F (3). Of the acetophenones isolated in this study, prenylacronylin (4) and acronyculatin D. (10) exhibited significant inhibitory activity against 12-0-tetradecanoylphorbol 13-acetate-induced Epstein-Barr virus early antigen activation in Raji cells.

Chemical constituents of Murraya siamensis: three coumarins and their anti-tumor promoting effect.

Isolation and structure elucidation of three coumarins, murrayacoumarins A, B, and C, together with eight known coumarins, from the leaves of Murraya siamensis Craib collected in Thailand are described. Results of a primary screening of inhibitory effects of seven of these compounds on 12-O-tetradecanoylphorbol-13-acetate-induced Epstein-Barr virus early antigen activation in Raji cells are also presented.