Cancer preventive agents. Part 1: chemopreventive potential of cimigenol, cimigenol-3,15-dione, and related compounds.

In continuation of our previous report, cimigenol (1) and 15 related compounds were screened as potential antitumor promoters by using the in vitro short-term 12-O-tetradecanoylphorbol-13-acetate (TPA)--induced Epstein-Barr virus early antigen (EBV-EA) activation assay. Cimigenol-3,15-dione (2) displayed the greatest potency (100% inhibition at 1000 mol ratio/TPA) and consequently was further examined for antitumor-promoting activity in a two-stage carcinogenesis assay of mouse skin tumors (DMBA/TPA). In this assay, compound 2 showed significant activity, reducing the number of papillomas per mouse to 48% of the control group at 20 weeks. In addition, compounds 1 and 2 were examined for antitumor-initiating activity in a two-stage carcinogenesis assay of mouse skin tumors induced by peroxynitrite as an initiator and TPA as a promoter. Results showed that these two triterpenoids were almost equipotent with epigallocatechin gallate (EGCG) and slightly more potent than tocinol (group V), the positive controls. Thus, compounds 1 and 2 exhibited not only strong antitumor-promoting activity but also significant antitumor-initiating effect on mouse skin. These data suggest that both compounds might be valuable chemopreventors.

Multidrug-resistant cancer cell susceptibility to cytotoxic quassinoids, and cancer chemopreventive effects of quassinoids and canthin alkaloids.

Twenty-three quassinoids (1-23), which were isolated previously from Simaroubaceous plants, were evaluated for cytotoxicity against three multidrug-resistant cancer cell lines, KB-VIN, KB-7d, and KB-CPT. Nine compounds (2-7 and 9-11) showed significant cytotoxicity in all three cell lines. Compounds 1, 12-14, 17, and 20 demonstrated significant activity against the KB-7d and KB-CPT cell lines, and compounds 18, 19, and 23 revealed notable activity only against KB-7d cells. Structure-activity relationships were drawn based on these data. In addition, six quassinoid derivatives (24-29) and four canthin alkaloids (30-33), which were isolated from Brucea antidysenterica, were examined for their inhibitory effects on 12-O-tetradecanoylphorbol-13-acetate (TPA) induced Epstein-Barr virus early antigen (EBV-EA) activation as cancer chemopreventive agents. All of these compounds demonstrated significant inhibitory effects against EBV-EA activation.

Chemical constituents of Glycosmis arborea: three new carbazole alkaloids and their biological activity.

As part of the phytochemical studies of the plant genus Glycosmis, the constituents of the plant Glycosmis arborea were investigated. Three new carbazole alkaloids named glybomines A (1), B (2), and C (3), along with known monomeric alkaloids belonging to the carbazole, quinazoline, furoquinoline, quinolone, and acridone classes, were isolated from stems of the plant collected at Mymensing in Dhaka, Bangladesh. Glybomine A (1) is the first example of a 2,5-oxygenated carbazole alkaloid from natural sources. As a primary screening test for anti-tumor promoters, nine alkaloids isolated from this plant have been tested for their inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) induction by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). All alkaloids tested showed inhibitory activity.

Cancer chemopreventive effect of phenothiazines and related tri-heterocyclic analogues in the 12-O-tetradecanoylphorbol-13-acetate promoted Epstein-Barr virus early antigen activation and the mouse skin two-stage carcinogenesis models.

In continuation of our search for novel agents, we have investigated 29 phenothiazines and related tri-heterocyclic compounds as potential cancer chemopreventive agents in a short-term in vitro assay of Epstein-Barr virus early antigen (EBV-EA) activation induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Among the evaluated compounds, chlorpromazine, phenoxazine, ethylpropazine, 9-oxo-9H-thioxanthene-3-carbonitrile-10,10-dioxide, thiothixene and phenothiazine showed profound inhibition of EBV-EA in the in vitro assay. This activity was influenced by a modification of the phenothiazine ring. Replacement of nitrogen in the phenothiazine ring with sulfur atoms decreased the anti-tumor activity. Overall analysis showed that the simple tri-cyclic compound phenoxazine was the most active anti-tumor promoting compound in the test system. Therefore, we assessed the anti-tumor promoting effect of phenoxazine in vivo in two different chemical carcinogen-induced-promotion experimental models in mice namely the 7,12-dimethylbenz(a)anthracene (DMBA) initiated and TPA-promoted ICR mouse skin two-stage carcinogenesis protocol and the peroxynitrite (PN)-induced and TPA-promoted skin carcinogenesis in HOS:HR-1 mouse. Following tumor initiation with DMBA, topical application of 0.0025% phenoxazine to the dorsal initiated mouse skin resulted in a highly significant inhibition of TPA tumor promotion. The compound exhibited remarkable inhibitory effects on the mouse skin tumor promotion in terms of a reduction in tumor multiplicity (>50%) and incidence, accompanied by an extension of the tumor latency. In the PN-induced and TPA-promoted two-stage mouse skin carcinogenesis, oral administration of phenoxazine (0.0025%) for 2 weeks showed profound decrease in both the tumor incidence and burden by more than 20 and 80%, respectively, at 10 weeks of treatment. This was also accompanied by a 20% delay in the tumor latency period. In all the treatment groups, there was no toxicity due to phenoxazine in the treatment groups as compared to the control animals. These significant anti-tumor potentials of phenoxazine either via topical or oral administration might be due to the inherent cytotoxicity of these classes of compounds, which can be utilized in the prevention of development of overt tumors, immunopotentiation, induction of differentiation and apoptosis. In addition, since phenoxazine derivatives and other related phenothiazine compounds in use, as anti-psychotic agents without any reported adverse effect are known to pass the blood-brain barrier, they represent a new class of cancer chemopreventive agents with greater implication in the prevention of brain cancers.

Tumor chemopreventive activity of 3-O-acylated (--)-epigallocatechins.

In order to seek promising cancer chemopreventive agents, we assessed the antitumor promoting activities of 3-O-octanoyl- or 3-O-(2-methyloctanoyl)-(--)-epigallocatechins, inhibiting markedly the activation of Epstein-Barr virus early antigen, in a two-stage mouse skin carcinogenesis assay. As a result, these derivatives inhibited a papilloma formation 1.3-1.6-fold more strongly than (--)-epigallocatechin gallate well established as anti-tumor promoter.

Isoflavonoids from Dalbergia olivari.

Two isoflavonoids, named olibergin A (1) and B (2) were isolated from the stem bark of Dalbergia oliveri (Leguminosae). Along with three previously known compounds, they are inhibitors of Epstein-Barr virus early antigen activation induced by 12-O-tetradecanoylphorbol-13-acetate in Raji cells. Their structures were elucidated on the basis of spectroscopic analyses.

Chalcones, coumarins, and flavanones from the exudate of Angelica keiskei and their chemopreventive effects.

From an ethyl acetate-soluble fraction of the exudate obtained from the stems of Angelica keiskei (Umbelliferae), 17 compounds, viz. five chalcones (1-5), seven coumarins (6-12), three flavanones (13-15), one diacetylene (16), and one 5-alkylresorcinol (17), were isolated. These compounds were evaluated with respect to their inhibitory effects on the induction of Epstein-Barr virus early antigen (EBV-EA) by 12-O-tetradecanoylphorbol-13-acetate (TPA) in Raji cells, which is known to be a primary screening test for antitumor-promoters. With the exception of three compounds (10, 16, and 17), all other compounds tested showed potent inhibitory effects on EBV-EA induction (92-100% inhibition at 1x10(3)mol ratio/TPA). In addition, upon evaluation of these compounds for the inhibitory effects against activation of (+/-)-(E)-methyl-2-[(E)-hydroxy-imino]-5-nitro-6-methoxy-3-hexemide (NOR 1), a nitric oxide (NO) donor, as a primary screening test for antitumor-initiators, two chalcones (2 and 3) and six coumarins (6-11) exhibited potent inhibitory effects.

Sterol ferulates, sterols, and 5-alk(en)ylresorcinols from wheat, rye, and corn bran oils and their inhibitory effects on Epstein-Barr virus activation.

Sterol ferulate, free sterol, and 5-alk(en)ylresorcinol constituents of wheat, rye, and corn bran oils were studied. Among the sterol ferulates, one novel compound, 24-methylenecholestanol ferulate (7), along with six known compounds, namely, 24-methylcholestanol ferulate (1), 24-methylcholesterol ferulate (2), 2-methyllathosterol ferulate (3), stigmastanol ferulate (4), sitosterol ferulate (5), and schottenol ferulate (6), were isolated and characterized. Five known free sterols, namely, 24-methylcholesterol (8), stigmastanol (9), sitosterol (10), schottenol (11), and stigmasterol (12), were isolated and identified. 5-Alk(en)ylresorcinols were found in wheat and rye bran oils but not in corn bran oil. Of these, one new compound, 5-n-(2'-oxo-14'-Z-heneicosenyl) resorcinol (19), and seven known compounds, namely, 5-n-heptadecyl- (13), 5-n-nonadecyl- (14), 5-n-heneicosyl- (15), 5-n-tricosyl- (16), 5-n-pentacosyl- (17), 5-n-(14'-Z-nonadecenyl)- (18), and 5-n-(2'-oxoheneicosyl)resorcinols (20), were isolated and characterized. These compounds were evaluated with respect to their inhibitory effects on the induction of Epstein-Barr virus early antigen (EBV-EA) by 12-O-tetradecanoylphorbol-13-acetate (TPA) in Raji cells, which is known to be a primary screening test for antitumor promoters. Four compounds, 1, 2, 4, and 11, showed potent inhibitory effects on EBV-EA induction.

Ichthyotoxic and anticarcinogenic effects of triterpenoids from Sandoricum koetjape bark.

After bioassay-guided fractionation of the extract from Sandoricum koetjape bark, which exhibited significant toxicity to killifish (Oryzias latipes), two ichthyotoxic triterpenoids were isolated and characterized as koetjapic acid and 3-oxo-olean-12-en-29-oic acid. These constituents, along with non-toxic katonic acid, had a remarkable inhibitory effect on Epstein-Barr virus early antigen (EBV-EA) activation induced by 12-O-tetradecanoylphorbol 13-acetate (TPA), which is a preliminary in vitro screening method for possible anti-tumor-promoting agents. Of the triterpenoids active in vitro, koetjapic acid appears to be a promising cancer chemopreventive agent, since it significantly delayed tumor promotion in two-stage mouse skin carcinogenesis induced by 7,12-dimethylbenz(a)anthracene and promoted by TPA.

Absolute configuration of sesquiterpenes from Crossopetalum tonduzii and their inhibitory effects on Epstein-Barr virus early antigen activation in Raji cells.

Two new sesquiterpenoids (1 and 2) with a dihydro-beta-agarofuran skeleton were isolated from Crossopetalum tonduzii. Their structures were elucidated on the basis of spectral analysis, including homonuclear and heteronuclear correlation NMR experiments (COSY, ROESY, HSQC, and HMBC). Their absolute configurations were determined by CD studies on 3, the benzoylated derivative of 1. Chemical correlations have allowed the absolute configurations of 4 and 5, two previously known dihydro-beta-agarofuran analogues, to be reported for the first time. Compounds 1, 2, and 5 showed strong antitumor-promoting effects on Epstein-Barr virus early antigen (EBV-EA) activation.

Polyprenylated benzophenones from Garcinia assigu and their potential cancer chemopreventive activities.

In a further study on the chemical constituents of Garcinia assigu, two new benzophenones corresponding to the 13-O-methyl ethers (1 and 2) of the known isogarcinol and garcinol, respectively, were isolated and characterized, along with known benzophenones (3-6). Inhibitory effects of the benzophenones isolated from this plant on Epstein-Barr virus early antigen (EBV-EA) activation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in Raji cells and their radical-scavenging ability against 1,1-diphenyl-2-picrylhydrazyl (DPPH) were demonstrated. The cyclized polyprenylbenzophenones (1-5) showed comparable or stronger potential cancer chemopreventive activity when compared to glycyrrhetic acid, a known anti-tumor promoter.

Anti-tumor promoting effect of glycosides from Prunus persica seeds.

Four minor components, along with the major cyanogenic glycosides, amygdalin and prunasin, were isolated from Prunus persica seeds (Persicae Semen; Tounin), and characterized as mandelic acid glycosides (beta-gentiobioside and beta-D-glucoside) and benzyl alcohol glycosides (beta-gentiobioside and beta-D-glucoside). The anti-tumor promoting activity of these compounds was examined in both in vitro and in vivo assays. All of the compounds significantly inhibited the Epstein-Barr virus early antigen activation induced by tumor promoter. In addition, they produced a delay of two-stage carcinogenesis on mouse skin that was comparable in potency to (-)-epigallocatechin gallate from green tea. Structure-activity relationships indicated that a substituent at the benzylic position with glycosidic linkage affected the in vitro and in vivo activities with an order of enhancing potency, CN

Inhibitory effects of lapachol derivatives on epstein-barr virus activation.

Sixteen derivatives (2-17) synthesized from the naphthoquinone lapachol (1), were tested for their inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) activation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA), as a test for potential cancer chemopreventive agents. They exhibited a variety of inhibitory activities from very high to moderate, which allow us to suggest structure-activity relationships. Ten of these derivatives are reported for the first time, their structures being thoroughly determined by spectroscopic methods.

Inhibitory effects of cucurbitane glycosides and other triterpenoids from the fruit of Momordica grosvenori on epstein-barr virus early antigen induced by tumor promoter 12-O-tetradecanoylphorbol-13-acetate.

Two new triterpene benzoates, 5-dehydrokarounidiol dibenzoate (1) and karounidiol dibenzoate (2), and two new triterpene glycosides, 5alpha,6alpha-epoxymogroside IE(1) (8) and 11-oxomogroside A(1) (9), along with 15 known triterpenoids (one triterpene benzoate, 3; three triterpene mono-ols, 4-6; one triterpene aglycon, 7; and 10 triterpene glycosides, 10-19), were isolated from the ethanol extract of the fruit of Momordica grosvenori. The structures of 1, 2, 8, and 9 were determined on the basis of spectroscopic and chemical methods. Among the known triterpene glycosides, mogroside I E(1) (12) was a new naturally occurring compound. Eighteen triterpenoids (2-19) and 11-oxomogrol (20), a hydrolysis product of 9, were evaluated with respect to their inhibitory effects on the induction of Epstein-Barr virus early antigen (EBV-EA) by 12-O-tetradecanoylphorbol-13-acetate (TPA) in Raji cells, which is known to be a primary screening test for antitumor promoters. All of the compounds tested showed potent inhibitory effects on EBV-EA induction (70-100% inhibition at 1 x 10(3) mol ratio/TPA).

Interiotherins C and D, two new lignans from Kadsura interior and antitumor-promoting effects of related neolignans on Epstein-Barr virus activation.

Two new lignans, interiotherins C (1) and D (2), together with the known compounds interiorin (3), heteroclitin F (4), neokadsuranin (5), heteroclitin D (6), kadsurin (7), gomisin A (8), schisandrin C (9), interiotherin A (10), angeloylgomisin R (11), gomisin G (12), interiotherin B (13), and gomisin C (14), were isolated from the stems of Kadsura interior. The structures and stereochemistries of the new compounds were determined from mass, CD, and NMR spectral data. Fourteen neolignans were screened as potential antitumor promoters by examining their ability to inhibit Epstein-Barr virus early antigen (EBV-EA) activation (induced by 12-O-tetradecanoylphorbol-13-acetate) in Raji cells. Neokadsuranin (5) and schisandrin C (9) were the most potent compounds. These data suggest that some neolignans might be valuable antitumor promoters or chemopreventors.

A newly established neuronal rho-0 cell line highly susceptible to oxidative stress accumulates iron and other metals. Relevance to the origin of metal ion deposits in brains with neurodegenerative disorders.

From human neuroblastoma-derived SILA cells we have established a rho-0 cell line that is deficient in both respiration and mitochondrial DNA. Lactate dehydrogenase activity, lactate production, and growth in the medium without glucose indicate that these cells shift from aerobic to anaerobic metabolism. Electron microscopic observations revealed abnormal mitochondria with unique cristae structures. Staining with MitoTracker dye showed that the mitochondrial transmembrane potential was reduced by 30-40% from the parent cell levels. These cells were markedly susceptible to H(2)O(2) and died apparently by a necrotic mechanism, a process blocked by deferoxamine in the parent cells but not rho-0 cells. Analysis by inductively coupled plasma-mass spectrometry revealed an approximately 3-fold accumulation of iron in the rho-0 cells at confluence (n = 4-6, three clones, *p < 0.05). Iron and four other metals were all elevated in the cells of one of the rho-0 clones and were similar to control levels in the control cybrid cells, which were replenished with normal mitochondrial DNA. Their sensitivity to H(2)O(2) was also similar to that of the parent cells. These results indicate that a newly established neuronal related rho-0 cell line is highly susceptible to active oxygen species and that these toxicity effects appear to be related to an accumulation of transition metals, which probably occurs through the respiratory impairment.

Chemopreventive effect of resveratrol, sesamol, sesame oil and sunflower oil in the Epstein-Barr virus early antigen activation assay and the mouse skin two-stage carcinogenesis.

Resveratrol, sesamol, sesame oil and sunflower oil are known natural dietary components with intrinsic cancer chemopreventive potentials. As a part of our study of dietary constituents as potential cancer chemopreventive agents, we have assessed the anti-cancer potentials of these products in the promotion stage of cancer development employing the in vitro Epstein-Barr virus early antigen activation assay induced by the tumor promoter 12-O-tetradecanoylphorbol 13-acetate (TPA). Further, we studied the activities of these compounds in the brine shrimp cytotoxicity assay as well as on the stable 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging bioassay with a view to comparing some of the mechanisms of their anti-cancer activity. Finally, we compared the observed chemoprotective capabilities of the four products in the in vivo 7,12 dimethylbenz(a)anthracene initiated and TPA-promoted mouse skin two-stage carcinogenesis protocols. All the products tested showed a profound inhibitory effect on the Epstein-Barr virus early antigen induction using Raji cells. Comparatively, sesame oil was the most potent followed by sesamol and then resveratrol. Only sesamol and resveratrol showed a remarkable cytotoxic activity in the brine shrimp lethality assays as well as profound free radical scavenging activity in the DPPH bioassay. In both test systems, sesamol exhibited a more remarkable activity than resveratrol while sesame oil and sunflower oil did not exhibit any appreciable activity even at the highest concentrations tested (4000 microg ml(-1) ). In our in vivo assay at a 50-fold molar ratio to TPA, sesamol offered 50% reduction in mouse skin papillomas at 20 weeks after promotion with TPA. Under an identical molar ratio to TPA, resveratrol offered a 60% reduction in the papillomas in mouse at 20 weeks. Thus sesamol seems to be an almost equally potent chemopreventive agent. Sesame oil and sunflower oil offered 20 and 40% protection, respectively, in the mouse skin tumor model. The anti-oxidant capabilities of these compounds could not solely explain the observed anti-cancer characteristics. Resveratrol is present in grapes. Sesamol, a constituent of sesame oil and sunflower oil are regularly consumed dietary natural products. The observed chemopreventive effect of these products particularly warrants more attention since they already exist in the population with no known adverse effects.

Cancer chemopreventive effect of quassinoid derivatives. Introduction of side chain to shinjulactone C for enhancement of inhibitory effect on Epstein-Barr virus activation.

A series of shinjulactone C (1) derivatives (2-8) were synthesized and evaluated for their anti-tumor promoting effects against Epstein-Barr virus early antigen activation introduced by 12-O-tetradecanoylphorbol-13-acetate in Raji cells. The succinate and 3',3'-dimethylsuccinate derivatives of 1 exhibited higher inhibitory effects than 1. From the point of view of structure-activity relationships, the succinate derivatives (2, 4) demonstrated better potency than the glutarate derivatives (3, 5-8). As substituted moieties of 3'-position became bulky, the inhibitory effects of the glutarate derivatives (7, 8) significantly decreased.

Chemopreventive effects of emodin and cassiamin B in mouse skin carcinogenesis.

In continuation of our works of natural and synthetic products as cancer chemopreventive agents, we have examined emodin and cassiamin B, which were isolated from Cassia siamea. These compounds exhibited the remarkable anti-tumor promoting effect on two-stage carcinogenesis test of mouse skin tumors induced by 7,12-dimethylbenz[a]anthracene as an initiator and 12-O-tetradecanoylphorbol-13-acetate (TPA) as a promoter by both topical application. Furthermore, emodin exhibited potent inhibitory activity on two-stage carcinogenesis test of mouse skin tumors induced by nitric oxide donor, (+/-)-(E)-methyl-2-[(E)-hydroxyimino]-5-nitro-6-methoxy-3-hexeneamide as an initiator and TPA as a promoter.

Cancer chemopreventive effects of a Brazilian folk medicine, Juca, on in vivo two-stage skin carcinogenesis.

Gallic acid (1) and methyl gallate (2) were isolated from Juca, a Brazilian folk medicine, fruits of Caesalpinia ferrea MART (Leguminosae), decreased significantly the average number of papillomas per mouse in the experiment of the promoting effects of 12-O-tetra- decanoylphorbol-13-acetate (TPA) on skin tumor formation in mice initiated with 7,12-dimethylbenz[a]anthracene (DMBA).