Anticancer and multidrug resistance-reversal effects of solanidine analogs synthetized from pregnadienolone acetate.

A set of solanidine analogs with antiproliferative properties were recently synthetized from pregnadienolone acetate, which occurs in Nature. The aim of the present study was an in vitro characterization of their antiproliferative action and an investigation of their multidrug resistance-reversal activity on cancer cells. Six of the compounds elicited the accumulation of a hypodiploid population of HeLa cells, indicating their apoptosis-inducing character, and another one caused cell cycle arrest at the G2/M phase. The most effective agents inhibited the activity of topoisomerase I, as evidenced by plasmid supercoil relaxation assays. One of the most potent analogs down-regulated the expression of cell-cycle related genes at the mRNA level, including tumor necrosis factor alpha and S-phase kinase-associated protein 2, and induced growth arrest and DNA damage protein 45 alpha. Some of the investigated compounds inhibited the ABCB1 transporter and caused rhodamine-123 accumulation in murine lymphoma cells transfected by human MDR1 gene, expressing the efflux pump (L5178). One of the most active agents in this aspect potentiated the antiproliferative action of doxorubicin without substantial intrinsic cytostatic capacity. The current results indicate that the modified solanidine skeleton is a suitable substrate for the rational design and synthesis of further innovative drug candidates with anticancer activities.

Bioactivity-guided isolation of cytotoxic sesquiterpenes and flavonoids from Anthemis ruthenica.

A new eudesmanolide sesquiterpene, sivasinolide 6-O-angelate (1), was isolated from the aerial parts of Anthemis ruthenica together with the known compounds chrysanin (2), tanacin (3), 3 beta-hydroxycostunolide (4), centauridin (5), and centaureidin (6). The compounds were obtained by means of bioactivity-guided fractionation from the CHCl (3) extract of the herb, which displayed high cytotoxic activity. The structures were determined by UV, HR-ESI-MS, and high-field 1D and 2D NMR spectral analyses, affording complete (1)H- and (13)C-NMR assignments for all compounds. The cytotoxic activities of the isolated sesquiterpenes and flavonoids were assessed against cervical adenocarcinoma HeLa, breast adenocarcinoma MCF7, and skin epidermoid carcinoma A431 cells using the MTT assay. It was found that, apart from centaureidin (6), which is extremely active (IC(50) 0.082, 0.13, and 0.35 microM on the HeLa, MCF7, and A431 cell lines, respectively), all these compounds exert high or moderate tumor cell-growth inhibitory activity (IC(50) 3.42-58.15 microM).

Xanthanolides with antitumour activity from Xanthium italicum.

Bioassay-guided fractionation of a CHCl3 extract of the leaves of Xanthium italicum Moretti led to the isolation of four xanthanolides: xanthatin (1), 4-epixanthanol (2), 4-epi-isoxanthanol (3), and 2-hydroxyxanthinosin (4). Their structures were determined by means of 1D and 2D NMR spectroscopy, including 1H-1H COSY, NOESY, HSQC and HMBC experiments, which resulted in complete and unambiguous 1H and 13C NMR chemical shift assignments. The isolated compounds 1-4 were evaluated for their antiproliferative activities, and were demonstrated to exert significant cell growth inhibitory activity against human cervix adenocarcinoma (HeLa), skin carcinoma (A431), and breast adenocarcinoma (MCF7) cells.

Antitumor activity of alkaloids derived from Amaryllidaceae species.

The aim of the present study was to investigate the anticancer properties of five alkaloids isolated from Amaryllidaceae, including the inhibitory effect on P-glycoprotein (P-gp) and the apoptosis-inducing capacity. The tested alkaloids were evaluated for their multidrug resistance (MDR)-reversing activity on human MDR1-gene-transfected L5178 mouse lymphoma cells, using the rhodamine-123 (Rh-123) assay. Trisphaeridine and pretazettine increased the intracellular Rh-123 concentration 30- and 50-fold, respectively, as compared to the non-treated cells, and 2-O-acetyllycorine and trisphaeridine were demonstrated by means of the checkerboard method to enhance the antiproliferative activity of doxorubicin on L5178 MDR mouse lymphoma cells. The MTT assay revealed that pretazettine, trisphaeridine and 2-O-acetyllycorine displayed excellent antiproliferative effects on both the human and the mouse cell lines. The apoptosis-inducing activities of selected agents (2-O-acetyllycorine and trisphaeridine) were measured via acridine orange and ethidium bromide dual staining and flow cytometry of the subG1 population.

Efficient approach to androstene-fused arylpyrazolines as potent antiproliferative agents. Experimental and theoretical studies of substituent effects on BF(3)-catalyzed intramolecular [3 + 2] cycloadditions of olefinic phenylhydrazones.

Highly diastereoselective Lewis acid induced intramolecular 1,3-dipolar cycloadditions of alkenyl phenylhydrazones (containing various substituents on the aromatic ring) obtained from a d-secopregnene aldehyde were carried out under fairly mild conditions to furnish androst-5-ene-fused arylpyrazolines in good to excellent yields. The ability of phenylhydrazones to undergo cyclization was found to be affected significantly by the electronic features of the substituents on the aromatic moiety. The rates of the ring-closure reactions were observed to be increased by electron-donating and decreased by electron-withdrawing groups. The experimental findings on the BF(3)-catalyzed transformations were supported by calculations of the proposed mechanism at the BLYP/6-31G(d) level of theory, indicating a noteworthy dependence, mainly of the initial complexation step, and hence of the whole process, on the character of the substituent. The cycloaddition was estimated to occur via a zwitterionic intermediate rather than involving a pure concerted mechanism. The antiproliferative activities of the structurally related pyrazoline derivatives were tested in vitro on three malignant human cell lines (HeLa, MCF7, and A431): the microculture tetrazolium assay revealed that several compounds exerted marked cell growth-inhibitory effects. The highest cytotoxic activities, displayed by the p-methoxyphenylpyrazoline derivative 7d (IC(50) values: 2.01, 2.16, and 1.41 microM on HeLa, MCF7, and A341 cells, respectively), were better than those of cisplatin (IC(50) values: 12.43, 9.63, and 2.84 microM, respectively).

Antiproliferative activity of Hungarian Asteraceae species against human cancer cell lines. Part II.

The antiproliferative activities of aqueous and organic extracts prepared from 26 Hungarian species of the tribes Cynereae and Lactuceae (Asteraceae) were tested in vitro against HeLa (cervix epithelial adenocarcinoma), A431 (skin epidermoid carcinoma) and MCF7 (breast epithelial adenocarcinoma) cells by using the MTT assay. Of the tested 200 extracts of different plant parts obtained with n-hexane, chloroform, 50% methanol and water, 16 extracts displayed noteworthy cell growth inhibitory activity (>50% inhibition at a concentration of 10 microg/mL). The IC50 values of these extracts were determined, and their direct cytotoxic effects were measured. High differences between the antiproliferative and cytotoxic activities, demonstrating a real cell proliferation inhibitory activity rather than direct killing effects, were found for some Centaurea, Cirsium, Cichorium, Lactuca, Onopordum and Scorsonera extracts.

Biological activity of bis-benzimidazole derivatives on DNA topoisomerase I and HeLa, MCF7 and A431 cells.

Benzimidazoles of both natural and synthetic sources are the key components of many bio-active compounds. Several reports have shown antifungal, antiviral, H(2) receptor blocker and antitumor activities for benzimidazoles and their derivatives. In this study, we synthesized twelve bis-benzimidazole derivatives by selecting di(1H-benzo[d]imidazol-2-yl)methane as the main compound. The numbers of carbons at 2 positions of bis-benzimidazole derivatives were changed from 1 to 4, and derivatives were synthesized with methyl substitutions at 5- and/or 6- positions. The compounds were screened via in vitro plasmid superciol relaxation assays using mammalian DNA topoisomerase I and cytostatic assays were carried out against HeLa (cervix adenocarcinoma), MCF7 (breast adenocarcinoma) and A431 (skin epidermoid carcinoma) cells for selected derivatives. Our results suggest that the malonic acid derivatives of bis-benzimidazoles, namely, bis(5-methyl-1H-benzo[d]imidazol-2-yl)methane and bis(5,6-dimethyl-1H-benzo[d]imidazol-2-yl)methane, were remarkably active compounds in interfering with DNA topoisomerase I and the former compound was also found to be cytotoxic against MCF7 and A431 cells. The inhibitory effects obtained with these derivatives are significant as these compounds can be potential sources of anticancer agents.

Antiproliferative effect of flavonoids and sesquiterpenoids from Achillea millefolium s.l. on cultured human tumour cell lines.

The antiproliferative activities of n-hexane, chloroform, aqueous-methanol and aqueous extracts of the aerial parts of the Achillea millefolium aggregate on three human tumour cell lines were investigated by means of MTT assays. The chloroform-soluble extract exerted high tumour cell proliferation inhibitory activities on HeLa and MCF-7 cells, and a moderate effect on A431 cells; accordingly, it was subjected to detailed bioactivity-guided fractionation. As a result of the multistep chromatographic purifications (VLC, CPC, PLC, gel filtration), five flavonoids (apigenin, luteolin, centaureidin, casticin and artemetin) and five sesquiterpenoids (paulitin, isopaulitin, psilostachyin C, desacetylmatricarin and sintenin) were isolated and identified by spectroscopic methods. The antiproliferative assay demonstrated that centaureidin is the most effective constituent of the aerial parts of yarrow: high cell growth inhibitory activities were observed especially on HeLa (IC(50) 0.0819 microm) and MCF-7 (IC(50) 0.1250 microm) cells. Casticin and paulitin were also highly effective against all three tumour cell lines (IC(50) 1.286-4.76 microm), while apigenin, luteolin and isopaulitin proved to be moderately active (IC(50) 6.95-32.88 microm). Artemetin, psilostachyin C, desacetylmatricarin and sintenin did not display antiproliferative effects against these cell lines. This is the first report on the occurrence of seco-pseudoguaianolides (paulitin, isopaulitin and psilostachyin C) in the Achillea genus.

Synthesis and biological activity evaluation of 1H-benzimidazoles via mammalian DNA topoisomerase I and cytostaticity assays.

Benzimidazoles are important compounds because of their antibacterial, antifungal, antimicrobial, antiprotozoal and antihelmintic activities. Some benzimidazole derivatives also interfere with the reactions of DNA topoisomerases, enzymes functioning at almost all stages of the cell cycle. In this study, nine 1H-benzimidazole derivatives with substituents at positions 2 and 5 were synthesized and the structure of the compounds was elucidated by instrumental methods. The characterized compounds were screened to identify if they interfered with mammalian type I DNA topoisomerase activity via in vitro supercoil relaxation assays. Selected compounds were subjected to cytostatic assays using HeLa (cervix adenocarcinoma), MCF7 (breast adenocarcinoma) and A431 (skin epidermoid carcinoma) cells. Our results showed that 5-chloro-2-(2-hydroxyphenyl)-1H-benzimidazole exerted the most profound topoisomerase I inhibition and cytotoxicity.

[Antioxidant activity of the fruits and hydrophilic compounds of Physalis alkekengi]. / A Physalis alkekengi termésének és hidrofil anyagainak antioxidáns hatása.

Physalis alkekengi L. (bladder cherry, Chinese lantern, winter cherry) is an unusual species of the family Solanaceae. Although accumulation of alkaloids is characteristic to Solanaceae species, and accordingly the root and above ground parts of P. alkekengi are toxic, its fruits are in exceptionally edible. The present paper deals with the investigation of antioxidant hydrophilic compounds of the fruits in order to find correlation between the quantity of the constituents and antioxidant capacity of the extracts. Dried and fresh, freeze stored fruits were extracted with water, and the ascorbic acid and total polyphenol content of the fruits was determined. Furthermore, the antioxidant effect was investigated by DPPH test, and in vitro using the rat-brain homogenate method. The antioxidant activity measured by DPPH (fresh fruit: IC50 = 2.48 mg/ml; dried fruit: IC50 = 22.32 mg/ml) showed good correlation with the ascorbic acid content of the fruit (fresh fruit: 1.095%; dried fruit: 0.162%), and exhibited substantial decrease due the drying process. Lipid peroxidation inhibitory activity was found to be weaker as the DPPH radical scavenger capacity, however, also showed a decrease during the drying process of the fruit (fresh fruit: IC50 = 6.43 mg/ml; dried fruit: IC50 = 15.59 mg/ml). Our results clearly demonstrated the radical scavenger and lipid peroxidation inhibitory activity of aqueous extracts of bladder cherry, and indicate that the conservation and processing technology significantly influenced the antioxidant activity and the content of the active ingredients.

Antitumour properties of acridone alkaloids on a murine lymphoma cell line.

The aim of the present study was to investigate the anticancer properties of a set of furanoacridone alkaloids, arborinine and evoxanthine, including the inhibitory effect of P-glycoprotein (Pgp) and the apoptosis-inducing capacity. The tested alkaloids were evaluated for multidrug resistance (MDR)-reversing activity on human Pgp-transfected L5178 mouse lymphoma cells, using the rhodamine-123 (Rh-123) assay. The antiproliferative effects of natural compounds and their interactions with doxorubicin were determined in MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. Apoptosis-inducing activity was additionally measured by means of dual annexin V and propidium iodide staining. RT-PCR was used to test the expression of Pgp mRNA after acridone treatment. All of the acridones investigated increased the accumulation of Rh-123. Gravacridonetriol and gravacridonediol monomethyl ether increased the antiproliferative effect of doxorubicin on resistant L5178 cells. Treatment with these agents resulted in a decrease in Pgp mRNA levels. Naturally occurring acridone alkaloids exhibit a beneficial combination of anticancer effects and, accordingly, the acridone skeleton can be considered useful in the design of novel antiproliferative agents.

Antiproliferative activity of Hungarian Asteraceae species against human cancer cell lines. Part I.

Aqueous and organic extracts of 25 selected species from four tribes of Hungarian Asteraceae were screened in vitro for antiproliferative activity against HeLa (cervix epithelial adenocarcinoma), A431 (skin epidermoid carcinoma) and MCF7 (breast epithelial adenocarcinoma) cells, using the MTT assay. Twenty five of the 228 tested extracts from different parts of the species of Astereae (6), Inuleae (3), Heliantheae (5) and Anthemideae (11) demonstrated a substantial antiproliferative effect (at least 50% inhibition of cell proliferation) at 10 microg/mL against one or more of the cell lines. Complete dose-response curves were generated and IC(50) values were calculated for these active extracts, and their direct cytotoxic effects were determined. In summary, 11 of the tested 25 plants were found to be active and 4 of them (Anthemis ruthenica, Erigeron canadensis, Erigeron annuus and Inula ensifolia) had not been studied previously for either active compounds or anticancer properties.

Phenanthrenes and a dihydrophenanthrene from Tamus communis and their cytotoxic activity.

From the petroleum ether extract of the rhizomes of Tamus communis, the 7-hydroxy-2,3,4,8-tetramethoxyphenanthrene (1) was isolated, together with the known 2,3,4-trimethoxy-7,8-methylenedioxyphenanthrene (2), 3-hydroxy-2,4,-dimethoxy-7,8-methylenedioxyphenanthrene (3), 2-hydroxy-3,5,7-trimethoxyphenanthrene (4) and 2-hydroxy-3,5,7-trimethoxy-9,10-dihydrophenanthrene (5), through cytotoxic assay guidance. The structures were determined by means of HREIMS, (1)H NMR, JMOD and NOESY experiments. The cytotoxic effects of the isolated compounds were tested on cervix adenocarcinoma (HeLa) cells, with the MTT assay. The results demonstrated that, with the exception of 2, all these compounds displayed pronounced cytotoxic activity; especially 1 and 3 exhibited significant cell growth inhibitory effects, with IC(50)=8.52+/-0.70 and 3.64+/-0.12 microM, respectively.

Investigation of cytotoxic activity on human cancer cell lines of arborinine and furanoacridones isolated from Ruta graveolens.

The cytotoxic effects of a series of furanoacridones isolated from Ruta graveolens L. (Rutaceae) and of two further acridone alkaloids (arborinine and evoxanthine) were investigated by means of the MTT assay, using the human cell lines HeLa, MCF7 and A431. Arborinine proved best in inhibiting the proliferation of all three cell lines. The cytotoxic potency of the furacridone alkaloids was a function of their lipid solubility, which was determined by means of PAMPA. The capacity of the most effective furanoacridones to induce apoptosis was demonstrated by flow cytometric cell cycle analysis and by staining with ethidium bromide and acridine orange. This finding was reinforced by determining the apoptosis-regulating factors Bcl-2 and Bax, which were revealed by means of RT-PCR to change dose-dependently. The data presented here indicate that naturally occurring furanoacridones can be regarded as excellent starting structures for the potential development of new anticancer agents.

Isobrassinin and its analogues: novel types of antiproliferative agents.

Isobrassinin (2-(S-methyldithiocarbamoylaminomethyl)indole (7a), a regioisomer of the cruciferous phytoalexin brassinin (1), exerted marked antiproliferative effects on the HeLa, A431 and MCF7 cell lines (>78.6% inhibition at 30muM). For structure-activity relationships, further analogues were synthesized. The highest cytotoxic effect was displayed by 2-phenylimino-1,3-thiazino[5,6-b]indole (10) (10 microM, 76.8%-HeLa and 46.3%-MCF7). The effect of the natural phytoalexin brassinin was also determined.

Cytotoxic phenanthrenes from the rhizomes of Tamus communis.

From the fresh rhizomes of Tamus communis five phenanthrenes (1 - 5) were isolated under the guidance of cytotoxic assays in HeLa cells. The compounds were obtained from the highly active CHCl (3) fraction of the MeOH extract by using multistep chromatographic purifications, including VLC, preparative TLC, HPLC and gel filtration. The compounds were identified by means of EI-mass, UV and NMR spectroscopy as 7-hydroxy-2,3,4-trimethoxyphenanthrene (1), 2,7-dihydroxy-3,4-dimethoxyphenanthrene (nudol) (2), 2,7-dihydroxy-3,4,8-trimethoxyphenanthrene (3), 3,7-dihydroxy-2,4,8-trimethoxyphenanthrene (confusarin) (4), and 3,7-dihydroxy-2,4-dimethoxyphenanthrene (5). Compound 1 is a new natural product, and 2 - 4 were isolated for the first time from T. communis. In the cytotoxic assays, compounds 1 - 3 and 5 significantly inhibited the growth of HeLa cells (IC (50) = 0.97 - 20.18 microM). Compound 3, with an IC (50) value of 0.97 microM, is of special interest because of its high activity.