Inhibition of human topoisomerase I and activation of caspase-3 by aza-angucyclinones and arylaminopyrimido[4,5-c]isoquinoline-7,10-quinones.

Cancer is the second cause of death in the world after cardiovascular diseases. Cancer cells acquire capacities not present in normal cells, such as self-sufficiency, resistance to antiproliferative stimuli, evasion of apoptosis, unlimited replication, invasiveness and metastasis. Consequently, it is of major interest to explore and develop molecules with anticancer activity directed to specific targets. In this study, we aimed to evaluate two series of polycyclic quinones: aza-angucyclinone and arylaminopyrimido[4,5-c]isoquinoline-7,10-quinones, in their capacity to inhibit human topoisomerase I (TOP1) and to trigger apoptosis through activation of caspase-3. We evaluated the capacity of the two series of polycyclic quinones to inhibit TOP1, using a DNA supercoiled relaxation assay and their capacity to induce apoptosis through the activation of caspase-3 in HL60 cells. Both series of quinones inhibited TOP1 activity over 50%. When we evaluated the pro-apoptotic capacity of both series of quinones, at therapeutically relevant concentrations, the arylaminoquinones ADPA-1CC (methyl 7-(4-methoxyphenyl)amino-1,3-dimethyl-5,8-dioxo-5,8-dihydroisoquinoline-4-carboxylate), P4 (9-phenylamino-3,4-dihydrophenanthridine-1,7,10(2H)-trione) and the aza-angucyclinone OH-6H (8-hydroxy-2,4-dimethyl-2H,4H-benzo[g]pyrimido[4,5-c]isoquinoline-1,3,7,12-tetraone) increased the caspase-3 activity by approximately 2-fold over the control. The series of the arylaminoquinones and aza-angucyclinones showed differential antiproliferative capacity. We further identified a group of them that showed antiproliferative capacity possibly through inhibition of TOP1 and by activation of caspase-3. This group of molecules may represent a potential pharmacological tool in the treatment against cancer.

Studies on quinones. Part 47. Synthesis of novel phenylaminophenanthridinequinones as potential antitumor agents.

In our search for potential anticancer agents, a series of 8- and 9-phenylamino-3,4-tetrahydro-phenanthridine-1,7,10(2H)-triones with substituent variations at 6-, 8- and 9-positions were prepared using a highly efficient sequence involving: a) solar photoacylation reactions of benzoquinone with arylaldehydes, b) one-pot procedure for the synthesis of 3,4-dihydrophenanthridine-1,7,10(2H)-trione intermediates from acylhydroquinones and c) highly regiocontrolled acid-induced amination reaction of phenanthridinequinones with phenylamines. The members of this series were in vitro evaluated using the MTT colorimetric method against one normal cell line and three human cancer cell lines. The SAR analysis indicates that the location of nitrogen substituents on the quinone nucleus, the presence of methyl, phenyl, furyl and thienyl groups at the 6-position and the aromatization of the angular cycloaliphatic ring of the phenylamino-3,4-tetrahydrophenanthridine-1,7,10(2H)-trione pharmacophore play key roles in the antitumor activity.

Gastroprotective effect and cytotoxicity of carnosic acid derivatives.

Carnosic acid (CA) is the main phenolic diterpene of rosemary (Rosmarinus officinalis L., Lamiaceae) and presents gastroprotective effect in vitro and in vivo. To determine structure-activity relationships, seventeen esters and ethers of CA were prepared, comprising aliphatic, aromatic, and heterocyclic compounds. The naturally occurring 12-O-methylcarnosic acid (14) was also included in the study. The compounds were evaluated for their gastroprotective activity in the HCl/EtOH-induced gastric lesions model in mice, and for cytotoxicity in human adenocarcinoma AGS cells, Hep G2 hepatocellular carcinoma cells, and human lung fibroblasts. At 10 mg/kg, some of the CA derivatives (5, 8, 9, 12, 14, and 18) were more effective preventing gastric lesions than the reference compound lansoprazole at the same dose. The dibenzoate 9, diindoleacetate 12, and the derivative 18 showed the best gastroprotective effect combined with the lowest cytotoxicity.

Gastroprotective effect and cytotoxicity of labdeneamides with amino acids.

Semisynthetic aromatic amides from ARAUCARIA ARAUCANA diterpene acids have been shown to display a relevant gastroprotective effect with low cytotoxicity. The aim of this work was to assess the gastroprotective effect of amino acid amides from imbricatolic acid and its 8(9)-en isomer in the ethanol/HCl-induced gastric lesions model in mice as well as to determine the cytotoxicity of the obtained compounds on the following human cell lines: normal lung fibroblasts (MRC-5), gastric adenocarcinoma (AGS), and liver hepatocellular carcinoma (Hep G2). The diterpenes 15-acetoxyimbricatolic acid, its 8(9)-en isomer, 15-hydroxyimbricatolic acid, and the 8(9)-en derivative, bearing a COOH function at C-19, were used as starting compounds. New amides with C-protected amino acids were prepared. The study reports the effect of a single oral administration of either compound 50 min before the induction of gastric lesions by ethanol/HCl. Some 20 amino acid monoamides were obtained. Dose-response experiments on the glycyl derivatives showed that at a single oral dose of 100 mg/kg, the compounds presented an effect comparable to the reference drug lansoprazole at 20 mg/kg and at 50 mg/kg reduced gastric lesions by about 50%. All derivatives obtained in amounts > 30 mg were compared at a single oral dose of 50 mg/kg. The best gastroprotective effect was observed for the exomethylene derivatives bearing a valine residue at C-19 either with an acetoxy or free hydroxy group at C-15. The tryptophanyl derivative from the acetate belonging to the 8,9-en series presented selective cytotoxicity against hepatocytes. The glycyl amide of 15-acetoxyimbricatolic acid was the most cytotoxic and less selective compound with IC50 values between 47 and 103 µM for the studied cell lines. This is the first report on the obtention of semisynthetic amino acid amides from labdane diterpenes.

Synthesis, gastroprotective effect and cytotoxicity of new amino acid diterpene monoamides and diamides.

Following our studies on the gastroprotective effect and cytotoxicity of terpene derivatives, new amides were prepared from the diterpene 8(17)-labden-15,19-dioic acid (junicedric acid) and its 8(9)-en isomer with C-protected amino acids (amino acid esters). The new compounds were evaluated for their gastroprotective effect in the ethanol/HCl-induced gastric lesions model in mice, as well as for cytotoxicity using the following human cell lines: normal lung fibroblasts (MRC-5), gastric adenocarcinoma cells (AGS) and liver hepatocellular carcinoma (Hep G2). A dose-response experiment showed that at 25 mg/kg the C-15 leucyl and C-15,19-dileucylester amides of junicedric acid reduced gastric lesions by about 65.6 and 49.6%, respectively, with an effect comparable to lansoprazole at 20 mg/kg (79.3% lesion reduction). The comparison of the gastroprotective effect of 18 new amino acid ester amides was carried out at a single oral dose of 25 mg/kg. Several compounds presented a strong gastroprotective effect, reducing gastric lesions in the 70.9-87.8% range. The diprolyl derivative of junicedric acid, the most active product of this study (87.8% lesion reduction at 25 mg/kg) presented a cytotoxicity value comparable with that of the reference compound lansoprazole. The structure-activity relationships are discussed.

Studies on quinones. Part 46. Synthesis and in vitro antitumor evaluation of aminopyrimidoisoquinolinequinones.

In the search of structure-activity relationship studies and to explore the antitumor effect associated with the pyrimidoisoquinolinequinone scaffold, several diversily substituted 8-aminopyrimido[4,5-c]isoquinolinequinones were regioselectively synthesized. Variation in the structure of the nitrogen substituent bonded to the 8-position of the pyrimidoisoquinolinequinone system led to a set of alkylamino-, phenylamino- and alkyphenylamino derivatives. The cytotoxic activity of the aminoquinone derivatives was evaluated in vitro using the MTT colorimetric method against one normal cell line (MRC-5 lung fibroblasts) and four human cancer cell lines (AGS human gastric adenocarcinoma; SK-MES-1 human lung cancer cells, and J82 human bladder carcinoma; HL-60 human leukemia) in 72-h drug exposure assays. Among the series, five compounds exhibited interesting antitumor activity against AGS human gastric adenocarcinoma and human lung cancer cells. The SAR studies revealed that both the nature of the nitrogen substituent into the quinone ring and the methyl group at the 6-position play key roles in the antitumor activity.

Gastroprotective effect of carnosic acid gamma-lactone derivatives.

Carnosic acid (1) has been shown to possess gastroprotective activity in vitro and in vivo. However, little is known of the gastroprotective effect or cytotoxicity of carnosic acid gamma-lactone (3). To determine structure-activity relationships, a series of 17 esters of 3 were prepared including aliphatic, aromatic, and heterocyclic derivatives. Also, two units of 3 were coupled with succinic and phthalic acid as linkers. The compounds were assessed for their gastroprotective effect in the HCl/EtOH-induced gastric lesions model in mice and for cytotoxicity in human lung fibroblasts, human adenocarcinoma AGS cells, and Hep G2 hepatocellular carcinoma cells. At a single oral dose of 40 mg/kg, the gastroprotective effect increased moderately with the length of the alkyl chain. The best effects were observed for the butyrate (9) and chloroacetate (6) derivatives. Activity of fatty acid esters increased with chain length but decreased with unsaturation. The best gastroprotective effect, with lowest cytotoxicity, was found for the palmitate (11) and oleate (12) derivatives.

Gastroprotective effect and cytotoxicity of abietane diterpenes from the Chilean Lamiaceae Sphacele chamaedryoides (Balbis) Briq.

OBJECTIVES: The aim of this report was to isolate, identify and assess the gastroprotective effect and cytotoxicity of abietane diterpenes from the Chilean medicinal plant Sphacele chamaedryoides (Balbis) Briq. (Lamiaceae). METHODS: The isolated compounds were identified by spectroscopic means. The gastroprotective effect of the compounds was studied on the HCl/EtOH-induced gastric lesions model in mice. The cytotoxicity of the compounds was assessed on human normal lung fibroblasts (MRC-5) and gastric adenocarcinoma cells (AGS). KEY FINDINGS: From the aerial parts of the plant, five phenolic and five p-quinone abietanes, the sesquiterpene spathulenol and two flavonoids were obtained. The main diterpene from the plant was carnosol (7:). Lansoprazole at 20 mg/kg reduced gastric lesions by 64.7% (P < 0.01), being statistically similar to carnosol at doses of 10 and 20 mg/kg; the percent lesion reduction with 7: at 5 mg/kg was 49.3%. At a single oral dose of 5 mg/kg, the diterpenes bearing a p-quinone moiety - 6,7-dehydroroyleanone (1), royleanone (2), 7,20-epoxyroyleanone (3), taxoquinone (5) and horminone (6) - presented a gastroprotective effect of 54.4, 70.8, 65.0, 35.8 and 52.7%, respectively. Of the C-7 hydroxy derivatives, the activity was much lower for the 7beta-OH isomer. The phenolic diterpenes 7 and 7-oxo-11,12,14-trihydroxy-8,11,13-abietatrien-20-al (8) inhibited gastric lesions by 49.3 and 53.0%, respectively. Royleanone (2), 7,20-epoxyroyleanone (3), horminone (6), 8 and spathulenol proved to be cytotoxic with IC50 values in the range of 11-67 microm. The selective cytotoxicity of compounds 1 (IC50: 61 and 366 microm) and 5 (IC50: 310 and 27 microm) against AGS cells and fibroblasts, respectively, merit additional studies. CONCLUSIONS: All the abietanes obtained from S. chamaedryoides present either one or two phenolic OH groups, a quinone system, or both. Several compounds present in the plant showed higher gastroprotective effect than lansoprazole. The cytotoxic effect of most compounds was found at fairly high concentrations and lacked cell specificity. Further studies are required using different tumour cell lines and viability/proliferation assays to assess the specificity of the isolated compounds. The selective cytotoxicity of compounds 1 and 5 against AGS cells and fibroblasts, respectively, merit additional studies.

Synthesis and antitumor evaluation of 8-phenylaminopyrimido[4,5-c]isoquinolinequinones.

A series of 8-phenylaminopyrimido[4,5-c]isoquinoline-7,10-quinone derivatives were prepared by regioselective amination reaction of pyrimido[4,5-c]isoquinoline-7,10-quinones with arylamines in the presence of a Lewis acid catalyst. Preliminary evaluation of the members of the series against cancer cell lines and assays of activation of their cytotoxic activity on K562 cells with ascorbic acid are reported.

Antiproliferative activity of the diterpenes jatrophone and jatropholone and their derivatives.

The antiproliferative activity of the diterpenes jatropholone A and B, 16 semi-synthetic derivatives thereof, and that of jatrophone and its three derivatives was assessed on human cell cultures. The cells used comprised normal lung fibroblasts (MRC-5), gastric adenocarcinoma (AGS), leukemia (HL-60), lung cancer (SK-MES-1), and bladder carcinoma (J82). Jatropholone A ( 1) was inactive against all the tumor cell lines; however, its acetylation rendered a compound with antiproliferative activity. The epimeric jatropholone B ( 8) was active against all the cancer cell lines, and its derivatives presented different effects on the selected cell lines. While jatrophone ( 19) showed strong anticancer activity, its derivatives 9beta,13alpha-dihydroxyisabellione and 13alpha-hydroxy-9 beta-acetoxyisabellione were less active.

Studies on quinones. Part 44: Novel angucyclinone N-heterocyclic analogues endowed with antitumoral activity.

In the search for new potentially anticancer drugs, series of angucyclinone aza-analogues containing pyridine and pyridopyridazine rings have been designed and synthesized by a highly efficient sequence involving a one-pot step for the synthesis of tricyclic quinone intermediate and highly regiocontrolled cycloaddition reactions with polarized 1,3-dienes. The new N-heterocyclic angular quinones were evaluated in vitro on normal human fibroblasts and on a panel of four distinct human cancer cell lines. All tested compounds showed high to moderate antitumor activity. Among the compounds, those with one and two pyridine moieties fused to the quinone system have shown the best effect. Structure-activity relationships established the main structural requirement for the activity of the new potential anticancer drugs.

New gastroprotective ferruginol derivatives with selective cytotoxicity against gastric cancer cells.

The diterpene ferruginol has shown a strong protective effect in animal gastric ulcer models. In the present work, we report the gastroprotective effect and cytotoxicity of 16 new semisynthetic ester derivatives of ferruginol. The gastroprotective effect of these compounds was assessed with the HCl/EtOH-induced gastric lesions model in mice and the cytotoxicity was measured using MRC-5 fibroblasts, gastric adenocarcinoma (AGS) and liver hepatoma Hep G2 cells. The compounds were tested for a gastroprotective effect at a single oral dose of 20 mg/kg. The best gastroprotective effect was elicited by ferruginyl nicotinate ( 13), reducing the lesion index by 71 %, while the derivatives ferruginyl chloroacetate ( 2), ferruginyl palmitate ( 6), ferruginyl oleate ( 7), ferruginyl 3,5-dinitrobenzoate ( 11), ferruginyl 3-methylbenzofuran-2-carbonyl ester ( 12), ferruginyl indoleacetate ( 14), ferruginyl indolebutyrate ( 15) and ferruginyl pthalate ( 16) reduced the lesions by 49 - 66 %. The most promising compounds were 11, 13 and 14, presenting a gastroprotective effect higher or similar to that of ferruginol but with a high selectivity towards the tumor AGS cells. Among the three products, the most selective towards AGS cells was 14, followed by 13, and 11 (IC (50) values of 12, 22 and 29 microM, respectively). The isobutyrate 4, inactive as a gastroprotective agent, showed selective cytotoxicity against AGS and Hep G2 cells (IC (50) values of 60 and 39.2 microM, respectively). The cytotoxicity of the above cited compounds towards fibroblasts was >1000 microM. Considering the aliphatic esters of ferruginol, the best gastroprotective activity was found in the C (16) and C (18) derivatives but tended to decrease with increasing aliphatic chain unsaturation. For short-chain esters, the gastroprotective effect could be observed when the chain contained a chlorine atom. For aromatic esters, the presence of nitro groups or a nitrogen atom in the aromatic ring enhanced the gastroprotective activity. The compounds with the best gastroprotective effect and the highest selectivity against tumor cells bear an amino group (indoleacetate and nicotinate) or nitro group (3,5-dinitrobenzoate).

Studies on quinones. Part 43: Synthesis and cytotoxic evaluation of polyoxyethylene-containing 1,4-naphthoquinones.

A series of naphthoquinones 2,3-disubstituted with chlorine and oxyethylene groups have been prepared from 2,3-dichloro- and 2,3-dimethoxy-1,4-naphthoquinone. The members of these series were tested on normal human fibroblasts and on a panel of four human cancer cell lines. Antitumor activities, which were in the range of IC(50) 1.3-89.5 microM, discussed in terms of LUMO energy, lipophilicity and size of the polyoxyethylene moiety.

Ferruginol suppresses survival signaling pathways in androgen-independent human prostate cancer cells.

Ferruginol, a bioactive compound isolated from a Chilean tree (Podocarpaceae), attracts attention as a consequence of its pharmacological properties, which include anti-fungal, anti-bacterial, cardioprotective, anti-oxidative, anti-plasmodial and anti-ulcerogenic actions. Nevertheless, the molecular basis for these actions remains only partly understood and hence we investigated the effects of ferruginol on androgen-independent human prostate cancer cells (PC3), a known model for solid tumor cells with an exceptional resistance to therapy. The results show that ferruginol induces PC3 cell death via activation of caspases as well as apoptosis-inducing factor (AIF) as confirmed by its translocation into the nucleus. In order to clarify the biochemical mechanism responsible for the anti-tumor activity of ferruginol, we analyzed a set of molecular mediators involved in tumor cell survival, progression and aggressiveness. Ferruginol was able to trigger inhibition/downregulation of Ras/PI3K, STAT 3/5, protein tyrosine phosphatase and protein kinases related to cell cycle regulation. Importantly, the toxic effect of ferruginol was dramatically impeded in a more reducing environment, which indicates that at least in part, the anti-tumoral activity of ferruginol might be related to redox status modulation. This study supports further examination of ferruginol as a potential agent for both the prevention and treatment of prostate cancer.

Gastroprotective activity of ferruginol in mice and rats: effects on gastric secretion, endogenous prostaglandins and non-protein sulfhydryls.

The gastroprotective mechanism of the natural diterpene ferruginol was assessed in mice and rats. The involvement of gastric prostaglandins (PGE(2)), reduced glutathione, nitric oxide or capsaicin receptors was evaluated in mice either treated or untreated with indometacin, N-ethylmaleimide (NEM), N-nitro-L-arginine methyl ester (L-NAME) or ruthenium red, respectively, and then orally treated with ferruginol or vehicle. Gastric lesions were induced by oral administration of ethanol. The effects of ferruginol on the parameters of gastric secretion were assessed in pylorus-ligated rats. Gastric PGE(2) content was determined in rats treated with ferruginol and/or indometacin. The reduction of gastric glutathione (GSH) content was determined in rats treated with ethanol after oral administration of ferruginol, lansoprazole or vehicle. Finally, the acute oral toxicity was assessed in mice. Indometacin reversed the gastroprotective effect of ferruginol (25 mg kg(-1)) but not NEM, ruthenium red or L-NAME. The diterpene (25 mg kg(-1)) increased the gastric juice volume and its pH value, and reduced the titrable acidity but was devoid of effect on the gastric mucus content. Ferruginol (25, 50 mg kg(-1)) increased gastric PGE(2) content in a dose-dependent manner and prevented the reduction in GSH observed due to ethanol-induced gastric lesions in rats. Single oral doses up to 3 g kg(-1) ferruginol did not elicit mortality or acute toxic effects in mice. Our results showed that ferruginol acted as a gastroprotective agent stimulating the gastric PGE(2) synthesis, reducing the gastric acid output and improving the antioxidant capacity of the gastric mucosa by maintaining the GSH levels.

Biotransformations of imbricatolic acid by Aspergillus niger and Rhizopus nigricans cultures.

Microbial transformation of imbricatolic acid (1) by Aspergillus niger afforded 1alpha-hydroxyimbricatolic acid (2), while transformation with Rhizopus nigricans yielded 15-hydroxy-8,17-epoxylabdan-19-oic acid (3). When the diterpene 1 was added to a Cunninghamella echinulata culture, the main products were the microbial metabolites mycophenolic acid (4) and its 3-hydroxy derivative 5. All the structures were elucidated by spectroscopic methods. The cytotoxicity of these compounds towards human lung fibroblasts and AGS cells was assessed. While 4 and 5 showed low cytotoxicity, with IC50 values > 1000 microM against AGS cells and fibroblasts, 1alpha-hydroxyimbricatolic acid (2) presented moderate toxicity towards these targets, with IC50 values of 307 and 631 microM, respectively. The structure of 2 is presented for the first time.

Gastroprotective and cytotoxic effect of semisynthetic ferruginol derivatives.

The gastroprotective abietane diterpene ferruginol has been shown to present high cytotoxicity. In order to obtain active compounds with less cytotoxicity, 18 semisynthetic ferruginol derivatives and totarol were assessed for their gastroprotective effects in the HCl/ethanol-induced gastric lesion model in mice, as well as for cytotoxicity in human gastric epithelial cells (AGS) and human lung fibroblasts (MRC-5). At 20 mg kg(-1), the greatest gastroprotective effects were provided by abieta-8,11,13-triene (1), abieta-8,11,13-trien-12-yl-2-chloropropanoate (8), abieta-8,11,13-trien-12-yl propenoate (9), 12-(2,3,4,6-tetra-O-acetyl-beta-D-glucopyranosyloxy)-abieta-8,11,13-triene (17) and 12-(beta-D-galactopyranosyloxy)-abieta-8,11,13-triene (18), all of which were as active as the reference drug lansoprazole at 20 mg kg(-1), reducing gastric lesions by 69, 76, 67, 72 and 61%, respectively. No relation was observed between lipophilicity and the gastroprotective effect. Compounds that showed the greatest cytotoxicity towards AGS cells were ferruginol (2), the corresponding formate (5), acetate (6), propionate (7), 8, 9, 12-(beta-D-glucopyranosyloxy)-abieta-8,11,13-triene (16), 18 and totarol (20) (IC50 18-44 microM). Ferruginol and compounds 5-9, 16, 18 and 20 were the most toxic compounds against fibroblasts (IC50 19-56 microM), with a correlation to AGS cells. The derivative 19 was much more active against AGS cells than towards fibroblasts. The best activity/cytotoxicity ratio was found for compound 17, with a lesion index comparable with lansoprazole at 20 mg kg(-1) and cytotoxicity >1000 microM towards MRC-5 and AGS cells, respectively. In conclusion, some derivatives showed a better gastroprotective effect/cytotoxicity ratio than the parent compound ferruginol. A total of 13 new compounds are reported here for the first time.

Gastroprotective and ulcer-healing activity of oleanolic acid derivatives: in vitro-in vivo relationships.

The triterpene oleanolic acid 1 and its semisynthetic derivatives 2-7 were assessed for gastroprotective and ulcer-healing effect using human epithelial gastric cells (AGS) and human lung fibroblasts (MRC-5). The ability of the compounds to protect the AGS cells against the damage induced by sodium taurocholate (NaT), to stimulate the cellular reduced glutathione (GSH) and prostaglandin E(2) content, to enhance AGS and MRC-5 cell proliferation and to scavenge superoxide anion in vitro was studied. The cytotoxicity of the compounds was assessed towards MRC-5 and AGS cells. In addition, the gastroprotective activity of the compounds was assessed in vivo using the HCl/EtOH-induced ulcer model in mice. All the assayed compounds displayed a significant reduction of AGS cells damage after incubation with NaT. None of the studied compounds was active as a superoxide anion scavenger nor stimulated the GSH content in AGS cell cultures. Compounds 1, 2, 4 and 6 were able to increase the prostaglandin content in AGS cell cultures. Concerning the proliferation assays, a significant stimulating effect was observed for compounds 3 and 7 on AGS cells and for 1 and 7 on MRC-5 fibroblasts. Regarding cytotoxicity, derivatives 2, 4, 6 and 7 were less toxic than the parent compound oleanolic acid. Our results strongly support the predictive capacity of the in vitro assessment of gastroprotective activity allowing the reduction of experimental animals.

Studies on quinones. Part 41: synthesis and cytotoxicity of isoquinoline-containing polycyclic quinones.

In the search for new potentially anticancer drugs, isoquinolinequinone-containing polycyclic compounds have been designed and synthesized through highly regiocontrolled cycloaddition reactions of methyl 1,3-dimethyl-5,8-dioxo-5,8-dihydroisoquinoline-4-carboxylate with polarized 1,3-dienes and a thiazole-o-quinodimethane. The new N-heterocyclic quinones were tested on normal human fibroblasts and four distinct human cancer cell lines. Two of the evaluated compounds displayed significant in vitro activity (IC50: 0.44-5.9 microM) comparable to that of the reference drug etoposide.

Gastroprotective and ulcer healing effect of ferruginol in mice and rats: assessment of its mechanism of action using in vitro models.

The gastroprotective activity of the diterpene ferruginol isolated from Prumnopitys andina wood and bark was determined on HCl/EtOH-induced gastric lesions in mice. The effect of the compound on the healing of subacute gastric lesions in rats was also studied. The mode of action of the diterpene was assessed using human gastric epithelial cells (AGS) and MRC-5 fibroblasts. The effect of ferruginol on the prostaglandin E2 content, protection against sodium taurocholate induced-damage and reduced glutathione content was evaluated on AGS cells as well as on the growth of AGS and fibroblast cultures. The free radical scavenging effect of ferruginol was assessed by the 1,1-diphenyl-2-picryl-hydrazil radical and superoxide anion assays. The effect of ferruginol on human erythrocyte membrane lipoperoxidation was determined. The cytotoxicity of the compound was assessed by means of the neutral red uptake. At 25 mg/kg, ferruginol inhibited the appearance of gastric lesions by 60% showing similar effects than lansoprazole at 20 mg/kg. Additionally, the compound displayed a significant ulcer healing activity in rats at 25 and 50 mg/kg with curative ratios of 36.0% and 92.5%, respectively, while the reference compound ranitidine at 50 mg/kg showed a curative ratio of 79.6%. At 6 and 12 microM, ferruginol increased significantly the prostaglandin E2 content. A strong inhibition of lipoperoxidation was found (IC50: 1.4 microM), but no effect was observed on the sodium taurocholate induced-damage or reduced glutathione content. Ferruginol stimulated cell proliferation at 1-2 microM in AGS cells and at 4-8 microM in fibroblasts, with cytotoxicities (IC50) of 24 and 26 microM, respectively. Our results support that ferruginol acts as gastroprotective increasing the PGs content, protecting the cells against lipid peroxidation and improving the gastric ulcer healing by a stimulating effect on the cell proliferation. These findings encourage further pharmacological studies of ferruginol as a potential new anti-ulcerogenic drug.