The plant diterpene epoxysiderol targets Hsp70 in cancer cells, affecting its ATPase activity and reducing its translocation to plasma membrane.

The ATP-dependent molecular chaperone Hsp70 is over-expressed in cancer cells where it plays pivotal roles in stabilization of onco-proteins, promoting cell proliferation and protecting cells from apoptosis and necrosis. Moreover, a relationship between the ability of cancer cells to migrate and the abundance of membrane-associated Hsp70 was shown. However, although Hsp70 is a promising target for cancer therapy, there is a still unsatisfied requirement of inhibitors possibly blocking its cancer-associated activities. Moving from the evidence that the plant diterpene oridonin efficiently targets Hsp70 1A in cancer cells, we set up a small kaurane diterpenoids collection and subjected it to a Surface Plasmon Resonance-screening, to identify new putative inhibitors of this chaperone. The results obtained suggested epoxysiderol as an effective Hsp70 1A interactor; therefore, using a combination of bioanalytical, biochemical and bioinformatics approaches, this compound was shown to bind the nucleotide-binding-domain of the chaperone, thus affecting its ATPase activity. The interaction between epoxysiderol and Hsp70 1A was also demonstrated to actually occur inside cancer cells, significantly reduced the translocation of the chaperone to the cell membrane, thus suggesting a possible role of epoxysiderol as an anti-metastasis agent.

Correction to: Increasing the synthesis of bioactive abietane diterpenes in Salvia sclarea hairy roots by elicited transcriptional reprogramming.

Unfortunately, the second author name was wrongly published in the original publication. The correct author name should read as follows.

Increasing the synthesis of bioactive abietane diterpenes in Salvia sclarea hairy roots by elicited transcriptional reprogramming.

KEY MESSAGE: Transcriptional activation of genes belonging to the plastidial MEP-derived isoprenoid pathway by elicitation with methyl jasmonate and coronatine enhanced the content of bioactive abietane diterpenes in Salvia sclarea hairy roots. We have shown that aethiopinone, an abietane diterpene synthesized in Salvia sclarea roots is cytotoxic and induces apoptosis in human melanoma cells. To develop a production platform for this compound and other abietane diterpenes, hairy root technology was combined with the elicitation of methyl jasmonate (MeJA) or the phytotoxin coronatine (Cor). Both MeJA and Cor induced a significant accumulation of aethiopinone, but prolonged exposure to MeJA irremediably caused inhibition of hairy root growth, which was unaffected by Cor treatment. Considering together the fold increase in aethiopinone content and the final hairy root biomass, the best combination was a Cor treatment for 28 days, which allowed to obtain up to 105.34 ± 2.30 mg L-1 of this compound to be obtained, corresponding to a 24-fold increase above the basal content in untreated hairy roots. MeJA or Cor elicitation also enhanced the synthesis of other bioactive abietane-quinone diterpenes. The elicitor-dependent steering effect was due to a coordinated transcriptional activation of several biosynthetic genes belonging to the plastidial MEP-derived isoprenoid pathway. High correlations between aethiopinone content and MeJA or Cor-elicited level of gene transcripts were found for DXS2 (r 2 = 0.99), DXR (r 2 = 0.99), and GGPPS (r 2 = 0.98), encoding enzymes acting upstream of GGPP, the common precursor of diterpenes and other plastidial-derived terpenes, as well as CPPS (r 2 = 0.99), encoding the enzyme involved in the first cyclization steps leading to copalyl-diphosphate, the precursor of abietane-like diterpenes. These results point to these genes as possible targets of metabolic engineering approaches to establish a more efficient production platform for such promising anti-proliferative plant-derived compounds.

Antioxidant activities and quali-quantitative analysis of different Smallanthus sonchifolius [(Poepp. and Endl.) H. Robinson] landrace extracts.

Five landraces of Smallanthus sonchifolius [(Poepp. and Endl.) H. Robinson], known as yacon, were investigated in total phenolic content, antioxidant activity and chemical composition of ethanol extracts (EEs) and decoction extracts (DEs). The results demonstrated that DEs are rich in phenolic acids as caffeic acid, while the EEs show an higher amount of flavonoids, as luteolin 3',7-O-diglucoside and luteolin 7-O-glucoside. These flavonoid glycosides were identified for the first time in yacon extracts, together with apigenin and luteolin. The phytochemical profile explains the different antioxidant activities shown in our study. The landraces PER6-DE and PER4-DE showed the highest radical-scavenging activity and reducing power related to their polyphenolic contents. Results also show that yacon can be considered an important source of bioactive compounds with significant differences among the analysed landraces.

Effect of sesquiterpene lactone coronopilin on leukaemia cell population growth, cell type-specific induction of apoptosis and mitotic catastrophe.

OBJECTIVES: The aim of this study was to investigate anti-leukaemic potential of coronopilin, a sesquiterpene lactone from Ambrosia arborescens, and to characterize mechanism(s) underlying its activity. MATERIALS AND METHODS: The study was conducted on Jurkat and U937, two leukaemia-derived cell lines. Apoptosis and impairment of cell cycle progression were evaluated by flow cytometry and by microscopic analysis. Changes in protein expression and activation were evaluated by western blot analysis. Coronopilin-tubulin covalent adducts were demonstrated by mass spectrometry. RESULTS: Coronopilin inhibited (IC(50) ≤ 20 µm) leukaemia cell population growth, but displayed poor cytotoxicity to normal white blood cells. On Jurkat cells, coronopilin exerted cell population growth inhibition activity, mainly by triggering caspase-dependent apoptosis. Conversely, in U937 cells, coronopilin's primary response was a robust arrest in G(2) /M. Marked increase in mitotic index and presence of activated cyclin B1/Cdk1 complex, phosphorylated histone H3 at Ser10, and hyperpolymerized tubulin indicated that cells accumulated in mitosis. Prolonged mitotic arrest ultimately resulted in U937 mitotic catastrophe, and dying cells exhibited the features of non-caspase-dependent death. CONCLUSIONS: This study demonstrated that coronopilin efficiently inhibited leukaemia cell population growth by triggering cell type-specific responses. Moreover, coronopilin-mediated cell population expansion inhibition was specific to neoplastic cells, as normal white blood cell viability was not significantly affected. Thus, coronopilin may represent an interesting new chemical scaffold upon which to develop new anti-leukaemic agents.

Data collection and advanced statistical analysis in phytotoxic activity of aerial parts exudates of Salvia spp

In order to define the phytotoxic potential of Salvia species a database was developed for fast and efficient data collection in screening studies of the inhibitory activity of Salvia exudates on the germination of Papaver rhoeas L. and Avena sativa L.. The structure of the database is associated with the use of algorithms for calculating the usual germination indices reported in the literature, plus the newly defined indices (Weighted Average Damage, Differential Weighted Average Damage, Germination Weighted Average Velocity) and other variables usually recorded in experiments of phytotoxicity (LC50, LC90). Furthermore, other algorithms were designed to calculate the one-way ANOVA followed by Duncan's multiple range test to highlight automatically significant differences between the species. The database model was designed in order to be suitable also for the development of further analysis based on the artificial neural network approach, using Self-Organising Maps (SOM).

Inhibition of bone resorption by Tanshinone VI isolated from Salvia miltiorrhiza Bunge.

During the last decade, a more detailed knowledge of molecular mechanisms involved in osteoclastogenesis has driven research efforts in the development and screening of compound libraries of several small molecules that specifically inhibit the pathway involved in the commitment of the osteoclast precursor cells. Natural compounds that suppress osteoclast differentiation may have therapeutic value in treating osteoporosis and other bone erosive diseases such as rheumatoid arthritis or metastasis associated with bone loss. In ongoing investigation into anti-osteoporotic compounds from natural products we have analyzed the effect of Tanshinone VI on osteoclasts differentiation, using a physiologic three-dimensional osteoblast/bone marrow model of cell co-culture. Tanshinone VI is an abietane diterpene extracted from the root of Salvia miltiorrhiza Bunge (Labiatae), a Chinese traditional crude drug, "Tan-Shen". Tashinone has been widely used in clinical practice for the prevention of cardiac diseases, arthritis and other inflammation-related disorders based on its pharmacological actions in multiple tissues. Although Tanshinone VI A has been used as a medicinal agent in the treatment of many diseases, its role in osteoclast-related bone diseases remains unknown. We showed previously that Tanshinone VI greatly inhibits osteoclast differentiation and suppresses bone resorption through disruption of the actin ring; subsequently, we intended to examine the precise inhibitory mechanism of Tanshinone VI on osteoclast differentiating factor. This study shows, for the first time, that Tanshinone VI prevents osteoclast differentiation by inhibiting RANKL expression and NFkB induction.

New polyhydroxylated triterpenes and anti-inflammatory activity of Salvia hierosolymitana.

Salvia hierosolymitana (Lamiaceae) leaves were investigated for their topical anti-inflammatory properties following a bioassay-guided fractionation. The chloroform extract showed a strong inhibition of the Croton oil-induced ear oedema in mice, comparable to that of indomethacin. Phytochemical and pharmacological investigations of this extract led to the isolation of eight anti-inflammatory polyhydroxylated triterpenes of the ursane and oleanane series. Four of them are new compounds, whose structures were elucidated by NMR and mass spectroscopy as 3beta, 6alpha,23-trihydroxyurs-12,19(29)-dien-28-oic acid, 23-( TRANS-P-coumaroyloxy)-3beta, 6alpha,30-trihydroxyurs-12-en-28-oic acid, 2alpha,3beta-dihydroxyolean-28-oic acid and 24-nor-2alpha,3beta-dihydroxyolean-4(23),12-ene.

The effect of isoquinoline alkaloids on opiate withdrawal.

Our interest has been centered on isoquinoline alkaloids obtained from Argemone mexicana (Papaveraceae), Aristolochia constricta (Aristolochiaceae) and the opium alkaloid, papaverine. In this respect, the effect of these isoquinoline alkaloids was investigated on contractions induced by naloxone of isolated guinea pig ileum acutely exposed to morphine in vitro. The activity of these alkaloids was compared to the control compound, papaverine. Furthermore, the effect of these isoquinoline alkaloids was also determined on naloxone-precipitated withdrawal in isolated guinea pig ileum exposed to DAMGO (highly selective mu opioid receptor agonist) and U50-488H (highly selective kappa opioid receptor agonist) to test whether the possible interaction of isoquinoline alkaloids on opioid withdrawal involves mu- and/or kappa-opioid receptors. Isoquinoline alkaloids from A. mexicana (from 5 x 10(-6) to 1 x 10(-4) M), from A. constricta (1 x 10(-5) x 10(-5)-1 x 10(-4) M) as well as papaverine treatment (1 x 10(-7)-5 x 10(-6)-1 x 10(-6) M) before or after the opioid agonists were able of both preventing and reversing the naloxone-induced contraction after exposure to mu (morphine and DAMGO) or kappa (U50-488H) opiate receptor agonists in a concentration-dependent manner. Both acetylcholine response and electrical stimulation were also reduced by isoquinoline alkaloids and papaverine treatment as well as the final opiate withdrawal was still reduced. The results of the present study indicate that isoquinoline alkaloids as well as papaverine were able to produce significant influence on the opiate withdrawal in vitro and these compounds were able to exert their effects both at mu and kappa opioid agonists.

Triterpenoids from Salvia wagneriana.

From the exudate of Salvia wagneriana Polak, beside the known ursolic acid, two pentaoxygenated triterpenoids were isolated. Their structures were determined as 3-oxo-11alpha,19beta,20,22beta-tetrahydroxy-lupane (1) and 3beta,11alpha,19beta,20,22beta-pentahydroxy-lupane (2) using a combination of one- and two-dimensional NMR techniques.

Two new sesquiterpene lactones from Montanoa tomentosa ssp. microcephala.

Two new sesquiterpene lactones: 8alpha-(4'-acetoxymethacryloyloxy)-3alpha,9beta-dihydroxy-1(10)E,4Z,11(13)-germacratrien-12,6alpha-olide (1) and 8alpha-(2'E)-(2'-acetoxymethyl-2'-butenoyloxy)-3alpha,9beta-dihydroxy-1(10)E,4Z,11(13)-germacratrien-12,6alphaolide (2), together with the known zoapatanolide A were isolated from the aerial parts of Montanoa tomentosa Cerv. in La Llave et Lex ssp. microcephala (Sch. Bip. In K. Koch) V.A. Funk (Asteraceae). The structures of all compounds were established on the basis of 1D, 2D NMR, and EIMS analysis.

The antiproliferative effects of Uncaria tomentosa extracts and fractions on the growth of breast cancer cell line.

Uncaria tomentosa, also known as "Uña de gato", is a Rubiaceae species widely used in South-American folk medicine for the treatment of cancer, arthritis, gastritis and epidemic diseases. Extracts of the plant have been shown to possess cytostatic and anti-inflammatory activity as well as mutagenic and antimutagenic properties. However, to date no studies have been carried out to verify the direct antitumor activity of the extracts. The present study investigates the effects of some extracts and their chromatographic fractions from the bark of U. tomentosa on the growth of a human breast cancer cell line (MCF7). Our data indicated that, in addition to the antimutagenic activity, U. tomentosa extracts and fractions exert a direct antiproliferative activity on MCF7. The bioassay-directed fractionation from barks and leaves resulted in the isolation of two active fractions, which displayed an IC50 of 10 mg/ml and 20 mg/ml, respectively and an antiproliferative effect, with about 90% of inhibition at a concentration of 100 mg/ml.

Studies on the constituents of Cyclanthera pedata fruits: isolation and structure elucidation of new flavonoid glycosides and their antioxidant activity.

The isolation of six flavon glycosides (1-6), among them four new natural compounds (1-4), from the CHCl(3)/MeOH extract of the fruits of Cyclanthera pedata is reported. All of the structures were elucidated by spectroscopic methods, including the concerted application of one-dimensional ((1)H, (1)H TOCSY, (13)C, and (13)C DEPT-NMR) and two-dimensional NMR techniques (DQF-COSY, HSQC, and HMBC). For all of the isolated compounds the antioxidant activity was determined by measuring the free radical scavenging activity, using the Trolox equivalent antioxidant capacity (TEAC) method, and the coupled oxidation of beta-carotene and linoleic acid.

Antioxidant principles from Bauhinia tarapotensis.

A new cyclohexenone (1) and a new caffeoyl ester derivative (2), together with the known compounds (-)-isolariciresinol 3-alpha-O-beta-D-glucopyranoside (3), (+)-1-hydroxypinoresinol 1-O-beta-D-glucopyranoside (4), isoacteoside (5), luteolin 4'-O-beta-D-glucopyranoside (6), and indole-3-carboxylic acid (7), were isolated from the leaves of Bauhinia tarapotensis. The structures of these new compounds were determined by spectroscopic data analysis. The antioxidant activities of 1-7 were determined by measuring their free radical scavenging effects, using the 1,1-diphenyl-2-dipicrylhydrazyl free radical (DPPH) and Trolox equivalent antioxidant activity (TEAC) methods, and the coupled oxidation of beta-carotene and linoleic acid. Compounds 3-5 showed good activities in the DPPH and TEAC tests, while compounds 1 and 2 were active in the coupled oxidation of beta-carotene and linoleic acid bioassay.

Flavonoids from Aconitum napellus subsp. neomontanum.

Three flavonol glycosides quercetin 7-O-(6-trans-caffeoyl)-beta-glucopyranosyl-(1-->3)-alpha-rhamnopyranoside-3-O-beta-glucopyranoside (1), kaempferol 7-O-(6-trans-caffeoyl)-beta-glucopyranosyl-(1-->3)-alpha-rhamnopyranoside-3-O-beta-glucopyranoside (2), and kaempferol 7-O-(6-trans-p-coumaroyl)-beta-glucopyranosyl-(1-->3)-alpha-rhamnopyranoside-3-O-beta-glucopyranoside (3), together with the known beta-3,4-dihydroxyphenethyl beta-glucopyranoside, were isolated from the flowers of Aconitum napellus subsp. neomontanum. Their structures were elucidated by spectroscopic methods, including 2D NMR spectral techniques.

Triterpene saponins from Tupidanthus calyptratus.

Five new bisdesmosidic saponins (1--5) were isolated from the aerial parts of Tupidanthus calyptratus. Their structures were determined by (1)H--(1)H correlation spectroscopy (COSY, TOCSY, ROESY) and (1)H--(13)C correlation (HSQC, HMBC) NMR experiments, FABMS, and chemical data.

A new diterpenoid with antispasmodic activity from Salvia cinnabarina.

From the leaf surface exudate of the aerial parts of Salvia cinnabarina a new secoisopimarane diterpenoid with a non-specific spasmolytic activity on histamine-, acetylcholine-, and barium chloride-induced contractions in the isolated guinea-pig ileum was obtained. The IC50 value obtained was comparable with that obtained for papaverine. The structure of 3,4-secoisopimara-4(18),7,15-triene-3-oic acid was established by 1D and 2D NMR spectroscopic techniques.

New protopine alkaloids from Aristolochia constricta reduce morphine withdrawal in vitro.

The present study examines the effect of four new protopine alkaloids (1-4) isolated and purified from the aerial parts of Aristolochia constricta (Aristolichiaceae) on morphine withdrawal in vitro. The results of our experiments indicate that the pure compounds (1-4) significantly and in a concentration-dependent manner reduced the morphine withdrawal. The results of the present study suggest that these new protopine alkaloids may be potential anti-addictive agents.