Geobarrettin D, a Rare Herbipoline-Containing 6-Bromoindole Alkaloid from Geodia barretti.

Geobarrettin D (1), a new bromoindole alkaloid, was isolated from the marine sponge Geodia barretti collected from Icelandic waters. Its structure was elucidated by 1D, and 2D NMR (including 1H-15N HSQC, 1H-15N HMBC spectra), as well as HRESIMS data. Geobarrettin D (1) is a new 6-bromoindole featuring an unusual purinium herbipoline moiety. Geobarrettin D (1) decreased secretion of the pro-inflammatory cytokine IL-12p40 by human monocyte derived dendritic cells, without affecting secretion of the anti-inflammatory cytokine IL-10. Thus, compound 1 shows anti-inflammatory activity.

Evaluating the Effects of an Organic Extract from the Mediterranean Sponge Geodia cydonium on Human Breast Cancer Cell Lines.

Marine sponges are an excellent source of bioactive secondary metabolites for pharmacological applications. In the present study, we evaluated the chemistry, cytotoxicity and metabolomics of an organic extract from the Mediterranean marine sponge Geodia cydonium, collected in coastal waters of the Gulf of Naples. We identified an active fraction able to block proliferation of breast cancer cell lines MCF-7, MDA-MB231, and MDA-MB468 and to induce cellular apoptosis, whereas it was inactive on normal breast cells (MCF-10A). Metabolomic studies showed that this active fraction was able to interfere with amino acid metabolism, as well as to modulate glycolysis and glycosphingolipid metabolic pathways. In addition, the evaluation of the cytokinome profile on the polar fractions of three treated breast cancer cell lines (compared to untreated cells) demonstrated that this fraction induced a slight anti-inflammatory effect. Finally, the chemical entities present in this fraction were analyzed by liquid chromatography high resolution mass spectrometry combined with molecular networking.

Metabolomic Profiling Reveals the N-Acyl-Taurine Geodiataurine in Extracts from the Marine Sponge Geodia macandrewii (Bowerbank).

A metabolomic approach was used to identify known and new natural products from the marine sponges Geodia baretti and G. macandrewii. G. baretti is known to produce bioactive natural products such as barettin (1), 8,9-dihydrobarettin (2), and bromobenzisoxazolone barettin (3), while secondary metabolites from G. macandrewii are not reported in the literature. Specimens of the two sponges were collected from different sites along the coast of Norway, and their extracts were analyzed using UHPLC-HR-MS. Metabolomic analyses revealed that extracts from both species contained barettin (1) and 8,9-dihydrobarettin (2), and all samples of G. baretti contained higher amounts of both compounds compared to G. macandrewii. The analysis of the MS data also revealed that samples of G. macandrewii contained a compound that was not present in any of the G. baretti samples. This new compound was isolated and identified as the N-acyl-taurine geodiataurine (4), and it was tested for antioxidant, anticancer, and antibacterial properties.

Marine AChE inhibitors isolated from Geodia barretti: natural compounds and their synthetic analogs.

Barettin, 8,9-dihydrobarettin, bromoconicamin and a novel brominated marine indole were isolated from the boreal sponge Geodia barretti collected off the Norwegian coast. The compounds were evaluated as inhibitors of electric eel acetylcholinesterase. Barettin and 8,9-dihydrobarettin displayed significant inhibition of the enzyme, with inhibition constants (Ki) of 29 and 19 µM respectively towards acetylcholinesterase via a reversible noncompetitive mechanism. These activities are comparable to those of several other natural acetylcholinesterase inhibitors of marine origin. Bromoconicamin was less potent against acetylcholinesterase, and the novel compound was inactive. Based on the inhibitory activity, a library of 22 simplified synthetic analogs was designed and prepared to probe the role of the brominated indole, common to all the isolated compounds. From the structure-activity investigation it was shown that the brominated indole motif is not sufficient to generate a high acetylcholinesterase inhibitory activity, even when combined with natural cationic ligands for the acetylcholinesterase active site. The four natural compounds were also analysed for their butyrylcholinesterase inhibitory activity in addition and shown to display comparable activities. The study illustrates how both barettin and 8,9-dihydrobarettin display additional bioactivities which may help to explain their biological role in the producing organism. The findings also provide new insights into the structure-activity relationship of both natural and synthetic acetylcholinesterase inhibitors.

Anti-Inflammatory Effects of a Methanol Extract from the Marine Sponge Geodia cydonium on the Human Breast Cancer MCF-7 Cell Line.

Many research groups are working to find new possible anti-inflammatory molecules, and marine sponges represent a rich source of biologically active compounds with pharmacological applications. In the present study, we tested different concentrations of the methanol extract from the marine sponge, Geodia cydonium, on normal human breast epithelial cells (MCF-10A) and human breast cancer cells (MCF-7). Our results show that this extract has no cytotoxic effects on both cell lines whereas it induces a decrease in levels of VEGF and five proinflammatory cytokines (CCL2, CXCL8, CXCL10, IFN-γ, and TNF-α) only in MCF-7 cells in a dose-dependent manner, thereby indicating an anti-inflammatory effect. Moreover, interactomic analysis suggests that all six cytokines are involved in a network and are connected with some HUB nodes such as NF-kB subunits and ESR1 (estrogen receptor 1). We also report a decrease in the expression of two NFKB1 and c-Rel subunits by RT-qPCR experiments only in MCF-7 cells after extract treatment, confirming NF-kB inactivation. These data highlight the potential of G. cydonium for future drug discovery against major diseases, such as breast cancer.

Isolation, Characterization, and Synthesis of the Barrettides: Disulfide-Containing Peptides from the Marine Sponge Geodia barretti.

Two disulfide-containing peptides, barrettides A (1) and B (2), from the cold-water marine sponge Geodia barretti are described. Those 31 amino acid residue long peptides were sequenced using mass spectrometry methods and structurally characterized using NMR spectroscopy. The structure of 1 was confirmed by total synthesis using the solid-phase peptide synthesis approach that was developed. The two peptides were found to differ only at a single position in their sequence. The three-dimensional structure of 1 revealed that these peptides possess a unique fold consisting of a long ß-hairpin structure that is cross-braced by two disulfide bonds in a ladder-like arrangement. The peptides are amphipathic in nature with the hydrophobic and charged residues clustered on separate faces of the molecule. The barrettides were found not to inhibit the growth of either Escherichia coli or Staphylococcus aureus but displayed antifouling activity against barnacle larvae (Balanus improvisus) without lethal effects in the concentrations tested.

Antioxidant and anti-inflammatory activities of barettin.

In this paper, we present novel bioactivity for barettin isolated from the marine sponge Geodia barretti. We found that barettin showed strong antioxidant activity in biochemical assays as well as in a lipid peroxidation cell assay. A de-brominated synthetic analogue of barettin did not show the same activity in the antioxidant cell assay, indicating that bromine is important for cellular activity. Barettin was also able to inhibit the secretion of the inflammatory cytokines IL-1ß and TNFα from LPS-stimulated THP-1 cells. This combination of anti-inflammatory and antioxidant activities could indicate that barettin has an atheroprotective effect and may therefore be an interesting product to prevent development of atherosclerosis.

A novel interdomain interface in crystallins: structural characterization of the ßγ-crystallin from Geodia cydonium at 0.99 Å resolution.

The ßγ-crystallin superfamily includes highly diverse proteins belonging to all of the kingdoms of life. Based on structural topology, these proteins are considered to be evolutionarily related to the long-lived ßγ-crystallins that constitute the vertebrate eye lens. This study reports the crystallographic structure at 0.99 Å resolution of the two-domain ßγ-crystallin (geodin) from the sponge Geodia cydonium. This is the most ancient member of the ßγ-crystallin superfamily in metazoans. The X-ray structure shows that the geodin domains adopt the typical ßγ-crystallin fold with a paired Greek-key motif, thus confirming the hypothesis that the crystallin-type scaffold used in the evolution of bacteria and moulds was recruited very early in metazoans. As a significant new structural feature, the sponge protein possesses a unique interdomain interface made up by pairing between the second motif of the first domain and the first motif of the second domain. The atomic resolution also allowed a detailed analysis of the calcium-binding site of the protein.

Stellettin A induces endoplasmic reticulum stress in murine B16 melanoma cells.

Isomalabaricanes are a small class of rearranged triterpenoids obtained from marine sponges. Most of these are cytotoxic to tumor cells, but the underlying mechanism is not clear. In this study, it was demonstrated that stellettin A (1), obtained from Geodia japonica, inhibited the growth of B16F10 murine melanoma cells by the induction of endoplasmic reticulum stress and accumulation of unfolded proteins. Immunoblotting analysis revealed abnormal glycosylation patterns of two melanoma marker proteins, tyrosinase and tyrosinase-related protein 1, and the retention of these proteins in the endoplasmic reticulum. Compound 1 induced the upregulation of the unfolded protein chaperone, glucose-regulated protein 78, in a dose-dependent manner. Increase of autophagosome-associated protein light chain 3 (LC3) in a membrane-bound form (LC3II) and its immunofluorescence co-localization with tyrosinase suggest the possible removal of deglycosylated and unfolded proteins by autophagy of the cells. There was no change in either the expression of the apoptosis marker protein Bcl-2 or the appearance of apoptotic nuclei in 1-treated cells. Taken together, 1 is an endoplasmic reticulum stressor that inhibits the growth of B16 melanoma cells by induction of abnormal protein glycosylation and autophagy.

Two brominated cyclic dipeptides released by the coldwater marine sponge Geodia barretti act in synergy as chemical defense.

The current work shows that two structurally similar cyclodipeptides, barettin (1) and 8,9-dihydrobarettin (2), produced by the coldwater marine sponge Geodia barretti Bowerbank act in synergy to deter larvae of surface settlers and may also be involved in defense against grazers. Previously, 1 and 2 were demonstrated to bind specifically to serotonergic 5-HT receptors. It may be suggested that chemical defense in G. barretti involves a synergistic action where one of the molecular targets is a 5-HT receptor. A mixture of 1 and 2 lowered the EC(50) of larval settlement as compared to the calculated theoretical additive effect of the two compounds. Moreover, an in situ sampling at 120 m depth using a remotely operated vehicle revealed that the sponge releases these two compounds to the ambient water. Thus, it is suggested that the synergistic action of 1 and 2 may benefit the sponge by reducing the expenditure of continuous production and release of its chemical defense substances. Furthermore, a synergistic action between structurally closely related compounds produced by the same bioenzymatic machinery ought to be the most energy effective for the organism and, thus, is more common than synergy between structurally indistinct compounds.

Total synthesis and biological assessment of (-)-exiguolide and analogues.

We describe herein an enantioselective total synthesis of (-)-exiguolide, the natural enantiomer. The methylene bis(tetrahydropyran) substructure was efficiently synthesized by exploiting olefin cross-metathesis for the assembly of readily available acyclic segments and intramolecular oxa-conjugate cyclization and reductive etherification for the formation of the tetrahydropyran rings. The 20-membered macrocyclic framework was constructed in an efficient manner by means of Julia-Kocienski coupling and Yamaguchi macrolactonization. Finally, the (E,Z,E)-triene side chain was introduced stereoselectively via Suzuki-Miyaura coupling to complete the total synthesis. Assessment of the growth inhibitory activity of synthetic (-)-exiguolide against a panel of human cancer cell lines elucidated for the first time that this natural product is an effective antiproliferative agent against the NCI-H460 human lung large cell carcinoma and the A549 human lung adenocarcinoma cell lines. Moreover, we have investigated structure-activity relationships of (-)-exiguolide, which elucidated that the C5-methoxycarbonylmethylidene group and the length of the side chain are important for the potent activity.

Antifouling activity of a dibrominated cyclopeptide from the marine sponge Geodia barretti.

Many sessile suspension-feeding marine organisms rely on chemical defense to keep their surfaces free from fouling organisms. The brominated cyclopeptides barettin (cyclo[(6-bromo-8-entryptophan)arginine]) ( 1) and 8,9-dihydrobarettin (cyclo[(6-bromotryptophan)arginine]) ( 2) from the cold-water sponge Geodia barretti have previously displayed settlement inhibition of barnacle larvae in a dose-dependent manner. In this paper, we describe a novel dibrominated cyclopeptide, bromobenzisoxazolone barettin (cyclo[(6-bromo-8-(6-bromobenzioxazol-3(1 H)-one)-8-hydroxy)tryptophan)]arginine) ( 3), which we have isolated from G. barretti and which displays settlement inhibition of barnacle larvae ( Balanus improvisus) with an EC 50 value of 15 nM. The chemical structure was determined using MS and 2D-NMR.

Apoptotic activity of isomalabaricane triterpenes on human promyelocytic leukemia HL60 cells.

Four isomalabaricane triterpenes were isolated from marine sponge Geodia japonica [W.H. Zhang, C.T. Che, Isomalabaricane-type nortriterpenoids and other constituents of the marine sponge Geodia japonica, J. Nat. Prod. 64 (2001) 1489-1492. ] and their cytotoxicity was evaluated using a human promyelocytic leukemia HL60 cell line. Of the four triterpenes tested, geoditin A was the most cytotoxic to HL60 cells [IC50=3 microg/ml (<6.6 microM)], followed by stellettins A and B, whereas geoditin B exhibited relatively weak cytotoxicity. The treated cells manifested nuclear changes characteristic for apoptosis, and associated with dissipation of mitochondrial membrane potential, activation of caspase 3, and decrease of cytoplasmic proliferating cell nuclear antigen (PCNA), as demonstrated by fluorescence and immunofluorescence microscopy. When the HL60 cells were exposed to geoditin A ranging from 1.25 to 25 microg/ml, a dose-dependent increase of reactive oxygen species, a progressive dissipation of mitochondrial membrane potential, and an increase in annexin V-FITC binding were measured by flow cytometry. Taken together our results suggest that geoditin A markedly induced reactive oxygen species, decreased mitochondrial membrane potential and mediated a caspases 3 apoptosis pathway.