Chemical Investigation of the Calcareous Marine Sponge Pericharax heteroraphis, Clathridine-A Related Derivatives Isolation, Synthesis and Osteogenic Activity.

As a result of screening a panel of marine organisms to identify lead molecules for the stimulation of endochondral bone formation, the calcareous sponge Pericharax heteroraphis was identified to exhibit significant activity during endochondral differentiation. On further molecular networking analysis, dereplication and chemical fractionation yielded the known clathridine A-related metabolites 3-6 and the homodimeric complex (clathridine A)2 Zn2+ (9), together with the new unstable heterodimeric complex (clathridine A-clathridimine)Zn2+ (10). With the presence of the zinc complexes annotated through the LC-MS analysis of the crude extract changing due to the instability of some metabolites and complexes constituting the mixture, we combined the isolation of the predicted molecules with their synthesis in order to confirm their structure and to understand their reactivity. Interestingly, we also found a large quantity of the contaminant benzotriazoles BTZ (7) and its semi-dimer (BTZ)2CH2 (8), which are known to form complexes with transition metals and are used for preventing corrosion in water. All isolated 2-aminoimidazole derivatives and complexes were synthesized not only for structural confirmation and chemical understanding but to further study their bioactivity during endochondral differentiation, particularly the positively screened imidazolone derivatives. Compounds leucettamine B, clathridine A and clathridimine were found to increase type X collagen transcription and stimulate endochondral ossification in the ATDC5 micromass model.

Isolation, Synthesis and Absolute Configuration of the Pericharaxins A and B, Epimeric Hydroxy-Polyene Glycerol Ethers from the Calcarean Sponge Pericharax heteroraphis.

Naturally occurring epimeric hydroxy-polyene glycerol ether pericharaxins A (1a) and B (1b) were isolated from the calcarean sponge Pericharax heteroraphis. The structural and stereochemical characterization of both diastereoisomers were established on the basis of spectroscopic data analysis and total synthesis in seven steps. The mixture of pericharaxins A (1a) and B (1b) was proven to be epimeric by chiral-phase HPLC analysis of both synthetic and natural samples. Further separation of the epimers and application of Mosher's method to the synthetic compounds allowed unequivocal absolute configuration assignment. While natural products and the synthetic intermediates were shown to be non-cytotoxic on the HCT116 cell line, the endochondral differentiation activity using human type X collagen transcription activity in ATDC5 cells is interesting.

Bioactive Bromotyrosine Derivatives from the Pacific Marine Sponge Suberea clavata (Pulitzer-Finali, 1982).

Chemical investigation of the South-Pacific marine sponge Suberea clavata led to the isolation of eight new bromotyrosine metabolites named subereins 1-8 (2-9) along with twelve known co-isolated congeners. The detailed configuration determination of the first representative major compound of this family 11-epi-fistularin-3 (11R,17S) (1) is described. Their chemical characterization was achieved by HRMS and integrated 1D and 2D NMR (nuclear magnetic resonance) spectroscopic studies and extensive comparison with literature data. For the first time, a complete assignment of the absolute configurations for stereogenic centers C-11/17 of the known members (11R,17S) 11-epi-fistularin-3 (1) and 17-deoxyfistularin-3 (10) was determined by a combination of chemical modifications, Mosher's technology, and ECD spectroscopy. Consequently, the absolute configurations of all our new isolated compounds 2-9 were determined by the combination of NMR, Mosher's method, ECD comparison, and chemical modifications. Interestingly, compounds 2-7 were obtained by chemical transformation of the major compound 11-epi-fistularin-3 (1). Evaluation for acetylcholinesterase inhibition (AChE), DNA methyltransferase 1 (DNMT1) modulating activity and antifouling activities using marine bacterial strains are also presented.

Quorum Sensing Inhibitory and Antifouling Activities of New Bromotyrosine Metabolites from the Polynesian Sponge Pseudoceratina n. sp.

Four new brominated tyrosine metabolites, aplyzanzines C-F (1-4), were isolated from the French Polynesian sponge Pseudoceratina n. sp., along with the two known 2-aminoimidazolic derivatives, purealidin A (5) and 6, previously isolated, respectively, from the sponges Psammaplysilla purpurea and Verongula sp. Their structures were assigned based on the interpretation of their NMR and HRMS data. The compounds exhibited quorum sensing inhibition (QSi) and antifouling activities against several strains of bacteria and microalgae. To our knowledge, the QSi activity of this type of bromotyrosine metabolite is described here for the first time.

New Antimalarial and Antimicrobial Tryptamine Derivatives from the Marine Sponge Fascaplysinopsis reticulata.

Chemical study of the CH2Cl2-MeOH (1:1) extract of the sponge Fascaplysinopsis reticulata collected in Mayotte highlighted three new tryptophan derived alkaloids, 6,6'-bis-(debromo)-gelliusine F (1), 6-bromo-8,1'-dihydro-isoplysin A (2) and 5,6-dibromo-8,1'-dihydro-isoplysin A (3), along with the synthetically known 8-oxo-tryptamine (4) and the three known molecules from the same family, tryptamine (5), (E)-6-bromo-2'-demethyl-3'-N-methylaplysinopsin (6) and (Z)-6-bromo-2'-demethyl-3'-N-methylaplysinopsin (7). Their structures were elucidated by 1D and 2D NMR spectra and HRESIMS data. All compounds were evaluated for their antimicrobial and their antiplasmodial activities. Regarding antimicrobial activities, the best compounds are (2) and (3), with minimum inhibitory concentration (MIC) of 0.01 and 1 µg/mL, respectively, towards Vibrio natrigens, and (5), with MIC values of 1 µg/mL towards Vibrio carchariae. In addition the known 8-oxo-tryptamine (4) and the mixture of the (E)-6-bromo-2'-demethyl-3'-N-methylaplysinopsin (6) and (Z)-6-bromo-2'-demethyl-3'-N-methylaplysinopsin (7) showed moderate antiplasmodial activity against Plasmodium falciparum with IC50 values of 8.8 and 8.0 µg/mL, respectively.

C2'-F Stereoconfiguration As a Puckering Switch for Base Stacking at the Dinucleotide Level.

Fluorine configuration at C2' of the bis(2'-fluorothymidine) dinucleotide is demonstrated to drive intramolecular base stacking. 2'-ß F-Configuration drastically reduces stacking compared to the 2'-α series. Hence, base stacking emerges as being tunable by the C2'-F stereoconfiguration through dramatic puckering variations scrutinized by NMR and natural bond orbital analysis. Accordingly, 2'-ß F-isomer photoreactivity is significantly reduced compared to that of the 2'-α F-isomer.

Synergistic AML Cell Death Induction by Marine Cytotoxin (+)-1(R), 6(S), 1'(R), 6'(S), 11(R), 17(S)-Fistularin-3 and Bcl-2 Inhibitor Venetoclax.

Treatment of acute myeloid leukemia (AML) patients is still hindered by resistance and relapse, resulting in an overall poor survival rate. Recently, combining specific B-cell lymphoma (Bcl)-2 inhibitors with compounds downregulating myeloid cell leukemia (Mcl)-1 has been proposed as a new effective strategy to eradicate resistant AML cells. We show here that 1(R), 6(S), 1'(R), 6'(S), 11(R), 17(S)-fistularin-3, a bromotyrosine compound of the fistularin family, isolated from the marine sponge Suberea clavata, synergizes with Bcl-2 inhibitor ABT-199 to efficiently kill Mcl-1/Bcl-2-positive AML cell lines, associated with Mcl-1 downregulation and endoplasmic reticulum stress induction. The absolute configuration of carbons 11 and 17 of the fistularin-3 stereoisomer was fully resolved in this study for the first time, showing that the fistularin we isolated from the marine sponge Subarea clavata is in fact the (+)-11(R), 17(S)-fistularin-3 stereoisomer keeping the known configuration 1(R), 6(S), 1'(R), and 6'(S) for the verongidoic acid part. Docking studies and in vitro assays confirm the potential of this family of molecules to inhibit DNA methyltransferase 1 activity.

Bioactive Bromotyrosine-Derived Alkaloids from the Polynesian Sponge Suberea ianthelliformis.

Herein, we describe the isolation and spectroscopic identification of eight new tetrabrominated tyrosine alkaloids 2⁻9 from the Polynesian sponge Suberea ianthelliformis, along with known major compound psammaplysene D (1), N,N-dimethyldibromotyramine, 5-hydroxy xanthenuric acid, and xanthenuric acid. Cytotoxicity and acetylcholinesterase inhibition activities were evaluated for some of the isolated metabolites. They exhibited moderate antiproliferative activity against KB cancer cell lines, but psammaplysene D (1) displayed substantial cytotoxicity as well as acetylcholinesterase inhibition with IC50 values of 0.7 μM and 1.3 μM, respectively.

Orbicularisine: A Spiro-Indolothiazine Isolated from Gills of the Tropical Bivalve Codakia orbicularis.

A novel spiro-indolofuranone fused to a thiazine skeleton, orbicularisine (1), was isolated from gills of the mollusk Codakia orbicularis. The isolation and structure elucidation using spectroscopic evidence including mass and NMR spectroscopy are described. The final structure of 1 was supported by key HMBC correlation.

Aedes aegypti Larvicidal Sesquiterpene Alkaloids from Maytenus oblongata.

Four new sesquiterpene alkaloids (1-4) with a ß-dihydroagrofuran skeleton and a new triterpenoid (5) were isolated from an ethyl acetate extract of Maytenus oblongata stems. Their structures were elucidated using 1D and 2D NMR spectroscopy as well as MS and ECD experiments. The M. oblongata stem EtOAc extract and the pure compounds isolated were tested for larvicidal activity against Aedes aegypti under laboratory conditions, and compounds 2 and 3 were found to be active.

Cytotoxic Guanidine Alkaloids from a French Polynesian Monanchora n. sp. Sponge.

Four bicyclic and three pentacyclic guanidine alkaloids (1-7) were isolated from a French Polynesian Monanchora n. sp. sponge, along with the known alkaloids monalidine A (8), enantiomers 9-11 of known natural product crambescins, and the known crambescidins 12-15. Structures were assigned by spectroscopic data interpretation. The relative and absolute configurations of the alkaloids were established by analysis of (1)H NMR and NOESY spectra and by circular dichroism analysis. The new norcrambescidic acid (7) corresponds to interesting biosynthetic variation within the pentacyclic core. All compounds exhibited antiproliferative and cytotoxic efficacy against KB, HCT116, HL60, MRC5, and B16F10 cancer cells, with IC50 values ranging from 4 nM to 10 µM.

A minute amount of s-puckered sugars is sufficient for (6-4) photoproduct formation at the dinucleotide level.

The di-2'-α-fluoro analogue of thymidylyl(3',5')thymidine, synthesized to probe the effect of a minimum amount of S conformer on the photoreactivity of dinucleotides, is endowed with only 3% and 8% of S sugar conformation at its 5'- and 3'-end, respectively. This analogue gives rise to the (6-4) photoproduct as efficiently as the dithymine dinucleotide (74% and 66% at the 5'- and 3'-end, respectively) under 254 nm. Our results suggest that the 5'-N, 3'-S conformer gives rise to the (6-4) photoproduct.

Netamines O-S, Five New Tricyclic Guanidine Alkaloids from the Madagascar Sponge Biemna laboutei, and Their Antimalarial Activities.

In our continuing program to isolate new compounds from the Madagascar sponge Biemna laboutei, five new tricyclic guanidine alkaloids, netamines O - S (1-5, resp.), have been identified together with the known compounds netamine E (6) and mirabilin J (7). The structures of all new netamines were assigned on the basis of spectroscopic analyses. Their relative configurations were established by analysis of ROESY data and comparison with literature data. Netamines O, P, and Q, which were isolated in sufficient quantities, were tested for their cytotoxic activities against KB cells and their activities against the malaria parasite Plasmodium falciparum. Netamines O and Q were found to be moderately cytotoxic. Netamines O, P, and Q exhibited antiplasmodial activities with IC50 values of 16.99 ± 4.12, 32.62 ± 3.44, and 8.37 ± 1.35 µM, respectively.

Agelastatin E, agelastatin F, and benzosceptrin C from the marine sponge Agelas dendromorpha.

The study of the n-butanol extract of the New Caledonian sponge Agelas dendromorpha led to the isolation and identification of three new pyrrole-2-aminoimidazole (P-2-AI) alkaloids, named agelastatins E (3) and F (4) and benzosceptrin C (5), together with 10 known metabolites, agelastatin A (1), agelastatin D (2), sceptrin (6), manzacidin A, tauroacidin A, taurodispacamide A, nortopsentin D, thymine, longamide, and 4,5-dibromopyrrole-2-carboxamide. Their structures were assigned by spectroscopic data interpretation. All the compounds were tested for cytotoxic activity.

Axiphenylalaninium and axityrosinium, modified amino acids from the Mediterranean marine sponge Axinella polypoides.

Two new modified amino acids, axiphenylalaninium (1) and axityrosinium (2), along with four known metabolites, C2-alpha-D-mannosylpyranosyl-L-tryptophan (3), N3,5'-cycloxanthosine (4), palythine (5), and taurine, were isolated from the marine sponge Axinella polypoides collected in the Mediterranean Sea. The structures were determined by spectroscopic studies and confirmed by X-ray analysis and chemical modifications.

The sugar conformation governs (6-4) photoproduct formation at the dinucleotide level.

The LNA dinucleotide mimic of TpT whose two-sugar puckers are locked in the C3'-endo conformation selectively produces the corresponding cyclobutane pyrimidine dimer under 254 nm irradiation. In the natural series (TpT) the sugar puckers are in a major C2'-endo sugar conformation and the (6-4) photoproduct is also produced. Consequently, this study demonstrates that the C2'-endo conformation of the sugar pucker is necessary for (6-4) photoproduct formation.

Unguiculins A-C: cytotoxic bis-guanidine alkaloids from the French Polynesian sponge, Monanchora n. sp.

Two new acyclic bis-guanidine alkaloids, unguiculins B-C (2-3), were isolated from a French Polynesian sponge Monanchora n. sp. together with the known compound unguiculin A (1). Their structures were established by spectroscopic data interpretation and comparison with the literature. Unguiculins A-C displayed antiproliferative and cytotoxic efficacy against several human cancer cells with IC50 values in the micromolar range.

Dinucleotide TpT and its 2'-O-Me analogue possess different backbone conformations and flexibilities but similar stacked geometries.

UV irradiation at 254 nm of 2'-O,5-dimethyluridylyl(3'-5')-2'-O,5-dimethyluridine (1a) and of natural thymidylyl(3'-5')thymidine (1b) generates the same photoproducts (CPD and (6-4)PP; responsible for cell death and skin cancer). The ratios of quantum yields of photoproducts obtained from 1a (determined herein) to that from 1b are in a proportion close to the approximately threefold increase of stacked dinucleotides for 1a compared with those of 1b (from previous circular dichroism results). 1a and 1b however are endowed with different predominant sugar conformations, C3'-endo (1a) and C2'-endo (1b). The present investigation of the stacked conformation of these molecules, by unrestrained state-of-the-art molecular simulation in explicit solvent and salt, resolves this apparent paradox and suggests the following main conclusions. Stacked dinucleotides 1a and 1b adopt the main characteristic features of a single-stranded A and B form, respectively, where the relative positions of the backbone and the bases are very different. Unexpectedly, the geometry of the stacking of two thymine bases, within each dinucleotide, is very similar and is in excellent agreement with photochemical and circular dichroism results. Analyses of molecular dynamics trajectories with conformational adiabatic mapping show that 1a and 1b explore two different regions of conformational space and possess very different flexibilities. Therefore, even though their base stacking is very similar, these molecules possess different geometrical, mechanical, and dynamical properties that may account for the discrepancy observed between increased stacking and increased photoproduct formations. The computed average stacked conformations of 1a and 1b are well-defined and could serve as starting models to investigate photochemical reactions with quantum dynamics simulations.

Metabolomics approach to chemical diversity of the Mediterranean marine sponge Agelas oroides.

The Mediterranean marine sponge Agelas oroides is known to contain a large quantity of oroidin, a deterrent, antifouling and antibiofilm pyrrole-2-aminoimidazole. In contrast with other tropical specimens, the chemical composition of Mediterranean Agelas oroides is surprisingly relatively poor in other related metabolites. In the course of finding novel marine natural products, LC-MS based metabolomics study of the Mediterranean Agelas oroides, however, revealed that next to the major compound oroidin, the sponge contains in fact a great diversity of known pyrrole-imidazole alkaloids in minute amounts. Here, we describe identification of 13 known oroidin class alkaloids along with one new monobromoagelaspongin (24). Five betaines and one amine were also identified from the aqueous fraction. One of those compounds (-)-equinobetaine B (30) was found to be an enantiomer of the known natural product (+)-equinobetaine B.