Bromo-spiroisoxazoline Alkaloids, Including an Isoserine Peptide, from the Caribbean Marine Sponge Aplysina lacunosa.

Three new bromotyrosine spiroisoxazoline alkaloids, lacunosins A and B (1 and 2) and desaminopurealin (3), were isolated from a MeOH extract of the marine sponge Aplysina lacunosa that showed modest α-chymotrypsin inhibitory activity. The structures of 1-3 share the spirocyclohexadienyl-isoxazoline ring system found in purealidin-R and several other Verongid sponge secondary metabolites. Compounds 1 and 2 are coupled to a glycine and an isoserine methyl ester, respectively. Alkaloid 3 is linked, contiguously, to an O-1-aminopropyl 3,5-dibromotyrosyl ether and, finally, to histamine through an amide bond. The planar structures of all three compounds were obtained from analysis of MS and 1D and 2D NMR data. The absolute configuration of the SIO unit of 1-3 was assigned by electronic circular dichroism (ECD). The isoserine amino acid residue in 2 was found to be a 1:1 mixture of epimers using a new Marfey's type reagent, derived from Trp-NH2. Allylic O-naphthoylation of the SIO subunit enhances the ECD spectrum of SIOs and improves discrimination of enantiomorphs. A unifying hypothesis is proposed that links the biosynthesis of several of the new compounds with previously reported analogues.

Cyclic azole-homologated peptides from Marine sponges.

This review discusses the chemistry of cyclic azole-homologated peptides (AHPs) from the marine sponges, Theonella swinhoei, other Theonella species, Calyx spp. and Plakina jamaicensis. The origin, distribution of AHPs and molecular structure elucidations of AHPs are described followed by their biosynthesis, bioactivity, and synthetic efforts towards their total synthesis. Reports of partial and total synthesis of AHPs extend beyond peptide coupling reactions and include creative construction of the non-proteinogenic amino acid components, mainly the homologated heteroaromatic and α-keto-ß-amino acids. A useful conclusion is drawn regarding AHPs: despite their rarity, exotic structures and the potent protease inhibitory properties of some members, their synthesis is under-developed and beckons solutions for outstanding problems towards their efficient assembly.

Cyclic Hexapeptide Dimers, Antatollamides A and B, from the Ascidian Didemnum molle. A Tryptophan-Derived Auxiliary for l- and d-Amino Acid Assignments.

Two dimerized cyclic hexapeptides, antatollamides A (1) and B (2), were isolated from the colonial ascidian Didemnum molle collected in Pohnpei. The amino acid compositions and sequences were determined by interpretation of MS and 1D and 2D NMR data. Raney Ni reduction of antatollamide A cleaved the dimer to the corresponding monomeric cyclic hexapeptide with replacement of Cys by Ala. The amino acid configuration of 1 was established, after total hydrolysis, by derivatization with a new chiral reagent, (5-fluoro-2,4-dinitrophenyl)-Nα-l-tryptophanamide (FDTA), prepared from l-Trp, followed by LCMS analysis; all amino acids were found to be l-configured except for d-Ala.

Peroxide Natural Products from Plakortis zyggompha and the Sponge Association Plakortis halichondrioides-Xestospongia deweerdtae: Antifungal Activity against Cryptococcus gattii.

Cryptococcus gattii is a human pathogen and causative agent of a pernicious, sometimes fatal, disseminated fungal disease. Investigation of antifungal extracts of the marine sponge association Plakortis halichondrioides-Xestospongia deweerdtae and the sponge Plakortis zyggompha from the Bahamas led to the discovery and isolation of 6-epi-7,8-dihydroplakortide K (1), plakortide AA (2), and three new plakinic acids, N-P (4-6; unstable 1,2-dioxolanes bearing benzyl-substituted conjugated dienes), along with known plakinic acids L, K, and M.5 Chiroptical comparisons and DFT calculations of (13)C NMR chemical shifts were used to assign the absolute stereostructure of 4. The stereospecific base-promoted rearrangement-saponification of 1 to 10 was briefly investigated and showed tight kinetic control and stereospecific formation of the new C-2 stereocenter with inversion at C-3. Plakinic acid M and plakortides 9 and 11 exhibited antifungal activity against C. gattii (MIC90 = 2.4 to 36 µM), but plakinic acids N-P were inactive under the same conditions.

Cytotoxic effects on tumour cell lines of fatty acids from the marine sponge Scopalina ruetzleri.

OBJECTIVES: Marine sponges are among the most promising sources of chemically diversified fatty acids (FAs). In addition, several studies have shown the effect of polyunsaturated FAs on cancer therapy. This research carried out a biological and chemical evaluation of the sponge Scopalina ruetzleri collected on the South Brazilian coastline. METHODS: Bioassay-guided fractionation of S. ruetzleri was performed in human glioma (U87) and neuroblastoma (SH-SY5Y) cell lines, and the in-vitro effects on free radicals were evaluated. KEY FINDINGS: The ethyl acetate fraction of S. ruetzleri showed promising cytotoxic effects in cancer cell lines, with IC50 < 20 µg/ml. Fingerprint (1) H Nuclear Magnetic Resonance (NMR) analysis showed that this fraction is mainly constituted of FAs. Through FA methyl ester analysis, it was possible to identify 32 FAs. In addition, some minor unusual FAs for the marine biosphere were identified. The results of conjugated dienes method showed that FAs fraction, at concentrations above 50 µg/ml, has a pro-oxidant effect, indicating that lipid peroxidation may be partially responsible for the mechanism of cytotoxicity on cancer cells. CONCLUSION: This work also contributes to studies that focus on the application of FAs on cancer therapy as a new adjuvant to radio or chemotherapy, or as a chemotherapeutic agent.

Potent fluorinated agelastatin analogues for chronic lymphocytic leukemia: design, synthesis, and pharmacokinetic studies.

Chronic lymphocytic leukemia (CLL) is the most common lymphoid neoplasia in Western societies and is currently incurable. Multiple treatment options are practiced, but the available small molecule drugs suffer from dose-limiting toxicity and undesirable side effects. The need for new, less toxic treatments is a pressing concern. Here, we demonstrate that (-)-agelastatin A (1a), a pyrrole-imidazole alkaloid obtained from a marine sponge, exhibits potent in vitro activity against primary cell lines of CLL and disclose the synthesis of several analogues that are equipotent or exceed the potency of the natural product. The novel synthetic analogue, 13-debromo-13-trifluoromethyl agelastatin A (1j), showed higher activity than the natural product when tested against the same cell lines and is the most potent agelastatin derivative reported to date. A detailed in vitro structure-activity relationship of 1a in CLL compared to that of 22 synthetic analogues is described along with preliminary in vivo pharmacokinetic and metabolism studies on the most potent compounds.

Malignant hyperthermia susceptibility arising from altered resting coupling between the skeletal muscle L-type Ca2+ channel and the type 1 ryanodine receptor.

Malignant hyperthermia (MH) susceptibility is a dominantly inherited disorder in which volatile anesthetics trigger aberrant Ca(2+) release in skeletal muscle and a potentially fatal rise in perioperative body temperature. Mutations causing MH susceptibility have been identified in two proteins critical for excitation-contraction (EC) coupling, the type 1 ryanodine receptor (RyR1) and Ca(V)1.1, the principal subunit of the L-type Ca(2+) channel. All of the mutations that have been characterized previously augment EC coupling and/or increase the rate of L-type Ca(2+) entry. The Ca(V)1.1 mutation R174W associated with MH susceptibility occurs at the innermost basic residue of the IS4 voltage-sensing helix, a residue conserved among all Ca(V) channels [Carpenter D, et al. (2009) BMC Med Genet 10:104-115.]. To define the functional consequences of this mutation, we expressed it in dysgenic (Ca(V)1.1 null) myotubes. Unlike previously described MH-linked mutations in Ca(V)1.1, R174W ablated the L-type current and had no effect on EC coupling. Nonetheless, R174W increased sensitivity of Ca(2+) release to caffeine (used for MH diagnostic in vitro testing) and to volatile anesthetics. Moreover, in Ca(V)1.1 R174W-expressing myotubes, resting myoplasmic Ca(2+) levels were elevated, and sarcoplasmic reticulum (SR) stores were partially depleted, compared with myotubes expressing wild-type Ca(V)1.1. Our results indicate that Ca(V)1.1 functions not only to activate RyR1 during EC coupling, but also to suppress resting RyR1-mediated Ca(2+) leak from the SR, and that perturbation of Ca(V)1.1 negative regulation of RyR1 leak identifies a unique mechanism that can sensitize muscle cells to MH triggers.

Symplocin A, a linear peptide from the Bahamian cyanobacterium Symploca sp. Configurational analysis of N,N-dimethylamino acids by chiral-phase HPLC of naphthacyl esters.

The absolute stereostructures of the components of symplocin A (3), a new N,N-dimethyl-terminated peptide from the Bahamian cyanobacterium Symploca sp., were assigned from spectroscopic analysis, including MS, 2D NMR, and Marfey's analysis. The complete absolute configuration of symplocin A, including the unexpected D-configurations of the terminal N,N-dimethylisoleucine and valic acid residues, was assigned by chiral-phase HPLC of the corresponding 2-naphthacyl esters, a highly sensitive, complementary strategy for assignment of N-blocked peptide residues where Marfey's method is ineffectual or other methods fall short. Symplocin A exhibited potent activity as an inhibitor of cathepsin E (IC(50) 300 pM).

Antifungal Diterpene Alkaloids from the Caribbean Sponge Agelas citrina: Unified Configurational Assignments of Agelasidines and Agelasines.

Three new diterpene alkaloids - the hypotaurocyamines, (-)-agelasidines E and F (5-6), and the adeninium salt, agelasine N (9) - were isolated from the Caribbean sponge Agelas citrina along with six known natural products agelasines B-E (7, 10-12), 2-oxo-agelasine B (8), and (-)-agelasidine C (3). The chemical structures of 5, 6 and 9 were elucidated by analysis of NMR spectra and mass spectrometry. This represents the first report of natural products from the sponge A. citrina. Unified assignment of absolute configurations of the new compounds and known compounds were achieved by chemical correlation, quantitative measurements of molar rotations, and comparative analysis by van't Hoff's principle of optical superposition. (-)-Agelasidine C (3) exhibited potent antifungal and modest cytotoxic activity against human chronic lymphocytic leukemia (CLL) cells.

Mollenyne A, a long-chain chlorodibromohydrin amide from the sponge Spirastrella mollis.

The structure of mollenyne A, a cytotoxic nitrogenous halogenated long-chain carboxamide from the sponge Spirastrella mollis, was elucidated by integrated spectroscopic analysis, including CD, and chemical conversion.

N,N'-methyleno-didemnin A from the ascidian Trididemnum solidum. Complete NMR assignments and confirmation of the imidazolidinone ring by strategic analysis of 1J(CH).

The complete NMR assignments of N,N'-methyleno-didemnin A from Trididemnum solidum Van Name 1902 are reported along with a strategic analysis of (1)J(CH) coupling constants that confirm the presence of the imidazolidinone ring.

Zwittermicin A: synthesis of analogs and structure-activity studies.

Analogs and diastereomers of the natural product zwittermicin A were prepared. SAR studies of these compounds reveal the antifungal activity to be dependent singularly upon the natural constitution and configuration.

Isorhizochalin: a minor unprecedented bipolar sphingolipid of stereodivergent biogenesis from the Rhizochalina incrustata.

Isorhizochalin (1) was isolated as its peracetate from the EtOH extract of the sponge Rhizochalina incrustata. Its structure and absolute stereochemistry were elucidated as (2S,3R,26R,27R)-2,27-diamino-3-O-beta-D: -galactopyranosyl-oxy-26-hydroxyoctacosan-18-one by extensive NMR, MS studies, chemical transformations, including micromolar-scale Baeyer-Villiger oxidation, and by analysis of CD spectra of isorhizochalinin perbenzoate (2b). Isorhizochalin is an unprecedented C-2 epimer of rhizochalin having an erythro configuration at the glycosylated 2-amino-3-alkanol alpha-terminus in contrast with a canonical threo configuration for other representatives of this structural group. Probable biogenesis of 1 is discussed in the context of known sphingolipid biosynthesis beginning with condensation of alanine with a fatty acyl CoA thioester. The aglycone, isorhizochalinin (2a), shows cytotoxicity against human leukemia HL-60 and THP-1 cells with IC(50) values of 2.90 and 2.20 microM, respectively.

(+)-Zwittermicin A. Rapid assembly of C9-C15 and a formal total synthesis.

A short, enantioselective synthesis of the C9-C15 portion of (+)-zwittermicin A is reported that exploits directional functionalization of the known hepta-2,5-diyne-1,7-diol by partial reduction of the two triple bonds followed by Sharpless asymmetric epoxidation and boron-directed double ring-opening with sodium azide under Miyashita conditions. Subsequent desymmetrization of the C(2)-symmetric diazidotetraol product converges upon (-)-3--the enantiomer of the key intermediate of our earlier structural proof and synthesis of (-)-zwittermicin A--and constitutes a formal synthesis of (+)-zwitttermicin A.

Hemi-phorboxazole a: structure confirmation, analogue design and biological evaluation.

A synthesis providing totally synthetic (+)-hemi-phorboxazole A (1), proceeding in two steps (85% yield) from known vinyl iodide precursor (+)-2, has been achieved in conjunction with the design, synthesis, and biological evaluation of two hemi-phorboxazole analogues [(+)-3 and (-)-4] featuring ring replacements inscribed within the macrolide. Although hemi-phorboxazole A (1) displayed no activity when tested against Candida albicans and two human cancer cell lines, analogue (-)-4 exhibited significant tumor cell growth inhibitory activity in the nanomolar range against HCT-116 (colon) and SK-BR-3 (breast), while (+)-3 displayed promising antifungal activity against C. albicans.

Drug development from marine natural products.

Drug discovery from marine natural products has enjoyed a renaissance in the past few years. Ziconotide (Prialt; Elan Pharmaceuticals), a peptide originally discovered in a tropical cone snail, was the first marine-derived compound to be approved in the United States in December 2004 for the treatment of pain. Then, in October 2007, trabectedin (Yondelis; PharmaMar) became the first marine anticancer drug to be approved in the European Union. Here, we review the history of drug discovery from marine natural products, and by describing selected examples, we examine the factors that contribute to new discoveries and the difficulties associated with translating marine-derived compounds into clinical trials. Providing an outlook into the future, we also examine the advances that may further expand the promise of drugs from the sea.

Phorbasides A-E, cytotoxic chlorocyclopropane macrolide glycosides from the marine sponge Phorbas sp. CD determination of C-methyl sugar configurations.

Five new cytotoxic macrolide glycosides phorbasides A-E (3-7), each possessing a macrolide ring appended to a rare ene-yne-trans-2-chlorocyclopropane, were isolated from the same Western Australian sponge (Phorbas sp.) that provided phorboxazoles A and B. The structures of 3-7 were solved by analysis of spectroscopic data including NMR, MS, and CD. A synthesis of methyl 2-O-methyl-alpha-L-evalose from L-rhamnose was completed and used for configurational assignment of the sugar residue in 3. Acid-catalyzed methanolysis of 3 followed by two-step derivatization of the liberated O-methyl glycoside gave a vicinal 4-O-naphthoyl/tertiary 3-N-(2-aminonaphthyl)carbamate derivative that exhibited exciton coupled CD identical with that of the derivative prepared from synthetic 1,2- O-dimethyl-alpha-L-evalose.

Long-chain 2H-azirines with heterogeneous terminal halogenation from the marine sponge Dysidea fragilis.

Three new omega-halogenated long-chain 2H-azirines were isolated from the sponge Dysidea fragilis. Their structures revealed heterogeneity in both the composition of the terminal 1,1-dihalo-vinyl group and enantiomeric ratios at C2 of the azirine-2-carboxylate ester terminus. Azirine-2-carboxylate esters were shown to racemize spontaneously. A hypothesis is proposed for the biosynthesis of the azirinecarboxylate family of natural products that involves enzyme-catalyzed free radical halogenation followed by elimination of hydrohalic acid.

Asymmetric synthesis of diastereomeric diaminoheptanetetraols. A proposal for the configuration of (+)-zwittermicin a.

[structure: see text] A proposed absolute configuration for the 7 stereocenters in (+)-zwittermicin A is described based on asymmetric synthesis of six diastereomeric 2,6-diamino-1,3,5,7-heptanetetraols corresponding to the C9-C15 segment, pairwise 13C NMR chemical shift difference analysis of the models with the natural product, interpretation of enantiospecificity of the serine loading domain of the zwittermicin A biosynthetic gene cluster, and degradation of the natural product.