Australian Marine Sponge Research Articles

Identification of Axinellamines A and B as Anti-Tubercular Agents.

Tuberculosis remains a significant global health pandemic. There is an urgent need for new anti-tubercular agents to combat the rising incidence of drug resistance and to offer effective and additive therapeutic options. High-throughput screening of a subset of the NatureBank marine fraction library (n = 2000) identified a sample derived from an Australian marine sponge belonging to the order Haplosclerida that displayed promising anti-mycobacterial activity. Bioassay-guided fractionation of the organic extract from this Haplosclerida sponge led to the purification of previously identified antimicrobial pyrrole alkaloids, axinellamines A (1) and B (2). The axinellamine compounds were found to have a 90% minimum inhibitory concentration (MIC90) of 18 µM and 15 µM, respectively. The removal of protein and complex carbon sources reduced the MIC90 of 1 and 2 to 0.6 and 0.8 µM, respectively. The axinellamines were not toxic to mammalian cells at 25 µM and significantly reduced the intracellular bacterial load by >5-fold. These data demonstrate that axinellamines A and B are effective anti-tubercular agents and promising targets for future medicinal chemistry efforts.

Cyclotheonellazoles D-I, Potent Elastase Inhibitory Thiazole-Containing Cyclic Peptides from Theonella sp. (2131).

Six new thiazole-containing cyclic peptides, the cyclotheonellazoles D-I (1-6), were isolated from the Australian marine sponge Theonella sp. (2131) with their structures assigned by comprehensive 1D and 2D NMR spectroscopic and MS spectrometric analyses, Marfey's derivatization studies, and comparison with time-dependent density functional theory (TDDFT) calculated ECD data. The Type 2 azole-homologated peptides herein comprise up to five nonproteinogenic amino acids, including the protease transition state mimic α-keto-β-amino acid residue 3-amino-4-methyl-2-oxohexanoic acid (Amoha), while 1-3 also contain a terminal hydantoin residue not previously found in cyclotheonellazoles. The keramamides A (7) and L (8) were reisolated affording expanded exploration of their biological activities. The peptides were examined for protease inhibitory activities against two mammalian serine proteases (elastase and chymotrypsin) and SARS-CoV-2 3-chymotrypsin-like protease (3CLpro), a validated antiviral therapeutic target for COVID-19. Peptides 1-6 and keramamide A (7) displayed potent nanomolar inhibition of elastase (IC50 16.0 to 61.8 nM), while 7 also contained modest inhibition of chymotrypsin and SARS-CoV-2 3CLpro (IC50 0.73 and 1.1 μM, respectively). The cyclotheonellazoles D-E (1-3) do not affect the viability of human breast, ovarian, and colon cancer cells (>100 μM), with the cytotoxicity previously reported for keramamide L (8) not replicated (inactive >20 μM).

Using UHPLC-MS profiling for the discovery of new sponge-derived metabolites and anthelmintic screening of the NatureBank bromotyrosine library.

In order to further expand the NatureBank open access compound library, chemical investigations of the Australian marine sponge, Ianthella basta, were undertaken since UHPLC-MS analysis of the extract from this sponge indicated the presence of a new alkaloid. Large-scale extraction and mass-directed isolation studies on the CH2Cl2/MeOH I. basta extract resulted in the purification of a new bromotyrosine-derived alkaloid, 5-debromopurealidin H (1), along with the known marine natural product, ianthesine E (2). The chemical structure of the new compound was determined following detailed spectroscopic and spectrometric data analysis. These two compounds (1 and 2) along with seven previously reported marine bromotyrosine alkaloids from the NatureBank open access library, which included psammaplysins F (3) and H (4), bastadins 4 (5), 8 (6) and 13 (7), aerothionin (8) and hexadellin A (9), were evaluated for their nematocidal activity against exsheathed third-stage larvae of Haemonchus contortus, a highly pathogenic parasite of ruminants. Of the nine compounds, bastadin 8 (6), hexadellin A (9) and bastadin 4 (5) showed inhibition towards larval motility after 72 h of exposure with IC50 values of 1.6 µM, 10.0 µM and 33.3 µM, respectively.

Identification of Anthelmintic Bishomoscalarane Sesterterpenes from the Australian Marine Sponge Phyllospongia bergquistae and Structure Revision of Phyllolactones A-D.

High-throughput screening of the NatureBank marine extract library (7616 samples) identified an extract derived from the Australian marine sponge Phyllospongia bergquistae with activity against Hemonchus contortus (barber's pole worm), an economically important parasitic nematode. Bioassay-guided fractionation of the CH2Cl2/MeOH extract from P. bergquistae led to the purification of four known bishomoscalarane sesterterpenes, phyllolactones A-D (1-4). The absolute configurations of phyllolactones B (2) and C (3) were determined by single-crystal X-ray diffraction analysis; literature and data analyses revealed the need for these chemical structures to be revised. Compounds 2-4 induced a lethal, skinny (Ski) phenotype in larvae of H. contortus at concentrations between 5.3 and 10.1 μM. These data indicate that the bishomoscalarane sesterterpene structure class warrants further investigation for nematocidal or nematostatic activity.

Dysidenin from the Marine Sponge Citronia sp. Affects the Motility and Morphology of Haemonchus contortus Larvae In Vitro.

High-throughput screening of the NatureBank marine extract library (n = 7616) using a phenotypic assay for the parasitic nematode Haemonchus contortus identified an active extract derived from the Australian marine sponge Citronia sp. Bioassay-guided fractionation of the CH2Cl2/MeOH extract from Citronia sp. resulted in the purification of two known hexachlorinated peptides, dysidenin (1) and dysideathiazole (2). Compound 1 inhibited the growth/development of H. contortus larvae and induced multiple phenotypic changes, including a lethal evisceration (Evi) phenotype and/or somatic cell and tissue destruction. This is the first report of anthelmintic activity for these rare and unique polychlorinated peptides.

Sycosterol A, an α-Synuclein Inhibitory Sterol from the Australian Ascidian Sycozoa cerebriformis.

During a recent biodiscovery study to identify new α-synuclein (α-syn) aggregation inhibitors, we screened 29 Australian marine sponge and ascidian extracts in an MS binding assay. This resulted in an extract from the ascidian Sycozoa cerebriformis showing activity toward α-syn. The bioassay and MS guided isolation process led to the identification of one new polyoxygenated sterol sulfate, sycosterol A (1). The structure of this low-yielding steroid was elucidated from HRMS and NMR analysis. Sycosterol A displayed moderate antiaggregation activity with 46.2% (±1.8) inhibition when screened against α-syn at a 5:1 (sycosterol A:α-syn) molar ratio. The α-syn antiaggregation activity displayed by 1 and the recent discovery of similar sterols with α-syn antiaggregation activity and potent antiprion activity suggest this unique class may be useful antineurodegenerative compounds.

New from Old: Thorectandrin Alkaloids in a Southern Australian Marine Sponge, Thorectandra choanoides (CMB-01889)

Thorectandra choanoides (CMB-01889) was prioritized as a source of promising new chemistry from a library of 960 southern Australian marine sponge extracts, using a global natural products social (GNPS) molecular networking approach. The sponge was collected at a depth of 45 m. Chemical fractionation followed by detailed spectroscopic analysis led to the discovery of a new tryptophan-derived alkaloid, thorectandrin A (1), with the GNPS cluster revealing a halo of related alkaloids 1a–1n. In considering biosynthetic origins, we propose that Thorectandra choanoides (CMB-01889) produces four well-known alkaloids, 6-bromo-1′,8-dihydroaplysinopsin (2), 6-bromoaplysinopsin (3), aplysinopsin (4), and 1′,8-dihydroaplysinopsin (10), all of which are susceptible to processing by a putative indoleamine 2,3-dioxygenase-like (IDO) enzyme to 1a–1n. Where the 1′,8-dihydroalkaloids 2 and 10 are fully transformed to stable ring-opened thorectandrins 1 and 1a–1b, and 1h–1j, respectively, the conjugated precursors 3 and 4 are transformed to highly reactive Michael acceptors that during extraction and handling undergo complete transformation to artifacts 1c–1g, and 1k–1n, respectively. Knowledge of the susceptibility of aplysinopsins as substrates for IDOs, and the relative reactivity of Michael acceptor transformation products, informs our understanding of the pharmaceutical potential of this vintage marine pharmacophore. For example, the cancer tissue specificity of IDOs could be exploited for an immunotherapeutic response, with aplysinopsins transforming in situ to Michael acceptor thorectandrins, which covalently bind and inhibit the enzyme.

Citronamine A, an Antiplasmodial Isoquinoline Alkaloid from the Australian Marine Sponge Citronia astra.

Citronamine A (1), an isoquinoline alkaloid containing an unprecedented pentacyclic ring system, was isolated from the Australian marine sponge Citronia astra. Based on the combination of MS and NMR analyses and comparison of experimental and TDDFT calculated ECD spectra, the absolute structure of 1 was determined. Compound 1 displayed moderate activity against drug sensitive (3D7) and drug resistant (Dd2) strains of the parasite, Plasmodium falciparum, responsible for malaria.

Dysidealactams and Dysidealactones: Sesquiterpene Glycinyl-Lactams, Imides, and Lactones from a Dysidea sp. Marine Sponge Collected in Southern Australia.

A GNPS molecular networking approach mapped a library of 960 southern Australian marine sponges and prioritized Dysidea sp. (CMB-01171) for chemical investigation. Although the published natural products literature on Australian Dysidea sponges extends back over half a century and suffers from the perception of being near exhausted, fractionation of Dysidea sp. (CMB-01171) led to the discovery of a family of 10 new biosynthetically and chemically related sesquiterpenes. Detailed spectroscopic analysis guided structure elucidation identified dysidealactams A-F (1-6), dysidealactones A and B (7 and 8), and two solvolysis artifacts, 9 and 10. The dysidealactams A-D (1-4) incorporate a rare glycinyl-lactam functionality, while dysidealactam E (5) is particularly noteworthy in incorporating an unprecedented glycinyl-imide moiety. In addition to expanding knowledge of Dysidea natural products, this study demonstrates the value of applying GNPS molecular networking to map chemical diversity and prioritize the selection of marine sponge extracts for more detailed chemical analysis.

Structure Reassignment of Echinosulfone A and the Echinosulfonic Acids A-D Supported by Single-Crystal X-ray Diffraction and Density Functional Theory Analysis.

Two previously reported bis-indole alkaloids, echinosulfone A and echinosulfonic acid B, have been isolated for the first time from a Western Australian marine sponge, Crella sp. (order: Poecilosclerida, family: Crellidae). Single-crystal X-ray diffraction of a decomposition product of echinosulfone A prompted our investigation and subsequent structure reassignment of the echinosulfonic acid natural product family, which we report here. The reassignments are supported by analysis of 1D and 2D NMR data, MS fragmentation, and DFT calculations of 13C NMR shifts.

Trachycladindoles H–M: Molecular Networking Guided Exploration of a Library of Southern Australian Marine Sponges

A global natural product social (GNPS) molecular network guided search of a library of 960 southern Australian marine sponge extracts successfully detected a deep-water Great Australian Bight sponge, Geodia sp. (CMB-01063), as a new source of a rare class of indolo-imidazole alkaloids previously believed to be unique to a single specimen of Trachycladus laevispirulifer (CMB-03397). Chemical analysis of CMB-01063 detected the known trachycladindoles A–G (1–7), and led to the isolation, characterisation, and structure elucidation of the new trachycladindoles H–M (8–13). Structures for 8–13 were assigned on the basis of detailed spectroscopic analysis, with comparison to authentic standards of 1–7.

Identification of Fromiamycalin and Halaminol A from Australian Marine Sponge Extracts with Anthelmintic Activity against Haemonchus contortus.

There is an urgent need to discover and develop new anthelmintics for the treatment of parasitic nematodes of veterinary importance to circumvent challenges linked to drug resistant parasites. Being one of the most diverse natural ecosystems, the marine environment represents a rich resource of novel chemical entities. This study investigated 2000 extracts from marine invertebrates, collected from Australian waters, for anthelmintic activity. Using a well-established in vitro bioassay, these extracts were screened for nematocidal activity against Haemonchus contortus — a socioeconomically important parasitic nematode of livestock animals. Extracts (designated Mu-1, Ha-1 and Ha-2) from two marine sponges (Monanchora unguiculata and Haliclona sp.) each significantly affected larvae of H. contortus. Individual extracts displayed a dose-dependent inhibition of both the motility of exsheathed third-stage larvae (xL3s) and the development of xL3s to fourth-stage larvae (L4s). Active fractions in each of the three extracts were identified using bioassay-guided fractionation. From the active fractions from Monanchora unguiculata, a known pentacyclic guanidine alkaloid, fromiamycalin (1), was purified. This alkaloid was shown to be a moderately potent inhibitor of L4 development (half-maximum inhibitory concentration (IC50) = 26.6 ± 0.74 µM) and L4 motility (IC50 = 39.4 ± 4.83 µM), although it had a relatively low potency at inhibiting of xL3 motility (IC50 ≥ 100 µM). Investigation of the active fractions from the two Haliclona collections led to identification of a mixture of amino alcohol lipids, and, subsequently, a known natural product halaminol A (5). Anthelmintic profiling showed that 5 had limited potency at inhibiting larval development and motility. These data indicate that fromiamycalin, other related pentacyclic guanidine alkaloids and/or halaminols could have potential as anthelmintics following future medicinal chemistry efforts.

Solvolysis Artifacts: Leucettazoles as Cryptic Macrocyclic Alkaloid Dimers from a Southern Australian Marine Sponge, Leucetta sp.

Chemical analysis of a southern Australian sponge, Leucetta sp., led to the discovery of a pair of solvolysis adducts, leucettazoles A1 (1a) and B1 (2a), as artifacts of an unprecedented family of 15-membered macrocyclic alkaloid dimers featuring a pair of imino bridged 2-aminoimidazoles, together with a putative monomeric precursor, leucettazine A (3). The dimeric alkaloids 1a and 2a, and monomer 3, were identified by detailed spectroscopic analysis, supported by chemical transformations, analytical mass spectrometry, and biosynthetic considerations. Global natural product social networking (GNPS) molecular analysis of crude sponge extracts and solvent partitions, supported by single ion extraction (SIE) and diagnostic MS/MS fragmentations, revealed the associated natural products, leucettazoles A (1) and B (2). This study highlights that the study of natural product artifacts can be useful, and can on occasion serve as a pathway to discover cryptic new classes of natural products.

Three New Spongian Diterpenes from the Marine Sponge Dendrilla rosea

Three new diterpenes, aplyroseols 20–22 (2–4), were isolated from two specimens of the Australian marine sponge Dendrilla rosea. The structures and relative configurations of the new metabolites were elucidated from analysis of 2D NMR data. The compounds were screened for activity against Staphylococcus aureus, but were inactive at 64µgmL−1.

Cacolides: Sesterterpene Butenolides from a Southern Australian Marine Sponge, Cacospongia sp.

Chemical analysis of a marine sponge, Cacospongia sp. (CMB-03404), obtained during deep sea commercial fishing activities off the southern coast of Australia, yielded an unprecedented family of sesterterpene α-methyl-γ-hydroxybutenolides, cacolides A–L (1–12), together with biosynthetically related norsesterterpene carboxylic acids, cacolic acids A–C (13–15). Structures were assigned on the basis of detailed spectroscopic analysis with comparisons to known natural products and biosynthetic considerations. In addition to revealing new chemical diversity, this study provided a valuable platform for comparing and contrasting the capabilities of the traditional dereplication technologies of HPLC-DAD, HPLC-MS and NMR, with those of the emerging HPLC-MS/MS approach known as global natural products social molecular networking (GNPS), as applied to marine sponge sesterterpene tetronic acids.

In vitro protective activity of South Australian marine sponge and macroalgae extracts against amyloid beta (Aβ1–42) induced neurotoxicity in PC-12 cells

South Australia is a biodiversity hotspot of marine sponges and macroalgae. This study aimed to evaluate the potential neuroprotective activity of extracts from these two marine sources by reducing the toxicity of human amyloid beta Aβ1–42 in a cell model assay using PC-12 cells. A total of 92 extracts (43, 13, 16, and 20 extracts from sponge of 8 orders and 17 families, green algae of 3 orders and 4 families, brown algae of 6 orders and 8 families, and red algae of 5 orders and 10 families, respectively) were initially screened at three different concentrations (0.25, 2.5 and 25 μg/mL) to evaluate their toxicity using the MTT assay. About half of these extracts (26, 6, 5, and 10 extracts from sponge, green algae, brown algae, and red algae, respectively) showed some cytotoxicity, and were hence excluded from further assays. The rest of extracts (45 extracts in total) at 0.25 and 25 μg/mL were subsequently screened in a neuroprotection assay against Aβ1–42 cytotoxicity. A cell viability reduction of 30% was observed in the MTT assay when the cells were treated with 1 μM Aβ1–42. 29 extracts (13, 4, 7, and 5 extracts from sponge, green algae, brown algae, and red algae, respectively) reduced the toxicity induced by Aβ1–42 (P < 0.05), indicating neuroprotective activity. These results demonstrate that marine sponge and macroalgae form a broad spectrum are promising sources of neuroprotective compounds against the hallmark neurotoxic protein in Alzheimer's disease (AD).

Enantiodivergence in the Biosynthesis of Bromotyrosine Alkaloids from Sponges?

The isolation of bromotyrosine alkaloids, some of which are enantiomers of previously isolated compounds, has highlighted a possible enantiodivergence in their biosynthesis. Two new (1, 2) and six known bromotyrosine alkaloids (4-9), and the enantiomer (10) of a known compound, have been isolated from a Western Australian marine sponge, Pseudoceratina cf. verrucosa. The compounds inhibited the growth of multidrug-resistant and methicillin-resistant Staphylococcus aureus with comparable activity to vancomycin. In addition, one possible artifact of extraction (3) containing an ethoxy group was isolated. From analysis of the known bromotyrosine alkaloids, a biogenesis is proposed that explains the formation of antipodal natural products within this family of sponges.

Computational strategies to explore antimalarial thiazine alkaloid lead compounds based on an Australian marine sponge Plakortis Lita

In this work, an attempt was made to propose new leads based on the natural scaffold Thiaplakortone-A active against malaria. The 2D QSAR studies suggested that three descriptors correlate with the anti-malarial activity with an R2 value of 0.814. Robustness, reliability, and predictive power of the model were tested by internal validation, external validation, Y-scrambling, and applicability domain analysis. HQSAR studies were carried out as an additional tool to find the sub-structural fingerprints. The CoMFA and CoMSIA models gave Q2 values of 0.813 and 0.647, and values of 0.994 and 0.984, respectively. Using the 2D-QSAR equation, the activity values of the seven modified compounds were calculated and it was found that three molecules showed good anti-malarial activity. Molecular docking of the 42 Thiaplakortone-A derivatives with Plasmodium falciparum calcium-dependent protein kinase 1 (PfCDPK1) was carried out to find out protein-ligand interactions. Data mining of the bioassay data-set AID: 504850 using the classifier based on Random Forest of Weka suggested that all of the eight molecules selected and three out of the seven virtual molecules were anti-malarial active. Both the virtual molecules and drug molecules were docked with CYP3A4, indicating that the virtual molecules could metabolize easily. Toxicity studies using Osiris shows that three molecules showed no toxic characters.

Synthesis and anticancer evaluation of spermatinamine analogues.

Spermatinamine was isolated from an Australian marine sponge, Pseudoceratina sp. as an inhibitor of isoprenylcysteine carboxyl methyltransferase (Icmt), an attractive and novel anticancer target. Herein, we report the synthesis of spermatinamine analogues and their cytotoxic evaluation against three human cancer cell lines, that is, cervix adenocarcinoma (HeLa), breast adenocarcinoma (MCF-7), and prostate carcinoma (DU145). Analogues 12, 14 and 15 were found to be the most potent against one or more cell lines with the IC50 values in the range of 5-10 μM. The obtained results suggested that longer polyamine linker along with aromatic oxime substitution provided the most potent analogue compounds against cancer cell lines.

Concise Total Synthesis of Aplysinellamides A and B

Concise and efficient total syntheses of bromotyrosine-derived metabolites aplysinellamides A and B, isolated from Australian marine sponge Aplysinella sp., have been accomplished in seven steps. A condensation between cinnamic acid and Boc-D-lysine methyl ester was applied to form the amide skeleton as a key step.